HIGH THROUGHPUT ENGINEERING OF FUNCTIONAL AAV CAPSIDS

2. SEQUENCE LISTING

The instant application contains a Sequence Listing with XX sequences, which has been submitted via EFS-Web and is hereby incorporated herein by reference in its entirety. Said ASCII copy, created on XXXX, is named 45736WO_sequencelisting.txt, and is XXX bytes in size.

3. BACKGROUND

Recombinant adeno-associated viruses (rAAV) provide the leading platform for in vivo delivery of gene therapies. Current clinical trials employ a limited number of AAV capsids, primarily from naturally occurring human or primate serotypes such as AAV1, AAV2, AAV5, AAV6, AAV8, AAV9, AAVrh.10, AAV4rh.74, and AAVhu.67. These capsids often provide suboptimal targeting to tissues of interest, both due to poor infectivity of the tissue of interest and competing liver tropism. Increasing the dose to ensure infection of desired tissues can lead to dose-dependent liver toxicity. In addition, use of naturally-occurring capsids presents an immunological memory challenge—pre-immune patient populations are excluded from treatment and repeat dosing in a previously immune naïve patient is often not possible. Thus, there is a need for additional AAV capsids for use in gene therapy, in particular capsids that confer upon the rAAV high infectivity for specific tissues and low liver tropism

4. SUMMARY OF THE INVENTION

We have designed a system for high throughput engineering of functional AAV capsids with altered tropism for various tissues, and using this system have identified capsid variants that have either increased or reduced liver tropism, and increased tropism for target tissues, such as liver or central nervous system (CNS) tissues.

Provided herein is an engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide having an amino acid sequence at least 70% identical to SEQ ID NO: 1, wherein the engineered AAV VP capsid polypeptide has at least one mutation as compared to SEQ ID NO: 1 in the region from a residue corresponding to residue 581 of SEQ ID NO: 1 to a residue corresponding to residue 589 of SEQ ID NO: 1, inclusive, wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV), wherein the at least one mutation confers higher tropism for a central nervous system (CNS) tissue on the rAAV as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1, and wherein the engineered AAV VP capsid polypeptide does not have the sequence of any of SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8. In certain embodiments, the engineered AAV VP capsid polypeptide has a sequence of SEQ ID NO: 2 and wherein Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from any amino acid.

Additionally, provided herein is an engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide having an amino acid sequence of SEQ ID NO: 2, wherein amino acid residues Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V, wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV), and wherein the engineered AAV VP capsid polypeptide does not have the sequence of any of SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8. In certain embodiments, the rAAV has higher tropism for a central nervous system (CNS) tissue as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1.

5. BRIEF DESCRIPTION OF THE DRAWINGS

These and other features, aspects, and advantages of the present invention will become better understood with regard to the following description, and accompanying drawings where:

FIG. 1 is a schematic of the high throughput AAV capsid engineering system.

FIG. 2A provides a side view (top panel) and top view (bottom panel) of key residues of known AAV capsids that have been shown to interact with target cells. FIG. 2B illustrates salient features of the AAV capsid library described in Example 1, showing the region of introduced diversity, with residue numbering corresponding to the numbering of amino acids in AAV5 VP1.

FIG. 3 is a gel photograph showing effective removal of the linear insert and linear plasmid backbone (“BB”) following digestion with PS-DNAse, enriching “relaxed form” (“RF”) circular species for highly efficient subsequent bacterial transformation.

FIG. 4 is a schematic of the high throughput AAV capsid engineering system. The column graph on the left shows the size of the AAV capsid library at various stages of preparation, tracking the size of the library from theoretical diversity, synthesized capsid genes, total cloned variants, to sequenced assembled viruses (error bars denote minimum/maximum predicted diversity from NGS sequencing analysis). The pooled viral library is injected into non-human primates and following a period of time sufficient to ensure stable transduction, the animals are euthanized, and tissues are harvested. DNA is purified from the tissues (1), and (2) a unique molecular identifier (UMI) is appended. Exonuclease I is added to digest excess UMI-containing primers (3). Subsequent PCR amplification adds sequencing indexes/tissue barcoding and adapters for next generation sequencing (NGS) sequencing. The addition of UMIs allows for high resolution frequency analysis of capsid variants in the tissues.

FIGS. 5A-5C illustrates the packaging approach used to maximize rAAV production from the library, with FIG. 5A comparing standard triple transfection (top panel) to the two-plasmid cis packaging approach used herein (bottom panel), and FIG. 5B showing relative production of wild type AAV5 using the two transfection approaches. FIG. 5C is an example of UMI distribution in a liver specimen and shows a majority of capsid variants are found a single time (single UMI). However, a subset of capsid variants is enriched in a given tissue and has increased numbers of UMIs, corresponding to their increased frequency in the target tissue.

FIGS. 6A-6B are heat maps and a statistical analysis showing clear functional selection through the stages of viral assembly and tissue transduction. FIG. 6A is a heat map prepared at various stages of the high throughput system from pre-assembly, assembled virus, to liver transduction. For the pre-assembly heat map, the library of capsid variants was cloned into plasmids and transformed into electrocompetent bacteria. The resultant library was sequenced and amino acid diversity was measured at each position, with the low levels of positional variation likely arising from synthesis error. Subsequent assembly into virus shows clear amino acid residue/positional biases that highlight selection for viral assembly. Liver transducing viruses show even greater patterns of AA residue/positional selection, with distinctly favored/disfavored variants. Note that each of these heatmaps is normalized by their respective input frequencies. FIG. 6B is a table of statistical analysis (ANOVA) showing that in three liver samples, amino acid residue distribution and residue-position vary significantly, but inter-sample variation is not significant.

FIGS. 7A-7B show analysis of repeat observed (high UMI) capsid variants between multiple samples. FIG. 7A is a Venn diagram illustrating counts of overlapping variant identification in two liver samples: ˜38% of variants with >10 UMIs were observed in both samples. FIG. 7B is a heat map showing positional amino acid distribution illustrating the strongly selected residues/positions of repeat observed capsid variants.

FIG. 8 is a schematic of machine learning-based clustering of capsid variants. The example utilizes capsid variant sequences upon which machine learning algorithms were used to map the similarity of capsids among those that infected the liver. This output can then be used to inform selection of candidate variants to selectively target the tissue of interest.

FIG. 9 illustrates an example of the performance of the library as a whole infecting cortex at a higher relative level than liver after the intravenous administration to a non-human primate (NHP). DNA was isolated from the liver and cortex and either qPCR (at left) or droplet digital PCR (ddPCR) (at right) were used to quantify viral genomes recovered from the respective tissues. By these methods of quantification, the library as a whole has ˜2-fold increased CNS targeting over the liver.

FIG. 10 illustrates wild-type crystal structure of AAV5 capsid emphasizing electrostatic potential, and two exemplary recombinant VP1 capsid variants obtained by the methods described herein.

FIGS. 11A-D show Venn diagrams and analytic tables depicting the number of unique sequence variants found in liver tissue only, liver and brain cortex tissues, and cortex tissue only for three different sequencing analysis filters. Next-generation sequencing was performed on viral genomic capsid variants recovered from liver or cortical tissue samples. Unique Molecular Identifiers (UMIs) were appended as part of the molecular recovery process as described herein. This allows for increased confidence that a given capsid variant is present in the tissue, and may be correlated to abundance. FIG. 11A shows a Venn diagram in which the sequencing analysis filter applied was 4 or greater distinct UMIs (also referred to herein as “count”), per each distinct capsid sequence variant. FIG. 11B shows a Venn diagram in which the sequencing analysis filter applied was 50 or greater UMIs. FIG. 11C shows a Venn diagram in which the sequencing analysis filter applied was 100 or greater distinct UMIs. FIG. 11D shows a histogram analysis of the distribution of UMIs in the population of capsid sequence variants found in cortex only in which the sequencing analysis filter applied was 50 or greater UMIs as shown in FIG. 11B.

FIG. 12 shows heatmaps of normalized amino acid residue frequency in the rAAV5 capsid polypeptide sequences of the present disclosure and how they are enriched moving from pre-assembly to post-assembly to CNS-transducing subsets of variants. The x-axes of the heatmaps indicate the position in the 581-589 region and the y-axis includes all amino acid residues.

FIG. 13 shows an example of machine learning (ML) model performance validation. Distinct variants are found in CNS and non-CNS tissues, as well as some shared variants found in both (left). Random Forest (RF) ML models show good performance at predicting CNS targeting. At high predicted class probability values, the ML model can resolve CNS-targeting from non-CNS targeting variant sequences (middle, right).

FIG. 14 illustrates top 20 positional features contributing to model output probability (at left). Shapley Additive Explanations (SHAP) values can be used to interrogate the relative contribution of features to model predictions. These features can be further compared between tissue targeting and non-targeting variants. At right is a model of a CNS variant having a sequence of KRLQQMETM (SEQ ID NO: 1117), representing some features predicted to increase CNS-targeting.

FIG. 15 shows recovery of rAAV5 variants of the present disclosure from two NHPs across all tissue types including skin, liver, lung, heart, spleen, lymph node, thyroid gland, skeletal muscle, bone marrow, mammary gland, adrenal gland, colon, sciatic nerve (a peripheral nerve), and a number of CNS tissues (forebrain cortex, occipital cortex, temporal cortex, thalamus, hypothalamus, substantia nigra, hippocampus DG, hippocampus CA1, hippocampus CA3, cerebellum, and spinal cord). Analysis of tissue samples enabled the ability to enrich favored properties of a particular variant (e.g., enriched in a first tissue and not enriched in one or more other tissues). Machine learning algorithms were applied to discover determinants of tissue tropism.

FIG. 16 shows that rAAV5 variants include variants present in the CNS at large, as well as found in the substantia nigra, a subregion of the brain particularly affected in Parkinson's disease. Additionally, FIG. 16 shows that machine learning models exhibited good performance at predicting determinants of high CNS specificity (at right).

FIG. 17 shows that rAAV5 variants include variants present in muscle tissue, including variants that target heart (cardiac tissue) and skeletal tissue while de-targeted to liver tissue (at left). Additionally, FIG. 17 shows that machine learning models exhibited good performance at predicting determinants of liver detargeting (at right).

FIG. 18A shows a heatmap of amino acid positional frequencies in a CNS tissue compared to all other analyzed tissues. FIG. 18B shows a heatmap of amino acid positional frequencies in a liver tissue compared to all other analyzed tissues. FIG. 18C shows a heatmap of amino acid positional frequencies in a skeletal muscle tissue compared to all other analyzed tissues. FIG. 18D shows a heatmap of amino acid positional frequencies in a heart tissue compared to all other analyzed tissues. FIG. 18E shows a heatmap of amino acid positional frequencies in a lung tissue compared to all other analyzed tissues. FIG. 18F shows a heatmap of amino acid positional frequencies in a spleen tissue compared to all other analyzed tissues. FIG. 18G shows a heatmap of amino acid positional frequencies in a lymph node tissue compared to all other analyzed tissues. FIG. 18H shows a heatmap of amino acid positional frequencies in a bone marrow tissue compared to all other analyzed tissues. FIG. 18I shows a heatmap of amino acid positional frequencies in a mammary gland tissue compared to all other analyzed tissues. FIG. 18J shows a heatmap of amino acid positional frequencies in a skin tissue compared to all other analyzed tissues. FIG. 18K shows a heatmap of amino acid positional frequencies in an adrenal gland tissue compared to all other analyzed tissues. FIG. 18L shows a heatmap of amino acid positional frequencies in a thyroid tissue compared to all other analyzed tissues. FIG. 18M shows a heatmap of amino acid positional frequencies in a colon tissue compared to all other analyzed tissues. FIG. 18N shows a heatmap of amino acid positional frequencies in a sciatic nerve tissue compared to all other analyzed tissues. FIG. 18O shows a heatmap of amino acid positional frequencies in a spinal cord tissue compared to all other analyzed tissues.

FIG. 19A shows the results at various steps of bioinformatics pre-processing of the NGS data. Stepwise read count for 48 individual samples through filters is shown. FIG. 19B shows a positional comparison of CNS/non-CNS variant sequences.

FIG. 20A shows an example of peripheral and CNS tissue samples analyzed by the methods described herein. FIG. 20B shows amino acid positional abundance in the top 1000 machine learning predicted/filtered variants recovered from the CNS compared to non-CNS variants.

FIG. 21A shows an example of an ensemble of both ML models: Random Forest (RF) and Histogram-based Gradient Boosting Tree (HGB) for averaged predicted CNS probability. Outputs of CNS-targeting probability from both ML models showed good concordance. FIG. 21B shows the distribution of average CNS-targeting probability for all CNS variants.

FIG. 22 shows an example of machine learning model performance validation. Both Histogram-based Gradient Boosting Tree (HGB) (top) and Random Forest (RF) (bottom) machine learning models showed good performance at predicting CNS targeting. At high predicted class probability values, both machine learning models resolved CNS-targeting from non-CNS targeting.

FIG. 23 shows how a set of top 20 positional features contributed to model output probability. Shapley Additive Explanations (SHAP) values were used to interrogate the relative contribution of features to model predictions. These features can be further compared between tissue targeting and non-targeting variants (as in FIGS. 24A-C).

FIGS. 24A, 24B, and 24C shows a comparison of a set of top predictive features positionally. Features were selected if they were found to be important to both the HGB & RF models. Summaries are shown of the features in the top 1000 machine learning-predicted CNS variants compared to a random 2% of CNS variants.

FIG. 25 shows a set of top 20 positional features contributing to model output probability in a machine learning analysis of sequences that target liver tissue.

FIGS. 26A, 26B, and 26C show a comparison of top predictive features positionally in a machine learning analysis of sequences that target liver tissue. Features were compared between tissue targeting and non-targeting variants.

FIG. 27 shows a set of top 20 positional features contributing to model output probability in a machine learning analysis of sequences that are liver-detargeted.

FIGS. 28A and 28B show a comparison of top predictive features positionally for liver-detargeted variants.

FIG. 29 shows a set of top 20 positional features contributing to model output probability in a machine learning analysis of sequences that target muscle tissue.

FIGS. 30A-30B show a comparison of top predictive features positionally in a machine learning analysis of sequences that target muscle tissue. Features were compared between tissue targeting and non-targeting variants.

6. DETAILED DESCRIPTION OF THE INVENTION

Unless described otherwise, all technical and scientific terms used herein have the meaning commonly understood by one of ordinary skill in the art to which the invention pertains.

Unless otherwise stated, whenever a range is recited, the range is inclusive of the recited endpoints. For example, the region from amino acid residue 581 to amino acid residue 589 of SEQ ID NO: 1 includes amino acid residues 581 and 589.

“Homology” or “identity” or “similarity” can refer to sequence similarity between two peptides or between two nucleic acid molecules. Homology can be determined by comparing a position in each sequence which can be aligned for purposes of comparison. When a position in the compared sequence can be occupied by the same base or amino acid, then the molecules can be homologous at that position. A degree of homology between sequences can be a function of the number of matching or homologous positions shared by the sequences. An “unrelated” or “non-homologous” sequence shares less than 40% identity, or alternatively less than 25% identity, with one of the sequences of the disclosure. Sequence homology can refer to a % identity of a sequence to a reference sequence. As a practical matter, whether any particular sequence can be at least 50%, 60%, 70%, 80%, 85%, 90%, 92%, 95%, 96%, 97%, 98% or 99% identical to any sequence described herein (which can correspond with a particular nucleic acid sequence described herein), such particular polypeptide sequence can be determined conventionally using known computer programs such the Bestfit program (Wisconsin Sequence Analysis Package, Version 8 for Unix, Genetics Computer Group, University Research Park, 575 Science Drive, Madison, Wis. 53711). When using Bestfit or any other sequence alignment program to determine whether a particular sequence is, for instance, 95% identical to a reference sequence, the parameters can be set such that the percentage of identity can be calculated over the full length of the reference sequence and that gaps in sequence homology of up to 5% of the total reference sequence can be allowed. The term percent “identity” or percent “homology,” in the context of two or more nucleic acid or polypeptide sequences, refer to two or more sequences or subsequences that have a specified percentage of nucleotides or amino acid residues that are the same, when compared and aligned for maximum correspondence, as measured using one of the sequence comparison algorithms described below (e.g., BLASTP and BLASTN or other algorithms available to persons of skill) or by visual inspection. Depending on the application, the percent “identity” can exist over a region of the sequence being compared, e.g., over a functional domain, or, alternatively, exist over the full length of the two sequences to be compared. For sequence comparison, typically one sequence acts as a reference sequence to which test sequences are compared. When using a sequence comparison algorithm, test and reference sequences are input into a computer, subsequence coordinates are designated, if necessary, and sequence algorithm program parameters are designated. The sequence comparison algorithm then calculates the percent sequence identity for the test sequence(s) relative to the reference sequence, based on the designated program parameters. For purposes herein, percent identity and sequence similarity is performed using the BLAST algorithm, which is described in Altschul et al., J. Mol. Biol. 215:403-410 (1990). Software for performing BLAST analyses is publicly available through the National Center for Biotechnology Information (www.ncbi.nlm.nih.gov/).

In some cases, the identity between a reference sequence (query sequence, e.g., a sequence of the disclosure) and a subject sequence, also referred to as a global sequence alignment, can be determined using the FASTDB computer program. In some embodiments, parameters for a particular embodiment in which identity can be narrowly construed, used in a FASTDB amino acid alignment, can include: Scoring Scheme=PAM (Percent Accepted Mutations) 0, k-tuple=2, Mismatch Penalty=1, Joining Penalty=20, Randomization Group Length=0, Cutoff Score=1, Window Size=sequence length, Gap Penalty=5, Gap Size Penalty=0.05, Window Size=500 or the length of the subject sequence, whichever can be shorter. According to this embodiment, if the subject sequence can be shorter than the query sequence due to N- or C-terminal deletions, not because of internal deletions, a manual correction can be made to the results to take into consideration the fact that the FASTDB program does not account for N- and C-terminal truncations of the subject sequence when calculating global percent identity. For subject sequences truncated at the N- and C-termini, relative to the query sequence, the percent identity can be corrected by calculating the number of residues of the query sequence that can be lateral to the N- and C-terminal of the subject sequence, which can be not matched/aligned with a corresponding subject residue, as a percent of the total bases of the query sequence. A determination of whether a residue can be matched/aligned can be determined by results of the FASTDB sequence alignment. This percentage can be then subtracted from the percent identity, calculated by the FASTDB program using the specified parameters, to arrive at a final percent identity score. This final percent identity score can be used for the purposes of this embodiment. In some cases, only residues to the N- and C-termini of the subject sequence, which can be not matched/aligned with the query sequence, can be considered for the purposes of manually adjusting the percent identity score. That is, only query residue positions outside the farthest N- and C-terminal residues of the subject sequence can be considered for this manual correction. For example, a 90-residue subject sequence can be aligned with a 100-residue query sequence to determine percent identity. The deletion occurs at the N-terminus of the subject sequence, and therefore, the FASTDB alignment does not show a matching/alignment of the first 10 residues at the N-terminus. The 10 unpaired residues represent 10% of the sequence (number of residues at the N- and C-termini not matched/total number of residues in the query sequence) so 10% can be subtracted from the percent identity score calculated by the FASTDB program. If the remaining 90 residues were perfectly matched, the final percent identity can be 90%. In another example, a 90-residue subject sequence can be compared with a 100-residue query sequence. This time the deletions can be internal deletions, so there can be no residues at the N- or C-termini of the subject sequence which can be not matched/aligned with the query. In this case, the percent identity calculated by FASTDB can be not manually corrected. Once again, only residue positions outside the N- and C-terminal ends of the subject sequence, as displayed in the FASTDB alignment, which can be not matched/aligned with the query sequence can be manually corrected for.

As used herein, “tropism” of a rAAV for a tissue is defined as the ability of a given rAAV to preferentially infect a given cell or tissue. Altered or engineered tropism includes increased or decreased targeting ability for desired tissues, with a corresponding increased or decreased infection of the target tissue.

For simplicity throughout this disclosure, viral capsid protein is generally referred to as “VP.” Viral capsid protein is referred to as VP1 when referencing AAV5 VP1 positional notation. In all cases, viral capsid sequences and mutations disclosed herein should be understood as pertaining to all isoforms of the capsid protein (VP1, VP2, and VP3), as a mixture of these isoforms assemble to form virions. The positional amino acid residue designations “581-589” are relative to the translational start of the VP1 polypeptide and should be adjusted accordingly to the relative start sites of VP2 and VP3. It should be understood that the present disclosure, when describing any particular VP1 sequence with mutations at particular amino acid residue positions, necessarily also encompasses corresponding mutations in VP2 and VP3. For example, any consensus sequence or specific sequence of a VP1 capsid protein having one or more mutations in the amino acid residue s of the 581-589 region also encompasses VP2 and VP3 capsid proteins having said one or more mutations in an amino acid residue region in VP2 and VP3 corresponding to the amino acid residues of the VP1 581-589 region. For example, the amino acid residues of the 581 to 589 region of VP1 (SEQ ID NO: 1) correspond to the amino acid residues of the 445 to 453 region of VP2 (SEQ ID NO: 1115) and to the amino acid residues of 389 to 397 region of VP3 (SEQ ID NO: 1116).

It should be understood that the present disclosure includes polynucleotide sequences encoding for any sequence disclosed herein. For example, if an amino acid sequence is provided, the present disclosure also encompasses a polynucleotide sequence encoding for said amino acid sequence.

It should be understood that further embodiments include mutations in VP1. VP2, VP3, or any combination thereof that do not alter the desired properties (e.g., a particular tissue tropism) or affect viral assembly, as described herein.

As used herein, “tissue tropism” refers to a preference of a virus having an engineered VP capsid polypeptide of the present disclosure to infect a given tissue or be enriched in or accumulate in a given tissue. Tissue tropism, when used as a relative term and depending on the context in which it is described herein, refers to an increase or decrease in tissue tropism of a given rAAV virion having a first capsid polypeptide in a first tissue as compared to a second tissue and/or refers to an increase or decrease in tissue tropism of a given rAAV virion having a first capsid polypeptide to an rAAV virion having a second capsid polypeptide. In some embodiments, the first tissue can be a group of tissues. In some embodiments, the second tissue can be a group of tissues. For example, the first tissue may be CNS tissues, which comprise cortex forebrain, cortex occipital, cortex temporal, thalamus, hypothalamus, substantia nigra, hippocampus DG, hippocampus CA1, hippocampus CA3, and cerebellum and the second tissue may be a non-CNS tissue consisting collectively of liver, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord tissues. As another example, the first tissue may be liver tissue and the second tissue may be non-liver tissue consisting collectively of CNS tissues, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord tissues.

As used herein, the word “recombinant” in the context of an AAV capsid polypeptide, interchangeably refers to an “engineered” or “variant” AAV capsid polypeptide. As used herein, the word “recombinant” in the context of an AAV virion, abbreviated to rAAV, refers to a recombinant virus particle. Said rAAV virion is made of a capsid that may include the engineered AAV5 VP capsid polypeptides disclosed herein.

6.1. Capsid Engineering Methods

Disclosed herein is a system for high throughput engineering of engineered AAV capsids with modified function, including increased or decreased infectivity of desired tissues, such as increased or decreased liver tropism, or increased targeting of the central nervous system (CNS). A general schematic of the process is shown in FIG. 1, however, it should be understood that the present disclosure also encompasses reasonable variations or extensions to the method that are understood to those of ordinary skill in the art. As shown in FIG. 1, the method may begin with production of a capsid library with theoretical diversity of 5×10¹¹unique sequence variants. Higher or lower theoretical diversities are also encompassed herein. For example, a capsid library may have a theoretical diversity of from about 1×10{circumflex over ( )}3 to about 1×10{circumflex over ( )}20. The library may then be cloned into plasmids, transformed into bacteria, and subsequently library plasmids are screened for productive virion assembly in a production cell line. The assembled virions may then be administered intravenously into non-human primates (NHP). After a period sufficient for distribution, infection, and stable transduction, the NHP may be sacrificed, organs harvested, and sequences of AAV capsids in each tissue may be determined by deep sequencing.

FIG. 2A provides a side view (top panel) and top view (bottom panel) of the surface of a prototype AAV virion, identifying residues of known AAV capsids—including AAV2, AAV5, AAV6, and AAV9—that have been shown in the research literature to interact with target cells. These target-interacting residues correspond to amino acids 581-589 in the AAV5 VP1 capsid protein.

FIG. 2B shows the salient elements of the library plasmid, illustrating rep and cap coding sequences positioned between AAV ITRs. In the illustrated embodiment, further described in Example 1, variation is introduced into each of residues 581-589 of the AAV5 cap protein (“Library variant region”). Each of the 20 natural amino acids is introduced at each of the 9 positions, providing a theoretical library diversity of 20⁹(20{circumflex over ( )}9; approximately 5×10¹¹) unique sequence variants.

The area targeted for engineering is the most likely to interact with target cell receptors, and relatively tolerant to changes without disrupting virion assembly. Unlike earlier approaches that add unstructured peptides that protrude above the virion 3-fold axis of symmetry, the current approach introduces sequence diversity that alters the characteristics of the binding pocket. In addition, this approach may change the overall structure of the receptor-binding trimer, allowing for altered allosteric interactions outside the binding pocket (e.g., AAVR PKD1). Introduced diversity is non-random, thereby reducing missense and frameshifts of randomized libraries.

By cloning the polynucleotide encoding the capsid variants into the packaged viral genome (between the ITRs), the recombinant virions with variant capsids carry polynucleotides having their cognate mutation, so the unique variant providing the desired function can be identified by sequencing packaged virus or infected cells.

In some embodiments, the capsid is a capsid selected from AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV 10, AAV11, AAV 12, AAV13, AAV 14, AAV 15 and AAV 16, AAV.rh8, AAV.rh10, AAV.rh20, AAV.rh39, AAV.Rh74, AAV.RHM4-1, AAV.hu37, AAV.Anc80, AAV.Anc80L65, AAV.7m8, AAV.PHP.B, AAV2.5, AAV2tYF, AAV3B, AAV.LK03, AAV.HSC1, AAV.HSC2, AAV.HSC3, AAV.HSC4, AAV.HSC5, AAV.HSC6, AAV.HSC7, AAV.HSC8, AAV.HSC9, AAV.HSC10, AAV.HSC11, AAV.HSC12, AAV.HSC13, AAV.HSC14, AAV.HSC15, AAV.HSC16 or AAVhu68 (described in WO2020/033842, incorporated herein by reference in its entirety). The hu68 capsid is described in WO 2018/160582, incorporated herein by reference in its entirety.

Such capsids may comprise a region corresponding to the 581-589 region of the AAV5 VP1, and as such analogous engineered VP capsids with desired tissue tropism, ability to assemble, and exhibit various other desired traits are encompassed herein. Thus, any one of the engineered AAV5 VP capsid polypeptides disclosed herein having a mutation in a region corresponding to the 581 to 589 region of AAV5 VP1 may be inserted into the corresponding region in any one of the other AAV capsids described herein and the present disclosure encompasses such variants.

In some embodiments, the capsid is a derivative, modification, or pseudotype of AAV1, AAV2, AAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV 10, AAV11, AAV 12, AAV 13, AAV 14, AAV 15 and AAV 16, AAV.rh8, AAV.rh0. AAV.rh20, AAV.rh39, AAV.Rh74, AAV.RHM4-1, AAV.hu37, AAV.Anc80, AAV.Anc80L65, AAV.7m8, AAV.PHP.B, AAV2.5, AAV2tYF, AAV3B, AAV.LK03, AAV.HSC1, AAV.HSC2, AAV.HSC3, AAV.HSC4, AAV.HSC5, AAV.HSC6, AAV.HSC7, AAV.HSC8, AAV.HSC9, AAV.HSC10, AAV.HSC11, AAV.HSC12, AAV.HSC13, AAV.HSC14, AAV.HSC15, AAV.HSC16 or AAVhu68.

In some embodiments, capsid protein is a chimera of capsid proteins from two or more serotype selected from AAV1, AAV2, rAAV3, AAV4, AAV5, AAV6, AAV7, AAV8, AAV9, AAV 10, AAV 11, AAV 12, AAV13, AAV 14, AAV 15 and AAV 16, AAV.rh8, AAV.rh10, AAV.rh20, AAV.rh39, AAV.Rh74, AAV.RHM4-1, AAV.hu37, AAV.Anc80, AAV.Anc80L65, AAV.7m8, AAV.PHP.B, AAV2.5, AAV2tYF, AAV3B, AAV.LK03, AAV.HSC1, AAV.HSC2, AAV.HSC3, AAV.HSC4, AAV.HSC5, AAV.HSC6, AAV.HSC7, AAV.HSC8, AAV.HSC9, AAV.HSC10, AAV.HSC11, AAV.HSC12, AAV.HSC13, AAV.HSC14, AAV.HSC15, and AAV.HSC16 (described in WO2020/033842, incorporated herein by reference in its entirety). In certain embodiments, the capsid is an rh32.33 capsid, described in U.S. Pat. No. 8,999,678, incorporated herein by reference in its entirety.

Such capsids may comprise a region corresponding to 581-589 of the AAV5 VP1, and as such analogous engineered VP capsids with desired tissue tropism, ability to assemble, and exhibit various other desired traits are encompassed herein.

6.2. VP-Encoding Polynucleotides, Vectors, and Vector Libraries

Accordingly, in a first aspect, polynucleotides are provided. The polynucleotides encode an adeno-associated virus (AAV) VP1 capsid polypeptide having the amino acid sequence of SEQ ID NO:2, wherein Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from the 20 naturally occurring amino acids—using standard one letter codes, from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V. The polypeptide includes at least one mutation of the native AAV5 capsid and thus does not have the sequence of SEQ ID NO: 1. In addition, the polypeptide does not have the sequence of SEQ ID NO: 3, SEQ ID NO:4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, or SEQ ID NO: 8.

In some embodiments, the polynucleotide encodes an AAV VP1 capsid polypeptide that further comprises one or more mutations at an amino acid residue outside of the 581-589 region, with reference to SEQ ID NO: 1, wherein the resulting recombinant capsid is capable of forming an assembled virion that exhibits desired tissue targeting.

In another aspect, a vector capable of replication in prokaryotic cells is provided, wherein the vector comprises the polynucleotide described immediately above. In typical embodiments, the vector is a plasmid encoding a replication-competent AAV genome.

In a further aspect, a library is provided. The library comprises a plurality of vectors comprising the AAV capsid-encoding polynucleotides. In some embodiments, the vectors are plasmids, and the plurality of plasmids comprise a plurality of different AAV VP-encoding polynucleotides.

In various library embodiments, at least one of Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 of SEQ ID NO: 2 is invariant. Such invariant residues are also referred to herein as “framework” residues. Framework residues may contribute to competence of the capsid to assemble into functional virions or infect a particular target cell or tissue

In some library embodiments, one residue of Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 of SEQ ID NO: 2 is invariant. In particular embodiments, the invariant residue is the native amino acid of AAV5 VP1 at that position within the VP1 primary amino acid sequence. In particular embodiments, the invariant residue is an amino acid other than the native amino acid of AAV5 VP1 at that position. In some embodiments, two of Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 of SEQ ID NO: 2 are invariant. In particular embodiments, each invariant residue is the native amino acid of AAV5 VP1 at the respective positions. In particular embodiments, each invariant residue is an amino acid other than the native amino acid of AAV5 VP1 at the respective positions. In some embodiments, three of Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 of SEQ ID NO: 2 are invariant. In particular embodiments, each invariant residue is the native amino acid of AAV5 VP1 at the respective positions. In particular embodiments, each invariant residue is an amino acid other than the native amino acid of AAV5 VP1 at the respective positions. In some embodiments, four of Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 or SEQ ID NO: 2 are invariant. In particular embodiments, each invariant residue is the native amino acid of AAV5 VP1 at the respective positions. In particular embodiments, each invariant residue is an amino acid other than the native amino acid of AAV5 VP1 at the respective positions. In some embodiments, five of Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 of SEQ ID NO: 2 are invariant. In particular embodiments, each invariant residue is the native amino acid of AAV5 VP1 at the respective positions. In particular embodiments, each invariant residue is an amino acid other than the native amino acid of AAV5 VP1 at the respective positions. In some embodiments, six of Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 of SEQ ID NO: 2 are invariant. In particular embodiments, each invariant residue is the native amino acid of AAV5 VP1 at the respective positions. In particular embodiments, each invariant residue is an amino acid other than the native amino acid of AAV5 VP1 at the respective positions.

In particular embodiments the rAAV VP1 capsid at position 587 (Xaa7) is not A, C, D, E, F, G, H, I, K, M, P, Q, R, V, W, or Y. In some embodiments, position 587 can be N, S, or T. In particular embodiments, the rAAV VP1 capsid at position 582 (Xaa2) is not G, V, L. or I.

In various embodiments, the library encodes at least 1× 10⁹different AAV VP capsid polypeptides, at least 2.5×10⁹different AAV VP capsid polypeptides, at least 5×10⁹different AAV VP capsid polypeptides, at least 7.5×10⁹different AAV VP capsid polypeptides, at least 1×10¹⁰different AAV VP capsid polypeptides, at least 2.5×10¹⁰different AAV VP capsid polypeptides, at least 5×10¹⁰different AAV VP capsid polypeptides, at least 7.5×10¹⁰different AAV VP capsid polypeptides, at least 1×10¹¹different AAV VP capsid polypeptides, at least 2.5×10¹¹different AAV VP capsid polypeptides, or at least 5×10¹¹different AAV VP capsid polypeptides.

In another aspect, prokaryotic cells comprising the vectors are provided. In some embodiments, the prokaryotic cell is an E. coli cell and the vector is a plasmid.

In a related aspect, libraries are provided, the library comprising a plurality of E. coli cells, wherein the plurality of cells comprise a plurality of plasmids, wherein the plurality of plasmids comprise a plurality of different AAV VP-encoding polynucleotides.

In various library embodiments, at least one of Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 of SEQ ID NO: 2 is invariant.

In some library embodiments, one of Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 is invariant of SEQ ID NO: 2. In particular embodiments, the invariant residue is the native amino acid of AAV5 VP1 at that position within the VP1 primary amino acid sequence. In particular embodiments, the invariant residue is an amino acid other than the native amino acid of AAV5 VP1 at that position. In some embodiments, two of Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 of SEQ ID NO: 2 are invariant. In particular embodiments, each invariant residue is the native amino acid of AAV5 VP1 at the respective positions. In particular embodiments, each invariant residue is an amino acid other than the native amino acid of AAV5 VP1 at the respective positions. In some embodiments, three of Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 of SEQ ID NO: 2 are invariant. In particular embodiments, each invariant residue is the native amino acid of AAV5 VP1 at the respective positions. In particular embodiments, each invariant residue is an amino acid other than the native amino acid of AAV5 VP1 at the respective positions. In some embodiments, four of Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 of SEQ ID NO: 2 are invariant. In particular embodiments, each invariant residue is the native amino acid of AAV5 VP1 at the respective positions. In particular embodiments, each invariant residue is an amino acid other than the native amino acid of AAV5 VP1 at the respective positions. In some embodiments, five of Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 of SEQ ID NO: 2 are invariant. In particular embodiments, each invariant residue is the native amino acid of AAV5 VP1 at the respective positions. In particular embodiments, each invariant residue is an amino acid other than the native amino acid of AAV5 VP1 at the respective positions. In some embodiments, six of Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 of SEQ ID NO: 2 are invariant. In particular embodiments, each invariant residue is the native amino acid of AAV5 VP1 at the respective positions. In particular embodiments, each invariant residue is an amino acid other than the native amino acid of AAV5 VP1 at the respective positions.

In some embodiments, the library encodes at least 1×10⁹different AAV VP capsid polypeptides, at least 2.5×10⁹different AAV VP capsid polypeptides, at least 5×10⁹different AAV VP capsid polypeptides, at least 7.5×10⁹different AAV VP capsid polypeptides, at least 1×10¹⁰different AAV VP capsid polypeptides, at least 5×10¹⁰different AAV VP capsid polypeptides, at least 7.5×10¹⁰different AAV VP capsid polypeptides, at least 1×10¹¹different AAV VP capsid polypeptides, at least 2.5×10¹¹different AAV VP capsid polypeptides, or at least 5×10¹¹different AAV VP capsid polypeptides.

6.3. VP Polypeptides, Peptide Libraries

In another aspect, AAV VP1 capsid polypeptides are provided. The polypeptide has the amino acid sequence of SEQ ID NO: 2, wherein Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V. The polypeptide includes at least one mutation as compared to native AAV VP1, and thus does not have the sequence of SEQ ID NO: 1. In addition, the polypeptide does not have the sequence of SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, or SEQ ID NO: 8.

In some embodiments, the polypeptide further comprises one or more mutations at an amino acid residue outside of the 581-589 region, with reference to SEQ ID NO: 1, wherein the resulting recombinant capsid is capable of forming an assembled virion that exhibits desired tissue targeting.

In a further aspect, libraries are provided, the libraries comprising a plurality of polypeptides as described immediately above, the plurality having different primary amino acid sequences.

In various library embodiments, at least one of Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 of SEQ ID NO: 2 is invariant.

In some library embodiments, one of Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 of SEQ ID NO: 2 is invariant. Such invariant residues are also referred to herein as “framework” residues. In particular embodiments, the invariant residue is the native amino acid of AAV5 VP1 at that position within the VP1 primary amino acid sequence. In particular embodiments, the invariant residue is an amino acid other than the native amino acid of AAV5 VP1 at that position. In some embodiments, two of Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 of SEQ ID NO: 2 are invariant. In particular embodiments, each invariant residue is the native amino acid of AAV5 VP1 at the respective positions. In particular embodiments, each invariant residue is an amino acid other than the native amino acid of AAV5 VP1 at the respective positions. In some embodiments, three of Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 of SEQ ID NO: 2 are invariant. In particular embodiments, each invariant residue is the native amino acid of AAV5 VP1 at the respective positions. In particular embodiments, each invariant residue is an amino acid other than the native amino acid of AAV5 VP1 at the respective positions. In some embodiments, four of Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 of SEQ ID NO: 2 are invariant. In particular embodiments, each invariant residue is the native amino acid of AAV5 VP1 at the respective positions. In particular embodiments, each invariant residue is an amino acid other than the native amino acid of AAV5 VP1 at the respective positions. In some embodiments, five of Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 of SEQ ID NO: 2 are invariant. In particular embodiments, each invariant residue is the native amino acid of AAV5 VP1 at the respective positions. In particular embodiments, each invariant residue is an amino acid other than the native amino acid of AAV5 VP1 at the respective positions. In some embodiments, six of Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 of SEQ ID NO: 2 are invariant. In particular embodiments, each invariant residue is the native amino acid of AAV5 VP1 at the respective positions. In particular embodiments, each invariant residue is an amino acid other than the native amino acid of AAV5 VP1 at the respective positions.

In some embodiments, library comprises at least 1×10⁹different AAV VP capsid polypeptides, at least 2.5×10⁹different AAV VP capsid polypeptides, at least 5×10⁹different AAV VP capsid polypeptides, at least 7.5×10⁹different AAV VP capsid polypeptides, at least 1×10¹⁰different AAV VP capsid polypeptides, at least 2.5×10¹⁰different AAV VP capsid polypeptides, at least 5×10¹⁰different AAV VP capsid polypeptides, at least 7.5×10¹⁰different AAV VP capsid polypeptides, at least 1×10¹¹different AAV VP capsid polypeptides, at least 2.5×10¹¹different AAV VP capsid polypeptides, or at least 5×10¹¹different AAV VP capsid polypeptides.

In certain embodiments, the library comprises at least from about 1×10⁵to at least about 5×10¹¹different AAV VP capsid polypeptides. In certain embodiments, the library comprises at least about 1×10⁵, at least about 2×10⁵, at least about 3×10⁵, at least about 4×10⁵, at least about 5×10⁵, at least about 6×10⁵, at least about 7×10⁵, at least about 8×10⁵, at least about 9×10⁵, at least about 1× 10⁶, at least about 2×10 at least about 3×10⁶, at least about 4×10⁶, at least about 5×10⁶, at least about 6×10⁶, at least about 7×10⁶, at least about 8×10⁶, at least about 9×10⁶, at least about 1× 10⁷, at least about 2×10⁷, at least about 3×10⁷, at least about 4×10⁷, at least about 5×10⁷, at least about 6×10⁷, at least about 7×10⁷, at least about 8×10⁷, at least about 9×10⁷, at least about 1×10⁸, at least about 2×10⁸, at least about 3×10⁸, at least about 4×10⁸, at least about 5×10⁸, at least about 6×10, at least about 7×10⁸, at least about 8×10⁸, at least about 9×10′, at least about 1×10⁹, at least about 2×10⁹, at least about 3×10⁹, at least about 4×10⁹, at least about 5×10⁹, at least about 6×10⁹, at least about 7×10⁹, at least about 8×10⁹, at least about 9×10⁹, at least about 1×10¹⁰, at least about 2×10¹⁰, at least about 3×10¹⁰, at least about 4×10¹⁰, at least about 5×10¹⁰, at least about 6×10¹⁰, at least about 7×10¹⁰, at least about 8×10¹⁰, at least about 9×10¹⁰, at least about 1×10¹¹, at least about 2×10¹¹, at least about 3×10¹¹, at least about 4×10¹¹, or at least about 5×10¹¹AAV VP capsid polypeptides.

In certain embodiments, provided herein is a recombinant adeno-associated virus AAV VP1 capsid polypeptide having at least one mutation in a residue of region 581 to residue 589 in SEQ ID NO: 1, inclusive, wherein the mutation confers at least about a two-fold increased accumulation of an AAV virion having said AAV VP1 capsid polypeptide in a non-liver tissue as compared to a liver tissue, as compared to wildtype AAV virion having a wildtype AAV5 VP1 capsid polypeptide, and wherein the AAVVP1 capsid polypeptide does not have the sequence of any of SEQ ID NO: 1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, and SEQ ID NO:8.

6.4. rAAV Virions, Virion Libraries

In another aspect, recombinant AAV virions (rAAV) are provided. The virion comprises an AAV VP capsid polypeptide as described above.

In some embodiments, the rAAV has increased tropism for primate and human liver as compared to a rAAV having the native AAV5 VP1 capsid polypeptide (SEQ ID NO:1). In some embodiments, the rAAV has increased ability to assemble, or exhibits greater virion stability, as compared to a rAAV having the native AAV5 VP1 capsid polypeptide (SEQ ID NO:1).

In some embodiments, the rAAV has reduced tropism for human liver as compared to a rAAV having the native AAV5 VP1 capsid polypeptide (SEQ ID NO:1).

In some embodiments, the rAAV has increased ability to cross the blood-brain barrier following intravenous administration as compared to a rAAV having the native AAV5 VP1 capsid polypeptide (SEQ ID NO:1).

In certain of these embodiments, the rAAV has increased ability to infect one or more brain regions selected from hippocampus, dentate gyrus, cerebral cortex, temporal cortex, occipital cortex, thalamus, forebrain, substantia nigra, hypothalamus, and cerebellum following intravenous, intrathecal, intracerebral ventricular, or intracisternal magna administration, as compared to a rAAV having the native AAV5 VP1 capsid polypeptide (SEQ ID NO 1).

In some embodiments, the rAAV has increased ability to infect one or more brain regions selected from hippocampus, dentate gyrus, cerebral cortex, temporal cortex, occipital cortex, thalamus, forebrain, substantia nigra, hypothalamus, and cerebellum following intravenous, intrathecal, intracerebral ventricular, or intracisternal magna administration and also has reduced tropism for all non CNS tissues, including being detargeted for cardiac tissue, as compared to a rAAV having the native AAV5 VP1 capsid polypeptide (SEQ ID NO:1).

In some embodiments, the rAAV has increased ability to infect human retinal cells following intravitreal injection as compared to a rAAV having the native AAV5 VP1 capsid polypeptide (SEQ ID NO:1).

In some embodiments, the rAAV has increased ability to infect human skeletal muscle following intravenous administration as compared to a rAAV having a VP1 capsid polypeptide having the native AAV5 VP1 capsid polypeptide (SEQ ID NO:1).

In some embodiments, the rAAV has increased ability to infect a tissue selected from adipose, adrenal gland, aorta, brain (including hippocampus: dentate gyrus, CA1 and CA3; cerebellum, caudate, putamen, midbrain, pons, hypothalamus, cortex-including occipital, temporal and forebrain; substantia nigra, and thalamus), bone marrow, cecum, colon, dorsal root ganglion, duodenum, epididymis, esophagus, eye, gallbladder, heart, ileum, jejunum, kidney, lung, lymph nodes, mammary gland, ovary, pancreas, parathyroid gland, peripheral nerve, pituitary, prostate, salivary gland, seminal vesicle, skeletal muscle, skin, spinal cord, spleen, stomach, testis, thymus, thyroid, trachea, urinary bladder, uterus, and vagina following intravenous administration, as compared to a rAAV having the native AAV5 capsid polypeptide (SEQ ID NO:1).

Additionally, provided are polynucleotide sequences encoding the rAAV capsid VP proteins described herein.

In a further aspect, libraries are provided that comprise a plurality of rAAV as described above. The plurality of rAAV comprise a plurality of VP capsid polypeptides having different primary amino acid sequences.

In various library embodiments, at least one of Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 of SEQ ID NO: 2 is invariant.

In some embodiments, the library comprises at least 1×10⁹different AAV VP capsid polypeptides, at least 2.5×10⁹different AAV VP capsid polypeptides, at least 5×10⁹different AAV VP capsid polypeptides, at least 7.5×10 different AAV VP capsid polypeptides, at least 1×10¹⁰different AAV VP capsid polypeptides, at least 2.5×10¹⁰different AAV VP capsid polypeptides, at least 5×10¹⁰different AAV VP capsid polypeptides, at least 7.5×10¹⁰different AAV VP capsid polypeptides, at least 1×10¹¹different AAV VP capsid polypeptides, at least 2.5×10¹¹different AAV VP capsid polypeptides, or at least 5×10¹¹different AAV VP capsid polypeptides.

6.5. Pharmaceutical Compositions

In another aspect, pharmaceutical compositions are provided. The pharmaceutical composition comprises a rAAV as described above and a pharmaceutically acceptable carrier.

A pharmaceutical composition can comprise a first active ingredient. The first active ingredient can comprise a viral vector as described herein and/or any payload as described herein. The pharmaceutical composition can be formulated in unit dose form. The pharmaceutical composition can comprise a pharmaceutically acceptable excipient, diluent, or carrier. The pharmaceutical composition can comprise a second, third, or fourth active ingredient—such as to facilitate enhanced gene replacement, RNA editing. DNA editing, or imaging.

A pharmaceutical composition described herein can compromise an excipient. An excipient can comprise a cryo-preservative, such as DMSO, glycerol, polyvinylpyrrolidone (PVP), or any combination thereof. An excipient can comprise a cryo-preservative, such as a sucrose, a trehalose, a starch, a salt of any of these, a derivative of any of these, or any combination thereof. An excipient can comprise a pH agent (to minimize oxidation or degradation of a component of the composition), a stabilizing agent (to prevent modification or degradation of a component of the composition), a buffering agent (to enhance temperature stability), a solubilizing agent (to increase protein solubility), or any combination thereof. An excipient can comprise a surfactant, a sugar, an amino acid, an antioxidant, a salt, a non-ionic surfactant, a solubilizer, a triglyceride, an alcohol, or any combination thereof. An excipient can comprise sodium carbonate, acetate, citrate, phosphate, poly-ethylene glycol (PEG), human serum albumin (HSA), sorbitol, sucrose, trehalose, polysorbate 80, sodium phosphate, sucrose, disodium phosphate, mannitol, polysorbate 20, histidine, citrate, albumin, sodium hydroxide, glycine, sodium citrate, trehalose, arginine, sodium acetate, acetate, HCl, disodium edetate, lecithin, glycerol, xanthan rubber, soy isoflavones, polysorbate 80, ethyl alcohol, water, teprenone, or any combination thereof.

Compositions and methods provided herein can utilize pharmaceutical compositions. The compositions described throughout can be formulated into a pharmaceutical and be used to treat a human or mammal, in need thereof, diagnosed with a disease. In some cases, pharmaceutical compositions can be used prophylactically.

The compositions provided herein can be utilized in methods provided herein. Any of the provided compositions provided herein can be utilized in methods provided herein. In some cases, a method comprises at least partially preventing, reducing, ameliorating, and/or treating a disease or condition, or a symptom of a disease or condition. A subject can be a human or non-human. A subject can be a mammal (e.g., rat, mouse, cow, dog, pig, sheep, horse). A subject can be a vertebrate or an invertebrate. A subject can be a laboratory animal A subject can be a patient. A subject can be suffering from a disease. A subject can display symptoms of a disease. A subject may not display symptoms of a disease, but still have a disease. A subject can be under medical care of a caregiver (e.g., the subject is hospitalized and is treated by a physician).

6.6. Methods of Treatment or Detection

In some aspects, the present disclosure provides for methods of treatment using an rAAV virion having any one of the engineered AAV VP capsid polypeptide sequences disclosed herein. In some aspects, the present disclosure provides for methods of detection using an rAAV virion having any one of the engineered AAV VP capsid polypeptide sequences disclosed herein. The method comprises administering an effective amount of the pharmaceutical composition comprising rAAV virions having any one of the AAV VP capsid polypeptide sequences disclosed herein to a subject in need thereof. The rAAV virions encapsidate any payload, including those payloads disclosed herein.

In some embodiments, the effective amount is at least 1×10⁸viral genomes per dose. In some embodiments, the effective amount is at least 5×10⁸viral genomes/dose, 7.5×10⁸viral genomes/dose, at least 1×10¹¹viral genomes/dose, at least 2.5×10⁹viral genomes/dose, at least 5×10⁹viral genomes/dose.

In some embodiments, the effective amount is at least 1×10¹¹viral genomes/kg patient weight, at least 5×10¹¹viral genomes/kg, at least 1×10¹¹viral genomes/kg, at least 5×10¹²viral genomes/kg, at least 1×10¹³viral genomes/kg, at least 1×10¹⁴viral genomes/kg, or at least 5×10¹⁴.

In some embodiments, the rAAV virion is administered via a systemic administration route including enteral routes of administration and parenteral routes of administration. The rAAV virion may be administered intravenously. In some embodiments, the rAAV may be administered intramuscularly. In some embodiments, the rAAV may be administered intraperitoneally. In some embodiments, the rAAV may be administered topically. In some embodiments, the rAAV may be administered orally. In particular embodiments, the rAAV virion is administered intravenously. In some embodiments, the rAAV is administered intrathecally. In some embodiments, the rAAV is administered by intracerebral ventricular injection. In some embodiments, the rAAV is administered by intracisternal magna administration. In some embodiments, the rAAV is administered by intravitreal injection.

In various embodiments, the patient suffers from one of the conditions listed in TABLE 1, below. In particular embodiments, the patient suffers from one of the conditions listed in TABLE 1 and the rAAV includes the transgene product associated therewith in TABLE 1.

In some embodiments, the therapeutic polynucleotide encodes a guide RNA, a tRNA, a suppressor tRNA, a siRNA, a miRNA, an mRNA, a shRNA, a circular RNA, or an antisense oligonucleotide (ASO), a ribozyme, a DNAzyme, an aptamer, or any combination thereof. In some embodiments, the therapeutic polynucleotide encodes a linear therapeutic polynucleotide or a circular therapeutic polynucleotide.

In some embodiments, the therapeutic polynucleotide encodes a therapeutic protein (a transgene). In particular embodiments, the transgene encodes a protein selected from the targets suitable for modification or transgene products of TABLE 1.

TABLE 1

Suitable Therapeutic Targets

Target of a Therapeutic

Primary gene delivery target
Condition
Polynucleotide

Brain/CNS
AADC deficiency
AADC

Alzheimer's Disease
Multiple, including APP, SNCA,

MAPT, ApoE, NGF, TERT

Tauopathies
MAPT

Synucleinopathies
SNCA

Batten disease (CLN2)
CLN2

Batten disease (CLN3)
CLN3

Batten disease (CLN6)
CLN6

MPS-IIIB
NAGLU

Frontotemporal dementia
GRN

with GRN mutations (FTD-

GRN)

Parkinson's Disease with
GBA1

GBA1 mutations (PD-

GBA) and neuronpathic

Gaucher's disease

Synucleinopathies
GBA1 + alpha-synuclein

Gaucher disease type 2
GBA

Canavan Disease
ASPA

Parkinson disease
AADC

Parkinson disease
GDNF

Parkinson disease
Neurturin

Parkinson disease
GAD

Parkinson disease
ntn

Parkinson disease
hFOXG1

Parkinson disease
hKCNQ2

Parkinson disease
hFMR1

Parkinson disease
anti-Tau/miRNA

Parkinson disease
EPM2A or EPM2B

Parkinson disease
LRRK2

Parkinson's Disease
LRRK2

Parkinson's Disease
SNCA

Tay-Sachs Disease
HEXA

Huntington's disease
IT15

Huntington's disease
CYP46A1

Huntington's disease
HTT

Protocki-Lupski Syndrome
IT15

Amyotrophic lateral
C9orf72

sclerosis

Amyotrophic lateral
SOD1

sclerosis

Down syndrome
DYRK1A

Sanfilippo disease type A
SGSH

Sanfilippo disease type B
hNAGLU

(Nervous system)
HEXB and HEXA

(Nervous system)
human codon-optimized CLN1

complementary DNA

(Nervous system)
SURF1

(Nervous system)
anti-UBE3A-ATS shRNA

(Nervous system)
hSLC6A1

Rett syndrome
MECP2

Spinal cord
spinal muscular atrophy
SMN

(SMA)

Giant axonal neuropathy
GAN

Chronic Pain
Nav1.7

spinocerebellar ataxias
ATXN3

(SCAs),

Eye
Achromatopsia
CNGB3

Choroideraemia
REP1

ad Retinitis Pigmentosa
NRL, RDH12, PRPH2 (RDS),

RHO, RPGR, SNRNP200,

NR2E3, IMPDH1, CRX, HK1,

IMPDH2, SNRNP200

Stargardt disease
ABCA4

Usher Syndrome 2A
USH2A

Wet AMD, Dry AMD
NRP1

Leber congenital amaurosis
RPE65

(LCA)

Leber hereditary optic
ND4

neuropathy (LHON)

retinitis pigmentosa (RP,
RLBP1

including RLBP1)

Wet AMD
Anti-VEGF antibody

X-linked retinitis
RPGR

pigmentosa (X-linked RP)

X-linked retinoschisis
RS1

Liver
Crigler-Najjar syndrome
UGT1A1

Familial
LDLR

Hypercholesterolemia (FH

homozygous)

Glycogen storage disease
G6PC

type 1A (GSD1a)

Haemophilia A
FVIII

Haemophilia B
FIX

Mucopolysaccharidosis I
ZFN1, ZFN2 and IDUA donor

(MPS-I)

Mucopolysaccharidosis II
ZFN1, ZFN2 and IDS donor

(MPS-II)

Mucopolysaccharidosis
SGSH

IIIA (MPS-IIIA)

Mucopolysaccharidosis
NAGLU

IIIB (MPS-IIIB)

Mucopolysaccharidosis VI
ARSB

(MPS-VI)

hydroxylase deficiency
CYP21A2

Cardiovascular disease
PCSK9

Porphyria and Acute
ALAS1

hepatic porphyria

Hemochromatosis
HFE

Cholesteryl ester storage
LIPA

disease

Wilson disease
ATP7B

Adult polyglucosan body-
GBE1 (also muscle cells)

disease

hepatic steatosis
hSLC13A5

Alpha-1 antitrypsin
SERPINA1

deficiency

Ornithine
OTC

Transcarbamylase

Deficiency (OTC

deficiency)

Alpha-1 antitrypsin
A1AT

deficiency (A1AT

deficiency)

Muscle
Charcot-Marie-Tooth
NTF3

disease type 1A (CMT1A)

Duchenne muscular
Micro-dystrophin

dystrophy (DMD)

Duchenne muscular
Mini-dystrophin

dystrophy (DMD)

Dysferlinopathy
DYSF

Pompe disease
GAA

Limb-girdle muscular
FKRP

dystrophies (LGMD)

(2i/R9)

Duchenne muscular
DMD

dystrophy (DMD)

Facioscapulohumeral
DUX4

Dystrophy

Myotonic Dystrophy
DMPK

Glycogen storage disorders
anti-GYS1 miRNA

X-linked myotubular
MTM1

myopathy (X-linked MTM)

euchromatic histone-lysine
anti-EHMT2 shRNA

N-methyltransferase 2

Other
(Associated with hearing
TMC1

loss)

Obesity (adipose tissue)
CIDEC, SCD1, GNB3

Bone (osteoclasts)
CLCN7

Chondrocytes
FGFR3

Primary Hyperoxaluria
HAO1

Type 1 (kidney)

Primary Hyperoxaluria
LDHA

(kidney)

Acromegaly (multi-organ)
GHR

Asthma (WBCs;
Mex3B

neutrophils, eosinophils)

Alport syndrome (kidney)
COL4A5

Transthyretin amyloidosis
TTR

(familial) (multi-organ)

Charcot-Marie Tooth
PMP22

Syndrome (PNS/Sciatic

Nerve; Schwann Cells)

Angelman syndrome
UBE3A

(nervous system)

Congestive heart failure
I-1c

(heart)

Methylmalonic acidemia
MMUT, MMAA, MMAB,

(MMA) (Kidneys)
MMADHC, MCEE

Cystic fibrosis (lung)
CFTR

HIV infections
PG9 antibody

HIV infections
VRC07 antibody

Anemia-related disorders
Hemophilia
F8 (Factor VIII), F9 (Factor IX)

Sickle-cell related disorders
sickle cell anemia
HBB

sickle cell hemoglobin C
Hemoglobin

disease

sickle cell thalassemia
beta thalassemia

disease

In some embodiments, the therapeutic polynucleotide encodes a therapeutic RNA. In some embodiments the therapeutic polynucleotide encodes an RNA, such as a guide RNA (including an engineered or synthetic guide RNA) for genome editing or for RNA editing.

In some embodiments, the therapeutic polynucleotide encodes a tRNA or a modified tRNA (engineered or synthetic tRNA). For example, the tRNA or modified tRNA can be a suppressor tRNA. The suppressor tRNA can be engineered to have an anticodon region that recognizes a stop codon, such as any premature stop codon (opal, ochre, or amber stop codons).

In some embodiments, the therapeutic polynucleotide (e.g., a therapeutic RNA, a tRNA, or a genome modifying entity) can target a gene listed in TABLE 1 or any gene associated with a neurologic disease, Parkinson's disease, Alzheimer's disease, a Tauopathy. Stargardt disease, alpha-1 antitrypsin deficiency, Duchenne's muscular dystrophy, Rett syndrome, cystic fibrosis, or any genetic disease. In some embodiments, the targeted gene may be ABCA4, AAT, SERPINA1, SERPINA1 E342K, HEXA, LRRK2, SNCA, DMD, APP, Tau, GBA, PINK1, RAB7A, CFTR, ALAS1, ATP7B, ATP7B G1226R, HFE C282Y, LIPA c.894 G>A, PCSK9 start site, or SCNN1A start site, a fragment any of these, or any combination thereof. In some embodiments, the therapeutic polynucleotide is a gene therapy payload (e.g., a transgene) and, thus, may itself be one of the genes listed in TABLE 1 or any gene associated with a neurologic disease, Parkinson's disease, Alzheimer's disease, a Tauopathy, Stargardt disease, alpha-1 antitrypsin deficiency, Duchenne's muscular dystrophy, Rett syndrome, cystic fibrosis, or any genetic disease. In some embodiments, the transgene may be ABCA4, AAT, SERPINA1, SERPINA1 E342K, HEXA, LRRK2, SNCA, DMD. APP, Tau, GBA, PINK1, RAB7A, CFTR, ALAS1, ATP7B, ATP7B G1226R, HFE C282Y, LIPA c.894 G>A, PCSK9 start site, or SCNN1A start site, a fragment any of these, or any combination thereof.

In some embodiments, the therapeutic polynucleotide encodes genome modifying entities. For example, a genome modifying entity may be a DNA editing enzyme, an RNA editing enzyme, a transcriptional activator, or a transcriptional repressor. The DNA editing enzyme may be any DNA editing enzyme, including any CRISPR/Cas systems, meganucleases, zinc-finger nucleases, (ZFNs). TALE Nucleases (TALENs and megaTALENS). The CRISPR/Cas system can be a Cas3, Cas8, Cas10, Cas9, Cas4, Cas12, or Cas13. The RNA editing enzyme may be ADAR. In some embodiments, the ADAR is a human ADAR1 or human ADAR2. The transcriptional activator may be VP64. A transcriptional repressor may be KRAB. Such genome modifying entities may target any gene listed in TABLE 1 for editing.

In some embodiments, the therapeutic polynucleotide is a polynucleotide capable of serving as a homology template for homology-directed repair.

In some embodiments, an rAAV virion of the present disclosure, having any of the engineered AAV VP capsid polypeptide sequences disclosed herein, comprises a vector genome, the vector genome comprising a detectable polynucleotide or payload. In further embodiments, said payload may be under control of regulatory sequences that direct expression in infected human cells. Examples of detectable polynucleotides include, but are not limited to, any genetically encodable detectable moiety. For example, a genetically encodable detectable moiety may be a fluorescent protein such as EGFP, GFP, YFP, RFP, CFP, or any variants thereof. In some embodiments, the present disclosure provides for rAAV virions having an engineered AAV VP capsid polypeptide, where the virion encapsidates any one of or any combination of the detectable payloads disclosed herein. In some embodiments, multiple copies of the detectable payload are encapsidated.

In some embodiments, the present disclosure provides for rAAV virions having an engineered AAV VP capsid polypeptide, where the virion encapsidates any one of or any combination of the therapeutic payloads and detectable payloads disclosed herein. For example, an rAAV of the present disclosure having an engineered AAV VP capsid polypeptide may encapsidate a transgene and a fluorescent protein. As another example, an rAAV of the present disclosure having an engineered AAV VP capsid polypeptide may encapsidate a therapeutic RNA (e.g., a guide RNA) and a fluorescent protein.

6.7. In Vivo Selected VP Polypeptides

In a further aspect, engineered (synonymously, recombinant) adeno-associated virus (AAV) VP capsid polypeptides identified using the methods described herein are provided.

In some embodiments, the engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide has an amino acid sequence at least 70% identical to SEQ ID NO: 1, wherein the engineered AAV VP capsid polypeptide has at least one substitution as compared to SEQ ID NO: 1 in the region from residue 581 to residue 589 of SEQ ID NO: 1, inclusive, wherein the capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV), and wherein the VP capsid polypeptide does not have the sequence of any of SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8.

In some embodiments, the engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide has an amino acid sequence at least 70% identical to SEQ ID NO: 1, wherein the engineered AAV VP capsid polypeptide has at least one substitution as compared to SEQ ID NO: 1 in the region from residue 581 to residue 589 of SEQ ID NO: 1, inclusive, wherein the capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV), wherein the at least one substitution confers higher tropism for a central nervous system (CNS) tissue on the rAAV as compared to an rAAV virion having an AAV5 VP capsid polypeptide of SEQ ID NO: 1, and wherein the VP capsid polypeptide does not have the sequence of any of SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8.

In particular embodiments, the engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide has an amino acid sequence at least 75%, 80%, 85%, 90%, 91%, 92%, 93%, 94%, 95%, 96%, 97%, 98% identical to the sequence of SEQ ID NO: 1.

In some embodiments, the AAV VP capsid polypeptide has an amino acid sequence of SEQ ID NO: 2, wherein Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from any amino acid, wherein the capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV), and wherein the VP capsid polypeptide does not have the sequence of any of SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8, optionally with further mutations elsewhere in the VP capsid polypeptide

In some embodiments, the AAV VP capsid polypeptide has an amino acid sequence of SEQ ID NO: 2, wherein Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from any amino acid, wherein the capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV), wherein the at least one substitution confers higher tropism for a central nervous system (CNS) tissue on the rAAV as compared to an rAAV virion having an AAV5 VP capsid polypeptide of SEQ ID NO: 1, and wherein the VP capsid polypeptide does not have the sequence of any of SEQ ID NO: 3. SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8, optionally with further mutations elsewhere in the VP capsid polypeptide

In some embodiments, the engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide has an amino acid sequence of SEQ ID NO: 2, wherein amino acid residues Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V; wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV); and wherein the rAAV VP capsid polypeptide does not have the sequence of any of SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8.

In some embodiments, the region of the engineered VP capsid polypeptide from residue 581 to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, or at least 99% identical to any one of SEQ ID NO:7118-SEQ ID NO:10,117. In particular embodiments, the region of the engineered VP capsid polypeptide from residue 581 to residue 589, inclusive, has a sequence that is identical to any one of SEQ ID NO:7118-SEQ ID NO:10,117.

In some embodiments, the engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide is an engineered AAV5 viral capsid protein, wherein the engineered AAV VP5 capsid polypeptide has at least one substitution as compared to SEQ ID NO: 1 in the region from residue 581 to residue 589 of SEQ ID NO: 1, inclusive; wherein the capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV); wherein the at least one substitution confers higher tropism for a central nervous system (CNS) tissue on the rAAV as compared to an rAAV virion having an AAV5 VP capsid polypeptide of SEQ ID NO: 1, and wherein the VP capsid polypeptide does not have the sequence of any of SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8, optionally with further mutations elsewhere in the VP protein.

In some embodiments, the AAV VP capsid polypeptides have an amino acid sequence of SEQ ID NO: 2, wherein Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V; and wherein the polypeptide does not have the sequence of any of SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8.

In some embodiments, the engineered AAV VP capsid polypeptide comprises a polypeptide sequence represented by the formula: (A)-(X)-(B)

wherein:

(A) is the polypeptide sequence of SEQ ID NO: 47438

(VAYNVGGQMATNNQSSTTAP residues 561-580 of SEQ ID

NO: 2);

(X) is the polypeptide sequence comprising amino acid residues Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 of SEQ ID NO: 2; and

(B) is the polypeptide sequence of SEQ ID NO:47439 (IVPGSVWMERDVYLQGPIWA residues 590-609 of SEQ ID NO: 2;

wherein Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V; and wherein the capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV); and;

wherein the polypeptide does not have the sequence of any of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4. SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, and SEQ ID NO:8.

In some embodiments, the engineered AAV VP capsid polypeptide comprises a polypeptide sequence represented by the formula: (A)-(X)-(B) wherein:

(A) is the polypeptide sequence of SEQ ID NO: 47438

(residues 561-580 of SEQ ID NO: 2 VAYNVGGQMATNNQSST

TAP);

(X) is a polypeptide sequence selected from the list of polypeptides in Table 8 (SEQ ID NOs:115-1114) or Table 10 (SEQ ID NOs: 7118-8117) that confers CNS tissue tropism on a recombinant AAV virion (rAAV); and

(B) is the polypeptide sequence of SEQ ID NO: 47439 (residues 590-609 of SEQ ID NO: 2: (IVPGSVWMERDVYLQGPIWA)); and

wherein the capsid polypeptide is capable of assembling into the rAAV and,

the capsid does not have the sequence of any of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, and SEQ ID NO:8.

In some embodiments, the engineered AAV VP capsid polypeptide confers CNS tissue tropism, wherein the CNS tissue is selected from the group consisting of hippocampus: (dentate gyrus. CA1 and CA3); cerebellum, hypothalamus, cortex: (occipital, temporal and forebrain); substantia nigra, thalamus, and any combination thereof.

In some embodiments, the engineered AAV VP capsid polypeptide comprises a polypeptide sequence represented by the formula: (A)-(X)-(B) wherein:

(A) is the polypeptide sequence of SEQ ID NO: 47438 (residues 561-580 of SEQ ID NO: 2: (VAYNVGGQMATNNQSSTTAP));

(X) is a polypeptide sequence selected from the polypeptides of SEQ ID NO: 115-1114 or SEQ ID NO: 1118-47437 that confer corresponding tissue tropism on a recombinant AAV virion (rAAV); and

(B) is the polypeptide sequence of SEQ ID NO: 47439 (residues 590-609 of SEQ ID NO: 2: (IVPGSVWMERDVYLQGPIWA)); and

wherein the capsid polypeptide is capable of assembling into the rAAV and,

the capsid does not have the sequence of any of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, and SEQ ID NO:8.

Described below are engineered mutated AAV5 VP1 polypeptide sequences that confer stable or improved virion assembly, tissue tropism, or both. In some embodiments, the present disclosure provides an AAV5 VP1 capsid polypeptide having a sequence homology of no more than 98.7% to SEQ ID NO: 1, wherein the AAV5 capsid polypeptide sequence has at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of SEQ ID NO: 1.

Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the Tables of the Examples (e.g., Table 7, 10, 12, 14, 16, 18, 20, 22, 24, 26, 28, 30, 32, 34, 36, 39, 41, 43, 45, 47, 49, 51, 53, 55, 57, 59, 61, 63, 65, 67, 69, 71, 72, 73, 74, 75, 76, 77, 78, 79, 80, 81, 82, 83, 84, 85, and 86) at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

6.7.1. In Vivo Selected VP Polypeptides that Confer Increased Liver Tropism

In various embodiments, the present disclosure provides a mutated VP polypeptide capable of forming an assembled virion that exhibits increased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid polypeptide of SEQ ID NO: 1. In this section of the disclosure, liver tissue tropism is determined by the frequency of a given amino acid residue occurring at a specified position corresponding to position 581 to position 589 of SEQ ID NO: 1 (generalized in SEQ ID NO: 2) over the frequency of that given amino acid residue in the total library of virus administered to NHP.

In some embodiments, Xaa1 is selected from A, G, K, M, N, Q, R, S, or T.

In some embodiments, Xaa1 is selected from A, K, M, or T.

In some embodiments, Xaa1 is K.

In additional embodiments, Xaa2 is selected from A, C, H, I, K, S, T, or V.

In some embodiments, Xaa2 is selected from A, S, T, or V.

In some embodiments, Xaa2 is T.

In additional embodiments, Xaa3 is selected from A, G, H, K, M, N, Q, R, S, T, or V.

In some embodiments, Xaa3 is selected from A, M, or T.

In some embodiments. Xaa3 is A or T.

In additional embodiments, Xaa4 is selected from L, M, P, Q, R, T, or W.

In some embodiments, Xaa4 is selected from L, P, Q, or T.

In some embodiments, Xaa4 is P.

In additional embodiments, Xaa5 is selected from F, H, I, K, M, T, or Y.

In some embodiments, Xaa5 is selected from H, I, or Y.

In some embodiments, Xaa5 is Y.

In additional embodiments, Xaa6 is selected from E, G, H, L, M, N, Q, T, or W.

In some embodiments, Xaa6 is selected from N, or Q.

In some embodiments, Xaa6 is N.

In additional embodiments, Xaa7 is selected from A, C, G, H, L, M, R or S.

In some embodiments, Xaa7 is selected from A, C, H or M.

In some embodiments, Xaa7 is A.

In additional embodiments, Xaa8 is selected from A, C, D, F, G, H, M, Q, S, V, W, or Y.

In some embodiments, Xaa8 is selected from G, M, Q, or S.

In some embodiments, Xaa8 is G.

In additional embodiments, Xaa9 is selected from A, C, E, G, H, M, N, P, Q, S, V, or W.

In some embodiments, Xaa9 is selected from E, G, or P.

In some embodiments, Xaa9 is G.

In particular embodiments, the sequence of Xaa1-Xaa9 of the engineered (recombinant) capsid polypeptide is selected from the amino acid sequence provided in TABLE 2.

In some embodiments, the engineered AAV capsid and corresponding virion exhibits increased liver tropism, when compared with AAV5 wildtype capsid and corresponding virion. This increased tropism can range from about 1.5, 2.0, 2.5, 3.0, 3.5, 4.0, 4.5, 5.0, 5.5, 6.0, 6.5, 7.0, 7.5, 8.0, 8.5, 9.0, 9.5, to about 10.0-fold when compared to a virion that comprises the AAV5 VP1 capsid polypeptide of SEQ ID NO: 1.

TABLE 2

Sequences of the 581 to 589 Region in

AAV5 VP1 Capsid Polypeptide that Drive

Liver Tropism

Var #
Translation

SEQ ID NO: 10
IAVASHAHG

SEQ ID NO: 11
SAYPKSEDV

SEQ ID NO: 12
SSMQCAMRP

SEQ ID NO: 13
ITVVVGEVN

SEQ ID NO: 14
SEAGDRCSD

SEQ ID NO: 15
ECACGHVHL

SEQ ID NO: 16
TFTNAGKMC

SEQ ID NO: 17
LASPYLIDA

SEQ ID NO: 18
TTQLYEVGS

SEQ ID NO: 19
TVHPIDHAT

SEQ ID NO: 20
TWMMVHRYV

SEQ ID NO: 21
WVYRCDVRH

SEQ ID NO: 22
HWADKALDW

SEQ ID NO: 23
MHTMKNCGW

SEQ ID NO: 24
LCNRMQADQ

SEQ ID NO: 25
WKATPGGQC

SEQ ID NO: 26
ARETTPYQT

SEQ ID NO: 27
MHMSYHWRE

SEQ ID NO: 28
MPKAEIFVP

SEQ ID NO: 29
DKTICQHQL

SEQ ID NO: 30
TTRGSERCS

SEQ ID NO: 31
HTPLAWHGD

SEQ ID NO: 32
SCIPFQQHG

SEQ ID NO: 33
AHQLAMVMN

SEQ ID NO: 34
AAAQRSRYV

SEQ ID NO: 35
DCMTSMLGP

SEQ ID NO: 36
RYLFAVWRY

SEQ ID NO: 37
MMNMQMDRM

SEQ ID NO: 38
AVQWIQWVG

SEQ ID NO: 39
QSQPHVQYN

SEQ ID NO: 40
THIFAYENN

SEQ ID NO: 41
GWQDEQMHL

SEQ ID NO: 42
TSKQETPQQ

SEQ ID NO: 43
CSNSPACLC

SEQ ID NO: 44
VDAHLETNG

SEQ ID NO: 45
CNEKEYEWH

SEQ ID NO: 46
IADIGRMQW

SEQ ID NO: 47
TEWPYGVAF

SEQ ID NO: 48
QCQYQTAWW

SEQ ID NO: 49
GPSGIFHCG

SEQ ID NO: 50
MAHAIDALQ

SEQ ID NO: 51
MHHCYQTFA

SEQ ID NO: 52
GQTVFHYDG

SEQ ID NO: 53
CNAIIAMPC

SEQ ID NO: 54
GTITLMPTQ

SEQ ID NO: 55
TVQCEMDVC

SEQ ID NO: 56
CQHNTPFVQ

SEQ ID NO: 57
RWTDTNFRG

SEQ ID NO: 58
KYEQKWMMD

SEQ ID NO: 59
MLYSTTCWK

SEQ ID NO: 60
ASWVGSFAQ

SEQ ID NO: 61
TTKQYHTME

SEQ ID NO: 62
PGGCYRMME

SEQ ID NO: 63
RNAVCLYRH

SEQ ID NO: 64
VIHEMLSAC

SEQ ID NO: 65
YGEQKYNHS

SEQ ID NO: 66
ANQWENAVK

SEQ ID NO: 67
TSAWWWCSP

SEQ ID NO: 68
EQTMMDLVY

SEQ ID NO: 69
YAMPRNPNV

SEQ ID NO: 70
MHKVWWMKN

SEQ ID NO: 71
AILQMYTAQ

SEQ ID NO: 72
CSTPANSCP

SEQ ID NO: 73
KVVEAELWQ

SEQ ID NO: 74
IANIDPSRV

SEQ ID NO: 75
QLTTTKPLR

SEQ ID NO: 76
ACWRFECDD

SEQ ID NO: 77
QINPEHGGC

SEQ ID NO: 78
MAPPFHVYE

SEQ ID NO: 79
DNWGTWTWT

SEQ ID NO: 80
DESSCVAHG

SEQ ID NO: 81
VAATFNNKV

SEQ ID NO: 82
EVVPFPWAF

SEQ ID NO: 83
TQMPALAEW

SEQ ID NO: 84
NVTNQQYDH

SEQ ID NO: 85
NMTEYVAYR

SEQ ID NO: 86
NVSTVHQPL

SEQ ID NO: 87
QTGPMTHSW

SEQ ID NO: 88
ECMEKHASY

SEQ ID NO: 89
LQAHVVRCT

SEQ ID NO: 90
VGDRYSSMG

SEQ ID NO: 91
PQGLIPMWA

SEQ ID NO: 92
MYVHKGYRS

SEQ ID NO: 93
AVPQYQKAE

SEQ ID NO: 94
ERMMILCSP

SEQ ID NO: 95
NFGFTCPVY

SEQ ID NO: 96
SQIWNVAAY

SEQ ID NO: 97
MWGQQGTWA

SEQ ID NO: 98
QAMMMTMMN

SEQ ID NO: 99
AHTANEFSP

SEQ ID NO: 100
DAHYVYEKG

SEQ ID NO: 101
CNNWIWAHE

SEQ ID NO: 102
NHNLMWVVS

SEQ ID NO: 103
ATMWGDCDY

SEQ ID NO: 104
EWMQEFAGP

SEQ ID NO: 105
QDGSVEWAF

SEQ ID NO: 106
WCPQPPGGN

SEQ ID NO: 107
AECQIWYDW

SEQ ID NO: 108
NAVKFVCED

SEQ ID NO: 109
TQCFASCVA

SEQ ID NO: 110
TVNNHDIGY

6.7.2. In Vivo Selected Engineered VP Polypeptides that are Competent for Assembly into rAAV

In various preferred embodiments, the mutated (engineered, recombinant) VP capsid polypeptides of the present disclosure are capable of forming an assembled virion, and in some instances that exhibit similar or improved stability when compared to a virion that comprises the AAV5 VP1 capsid polypeptide of SEQ ID NO: 1.

The frequency of a given amino acid residue occurring in assembled, purified viruses at a specified position corresponding to position 581 to position 589 of SEQ ID NO: 1 (generalized in SEQ ID NO: 2) over the frequency of that given amino acid residue occurring at the specified position in the entire plasmid library was analyzed to identify sequence rules for capsids that preferentially virally assembly.

Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that may exhibit similar or improved stability as compared to wildtype AAV5 VP capsid polypeptide, wherein the engineered variant AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein Xaa1 is selected from A, D, E, G, L, M, N, Q, S, T, or V, or Xaa1 is selected from A, D, E, M, or T. In some embodiments, Xaa1 is E; or Xaa2 is selected from A, C, D, E, G, H, I, N, P, Q, S, T, or V. or Xaa2 is selected from A, S, T, or V, or Xaa2 is A; or wherein Xaa3 is selected from A, D, E, G, H, M, N, Q, S, T, or V, or Xaa3 is selected from D, E, N, Q or T, or Xaa3 is D or T; or wherein Xaa4 is selected from A, D, E, G, H, N, P, Q, S, or T, or Xaa4 is selected from D, E, P, or Q, or Xaa4 is E; or wherein Xaa5 is selected from A, C, D, E, G, H, N, Q, S, T, or Y, or Xaa5 is selected from D, E, N, Q or T, or Xaa5 is N; or wherein Xaa6 is selected from A, D, E, G, H, N, P, Q, S, or T, or Xaa6 is selected from D, N, or Q, or Xaa6 is D; or wherein Xaa7 is selected from A, C, D, E, G, H, N, Q, S, or T, or Xaa7 is selected from A, D, E or G, or Xaa7 is A; or wherein Xaa8 is selected from A, C, D, E, G, H, N, Q, S, or T, or Xaa8 comprises A, D, G, or S, or Xaa8 is G; or wherein Xaa9 is selected from A, D, E, G, H, N, P, Q, S, or T, or Xaa9 is selected from A, D, G, or P, or Xaa9 is G.

In various embodiments, the VP polypeptide is capable of forming an assembled virion, and in some instances exhibits similar or improved stability when compared to a virion that comprises the AAV5 VP1 capsid polypeptide of SEQ ID NO:1.

In some embodiments, Xaa1 is selected from A, D, E, G, L, M, N, Q, S, T, or V.

In some embodiments, Xaa1 is selected from A, D, E, M, or T. In some embodiments, Xaa1 is E.

In some embodiments, Xaa2 is selected from A, C, D, E, G, H, I, N, P, Q, S, T, or V. In some embodiments, Xaa2 is selected from A, S, T, or V. In some embodiments, Xaa2 is A.

In some embodiments, Xaa3 is selected from A, D, E, G, H, M, N, Q, S, T, or V. In some embodiments, Xaa3 is selected from D, E, N, Q or T. In some embodiments, Xaa3 is D or T.

In some embodiments, Xaa4 is selected from A, D, E, G, H, N, P, Q, S, or T. In some embodiments, Xaa4 is selected from D, E, P, or Q. In some embodiments, Xaa4 is E.

In some embodiments, Xaa5 is selected from A, C, D, E, G, H, N, Q, S, T, or Y. In some embodiments, Xaa5 is selected from D, E, N, Q or T. In some embodiments, Xaa5 is N.

In some embodiments, Xaa6 is selected from A, D, E, G, H, N, P, Q, S, or T. In some embodiments, Xaa6 is selected from D, N, or Q. In some embodiments, Xaa6 is D.

In some embodiments. Xaa7 is selected from A, C, D, E, G, H, N, Q, S, or T. In some embodiments, Xaa7 is selected from A, D, E or G. In some embodiments, Xaa7 is A.

In some embodiments, Xaa8 is selected from A, C, D, E, G, H, N, Q, S, or T. In some embodiments, Xaa8 comprises A, D, G, or S. In some embodiments, Xaa8 is G.

In some embodiments, Xaa9 is selected from A, D, E, G, H, N, P, Q, S, or T. In some embodiments, Xaa9 is selected from A, D, G, or P. In some embodiments, Xaa9 is G.

6.7.3. In Vivo Selected Mutated VP Polypeptides that are Competent for Assembly into rAAV Virions and Exhibit Decreased Liver Tropism

The present disclosure provides AAV5 virions with a VP capsid polypeptide having at least one mutation in a region with residues that interact with target cells, where the at least one mutation confers decreased liver tissue tropism as compared to a wildtype VP capsid polypeptide. In some embodiments, provided herein are AAV5 VP1 capsid polypeptide having a sequence homology of at least 80% to SEQ ID NO: 1, wherein the AAV5 VP1 capsid polypeptide has at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of SEQ ID NO: 1 and wherein said at least one mutation drives decreased liver tropism.

Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits decreased liver tissue tropism as compared to wildtype AAV5 VP capsid polypeptide, wherein the engineered variant AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein Xaa1 is not K, or Xaa1 is not A, K, M, or T, or Xaa1 is not A, G, K, M, N, Q, R, S, or T; or wherein Xaa2 is not T, or Xaa2 is not A, S, T, or V, or Xaa2 is not A, C, H, I, K, S, T, or V; or wherein Xaa3 is not A or T, or Xaa3 is not A, M, or T, or Xaa3 is not A, G, H, K, M, N, Q, R, S, T, or V; or wherein Xaa4 is not P, or wherein Xaa4 is not L, P, Q, or T, or Xaa4 is not L, M, P, Q, R, T, or W; or wherein Xaa5 is not Y, or Xaa5 is not H, I, or Y, or Xaa5 is not F, H, I, K, M, T, or Y; or wherein Xaa6 is not N, or Xaa6 is not N, or Q, or Xaa6 is not E, G, H, L, M, N, Q, T, or W; or wherein Xaa7 is not A, or Xaa7 is not A, C, H or M, or Xaa7 is not A, C, G, H, L, M, R or S; or wherein Xaa8 is not G, or Xaa8 is not G, M, Q, or S, or Xaa8 is not A, C, D, F, G, H, M, Q, S, V, W, or Y; or wherein Xaa9 is not G, or Xaa9 is not E, G, or P, or Xaa9 is not A, C, E, G, H, M, N, P, Q, S, V, or W.

In certain embodiments, Xaa1 is not K, and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO: 1.

In certain embodiments, Xaa1 is not A, K, M, or T, and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, Xaa1 is not A, G, K, M, N, Q, R, S, or T, and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, Xaa2 is not T, and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, Xaa2 is not A, S, T, or V, and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, Xaa2 is not A, C, H, I, K, S, T, or V, and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, Xaa3 is not A or T, and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, Xaa3 is not A, M, or T, and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, Xaa3 is not A, G, H, K, M, N, Q, R, S, T, or V, and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, Xaa4 is not P, and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, wherein Xaa4 is not L, P, Q, or T. and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, Xaa4 is not L, M, P, Q, R, T, or W, and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, Xaa5 is not Y. and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, Xaa5 is not H, I, or Y, and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, Xaa5 is not F, H, I, K, M, T, or Y. and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, Xaa6 is not N, and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, Xaa6 is not N, or Q. and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, Xaa6 is not E, G, H, L, M, N, Q, T, or W, and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, Xaa7 is not A, and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, Xaa7 is not A, C, H or M, and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, Xaa7 is not A, C, G, H, L, M, R or S, and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, Xaa8 is not G, and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, Xaa8 is not G, M, Q, or S, and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, Xaa8 is not A, C, D, F, G, H, M, Q, S, V, W, or Y, and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, Xaa9 is not G, and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, Xaa9 is not E, G, or P. and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

In certain embodiments, Xaa9 is not A, C, E, G, H, M, N, P, Q, S, V, or W, and wherein the VP capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1.

The present disclosure encompasses variant VP capsids that have increased tissue tropism, compared to the AAV5 VP1 capsid of SEQ ID NO:1, for any of the following tissues: adipose, adrenal gland, aorta, brain (including hippocampus: dentate gyrus, CA1 and CA3; cerebellum, caudate, putamen, midbrain, pons, hypothalamus, cortex-including occipital, temporal and forebrain; substantia nigra, and thalamus), bone marrow, cecum, colon, dorsal root ganglion, duodenum, epididymis, esophagus, eye, gallbladder, heart, ileum, jejunum, kidney, lung, lymph nodes, mammary gland, ovary, pancreas, parathyroid gland, peripheral nerve, pituitary, prostate, salivary gland, seminal vesicle, skeletal muscle, skin, spinal cord, spleen, stomach, testis, thymus, thyroid, trachea, urinary bladder, uterus, and vagina.

6.7.4. In Vivo Selected Mutated VP Polypeptides that Detarget Liver Tissue

A. Positional Frequency Rules

In this section, unless otherwise specified, the frequency of a given amino acid residue occurring at a specified position corresponding to position 581 to position 589 of SEQ ID NO: 1 (generalized in SEQ ID NO: 2) in variants identified in non-liver over the frequency of that given amino acid residue occurring at the specified position in variants identified in all other harvested tissues (CNS (cortex forebrain, cortex occipital, cortex temporal, thalamus, hypothalamus, substantia nigra, hippocampus DG, hippocampus CA1, hippocampus CA3, cerebellum), skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord tissues, liver) was analyzed to identify a set of sequence rules for capsids that preferentially detarget liver tissue. Identification of positional frequency rules from in vivo data is described in detail in EXAMPLE 4. With reference to TABLE 6B in EXAMPLE 4, and SEQ ID NO: 2 (AAV5 VP1), the following amino acids can, thus, be independently mutated, in any combination, at any one or more positions Xaa1-Xaa9, to provide a VP1 capsid with reduced liver tropism as compared wildtype AAV5 VP1 capsid (SEQ ID NO: 1), where liver tropism here refers to properties that are deterministic for liver transduction over properties that are deterministic for transduction of all other harvested tissues.

Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits decreased liver tissue tropism as compared to wildtype AAV5 VP capsid polypeptide, wherein the engineered variant AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein Xaa1 excludes K; or Xaa1 excludes A, K, M, or T; or Xaa1 excludes A, G, K, M, N, Q, R, S, or T; or Xaa2 excludes T; or Xaa2 excludes A, S, T, or V; or Xaa2 excludes A, C, H, I, K, S, T, or V; or Xaa3 excludes A or T; or Xaa3 excludes A, M, or T; or Xaa3 excludes A, G, H, K, M, N, Q, R, S, T, or V; or Xaa4 excludes P; or Xaa4 excludes L, P, Q, or T; or Xaa4 excludes L, M, P, Q, R, T, or W; or Xaa5 excludes Y; or Xaa5 excludes H, I, or Y; or Xaa5 excludes F, H, I, K, M, T, or Y; or Xaa6 excludes N; or Xaa6 excludes N, or Q; or Xaa6 excludes E, G, H, L, M, N, Q, T, or W; or Xaa7 excludes A; or Xaa7 excludes A, C, H or M; or Xaa7 excludes A, C, G, H, L, M, R or S; or Xaa8 excludes G; or Xaa8 excludes G, M, Q, or S; or Xaa8 excludes A, C, D, F, G, H, M, Q, S, V, W, or Y; or Xaa9 excludes G; or Xaa9 excludes E, G, or P; or Xaa9 excludes A, C, E, G, H, M, N, P, Q, S, V, or W.

Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits decreased liver tissue tropism as compared to wildtype AAV5 VP capsid polypeptide, wherein the engineered variant AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein said one or more mutations are selected from the following rules. In some embodiments, Xaa1 excludes K. In some embodiments, Xaa1 excludes A, K, M, or T. In some embodiments, Xaa1 excludes A, G, K, M, N, Q, R, S, or T. In some embodiments, Xaa2 excludes T. In some embodiments, Xaa2 excludes A, S, T, or V. In some embodiments, Xaa2 excludes A, C, H, I, K, S, T, or V. In some embodiments, Xaa3 excludes A or T. In some embodiments, Xaa3 excludes A, M, or T. In some embodiments, Xaa3 excludes A, G, H, K, M, N, Q, R, S, T, or V. In some embodiments. Xaa4 excludes P. In some embodiments. Xaa4 excludes L, P, Q, or T. In some embodiments, Xaa4 excludes L, M, P, Q, R, T, or W. In some embodiments, Xaa5 excludes Y. In some embodiments, Xaa5 excludes H, I, or Y. In some embodiments, Xaa5 excludes F, H, I, K, M, T, or Y. In some embodiments, Xaa6 excludes N. In some embodiments, Xaa6 excludes N, or Q. In some embodiments, Xaa6 excludes E, G, H, L, M, N, Q, T, or W. In some embodiments, Xaa7 excludes A. In some embodiments, Xaa7 excludes A, C, H or M. In some embodiments, Xaa7 excludes A, C, G, H, L, M, R or S. In some embodiments, Xaa8 excludes G. In some embodiments, Xaa8 excludes G, M, Q, or S. In some embodiments, Xaa8 excludes A, C, D, F, G, H, M, Q, S, V, W, or Y. In some embodiments, Xaa9 excludes G. In some embodiments, Xaa9 excludes E, G, or P. In some embodiments, Xaa9 excludes A, C, E, G, H, M, N, P, Q, S, V, or W.

B. ML Rules

For the following set of rules described in this paragraph, favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an engineered AAV5 VP1 capsid polypeptide, were determined using in vivo data and two ML models, which are described in EXAMPLE 21. Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits decreased liver tissue tropism as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide, wherein the engineered AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein Xaa1 is selected from an amino acid of low solubility at position Xaa1 (e.g., Xaa1 is selected from D, or P); or wherein Xaa1 is selected from an amino acid of low mutability at position Xaa1 (e.g., Xaa1 is selected from C, K, or L); or wherein Xaa2 is selected from an amino acid of low solubility at position Xaa2 (e.g., Xaa2 is selected from N, K, P, E, or D); or wherein Xaa2 is selected from an amino acid of low hydropathy at position Xaa2 (e.g., Xaa2 is selected from D, E, R, K, H, N, or Q); or wherein Xaa2 is selected from an amino acid of low charge at position Xaa2 (e.g., Xaa2 is selected from D or E); or wherein Xaa2 is selected from an amino acid of high number of total potential hydrogen bonds at position Xaa2 (e.g., Xaa2 is selected from H, N, Q, D, E, or R); or wherein Xaa2 is selected from an amino acid of medium volume at position Xaa2 (e.g., Xaa2 is selected from D, E, V, P, N, or T); or wherein Xaa3 is selected from an amino acid of low solubility at position Xaa3 (e.g., Xaa3 is selected from P or D); or wherein Xaa4 is selected from an amino acid of medium volume at position Xaa4 (e.g., Xaa4 is selected from D, E, V, P, N, or T) or wherein Xaa5 is selected from an amino acid of low solubility at position Xaa5 (e.g., Xaa5 is selected from N, P, E, or D); or wherein Xaa8 is selected from an amino acid of low solubility at position Xaa8 (e.g., Xaa8 is selected from K or Q); or wherein Xaa8 is selected from an amino acid of low hydropathy at position Xaa8 (e.g., Xaa8 is selected from K or R); or wherein Xaa8 is selected from an amino acid of high surface accessibility at position Xaa8 (e.g., Xaa8 is selected from E, R, or K); or any combination thereof.

In some embodiments, Xaa1 is selected from an amino acid of low solubility at position Xaa1. In some embodiments, Xaa1 is selected from D or P. In some embodiments, Xaa1 is selected from an amino acid of low mutability at position Xaa1. In some embodiments. Xaa1 is selected from C, K, or L. In some embodiments, Xaa2 is selected from an amino acid of low solubility at position Xaa2. In some embodiments, Xaa2 is selected from N, K, P, E, or D. In some embodiments, Xaa2 is selected from an amino acid of low hydropathy at position Xaa2. In some embodiments, Xaa2 is selected from D, E, R, K, H, N, or Q. In some embodiments, Xaa2 is selected from an amino acid of low charge at position Xaa2. In some embodiments, Xaa2 is selected from D, E. In some embodiments, Xaa2 is selected from an amino acid of high number of total potential hydrogen bonds at position Xaa2. In some embodiments, Xaa2 is selected from H, N, Q, D, E, or R. In some embodiments, Xaa2 is selected from an amino acid of medium volume at position Xaa2. In some embodiments, Xaa2 is selected from D, E, V, P, N, or T. In some embodiments, Xaa3 is selected from an amino acid of low solubility at position Xaa3. In some embodiments, Xaa3 is selected from P or D. In some embodiments, Xaa4 is selected from an amino acid of medium volume at position Xaa4. In some embodiments, Xaa4 is selected from D, E, V, P, N, or T. In some embodiments, Xaa5 is selected from an amino acid of low solubility at position Xaa5. In some embodiments. Xaa5 is selected from N, P, E, or D. In some embodiments, Xaa8 is selected from an amino acid of low solubility at position Xaa8. In some embodiments, Xaa8 is selected from K or Q. In some embodiments, Xaa8 is selected from an amino acid of low hydropathy at position Xaa8. In some embodiments, Xaa8 is selected from K or R. In some embodiments. Xaa8 is selected from an amino acid of high surface accessibility at position Xaa8. In some embodiments, Xaa8 is selected from E, R, or K.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 46438-SEQ ID NO: 47437, wherein said at least one mutation drives liver detargeting tissue tropism.

6.7.5. In Vivo Selected Mutated VP Polypeptides that Confer Increased Liver Tropism

The present disclosure provides AAV5 virions with a VP capsid polypeptide having at least one mutation in a region with residues that interact with target cells (e.g., a target liver cell in a target liver tissue of interest), where the at least one mutation confers increased liver tissue tropism as compared to a wildtype VP capsid polypeptide. In some embodiments, provided herein are AAV5 VP1 capsid polypeptide having a sequence homology of at least 80% to SEQ ID NO: 1, wherein the AAV5 VP1 capsid polypeptide has at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of SEQ ID NO: 1 and wherein said at least one mutation drives increased liver tropism. The following sequences rules and sequences also apply to the region in AAV5 VP2 (amino acid residues 445 to 453; VP2 sequence shown in SEQ ID NO: 1115) and AAV5 VP3 (amino acid residues 389 to 397; VP3 sequences shown in SEQ ID NO: 1116) corresponding to AAV5 VP1 amino acid residues 581 to 589. Thus, the present disclosure encompasses AAV5 VP2 capsid polypeptides and AAV5 VP3 capsid polypeptides having one or more mutations in the VP2 and VP3 regions corresponding to the AAV5 VP1 amino acid residues of the 581 to 589 region, where the one or more mutations comport to the rules or sequences in the following section.

A. Positional Frequency Rules

Xaa1 is selected from A, K, Q, or R, or Xaa1 is K; or wherein Xaa2 is selected from A, C, I, K, S, T, or V, or Xaa2 is selected from A, K, S, or T, or Xaa2 is A; or wherein Xaa3 is selected from A, G, I, K, M, Q, R, S, T, or V, or Xaa3 is selected from A, K, Q, S, or T, or Xaa3 is selected from K, Q, or T, or Xaa3 is K; or wherein Xaa4 is selected from A, I, K, L, P, Q, R, S, T, or V, or Xaa4 is selected from K, I, S, or V, or Xaa4 is K; or Xaa5 is selected from F, I, L, M, T, V, or Y, or wherein Xaa5 is selected from F, L, or Y, or Xaa5 is F; or Xaa6 is selected from F, H, M, N, Q, S, or Y, or wherein Xaa6 is selected from M or N, or Xaa6 is N; or Xaa7 is selected from A, C, K, M, Q or S, or wherein Xaa7 is selected from A, C, or S, or Xaa7 is S; or wherein Xaa8 is selected from A, C, F, G, M, Q, or S, or Xaa8 is selected from A, C, M, or S, or Xaa8 is C; or wherein Xaa9 is selected from E, F, L, Q, R, or Y, or Xaa9 is selected from L, Q, or R, or Xaa9 is R.

Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased liver tissue tropism as compared to wildtype AAV5 VP capsid polypeptide, wherein the engineered variant AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein said one or more mutations are selected from the following rules. In some embodiments, Xaa1 is selected from A, G, K, M, N, Q, R, S, or T. In some embodiments, Xaa1 is selected from A, K, Q, or R. In some embodiments, Xaa1 is K. In some embodiments, Xaa2 is selected from A, C, I, K, S, T, or V. In some embodiments, is selected from A, K, S, or T, or Xaa2 is A. In some embodiments, wherein Xaa3 is selected from A, G, I, K, M, Q, R, S, T, or V, or Xaa3 is selected from A, K, Q, S, or T. In some embodiments, Xaa3 is selected from K, Q, or T. In some embodiments, Xaa3 is K. In some embodiments, Xaa4 is selected from A, I, K, L, P, Q, R, S, T, or V. In some embodiments, Xaa4 is selected from K, I, S, or V. In some embodiments, Xaa4 is K. In some embodiments, Xaa5 is selected from F, I, L, M, T, V, or Y. In some embodiments, Xaa5 is selected from F, L, or Y, or Xaa5 is F. In some embodiments, Xaa6 is selected from F, H, M, N, Q, S, or Y. In some embodiments, wherein Xaa6 is selected from M or N, or Xaa6 is N. In some embodiments, Xaa7 is selected from A, C, K, M, Q or S. In some embodiments, Xaa7 is selected from A, C, or S, or Xaa7 is S. In some embodiments. Xaa8 is selected from A, C, F, G, M, Q, or S. In some embodiments. Xaa8 is selected from A, C, M, or S, or Xaa8 is C. In some embodiments, Xaa9 is selected from E, F, L, Q, R, or Y. In some embodiments, Xaa9 is selected from L, Q, or R, or Xaa9 is R.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 43438-SEQ ID NO: 44437, wherein said at least one mutation drives increased liver tissue tropism.

B. ML Rules

For the following set of rules described in this paragraph, favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an engineered AAV5 VP1 capsid polypeptide, were determined using in vivo data and two ML models, which are described in EXAMPLE 20. Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased liver tissue tropism as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide, wherein the engineered AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein Xaa1 is selected from an amino acid of high surface accessibility (e.g., Xaa1 is selected from K, R, or E); or wherein Xaa1 is selected from an amino acid of low hydropathy (e.g., Xaa1 is selected from K, R); or wherein Xaa1 is selected from an amino acid of low amino acid mutability (e.g., Xaa1 is selected from H, P, K, or R); or wherein Xaa1 is selected from an amino acid of low amino acid solubility (e.g., Xaa1 is selected from Q, K, R); or wherein Xaa2 is selected from an amino acid of high surface accessibility (e.g., Xaa2 is selected from E, R, or K); or wherein Xaa2 is selected from an amino acid of low hydropathy (e.g., Xaa2 is selected from K. R); or wherein Xaa2 is selected from an amino acid of high amino acid volume (e.g., Xaa2 is selected from S, L, I, A, R, or K); or wherein Xaa3 is selected from an amino acid of high mutability (e.g., Xaa3 is selected from N, I, A, M, E, or D); or wherein Xaa3 is selected from an amino acid of low solubility (e.g., Xaa3 is selected from N, K, R, or E); or wherein Xaa4 is selected from an amino acid of low hydropathy (e.g., Xaa4 is selected from K or R); or wherein Xaa4 is selected from an amino acid of high amino acid volume (e.g., Xaa4 is selected from K, R, I, or L); or wherein Xaa5 is selected from an amino acid of medium amino acid solubility (e.g., Xaa5 is selected from H or T); or wherein Xaa8 is selected from an amino acid of low surface accessibility (e.g., Xaa8 is selected from V or C); or wherein Xaa8 is selected from an amino acid of low average flexibility index (e.g., Xaa8 is selected from W, V, M, A, F, L, H, or C); or any combination thereof.

In some embodiments, Xaa1 is selected from an amino acid of high surface accessibility. In some embodiments, Xaa1 is selected from K, R, or E. In some embodiments, Xaa1 is selected from an amino acid of low hydropathy. In some embodiments, Xaa1 is selected from K or R. In some embodiments, Xaa1 is selected from an amino acid of low amino acid mutability. In some embodiments, Xaa1 is selected from H, P, K, or R. In some embodiments, Xaa1 is selected from an amino acid of low amino acid solubility. In some embodiments, Xaa1 is selected from Q, K, or R. In some embodiments, Xaa2 is selected from an amino acid of high surface accessibility. In some embodiments, Xaa2 is selected from E, R, or K. In some embodiments, Xaa2 is selected from an amino acid of low hydropathy. In some embodiments, Xaa2 is selected from K or R. In some embodiments, Xaa2 is selected from an amino acid of high amino acid volume. In some embodiments. Xaa2 is selected from S, L, I, A, R, or K. In some embodiments, Xaa3 is selected from an amino acid of high mutability. In some embodiments, Xaa3 is selected from N, I, A, M, E, or D. In some embodiments, Xaa3 is selected from an amino acid of low solubility. In some embodiments, Xaa3 is selected from N, K, R, or E. In some embodiments, Xaa4 is selected from an amino acid of low hydropathy. In some embodiments, Xaa4 is selected from K, R In some embodiments, Xaa4 is selected from an amino acid of high amino acid volume. In some embodiments, Xaa4 is selected from K, R, I, or L. In some embodiments, Xaa5 is selected from an amino acid of medium amino acid solubility. In some embodiments, Xaa5 is selected from H, T. In some embodiments, Xaa8 is selected from an amino acid of low surface accessibility. In some embodiments, Xaa8 is selected from V or C. In some embodiments, Xaa8 is selected from an amino acid of low average flexibility index. In some embodiments, Xaa8 is selected from W, V, M, A, F, L, H, or C.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 44438-SEQ ID NO: 45437, wherein said at least one mutation drives increased liver tissue tropism.

C. Enriched Liver Sequences

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 45438-SEQ ID NO: 46437, wherein said at least one mutation drives increased liver tissue tropism.

6.7.6. In Vivo Selected Mutated VP Polypeptides that Confer Increased Central Nervous System Tropism, Positional Frequency Based Rules and ML Rules

The present disclosure provides AAV5 virions with a VP capsid polypeptide having at least one mutation in a region with residues that interact with target cells (e.g., a target CNS cell in a target CNS tissue of interest), where the at least one mutation confers increased CNS tissue tropism as compared to a wildtype VP capsid polypeptide. In some embodiments, provided herein are AAV5 VP1 capsid polypeptide having a sequence homology of at least 80% to SEQ ID NO: 1, wherein the AAV5 VP1 capsid polypeptide has at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of SEQ ID NO: 1 and wherein said at least one mutation drives increased central nervous system tropism. The following sequences rules and sequences also apply to the region in AAV5 VP2 (amino acid residues 445 to 453; VP2 sequence shown in SEQ ID NO: 1115) and AAV5 VP3 (amino acid residues 389 to 397; VP3 sequences shown in SEQ ID NO: 1116) corresponding to AAV5 VP1 amino acid residues 581 to 589. Thus, the present disclosure encompasses AAV5 VP2 capsid polypeptides and AAV5 VP3 capsid polypeptides having one or more mutations in the VP2 and VP3 regions corresponding to the AAV5 VP1 amino acid residues of the 581 to 589 region, where the one or more mutations comport to the rules or sequences in the following section.

A. Positional Frequency Rules

In this section, unless otherwise specified, the frequency of a given amino acid residue occurring at a specified position corresponding to position 581 to position 589 of SEQ ID NO: 1 (generalized in SEQ ID NO: 2) in variants identified in central nervous system (cortex forebrain, cortex occipital, cortex temporal, thalamus, hypothalamus, substantia nigra, hippocampus DG, hippocampus CA1, hippocampus CA3, cerebellum) over the frequency of that given amino acid residue occurring at the specified position in variants identified in all other harvested tissues (liver, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord tissues) was analyzed to identify a set of sequence rules for capsids that preferentially target central nervous system tissues. Identification of positional frequency rules from in vivo data is described in detail in EXAMPLE 5.

Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased central nervous system tissue tropism as compared to wildtype AAV5 VP capsid polypeptide, wherein the engineered variant AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein Xaa1 is selected from A, C, K, M, Q, R, T, or W, or Xaa1 is selected from K, Q, R, or W, or Xaa1 is K; or Xaa2 is selected from F, I, K, R, T, or W, or Xaa2 is selected from F, I, R or T, or Xaa2 is R; or Xaa3 is selected from A, H, N, R, or W, or Xaa3 is selected from A, R, or W, or Xaa3 is R; or Xaa4 is selected from E, G, I, M, Q, or R, or Xaa4 is selected from E, M, or R, or Xaa4 is R; or Xaa5 is selected from C, G, K, I, M, or R, or Xaa5 is selected from K, I, or R, or Xaa5 is I; or Xaa6 is selected from I, K, L, P, Q, R, Y, or Xaa6 is selected from K, R, or Y, or Xaa6 is R; or Xaa7 is selected from D, I, K, R, V, or W, or Xaa7 is selected from I, R, or V, or Xaa7 is V; or Xaa8 is selected from C, G, H, K, L, or V, or Xaa8 is selected from H. K, or V, or Xaa8 is H; or Xaa9 is selected from I, K, L, R, or V, or Xaa9 is selected from I, K, or R, or Xaa9 is R.

Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased central nervous system tissue tropism as compared to wildtype AAV5 VP capsid polypeptide, wherein the engineered variant AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein said one or more mutations are selected from the following rules. In some embodiments, Xaa1 is selected from A, C, K, M, Q, R, T, or W. In some embodiments. Xaa1 is selected from K, Q, R, or W. In some embodiments, Xaa1 is K. In some embodiments, Xaa2 is selected from F, I, K, R, T, or W. In some embodiments, Xaa2 is selected from F, I, R or T. In some embodiments, Xaa2 is R. In some embodiments, Xaa3 is selected from A, H, N, R, or W. In some embodiments, Xaa3 is selected from A, R, or W. In some embodiments, Xaa3 is R. In some embodiments, Xaa4 is selected from E, G, I, M, Q, or R. In some embodiments, Xaa4 is selected from E, M, or R. In some embodiments, Xaa4 is R. In some embodiments, Xaa5 is selected from C, G, K, I, M, or R. In some embodiments, Xaa5 is selected from K, I, or R. In some embodiments, Xaa5 is I. In some embodiments, Xaa6 is selected from I, K, L, P, Q, R, Y. In some embodiments, Xaa6 is selected from K, R, or Y. In some embodiments, Xaa6 is R. In some embodiments, Xaa7 is selected from D, I, K, R, V, or W. In some embodiments, Xaa7 is selected from I, R, or V. In some embodiments. Xaa7 is V. In some embodiments, Xaa8 is selected from C, G, H, K, L, or V. In some embodiments, Xaa8 is selected from H, K, or V. In some embodiments, Xaa8 is H. In some embodiments, Xaa9 is selected from I, K, L, R, or V. In some embodiments, Xaa9 is selected from I, K, or R. In some embodiments, Xaa9 is R.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 7118-SEQ ID NO: 8117, wherein said at least one mutation drives increased central nervous system tissue tropism.

B. ML Rules

For the following set of rules described in the subsequent paragraphs in this section, favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an engineered AAV5 VP1 capsid polypeptide, were determined using in vivo data and two ML models, which are described in EXAMPLE 19. Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased central nervous system tissue tropism as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide, wherein the engineered variant AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein Xaa1 is selected from an amino acid of low amino acid solubility (e.g., Xaa1 is selected from K, R, or Q); or wherein Xaa1 is selected from an amino acid of low amino acid hydropathy (e.g., Xaa1 is selected from K or R); or wherein Xaa1 is selected from an amino acid of high average amino acid flexibility index (e.g., Xaa1 is selected from D, E, R, K, G, I, N, Q, or S); or wherein Xaa1 is selected from an amino acid of high hydrogen bond donors (e.g., Xaa1 is selected from K or R); or wherein Xaa1 is selected from an amino acid of amino acid mutability (e.g., Xaa1 is selected from K, R, P, or H); or wherein Xaa2 is selected from an amino acid of low amino acid solubility (e.g., Xaa2 is selected from R, K, Q, or S); or wherein Xaa2 is selected from an amino acid of low amino acid hydropathy (e.g., Xaa2 is selected from R, K, D, E, N, Q, H, P, Y, W, S, or T); or wherein Xaa2 is selected from an amino acid of high amino acid charge (e.g., Xaa2 is selected from R, K, or H); or wherein Xaa3 is selected from an amino acid of high amino acid solubility (e.g., Xaa3 is selected from A, M, V, W, L, or I); or wherein Xaa5 is selected from an amino acid of high amino acid solubility (e.g., Xaa5 is selected from C, M, V, W, L, or I); or wherein Xaa5 is selected from an amino acid of high hydropathy (e.g., Xaa5 is selected from M, V, or I); or wherein Xaa5 is selected from an amino acid of low average amino acid flexibility index (e.g., Xaa5 is selected from M, W, F, or C); or wherein Xaa8 is selected from an amino acid of high amino acid solubility (e.g., Xaa8 is selected from H, V, or I); or any combination thereof.

In some embodiments, Xaa1 is selected from an amino acid of low amino acid solubility. In some embodiments, Xaa1 is selected from K, R, Q. In some embodiments, Xaa1 is selected from an amino acid of low amino acid hydropathy. In some embodiments, Xaa1 is selected from K or R. In some embodiments, Xaa1 is selected from an amino acid of high average amino acid flexibility index. In some embodiments, Xaa1 is selected from D, E, R, K, G, I, N, Q, or S. In some embodiments, Xaa1 is selected from an amino acid of high hydrogen bond donors. In some embodiments, Xaa1 is selected from K or R. In some embodiments, Xaa1 is selected from an amino acid of amino acid mutability. In some embodiments, Xaa1 is selected from K, R, P, or H. In some embodiments, Xaa2 is selected from an amino acid of low amino acid solubility. In some embodiments, Xaa2 is selected from R, K, Q, or S. In some embodiments, Xaa2 is selected from an amino acid of low amino acid hydropathy. In some embodiments, Xaa2 is selected from R, K, D, E, N, Q, H, P, Y, W, S, or T. In some embodiments, Xaa2 is selected from an amino acid of high amino acid charge. In some embodiments, Xaa2 is selected from R, K, H. In some embodiments. Xaa3 is selected from an amino acid of high amino acid solubility. In some embodiments, Xaa3 is selected from A, M, V, W, L, or I. In some embodiments, Xaa5 is selected from an amino acid of high amino acid solubility. In some embodiments, Xaa5 is selected from C, M, V, W, L, or I. In some embodiments, Xaa5 is selected from an amino acid of high hydropathy. In some embodiments, Xaa5 is selected from M, V, or I. In some embodiments, Xaa5 is selected from an amino acid of low average amino acid flexibility index. In some embodiments, Xaa5 is selected from M, W, F, or C. In some embodiments, Xaa8 is selected from an amino acid of high amino acid solubility. In some embodiments, Xaa8 is selected from H, V, or I.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 8118-SEQ ID NO: 9117, wherein said at least one mutation drives increased CNS tissue tropism.

C. Enriched CNS Sequences

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 9118-SEQ ID NO: 10117, wherein said at least one mutation drives increased CNS tissue tropism.

6.7.7. In Vivo Selected Mutated VP Polypeptides that Confer Increased Spleen Tropism

The present disclosure provides AAV5 virions with a VP capsid polypeptide having at least one mutation in a region with residues that interact with target cells (e.g., a target spleen cell in a target spleen tissue of interest), where the at least one mutation confers increased spleen tissue tropism as compared to a wildtype VP capsid polypeptide. In some embodiments, provided herein are AAV5 VP1 capsid polypeptide having a sequence homology of at least 80% to SEQ ID NO: 1, wherein the AAV5 VP1 capsid polypeptide has at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of SEQ ID NO: 1 and wherein said at least one mutation drives increased spleen tropism. The following sequences rules and sequences also apply to the region in AAV5 VP2 (amino acid residues 445 to 453; VP2 sequence shown in SEQ ID NO: 1115) and AAV5 VP3 (amino acid residues 389 to 397; VP3 sequences shown in SEQ ID NO: 1116) corresponding to AAV5 VP1 amino acid residues 581 to 589. Thus, the present disclosure encompasses AAV5 VP2 capsid polypeptides and AAV5 VP3 capsid polypeptides having one or more mutations in the VP2 and VP3 regions corresponding to the AAV5 VP1 amino acid residues of the 581 to 589 region, where the one or more mutations comport to the rules or sequences in the following section.

A. Positional Frequency Rules

Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased spleen tissue tropism as compared to wildtype AAV5 VP capsid polypeptide, wherein the engineered variant AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein said one or more mutations are selected from the following rules. In some embodiments, Xaa1 is selected from C, F, H, I, L, P, W, or Y. In some embodiments, Xaa1 is selected from C, F, P, W, or Y. In some embodiments, Xaa1 is selected from P, W, or Y. In some embodiments, Xaa1 is P. In some embodiments, Xaa2 is selected from D, E, L, N, P, R, or W. In some embodiments, Xaa2 is selected from D, E, or W. In some embodiments, Xaa2 is D. In some embodiments, Xaa3 is selected from C, D, E, P, or W. In some embodiments, Xaa3 is selected from D, P, or W. In some embodiments, Xaa3 is P. In some embodiments, Xaa4 is selected from C, F, G, H, R, W or Y. In some embodiments, Xaa4 is selected from C, H, or W. In some embodiments. Xaa4 is C. In some embodiments. Xaa5 is selected from A, D, E, G, P, R, or W. In some embodiments, Xaa5 is selected from D, E, G, or P. In some embodiments, Xaa5 is D. In some embodiments, Xaa6 is selected from A, C, D, E, K, R, W. In some embodiments, Xaa6 is selected from C, K, or R. In some embodiments, Xaa6 is K. In some embodiments, Xaa7 is selected from F, L, P, R, W, Y. In some embodiments, Xaa7 is selected from L, P, or W. In some embodiments, Xaa7 is P. In some embodiments, Xaa8 is selected from E, I, K, L, P, R, or T. In some embodiments, Xaa8 is selected from P, R, or K. In some embodiments, Xaa8 is K. In some embodiments, Xaa9 is selected from C, H, M, T, V, or W. In some embodiments, Xaa9 is selected from C, T, or V. In some embodiments, Xaa9 is V.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 37438-SEQ ID NO: 38437, wherein said at least one mutation drives increased spleen tissue tropism.

B. ML Rules

For the following set of rules described in this paragraph, favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an engineered AAV5 VP1 capsid polypeptide, were determined using in vivo data and two ML models, which are described in EXAMPLE 42. Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased spleen tissue tropism as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide, wherein the engineered AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the engineered AAV5 VP1 capsid polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein Xaa1 is selected from an amino acid of low solubility (e.g., Xaa1 is selected from D or P); or wherein Xaa1 is selected from an amino acid of high solubility (e.g., Xaa1 is selected from F, I, L); or wherein Xaa1 is selected from an amino acid of low hydropathy (e.g., Xaa1 is selected from Y or P); or wherein Xaa1 is selected from an amino acid of low mutability (e.g., Xaa1 is selected from C, K, or P); or wherein Xaa2 is selected from an amino acid of low solubility (e.g., Xaa2 is selected from D, Q, or R); or wherein Xaa2 is selected from an amino acid of low hydropathy (e.g., Xaa2 is selected from D, E, R, K, H, N, or Q); or wherein Xaa2 is selected from an amino acid of low charge (e.g., Xaa2 is selected from D or E); or wherein Xaa2 is selected from an amino acid of low volume (e.g., Xaa2 is selected from T, N, P, or D); or wherein Xaa2 is selected from an amino acid of high average flexibility (e.g., Xaa2 is selected from D, E, R, P, G, Q, or S); or wherein Xaa3 is selected from an amino acid of low solubility (e.g., Xaa3 is selected from D, E, P, or N); or wherein Xaa3 is selected from an amino acid of low hydropathy (e.g., Xaa3 is selected from D, E, H, N, Q, or P); or wherein Xaa4 is selected from an amino acid of low hydropathy (e.g., Xaa4 is selected from K or R); or wherein Xaa5 is selected from an amino acid of low solubility (e.g., Xaa5 is selected from D, E, P, or N); or wherein Xaa5 is selected from an amino acid of high average flexibility (e.g., Xaa5 is selected from D, E, R, P, G, Q, or S); or wherein Xaa6 is selected from an amino acid of low mutability (e.g., Xaa6 is selected from C); or wherein Xaa8 is selected from an amino acid of high surface accessibility (e.g., Xaa8 is selected from E, R, or K); or wherein Xaa8 is selected from an amino acid of low solubility (e.g., Xaa8 is selected from E, P, R, K, N, or Q); or wherein Xaa8 is selected from an amino acid of medium volume (e.g., Xaa8 is selected from E, D, R, K, V, P, M, I, L, H, N, Q, or T); or wherein Xaa9 is selected from an amino acid of medium mol mass (e.g., Xaa9 is selected from E, D, K, M, I, L, H, or N); or any combination thereof.

In some embodiments, Xaa1 is selected from an amino acid of low solubility. In some embodiments, Xaa1 is selected from D or P. In some embodiments. Xaa1 is selected from an amino acid of high solubility. In some embodiments, Xaa1 is selected from F, I, or L. In some embodiments, Xaa1 is selected from an amino acid of low hydropathy. In some embodiments, Xaa1 is selected from Y or P. In some embodiments, Xaa1 is selected from an amino acid of low mutability. In some embodiments. Xaa1 is selected from C, K, or P. In some embodiments, Xaa2 is selected from an amino acid of low solubility. In some embodiments, Xaa2 is selected from D, Q, or R. In some embodiments, Xaa2 is selected from an amino acid of low hydropathy. In some embodiments, Xaa2 is selected from D, E, R, K, H, N, or Q. In some embodiments, Xaa2 is selected from an amino acid of low charge. In some embodiments, Xaa2 is selected from D or E. In some embodiments, Xaa2 is selected from an amino acid of low volume. In some embodiments, Xaa2 is selected from T, N, P, or D. In some embodiments, Xaa2 is selected from an amino acid of high average flexibility. In some embodiments, Xaa2 is selected from D, E, R, P, G, Q, or S. In some embodiments. Xaa3 is selected from an amino acid of low solubility. In some embodiments, Xaa3 is selected from D, E, P, or N. In some embodiments, Xaa3 is selected from an amino acid of low hydropathy. In some embodiments, Xaa3 is selected from D, E, H, N, Q, or P. In some embodiments, Xaa4 is selected from an amino acid of low hydropathy. In some embodiments, Xaa4 is selected from K or R. In some embodiments, Xaa5 is selected from an amino acid of low solubility. In some embodiments, Xaa5 is selected from D, E, P, or N. In some embodiments. Xaa5 is selected from an amino acid of high average flexibility. In some embodiments. Xaa5 is selected from D, E, R, P, G, Q, or S. In some embodiments, Xaa6 is selected from an amino acid of low mutability. In some embodiments, Xaa6 is selected from C. In some embodiments, Xaa8 is selected from an amino acid of high surface accessibility. In some embodiments, Xaa8 is selected from E, R, or K. In some embodiments, Xaa8 is selected from an amino acid of low solubility. In some embodiments, Xaa8 is selected from E, P, R, K, N, or Q. In some embodiments, Xaa8 is selected from an amino acid of medium volume. In some embodiments, Xaa8 is selected from E, D, R, K, V, P, M, I, L, H, N, Q, or T. In some embodiments, Xaa9 is selected from an amino acid of medium mol mass. In some embodiments, Xaa9 is selected from E, D, K, M, I, L, H, or N.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 38438-SEQ ID NO: 39437, wherein said at least one mutation drives increased spleen tissue tropism.

C. Enriched Spleen Sequences

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 39438-SEQ ID NO: 40437, wherein said at least one mutation drives increased spleen tissue tropism.

6.7.8. In Vivo Selected Mutated VP Polypeptides that Confer Increased Adrenal Gland Tropism

The present disclosure provides AAV5 virions with a VP capsid polypeptide having at least one mutation in a region with residues that interact with target cells (e.g., a target adrenal gland cell in a target adrenal gland tissue of interest), where the at least one mutation confers increased adrenal gland tissue tropism as compared to a wildtype VP capsid polypeptide. In some embodiments, provided herein are AAV5 VP1 capsid polypeptide having a sequence homology of at least 80% to SEQ ID NO: 1, wherein the AAV5 VP1 capsid polypeptide has at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of SEQ ID NO: 1 and wherein said at least one mutation drives increased adrenal gland tropism. The following sequences rules and sequences also apply to the region in AAV5 VP2 (amino acid residues 445 to 453; VP2 sequence shown in SEQ ID NO: 1115) and AAV5 VP3 (amino acid residues 389 to 397; VP3 sequences shown in SEQ ID NO: 1116) corresponding to AAV5 VP1 amino acid residues 581 to 589. Thus, the present disclosure encompasses AAV5 VP2 capsid polypeptides and AAV5 VP3 capsid polypeptides having one or more mutations in the VP2 and VP3 regions corresponding to the AAV5 VP1 amino acid residues of the 581 to 589 region, where the one or more mutations comport to the rules or sequences in the following section.

A. Positional Frequency Rules

Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased adrenal gland tissue tropism as compared to wildtype AAV5 VP capsid polypeptide, wherein the engineered variant AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein said one or more mutations are selected from the following rules. In some embodiments, Xaa1 is selected from A, C, K, Q, R, S, or T. In some embodiments, Xaa1 is selected from C, K, or R. In some embodiments, Xaa1 is C. In some embodiments, Xaa2 is selected from A, C, I, S, T, or V. In some embodiments, Xaa2 is selected from A, V, or T. In some embodiments, Xaa2 is V. In some embodiments, Xaa3 is selected from A, F, G, K, M, Q, R, T, or V. In some embodiments, Xaa3 is selected from A, G, or M. In some embodiments, Xaa3 is M. In some embodiments, Xaa4 is selected from A, K, M, Q, R, or V. In some embodiments, Xaa4 is selected from A, R, or K. In some embodiments, Xaa4 is K. In some embodiments, Xaa5 is selected from F, I, L, M, R, T, V, or Y. In some embodiments. Xaa5 is selected from R, V, or Y. In some embodiments, Xaa5 is V. In some embodiments, Xaa6 is selected from G, H, M, N, R, or S. In some embodiments, Xaa6 is selected from H or N. In some embodiments, Xaa6 is N. In some embodiments, Xaa7 is selected from A, H, K, Q, R, S or V. In some embodiments, Xaa7 is selected from H, Q, or V. In some embodiments, Xaa7 is H. In some embodiments, Xaa8 is selected from A, G, H, M, Q, or S. In some embodiments, Xaa8 is selected from A, G, M, or S. In some embodiments, Xaa8 is S. In some embodiments, Xaa9 is selected from A, E, N, P, R, S, or Y. In some embodiments, Xaa9 is selected from P or E. In some embodiments, Xaa9 is P.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected SEQ ID NO: 1118-SEQ ID NO: 2117, wherein said at least one mutation drives increased adrenal gland tissue tropism.

B. ML Rules

For the following set of rules described in this paragraph, favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an engineered AAV5 VP1 capsid polypeptide, were determined using in vivo data and two ML models, which are described in EXAMPLE 31. Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased adrenal gland tissue tropism as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide, wherein the engineered AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the engineered AAV5 VP1 capsid polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein Xaa1 is selected from an amino acid of low mol mass at Xaa1 (e.g., Xaa1 is selected from V, P, S, or C); or wherein Xaa1 is selected from an amino acid of low hydropathy (e.g., Xaa1 is selected from T, S, W, or Y); or wherein Xaa2 is selected from an amino acid of low hydropathy (e.g., Xaa2 is selected from R); or wherein Xaa2 is selected from an amino acid of low mutability (e.g., Xaa2 is selected from C); or wherein Xaa2 is selected from an amino acid of low solubility (e.g., Xaa2 is selected from K); or wherein Xaa3 is selected from an amino acid of low average flexibility (e.g., Xaa3 is selected from W, M, or F); or wherein Xaa3 is selected from an amino acid of high solubility (e.g., Xaa3 is selected from M); or wherein Xaa4 is selected from an amino acid of high surface accessibility (e.g., Xaa4 is selected from K or R); or wherein Xaa4 is selected from an amino acid of high average flexibility (e.g., Xaa4 is selected from K, I, or N); or wherein Xaa5 is selected from an amino acid of medium mutability (e.g., Xaa5 is selected from R or H); or wherein Xaa5 is selected from an amino acid of high goldman engelman steitz (e.g., Xaa5 is selected from V or L); or wherein Xaa5 is selected from an amino acid of low hydropathy (e.g., Xaa5 is selected from R); or wherein Xaa5 is selected from an amino acid of high volume (e.g., Xaa5 is selected from Y, R, or F); or wherein Xaa6 is selected from an amino acid of high solubility (e.g., Xaa6 is selected from Y, V, M, A, or C); or wherein Xaa7 is selected from an amino acid of medium mutability (e.g., Xaa7 is selected from V, H, or R); or wherein Xaa7 is selected from an amino acid of low solubility (e.g., Xaa7 is selected from R); or wherein Xaa8 is selected from an amino acid of high average flexibility (e.g., Xaa8 is selected from K, I, or N); or wherein Xaa8 is selected from an amino acid of high mol mass (e.g., Xaa8 is selected from R or Y); or wherein Xaa9 is selected from an amino acid of high mutability (e.g., Xaa9 is selected from N); or any combination thereof.

In some embodiments, Xaa1 is selected from an amino acid of low mol mass. In some embodiments, Xaa1 is selected from V, P, S, or C. In some embodiments, Xaa1 is selected from an amino acid of low hydropathy. In some embodiments, Xaa1 is selected from T, S, W, or Y. In some embodiments, Xaa2 is selected from an amino acid of low hydropathy. In some embodiments, Xaa2 is selected from R. In some embodiments. Xaa2 is selected from an amino acid of low mutability. In some embodiments, Xaa2 is selected from C. In some embodiments. Xaa2 is selected from an amino acid of low solubility. In some embodiments, Xaa2 is selected from K. In some embodiments, Xaa3 is selected from an amino acid of low average flexibility. In some embodiments, Xaa3 is selected from W, M, or F. In some embodiments, Xaa3 is selected from an amino acid of high solubility. In some embodiments, Xaa3 is selected from M. In some embodiments, Xaa4 is selected from an amino acid of high surface accessibility. In some embodiments, Xaa4 is selected from K or R. In some embodiments, Xaa4 is selected from an amino acid of high average flexibility. In some embodiments. Xaa4 is selected from K, I, or N. In some embodiments, Xaa5 is selected from an amino acid of medium mutability. In some embodiments, Xaa5 is selected from R, H. In some embodiments, Xaa5 is selected from an amino acid of high goldman engelman steitz. In some embodiments, Xaa5 is selected from V, L. In some embodiments, Xaa5 is selected from an amino acid of low hydropathy. In some embodiments, Xaa5 is selected from R. In some embodiments, Xaa5 is selected from an amino acid of high volume. In some embodiments, Xaa5 is selected from Y, R, or F. In some embodiments, Xaa6 is selected from an amino acid of high solubility. In some embodiments, Xaa6 is selected from Y, V, M, A, or C. In some embodiments, Xaa7 is selected from an amino acid of medium mutability. In some embodiments, Xaa7 is selected from V, H, or R. In some embodiments, Xaa7 is selected from an amino acid of low solubility. In some embodiments, Xaa7 is selected from R. In some embodiments, Xaa8 is selected from an amino acid of high average flexibility. In some embodiments, Xaa8 is selected from K, I, or N. In some embodiments, Xaa8 is selected from an amino acid of high mol mass. In some embodiments, Xaa8 is selected from R or Y. In some embodiments, Xaa9 is selected from an amino acid of high mutability. In some embodiments, Xaa9 is selected from N.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 2118-SEQ ID NO: 3117, wherein said at least one mutation drives increased adrenal gland tissue tropism.

C. Enriched Adrenal Gland Sequences

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 3118-SEQ ID NO: 4117, wherein said at least one mutation drives increased adrenal gland tissue tropism.

6.7.9. In Vivo Selected Mutated VP Polypeptides that Confer Increased Sciatic Nerve Tropism

The present disclosure provides AAV5 virions with a VP capsid polypeptide having at least one mutation in a region with residues that interact with target cells (e.g., a target sciatic nerve cell in a target sciatic nerve tissue of interest), where the at least one mutation confers increased sciatic nerve tissue tropism as compared to a wildtype VP capsid polypeptide. In some embodiments, provided herein are AAV5 VP1 capsid polypeptide having a sequence homology of at least 80% to SEQ ID NO: 1, wherein the AAV5 VP1 capsid polypeptide has at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of SEQ ID NO: 1 and wherein said at least one mutation drives increased sciatic nerve tropism. The following sequences rules and sequences also apply to the region in AAV5 VP2 (amino acid residues 445 to 453; VP2 sequence shown in SEQ ID NO: 1115) and AAV5 VP3 (amino acid residues 389 to 397; VP3 sequences shown in SEQ ID NO: 1116) corresponding to AAV5 VP1 amino acid residues 581 to 589. Thus, the present disclosure encompasses AAV5 VP2 capsid polypeptides and AAV5 VP3 capsid polypeptides having one or more mutations in the VP2 and VP3 regions corresponding to the AAV5 VP1 amino acid residues of the 581 to 589 region, where the one or more mutations comport to the rules or sequences in the following section.

A. Positional Frequency Rules

Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased sciatic nerve tissue tropism as compared to wildtype AAV5 VP capsid polypeptide, wherein the engineered variant AAV5 VP1 capsid polypeptide sequence has one or more mutations, wherein Xaa1 is selected from C, G, K, M, Q, R, or Y, or Xaa1 is selected from C, R, or Q, or Xaa1 is C; or Xaa2 is selected from A, C, F, I, Q, T, or V, or Xaa2 is selected from A, C, or I, or Xaa2 is A; or Xaa3 is selected from A, F, I, M, R, S, or T, or Xaa3 is selected from F, M, R, or S, or Xaa3 is R; or Xaa4 is selected from E, N, T, Q, or V, or Xaa4 is selected from E, T, or V, or Xaa4 is T; or Xaa5 is selected from F, H, Q, S, V, or Y, or Xaa5 is selected from F, V, or Y, or Xaa5 is V; or Xaa6 is selected from K, M, N, Q, S, or V, or Xaa6 is selected from M, N, or S, or Xaa6 is N; or Xaa7 is selected from K, M, Q, R, or T, or Xaa7 is selected from M, Q, or T, or Xaa7 is M; or Xaa8 is selected from A, G, H, Q, S, or V, or Xaa8 is selected from H or S, or Xaa8 is H; or Xaa9 is selected from C, E, I, K, or R, or Xaa9 is selected from C, I, or K, or Xaa9 is I.

herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased sciatic nerve tissue tropism as compared to wildtype AAV5 VP capsid polypeptide, wherein the engineered variant AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein said one or more mutations are selected from the following rules. In some embodiments, Xaa1 is selected from C, G, K, M, Q, R, or Y. In some embodiments, Xaa1 is selected from C, R, or Q. In some embodiments, Xaa1 is C. In some embodiments, Xaa2 is selected from A, C, F, I, Q, T, or V. In some embodiments, Xaa2 is selected from A, C, or I. In some embodiments, Xaa2 is A. In some embodiments, Xaa3 is selected from A, F, I, M, R, S, or T. In some embodiments, Xaa3 is selected from F, M, R, or S. In some embodiments, Xaa3 is R. In some embodiments, Xaa4 is selected from E, N, T, Q, or V. In some embodiments, Xaa4 is selected from E, T, or V. In some embodiments, Xaa4 is T. In some embodiments, Xaa5 is selected from F, H, Q, S. V, or Y. In some embodiments, Xaa5 is selected from F, V, or Y. In some embodiments, Xaa5 is V. In some embodiments, Xaa6 is selected from K, M, N, Q, S, or V. In some embodiments, Xaa6 is selected from M, N, or S. In some embodiments, Xaa6 is N. In some embodiments, Xaa7 is selected from K, M, Q, R, or T. In some embodiments, Xaa7 is selected from M, Q, or T. In some embodiments, Xaa7 is M. In some embodiments, Xaa8 is selected from A, G, H, Q, S, or V. In some embodiments, Xaa8 is selected from H or S. In some embodiments, Xaa8 is H. In some embodiments, Xaa9 is selected from C, E, I, K, or R. In some embodiments, Xaa9 is selected from C, I, or K. In some embodiments, Xaa9 is I.

Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased sciatic nerve tissue tropism as compared to wildtype AAV5 VP capsid polypeptide, wherein the engineered variant AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein said one or more mutations are selected from the following rules: Xaa1 is selected from C, G, K, M, Q, R, or Y, Xaa2 is selected from A, C, F, I, Q, T, or V, Xaa3 is selected from A, F, I, M, R, S, or T, Xaa4 is selected from E, N, T, Q, or V, Xaa5 is selected from F, H, Q, S, V, or Y, Xaa6 is selected from K, M, N, Q, S, or V, Xaa7 is selected from K, M, Q, R, or T, Xaa8 is selected from A, G, H, Q, S, or V, and Xaa9 is selected from C, E, I, K, or R.

Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased sciatic nerve tissue tropism as compared to wildtype AAV5 VP capsid polypeptide, wherein the engineered variant AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein said one or more mutations are as follows: Xaa1 is selected from C, G, K, M, Q, R, or Y, Xaa2 is selected from A, C, F, I, Q, T, or V, Xaa3 is selected from A, F, I, M, R, S, or T, Xaa4 is selected from E, N, T, Q, or V, Xaa5 is selected from F, H, Q, S, V, or Y, Xaa6 is selected from K, M, N, Q, S, or V, Xaa7 is selected from K, M, Q, R, or T, Xaa8 is selected from A, G, H, Q, S, or V, Xaa9 is selected from C, E, I, K, or R, or any combination thereof.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 26118-SEQ ID NO: 26990, wherein said at least one mutation drives increased sciatic nerve tissue tropism.

B. ML Rules

For the following set of rules described in this paragraph, favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an engineered AAV5 VP1 capsid polypeptide, were determined using in vivo data and two ML models, which are described in EXAMPLE 38. Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased sciatic nerve tissue tropism as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide, wherein the engineered AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the engineered AAV5 VP1 capsid polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein Xaa1 is selected from an amino acid of high average flexibility (e.g., Xaa1 is selected from G or R); or wherein Xaa1 is selected from an amino acid of low solubility (e.g., Xaa1 is selected from R, or Q); or wherein Xaa1 is selected from an amino acid of low mutability (e.g., Xaa1 is selected from C, L, F, Y, R, K, P, or H); or wherein Xaa1 is selected from an amino acid of high volume (e.g., Xaa1 is selected from Y, F); or wherein Xaa2 is selected from an amino acid of high surface accessibility (e.g., Xaa2 is selected from E, R, or K); or wherein Xaa3 is selected from an amino acid of medium mutability (e.g., Xaa3 is selected from H or R); or wherein Xaa3 is selected from an amino acid of medium average flexibility (e.g., Xaa3 is selected from V or Y); or wherein Xaa4 is selected from an amino acid of high mutability (e.g., Xaa4 is selected from N); or wherein Xaa4 is selected from an amino acid of high average flexibility (e.g., Xaa4 is selected from I, N, G, or R); or wherein Xaa4 is selected from an amino acid of low solubility (e.g., Xaa4 is selected from N); or wherein Xaa6 is selected from an amino acid of low mutability (e.g., Xaa6 is selected from C, L, F, or Y); or wherein Xaa6 is selected from an amino acid of high volume (e.g., Xaa6 is selected from K, M, I, or L); or wherein Xaa7 is selected from an amino acid of low mutability (e.g., Xaa7 is selected from L, F, or Y); or wherein Xaa7 is selected from an amino acid of medium mol mass (e.g., Xaa7 is selected from D, I, L, or N); or wherein Xaa8 is selected from an amino acid of high surface accessibility (e.g., Xaa8 is selected from S, Y, T, D, P, H, or N); or wherein Xaa9 is selected from an amino acid of low mutability (e.g., Xaa9 is selected from C, H, R); or wherein Xaa9 is selected from an amino acid of medium solubility (e.g., Xaa9 is selected from Q, T, or C); or wherein Xaa9 is selected from an amino acid of low surface accessibility (e.g., Xaa9 is selected from C); or any combination thereof.

In some embodiments, Xaa1 is selected from an amino acid of high average flexibility. In some embodiments, Xaa1 is selected from G or R. In some embodiments, Xaa1 is selected from an amino acid of low solubility. In some embodiments, Xaa1 is selected from R or Q. In some embodiments, Xaa1 is selected from an amino acid of low mutability. In some embodiments, Xaa1 is selected from C, L, F, Y, R, K, P, or H. In some embodiments, Xaa1 is selected from an amino acid of high volume. In some embodiments, Xaa1 is selected from Y or F. In some embodiments, Xaa2 is selected from an amino acid of high surface accessibility. In some embodiments, Xaa2 is selected from E, R, or K. In some embodiments, Xaa3 is selected from an amino acid of medium mutability. In some embodiments, Xaa3 is selected from H or R. In some embodiments, Xaa3 is selected from an amino acid of medium average flexibility. In some embodiments, Xaa3 is selected from V or Y. In some embodiments, Xaa4 is selected from an amino acid of high mutability. In some embodiments, Xaa4 is selected from N. In some embodiments, Xaa4 is selected from an amino acid of high average flexibility. In some embodiments, Xaa4 is selected from I, N, G, or R. In some embodiments. Xaa4 is selected from an amino acid of low solubility. In some embodiments, Xaa4 is selected from N. In some embodiments, Xaa6 is selected from an amino acid of low mutability. In some embodiments, Xaa6 is selected from C, L, F, or Y. In some embodiments, Xaa6 is selected from an amino acid of high volume. In some embodiments, Xaa6 is selected from K, M, I, or L. In some embodiments, Xaa7 is selected from an amino acid of low mutability. In some embodiments, Xaa7 is selected from L, F, or Y. In some embodiments, Xaa7 is selected from an amino acid of medium mol mass. In some embodiments, Xaa7 is selected from D, I, L, or N. In some embodiments, Xaa8 is selected from an amino acid of high surface accessibility. In some embodiments, Xaa8 is selected from S, Y, T, D, P, H, or N. In some embodiments, Xaa9 is selected from an amino acid of low mutability. In some embodiments, Xaa9 is selected from C, H, or R In some embodiments, Xaa9 is selected from an amino acid of medium solubility. In some embodiments, Xaa9 is selected from Q, T, or C. In some embodiments, Xaa9 is selected from an amino acid of low surface accessibility. In some embodiments, Xaa9 is selected from C.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 26991-SEQ ID NO: 27990, wherein said at least one mutation drives increased sciatic nerve tissue tropism.

C. Enriched Sciatic Nerve Sequences

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 27991-SEQ ID NO: 28990, wherein said at least one mutation drives increased sciatic nerve tissue tropism.

6.7.10. In Vivo Selected Mutated VP Polypeptides that Confer Increased Skeletal Muscle Tropism

The present disclosure provides AAV5 virions with a VP capsid polypeptide having at least one mutation in a region with residues that interact with target cells (e.g., a target skeletal muscle cell in a target skeletal muscle tissue of interest), where the at least one mutation confers increased skeletal muscle tissue tropism as compared to a wildtype VP capsid polypeptide. In some embodiments, provided herein are AAV5 VP1 capsid polypeptide having a sequence homology of at least 80% to SEQ ID NO: 1, wherein the AAV5 VP1 capsid polypeptide has at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of SEQ ID NO: 1 and wherein said at least one mutation drives increased skeletal muscle tropism. The following sequences rules and sequences also apply to the region in AAV5 VP2 (amino acid residues 445 to 453; VP2 sequence shown in SEQ ID NO: 1115) and AAV5 VP3 (amino acid residues 389 to 397; VP3 sequences shown in SEQ ID NO: 1116) corresponding to AAV5 VP1 amino acid residues 581 to 589. Thus, the present disclosure encompasses AAV5 VP2 capsid polypeptides and AAV5 VP3 capsid polypeptides having one or more mutations in the VP2 and VP3 regions corresponding to the AAV5 VP1 amino acid residues of the 581 to 589 region, where the one or more mutations comport to the rules or sequences in the following section.

A. Positional Frequency Rules

Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased skeletal muscle tissue tropism as compared to wildtype AAV5 VP capsid polypeptide, wherein the engineered variant AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein said one or more mutations are selected from the following rules. In some embodiments, Xaa1 is selected from A, E, H, M, P, Q, or S. In some embodiments, Xaa1 is selected from P or Q. In some embodiments, Xaa1 is Q. In some embodiments, Xaa2 is selected from F, H, I, T, or V. In some embodiments, Xaa2 is selected from T or V. In some embodiments, Xaa2 is V. In some embodiments. Xaa3 is selected from A, G, I, K, M, Q, R, S, T, or V. In some embodiments, Xaa3 is selected from A, L, P, R, or T. In some embodiments, Xaa3 is selected from L, P, or T. In some embodiments, Xaa3 is P. In some embodiments, Xaa4 is selected from D, E, G, P, or S. In some embodiments, Xaa4 is selected from D, E, or S. In some embodiments, Xaa4 is E. In some embodiments, Xaa5 is selected from H, L, M, P, or V. In some embodiments, Xaa5 is selected from L, M, or V. In some embodiments, Xaa5 is L. In some embodiments, Xaa6 is selected from E, H, N, or P. In some embodiments, Xaa6 is P. In some embodiments, Xaa7 is selected from A, H, N, Q or T. In some embodiments, Xaa7 is H. In some embodiments, Xaa8 is selected from I, K, M, P, or W. In some embodiments, Xaa8 is selected from I, P, or W. In some embodiments. Xaa8 is P. In some embodiments, Xaa9 is selected from A, I, M, P, or V. In some embodiments, Xaa9 is selected from A. M, or P. In some embodiments, Xaa9 is M.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 28991-SEQ ID NO: 29990, wherein said at least one mutation drives increased skeletal muscle tissue tropism.

B. ML Rules

For the following set of rules described in this paragraph, favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an engineered AAV5 VP1 capsid polypeptide, were determined using in vivo data and two ML models, which are described in EXAMPLE 39. Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased skeletal muscle tissue tropism as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide, wherein the engineered AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the engineered AAV5 VP1 capsid polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein Xaa1 is selected from an amino acid of high average flexibility (e.g., Xaa1 is selected from G or R); or wherein Xaa1 is selected from an amino acid of low average flexibility (e.g., Xaa1 is selected from W, M, F, or H); or wherein Xaa1 is selected from an amino acid of high mol mass (e.g., Xaa1 is selected from R, F, or W); or wherein Xaa2 is selected from an amino acid of low hydropathy (e.g., Xaa2 is selected from K, or R); or wherein Xaa2 is selected from an amino acid of low mutability (e.g., Xaa2 is selected from C, R, or H); or wherein Xaa2 is selected from an amino acid of high average flexibility (e.g., Xaa2 is selected from G or R); or wherein Xaa3 is selected from an amino acid of high average flexibility (e.g., Xaa3 is selected from G or R); or wherein Xaa4 is selected from an amino acid of high hydrophilicity (e.g., Xaa4 is selected from D, E, R, K, or N); or wherein Xaa4 is selected from an amino acid of low mutability (e.g., Xaa4 is selected from C, R, H); or wherein Xaa5 is selected from an amino acid of low mol mass (e.g., Xaa5 is selected from A); or wherein Xaa5 is selected from an amino acid of low average flexibility (e.g., Xaa5 is selected from A or L); or wherein Xaa5 is selected from an amino acid of high mutability (e.g., Xaa5 is selected from D, A, or E); or wherein Xaa6 is selected from an amino acid of low average flexibility (e.g., Xaa6 is selected from W, M, or F); or wherein Xaa6 is selected from an amino acid of low mutability (e.g., Xaa6 is selected from C); or wherein Xaa6 is selected from an amino acid of high mol mass (e.g., Xaa6 is selected from W); or wherein Xaa7 is selected from an amino acid of low goldman engelman steitz (e.g., Xaa7 is selected from R); or wherein Xaa7 is selected from an amino acid of high average flexibility (e.g., Xaa7 is selected from D, R, P, G, or S); or wherein Xaa7 is selected from an amino acid of high mutability (e.g., Xaa7 is selected from R, H, or N); or wherein Xaa7 is selected from an amino acid of low solubility (e.g., Xaa7 is selected from R or Q); or wherein Xaa8 is selected from an amino acid of high hydrophilicity (e.g., Xaa8 is selected from D, E, R, K, or N); or wherein Xaa9 is selected from an amino acid of low mutability (e.g., Xaa9 is selected from Y, F, or L); or any combination thereof.

In some embodiments, Xaa1 is selected from an amino acid of high average flexibility. In some embodiments, Xaa1 is selected from G or R. In some embodiments, Xaa1 is selected from an amino acid of low average flexibility. In some embodiments, Xaa1 is selected from W, M, F, or H. In some embodiments, Xaa1 is selected from an amino acid of high mol mass. In some embodiments, Xaa1 is selected from R, F, or W. In some embodiments, Xaa2 is selected from an amino acid of low hydropathy. In some embodiments, Xaa2 is selected from K or R. In some embodiments, Xaa2 is selected from an amino acid of low mutability. In some embodiments, Xaa2 is selected from C, R, or H. In some embodiments, Xaa2 is selected from an amino acid of high average flexibility. In some embodiments, Xaa2 is selected from G or R. In some embodiments, Xaa3 is selected from an amino acid of high average flexibility. In some embodiments, Xaa3 is selected from G or R. In some embodiments, Xaa4 is selected from an amino acid of high hydrophilicity. In some embodiments, Xaa4 is selected from D, E, R, K, or N. In some embodiments, Xaa4 is selected from an amino acid of low mutability. In some embodiments, Xaa4 is selected from C, R, or H. In some embodiments, Xaa5 is selected from an amino acid of low mol mass. In some embodiments, Xaa5 is selected from A. In some embodiments, Xaa5 is selected from an amino acid of low average flexibility. In some embodiments, Xaa5 is selected from A or L. In some embodiments, Xaa5 is selected from an amino acid of high mutability. In some embodiments, Xaa5 is selected from D, A, or E. In some embodiments, Xaa6 is selected from an amino acid of low average flexibility. In some embodiments, Xaa6 is selected from W, M, or F. In some embodiments, Xaa6 is selected from an amino acid of low mutability. In some embodiments, Xaa6 is selected from C. In some embodiments, Xaa6 is selected from an amino acid of high mol mass. In some embodiments, Xaa6 is selected from W. In some embodiments, Xaa7 is selected from an amino acid of low goldman engelman steitz. In some embodiments, Xaa7 is selected from R. In some embodiments, Xaa7 is selected from an amino acid of high average flexibility. In some embodiments, Xaa7 is selected from D, R, P, G, or S. In some embodiments, Xaa7 is selected from an amino acid of high mutability. In some embodiments, Xaa7 is selected from R, H, or N. In some embodiments, Xaa7 is selected from an amino acid of low solubility. In some embodiments, Xaa7 is selected from R or Q. In some embodiments, Xaa8 is selected from an amino acid of high hydrophilicity. In some embodiments, Xaa8 is selected from D, E, R, K, or N. In some embodiments, Xaa9 is selected from an amino acid of low mutability. In some embodiments, Xaa9 is selected from Y, F, or L.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 29991-SEQ ID NO: 30990, wherein said at least one mutation drives increased skeletal muscle tissue tropism.

C. Enriched Skeletal Muscle Sequences

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 30991-SEQ ID NO: 31990, wherein said at least one mutation drives increased skeletal muscle tissue tropism.

6.7.11. In Vivo Selected Mutated VP Polypeptides that Confer Increased Spinal Cord Tropism

The present disclosure provides AAV5 virions with a VP capsid polypeptide having at least one mutation in a region with residues that interact with target cells (e.g., a target spinal cord cell in a target spinal cord tissue of interest), where the at least one mutation confers increased CNS tissue tropism as compared to a wildtype VP capsid polypeptide. In some embodiments, provided herein are AAV5 VP1 capsid polypeptide having a sequence homology of at least 80% to SEQ ID NO: 1, wherein the AAV5 VP1 capsid polypeptide has at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of SEQ ID NO: 1 and wherein said at least one mutation drives increased spinal cord tropism. The following sequences rules and sequences also apply to the region in AAV5 VP2 (amino acid residues 445 to 453; VP2 sequence shown in SEQ ID NO: 1115) and AAV5 VP3 (amino acid residues 389 to 397; VP3 sequences shown in SEQ ID NO: 1116) corresponding to AAV5 VP1 amino acid residues 581 to 589. Thus, the present disclosure encompasses AAV5 VP2 capsid polypeptides and AAV5 VP3 capsid polypeptides having one or more mutations in the VP2 and VP3 regions corresponding to the AAV5 VP1 amino acid residues of the 581 to 589 region, where the one or more mutations comport to the rules or sequences in the following section.

A. Positional Frequency Rules

Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased spinal cord tissue tropism as compared to wildtype AAV5 VP capsid polypeptide, wherein the engineered variant AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein said one or more mutations are selected from the following rules. In some embodiments, Xaa1 is selected from A, C, K, Q, R, S, or W. In some embodiments, Xaa1 is selected from K, R, or W. In some embodiments, Xaa1 is K. In some embodiments, Xaa2 is selected from H, I, K, L, T, V, or W. In some embodiments, Xaa2 is selected from H, I, or T. In some embodiments, Xaa2 is I. In some embodiments, Xaa3 is selected from C, F, G, H, I, K, N, or R. In some embodiments, Xaa3 is selected from F, I, or R. In some embodiments, Xaa3 is I. In some embodiments, Xaa4 is selected from 1, M, Q, S, or V. In some embodiments, Xaa4 is selected from I, M, or V. In some embodiments, Xaa4 is V. In some embodiments, Xaa5 is selected from H, K, Q, T, W, or Y. In some embodiments, Xaa5 is selected from T, W, or Y. In some embodiments, Xaa5 is Y. In some embodiments, Xaa6 is selected from H, L, N, Q, R, W, or Y. In some embodiments, Xaa6 is selected from L, N, R, or Y. In some embodiments, Xaa6 is Y. In some embodiments, Xaa7 is selected from D, H, P, Q, or R. In some embodiments, Xaa7 is R. In some embodiments, Xaa8 is selected from D, F, L, S, T, or Y. In some embodiments, Xaa8 is selected from S, T, or Y. In some embodiments, Xaa8 is T. In some embodiments, Xaa9 is selected from C, I, N, P, R, S, or Y. In some embodiments, Xaa9 is selected from I, P, or R. In some embodiments, Xaa9 is I.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 34991-SEQ ID NO: 35437, wherein said at least one mutation drives increased spinal cord tissue tropism.

B. ML Rules

For the following set of rules described in this paragraph, favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an engineered AAV5 VP1 capsid polypeptide, were determined using in vivo data and two ML models, which are described in EXAMPLE 41. Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased spinal cord tissue tropism as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide, wherein the engineered AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the engineered AAV5 VP1 capsid polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein Xaa1 is selected from an amino acid of high volume (e.g., Xaa1 is selected from F, W, or Y); or wherein Xaa1 is selected from an amino acid of low mutability (e.g., Xaa1 is selected from Y, F, L, or C); or wherein Xaa1 is selected from an amino acid of high solubility (e.g., Xaa1 is selected from W, F, I, or L); or wherein Xaa1 is selected from an amino acid of low average flexibility (e.g., Xaa1 is selected from F, M, or W); or wherein Xaa2 is selected from an amino acid of low hydropathy (e.g., Xaa2 is selected from P or Y); or wherein Xaa3 is selected from an amino acid of low hydrophilicity (e.g., Xaa3 is selected from Y, W, V, M, F, I, or L); or wherein Xaa3 is selected from an amino acid of high solubility (e.g., Xaa3 is selected from W, F, I, or L); or wherein Xaa6 is selected from an amino acid of high volume (e.g., Xaa6 is selected from W, R, K, M, I, or L); or wherein Xaa6 is selected from an amino acid of high mol mass (e.g., Xaa6 is selected from W); or wherein Xaa8 is selected from an amino acid of high mol mass (e.g., Xaa8 is selected from W, E, K, M, H, or Q); or wherein Xaa8 is selected from an amino acid of high volume (e.g., Xaa8 is selected from W, K, M, I, or L); or wherein Xaa8 is selected from an amino acid of high goldman engelman steitz. (e.g., Xaa8 is selected from V or L); or wherein Xaa9 is selected from an amino acid of high hydropathy (e.g., Xaa9 is selected from V, or I); or wherein Xaa9 is selected from an amino acid of high solubility (e.g., Xaa9 is selected from W, F, I, or L); or any combination thereof.

In some embodiments, Xaa1 is selected from an amino acid of high volume. In some embodiments, Xaa1 is selected from F, W, or Y. In some embodiments, Xaa1 is selected from an amino acid of low mutability. In some embodiments, Xaa1 is selected from Y, F, L, or C. In some embodiments, Xaa1 is selected from an amino acid of high solubility. In some embodiments, Xaa1 is selected from W, F, I, or L. In some embodiments, Xaa1 is selected from an amino acid of low average flexibility. In some embodiments, Xaa1 is selected from F, M, or W. In some embodiments, Xaa2 is selected from an amino acid of low hydropathy. In some embodiments, Xaa2 is selected from P or Y. In some embodiments, Xaa3 is selected from an amino acid of low hydrophilicity. In some embodiments, Xaa3 is selected from Y, W, V, M, F, I, or L. In some embodiments, Xaa3 is selected from an amino acid of high solubility. In some embodiments, Xaa3 is selected from W, F, I, L. In some embodiments, Xaa6 is selected from an amino acid of high volume. In some embodiments, Xaa6 is selected from W, R, K, M, I, or L. In some embodiments, Xaa6 is selected from an amino acid of high mol mass. In some embodiments, Xaa6 is selected from W. In some embodiments, Xaa8 is selected from an amino acid of high mol mass. In some embodiments, Xaa8 is selected from W, E, K, M, H, or Q. In some embodiments, Xaa8 is selected from an amino acid of high volume. In some embodiments, Xaa8 is selected from W, K, M, I, L. In some embodiments, Xaa8 is selected from an amino acid of high goldman engelman steitz. In some embodiments, Xaa8 is selected from V or L. In some embodiments, Xaa9 is selected from an amino acid of high hydropathy. In some embodiments, Xaa9 is selected from V or I. In some embodiments, Xaa9 is selected from an amino acid of high solubility. In some embodiments, Xaa9 is selected from W, F, I, or L.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 35438-SEQ ID NO: 36437, wherein said at least one mutation drives increased spinal cord tissue tropism.

C. Enriched Spinal Cord Sequences

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 36438-SEQ ID NO: 37437, wherein said at least one mutation drives increased spinal cord tissue tropism.

6.7.12. In Vivo Selected Mutated VP Polypeptides that Confer Increased Mammary Gland Tropism

The present disclosure provides AAV5 virions with a VP capsid polypeptide having at least one mutation in a region with residues that interact with target cells (e.g., a target mammary gland cell in a target mammary gland tissue of interest), where the at least one mutation confers increased mammary gland tissue tropism as compared to a wildtype VP capsid polypeptide. In some embodiments, provided herein are AAV5 VP1 capsid polypeptide having a sequence homology of at least 80% to SEQ ID NO: 1, wherein the AAV5 VP1 capsid polypeptide has at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of SEQ ID NO: 1 and wherein said at least one mutation drives increased mammary gland tropism. The following sequences rules and sequences also apply to the region in AAV5 VP2 (amino acid residues 445 to 453; VP2 sequence shown in SEQ ID NO: 1115) and AAV5 VP3 (amino acid residues 389 to 397; VP3 sequences shown in SEQ ID NO: 1116) corresponding to AAV5 VP1 amino acid residues 581 to 589. Thus, the present disclosure encompasses AAV5 VP2 capsid polypeptides and AAV5 VP3 capsid polypeptides having one or more mutations in the VP2 and VP3 regions corresponding to the AAV5 VP1 amino acid residues of the 581 to 589 region, where the one or more mutations comport to the rules or sequences in the following section.

A. Positional Frequency Rules

Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased mammary gland tissue tropism as compared to wildtype AAV5 VP capsid polypeptide, wherein the engineered variant AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein said one or more mutations are selected from the following rules. In some embodiments, Xaa1 is selected from C, K, M, Q, R, or Y. In some embodiments, Xaa1 is selected from C, Q, or R. In some embodiments, Xaa1 is C. In some embodiments, Xaa2 is selected from A, F, I, K, S, T, or V. In some embodiments, Xaa2 is selected from A, S, or V. In some embodiments, Xaa2 is V. In some embodiments, Xaa3 is selected from A, F, G, I, K, L, R, T, or Y. In some embodiments, Xaa3 is selected from F, G, K, R, or Y. In some embodiments, Xaa3 is selected from F, K, or Y. In some embodiments, Xaa3 is F. In some embodiments, Xaa4 is selected from A, I, K, Q, R, or T. In some embodiments, Xaa4 is selected from A, I, or R. In some embodiments, Xaa4 is I. In some embodiments, Xaa5 is selected from I, L, M, Q, R, T, V, or Y. In some embodiments, Xaa5 is selected from I, M, or Y. In some embodiments, Xaa5 is Y. In some embodiments, Xaa6 is selected from H, N, S, or V. In some embodiments, Xaa6 is H. In some embodiments, Xaa7 is selected from A, H, I, N, S or Y. In some embodiments, Xaa7 is N or S. In some embodiments, Xaa7 is N. In some embodiments, Xaa8 is selected from A, C, D, G, H, M, Q, or S. In some embodiments, Xaa8 is selected from G, M, or Q. In some embodiments, Xaa8 is G. In some embodiments, Xaa9 is selected from A, E, L, W, or Y. In some embodiments, Xaa9 is selected from A, L, or W. In some embodiments, Xaa9 is A.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 22118-SEQ ID NO: 23117, wherein said at least one mutation drives increased mammary gland tissue tropism.

B. ML Rules

For the following set of rules described in this paragraph, favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an engineered AAV5 VP1 capsid polypeptide, were determined using in vivo data and two ML models, which are described in EXAMPLE 36. Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased mammary gland tissue tropism as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide, wherein the engineered AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the engineered AAV5 VP1 capsid polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein Xaa1 is selected from an amino acid of low surface accessibility (e.g., Xaa1 is selected from C); or wherein Xaa1 is selected from an amino acid of medium mol mass (e.g., Xaa1 is selected from C); or wherein Xaa2 is selected from an amino acid of high surface accessibility (e.g., Xaa2 is selected from D, N, or Q); or wherein Xaa2 is selected from an amino acid of low hydropathy (e.g., Xaa2 is selected from D, E, R, K, H, N, or Q); or wherein Xaa3 is selected from an amino acid of high average flexibility (e.g., Xaa3 is selected from D, E, R, P, G, or S); or wherein Xaa3 is selected from an amino acid of medium mutability (e.g., Xaa3 is selected from R or H); or wherein Xaa4 is selected from an amino acid of high mutability (e.g., Xaa4 is selected from M, I, Q, or T); or wherein Xaa4 is selected from an amino acid of high solubility (e.g., Xaa4 is selected from W, F, I, or L); or wherein Xaa4 is selected from an amino acid of high surface accessibility (e.g., Xaa4 is selected from E, R, or K); or wherein Xaa5 is selected from an amino acid of high solubility (e.g., Xaa5 is selected from W, F, I, or L); or wherein Xaa5 is selected from an amino acid of low mutability (e.g., Xaa5 is selected from Y, F, or L); or wherein Xaa6 is selected from an amino acid of high hydropathy (e.g., Xaa6 is selected from V, I, or L); or wherein Xaa6 is selected from an amino acid of medium mol mass (e.g., Xaa6 is selected from D, I, L, or N); or wherein Xaa8 is selected from an amino acid of low surface accessibility (e.g., Xaa8 is selected from C); or wherein Xaa8 is selected from an amino acid of low mutability (e.g., Xaa8 is selected from C, R, or H); or wherein Xaa9 is selected from an amino acid of medium mutability (e.g., Xaa9 is selected from R or H); or any combination thereof.

[In some embodiments, Xaa1 is selected from an amino acid of low surface accessibility. In some embodiments, Xaa1 is selected from C. In some embodiments, Xaa1 is selected from an amino acid of medium mol mass. In some embodiments, Xaa1 is selected from C. In some embodiments, Xaa2 is selected from an amino acid of high surface accessibility. In some embodiments, Xaa2 is selected from D, N, or Q. In some embodiments, Xaa2 is selected from an amino acid of low hydropathy. In some embodiments, Xaa2 is selected from D, E, R, K, H, N, or Q. In some embodiments, Xaa3 is selected from an amino acid of high average flexibility. In some embodiments, Xaa3 is selected from D, E, R, P, G, or S. In some embodiments, Xaa3 is selected from an amino acid of medium mutability. In some embodiments, Xaa3 is selected from R or H. In some embodiments, Xaa4 is selected from an amino acid of high mutability. In some embodiments, Xaa4 is selected from M, I, Q, or T. In some embodiments, Xaa4 is selected from an amino acid of high solubility. In some embodiments, Xaa4 is selected from W, F, I, or L. In some embodiments, Xaa4 is selected from an amino acid of high surface accessibility. In some embodiments, Xaa4 is selected from E, R, or K. In some embodiments, Xaa5 is selected from an amino acid of high solubility. In some embodiments, Xaa5 is selected from W, F, I, or L. In some embodiments, Xaa5 is selected from an amino acid of low mutability. In some embodiments, Xaa5 is selected from Y, F, or L. In some embodiments, Xaa6 is selected from an amino acid of high hydropathy. In some embodiments, Xaa6 is selected from V, I, or L. In some embodiments, Xaa6 is selected from an amino acid of medium mol mass. In some embodiments, Xaa6 is selected from D, I, L, or N. In some embodiments, Xaa8 is selected from an amino acid of low surface accessibility. In some embodiments, Xaa8 is selected from C. In some embodiments, Xaa8 is selected from an amino acid of low mutability. In some embodiments, Xaa8 is selected from C, R, or H. In some embodiments, Xaa9 is selected from an amino acid of medium mutability. In some embodiments, Xaa9 is selected from R or H.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 23118-SEQ ID NO: 24117, wherein said at least one mutation drives increased mammary gland tissue tropism.

C. Enriched Mammary Gland Sequences

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 24118-SEQ ID NO: 25117, wherein said at least one mutation drives increased mammary gland tissue tropism.

6.7.13. In Vivo Selected Mutated VP Polypeptides that Confer Increased Lung Tropism

The present disclosure provides AAV5 virions with a VP capsid polypeptide having at least one mutation in a region with residues that interact with target cells (e.g., a target lung cell in a target lung tissue of interest), where the at least one mutation confers increased lung tissue tropism as compared to a wildtype VP capsid polypeptide. In some embodiments, provided herein are AAV5 VP1 capsid polypeptide having a sequence homology of at least 80% to SEQ ID NO: 1, wherein the AAV5 VP1 capsid polypeptide has at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of SEQ ID NO: 1 and wherein said at least one mutation drives increased lung tropism. The following sequences rules and sequences also apply to the region in AAV5 VP2 (amino acid residues 445 to 453; VP2 sequence shown in SEQ ID NO: 1115) and AAV5 VP3 (amino acid residues 389 to 397; VP3 sequences shown in SEQ ID NO: 1116) corresponding to AAV5 VP1 amino acid residues 581 to 589. Thus, the present disclosure encompasses AAV5 VP2 capsid polypeptides and AAV5 VP3 capsid polypeptides having one or more mutations in the VP2 and VP3 regions corresponding to the AAV5 VP1 amino acid residues of the 581 to 589 region, where the one or more mutations comport to the rules or sequences in the following section.

A. Positional Frequency Rules

Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased lung tissue tropism as compared to wildtype AAV5 VP capsid polypeptide, wherein the engineered variant AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein said one or more mutations are selected from the following rules. In some embodiments, Xaa1 is selected from A, E, K, M, Q, R, S, or T. In some embodiments, Xaa1 is selected from A, E, or Q. In some embodiments, Xaa1 is E. In some embodiments, Xaa2 is selected from A, I, K, S, T, or V. In some embodiments, Xaa2 is selected from S, T, or V. In some embodiments, Xaa2 is T. In some embodiments, Xaa3 is selected from A, E, K, M, Q, R, S, T, or V. In some embodiments, Xaa3 is selected from A, K, R, or S. In some embodiments, Xaa3 is R. In some embodiments, Xaa4 is selected from M, P, R, S, or T. In some embodiments, Xaa4 is selected from P, Q, or T. In some embodiments, Xaa4 is Q. In some embodiments, Xaa5 is selected from I, K, L, M, T, V, or Y. In some embodiments, Xaa5 is selected from L, M, or Y. In some embodiments, Xaa5 is L. In some embodiments, Xaa6 is selected from D, G, H, M, N, R, or S. In some embodiments, Xaa6 is selected from H or N. In some embodiments, Xaa6 is N. In some embodiments, Xaa7 is selected from A, K, M, Q, or R. In some embodiments, Xaa7 is selected from A, K or R. In some embodiments, Xaa7 is R. In some embodiments, Xaa8 is selected from A, F, G, S, W, or Y. In some embodiments, Xaa8 is selected from A, F, or G. In some embodiments, Xaa8 is F. In some embodiments, Xaa9 is selected from A, E, G, P, R, or Y. In some embodiments, Xaa9 is selected from G, P, or R. In some embodiments, Xaa9 is G.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 16118-SEQ ID NO: 17117, wherein said at least one mutation drives increased lung tissue tropism.

B. ML Rules

For the following set of rules described in this paragraph, favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an engineered AAV5 VP1 capsid polypeptide, were determined using in vivo data and two ML models, which are described in EXAMPLE 37. Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased lung tissue tropism as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide, wherein the engineered AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the engineered AAV5 VP1 capsid polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein Xaa1 is selected from an amino acid of high mutability (e.g., Xaa1 is selected from D, E, M, A, I, Q, or T); or wherein Xaa2 is selected from an amino acid of high mol mass (e.g., Xaa2 is selected from F); or wherein Xaa2 is selected from an amino acid of low mutability (e.g., Xaa2 is selected from Y, F, or L); or wherein Xaa3 is selected from an amino acid of low mutability (e.g., Xaa3 is selected from K, V, P, or H); or wherein Xaa3 is selected from an amino acid of low hydropathy (e.g., Xaa3 is selected from K or R); or wherein Xaa4 is selected from an amino acid of low mutability (e.g., Xaa4 is selected from K or P); or wherein Xaa4 is selected from an amino acid of high average flexibility (e.g., Xaa4 is selected from D, E, P, or S); or wherein Xaa5 is selected from an amino acid of low average flexibility (e.g., Xaa5 is selected from W, M, or F); or wherein Xaa5 is selected from an amino acid of high solubility (e.g., Xaa5 is selected from W, F, I, or L); or wherein Xaa6 is selected from an amino acid of medium mutability (e.g., Xaa6 is selected from R or H); or wherein Xaa6 is selected from an amino acid of high surface accessibility (e.g., Xaa6 is selected from T); or wherein Xaa7 is selected from an amino acid of low mutability (e.g., Xaa7 is selected from C); or wherein Xaa7 is selected from an amino acid of high solubility (e.g., Xaa7 is selected from W, V, M, F, I, or L); or wherein Xaa8 is selected from an amino acid of high mutability (e.g., Xaa8 is selected from D, E, M, A, I, Q, or T); or wherein Xaa8 is selected from an amino acid of low hydropathy (e.g., Xaa8 is selected from R or K); or wherein Xaa9 is selected from an amino acid of high average flexibility (e.g., Xaa9 is selected from R or G); or any combination thereof.

In some embodiments, Xaa1 is selected from an amino acid of high mutability. In some embodiments, Xaa1 is selected from D, E, M, A, T, Q, or T. In some embodiments, Xaa2 is selected from an amino acid of high mol mass. In some embodiments, Xaa2 is selected from F. In some embodiments, Xaa2 is selected from an amino acid of low mutability. In some embodiments, Xaa2 is selected from Y, F, or L. In some embodiments, Xaa3 is selected from an amino acid of low mutability. In some embodiments, Xaa3 is selected from K, V, P, or H. In some embodiments, Xaa3 is selected from an amino acid of low hydropathy. In some embodiments, Xaa3 is selected from K or R. In some embodiments, Xaa4 is selected from an amino acid of low mutability. In some embodiments, Xaa4 is selected from K or P. In some embodiments, Xaa4 is selected from an amino acid of high average flexibility. In some embodiments, Xaa4 is selected from D, E, P, or S. In some embodiments, Xaa5 is selected from an amino acid of low average flexibility. In some embodiments, Xaa5 is selected from W, M, or F. In some embodiments, Xaa5 is selected from an amino acid of high solubility. In some embodiments, Xaa5 is selected from W, F, I, or L. In some embodiments, Xaa6 is selected from an amino acid of medium mutability. In some embodiments, Xaa6 is selected from R or H. In some embodiments, Xaa6 is selected from an amino acid of high surface accessibility. In some embodiments, Xaa6 is selected from T. In some embodiments, Xaa7 is selected from an amino acid of low mutability. In some embodiments, Xaa7 is selected from C. In some embodiments, Xaa7 is selected from an amino acid of high solubility. In some embodiments, Xaa7 is selected from W, V, M, F, I, or L. In some embodiments, Xaa8 is selected from an amino acid of high mutability. In some embodiments, Xaa8 is selected from D, E, M, A, I, Q, or T. In some embodiments, Xaa8 is selected from an amino acid of low hydropathy. In some embodiments, Xaa8 is selected from R or K. In some embodiments, Xaa9 is selected from an amino acid of high average flexibility. In some embodiments, Xaa9 is selected from R or G.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 17118-SEQ ID NO: 18117, wherein said at least one mutation drives increased lung tissue tropism.

C. Enriched Lung Sequences

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 18118-SEQ ID NO: 19117, wherein said at least one mutation drives increased lung tissue tropism.

6.7.14. In Vivo Selected Mutated VP Polypeptides that Confer Increased Heart Tropism

The present disclosure provides AAV5 virions with a VP capsid polypeptide having at least one mutation in a region with residues that interact with target cells (e.g., a target heart cell in a target heart tissue of interest), where the at least one mutation confers increased heart tissue tropism as compared to a wildtype VP capsid polypeptide. In some embodiments, provided herein are AAV5 VP1 capsid polypeptide having a sequence homology of at least 80% to SEQ ID NO: 1, wherein the AAV5 VP1 capsid polypeptide has at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of SEQ ID NO: 1 and wherein said at least one mutation drives increased heart tropism. The following sequences rules and sequences also apply to the region in AAV5 VP2 (amino acid residues 445 to 453; VP2 sequence shown in SEQ ID NO: 1115) and AAV5 VP3 (amino acid residues 389 to 397; VP3 sequences shown in SEQ ID NO: 1116) corresponding to AAV5 VP1 amino acid residues 581 to 589. Thus, the present disclosure encompasses AAV5 VP2 capsid polypeptides and AAV5 VP3 capsid polypeptides having one or more mutations in the VP2 and VP3 regions corresponding to the AAV5 VP1 amino acid residues of the 581 to 589 region, where the one or more mutations comport to the rules or sequences in the following section. In some embodiments, “heart” and “cardiac” may be used interchangeably herein.

A. Positional Frequency Rules

Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased heart tissue tropism as compared to wildtype AAV5 VP capsid polypeptide, wherein the engineered variant AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein said one or more mutations are selected from the following rules. In some embodiments, Xaa1 is selected from I, K, L, M, T, or V. In some embodiments, Xaa1 is selected from K or L. In some embodiments, Xaa1 is K. In some embodiments, Xaa2 is selected from A, C, G, I, K, or S. In some embodiments, Xaa2 is selected from A, C, or S. In some embodiments, Xaa2 is A. In some embodiments, Xaa3 is selected from A, D, E, G, K, M, or V. In some embodiments, Xaa3 is selected from E or V. In some embodiments, Xaa3 is E. In some embodiments, Xaa4 is selected from F, H, R, T, W, or Y. In some embodiments, Xaa4 is selected from F, R, or T. In some embodiments, Xaa4 is R. In some embodiments, Xaa5 is selected from F, L, M, or R. In some embodiments, Xaa5 is L. In some embodiments, Xaa6 is selected from A, H, N, W, or Y. In some embodiments, Xaa6 is selected from H, N, or Y. In some embodiments, Xaa6 is H. In some embodiments, Xaa7 is selected from A, C, E, F, K, or T. In some embodiments, Xaa7 is selected from C, F, or T. In some embodiments, Xaa7 is F. In some embodiments, Xaa8 is selected from A, C, M, S, or T. In some embodiments, Xaa8 is selected from C, M, or S. In some embodiments, Xaa8 is C. In some embodiments, Xaa9 is selected from A, D, G, or P. In some embodiments, Xaa9 is selected from A or G. In some embodiments, Xaa9 is A.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 13118-SEQ ID NO: 14117, wherein said at least one mutation drives increased heart tissue tropism.

B. ML Rules

For the following set of rules described in this paragraph, favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an engineered AAV5 VP1 capsid polypeptide, were determined using in vivo data and two ML models, which are described in EXAMPLE 34. Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased heart tissue tropism as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide, wherein the engineered AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the engineered AAV5 VP1 capsid polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein Xaa1 is selected from an amino acid of low solubility (e.g., Xaa1 is selected from N or E); or wherein Xaa1 is selected from an amino acid of low hydropathy (e.g., Xaa1 is selected from H, N, Q, P, Y, D, or E); or wherein Xaa1 is selected from an amino acid of high mutability (e.g., Xaa1 is selected from A or E); or wherein Xaa2 is selected from an amino acid of high hydropathy (e.g., Xaa2 is selected from V or I); or wherein Xaa2 is selected from an amino acid of medium mutability (e.g., Xaa2 is selected from V); or wherein Xaa2 is selected from an amino acid of medium volume (e.g., Xaa2 is selected from V, E, or Q); or wherein Xaa2 is selected from an amino acid of high solubility (e.g., Xaa2 is selected from V or M); or wherein Xaa3 is selected from an amino acid of low solubility (e.g., Xaa3 is selected from R or Q); or wherein Xaa4 is selected from an amino acid of low surface accessibility (e.g., Xaa4 is selected from C); or wherein Xaa4 is selected from an amino acid of high solubility (e.g., Xaa4 is selected from C); or wherein Xaa4 is selected from an amino acid of low charge (e.g., Xaa4 is selected from D, E, Y, W, V, P, M, A, G, F, I, L, N, Q, S, T, or C); or wherein Xaa4 is selected from an amino acid of high hydropathy (e.g., Xaa4 is selected from C); or wherein Xaa5 is selected from an amino acid of high surface accessibility (e.g., Xaa5 is selected from D, E, R, K, N, or Q); or wherein Xaa5 is selected from an amino acid of low solubility (e.g., Xaa5 is selected from D); or wherein Xaa6 is selected from an amino acid of low mutability (e.g., Xaa6 is selected from C); or wherein Xaa6 is selected from an amino acid of low solubility (e.g., Xaa6 is selected from D); or wherein Xaa8 is selected from an amino acid of high surface accessibility (e.g., Xaa8 is selected from D or N); or wherein Xaa8 is selected from an amino acid of high average flexibility (e.g., Xaa8 is selected from D, R, P, G, or S); or wherein Xaa9 is selected from an amino acid of medium mol mass (e.g., Xaa9 is selected from N, D, L, or I); or any combination thereof.

In some embodiments, Xaa1 is selected from an amino acid of low solubility. In some embodiments, Xaa1 is selected from N or E. In some embodiments, Xaa1 is selected from an amino acid of low hydropathy. In some embodiments, Xaa1 is selected from H, N, Q, P, Y, D, or E. In some embodiments, Xaa1 is selected from an amino acid of high mutability. In some embodiments, Xaa1 is selected from A or E. In some embodiments, Xaa2 is selected from an amino acid of high hydropathy. In some embodiments, Xaa2 is selected from V or I. In some embodiments, Xaa2 is selected from an amino acid of medium mutability. In some embodiments, Xaa2 is selected from V. In some embodiments, Xaa2 is selected from an amino acid of medium volume. In some embodiments, Xaa2 is selected from V, E, or Q. In some embodiments, Xaa2 is selected from an amino acid of high solubility. In some embodiments, Xaa2 is selected from V or M. In some embodiments, Xaa3 is selected from an amino acid of low solubility. In some embodiments, Xaa3 is selected from R or Q. In some embodiments, Xaa4 is selected from an amino acid of low surface accessibility. In some embodiments, Xaa4 is selected from C. In some embodiments, Xaa4 is selected from an amino acid of high solubility. In some embodiments, Xaa4 is selected from C. In some embodiments, Xaa4 is selected from an amino acid of low charge. In some embodiments, Xaa4 is selected from D, E, Y, W, V, P, M, A, G, F, I, L, N, Q, S, T, or C. In some embodiments, Xaa4 is selected from an amino acid of high hydropathy. In some embodiments, Xaa4 is selected from C. In some embodiments, Xaa5 is selected from an amino acid of high surface accessibility. In some embodiments, Xaa5 is selected from D, E, R, K, N, or Q. In some embodiments, Xaa5 is selected from an amino acid of low solubility. In some embodiments, Xaa5 is selected from D. In some embodiments, Xaa6 is selected from an amino acid of low mutability. In some embodiments, Xaa6 is selected from C. In some embodiments, Xaa6 is selected from an amino acid of low solubility. In some embodiments, Xaa6 is selected from D. In some embodiments, Xaa8 is selected from an amino acid of high surface accessibility. In some embodiments, Xaa8 is selected from D or N. In some embodiments, Xaa8 is selected from an amino acid of high average flexibility. In some embodiments, Xaa8 is selected from D, R, P, G, or S. In some embodiments, Xaa9 is selected from an amino acid of medium mol mass. In some embodiments, Xaa9 is selected from N, D, L, or I.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 14118-SEQ ID NO: 15117, wherein said at least one mutation drives increased heart tissue tropism.

C. Enriched Heart Sequences

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 15118-SEQ ID NO: 16117, wherein said at least one mutation drives increased heart tissue tropism.

6.7.15. In Vivo Selected Mutated VP Polypeptides that Confer Increased Colon Tropism

The present disclosure provides AAV5 virions with a VP capsid polypeptide having at least one mutation in a region with residues that interact with target cells (e.g., a target colon cell in a target colon tissue of interest), where the at least one mutation confers increased colon tissue tropism as compared to a wildtype VP capsid polypeptide. In some embodiments, provided herein are AAV5 VP1 capsid polypeptide having a sequence homology of at least 80% to SEQ ID NO: 1, wherein the AAV5 VP1 capsid polypeptide has at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of SEQ ID NO: 1 and wherein said at least one mutation drives increased colon tropism. The following sequences rules and sequences also apply to the region in AAV5 VP2 (amino acid residues 445 to 453; VP2 sequence shown in SEQ ID NO: 1115) and AAV5 VP3 (amino acid residues 389 to 397; VP3 sequences shown in SEQ ID NO: 1116) corresponding to AAV5 VP1 amino acid residues 581 to 589. Thus, the present disclosure encompasses AAV5 VP2 capsid polypeptides and AAV5 VP3 capsid polypeptides having one or more mutations in the VP2 and VP3 regions corresponding to the AAV5 VP1 amino acid residues of the 581 to 589 region, where the one or more mutations comport to the rules or sequences in the following section.

A. Positional Frequency Rules

Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased colon tissue tropism as compared to wildtype AAV5 VP capsid polypeptide, wherein the engineered variant AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein said one or more mutations are selected from the following rules. In some embodiments, Xaa1 is selected from C, F, H, N, P, W, or Y. In some embodiments, Xaa1 is selected from F, P, or W. In some embodiments, Xaa1 is P. In some embodiments, Xaa2 is selected from D, E, F, L, or P. In some embodiments, Xaa2 is selected from D, E, L, or P. In some embodiments, Xaa2 is P. In some embodiments, Xaa3 is selected from C, F, H, I, L, P, or Y. In some embodiments, Xaa3 is selected from C, H, or P. In some embodiments, Xaa3 is P. In some embodiments, Xaa4 is selected from C, D, E, N, or P. In some embodiments, Xaa4 is selected from C, D, or E. In some embodiments, Xaa4 is C. In some embodiments, Xaa5 is selected from D, E, G, P, or W. In some embodiments, Xaa5 is selected from G, P, or W. In some embodiments, Xaa5 is P. In some embodiments, Xaa6 is selected from C, K, R, or V. In some embodiments, Xaa6 is selected from K or R. In some embodiments, Xaa6 is R. In some embodiments, Xaa7 is selected from D, M, P, or V. In some embodiments, Xaa7 is P. In some embodiments, Xaa8 is selected from D, I, K, L, P, R, or V. In some embodiments, Xaa8 is selected from K, P, or R. In some embodiments, Xaa8 is P. In some embodiments, Xaa9 is selected from C, H, I, K, L, M, or W. In some embodiments, Xaa9 is selected from I, L, or M. In some embodiments, Xaa9 is I.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 10118-SEQ ID NO: 11117, % wherein said at least one mutation drives increased colon tissue tropism.

B. ML Rules

For the following set of rules described in this paragraph, favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an engineered AAV5 VP1 capsid polypeptide, were determined using in vivo data and two ML models, which are described in EXAMPLE 33. Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased colon tissue tropism as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide, wherein the engineered AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the engineered AAV5 VP1 capsid polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein Xaa1 is selected from an amino acid of high mol mass (e.g., Xaa1 is selected from Y or W); or wherein Xaa1 is selected from an amino acid of high solubility (e.g., Xaa1 is selected from W, F, I, or L); or wherein Xaa2 is selected from an amino acid of low solubility (e.g., Xaa2 is selected from D); or wherein Xaa2 is selected from an amino acid of low mutability (e.g., Xaa2 is selected from P or K); or wherein Xaa2 is selected from an amino acid of medium mol mass (e.g., Xaa2 is selected from D, E, N, K, M, Q, I, or L); or wherein Xaa2 is selected from an amino acid of low hydropathy (e.g., Xaa2 is selected from D, E, R, K, H, N, or Q); or wherein Xaa3 is selected from an amino acid of low mutability (e.g., Xaa3 is selected from K, V, P, or C); or wherein Xaa3 is selected from an amino acid of high solubility (e.g., Xaa3 is selected from W, F, I, or L); or wherein Xaa5 is selected from an amino acid of high average flexibility (e.g., Xaa5 is selected from S, P, G, R, E, or D); or wherein Xaa5 is selected from an amino acid of high surface accessibility (e.g., Xaa5 is selected from D or N); or wherein Xaa6 is selected from an amino acid of low hydropathy (e.g., Xaa6 is selected from R); or wherein Xaa6 is selected from an amino acid of low mutability (e.g., Xaa6 is selected from Y, R, F, or L); or wherein Xaa6 is selected from an amino acid of low solubility (e.g., Xaa6 is selected from R or Q); or wherein Xaa6 is selected from an amino acid of high surface accessibility (e.g., Xaa6 is selected from E, R, or K); or wherein Xaa6 is selected from an amino acid of high average flexibility (e.g., Xaa6 is selected from G or R); or wherein Xaa8 is selected from an amino acid of low solubility (e.g., Xaa8 is selected from D); or any combination thereof.

In some embodiments, Xaa1 is selected from an amino acid of high mol mass. In some embodiments, Xaa1 is selected from Y or W. In some embodiments, Xaa1 is selected from an amino acid of high solubility. In some embodiments, Xaa1 is selected from W, F, I, or L. In some embodiments, Xaa2 is selected from an amino acid of low solubility. In some embodiments, Xaa2 is selected from D. In some embodiments, Xaa2 is selected from an amino acid of low mutability. In some embodiments, Xaa2 is selected from P, K. In some embodiments, Xaa2 is selected from an amino acid of medium mol mass. In some embodiments, Xaa2 is selected from D, E, N, K, M, Q, I, or L. In some embodiments, Xaa2 is selected from an amino acid of low hydropathy. In some embodiments, Xaa2 is selected from D, E, R, K, H, N, or Q. In some embodiments, Xaa3 is selected from an amino acid of low mutability. In some embodiments, Xaa3 is selected from K, V, P, or C. In some embodiments, Xaa3 is selected from an amino acid of high solubility. In some embodiments, Xaa3 is selected from W, F, I, or L. In some embodiments, Xaa5 is selected from an amino acid of high average flexibility. In some embodiments, Xaa5 is selected from S, P, G, R, E, or D. In some embodiments, Xaa5 is selected from an amino acid of high surface accessibility. In some embodiments, Xaa5 is selected from D or N. In some embodiments, Xaa6 is selected from an amino acid of low hydropathy. In some embodiments, Xaa6 is selected from R. In some embodiments, Xaa6 is selected from an amino acid of low mutability. In some embodiments, Xaa6 is selected from Y, R, F, or L. In some embodiments, Xaa6 is selected from an amino acid of low solubility. In some embodiments, Xaa6 is selected from R or Q. In some embodiments, Xaa6 is selected from an amino acid of high surface accessibility. In some embodiments, Xaa6 is selected from E, R, or K. In some embodiments, Xaa6 is selected from an amino acid of high average flexibility. In some embodiments, Xaa6 is selected from G or R. In some embodiments, Xaa8 is selected from an amino acid of low solubility. In some embodiments, Xaa8 is selected from D.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 11118-SEQ ID NO: 12117, wherein said at least one mutation drives increased colon tissue tropism.

C. Enriched Colon Sequences

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 12118-SEQ ID NO: 13117, wherein said at least one mutation drives increased colon tissue tropism.

6.7.16. In Vivo Selected Mutated VP Polypeptides that Confer Increased Thyroid Gland Tropism

The present disclosure provides AAV5 virions with a VP capsid polypeptide having at least one mutation in a region with residues that interact with target cells (e.g., a target thyroid cell in a target thyroid gland tissue of interest), where the at least one mutation confers increased thyroid gland tissue tropism as compared to a wildtype VP capsid polypeptide. In some embodiments, provided herein are AAV5 VP1 capsid polypeptide having a sequence homology of at least 80% to SEQ ID NO: 1, wherein the AAV5 VP1 capsid polypeptide has at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of SEQ ID NO: 1 and wherein said at least one mutation drives increased thyroid gland tropism. The following sequences rules and sequences also apply to the region in AAV5 VP2 (amino acid residues 445 to 453; VP2 sequence shown in SEQ ID NO: 1115) and AAV5 VP3 (amino acid residues 389 to 397; VP3 sequences shown in SEQ ID NO: 1116) corresponding to AAV5 VP1 amino acid residues 581 to 589. Thus, the present disclosure encompasses AAV5 VP2 capsid polypeptides and AAV5 VP3 capsid polypeptides having one or more mutations in the VP2 and VP3 regions corresponding to the AAV5 VP1 amino acid residues of the 581 to 589 region, where the one or more mutations comport to the rules or sequences in the following section.

A. Positional Frequency Rules

Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased thyroid gland tissue tropism as compared to wildtype AAV5 VP capsid polypeptide, wherein the engineered variant AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein said one or more mutations are selected from the following rules. In some embodiments, Xaa1 is selected from A, K, M, N, Q, or R. In some embodiments, Xaa1 is selected from K, N or Q. In some embodiments, Xaa1 is K. In some embodiments, Xaa2 is selected from A, F, K, L, M, T, V, or W. In some embodiments, Xaa2 is selected from F, V, or W. In some embodiments, Xaa2 is W. In some embodiments, Xaa3 is selected from A, I, K, R, S, T, V, or W. In some embodiments, Xaa3 is selected from A, R or T. In some embodiments, Xaa3 is R In some embodiments, Xaa4 is selected from A, D, E, I, P, or V. In some embodiments, Xaa4 is selected from A, E, or I. In some embodiments, Xaa4 is A. In some embodiments, Xaa5 is selected from F, I, M, Q, V, or Y. In some embodiments, Xaa5 is M, V, Y. In some embodiments, Xaa5 is M. In some embodiments, Xaa6 is selected from H, M, N, or Y. In some embodiments, Xaa6 is N. In some embodiments, Xaa7 is selected from H, I, N, Q, S, or W. In some embodiments, Xaa7 is selected from H, I, or N. In some embodiments, Xaa7 is H. In some embodiments, Xaa8 is selected from A, D, F, Q, S, or Y. In some embodiments, Xaa8 is selected from A, F, or S. In some embodiments, Xaa8 is F. In some embodiments, Xaa9 is selected from A, Q, S, or Y. In some embodiments, Xaa9 is selected from A or S. In some embodiments, Xaa9 is A.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 40438-SEQ ID NO: 41437, wherein said at least one mutation drives increased thyroid gland tissue tropism.

B. ML Rules

For the following set of rules described in this paragraph, favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an engineered AAV5 VP1 capsid polypeptide, were determined using in vivo data and two ML models, which are described in EXAMPLE 43. Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased thyroid gland tissue tropism as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide, wherein the engineered AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the engineered AAV5 VP1 capsid polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein Xaa1 is selected from an amino acid of high mutability (e.g., Xaa1 is selected from N); or wherein Xaa2 is selected from an amino acid of low surface accessibility (e.g., Xaa2 is selected from F, G, or M); or wherein Xaa3 is selected from an amino acid of high solubility (e.g., Xaa3 is selected from F); or wherein Xaa3 is selected from an amino acid of low mutability (e.g., Xaa3 is selected from Y, F, L, or C); or wherein Xaa3 is selected from an amino acid of medium mol mass (e.g., Xaa3 is selected from D, E, R, K, V, P, M, I, L, N, Q, T, or C); or wherein Xaa3 is selected from an amino acid of low surface accessibility (e.g., Xaa3 is selected from V, I, L, or C); or wherein Xaa4 is selected from an amino acid of high goldman engelman steitz (e.g., Xaa4 is selected from L or V); or wherein Xaa4 is selected from an amino acid of low surface accessibility (e.g., Xaa4 is selected from V, M, A, G, F, I, or L); or wherein Xaa4 is selected from an amino acid of low mol mass (e.g., Xaa4 is selected from D, A, G, I, L, or N); or wherein Xaa5 is selected from an amino acid of high solubility (e.g., Xaa5 is selected from C, L, F, M, V, or Y); or wherein Xaa5 is selected from an amino acid of low solubility (e.g., Xaa5 is selected from D); or wherein Xaa5 is selected from an amino acid of low average flexibility (e.g., Xaa5 is selected from F, M, or W); or wherein Xaa6 is selected from an amino acid of low average flexibility (e.g., Xaa6 is selected from F, M, or W); or wherein Xaa7 is selected from an amino acid of high mutability (e.g., Xaa7 is selected from N); or wherein Xaa7 is selected from an amino acid of low volume (e.g., Xaa7 is selected from P, N, or T); or wherein Xaa8 is selected from an amino acid of low average flexibility (e.g., Xaa8 is selected from F, M, or W); or wherein Xaa8 is selected from an amino acid of low surface accessibility (e.g., Xaa8 is selected from M, G, or F); or wherein Xaa9 is selected from an amino acid of low mutability (e.g., Xaa9 is selected from R, K, P, H, or C); or wherein Xaa9 is selected from an amino acid of low hydropathy (e.g., Xaa9 is selected from R); or any combination thereof.

In some embodiments, Xaa1 is selected from an amino acid of high mutability. In some embodiments, Xaa1 is selected from N. In some embodiments, Xaa2 is selected from an amino acid of low surface accessibility. In some embodiments, Xaa2 is selected from F, G, or M. In some embodiments, Xaa3 is selected from an amino acid of high solubility. In some embodiments, Xaa3 is selected from F. In some embodiments, Xaa3 is selected from an amino acid of low mutability. In some embodiments, Xaa3 is selected from Y, F, L, or C. In some embodiments, Xaa3 is selected from an amino acid of medium mol mass. In some embodiments, Xaa3 is selected from D, E, R, K, V, P, M, I, L, N, Q, T, or C. In some embodiments, Xaa3 is selected from an amino acid of low surface accessibility. In some embodiments, Xaa3 is selected from V, I, L, or C. In some embodiments, Xaa4 is selected from an amino acid of high goldman engelman steitz. In some embodiments, Xaa4 is selected from L or V. In some embodiments, Xaa4 is selected from an amino acid of low surface accessibility. In some embodiments, Xaa4 is selected from V, M, A, G, F, I, or L. In some embodiments, Xaa4 is selected from an amino acid of low mol mass. In some embodiments, Xaa4 is selected from D, A, G, I, L, or N. In some embodiments, Xaa5 is selected from an amino acid of high solubility. In some embodiments, Xaa5 is selected from C, L, F, M, V, or Y. In some embodiments, Xaa5 is selected from an amino acid of low solubility. In some embodiments, Xaa5 is selected from D. In some embodiments, Xaa5 is selected from an amino acid of low average flexibility. In some embodiments, Xaa5 is selected from F, M, or W. In some embodiments, Xaa6 is selected from an amino acid of low average flexibility. In some embodiments, Xaa6 is selected from F, M, or W. In some embodiments, Xaa7 is selected from an amino acid of high mutability. In some embodiments, Xaa7 is selected from N. In some embodiments, Xaa7 is selected from an amino acid of low volume. In some embodiments, Xaa7 is selected from P, N, or T. In some embodiments, Xaa8 is selected from an amino acid of low average flexibility. In some embodiments, Xaa8 is selected from F, M, or W. In some embodiments, Xaa8 is selected from an amino acid of low surface accessibility. In some embodiments, Xaa8 is selected from M, G, or F. In some embodiments, Xaa9 is selected from an amino acid of low mutability. In some embodiments, Xaa9 is selected from R, K, P, H, or C. In some embodiments, Xaa9 is selected from an amino acid of low hydropathy. In some embodiments, Xaa9 is selected from R.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 41438-SEQ ID NO: 42437, wherein said at least one mutation drives increased thyroid gland tissue tropism.

C. Enriched Thyroid Gland Sequences

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 42438-SEQ ID NO: 43437, wherein said at least one mutation drives increased thyroid gland tissue tropism.

6.7.17. In Vivo Selected Mutated VP Polypeptides that Confer Increased Lymph Node Tropism

The present disclosure provides AAV5 virions with a VP capsid polypeptide having at least one mutation in a region with residues that interact with target cells (e.g., a target lymph node cell in a target lymph node tissue of interest), where the at least one mutation confers increased lymph node tissue tropism as compared to a wildtype VP capsid polypeptide. In some embodiments, provided herein are AAV5 VP1 capsid polypeptide having a sequence homology of at least 80% to SEQ ID NO: 1, wherein the AAV5 VP1 capsid polypeptide has at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of SEQ ID NO: 1 and wherein said at least one mutation drives increased lymph node tropism. The following sequences rules and sequences also apply to the region in AAV5 VP2 (amino acid residues 445 to 453; VP2 sequence shown in SEQ ID NO: 1115) and AAV5 VP3 (amino acid residues 389 to 397; VP3 sequences shown in SEQ ID NO: 1116) corresponding to AAV5 VP1 amino acid residues 581 to 589. Thus, the present disclosure encompasses AAV5 VP2 capsid polypeptides and AAV5 VP3 capsid polypeptides having one or more mutations in the VP2 and VP3 regions corresponding to the AAV5 VP1 amino acid residues of the 581 to 589 region, where the one or more mutations comport to the rules or sequences in the following section.

A. Positional Frequency Rules

Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased lymph node tissue tropism as compared to wildtype AAV5 VP capsid polypeptide, wherein the engineered variant AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein said one or more mutations are selected from the following rules. In some embodiments, Xaa1 is selected from A, D, E, Q, S, or T. In some embodiments, Xaa1 is selected from D, E, or T. In some embodiments, Xaa1 is E. In some embodiments, Xaa2 is selected from A, H, I, S, T, or V. In some embodiments, Xaa2 is selected from I, T, or V. In some embodiments, Xaa2 is V. In some embodiments, Xaa3 is selected from A, E, H, I, T, or V. In some embodiments, Xaa3 is selected from A, I, T, or V. In some embodiments, Xaa3 is T. In some embodiments, Xaa4 is selected from A, D, E, or P. In some embodiments, Xaa4 is selected from D, or E. In some embodiments, Xaa4 is E. In some embodiments, Xaa5 is selected from I, L, M, V, or Y. In some embodiments, Xaa5 is selected from I, L, V, or Y. In some embodiments, Xaa5 is L. In some embodiments, Xaa6 is selected from D, E, I, N, or Q. In some embodiments, Xaa6 is selected from D, E, or I. In some embodiments, Xaa6 is D. In some embodiments, Xaa7 is selected from A, E, G, Q, or V. In some embodiments, Xaa7 is A, Q, or V. In some embodiments, Xaa7 is V. In some embodiments, Xaa8 is selected from F, G, M, or W. In some embodiments, Xaa8 is selected from F or W. In some embodiments, Xaa8 is W. In some embodiments, Xaa9 is selected from I, P, T, or Y. In some embodiments, Xaa9 is I or P.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 19118-SEQ ID NO: 20117, wherein said at least one mutation drives increased lymph node tissue tropism.

B. ML Rules

For the following set of rules described in this paragraph, favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an engineered AAV5 VP1 capsid polypeptide, were determined using in vivo data and two ML models, which are described in EXAMPLE 35. Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased lymph node tissue tropism as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide, wherein the engineered AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the engineered AAV5 VP1 capsid polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein Xaa1 is selected from an amino acid of high average flexibility (e.g., Xaa1 is selected from D, E, P, G, Q, S, or R); or wherein Xaa1 is selected from an amino acid of high hbond donors (e.g., Xaa1 is selected from R); or wherein Xaa1 is selected from an amino acid of high mol mass (e.g., Xaa1 is selected from Y, W, R, or F); or wherein Xaa2 is selected from an amino acid of low solubility (e.g., Xaa2 is selected from N or E); or wherein Xaa3 is selected from an amino acid of low average flexibility (e.g., Xaa3 is selected from W, M, or F); or wherein Xaa3 is selected from an amino acid of low mutability (e.g., Xaa3 is selected from R, H, K, P, Y, F, L, or C); or wherein Xaa4 is selected from an amino acid of low mutability (e.g., Xaa4 is selected from C); or wherein Xaa5 is selected from an amino acid of high mutability (e.g., Xaa5 is selected from N); or wherein Xaa5 is selected from an amino acid of medium mol mass (e.g., Xaa5 is selected from D, I, L, or N); or wherein Xaa6 is selected from an amino acid of high mol mass (e.g., Xaa6 is selected from Y, W, R, or F); or wherein Xaa6 is selected from an amino acid of high average flexibility (e.g., Xaa6 is selected from G or R); or wherein Xaa7 is selected from an amino acid of high average flexibility (e.g., Xaa7 is selected from D, E, K, P, I, N, Q, or S); or wherein Xaa7 is selected from an amino acid of low solubility (e.g., Xaa7 is selected from N, E); or wherein Xaa8 is selected from an amino acid of low solubility (e.g., Xaa8 is selected from N, E, or D); or wherein Xaa8 is selected from an amino acid of medium mutability (e.g., Xaa8 is selected from R or H); or wherein Xaa9 is selected from an amino acid of low mutability (e.g., Xaa9 is selected from P or K); or wherein Xaa9 is selected from an amino acid of high average flexibility (e.g., Xaa9 is selected from D, E, P, or S); or wherein Xaa9 is selected from an amino acid of high solubility (e.g., Xaa9 is selected from M or V); or any combination thereof.

In some embodiments, Xaa1 is selected from an amino acid of high average flexibility. In some embodiments, Xaa1 is selected from D, E, P, G, Q, S, or R. In some embodiments, Xaa1 is selected from an amino acid of high hbond donors. In some embodiments, Xaa1 is selected from R. In some embodiments, Xaa1 is selected from an amino acid of high mol mass. In some embodiments, Xaa1 is selected from Y, W, R, or F. In some embodiments, Xaa2 is selected from an amino acid of low solubility. In some embodiments, Xaa2 is selected from N or E. In some embodiments, Xaa3 is selected from an amino acid of low average flexibility. In some embodiments, Xaa3 is selected from W, M, or F. In some embodiments, Xaa3 is selected from an amino acid of low mutability. In some embodiments, Xaa3 is selected from R, H, K, P, Y, F, L, or C. In some embodiments, Xaa4 is selected from an amino acid of low mutability. In some embodiments, Xaa4 is selected from C. In some embodiments, Xaa5 is selected from an amino acid of high mutability. In some embodiments, Xaa5 is selected from N. In some embodiments, Xaa5 is selected from an amino acid of medium mol mass. In some embodiments, Xaa5 is selected from D, I, L, or N. In some embodiments, Xaa6 is selected from an amino acid of high mol mass. In some embodiments, Xaa6 is selected from Y, W, R, or F. In some embodiments, Xaa6 is selected from an amino acid of high average flexibility. In some embodiments, Xaa6 is selected from G, R. In some embodiments, Xaa7 is selected from an amino acid of high average flexibility. In some embodiments, Xaa7 is selected from D, E, K, P, I, N, Q, or S. In some embodiments, Xaa7 is selected from an amino acid of low solubility. In some embodiments, Xaa7 is selected from N or E. In some embodiments, Xaa8 is selected from an amino acid of low solubility. In some embodiments, Xaa8 is selected from N, E, or D. In some embodiments, Xaa8 is selected from an amino acid of medium mutability. In some embodiments, Xaa8 is selected from R or H. In some embodiments, Xaa9 is selected from an amino acid of low mutability. In some embodiments, Xaa9 is selected from P or K. In some embodiments, Xaa9 is selected from an amino acid of high average flexibility. In some embodiments, Xaa9 is selected from D, E, P, or S. In some embodiments, Xaa9 is selected from an amino acid of high solubility. In some embodiments, Xaa9 is selected from M or V.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 20118-SEQ ID NO: 21117, wherein said at least one mutation drives increased lymph node tissue tropism.

C. Enriched Lymph Node Sequences

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 21118-SEQ ID NO: 22117, wherein said at least one mutation drives increased lymph node tissue tropism.

6.7.18. In Vivo Selected Mutated VP Polypeptides that Confer Increased Skin Tropism

The present disclosure provides AAV5 virions with a VP capsid polypeptide having at least one mutation in a region with residues that interact with target cells (e.g., a target skin cell in a target skin tissue of interest), where the at least one mutation confers increased skin tissue tropism as compared to a wildtype VP capsid polypeptide. In some embodiments, provided herein are AAV5 VP1 capsid polypeptide having a sequence homology of at least 80% to SEQ ID NO: 1, wherein the AAV5 VP1 capsid polypeptide has at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of SEQ ID NO: 1 and wherein said at least one mutation drives increased skin tropism. The following sequences rules and sequences also apply to the region in AAV5 VP2 (amino acid residues 445 to 453; VP2 sequence shown in SEQ ID NO: 1115) and AAV5 VP3 (amino acid residues 389 to 397; VP3 sequences shown in SEQ ID NO: 1116) corresponding to AAV5 VP1 amino acid residues 581 to 589. Thus, the present disclosure encompasses AAV5 VP2 capsid polypeptides and AAV5 VP3 capsid polypeptides having one or more mutations in the VP2 and VP3 regions corresponding to the AAV5 VP1 amino acid residues of the 581 to 589 region, where the one or more mutations comport to the rules or sequences in the following section.

A. Positional Frequency Rules

Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased skin tissue tropism as compared to wildtype AAV5 VP capsid polypeptide, wherein the engineered variant AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein said one or more mutations are selected from the following rules. In some embodiments, Xaa1 is selected from A, C, K, Q, R, or T. In some embodiments, Xaa1 is selected from C, K, or R. In some embodiments, Xaa1 is C. In some embodiments, Xaa2 is selected from A, C, I, S, T, or V. In some embodiments, Xaa2 is selected from A, S, T, or V. In some embodiments, Xaa2 is V. In some embodiments, Xaa3 is selected from A, C, F, G, M, Q, S, or V. In some embodiments, Xaa3 is selected from A, C, F, M, or Q. In some embodiments, Xaa3 is C. In some embodiments, Xaa4 is selected from C, K, L, P, R, or W. In some embodiments, Xaa4 is selected from L, P, or R. In some embodiments, Xaa4 is R. In some embodiments, Xaa5 is selected from F, H, I, M, V, or Y. In some embodiments, Xaa5 is selected from M, V, or Y. In some embodiments, Xaa5 is Y. In some embodiments, Xaa6 is selected from F, H, I, M, N, Q, or S. In some embodiments, Xaa6 is selected from M, N, or Q. In some embodiments, Xaa6 is N. In some embodiments, Xaa7 is selected from A, H, K, M, N, R, or V. In some embodiments, Xaa7 is A, H, K, or R. In some embodiments, Xaa7 is K. In some embodiments, Xaa8 is selected from A, F, G, H, S, or Y. In some embodiments, Xaa8 is selected from A, F, or S. In some embodiments, Xaa8 is S. In some embodiments, Xaa9 is selected from A, E, G, P, Q, R, or S. In some embodiments, Xaa9 is selected from A, Q, or S. In some embodiments, Xaa9 is A.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 31991-SEQ ID NO: 32990, wherein said at least one mutation drives increased skin tissue tropism.

B. ML Rules

For the following set of rules described in this paragraph, favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an engineered AAV5 VP1 capsid polypeptide, were determined using in vivo data and two ML models, which are described in EXAMPLE 40. Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased skin tissue tropism as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide, wherein the engineered AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the engineered AAV5 VP1 capsid polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein Xaa1 is selected from an amino acid of low surface accessibility (e.g., Xaa1 is selected from C); or wherein Xaa1 is selected from an amino acid of low volume (e.g., Xaa1 is selected from C); or wherein Xaa1 is selected from an amino acid of low mutability (e.g., Xaa1 is selected from C); or wherein Xaa2 is selected from an amino acid of high surface accessibility (e.g., Xaa2 is selected from R or K); or wherein Xaa2 is selected from an amino acid of high average flexibility (e.g., Xaa2 is selected from K, I, or N); or wherein Xaa2 is selected from an amino acid of low mutability (e.g., Xaa2 is selected from P or K); or wherein Xaa3 is selected from an amino acid of high hydropathy (e.g., Xaa3 is selected from I or V); or wherein Xaa4 is selected from an amino acid of low mutability (e.g., Xaa4 is selected from L, F, or Y); or wherein Xaa4 is selected from an amino acid of low average flexibility (e.g., Xaa4 is selected from W, H, F, or M); or wherein Xaa5 is selected from an amino acid of high average flexibility (e.g., Xaa5 is selected from G, R, K, I, or N); or wherein Xaa6 is selected from an amino acid of high average flexibility (e.g., Xaa6 is selected from G, R, K, I, or N); or wherein Xaa8 is selected from an amino acid of high surface accessibility (e.g., Xaa8 is selected from M, G, or F); or wherein Xaa8 is selected from an amino acid of low average flexibility (e.g., Xaa8 is selected from H, F, M, or W); or wherein Xaa8 is selected from an amino acid of low mutability (e.g., Xaa8 is selected from L, F, Y); or wherein Xaa9 is selected from an amino acid of high average flexibility (e.g., Xaa9 is selected from D, E, R, K, P, or G); or wherein Xaa9 is selected from an amino acid of high mutability (e.g., Xaa9 is selected from D, E, R, V, A, or H); or any combination thereof.

In some embodiments, Xaa1 is selected from an amino acid of low surface accessibility. In some embodiments, Xaa1 is selected from C. In some embodiments, Xaa1 is selected from an amino acid of low volume. In some embodiments, Xaa1 is selected from C. In some embodiments, Xaa1 is selected from an amino acid of low mutability. In some embodiments, Xaa1 is selected from C. In some embodiments, Xaa2 is selected from an amino acid of high surface accessibility. In some embodiments, Xaa2 is selected from R or K. In some embodiments, Xaa2 is selected from an amino acid of high average flexibility. In some embodiments, Xaa2 is selected from K, I, or N. In some embodiments, Xaa2 is selected from an amino acid of low mutability. In some embodiments, Xaa2 is selected from P or K. In some embodiments, Xaa3 is selected from an amino acid of high hydropathy. In some embodiments, Xaa3 is selected from 1 or V. In some embodiments, Xaa4 is selected from an amino acid of low mutability. In some embodiments, Xaa4 is selected from L, F, or Y. In some embodiments, Xaa4 is selected from an amino acid of low average flexibility. In some embodiments, Xaa4 is selected from W, H, F, or M. In some embodiments, Xaa5 is selected from an amino acid of high average flexibility. In some embodiments, Xaa5 is selected from G, R, K, I, or N. In some embodiments, Xaa6 is selected from an amino acid of high average flexibility. In some embodiments, Xaa6 is selected from G, R, K, I, or N. In some embodiments, Xaa8 is selected from an amino acid of high surface accessibility. In some embodiments, Xaa8 is selected from M, G. or F. In some embodiments, Xaa8 is selected from an amino acid of low average flexibility. In some embodiments, Xaa8 is selected from H, F, M, or W. In some embodiments, Xaa8 is selected from an amino acid of low mutability. In some embodiments, Xaa8 is selected from L, F, or Y. In some embodiments, Xaa9 is selected from an amino acid of high average flexibility. In some embodiments, Xaa9 is selected from D, E, R, K, P, or G. In some embodiments, Xaa9 is selected from an amino acid of high mutability. In some embodiments, Xaa9 is selected from D, E, R, V, A, or H.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 32991-SEQ ID NO: 33990, wherein said at least one mutation drives increased skin tissue tropism.

C. Enriched Skin Sequences

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 33991-SEQ ID NO: 34990, wherein said at least one mutation drives increased skin tissue tropism.

6.7.19. In Vivo Selected Mutated VP Polypeptides that Confer Increased Bone Marrow Tropism

The present disclosure provides AAV5 virions with a VP capsid polypeptide having at least one mutation in a region with residues that interact with target cells (e.g., a target bone marrow cell in a target bone marrow tissue of interest), where the at least one mutation confers increased bone marrow tissue tropism as compared to a wildtype VP capsid polypeptide. In some embodiments, provided herein are AAV5 VP1 capsid polypeptide having a sequence homology of at least 80% to SEQ ID NO: 1, wherein the AAV5 VP1 capsid polypeptide has at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of SEQ ID NO: 1 and wherein said at least one mutation drives increased bone marrow tropism. The following sequences rules and sequences also apply to the region in AAV5 VP2 (amino acid residues 445 to 453; VP2 sequence shown in SEQ ID NO: 1115) and AAV5 VP3 (amino acid residues 389 to 397; VP3 sequences shown in SEQ ID NO: 1116) corresponding to AAV5 VP1 amino acid residues 581 to 589. Thus, the present disclosure encompasses AAV5 VP2 capsid polypeptides and AAV5 VP3 capsid polypeptides having one or more mutations in the VP2 and VP3 regions corresponding to the AAV5 VP1 amino acid residues of the 581 to 589 region, where the one or more mutations comport to the rules or sequences in the following section.

A. Positional Frequency Rules

Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased bone marrow tissue tropism as compared to wildtype AAV5 VP capsid polypeptide, wherein the engineered variant AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the VP1 polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO. 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein said one or more mutations are selected from the following rules. In some embodiments, Xaa1 is selected from A, E, G, Q, S, or T. In some embodiments, Xaa1 is selected from A, E, or T. In some embodiments, Xaa1 is E. In some embodiments, Xaa2 is selected from A, I, Q, S, T, V, or Y. In some embodiments, Xaa2 is selected from A, S, T. In some embodiments, Xaa2 is A. In some embodiments, Xaa3 is selected from A, G, I, M, Q, S, or T. In some embodiments, Xaa3 is selected from A, Q, or T. In some embodiments, Xaa3 is Q. In some embodiments, Xaa4 is selected from A, E, P, Q, T, or V. In some embodiments, Xaa4 is selected from A, P, or Q. In some embodiments, Xaa4 is Q. In some embodiments, Xaa5 is selected from F, I, L, M, Q, V, or Y. In some embodiments, Xaa5 is selected from F, V, or Y. In some embodiments, Xaa5 is V. In some embodiments, Xaa6 is selected from F, I, N, Q, S, or V. In some embodiments, Xaa6 is selected from I, N, Q, or S. In some embodiments, Xaa6 is S. In some embodiments, Xaa7 is selected from A, C, M, S, or V. In some embodiments, Xaa7 is A, C, or V. In some embodiments, Xaa7 is C. In some embodiments, Xaa8 is selected from A, C, D, G, M, S, or Y. In some embodiments, Xaa8 is selected from A, M, S, or Y. In some embodiments, Xaa8 is M. In some embodiments, Xaa9 is selected from D, E, G, L, P, S, or Y. In some embodiments, Xaa9 is selected from D, E, or P. In some embodiments, Xaa9 is P.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 4118-SEQ ID NO: 5117, wherein said at least one mutation drives increased bone marrow tissue tropism.

B. ML Rules

For the following set of rules described in this paragraph, favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an engineered AAV5 VP1 capsid polypeptide, were determined using in vivo data and two ML models, which are described in EXAMPLE 32. Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased bone marrow tissue tropism as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide, wherein the engineered AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the engineered AAV5 VP1 capsid polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein Xaa1 is selected from an amino acid of high hydropathy (e.g., Xaa1 is selected from V, I, or L); or wherein Xaa1 is selected from an amino acid of low mutability (e.g., Xaa1 is selected from Y, L, F, or C); or wherein Xaa2 is selected from an amino acid of low hydropathy (e.g., Xaa2 is selected from Y or W); or wherein Xaa2 is selected from an amino acid of high mol mass (e.g., Xaa2 is selected from W); or wherein Xaa2 is selected from an amino acid of low surface accessibility (e.g., Xaa2 is selected from W or A); or wherein Xaa2 is selected from an amino acid of low hydrophilicity (e.g., Xaa2 is selected from W); or wherein Xaa2 is selected from an amino acid of low mutability (e.g., Xaa2 is selected from C); or wherein Xaa2 is selected from an amino acid of low average flexibility (e.g., Xaa2 is selected from W, M, or F); or wherein Xaa5 is selected from an amino acid of low average flexibility (e.g., Xaa5 is selected from W, M, or F); or wherein Xaa6 is selected from an amino acid of low average flexibility (e.g., Xaa6 is selected from W, M, or F); or wherein Xaa6 is selected from an amino acid of low mutability (e.g., Xaa6 is selected from Y, F, L, or C); or wherein Xaa6 is selected from an amino acid of high solubility (e.g., Xaa6 is selected from W, F, I, or L); or wherein Xaa7 is selected from an amino acid of low surface accessibility (e.g., Xaa7 is selected from C); or wherein Xaa7 is selected from an amino acid of high surface accessibility (e.g., Xaa7 is selected from D or N); or wherein Xaa7 is selected from an amino acid of low mutability (e.g., Xaa7 is selected from C); or wherein Xaa7 is selected from an amino acid of high solubility (e.g., Xaa7 is selected from C); or wherein Xaa7 is selected from an amino acid of low solubility (e.g., Xaa7 is selected from D); or wherein Xaa8 is selected from an amino acid of low charge (e.g., Xaa8 is selected from D or E); or wherein Xaa8 is selected from an amino acid of high mutability (e.g., Xaa8 is selected from D, E, A, or T); or wherein Xaa9 is selected from an amino acid of high mol mass (e.g., Xaa9 is selected from H or F); or wherein Xaa9 is selected from an amino acid of low mutability (e.g., Xaa9 is selected from Y, F, or L); or any combination thereof.

In some embodiments, Xaa1 is selected from an amino acid of high hydropathy. In some embodiments, Xaa1 is selected from V, I, or L. In some embodiments, Xaa1 is selected from an amino acid of low mutability. In some embodiments, Xaa1 is selected from Y, L, F, or C. In some embodiments, Xaa2 is selected from an amino acid of low hydropathy. In some embodiments, Xaa2 is selected from Y or W. In some embodiments, Xaa2 is selected from an amino acid of high mol mass. In some embodiments, Xaa2 is selected from W. In some embodiments, Xaa2 is selected from an amino acid of low surface accessibility. In some embodiments, Xaa2 is selected from W or A. In some embodiments, Xaa2 is selected from an amino acid of low hydrophilicity. In some embodiments, Xaa2 is selected from W. In some embodiments, Xaa2 is selected from an amino acid of low mutability. In some embodiments, Xaa2 is selected from C. In some embodiments, Xaa2 is selected from an amino acid of low average flexibility. In some embodiments, Xaa2 is selected from W, M, or F. In some embodiments, Xaa5 is selected from an amino acid of low average flexibility. In some embodiments, Xaa5 is selected from W, M, or F. In some embodiments, Xaa6 is selected from an amino acid of low average flexibility. In some embodiments, Xaa6 is selected from W, M, or F. In some embodiments, Xaa6 is selected from an amino acid of low mutability. In some embodiments, Xaa6 is selected from Y, F, L, or C. In some embodiments, Xaa6 is selected from an amino acid of high solubility. In some embodiments, Xaa6 is selected from W, F, I, or L. In some embodiments, Xaa7 is selected from an amino acid of low surface accessibility. In some embodiments, Xaa7 is selected from C. In some embodiments, Xaa7 is selected from an amino acid of high surface accessibility. In some embodiments, Xaa7 is selected from D or N. In some embodiments, Xaa7 is selected from an amino acid of low mutability. In some embodiments, Xaa7 is selected from C. In some embodiments, Xaa7 is selected from an amino acid of high solubility. In some embodiments, Xaa7 is selected from C. In some embodiments, Xaa7 is selected from an amino acid of low solubility. In some embodiments, Xaa7 is selected from D. In some embodiments, Xaa8 is selected from an amino acid of low charge. In some embodiments, Xaa8 is selected from D or E. In some embodiments, Xaa8 is selected from an amino acid of high mutability. In some embodiments, Xaa8 is selected from D, E, A, or T. In some embodiments, Xaa9 is selected from an amino acid of high mol mass. In some embodiments, Xaa9 is selected from H or F. In some embodiments, Xaa9 is selected from an amino acid of low mutability. In some embodiments, Xaa9 is selected from Y, F, or L.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 5118-SEQ ID NO: 6117, wherein said at least one mutation drives increased bone marrow tissue tropism.

C. Enriched Bone Marrow Sequences

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 6118-SEQ ID NO: 7117, wherein said at least one mutation drives increased bone marrow tissue tropism.

6.7.20. In Vivo Selected Mutated VP Polypeptides that Confer Increased Skeletal Muscle Tropism or Cardiac Muscle Tropism

A. ML Rules

For the following set of rules described in this paragraph, favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an engineered AAV5 VP1 capsid polypeptide, were determined using in vivo data and two ML models, which are described in EXAMPLE 22. Disclosed herein are engineered AAV5 VP capsid polypeptides capable of forming an assembled virion that exhibits increased skeletal muscle tissue tropism or cardiac muscle tissue tropism as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide, wherein the engineered AAV5 VP capsid polypeptide sequence has one or more mutations, wherein the engineered AAV5 VP1 capsid polypeptide sequence has said one or more mutations in a region from a position corresponding to 581 in SEQ ID NO: 2 to a position corresponding to 589 in SEQ ID NO: 2 and wherein Xaa1 is selected from an amino acid of low solubility (e.g., Xaa1 is selected from D, E, R, K, P, N, or Q); or wherein Xaa1 is selected from an amino acid of low hydropathy (e.g., Xaa1 is selected from D, E, R, K, Q, N, Y, or P); or wherein Xaa1 is selected from an amino acid of high surface accessibility (e.g., Xaa1 is selected from E, R, or K); or wherein Xaa2 is selected from an amino acid of high hydropathy (e.g., Xaa2 is selected from V, I, F, L, or C); or wherein Xaa2 is selected from an amino acid of low mutability (e.g., Xaa2 is selected from R, V, I, H, or C); or wherein Xaa2 is selected from an amino acid of medium volume (e.g., Xaa2 is selected from E, V, or Q); or wherein Xaa3 is selected from an amino acid of low solubility (e.g., Xaa3 is selected from D, R, or Q); or wherein Xaa4 is selected from an amino acid of low solubility (e.g., Xaa4 is selected from D, E, P, or N); or wherein Xaa4 is selected from an amino acid of low charge (e.g., Xaa4 is selected from D or E); or wherein Xaa5 is selected from an amino acid of low amino acid solubility (e.g., Xaa5 is selected from D, E, R, K, N, or Q); or wherein Xaa8 is selected from an amino acid of low solubility (e.g., Xaa8 is selected from D, E, K, P, or N); or wherein Xaa8 is selected from an amino acid of high flexibility index (e.g., Xaa8 is selected from Q, S, P, E, or D); or wherein Xaa8 is selected from an amino acid of high surface accessibility (e.g., Xaa8 is selected from S, D, P, N, E, R, or K); or any combination thereof.

In some embodiments, Xaa1 is selected from an amino acid of low solubility. In some embodiments, Xaa1 is selected from D, E, R, K, P, N, or Q. In some embodiments, Xaa1 is selected from an amino acid of low hydropathy. In some embodiments, Xaa1 is selected from D, E, R, K, Q, N, Y, or P. In some embodiments, Xaa1 is selected from an amino acid of high surface accessibility. In some embodiments, Xaa1 is selected from E, R, or K. In some embodiments, Xaa2 is selected from an amino acid of high hydropathy. In some embodiments, Xaa2 is selected from V, I, F, L, or C. In some embodiments, Xaa2 is selected from an amino acid of low mutability. In some embodiments, Xaa2 is selected from R, V, I, H, or C. In some embodiments, Xaa2 is selected from an amino acid of medium volume. In some embodiments, Xaa2 is selected from E, V, or Q. In some embodiments, Xaa3 is selected from an amino acid of low solubility. In some embodiments, Xaa3 is selected from D, R, or Q. In some embodiments, Xaa4 is selected from an amino acid of low solubility. In some embodiments, Xaa4 is selected from D, E, P, or N. In some embodiments, Xaa4 is selected from an amino acid of low charge. In some embodiments, Xaa4 is selected from D or E. In some embodiments, Xaa5 is selected from an amino acid of low amino acid solubility. In some embodiments, Xaa5 is selected from D, E, R, K, N, or Q. In some embodiments, Xaa8 is selected from an amino acid of low solubility. In some embodiments, Xaa8 is selected from D, E, K, P, or N. In some embodiments, Xaa8 is selected from an amino acid of high flexibility index. In some embodiments, Xaa8 is selected from Q, S, P, E, or D. In some embodiments, Xaa8 is selected from an amino acid of high surface accessibility. In some embodiments, Xaa8 is selected from S, D, P, N, E, R, or K.

In some embodiments, provided herein are AAV5 VP capsid polypeptide having at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 of AAV5 VP1 and having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 99%, or 100% sequence identity to any sequence selected from SEQ ID NO: 25118-SEQ ID NO: 26117, wherein said at least one mutation drives increased skeletal muscle tissue tropism or cardiac muscle tissue tropism.

1.1. Numbered Embodiments

A number of compositions, and methods are disclosed herein. Specific exemplary embodiments of these compositions and methods are disclosed below. The following embodiments recite non-limiting permutations of combinations of features disclosed herein. Other permutations of combinations of features are also contemplated. In particular, each of these numbered embodiments is contemplated as depending from or relating to every previous or subsequent numbered embodiment, independent of their order as listed.

In a further aspect, the following embodiments are provided. All numerical references to a preceding embodiment refer to the embodiment so numbered within the same subsection. In yet a further aspect, rAAV comprising the recombinant or engineered VP capsid polypeptides of the following numbered embodiments are provided, as are methods of using pharmaceutical compositions comprising the rAAV for treatment of a subject in need thereof.

Series A Embodiments

In the Series A embodiments, “recombinant” adeno-associated (AAV) VP1 capsid polypeptide is synonymous with “engineered” adeno-associated (AAV) VP1 capsid polypeptide.

1. A recombinant adeno-associated virus (AAV) VP1 capsid polypeptide having the amino acid sequence of SEQ ID NO:2, wherein Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V; and wherein the polypeptide does not have the sequence of any of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, and SEQ ID NO:8. 2. A recombinant adeno-associated virus (AAV) VP1 capsid polypeptide having at least one mutation in a residue corresponding to residue 581 to residue 589 in SEQ ID NO: 1, wherein the mutation confers tissue tropism for a first tissue as compared to a second tissue and wherein the AAV VP1 capsid polypeptide does not have the sequence of any of SEQ ID NO:1. SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, and SEQ ID NO:8. 3. The recombinant AAV VP1 capsid polypeptide of embodiment 2, wherein the AAV VP1 capsid polypeptide is an AAV5 VP1 capsid polypeptide. 4. The recombinant AAV VP1 capsid polypeptide of any one of embodiments 1-3, wherein a specific order of the residues at residue 581-589 corresponding to SEQ ID NO: 1 results in a specific tissue tropism. 5. The recombinant AAV VP1 capsid polypeptide of any one of embodiments 1-4, wherein the first tissue is selected from adipose, adrenal gland, aorta, brain (including hippocampus: dentate gyrus, CA1 and CA3; cerebellum, caudate, putamen, midbrain, pons, hypothalamus, cortex-including occipital, temporal and forebrain; substantia nigra, and thalamus), bone marrow, cecum, colon, dorsal root ganglion, duodenum, epididymis, esophagus, eye, gallbladder, heart, ileum, jejunum, kidney, lung, lymph nodes, mammary gland, ovary, pancreas, parathyroid gland, peripheral nerve, pituitary, prostate, salivary gland, seminal vesicle, skeletal muscle, skin, spinal cord, spleen, stomach, testis, thymus, thyroid, trachea, urinary bladder, uterus, and vagina; wherein the second tissue is selected from: adipose, adrenal gland, aorta, brain (including hippocampus: dentate gyrus, CA1 and CA3; cerebellum, caudate, putamen, midbrain, pons, hypothalamus, cortex-including occipital, temporal and forebrain; substantia nigra, and thalamus), bone marrow, cecum, colon, dorsal root ganglion, duodenum, epididymis, esophagus, eye, gallbladder, heart, ileum, jejunum, kidney, lung, lymph nodes, mammary gland, ovary, pancreas, parathyroid gland, peripheral nerve, pituitary, prostate, salivary gland, seminal vesicle, skeletal muscle, skin, spinal cord, spleen, stomach, testis, thymus, thyroid, trachea, urinary bladder, uterus, and vagina.; and wherein the first tissue and the second tissue are different. 6. The recombinant AAV VP1 capsid polypeptide of any one of embodiments 2-4, wherein the rAAV has increased ability to infect a tissue selected from adipose, adrenal gland, aorta, brain (including hippocampus: dentate gyrus, CA1 and CA3; cerebellum, caudate, putamen, midbrain, pons, hypothalamus, cortex-including occipital, temporal and forebrain; substantia nigra, and thalamus), bone marrow, cecum, colon, dorsal root ganglion, duodenum, epididymis, esophagus, eye, gallbladder, heart, ileum, jejunum, kidney, lung, lymph nodes, mammary gland, ovary, pancreas, parathyroid gland, peripheral nerve, pituitary, prostate, salivary gland, seminal vesicle, skeletal muscle, skin, spinal cord, spleen, stomach, testis, thymus, thyroid, trachea, urinary bladder, uterus, and vagina. following intravenous administration as compared to a rAAV having a VP1 capsid polypeptide of SEQ ID NO:1. 7. The recombinant AAV VP1 capsid poly peptide of any one of embodiments 2-6, wherein the rAAV exhibits from about a 1.0005-fold to about a 1000-fold increased accumulation in the first tissue as compared to the second tissue. 8. The recombinant AAV VP1 capsid polypeptide of any one of embodiments 2-6, wherein the rAAV exhibits at least about a 1.0005-fold, at least about a two-fold, at least about a three-fold, at least about a four-fold, at least about a five-fold, at least about a ten-fold, at least about a twenty-fold, at least about a 50-fold, at least about a 75-fold, at least about a 100-fold, or at least about a 1000-fold increased accumulation in the first tissue as compared to the second tissue. 9. The recombinant capsid polypeptide of any preceding embodiment, further comprising one or more mutations at an amino acid residue outside of the 581-589 region, wherein the one or more mutations at an amino acid residue outside of the 581-589 region confers improved manufacturability, improved viral assembly, improved tissue targeting/tropism, or any combination thereof. 10. The recombinant AAV VP1 capsid polypeptide of any of embodiments 1-9, wherein Xaa1 is selected from A, G, K, M, N, Q, R, S, or T. 11. The recombinant capsid polypeptide of any preceding embodiment, wherein Xaa1 is selected from A, K, M, or T. 12. The recombinant capsid polypeptide of any preceding embodiment, wherein Xaa1 is K. 13. The recombinant capsid polypeptide of any preceding embodiment, wherein Xaa2 is selected from A, C, H, I, K, S, T, or V. 14. The recombinant capsid polypeptide of any preceding embodiment, wherein Xaa2 is selected from A, S, T, or V. 15. The recombinant capsid polypeptide of any preceding embodiment, wherein Xaa2 is T. 16. The recombinant capsid poly peptide of any preceding embodiment, wherein Xaa3 is selected from A, G, H, K, M, N, Q, R, S, T, or V. 17. The recombinant capsid polypeptide of any preceding embodiment, wherein Xaa3 is selected from A, M, or T. 18. The recombinant capsid polypeptide of any preceding embodiment, wherein Xaa3 comprises A or T. 19. The recombinant capsid polypeptide of any preceding embodiment, wherein Xaa4 is selected from L, M, P, Q, R, T, or W. 20. The recombinant capsid polypeptide of any preceding embodiment, wherein Xaa4 is selected from L, P, Q, or T. 21. The recombinant capsid polypeptide of any preceding embodiment, wherein Xaa4 is P; 22. The recombinant capsid polypeptide of any preceding embodiment, wherein Xaa5 is selected from F, H, I, K, M, T, or Y. 23. The recombinant capsid polypeptide of any preceding embodiment, wherein Xaa5 is selected from H, I, or Y. 24. The recombinant capsid polypeptide of any preceding embodiment, wherein Xaa5 is Y. 25. The recombinant capsid polypeptide of any preceding embodiment, wherein Xaa6 is selected from E, G, H, L, M, N, Q, T, or W. 26. The recombinant capsid polypeptide of any preceding embodiment, wherein Xaa6 is selected from N, or Q. 27. The recombinant capsid polypeptide of any preceding embodiment, wherein Xaa6 is N. 28. The recombinant capsid polypeptide of any preceding embodiment, wherein Xaa7 is selected from A, C, G, H, L, M, R or S. 29. The recombinant capsid polypeptide of any preceding embodiment, wherein Xaa7 is selected from A, C, H or M. 30. The recombinant capsid polypeptide of any preceding embodiment, wherein Xaa7 is A. 31. The recombinant capsid poly peptide of any preceding embodiment, wherein Xaa8 is selected from A, C, D, F, G, H, M, Q, S, V, W, or Y. 32. The recombinant capsid polypeptide of any preceding embodiment, wherein Xaa8 is selected from G, M, Q, or S. 33. The recombinant capsid polypeptide of any preceding embodiment, wherein Xaa8 is G. 34. The recombinant capsid polypeptide of any preceding embodiment, wherein Xaa9 is selected from A, C, E, G, H, M, N, P, Q, S, V, or W. 35. The recombinant capsid polypeptide of any preceding embodiment, wherein Xaa9 is selected from E, G, or P. 36. The recombinant capsid polypeptide of any preceding embodiment, wherein Xaa9 is G. 37. The recombinant capsid polypeptide of any of embodiments 1-9, wherein Xaa1 is selected from A, D, E, G, L, M, N, Q, S, T, or V. 38. The recombinant capsid polypeptide of embodiment 37, wherein Xaa1 is selected from A, D, E, M, or T. 39. The recombinant capsid polypeptide of embodiment 37, wherein Xaa1 is E. 40. The recombinant capsid polypeptide of any of embodiments 37-39, wherein Xaa2 is selected from A, C, D, E, G, H, I, N, P, Q, S, T, or V. 41. The recombinant capsid polypeptide of any of embodiments 37-40, wherein Xaa2 is selected from A, S, T, or V. 42. The recombinant capsid polypeptide of any of embodiments 37-41, wherein Xaa2 is A. 43. The recombinant capsid polypeptide of any of embodiments 37-42, wherein Xaa3 is selected from A, D, E, G, H, M, N, Q, S, T, or V. 44. The recombinant capsid polypeptide of any of embodiments 37-43, wherein Xaa3 is selected from D, E, N, Q or T. 45 The recombinant capsid polypeptide of any of embodiments 37-44, wherein Xaa3 is D or T. 46. The recombinant capsid polypeptide of any of embodiments 37-45, wherein Xaa4 is selected from A, D, E, G, H, N, P, Q, S, or T. 47. The recombinant capsid polypeptide of any of embodiments 37-46, wherein Xaa4 is selected from D, E, P, or Q. 48. The recombinant capsid polypeptide of any of embodiments 37-47, wherein Xaa4 is E. 49. The recombinant capsid polypeptide of any of embodiments 37-48, wherein Xaa5 is selected from A, C, D, E, G, H, N, Q, S, T, or Y. 50. The recombinant capsid polypeptide of any of embodiments 37-49, wherein Xaa5 is selected from D, E, N, Q or T. 51. The recombinant capsid polypeptide of any of embodiments 37-50, wherein Xaa5 is N. 52. The recombinant capsid polypeptide of any of embodiments 37-51, wherein Xaa6 is selected from A, D, E, G, H, N, P, Q, S, or T. 53. The recombinant capsid polypeptide of any of embodiments 37-52, wherein Xaa6 is selected from D, N, or Q. 54. The recombinant capsid polypeptide of any of embodiments 37-53, wherein Xaa6 is D. 55. The recombinant capsid polypeptide of any of embodiments 37-54, wherein Xaa7 is selected from A, C, D, E, G, H, N, Q, S, or T. 56. The recombinant capsid polypeptide of any of embodiments 37-55, wherein Xaa7 is selected from A. D. E or G. 57. The recombinant capsid polypeptide of any of embodiments 37-56, wherein Xaa7 is A. 58. The recombinant capsid polypeptide of any of embodiments 37-57, wherein Xaa8 is selected from A, C, D, E, G, H, N, Q, S, or T. 59. The recombinant capsid polypeptide of any of embodiments 37-58, wherein Xaa8 comprises A, D, G, or S. 60. The recombinant capsid polypeptide of any of embodiments 37-59, wherein Xaa8 is G. 61. The recombinant capsid polypeptide of any of embodiments 37-60, wherein Xaa9 is selected from A, D, E, G, H, N, P, Q, S, or T. 62. The recombinant capsid polypeptide of any of embodiments 37-61, wherein Xaa9 is selected from A, D, G, or P. 63. The recombinant capsid polypeptide of any of embodiments 37-62, wherein Xaa9 is G. 64. A recombinant capsid polypeptide of any of embodiments 1-36 combined with the recombinant capsid polypeptide of any of embodiments 37-63, wherein the VP1 capsid is capable of forming an assembled virion that exhibits increased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid polypeptide of SEQ ID NO:1. 65. The recombinant capsid polypeptide of embodiment 1, wherein Xaa1 is not K. and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 66. The recombinant capsid polypeptide of embodiment 1, wherein Xaa1 is not A, K, M, or T, and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 67. The recombinant capsid polypeptide of embodiment 1, wherein Xaa1 is not A, G, K, M, N, Q, R, S, or T, and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 68. The recombinant capsid polypeptide of embodiment 1, or any of embodiments 65-67, wherein Xaa2 is not T, and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 69. The recombinant capsid polypeptide of embodiment 1, or any of embodiments 65-68, wherein Xaa2 is not A, S, T, or V, and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 70. The recombinant capsid polypeptide of embodiment 1, or any of embodiments 65-69, wherein Xaa2 is not A, C, H, I, K, S, T, or V, and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 71. The recombinant capsid polypeptide of embodiment 1, or any of embodiments 65-70, wherein Xaa3 is not A or T, and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 72. The recombinant capsid polypeptide of embodiment 1, or any of embodiments 65-71, Xaa3 is not A, M, or T, and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 73. The recombinant capsid polypeptide of embodiment 1, or any of embodiments 65-72, wherein Xaa3 is not A, G, H, K, M, N, Q, R, S, T, or V, and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 74. The recombinant capsid polypeptide of embodiment 1, or any of embodiments 65-73, wherein Xaa4 is not P, and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 75. The recombinant capsid polypeptide of embodiment 1, or any of embodiments 65-74, wherein Xaa4 is not L, P, Q, or T, and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 76. The recombinant capsid polypeptide of embodiment 1, or any of embodiments 65-75, wherein Xaa4 is not L, M, P, Q, R, T, or W, and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 77. The recombinant capsid polypeptide of embodiment 1, or any of embodiments 65-76, wherein Xaa5 is not Y, and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 78. The recombinant capsid polypeptide of embodiment 1, or any of embodiments 65-77, wherein Xaa5 is not H, I, or Y, and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 79. The recombinant capsid polypeptide of embodiment 1, or any of embodiments 65-78, wherein Xaa5 is not F, H, I, K, M, T, or Y, and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 80. The recombinant capsid polypeptide of embodiment 1, or any of embodiments 65-79, wherein Xaa6 is not N. and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 81. The recombinant capsid polypeptide of embodiment 1, or any of embodiments 65-80, wherein Xaa6 is not N, or Q. and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 82. The recombinant capsid polypeptide of embodiment 1, or any of embodiments 65-81, wherein Xaa6 is not E, G, H, L, M, N, Q, T, or W, and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 83. The recombinant capsid polypeptide of embodiment 1, or any of embodiments 65-82, wherein Xaa7 is not A, and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 84. The recombinant capsid polypeptide of embodiment 1, or any of embodiments 65-83, wherein Xaa7 is not A, C, H or M, and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 85. The recombinant capsid polypeptide of embodiment 1, or any of embodiments 65-84, wherein Xaa7 is not A, C, G, H, L, M, R or S, and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 86. The recombinant capsid polypeptide of embodiment 1, or any of embodiments 65-85, wherein Xaa8 is not G, and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 87. The recombinant capsid polypeptide of embodiment 1, or any of embodiments 65-86, wherein Xaa8 is not G, M, Q, or S, and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 88. The recombinant capsid polypeptide of embodiment 1, or any of embodiments 65-87, wherein Xaa8 is not A, C, D, F, G, H, M, Q, S, V, W, or Y. and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 89. The recombinant capsid polypeptide of embodiment 1, or any of embodiments 65-88, Xaa9 is not G. and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 90. The recombinant capsid polypeptide of embodiment 1, or any of embodiments 65-89, wherein Xaa9 is not E, G, or P, and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue w % ben compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 91. The recombinant capsid polypeptide of embodiment 1, or any of embodiments 65-90, wherein Xaa9 is not A, C, E, G, H, M, N, P, Q, S, V, or W, and wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue when compared to a virion that comprises the AAV5 VP1 capsid of SEQ ID NO:1. 92. A recombinant capsid polypeptide of any of embodiments 65-91 combined with the recombinant capsid polypeptide of any of embodiments 37-63, wherein the VP1 capsid is capable of forming an assembled virion that exhibits decreased tropism for liver tissue w % ben compared to a virion that comprises the AAV5 VP1 capsid polypeptide of SEQ ID NO:1. 93. A recombinant adeno-associated virus (AAV) VP1 capsid polypeptide having at least one residue corresponding to residue 581 to residue 589 in SEQ ID NO: 2, wherein the at least one residue is: Xaa1 and Xaa1 is selected from A, G, K, M, N, Q, R, S, or T; Xaa2 and Xaa2 is selected from A, C, H, I, K, S, T, or V; Xaa3 and Xaa3 is selected from A, G, H, K, M, N, Q, R, S, T, or V; Xaa4 and Xaa4 is selected from L, M, P, Q, R, T, or W; Xaa5 and Xaa5 is selected from F, H, I, K, M, T, or Y; Xaa6 and Xaa6 is selected from E, G, H, L, M, N, Q, T, or W; Xaa7 and Xaa7 is selected from A, C, G, H, L, M, R or S; Xaa8 and Xaa8 is selected from A, C, D, F, G, H, M, Q, S, V, W, or Y; Xaa9 and Xaa9 is selected from A, C, E, G, H, M, N, P, Q, S, V, or W; or any combination thereof, wherein the AAV VP1 capsid polypeptide is capable of exhibiting tissue tropism for liver tissue. 94. A recombinant adeno-associated virus AAV VP1 capsid polypeptide having at least one mutation in a residue of region 581 to residue 589 in SEQ ID NO: 1, wherein the mutation confers at least about a two-fold increased accumulation in a non-liver tissue as compared to a liver tissue, as compared to AAV5 VP1, and wherein the AAV VP1 capsid polypeptide does not have the sequence of any of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, and SEQ ID NO:8. 95. The recombinant AAVVP1 capsid polypeptide of embodiment 94, wherein the mutation confers at least about a three-fold, at least about a four-fold, at least about a five-fold, at least about a ten-fold, at least about a twenty-fold, at least about a fifty-fold, at least about a 75-fold, at least about a 100-fold increased accumulation in a non-liver tissue as compared to a liver tissue. 96. The recombinant AAVVP1 capsid poly peptide of embodiment 94, wherein the mutation confers from about a 1.0005-fold to about a 1000-fold increased accumulation in a non-liver tissue as compared to a liver tissue. 97. The recombinant capsid polypeptide of any preceding embodiment, further comprising one or more mutations at an amino acid residue outside of the 581-589 region, with reference to SEQ ID NO:1, wherein the resulting recombinant capsid is capable of forming an assembled virion that exhibits desired tissue targeting/tropism. 98. The recombinant capsid polypeptide of embodiment 97, wherein the one or more mutations at an amino acid residue outside of the 581-589 region confers improved manufacturability, improved viral assembly, improved tissue targeting/tropism, or any combination thereof. 99. A vector capable of replication in prokaryotic cells, wherein the vector comprises a polynucleotide encoding the recombinant capsid polypeptide of any preceding embodiment. 100. The vector of embodiment 99, wherein the vector is a plasmid. 101. A library comprising a plurality of plasmids of embodiment 100, the plurality of plasmids comprising a plurality of different AAV VP1-encoding polynucleotides. 102. The plasmid library of embodiment 101, wherein the library encodes at least 1×109 different AAV VP1 capsid polypeptides. 103. The plasmid library of embodiment 102, wherein the library encodes at least 5×109 different AAV VP1 capsid polypeptides. 104. The plasmid library of embodiment 103, wherein the library encodes at least 1×1010 different AAV VP1 capsid polypeptides. 105. The plasmid library of embodiment 104, wherein the library encodes at least 5×1010 different AAV VP1 capsid polypeptides. 106. The plasmid library of embodiment 105, wherein the library encodes at least 7.5×1010 different AAV VP1 capsid polypeptides. 107. The plasmid library of embodiment 106, wherein the library encodes at least 1×1011 different AAV VP1 capsid polypeptides. 108. The plasmid library of embodiment 107, wherein the library encodes at least 2.5×1011 different AAV VP1 capsid polypeptides. 109. The plasmid library of embodiment 108, wherein the library encodes at least 5×1011 different AAV VP1 capsid polypeptides. 110. A prokaryotic cell comprising the vector of embodiment 100. 111. The prokaryotic cell of embodiment 110, wherein prokaryotic cell is an E. coli cell and the vector is a plasmid. 112. A library comprising a plurality of E. coli cells of embodiment 111, wherein the plurality of cells comprises a plurality of plasmids, wherein the plurality of plasmids comprises a plurality of different AAV VP1-encoding polynucleotides. 113. A library comprising a plurality of polypeptides of any of embodiments 1-98, the plurality having different primary amino acid sequences. 114. The library of embodiment 113, wherein the library comprises at least from about 1×105 to at least about 5×1011 different AAV VP1 capsid polypeptides. 115. A recombinant AAV virion (rAAV), the virion comprising an AAV VP1 capsid polypeptide of any of embodiments 1-98. 116. The rAAV virion of embodiment 115, wherein the rAAV has reduced tropism for human liver as compared to a rAAV having a VP1 capsid polypeptide having the sequence of SEQ ID NO:1. 117. The rAAV virion of embodiment 115 or embodiment 116, wherein the rAAV has increased ability to cross the blood-brain barrier following intravenous administration as compared to a rAAV having a VP1 capsid polypeptide having the sequence of SEQ ID NO: 1. 118. The rAAV virion of any one of embodiments 115-117, wherein the rAAV has increased ability to infect one or more brain regions selected from hippocampus, dentate gyrus, cerebral cortex, temporal cortex, occipital cortex, thalamus, forebrain, substantia nigra, hypothalamus, and cerebellum, following intravenous, intrathecal, intracerebral ventricular, or intracisternal magna administration as compared to a rAAV having a VP1 capsid polypeptide having the sequence of SEQ ID NO:1. 119. The rAAV virion of any one of embodiments 115-118, wherein the rAAV has increased ability to infect human retinal cells following intravitreal injection as compared to a rAAV having a VP1 capsid polypeptide having the sequence of SEQ ID NO:1. 120. The rAAV virion of any one of embodiments 115-119, wherein the rAAV has increased ability to infect human skeletal muscle following intravenous administration as compared to a rAAV having a VP1 capsid polypeptide having the sequence of SEQ ID NO:1. 121. The rAAV virion of any one of embodiments 115, and 117-120, wherein the rAAV has increased tropism for human liver as compared to a rAAV having a VP1 capsid polypeptide having the sequence of SEQ ID NO:1. 122. The rAAV virion of embodiment 115, 116, or 121, wherein the rAAV has increased ability to infect a tissue selected from adipose, adrenal gland, aorta, brain (including hippocampus: dentate gyrus, CA1 and CA3; cerebellum, caudate, putamen, midbrain, pons, hypothalamus, cortex-including occipital, temporal and forebrain; substantia nigra, and thalamus), bone marrow, cecum, colon, dorsal root ganglion, duodenum, epididymis, esophagus, eye, gallbladder, heart, ileum, jejunum, kidney, lung, lymph nodes, mammary gland, ovary, pancreas, parathyroid gland, peripheral nerve, pituitary, prostate, salivary gland, seminal vesicle, skeletal muscle, skin, spinal cord, spleen, stomach, testis, thymus, thyroid, trachea, urinary bladder, uterus, and vagina, following intravenous administration as compared to a rAAV having a VP1 capsid polypeptide of SEQ ID NO:1. 123. The rAAV virion of any one of embodiments 115-122, wherein the virion further comprises a vector genome, the vector genome comprising a therapeutic polynucleotide encoding any of the following: a therapeutic RNA selected from a guide RNA or a tRNA, or transgene encoding a protein under control of regulatory sequences that direct transgene expression in infected human cells. 124. The rAAV virion of embodiment 123, wherein the transgene encodes a protein selected from the transgene products of Table 1. 125. A library comprising a plurality of rAAV virions of any one of embodiments 115-124, wherein the plurality of rAAV virions comprise a plurality of VP1 capsid polypeptides with different primary amino acid sequences. 126. The library of embodiment 125, wherein the library comprises at least about 1×10⁵to at least about 5×10¹¹different AAV VP1 capsid polypeptides different AAV VP1 capsid polypeptides. 127. A pharmaceutical composition comprising the rAAV of embodiment 123 or embodiment 124 and a pharmaceutically acceptable carrier. 128. A method of treatment, comprising: administering an effective amount of the pharmaceutical composition of embodiment 127 to a patient in need thereof. 129. The method of embodiment 128, wherein the effective amount of the rAAV is less than the effective amount of a wild type rAAV. 130. The method of embodiment 128, wherein the effective amount of the rAAV is less than the effective amount of an otherwise comparable rAAV lacking one or more than one mutation at a position corresponding to residue 581 to residue 589 of SEQ ID NO: 1. 131. The method of any one of embodiments 128-130, wherein the effective amount of the results in lower toxicity in the patient as compared to the effective amount of the wild type rAAV, the otherwise comparable rAAV, or both. 132. The method of embodiment 128, wherein the effective amount is at least from 1×10⁵viral genomes/kg patient weight to 5×10¹⁴viral genomes/kg. 133. The method of any one of embodiments 128-132, wherein the rAAV is administered intravenously. 134. The method of any one of embodiments 128-132, wherein the rAAV is administered intrathecally. 135. The method of any one of embodiments 128-132, wherein the rAAV is administered by intracisternal magna administration. 136. The method of any one of embodiments 128-132, wherein the rAAV is administered by intravitreal injection. 137. A method of identifying an AAV VP1 capsid polypeptide that confers tropism for a desired tissue, comprising: administering an aliquot of the library of any one of embodiments 101-109, or 112-144, or 125-126 to a non-human primate; and identifying the sequences of AAV capsid sequence of rAAV that had infected the desired tissue. 138. The method of embodiment 137, wherein the library aliquot is administered intravenously. 139. The method of embodiment 137 wherein the library aliquot is administered intrathecally. 140. The method of embodiment 137, wherein the library aliquot is administered by intra-cisterna magna administration. 141. The method of embodiment 137, wherein the library aliquot is administered by intracerebral ventricular injection. 142. A method of formulating the therapeutic polynucleotide of any one of embodiments 123-124 in a virion, the method comprising: transfecting a cell with plasmid encoding for the recombinant capsid polypeptide of any one of embodiments 1-98 and transfecting the cell with a plasmid encoding for the therapeutic polynucleotide, wherein upon transfection, the cell produces the virion within which is packaged the therapeutic polynucleotide. 143. A composition comprising an AAV virion comprising the recombinant capsid polypeptide of any one of embodiments 1-98 within which is packaged the therapeutic polynucleotide of any one of embodiments 123-124. 144. A recombinant AAV VP1 capsid polypeptide having any one of the VP1 capsid mutations recited in Table 8 (SEQ ID NO:115-1114), and wherein the VP1 capsid polypeptide does not have the sequence of any of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, and SEQ ID NO:8. 145. A recombinant AAV VP1 capsid polypeptide having any of the VP1 capsid mutations recited in Table 8 (SEQ ID NO:115-1114), wherein the mutation confers tissue tropism for a first tissue as compared to a second tissue and wherein the AAV VP1 capsid polypeptide does not have the sequence of any of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4. SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, and SEQ ID NO:8. 146. The recombinant AAV VP1 capsid polypeptide of embodiment 145, wherein the mutation confers at least about a two-fold increased accumulation of rAAV comprising the mutated VP1 protein in a non-liver tissue as compared to a liver tissue as compared to accumulation of rAAV comprising AAV5 VP1 (SEQ ID NO:1), wherein the mutated rAAV and AAV5 rAAV are each administered intravenously at the same titer, and wherein the VP1 capsid polypeptide does not have the sequence of any of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, and SEQ ID NO:8. 147. A composition comprising an AAV virion comprising the recombinant capsid polypeptide of any one of embodiments 144-146, within which is packaged a therapeutic polynucleotide encoding any of the following: a therapeutic RNA selected from a guide RNA or a tRNA, or transgene encoding a protein under control of regulatory sequences that direct transgene expression in infected human cells.

Series B Embodiments—CNS Tropic Capsids

1. An engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide having an amino acid sequence at least 70% identical to SEQ ID NO: 1,

wherein the engineered AAV VP capsid polypeptide has at least one mutation as compared to SEQ ID NO: 1 in the region from a residue corresponding to residue 581 of SEQ ID NO: 1 to a residue corresponding to residue 589 of SEQ ID NO: 1, inclusive,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV),

wherein the at least one mutation confers higher tropism for a central nervous system (CNS) tissue on the rAAV as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1, and

wherein the engineered AAV VP capsid polypeptide does not have the sequence of any of SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8. 2. The engineered AAV VP capsid polypeptide of embodiment 1, wherein the engineered AAV VP capsid polypeptide has a sequence of SEQ ID NO: 2 and wherein Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from any amino acid. 3. An engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide having an amino acid sequence of SEQ ID NO: 2,

wherein amino acid residues Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV), and

wherein the engineered AAV VP capsid polypeptide does not have the sequence of any of SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8. 4. The engineered AAV VP capsid polypeptide of embodiment 3, wherein the rAAV has higher tropism for a central nervous system (CNS) tissue as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1. 5. The engineered AAV VP capsid polypeptide of any one of embodiments 1-2, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 7118-SEQ ID NO: 10117. 6. The engineered AAV VP capsid polypeptide of embodiment 4, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence of any one of SEQ ID NO: 7118-SEQ ID NO: 10117. 7. The engineered AAV VP capsid polypeptide of embodiment 3, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 7118-SEQ ID NO: 10117. 8. The engineered AAV VP capsid polypeptide of embodiment 6, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence of any one of SEQ ID NO: 7118-SEQ ID NO: 10117.9. The engineered AAV VP capsid polypeptide of any one of embodiments 2-3, wherein:

Xaa1 is selected from the group consisting of A, C, K, M, Q, R, T, and W; or

Xaa2 is selected from the group consisting of F, I, K, R, T, and W; or

Xaa3 is selected from the group consisting of A, H, N, R, and W; or

Xaa4 is selected from the group consisting of E, G, I, M, Q, and R; or

Xaa5 is selected from the group consisting of C, G, K, I, M, and R; or

Xaa6 is selected from the group consisting of I, K, L, P, Q, R, and Y; or

Xaa7 is selected from the group consisting of D, I, K, R, V, and W; or

Xaa8 is selected from the group consisting of C, G, H, K, L, and V; or

Xaa9 is selected from the group consisting of I, K, L, R, and V; or

any combination thereof.

10. The engineered AAV VP capsid polypeptide of embodiment 5, wherein Xaa1 is selected from K, Q, R, or W. 11. The engineered AAV VP capsid polypeptide of embodiment 5, wherein Xaa2 is selected from F, I, R or T. 12. The engineered AAV VP capsid polypeptide of embodiment 5, wherein Xaa3 is selected from A, R, or W. 13. The engineered AAV VP capsid polypeptide of embodiment 5, wherein Xaa4 is selected from E, M, or R. 14. The engineered AAV VP capsid polypeptide of embodiment 5, wherein Xaa5 is selected from K, I, or R. 15. The engineered AAV VP capsid polypeptide of embodiment 5, wherein Xaa6 is selected from K, R, or Y. 16. The engineered AAV VP capsid polypeptide of embodiment 5, wherein Xaa7 is selected from I, R, or V. 17. The engineered AAV VP capsid polypeptide of embodiment 5, wherein Xaa8 is selected from H, K, or V. 18. The engineered AAV VP capsid polypeptide of embodiment 5, wherein Xaa9 is selected from I, K, or R. 19. The engineered AAV VP capsid polypeptide of embodiment 5, wherein Xaa1 is K. 20. The engineered AAV VP capsid polypeptide of embodiment 5, wherein Xaa2 is R. 21. The engineered AAV VP capsid polypeptide of embodiment 5, wherein Xaa3 is R. 22. The engineered AAV VP capsid polypeptide of embodiment 5, wherein Xaa4 is R. 23. The engineered AAV VP capsid polypeptide of embodiment 5, wherein Xaa5 is I. 24. The engineered AAV VP capsid polypeptide of embodiment 5, wherein Xaa6 is R. 25. The engineered AAV VP capsid polypeptide of embodiment 5, wherein Xaa7 is V. 26. The engineered AAV VP capsid polypeptide of embodiment 5, wherein Xaa8 is H. 27. The engineered AAV VP capsid polypeptide of embodiment 5, wherein Xaa9 is R. 28. The engineered AAV VP capsid poly peptide of any one of embodiments 1-8, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 98%, or at least 99% identical to any one of SEQ ID NO: 7118-SEQ ID NO: 8117. 29. The engineered AAV VP capsid polypeptide of embodiment 27, wherein the region from residue 581 to residue 589 of SEQ ID NO: 1 has a sequence of any one of SEQ ID NO: 7118-SEQ ID NO: 8117. 30. The engineered AAV VP capsid polypeptide of any one of embodiments 2-3, wherein Xaa1 has low amino acid solubility. 31. The engineered AAV VP capsid polypeptide of embodiment 25, wherein Xaa1 is selected from K, R, or Q. 32. The engineered AAV VP capsid polypeptide of any one of embodiments 2-3, wherein Xaa1 has low amino acid hydropathy. 33. The engineered AAV VP capsid polypeptide of embodiment 27, wherein Xaa1 is selected from K or R. 34. The engineered AAV VP capsid polypeptide of any one of embodiments 2-3, wherein Xaa1 has a high average amino acid flexibility index. 35. The engineered AAV VP capsid polypeptide of embodiment 29, wherein Xaa1 is selected from D, E, R, K, G, I, N, Q, or S. 36. The engineered AAV VP capsid polypeptide of any one of embodiments 2-3, wherein Xaa1 has high hydrogen bond donors. 37. The engineered AAV VP capsid polypeptide of embodiment 31, wherein Xaa1 is selected from K. R. 38. The engineered AAV VP capsid polypeptide of any one of embodiments 2-3, wherein Xaa1 has low amino acid mutability. 39. The engineered AAV VP capsid polypeptide of embodiment 33, wherein Xaa1 is selected from K, R, P, or H. 40. The engineered AAV VP capsid polypeptide of any one of embodiments 2-3, wherein Xaa2 has low amino acid solubility. 41. The engineered AAV VP capsid polypeptide of embodiment 35, wherein Xaa2 is selected from R, K, Q, or S. 42. The engineered AAV VP capsid polypeptide of any one of embodiments 2-3, wherein Xaa2 has low amino acid hydropathy. 43. The engineered AAV VP capsid polypeptide of embodiment 37, wherein Xaa2 is selected from R, K, D, E, N, Q, H, P, Y, W, S, or T. 44. The engineered AAV VP capsid polypeptide of any one of embodiments 2-3, wherein Xaa2 has high amino acid charge. 45. The engineered AAV VP capsid polypeptide of embodiment 39, wherein Xaa2 is selected from R, K, or H. 46. The engineered AAV VP capsid polypeptide of any one of embodiments 2-3, wherein Xaa3 has high amino acid solubility. 47. The engineered AAV VP capsid polypeptide of embodiment 41, wherein Xaa3 is selected from A, M, V, W, L, or I. 48. The engineered AAV VP capsid polypeptide of any one of embodiments 2-3, wherein Xaa5 has high amino acid solubility. 49. The engineered AAV VP capsid polypeptide of embodiment 43, wherein Xaa5 is selected from C, M, V, W, L, or I. 50. The engineered AAV VP capsid polypeptide of any one of embodiments 2-3, wherein Xaa5 has high hydropathy. 51. The engineered AAV VP capsid polypeptide of embodiment 45, wherein Xaa5 is selected from M, V, or I. 52. The engineered AAV VP capsid polypeptide of any one of embodiments 2-3, wherein Xaa5 has low average amino acid flexibility index. 53. The engineered AAV VP capsid polypeptide of embodiment 47, wherein Xaa5 is selected from M, W, F, or C. 54. The engineered AAV VP capsid polypeptide of any one of embodiments 2-3, wherein Xaa8 has high amino acid solubility. 55. The engineered AAV VP capsid polypeptide of embodiment 49, wherein Xaa8 is selected from H, V, or I. 56. The engineered AAV VP capsid polypeptide of any one of embodiments 29-54, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 8118-SEQ ID NO: 9117. 57. The engineered AAV VP capsid polypeptide of any one of embodiments 25-50, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence of any one of SEQ ID NO: 8118-SEQ ID NO: 9117. 58. The engineered AAV VP capsid polypeptide of any one of embodiments 1-56, wherein the CNS tissue is selected from forebrain cortex, occipital cortex, temporal cortex, thalamus, hypothalamus, substantia nigra, hippocampus DG, hippocampus CA1, hippocampus CA3, cerebellum, and any combination thereof. 59. The engineered AAV VP capsid polypeptide of any one of embodiments 1-57, wherein tropism for CNS tissue is measured as a relative accumulation of the rAAV virion in a CNS tissue as compared to a non-CNS tissue, wherein the non-CNS tissue consists collectively of liver, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord. 60. The engineered AAV VP capsid polypeptide of any one of embodiments 1-58, wherein the higher tissue tropism is a 1.0005-fold to about a 1000-fold increased accumulation in the CNS tissue as compared to a non-CNS tissue. 61. The engineered AAV VP capsid polypeptide of embodiment 59, wherein the higher tissue tropism is at least about a 1.0005-fold, at least about a two-fold, at least about a three-fold, at least about a four-fold, at least about a five-fold, at least about a ten-fold, at least about a twenty-fold, at least about a 50-fold, at least about a 75-fold, at least about a 100-fold, or at least about a 1000-fold increased accumulation in the CNS tissue as compared to a non-CNS tissue.

Series C Embodiments—Liver De-Targeted Capsids

1. An engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide having an amino acid sequence at least 70% identical to SEQ ID NO: 1,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV),

wherein the at least one mutation confers higher tropism for a non-liver tissue on the rAAV as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1, and

wherein amino acid residues Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV), and

wherein the engineered AAV VP capsid polypeptide does not have the sequence of any of SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8. 4. The engineered AAV VP capsid polypeptide of embodiment 3, wherein the rAAV has higher tropism for a non-liver tissue as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1. 5. The engineered AAV VP capsid poly peptide of embodiment 1, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 98%, or at least 99% identical to any one of SEQ ID NO: 46438-SEQ ID NO: 47437. 6. The engineered AAV VP capsid polypeptide of embodiment 5, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence of any one of SEQ ID NO: 46438-SEQ ID NO: 47437. 7. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 46438-SEQ ID NO: 47437. 8. The engineered AAV VP capsid polypeptide of embodiment 7, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence of any one of SEQ ID NO: 46438-SEQ ID NO: 47437. 9. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein:

Xaa1 excludes A, G, K, M, N, Q, R, S, or T; or

Xaa2 excludes A, C, I, K, S, T, or V; or

Xaa3 excludes A, G, I, K, M, Q, R, S, T, or V; or

Xaa4 excludes A, I, K, L, P, Q, R, S, T, or V; or

Xaa5 excludes F, I, L, M, T, V, or Y; or

Xaa6 excludes F, H, M, N, Q, S, or Y; or

Xaa7 excludes A, C, K, M, Q or S; or

Xaa8 excludes A, C, F, G, M, Q, or S; or

Xaa9 excludes E, F, L, Q, R, or Y; or

any combination thereof.

10. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 excludes A, K, Q, or R. 11. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 excludes A, K, S, or T. 12. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 excludes A, K, Q, S, or T. 13. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 excludes K, I, S, or V. 14. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 excludes F, L, or Y. 15. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 excludes M or N. 16. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 excludes A, C, or S. 17. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 excludes A, C, M, or S. 18. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 excludes L, Q, or R. 19. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 excludes K. 20. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 excludes A. 21. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 excludes K, Q, or T. 22. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 excludes K. 23. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 excludes K. 24. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 excludes F. 25. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 excludes N. 26. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 excludes S. 27. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 excludes C. 28. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 excludes R. 29. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has low solubility. 30. The engineered AAV VP capsid polypeptide of embodiment 29, wherein Xaa1 is selected from D and P. 31. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has low mutability. 32. The engineered AAV VP capsid polypeptide of embodiment 31, wherein Xaa1 is selected from Xaa1 is selected from C, K, and L. 33. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has low solubility. 34. The engineered AAV VP capsid polypeptide of embodiment 33, wherein Xaa2 is selected from Xaa2 is selected from N, K, P, E, and D. 35. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has low hydropathy. 36.The engineered AAV VP capsid polypeptide of embodiment 35, wherein Xaa2 is selected from Xaa2 is selected from D, E, R, K, H, N, and Q. 37. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has low charge. 38. The engineered AAV VP capsid polypeptide of embodiment 37, wherein Xaa2 is selected from D and E. 39. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has high number of total potential hydrogen bonds. 40. The engineered AAV VP capsid polypeptide of embodiment 39, wherein Xaa2 is selected from H, N, Q, D, E. and R. 41. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has medium volume. 42. The engineered AAV VP capsid polypeptide of embodiment 41, wherein Xaa2 is selected from D, E, V, P, N, and T. 43. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has low solubility. 44. The engineered AAV VP capsid polypeptide of embodiment 43, wherein Xaa3 is selected from P and D. 45. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has medium volume. 46. The engineered AAV VP capsid polypeptide of embodiment 45, wherein Xaa4 is selected from D, E, V, P, N, and T. 47. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has low solubility. 48. The engineered AAV VP capsid polypeptide of embodiment 47, wherein Xaa5 is selected from N, P, E, and D. 49. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has low solubility. 50. The engineered AAV VP capsid polypeptide of embodiment 49, wherein Xaa8 is selected from K and Q. 51. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has low hydropathy. 52. The engineered AAV VP capsid polypeptide of embodiment 51, wherein Xaa8 is selected from K and R. 53. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has high surface accessibility. 54. The engineered AAV VP capsid polypeptide of embodiment 53, wherein Xaa8 is selected from E, R, and K. 55. The engineered AAV VP capsid polypeptide of any one of embodiments 1-54, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 46438-SEQ ID NO: 47437. 56. The engineered AAV VP capsid polypeptide of embodiment 55, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence of any one of SEQ ID NO: 46438-SEQ ID NO: 47437. 57. The engineered AAV VP capsid polypeptide of any one of embodiments 1-56, wherein tropism for a non-liver tissue is measured as a relative accumulation of the rAAV virion in a non-liver tissue as compared to a liver tissue, wherein the non-liver tissue consists collectively of CNS tissue, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord. 58. The engineered AAV VP capsid polypeptide of embodiment 57, wherein the CNS tissue is selected from forebrain cortex, occipital cortex, temporal cortex, thalamus, hypothalamus, substantia nigra, hippocampus DG, hippocampus CA1, hippocampus CA3, cerebellum, and any combination thereof. 59. The engineered AAV VP capsid polypeptide of any one of embodiments 57-58, wherein the higher tissue tropism is a 1.0005-fold to about a 1000-fold increased accumulation in the non-liver tissue as compared to a liver tissue. 60. The engineered AAV VP capsid polypeptide of embodiment 59, wherein the higher tissue tropism is at least about a 1.0005-fold, at least about a two-fold, at least about a three-fold, at least about a four-fold, at least about a five-fold, at least about a ten-fold, at least about a twenty-fold, at least about a 50-fold, at least about a 75-fold, at least about a 100-fold, or at least about a 1000-fold increased accumulation in the non-liver tissue as compared to a liver tissue.

Series D Embodiments—Liver Tropic Capsids

1. An engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide having an amino acid sequence at least 70% identical to SEQ ID NO: 1,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV),

wherein the at least one mutation confers higher tropism for a liver tissue on the rAAV as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1, and

wherein amino acid residues Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV), and

wherein the engineered AAV VP capsid polypeptide does not have the sequence of any of SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8. 4. The engineered AAV VP capsid polypeptide of embodiment 3, wherein the rAAV has higher tropism for a liver tissue as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1. 5. The engineered AAV VP capsid polypeptide of embodiment 1, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 43438-SEQ ID NO: 46437. 6. The engineered AAV VP capsid polypeptide of embodiment 5, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence of any one of SEQ ID NO: 43438-SEQ ID NO: 46437. 7. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 43438-SEQ ID NO: 46437. 8. The engineered AAV VP capsid polypeptide of embodiment 7, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence of any one of SEQ ID NO: 43438-SEQ ID NO: 46437. 9. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein:

Xaa1 is selected from the group consisting of A, G, K, M, N, Q, R, S, and T; or

Xaa2 is selected from the group consisting of A, C, I, K, S, T, and V; or

Xaa3 is selected from the group consisting of A, G, I, K, M, Q, R, S, T, and V; or

Xaa4 is selected from the group consisting of A, I, K, L, P, Q, R, S, T, and V; or

Xaa5 is selected from the group consisting of F, I, L, M, T, V, and Y; or

Xaa6 is selected from the group consisting of F, H, M, N, Q, S, and Y; or

Xaa7 is selected from the group consisting of A, C, K, M, Q and S; or

Xaa8 is selected from the group consisting of A, C, F, G, M, Q, and S; or

Xaa9 is selected from the group consisting of E, F, L, Q, R, and Y; or

any combination thereof.

10. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is selected from A, K, Q, and R. 11. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is K. 12. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is selected from A, K, S, and T. 13. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is A. 14. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is selected from A, K, Q, S, and T. 15. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is selected from K, Q, and T. 16. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is K. 17. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is selected from K, I, S, and V. 18. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is K. 19. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is selected from F, L, and Y. 20. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is F. 21. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is selected from M and N. 22. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is N. 23. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 is selected from A, C, and S. 24. The engineered AAV VP capsid poly peptide of embodiment 9, wherein Xaa7 is S. 25. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is selected from A, C, M, and S. 26. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is C. 27. The engineered AAV VP capsid poly peptide of embodiment 9, wherein Xaa9 is selected from L, Q, and R. 28. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 is R. 29. The engineered AAV VP capsid polypeptide of any one of embodiments 1-28, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 43438-SEQ ID NO: 44437. 30. The engineered AAV VP capsid polypeptide of embodiment 29, wherein the region from residue 581 to residue 589 of SEQ ID NO: 1 has a sequence of any one of SEQ ID NO: 43438-SEQ ID NO: 44437. 31. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has high surface accessibility. 32. The engineered AAV VP capsid polypeptide of embodiment 31, wherein Xaa1 is selected from K, R, and E. 33. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has Low hydropathy (<−3.5). 34. The engineered AAV VP capsid polypeptide of embodiment 33, wherein Xaa1 is selected from K and R. 35. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has Low amino acid mutability. 36. The engineered AAV VP capsid polypeptide of embodiment 35, wherein Xaa1 is selected from H, P, K, and R 37. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has Low amino acid solubility. 38. The engineered AAV VP capsid polypeptide of embodiment 37, wherein Xaa1 is selected from Q, K, and R. 39. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has High surface accessibility.

40. The engineered AAV VP capsid polypeptide of embodiment 39, wherein Xaa2 is selected from E, R, and K. 41. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has Low hydropathy. 42. The engineered AAV VP capsid polypeptide of embodiment 41, wherein Xaa2 is selected from K and R. 43. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has High amino acid volume. 44. The engineered AAV VP capsid polypeptide of embodiment 43, wherein Xaa2 is selected from S, L, I, A, R, and K. 45. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has High mutability. 46. The engineered AAV VP capsid polypeptide of embodiment 45, wherein Xaa3 is selected from N, I, A, M, E, and D. 47. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has Low solubility. 48. The engineered AAV VP capsid polypeptide of embodiment 47, wherein Xaa3 is selected from N, K, R, and E. 49. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has Low hydropathy. 50. The engineered AAV VP capsid polypeptide of embodiment 49, wherein Xaa4 is selected from K and R. 51. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has High amino acid volume. 52. The engineered AAV VP capsid polypeptide of embodiment 51, wherein Xaa4 is selected from K, R, I, and L. 53. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has Medium amino acid solubility. 54. The engineered AAV VP capsid polypeptide of embodiment 53, wherein Xaa5 is selected from H and T. 55. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has Low surface accessibility. 56. The engineered AAV VP capsid polypeptide of embodiment 55, wherein Xaa8 is selected from V and C. 57. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has Low average flexibility index. 58. The engineered AAV VP capsid polypeptide of embodiment 57, wherein Xaa8 is selected from W, V, M, A, F, L, H, and C. 59. The engineered AAV VP capsid polypeptide of any one of embodiments 1-58, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any, one of SEQ ID NO: 44438-SEQ ID NO: 45437. 60. The engineered AAV VP capsid polypeptide of embodiment 59, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence of any one of SEQ ID NO: 44438-SEQ ID NO: 45437. 61. The engineered AAV VP capsid polypeptide of any one of embodiments 1-60, wherein tropism for liver tissue is measured as a relative accumulation of the rAAV virion in a liver tissue as compared to a non-liver tissue, wherein the non-liver tissue consists collectively of CNS, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord. 62. The engineered AAV VP capsid polypeptide of embodiment 61, wherein the CNS tissue is selected from forebrain cortex, occipital cortex, temporal cortex, thalamus, hypothalamus, substantia nigra, hippocampus DG, hippocampus CA1, hippocampus CA3, cerebellum, and any combination thereof. 63. The engineered AAV VP capsid polypeptide of any one of embodiments 61-62, wherein the higher tissue tropism is a 1.0005-fold to about a 1000-fold increased accumulation in the liver tissue as compared to a non-liver tissue. 64. The engineered AAV VP capsid polypeptide of embodiment 63, wherein the higher tissue tropism is at least about a 1.0005-fold, at least about a two-fold, at least about a three-fold, at least about a four-fold, at least about a five-fold, at least about a ten-fold, at least about a twenty-fold, at least about a 50-fold, at least about a 75-fold, at least about a 100-fold, or at least about a 1000-fold increased accumulation in the liver tissue as compared to a non-liver tissue.

Series E Embodiments—Adrenal Gland Tropic Capsids

1. An engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide having an amino acid sequence at least 70% identical to SEQ ID NO: 1,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV),

wherein the at least one mutation confers higher tropism for an adrenal gland tissue on the rAAV as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1, and

wherein the engineered AAV VP capsid polypeptide does not have the sequence of any of SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8. 2. The engineered AAV VP capsid polypeptide of embodiment 1, wherein the engineered AAV VP capsid polypeptide has a sequence of SEQ ID NO: 2 and wherein Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from any amino acid. 3.An engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide having an amino acid sequence of SEQ ID NO: 2,

wherein amino acid residues Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV), and

wherein the engineered AAV VP capsid polypeptide does not have the sequence of any of SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8. 4. The engineered AAV VP capsid polypeptide of embodiment 3, wherein the rAAV has higher tropism for an adrenal gland tissue as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1. 5. The engineered AAV VP capsid poly peptide of embodiment 1, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 1118-SEQ ID NO: 4117. 6. The engineered AAV VP capsid polypeptide of embodiment 5, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence of any one of SEQ ID NO: 1118-SEQ ID NO: 4117. 7. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 1118-SEQ ID NO: 4117. 8. The engineered AAV VP capsid polypeptide of embodiment 7, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence of any one of SEQ ID NO: 1118-SEQ ID NO: 4117. 9. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein:

Xaa1 is selected from the group consisting of A, C, K, Q, R S, and T; or

Xaa2 is selected from the group consisting of A, C, I, S, T, and V; or

Xaa3 is selected from the group consisting of A, F, G, K, M, Q, R, T, and V; or

Xaa4 is selected from the group consisting of A, K, M, Q, R, and V; or

Xaa5 is selected from the group consisting of F, I, L, M, R, T, V, and Y; or

Xaa6 is selected from the group consisting of G, H, M, N, R, and S; or

Xaa7 is selected from the group consisting of A, H, K, Q, R, S and V; or

Xaa8 is selected from the group consisting of A, G, H, M, Q, and S; or

Xaa9 is selected from the group consisting of A, E, N, P, R, S, and Y; or

any combination thereof. 10. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is selected from C, K, and R. 11. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is C. 12. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is selected from A, V, and T. 13. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is V. 14. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is selected from A, G, and M. 15. The engineered AAV VP capsid poly peptide of embodiment 9, wherein Xaa3 is M. 16. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is selected from A, R, and K. 17. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is K. 18. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is selected from R, V, and Y. 19. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is V. 20. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is selected from H and N. 21. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is N. 22. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 is selected from H, Q, and V. 23. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 is H. 24. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is selected from A, G, M, and S. 25. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is S. 26. The engineered AAV VP capsid poly peptide of embodiment 9, wherein Xaa9 is selected from P and E. 27. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 is P. 28. The engineered AAV VP capsid polypeptide of any one of embodiments 1-27, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 1118-SEQ ID NO: 2117. 29. The engineered AAV VP capsid polypeptide of embodiment 28, wherein the region from residue 581 to residue 589 of SEQ ID NO: 1 has a sequence of any one of SEQ ID NO: 1118-SEQ ID NO: 2117. 30. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has low mol mass. 31. The engineered AAV VP capsid polypeptide of embodiment 30, wherein Xaa1 is selected from V, P, S, and C. 32. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has low hydropathy. 33. The engineered AAV VP capsid polypeptide of embodiment X, wherein Xaa1 is selected from T, S, W, and Y. 34. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has low hydropathy. 35. The engineered AAV VP capsid polypeptide of embodiment 34, wherein Xaa2 is R. 36. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has low mutability. 37. The engineered AAV VP capsid polypeptide of embodiment 36, wherein Xaa2 is C. 38. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has low solubility. 39. The engineered AAV VP capsid polypeptide of embodiment 38, wherein Xaa2 is K. 40. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has low average flexibility. 41. The engineered AAV VP capsid polypeptide of embodiment 40, wherein Xaa3 is selected from W, M, and F. 42. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has high solubility. 43. The engineered AAV VP capsid polypeptide of embodiment 42, wherein Xaa3 is M. 44. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has high surface accessibility. 45. The engineered AAV VP capsid polypeptide of embodiment 44, wherein Xaa4 is selected from K and R. 46. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has high average flexibility. 47. The engineered AAV VP capsid polypeptide of embodiment 46, wherein Xaa4 is selected from K, I, and N. 48. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has medium mutability. 49. The engineered AAV VP capsid polypeptide of embodiment 48, wherein Xaa5 is selected from R and H. 50. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has high goldman engelman steitz. 51. The engineered AAV VP capsid polypeptide of embodiment 50, wherein Xaa5 is selected from V and L. 52. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has low hydropathy. 53. The engineered AAV VP capsid polypeptide of embodiment 52, wherein Xaa5 is R. 54. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has high volume. 55. The engineered AAV VP capsid polypeptide of embodiment 54, wherein Xaa5 is selected from Y, R, and F. 56. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has high solubility. 57. The engineered AAV VP capsid polypeptide of embodiment 56, wherein Xaa6 is selected from Y, V, M, A, and C. 58. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa7 has medium mutability. 59. The engineered AAV VP capsid polypeptide of embodiment 58, wherein Xaa7 is selected from V, H, and R. 60. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa7 has low solubility. 61. The engineered AAV VP capsid polypeptide of embodiment 59, wherein Xaa7 is R. 62. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has high average flexibility. 64. The engineered AAV VP capsid polypeptide of embodiment 62, wherein Xaa8 is selected from K, I, and N. 65. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has high mol mass. 66. The engineered AAV VP capsid polypeptide of embodiment 65, wherein Xaa8 is selected from R and Y. 67. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa9 has high mutability. 68. The engineered AAV VP capsid polypeptide of embodiment X, wherein Xaa9 is N. 69. The engineered AAV VP capsid poly peptide of any one of embodiments 1-68, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 2118-SEQ ID NO: 3117. 70. The engineered AAV VP capsid polypeptide of embodiment 69, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence of any one of SEQ ID NO: 2118-SEQ ID NO: 3117. 71. The engineered AAV VP capsid polypeptide of any one of embodiments 1-70, wherein tropism for adrenal gland tissue is measured as a relative accumulation of the rAAV virion in an adrenal gland tissue as compared to a non-adrenal gland tissue, wherein the non-adrenal gland tissue consists collectively of CNS, liver, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, thyroid, colon, sciatic nerve, and spinal cord. 72. The engineered AAV VP capsid polypeptide of embodiment 71, wherein the CNS tissue is selected from forebrain cortex, occipital cortex, temporal cortex, thalamus, hypothalamus, substantia nigra, hippocampus DG, hippocampus CA1, hippocampus CA3, cerebellum, and any combination thereof. 73. The engineered AAV VP capsid polypeptide of any one of embodiments 71-72, wherein the higher tissue tropism is a 1.0005-fold to about a 1000-fold increased accumulation in the adrenal gland tissue as compared to a non-adrenal gland tissue. 74. The engineered AAV VP capsid polypeptide of embodiment 73, wherein the higher tissue tropism is at least about a 1.0005-fold, at least about a two-fold, at least about a three-fold, at least about a four-fold, at least about a five-fold, at least about a ten-fold, at least about a twenty-fold, at least about a 50-fold, at least about a 75-fold, at least about a 100-fold, or at least about a 1000-fold increased accumulation in the adrenal gland tissue as compared to a non-adrenal gland tissue.

Series F Embodiments—Bone Marrow Tropic Capsids

1. An engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide having an amino acid sequence at least 70% identical to SEQ ID NO: 1,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV),

wherein the at least one mutation confers higher tropism for a bone marrow tissue on the rAAV as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1, and

wherein amino acid residues Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV), and

wherein the engineered AAV VP capsid polypeptide does not have the sequence of any of SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8. 4. The engineered AAV VP capsid polypeptide of embodiment 3, wherein the rAAV has higher tropism for a bone marrow tissue as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1. 5. The engineered AAV VP capsid polypeptide of embodiment 1, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 4118-SEQ ID NO: 7117. 6. The engineered AAV VP capsid polypeptide of embodiment 5, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence of any one of SEQ ID NO: 4118-SEQ ID NO: 7117. 7. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, at least 98%, or at least 99% identical to any one of SEQ ID NO: 4118-SEQ ID NO: 7117. 8. The engineered AAV VP capsid polypeptide of embodiment 7, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence of any one of SEQ ID NO: 4118-SEQ ID NO: 7117. 9. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein:

Xaa1 is selected from the group consisting of A, E, G, Q, S, and T; or

Xaa2 is selected from the group consisting of A, I, Q, S, T, V, and Y; or

Xaa3 is selected from the group consisting of A, G, I, M, Q, S, and T; or

Xaa4 is selected from the group consisting of A, E, P, Q, T, and V; or

Xaa5 is selected from the group consisting of F, I, L, M, Q, V, and Y; or

Xaa6 is selected from the group consisting of F, I, N, Q, S, and V; or

Xaa7 is selected from the group consisting of A, C, M, S, and V; or

Xaa8 is selected from the group consisting of A, C, D, G, M, S, and Y; or

Xaa9 is selected from the group consisting of D, E, G, L, P, S, and Y; or

any combination thereof. 10. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is selected from A, E. and T. 11. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is E. 12. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is selected from A, S, and T. 13. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is A. 14. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is selected from A, Q, and T. 15. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is Q. 16. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is selected from A, P, and Q. 17. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is Q. 18. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is selected from F, V, and Y. 19. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is V. 20. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is selected from I, N, Q, and S. 21. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is S. 22. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 is A, C, and V. 23. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 is C. 24. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is selected from A, M, S, and Y. 25. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is M. 26. The engineered AAV VP capsid poly peptide of embodiment 9, wherein Xaa9 is selected from D, E, and P. 27. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 is P. 28. The engineered AAV VP capsid polypeptide of any one of embodiments 1-27, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 4118-SEQ ID NO: 5117. 29. The engineered AAV VP capsid polypeptide of embodiment 28, wherein the region from residue 581 to residue 589 of SEQ ID NO: 1 has a sequence of any one of SEQ ID NO: 4118-SEQ ID NO: 5117. 30. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has high hydropathy. 31. The engineered AAV VP capsid polypeptide of embodiment 30, wherein Xaa1 is selected from V, I, and L. 32. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has low mutability. 33. The engineered AAV VP capsid polypeptide of embodiment 32, wherein Xaa1 is selected from Y, L, F, and C. 34. The engineered AAV VP capsid poly peptide of any one of embodiments 2-4, wherein Xaa2 has low hydropathy. 35. The engineered AAV VP capsid polypeptide of embodiment 34, wherein Xaa2 is selected from Y and W. 36. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has high mol mass. 37. The engineered AAV VP capsid polypeptide of embodiment 36, wherein Xaa2 is W. 38. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has low surface accessibility. 39. The engineered AAV VP capsid polypeptide of embodiment 38, wherein Xaa2 is selected from W and A. 40. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has low hydrophilicity. 41. The engineered AAV VP capsid polypeptide of embodiment 40, wherein Xaa2 is W. 42. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has low mutability. 43. The engineered AAV VP capsid polypeptide of embodiment 42, wherein Xaa2 is C. 44. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has low average flexibility. 45. The engineered AAV VP capsid polypeptide of embodiment 44, wherein Xaa2 is selected from W, M, and F. 46. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has low average flexibility. 47. The engineered AAV VP capsid polypeptide of embodiment 46, wherein Xaa5 is selected from W, M, and F. 48. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has low average flexibility. 49. The engineered AAV VP capsid polypeptide of embodiment 48, wherein Xaa6 is selected from W, M, and F. 50. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has low mutability. 51. The engineered AAV VP capsid polypeptide of embodiment 50, wherein Xaa6 is selected from Y, F, L, and C. 52. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has high solubility. 53. The engineered AAV VP capsid polypeptide of embodiment 52, wherein Xaa6 is selected from W, F, I, and L. 54. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa7 has low surface accessibility. 55. The engineered AAV VP capsid polypeptide of embodiment 54, wherein Xaa7 is C. 56. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa7 has high surface accessibility. 57. The engineered AAV VP capsid polypeptide of embodiment 56, wherein Xaa7 is selected from D and N.

58. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa7 has low mutability.

59. The engineered AAV VP capsid polypeptide of embodiment 58, wherein Xaa7 is C.

60. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa7 has high solubility.

61. The engineered AAV VP capsid polypeptide of embodiment 60, wherein Xaa7 is C.

62. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa7 has low solubility.

63. The engineered AAV VP capsid polypeptide of embodiment X, wherein Xaa7 is D.

64. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has low charge.

65. The engineered AAV VP capsid polypeptide of embodiment 64, wherein Xaa8 is selected from D and E.

66. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has high mutability.

67. The engineered AAV VP capsid polypeptide of embodiment 66, wherein Xaa8 is selected from D, E. A, and T.

68. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa9 has high mol mass.

69. The engineered AAV VP capsid polypeptide of embodiment 68, wherein Xaa9 is selected from H and F.

70. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa9 has low mutability.

71. The engineered AAV VP capsid polypeptide of embodiment X, wherein Xaa9 is selected from Y, F, and L.

72. The engineered AAV VP capsid polypeptide of any one of embodiments 1-71, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 5118-SEQ ID NO: 6117.

73. The engineered AAV VP capsid polypeptide of embodiment 72, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence of any one of SEQ ID NO: 5118-SEQ ID NO: 6117.

74. The engineered AAV VP capsid polypeptide of any one of embodiments 1-73, wherein tropism for bone marrow tissue is measured as a relative accumulation of the rAAV virion in a bone marrow tissue as compared to a non-bone marrow tissue, wherein the non-bone marrow tissue consists collectively of CNS, liver, skeletal muscle, heart, lung, spleen, lymph node, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord.

75. The engineered AAV VP capsid polypeptide of embodiment 74, wherein the CNS tissue is selected from forebrain cortex, occipital cortex, temporal cortex, thalamus, hypothalamus, substantia nigra, hippocampus DG, hippocampus CA1, hippocampus CA3, cerebellum, and any combination thereof.

76. The engineered AAV VP capsid polypeptide of any one of embodiments 74-75, wherein the higher tissue tropism is a 1.0005-fold to about a 1000-fold increased accumulation in the bone marrow tissue as compared to a non-bone marrow tissue.

77. The engineered AAV VP capsid polypeptide of embodiment 76, wherein the higher tissue tropism is at least about a 1.0005-fold, at least about a two-fold, at least about a three-fold, at least about a four-fold, at least about a five-fold, at least about a ten-fold, at least about a twenty-fold, at least about a 50-fold, at least about a 75-fold, at least about a 100-fold, or at least about a 1000-fold increased accumulation in the bone marrow tissue as compared to a non-bone marrow tissue.

Series G Embodiments—Colon Tropic Capsids

1. An engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide having an amino acid sequence at least 70% identical to SEQ ID NO: 1,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV),

wherein the at least one mutation confers higher tropism for a colon tissue on the rAAV as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1, and

wherein amino acid residues Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV), and

Xaa1 is selected from the group consisting of C, F, H, N, P, W, and Y; or

Xaa2 is selected from the group consisting of D, E, F, L, and P; or

Xaa3 is selected from the group consisting of C, F, H, I, L, P, and Y; or

Xaa4 is selected from the group consisting of C, D, E, N, and P; or

Xaa5 is selected from the group consisting of D, E, G, P, and W; or

Xaa6 is selected from the group consisting of C, K, R, and V; or

Xaa7 is selected from the group consisting of D, M, P, and V; or

Xaa8 is selected from the group consisting of D, I, K, L, P, R, and V; or

Xaa9 is selected from the group consisting of C, H, I, K, L, M, and W; or

any combination thereof.

10. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is selected from F, P, and W.

11. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is P.

12. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is selected from D, E, L, and P.

13. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is P.

14. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is selected from C, H, and P.

15. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is P.

16. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is selected from C, D, and E.

17. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is C.

18. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is selected from G, P, and W.

19. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is P.

20. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is selected from K and R.

21. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is R.

22. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 is P.

23. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is selected from K, P, and R.

24. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is P.

25. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 is selected from I, L, and M.

26. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 is I.

27. The engineered AAV VP capsid polypeptide of any one of embodiments 1-26, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 10118-SEQ ID NO: 11117.

28. The engineered AAV VP capsid polypeptide of embodiment 27, wherein the region from residue 581 to residue 589 of SEQ ID NO: 1 has a sequence of any one of SEQ ID NO. 10118-SEQ ID NO: 11117.

29. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has High mol mass.

30. The engineered AAV VP capsid polypeptide of embodiment 29, wherein Xaa1 is selected from Y, W.

31. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has High solubility.

32. The engineered AAV VP capsid polypeptide of embodiment 31, wherein Xaa1 is selected from W, F, I, and L.

33. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has Low solubility.

34. The engineered AAV VP capsid polypeptide of embodiment 33, wherein Xaa2 is D.

35. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has Low mutability.

36. The engineered AAV VP capsid polypeptide of embodiment 35, wherein Xaa2 is selected from P and K.

37. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has Medium mol mass.

38. The engineered AAV VP capsid polypeptide of embodiment 37, wherein Xaa2 is selected from D, E, N, K, M, Q, I, and L.

39. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has Low hydropathy.

40. The engineered AAV VP capsid polypeptide of embodiment 39, wherein Xaa2 is selected from D, E, R, K, H, N, and Q.

41. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has Low mutability.

42. The engineered AAV VP capsid polypeptide of embodiment 41, wherein Xaa3 is selected from K, V, P, and C.

43. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has High solubility.

44. The engineered AAV VP capsid polypeptide of embodiment 43, wherein Xaa3 is selected from W, F, I, and L.

45. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has High average flexibility.

46. The engineered AAV VP capsid polypeptide of embodiment 45, wherein Xaa5 is selected from S, P, G, R, E, and D.

47. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has High surface accessibility.

48. The engineered AAV VP capsid polypeptide of embodiment 47, wherein Xaa5 is selected from D and N.

49. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has Low hydropathy.

50. The engineered AAV VP capsid polypeptide of embodiment 49, wherein Xaa6 is selected from R.

51. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has Low mutability.

52. The engineered AAV VP capsid polypeptide of embodiment 51, wherein Xaa6 is selected from Y, R, F, and L.

53. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has Low solubility.

54. The engineered AAV VP capsid polypeptide of embodiment 53, wherein Xaa6 is selected from R and Q.

55. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has High surface accessibility.

56. The engineered AAV VP capsid polypeptide of embodiment 55, wherein Xaa6 is selected from E, R, K.

57. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has High average flexibility.

58. The engineered AAV VP capsid polypeptide of embodiment 57, wherein Xaa6 is selected from G and R.

59. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has Low solubility.

60. The engineered AAV VP capsid polypeptide of embodiment 59, wherein Xaa8 is D.

61. The engineered AAV VP capsid polypeptide of any one of embodiments 1-60, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 11118-SEQ ID NO: 12117.

62. The engineered AAV VP capsid polypeptide of embodiment 61, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence of any one of SEQ ID NO: 11118-SEQ ID NO: 12117.

63. The engineered AAV VP capsid polypeptide of any one of embodiments 1-62, wherein tropism for colon tissue is measured as a relative accumulation of the rAAV virion in a colon tissue as compared to a non-colon tissue, wherein the non-colon tissue consists collectively of CNS, liver, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, sciatic nerve, and spinal cord.

64. The engineered AAV VP capsid polypeptide of embodiment 63, wherein the CNS tissue is selected from forebrain cortex, occipital cortex, temporal cortex, thalamus, hypothalamus, substantia nigra, hippocampus DG, hippocampus CA1, hippocampus CA3, cerebellum, and any combination thereof.

65. The engineered AAV VP capsid polypeptide of any one of embodiments 63-64, wherein the higher tissue tropism is a 1.0005-fold to about a 1000-fold increased accumulation in the colon tissue as compared to a non-colon tissue.

66. The engineered AAV VP capsid polypeptide of embodiment 65, wherein the higher tissue tropism is at least about a 1.0005-fold, at least about a two-fold, at least about a three-fold, at least about a four-fold, at least about a five-fold, at least about a ten-fold, at least about a twenty-fold, at least about a 50-fold, at least about a 75-fold, at least about a 100-fold, or at least about a 1000-fold increased accumulation in the colon tissue as compared to a non-colon tissue.

Series H Embodiments—Heart Tropic Capsids

1. An engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide having an amino acid sequence at least 70% identical to SEQ ID NO: 1,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV),

wherein the at least one mutation confers higher tropism for a heart tissue on the rAAV as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1, and

wherein amino acid residues Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV), and

5. The engineered AAV VP capsid polypeptide of embodiment 1, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 13118-SEQ ID NO: 16117.

6. The engineered AAV VP capsid polypeptide of embodiment 5, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence of any one of SEQ ID NO: 13118-SEQ ID NO: 16117.

7. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 13118-SEQ ID NO: 16117.

8. The engineered AAV VP capsid polypeptide of embodiment 7, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence of any one of SEQ ID NO: 13118-SEQ ID NO: 16117.

9. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein:

Xaa1 is selected from the group consisting of I, K, L, M, T, and V; or

Xaa2 is selected from the group consisting of A, C, G, I, K, and S; or

Xaa3 is selected from the group consisting of A, D, E, G, K, M, and V; or

Xaa4 is selected from the group consisting of F, H, R, T, W, and Y; or

Xaa5 is selected from the group consisting of F, L, M, and R; or

Xaa6 is selected from the group consisting of A, H, N, W, and Y; or

Xaa7 is selected from the group consisting of A, C, E, F, K, and T; or

Xaa8 is selected from the group consisting of A, C, M, S, and T; or

Xaa9 is selected from the group consisting of A, D, G, and P; or

any combination thereof.

10. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is selected from K and L.

11. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is K.

12. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is selected from A, C, and S.

13. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is A.

14. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is selected from E and V.

15. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is E.

16. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is selected from F, R, and T.

17. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is R.

18. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is L.

19. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is selected from H, N, and Y.

20. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is H.

21. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 is selected from C, F, and T.

22. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 is F.

23. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is selected from C, M, and S.

24. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is C.

25. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 is selected from A and G.

26. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 is A.

27. The engineered AAV VP capsid polypeptide of any one of embodiments 1-26, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 13118-SEQ ID NO: 14117.

28. The engineered AAV VP capsid polypeptide of embodiment 27, wherein the region from residue 581 to residue 589 of SEQ ID NO: 1 has a sequence of any one of SEQ ID NO: 13118-SEQ ID NO: 14117.

29. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has low solubility.

30. The engineered AAV VP capsid polypeptide of embodiment 29, wherein Xaa1 is selected from N and E.

31. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has low hydropathy.

32. The engineered AAV VP capsid polypeptide of embodiment 31, wherein Xaa1 is selected from H, N, Q, P, Y, D, and E.

33. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has high mutability.

34. The engineered AAV VP capsid polypeptide of embodiment 33, wherein Xaa1 is selected from A and E.

35. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has high hydropathy.

36. The engineered AAV VP capsid polypeptide of embodiment 35, wherein Xaa2 is selected from V and I.

37. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has medium mutability.

38. The engineered AAV VP capsid polypeptide of embodiment 37, wherein Xaa2 is V.

39. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has medium volume.

40. The engineered AAV VP capsid polypeptide of embodiment 39, wherein Xaa2 is selected from V, E, and Q.

41. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has high solubility.

42. The engineered AAV VP capsid polypeptide of embodiment 41, wherein Xaa2 is selected from V and M.

43. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has low solubility.

44. The engineered AAV VP capsid polypeptide of embodiment 43, wherein Xaa3 is selected from R and Q.

45. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has low surface accessibility.

46. The engineered AAV VP capsid polypeptide of embodiment 45, wherein Xaa4 is C.

47. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has high solubility.

48. The engineered AAV VP capsid polypeptide of embodiment 47, wherein Xaa4 is C.

49. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has low charge.

50. The engineered AAV VP capsid polypeptide of embodiment 49, wherein Xaa4 is selected from D, E, Y, W, V, P, M, A, G, F, I, L, N, Q, S, T, and C.

51. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has high hydropathy.

52. The engineered AAV VP capsid polypeptide of embodiment 51, wherein Xaa4 is C.

53. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has high surface accessibility.

54. The engineered AAV VP capsid polypeptide of embodiment 53, wherein Xaa5 is selected from D, E, R, K, N, and Q.

55. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has low solubility.

56. The engineered AAV VP capsid polypeptide of embodiment 55, wherein Xaa5 is D.

57. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has low mutability.

58. The engineered AAV VP capsid polypeptide of embodiment 57, wherein Xaa6 is C.

59. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has low solubility.

60. The engineered AAV VP capsid polypeptide of embodiment 59, wherein Xaa6 is D.

61. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has high surface accessibility.

62. The engineered AAV VP capsid polypeptide of embodiment 61, wherein Xaa8 is selected from D and N.

63. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has high average flexibility.

64. The engineered AAV VP capsid polypeptide of embodiment 63, wherein Xaa8 is selected from D, R, P, G, and S.

65. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa9 has medium mol mass.

66. The engineered AAV VP capsid polypeptide of embodiment 65, wherein Xaa9 is selected from N, D, L, and I.

67. The engineered AAV VP capsid polypeptide of any one of embodiments 1-66, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 900%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 14118-SEQ ID NO: 15117.

68. The engineered AAV VP capsid polypeptide of embodiment 67, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence of any one of SEQ ID NO: 14118-SEQ ID NO: 15117.

69. The engineered AAV VP capsid polypeptide of any one of embodiments 1-68, wherein tropism for heart tissue is measured as a relative accumulation of the rAAV virion in a heart tissue as compared to a non-heart tissue, wherein the non-heart tissue consists collectively of CNS, liver, skeletal muscle, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord.

70. The engineered AAV VP capsid polypeptide of embodiment 69, wherein the CNS tissue is selected from forebrain cortex, occipital cortex, temporal cortex, thalamus, hypothalamus, substantia nigra, hippocampus DG, hippocampus CA1, hippocampus CA3, cerebellum, and any combination thereof.

71. The engineered AAV VP capsid polypeptide of any one of embodiments 69-70, wherein the higher tissue tropism is a 1.0005-fold to about a 1000-fold increased accumulation in the heart tissue as compared to a non-heart tissue.

72. The engineered AAV VP capsid polypeptide of embodiment 71, wherein the higher tissue tropism is at least about a 1.0005-fold, at least about a two-fold, at least about a three-fold, at least about a four-fold, at least about a five-fold, at least about a ten-fold, at least about a twenty-fold, at least about a 50-fold, at least about a 75-fold, at least about a 100-fold, or at least about a 1000-fold increased accumulation in the heart tissue as compared to a non-heart tissue.

Series I Embodiments—Lung Tropic Capsids

1. An engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide having an amino acid sequence at least 70% identical to SEQ ID NO: 1,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV),

wherein the at least one mutation confers higher tropism for a lung tissue on the rAAV as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1, and

wherein amino acid residues Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV), and

wherein the engineered AAV VP capsid polypeptide does not have the sequence of any of SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8.

4. The engineered AAV VP capsid polypeptide of embodiment 3, wherein the rAAV has higher tropism for a lung tissue as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1.

5. The engineered AAV VP capsid polypeptide of embodiment 1, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 16118-SEQ ID NO: 19117.

6. The engineered AAV VP capsid poly peptide of embodiment 5, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence of any one of SEQ ID NO: 16118-SEQ ID NO: 19117.

7. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence that is at least 7M/%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 16118-SEQ ID NO: 19117.

8. The engineered AAV VP capsid polypeptide of embodiment 7, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence of any one of SEQ ID NO: 16118-SEQ ID NO: 19117.

9. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein:

Xaa1 is selected from the group consisting of A, E, K, M, Q, R, S, and T; or

Xaa2 is selected from the group consisting of A, I, K, S, T, and V; or

Xaa3 is selected from the group consisting of A, E, K, M, Q, R, S, T, and V; or

Xaa4 is selected from the group consisting of M, P, R, S, and T; or

Xaa5 is selected from the group consisting of I, K, L, M, T, V, and Y; or

Xaa6 is selected from the group consisting of D, G, H, M, N, R, and S; or

Xaa7 is selected from the group consisting of A, K, M, Q, and R; or

Xaa8 is selected from the group consisting of A, F, G, S, W, and Y; or

Xaa9 is selected from the group consisting of A, E, G, P, R, and Y; or

any combination thereof.

10. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is selected from A, E, and Q.

11. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is E.

12. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is selected from S, T, and V.

13. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is T.

14. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is selected from A, K, R, and S.

15. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is R.

16. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is selected from P, Q, and T.

17. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is Q.

18. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is selected from L, M, and Y.

19. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is L.

20. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is selected from H and N.

21. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is N.

22. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 is selected from A, K and R.

23. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 is R.

24. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is selected from A, F, and G.

25. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is F.

26. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 is selected from G, P, and R.

27. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 is G.

28. The engineered AAV VP capsid polypeptide of any one of embodiments 1-27, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 16118-SEQ ID NO: 17117.

29. The engineered AAV VP capsid polypeptide of embodiment 28, wherein the region from residue 581 to residue 589 of SEQ ID NO: 1 has a sequence of any one of SEQ ID NO: 16118-SEQ ID NO: 17117.

30. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has high mutability.

31. The engineered AAV VP capsid polypeptide of embodiment 30, wherein Xaa1 is selected from D, E, M, A, I, Q, and T.

32. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has high mol mass.

33. The engineered AAV VP capsid polypeptide of embodiment 32, wherein Xaa2 is F.

34. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has low mutability.

35. The engineered AAV VP capsid polypeptide of embodiment 34, wherein Xaa2 is selected from Y, F, and L.

36. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has low mutability.

37. The engineered AAV VP capsid polypeptide of embodiment 36, wherein Xaa3 is selected from K, V, P, and H.

38. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has low hydropathy.

39. The engineered AAV VP capsid polypeptide of embodiment 38, wherein Xaa3 is selected from K and R.

40. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has low mutability.

41. The engineered AAV VP capsid polypeptide of embodiment 40, wherein Xaa4 is selected from K and P.

42. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has high average flexibility.

43. The engineered AAV VP capsid polypeptide of embodiment 42, wherein Xaa4 is selected from D, E, P, and S.

44. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has low average flexibility.

45. The engineered AAV VP capsid polypeptide of embodiment 44, wherein Xaa5 is selected from W, M, and F.

46. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has high solubility.

47. The engineered AAV VP capsid polypeptide of embodiment 46, wherein Xaa5 is selected from W, F, I, and L.

48. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has medium mutability.

49. The engineered AAV VP capsid polypeptide of embodiment 48, wherein Xaa6 is selected from R, and H.

50. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has high surface accessibility.

51. The engineered AAV VP capsid polypeptide of embodiment 50, wherein Xaa6 is selected from T.

52. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa7 has low mutability.

53. The engineered AAV VP capsid polypeptide of embodiment 52, wherein Xaa7 is C.

54. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa7 has high solubility.

55. The engineered AAV VP capsid polypeptide of embodiment 54, wherein Xaa7 is selected from W, V, M, F, I, and L.

56. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has high mutability.

57. The engineered AAV VP capsid polypeptide of embodiment 56, wherein Xaa8 is selected from D, E, M, A, I, Q, and T.

58. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has low hydropathy.

59. The engineered AAV VP capsid polypeptide of embodiment 58, wherein Xaa8 is selected from R and K.

60. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa9 has high average flexibility.

61. The engineered AAV VP capsid polypeptide of embodiment 60, wherein Xaa9 is selected from R and G.

62. The engineered AAV VP capsid polypeptide of any one of embodiments 1-61, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 17118-SEQ ID NO: 18117.

63. The engineered AAV VP capsid polypeptide of embodiment 62, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence of any one of SEQ ID NO: 17118-SEQ ID NO: 18117.

64. The engineered AAV VP capsid polypeptide of any one of embodiments 1-63, wherein tropism for lung tissue is measured as a relative accumulation of the rAAV virion in a lung tissue as compared to a non-lung tissue, wherein the non-lung tissue consists collectively of CNS, liver, skeletal muscle, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord.

65. The engineered AAV VP capsid polypeptide of embodiment 64, wherein the CNS tissue is selected from forebrain cortex, occipital cortex, temporal cortex, thalamus, hypothalamus, substantia nigra, hippocampus DG, hippocampus CA1, hippocampus CA3, cerebellum, and any combination thereof.

66. The engineered AAV VP capsid polypeptide of any one of embodiments 64-65, wherein the higher tissue tropism is a 1.0005-fold to about a 1000-fold increased accumulation in the lung tissue as compared to a non-lung tissue.

67. The engineered AAV VP capsid polypeptide of embodiment 66, wherein the higher tissue tropism is at least about a 1.0005-fold, at least about a two-fold, at least about a three-fold, at least about a four-fold, at least about a five-fold, at least about a ten-fold, at least about a twenty-fold, at least about a 50-fold, at least about a 75-fold, at least about a 100-fold, or at least about a 1000-fold increased accumulation in the lung tissue as compared to a non-lung tissue.

Series J Embodiments—Lymph Node Tropic Capsids

1. An engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide having an amino acid sequence at least 70% identical to SEQ ID NO: 1,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV),

wherein the at least one mutation confers higher tropism for a lymph node tissue on the rAAV as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1, and

wherein amino acid residues Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV), and

wherein the engineered AAV VP capsid polypeptide does not have the sequence of any of SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8. 4. The engineered AAV VP capsid polypeptide of embodiment 3, wherein the rAAV has higher tropism for a lymph node tissue as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1. 5. The engineered AAV VP capsid polypeptide of embodiment 1, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 19118-SEQ ID NO: 22117. 6. The engineered AAV VP capsid polypeptide of embodiment 5, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence of any one of SEQ ID NO: 19118-SEQ ID NO: 22117. 7. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 19118-SEQ ID NO: 22117. 8. The engineered AAV VP capsid polypeptide of embodiment 7, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence of any one of SEQ ID NO: 19118-SEQ ID NO: 22117. 9. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein:

Xaa1 is selected from the group consisting of A, D, E, Q, S, and T; or

Xaa2 is selected from the group consisting of A, H, I, S, T, and V; or

Xaa3 is selected from the group consisting of A, E, H, I, T, and V; or

Xaa4 is selected from the group consisting of A, D, E, and P; or

Xaa5 is selected from the group consisting of I, L, M, V, and Y; or

Xaa6 is selected from the group consisting of D, E, I, N, and Q; or

Xaa7 is selected from the group consisting of A, E, G, Q, and V; or

Xaa8 is selected from the group consisting of F, G, M, and W; or

Xaa9 is selected from the group consisting of I, P, T, and Y; or

any combination thereof.

10. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is selected from D, E, and T.

11. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is E.

12. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is selected from I, T, and V.

13. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is V.

14. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is selected from A, I, T, and V.

15. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is T.

16. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is selected from D and E.

17. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is E.

18. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is selected from I, L, V, and Y.

19. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is L.

20. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is selected from D, E, and I.

21. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is D.

22. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 is A, Q, or V.

23. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 is V.

24. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is selected from F and W.

25. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is W.

26. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 is I or P.

27. The engineered AAV VP capsid polypeptide of any one of embodiments 1-26, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 19118-SEQ ID NO: 20117.

28. The engineered AAV VP capsid polypeptide of embodiment 27, wherein the region from residue 581 to residue 589 of SEQ ID NO: 1 has a sequence of any one of SEQ ID NO: 19118-SEQ ID NO: 20117.

29. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has high average flexibility.

30. The engineered AAV VP capsid polypeptide of embodiment 29, wherein Xaa1 is selected from D, E, P, G, Q, S, and R.

31. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has high hbond donors.

32. The engineered AAV VP capsid poly peptide of embodiment 31, wherein Xaa1 is R.

33. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has high mol mass.

34. The engineered AAV VP capsid polypeptide of embodiment 33, wherein Xaa1 is selected from Y, W, R, and F.

35. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has low solubility.

36. The engineered AAV VP capsid polypeptide of embodiment 35, wherein Xaa2 is selected from N and E.

37. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has low average flexibility.

38. The engineered AAV VP capsid polypeptide of embodiment 37, wherein Xaa3 is selected from W, M, and F.

39. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has low mutability.

40. The engineered AAV VP capsid polypeptide of embodiment 39, wherein Xaa3 is selected from R, H, K, P, Y, F, L, and C.

41. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has low mutability.

42. The engineered AAV VP capsid polypeptide of embodiment 41, wherein Xaa4 is C.

43. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has high mutability.

44. The engineered AAV VP capsid polypeptide of embodiment 43, wherein Xaa5 is N.

45. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has medium mol mass.

46. The engineered AAV VP capsid polypeptide of embodiment 45, wherein Xaa5 is selected from D, I, L, and N.

47. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has high mol mass.

48. The engineered AAV VP capsid polypeptide of embodiment 47, wherein Xaa6 is selected from Y, W, R, and F.

49. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has high average flexibility.

50. The engineered AAV VP capsid polypeptide of embodiment 49, wherein Xaa6 is selected from G and R.

51. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa7 has high average flexibility.

52. The engineered AAV VP capsid polypeptide of embodiment 51, wherein Xaa7 is selected from D, E, K, P, I, N, Q, and S.

53. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa7 has low solubility.

54. The engineered AAV VP capsid polypeptide of embodiment 53, wherein Xaa7 is selected from N and E.

55. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has low solubility.

56. The engineered AAV VP capsid polypeptide of embodiment 55, wherein Xaa8 is selected from N, E, and D.

57. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has medium mutability.

58. The engineered AAV VP capsid polypeptide of embodiment 57, wherein Xaa8 is selected from R and H.

59. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa9 has low mutability.

60. The engineered AAV VP capsid polypeptide of embodiment 59, wherein Xaa9 is selected from P and K.

61. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa9 has high average flexibility.

62. The engineered AAV VP capsid polypeptide of embodiment 61, wherein Xaa9 is selected from D, E, P, and S.

63. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa9 has high solubility.

64. The engineered AAV VP capsid polypeptide of embodiment 63, wherein Xaa9 is selected from M and V.

65. The engineered AAV VP capsid polypeptide of any one of embodiments 1-64, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 20118-SEQ ID NO: 21117.

66. The engineered AAV VP capsid polypeptide of embodiment 65, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence of any one of SEQ ID NO: 20118-SEQ ID NO: 21117.

67. The engineered AAV VP capsid polypeptide of any one of embodiments 1-66, wherein tropism for lymph node tissue is measured as a relative accumulation of the rAAV virion in a lymph node tissue as compared to a non-lymph node tissue, wherein the non-lymph node tissue consists collectively of CNS, liver, skeletal muscle, heart, lung, spleen, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord.

68. The engineered AAV VP capsid polypeptide of embodiment 67, wherein the CNS tissue is selected from forebrain cortex, occipital cortex, temporal cortex, thalamus, hypothalamus, substantia nigra, hippocampus DG, hippocampus CA1, hippocampus CA3, cerebellum, and any combination thereof.

69. The engineered AAV VP capsid polypeptide of any one of embodiments 67-68, wherein the higher tissue tropism is a 1.0005-fold to about a 1000-fold increased accumulation in the lymph node tissue as compared to a non-lymph node tissue.

70. The engineered AAV VP capsid polypeptide of embodiment 69, wherein the higher tissue tropism is at least about a 1.0005-fold, at least about a two-fold, at least about a three-fold, at least about a four-fold, at least about a five-fold, at least about a ten-fold, at least about a twenty-fold, at least about a 50-fold, at least about a 75-fold, at least about a 100-fold, or at least about a 1000-fold increased accumulation in the lymph node tissue as compared to a non-lymph node tissue.

Series K Embodiments—Mammary Gland Tropic Capsids

1. An engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide having an amino acid sequence at least 70% identical to SEQ ID NO: 1,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV),

wherein the at least one mutation confers higher tropism for a mammary gland tissue on the rAAV as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1, and

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV), and

wherein the engineered AAV VP capsid polypeptide does not have the sequence of any of SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6. SEQ ID NO: 7, and SEQ ID NO: 8. 4. The engineered AAV VP capsid polypeptide of embodiment 3, wherein the rAAV has higher tropism for a mammary gland tissue as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1.

5. The engineered AAV VP capsid polypeptide of embodiment 1, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 22118-SEQ ID NO: 25117.

6. The engineered AAV VP capsid polypeptide of embodiment 5, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence of any one of SEQ ID NO: 22118-SEQ ID NO: 25117.

7. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 22118-SEQ ID NO: 25117.

8. The engineered AAV VP capsid polypeptide of embodiment 7, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence of any one of SEQ ID NO: 22118-SEQ ID NO: 25117.

9. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein:

Xaa1 is selected from the group consisting of C, K, M, Q, R, and Y; or

Xaa2 is selected from the group consisting of A, F, I, K, S, T, and V; or

Xaa3 is selected from the group consisting of A, F, G, I, K, L, R, T, and Y; or

Xaa4 is selected from the group consisting of A, I, K, Q, R, and T; or

Xaa5 is selected from the group consisting of I, L, M, Q, R, T, V, and Y; or

Xaa6 is selected from the group consisting of H, N, S, and V; or

Xaa7 is selected from the group consisting of A, H, I, N, S and Y; or

Xaa8 is selected from the group consisting of A, C, D, G, H, M, Q, and S; or

Xaa9 is selected from the group consisting of A, E, L, W, and Y; or

any combination thereof.

10. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is selected from C, Q, and R.

11. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is C.

12. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is selected from A, S, and V.

13. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is V.

14. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is selected from F, G, K, R, and Y.

15. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is selected from F, K, and Y.

16. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is F.

17. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is selected from A, I, and R.

18. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is 1.

19. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is selected from I, M, and Y.

20. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is Y.

21. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is H.

22. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 is N or S.

23. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 is N.

24. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is selected from G, M, and Q.

25. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is G.

26. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 is selected from A, L, and W.

27. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 is A.

28. The engineered AAV VP capsid polypeptide of any one of embodiments 1-27, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 700%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 22118-SEQ ID NO: 23117.

29. The engineered AAV VP capsid polypeptide of embodiment 28, wherein the region from residue 581 to residue 589 of SEQ ID NO: 1 has a sequence of any one of SEQ ID NO: 22118-SEQ ID NO: 23117.

30. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has low surface accessibility.

31. The engineered AAV VP capsid polypeptide of embodiment 30, wherein Xaa1 is C.

32. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has medium mol mass.

33. The engineered AAV VP capsid polypeptide of embodiment 32, wherein Xaa1 is C.

34. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has high surface accessibility.

35. The engineered AAV VP capsid polypeptide of embodiment 34, wherein Xaa2 is selected from D, N, and Q.

36. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has low hydropathy.

37. The engineered AAV VP capsid polypeptide of embodiment 36, wherein Xaa2 is selected from D, E, R, K, H, N, and Q.

38. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has high average flexibility.

39. The engineered AAV VP capsid polypeptide of embodiment 38, wherein Xaa3 is selected from D, E, R, P, G, and S.

40. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has medium mutability.

41. The engineered AAV VP capsid polypeptide of embodiment 40, wherein Xaa3 is selected from R and H.

42. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has high mutability.

43. The engineered AAV VP capsid polypeptide of embodiment 42, wherein Xaa4 is selected from M, I, Q, and T.

44. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has high solubility.

45. The engineered AAV VP capsid polypeptide of embodiment 44, wherein Xaa4 is selected from W, F, I, and L.

46. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has high surface accessibility.

47. The engineered AAV VP capsid polypeptide of embodiment 46, wherein Xaa4 is selected from E, R, and K.

48. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has high solubility.

49. The engineered AAV VP capsid polypeptide of embodiment 48, wherein Xaa5 is selected from W, F, I, and L.

50. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has low mutability.

51. The engineered AAV VP capsid polypeptide of embodiment 50, wherein Xaa5 is selected from Y, F, and L.

52. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has high hydropathy.

53. The engineered AAV VP capsid polypeptide of embodiment 52, wherein Xaa6 is selected from V, I, and L.

54. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has medium mol mass.

55. The engineered AAV VP capsid polypeptide of embodiment 54, wherein Xaa6 is selected from D, I, L, and N.

56. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has low surface accessibility.

57. The engineered AAV VP capsid polypeptide of embodiment 56, wherein Xaa8 is C.

58. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has low mutability.

59. The engineered AAV VP capsid polypeptide of embodiment 58, wherein Xaa8 is selected from C, R, and H.

60. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has medium mutability.

61. The engineered AAV VP capsid polypeptide of embodiment 60, wherein Xaa9 is selected from R and H.

62. The engineered AAV VP capsid polypeptide of any one of embodiments 1-61, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 23118-SEQ ID NO: 24117.

63. The engineered AAV VP capsid polypeptide of embodiment 62, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence of any one of SEQ ID NO: 23118-SEQ ID NO: 24117.

64. The engineered AAV VP capsid polypeptide of any one of embodiments 1-63, wherein tropism for mammary gland tissue is measured as a relative accumulation of the rAAV virion in a mammary gland tissue as compared to a non-mammary gland tissue, wherein the non-mammary gland tissue consists collectively of CNS, liver, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord.

65. The engineered AAV VP capsid polypeptide of embodiment 65, wherein the CNS tissue is selected from forebrain cortex, occipital cortex, temporal cortex, thalamus, hypothalamus, substantia nigra, hippocampus DG, hippocampus CA1, hippocampus CA3, cerebellum, and any combination thereof.

66. The engineered AAV VP capsid polypeptide of any one of embodiments 64-65, wherein the higher tissue tropism is a 1.0005-fold to about a 1000-fold increased accumulation in the mammary gland tissue as compared to a non-mammary gland tissue.

67. The engineered AAV VP capsid polypeptide of embodiment 66, wherein the higher tissue tropism is at least about a 1.0005-fold, at least about a two-fold, at least about a three-fold, at least about a four-fold, at least about a five-fold, at least about a ten-fold, at least about a twenty-fold, at least about a 50-fold, at least about a 75-fold, at least about a 100-fold, or at least about a 1000-fold increased accumulation in the mammary gland tissue as compared to a non-mammary gland tissue.

Series L Embodiments—Skeletal or Cardiac Muscle Tropic Capsids

1. An engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide having an amino acid sequence at least 70% identical to SEQ ID NO: 1,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV),

wherein the at least one mutation confers higher tropism for a skeletal muscle tissue or a cardiac muscle tissue on the rAAV as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1, and

wherein amino acid residues Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV), and

Series M Embodiments—Sciatic Nerve Tropic Capsids

1. An engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide having an amino acid sequence at least 70% identical to SEQ ID NO: 1,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV),

wherein the at least one mutation confers higher tropism for a sciatic nerve tissue on the rAAV as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1, and

wherein amino acid residues Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV), and

wherein the engineered AAV VP capsid polypeptide does not have the sequence of any of SEQ ID NO: 1. SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8. 4. The engineered AAV VP capsid polypeptide of embodiment 3, wherein the rAAV has higher tropism for a sciatic nerve tissue as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1. 5. The engineered AAV VP capsid polypeptide of embodiment 1, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 26118-SEQ ID NO: 28990.

6. The engineered AAV VP capsid polypeptide of embodiment 5, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence of any one of SEQ ID NO: 26118-SEQ ID NO: 28990.

7. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 26118-SEQ ID NO: 28990.

8. The engineered AAV VP capsid polypeptide of embodiment 7, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence of any one of SEQ ID NO: 26118-SEQ ID NO: 28990.

9. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein:

Xaa1 is selected from the group consisting of C, G, K, M, Q, R, and Y; or

Xaa2 is selected from the group consisting of A, C, F, I, Q, T, and V; or

Xaa3 is selected from the group consisting of A, F, I, M, R, S, and T; or

Xaa4 is selected from the group consisting of E, N, T, Q, and V; or

Xaa5 is selected from the group consisting of F, H, Q, S, V, and Y; or

Xaa6 is selected from the group consisting of K, M, N, Q, S, and V; or

Xaa7 is selected from the group consisting of K, M, Q, R, and T; or

Xaa8 is selected from the group consisting of A, G, H, Q, S, and V; or

Xaa9 is selected from the group consisting of C, E, I, K, and R; or

any combination thereof.

10. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is selected from C, R, and Q.

11. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is C.

12. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is selected from A, C, and I.

13. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is A.

14. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is selected from F, M, R, and S.

15. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is R.

16. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is selected from E, T, and V.

17. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is T.

18. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is selected from F, V, and Y.

19. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is V.

20. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is selected from M, N, and S.

21. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is N.

22. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 is selected from M, Q, and T.

23. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 is M.

24. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is selected from H and S.

25. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is H.

26. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 is selected from C, I, and K.

27. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 is I.

28. The engineered AAV VP capsid polypeptide of any one of embodiments 1-27, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 26118-SEQ ID NO: 26990.

29. The engineered AAV VP capsid polypeptide of embodiment 28, wherein the region from residue 581 to residue 589 of SEQ ID NO: 1 has a sequence of any one of SEQ ID NO. 26118-SEQ ID NO: 26990.

30. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has high average flexibility.

31. The engineered AAV VP capsid polypeptide of embodiment 30, wherein Xaa1 is selected from G and R.

32. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has low solubility.

33. The engineered AAV VP capsid polypeptide of embodiment 32, wherein Xaa1 is selected from R and Q.

34. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has low mutability.

35. The engineered AAV VP capsid polypeptide of embodiment 34, wherein Xaa1 is selected from C, L, F, Y, R, K, P, and H.

36. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has high volume.

37. The engineered AAV VP capsid polypeptide of embodiment 36, wherein Xaa1 is selected from Y and F.

38. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has high surface accessibility.

39. The engineered AAV VP capsid polypeptide of embodiment 38, wherein Xaa2 is selected from E, R, and K.

40. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has medium mutability.

41. The engineered AAV VP capsid polypeptide of embodiment 40, wherein Xaa3 is selected from H and R.

42. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has medium average flexibility.

43. The engineered AAV VP capsid polypeptide of embodiment 42, wherein Xaa3 is selected from V and Y.

44. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has high mutability.

45. The engineered AAV VP capsid poly peptide of embodiment 44, wherein Xaa4 is N.

46. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has high average flexibility.

47. The engineered AAV VP capsid polypeptide of embodiment 46, wherein Xaa4 is selected from I, N, G, and R.

48. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has low solubility.

49. The engineered AAV VP capsid polypeptide of embodiment 48, wherein Xaa4 is N.

50. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has low mutability.

51. The engineered AAV VP capsid polypeptide of embodiment 50, wherein Xaa6 is selected from C, L, F, and Y.

52. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has high volume.

53. The engineered AAV VP capsid polypeptide of embodiment 52, wherein Xaa6 is selected from K, M, I, and L.

54. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa7 has low mutability.

55. The engineered AAV VP capsid polypeptide of embodiment 54, wherein Xaa7 is selected from L, F, and Y.

56. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa7 has medium mol mass.

57. The engineered AAV VP capsid polypeptide of embodiment 56, wherein Xaa7 is selected from D, I, L, and N.

58. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has high surface accessibility.

59. The engineered AAV VP capsid polypeptide of embodiment 58, wherein Xaa8 is selected from S, Y, T, D, P, H, and N.

60. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa9 has low mutability.

61. The engineered AAV VP capsid polypeptide of embodiment 60, wherein Xaa9 is selected from C, H, and R.

62. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa9 has medium solubility.

63. The engineered AAV VP capsid polypeptide of embodiment 62, wherein Xaa9 is selected from Q, T, and C.

64. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa9 has low surface accessibility.

65. The engineered AAV VP capsid polypeptide of embodiment 64, wherein Xaa9 is C.

66. The engineered AAV VP capsid polypeptide of any one of embodiments 1-65, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 26991-SEQ ID NO: 27990.

67. The engineered AAV VP capsid polypeptide of embodiment 66, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence of any one of SEQ ID NO: 26991-SEQ ID NO: 27990.

68. The engineered AAV VP capsid polypeptide of any one of embodiments 1-67, wherein tropism for sciatic nerve tissue is measured as a relative accumulation of the rAAV virion in a sciatic nerve tissue as compared to a non-sciatic nerve tissue, wherein the non-sciatic nerve tissue consists collectively of CNS, liver, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, and spinal cord.

69. The engineered AAV VP capsid polypeptide of any one of embodiments 68, wherein the CNS tissue is selected from forebrain cortex, occipital cortex, temporal cortex, thalamus, hypothalamus, substantia nigra, hippocampus DG, hippocampus CA1, hippocampus CA3, cerebellum, and any combination thereof.

70. The engineered AAV VP capsid polypeptide of any one of embodiments 68-69, wherein the higher tissue tropism is a 1.0005-fold to about a 1000-fold increased accumulation in the sciatic nerve tissue as compared to a non-sciatic nerve tissue.

71. The engineered AAV VP capsid polypeptide of embodiment 70, wherein the higher tissue tropism is at least about a 1.0005-fold, at least about a two-fold, at least about a three-fold, at least about a four-fold, at least about a five-fold, at least about a ten-fold, at least about a twenty-fold, at least about a 50-fold, at least about a 75-fold, at least about a 100-fold, or at least about a 1000-fold increased accumulation in the sciatic nerve tissue as compared to a non-sciatic nerve tissue.

Series N Embodiments—Skeletal Muscle Tropic Capsids

1. An engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide having an amino acid sequence at least 70% identical to SEQ ID NO: 1,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV),

wherein the at least one mutation confers higher tropism for a skeletal muscle tissue on the rAAV as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1, and

wherein amino acid residues Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV), and

wherein the engineered AAV VP capsid polypeptide does not have the sequence of any of SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8. 4. The engineered AAV VP capsid polypeptide of embodiment 3, wherein the rAAV has higher tropism for a skeletal muscle tissue as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1. 5. The engineered AAV VP capsid polypeptide of any one of embodiment 1, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 700%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 28991-SEQ ID NO: 31990. 6. The engineered AAV VP capsid polypeptide of embodiment 5, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence of any one of SEQ ID NO: 28991-SEQ ID NO: 31990. 7. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 28991-SEQ ID NO: 31990.

8. The engineered AAV VP capsid polypeptide of embodiment 7, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence of any one of SEQ ID NO: 28991-SEQ ID NO: 31990.

9. The engineered AAV VP capsid polypeptide of any one of embodiments X-Y, wherein:

Xaa1 is selected from the group consisting of A, E, H, M, P, Q, and S; or

Xaa2 is selected from the group consisting of F, H, I, T, and V; or

Xaa3 is selected from the group consisting of A, G, I, K, M, Q, R, S, T, and V; or

Xaa4 is selected from the group consisting of D, E, G, P, and S; or

Xaa5 is selected from the group consisting of H, L, M, P, and V; or

Xaa6 is selected from the group consisting of E, H, N, and P; or

Xaa7 is selected from the group consisting of A, H, N, Q and T; or

Xaa8 is selected from the group consisting of I, K, M, P, and W; or

Xaa9 is selected from the group consisting of A, I, M, P, and V; or any combination thereof.

10. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is selected from P and Q.

11. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is Q.

12. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is selected from T and V.

13. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is V.

14. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is selected from A, L, P, R, and T.

15. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is selected from L, P, and T.

16. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is P.

17. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is selected from D, E, and S.

18. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is E.

19. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is selected from L. M, and V.

20. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is L.

21. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is P.

22. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 is H.

23. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is selected from I, P, and W.

24. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is P.

25. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 is selected from A, M, and P.

26. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 is M.

27. The engineered AAV VP capsid polypeptide of any one of embodiments 1-26, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 28991-SEQ ID NO: 29990.

28. The engineered AAV VP capsid polypeptide of embodiment 27, wherein the region from residue 581 to residue 589 of SEQ ID NO: 1 has a sequence of any one of SEQ ID NO: 28991-SEQ ID NO: 29990.

29. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has high average flexibility.

30. The engineered AAV VP capsid polypeptide of embodiment 29, wherein Xaa1 is selected from G and R.

31. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has low average flexibility.

32. The engineered AAV VP capsid polypeptide of embodiment 31, wherein Xaa1 is selected from W, M, F, and H.

33. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has high mol mass.

34. The engineered AAV VP capsid polypeptide of embodiment 33, wherein Xaa1 is selected from R, F, and W.

35. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has low hydropathy.

36. The engineered AAV VP capsid polypeptide of embodiment 35, wherein Xaa2 is selected from K and R.

37. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has low mutability.

38. The engineered AAV VP capsid polypeptide of embodiment 37, wherein Xaa2 is selected from C, R, and H.

39. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has high average flexibility.

40. The engineered AAV VP capsid polypeptide of embodiment 39, wherein Xaa2 is selected from G and R.

41. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has high average flexibility.

42. The engineered AAV VP capsid polypeptide of embodiment 41, wherein Xaa3 is selected from G and R.

43. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has high hydrophilicity.

44. The engineered AAV VP capsid polypeptide of embodiment 43, wherein Xaa4 is selected from D, E, R, K, and N.

45. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has low mutability.

46. The engineered AAV VP capsid polypeptide of embodiment 45, wherein Xaa4 is selected from C, R, and H.

47. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has low mol mass.

48. The engineered AAV VP capsid polypeptide of embodiment 47, wherein Xaa5 is A.

49. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has low average flexibility.

50. The engineered AAV VP capsid polypeptide of embodiment 49, wherein Xaa5 is selected from A and L.

51. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has high mutability.

52. The engineered AAV VP capsid polypeptide of embodiment 51, wherein Xaa5 is selected from D. A, and E.

53. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has low average flexibility.

54. The engineered AAV VP capsid polypeptide of embodiment 53, wherein Xaa6 is selected from W, M, and F.

55. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has low mutability.

56. The engineered AAV VP capsid polypeptide of embodiment 55, wherein Xaa6 is C.

57. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has high mol mass.

58. The engineered AAV VP capsid polypeptide of embodiment 57, wherein Xaa6 is W.

59. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa7 has low goldman engelman steitz.

60. The engineered AAV VP capsid polypeptide of embodiment 59, wherein Xaa7 is R.

61. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa7 has high average flexibility.

62. The engineered AAV VP capsid polypeptide of embodiment 61, wherein Xaa7 is selected from D, R, P, G, and S.

63. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa7 has high mutability.

64. The engineered AAV VP capsid polypeptide of embodiment 63, wherein Xaa7 is selected from R, H, and N.

65. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa7 has low solubility.

66. The engineered AAV VP capsid polypeptide of embodiment 65, wherein Xaa7 is selected from R and Q.

67. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has high hydrophilicity.

68. The engineered AAV VP capsid polypeptide of embodiment 67, wherein Xaa8 is selected from D, E, R, K, and N.

69. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa9 has low mutability.

70. The engineered AAV VP capsid polypeptide of embodiment 69, wherein Xaa9 is selected from Y. F, and L.

71. The engineered AAV VP capsid polypeptide of any one of embodiments 1-70, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 900%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 29991-SEQ ID NO: 30990.

72. The engineered AAV VP capsid polypeptide of embodiment 71, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence of any one of SEQ ID NO: 29991-SEQ ID NO: 30990.

73. The engineered AAV VP capsid polypeptide of any one of embodiments 1-72, wherein tropism for skeletal muscle tissue is measured as a relative accumulation of the rAAV virion in a skeletal muscle tissue as compared to a non-skeletal muscle tissue, wherein the non-skeletal muscle tissue consists collectively of CNS, liver, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord.

74. The engineered AAV VP capsid polypeptide of embodiment 73, wherein the CNS tissue is selected from forebrain cortex, occipital cortex, temporal cortex, thalamus, hypothalamus, substantia nigra, hippocampus DG, hippocampus CA1, hippocampus CA3, cerebellum, and any combination thereof.

75. The engineered AAV VP capsid polypeptide of any one of embodiments 73 and 74, wherein the higher tissue tropism is a 1.0005-fold to about a 1000-fold increased accumulation in the skeletal muscle tissue as compared to a non-skeletal muscle tissue.

76. The engineered AAV VP capsid polypeptide of embodiment 75, wherein the higher tissue tropism is at least about a 1.0005-fold, at least about a two-fold, at least about a three-fold, at least about a four-fold, at least about a five-fold, at least about a ten-fold, at least about a twenty-fold, at least about a 50-fold, at least about a 75-fold, at least about a 100-fold, or at least about a 1000-fold increased accumulation in the skeletal muscle tissue as compared to a non-skeletal muscle tissue.

Series O Embodiments—Skin Tropic Capsids

1. An engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide having an amino acid sequence at least 70% identical to SEQ ID NO: 1,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV),

wherein the at least one mutation confers higher tropism for a skin tissue on the rAAV as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1, and

wherein amino acid residues Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV), and

6. The engineered AAV VP capsid polypeptide of embodiment 5, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence of any one of SEQ ID NO: 31991-SEQ ID NO: 34990.

7. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 31991-SEQ ID NO: 34990.

8. The engineered AAV VP capsid polypeptide of embodiment 7, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence of any one of SEQ ID NO: 31991-SEQ ID NO: 34990

9. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein:

Xaa1 is selected from the group consisting of A, C, K, Q, R, and T; or

Xaa2 is selected from the group consisting of A, C, I, S, T, and V; or

Xaa3 is selected from the group consisting of A, C, F, G, M, Q, S, and V; or

Xaa4 is selected from the group consisting of C, K, L, P, R, and W; or

Xaa5 is selected from the group consisting of F, H, I, M, V, and Y; or

Xaa6 is selected from the group consisting of F, H, I, M, N, Q, and S; or

Xaa7 is selected from the group consisting of A, H, K, M, N, R, and V; or

Xaa8 is selected from the group consisting of A, F, G, H, S, and Y; or

Xaa9 is selected from the group consisting of A, E, G, P, Q, R, and S; or

any combination thereof.

10. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is selected from C, K, and R.

11. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is C.

12. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is selected from A, S, T, and V.

13. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is V.

14. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is selected from A, C, F, M, and Q.

15. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is C.

16. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is selected from L, P, and R.

17. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is R.

18. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is selected from M, V, and Y.

19. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is Y.

20. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is selected from M, N, and Q.

21. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is N.

22. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 is A, H, K, or R.

23. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 is K.

24. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is selected from A, F, and S.

25. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is S.

26. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 is selected from A, Q, and S.

27. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 is A.

28. The engineered AAV VP capsid polypeptide of any one of embodiments 1-27, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 31991-SEQ ID NO: 32990.

29. The engineered AAV VP capsid polypeptide of embodiment 28 wherein the region from residue 581 to residue 589 of SEQ ID NO: 1 has a sequence of any one of SEQ ID NO: 31991-SEQ ID NO: 32990.

30. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has low surface accessibility.

31. The engineered AAV VP capsid polypeptide of embodiment 30, wherein Xaa1 is C.

32. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has low volume.

33. The engineered AAV VP capsid polypeptide of embodiment 32, wherein Xaa1 is C.

34. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has low mutability.

35. The engineered AAV VP capsid polypeptide of embodiment 34, wherein Xaa1 is C.

36. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has high surface accessibility.

37. The engineered AAV VP capsid polypeptide of embodiment 36, wherein Xaa2 is selected from R and K.

38. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has high average flexibility.

39. The engineered AAV VP capsid polypeptide of embodiment 38, wherein Xaa2 is selected from K, I, and N.

40. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has low mutability.

41. The engineered AAV VP capsid polypeptide of embodiment 40, wherein Xaa2 is selected from P and K.

42. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has high hydropathy.

43. The engineered AAV VP capsid polypeptide of embodiment 42, wherein Xaa3 is selected from I and V.

44. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has low mutability.

45. The engineered AAV VP capsid polypeptide of embodiment 44, wherein Xaa4 is selected from L, F, and Y.

46. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has low average flexibility.

47. The engineered AAV VP capsid polypeptide of embodiment 46, wherein Xaa4 is selected from W, H, F, and M.

48. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has high average flexibility.

49. The engineered AAV VP capsid polypeptide of embodiment 48, wherein Xaa5 is selected from G, R, K, I, and N.

50. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has high average flexibility.

51. The engineered AAV VP capsid polypeptide of embodiment 50, wherein Xaa6 is selected from G, R, K, I, and N.

52. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has high surface accessibility.

53. The engineered AAV VP capsid polypeptide of embodiment 52, wherein Xaa8 is selected from M, G, and F.

54. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has low average flexibility.

55. The engineered AAV VP capsid polypeptide of embodiment 54, wherein Xaa8 is selected from H, F, M, and W.

56. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has low mutability.

57. The engineered AAV VP capsid polypeptide of embodiment 56, wherein Xaa8 is selected from L, F, and Y.

58. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa9 has high average flexibility.

59. The engineered AAV VP capsid polypeptide of embodiment 58, wherein Xaa9 is selected from D, E, R, K, P, and G.

60. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa9 has high mutability.

61. The engineered AAV VP capsid polypeptide of embodiment 60, wherein Xaa9 is selected from D, E, R, V, A, and H.

62. The engineered AAV VP capsid polypeptide of any one of embodiments 1-61, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 32991-SEQ ID NO: 33990.

63. The engineered AAV VP capsid polypeptide of embodiment 62, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence of any one of SEQ ID NO: 32991-SEQ ID NO: 33990.

64. The engineered AAV VP capsid polypeptide of any one of embodiments 1-61, wherein tropism for skin tissue is measured as a relative accumulation of the rAAV virion in a skin tissue as compared to a non-skin tissue, wherein the non-skin tissue consists collectively of CNS, liver, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord.

65. The engineered AAV VP capsid polypeptide of embodiment 64, wherein the CNS tissue is selected from forebrain cortex, occipital cortex, temporal cortex, thalamus, hypothalamus, substantia nigra, hippocampus DG, hippocampus CA1, hippocampus CA3, cerebellum, and any combination thereof.

66. The engineered AAV VP capsid polypeptide of any one of embodiments 64-65, wherein the higher tissue tropism is a 1.0005-fold to about a 1000-fold increased accumulation in the skin tissue as compared to a non-skin tissue.

67. The engineered AAV VP capsid polypeptide of embodiment 66, wherein the higher tissue tropism is at least about a 1.0005-fold, at least about a two-fold, at least about a three-fold, at least about a four-fold, at least about a five-fold, at least about a ten-fold, at least about a twenty-fold, at least about a 50-fold, at least about a 75-fold, at least about a 100-fold, or at least about a 1000-fold increased accumulation in the skin tissue as compared to a non-skin tissue.

Series P Embodiments—Spinal Cord Tropic Capsids

1. An engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide having an amino acid sequence at least 70% identical to SEQ ID NO: 1,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV),

wherein the at least one mutation confers higher tropism for a spinal cord tissue on the rAAV as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1, and

wherein amino acid residues Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV), and

wherein the engineered AAV VP capsid polypeptide does not have the sequence of any of SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8. 4. The engineered AAV VP capsid polypeptide of embodiment 3, wherein the rAAV has higher tropism for a spinal cord tissue as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1. 5. The engineered AAV VP capsid polypeptide of embodiment 1, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 34991-SEQ ID NO: 37437. 6. The engineered AAV VP capsid polypeptide of embodiment 5, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence of any one of SEQ ID NO: 34991-SEQ ID NO: 37437. 7. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 34991-SEQ ID NO: 37437.

8. The engineered AAV VP capsid polypeptide of embodiment 7, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence of any one of SEQ ID NO: 34991-SEQ ID NO: 37437.

9. The engineered AAV VP capsid polypeptide of any one of embodiments X-Y, wherein:

Xaa1 is selected from the group consisting of A, C, K, Q, R, S, and W; or

Xaa2 is selected from the group consisting of H, I, K, L, T, V, and W; or

Xaa3 is selected from the group consisting of C, F, G, H, I, K, N, and R; or

Xaa4 is selected from the group consisting of I, M, Q, S, and V; or

Xaa5 is selected from the group consisting of H, K, Q, T, W, and Y; or

Xaa6 is selected from the group consisting of H, L, N, Q, R, W, and Y; or

Xaa7 is selected from the group consisting of D, H, P, Q, and R; or

Xaa8 is selected from the group consisting of D, F, L, S, T, and Y; or

Xaa9 is selected from the group consisting of C, I, N, P, R, S, and Y; or

any combination thereof.

10. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is selected from K, R, and W.

11. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is K.

12. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is selected from H, I, and T.

13. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is I.

14. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is selected from F. 1, and R.

15. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is I.

16. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is selected from 1, M, and V.

17. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is V.

18. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is selected from T, W, and Y.

19. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is Y.

20. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is selected from L, N, R, and Y.

21. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is Y.

22. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 is R.

23. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is selected from S, T, and Y.

24. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is T.

25. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 is selected from I, P, and R.

26. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 is I.

27. The engineered AAV VP capsid polypeptide of any one of embodiments 1-26, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 34991-SEQ ID NO: 35437.

28. The engineered AAV VP capsid polypeptide of embodiment 27, wherein the region from residue 581 to residue 589 of SEQ ID NO: 1 has a sequence of any one of SEQ ID NO: 34991-SEQ ID NO: 35437.

29. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has high volume.

30. The engineered AAV VP capsid polypeptide of embodiment 29, wherein Xaa1 is selected from F. W, and Y.

31. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has low mutability.

32. The engineered AAV VP capsid polypeptide of embodiment 31, wherein Xaa1 is selected from Y, F, L, and C.

33. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has high solubility.

34. The engineered AAV VP capsid polypeptide of embodiment 33, wherein Xaa1 is selected from W, F, I, and L.

35. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has low average flexibility.

36. The engineered AAV VP capsid polypeptide of embodiment 35, wherein Xaa1 is selected from F, M, and W.

37. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has low hydropathy.

38. The engineered AAV VP capsid polypeptide of embodiment 37, wherein Xaa2 is selected from P and Y.

39. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has low hydrophilicity.

40. The engineered AAV VP capsid polypeptide of embodiment 39, wherein Xaa3 is selected from Y, W, V, M, F, I, and L.

41. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has high solubility.

42. The engineered AAV VP capsid polypeptide of embodiment 41, wherein Xaa3 is selected from W, F, I, and L.

43. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has high volume.

44. The engineered AAV VP capsid polypeptide of embodiment 43, wherein Xaa6 is selected from W, R, K, M, I, and L.

45. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has high mol mass.

46. The engineered AAV VP capsid polypeptide of embodiment 45, wherein Xaa6 is selected from W.

47. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has high mol mass.

48. The engineered AAV VP capsid polypeptide of embodiment 47, wherein Xaa8 is selected from W, E, K, M, H, and Q.

49. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has high volume.

50. The engineered AAV VP capsid polypeptide of embodiment 49, wherein Xaa8 is selected from W, K, M, I, and L.

51. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has high goldman engelman steitz.

52. The engineered AAV VP capsid polypeptide of embodiment 51, wherein Xaa8 is selected from V and L.

53. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa9 has high hydropathy.

54. The engineered AAV VP capsid polypeptide of embodiment 53, wherein Xaa9 is selected from V and I.

55. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa9 has high solubility.

56. The engineered AAV VP capsid polypeptide of embodiment 55, wherein Xaa9 is selected from W, F, I, and L.

57. The engineered AAV VP capsid polypeptide of any one of embodiments 1-56, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 35438-SEQ ID NO: 36437.

58. The engineered AAV VP capsid polypeptide of embodiment 57, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence of any one of SEQ ID NO: 35438-SEQ ID NO: 36437.

59. The engineered AAV VP capsid polypeptide of any one of embodiments 1-58, wherein tropism for spinal cord tissue is measured as a relative accumulation of the rAAV virion in a spinal cord tissue as compared to a non-spinal cord tissue, wherein the non-spinal cord tissue consists collectively of CNS, liver, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, and sciatic nerve.

60. The engineered AAV VP capsid polypeptide of embodiment 59, wherein the CNS tissue is selected from forebrain cortex, occipital cortex, temporal cortex, thalamus, hypothalamus, substantia nigra, hippocampus DG, hippocampus CA1, hippocampus CA3, cerebellum, and any combination thereof.

61. The engineered AAV VP capsid polypeptide of any one of embodiments 59-60, wherein the higher tissue tropism is a 1.0005-fold to about a 1000-fold increased accumulation in the spinal cord tissue as compared to a non-spinal cord tissue.

62. The engineered AAV VP capsid polypeptide of embodiment 61, wherein the higher tissue tropism is at least about a 1.0005-fold, at least about a two-fold, at least about a three-fold, at least about a four-fold, at least about a five-fold, at least about a ten-fold, at least about a twenty-fold, at least about a 50-fold, at least about a 75-fold, at least about a 100-fold, or at least about a 1000-fold increased accumulation in the spinal cord tissue as compared to a non-spinal cord tissue.

Series Q Embodiments—Spleen Tropic Capsids

1. An engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide having an amino acid sequence at least 70% identical to SEQ ID NO: 1,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV),

wherein the at least one mutation confers higher tropism for a spleen tissue on the rAAV as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1, and

wherein the engineered AAV VP capsid polypeptide does not have the sequence of any of SEQ ID NO: 3. SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8. 2. The engineered AAV VP capsid polypeptide of embodiment 1, wherein the engineered AAV VP capsid polypeptide has a sequence of SEQ ID NO: 2 and wherein Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from any amino acid. 3. An engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide having an amino acid sequence of SEQ ID NO: 2,

wherein amino acid residues Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV), and

7. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 37438-SEQ ID NO: 40437.

8. The engineered AAV VP capsid polypeptide of embodiment 7, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence of any one of SEQ ID NO: 37438-SEQ ID NO: 40437.

9. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein:

Xaa1 is selected from the group consisting of C, F, H, I, L, P, W, and Y; or

Xaa2 is selected from the group consisting of D, E, L, N, P, R, and W; or

Xaa3 is selected from the group consisting of C, D, E, P, and W; or

Xaa4 is selected from the group consisting of C, F, G, H, R, W and Y; or

Xaa5 is selected from the group consisting of A, D, E, G, P, R, and W; or

Xaa6 is selected from the group consisting of A, C, D, E, K, R, and W; or

Xaa7 is selected from the group consisting of F, L, P, R, W, and Y; or

Xaa8 is selected from the group consisting of E, I, K, L, P, R, and T; or

Xaa9 is selected from the group consisting of C, H, M, T, V, and W; or

any combination thereof.

10. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is selected from C, F, P, W, and Y.

11. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is selected from P, W, and Y.

12. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is P.

13. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is selected from D, E, and W.

14. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is D.

15. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is selected from D, P, and W.

16. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is P.

17. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is selected from C, H, and W.

18. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is C.

19. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is selected from D, E, G, and P.

20. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is D.

21. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is selected from C, K, and R.

22. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is K.

23. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 is selected from L, P, and W.

24. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 is P.

25. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is selected from P, R, and K.

26. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is K.

27. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 is selected from C, T, and V.

28. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 is V.

29. The engineered AAV VP capsid polypeptide of any one of embodiments 1-28, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 37438-SEQ ID NO: 38437.

30. The engineered AAV VP capsid polypeptide of embodiment 29, wherein the region from residue 581 to residue 589 of SEQ ID NO: 1 has a sequence of any one of SEQ ID NO: 37438-SEQ ID NO: 38437.

31. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has low solubility.

32. The engineered AAV VP capsid polypeptide of embodiment 31, wherein Xaa1 is selected from D and P.

33. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has high solubility.

34. The engineered AAV VP capsid polypeptide of embodiment 33, wherein Xaa1 is selected from F, I, and L.

35. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has low hydropathy.

36. The engineered AAV VP capsid polypeptide of embodiment 35, wherein Xaa1 is selected from Y and P.

37. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has low mutability.

38. The engineered AAV VP capsid polypeptide of embodiment 37, wherein Xaa1 is selected from C, K, and P.

39. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has low solubility.

40. The engineered AAV VP capsid polypeptide of embodiment 39, wherein Xaa2 is selected from D, Q, and R.

41. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has low hydropathy.

42. The engineered AAV VP capsid polypeptide of embodiment 41, wherein Xaa2 is selected from D, E, R, K, H, N, and Q.

43. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has low charge.

44. The engineered AAV VP capsid polypeptide of embodiment 43, wherein Xaa2 is selected from D and E.

45. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has low volume.

46. The engineered AAV VP capsid polypeptide of embodiment 45, wherein Xaa2 is selected from T, N, P, and D.

47. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has high average flexibility.

48. The engineered AAV VP capsid polypeptide of embodiment 47, wherein Xaa2 is selected from D, E, R, P, G, Q, and S.

49. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has low solubility.

50. The engineered AAV VP capsid polypeptide of embodiment 49, wherein Xaa3 is selected from D, E, P, and N.

51. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has low hydropathy.

52. The engineered AAV VP capsid polypeptide of embodiment 51, wherein Xaa3 is selected from D, E, H, N, Q, and P.

53. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has low hydropathy.

54. The engineered AAV VP capsid polypeptide of embodiment 53, wherein Xaa4 is selected from K and R.

55. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has low solubility.

56. The engineered AAV VP capsid polypeptide of embodiment 55, wherein Xaa5 is selected from D, E, P, and N.

57. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has high average flexibility.

58. The engineered AAV VP capsid polypeptide of embodiment 57, wherein Xaa5 is selected from D, E, R, P, G, Q, and S.

59. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has low mutability.

60. The engineered AAV VP capsid polypeptide of embodiment 59, wherein Xaa6 is selected from C.

61. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has high surface accessibility.

62. The engineered AAV VP capsid polypeptide of embodiment 61, wherein Xaa8 is selected from E, R, and K.

63. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has low solubility.

64. The engineered AAV VP capsid polypeptide of embodiment 63, wherein Xaa8 is selected from E, P, R, K, N, Q.

65. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has medium volume.

66. The engineered AAV VP capsid polypeptide of embodiment 65, wherein Xaa8 is selected from E, D, R, K, V, P, M, I, L, H, N, Q, and T.

67. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa9 has medium mol mass.

68. The engineered AAV VP capsid polypeptide of embodiment 67, wherein Xaa9 is selected from E, D, K, M, I, L, H, N.

69. The engineered AAV VP capsid polypeptide of any one of embodiments 1-68, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 38438-SEQ ID NO: 39437.

70. The engineered AAV VP capsid polypeptide of embodiment 69, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence of any one of SEQ ID NO: 38438-SEQ ID NO: 39437.

71. The engineered AAV VP capsid polypeptide of any one of embodiments 1-70, wherein tropism for spleen tissue is measured as a relative accumulation of the rAAV virion in a spleen tissue as compared to a non-spleen tissue, wherein the non-spleen tissue consists collectively of CNS, liver, skeletal muscle, heart, lung, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord.

72. The engineered AAV VP capsid polypeptide of embodiment 71, wherein the CNS tissue is selected from forebrain cortex, occipital cortex, temporal cortex, thalamus, hypothalamus, substantia nigra, hippocampus DG, hippocampus CA1, hippocampus CA3, cerebellum, and any combination thereof.

73. The engineered AAV VP capsid polypeptide of any one of embodiments 71-72, wherein the higher tissue tropism is a 1.0005-fold to about a 1000-fold increased accumulation in the spleen tissue as compared to a non-spleen tissue.

74. The engineered AAV VP capsid polypeptide of embodiment 73, wherein the higher tissue tropism is at least about a 1.0005-fold, at least about a two-fold, at least about a three-fold, at least about a four-fold, at least about a five-fold, at least about a ten-fold, at least about a twenty-fold, at least about a 50-fold, at least about a 75-fold, at least about a 100-fold, or at least about a 1000-fold increased accumulation in the spleen tissue as compared to a non-spleen tissue.

Series R Embodiments—Thyroid Gland Tropic Capsids

1. An engineered adeno-associated virus (AAV) viral protein (VP) capsid polypeptide having an amino acid sequence at least 70% identical to SEQ ID NO: 1,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV),

wherein the at least one mutation confers higher tropism for a thyroid gland tissue on the rAAV as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1, and

wherein amino acid residues Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V,

wherein the engineered AAV VP capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV), and

wherein the engineered AAV VP capsid polypeptide does not have the sequence of any of SEQ ID NO: 1, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8. 4. The engineered AAV VP capsid polypeptide of embodiment 3, wherein the rAAV has higher tropism for a thyroid gland tissue as compared to an rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1. 5. The engineered AAV VP capsid polypeptide of embodiment 1, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 40438-SEQ ID NO: 43437. 6. The engineered AAV VP capsid polypeptide of embodiment 5, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence of any one of SEQ ID NO: 40438-SEQ ID NO: 43437. 7. The engineered AAV VP capsid polypeptide of embodiment 2-4, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 40438-SEQ ID NO: 43437. 8. The engineered AAV VP capsid polypeptide of embodiment 7, wherein the Xaa1 to Xaa9 region of SEQ ID NO: 2 has a sequence of any one of SEQ ID NO: 40438-SEQ ID NO: 43437.

9. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein:

Xaa1 is selected from the group consisting of A, K, M, N, Q, or R; or

Xaa2 is selected from the group consisting of A, F, K, L, M, T, V, or W; or

Xaa3 is selected from the group consisting of A, I, K, R, S, T, V, or W; or

Xaa4 is selected from the group consisting of A, D, E, I, P, or V; or

Xaa5 is selected from the group consisting of F, I, M, Q, V, or Y; or

Xaa6 is selected from the group consisting of H, M, N, or Y; or

Xaa7 is selected from the group consisting of H, I, N, Q, S, or W; or

Xaa8 is selected from the group consisting of A, D, F, Q, S, or Y; or

Xaa9 is selected from the group consisting of A, Q, S, or Y; or

any combination thereof.

10. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is selected from K, N and Q.

11. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa1 is K.

12. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is selected from F, V, and W.

13. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa2 is W.

14. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is selected from A, R and T.

15. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa3 is R.

16. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is selected from A, E, and I.

17. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa4 is A.

18. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is M, V, or Y.

19. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa5 is M.

20. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa6 is N.

21. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 is selected from H. I, and N.

22. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa7 is H.

23. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is selected from A, F, and S.

24. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa8 is F.

25. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 is selected from A and S.

26. The engineered AAV VP capsid polypeptide of embodiment 9, wherein Xaa9 is A.

27. The engineered AAV VP capsid polypeptide of any one of embodiments 1-26, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589, inclusive, has a sequence that is at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 40438-SEQ ID NO: 41437.

28. The engineered AAV VP capsid polypeptide of embodiment 27, wherein the region from residue 581 to residue 589 of SEQ ID NO: 1 has a sequence of any one of SEQ ID NO: 40438-SEQ ID NO: 41437.

29. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa1 has high mutability.

30. The engineered AAV VP capsid poly peptide of embodiment 29, wherein Xaa1 is N.

31. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa2 has low surface accessibility.

32. The engineered AAV VP capsid polypeptide of embodiment 31, wherein Xaa2 is selected from F, G, and M.

33. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has high solubility.

34. The engineered AAV VP capsid polypeptide of embodiment 33, wherein Xaa3 is selected from F.

35. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has low mutability.

36. The engineered AAV VP capsid polypeptide of embodiment 35, wherein Xaa3 is selected from Y, F, L, and C.

37. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has medium mol mass.

38. The engineered AAV VP capsid polypeptide of embodiment 37, wherein Xaa3 is selected from D, E, R, K, V, P, M, I, L, N, Q, T, and C.

39. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa3 has low surface accessibility.

40. The engineered AAV VP capsid polypeptide of embodiment 39, wherein Xaa3 is selected from V, I, L, and C.

41. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has high goldman engelman steitz.

42. The engineered AAV VP capsid polypeptide of embodiment 41, wherein Xaa4 is selected from L and V.

43. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has low surface accessibility.

44. The engineered AAV VP capsid polypeptide of embodiment 43, wherein Xaa4 is selected from V, M, A, G, F, I, and L.

45. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa4 has low mol mass.

46. The engineered AAV VP capsid polypeptide of embodiment 45, wherein Xaa4 is selected from D, A, G, I, L, and N.

47. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has high solubility.

48. The engineered AAV VP capsid polypeptide of embodiment 47, wherein Xaa5 is selected from C, L, F, M, V, and Y.

49. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has low solubility.

50. The engineered AAV VP capsid polypeptide of embodiment 49, wherein Xaa5 is selected from D.

51. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa5 has low average flexibility.

52. The engineered AAV VP capsid polypeptide of embodiment 51, wherein Xaa5 is selected from F, M, and W.

53. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa6 has low average flexibility.

54. The engineered AAV VP capsid polypeptide of embodiment 53, wherein Xaa6 is selected from F, M, and W.

55. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa7 has high mutability.

56. The engineered AAV VP capsid polypeptide of embodiment 55, wherein Xaa7 is selected from N.

57. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa7 has low volume.

58. The engineered AAV VP capsid polypeptide of embodiment 57, wherein Xaa7 is selected from P, N, and T.

59. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has low average flexibility.

60. The engineered AAV VP capsid polypeptide of embodiment 59, wherein Xaa8 is selected from F, M, and W.

61. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa8 has low surface accessibility.

62. The engineered AAV VP capsid polypeptide of embodiment 61, wherein Xaa8 is selected from M, G, and F.

63. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa9 has low mutability.

64. The engineered AAV VP capsid polypeptide of embodiment 63, wherein Xaa9 is selected from R, K, P, H, and C.

65. The engineered AAV VP capsid polypeptide of any one of embodiments 2-4, wherein Xaa9 has low hydropathy.

66. The engineered AAV VP capsid polypeptide of embodiment 65, wherein Xaa9 is selected from R.

67. The engineered AAV VP capsid polypeptide of any one of embodiments 1-66, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence having at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 92%, at least 95%, at least 97%, or at least 99% identical to any one of SEQ ID NO: 41438-SEQ ID NO: 42437.

68. The engineered AAV VP capsid polypeptide of embodiment 67, wherein the region from the residue corresponding to residue 581 to the residue corresponding to residue 589 inclusive has a sequence of any one of SEQ ID NO: 41438-SEQ ID NO: 42437.

69. The engineered AAV VP capsid polypeptide of any one of embodiments 1-68, wherein tropism for thyroid gland tissue is measured as a relative accumulation of the rAAV virion in a thyroid gland tissue as compared to a non-thyroid gland tissue, wherein the non-thyroid gland tissue consists collectively of CNS, liver, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, colon, sciatic nerve, and spinal cord.

70. The engineered AAV VP capsid polypeptide of embodiment 69, wherein the CNS tissue is selected from forebrain cortex, occipital cortex, temporal cortex, thalamus, hypothalamus, substantia nigra, hippocampus DG, hippocampus CA1, hippocampus CA3, cerebellum, and any combination thereof.

71. The engineered AAV VP capsid polypeptide of any one of embodiments 69-70, wherein the higher tissue tropism is a 1.0005-fold to about a 1000-fold increased accumulation in the thyroid gland tissue as compared to a non-thyroid gland tissue.

72. The engineered AAV VP capsid polypeptide of embodiment 71, wherein the higher tissue tropism is at least about a 1.0005-fold, at least about a two-fold, at least about a three-fold, at least about a four-fold, at least about a five-fold, at least about a ten-fold, at least about a twenty-fold, at least about a 50-fold, at least about a 75-fold, at least about a 100-fold, or at least about a 1000-fold increased accumulation in the thyroid gland tissue as compared to a non-thyroid gland tissue.

Series S Embodiments

1. An engineered adeno-associated virus (AAV) VP capsid polypeptide having the amino acid sequence of SEQ ID NO:2,

wherein amino acid residues Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V:

wherein the capsid polypeptide is capable of assembling into a recombinant AAV

virion (rAAV); and

wherein the polypeptide does not have the sequence of any of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, and SEQ ID NO:8.

2. An engineered adeno-associated virus (AAV) VP capsid polypeptide having at least one mutation in a residue corresponding to residue 581 to residue 589 in SEQ ID NO: 1, wherein the capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV) and wherein the polypeptide mutation confers tissue tropism on the recombinant rAAV for a first tissue as compared to a second tissue and wherein the AAV VP capsid polypeptide does not have the sequence of any of SEQ ID NO:3. SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, and SEQ ID NO:8.

3. An engineered AAV VP capsid polypeptide comprising a polypeptide sequence represented by the formula. (A)-(X)-(B)

- wherein:
- (A) is the polypeptide sequence of SEQ ID NO:47438 (VAYNVGGQMATNNQSSTTAP residues 561-580 of SEQ ID NO:2);
- (X) is the polypeptide sequence comprising amino acid residues Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 of SEQ ID NO:2; and
- (B) is the polypeptide sequence of SEQ ID NO:47439 (IVPGSVWMERDVYLQGPIWA residues 590-609 of SEQ ID NO:2;

wherein Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are each independently selected from A, R, N, D, C, E, Q, G, H, I, L, K, M, F, P, S, T, W, Y, and V; and

wherein the capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV); and;

wherein the polypeptide does not have the sequence of any of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, and SEQ ID NO:8.

4. The engineered AAV VP capsid polypeptide of embodiment 3, comprising a polypeptide sequence represented by the formula: (A)-(X)-(B) wherein:

- (A) is the polypeptide sequence of SEQ ID NO: 47438 (residues 561-580 of SEQ ID NO: 2 VAYNVGGQMATNNQSSTTAP);
- (X) is a polypeptide sequence selected from the list of polypeptides in Table 8 (SEQ ID NOs:115-1114)¹or Table 10 (SEQ ID NOs: 7118-8117) that confers CNS tissue tropism on a recombinant AAV virion (rAAV); and
- (B) is the polypeptide sequence of SEQ ID NO: 47439 (residues 590-609 of SEQ ID NO: 2: (IVPGSVWMERDVYLQGPIWA)); and

wherein the capsid polypeptide is capable of assembling into the rAAV and, the capsid does not have the sequence of any of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4. SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, and SEQ ID NO:8. 5. The engineered AAV VP capsid polypeptide of embodiment 4, wherein the CNS tissue is selected from the group consisting of hippocampus: (dentate gyrus, CA1 and CA3); cerebellum, hypothalamus, cortex: (occipital, temporal and forebrain); substantia nigra, thalamus, and any combination thereof.

6. The recombinant AAV VP capsid polypeptide of embodiment 3, comprising a polypeptide sequence represented by the formula: (A)-(X)-(B) wherein:

- (A) is the polypeptide sequence of SEQ ID NO: 47438 (residues 561-580 of SEQ ID NO: 2: (VAYNVGGQMATNNQSSTTAP));
- (X) is a polypeptide sequence selected from any of the polypeptides of SEQ ID NO:115-1114 or 1118-47437 that confer corresponding tissue tropism on a recombinant AAV virion (rAAV); and
- (B) is the polypeptide sequence of SEQ ID NO: 47439 (residues 590-609 of SEQ ID NO: 2: (IVPGSVWMERDVYLQGPIWA)); and

wherein the capsid polypeptide is capable of assembling into the rAAV and, the capsid does not have the sequence of any of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4. SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, and SEQ ID NO:8. 7. The engineered AAV VP capsid polypeptide of any of the previous embodiments, wherein the AAV VP capsid polypeptide is an AAV5 VP1 capsid polypeptide. 8. The engineered AAV VP capsid polypeptide of any of the previous embodiments, wherein the capsid confers upon a recombinant AAV virion (rAAV), increased tissue targeting for any tissue selected from the following tissues:

- adipose, adrenal gland, aorta, brain (including hippocampus: dentate gyrus, CA1 and CA3; cerebellum, caudate, putamen, midbrain, pons, hypothalamus, cortex-including occipital, temporal and forebrain; substantia nigra, thalamus, and any combinations thereof), bone marrow, cecum, colon, dorsal root ganglion, duodenum, epididymis, esophagus, eye, gallbladder, heart, ileum, jejunum, kidney, lung, lymph nodes, mammary gland, ovary, pancreas, parathyroid gland, peripheral nerve, pituitary, prostate, salivary gland, seminal vesicle, skeletal muscle, skin, spinal cord, spleen, stomach, testis, thymus, thyroid, trachea, urinary bladder, uterus, and vagina.
  
  compared to a control virion comprising a capsid with any of the following sequences: SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, and SEQ ID NO:8. 9. The engineered AAV VP capsid polypeptide of embodiment 2, wherein the rAAV exhibits from about a 1.0005-fold to about a 1000-fold increased accumulation in the first tissue as compared to the second tissue. 10. The engineered AAV VP capsid polypeptide of embodiment 9, wherein the rAAV exhibits at least about a 1.0005-fold, at least about a two-fold, at least about a three-fold, at least about a four-fold, at least about a five-fold, at least about a ten-fold, at least about a twenty-fold, at least about a 50-fold, at least about a 75-fold, at least about a 100-fold, or at least about a 1000-fold increased accumulation in the first tissue as compared to the second tissue. 11. The engineered AAV VP capsid polypeptide of any of embodiments 1 or 3-8, wherein the rAAV exhibits from about a 1.0005-fold to about a 1000-fold increased accumulation in the tissue. 12. The engineered AAV VP capsid polypeptide of embodiment 11, wherein the rAAV exhibits at least about a 1.0005-fold, at least about a two-fold, at least about a three-fold, at least about a four-fold, at least about a five-fold, at least about a ten-fold, at least about a twenty-fold, at least about a 50-fold, at least about a 75-fold, at least about a 100-fold, or at least about a 1000-fold increased accumulation in the tissue. 13. The engineered AAV VP capsid polypeptide of any of embodiments 1-3 or 6-12, wherein at least one mutant residue is:

Xaa1 and Xaa1 is selected from A, G, K, M, N, Q, R, S, or T;

Xaa2 and Xaa2 is selected from A, C, H, I, K, S, T, or V;

Xaa3 and Xaa3 is selected from A, G, H, K, M, N, Q, R, S, T, or V;

Xaa4 and Xaa4 is selected from L, M, P, Q, R, T, or W;

Xaa5 and Xaa5 is selected from F, H, I, K, M, T, or Y;

Xaa6 and Xaa6 is selected from E, G, H, L, M, N, Q, T, or W;

Xaa7 and Xaa7 is selected from A, C, G, H, L, M, R or S;

Xaa8 and Xaa8 is selected from A, C, D, F, G, H, M, Q, S, V, W, or Y;

Xaa9 and Xaa9 is selected from A, C, E, G, H, M, N, P, Q, S, V, or W;

or any combination thereof,

wherein the capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV) and wherein the rAAV is capable of exhibiting tissue tropism for liver tissue.

14. The engineered AAV VP capsid polypeptide of any of embodiments 1-12, wherein:

Xaa1 excludes A, G, K, M, N, Q, R, S, or T;

Xaa2 excludes A, C, H, I, K, S, T, or V;

Xaa3 excludes A, G, H, K, M, N, Q, R, S, T, or V:

Xaa4 excludes L, M, P, Q, R, T, or W;

Xaa5 excludes F, H, I, K, M, T, or Y;

Xaa6 excludes E, G, H, L, M, N, Q, T, or W;

Xaa7 excludes A, C, G, H, L, M, R or S.

Xaa8 excludes A, C, D, F, G, H, M, Q, S, V, W, or Y;

Xaa9 excludes A, C, E, G, H, M, N, P, Q, S, V, or W.

or any combination thereof,

wherein the capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV) and exhibits less targeting or tropism to liver tissue compared to a rAAV having a VP1 capsid polypeptide having the sequence of SEQ ID NO:1. 15. The engineered AAV VP capsid polypeptide of any of the previous embodiments, having at least one residue corresponding to residue 581 to residue 589 in SEQ ID NO: 2, wherein:

Xaa1 is A, D, E, G, L, M, N, Q, S, T, or V;

Xaa2 is A, C, D, E, G, H, I, N, P, Q, S, T, or V;

Xaa3 is A, D, E, G, H, M, N, Q, S, T, or V;

Xaa4 is A, D, E, G, H, N, P, Q, S, or T;

Xaa5 is A, C, D, E, G, H, N, Q, S, T, or Y;

Xaa6 is A, D, E, G, H, N, P, Q, S, or T;

Xaa7 is s A, C, D, E, G, H, N, Q, S, or T;

Xaa8 is A, C, D, E, G, H, N, Q, S, or T;

Xaa9 is A, D, E, G, H, N, P, Q, S, or T;

or any combination thereof,

wherein the capsid polypeptide is capable of assembling into a recombinant AAV virion (rAAV).

16. The recombinant capsid polypeptide of any preceding embodiment, further comprising one or more mutations or substitutions at an amino acid residue outside of the 581-589 region, wherein the one or more mutations or substitutions at an amino acid residue outside of the 581-589 region confers improved manufacturability, improved viral assembly, improved tissue targeting/tropism, or any combination thereof. 17. A polynucleotide encoding any of the recombinant adeno-associated virus (AAV) VP capsid polypeptides of embodiments 1-16. 18. A recombinant AAV virion (rAAV), the virion comprising an AAV VP capsid polypeptide of any of embodiments 1-15. 19. The rAAV virion of embodiment 18, wherein the rAAV has reduced tropism for human liver as compared to a rAAV having a VP1 capsid polypeptide having the sequence of SEQ ID NO:1. 20. The rAAV virion of embodiment 18 or embodiment 19, wherein the rAAV has increased ability to cross the blood-brain barrier following intravenous administration as compared to a rAAV having a VP1 capsid polypeptide having the sequence of SEQ ID NO:1. 21. The rAAV virion of any one of embodiments 18-20 wherein the rAAV has increased ability to infect one or more brain regions selected from hippocampus, dentate gyrus, cerebral cortex, temporal cortex, occipital cortex, thalamus, forebrain, substantia nigra, hypothalamus, and cerebellum, following intravenous, intrathecal, intracerebral ventricular, or intracisternal magna administration as compared to a rAAV having a VP1 capsid polypeptide having the sequence of SEQ ID NO:1.

22. The rAAV virion of any one of embodiments 18-21, wherein the rAAV has increased ability to infect human retinal cells following intravitreal injection as compared to a rAAV having a VP1 capsid polypeptide having the sequence of SEQ ID NO:1.

23. The rAAV virion of any one of embodiments 18, wherein the rAAV has increased ability to infect human skeletal muscle following intravenous administration as compared to a rAAV having a VP1 capsid polypeptide having the sequence of SEQ ID NO:1.

24. The rAAV virion of any one of embodiments 18, wherein the rAAV has increased tropism for human liver as compared to a rAAV having a VP1 capsid poly peptide having the sequence of SEQ ID NO: 1. 25. The rAAV virion of any one of embodiments 18-24, wherein the virion further comprises a vector genome, the vector genome comprising a therapeutic polynucleotide encoding any of the following: a therapeutic RNA selected from a guide RNA or a tRNA, or transgene encoding a protein under control of regulatory sequences that direct transgene expression in infected human cells. 26. The rAAV virion of embodiment 25, wherein the transgene encodes a protein selected from the transgene products of Table 1. 27. A composition comprising an AAV virion comprising the engineered capsid polypeptide of any one of embodiments 1-16, within which is packaged the therapeutic polynucleotide of embodiment 25. 28. A composition comprising an AAV virion comprising the engineered capsid polypeptide of any one of embodiments 1-16, within which is packaged a therapeutic polynucleotide encoding any of the following: a therapeutic RNA selected from a guide RNA or a tRNA, or transgene encoding a protein under control of regulatory sequences that direct transgene expression in infected human cells. 29. The rAAV virion of any one of embodiments 18-26, wherein the virion further comprises a vector genome, the vector genome comprising a therapeutic polynucleotide encoding any of the following: a therapeutic RNA selected from a guide RNA or a tRNA, or transgene encoding a protein under control of regulatory sequences that direct transgene expression in infected human cells. 30. The rAAV virion of embodiment 29, wherein the transgene encodes a protein selected from the transgene products of Table 1.

31. A pharmaceutical composition comprising the rAAV of embodiment 29 or embodiment 30 and a pharmaceutically acceptable carrier. 32. A method of treatment, comprising:

administering an effective amount of the pharmaceutical composition of embodiment 31 to a patient in need thereof. 33. The method of embodiment 32, wherein the effective amount of the rAAV is less than the effective amount of a wild type rAAV. 34. The method of embodiment 32, wherein the effective amount of the rAAV is less than the effective amount of an otherwise comparable rAAV lacking one or more than one mutation or substitution at a position corresponding to residue 581 to residue 589 of SEQ ID NO: 1.

35. The method of embodiment 34, wherein the effective amount of the rAAV results in lower toxicity in the patient as compared to the effective amount of the wild type rAAV, the otherwise comparable rAAV, or both.

36. The method of embodiment 32, wherein the effective amount is at least from 1×10⁵viral genomes/kg patient weight to 5×10¹⁴viral genomes/kg.

37. The method of any one of embodiments 32-36, wherein the rAAV is administered intravenously.

38. The method of any one of embodiments 32-36, wherein the rAAV is administered intrathecally.

39. The method of any one of embodiments 32-36, wherein the rAAV is administered by intracisternal magna administration.

40. The method of any one of embodiments 32-36, wherein the rAAV is administered by intravitreal injection.

41. The rAAV virion according to any one of embodiments 18-26 or 29-30, for use in treating a disease in a patient.

42. The rAAV virion for use according to embodiment 41, for use in treating any of the diseases in Table 1.

43. The rAAV virion for use according to embodiment 41, for use in treating a CNS disease.

44. The rAAV virion for use according to embodiment 43, wherein the CNS disease is selected from the conditions listed in Table 1.

45. The rAAV virion for use according to embodiment 43 or 44, wherein the engineered AAV VP capsid polypeptide comprises any one of the mutations recited in Table 8 (SEQ ID NO:115-1114). Table 10 (SEQ ID NO: 7118-8117), Table 39 (SEQ ID NO: 8118-9117) or Table 74 (SEQ ID NO: 9118-10117).

46. The rAAV virion for use according to embodiment 45, wherein the engineered VP1 capsid polypeptide does not have the sequence of any of SEQ ID NO:1, SEQ ID NO:3, SEQ ID NO:4, SEQ ID NO:5, SEQ ID NO:6, SEQ ID NO:7, and SEQ ID NO:8.

47. The rAAV virion for use according to embodiment 42, wherein the virion comprises a therapeutic polynucleotide encoding a target from any of the targets listed in Table 1.

Series T Embodiments—AAV5

- 1. The engineered AAV VP capsid polypeptide of any of the embodiments described herein, wherein the engineered AAV VP capsid polypeptide is an engineered AAV5 VP capsid polypeptide.
- 2. The engineered AAV VP capsid polypeptide of embodiment 1, wherein the mutation is a substitution.

Series U Embodiments—Assembly

- 1. The engineered AAV VP capsid polypeptide of any of the embodiments described herein, wherein:
  - Xaa1 is selected from the group consisting of A, D, E, G, L, M, N, Q, S, T, or V; or
    - Xaa2 is selected from the group consisting of A, C, D, E, G, H, I, N, P, Q, S, T, or V; or
    - Xaa3 is selected from the group consisting of A, D, E, G, H, M, N, Q, S, T, or V; or
    - Xaa4 is selected from the group consisting of A, D, E, G, H, N, P, Q, S, or T; or
    - Xaa5 is selected from the group consisting of A, C, D, E, G, H, N, Q, S, T, or Y; or
    - Xaa6 is selected from the group consisting of A, D, E, G, H, N, P, Q, S, or T; or
    - Xaa7 is selected from the group consisting of A, C, D, E, G, H, N, Q, S, or T; or
    - Xaa8 is selected from the group consisting of A, C, D, E, G, H, N, Q, S, or T; or
    - Xaa9 is selected from the group consisting of A, D, E, G, H, N, P, Q, S, or T; or any combination thereof.
- 2. The engineered AAV VP capsid polypeptide of embodiment 1, wherein Xaa1 is A, D, E, M, or T.
- 3. The engineered AAV VP capsid polypeptide of embodiment 1, wherein Xaa1 is E.
- 4. The engineered AAV VP capsid polypeptide of embodiment 1, wherein Xaa2 is A, S, T, or V.
- 5. The engineered AAV VP capsid polypeptide of embodiment 1, wherein Xaa2 is A.
- 6. The engineered AAV VP capsid polypeptide of embodiment 1, wherein Xaa3 is D, E, N, Q or T.
- 7. The engineered AAV VP capsid polypeptide of embodiment 1, wherein Xaa3 is D or T.
- 8. The engineered AAV VP capsid polypeptide of embodiment 1, wherein Xaa4 is D, E. P, or Q.
- 9. The engineered AAV VP capsid polypeptide of embodiment 1, wherein Xaa4 is E.
- 10. The engineered AAV VP capsid polypeptide of any one of claims X-Y, wherein Xaa5 is D, E, N, Q or T.
- 11. The engineered AAV VP capsid polypeptide of embodiment 1, wherein Xaa5 is N.
- 12. The engineered AAV VP capsid polypeptide of embodiment 1, wherein Xaa6 is D, N, or Q.
- 13. The engineered AAV VP capsid polypeptide of embodiment 1, wherein Xaa6 is D.
- 14. The engineered AAV VP capsid polypeptide of any one of claims X-Y, wherein Xaa7 is A, D, E or G.
- 15. The engineered AAV VP capsid polypeptide of embodiment 1, wherein Xaa7 is A.
- 16. The engineered AAV VP capsid polypeptide of any one of claims X-Y, wherein Xaa8 is A, D, G, or S.
- 17. The engineered AAV VP capsid polypeptide of embodiment 1, wherein Xaa8 is G.
- 18. The engineered AAV VP capsid polypeptide of embodiment 1, wherein Xaa9 is A, D, G, or P.
- 19. The engineered AAV VP capsid polypeptide of embodiment 1, wherein Xaa9 is G.

Series V Embodiments-Recombinant AAV Virions

- 1. A recombinant AAV virion (rAAV), comprising:
  - the engineered AAV VP capsid polypeptide of embodiments described herein assembled into a capsid; and
  - a payload,
  - wherein the engineered AAV VP capsid polypeptide encapsidates the payload.
- 2. The recombinant rAAV of embodiment 1, wherein the payload comprises a therapeutic polynucleotide, optionally wherein the therapeutic polynucleotide encodes a transgene or a genome modifying entity.
- 3. The rAAV of embodiment 2, wherein the therapeutic polynucleotide encodes a guide RNA, a tRNA, a suppressor tRNA, a siRNA, a miRNA, an mRNA, a shRNA, a circular RNA, or an antisense oligonucleotide (ASO), a ribozyme, a DNAzyme, an aptamer, or any combination thereof.
- 4. The rAAV of embodiment 3, wherein the therapeutic polynucleotide encodes a linear therapeutic polynucleotide or a circular therapeutic polynucleotide.
- 5. The rAAV of embodiment 2, wherein the transgene is selected from TABLE 1.
- 6. The rAAV of embodiment 2, wherein the genome modifying entity comprises a CRISPR/Cas system, an adenosine deaminase acting on RNA (ADAR) enzyme, a transcriptional activator, or a transcriptional repressor.
- 7. The rAAV of embodiment 6, wherein the CRISPR/Cas system comprises a Cas3. Cas8, Cas10, Cas9, Cas4, Cas12, or Cas13.
- 8. The rAAV of embodiment 6, wherein the ADAR is human ADAR1 or human ADAR2.
- 9. The rAAV of embodiment 6, wherein the transcriptional activator is VP64.
- 10. The rAAV of embodiment 6, wherein the transcriptional repressor is KRAB.
- 11. A pharmaceutical composition comprising the rAAV virion of any one of embodiments 1-10, and a pharmaceutically acceptable carrier.
- 12. A method of treatment, comprising administering a therapeutically effective amount of the pharmaceutical composition of embodiment 11, to a subject in need thereof.
- 13. The pharmaceutical composition of embodiment 12, for use in treating a subject in need thereof, wherein the use comprises administering a therapeutically effective amount of the pharmaceutical composition of embodiment 11, to the subject.
- 14. The rAAV virion of any one of the embodiments 1-10, for use in the manufacture of a medicament for treating a subject in need thereof by administration of a therapeutically effective amount thereof
- 15. The method or use of any one of the embodiments 12-14, wherein the therapeutically effective amount of the rAAV virion is less than the therapeutically effective amount of a rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1 encapsidating the same payload, wherein the rAAV virion having the wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1 is administered by the same route of administration.
- 16. The method or use of any one of the embodiments 12-15, wherein the therapeutically effective amount of the rAAV virion results in lower toxicity in the subject as compared to a therapeutically effective amount of a rAAV virion having a wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1 encapsidating the same payload, wherein the rAAV virion having the wildtype AAV5 VP capsid polypeptide of SEQ ID NO: 1 is administered by the same route of administration.
- 17. The method or use of any one of the embodiments 12-16, wherein the therapeutically effective amount is at least from 1×10⁵viral genomes/kg subject weight to 5×10¹⁴viral genomes/kg subject weight.
- 18. The method or use of any one of the embodiments 12-17, wherein the rAAV virion is administered via systemic administration.
- 19. The method or use of embodiment 18, wherein the systemic administration is intravenous administration, intramuscular administration, intraperitoneal administration, or oral administration.
- 20. The method or use of any one of the embodiments 12-19, wherein the subject is a human or a non-human animal.
- 21. The method or use of any one of the embodiments 12-20, wherein the subject has a disease.
- 22. The method or use of embodiment 21, wherein the disease is a neurological condition.
- 23. The method of embodiment 22, wherein the neurological condition is AADC deficiency, Alzheimer's Disease, tauopathies, synucleinopathies, Batten Disease. MPS-IIIB, frontotemporal dementia with GBA1 mutations (PD-GBA), neuronpathic Gaucher's Disease, Gaucher Disease Type 2, Canavan Disease, Parkinson's Disease, Tay-Sachs Disease, Huntington's Disease, Protocki-Lupski Syndrome, amyotrophic lateral sclerosis, down syndrome, Sanfilippo Disease Type A. Sanfilippo Disease Type B, or Rett syndrome.
- 24. The method of embodiment 23, wherein the dementia is frontotemporal dementia (FTD).

Series W Embodiments

- 1. The method or use of any embodiment disclosed herein, wherein the disease is selected from Table 1.
- 2. The method or use of any embodiment disclosed herein, wherein the rAAV VP1 capsid at position 587 (Xaa7) is not A, C, D, E, F, G, H, I, K, M, P, Q, R, V, W, or Y.
- 3. The method or use of any embodiment disclosed herein, wherein the rAAV VP1 capsid at position 587 can be N, S, or T.
- 4. The method or use of any embodiment disclosed herein, wherein the rAAV VP1 capsid at position 582 (Xaa2) is not G, V, L, or I.
- 5. The method of use of any embodiment disclosed herein, wherein the rAAV VP1 capsid comprises one or more mutations outside of the 581-589 (Xaa1-Xaa9) region wherein the rAAV VP1 capsid is capable of assembling and exhibiting a desired tissue tropism.

1.2. Examples

Below are examples of specific embodiments for carrying out the present invention. The examples are offered for illustrative purposes only and are not intended to limit the scope of the present invention in any way. Efforts have been made to ensure accuracy with respect to numbers used (e.g., amounts, temperatures, etc.), but some experimental error and deviation should, of course, be allowed for.

The practice of the present invention will employ, unless otherwise indicated, conventional methods of protein chemistry, biochemistry, recombinant DNA techniques and pharmacology, within the skill of the art. Such techniques are explained fully in the literature.

Example 1
High Throughput Capsid Engineering Systems

This example describes the high throughput capsid engineering systems and methods disclosed herein to discover engineered tissue tropic AAV variants. The high throughput capsid engineering system is schematized in FIG. 1. As shown in FIG. 1, in the pre-assembly phase, processes described herein allow for generation of plasmid libraries containing variants of the AAV capsid gene with massive diversity at specific locations in the capsid gene encoding sequence. This library of variants is assembled into virus using the 2-plasmid system described herein, then assembled viruses are directly injected into non-human primates (NHP) for in vivo selection. Tissues are harvested and tissue-transducing capsid genes are labeled with unique molecular identifiers and barcodes. These variant sequences/UMIs/barcodes are parameterized and machine learning algorithms are used to identify deterministic features of specific tissue targeting/de-targeting capsids. “De-targeting” may be referred to herein as “sparing” with respect to a particular tissue. For example, liver detargeting or liver sparing may both be used herein to refer to variants or properties of variants that preferentially exhibit reduced homing to liver tissue.

Processes described herein allow for cloning of synthetic oligonucleotide variants of the AAV capsid gene with massive diversity. This library of variants is assembled into virus using the 2-plasmid system described herein, then assembled viruses are directly injected into non-human primates for n vivo selection. The process was carried out as follows, but it is understood that reasonable and scientifically standard modifications to these methods are encompassed herein. The library template was primary PCR amplified (8-10 cycles) and DNA is purified using a column (pool library). Optionally, a second 8 cycle amplification is performed from the library pool. DNA is purified using a suitable column. Next, a digestion was performed with NEB BsaI HF-V2 (library—3 hours; backbone—2 hours+1 hour with calf intestinal alkaline phosphatase) and DNA is purified using a column. The resulting digest was combined at a 3:1 ratio and ligated with NEB T4 ligase ON a 16° C. and more ATP (1 μM) and T4 ligase was added the following morning. A “ligase cycle” was carried out 20 times. Ligase cycling refers to 10 seconds at 16 degrees C., followed by 30 seconds at 25 degrees C. for each “cycle”. The ligase was heat inactivated and more ATP (1 μM) was added. PS-DNAse was added for 2 hours at 37° C. to digest linear (un-ligated) species, enriching for relaxed form (rf) circular DNA (FIG. 3). DNA was purified using a column and DNA was eluted in water. Electrocompetent recombinase-defective bacteria was transformed at massive scale by performing many (>100) electroporation reactions to incorporate library plasmids. Transformed bacteria was selected using antibiotic selection in liquid shaking culture overnight at 37° C. Transformation efficiency was confirmed and library coverage was measured by plating limiting dilutions of transformed bacteria on antibiotic selective agar plates. Library plasmid DNA was prepared for sequencing and transfection for viral production using a Megaprep kit. Virions were produced via “double transfection”: viral producer eukaryotic cell lines (e.g., HEK293) were co-transfected with a mixture of Adenovirus-derived “helper” gene plasmid and library plasmids. Virus from cells/supernatant was purified the following 3-7 days using density centrifugation or FPLC methods. DNA from virions was amplified for NGS sequencing analysis of assembly. Optionally, cloning may be repeated with assembly-competent variants. Purified library virus was intravenously administered to NHPs at a dose range between 1-5×10¹³viral genomes/kg. Tissues were harvested after 4-6 weeks and library variants were sequenced.

Tissues were harvested and transducing capsid genes are labeled with unique molecular identifiers and barcodes. These variant sequences/UMIs/barcodes were parameterized, and machine learning algorithms were used to identify deterministic features of specific tissue targeting/de-targeting capsids.

As noted above, Plasmid-Safe DNAse (PS-DNAse) was used to remove linear DNA after ligation. Use of this linear-selective DNAse enzyme to degrade un-ligated DNA maximizes transformation efficiency to maximize library diversity. FIG. 3 is a photograph of a gel showing effective removal of the linear insert and linear plasmid backbone (“BB”) following digestion with PS-DNAse. Total efficiency of plasmid library generation was 4.23×10⁹colony-forming units per μg plasmid DNA in one example of this method.

FIG. 4 is a schematic of the high throughput AAV capsid engineering system described in EXAMPLE 1. The histogram shows the size of the AAV capsid library at various stages of preparation, tracking the size of the library from theoretical diversity, synthesized capsid genes, total cloned variants, to sequenced assembled viruses (error bars denote minimum/maximum predicted diversity from NGS sequencing analysis). The pooled viral library was injected into non-human primates and following a period of time sufficient to ensure stable transduction, the animals were euthanized, and tissues were harvested. DNA was purified from the tissues (1), and (2) a unique molecular identifier (UMI) was appended. Exonuclease I was added to digest excess UMI-containing primers (3). Subsequent PCR amplification added sequencing indexes/tissue barcoding and adapters for sequencing. The addition of UMIs allows for high resolution frequency analysis of capsid variants in the tissues.

FIG. 5A illustrates the packaging approach used to maximize rAAV production from the library, with FIG. 5A comparing standard triple transfection (top panel) to the two-plasmid cis packaging approach used in EXAMPLE 1 (bottom panel), and FIG. 5B showing relative production of wild type AAV5 using the two transfection approaches. FIG. 5C is an example of UMI distribution in a liver specimen and shows a majority of capsid variants are found a single time (single UMI). However, a subset of capsid variants is enriched in a given tissue and may have increased numbers of UMIs, corresponding to their increased frequency in the target tissue.

Example 2
Identification and Analysis of AAV5 Variants in NHPs

This example describes identification and analysis of tissue targeting AAV5 capsid variants discovered using the general methods described in EXAMPLE 1. To identify variants with preferential infection of specific tissues (tropism), the biodistribution of the AAV5-based variant capsid library was evaluated in non-human primates (NHPs). The biodistribution investigation was performed according to Primate Products, LLC Standard Operating Procedures and authorized veterinary standards. Prior to library dose injection, a pre-study schedule of activities was performed to benchmark NHP clinical pathology, general health, and AAV5 seronegativity. Study animals were selected based on seronegativity for antibodies against AAV5.

NHPs received an IV Bolus injection of the AAV5-based capsid library at a dose range between 1-5×10¹³viral genomes/kg. The final diluted AAV5-based capsid library sample was mixed and transferred to sterile syringes for intravenous administration with a dose volume of 5 mL/kg. Dose preparations were made using common pipetting and transferring techniques of the AAV5-based capsid library. From a common stock of diluted stock of AAV5-based capsid library, dose volumes were prepared separately for each animal based on their respective body weight in order to deliver equivalent amount of vector to each animal on a per kg basis.

Following Day 1 library injection, clinical observations were made twice a day (Days 1-29) and body weight was checked weekly. At Day 29, all study animals were euthanized and underwent tissue collection in accordance with Primate Products, LLC Standard euthanasia procedures.

A diverse array of tissues were collected, including: adrenal gland, aorta, bone with bone marrow, brain (cerebellum), brain (hippocampus, dentate gyrus), brain (hippocampus, CA1), brain (hippocampus, CA3), brain (hypothalamus), brain (cortex, temporal), brain (cortex, forebrain), brain (cortex, occipital), brain (substantia nigra), brain (thalamus), cecum, colon, duodenum, epididymis, esophagus, eye, gallbladder, heart, ileum, jejunum, kidney, liver, lung, lymph node(s), mammary gland, ovary, pancreas, parathyroid gland, peripheral nerve (sciatic), pituitary, prostate, salivary gland, seminal vesicle, skeletal muscle, skin, spinal cord, spleen, stomach, testis, thymus, thyroid gland, trachea, urinary bladder, uterus, and vagina. Tissues were collected as two 1 cm³blocks (or whole organ) and preserved in liquid nitrogen. Liver was specifically collected in five 1 cm³blocks. For Brain, two 1 cm³blocks were collected from each region as noted in the above list, with the exception of smaller regions, which were collected in their entirety.

FIG. 9 illustrates an example of the performance of the library as a whole infecting cortex at a higher relative level than liver after the intravenous administration to an NHP. DNA was isolated from the liver and cortex and either qPCR (at left) or droplet digital PCR (ddPCR) (at right) were used to quantify viral genomes recovered from the respective tissues. By these methods of quantification, the library as a whole has ˜2-fold increased CNS targeting over the liver.

FIGS. 11A-D show Venn diagrams and analytic tables depicting the number of unique sequence variants found in liver tissue only, liver and cortex tissues, and cortex tissue only for three different sequencing analysis filters. Next-generation sequencing was performed on viral genomic capsid variants recovered from liver or cortical tissue samples. Unique Molecular Identifiers (UMIs) were appended as part of the molecular recovery process as described herein. This allows for increased confidence that a given capsid variant is present in the tissue, and may be correlated to abundance. FIG. 11A shows a Venn diagram in which the sequencing analysis filter applied was 4 or greater distinct UMIs—also referred to herein as “count”—per each distinct capsid sequence variant. FIG. 11B shows a Venn diagram in which the sequencing analysis filter applied was 50 or greater UMIs. FIG. 11C shows a Venn diagram in which the sequencing analysis filter applied was 100 or greater distinct UMIs. FIG. 11D shows a histogram analysis of the distribution of UMIs in the population of capsid sequence variants found in cortex only in which the sequencing analysis filter applied was 50 or greater UMIs as shown in FIG. 11B.

Example 3
AAV5 Variants that Functionally Assemble in Virus

This example describes engineered AAV5 variants that assemble into virions discovered using the methods and systems described in EXAMPLE 1-EXAMPLE 2. Following production and purification of the library virus as described in EXAMPLE 1, assembly of the AAV5 library virus was assessed. The composition of capsid variants was measured by NGS of an amplicon spanning the variant region. The frequency of a given amino acid residue occurring at a specified position corresponding to position 581 to position 589 of SEQ ID NO: 1 (generalized in SEQ ID NO: 2) over the frequency of that given amino acid residue occurring at the specified position in the library plasmid was analyzed to identify sequence rules that are preferred for viral assembly. With reference to TABLE 3 below, and SEQ ID NO: 2, the following amino acids can be independently mutated, in any combination, at any one or more positions Xaa1-Xaa9, to provide an AAV VP1 capsid that is capable of assembling. Additionally, one or more mutations outside of the Xaa1-Xaa9 region can be allowed, as long as the capsid is still capable of assembling.

TABLE 3

Viral Assembly Options

Xaa1 is A, D, E, G, L, M, N, Q, S, T, or V

Xaa1 is A, D, E, M, or T

Xaa1 is E

Xaa2 is A, C, D, E, G, H, I, N, P, Q, S, T, or V

Xaa2 is A, S, T, or V

Xaa2 is A

Xaa3 is A, D, E, G, H, M, N, Q, S, T, or V

Xaa3 is D, E, N, Q or T

Xaa3 is D or T

Xaa4 is A, D, E, G, H, N, P, Q, S, or T

Xaa4 is D, E, P, or Q

Xaa4 is E

Xaa5 is A, C, D, E, G, H, N, Q, S, T, or Y

Xaa5 is D, E, N, Q or T

Xaa5 is N.

Xaa6 is A, D, E, G, H, N, P, Q, S, or T;

Xaa6 is D, N, or Q; or

Xaa6 is D.

Xaa7 is s A, C, D, E, G, H, N, Q, S, or T;

Xaa7 is A, D, E or G; or

Xaa7 is A.

Xaa8 is A, C, D, E, G, H, N, Q, S, or T;

Xaa8 is A, D, G, or S; or

Xaa8 is G.

Xaa9 is A, D, E, G, H, N, P, Q, S, or T;

Xaa9 is A, D, G, or P; or

Xaa9 is G.

Example 4
AAV5 Variants with Liver Tissue Targeting and AAV5 Variants with Liver-Detargeting Properties

This example describes engineered AAV5 variants with tissue tropism in liver and AAV5 variants that preferentially detarget liver tissue that were discovered using the methods and systems described in EXAMPLE 1-EXAMPLE 2.

FIGS. 6A-B are heat maps and a statistical analysis showing clear functional selection through viral assembly and tissue transduction in liver. FIG. 6A is a heat map prepared at various stages of the high throughput system described in EXAMPLES 1-2 from pre-assembly, assembled virus, to liver transduction. Thus, FIG. 6A, at right, shows a heatmap of favored positional residues for all liver transducing variants divided by all assembled virus. For the pre-assembly heat map, the library of capsid variants was cloned into plasmids and transformed into electrocompetent bacteria. The resultant library was sequenced and amino acid diversity was measured at each position, with the low levels of positional variation likely arising from synthesis error. Subsequent assembly into virus showed clear amino acid residue/positional biases that highlight selection for viral assembly. Liver transducing viruses showed even greater patterns of AA residue/positional selection, with distinctly favored/disfavored variants. Each of these heatmaps was normalized by their respective input frequencies. FIG. 6B is a table of statistical analysis (ANOVA) showing that in three liver samples, amino acid residue distribution and residue-position varied significantly, but inter-sample variation was not significant.

FIGS. 7A-B illustrate analysis of repeat observed (high UMI) capsid variants between multiple samples. FIG. 7A is a Venn diagram illustrating counts of overlapping variant identification in two liver samples: ˜38% of variants with >10 UMIs were observed in both samples. FIG. 7B is a heat map showing positional amino acid distribution illustrating the strongly selected residues/positions of repeat observed capsid variants.

FIG. 10 provides structure illustrations with the illustration on the left representing the crystal structure of wildtype AAV5 [SEQ NO:1] (PDB: 6JCT from Zhang et al. Nat Commun. 2019 Aug. 21; 10(1):3760), oriented to the 3-fold axis of symmetry. The middle and right illustrations represent variants recovered from NHP liver replacing the 581-589 residues with NPAMFNCDY (middle panel, amino acid SEQ ID NO:112), and KLTAHIYAL (right panel, amino acid SEQ ID NO:114). These illustrations were generated using PyMOL to replace the variant sequences and are colored by predicted electrostatic charge to highlight the differences in charge among liver variants recovered from the same tissue.

The frequency of a given amino acid residue occurring at a specified position corresponding to position 581 to position 589 of SEQ ID NO: 1 (generalized in SEQ ID NO: 2) in variants identified in liver over the frequency of that given amino acid residue occurring at the specified position in variants forming assembled virus was analyzed to identify a set of sequence rules for capsids that preferentially target liver tissue. With reference to TABLE 4 below, and SEQ ID NO: 2 (AAV5 VP1), the following amino acids can be independently mutated, in any combination, at any one or more positions Xaa1-Xaa9, to provide a VP1 capsid with enhanced liver tropism as compared wildtype AAV5 VP1 capsid (SEQ ID NO: 1), where liver tropism here refers to properties that are preferred for liver transduction over properties that are preferred for virion assembly. Additionally, one or more mutations outside of the Xaa1-Xaa9 region can be allowed, as long as the capsid is still capable of assembling and exhibiting the desired liver targeting property.

TABLE 4

Liver Tropism Options and Rules

Xaa1 is A, G, K, M, N, Q, R, S, or T;

Xaa1 is A, K, M, or T; or

Xaa1 is K.

Xaa2 is A, C, H, I, K, S, T, or V;

Xaa2 is A, S, T, or V; or

Xaa2 is T.

Xaa3 is A, G, H, K, M, N, Q, R, S, T, or V;

Xaa3 is A, M, or T; or

Xaa3 is s A or T.

Xaa4 is L, M, P, Q, R, T, or W;

Xaa4 is L, P, Q, or T; or

Xaa4 is P.

Xaa5 is F, H, I, K, M, T, or Y;

Xaa5 is H, I, or Y; or

Xaa5 is Y.

Xaa6 is E, G, H, L, M, N, Q, T, or W;

Xaa6 is N, or Q; or

Xaa6 is N,

Xaa7 is A, C, G, H, L, M, R or S;

Xaa7 is A, C, H or M; or

Xaa7 is A.

Xaa8 is A, C, D, F, G, H, M, Q, S, V, W, or Y;

Xaa8 is G, M, Q, or S; or

Xaa8 is G.

Xaa9 is A, C, E, G, H, M, N, P, Q, S, V, or W;

Xaa9 is E, G, or P; or

Xaa9 is G.

The frequency of a given amino acid residue occurring at a specified position corresponding to position 581 to position 589 of SEQ ID NO: 1 (generalized in SEQ ID NO: 2) in variants not identified in liver over the frequency of that given amino acid residue occurring at the specified position in variants forming assembled virus was analyzed to identify a set of sequence rules that are preferred for liver detargeting. With reference to Table 5 below and AAV5 VP1 (SEQ ID NO: 2), the following amino acids can be independently excluded, in any combination, to provide an AAV VP1 capsid that is capable of assembling and is less targeted to liver tissue (“liver-sparing”, or “liver-detargeted”) than the wildtype AAV5 VP1 capsid (SEQ ID NO: 1), where liver-detargeting here refers to properties that are preferred among viruses that have decreased liver transduction over properties that are preferred for virion assembly. Additionally, one or more mutations outside of the Xaa1-Xaa9 region can be allowed, as long as the capsid is still capable of assembling and exhibits the desired features.

TABLE 5

Liver Detargeting Rules

Xaa1 excludes K;

Xaa1 excludes A, K, M, or T; or

Xaa1 excludes A, G, K, M, N, Q, R, S, or T.

Xaa2 excludes T;

Xaa2 excludes A, S, T, or V; or

Xaa2 excludes A, C, H, I, K, S, T, or V.

Xaa3 excludes A or T;

Xaa3 excludes A, M, or T; or

Xaa3 excludes A, G, H, K, M, N, Q, R, S, T, or V.

Xaa4 excludes P;

Xaa4 excludes L, P, Q, or T; or

Xaa4 excludes L, M, P, Q, R, T, or W.

Xaa5 excludes Y;

Xaa5 excludes H, I, or Y; or

Xaa5 excludes F, H, I, K, M, T, or Y.

Xaa6 excludes N;

Xaa6 excludes N, or Q; or

Xaa6 excludes E, G, H, L, M, N, Q, T, or W.

Xaa7 excludes A;

Xaa7 excludes A, C, H or M; or

Xaa7 excludes A, C, G, H, L, M, R or S.

Xaa8 excludes G;

Xaa8 excludes G, M, Q, or S; or

Xaa8 excludes A, C, D, F, G, H, M, Q, S, V, W, or Y.

Xaa9 excludes G;

Xaa9 excludes E, G, or P; or

Xaa9 excludes A, C, E, G, H, M, N, P, Q, S, V, or W.

Liver, CNS, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord tissues were analyzed to identify, amino acid residues in the AAV5 VP1 581-589 region that drive liver tropism. The results are shown in FIG. 18B which shows the positional frequency of each amino acid in the 581 to 589 region of AAV5 VP1 in liver tissue as compared to all other analyzed tissues, wherein the capsid polypeptide has the sequence of SEQ ID NO: 2. The position (“Xaan”, n=1-9) is denoted on the x-axis and amino acid residues are denoted on the y-axis. The frequency of a particular amino acid residue in a particular position is indicated in the heatmap as intensity. Darker squares indicate a higher frequency of a particular amino acid in a particular position and lighter squares indicate a lower frequency of a particular amino acid in a particular position.

The frequency of a given amino acid residue occurring at a specified position corresponding to position 581 to position 589 of SEQ ID NO: 1 (generalized in SEQ ID NO: 2) in variants identified in liver over the frequency of that given amino acid residue occurring at the specified position in variants identified in all other analyzed tissues was compared to identify a set of sequence rules that are preferred for liver targeting. With reference to TABLE 6A below, and SEQ ID NO: 2 (AAV5 VP1), the following amino acids can, thus, be independently mutated, in any combination, at any one or more positions Xaa1-Xaa9, to provide a VP1 capsid with enhanced liver tropism as compared wildtype AAV5 VP1 capsid (SEQ ID NO: 1), where liver tropism here refers to properties that are preferred for liver transduction over properties that are preferred for transduction of all other harvested tissues.

TABLE 6A

Liver Tropism Rules

Xaa1 is selected from A, G, K, M, Q, R, S, or T

Xaa1 is selected from A, K, Q, or R

Xaa1 is K

Xaa2 is selected from A, C, I, K, S, T, or V

Xaa2 is selected from A, K, S, or T

Xaa2 is A

Xaa3 is selected from A, G, I, K, M, Q, R, S, T, or V

Xaa3 is selected from A, K, Q, S, or T

Xaa3 is selected from K, Q, or T

Xaa3 is K

Xaa4 is selected from A, I, K, L, P, Q, R, S, T, or V

Xaa4 is selected from K, I, S, or V

Xaa4 is K

Xaa5 is selected from F, I, L, M, T, V, or V.

Xaa5 is selected from F, L, or Y

Xaa5 is F

Xaa6 is selected from F, H, M, N, Q, S, or Y

Xaa6 is selected from M or N

Xaa6 is N

Xaa7 is selected from A, C, K, M, Q or S

Xaa7 is selected from A, C, or S

Xaa7 is S

Xaa8 is selected from A, C, F, G, M, Q, or S

Xaa8 is selected from A, C, M, or S

Xaa8 is C

Xaa9 is selected from E, F, L, Q, R, or Y

Xaa9 is selected from L, Q, or R

Xaa9 is R

With reference to TABLE 6B below, and SEQ ID NO: 2 (AAV5 VP1), the following amino acids can, thus, be independently mutated, in any combination, at any one or more positions Xaa1-Xaa9, to provide a VP1 capsid with reduced liver tropism as compared wildtype AAV5 VP1 capsid (SEQ ID NO: 1), where liver tropism here refers to properties that are deterministic for liver transduction over properties that are deterministic for transduction of all other harvested tissues.

TABLE 6B

Liver Detargeting Rules

Xaa1 is not selected from A, G, K, M, N, Q, R, S, or T

Xaa1 is not selected from A, K, Q, or R

Xaa1 is not K

Xaa2 is not selected from A, C, I, K, S, T, or V

Xaa2 is not selected from A, K, S, or T

Xaa2 is not A

Xaa3 is not selected from A, G, I, K, M, Q, R, S, T, or V

Xaa3 is not selected from A, K, Q, S, or T

Xaa3 is not selected from K, Q, or T

Xaa3 is not K

Xaa4 is not selected from A, I, K, L, P, Q, R, S, T, or V

Xaa4 is not selected from K, I, S, or V

Xaa4 is not K

Xaa5 is not selected from F, I, L, M, T, V, or Y.

Xaa5 is not selected from F, L, or Y

Xaa5 is not F

Xaa6 is not selected from F, H, M, N, Q, S, or Y

Xaa6 is not selected from M or N

Xaa6 is not N

Xaa7 is not selected from A, C, K, M, Q or S

Xaa7 is not selected from A, C, or S

Xaa7 is not S

Xaa8 is not selected from A, C, F, G, M, Q, or S

Xaa8 is not selected from A, C, M, or S

Xaa8 is not C

Xaa9 is not selected from E, F, L, Q, R, or Y

Xaa9 is not selected from L, Q, or R

Xaa9 is not R

TABLE 7 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that were found in liver tissue and comport to one or more of the rules provided in TABLE 6. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 43438-SEQ ID NO: 44437, as disclosed in TABLE 7. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amino acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 7

Sequences of the 581 to 589 Region in AAV5 VP1

Capsid Polypeptide that Drive Liver Tissue Tropism

SEQ

ID
581-589

NO
Sequence

43438
KADPATSIG

43439
KAEAMRNDR

43440
KAECYYGYE

43441
KAEGENVAK

43442
KAFPAPQGP

43443
KAGCMFWCE

43444
KAQHDTAVG

43445
KARVNCHGS

43446
KASDLNTQP

43447
KCCEYCDIA

43448
KCCTCVSFP

43449
KCDFWMGWM

43450
KCICSDDLK

43451
KCLSESFWG

43452
KCLWPFDWG

43453
KCLWPFNWG

43454
KCMDAKEHA

43455
KCQPCIVKC

43456
KCQSKDTHF

43457
KCSTDKAVE

43458
KCVVLENTQ

43459
KDHWECAGY

43460
KDNCVMWHW

43461
KDPKEQFFQ

43462
KDQQDQCND

43463
KEDERKSGT

43464
KETCTNMDL

43465
KFDLCQDSM

43466
KFWMPEMEF

43467
KGDLLDCLE

43468
KGEITPSGD

43469
KGGDGNYPF

43470
KGLDVHALN

43471
KGWKPDMTN

43472
KHAAYNACE

43473
KHQDIMHRG

43474
KIAQVDTWV

43475
KIDSQQSNY

43476
KIDSYGTSM

43477
KIEMQYRQS

43478
KIGYWWPQA

43479
KITPPLGNY

43480
KIYPECHYS

43481
KLEPNMADR

43482
KLLIHGPYE

43483
KMCGNCEAG

43484
KMEHMQHQS

43485
KMESRQLYN

43486
KMQHYDAEA

43487
KNDEYRMQN

43488
KNMLGDWPN

43489
KNMYYMYEA

43490
KPATQLDCA

43491
KPCKHMGGR

43492
KPDHMRWAL

43493
KPDQGGICK

43494
KPEGCLNSR

43495
KPFQALMMS

43496
KPGVHSHHG

43497
KPIAETVNW

43498
KPMGCHASS

43499
KPTPHAYDA

43500
KPWIQIASH

43501
KPWYERYQT

43502
KQATYPHAW

43503
KQDTKNLDE

43504
KQEMFGRFL

43505
KOAPFVSGD

43506
KQQMDHCAM

43507
KQTPYEMFH

43508
KRWVAEWIV

43509
KSDCQFRDH

43510
KSDRWAWGC

43511
KSELSTWGI

43512
KSEPPQVVT

43513
KSGYIDLYH

43514
KSIFLDSWE

43515
KSILKEDDG

43516
KSNGDMSGL

43517
KSNRDMNFA

43518
KSQMEMCIG

43519
KSRDNDGVL

43520
KSTTDSGCQ

43521
KSVQVPDAM

43522
KSWINEWLS

43523
KTDGPGTSW

43524
KTKPLDTVD

43525
KTLSDLAVQ

43526
KTLWALSCN

43527
KTNAEENFI

43528
KTNAWDTPC

43529
KTNPVKYVK

43530
KTQCTAQQN

43531
KTONEKVWL

43532
KTQTPCFQH

43533
KTSWMMEAV

43534
KTTDGKMID

43535
KITQWHEWA

43536
KVAQGLGPE

43537
KVCEIGTQG

43538
KVDAFEGRM

43539
KVECAQFDV

43540
KVEPSMLLD

43541
KVGINELFT

43542
KVHLSLWPQ

43543
KVIMYQHNG

43544
KVMWAGSKR

43545
KVNADGSRC

43546
KVPFACCSS

43547
KVRNKDPTM

43548
KVSVOGQVF

43549
KVTHCEAMN

43550
KWELYNNDP

43551
KWHTEDRNC

43552
KYNYKTYKT

43553
KACSAMDFC

43554
KALHQINED

43555
KAPMGWMNE

43556
KAQCALDTM

43557
KCENKDVGY

43558
KCNMNQEAE

43559
KCPQLNDNQ

43560
KCQQEQQYN

43561
KEGVENTGW

43562
KEMWNCDPA

43563
KETPFLMQS

43564
KFHVKPVDM

43565
KHMQSDHFT

43566
KHVYSEMGL

43567
KIHKNEDLF

43568
KKLDGHDKY

43569
KKVMGMGIY

43570
KLVQRCEVD

43571
KMLDWKNCS

43572
KNGVDDAHN

43573
KNVVHEKQA

43574
KQALMENFF

43575
KQTTAEIGY

43576
KRDDCHNDN

43577
KRTVIHEMM

43578
KSHYGAGVT

43579
KSIDNYLCE

43580
KSKRWNWDV

43581
KSVCCQINN

43582
KTFCTQYMD

43583
KTFLVKTFI

43584
KTRNCPKEA

43585
KTSLVVRRG

43586
KTTLCVQWG

43587
KTVQQVQMG

43588
KTWWTEQSA

43589
KVAADLGHL

43590
KVEHYYDIG

43591
KVISHCDHY

43592
KWRVSVYVM

43593
KYEEFECMY

43594
KYICTRWHC

43595
KYSPTPFMY

43596
KYTPWDATF

43597
IACRCNCFD

43598
IADSKRLQK

43599
IADVQRCHY

43600
IAGHYDSWK

43601
IAGMKVITP

43602
IAHSVFTDH

43603
IAQRTRQQQ

43604
IAQTTLLPG

43605
IATIKVHAS

43606
LAAPKTMCH

43607
LACFSYTNV

43608
LAEPKKWHT

43609
LAFQYCYPP

43610
LAGLKHAYK

43611
LAHWQGLTY

43612
LALCIRHSW

43613
LAPPHKLDL

43614
LASFTSPDP

43615
LASTYWYSQ

43616
LATLNAFQN

43617
LAVVRATLD

43618
MAASDWHVM

43619
MAAWLEWGL

43620
MAEIRTRHV

43621
MAHENTKAG

43622
MAMTLEISD

43623
MASTYIERE

43624
MAVPMKMHC

43625
MAYLVVRQH

43626
NAARMEQAT

43627
NACTTKVGS

43628
NADYCSFPI

43629
NAENHTKTC

43630
NAFSMDIHV

43631
NAQSDPHQG

43632
NASKYGDFL

43633
NASSTVINT

43634
NATANYCLG

43635
NATTHSHTS

43636
NAVDRGIAT

43637
NAVPIKYPY

43638
NAWSSNEEA

43639
PAQRVIANL

43640
QAALMTEQF

43641
QADRMLKTG

43642
QADRWTGII

43643
QAEWPGTMT

43644
QAGLGSDAP

43645
QAHCVNLSS

43646
QAMNSARTH

43647
QAMTMLMQH

43648
QANNYNRMD

43649
QANTGRVYS

43650
QAQCAMGAR

43651
QAQWMGCKW

43652
QARIENNQF

43653
QATLQPVCI

43654
QATQTYDWH

43655
QATYP1TSE

43656
RADLWHIQN

43657
RAEDKWRSC

43658
RAERNSAEQ

43659
RAMAMDSSH

43660
RAMDDQHYG

43661
RAMLVEQWG

43662
RAQLFHQKT

43663
RAQPTSCCG

43664
RASVMPEAV

43665
SAAFQARLG

43666
SAAFSAFNV

43667
SAAWQLQMT

43668
SACQKEWCP

43669
SAEHKTHTM

43670
SAGRRNIGM

43671
SAHAGMHSN

43672
SAHKEDISG

43673
SAKSHLSNV

43674
SAKYFSFGV

43675
SAWAFVFGK

43676
SANYYMTNM

43677
SARVCDCPW

43678
SASDPKNCY

43679
SASKFGWFS

43680
SASWLDDQS

43681
SATGMRDYA

43682
SAWFTIVSG

43683
TAAGYHFCK

43684
TAAPWRHQS

43685
TAAQFMDEL

43686
TACLCLERP

43687
TADPLCRGA

43688
TAGILDHYL

43689
TAHCVMNKQ

43690
TAHGIYTQF

43691
TAHPGWEGY

43692
TAKFFADWP

43693
TALIISQNL

43694
TALLLDANP

43695
TAMPITAIP

43696
TAMQSGKVS

43697
TANPEALVD

43698
TANYGAAFA

43699
TAQKSFMNC

43700
TATQCKAKE

43701
TAVTTWNMP

43702
VADSWAYNP

43703
VAEWIMEKC

43704
VAKQSADPI

43705
VALNVAQKG

43706
VAMKKMTDD

43707
VAMPTMKYK

43708
VAQLALDEK

43709
VARFTELCP

43710
VARTFGEEK

43711
VASKFAGSV

43712
VATAQWLSC

43713
VAVQFREWL

43714
WAEHRDMRN

43715
WALTGYNIT

43716
WAQVCHCGE

43717
YAAGAMEAS

43718
YADMPSERG

43719
YAELLQKMF

43720
YAGCMLAIQ

43721
YAGSHEENC

43722
YAHAIMGCQ

43723
YAHENQQTS

43724
YANMKMICE

43725
YAQTYYNKD

43726
YARRFWTCV

43727
YARTTLANK

43728
YASILTNQS

43729
YASQSWQQE

43730
YATELGEYG

43731
YAVFGHQSR

43732
YAVLQNHYG

43733
AALIGLIGD

43734
AANPTQWMT

43735
AATMQTVPT

43736
AAYMAKLRA

43737
CAEQIMRNQ

43738
CAQRSMRSM

43739
CAQSFTVKG

43740
CASHRKGAV

43741
DAENFVFKG

43742
DANMTRTRP

43743
DANVIAHNP

43744
EASFVVEPV

43745
FAIWEAHGR

43746
FASHNYAPV

43747
FASRVCVDG

43748
GAEQNNECT

43749
GAFTVEFSP

43750
HAMQEELPP

43751
HAQFRGSAT

43752
IACTOQISV

43753
IADWFDYEC

43754
IAMQYPEQY

43755
IAPSVNQAT

43756
IAPWWSTTT

43757
LAEAGQNQV

43758
LAEGLKRTG

43759
LAGKSGQQV

43760
LAIEMVDRS

43761
LAISFKGKW

43762
LARGKDVAE

43763
MAKIGYYMI

43764
MAKPNPPLG

43765
NAAKYRNKQ

43766
NADLQIFFR

43767
NALPESMTC

43768
NAYPFVDHR

43769
PAAMDYLMD

43770
PACQETGMF

43771
PAEAEKIGR

43772
QAEFRCGNH

43773
QAFCEPNTP

43774
QAGKWAVCE

43775
QALWYEDMT

43776
QASCTHVWD

43777
RAEWKQWSC

43778
SAEWMQSWH

43779
SAEWTHNNQ

43780
SAGMYARVA

43781
SATMCDSQS

43782
SAYWFQMAG

43783
TAEHIKVDL

43784
TAISEIPAP

43785
TAKMVAAME

43786
TALMLNMAS

43787
TAVTTNKWA

43788
VAAKNYKER

43789
VAMMVAGFQ

43790
VAMTDAKMC

43791
VANVAIADM

43792
VASGSTEVC

43793
VATMIATQL

43794
WAHKDWSNW

43795
WAMEDGKSA

43796
YAALTYYYS

43797
ICKQNYSRT

43798
IFKEGIHYQ

43799
EPKNMGGCE

43800
IRKEVAVVP

43801
IRKNYRMKQ

43802
ITKNRDRWP

43803
IVKLVLPVG

43804
IVKQEHESY

43805
IYKPNTQLQ

43806
IYKTRHGWW

43807
IYKVLKPVH

43808
LCKQLGKQE

43809
LGKQEAVTN

43810
LGKWKIQGT

43811
LIKPLEISD

43812
LMKCLNWCC

43813
LMKCNEAFT

43814
LQKSWNMLQ

43815
LVKRKELET

43816
LVKWTNVLP

43817
LYKMMYAFR

43818
MCKLKLEIP

43819
MCKNQNKPA

43820
MHKPEGMNG

43821
MIKPSVEGM

43822
MLKCTHDLP

43823
MNKTMINYA

43824
MQKLMALQA

43825
MQKTIQQWM

43826
MWKVYEKSE

43827
NHKWICLTR

43828
NIKSRMIEF

43829
NMKSQWQPS

43830
NNKFWDYDP

43831
NQKATTASY

43832
NQKWSTWNN

43833
NVKCMQHMV

43834
PRKNQVDLG

43835
QCKQPLVIN

43836
QDKTKSCMY

43837
QGKYSANWY

43838
QKKFIQCYG

43839
QLKAPPAAH

43840
QMKMLWFNV

43841
QNKPASMHP

43842
QQKPQECYS

43843
QVKDVVGNV

43844
RGKTYEHGN

43845
RPKTFAALR

43846
RSKQARISH

43847
SCKLNLHEG

43848
SFKFYAEEA

43849
SGKQGNWRL

43850
SIKELVGCG

43851
SIKMIDCFG

43852
SMKWFITDD

43853
SVKYCQSKD

43854
TFKSYEMET

43855
TFKYHYNAC

43856
THKWGWWIA

43857
TIKPKNEWC

43858
TIKPYYPYQ

43859
TKKKRNGHL

43860
TRKDTLVAS

43861
TSKTCFAEK

43862
TTKEDEVCS

43863
TTKYTQSIF

43864
TVKPAVPGE

43865
VEKSSIMNR

43866
VFKLPEPVV

43867
VSKCGVSHI

43868
VTKYKYEGM

43869
WKKMKYTWK

43870
WMKPEFHYD

43871
YSKYKRLWI

43872
AEKPWNAMN

43873
AIKFKYAPD

43874
ANKVGTAGN

43875
AQKEYSKYL

43876
CKKMGLHVM

43877
CSKFRECCG

43878
DFKTMNYMV

43879
DFKWIQDQG

43880
DHKILGAIP

43881
ELKVAISSG

43882
ENKTQFAYL

43883
FRKMGCGKT

43884
GLKLNLFQA

43885
GSKTCQQVM

43886
LNKGQPLEM

43887
LTKETGQVV

43888
MLKTARWRD

43889
MSKPRVCCA

43890
MVKTGVEEM

43891
MWKFAQLRL

43892
MYKPEQWCM

43893
QIKSCKNTD

43894
STKQHPHLN

43895
STKVPWWFP

43896
TMKFSHESA

43897
TYKYKESSE

43898
VQKSWEYDN

43899
WSKKDIWAD

43900
IIPKDNKWR

43901
IPAKFRSWW

43902
ISVKGLTTH

43903
IYCKIKEWG

43904
LDHKCAAGM

43905
LHLKLINHC

43906
LLEKKQASG

43907
LSQKPSNLI

43908
LYAKKMAGY

43909
LYAKKMVGY

43910
MGSKIHFMA

43911
MQGKNMAVT

43912
NCVKAMDWY

43913
NHEKTIASF

43914
NIDKRGPMM

43915
NMLKHMFGQ

43916
NMVKYIHMG

43917
NRAKEHWHQ

43918
NSYKLDWGW

43919
NTGKMEEKR

43920
PFEKWNSQS

43921
PFRKSAGYM

43922
QHTKPNCRD

43923
QLCKVGRDN

43924
QTEKLMRLD

43925
QTNKWDVVT

43926
QTWKLITHG

43927
QVHKGQKFS

43928
QVNKITWVN

43929
QVTKFVTEG

43930
QWCKLINMC

43931
QWVKYHDIW

43932
QYIKKYWNH

43933
RVHKGQKFS

43934
SCAKFDVVE

43935
SDSKLHQVA

43936
SGTKYCCWN

43937
SIGKWPAYE

43938
SMMKCPTQY

43939
SVWKCERIS

43940
TCTKAVDHC

43941
TYEKFNKHQ

43942
TYIKSYQMA

43943
VRDKGVQCA

43944
VSQKSPTAW

43945
WCTKCNYNQ

43946
WCVKRMENH

43947
WVEKHLMLG

43948
YSQKPEEAN

43949
CTYKTGSPA

43950
CVLKWATMW

43951
DQMKWSQMG

43952
DWVKATQMC

43953
ESDKEERMT

43954
EVVKMPNHR

43955
EYIKWDGTP

43956
FCQKIEYYH

43957
FGPKLTWHT

43958
GCGKCECWN

43959
GTFKNPVHQ

43960
GVTKLEVWP

43961
GWDKKRDFR

43962
IPIKLPPQG

43963
LEAKVAQYY

43964
MDCKYPRDQ

43965
NDGKIQFSG

43966
NLSKLAQMP

43967
NLSKLTQMP

43968
NYAKLFQDI

43969
QKRKLQVYS

43970
OPAKKFWDF

43971
QSSKWMWPL

43972
QWYKMGYNR

43973
RGPKDTNVK

43974
RSLKYEGQQ

43975
SPNKSPFPG

43976
SSIKPMLSP

43977
SYHKQERCQ

43978
TCVKAGLCL

43979
TGLKVIGQH

43980
TIMKPLCFA

43981
TNHKFRCDR

43982
TSSKCTMDA

43983
VHPKVQRVT

43984
YSDKMESSN

43985
IHCMFGAGA

43986
IQQYFNWCE

43987
IRDMFEVGR

43988
LEFEFYWDE

43989
LIHPFARGN

43990
LIQAFPRDT

43991
LPTPFMAHL

43992
LSRDFCNWF

43993
LVTSFASML

43994
MCDHFQMAY

43995
MPRCFDAAA

43996
MPVMFGLHC

43997
NMYPFMKYQ

43998
NWITFSGAV

43999
QCDLFSEGY

44000
QHSVFMQIP

44001
QNIRFTTDM

44002
QSNHFEQEK

44003
QYGSFNQNV

44004
REEIFSYMV

44005
RGIVFSQDN

44006
RKADFMHIT

44007
RPIWFIFVL

44008
RQQEFDRCR

44009
RTDHFGHWE

44010
RVEMFKWRT

44011
SCHGFQNWP

44012
SIGLFMKDS

44013
SKEWFSGTS

44014
SNAPFEQCN

41015
SNLSFGLTA

44016
SQGYFDANL

44017
STQMFANVQ

44018
TCMYFSCAL

44019
TEAWFQSAF

44020
TIRNFWLQN

44021
TNMLFLVWP

44022
TPCRFNSQR

44023
TQSCFHLEN

44024
TRQFFHHGD

44025
TVEQFTNSV

44026
TVRMFCRAI

44027
TWHSFPNQE

41028
TYISFQDEK

44029
VSQSFDHVN

44030
VWQPFSGDH

44031
YCCWFEHRL

44032
YGCHFQRQE

44033
YHMGFTGMG

44034
YMSSFTCHW

44035
YPMTFRAAC

44036
YYNNFICDK

44037
ACVAFSQQM

44038
AHEIFNLQV

44039
AHTQFLSAD

44040
AMQSFDMPG

44041
ARFLFYFDV

44042
CCVYFNRQM

44043
CNIDFTADY

44044
CSQIFWERG

44045
CTPQFNRGF

44046
DEIAFWDGE

44047
DGLMFHQQR

44048
DIDQFASVN

44049
DREAFDMAS

44050
DSQWFEHNA

44051
DTGPFMLFQ

44052
ETRPFEQVW

44053
FTNRFQLMA

44054
GCCHFCDHL

44055
HRMYFLRWP

44056
IICGFFAWS

44057
LGHQFNNWT

44058
MDGYFGFAV

44059
MGSNFVKQF

44060
NICGFFAWS

44061
QHCQFEQAI

44062
QKDIFPMQM

44063
QMELFCQQD

44064
QNTEFMDHY

44065
QTSDFAMSS

44066
QVMPFVIEG

44067
SKMNFGTAN

44068
SKTEFSLKM

44069
TGDLFCCHL

44070
TQGLFAQVG

44071
VCFHFRDAA

44072
VCMLFQHEA

44073
VHSRFGQYT

44074
WQPRFYHIW

44075
YEDPFRHYS

44076
ICHSGNRLL

44077
ICQGSNSMA

44078
IGTVRNWCW

44079
IICPTNEYM

44080
EKILLNEMT

44081
EQCFFINMEE

44082
EVMCHNVYP

44083
EVNLNNHHW

44084
LCAQHNEVC

44085
LETEGNPFY

44086
LGIGNNWEQ

44087
LITPPNWWH

44088
LLEEMNEQC

44089
LLFWYNHMS

44090
LMILRNTHQ

44091
LQMPYNWQT

44092
LRHICNCGQ

44093
LSLSKNCAG

44094
LTPLANQYC

44095
LTRFNNDLP

44096
LVPRENMLW

44097
LYELHNTWD

44098
MFEQDNAPT

44099
MLLYPNNTN

44100
MMCSNNSHR

44101
MQSPENTVY

44102
MTCLQNQSE

44103
MTIANNIRN

44104
MTYNYNSKD

44105
MVWICNAPR

44106
NCGYKNNKA

44107
NICFVNDAL

44108
NILSPNCYE

44109
NKVGMNYMQ

44110
NMCHHNWRD

44111
NPAYKNWGW

44112
NQIYTNPWG

44113
NSLESNAWT

44114
NTMTPNFRK

44115
NYVAKNYMV

44116
PKFIANRQE

44117
PNYWINYQN

44118
PRCYENWRY

44119
PWQFTNLEK

44120
QCCHPNGYQ

44121
QNCWYNFFR

44122
QQCNTNTDS

44123
QVVTVNASD

44124
REGIANDWV

44125
SDGTENRMN

44126
SHLQCNYMR

44127
SLTVWNQNG

44128
STMWRNRES

44129
STVYWNMET

44130
SYTSGNESW

44131
TGAPGNWFH

44132
TRNAYNCHQ

44133
TSSWVNWDP

44134
TTMPENIPL

44135
TTRFHNNVM

44136
TVWNTNTGW

44137
VCRNGNRQA

44138
VEFPENSHY

44139
VLTWDNPDE

44140
VTHATNVLE

44141
WNGFMNQGY

44142
WPVRLNEYK

44143
YEAVPNKMS

44144
YYECRNYSD

44145
AKVYSNLYG

44146
ATILVNRDG

44147
AWFTHNALP

44148
CIHHRNDKV

44149
CNYDTNLRP

44150
CTOSINGLI

44151
CTVLINCRD

44152
CVTWENVMT

44153
DLVHCNNHM

44154
DRGYYNSMM

44155
DTEHENEGV

44156
EPHIMNSHE

44157
EQRVYNHQY

44158
FTVTMNCTA

44159
GGALDNGQC

44160
GPDRYNEES

44161
GSLALNVRQ

44162
GTQRDNTGF

44163
HCICNNSGA

44164
HMTLGNLCP

44165
ICGWENCSE

44166
LCENVNPSE

44167
LFTEINGCY

44168
LMCQMNNRS

44169
MPGFENKMP

44170
MPRDENHIG

44171
NFDCSNDQT

41172
NLLVSNVIT

44173
PDGPMNLLG

44174
PMMMANRIA

44175
QHRQANGIH

44176
QMSYVNQYR

44177
QSVSNNRRA

44178
QTFMTNFDT

44179
QYSPANQWD

44180
RTFLTNPKD

44181
SMHDYNENV

44182
STELKNMER

41183
SVLYSNYFA

41184
TGAMTNYCY

44185
TMMQNNTEV

44186
TRDTHNSVD

44187
TRIHVNRYG

44188
TTWLDNEDA

44189
VERPENQEW

44190
WQAPHNOSY

44191
YVITANNHQ

44192
ICQHIGSWG

44193
IDHDYASDS

44194
ILIMILSHS

44195
EPRFVYSPV

44196
ITVADRSSH

44197
LEGATESLW

44198
LLPLHGSWH

44199
MCLAPTSMH

44200
MCSPEPSLQ

44201
MEHQQDSRF

44202
MGDDSMSEH

44203
MPRTCMSTN

44204
NCEQWKSMF

44205
NDQTSSSAW

44206
NIQFHTSGQ

44207
NKDVDRSHV

44208
NLVRWESGW

44209
NMSPWMSDR

44210
NPCIVISSV

44211
NSYHLESMD

44212
NVAMYASSE

44213
NVLCVKSVQ

44214
NVMECHSPA

44215
NYCTLMSAC

44216
PILETHSVE

44217
QIGWEASSL

44218
QIVELHSLG

44219
QIVQAMSSE

44220
QLADNHSRP

44221
QSLYDRSGT

44222
QSTCYFSNC

44223
QTREGVSYY

44224
QVDMARSSF

44225
QVMENSSWA

44226
RECVHLSCP

44227
REGLLWSNS

44228
RISQQWSRS

44229
RTIQKESHV

44230
RWCMKISAN

44231
SCTHVMSNA

44232
SHELQLSGL

44233
SHMNQASMP

44234
SITRWVSAE

44235
SLYMEMSWQ

44236
SWCSEHSPN

44237
SYIEWGSWM

44238
TCERKRSDK

44239
TCTYDKSQI

44240
THIQCRSMM

44241
THWWQPSSG

44242
TKHMIQSYG

44243
TSARCISAE

44244
TSTHVKSYQ

44245
TTHFAVSGI

44246
TTSHCVSPS

44247
TVHMCQSFV

44248
VGTAAHSQR

44249
VIAQELSAK

44250
VSDWGHSDS

44251
VWWREDSTN

44252
WLEPCVSSL

44253
WPETVGSSE

44254
WYAPAESSN

44255
YCCRPVSDF

44256
YHMDGDSRH

44257
ADETQYSCH

44258
AFNQHTSMD

44259
AISPWMSCA

44260
AMYNVSSAA

44261
AMYQNPSPC

44262
ARAEWKSKD

44263
CVCADTSVF

44264
DEEENTSEQ

44265
ELAAPHSQV

44266
ERATLKSMC

44267
ERMWKTSDN

44268
ESIRLRSSQ

44269
FNAVHMSLE

44270
GGRFNQSWA

44271
GHFMTLSKG

44272
GNERMFSSA

44273
HLTMWESFD

44274
LCTFNGSNG

44275
LKGTMMSPI

44276
LTYLKRSIG

44277
MLNLHGSDM

44278
MSIPASSEV

44279
NIGYRKSSG

44280
NRESDVSTG

44281
PCQWPVSRA

44282
PCVYKSLEI

44283
PTIWLHSYL

44284
PVCPWGSFD

44285
PWVAVESAE

44286
QQTRHRSCV

44287
RTEWHQSQA

44288
SMQPTGSGW

44289
STQWEQSMQ

44290
SWVSKESEE

44291
TNMPMPSYE

44292
TQGRMYSCW

44293
TWGFNRSYC

44294
VQHCSHSLD

44295
VTYEAKSQW

44296
WDSQPQSGV

44297
WELNNPSVM

44298
WLTWHFSSF

44299
IIATGWECD

44300
ISLPMLECI

44301
LKIYTKCCD

44302
LSSHSEECQ

44303
MCFDIATCA

44304
MEHFLQVCN

44305
MKWPAFACA

44306
MMNVHQNCE

44307
MPVRLTKCS

44308
MWVDCCECH

44309
NGGQKATCP

44310
NGWFVRICG

44311
NNASPSYCG

44312
NRPHQWLCC

44313
NSHACSECS

44314
PNNQCSYCC

44315
PRTEDLQCI

44316
QCMAHIQCD

44317
QEMDGWYCC

44318
QGAIYEVCL

44319
QEFLWFYCG

44320
QKQSTVICE

44321
QLNVTPICC

44322
QMAPYSICQ

44323
QNGNIEKCQ

44324
QQNPNGCCH

44325
QVSSHVECV

44326
RLSAGDACY

44327
RPVLECNCE

44328
RQYEQDVCD

44329
RREPTKCCW

44330
RVTEWFQCG

44331
RWCGKGYCV

44332
SFSYAECCA

44333
SHENQWACI

44334
SHMFQHTCG

44335
SNTLYDTCY

44336
SQFMDDDCA

44337
SRNAMQYCP

44338
SSHIDGICN

44339
TELVYMTCQ

44340
TGDSRMDCQ

44341
TKGFMSVCR

44342
TNYPNWICH

44343
TQEAISDCF

44344
TQQFCHDCQ

44345
TTGMLENCN

44346
TWTQMIDCY

44347
TYVRRWMCL

44348
VIHGSLQCQ

44349
VIMMDGYCT

44350
VIMTQMNCW

44351
VPNTKDACF

44352
VWCGNQWCN

44353
WFTNVLWCG

44354
YPIMERWCH

44355
YSEICDRCI

44356
AFAQLGQCH

44357
ALVLRCICL

44358
CHPDKMECP

44359
CVMVEWLCS

44360
DGSCKWNCH

44361
DSMRKYQCR

44362
DTVMQAACW

44363
GGPHREFCI

44364
HTIDKMNCP

44365
IPYQQDCCA

44366
LILHMLECQ

44367
MHTSMLICC

44368
MKAATMVCL

44369
MKFTLPQCN

44370
NCHRNDACD

44371
NENQMMHCA

44372
NTAQQLCCE

44373
NTTQRALCG

44374
PTRTRRTCR

44375
QINLDSQCM

44376
QSAIEEVCR

44377
QSHPRITCA

44378
QVQMGRDCG

44379
RCETPTGCE

44380
SCCYCPTCC

44381
SMCFKGTCE

44382
SRNANDGCF

44383
SVFMRDECC

44384
TCWQWLKCE

44385
TMDRQPMCI

44386
TSSTEQMCV

44387
IGRITMGDR

44388
IHTLKHDYR

44389
IMFMVAFDR

44390
LHVPPSWAR

44391
LMCTMLTDR

44392
LMNELMHLR

44393
MDFLSCGLR

44394
MELDVSNDR

44395
MFMMMHDIR

44396
MNHWNMHMR

44397
NDPHQHDAR

44398
NFLDCSERR

44399
NPNLIEKHR

44400
QCLLTLQNR

44401
QEHHVEGVR

44402
QITHLQVGR

44403
QSSFNPDMR

44404
QYEVKMWSR

44405
RNDDNLERR

44406
RTFYKQVAR

44407
SCSQIQEIR

44408
SEGCRMPPR

44409
SFAVMMEDR

44410
SPNLEEKHR

44411
SSSSRRDYR

44412
SVYLKLRIR

44413
SYLQWHVTR

44414
TGYFYMAHR

44415
TPAFTHWER

44416
TPMSTQNVR

44417
TTPQYMRER

44418
VNMPLYKFR

44419
WEHHCAEWR

44420
WLPFAMTNR

44421
YCYRAKEQR

44422
YIEPPALGR

44423
YINMNAQER

44424
YKLCRVKGR

44425
YMLVDMIFR

44426
DHAHYWYGR

44427
DSVSQQVER

44428
ECCNYVWER

44429
ESQYSQGTR

44430
FMSMQGDQR

44431
GKNCWPIKR

44432
HTGPMLGDR

44433
HWCTIGKLR

44434
MGADYGVWR

44435
NFNQCTIYR

44436
QKQGTICER

44437
QSAIEEVYR

Example 5
AAV5 Variants with Tissue Tropism in CNS

This example describes engineered AAV5 variants with tissue tropism in CNS that were discovered using the methods and systems described in EXAMPLE 1-EXAMPLE 2.

TABLE 8 is a list of 1000 variants (CNS variants) that had the highest normalized UMI counts across two cortical sequencing samples and absent in two liver sequencing samples. Minimal inclusion criteria for this list included presence in both quality filtered cortex NGS data sets; and exclusion criteria were detection of a given variant in either of two independent liver sequencing samples. The total number of variants that meet these criteria following analysis of the four data sets is 2108. UMI counts were then normalized independently for the two separate cortex NGS data sets, and the variant sequences were then rank ordered by the sum of the two independently normalized UMI counts (maximum possible=200%), and the top 1000 are listed here (SEQ ID NO: 115-SEQ ID NO: 1114). Variant residues are at positions 581-589.

TABLE 8

norm 2

samples

(max

possible

SEQ ID NO
var_aa
score 200%)

SEQ ID NO: 115
RVTIMFTGT
112.28%

SEQ ID NO: 116
YTGCIDGFL
100.01%

SEQ ID NO: 117
TNMSTQPCW
97.08%

SEQ ID NO: 118
VWTTVCNDY
96.78%

SEQ ID NO: 119
DYANILKQS
91.84%

SEQ ID NO: 120
EFVLGCQGI
76.88%

SEQ ID NO: 121
TIMWTMMRE
76.32%

SEQ ID NO: 122
SPMVVTHSD
76.07%

SEQ ID NO: 123
DWYDIWTLV
69.59%

SEQ ID NO: 124
REFAISGGN
69.44%

SEQ ID NO: 125
SITNCNHOR
69.01%

SEQ ID NO: 126
HIQEELEEQ
64.92%

SEQ ID NO: 127
DINGKKNIC
64.05%

SEQ ID NO: 128
FLAEHFTSH
63.45%

SEQ ID NO: 129
QCQSQEWFH
60.94%

SEQ ID NO: 130
ARSGINGHE
60.24%

SEQ ID NO: 131
CIRQYDAID
59.65%

SEQ ID NO: 132
ILIFESRIG
58.77%

SEQ ID NO: 133
HFADNQVVQ
56.73%

SEQ ID NO: 134
NESHVERHC
56.14%

SEQ ID NO: 135
TMVTSVTLP
55.86%

SEQ ID NO: 136
PTLSCPYHK
54.39%

SEQ ID NO: 137
TVAKVLSGH
53.82%

SEQ ID NO: 138
KIFITTDEG
52.65%

SEQ ID NO: 139
KSAQVYWHP
52.63%

SEQ ID NO: 140
CSALFAQHD
52.05%

SEQ ID NO: 141
AGSACMPAH
50.59%

SEQ ID NO: 142
IVIWQGEWI
50.29%

SEQ ID NO: 143
SYCEVEDAP
50.00%

SEQ ID NO: 144
DCSWWLQLG
49.42%

SEQ ID NO: 145
NFCKQGNIC
47.08%

SEQ ID NO: 146
HIHDTMSDY
46.81%

SEQ ID NO: 147
QYVHMLGEC
46.50%

SEQ ID NO: 148
RQHHCLPVN
46.22%

SEQ ID NO: 149
AHVDHQCSG
45.43%

SEQ ID NO: 150
SFSQTHLGW
45.40%

SEQ ID NO: 151
AASQMWQRG
45.04%

SEQ ID NO: 152
NDCTCRAIM
44.16%

SEQ ID NO: 153
NSHEYLDRH
43.86%

SEQ ID NO: 154
DWIWGTDMW
43.60%

SEQ ID NO: 155
MIFYRTYGF
42.40%

SEQ ID NO: 156
HPAPLNMPH
42.11%

SEQ ID NO: 157
SQQHCQHDQ
41.58%

SEQ ID NO: 158
HEVMMCPAP
41.01%

SEQ ID NO: 159
KRTDLHAAD
40.95%

SEQ ID NO: 160
EYFKPKSQD
40.94%

SEQ ID NO: 161
KLAHSWCIW
40.94%

SEQ ID NO: 162
NAMANFSVR
40.64%

SEQ ID NO: 163
EVTTPRPPT
39.50%

SEQ ID NO: 164
YTHDEQIDY
38.90%

SEQ ID NO: 165
LTHQGGRQR
38.31%

SEQ ID NO: 166
AQWYMWCWD
37.73%

SEQ ID NO: 167
RMMWQSHCD
37.44%

SEQ ID NO: 168
EIHKRIAMT
36.55%

SEQ ID NO: 169
CMSCCKWPH
36.42%

SEQ ID NO: 170
DFQRPTETQ
35.67%

SEQ ID NO: 171
SFMPCHCRG
35.67%

SEQ ID NO: 172
LTHNTPFPK
35.40%

SEQ ID NO: 173
HYVVRHEDP
35.14%

SEQ ID NO: 174
FQEAPPHVH
34.80%

SEQ ID N0: 175
YAELEWKVC
33.92%

SEQ ID NO: 176
RCVAEPNEP
33.37%

SEQ ID NO: 177
GLIFHQSLS
33.34%

SEQ ID NO: 178
ASHGMDEYN
33.33%

SEQ ID NO: 179
PIACRAINW
33.33%

SEQ ID NO: 180
NCKPDQIAM
32.69%

SEQ ID NO: 181
FRMWFTECI
31.87%

SEQ ID NO: 182
MGYAKHDSH
31.59%

SEQ ID NO: 183
DTSCLFLMQ
31.29%

SEQ ID NO: 184
KVAGGHETT
31.16%

SEQ ID NO: 185
YSRYHHSLL
30.99%

SEQ ID NO: 186
VADTWQVHC
30.43%

SEQ ID NO: 187
LWGYGAVNM
29.25%

SEQ ID NO: 188
FGRQAYVAW
29.24%

SEQ ID NO: 189
EEVQFTQQD
29.24%

SEQ ID NO: 190
GTGHIGNAT
29.24%

SEQ ID NO: 191
AYTYNSKLY
28.95%

SEQ ID NO: 192
FIFRKCDHW
28.37%

SEQ ID NO: 193
QCVDCPPYC
27.79%

SEQ ID NO: 194
ETPPLCHGP
27.78%

SEQ ID NO: 195
RMTPNNCQR
27.28%

SEQ ID NO: 196
NSTPMACME
26.91%

SEQ ID NO: 197
STHPLNLYH
26.90%

SEQ ID NO: 198
TTVPDPGPR
26.32%

SEQ ID NO: 199
LIWSVYQDE
26.09%

SEQ ID NO: 200
YDNHQKARN
26.02%

SEQ ID NO: 201
SIYQKWVTN
25.92%

SEQ ID NO: 202
MQSMQHWHP
25.74%

SEQ ID NO: 203
TLVSCDQGN
25.73%

SEQ ID NO: 204
WSITGQPWF
25.73%

SEQ ID NO: 205
YQKVEYWNY
25.73%

SEQ ID NO: 206
AHYQQSTQR
25.44%

SEQ ID NO: 207
QVRVITHKS
25.17%

SEQ ID NO: 208
WAAKPGDPK
25.15%

SEQ ID NO: 209
RSDNDWCFL
25.15%

SEQ ID NO: 210
ETHRGTVGL
24.89%

SEQ ID NO: 211
KCSVQEKKA
24.89%

SEQ ID NO: 212
WVEPEQCFG
24.56%

SEQ ID NO: 213
NHQQCPKCP
24.28%

SEQ ID NO: 214
AAHSAPFFD
24.27%

SEQ ID NO: 215
KGQRRDSVS
24.07%

SEQ ID NO: 216
DVMQNQCKS
23.98%

SEQ ID NO: 217
ALVYSGWIC
23.72%

SEQ ID NO: 218
FAKARHNWR
23.69%

SEQ ID NO: 219
TYIQYPTNA
23.69%

SEQ ID NO: 220
VSKFIAASW
23.69%

SEQ ID NO: 221
RQREGKHGN
23.68%

SEQ ID NO: 222
MAAHTKYRF
23.40%

SEQ ID NO: 223
FVGLMNQCG
23.39%

SEQ ID NO: 224
TWQSMVNSE
23.15%

SEQ ID NO: 225
TWQSMVNSE
23.10%

SEQ ID NO: 226
SRHYEQFEC
23.10%

SEQ ID NO: 227
VTAGQLGLN
23.10%

SEQ ID NO: 228
DSQYHVKCG
22.82%

SEQ ID NO: 229
LSECSDRAN
22.51%

SEQ ID NO: 230
YAYPTEGQI
22.22%

SEQ ID NO: 231
KRFVDMPEM
21.95%

SEQ ID NO: 232
VHKFQGNFG
21.94%

SEQ ID NO: 233
RGHEPPMWL
21.93%

SEQ ID NO: 234
QKCYCDEQC
21.64%

SEQ ID NO: 235
RHAPTDISP
21.60%

SEQ ID NO: 236
MTGIGKCWN
21.05%

SEQ ID NO: 237
QAYCCMHQY
20.48%

SEQ ID NO: 238
KHDDCCTGT
20.47%

SEQ ID NO: 239
KWYQFGMVA
20.42%

SEQ ID NO: 240
LTWECKFGG
20.18%

SEQ ID NO: 241
AIVQQHYYP
19.88%

SEQ ID NO: 242
EENVWMQYQ
19.88%

SEQ ID NO: 243
NSQPMDFLP
19.88%

SEQ ID NO: 244
FDTAFQGDI
19.59%

SEQ ID NO: 245
QMEARMDCE
19.49%

SEQ ID NO: 246
NWVPQIEHW
19.33%

SEQ ID NO: 247
VAILGLNPT
19.31%

SEQ ID NO: 248
ANVPMCSLQ
19.31%

SEQ ID NO: 249
LIPMDWIGM
19.30%

SEQ ID NO: 250
IYCGNSDMQ
18.73%

SEQ ID NO: 251
PAETLMDGF
18.71%

SEQ ID NO: 252
FSYQIQCQN
18.43%

SEQ ID NO: 253
TMRLFWDMG
18.42%

SEQ ID NO: 254
QRVPMQLVE
18.42%

SEQ ID NO: 255
AMTRQNGWM
18.16%

SEQ ID NO: 256
GMRTHSVMN
18.13%

SEQ ID NO: 257
LITVQHIRN
18.13%

SEQ ID NO: 258
TTYPHNTHG
17.88%

SEQ ID NO: 259
RIEMDEWEA
17.85%

SEQ ID NO: 260
DNTPEHCYM
17.57%

SEQ ID NO: 261
SSVVMAKQP
17.55%

SEQ ID NO: 262
CLVRIQKDH
17.54%

SEQ ID NO: 263
TLPKMHMWP
17.54%

SEQ ID NO: 264
MPHHSPDCW
17.32%

SEQ ID NO: 265
QELECHACS
17.31%

SEQ ID NO: 266
HMHNLPVKP
17.31%

SEQ ID NO: 267
GIAVWFDFQ
17.25%

SEQ ID NO: 268
ANRSRPTWQ
17.25%

SEQ ID NO: 269
KTFPILEAW
17.25%

SEQ ID NO: 270
ALQYCQGKD
16.97%

SEQ ID NO: 271
YHFEYMTSF
16.97%

SEQ ID NO: 272
VCGHRNDGN
16.96%

SEQ ID NO: 273
EICAAMDSF
16.68%

SEQ ID NO: 274
YWFVLNPCV
16.41%

SEQ ID NO: 275
QQNPCLGGM
16.32%

SEQ ID NO: 276
FNWFVRAYI
16.14%

SEQ ID NO: 277
DCDPYNMVD
16.12%

SEQ ID NO: 278
RTGDAGMSP
16.10%

SEQ ID NO: 279
LEFNGHLFN
16.08%

SEQ ID NO: 280
VYVAAGGGC
15.92%

SEQ ID NO: 281
MTTGLMAYE
15.87%

SEQ ID NO: 282
MAIDNLECK
15.83%

SEQ ID NO: 283
QKAIVAQND
15.81%

SEQ ID NO: 284
GGDGRVHNK
15.52%

SEQ ID NO: 285
THDAARKTA
15.50%

SEQ ID NO: 286
TWNEPHQNQ
15.20%

SEQ ID NO: 287
ANRADWQHL
14.96%

SEQ ID NO: 288
CTKAYRKPG
14.93%

SEQ ID NO: 289
RTAVIQCIE
14.92%

SEQ ID NO: 290
WKVSPIQGF
14.92%

SEQ ID NO: 291
MCGEYQPGS
14.91%

SEQ ID NO: 292
NHHPYTFHQ
14.91%

SEQ ID NO: 293
NPRYCLSLS
14.64%

SEQ ID NO: 294
SRASQQSMW
14.64%

SEQ ID NO: 295
VLKSCDRHG
14.64%

SEQ ID NO: 296
KEARSQTGD
14.63%

SEQ ID NO: 297
FANGCTGSI
14.63%

SEQ ID NO: 298
GAMAAPYHD
14.62%

SEQ ID NO: 299
YEHPIMWPV
14.62%

SEQ ID NO: 300
CYGWEGCKC
14.33%

SEQ ID NO: 301
AAAMRDKTD
14.33%

SEQ ID NO: 302
KLRIGVVCN
14.33%

SEQ ID NO: 303
TPHSIHTDK
14.33%

SEQ ID NO: 304
QTETWEFGA
14.26%

SEQ ID NO: 305
KAEMKMGCN
14.16%

SEQ ID NO: 306
VPICCNLPM
14.11%

SEQ ID NO: 307
LPSHCNLGS
14.05%

SEQ ID NO: 308
SMNRTHLER
14.04%

SEQ ID NO: 309
FGEFTMDEN
13.84%

SEQ ID NO: 310
YTAHNANVV
13.78%

SEQ ID NO: 311
GTMKHNGRA
13.76%

SEQ ID NO: 312
EHYCKDNVL
13.75%

SEQ ID NO: 313
TKRMMEDSG
13.75%

SEQ ID NO: 314
CERDCNCTA
13.74%

SEQ ID NO: 315
TGRVTADMA
13.74%

SEQ ID NO: 316
YNGDRLFCF
13.53%

SEQ ID NO: 317
NHEVVAFMQ
13.47%

SEQ ID NO: 318
QNEICVHHE
13.47%

SEQ ID NO: 319
LRSGMIVSG
13.46%

SEQ ID NO: 320
QPKMWMMAS
13.45%

SEQ ID NO: 321
DVRHSWSEK
13.45%

SEQ ID NO: 322
EILQDVYYL
13.45%

SEQ ID NO: 323
SVVNNLKAS
13.45%

SEQ ID NO: 324
RCTDCSYFY
13.29%

SEQ ID NO: 325
GSTSRNQRD
13.17%

SEQ ID NO: 326
LDCNAMPST
13.16%

SEQ ID NO: 327
QGRWACWYS
13.16%

SEQ ID NO: 328
TIEWYPDSQ
13.16%

SEQ ID NO: 329
KAVSYHHDG
13.16%

SEQ ID NO: 330
TMEHCQRPQ
13.16%

SEQ ID NO: 331
RMFWDTSDR
12.89%

SEQ ID NO: 332
MCNPKMMSS
12.88%

SEQ ID NO: 333
DSHDMINYK
12.87%

SEQ ID NO: 334
LTCYHNELS
12.87%

SEQ ID NO: 335
AVVEHAVNR
12.87%

SEQ ID NO: 336
CGQMVICSE
12.87%

SEQ ID NO: 337
NLRDAHWCM
12.87%

SEQ ID NO: 338
ESSQEIKTC
12.77%

SEQ ID NO: 339
AKMWQLENP
12.73%

SEQ ID NO: 340
GTIPKQHEF
12.59%

SEQ ID NO: 341
ARDCYWKAR
12.58%

SEQ ID NO: 342
AADREQGWW
12.57%

SEQ ID NO: 343
DSVQQRWYV
12.57%

SEQ ID NO: 344
KCGQDWIHG
12.33%

SEQ ID NO: 345
QVYAPEIEG
12.32%

SEQ ID NO: 346
CCCVPRSIY
12.29%

SEQ ID NO: 347
AAHGEMKTA
12.28%

SEQ ID NO: 348
QFWLMHEWP
12.28%

SEQ ID NO: 349
KFAMDWVSD
12.00%

SEQ ID NO: 350
TSDSVINWL
12.00%

SEQ ID NO: 351
QIIYRCRST
12.00%

SEQ ID NO: 352
GYDLRGADE
11.99%

SEQ ID NO: 353
CIRFMMREG
11.99%

SEQ ID NO: 354
HSNTHGYPL
11.73%

SEQ ID NO: 355
HGNMPHCYP
11.72%

SEQ ID NO: 356
VGLSIKMYG
11.71%

SEQ ID NO: 357
ESWRYKHQA
11.70%

SEQ ID NO: 358
PKRYQAWTW
11.70%

SEQ ID NO: 359
SCSQKLMMQ
11.70%

SEQ ID NO: 360
ALVTYRSMQ
11.70%

SEQ ID NO: 361
ALNSDMAAW
11.60%

SEQ ID NO: 362
NHSLCWDSK
11.60%

SEQ ID NO: 363
KKEDYAKFF
11.41%

SEQ ID NO: 364
RQYLYKRQQ
11.41%

SEQ ID NO: 365
TAMPEHRWD
11.41%

SEQ ID NO: 366
GFCTIPVAD
11.40%

SEQ ID NO: 367
IAVQKRLFC
11.40%

SEQ ID NO: 368
LAEIKDWVP
11.40%

SEQ ID NO: 369
MLPMFMGON
11.40%

SEQ ID NO: 370
NVRTYIDDS
11.40%

SEQ ID NO: 371
YFWISKALY
11.13%

SEQ ID NO: 372
ISYQPNPME
11.12%

SEQ ID NO: 373
ACDDWCWMC
11.11%

SEQ ID NO: 374
QTEMKKCAR
11.11%

SEQ ID NO: 375
ATTHKVKDN
11.11%

SEQ ID NO: 376
DIEKCMNVS
11.11%

SEQ ID NO: 377
QSFSYDAKE
11.11%

SEQ ID NO: 378
YGPEILKLT
11.11%

SEQ ID NO: 379
YGPEILKLT
11.11%

SEQ ID NO: 380
WAAWMYQQE
11.09%

SEQ ID NO: 381
DIACLDQWQ
10.93%

SEQ ID NO: 382
TSLVMPSLP
10.89%

SEQ ID NO: 383
MHSQQRAIK
10.84%

SEQ ID NO: 384
VRQSVPHWA
10.83%

SEQ ID NO: 385
DTRPSRDSS
10.82%

SEQ ID NO: 386
RSGAKCYCD
10.82%

SEQ ID NO: 387
ECKTHRDVY
10.63%

SEQ ID NO: 388
DELHVISIM
10.55%

SEQ ID NO: 389
QTKFMDDMH
10.54%

SEQ ID NO: 390
TSMTERRTV
10.54%

SEQ ID NO: 391
VQTWAIPCC
10.53%

SEQ ID NO: 392
LPTLRVADK
10.53%

SEQ ID NO: 393
KTGTTMEPM
10.52%

SEQ ID NO: 394
TKQFVHNED
10.50%

SEQ ID NO: 395
TCQEPHNIT
10.28%

SEQ ID NO: 396
QKVASMGCW
10.27%

SEQ ID NO: 397
CKGHDAGEY
10.27%

SEQ ID NO: 398
FPWSRPKCW
10.26%

SEQ ID NO: 399
HAMSLSTFQ
10.25%

SEQ ID NO: 400
MTCVWHSDF
10.25%

SEQ ID NO: 401
VKAWWHDHQ
10.24%

SEQ ID NO: 402
DVLKCEINE
10.24%

SEQ ID NO: 403
PIKGVAVQE
10.24%

SEQ ID NO: 404
LSRLYPNTY
10.23%

SEQ ID NO: 405
KQTVMTDTS
9.96%

SEQ ID NO: 406
SATPHWYCH
9.95%

SEQ ID NO: 407
KITQLFOYE
9.94%

SEQ ID NO: 408
VTWINLRSG
9.92%

SEQ ID NO: 409
IHOLVGAFW
9.66%

SEQ ID NO: 410
CVNLMAVNN
9.66%

SEQ ID NO: 411
EKQKLTVSS
9.66%

SEQ ID NO: 412
RLDHWATCD
9.65%

SEQ ID NO: 413
AVERNLERM
9.65%

SEQ ID NO: 414
KTNNTVCHS
9.65%

SEQ ID NO: 415
HHLRNVHHC
9.44%

SEQ ID NO: 416
ARGTCNYCY
9.37%

SEQ ID NO: 417
STEENEALY
9.37%

SEQ ID NO: 418
KKQQISIRL
9.36%

SEQ ID NO: 419
SAAEKAMDQ
9.36%

SEQ ID NO: 420
TAPGGSTPQ
9.36%

SEQ ID NO: 421
QTNQVVVVE
9.36%

SEQ ID NO: 422
DSTPNLFDP
9.08%

SEQ ID NO: 423
DVASYCQLQ
9.08%

SEQ ID NO: 424
TQCKQLTVM
9.08%

SEQ ID NO: 425
TMEHHECMW
9.07%

SEQ ID NO: 426
DVLVCEILW
9.07%

SEQ ID NO: 427
NIYSGINNP
9.07%

SEQ ID NO: 428
MFPKKMCLH
8.83%

SEQ ID NO: 429
THONTWLFY
8.83%

SEQ ID NO: 430
NHGYRNNSM
8.78%

SEQ ID NO: 431
QMHICMYEG
8.74%

SEQ ID NO: 432
KWEVMSAAY
8.73%

SEQ ID NO: 433
QRMIVDALQ
8.50%

SEQ ID NO: 434
IMMISSYSM
8.50%

SEQ ID NO: 435
TGAGIKEAH
8.49%

SEQ ID NO: 436
VPWVEFRLQ
8.49%

SEQ ID NO: 437
OPWPHLINK
8.48%

SEQ ID NO: 438
SSLCRAYDN
8.22%

SEQ ID NO: 439
CVKSRODSD
8.21%

SEQ ID NO: 440
NYILTQLGM
8.20%

SEQ ID NO: 441
FGGVEDSWM
8.19%

SEQ ID NO: 442
SVKDPMDWG
8.19%

SEQ ID NO: 443
YSDRFQFQF
8.19%

SEQ ID NO: 444
NCPMYDMIS
8.12%

SEQ ID NO: 445
IWGIYVHYF
8.08%

SEQ ID NO: 446
FTPELWENY
7.91%

SEQ ID NO: 447
AHSRMDWTP
7.91%

SEQ ID NO: 448
OPFORKVEE
7.91%

SEQ ID NO: 449
DYENCFESY
7.91%

SEQ ID NO: 450
IEVYHFTNS
7.89%

SEQ ID NO: 451
KALTHDARN
7.89%

SEQ ID NO: 452
KTQMKRLRM
7.70%

SEQ ID NO: 453
KAPWAPMDS
7.67%

SEQ ID NO: 454
ETVMMPMSI
7.64%

SEQ ID NO: 455
DYVDQCCWS
7.63%

SEQ ID NO: 456
RACVGSPLW
7.62%

SEQ ID NO: 457
KIMHSWYND
7.62%

SEQ ID NO: 458
RHEPTQSVS
7.62%

SEQ ID NO: 459
DPKGOSIQA
7.61%

SEQ ID NO: 460
RTTSRDVED
7.61%

SEQ ID NO: 461
QCMVMTEAD
7.61%

SEQ ID NO: 462
VQHNSESNI
7.61%

SEQ ID NO: 463
WHESTTIGS
7.61%

SEQ ID NO: 464
RHEQYVVAT
7.60%

SEQ ID NO: 465
VVCTDTRAL
7.49%

SEQ ID NO: 466
EHHAMMLHE
7.47%

SEQ ID NO: 467
KACMSHQGR
7.42%

SEQ ID NO: 468
DSPAYTTQY
7.35%

SEQ ID NO: 469
ICELTHCNY
7.33%

SEQ ID NO: 470
KATDLMVNG
7.32%

SEQ ID NO: 471
PQSHELQLL
7.32%

SEQ ID NO: 472
CHCHYNPFC
7.32%

SEQ ID NO: 473
FAFCVAGER
7.32%

SEQ ID NO: 474
EFWTTHSKG
7.31%

SEQ ID NO: 475
GHVLLGWYP
7.31%

SEQ ID NO: 476
YCQQLFLCD
7.05%

SEQ ID NO: 477
AKEPVRLYP
7.04%

SEQ ID NO: 478
EERLFTGCH
7.04%

SEQ ID NO: 479
QTSPKFCMC
7.04%

SEQ ID NO: 480
WMNQCGCKD
7.04%

SEQ ID NO: 481
NYSRGKENE
7.03%

SEQ ID NO: 482
DIQRWEHEG
7.02%

SEQ ID NO: 483
MMAMKQGFY
7.02%

SEQ ID NO: 484
KVNACGNLT
6.89%

SEQ ID NO: 485
ETTHRREWS
6.75%

SEQ ID NO: 486
PTVEEVVVQ
6.75%

SEQ ID NO: 487
SIEPRKTVG
6.75%

SEQ ID NO: 488
QTIMRHAQT
6.74%

SEQ ID NO: 489
RIETVSVPL
6.74%

SEQ ID NO: 490
SRTICVHGN
6.74%

SEQ ID NO: 491
GACKYSMCL
6.73%

SEQ ID NO: 492
GWDQNCKDS
6.73%

SEQ ID NO: 493
ASINCHLTE
6.44%

SEQ ID NO: 494
HYMELLCGC
6.44%

SEQ ID NO: 495
ITRTYQEMQ
6.44%

SEQ ID NO: 496
RANENHPFT
6.44%

SEQ ID NO: 497
LNKMQSYSA
6.42%

SEQ ID NO: 498
NGYLVHACQ
6.21%

SEQ ID NO: 499
LMAYCYANN
6.18%

SEQ ID NO: 500
KESKEHRWA
6.16%

SEQ ID NO: 501
FTTMCEHTA
6.15%

SEQ ID NO: 502
ATMWVQAYG
6.15%

SEQ ID NO: 503
RGQFHLIMD
6.15%

SEQ ID NO: 504
DLVDLRSWM
6.14%

SEQ ID NO: 505
LGAVLNNMV
6.14%

SEQ ID NO: 506
MYRQKEMQT
6.14%

SEQ ID NO: 507
RLEDTVGGS
6.14%

SEQ ID NO: 508
ACHYRLDNC
6.06%

SEQ ID NO: 509
LNNAGYRPC
6.02%

SEQ ID NO: 510
SFSHTDLEA
5.89%

SEQ ID NO: 511
ATEPLKFQY
5.87%

SEQ ID NO: 512
NQNPYDMMG
5.87%

SEQ ID NO: 513
FKIEWSQDI
5.86%

SEQ ID NO: 514
QIGHQYKWD
5.86%

SEQ ID NO: 515
SWSQLQHTD
5.86%

SEQ ID NO: 516
QMMDFQHPA
5.86%

SEQ ID NO: 517
LFTQRGWGN
5.85%

SEQ ID NO: 518
TATQREAFV
5.59%

SEQ ID NO: 519
KENPSEYEM
5.58%

SEQ ID NO: 520
KEFQLQMEP
5.57%

SEQ ID NO: 521
KSAGIRTLL
5.57%

SEQ ID NO: 522
MSEKGYAWV
5.57%

SEQ ID NO: 523
VYCGKDFQA
5.57%

SEQ ID NO: 524
CVKLVMANC
5.56%

SEQ ID NO: 525
ALRPYMDDR
5.56%

SEQ ID NO: 526
CSIQINNCR
5.56%

SEQ ID NO: 527
GTRINTCNG
5.56%

SEQ ID NO: 528
FGAVELDAP
5.56%

SEQ ID NO: 529
YCQRGHLSC
5.56%

SEQ ID NO: 530
HRMNNHAWF
5.54%

SEQ ID NO: 531
NADFKYNNT
5.52%

SEQ ID NO: 532
MGATKSDDF
5.32%

SEQ ID NO: 533
SMHDYNFNV
5.31%

SEQ ID NO: 534
GNECFHNVE
5.29%

SEQ ID NO: 535
GVWMPHGWR
5.29%

SEQ ID NO: 536
EIWPAAWMD
5.28%

SEQ ID NO: 537
APKWQIVDP
5.27%

SEQ ID NO: 538
HAMHWWSPM
5.27%

SEQ ID NO: 539
QCKYSERGT
5.21%

SEQ ID NO: 540
TCCMWTFIW
5.01%

SEQ ID NO: 541
ETGYRENPC
5.00%

SEQ ID NO: 542
MEMFTMKSC
4.99%

SEQ ID NO: 543
PQGFCKPQC
4.98%

SEQ ID NO: 544
KTTFDVACY
4.98%

SEQ ID NO: 545
WCRTPQREN
4.98%

SEQ ID NO: 546
GSQQMQASQ
4.97%

SEQ ID NO: 547
ITGSPQNGC
4.97%

SEQ ID NO: 548
GTVEAPQSF
4.93%

SEQ ID NO: 549
LNCPSKDET
4.85%

SEQ ID NO: 550
CSNTETHGV
4.84%

SEQ ID NO: 551
QMEQWHRGG
4.78%

SEQ ID NO: 552
IPFNSMIYC
4.72%

SEQ ID NO: 553
RVEQWRGQQ
4.70%

SEQ ID NO: 554
WNMPWGFYH
4.70%

SEQ ID NO: 555
MISCMDSWN
4.69%

SEQ ID NO: 556
TSRALDVAA
4.69%

SEQ ID NO: 557
FFRWRWGLG
4.69%

SEQ ID NO: 558
FPKCAIWHY
4.69%

SEQ ID NO: 559
NMIKRCSQG
4.69%

SEQ ID NO: 560
GTSVNILMM
4.68%

SEQ ID NO: 561
MRQRTYICQ
4.68%

SEQ ID NO: 562
RGCMAECMP
4.68%

SEQ ID NO: 563
MPMPVYQRD
4.59%

SEQ ID NO: 564
ALNKTLAES
4.55%

SEQ ID NO: 565
EAGMWPSKA
4.41%

SEQ ID NO: 566
AAVKEQQVM
4.40%

SEQ ID NO: 567
SAQYSGFDT
4.40%

SEQ ID NO: 568
LILDFNKID
4.40%

SEQ ID NO: 569
MSAVSLGRS
4.39%

SEQ ID NO: 570
RWGSPMAVP
4.39%

SEQ ID NO: 571
KTVTNGDAG
4.39%

SEQ ID NO: 572
YTMSMQMQR
4.39%

SEQ ID NO: 573
AHEHYRIFP
4.11%

SEQ ID NO: 574
GKFPMPTHS
4.11%

SEQ ID NO: 575
KIENRGQCR
4.10%

SEQ ID NO: 576
MWEMRRAMG
4.10%

SEQ ID NO: 577
HGSSYNIVQ
4.09%

SEQ ID NO: 578
VIKAFQHFN
4.09%

SEQ ID NO: 579
EVRCNKCRS
3.88%

SEQ ID NO: 580
LALQFQYNS
3.85%

SEQ ID NO: 581
GVITVDAKG
3.84%

SEQ ID NO: 582
CLHHHYPVT
3.83%

SEQ ID NO: 583
AECRGHYQA
3.82%

SEQ ID NO: 584
KTGWDSHWS
3.82%

SEQ ID NO: 585
RYEMMDTSD
3.82%

SEQ ID NO: 586
DIFWSSMDF
3.81%

SEQ ID NO: 587
MSTHVLKWV
3.81%

SEQ ID NO: 588
CDVWWFPNL
3.81%

SEQ ID NO: 589
IVFHNKFQH
3.81%

SEQ ID NO: 590
SHIQSASNL
3.81%

SEQ ID NO: 591
VKQLRQDGF
3.81%

SEQ ID NO: 592
DKCLNNKFY
3.80%

SEQ ID NO: 593
RNDWQQVFS
3.80%

SEQ ID NO: 594
QRREQCTST
3.77%

SEQ ID NO: 595
YHWSYEGAT
3.75%

SEQ ID NO: 596
TDFDKRQVP
3.67%

SEQ ID NO: 597
QISMKGEHG
3.55%

SEQ ID NO: 598
LVGERKNAY
3.53%

SEQ ID NO: 599
DPMKIFVSH
3.53%

SEQ ID NO: 600
KQTRTDYAY
3.52%

SEQ ID NO: 601
AVAMIRPNQ
3.51%

SEQ ID NO: 602
ESTHMLYEM
3.51%

SEQ ID NO: 603
VDADYCSCS
3.51%

SEQ ID NO: 604
EMVRMQRGT
3.51%

SEQ ID NO: 605
NCDHRFGTL
3.50%

SEQ ID NO: 606
SVVESAAEV
3.48%

SEQ ID NO: 607
NARPMCGQV
3.40%

SEQ ID NO: 608
TEATEMTYY
3.33%

SEQ ID NO: 609
AYGSFEKDW
3.24%

SEQ ID NO: 610
MNSFYRAEW
3.24%

SEQ ID NO: 611
MIDNCKICL
3.23%

SEQ ID NO: 612
VCRGSPLAE
3.23%

SEQ ID NO: 613
DTRMDQCCY
3.22%

SEQ ID NO: 614
LVDCSNTCQ
3.22%

SEQ ID NO: 615
INKNMDAAY
3.22%

SEQ ID NO: 616
TGMTFNSFR
3.22%

SEQ ID NO: 617
AWKIAHNSF
3.22%

SEQ ID NO: 618
NFMQMSIMG
3.22%

SEQ ID NO: 619
WIKHCWPPH
3.22%

SEQ ID NO: 620
NLWNIAMQQ
3.18%

SEQ ID NO: 621
EHMDRADDA
3.11%

SEQ ID NO: 622
AQDVYYPGS
3.08%

SEQ ID NO: 623
FPQTMHAQA
2.96%

SEQ ID NO: 624
TFGCIESMQ
2.95%

SEQ ID NO: 625
KFFYTELMH
2.94%

SEQ ID NO: 626
GRLGPGTWK
2.93%

SEQ ID NO: 627
ASTDGTWKD
2.92%

SEQ ID NO: 628
DARPFLAWM
2.92%

SEQ ID NO: 629
KFHSKERMN
2.91%

SEQ ID NO: 630
SLQHFPECA
2.87%

SEQ ID NO: 631
VCVSAHHHF
2.87%

SEQ ID NO: 632
AVIFELDAT
2.84%

SEQ ID NO: 633
EQADQMQVD
2.83%

SEQ ID NO: 634
MSQFKAYLF
2.82%

SEQ ID NO: 635
EMSIQSSLG
2.76%

SEQ ID NO: 636
GIGYRTFIG
2.67%

SEQ ID NO: 637
KASHTVTGT
2.65%

SEQ ID NO: 638
NSVALENRG
2.64%

SEQ ID NO: 639
NEGPLMCWF
2.64%

SEQ ID NO: 640
NNELQSRPK
2.61%

SEQ ID NO: 641
NGTQCLNMD
2.52%

SEQ ID NO: 642
SSTWQNVDK
2.51%

SEQ ID NO: 643
YMVPYSNYP
2.50%

SEQ ID NO: 644
WFDRPMCMF
2.48%

SEQ ID NO: 645
LECTALVAM
2.38%

SEQ ID NO: 646
QVQSGQFIF
2.37%

SEQ ID NO: 647
KMFTMICFD
2.37%

SEQ ID NO: 648
QSDAVHEPA
2.36%

SEQ ID NO: 649
GYMCEDAGI
2.35%

SEQ ID NO: 650
EIHPGDLND
2.35%

SEQ ID NO: 651
EFSKACTNE
2.35%

SEQ ID NO: 652
KSTYYNLKW
2.34%

SEQ ID NO: 653
TSSRTNPWG
2.34%

SEQ ID NO: 654
SFKGFHLQN
2.28%

SEQ ID NO: 655
CDEWDNLTH
2.20%

SEQ ID NO: 656
DAGIGMKFY
2.16%

SEQ ID NO: 657
LGDQAGYHV
2.14%

SEQ ID NO: 658
IIVWMMMYR
2.13%

SEQ ID NO: 659
GADEWLLTM
2.10%

SEQ ID NO: 660
QSYPTMYQC
2.09%

SEQ ID NO: 661
DRTWTMTQM
2.08%

SEQ ID NO: 662
TSWTTEHRM
2.06%

SEQ ID NO: 663
TLRICELHG
2.06%

SEQ ID NO: 664
CQASHEHAH
2.06%

SEQ ID NO: 665
ESAEPRMIP
2.06%

SEQ ID NO: 666
FWMQLWCLP
2.03%

SEQ ID NO: 667
QGYCRNCTG
2.02%

SEQ ID NO: 668
ESCLVRRGW
2.00%

SEQ ID NO: 669
QIRCTCGWP
1.98%

SEQ ID NO: 670
HAYQHDHSS
1.97%

SEQ ID NO: 671
DMAVSEAHC
1.97%

SEQ ID NO: 672
WAGMAYHAQ
1.94%

SEQ ID NO: 673
FQQDIQIGQ
1.92%

SEQ ID NO: 674
VVHKGRWKE
1.91%

SEQ ID NO: 675
MQLRQSDKW
1.89%

SEQ ID NO: 676
YVRQFPIFL
1.87%

SEQ ID NO: 677
IIHRTEGAA
1.87%

SEQ ID NO: 678
GQYWGQGWL
1.85%

SEQ ID NO: 679
GSTHMNQEG
1.85%

SEQ ID NO: 680
INAGDKPNT
1.83%

SEQ ID NO: 681
LGTAMREMQ
1.81%

SEQ ID NO: 682
PGGGDRPAC
1.81%

SEQ ID NO: 683
QDVQMWYQI
1.80%

SEQ ID NO: 684
QARAVMTAD
1.79%

SEQ ID NO: 685
GAHDPKVWY
1.78%

SEQ ID NO: 686
GTLGYHITG
1.77%

SEQ ID NO: 687
SMLMMITGM
1.77%

SEQ ID NO: 688
RDDYGIYDN
1.77%

SEQ ID NO: 689
HLVTTRHSH
1.76%

SEQ ID NO: 690
ITDLRCWLK
1.76%

SEQ ID NO: 691
IMVRELMSS
1.75%

SEQ ID NO: 692
NAEHCFMGQ
1.73%

SEQ ID NO: 693
NNLQWMYAK
1.73%

SEQ ID NO: 694
SPMPFLNVL
1.72%

SEQ ID NO: 695
CEQQLIAEG
1.72%

SEQ ID NO: 696
NDKYAFADR
1.71%

SEQ ID NO: 697
WIKRHIPSG
1.71%

SEQ ID NO: 698
REGAKHCVN
1.69%

SEQ ID NO: 699
CDDWTFERQ
1.69%

SEQ ID NO: 700
GVGPSHGER
1.69%

SEQ ID NO: 701
ESTEWQHTF
1.69%

SEQ ID NO: 702
MFVCCTIIV
1.67%

SEQ ID NO: 703
QKYEPRFFG
1.67%

SEQ ID NO: 704
AHVKIQLSW
1.66%

SEQ ID NO: 705
TESHMLSVE
1.66%

SEQ ID NO: 706
LRPLQHIQL
1.65%

SEQ ID NO: 707
MANGNWGVW
1.65%

SEQ ID NO: 708
VTDAAMFMG
1.64%

SEQ ID NO: 709
WHEEKPANS
1.62%

SEQ ID NO: 710
KCORMSVQQ
1.62%

SEQ ID NO: 711
DDRIGHCRN
1.62%

SEQ ID NO: 712
IAPEARCGT
1.62%

SEQ ID NO: 713
GCSSSVNMR
1.61%

SEQ ID NO: 714
RKRVRAYSE
1.61%

SEQ ID NO: 715
QEPKEDHIV
1.60%

SEQ ID NO: 716
SILPYPIDV
1.59%

SEQ ID NO: 717
ILDIINTET
1.59%

SEQ ID NO: 718
RFMLESWLH
1.59%

SEQ ID NO: 719
NSAHVHYTH
1.58%

SEQ ID NO: 720
LVKGHMTTS
1.58%

SEQ ID NO: 721
DKYANVHNE
1.57%

SEQ ID NO: 722
RYCTPDHEV
1.57%

SEQ ID NO: 723
RYHRRHDKP
1.57%

SEQ ID NO: 724
SMRMFEHSC
1.56%

SEQ ID NO: 725
SRCNKPTVR
1.56%

SEQ ID NO: 726
TFHQSDYWE
1.55%

SEQ ID NO: 727
VLQMPWLYT
1.55%

SEQ ID NO: 728
VCTPPLNMQ
1.55%

SEQ ID NO: 729
LNSDRLIEY
1.52%

SEQ ID NO: 730
CETYVPVIY
1.52%

SEQ ID NO: 731
WVPTFHARY
1.52%

SEQ ID NO: 732
CEIRTYEWS
1.52%

SEQ ID NO: 733
KHDGFSVVG
1.51%

SEQ ID NO: 734
AWYHTQDDM
1.50%

SEQ ID NO: 735
WMCEFSCIQ
1.50%

SEQ ID NO: 736
AMVMSKTES
1.49%

SEQ ID NO: 737
NFCCLECCI
1.49%

SEQ ID NO: 738
RTSPALLCA
1.49%

SEQ ID NO: 739
DITLWMATE
1.48%

SEQ ID NO: 740
AVAEIRPEP
1.47%

SEQ ID NO: 741
SARVTTMLY
1.47%

SEQ ID NO: 742
TLRAEMSFH
1.47%

SEQ ID NO: 743
ASKQNGINC
1.47%

SEQ ID NO: 744
RRCPALTIR
1.47%

SEQ ID NO: 745
ARPLVCQAL
1.46%

SEQ ID NO: 746
MMLGYMGQD
1.46%

SEQ ID NO: 747
QEHIYKHSN
1.46%

SEQ ID NO: 748
DONTFPETG
1.46%

SEQ ID NO: 749
KSEGWLLGD
1.45%

SEQ ID NO: 750
SPCFDVFEE
1.44%

SEQ ID NO: 751
TINKFPWVC
1.44%

SEQ ID NO: 752
IRFTTQIVD
1.44%

SEQ ID NO: 753
SVWEKTQMR
1.43%

SEQ ID NO: 754
CPCHKWASC
1.43%

SEQ ID NO: 755
HDHAAKQLD
1.43%

SEQ ID NO: 756
RPVGKLYMI
1.43%

SEQ ID NO: 757
MWGCKLFVC
1.42%

SEQ ID NO: 758
KHNPSRHDI
1.42%

SEQ ID NO: 759
NPYQCIVAG
1.39%

SEQ ID NO: 760
CMRGGQKLT
1.36%

SEQ ID NO: 761
DVDTSQFDR
1.34%

SEQ ID NO: 762
VCVKWRNVN
1.34%

SEQ ID NO: 763
WASPSVWRR
1.34%

SEQ ID NO: 764
YFHYRYDYG
1.33%

SEQ ID NO: 765
KVYYIISHQ
1.32%

SEQ ID NO: 766
QTYRGDWQK
1.31%

SEQ ID NO: 767
RVDKAGAIF
1.28%

SEQ ID NO: 768
VFNLMLQDK
1.27%

SEQ ID NO: 769
VKKPQYDMH
1.26%

SEQ ID NO: 770
QYVQCAAKD
1.23%

SEQ ID NO: 771
EGCYFWKQV
1.22%

SEQ ID NO: 772
TMKTWIYRN
1.22%

SEQ ID NO: 773
GWESFPHCG
1.21%

SEQ ID NO: 774
QKAEHLFVG
1.21%

SEQ ID NO: 775
PDSIPNSWC
1.21%

SEQ ID NO: 776
CHGRMLCYI
1.20%

SEQ ID NO: 777
NQQRTYNSG
1.19%

SEQ ID NO: 778
LDTKTLANS
1.19%

SEQ ID NO: 779
FAPEFDNHC
1.19%

SEQ ID NO: 780
REFAISGGS
1.19%

SEQ ID NO: 781
AFKTFGHVN
1.18%

SEQ ID NO: 782
ASSRLAEVI
1.18%

SEQ ID NO: 783
RMSAEKERS
1.17%

SEQ ID NO: 784
TMGLGVNAI
1.17%

SEQ ID NO: 785
PADWDREHG
1.17%

SEQ ID NO: 786
PELRPCAPR
1.17%

SEQ ID NO: 787
PDTQSAFWI
1.17%

SEQ ID NO: 788
KFKWLVRMT
1.16%

SEQ ID NO: 789
MTNRCAKYK
1.15%

SEQ ID NO: 790
AQPRTCQII
1.13%

SEQ ID NO: 791
IVVFGQMGY
1.13%

SEQ ID NO: 792
KVVFMFRWY
1.12%

SEQ ID NO: 793
CQTHWEKGD
1.12%

SEQ ID NO: 794
SGTEHVALH
1.12%

SEQ ID NO: 795
LCDLIMTDE
1.11%

SEQ ID NO: 796
PTLDRWEYH
1.10%

SEQ ID NO: 797
QLPIYIQKS
1.08%

SEQ ID NO: 798
HLAWIVNED
1.00%

SEQ ID NO: 799
HYFKWLFVE
0.98%

SEQ ID NO: 800
KFTWKKYFA
0.97%

SEQ ID NO: 801
HIWPPLFYW
0.94%

SEQ ID NO: 802
TWMPKVEYK
0.91%

SEQ ID NO: 803
ALTAVHAGT
0.90%

SEQ ID NO: 804
TLNSWHPEC
0.90%

SEQ ID NO: 805
AHMSYCLSR
0.89%

SEQ ID NO: 806
TDYPWVLHG
0.89%

SEQ ID NO: 807
WFSIIMSKL
0.89%

SEQ ID NO: 808
SVKVKNGFT
0.89%

SEQ ID NO: 809
NHIFKKVTW
0.88%

SEQ I0 NO: 810
KPPACIGGK
0.88%

SEQ ID NO: 811
PPDARQSLR
0.88%

SEQ ID NO: 812
RQTGTGNME
0.88%

SEQ ID NO: 813
EQFADQAPR
0.88%

SEQ ID NO: 814
ISISAQDID
0.88%

SEQ ID NO: 815
KFTLQQMAS
0.88%

SEQ ID NO: 816
SFSGTHLGW
0.88%

SEQ ID NO: 817
NHSDTDTAQ
0.86%

SEQ ID NO: 818
AFCYSYYQN
0.80%

SEQ ID NO: 819
EGKFTKAAV
0.80%

SEQ ID NO: 820
VQCLWWHKE
0.77%

SEQ ID NO: 821
QNLMYCSSV
0.72%

SEQ ID NO: 822
RRWLNPAYN
0.69%

SEQ ID NO: 823
PWFYKDWPH
0.66%

SEQ ID NO: 824
QALWLESDN
0.62%

SEQ ID NO: 825
TLNLRHPEC
0.62%

SEQ ID NO: 826
APEFRNRLT
0.60%

SEQ ID NO: 827
DVMRMGHYL
0.60%

SEQ ID NO: 828
ERRAHNHYA
0.60%

SEQ ID NO: 829
TSHKMMVLS
0.59%

SEQ ID NO: 830
EVDRNTNWC
0.59%

SEQ ID NO: 831
MALPLRVLS
0.59%

SEQ ID NO: 832
PCKSTRSIL
0.59%

SEQ ID NO: 833
TTDGMMGDH
0.59%

SEQ ID NO: 834
YPVHLMPPL
0.59%

SEQ ID NO: 835
ECWMAVQSC
0.59%

SEQ ID NO: 836
ERTACKTKS
0.59%

SEQ ID NO: 837
GTARHNMQL
0.59%

SEQ ID NO: 838
GTVDAQDRE
0.59%

SEQ ID NO: 839
RQIGTGNTE
0.59%

SEQ ID NO: 840
SPDGRGLCG
0.59%

SEQ ID NO: 841
TLNSRHLEC
0.59%

SEQ ID NO: 842
VLTDGACAA
0.59%

SEQ ID NO: 843
AHMYYCLSR
0.58%

SEQ ID NO: 844
AMLMSDGGG
0.58%

SEQ ID NO: 845
AQGFKPSGG
0.58%

SEQ ID NO: 846
ATGTYNLEE
0.58%

SED ID NO: 847
AYAASKNAM
0.58%

SEQ ID NO: 848
CAWYMKQGN
0.58%

SEQ ID NO: 849
CCRQIWIQY
0.58%

SEQ ID NO: 850
CMKTFPMNA
0.58%

SEQ ID NO: 851
DRINLMWKD
0.58%

SEQ ID NO: 852
DRVDLMWKD
0.58%

SEQ ID NO: 853
DSATFESAD
0.58%

SEQ ID NO: 854
EFEMRDSGY
0.58%

SEQ ID NO: 855
EKYQIHRDR
0.58%

SEQ ID NO: 856
HNQRTYPAL
0.58%

SEQ ID NO: 857
KMTNSSENS
0.58%

SEQ ID NO: 858
KQPTFPMLA
0.58%

SEQ ID NO: 859
LMRWYNRCW
0.58%

SEQ ID NO: 860
LRCTVMDPG
0.58%

SEQ ID NO: 861
LYQKDQRFG
0.58%

SEQ ID NO: 862
MHVNRRIDQ
0.58%

SEQ ID NO: 863
NCKSDQIAM
0.58%

SEQ ID NO: 864
NNHRLNDAN
0.58%

SEQ ID NO: 865
QPKVHGNYL
0.58%

SEQ ID NO: 866
QVEDRYWSY
0.58%

SEQ ID NO: 867
RRGYVNIEW
0.58%

SEQ ID NO: 868
SPCPTWDGL
0.58%

SEQ ID NO: 869
SRLLVHECP
0.58%

SEQ ID NO: 870
SSMQCVMRP
0.58%

SEQ ID NO: 871
SVTYHEWHL
0.58%

SEQ ID NO: 872
TCICSHTYP
0.58%

SEQ ID NO: 873
TGMAVSEQS
0.58%

SEQ ID NO: 874
TIEHGAWKC
0.58%

SEQ ID NO: 875
TPLTKWLPI
0.58%

SEQ ID NO: 876
VSIPNLGWP
0.58%

SEQ ID NO: 877
YCHLEDRIT
0.58%

SEQ ID NO: 878
EWMVIMAKN
0.40%

SEQ ID NO: 879
RVAIMFTGT
0.35%

SEQ ID NO: 880
IPDIMRRCP
0.33%

SEQ ID NO: 881
ERTACKMRS
0.33%

SEQ ID NO: 882
YTTVEGRIT
0.33%

SEQ ID NO: 883
DLEFCVMER
0.32%

SEQ ID NO: 884
YAQRTCWSE
0.31%

SEQ ID NO: 885
DRVNLTWKD
0.31%

SEQ ID NO: 886
DSTPNLLDP
0.31%

SEQ ID NO: 887
EPLYSWVGV
0.31%

SEQ ID NO: 888
DQLFMKFWS
0.31%

SEQ ID NO: 889
FVGLMSQCG
0.31%

SEQ ID NO: 890
KPCSSMSIQ
0.31%

SEQ ID NO: 891
SIYQKWVTS
0.31%

SEQ ID NO: 892
TLSSRHPEC
0.31%

SEQ ID NO: 893
AQTNNTCSI
0.31%

SEQ ID NO: 894
IPSHVHLGE
0.31%

SEQ ID NO: 895
MALPPHVLS
0.31%

SEQ ID NO: 896
MGCIGNCIL
0.31%

SEQ ID NO: 897
QTQHYGSTT
0.31%

SEQ ID NO: 898
RVDRNTNWC
0.31%

SEQ ID NO: 899
TYDTRSHCL
0.31%

SEQ ID NO: 900
ARVDHQCSG
0.30%

SEQ ID NO: 901
NGSHYNAMR
0.30%

SEQ ID NO: 902
QTQLYQNDE
0.30%

SEQ ID NO: 903
SPYGRGLCG
0.30%

SEQ ID NO: 904
TYSLAMSLG
0.30%

SEQ ID NO: 905
WVGNANRPA
0.30%

SEQ ID NO: 906
AHVDHQCGG
0.30%

SEQ ID NO: 907
APEFNVARR
0.30%

SEQ ID NO: 908
CCQQIWVQY
0.30%

SEQ ID NO: 909
CITARSTAA
0.30%

SEQ ID NO: 910
CTIARSTAA
0.30%

SEQ ID NO: 911
DVIRTTHSG
0.30%

SEQ ID NO: 912
GSQQTTPSC
0.30%

SEQ ID NO: 913
GYATFPDSD
0.30%

SEQ ID NO: 914
ISSTMVEHG
0.30%

SEQ ID NO: 915
KLLFMKFWN
0.30%

SEQ ID NO: 916
KVDRNTYWC
0.30%

SEQ ID NO: 917
LFRAYAWSQ
0.30%

SEQ ID NO: 918
LITRDTGLA
0.30%

SEQ ID NO: 919
LSECSDRTN
0.30%

SEQ ID NO: 920
NCKPNQIAM
0.30%

SEQ ID NO: 921
PAVPADEFF
0.30%

SEQ ID NO: 922
QATDMHRWQ
0.30%

SEQ ID NO: 923
STARHNTQL
0.30%

SEQ ID NO: 924
THMNYCLSR
0.30%

SEQ ID NO: 925
TSHRMMVFS
0.30%

SEQ ID NO: 926
AMTRCMAGH
0.30%

SEQ ID NO: 927
CPEHNLVGV
0.30%

SEQ ID NO: 928
CPEHSQVGV
0.30%

SEQ ID NO: 929
DRANLMWKD
0.30%

SEQ ID NO: 930
DRVNLMWKG
0.30%

SEQ ID NO: 931
DWGYCHSKA
0.30%

SEQ ID NO: 932
EFVLGYQGI
0.30%

SEQ ID NO: 933
EKYQIHWDH
0.30%

SEQ ID NO: 934
EQFVDRAPR
0.30%

SEQ ID NO: 935
EROAHSHYA
0.30%

SEQ ID NO: 936
KIYQKWVTN
0.30%

SEQ ID NO: 937
GQFRSGAHR
0.30%

SEQ ID NO: 938
HTNEQYLQH
0.30%

SEQ ID NO: 939
HVNTSCKQS
0.30%

SEQ ID NO: 940
KYSPIQEHN
0.30%

SEQ ID NO: 941
LCLQGAKMD
0.30%

SEQ ID NO: 942
LMSITSAVE
0.30%

SEQ ID NO: 943
LTHNIPFPK
0.30%

SEQ ID NO: 944
MALPLHALS
0.30%

SEQ ID NO: 945
MCQSVEKEY
0.30%

SEQ ID NO: 946
NGNRYNAMR
0.30%

SEQ ID NO: 947
NHRSREIDI
0.30%

SEQ ID NO: 948
NIVRSRSIT
0.30%

SEQ ID NO: 949
NKTCYLTCP
0.30%

SEQ ID NO: 950
PQARVDMEF
0.30%

SEQ ID NO: 951
QAAGMHRWQ
0.30%

SEQ ID NO: 952
REFVISGGN
0.30%

SEQ ID NO: 953
SMREYQKHI
0.30%

SEQ ID NO: 954
SPHSCQHMG
0.30%

SEQ ID NO: 955
SRLSVHECP
0.30%

SEQ ID NO: 956
STTRHNMQL
0.30%

SEQ ID NO: 957
TFHFSILLT
0.30%

SEQ ID NO: 958
TPQPMNVAA
0.30%

SEQ ID NO: 959
TYIQHPTNA
0.30%

SEQ ID NO: 960
VAGRFGSNT
0.30%

SEQ ID NO: 961
VSMPSLGWP
0.30%

SEQ ID NO: 962
YDNHQKARI
0.30%

SEQ ID NO: 963
AHMIYCLSR
0.29%

SEQ ID NO: 964
AHMNYCLSH
0.29%

SEQ ID NO: 965
AQWIQQKHG
0.29%

SEQ ID NO: 966
ARTLREGWG
0.29%

SEQ ID NO: 967
ASKLEVQEQ
0.29%

SEQ ID NO: 968
AVEHGAWKC
0.29%

SEQ ID NO: 969
AVETGESAV
0.29%

SEQ ID NO: 970
CCRQIWVQH
0.29%

SEQ ID NO: 971
CGGQETGLA
0.29%

SEQ ID NO: 972
CNSWCTGSL
0.29%

SEQ ID NO: 973
DCRHTTLYV
0.29%

SEQ ID NO: 974
DESTCVAHG
0.29%

SEQ ID NO: 975
DGMHRELHS
0.29%

SEQ ID NO: 976
DKYGLGIFH
0.29%

SEQ ID NO: 977
DRVYLMWKD
0.29%

SEQ ID NO: 978
DTDYKPHFA
0.29%

SEQ ID NO: 979
DVHDGQLCY
0.29%

SEQ ID NO: 980
DVYDSQLCY
0.29%

SEQ ID NO: 981
DYAPTNTMM
0.29%

SEQ ID NO: 982
DYSQVNCER
0.29%

SEQ ID NO: 983
DYSQVNREK
0.29%

SEQ ID NO: 984
EEYYTWECH
0.29%

SEQ ID NO: 985
ETHHEYRDD
0.29%

SEQ ID NO: 986
EYACMLMTR
0.29%

SEQ ID NO: 987
FLRPKRHFS
0.29%

SEQ ID NO: 988
GCQLGFQIT
0.29%

SEQ ID NO: 989
GQFRRGAYR
0.29%

SEQ ID NO: 990
GSLSKDTPL
0.29%

SEQ ID NO: 991
GSQQTTLSR
0.29%

SEQ ID NO: 992
HCHQEDRIT
0.29%

SEQ ID NO: 993
HHECLYQIY
0.29%

SEQ ID NO: 994
HVDQHRIQA
0.29%

SEQ ID NO: 995
IADIGRKQW
0.29%

SEQ ID NO: 996
IAWPTDCSF
0.29%

SEQ ID NO: 997
IYCPCQDSD
0.29%

SEQ ID NO: 998
KAALFLKDC
0.29%

SEQ ID NO: 999
KAGMDQTAV
0.29%

SEQ ID NO: 1000
KCRMAVQSC
0.29%

SEQ ID NO: 1001
KPLASIGQH
0.29%

SEQ ID NO: 1002
KQLFMEFWN
0.29%

SEQ ID NO: 1003
KSLQEQGGA
0.29%

SEQ ID NO: 1004
KTQLSDHKH
0.29%

SEQ ID NO: 1005
LDHTNLMKL
0.29%

SEQ ID NO: 1006
LIQWYNRCW
0.29%

SEQ ID NO: 1007
LMKTSPCYW
0.29%

SEQ ID NO: 1008
LMLGYMEQD
0.29%

SEQ ID NO: 1009
LSECSDQAN
0.29%

SEQ ID NO: 1010
LVVIDPAHY
0.29%

SEQ ID NO: 1011
MALPLYVLS
0.29%

SEQ ID NO: 1012
MCOMSRRSI
0.29%

SEQ ID NO: 1013
MMLGYMERD
0.29%

SEQ I0 NO: 1014
MTQVHQALV
0.29%

SEQ ID NO: 1015
NACPQHDFG
0.29%

SEQ ID NO: 1016
NALWMTKAP
0.29%

SEQ ID NO: 1017
NCRHTILYV
0.29%

SEQ ID NO: 1018
NDCICRAIL
0.29%

SEQ ID NO: 1019
NFANHQPLP
0.29%

SEQ ID NO: 1020
NQRGLCKDE
0.29%

SEQ ID NO: 1021
PACLLRRML
0.29%

SEQ ID NO: 1022
PANPSFSYL
0.29%

SEQ ID NO: 1023
PCQQLMCFT
0.29%

SEQ ID NO: 1024
PDARQSLRH
0.29%

SEQ ID NO: 1025
PIYQKWVIN
0.29%

SEQ ID NO: 1026
PNGCRPFCC
0.29%

SEQ ID NO: 1027
PQHAGRAEH
0.29%

SEQ ID NO: 1028
QCFLWNDYQ
0.29%

SEQ ID NO: 1029
QHLMTLSHE
0.29%

SEQ ID NO: 1030
QPLSRNGAR
0.29%

SEQ ID NO: 1031
QPQLRNGAR
0.29%

SEQ ID NO: 1032
QRQSQEWFH
0.29%

SEQ ID NO: 1033
QTGVMAAIG
0.29%

SEQ ID NO: 1034
QVVIIECKN
0.29%

SEQ ID NO: 1035
RDRRVRDEC
0.29%

SEQ ID NO: 1036
RLETTRYIW
0.29%

SEQ ID NO: 1037
RMARVSVHG
0.29%

SEQ ID NO: 1038
RQHHCLPVI
0.29%

SEQ ID NO: 1039
RQVGTGNME
0.29%

SEQ ID NO: 1040
RSTRFIGIA
0.29%

SEQ ID NO: 1041
RTNEQYPQH
0.29%

SEQ ID NO: 1042
RTRISQMSG
0.29%

SEQ ID NO: 1043
RTRTSQMFG
0.29%

SEQ ID NO: 1044
RVNGSTCGH
0.29%

SEQ ID NO: 1045
SATTSGTAF
0.29%

SEQ ID NO: 1046
SATYHEWHP
0.29%

SEQ ID NO: 1047
SEAKYDTAT
0.29%

SEQ ID NO: 1048
SKRWYGHSA
0.29%

SEQ ID NO: 1049
SMQEYQKHV
0.29%

SEQ ID NO: 1050
SRCTVVDPG
0.29%

SEQ ID NO: 1051
SSWMQMQGP
0.29%

SEQ ID NO: 1052
TAMVSECID
0.29%

SEQ ID NO: 1053
TEEHMMIQT
0.29%

SEQ ID NO: 1054
TGRWLVYQA
0.29%

SEQ ID NO: 1055
TLNSRHPEY
0.29%

SEQ ID NO: 1056
TSEPHCSFA
0.29%

SEQ ID NO: 1057
TVDGMMGDH
0.29%

SEQ ID NO: 1058
TVPKWDNIS
0.29%

SEQ ID NO: 1059
TYHCYMSWM
0.29%

SEQ ID NO: 1060
TYIQCPTNA
0.29%

SEQ ID NO: 1061
TYIQYPANA
0.29%

SEQ ID NO: 1062
VHGQTDLSL
0.29%

SEQ ID NO: 1063
VPVPFLSLR
0.29%

SEQ ID NO: 1064
VSFQRMNNM
0.29%

SEQ ID NO: 1065
VSTPNLGWP
0.29%

SEQ ID NO: 1066
VSVQGRAQL
0.29%

SEQ ID NO: 1067
VTEVLNQDV
0.29%

SEQ ID NO: 1068
YATHMQHLE
0.29%

SEQ ID NO: 1069
YCHQEDRII
0.29%

SEQ ID NO: 1070
YETDSEGYD
0.29%

SEQ ID NO: 1071
YPAHMMPPL
0.29%

SEQ ID NO: 1072
YWCPSQRWR
0.29%

SEQ ID NO: 1073
QLKYAVGAN
0.23%

SEQ ID NO: 1074
MNHQCMLWD
0.07%

SEQ ID NO: 1075
RHQRVMPTY
0.05%

SEQ ID NO: 1076
NSGIATADH
0.05%

SEQ ID NO: 1077
ACHTGQSGM
0.04%

SEQ ID NO: 1078
SHWFFHFDL
0.03%

SEQ ID NO: 1079
DVTQTRYCY
0.03%

SEQ ID NO: 1080
FSEQNVSAA
0.03%

SEQ ID NO: 1081
GPLYSWVGI
0.03%

SEQ ID NO: 1082
NAGMKGVDV
0.03%

SEQ ID NO: 1083
CTEAGMTFG
0.02%

SEQ ID NO: 1084
PMNDMQFVP
0.02%

SEQ ID NO: 1085
SGRRSGGRP
0.02%

SEQ ID NO: 1086
DVAQYGLGS
0.02%

SEQ ID NO: 1087
NPTDLLQHE
0.02%

SEQ ID NO: 1088
QAQCIAMQR
0.02%

SEQ ID NO: 1089
DGMHRELCS
0.02%

SEQ ID NO: 1090
DGTHRELYS
0.02%

SEQ ID NO: 1091
KWCQFGMVA
0.02%

SEQ ID NO: 1092
LTHNTPFPE
0.02%

SEQ ID NO: 1093
PMPTADVLH
0.02%

SEQ ID NO: 1094
QGEEVWWEA
0.02%

SEQ ID NO: 1095
RMVTMERDR
0.02%

SEQ ID NO: 1096
RVNGSTCWH
0.02%

SEQ ID NO: 1097
RWYQFGMVA
0.02%

SEQ ID NO: 1098
TFLEYQALG
0.02%

SEQ ID NO: 1099
TTGVCSFAQ
0.02%

SEQ ID NO: 1100
YHKQMAVWW
0.02%

SEQ ID NO: 1101
AVSVACLNT
0.01%

SEQ ID NO: 1102
CDSPSLLQS
0.01%

SEQ ID NO: 1103
CMYLPALQY
0.01%

SEQ ID NO: 1104
CSSWANGCD
0.01%

SEQ ID NO: 1105
EDCPKVFTI
0.01%

SEQ ID NO: 1106
EFALGCQGI
0.01%

SEQ ID NO: 1107
ERGEMRATG
0.01%

SEQ ID NO: 1108
ERGGMKATG
0.01%

SEQ ID NO: 1109
GRVHGNVCI
0.01%

SEQ ID NO: 1110
HVWSLEWHL
0.01%

SEQ ID NO: 1111
IPGHVHFGF
0.01%

SEQ ID NO: 1112
IWGVYVHYF
0.01%

SEQ ID NO: 1113
NHDEKLWAP
0.01%

SEQ ID NO: 1114
NVWSLAVRD
0.01%

CNS (cortex forebrain, cortex occipital, cortex temporal, thalamus, hypothalamus, substantia nigra, hippocampus DG, hippocampus CA1, hippocampus CA3, cerebellum), liver, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord tissues were analyzed to identify amino acid residues in the AAV5 VP1 581-589 region that drive CNS tropism. The results are shown in FIG. 18B which shows the positional frequency of each amino acid in the 581 to 589 region of AAV5 VP1 in CNS tissue as compared to all other analyzed tissues, wherein the capsid polypeptide has the sequence of SEQ ID NO: 2. The position (“Xaan”, n=1-9) is denoted on the x-axis and amino acid residues are denoted on the y-axis. The frequency of a particular amino acid residue in a particular position is indicated in the heatmap as intensity. Darker squares indicate a higher frequency of a particular amino acid in a particular position and lighter squares indicate a lower frequency of a particular amino acid in a particular position.

The frequency of a given amino acid residue occurring at a specified position corresponding to position 581 to position 589 of SEQ ID NO: 1 (generalized in SEQ ID NO: 2) in variants identified in CNS over the frequency of that given amino acid residue occurring at the specified position in variants identified in all other harvested tissues was analyzed to identify a set of sequence rules for capsids that preferentially target CNS tissue. With reference to TABLE 9 below, and SEQ ID NO: 2 (AAV5 VP1), the following amino acids can, thus, be independently mutated, in any combination, at any one or more positions Xaa1-Xaa9, to provide a VP1 capsid with enhanced CNS tropism as compared wildtype AAV5 VP1 capsid (SEQ ID NO: 1), where CNS tropism here refers to properties that are preferred for CNS transduction over properties that are preferred for transduction of all other harvested tissues.

TABLE 9

CNS Tropism Rules

Xaa1 is selected from A, C, K, M, Q, R, T, or W

Xaa1 is selected from K, Q, R, or W

Xaa1 is K

Xaa2 is selected from F, I, K, R, T, or W

Xaa2 is selected from F, I, R or T

Xaa2 is R

Xaa3 is selected from A, H, N, R, or W

Xaa3 is selected from A, R, or W

Xaa3 is R

Xaa4 is selected from E, G, I, M, Q, or R

Xaa4 is selected from E, M, or R

Xaa4 is R

Xaa5 is selected from C, G, K, I, M, or R

Xaa5 is selected from K, I, or R

Xaa5 is I

Xaa6 is selected from I, K, L, P, Q, R, Y

Xaa6 is selected from K, R, or Y

Xaa6 is R

Xaa7 is selected from D, I, K, R, V, or W

Xaa7 is selected from I, R, or V

Xaa7 is V

Xaa8 is selected from C, G, H, K, L, or V

Xaa8 is selected from H, K, or V

Xaa8 is H

Xaa9 is selected from I, K, L, R, or V

Xaa9 is selected from I, K, or R

Xaa9 is R

TABLE 10 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that were found in CNS tissue and comport to one or more of the rules provided in TABLE 9. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 7118-SEQ ID NO: 8117 as disclosed in TABLE 10. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 10

Sequences of the 581 to 589 Region in AAV5 VP1

Capsid Polypeptide that Drive CNS Tissue Tropism

SEQ

ID
581-589

NO
Sequence

7118
KTTFDVACY

7119
KDGYREAWM

7120
KTSNVMDQM

7121
KTLHMLESK

7122
KSHDYHGRA

7123
KSEWTCCPQ

7124
KMHGNERCF

7125
KQTVEVDQE

7126
KVAHEMMSA

7127
KMNTWDQAR

7128
KFNTKDQLM

7129
KCQSKECGG

7130
KGLITENNY

7131
KVEWRWGSA

7132
KTFQYMGPT

7133
KCGLTPGVE

7134
KQRIAYEWQ

7135
KGQRRDSVS

7136
KQCSLAPTI

7137
KEWCVQQQC

7138
KKDPLHLGD

7139
KQTKSMVYM

7140
KISGLNDAS

7141
KPDILKTKT

7142
KWSAYFLIE

7143
KGESQCTSQ

7144
KQEYIMRQR

7145
KLDPRQADQ

7146
KIYMQCDQE

7147
KWAASHKHD

7148
KENKIMQAS

7149
KVAGGHETT

7150
KTDPIAEYC

7151
KPDHAMAQQ

7152
KSDKVWYVL

7153
KIEAEEMHA

7154
KKTECEMIV

7155
KGTPNWWGP

7156
KPAGKNTCW

7157
KSVHYETCE

7158
KCNVITYHG

7159
KQEQFHTGS

7160
KTMADMYNR

7161
KTVETYAAH

7162
KSWVWVDTE

7163
KAIWQGTEN

7164
KPLTYTVGS

7165
KYASMDIYS

7166
KCTMITPCF

7167
KEAYTDMSC

7168
KHMPTLQWQ

7169
KIEYGHWIH

7170
KYSPFQEHN

7171
KHTLAEMYG

7172
KIWAGVNLG

7173
KCDDRNTQH

7174
KSIMMSQLD

7175
KKIFCSWDY

7176
KQAGYDFFH

7177
KSDAQRNVA

7178
KPPSWPMSL

7179
KAHDITWNL

7180
KQATNNNGW

7181
KDPDCYDWP

7182
KHTEYCSHE

7183
KWYHYNEMT

7184
KGQMISETR

7185
KHEPMCVQL

7186
KVDTVRNPK

7187
KRVQQMETM

7188
KIIFELGMD

7189
KSGWGAPYE

7190
KIQMAVSLD

7191
KVPSQLVFE

7192
KSPDMTLLL

7193
KSMTCVYYP

7194
KPDKPNDYA

7195
KFICTTIAC

7196
KPAYEHRGF

7197
KNNTEDGGR

7198
KATREYDIP

7199
KCETKKRFN

7200
KTEKWASHE

7201
KATVEGSWR

7202
KRRDGTRSY

7203
KALHYAMIK

7204
KWEVMSAAY

7205
KALYLHTIG

7206
KLNPFSTGT

7207
KVEIINWHP

7208
KNDPIDNEG

7209
KTFTKEVTP

7210
KCGNEGRMS

7211
KNETFIAST

7212
KDNQSCEWN

7213
KAQVVPVEC

7214
KMMAFEKQC

7215
KWQEMPTFQ

7216
KYCPPQTPN

7217
KCAMFNAGE

7218
KSTEPQTYP

7219
KFTEALSCG

7220
KGEIEGGAK

7221
KQLTFPMLA

7222
KEFVGFQAM

7223
KHDGFSVVG

7224
KATRDPKFP

7225
KRWLHMYQG

7226
KHLDMTSCH

7227
KDEQFARCK

7228
KTLPCSNSE

7229
KYMQENVLS

7230
KSAPRENFN

7231
KQRMTRQGE

7232
KARQDDVNT

7233
KCGVEHEHR

7234
KSELFMKLH

7235
KNELENVEN

7236
KYWCKNEWD

7237
KASLGACVE

7238
KSCNDGQQN

7239
KVTPELEEV

7240
KQEWHVHTE

7241
KPKCDEQKS

7242
KWHGGPGVV

7243
KGNSVTHVQ

7244
KWDDMHTQM

7245
KQAHLEVAW

7246
KVNACGNLT

7247
KAIPQGSSC

7248
KENCRMDGC

7249
KQRDWENFA

7250
KMCWWPESM

7251
KQNLMCCHN

7252
KESPWDGFY

7253
KGMQMWNCE

7254
KAILNAMME

7255
KEDAHSNME

7256
KAPWAPMDS

7257
KRAIVGYAL

7258
KSGMIQEET

7259
KSYQMIMTW

7260
KFELHGTVG

7261
KSGTCGYGQ

7262
KSLQCASRH

7263
KTGTTMEPM

7264
KTDYSRLSD

7265
KHYKWGHCQ

7266
KLYGLNHSC

7267
KLKESMESA

7268
KAGYQEGSC

7269
KCWDCHEAG

7270
KPVQCACQN

7271
KDYWAGYQM

7272
KSEHHWINA

7273
KGGPWAYNC

7274
KYAYSLDCA

7275
KLWIDQRGH

7276
KLWIDQRGY

7277
QRVTKMHFE

7278
ERTACKMKS

7279
ERGEMKATG

7280
ARGTCNYCY

7281
QRNEMSASR

7282
FRDVCMHDV

7283
ERMMWEEYP

7284
HRMSYKWGW

7285
ERVADDMTC

7286
MRSECEHAM

7287
NRGEWLENF

7288
NRQFWYLNT

7289
PRLLRPNRV

7290
TRRVATHMP

7291
VRHSRNNTV

7292
SRWFQIGTE

7293
TRQWMCVHC

7294
HRVHSEKTD

7295
GRHTAFATG

7296
LRSRVEKDH

7297
ERMEGKACN

7298
NRLDQETRR

7299
SRSWFDSCG

7300
VRCQQHQGN

7301
YRSWMCWDI

7302
CRNTQYEQH

7303
ARKNCDHLN

7304
ARCQNHWSH

7305
MRQRTYICQ

7306
SRCTVMDPG

7307
PRNLVAMYW

7308
TRMQAVDHE

7309
FRAVYYYHS

7310
RREAMDDSP

7311
GRQSENEEA

7312
TRHPTIAMH

7313
YRLHVWMKV

7314
DRREKQLMK

7315
ERKAMRWSL

7316
LRMQKEGQE

7317
IRPQYAKMD

7318
SRTECVHGN

7319
NRCGVSKCC

7320
CRCNDAWNM

7321
FRKQGHGEK

7322
YRMEYGTCE

7323
SRPFNYFQR

7324
QRGPLQHQQ

7325
MRQCNYDFL

7326
MRQCNYDFW

7327
MRQCNYGFW

7328
SRTRMYMDH

7329
ERTLTKAGS

7330
WRLAALYQS

7331
CRCCPVMKD

7332
YRDRCWNRE

7333
LRNMRGDWD

7334
CRKCEISAR

7335
ARTQFHSDE

7336
ERSMKDADE

7337
YRRWQSNYT

7338
HRMNNHAWF

7339
QRTQCNFSI

7340
TRILNEFFC

7341
ARLEIEYMG

7342
WRFCDMRMS

7343
DRLYCKLAH

7344
ERKTTADWK

7345
GRSMVLHFV

7346
TRHDNGPSD

7347
PRAPERPPQ

7348
MRRKTPEDD

7349
ERFRVQEFQ

7350
ERYYPESSE

7351
GRAHGNVCI

7352
ARSGINGHE

7353
ARVALVKPG

7354
SRYVNNGTV

7355
ARAYNSYQA

7356
PRFKACHDP

7357
QRQGQLHYW

7358
SRCTIDVPS

7359
DRTMCKVQG

7360
LRVSQSFSN

7361
NRTNSPYVH

7362
CRGSLSNAN

7363
VRQSVPHWA

7364
HRQESPNNK

7365
TRNIGGVFQ

7366
RRWMNAKAT

7367
YRAHSAGYC

7368
YRAHSAGYY

7369
PPRTVARFK

7370
MTRSCQTEG

7371
SMRNSKGSG

7372
GGRSLHAYL

7373
GTREYGKVS

7374
EVRSTRKNH

7375
GLRHGFYAV

7376
LARTTTNSE

7377
VVREQFGIH

7378
ANRADWQHL

7379
WCRTPQREN

7380
WKRTAGVHF

7381
GYRNYFEWP

7382
TARNFGDPP

7383
EARESGMHD

7384
MARLYHPQM

7385
SNRYVSLFI

7386
PWRWHKSAC

7387
TLRAEMSFH

7388
EQRHISRAP

7389
TKRIKCCWE

7390
MKRAPTLDY

7391
ENREAVWFN

7392
AERDHKTIY

7393
ELRMSFTKV

7394
CVRIGMREP

7395
TARSTSDKC

7396
GKRVEDTSC

7397
NQRGFCKDE

7398
TPRCERADG

7399
DQRMLLKEL

7400
SNRYYSREF

7401
EDRMGLPHN

7402
EGRTESYGN

7403
ASRSCMTQE

7404
QSRQMMLLH

7405
MTRVSMTWR

7406
TFRHWEGHH

7407
VARQMQENQ

7408
INRPMGHGM

7409
CIRFMMREG

7410
FFRWRWGLG

7411
QGRWACWYS

7412
HHRVNYEMY

7413
SVRTCHQDE

7414
DDRVKMFHY

7415
GMRYSEWHW

7416
ETREHHLLV

7417
MFRQFYIQG

7418
TTRQISNID

7419
DCRHTILYV

7420
AIRQFMMHL

7421
LMRWSKDSN

7422
LMRGSKDSN

7423
LMRWSKDSS

7424
LMRWSRDSN

7425
ASRQFETCW

7426
SDRMVSESA

7427
NTREVCDLS

7428
EFRCRLEGS

7429
DCRCYDCHD

7430
TMRDSDYNW

7431
DVRACMEDL

7432
WCRCKRFEE

7433
WIRSAWRPG

7434
MGRPHAWWG

7435
NFRMQTGCH

7436
SMRDLIAEA

7437
TARDREHAM

7438
CSRNRCWSG

7439
DYREKYKCQ

7440
VCRQKTDSW

7441
YSRMEENDR

7442
PERYVKPAR

7443
QPRVYEGFY

7444
MDRMMYMLG

7445
VNRGDLSLF

7446
HTRKEGFNS

7447
LFRTIPIWQ

7448
APRREEQNC

7449
NTRQINGVD

7450
DARHKWCLE

7451
AERQACTMG

7452
ESRMLRQSS

7453
ETRVREDAV

7454
AMRAGLENS

7455
TCRWFAHYS

7456
QVRFEQNPA

7457
NERVYGWGM

7458
EKRLFDRTY

7459
VSRCGIDRS

7460
DARKTVNEW

7461
TIRYIGMSG

7462
FFRECGSYE

7463
NTRSDLRAG

7464
ANRMIWYPY

7465
AQRCFHRWG

7466
TCRYEELAD

7467
STRTTDWNM

7468
DARKTYECY

7469
SSRGQDGVM

7470
RTREPFWWW

7471
IERWWVIDV

7472
ENRSPNNPA

7473
DWRYFCVSL

7474
NPRQGFVPV

7475
NSRLLIKHL

7476
HTRVVKSTG

7477
EIRQFVKGA

7478
QERSMNAWW

7479
CARLEGRGN

7480
FMRDVFAWD

7481
MLRTQIHCA

7482
QLRCEYQHT

7483
CCRMDNKKS

7484
YERVGGNNE

7485
NVRNFFDHC

7486
ATRWVQMGQ

7487
MDRDIPHCR

7488
CYRRSDSFC

7489
EYSRCCHQE

7490
VHLRSHTWF

7491
QQLRCHCFS

7492
AAIRYDPGF

7493
MSNRIIDIQ

7494
VTQRPLHHS

7495
NQQRTYNNG

7496
SLGRIECDF

7497
SADRTAMFH

7498
NILRYLICD

7499
TFMRKICGM

7500
CWARSVAQN

7501
ELPRYYGQM

7502
EYSRMNKAS

7503
DKVRQGQKD

7504
EFNRYPHGN

7505
HEMRRDYKS

7506
QVYRMMMQP

7507
TVNRAMREE

7508
YKLRSSFRW

7509
FANRCPPDD

7510
NKIRDVAVE

7511
PVARKMSTC

7512
VIQRKFDSG

7513
ALDRERLIF

7514
AGSRMQWAM

7515
WSSREYWHK

7516
THARFKMKS

7517
NCARGPMEK

7518
LWSRFISMD

7519
IAMRCWGNE

7520
LVDRSGRTA

7521
SQSRTIIQE

7522
HEVRRYGQN

7523
NSTRNTSYY

7524
GPARSFEAC

7525
YCPRLVEAW

7526
VIGRTVDDP

7527
WFDRSWHIL

7528
CYIRNMVNL

7529
LIGRDSKVW

7530
QNERLGVPT

7531
VICRMKKHS

7532
VEDRVFACC

7533
GHLRDHPLY

7534
FEFRKCDHW

7535
RKTRCFCNY

7536
TSERHSCMV

7537
SMSRHMMHQ

7538
DEGRFTAYY

7539
GVSRLPYHA

7540
QVDRTMWMH

7541
CELRKPSYH

7542
NNDRHQHLK

7543
TVDRKVEIE

7544
NSARYAWVY

7545
AITRHVADT

7546
TDKRAHVGA

7547
LLWRRVKRT

7548
YVIRKTWFN

7549
FAERANYCC

7550
SYMRHPNFD

7551
GGDRMATDR

7552
FQTRPSCSF

7553
MMVRREFAD

7554
AVMRNDQWL

7555
LQERDHHMQ

7556
LTPRSEGLS

7557
LVGRTYQHQ

7558
DATRAQTGE

7559
MMVRELMSS

7560
CGHRHSSMG

7561
HHLRNVHHC

7562
NSTRWYHTG

7563
NHMRKQSSH

7564
DDCRLNVRT

7565
MCARGAMCA

7566
SWGRIPHES

7567
QLKRNEQTS

7568
RNQRTYPAL

7569
SFFREDYSE

7570
DSHRCLKAD

7571
SAPRNEITR

7572
DIQRWEHEG

7573
EQLRDLNIE

7574
NYARHDILA

7575
MGQRSAANY

7576
STARHNMQL

7577
PAYRMECCE

7578
ETHRGTVGL

7579
DVCRKPVSH

7580
DVMRMEHYL

7581
PAWRRYCWT

7582
DMHRGGMWT

7583
CTLRQIKMH

7584
YAQRTCWPE

7585
WCMRDWYQE

7586
ESQRGTLWC

7587
RMVRVSVHG

7588
EAWRNSSSG

7589
LWDRFCCPV

7590
NHTRGKQCV

7591
NCERWFQIE

7592
QAVRTRTFR

7593
AYLRRNVGN

7594
LTNRTQESW

7595
NVDRQHAQS

7596
DYERSIENH

7597
MPTRMQKGA

7598
MSARENTPD

7599
CSERGESAF

7600
MTERYYWCQ

7601
CHIRDCSPI

7602
NNVRKVGNT

7603
VEMRANKGL

7604
WDGRPDLST

7605
ESYQIQCQN

7606
TSSEITSPP

7607
CTKPIPCWM

7608
IMHEIPEID

7609
HQSPIHTNT

7610
REMMIVNCD

7611
IWPQIQYRM

7612
PEKWIHMHE

7613
ILIFIQNCP

7614
SACNIEYHT

7615
SGNQIVQAP

7616
HWELIRCHD

7617
NSHTIMNGM

7618
DVDIIGLPA

7619
QKHMILCHF

7620
ACEWINMYN

7621
ATYQIQWPG

7622
FIIWIEHRH

7623
GIEQIMGCP

7624
VIMGIQTDD

7625
VQPTESYSD

7626
YSPLIGRWK

7627
IHSNIPVAE

7628
NHDLIQFVK

7629
WSPSIPGGE

7630
CNYPIIWDK

7631
VSAYIERQW

7632
QVSHISEHW

7633
MMTTIQYHD

7634
WAEAIKHFT

7635
VSKFIAASW

7636
CSNDINGRA

7637
LNSMIMWGN

7638
HACVITLDG

7639
GTEPIMWRD

7640
LSTLITMMD

7641
GWALIVNGL

7642
MVNHILPNN

7643
RSLDIQYDI

7644
RVVPIEDSM

7645
LVNTIWKAA

7646
GYKFIGRSR

7647
QYLLIPHLP

7648
NNGPIWISS

7649
YGENISPSP

7650
QSDWISIPC

7651
CMAHIWEGH

7652
NWFGINSWH

7653
FQTSINYYG

7654
TMEYIGLDQ

7655
GKIPIDTFR

7656
WAYYIRLQP

7657
VAYPIQTDV

7658
LSIQINNQI

7659
EQTYISEWG

7660
TCGDIQLRA

7661
MAGHIGHHS

7662
GFKDILMVQ

7663
VWWMIIHTS

7664
NAAMIATVC

7565
SMHLICGEA

7666
ACSQIGQDD

7667
DSQEIGSES

7668
NWEVIGRWH

7669
HSGFIDRQA

7670
PVKPIIHAS

7671
IQAVIASGW

7672
DHLSICCQN

7673
CTKPIVMDW

7674
LNHPIADYD

7675
TQAKIECCS

7676
GTEHIPMQA

7677
SFPMILVNC

7678
GCQQIQLQT

7679
ATDMIFVSN

7680
RQCVIGDYD

7681
RLAWIEQSP

7682
YEHPIMWPV

7683
HEGLIMRLE

7684
TSYCIQTVD

7685
AMEMIVMHE

7686
MGCLIQFCE

7687
FSWPIHEPD

7688
ESTHIMKTP

7689
HPDKIEYWR

7690
TFGCIESMQ

7691
EDGLIVYGP

7692
DHIHIERAN

7693
GCMCIRHAY

7694
APIPINMEG

7695
QMNAIQLCV

7696
SQSQIRQAD

7697
PIVLIHYDA

7698
VADWIGHTP

7699
GSNSITRDT

7700
QWEDITKCS

7701
EDQWISNPF

7702
QCGFIRLNE

7703
DSDPITVDL

7704
EIGIIRHGH

7705
FQTLILSGL

7706
DNNMIVRCA

7707
MAPDIPHCR

7708
MDADIPHCR

7709
MDPAIPHCR

7710
MDPDIAHCR

7711
MDPDILHCR

7712
MDPDIPHCC

7713
MDPDIPHCG

7714
MDPDIPHCH

7715
MDPDIPHCL

7716
MDPDIPHCR

7717
MDPDIPHCS

7718
MDPDIPHFR

7719
MDPDIPHGR

7720
MDPDIPHWR

7721
MDPDIPNCR

7722
MDPDIPPCR

7723
MDPDIPRCR

7724
MDPDIPYCR

7725
MDPDIRHCR

7726
MDPGIPHCR

7727
MDPNIPHCR

7728
MDPVIPHCR

7729
MDPY1PHCR

7730
MDTDIPHCR

7731
MNPDIPHCR

7732
MYPD1PHCR

7733
VDPDIPHCR

7734
HVDQHRVQA

7735
MYLNYRHFA

7736
AVASERQLR

7737
LEQLDRCMY

7738
LFHQGREPM

7739
CVCEMRTVD

7740
NDCTCRAIM

7741
TIDVKRWGQ

7742
EATYTRYMT

7743
HSGYKRQMK

7744
TYMEYRQHK

7745
VDCWLRDPI

7746
YKTEFRTKL

7747
DPYNTRFRT

7748
LNYMRRAMW

7749
FAIWKRNES

7750
WKDDLRSTY

7751
ENQQRRNFG

7752
IMPPKRYFV

7753
VAADSRMMI

7754
IMCHARMMA

7755
RDIWERPCV

7756
EHSSPREDN

7757
EVHLFRGEG

7758
MYTDMRCWR

7759
LIHYCRCIN

7760
HQCQSRSYI

7761
GTAVSRQSK

7762
VMFYWRYNS

7763
FGDHLRVMY

7764
NSEWLRYTF

7765
AVHWHRKCE

7766
PMELWRTWY

7767
SQDCGRPES

7768
AAKKGRDLH

7769
QNLWYRPLT

7770
TCYCHRAHE

7771
LACTPRCIF

7772
VAFGMRWLL

7773
WAQCDRAVC

7774
FCDIGRSGG

7775
GHQWDRLLA

7776
WLTEKRHFE

7777
MTGQCRNSQ

7778
IHPMMRENG

7779
IFQLSRPAW

7780
IKETWRVYF

7781
NTNPGRYGC

7782
DLVDLRSWM

7783
CNEVCRNYL

7784
GFTPDRCRN

7785
DVPIHRVQL

7786
TMEAHRFGS

7787
GSATNRCGN

7788
NSAECREFQ

7789
VADTYRMWH

7790
DYYLNRASH

7791
NSKACRGCT

7792
LHNSCRTLE

7793
TVGSKRDLY

7794
FNWEVRAYI

7795
TNIKERTGS

7796
TTYHHRNTL

7797
MWTVARMFP

7798
EQFADRAPR

7799
LNELPRAEY

7800
RAEWFRCQH

7801
PANHARNPK

7802
SWQQHRKMS

7803
LPDHWRKFD

7804
EDWLRRNIW

7805
DAAMTRTVP

7806
LLWMGRRRR

7807
ATCWVRMGQ

7808
WVKQKDVLC

7809
MSLDTSVPF

7810
TSHKMMVFS

7811
NMKPKDVFR

7812
LLAKYMVHT

7813
QDCERQVFA

7814
PTMLPHVKH

7815
QGFSTDVKP

7816
EPLLPAVGN

7817
IAKDDAVRL

7818
MENLKGVYD

7819
QTNQVVVVE

7820
GTTGHAVNE

7821
AFLQTPVFS

7822
ETAFQQVQV

7823
TWHKQLVGV

7824
TKYHYHVKV

7825
RNDWQQVFS

7826
QVGVTVVQR

7827
DSHDMFVYK

7828
MYLSFPVHE

7829
PWEIEWVYW

7830
HFADNQVVQ

7831
MHQFLQVAQ

7832
NAGMKGVDV

7833
STQKVPVLA

7834
GWDMLSVAK

7835
RNLPTSVEA

7836
GMSDCEVSF

7837
GKLTVQVNH

7838
LILQRNVKD

7839
NAKEQVVNN

7840
GDHKCHVEW

7841
GFKYCEVDH

7842
MFDMMSVLK

7843
IVGINHVAW

7844
MMNQCQVWN

7845
QGQHVMVHS

7846
NNCNMNVMK

7847
EKHFMQVDR

7848
PIAQDQVWS

7849
ECHKCEVCV

7850
PHAYMQVWR

7851
MQVNLQVIE

7852
TPQPMNVAT

7853
AWMCRNVYC

7854
CGYWLVVPW

7855
QHTQQLVQN

7856
CPEHNQVGI

7857
HDEKQKVYG

7858
EPFLKEVNV

7859
AVTQLSVER

7860
QHKTKCVDN

7861
IHCNVLVNY

7862
MDGHFSVGC

7863
YHQLVQVNE

7864
GQPVWCVCM

7865
MIKPLLVVG

7866
GLGWWNVHT

7867
TMAQWTVHN

7868
EPMPMPVWN

7869
IPTFDGVEP

7870
ELPDTEVPW

7871
QAMLHHVWV

7872
FTEWKPVFF

7873
QAVTLEVVC

7874
HECDTVVVN

7875
CVNLMAVNN

7876
QHQAFFVFT

7877
ATEKQMVDH

7878
TGMHPFVRG

7879
QVMQEKVRC

7880
DFPHMHVGT

7881
NAGVVKVWP

7882
EITQHFVKH

7883
VADTWQVHC

7884
SIYQKWVTN

7885
PIKGVAVQE

7886
IFSHPSVSG

7887
ECSVHPVNC

7888
DKMSLQVQI

7889
EGEWLIVPV

7890
TQNACKVRT

7891
TGSLTDVIW

7892
GSAMLMVIR

7893
TAGEWIVQH

7894
TGMSRYVQD

7895
FAGFTPVWM

7896
NMWYSFVMI

7897
ETEHGSVYT

7898
PSILPVVMF

7899
PEYVKWVAD

7900
TIHPAWVIL

7901
QWFTKGVDE

7902
CYAEEPVWP

7903
QIMSAKVFE

7904
MTTFCDVEA

7905
FTGTPPVDM

7906
ESWMCPVDY

7907
DCTYCCVKA

7908
AVVEHAVNR

7909
MDYHCQVED

7910
QPGVVPVPC

7911
MPYPLNVKE

7912
NVYATPVVC

7913
NCHIDHVQD

7914
VDMGVHVGS

7915
VIKVNTVNP

7916
DFKALDVQC

7917
MSAQRYVQT

7918
FCKCFMKHC

7919
VIPEFMNHA

7920
AVIHEKPHG

7921
MLFDFEEHN

7922
RCVQAHFHK

7923
NPTDLFQHE

7924
GEMTRCFHN

7925
IAVASHAHG

7926
WKWYGLPHH

7927
SVTYHEWHP

7928
TDHPWVLHG

7929
AKEHMPHHM

7930
GIHQLIHHC

7931
EACMPWAHH

7932
DVMQYQNHM

7933
DVDFLKMHC

7934
MYAGWLNHG

7935
TNGVQIMHV

7936
DGLSYEAHD

7937
EHCYYKEHE

7938
HPSWPTMHK

7939
ISCIMKKHL

7940
IWGTRSAHR

7941
NQCWCPKHY

7942
SFKWCPGHE

7943
ADCFRPFHI

7944
IQDCSTLHW

7945
LQHFDLIHL

7946
PQQCGGFHR

7947
TWGLHHYHT

7948
VNPPDMIHT

7949
DKANKMMHG

7950
ETMSEGIHC

7951
FYPYTNCHH

7952
PAAPLILHY

7953
QCLNMMMHG

7954
QVGTGMYHS

7955
WDHYLPEHV

7956
TVQDNDEHA

7957
TKQFFTAHC

7958
NFSVCPRHL

7959
ITTDATGHF

7960
STFHFQSHD

7961
HMDHCQCHE

7962
QYWWPATHD

7963
LGEPYKAHD

7964
ISNTMVEHG

7965
VTAFTACHY

7966
EWLSLYCHW

7967
PIHTYKRHP

7968
GYNYYWQHA

7969
ATVHMPKHE

7970
MFSAFSMHD

7971
TCTNSEPHP

7972
TIDWCEPHC

7973
LMHGVMDHV

7974
YITCQYSHH

7975
LATTTSYHC

7976
GTCLFKTHK

7977
DSAKQWHHQ

7978
SYAAFLHHG

7979
PNCDPAHHD

7980
FAPEFDNHC

7981
QISMKGEHG

7982
QLTTNDAHG

7983
CMAQMVYHH

7984
CIDVLFKHL

7985
EAGKEGAHF

7986
QYCNTDNHI

7987
YLESKTIHA

7988
PHQYGDEHQ

7989
WPTMLSAHM

7990
DQFCPFDHY

7991
LPTQVYGHN

7992
DTHPMSNHR

7993
MGVITSIHQ

7994
NAMTCSMHN

7995
TIQHAFKHN

7996
NVFPAETHV

7997
TLEPAYRHA

7998
YEICKHGHF

7999
HSLVQQLHP

8000
VPVATMIHH

8001
RVDGADRHY

8002
CHILDGLHD

8003
MLGVAPGHL

8004
ANLISQGHC

8005
GSYIVQNHC

8006
WPEAFHLHQ

8007
CDFMFTRHI

8008
HVWNDVFHM

8009
ADYSMMWHM

8010
WNWLKVMHE

8011
PVTTKNCHE

8012
FCHEVFSHT

8013
IHKMNFDHL

8014
WAIFGDIHH

8015
QSGTCHYHD

8016
WVYSHEKHY

8017
DVKYATLHD

8018
GTEMMGDHA

8019
QTKVHQQHT

8020
ASSSQYPHN

8021
TSLCAKPHF

8022
GYTQQDEHF

8023
DHCHCIAHP

8024
IPACGIMHE

8025
TCNVSWKHE

8026
TKGMWQQHS

8027
SAGPFFDHY

8028
HTVNAESHM

8029
DDGSNTMHG

8030
GVAQYLHHC

8031
EFADYNFHF

8032
NTCEGWSHF

8033
FPKCAIWHY

8034
AFGLTTGHT

8035
STGSFSIHC

8036
HHEFINSHA

8037
QVVIDPAHY

8038
CAGMAWSHS

8039
TVQWNFAHM

8040
RCVQDHFHK

8041
LLWMGQRHR

8042
QQTWEQMQR

8043
CAMPHYQVR

8044
LGKVQTHIR

8045
CSVWMQRDR

8046
VFPKDEQFR

8047
EHITEQDLR

8048
DKTVQTSAR

8049
GGMWQCEVR

8050
LEKNSSQMR

8051
CDAEAEFRR

8052
MGGSCKQER

8053
TCVIGKGRR

8054
VAFFWQRCR

8055
FAESMWCDR

8056
FAQQDKQDR

8057
DANWTVRRR

8058
MCNGGIPAR

8059
EGATAWQDR

8060
SHIQLNAWR

8061
IPAEWDSWR

8062
DAHNDALAR

8063
QPQSRNGAR

8064
NWNHCHQWR

8065
CWHFFGADR

8066
CAMDNTMCR

8067
TFVYGARDR

8068
QLDTYMEGR

8069
AATTHHSAR

8070
YTNHYYNAR

8071
VGIWFIPCR

8072
WSEDMSQYR

8073
MCSQTISMR

8074
RGLAQDCCR

8075
FCNKLTSVR

8076
PTNYGWMWR

8077
HTLQWEAVR

8078
QCAHFYQTR

8079
EWEGVSQFR

8080
QAVIAVDGR

8081
GITEQPLAR

8082
GIVMLKCGR

8083
TLLLVGKDR

8084
GLWLNVAPR

8085
QTEMKKCAR

8086
GHIDLEIMR

8087
RMTPNNCQR

8088
TCGPKLTQR

8089
HIGWMGWLR

8090
QETHWAMMR

8091
PYHNPWECR

8092
YDLWGWSTR

8093
QHYNSMIDR

8094
TYMTNLGDR

8095
GKLTFGTER

8096
GVHDPLHDR

8097
TTYLSMRDR

8098
AHMNYCLSR

8099
NYCKYNKMR

8100
GVGPSHGER

8101
AKISATWDR

8102
INMEYLTDR

8103
YTGHTNSGR

8104
MHCLHQTMR

8105
YWCPSQWWR

8106
MLAFDPMGR

8107
IIAVSMRGR

8108
VWEDNPPFR

8109
CTLMHSHTR

8110
LLWMEQRRR

8111
LLWMGQQRR

8112
LLWMGQRCR

8113
LLWMGQRRR

8114
LLWMGQWRR

8115
MDPDMPHCR

8116
MDPDSPHCR

8117
MDPDVPHCR

Example 6
AAV5 Variants with Tissue Tropism in Spleen

This example describes engineered AAV5 variants with tissue tropism in spleen that were discovered using the methods and systems described in EXAMPLE 1-EXAMPLE 2.

CNS, liver, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord tissues were analyzed to identify amino acid residues in the AAV5 VP1 581-589 region that drive spleen tropism. The results are shown in FIG. 18F which shows the positional frequency of each amino acid in the 581 to 589 region of AAV5 VP1 in spleen tissue as compared to all other analyzed tissues, wherein the capsid polypeptide has the sequence of SEQ ID NO: 2. The position (“Xaan”, n=1-9) is denoted on the x-axis and amino acid residues are denoted on the y-axis. The frequency of a particular amino acid residue in a particular position is indicated in the heatmap as intensity. Darker squares indicate a higher frequency of a particular amino acid in a particular position and lighter squares indicate a lower frequency of a particular amino acid in a particular position.

The frequency of a given amino acid residue occurring at a specified position corresponding to position 581 to position 589 of SEQ ID NO: 1 (generalized in SEQ ID NO: 2) in variants identified in spleen over the frequency of that given amino acid residue occurring at the specified position in variants identified in all other harvested tissues was analyzed to identify a set of sequence rules for capsids that preferentially target spleen tissue. With reference to TABLE 11 below, and SEQ ID NO: 2 (AAV5 VP1), the following amino acids can, thus, be independently mutated, in any combination, at any one or more positions Xaa1-Xaa9, to provide a VP1 capsid with enhanced spleen tropism as compared wildtype AAV5 VP1 capsid (SEQ ID NO: 1), where spleen tropism here refers to properties that are preferred for spleen transduction over properties that are preferred for transduction of all other harvested tissues.

TABLE 11

Spleen Tropism Rules

Xaa1 is selected from C, F, H, I, L, P, W, or Y

Xaa1 is selected from C, F, P, W, or Y

Xaa1 is selected from P, W, or Y

Xaa1 is P

Xaa1 is selected from D, E, L, N, P, R, or W

Xaa2 is selected from D, E, or W

Xaa2 is D

Xaa3 is selected from C, D, E, P, or W

Xaa3 is selected from D, P, or W

Xaa3 is P

Xaa4 is selected from C, F, G, H, R, W or Y

Xaa4 is selected from C, H, or W

Xaa4 is C

Xaa5 is selected from A, D, E, G, P, R, or W

Xaa5 is selected from D, E, G, or P

Xaa5 is D

Xaa6 is selected from A, C, D, E, K, R, W

Xaa6 is selected from C, K, or R

Xaa6 is K

Xaa7 is selected from F, L, P, R, W, Y

Xaa7 is selected from L, P, or W

Xaa7 is P

Xaa8 is selected from E, I, K, L, P, R, or T

Xaa8 is selected from P, R, or K

Xaa8 is K

Xaa9 is selected from C, H, M, T, V, or W

Xaa9 is selected from C, T, or V

Xaa9 is V

TABLE 12 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that were found in spleen tissue and comport to one or more of the rules provided in TABLE 11. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 37438-SEQ ID NO: 38437, as disclosed in TABLE 12. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 12

Sequences of the 581 to 589 Region

in AAV5 VP1 Capsid Polypeptide

that Drive Spleen Tissue Tropism

SEQ

ID
581-589

NO
Sequence

37438
PANFGQAYP

37439
PCVQDLRA

37440
PCYGQTDWN

37441
PDAWNEMWH

37442
PDPRPATND

37443
PDTGHEWWW

37444
PDWREATVV

37445
PEDPTVTNH

37446
PEKHSCSTT

37447
PFILKQRNQ

37448
PGMTTTPSF

37449
PGSLECAAL

37450
PHVANMGPG

37451
PHVNYDTLC

37452
PIWGNCAKT

37453
PKGSQYGEQ

37454
PKGVLLGGA

37455
PKLQYHSES

37456
PLARIEPLW

37457
PLECKMWYN

37458
PMPFCKHQF

37459
PMQQKLWME

37460
PMRWNKFVP

37461
PMVKLEETS

37462
PNCKHSIAV

37463
PNTCTKDGL

37464
PPHRFRHIP

37465
PPWPPETAE

37466
PQASKAFSK

37467
PQQQYLSVC

37468
PQTAYMQAY

37469
PRYFVEACE

37470
PSHWFCGGA

37471
PTDFIRVNE

37472
PTRATMFID

37473
PVAWRENSL

37474
PVTRFECCK

37475
PYCICTQPY

37476
PYHDNSAPQ

37477
PYHLNQDNN

37478
PWAIEYSLV

37479
PYIMEFHPF

37480
PKEHFPTCM

37481
PAAAMFLPS

37482
PAAHKEWEW

37483
PAAHNACHC

37484
PAALDLYNG

37485
PACLGRTDA

37486
PACNCQIAS

37487
PACRKGYWN

37488
PADEAFVEC

37489
PADKSQLGA

37490
PADLNLSIP

37491
PADNLHCHG

37492
PADSAKQRK

37493
PADYGWCVG

37494
PADYRAYGD

37495
PAECKYIYA

37496
PAEFPFMIH

37497
PAEFRCTPF

37498
PAEPGQSMG

37499
PAEVTNFGV

37500
PAEWCLQHV

37501
PAFGCMLNH

37502
PAGCHGKTH

37503
PAGCVCASS

37504
PAGMAMPGT

37505
PAGNAAWTT

37506
PAGWNECAC

37507
PAHKVDLCF

37508
PAHQCTVHD

37509
PAHYVPGIE

37510
PAISNKFAG

37511
PAKLYITVR

37512
PAKQAADYG

37513
PAKYLPWQY

37514
PALRETPWT

37515
PALWRYYKQ

37516
PAMMMPTKQ

37517
PAMRYDSLY

37518
PANNCRIHT

37519
PAPCWWNVT

37520
PAPDNSDTC

37521
PAPHAWDVH

37522
PAPIWQVLD

37523
PAPKHSYQN

37524
PAPPLAQSN

37525
PAQNAPMCG

37526
PAQQLTCIK

37527
PARCAQFVA

37528
PAREDGNGW

37529
PARKAYCLP

37530
PARLISDYE

37531
PARSLWQTY

37532
PARSTYGAA

37533
PASCMWFSH

37534
PASLGIPKM

37535
PATEVQQFM

37536
PATNQCKRI

37537
PATQERGEK

37538
PAVADDMYK

37539
PAVHMQLDP

37540
PAVNANATN

37541
PAVNVPMST

37542
PAVQCSWHG

37543
PAVQGLPKM

37544
PAVSACPYW

37545
PAYGSPPWA

37546
PCADTMSVW

37547
PCAFETTQH

37548
PCAGKDICN

37549
PCAPYARSK

37550
PCAQWYCDC

37551
PCCEDVSNI

37552
PCCEGTMDN

37553
PCCQICNTW

37554
PCCYTQRPT

37555
PCDRTCPDM

37556
PCEFKSTET

37557
PCEGHEKPQ

37558
PCERALLQH

37559
PCEVQNAQT

37560
PCFAVMAGC

37561
PCFCHYTNC

37562
PCFGPSYAD

37563
PCFVKDSTA

37564
PCGGLTHPH

37565
PCGLMDDYP

37566
PCGQWKIQN

37567
PCHAFEPDH

37568
PCHMMVIAN

37569
PCHQQELDK

37570
PCHQYWIED

37571
PCHRYSKFM

37572
PCKCYPRKC

37573
PCLHMQYNP

37574
PCLHTCHRG

37575
PCLWPMGCY

37576
PCMHVQQWE

37577
PCMKKHSST

37578
PCMQNFDDG

37579
PCNKWNPKQ

37580
PCNLMRDQN

37581
PCNLVWNCE

37582
PCNTDPSIH

37583
PCPDWYVMS

37584
PCPHDRLNA

37585
PCPIIWESG

37586
PCPLSQYQT

37587
PCPSEDWFW

37588
PCQSIKCCW

37589
PCQSSWLMI

37590
PCRVGFSYE

37591
PCSESWSNT

37592
PCSHYCART

37593
PCSWFYHGG

37594
PCSYTIFDT

37595
PCTCCFNRQ

37596
PCTCVWYRC

37597
PCTGQLFMA

37598
PCTQKQKPF

37599
PCTWAHSHD

37600
PCVHGTQTW

37601
PCVKKDCEE

37602
PCVNKVGTM

37603
PCVWQWHCT

37604
PCWLKPTSW

37605
PCWRRDIHP

37606
PCWVNDHPM

37607
PCWWKWMCG

37608
PCYLCMLIL

37609
PCYWAIAGV

37610
PCYYFGMPH

37611
PDACSPFGA

37612
PDADQFVPQ

37613
PDAGWQNSP

37614
PDAMKDCVT

37615
PDANIEFLC

37616
PDAPWEGWH

37617
PDCCDMQCF

37618
PDCTERFME

37619
PDDEQVRVN

37620
PDDEVCSKV

37621
PDDGHKHPF

37622
PDDMWQYEA

37623
PDEEYAQYW

37624
PDEGFLYDN

37625
PDELHLEDG

37626
PDEPRSKCQ

37627
PDEQHMGYW

37628
PDFQRENFE

37629
PDGFGVSMP

37630
PDGRRYVCS

37631
PDHAMEYQC

37632
PDHEPNGDA

37633
PDHGIQFGW

37634
PDHIRMVTP

37635
PDHKLIVYD

37636
PDHMGLPYH

37637
PDHVHCIHA

37638
PDHVREHCM

37639
PDIHGPQSE

37640
PDKACSHHP

37641
PDKDNHVAY

37642
PDKSEAQRQ

37643
PDKVSDYQE

37644
PDLQPRDTT

37645
PDLVCDYCQ

37646
PDLVPIVGN

37647
PDLWSPFWY

37648
PDMFKERSQ

37649
PDMIVDPDN

37650
PDMLTDSTC

37651
PDMPHDRHC

37652
PDMQWNGEW

37653
PDMTCTINI

37654
PDNIFWGEA

37655
PDNLAPYVP

37656
PDNMITMQH

37657
PDNNFLSNQ

37658
PDPGHRMYA

37659
PDPHCEDRK

37660
PDPHIVWCG

37661
PDPLARRFE

37562
PDQHTHLAM

37663
PDQRGQTGQ

37664
PDRPCEDVN

37665
PDSCLHRTA

37666
PDSVPVQHN

37667
PDTAKQDMS

37658
PDTCWNWPR

37669
PDTSMECDH

37670
PDTTMKNTL

37671
PDWATLMGN

37672
PDWAVMKAP

37673
PDWFEENTV

37574
PDWGSHYCN

37675
PDWHASYGN

37676
PDWNMRMYP

37677
PDWRYGWYS

37678
PDWSGKYDI

37679
PDWSPPIQN

37680
PDWWAIEMG

37681
PDYHIGCNH

37682
PDYMDCAPM

37683
PDYWKSAFA

37684
PEAKWKEHD

37685
PEAMIGQEN

37585
PEANEVQTR

37687
PEARSWWAQ

37688
PEAVWCAFA

37689
PECAESPIT

37690
PECFLDSNV

37691
PECFSPENM

37692
PECLINYTC

37693
PECQSIDSV

37694
PECSNWRNH

37695
PECVIEYED

37696
PECVRYSKC

37697
PEDCCRQDS

37698
PEDMTLDGS

37699
PEDNNIRRN

37700
PEDSGYHWH

37701
PEEANQQVT

37702
PEEAPKQMF

37703
PEEETMYAC

37704
PEEHEVEQT

37705
PEEINDHCY

37706
PEEYEHMNC

37707
PEFFHWPHS

37708
PEFMSKGDS

37709
PEFVQYGSC

37710
PEGCTYPQL

37711
PEGHEEHPS

37712
PEGLVIHMT

37713
PEHGIHDGE

37714
PEHRAANVG

37715
PEHSNLFSY

37716
PEHTVIWES

37717
PEHVETHGQ

37718
PEIFGQMSW

37719
PEIIWQIAT

37720
PEITCKIAD

37721
PEIVNPARF

37722
PEKVLLGMV

37723
PELYTMFFT

37724
PEMALTKGL

37725
PEMCDPADQ

37726
PEMDKFKNM

37727
PEMGEATKV

37728
PEMPRHICR

37729
PEMTYEPMS

37730
PEMVEEMNV

37731
PEMWMILEG

37732
PENHKMGPW

37733
PENTFCSTY

37734
PENVGASGA

37735
PENWRPWVT

37736
PEPKLIGLF

37737
PEPLGQGTW

37738
PEPQTQDDG

37739
PEPRNVMKS

37740
PEQCSERIV

37741
PEQNDVNGM

37742
PEQRRRYDG

37743
PEQWSWGVC

37744
PEQWWRYQA

37745
PEQYKYYQC

37746
PEQYNAQAE

37747
PERINHLGC

37748
PERLFTCFV

37749
PERNKVWVA

37750
PERYNQNQL

37751
PESCGAAAP

37752
PESDRIYRP

37753
PESIKTIQR

37754
PESNKWEDH

37755
PESNWRCPE

37756
PETAAAKAM

37757
PETFDTYPH

37758
PETHSESCN

37759
PETWNDRCH

37760
PETYLGIPD

37761
PEVIQNESM

37762
PEVPSTNHA

37763
PEVTSRDIM

37764
PEVWGGRTQ

37765
PEWGHIGNH

37766
PEWIDGYVM

37767
PEWWGQNIH

37768
PEYHWDHYY

37769
PEYSTSFEQ

37770
PEYTEFEHQ

37771
PFAKPNIEE

37772
PFCCEMHIP

37773
PFCLQDVCH

37774
PFDTCFICT

37775
PFDVDGPIY

37776
PFEAQINFD

37777
PFECHKSSS

37778
PFESHDQYT

37779
PFFCSGVKC

37780
PFFEPDDEM

37781
PFGLVEINT

37782
PFGRMIVDD

37783
PFGWESQQP

37784
PFHEDCPTE

37785
PFHEPWAYA

37786
PFHGVILEN

37787
PFHLPRFPS

37788
PFHVHGEHY

37789
PFHYNKTLC

37790
PFKAHKPSL

37791
PFKHENLRH

37792
PFKPCPIEI

37793
PFKSDFWAI

37794
PFLAGDRPV

37795
PFLMEENYR

37796
PFLYVIDRR

37797
PFMAHGMYQ

37798
PFMISNALQ

37799
PFMQCCYSW

37800
PFNFMGMVQ

37801
PFNPDWGRF

37802
PFNRMTDYY

37803
PFNSDSYIR

37804
PFNYMFLIM

37805
PFPAMKIKL

37806
PFPWEWLRL

37807
PFQQFLERC

37808
PFQWVWREQ

37809
PFRECKNCT

37810
PFSFSQPGS

37811
PFSKGAYLA

37812
PFSQWIHSA

37813
PFTHPICWH

37814
PFTHQPAGM

37815
PFVGLLEHS

37816
PFVKPLDIA

37817
PFVQYKWDR

37818
PFWCEAISY

37819
PFWIHRMIE

37820
PFWLFHAKC

37821
PFWVRITPF

37822
PFYDQKLSF

37823
PFYKKYTAF

37824
PFYPLWENF

37825
PFYQHLSGY

37826
PFYTSDGCP

37827
PFYYFTCCQ

37828
PGADTHGPQ

37829
PGAGLGPPS

37830
PGANYWGCM

37831
PGAYKNQQH

37832
PGCLEIFHM

37833
PGCPNSNPH

37834
PGDCEEAII

37835
PGDCHEAWC

37836
PGDQYREWC

37837
PGDRGPIGR

37838
PGDWQTTPT

37839
PGDYRQTLP

37840
PGEAAFEKQ

37841
PGEAVCHRW

37842
PGEFKGLVS

37843
PGFGMNSIR

37844
PGGAYVWTR

37845
PGGCMWARD

37846
PGGGFCLNG

37847
PGGINNMTV

37848
PGGWLEHPA

37849
PGGWQVNPS

37850
PGHCYRMGW

37851
PGHFWWQHA

37852
PGHKFRSMD

37853
PGHWNLYHQ

37854
PGHYAIMKR

37855
PGIHSVMNQ

37856
PGIRAQCGE

37857
PGKLQTMGP

37858
PGKMINPQM

37859
PGKNLKWTD

37860
PGKVAWSQG

37861
PGLICSIDW

37862
PGLPYRNHH

37863
PGMEKNGDL

37864
PGNCIPAMG

37865
PGNEHMQMT

37866
PGNGRLYWT

37867
PGNRFMTYY

37868
PGNWAKPGG

37869
PGPHDMGSL

37870
PGPSDNTPN

37871
PGQGSDTEE

37872
PGQLTSRHF

37873
PGQSDSDMV

37874
PGQTEERVW

37875
PGRIDFWKP

37876
PGRPMWMTN

37877
PGRYCKTHS

37878
PGSQALQWA

37879
PGSSNKPYA

37880
PGSSSACLC

37881
PGSTYQAML

37882
PGSWYMHLP

37883
PGTFWHEQY

37884
PGTGVADRN

37885
PGTKACMKH

37886
PGTSWIPWM

37887
PGVTNCDVG

37888
PGVTVWGYH

37889
PGVWLSYNC

37890
PGWGKSFLT

37891
PGWHNFTAI

37892
PGWLEAWSP

37893
PGWPAPTGH

37894
PGYKTPWAK

37895
PHAEGLFCT

37896
PHAHPMFMT

37897
PHASSPMQP

37898
PHCFPAYCC

37899
PHCKICRWN

37900
PHCLADYFM

37901
PHCPMIRQD

37902
PHCREGEVQ

37903
PHCWWKWSG

37904
PHDMSMRFY

37905
PHDINNAKY

37906
PHEEGNKWG

37907
PHEGHDRKQ

37908
PHEHKEGVC

37909
PHENEAIHW

37910
PHEYIVVKS

37911
PHFKHHKTQ

37912
PHFKMEPHD

37913
PHFSRQNGT

37914
PHGGQRISN

37915
PHGLNEAAI

37916
PHHAGTQRS

37917
PHHDVSIPC

37918
PHHFTVYPF

37919
PHHIMWHHP

37920
PHHLMSCIG

37921
PHIEVAIGH

37922
PHIIPLEGL

37923
PHIKDWPRA

37924
PHISCAHAW

37925
PHKFIFMTS

37926
PHKPGTTKD

37927
PHKWIKFAG

37928
PHKWWNVHW

37929
PHMCYLLWR

37930
PHMWRDNHN

37931
PHMYKWTRM

37932
PHNCELNSA

37933
PHNQREHCA

37934
PHNYDWGCS

37935
PHPENMSKQ

37936
PHPFGHREA

37937
PHPHKQHNW

37938
PHPQCTHMM

37939
PHPRMGIQQ

37940
PHQCKQLTS

37941
PHRANSTQY

37942
PHRFPDRKM

37943
PHSCCCMKC

37944
PHSCLFMWA

37945
PHSEQIKCG

37946
PHSFNTIMS

37947
PHSIVATRD

37948
PHSNQAEFN

37949
PHSWAYHTS

37950
PHTMRVCLP

37951
PHTPQGTKA

37952
PHTREGLAQ

37953
PHTWIKQEG

37954
PHVMGGEWC

37955
PHVPHEIGI

37956
PHVRCPMDL

37957
PHWDKGYDL

37958
PHWFFCNMN

37959
PHWMPWNQV

37960
PHWQYETGS

37961
PHWRPPLGF

37962
PHYDSLAHS

37963
PHYLQVCCA

37964
PIAEDEVAV

37965
PIANDMHTY

37966
PIANKHPSI

37967
PIAPSGFVG

37968
PIAWVEQFP

37969
PICFEDCCH

37970
PICMIGRWT

37971
PIDEAINDS

37972
PIDPRQSGE

37973
PIDPVYHLI

37974
PIEMHVACS

37975
PIESPIHHS

37976
PIESTPVLV

37977
PIFMTEQNL

37978
PIGPSSTSC

37979
PIHHKQTYM

37980
PIHRESPRI

37981
PIIPQAQLF

37982
PIITNCAHS

37983
PIIWELGWA

37984
FDAGEEAYF

37985
FDLDQQVHT

37986
FDPQDDWCY

37987
FDRISVLGV

37988
FDTDHRWCW

37989
GDDLYVHGM

37990
GDECFCKLI

37991
GDFCFYKMT

37992
GDFHPDPLC

37993
GDGKSTKVT

37994
GDVHHLHSK

37995
GDVPTWPKC

37996
HDTSQAHNI

37997
IDEDIFFAA

37998
IDIHMSANI

37999
IDKNWKGLW

38000
IDQCAKFHR

38001
IDTFNCDTY

38002
IDWFAGACG

38003
KDAAYVTPD

38004
KDHCYTCID

38005
KDPRITWYV

38006
KDQFWGCLP

38007
LDHPMHMMC

38008
MDAHGRNHG

38009
MDHPRVEMQ

38010
MDLYYTLAN

38011
MDNFDRPAA

38012
MDNRELVVK

38013
NDQCQQHTC

38014
NDEKAAIDI

38015
NDGSSVHMN

38016
NDKHTTMEN

38017
NDPYNKQSQ

38018
QDHTNAWHQ

38019
QDQDLNAWA

38020
QDRDQSYHR

38021
RDAANLEFP

38022
RDDLMVKIV

38023
RDIDHKNDE

38024
SDANDRADW

38025
SDCAMTIHV

38026
SDCSDFTQF

38027
SDDHAMQMN

38028
SDDVKLPGL

38029
SDHTCCWTF

38030
SDMDPCSDK

38031
SDMWMERSC

38032
SDNCMEADN

38033
SDRNALMPN

38034
SDTKWNVCY

38035
TDDFGTPWD

38036
TDEHQGVFQ

38037
TDHDHHQEF

38038
TDKPMMGGH

38039
TDPTRSTVP

38040
TDRVGMPPY

38041
VDGNEEEKP

38042
VDHDVDTMQ

38043
VDNHGFNDW

38044
VDWGVDPWH

38045
VDYCKEFVW

38046
WDKAQDVRM

38047
WDSHCQKIQ

38048
WDSPFNYSF

38049
WDSRYKCPI

38050
YDAEHQQLA

38051
YDDCTTNQG

38052
YDFPIMTCE

38053
YDPDKMKPG

38054
CDWMANVSV

38055
LDTIQINEV

38056
EQPHADQKI

38057
FAPSGKFRD

38058
FVPFHWWSH

38059
GCPPDPWHN

38060
GMPVMAFQT

38061
GPPECHMML

38062
GSPQWLEYT

38063
HSPLEFSSP

38064
IAPLQCPVS

38065
IFPMADCRV

38066
IQPYYWNDT

38067
KTPIKCEQH

38068
KVPPYRTCP

38069
KWPGMMGEV

38070
LEPNLMNTE

38071
LMPHRTWKC

38072
LNPHFVFGR

38073
LPPTCANGT

38074
LVPMAHRAC

38075
MAPQPMSWG

38076
MCPMFFWCE

38077
MLPYVCDPG

38078
MPPPNHLDV

38079
MTPSWIVED

38080
NCPAMPLKI

38081
NRPIRAWKR

38082
NTPERPHRK

38083
NTPITIQPL

38084
NYPSAHQCG

38085
QFPEIHRGE

38086
QMPTTTDRC

38087
QQPMIAEHV

38088
RNPSIQYWS

38089
RVPYVETYD

38090
SCPEMDLHR

38091
SNPIMDSDC

38092
SPPISYPRM

38093
SQPVMELSE

38094
SVPFTNMQS

38095
SVPMWWCAP

38096
TRPCICCDV

38097
TSPQNDHYS

38098
TTPFEKCCA

38099
TVPLQFQDS

38100
TVPRYSRNQ

38101
TYPRMITPA

38102
VGPCVRSDW

38103
VGPPYDTQD

38104
VLPFGAVGT

38105
VMPANWFFS

38106
VQPCMQCWC

38107
VRPMHMNCT

38108
VTPRFQWLT

38109
VYPTHNEVV

38110
WNPVQWTHS

38111
WTPVPSSSN

38112
WWPMRKWVS

38113
YLPYQSICG

38114
YNPQQRSLS

38115
IVPQQKIDE

38116
HGPFCDHAY

38117
KGPIFHSAY

38118
ENKCFASTT

38119
EQLCGKKKQ

38120
ERTCMPSAY

38121
ESDCFMWLG

38122
EYECLDCNI

38123
FCYCSHFPW

38124
FHMCSCTRY

38125
FSSCFNAVE

38126
FWRCDPNKV

38127
FYNCCDNPL

38128
GCMCEGAEF

38129
GGFCPCDQK

38130
GGSCSWDKP

38131
GIICYDATN

38132
HRGCCESGR

38133
HSFCVEGIP

38134
IHFCMCCDF

38135
IHRCIAEAA

38136
IHTCPKNTQ

38137
ILDCHKHYS

38138
ITECHPRKT

38139
ITYCNVVYW

38140
IVVCMHCEQ

38141
KGMCLSQGD

38142
KGVCKGVLP

38143
KYECPFYYV

38144
LHDCQMWGD

38145
LHSCDTPWT

38146
LKECCFEDA

38147
LNECYYNRP

38148
LPDCTDYRK

38149
LPRCCTINY

38150
LVGCSYCQS

38151
MAGCPPHDS

38152
MASCIPQTS

38153
MFACTQFFG

38154
MGDCHRSPD

38155
MMRCNVDFE

38156
MMSCIAAVQ

38157
MVSCRSAAC

38158
NASCHWHFD

38159
NGLCVESTA

38160
NPACNKMPT

38161
NQACNGWHK

38162
NSHCTWRKF

38163
NVECLPMLP

38164
QAMCCSYPR

38165
QAMCVHWHM

38166
QCYCAKYWS

38167
QGDCYEMWE

38168
QMTCPSDLW

38169
QQICPVGEF

38170
QYACHQIGA

38171
RGACDEFET

38172
RLMCYFRTW

38173
RPACTVPGK

38174
SGDCRDWHD

38175
SGTCYHYAL

38176
STGCGLMWG

38177
SVYCEEFYT

38178
TIMCVASLA

38179
TLYCYDDYH

38180
TRICCKNRQ

38181
VKQCTKFDE

38182
VWCCSVNPG

38183
WRNCMQDSK

38184
WSICHKDSY

38185
QNWCGKNLE

38186
SQDCGRPES

38187
KVGCFVDVW

38188
FCKCFMKHC

38189
ESQRDDGWL

38190
ESRADIEVA

38191
ETCIDMQGK

38192
ETHTDYQGP

38193
ETNPDWEGR

38194
FCTSDQKAL

38195
FGDHDNMPE

38196
FRHHDVNNH

38197
GEYYDEVNH

38198
GMEPDTQLY

38199
GMVPDLLPH

38200
GVNMDMEVS

38201
HGELDCASC

38202
HHFTDVDAH

38203
HHHADFHLA

38204
HMHRDMHEE

38205
HPNRDSAHM

38206
HRDGDILIE

38207
HYTSDWFQA

38208
IAKEDICQP

38209
ICVNDNFHN

38210
ISAPDNQLS

38211
IVTADNTME

38212
KANWDVNHS

38213
KENGDSITL

38214
KVDPDCHMF

38215
KVLQDQFTI

38216
KVVIDRKWD

38217
KYEPDGFIP

38218
LVGLDLSCH

38219
MHYSDGFVS

38220
MIEYDVGYK

38221
MLKTDCPLH

38222
MPCMDRRRP

38223
MRCADMLRY

38224
MTESDDRCV

38225
NCGLDITMR

38226
NIQADMSQQ

38227
NTVMDRMNF

38228
QCDSDTQSR

38229
QTRKDGESH

38230
QTRWDLIGL

38231
QVCIDWSGG

38232
QVNEDDAAK

38233
REYMDDEKQ

38234
RHHIDIPTY

38235
RVLTDERMD

38236
SAREDDNPC

38237
SCRIDWTDW

38238
SFNYDMVSR

38239
SGRGDVNGA

38240
SMQVDVLQR

38241
SNQWDWHIF

38242
SVMLDEWIN

38243
SWCIDCEAD

38244
TIAVDGHHS

38245
TILPDSAEF

38246
TIVYDNWSR

38247
TTTADKPAS

38248
TTVLDHGLQ

38249
TWKEDWALL

38250
TYKRDIMKT

38251
VILKDRVNW

38252
VKKSDAKQC

38253
VQDGDYSPS

38254
VSMEDHCKH

38255
WEKFDRPWR

38256
WPSADCDIP

38257
YMSRDQIKS

38258
YPTRDNLFM

38259
YWNFDDYQQ

38260
YYTGDLTYK

38261
SVKQDCVED

38262
TVREDCCIN

32263
YFAADEQWG

38264
NQIKDVAAS

38265
DARFDQWHH

38266
ALTHDDGRT

38267
FAYFQKDLF

38268
FNADHKCPS

38269
GRDALKNGD

38270
GVATLKWEQ

38271
GVCSKKSGA

38272
HFAEIKYET

38273
HMRMCKATW

38274
HPRSTKWCQ

38275
IEEHLKVCY

38276
IVATPKQYG

38277
KLNHVKRTS

38278
MCNYHKLNK

38279
MGGPYKLMC

38280
MGNIVKPAD

38281
MIDRMKESY

38282
MRTNYKLEM

38283
NFWLAKPFK

38284
NLHGTKFGY

38285
QMMQTKLHD

38286
QMTIPKWGY

38287
QSCWTKCEA

38288
SGILSKDWK

38289
SIDEPKFTV

38290
TEHENKCVG

38291
TGLLRKEQS

38292
TKNEFKTND

38293
TMAQCKYPH

38294
TMFSHKPSD

38295
VMCVPKYKD

38296
VNEEMKFQP

38297
WNGHHKDML

38298
YWFLHKGCN

38299
EVARTKFGG

38300
QMSWGKYHQ

38301
SYYQSKNTD

38302
LWFFMKFAK

38303
GACLWKRHL

38304
THARFKMKS

38305
LMMMCKQQY

38306
ESSWLYPHN

38307
FSITEEPFW

38308
GVVDYWPTF

38309
HHTLNYPNE

38310
HWESFIPRV

38311
IGGWIWPGP

38312
IVSELCPSF

38313
KKQMPFPRD

38314
LGRAPCPCW

38315
LSNVPMPAQ

38316
MAAEAYPCI

38317
MHAEFGPLP

38318
MKFQSGPKS

38319
NQFRAEPIS

38320
NVTNFDPHP

38321
QAAPGPPYE

38322
QPNGWAPPC

38323
QQFQTDPQT

38324
QVEIQEPAI

38325
RMLVRVPDG

38326
RPDMLTPYR

38327
SQCTPFPFN

38328
SRDRRQPKK

38329
SSFHEDPCN

38330
THHGLQPND

38331
TKVVIEPNC

38332
TLYGHDPAH

38333
TMFSFQPVE

38334
VINNRVPPT

38335
VQNYADPTG

38336
YGTIPVPIT

38337
YIKAVTPPF

38338
IREPTEPYC

38339
IFQLSRPAW

38340
ISGWRMPCD

38341
QAEVFAPYH

38342
DAMWKFPST

38343
TVNPCIPDN

38344
KCLQMCPNG

38345
FCNTHCGKP

38346
GLDWVSVKA

38347
HGNAETHKD

38348
HITNPTSKP

38349
HPEWKMSKC

38350
IAKQEHVKS

38351
IVDALHHKW

38352
IWTHLCVKH

38353
LFIQYYHKS

38354
LQNDWDAKL

38355
LVIPRDTKK

38356
LYHETHKKC

38357
MAIPTGVKE

38358
MAVEYDRKG

38359
MMNWWMYKA

38360
MVTQTYFKN

38361
NCMAVWAKN

38362
NLDQWNVKV

38363
NTEGCRVKK

38364
QIGTPSQKA

38365
QIQMTDDKQ

38366
QSAQWNVKA

38367
RTFNRATKT

38368
SLVHFHAKT

38369
TFGFFCTKG

38370
TFSTGLSKK

38371
TKGFMQKKE

38372
TMDEHHMKI

38373
TSVDLQMKA

38374
VEMWVWEKK

38375
VKQFNAHKG

38376
VNLTTSVKF

38377
WCYQCILKF

38378
WSMQLSHKM

38379
YAWTTVCKT

38380
NMVWKCRKS

38381
VMWTWESKY

38382
EVLDSMSEV

38383
EVTHPIGAV

38384
FASRCPRTV

38385
FFFIRMARV

38386
FGVEAWLPV

38387
FHGERHQDV

38388
FVRVQQRAV

38389
HACIHTDEV

38390
HLSEFNMQV

38391
HTFYCTCRV

38392
HVSIATGSV

38393
HYDFQLTQV

38394
ICVNKQQSV

38395
IFHSHATCV

38396
IFSYSQRMV

38397
IRYPYGKMV

38398
KSSEIVKWV

38399
KSYQRTFLV

38400
KYTGGIVVV

38401
LTNWNGMHV

38402
MIETTGQQV

38403
MMGPPQHVV

38404
MPKWGREPV

38405
MVEATDWWV

38406
MWTTETMGV

38407
MWVMGPLCV

38408
NHCMWEGYV

38409
NHHPFMYYV

38410
NIMLGQNSV

38411
NWVHPITRV

38412
QGCHFNMSV

38413
QVVKFEIPV

38414
RRQWQDFMV

38415
SAGYMCYDV

38416
SGDGFHYGV

38417
SIMHLCEHV

38418
SNCSVVKCV

38419
SPVILNADV

38420
STRMCQMLV

38421
SWQQSPYFV

38422
TARYYDGPV

38423
TCGYFTRDV

38424
TWASRRKCV

38425
VKCLGAMNV

38426
VNCKHMNNV

38427
VNYSWNIQV

38428
VVMPFWSAV

38429
WTVELANGV

38430
WVQPRMSIV

38431
YHEHAVESV

38432
YLEANMCHV

38433
YQHWTTMGV

38434
YTTFYRFQV

38435
QAMLHHVWV

38436
QNLMYCSSV

38437
TCGQYVNGV

Example 7
AAV5 Variants with Tissue Tropism in Adrenal Gland

This example describes engineered AAV5 variants with tissue tropism in adrenal gland that were discovered using the methods and systems described in EXAMPLE 1-EXAMPLE 2.

CNS, liver, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord tissues were analyzed to identify amino acid residues in the AAV5 VP1 581-589 region that drive adrenal gland tropism. The results are shown in FIG. 18K which shows the positional frequency of each amino acid in the 581 to 589 region of AAV5 VP1 in adrenal gland tissue as compared to all other analyzed tissues, wherein the capsid polypeptide has the sequence of SEQ ID NO: 2. The position (“Xaan”, n=1-9) is denoted on the x-axis and amino acid residues are denoted on the y-axis. The frequency of a particular amino acid residue in a particular position is indicated in the heatmap as intensity. Darker squares indicate a higher frequency of a particular amino acid in a particular position and lighter squares indicate a lower frequency of a particular amino acid in a particular position.

The frequency of a given amino acid residue occurring at a specified position corresponding to position 581 to position 589 of SEQ ID NO: 1 (generalized in SEQ ID NO: 2) in variants identified in adrenal gland over the frequency of that given amino acid residue occurring at the specified position in variants identified in all other harvested tissues was analyzed to identify a set of sequence rules for capsids that preferentially target adrenal gland tissue. With reference to TABLE 13 below, and SEQ ID NO: 2 (AAV5 VP1), the following amino acids can, thus, be independently mutated, in any combination, at any one or more positions Xaa1-Xaa9, to provide a VP1 capsid with enhanced adrenal gland tropism as compared wildtype AAV5 VP1 capsid (SEQ ID NO: 1), where adrenal gland tropism here refers to properties that are preferred for adrenal gland transduction over properties that are preferred for transduction of all other harvested tissues.

TABLE 13

Adrenal Gland Tropism Rules

Xaa1 is selected from A, C, K, Q, R, S, or T

Xaa1 is selected from C, K, or R

Xaa1 is C

Xaa2 is selected from A, C, I, S, T, or V

Xaa2 is selected from A, V, or T

Xaa2 is

Xaa3 is selected from A, F, G, K, M, Q, R, T, or V

Xaa3 is selected from A, G, or M

Xaa3 is M

Xaa4 is selected from A, K, M, Q, R, or V

Xaa4 is selected from A, R, or K

Xaa4 is K

Xaa5 is selected from F, I, L, M, R, T, V, or Y

Xaa5 is selected from R, V, or Y

Xaa5 is V

Xaa6 is selected from G, H, M, N, R, or S

Xaa6 is selected from H or N

Xaa6 is N

Xaa7 is selected from A, H, K, Q, R, S or V

Xaa7 is selected from H, Q, or V

Xaa7 is H

Xaa8 is selected from A, G, H, M, Q, or S

Xaa8 is selected from A, G, M, or S

Xaa8 is S

Xaa9 is selected from A, E, N, P, R, S, or Y

Xaa9 is selected from P or E

Xaa9 is P

TABLE 14 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that were found in adrenal gland tissue and comport to one or more of the rules provided in TABLE 13. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 1118-SEQ ID NO: 2117, as disclosed in TABLE 14. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 14

Sequences of the 581 to 589 Region

in AAV5 VP1 Capsid Polypeptide that

Drive Adrenal Gland Tissue Tropism

SEQ

ID
581-589

NO
Sequence

1118
CAAQYWLTP

1119
CARFVERWP

1120
CCAKFQMCT

1121
CEADGNWMR

1122
CIDMVQSWS

1123
CMLLAEKMF

1124
CMYPKSTDR

1125
CSLPYVREG

1126
CSVKMPGFR

1127
CTANVWTAQ

1128
CTVVMMRQP

1129
CVIPLRMYM

1130
CVIQTSRGI

1131
CAAYRGLLE

1132
CAFQPVRWF

1133
CAFQRMMYQ

1134
CAHQPQRDR

1135
CANYAFMTE

1136
CAPLVNSFC

1137
CARMFAAQW

1138
CCHFQHKAG

1139
CCLQPELTM

1140
CCRPFVLHQ

1141
CDVKCKKED

1142
CEMIANTNN

1143
CFNIIQRYD

1144
CFRQVQAMS

1145
CGDWLWRNL

1146
CHGHRPFDA

1147
CHIDYDLGF

1148
CHMKFNKFG

1149
CIGKCWPPM

1150
CIGRVNGTR

1151
CIKLIMDDR

1152
CIKSKFSWQ

1153
CINEKQIRW

1154
CIVKFSNTQ

1155
CKMATNGMT

1156
CKMYMHESR

1157
CLMALKRSG

1158
CLSSYSRYQ

1159
CLVKYPQFG

1160
CMYFIAEWH

1161
CPDEPRWFP

1162
CPFKVFAHG

1163
CPMIDEFPT

1164
CPTWSCQQW

1165
CPWQIKVAA

1166
CQMEQHYSP

1167
CSAPLRHWY

1168
CSIFGPTYP

1169
CSKFIVDWE

1170
CSLRYFMAR

1171
CSRTMEVGL

1172
CSSVRWMSP

1173
CSSYSFQHS

1174
CTFMANWGF

1175
CTGQRGFAN

1176
CTMDQWQTA

1177
CTTHFYRAR

1178
CVAPVRLQD

1179
CVAYRLVNP

1180
CVFMRLTSG

1181
CVFVRNNMM

1182
CVGSLMRIQ

1183
CVHLTSSSA

1184
CVKQYMQPP

1185
CVRHTENWC

1186
CVTTRHKQC

1187
CVWSAPVEC

1188
CVYQSRFSC

1189
CWNENTYEA

1190
CWNGIQRQA

1191
CYTRFHYSE

1192
CYVFRHHYC

1193
HVASTSYVT

1194
HVFDTDRWM

1195
HVFHLDEAN

1196
HVKDTDVQI

1197
HVQARQNPG

1198
HVSDKNHAK

1199
HVVPQNWNW

1200
HVVQRLTIP

1201
IVEARLEFN

1202
IVEDFEIVI

1203
IVFYRFEFV

1204
IVLPTMLLA

1205
IVVEAKMTT

1206
IVVSTWHEQ

1207
IVWLQRVGD

1208
KVDHTINQM

1209
KVDTEKNMM

1210
KVTGHEQDV

1211
KVVLWEHFQ

1212
LVCTHTKAA

1213
LVDDDYQMD

1214
LVEIPCSNW

1215
LVHSCQHTE

1216
LVLQSFHHS

1217
LVNCYTYRA

1218
LVQVMDMKP

1219
LVVIFEGRH

1220
LVWMDPHWG

1221
MVAAKEKHD

1222
MVAHNGLGF

1223
MVCMLVHHC

1224
MVDHSYQQE

1225
MVEVIEGGQ

1226
MVHGNHECW

1227
MVNPMHCCN

1228
MVRKYEKMH

1229
MVSCNQHDS

1230
MVSHVVASE

1231
MVSVDQMSV

1232
MVWSRADWS

1233
NVAIILSTD

1234
NVAVEHNMG

1235
NVEDTRKGS

1236
NVFLAHQRA

1237
NVHNSGRFD

1238
NVIGTENMS

1239
NVKSEHRES

1240
NVLLEKAFW

1241
NVNRGHLLE

1242
NVNWQARND

1243
NVTHKQGCS

1244
NVYPDSMHG

1245
PVNFFEHPM

1246
QVDIDTQNA

1247
QVDPHGNMH

1248
QVDRWCTAA

1249
QVEYAHSRA

1250
QVFMQHDIE

1251
QVGNPLKEF

1252
QVIEWGRSG

1253
QVIIRKWFA

1254
QVLMAQFSR

1255
QVQKCMQYV

1256
QVQKGACPS

1257
QVQSGPYWG

1258
QVQTENMCD

1259
QVRCGIKTG

1260
QVTFHAYVC

1261
QVTIQNDYD

1262
QVWNPHLCM

1263
RVATEHFRE

1264
RVDNPFVSY

1265
RVSECTENY

1266
RVTEEVGSR

1267
SVDQHDHLL

1268
SVERYPLTA

1269
SVFDVVHEE

1270
SVHVRIVEP

1271
SVKQHHVDW

1272
SVLGMPAYR

1273
SVMDANTWR

1274
SVNKDVRHC

1275
SVNSYMDSV

1276
SVPRKSKCS

1277
SVRSYHSQP

1278
SVTQPLYCQ

1279
SVVKRNDDV

1280
TVAHFLNGG

1281
TVALSEQYR

1282
TVAVRYETL

1283
TVCSTIVDW

1284
TVDWRWGDF

1285
TVEHKMDAP

1286
TVISDYEDC

1287
TVKMLQHWT

1288
TVMKEQANM

1289
TVNDHGPCI

1290
TVNKWHQTA

1291
TVNQCMTHC

1292
TVYFVWDSQ

1293
VVGATATGM

1294
VVGPPGHHY

1295
VVMFWQEGD

1296
VVMQQYRSH

1297
VVNKDKWRI

1298
VVQWPTSLL

1299
VVSAAKSSG

1300
VVSHKQRCS

1301
VVSKYQIVP

1302
VVTCRHQGS

1303
VVVPMSNCE

1304
WVMCISGHY

1305
YVMFNTWRE

1306
YVTMSDERP

1307
AVGFMECAN

1308
DVKDRNELH

1309
EVSAEEGQN

1310
GVMPRWGLF

1311
GVVNLVKMQ

1312
IVMYDHFNQ

1313
KVILDSFRQ

1314
KVMPEPVTE

1315
LVHLFQYHL

1316
QVVACMHHT

1317
RVDYSAIHM

1318
SVIMHHDQH

1319
VVEMNGDDD

1320
VVEYAPWAW

1321
AVAKGVGVT

1322
AVMATGVTQ

1323
AVSGNWMMT

1324
AVSKSGCGM

1325
DVAAVQVLE

1326
DVASYEQAY

1327
DVEWQQSWT

1328
DVGGLNREY

1329
DVHEVMSCG

1330
DVHICTVHI

1331
DVKGMLDGF

1332
DVKMQQKYF

1333
DVMGHAKLA

1334
DVRAYLNKE

1335
DVRYNQNAW

1336
DVSFMGWDC

1337
DVTHHFAKQ

1338
EVAHFLANE

1339
EVDNMQMGE

1340
EVGVSAPMM

1341
EVSGMFHPA

1342
EVTTMLKAC

1343
EVWINMAPE

1344
EVYMEPANW

1345
GVFFRYEHG

1346
GVFSTGVVT

1347
GVGVGLGTQ

1348
GVHQESGQN

1349
GVNKLSYEN

1350
GVRIPMAAD

1351
GVSQFMAET

1352
HVGAHTRSD

1353
HVGKMPGGS

1354
HVNSFQPDR

1355
IVCTTFEDS

1356
IVDQTCMHA

1357
IVGIWQWMG

1358
IVKNLEQYM

1359
IVKTLICYN

1360
KVDVITVKD

1361
KVGAHNDEV

1362
KVTWLDLED

1363
LVEAVMNAT

1364
LVKACTDQH

1365
LVKQYGLIH

1366
LVLMDRQAY

1367
LVSQTPRKV

1368
LVVDHAEWS

1369
MVALTDQLG

1370
MVDSNACLC

1371
MVHPSQNVD

1372
MVKEAPYYR

1373
MVSEYATNA

1374
MVTGISKSS

1375
NVESNYNLI

1376
NVEWGTVQS

1377
NVFMTSFQC

1378
NVFQPEDHC

1379
NVTWEDVQE

1380
PVAAAEPRN

1381
QVAEPGGRF

1382
QVDSWPVLS

1383
QVGCTVGPG

1384
RVQHSGAHW

1385
RVINTWECM

1386
RVYTTGKCS

1387
SVHHSRWGY

1388
SVNQAMARK

1389
SVSVMTYSE

1390
TVAMDPDYA

1391
TVATEPAQQ

1392
TVDFFVMDC

1393
TVDHCVGTV

1394
TVGLNERNG

1395
TVHWWECVQ

1396
TVQTNFTSK

1397
TVTWEWMMK

1398
TVVGLITNA

1399
VVEKRMDRS

1400
VVHGYVCKQ

1401
VVHPLAYCT

1402
VVNKQQISE

1403
VVTSPDAFQ

1404
VVVHTQCNP

1405
WVTPTNHKP

1406
YVAEYGLGA

1407
YVEERGSQR

1408
YVIVHMKGM

1409
YVTPNIAVH

1410
HCMRQSESG

1411
HSMPEAVAY

1412
HTMEILPGL

1413
HTMHDSCFN

1414
HTMLWAKDE

1415
ICMSYLSWD

1416
IPMFSIHGN

1417
KAMLEGPYL

1418
KNMINEAAP

1419
KSMWEHAIF

1420
KTMEDVIHA

1421
KYMSYERES

1422
LAMTLAVGQ

1423
LCMDYALVD

1424
LCMLLASQQ

1425
LSMCFRDDY

1426
MCMVERRGS

1427
MFMSQHVED

1428
MNMHTDCTT

1429
NAMWTTEYE

1430
NEMKFSYSH

1431
NIMNWQNGD

1432
NLMPTNHAM

1433
NSMQHCVCP

1434
NTMHPVGMM

1435
NTMMVGRCE

1436
PIMFEGECK

1437
QCMMKSSYE

1438
QFMPYLGFA

1439
QGMENQWHQ

1440
QHMYANCLP

1441
QKMTRQHWG

1442
QSMEQEFVT

1443
WSMMALKDY

1444
QSMMTDRTY

1445
REMTKVGGR

1446
RGMYRCGWD

1447
RNMYISQWE

1448
RSMGNEQDP

1449
SEMKLVNLP

1450
SEMVKHNFE

1451
SGMKQVGAA

1452
SGMLVGIYN

1453
SHMVSNSAV

1454
SSMSKHEVA

1455
STMPGTDQK

1456
TAMHQTMLN

1457
TAMMSGHLV

1458
TAMVVEYDQ

1459
TCMKNINYQ

1460
TCMLGAEAA

1461
TCMQLATMQ

1462
TCMTLRHIG

1463
TCMYYWKQP

1464
THMLCPRMD

1465
TNMSYQRSD

1466
TRMPVNQSC

1467
TWMEWHANA

1468
TYMGTHNHS

1469
VFMPDYYDK

1470
VKMPYHYEG

1471
VQMQEVPHY

1472
WFMHYSWKH

1473
WKMEGDCFK

1474
EIMCAYVKA

1475
GAMPNPYHE

1476
GTMWSTKSC

1477
HPMFMIPGY

1478
HYMLKEYFA

1479
HPMFMIPGY

1480
IMMAQWFAH

1481
PSMPNIKPS

1482
PYMQYWYWE

1483
REMLRDKMP

1484
SAMQVMTMC

1485
TAMSICMYG

1486
TEMDKGLIR

1487
ANMFRNGMQ

1488
ASMFWNYQV

1489
DRMVIKSAE

1490
EAMRAMTPI

1491
EAMYWQCAI

1492
EIMWEGVGS

1493
ENMHKEYVQ

1494
EPMEHYLRS

1495
GAMTMCCEW

1496
GKMMGLHSH

1497
ILMEYDHDK

1498
IMMHRVMSD

1499
KAMEVHFQK

1500
KAMGLPSWY

1501
KSMWEDR1C

1502
LAMPWDTLQ

1503
LCMLRMMGH

1504
LIMAHWLDM

1505
LIMDSKMGQ

1506
LLMPVLHQM

1507
MGMKREGSG

1508
MQMSKNKHP

1509
MRMIHWMHC

1510
NKMEWHKYC

1511
NWMQNTEGH

1512
PEMSFEFTT

1513
PGMNMVRKE

1514
PTMWWYLLV

1515
QCMPVSGMT

1516
QGMHAMFYA

1517
QMMQLYCGA

1518
SAMVHSNKS

1519
SHMSYLMSH

1520
TCMPNIQCP

1521
TFMHQFGTY

1522
TQMMTPVGA

1523
TWMADWHQE

1524
VAMQSMQEH

1525
VAMQTHQCY

1526
WWMHGRVGR

1527
YCMLKHKAG

1528
YCMWQQTSN

1529
YKMDHNNTY

1530
YSMTWNQQT

1531
HTGKADVSV

1532
IMEKLRRSS

1533
IPFKAVMVG

1534
IRVKPGCRV

1535
ITGKYPMTA

1536
KLEKTVVYS

1537
LDIKMDFLC

1538
LMHKDQVSP

1539
LNVKTGNAW

1540
LTTKQPDMP

1541
MASKLHMAV

1542
NSLKYEFSE

1543
NTGKTGQFN

1544
PCAKSYDQS

1545
PKDKGVICE

1546
QEVKGQWWP

1547
QIFKQTEVQ

1548
QITKPILDR

1549
QLAKAMTQS

1550
QPVKYPQTN

1551
RISKDGSDR

1552
RIVKRYHNG

1553
SLCKMLTAK

1554
SYKKPVKYM

1555
SYNKVMTQH

1556
TACKTEGGP

1557
TCVKTTHGA

1558
TDGKTRMQQ

1559
TETKWQHLE

1560
TIHKYCHHQ

1561
TSTKHHWWC

1562
TSVKKPHLA

1563
TTLKCTNTW

1564
TWIKPHQQL

1565
VCEKSNDYA

1566
VCFKMLLGN

1567
VECKTMAHQ

1568
VNAKWQTAS

1569
VSCKIYHYA

1570
VSGKECSED

1571
VTYKYYCDF

1572
WYQKQAEHG

1573
YCSKSVRCC

1574
NFSKCAKYD

1575
VCTKGFLNS

1576
AAHKEMKDT

1577
ACGKMHGPN

1578
AFLKQSIIH

1579
AGSKRFDWF

1580
AILKMTKVL

1581
DTGKFMCKW

1582
EHQKNINMH

1583
ESEKVWMMT

1584
EWRKKKKRT

1585
FCRKIEQAN

1586
FICKTLHQH

1587
GCDKTRFEN

1588
GILKFRDFM

1589
ICWKFLVSN

1590
IEVKYWNHH

1591
IWYKWWVMF

1592
KCDKPMLKP

1593
KMEKVERMP

1594
LDLKHNMCD

1595
MCIKCDAND

1596
NCNKCKNRT

1597
NQCKINVGT

1598
QQLKTN1NF

1599
RIEKLQCDY

1600
STLKTYAYN

1601
TGTKITMQT

1602
TIHKDGEFS

1603
VLQKFGHSF

1604
VTGKHIAQN

1605
YCYKTAVAA

1606
HCALVNRLR

1607
HFAMVWAGE

1608
HIGYVEAMR

1609
HTKPVNAQP

1610
IHQYVNFVH

1611
INNTVAHQE

1612
KALMVQQFD

1613
KCLMVDWGV

1614
KHGQVSIHD

1615
KMDRVGYSA

1616
KWNNVENNE

1617
KYSPVEWMY

1618
LGQHVQCWQ

1619
LKSWVCATS

1620
LMKCVMCRD

1621
MASPVNSEF

1622
MCLFVRVSQ

1623
MHGMVHAFE

1624
MLIHVNHKS

1625
MQDQVCKIG

1626
MQQRINKHM

1627
MRQTVDRPE

1628
MYHAVQCMP

1629
MYYQVSKQD

1630
NAENVDNGV

1631
NFEFVERYR

1632
NGHEVPVEE

1633
NKQVVQMTA

1634
NLPLVHSLE

1635
NLQRVEMGV

1636
NMQNVARCY

1637
NSWDVGSFN

1638
NTYPVLSDW

1639
PEYFVVYMY

1640
QGGAVMHEE

1641
QGGAVMHEE

1642
QIQPVNALF

1643
QQLSVHMWH

1644
QSGWVQIAA

1645
QWQYVMTDK

1646
RCWSVEASQ

1647
RDQRVLKVM

1648
RTTGVGDIS

1649
SAWCVPGQL

1650
SCHLVIKHH

1651
SFAPVAHIE

1652
SGRHVLIEI

1653
SITAVWSDE

1654
SQNMVDSQC

1655
SSCIVNIAS

1656
SSNLVGAGE

1657
STELVKYID

1658
TMHPVCCPM

1659
TMREVQCEW

1660
TPAHVCRGY

1661
VNDMVLAVV

1662
VSFCVRKQS

1663
YALTVPHSE

1664
YTPNVHYMV

1665
ECANVIKFN

1666
GACTVQNMT

1667
HIEWVSMHR

1668
MGQNVNHAG

1669
QAAFVNYGP

1670
RGFHVIVDQ

1671
RYDLVSYHQ

1672
VFDTVFAYC

1673
VKKYVDVHF

1674
VQSAVSMCS

1675
APVFVEQMG

1676
ASTAVLVQR

1677
DAGFVREPV

1678
EAVPVESGD

1679
EEKEVICMC

1680
EFEQVGFMM

1681
EGTMVKQGE

1682
EHNCVYLTE

1683
EMHYVCCQT

1684
EPFRVTQCG

1685
FASIVFGGE

1686
FEYMVLNWH

1687
FQAPVTKAS

1688
GLGYVHRSN

1689
GWQSVWNAP

1690
GYNWVQEQR

1691
HAVMVYSDY

1692
HTKGVVHQP

1693
IARHVDHDL

1694
IQTEVHTVY

1695
KQQDVTMNA

1696
KYIDVWDAR

1697
LGVDVLADY

1698
MQVLVSMTP

1699
NTIDVQLGG

1700
QKAYVSVSG

1701
QSPAVNCAW

1702
RIEQVDVFN

1703
RIGGVADAD

1704
RWECVTGGY

1705
TCIRVWQWA

1706
TMRYVNTCV

1707
TSHNVANFG

1708
VEQMVMPPH

1709
VYTVVDEHS

1710
WFPVVWSHN

1711
YHVLVEGDR

1712
HGIALNLGT

1713
HRDYANSWN

1714
IGTRFNAQT

1715
ILDDCNCKC

1716
IPRMLNHVE

1717
ISYQPNPME

1718
KASELNQVL

1719
KHLDCNERE

1720
KLSNMNHPI

1721
KMEQGNMDS

1722
KMFHSNQEA

1723
KNWVCNLLT

1724
KTEVTNGPT

1725
LCIRKNYTH

1726
LCQFNNATD

1727
LEAECNSMQ

1728
LGDERNCGQ

1729
LHDNWNRYT

1730
LPFRSNNAQ

1731
LTVQNNGIR

1732
MCAYMNKKE

1733
MPNLSNSFQ

1734
NGAEMNWHE

1735
NGNQFNNCT

1736
NSTQPNLWF

1737
NYTIYNPAE

1738
PIETTNHMD

1739
PMYANNYFC

1740
PTQPLNQHG

1741
QAWCANTFH

1742
QCFYANRGD

1743
QHQQFNLTT

1744
QIDMMNCNR

1745
QQLPENLYQ

1746
QTDVINKFP

1747
QTGMENHQN

1748
QYSWSNINT

1749
RIEANNWGV

1750
RIICQNAVA

1751
SEKVINASC

1752
SIAVKNMCH

1753
SKRDNNCEV

1754
SMQRYNSKP

1755
SSLCRNMWP

1756
STYGRNLYP

1757
SWSNINHQT

1758
TCGEYNHRC

1759
TFQEKNVMH

1760
THQMTNTYS

1761
TIAQDNKEA

1762
TLITTNRQY

1763
TMVQNNMCF

1764
TRQPFNMSL

1765
TSSEFNRQR

1766
TSVACNSTD

1767
TTVFWNNFK

1768
TWVPDNADS

1769
TYQVNNWSD

1770
VANHHNEIQ

1771
VIGTANAAP

1772
VMGMINGAW

1773
VPLVINKEQ

1774
VSSQFNGPS

1775
VTHCKNVAC

1776
VWYPQNAVE

1777
WCNYLNQGM

1778
WQCTENWQN

1779
FYWAHNICE

1780
KYEILNCNH

1781
LYAMLNKFF

1782
MGNYQNTHY

1783
RKDEANGQT

1784
YFIIQNHLD

1785
YTRSNNWDY

1786
AAFMPNNYY

1787
AALFPNWSN

1788
AEECSNHNV

1789
AHSMTNLVG

1790
AWFDHNEHQ

1791
AYARSNRQD

1792
DATIANAQV

1793
DIAEKNRMM

1794
DWKTSNAEY

1795
EGDWYNYGT

1796
GIIFSNTCM

1797
GPPPYNAWF

1798
GSQRTNTSC

1799
GTSRYNAFI

1800
IALLTNDLA

1801
ICGRLNNNP

1802
INAPQNMSW

1803
ISGYHNVDE

1804
IYHTLNHCM

1805
KAEMSNAIT

1806
KAIPENSHR

1807
KIHSPNQDE

1808
LKSPHNENG

1809
LLEVKNHDH

1810
LSDLQNKGA

1311
LSHTSNMGH

1812
LTYLANQSD

1813
MGKSRNGSE

1814
MISMWNRSN

1815
MMDPTNFGS

1816
MNNITNWNP

1817
NTGRANKFD

1818
NNQWMNSLE

1819
NQIRNNMER

1820
NTYMHNREG

1821
NWNDSNDCC

1822
QCYTFNDTK

1823
QRDLANGQF

1824
QTASWNMEM

1825
RCEPFNWRE

1826
RMAQINQQG

1827
STALMNEYQ

1828
SWWLTNTAV

1829
TADFYNDQM

1830
TASSANLSW

1831
TFEICNSPQ

1832
TFLRLNDFV

1833
TFSQHNHKA

1834
TKCTTNQQC

1835
TSALNNNSK

1836
TSHDNNAQI

1837
TTFHINEYP

1838
VCVPKNGIH

1839
VGAQTNCLQ

1840
VYNLLNHYQ

1841
VYRIANTQY

1842
YAKSRNIVG

1843
HKESLGHHC

1844
HRHPCGHIP

1845
HSAPSFHTQ

1846
IDKLYWHWG

1847
IDQPIQHAS

1848
IGKMNDHGM

1849
IIQYNGHQA

1850
IKTELAHQY

1851
KAAGDSHTP

1852
KADAPIHWS

1853
KAECKPHHE

1854
KATELGHCG

1855
KCTAMRHKE

1856
KTEQATHTC

1857
KTSEWHHVG

1858
LATSRVHFG

1859
LGGWMEHSF

1860
LSDHLHHYT

1861
MCHHRHHAE

1862
MFRERIHHD

1863
NCIFIEHPT

1864
NKVGCQHSY

1865
NMKVDAHSN

1866
NSSYEEHDQ

1867
NTCFIGHNE

1868
PGSQYVHWI

1869
QASYRTHHD

1870
QCAIMRHYC

1871
QIAEIIHFG

1872
QIYPEHHKE

1873
QRKHYVHVF

1874
QWDTLRHTP

1875
RICFHQHWG

1876
RKECEDHGG

1877
RTLEWFHRG

1878
SFSWIYHQD

1879
TAVMYAHDC

1880
TREINDHGI

1881
TRGSMYHHC

1882
TTVRNFHAE

1883
TYCCYHHQG

1884
TYECAAHYP

1885
VAQPYIHSH

1886
VNKPAGHIQ

1887
VRDLLHHDM

1888
VYETLKHRG

1889
VYETMGHEE

1890
WYYYPAHHK

1891
YEVTIPHAI

1892
ETRWWPHIE

1893
MSCMKEHSP

1894
NCAPMEHHV

1895
QGLNYEHQE

1896
SCRHIYHEA

1897
SMNQAIHLE

1898
AAYMTDHMN

1899
ASTSIVHAH

1900
DSFLNRHTE

1901
DTCSRKHWM

1902
ESFYQRHVT

1903
ESVVGPHWD

1904
ETDVCRHGV

1905
HQNASMHMP

1906
HYIQIQHDY

1907
IMVALMHED

1908
IYNQFPHYG

1909
KHSPDHHFE

1910
KMAMLRHAT

1911
KSVGWCHAF

1912
LQQPIVHIE

1913
LTETEHHDQ

1914
MATHCLHWR

1915
MMPEIMHVL

1916
NMGLLYHTN

1917
NTEPMTHWN

1918
PMKPMHHGY

1919
QCTFYFHWR

1920
QHVQEIHGA

1921
QIYHNHHTA

1922
QSCVGLHQI

1923
SATSKSHTF

1924
SCDEYMHHP

1925
SCVLKVHFP

1926
SLCMHRHDF

1927
SMGPTVHQM

1928
TAGHEGHIC

1929
TCEWWIHMD

1930
TFGWFMHIP

1931
TFSQHSHKA

1932
TRQQLFHTQ

1933
TIGSTKHIG

1934
VMPCYWHTT

1935
WDGTKVHWL

1936
YHVILVHQA

1937
YPTVFEHSE

1938
HIGYMPRSY

1939
HLCPNQFSP

1940
HSNGLGPSC

1941
HSQIFMLSW

1942
IGSQIFTST

1943
IGSSIMSST

1944
IIHQQHNST

1945
ITSRRGMSE

1946
KAEPLRLSP

1947
KAFIRDESY

1948
KGDWTVKSY

1949
KGGHKDESP

1950
KIWAHMCSP

1951
KMFLLEQSD

1952
LASCNWRSQ

1953
LAYPCYASA

1954
LFSFLWRSA

1955
LIDQYTNST

1956
LIEQTCVSN

1957
LKVDELLSN

1958
LMEQFDYSQ

1959
LQGEQSASD

1960
LYEQYKASY

1961
MQQYAYISH

1962
MSIRHMISY

1963
NCGVSMKSN

1964
NCHSKCYSS

1965
NTAHCWESD

1966
QGSFAHVSQ

1967
QHKVMGRSW

1968
QTRCDFLSH

1969
RADHYRASK

1970
RCEAEGGSK

1971
REHVMETSM

1972
RSNIEHLSE

1973
RTDSFYESD

1974
RTIDPVQSS

1975
RTVEHQASA

1976
SLGCREISL

1977
SLHAWAISA

1978
SNFEFQKSH

1979
TCGLFSGSY

1980
TFTRPEQSQ

1981
TGKWISPSN

1982
TKENLAASY

1983
TMLVHHGSM

1984
TSFLLGVSS

1985
TTHDEGVSI

1986
VAAHPLTSD

1987
VERMATESE

1988
VGQVIDRSW

1989
VTTRNPFST

1990
VYQVCTISD

1991
WCQRFQTSS

1992
WITEIELSG

1993
YFALSGESS

1994
EIHHLPSSH

1995
GIAETYISA

1996
HSEQMHQSD

1997
LAKSCSSSQ

1998
NASLDMISV

1999
SSEPETKSN

2000
VADMGPWSG

2001
APHSYAMSY

2002
ASHIKWNSH

2003
DAQQAPNSS

2004
EDWIMHASF

2005
EPSFESDSN

2006
ESKWWIQSD

2007
FGKDTFVSS

2008
GANNIYVSM

2009
GKSLMMTSF

2010
GNTQQPMSF

2011
GTAAMEQSC

2012
IDASLDMSW

2013
ISEPCYVSI

2014
LMNSYCVSH

2015
LNVEAELSF

2016
LSLPGVPSM

2017
NMTGIHVSP

2018
NTFATGNSP

2019
QCYESMTSW

2020
QICQTDASA

2021
SCVFRLASG

2022
SDPHAGPSD

2023
SLWILRMSN

2024
SSCGNMVSN

2025
THWQYEASH

2026
TNAQFSNSW

2027
TTGVSEGSA

2028
VANYHPYSM

2029
VIRYGDLSQ

2030
VMGPQPVSV

2031
VSVMLAASD

2032
HFYSQSTLP

2033
HGAIHAVLP

2034
HHECMWMFP

2035
HTEARMVEP

2036
IGNYYHKWP

2037
IIRHHEFAP

2038
IIWQQDIGP

2039
IQSECVVVP

2040
KNESAKQCP

2041
KSVNLCVDP

2042
KTSFSEMPP

2043
KTSMTFYNP

2044
LARSDCRQP

2045
LFYPEWDGP

2046
LGCDTEQMP

2047
LHEFHKSYP

2048
LQLRSDWCP

2049
LTIHSQREP

2050
MACLFRACP

2051
MEGAIGQNP

2052
MGANIAQMP

2053
MGSSRMIYP

2054
MHIFHSLCP

2055
MKSECMSGP

2056
MWGHISEMP

2057
NFHMKSMCP

2058
NLHDFIFAP

2059
NWNLWDTYP

2060
PEFEFFTTP

2061
QCDEQEYNP

2062
QGQRFYNYP

2063
QYARLDCLP

2064
RADIGSLCP

2065
RCILKREQP

2066
RETVAMDMP

2067
RFEFLMAPP

2068
SAYIGEQGP

2069
SLRVCIGQP

2070
STHLKLSDP

2071
STNLIGQNP

2072
TCGWSHAMP

2073
TGNLWQSMP

2074
TSQAPMCNP

2075
YCDRSGIWP

2076
YFYEAKLMP

2077
YNGYTSVCP

2078
YRDRFFAVP

2079
YSHPRDGAP

2080
ACQLLHWQP

2081
DWDSMCWWP

2082
EDTMAQMWP

2083
LCHFRLVGP

2084
NQTGHMPAP

2085
QAPAYMWGP

2086
WHSRKQWPP

2087
YTKYLMTEP

2088
AFATCSMMP

2089
APVCGQDPP

2090
DADERQILP

2091
DERNSFWFP

2092
DIHTNTIEP

2093
DSGWWATMP

2094
GLTMNRCYP

2095
GMFWLTQAP

2096
GWWVATMLP

2097
HAELTMNVP

2098
IACCSEAIP

2099
KGDYQADAP

2100
LEKSMMTAP

2101
LRYLSDADP

2102
MCRANGDMP

2103
MCTLRHWQP

2104
MFELFVMGP

2105
MSTPNMTQP

2106
NAEQWCCGP

2107
NMTPLMTLP

2108
QAQPNVNYP

2109
QAYTDDTGP

2110
QWGGNQQTP

2111
QYEEMAWGP

2112
SAYMLRYQP

2113
SGQQFIFCP

2114
SPGHNMVGP

2115
SSVHTLYFP

2116
SYNYEHAWP

2117
TCNMQRAVP

Example 8
AAV5 Variants with Tissue Tropism in Sciatic Nerve

This example describes engineered AAV5 variants with tissue tropism in sciatic nerve that were discovered using the methods and systems described in EXAMPLE 1-EXAMPLE 2.

CNS, liver, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord tissues were analyzed to identify amino acid residues in the AAV5 VP1 581-589 region that drive sciatic nerve tropism. The results are shown in FIG. 18N which shows the positional frequency of each amino acid in the 581 to 589 region of AAV5 VP1 in sciatic nerve tissue as compared to all other analyzed tissues, wherein the capsid polypeptide has the sequence of SEQ ID NO: 2. The position (“Xaan”, n=1-9) is denoted on the x-axis and amino acid residues are denoted on the y-axis. The frequency of a particular amino acid residue in a particular position is indicated in the heatmap as intensity. Darker squares indicate a higher frequency of a particular amino acid in a particular position and lighter squares indicate a lower frequency of a particular amino acid in a particular position.

The frequency of a given amino acid residue occurring at a specified position corresponding to position 581 to position 589 of SEQ ID NO: 1 (generalized in SEQ ID NO: 2) in variants identified in sciatic nerve over the frequency of that given amino acid residue occurring at the specified position in variants identified in all other harvested tissues was analyzed to identify a set of sequence rules for capsids that preferentially target sciatic nerve tissue. With reference to TABLE 15 below, and SEQ ID NO: 2 (AAV5 VP1), the following amino acids can, thus, be independently mutated, in any combination, at any one or more positions Xaa1-Xaa9, to provide a VP1 capsid with enhanced sciatic nerve tropism as compared wildtype AAV5 VP1 capsid (SEQ ID NO: 1), where sciatic nerve tropism here refers to properties that are preferred for sciatic nerve transduction over properties that are preferred for transduction of all other harvested tissues.

TABLE 15

Sciatic Nerve Tropism Rules

Xaa1 is selected from C, G, K, M, Q, R, or Y.

Xaa1 is selected from C, R, or Q.

Xaa1 is C.

Xaa2 is selected from A, C, F, I, Q, T, or V.

Xaa2 is selected from A, C, or I.

Xaa2 is A.

Xaa3 is selected from A, F, I, M, R, S, or T.

Xaa3 is selected from F, M, R, or S.

Xaa3 is R.

Xaa4 is selected from E, N, T, Q, or V.

Xaa4 is selected from E, T, or V.

Xaa4 is T.

Xaa5 is selected from F, H, Q, S, V, or Y.

Xaa5 is selected from F, V, or Y.

Xaa5 is V.

Xaa6 is selected from K, M, N, Q, S, or V.

Xaa6 is selected from M, N, or S.

Xaa6 is N.

Xaa7 is selected from K, M, Q, R, or T.

Xaa7 is selected from M, Q, or T.

Xaa7 is M.

Xaa8 is selected from A, G, H, Q, S, or V.

Xaa8 is selected from H or S.

Xaa8 is H.

Xaa9 is selected from C, E, I, K, or R.

Xaa9 is selected from C, I, or K.

Xaa9 is I.

TABLE 16 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that were found in sciatic nerve tissue and comport to one or more of the rules provided in TABLE 15. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 26118-SEQ ID NO: 26990, as disclosed in TABLE 16. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 16

Sequences of the 581 to 589 Region

in AAV5 VP1 Capsid Polypeptide that

Drive Sciatic Nerve Tissue Tropism

SEQ

ID
581-589

NO
Sequence

26118
CQGYEHSQE

26119
CAHAMKWGS

26120
CGGQVEFGW

26121
CSSCYHRVF

26122
CCCEALMPQ

26123
CENPLCQSS

26124
CFGPEGYAG

26125
CHPQHHHSH

26126
CSSFYHRVF

26127
CKEPRKMRE

26128
CPVHQMDFT

26129
CIVKSNGSK

26130
CPGLQFDTP

26131
CHHFNMPSQ

26132
CCRQIWVQY

26133
CIIARSTAA

26134
CAEKQEYVQ

26135
CAHMQYAGP

26136
CANPLWAQT

26137
CAVKTHTCT

26138
CCNRNWNQQ

26139
CDYMHKLNE

26140
CEGRHTWGK

26141
CFHRHSTVP

26142
CFIAADNRN

26143
CFKHQMMMH

26144
CFSDTDNAM

26145
CFTNHWEMG

26146
CFVPMFNHN

26147
CFWLDWWCW

26148
CGYMHKLNE

26149
CHSIFNVKC

26150
CISAGMKND

26151
CKDLRCGCA

26152
CLGQMWAQW

26153
CLHQWLMPT

26154
CLIAPPHMD

26155
CLTSTSHCI

26156
CMCPGKEYP

26157
CMLAWQNDF

26158
CNPKCDAMM

26159
CNYDSHKMK

26160
CPLFLPALP

26161
CQNMKRIDW

26162
CQQPPRPEK

26163
CRIQSIHVH

26164
CSAGDEWGG

26165
CSKHATMAH

26166
CSKHVTMAH

26167
CSMFLHHTQ

26168
CTILNHQDA

26169
CTRPDEIKE

26170
CVAICQDGL

26171
CVHTISNCH

26172
CVTWTNRKL

26173
CVWSEDAGY

26174
CWCEKSHLG

26175
CWLCQCPRQ

26176
CYDHFQMKD

26177
CYQFHTGFW

26178
CHAAVANLY

26179
CTMCHDCSP

26180
CADPQERCD

26181
CMEVTCFVV

26182
CIGPFHCAN

26183
CWGPIMHPF

26184
CSESYRGPD

26185
CQQHFMFAP

26186
CGHLLRRAE

26187
CRRDGYAAM

26188
CPEHNQVGV

26189
CIHRFVSSP

26190
CDWMANVSV

26191
CCRMDNKKS

26192
CLNNSTCRG

26193
CDHVTKHPT

26194
CKYAFKMEK

26195
CDAEAEFRR

26196
CPFKLYSKG

26197
CSIQINNCR

26198
CMVSTQKLA

26199
CFCTYKIEH

26200
CVCWFWFFW

26201
CHMLNDCMS

26202
CMAHIWEGH

26203
CGCQVKCHT

26204
CSMPQQCME

26205
CWECCTDSG

26206
CIMLNGCFQ

26207
CIDVLFKHL

26208
CQMLCRPPV

26209
CAGIHLNEA

26210
CYMNFIVDG

26211
CYIRNMVNL

26212
CPKQANRSG

26213
CVLSRSTMR

26214
CEAGWCRSD

26215
CIQPTWDSY

26216
CYWPMTNCT

26217
CNGYWMCMS

26218
CSVWMQRDR

26219
CWILHHRCS

26220
CYNQFHTDN

26221
YAVMSPPAG

26222
YAVMSQPAG

26223
YAVMSRPAG

26224
YAVVSHPAG

26225
RAVEMQPCT

26226
IAEEGWACP

26227
QAPLATAIE

26228
FAPEFDNHC

26229
SAHMPKSVT

26230
EAYLYNIPM

26231
NAEQCMIGA

26232
RASMCDCFE

26233
SACPLWCGS

26234
DAMHKWIFA

26235
HAPCEQWAW

26236
MASIDKQHV

26237
AAQWPMEYT

26238
TAEWKLCGE

26239
QAEGGPNMP

26240
IATEHPDEW

26241
NALDEDCQH

26242
AAMLYNACM

26243
VADWIGHTP

26244
VAIQCNRSY

26245
GATILTKCL

26246
YASTMHSIP

26247
QALVGNAAE

26248
SAICQNYDA

26249
DAHNDALAR

26250
AADECMLHT

26251
AAEKLFAWE

26252
AAFLGQLSV

26253
AANSRENDW

26254
AATNMSNVH

26255
DAEANGCDA

26256
DAGKLGQHD

26257
DAGWMNSWV

26258
DALPMHGQR

26259
DAMCSIRKD

26260
DARANCDAA

26261
EAEIAYGFM

26262
EAGCINVPS

26263
EAQLYSTKA

26264
EASTYFIWD

26265
FADTCPRHA

26266
FAELHGMQG

26267
FAHDSCESG

26268
FANWLKSFP

26269
FAQIIVHEH

26270
FATGCKQGT

26271
GANKTCCEG

26272
GASPKLGPV

26273
HADEPYEWR

26274
HAEYQQQMG

26275
HAGYIASPN

26276
HAIPFNDSQ

26277
IAFHRGADH

26278
IAGPQQQMT

26279
IAHHRGCHT

26280
IAHVMDCLE

26281
IALVMTSWE

26282
IAMMENRQQ

26283
KADMCPYII

26284
KADMKNPCD

26285
KAELGKLIE

26286
LAMPKPDKQ

26287
LANEQIDFT

26288
LATMHSGQC

26289
MAAWCHKSQ

26290
NADPCMWMQ

26291
NAFQKEACI

26292
NAHMYTKHS

26293
NAMWHTKVG

26294
NANQKWYWS

26295
NASFSNHQA

26296
PAYYGPGSE

26297
QAAPPCAMS

26298
QAIRGTHDQ

26299
QAMRFSENG

26300
QATAEYACD

26301
QAWMPVCHT

26302
RAIMSDWEC

26303
RALECNWFL

26304
RALTFDCHK

26305
SAEYSNDES

26306
SAHTFCAPR

26307
SAHVVLGEM

26308
SALENVPKY

26309
SAMTSSHQK

26310
SAMYTENIS

26311
SASTKVASE

26312
TADLNHGEQ

26313
TADVNLALD

26314
TADVTAQWL

26315
TAGHFKWEQ

26316
TAIRADWQS

26317
TANRMTNQP

26318
TASEAHMEK

26319
TATLWARGY

26320
TATQRGMFQ

26321
TAVHNTHVQ

26322
VAFPAQLGA

26323
VAISTSPTT

26324
VASFNQICA

26325
VATDRMWMC

26326
VAWMMLAKD

26327
YAEVFRFKG

26328
YAIDSSRME

26329
YANWLYDCT

26330
NAELQQADI

26331
TAAGYIVDA

26332
SAFQSSDVP

26333
EAYITYNKA

26334
TAEVRWPCP

26335
YAQRTCWPE

26336
YAMPLYFCE

26337
KAGYQEGSC

26338
AAMWWQWDR

26339
GAYSLHDWR

26340
HAETDHVGL

26341
FAQQDKQDR

26342
YAYHPQCKP

26343
QATGMHRWQ

26344
IADGFAAVA

26345
QAGNSMWAK

26346
MAMWIADTH

26347
MATCYMYAD

26348
IAKIGNCVW

26349
NANYWFYYD

26350
EASWYYPAI

26351
KACMSHQGR

26352
QAEENSHWC

26353
SAAWQFGGS

26354
RACGMRGDY

26355
DADAWWMMH

26356
NACDWKNDY

26357
SAIGKTKKM

26358
QAVRRWWFF

26359
AAIRYDPGF

26360
NAGWQGQVD

26361
NALPAQQHY

26362
IAYHYDQQN

26363
DATTSTECH

26364
HAVNFDCKK

26365
EASIQASWN

26366
QAIPRENPM

26367
DAHAFHCQP

26368
LATTTSYHC

26369
DASQIPITF

26370
VAADSRMMI

26371
PACKEWCFC

26372
IAHWKCHYP

26373
AAMHNLFET

26374
FAQQIINAS

26375
GAKYIQNTD

26376
EAGPRKRDV

26377
PAHINDQYM

26378
MAHAPPNSD

26379
MAMWAVWNK

26380
MAEQMERDF

26381
SATNCDLYP

26382
HAKQANGMV

26383
AADIFAIND

26384
TATCGIEVE

26385
AAIGNQTLA

26386
AAQYMGIMK

26387
KAPWAPMDS

26388
NAENESSIG

26389
QAKMILDGT

26390
DMRKTWEAC

26391
TGRIKDAGP

36392
PWRCMQGYY

26393
MVRQCDYKM

26394
AIRTTMCDW

26395
KSRRGKLTW

26396
TDRQIDQYH

26397
TSRSEAADL

26398
IMRHCYSNG

26399
KRRDGTRSY

26400
YIRVGGNNE

26401
QERSMNAWW

26402
EKRESFDCG

26403
DYREKYKCQ

26404
DHRFKYYDH

26405
DIRTTGCQT

26406
DSRTETFFG

26407
EDRITECGA

26408
ESRNWDDTD

26409
ESRTMNMYE

26410
EVRSTRKNH

26411
FPRQASSQQ

26412
FWRQSGKLQ

26413
HCRWVKYHK

26414
HTRWPHIWH

26415
IERVEYAVN

26416
INRLLMTCD

26417
INRVEFTTS

26418
ITRMNSETV

26419
KIRVYCTPE

26420
LPRWADVQQ

26421
MERWWLVEG

26422
NCRPMEDMR

26423
QERITGMVM

26424
QSRTQGNDH

26425
SDRTAMCAY

26426
TCRWFAHYS

26427
TGREVDHTS

26428
VTRCIPTQW

26429
WCRPQQPDF

26430
YVRVIDDKW

26431
GKRVEDTSC

26432
WCRCKRFIE

26433
EIRQFVKGA

26434
DVRSHDEMT

26435
WKRTAGVHF

26436
GHRPKLCDQ

26437
GPRHIEEDG

26438
ENRSPNNPA

26439
ASRSCMTQE

26440
ECRMAVQSC

26441
NNRRQFGYV

26442
TCRYEELAD

26443
APRDMDACV

26444
PPRTVARFK

26445
KERMQVSKA

26446
TNRCHDDRG

26447
EFRCRLIGS

26448
GTRIYGKVS

26449
KIRDCDGNV

26450
TLRAEMSFH

26451
VDRLFRMEY

26452
KQRDWENFA

26453
SCRMVYGRA

26454
FFRICGSYI

26455
EKRLFDRTY

26456
ESRQVQDCY

26457
PWRWHKSAC

26458
MKRAPTLDY

26459
AKRECTHIP

26460
SYRALFNGW

26461
DLRWLPYDL

26462
IIRQSEQYH

26463
IHRVVCRDH

26464
EVRIQCKID

26465
WERCLELNQ

26466
EQRHISRAP

26467
DSRTQGGLC

26468
TQREVETSY

26469
KQRIAYEWQ

26470
NTRIVCDLS

26471
KTYTHECVA

26472
AVATTVMMC

26473
EKMTHTCLQ

26474
MQMTQEWMA

26475
YTKTYVDCF

26476
KSGTCGYGQ

26477
SKNTVGIGH

26478
TLTTDKTYS

26479
ACGTGVMAL

26480
YYDTISPDG

26481
QMLTTAHNL

26482
IKETWRVYF

26483
QLDTYMEGR

26484
ACDTPFQYH

26485
AEGTGMGNF

26486
ALNTQEMCN

26487
ANFTFVRSE

26488
ATGTGWLDN

26489
DFSTHPGTE

26490
DHMTPWERQ

26491
DMITGMQGS

26492
DPSTCCCFM

26493
DSDTMMNHN

26494
DWSTQEVNY

26495
EFHTKNRDM

26496
EMVTEDFWQ

26497
EWMTQCGWH

26498
GSHTCHLEV

26499
GSLTQNDNL

26500
GSNTIMTCL

26501
GTGTHEDYD

26502
IEGTCDRAG

26503
IQTTEMTSQ

26504
IWCTSCEIA

26505
KICTFGRES

26506
KIITFISPH

26507
LCCTEECVN

26508
LESTSMWMY

26509
LGITMCTTM

26510
LQSTGIFYA

26511
LSQTHQKLH

26512
LTPTINAWH

26513
MFCTSALRG

26514
MQETNQFMR

26515
MVATCFNND

26516
NMNTGQQPG

26517
NQGTCTQYS

26518
NQQTQVCMT

26519
PEATKPIKM

26520
RPCIYEYQD

26521
RWDTLYFDI

26522
SDETGVPAK

26523
SILTYQNVG

26524
SMATNHWAE

26525
SQFTQLTQA

26526
THYTDVLVV

26527
VCFTANLMG

26528
VGTTTMQSQ

26529
VHHTNTMAN

26530
VMSTDISQI

26531
VQTTYDRKG

26532
VRWTYMTNA

26533
VTPTLNRCQ

26534
YTHTPHLRE

26535
DMVTVNCID

26536
DSATFKSAD

26537
AVETGESAM

26538
QTNTMQMLS

26539
EIHTGRPGS

26540
EPETLWYVG

26541
ATDTYNLQE

26542
MEHTMDFGV

26543
EHITEQDLR

26544
ETSTHCCKV

26545
TYDTRNHCL

26546
PSNTGKFCQ

26547
KSQTVMYDF

26548
AECTEDNDD

26549
QNLTKEDQY

26550
WVPTPWRTH

26551
NWCTMYRYW

26552
AKPTHDVGS

26553
LFMTYSARN

26554
DRTTMVGWY

26555
GEMTRCFHN

26556
NDCTCRAIM

26557
IHFTSQSCL

26558
NSHTIMNGM

26559
YPDTADIFW

26560
SSMTAERFT

26561
SVNTMWYPD

26562
GRHTAFATG

26563
PWTTPENYT

26564
IWITCPYAA

26565
ACHTGQSGM

26566
QMVTQLNWE

26567
QWFTKGVGE

26568
QIKTHSNVF

26569
QTGTGHWEF

26570
NHWTTCDYS

26571
LMKTSPSYW

26572
NVWTNIAEM

26573
RQTTYDCLD

26574
GQNTTDRSN

26575
HVNTSCKQF

26576
NTHTGNDRL

26577
TYMTNLGDR

26578
KQKTQNRNA

26579
WKATPGGQC

26580
VTMTTKSVW

26581
TCLSVMNQY

26582
QSDAVHEPA

26583
YQERVRKDV

26584
DDLDVWQTS

26585
GCNHVHIGQ

26586
GIANVDDWE

26587
NQIDVQWLQ

26588
REVMVCILR

26589
LCHNVLDYY

26590
NQPCVNTYW

26591
TMAEVVINL

26592
QRMIVDALQ

26593
RMVRVSVHG

26594
FTVYVGARE

26595
ACAGVTWGI

26596
AFTNVTRWC

26597
ARNCVGSFC

26598
AVQSVLGCR

26599
DHCAVWNHE

26600
DISPVNRWH

26601
DSYRVLCLS

26602
DVIPVPKSC

26603
EQIHVTMAN

26604
EVYNVKGSA

26605
FLEFVHHGT

26606
FYSSVQRDQ

26607
GSHQVDILH

26608
MGDSVLTAH

26609
MMSNVPGEY

26610
NHVPVVGAM

26611
NTLLVDFDF

26612
PFTNVTRWC

26613
QNTCVIGAQ

26614
QQDAVCVSH

26615
QSFVVQGTY

26616
QWHAVTTNE

26617
RLDCVKESP

26618
RVVEVHAQV

26619
SDNFVITHS

26620
SQQFVHHIY

26621
SYYWVWMAG

26622
TNNVVIEWG

26623
TSMCVETQY

26624
TVIDVVRWH

26625
VFPDVSFWQ

26626
VIAQVFKYH

26627
VIGKVHHDN

26628
VTNRVAMPI

26629
VTSLVTNMG

26630
WYVPVNHLY

26631
WYVPVNQLY

26632
RWDFVWYIQ

26633
GVFYVGHWQ

26634
HGNMVLVIG

26635
NSVHVHASN

26636
NKEAVTGSS

26637
HPVKVDDFT

26638
SRLPVHECP

26639
WTLQVMKIN

26640
MRVQVFSDA

26641
SDSKVQWGM

26642
KSAQVYWHP

26643
QFMGVSSHS

26644
LFIKVKDTS

26645
HFHEVAWGS

26646
ALDRVSMAC

26647
WCEHVIMNP

26648
TKQPVHFLG

26649
YGNSVTCTS

26650
ESCLVRRGW

26651
EWEGVSQFR

26652
AEHMVVDHE

26653
TCNKVANMY

26654
GSYIVQNHC

26655
HHFHVNLDI

26656
KTSNVMDQM

26657
NVEDVGMIE

26658
VDMGVHVGS

26659
GGSYVSAPD

26660
VEDRVFACC

26661
WKAIVLEYW

26662
ASVMVAITG

26663
DSQSVGDSF

26664
INMPVSKDQ

26665
YRLHVWMKV

26666
YWEKVPCMK

26667
SGQPVDCMF

26668
FNWEVRAYI

26669
ACDQVNTVQ

26670
QTNQVVVVE

26671
FMMPVVDNY

26672
WCEAVGDYA

26673
SGFFVHTSN

26674
HFSHVDQCM

26675
NRCGVSKCC

26676
TSTFANEFT

26677
QRPIANDAM

26678
VQDKINYSY

26679
QTGHFNQRG

26680
EGTLWNVCV

26681
GVIPLNMPS

26682
MWTEFNEDG

26683
KMTNSNENS

26684
GRAHGNVCI

26685
QCEYSNCCY

26686
DVISRNGRC

26687
NMCEANHES

26688
EVAGENGNY

26689
YTGHTNSGR

26690
ACDFGNLGR

26691
ADVVTNINL

26692
AIEASNGGG

26693
ASESPNESG

26694
ATCQSNKCP

26695
ATTRHNTEA

26696
DGWSGNKMA

26697
DMKLQNDHP

26698
DMLKHNNAR

26699
DNNAMNDQQ

26700
DQQYSNVCC

26701
DRIWINWVG

26702
DVQKKNDKS

26703
EKDEINRAW

26704
ERFMANTGL

26705
ESSDTNYRQ

26706
EYHGLNLMA

26707
EYVWLNCGE

26708
FMHAGNLDA

26709
FVGCLNHYL

26710
GHIKLNVLP

26711
GIMQLNTEP

26712
GNCFQNKSN

26713
HPHVHNQKP

26714
HTVDGNVFP

26715
IDAVKNRND

26716
KEMAKNYEP

26717
KQDEWNCDR

26718
KQNCANLCE

26719
LLSDENTEC

26720
MKMEENSAS

26721
MLVQENNSY

26722
NLERCNTMM

26723
NQFFYNLQN

26724
NSCYMNSIE

26725
NYTQCNFSN

26726
QMAWENWFG

26727
QSCSDNYSG

26728
RCDQMNLHF

26729
RGDALNMVP

26730
SSFLLNHQY

26731
SWVFCNVGQ

26732
TTLNTNCSH

26733
YVYRMNGNC

26734
LMQWYNRCW

26735
VTEVFNQDV

26736
GNKLKNDQM

26737
YMGSTNIWM

26738
QLEECNPLS

26739
LVDCSNTCQ

26740
KLYGLNHSC

26741
SGALKNREE

26742
MIKLINCVN

26743
QPSRFNTAD

26744
MQEHKNFYG

26745
TTYPHNTHG

26746
IYKWDNKFH

26747
QRTQCNFSI

26748
KYWCKNEWD

26749
IEHCSNRYA

26750
MINWGNKCT

26751
VPLGFNFAG

26752
RENGYNATM

26753
QKSMLNSIQ

26754
EGQYWNYMT

26755
TFIYLNHFG

26756
DYFQYNEAP

26757
SCQLLNDGG

26758
AHEDPNTWC

26759
KNELENVIN

26760
VGGYSNQWV

26761
WDNWSNTQI

26762
HQYVHNWRQ

26763
VHHKTNRSY

26764
TMWEDNQNM

26765
DCDPYNMVD

26766
QISAWNEFF

26767
GNEDCNLAC

26768
MIMGYNFEF

26769
LPSHCNLGS

26770
TIDLSNKVT

26771
TTTITNRLC

26772
QCTAHNTCM

26773
DSMVYNRPQ

26774
KIQPTNHHE

26775
KGLIYNHHT

26776
NYGCHNCKY

26777
QEGRFAMSH

26778
RTGDAGMSP

26779
ILLPDDMLK

26780
HLDCLKMEM

26781
YSNSIQMSF

26782
YREHRTMSD

26783
GWYQKCMGD

26784
PTNYGWMWR

26785
GCYWNMMQP

26786
PNGHYWMIP

26787
ADYNNEMIQ

26788
AGSQMHMLS

26789
ASWSPQMNT

26790
ATGNKTMSY

26791
DLNQCQMYC

26792
EEVHSAMEG

26793
EGPALWMNT

26794
EGVHLPMGN

26795
EMEMEEMVG

26796
EQGYRVMSF

26797
EVFHDRMMG

26798
FCSEMVMNQ

26799
GNSKNDMHC

26800
GQCNTMMWQ

26801
HINAAVMHE

26802
IEPDCPMIP

26803
KIEAEEMHA

26804
KLVSKTMHH

26805
MMESALMDR

26806
NQDIMVMGQ

26807
NWACMGMGI

26808
PDWYNWMED

26809
PMKSPVMKD

26810
PQEEFMMTQ

26811
QELYLPMGC

26812
RFKHQMMMH

26813
RMDMKEMWG

26814
SQQLECMGP

26815
SRAEEYMPW

26816
TRVPSMMYK

26817
TYAANIMHV

26818
VCTHYVMQR

26819
VMKPCFMTP

26820
SKNLYTMAG

26821
LDGPIGMGT

26822
FCNYAVMSP

26823
TLPKMHMWP

26824
TKPLEDMPT

26825
ESAEPRMIP

26826
AEVPYVMQQ

26827
QHSRAYMWY

26828
TNFPTSMHG

26829
YISEYVMMV

26830
TTTNQFMFQ

26831
TVGIRHMKD

26832
QCFSTEMCQ

26833
ELEPDGMHV

26834
THARFKMKS

26835
YSVQAYMAN

26836
FQWRTEMNN

26837
PSIKHGMPK

26838
DVDFLKMHC

26839
IHSFMAMAQ

26840
ENCMSRMKP

26841
MLMHADMTQ

26842
FTEPMCMQG

26843
YHCCFAMQS

26844
PGGCYRMME

26845
SQKQFSMQN

26846
HMLHAIMES

26847
TNGVQIMHV

26848
ETEIRQMDG

26849
YSNSTIMNL

26850
YVKMCSMRN

26851
GTVECEMNA

26852
FWGMESMSC

26853
ASGHMTMCM

26854
NTSELLMAQ

26855
SEDCMDMYF

26856
STEFWVQHT

26857
WSSREYWHK

26858
TVMSAWSHM

26859
ETHYPYNHI

26860
FQIRFDRHH

26861
TSSHDEAHK

26862
TSYLRKIHS

26863
TRYRMRYHG

26864
GIWRTREHY

26865
RGSLMSFHE

26866
SYAAFLHHG

26867
LQFLTVVHL

26868
GVSRLPYHA

26869
AHFFEIPHS

26870
AILVEFPHG

26871
AWTPSHGHP

26872
DGCKMFNHC

26873
DGYIYHTHL

26874
DIGNQDRHS

26875
EDMPAVDHN

26876
EFDQYMTHC

26877
ERVPRYVHH

26878
ESVMQANHT

26879
EVCIYCSHV

26880
GEDEAWQHT

26881
GIVVADRHV

26882
GSLPWKNHV

26883
HIADKSTHR

26884
HILYAWWHS

26885
HSPFFMNHA

26886
HTYPLDHHF

26887
IYSDNQYHH

26888
KIVECVSHI

26889
KSMWSERHC

26890
KWWWAYTHK

26891
LIAMRIVHD

26892
LYCSPPNHC

26893
MEKRCGVHQ

26894
MIACDKVHY

26895
MMAKHTNHE

26896
NCVQWKYHQ

26897
NGCCLMTHL

26898
NYICTRKHP

26899
NYQNKLLHG

26900
QHMIREAHG

26901
QLSMLGVHF

26902
SCNMCVSHE

26903
SHKFMNYHW

26904
SHLNNITHQ

26905
SRNGSKRHA

26906
STKPWPEHN

26907
SWFECFHHE

26908
TIMMGVEHM

26909
VMFYTVHHE

26910
YSMEEHFHN

26911
YYVFMQLHA

26912
HWELIRCHD

26913
YEICKHGHF

26914
TCNVSWEHE

26915
HGNSFLVHW

26916
YITCQYSHH

26917
QVVIDPAHY

26918
RVWSLEWHL

26919
KYSPFQEHN

26920
YPQPCSCHS

26921
QVWSDGQHV

26922
SVTYHEWHP

26923
TCNVSWKHE

26924
MLFDFEEHN

26925
QTMCITIHP

26926
KTEKWASHE

26927
FCKCFMKHC

26928
QMSWGKYHQ

26929
KYINFDHHQ

26930
MKGYFVHHS

26931
NKPAMMQHE

26932
ITTDATGHF

26933
RDQKGLGHM

26934
MYLSTPVHE

26935
DVKYATLHD

26936
MYAGWLNHG

26937
AVIHEKPHG

26938
GTIRMPEHR

26939
QKHMILCHF

26940
QEAGYTRHN

26941
AKEHMPHHM

26942
TKQFFTAHC

26943
AFGLTTGHT

26944
YLESKTIHA

26945
TRDPFWHHD

26946
GVWMQGFHP

26947
AVQMQQQHI

26948
VKAWWHDHQ

26949
GIHQLIHHC

26950
TRQWMCVHC

26951
ILANPIRQI

26952
KHNPSRHDI

26953
NFYLSGFLI

26954
KQCSLAPTI

26955
GFHNTKLPI

26956
TILQQMEEI

26957
ACNPFWHSI

26958
AFTETKCSI

26959
AGSHFGRRI

26960
EVTRTQVQI

26961
FYLCNYGAI

26962
LEAKWQCSI

26963
LWMWQLSCI

26964
MKDRMHQVI

26965
NVAQPDFQI

26966
QITSGCHCI

26967
VTCPIQRTI

26968
VTHKLVDAI

26969
YPHKKCRCI

26970
YSLEKRFQI

26971
EPLYSWVGI

26972
ASIATMDRI

26973
TCGHTRFFI

26974
EHKGCMQAI

26975
VNDDYECGI

26976
NMPSLRTRI

26977
FKIEWSQDI

26978
EFVLGCQGI

26979
WDAYYTNGI

26980
QSMVACYTI

26981
ATNKYANAI

26982
WHFLMIFYI

26983
QQEAYILRI

26984
GGALARSSI

26985
EFDWLLGTI

26986
RRTDHGEPI

26987
WPNIKHQPI

26988
HQCQSRSYI

26989
EICMRSGFI

26990
IECSPRESI

Example 9
AAV5 Variants with Tissue Tropism in Skeletal Muscle

This example describes engineered AAV5 variants with tissue tropism in skeletal muscle that were discovered using the methods and systems described in EXAMPLE 1-EXAMPLE 2.

CNS, liver, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord tissues were analyzed to identify amino acid residues in the AAV5 VP1 581-589 region that drive skeletal muscle tropism. The results are shown in FIG. 18C which shows the positional frequency of each amino acid in the 581 to 589 region of AAV5 VP1 in skeletal muscle tissue as compared to all other analyzed tissues, wherein the capsid polypeptide has the sequence of SEQ ID NO: 2. The position (‘Xaan’, n=1-9) is denoted on the x-axis and amino acid residues are denoted on the y-axis. The frequency of a particular amino acid residue in a particular position is indicated in the heatmap as intensity. Darker squares indicate a higher frequency of a particular amino acid in a particular position and lighter squares indicate a lower frequency of a particular amino acid in a particular position.

The frequency of a given amino acid residue occurring at a specified position corresponding to position 581 to position 589 of SEQ ID NO: 1 (generalized in SEQ ID NO: 2) in variants identified in skeletal muscle over the frequency of that given amino acid residue occurring at the specified position in variants identified in all other harvested tissues was analyzed to identify a set of sequence rules for capsids that preferentially target skeletal muscle tissue. With reference to TABLE 17 below, and SEQ ID NO: 2 (AAV5 VP1), the following amino acids can, thus, be independently mutated, in any combination, at any one or more positions Xaa1-Xaa9, to provide a VP1 capsid with enhanced skeletal muscle tropism as compared wildtype AAV5 VP1 capsid (SEQ ID NO: 1), where skeletal muscle tropism here refers to properties that are preferred for skeletal muscle transduction over properties that are preferred for transduction of all other harvested tissues.

TABLE 17

Skeletal Muscle Tropism Rules

Xaa1 is selected from A, E, H, M, P, Q, or S

Xaa1 is selected from P or Q

Xaa1 is Q

Xaa2 is selected from F, H, I, T, or V

Xaa2 is selected from T or V

Xaa2 is V

Xaa3 is selected from A, G, I, K, M, Q, R, S, T, or V

Xaa3 is selected from A, L, P, R, or T

Xaa3 is selected from L, P, or T

Xaa3 is P

Xaa4 is selected from D, E, G, P, or S

Xaa4 is selected from D, E, or S

Xaa4 is E

Xaa5 is selected from H, L, M, P, or V

Xaa5 is selected from L, M, or V

Xaa5 is L

Xaa6 is selected from E, H, N, or P

Xaa6 is P

Xaa7 is selected from A, H, N, Q or T

Xaa7 is H

Xaa8 is selected from I, K, M, P, or W

Xaa8 is selected from I, P, or W

Xaa8 is P

Xaa9 is selected from A, I, M, P, or V

Xaa9 is selected from A, M, or P

Xaa9 is M

TABLE 18 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that were found in skeletal muscle tissue and comport to one or more of the rules provided in TABLE 17. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 28991-SEQ ID NO: 29990, as disclosed in TABLE 18. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 18

Sequences of the 581 to 589 Region

in AAV5 VP1 Capsid Polypeptide that

Drive Skeletal Muscle Tissue Tropism

SEQ

ID
581-589

NO
Sequence

28991
QAEIVCNSK

28992
QANGSMDHH

28993
QANWWVDAG

28994
QARTMDERP

28995
QAWEGCAGE

28996
QCAFIPNSA

28997
QCGNVMLMP

28998
CICHFLUCK

28999
QCMCVEIAA

29000
QCNAQAELF

29001
QCQLTVSMS

29002
QCQWQKMEH

29003
QECYEKMEC

29004
QEHCMNYGP

29005
QEMFIQARG

29006
QENLTNGDN

29007
QFIMLGETN

29008
QHVQIMAGQ

29009
QIEGVNHAN

29010
QIERYWHAT

29011
QIHGVTIWQ

29012
QIHIWDGQS

29013
QIHLRECSF

29014
QIRAMLDSS

29015
QISIADCYN

29016
QIWNEVHAQ

29017
QKNEYWTAF

29018
QKNLVPSYW

29019
QKSFPGDGI

29020
QLMVYMYES

29021
QMEYTWMGG

29022
QMFQDHNQP

29023
QMHHWDVWG

29024
QMQPTCGQF

29025
QNENVVWMS

29026
QPMHQQVTF

29027
QPMSFNWKN

29028
QPNYVEHCG

29029
QPTMHMPRG

29030
QPVVTWQSH

29031
QQITTEHSG

29032
QQLCQLDRK

29033
QQYLCMSSD

29034
QSCATTSAQ

29035
QSFQAPRLA

29036
QSHRVMDIP

29037
QSHYCHTFS

29038
QSMSYQKEG

29039
QSNLKSMYC

29040
QSSSMACDN

29041
QSSWAWMAE

29042
QTGNWWQYG

29043
QTIFWLNVG

29044
QTISGFQKD

29045
QTLHLHTNM

29046
QTMRLEPYH

29047
QTNLLVDTS

29048
QTTAILKLD

29049
QTTLKMKED

29050
QVETPWSGA

29051
QVQNVWCPK

29052
QVVTMPDPL

29053
QVWDMNDNA

29054
QWERGANYK

29055
QWLLLCHGN

29056
QYERLHTNM

29057
QYIPFERYW

29058
QYMTQAKKD

29059
QPSRFNTAD

29060
QKTANGWCH

29061
QMCVWMTRQ

29062
QAKMILDGT

29063
QTAEWYCGA

29064
QTNGHPYFP

29065
QFAQLWCAK

29066
QCTAHNTCM

29067
QAADSASAS

29068
QADRDGCYA

29069
QAGWINTLN

29070
QATFHATQK

29071
QCADKNVLE

29072
QCFNTCPCD

29073
QCKTVIMGA

29074
QDFQFDEVW

29075
QDTMMTYMS

29076
QDWFNLAHW

29077
QEEHQKFGM

29078
QEENENMGC

29079
QETQAAMRQ

29080
QEVTTTDPQ

29081
QFMVKNSNH

29082
QFNLTDYFH

29083
QFRGLEHMV

29084
QFVSTHHEA

29085
QGNTCHKDA

29086
QGRIYDQAT

29087
QGSWNFCPP

29088
QHAPLHNFI

29089
QHCPIVEQD

29090
QHREIVKLQ

29091
QHSTDSGDK

29092
QHVELQVMK

29093
QIACLQKQG

29094
QIHDQLKGN

29095
QIQEVQSSV

29096
QISIGCNSQ

29097
QISSSRDMY

29098
QITDWENFH

29099
QKDPLHVHS

29100
QKDQVWHAW

29101
QKEEQPKMG

29102
QKPELEQGP

29103
QLCNPLAPM

29104
QLPGIKMWD

29105
QMFGFQPGN

29106
QMHPMIRFF

29107
QMHSMTNCN

29108
QMRIWRMMW

29109
QMVPATPCN

29110
QNAFMDLLA

29111
QNCFRYECP

29112
QNETRQHCW

29113
QNMPVMQYR

29114
QNRPTECIG

29115
QNVKEYSAS

29116
QPADPNDGG

29117
QPAPGHRGQ

29118
QPILPNIWE

29119
QPQVIYDVR

29120
QQLFWDIGV

29121
QQPLMRTEE

29122
QQSTKMKWH

29123
QSDKKNRCA

29124
QSDQLSRIQ

29125
QSELGHELP

29126
QSLWQQLSM

29127
QSQPEWNEK

29128
QSSTDQEAW

29129
QSTENNWYS

29130
QSVQPCDDP

29131
QSVSQLTCC

29132
QTAATMTED

29133
QTAVKNIDN

29134
QTCNAEVFT

29135
QTCNCGKAF

29136
QTGQYATLK

29137
QTIPREAEC

29138
QTNLPVVKN

29139
QTQTWERQD

29140
QTWCKRIHF

29141
QTYMPHHAH

29142
QVCSMFGNA

29143
QVINVHGRW

29144
QVIRHECQY

29145
QVSNTATCL

29146
QVTQVQALG

29147
QVVEKHENR

29148
QVYHYCRRH

29149
QWHVGGTSA

29150
QWHVGGTSA

29151
QWRNPIYGP

29152
QWTQGYASP

29153
QYEWLNTCD

29154
QYFRFGFDP

29155
QYGNSLNAG

29156
QYVPAGYFA

29157
QADRIGQCL

29158
QANAMAEFP

29159
QCACTITMK

29160
QCASTMSMT

29161
QCELLNVLC

29162
QCIEYEATE

29163
QDGMEAKNR

29164
QDIKKWCSE

29165
QGFMEQQHT

29166
QHQAEMEGF

29167
QHTEMDCSN

29168
QIGGFQYYY

29169
QIKLQQWYD

29170
QKDEYLNWH

29171
QKDMHAYPL

29172
QKPRWFCCP

29173
QMSSGEWVR

29174
QNELRESYE

29175
QSKPMEHVQ

29176
QTDWSMFLP

29177
QTYFPWHWF

29178
QVCSRWTKS

29179
QWVEWPFPC

29180
AVEKVMEVQ

29181
AVKLNQTYK

29182
AVNGNVYGR

29183
CVEGMVFVP

29184
CVGQYNNDM

29185
CVLRQYDGG

29186
CVNVRGRSS

29187
CVRQKAETE

29188
CVTAASPWN

29189
DVAERHMHE

29190
DVCMATIGQ

29191
DVDPLPCEP

29192
DVGYQMKKQ

29193
DVICYSDHR

29194
DVISMSLMG

29195
DVLIDHDFF

29196
DVMHFLDLA

29197
DVQNMDIGV

29198
DVRDDEFGN

29199
EVCGMECSG

29200
EVDDKPSGH

29201
EVEYEAKTV

29202
EVFVADNNI

29203
EVGAWGASY

29204
EVKAANWQV

29205
EVQHHYVSG

29206
EVRMCNATH

29207
EVVKKPWDG

29208
FVFTHIDDY

29209
GVEPISRAA

29210
GVGQKCGHQ

29211
GVLAQQAQW

29212
GVLLPHHWM

29213
HVNQWGRYG

29214
HVQSIIHAE

29215
IVDWSNVLH

29216
IVGMANQEH

29217
IVHSLDAPG

29218
IVSFGYAQG

29219
IVSYLKTND

29220
IVYDMMADV

29221
KVDTQWKSV

29222
KVRPPCIYI

29223
LVDPHRPMM

29224
LVNHGDSYC

29225
LVTGNSWEQ

29226
MVIGTSLLA

29227
NVMGHDWVH

29228
NVQWFEQNI

29229
NVVWGMLYG

29230
NVYINLASS

29231
PVPHNYQWW

29232
PVRHCSVWF

29233
RVCTFCCCF

29234
RVDFNVYCK

29235
RVFMLEMYA

29236
RVNMNDIGE

29237
SVAMRQLDN

29238
SVFWPEHNP

29239
SVHICEDGW

29240
SVLSFVAQK

29241
SVNPYPHGI

29242
SVRTHMHLP

29243
SVTFDVGGP

29244
SVYICNMDQ

29245
TVAVNEHLA

29246
TVFVYDNPG

29247
TVLDAWTAE

29248
TVNSNRTGD

29249
TVNVFPIHE

29250
TVYDVNAFS

29251
VVMDAHCNL

29252
VVTDYMVGA

29253
WVKFANDCQ

29254
WVRMRNLEK

29255
YVAGLFNNR

29256
YVKFSVNIN

29257
YVQLRIADD

29258
YVQTTAHSS

29259
MVTERKPWS

29260
SVLHVGALA

29261
WVHPYSYCA

29262
DVKSWAVCD

29263
LVVTKFTYV

29264
RVNGSTCRH

29265
AVESSKGYV

29266
AVGCFGKEP

29267
AVMEYILYK

29268
AVNDQWQWG

29269
AVQLGNSMF

29270
AVSAPLSKE

29271
AVVQGMLYP

29272
CVAHFDQCF

29273
CVDYHRREQ

29274
CVESGTGHQ

29275
CVIHRMDMP

29276
CVMNVIWDA

29277
CVNETDTAA

29278
CVSYNPSIR

29279
CVTINTISG

29280
DVNCCITCG

29281
DVNTYANRD

29282
DVRAMTNDW

29283
DVVEKCSWF

29284
DVVHMWWWS

29285
EVACYDMTG

29286
EVAMMINNN

29287
EVGTECQWL

29288
EVGTNHKQH

29289
EVKMFYKQY

29290
EVLDHISAG

29291
EVREEGVLT

29292
EVTHQLKNA

29293
EVTVFIAPW

29294
EVWNEESMF

29295
EVYIKQQYY

29296
FVPPSCYNY

29297
GVAAQPCSP

29298
GVAPKPDFE

29299
GVASCNRFD

29300
GVMPKYFFG

29301
GVNAKQQTR

29302
GVYFMCDFM

29303
HVQTAEWIP

29304
IVDWPANRY

29305
IVIGRWYSC

29306
IVMRALANN

29307
IVMSFHRNH

29308
IVTDCRDGL

29309
IVTTKQLCR

29310
KVDDHICPG

29311
KVGWIEWYA

29312
KVYPLDFKM

29313
LVDGCVWMH

29314
LVDYTGNED

29315
LVFYTSVRG

29316
LVTKEDCAY

29317
MVAVDGTEP

29318
MVDMKQLMF

29319
MVEANEHAM

29320
MVEIHPDYD

29321
MVHLRGDGE

29322
NVDSSPFLT

29323
NVHGIMWDN

29324
NVQTKMHLP

29325
NVTPHEFQA

29326
NVTTIEPAH

29327
NVVQVTQNA

29328
NVVSKDHGG

29329
NVWHGYWLN

29330
PVAHVPFGM

29331
PVTAPGFAH

29332
RVYNPMYWH

29333
SVAQEMEST

29334
SVCEDAFFD

29335
SVDAQAFVY

29336
SVDQESAYR

29337
SVERHGSDK

29338
SVHQWCDVT

29339
SVIAFHIHV

29340
SVLHTMMVK

29341
SVNEVIDQV

29342
SVNQESAYR

29343
SVNSKNHTQ

29344
SVSYGNHSV

29345
SVTDRDHQY

29346
TVAPQSSLW

29347
TVCMDANFK

29348
TVKDLRVVT

29349
TVLWKHDKA

29350
TVSQCSFFP

29351
TVTLSEKWH

29352
TVYGFPNEV

29353
VVGVAPVMP

29354
WVKAVTLEQ

29355
WVVQTFTFD

29356
YVDCRNDAG

29357
YVELYEVDQ

29358
YVRDTALCF

29359
YVTSALDTF

29360
CVSFDACFA

29361
DVDRLCSIP

29362
DVSQTNFSE

29363
DVVWSTKDQ

29364
DVWSSISDS

29365
EVDGGGKMQ

29366
EVFLAGLMS

29367
EVLLAGIGP

29368
EVTPMYCSD

29369
FVRGNLQTP

29370
GVCHNTGCT

29371
HVFELDRTC

29372
HVIPDPRPA

29373
IVETDGSLM

29374
EVKFECCNH

29375
LVYHSFVHE

29376
MVCREQENG

29377
MVEVATMPH

29378
RVDDYERTF

29379
SVIHYSWAL

29380
SVSAWNPMS

29381
DKPFEWDLL

29382
EQPHDPSST

29383
FDPNRSQDV

29384
GEPKCVMME

29385
GGPDIKHYI

29386
GGPSEQPRP

29387
HHPKFAEKG

29388
HHPQVLRGM

29389
HNPVWVQQE

29390
ICPLEQYEM

29391
IGPMGQYGR

29392
IMPEAKNTP

29393
KCPDEMNNI

29394
KTPFTRWLR

29395
KTPLMVASI

29396
MLPSFGDEM

29397
NGPQGLMAC

29398
NNPTNTEAR

29399
PCPDHHRWL

29400
PLPCPRSGY

29401
RIPPMMWIL

29402
SSPPWGHPF

29403
SSPSHAEYL

29404
WMPLYCMHV

29405
WQPSHAYHF

29406
FCPCGQEMC

29407
SWPLPERPS

29408
LSPWPQTDQ

29409
EQPNMGDYG

29410
MLPLEMKNL

29411
FPPREGPEW

29412
AHPHCCRHP

29413
CAPQFNWQY

29414
CQPRYHWCY

29415
ESPNWGYMD

29416
GQPQQIWQA

29417
HHPHQANIY

29418
KDPLGESKW

29419
KMPLKQADF

29420
MFPQAPADG

29421
MFPVCQDRD

29422
MGPMKPMMD

29423
NPPLCQEYG

29424
PGPEQITAG

29425
PPPWFQSRS

29426
PQPCSWPAQ

29427
SGPWCGSAW

29428
SSPNVHDLD

29429
SWPFSWLIV

29430
TKPDSANWD

29431
VNPQEEFFD

29432
VQPPYIAMV

29433
VSPRDLHHM

29434
YFPQESENS

29435
YQPNWRSQT

29436
AHPWCLTPN

29437
DNPWLLTFR

29438
KQPISASPI

29439
MDPLGPAAD

29440
NEPQICRKC

29441
NQPGRFRTV

29442
VPPRMTIEL

29443
YSPTTNGGP

29444
ALSEHFWNV

29445
ANVELMTIK

29446
ATHEQGGQL

29447
CFCECSFCE

29448
CFDEFMLWG

29449
CITELINYT

29450
CLAEQITCQ

29451
CRSEGHIHT

29452
DGMEWDLCG

29453
DHVEIQHMS

29454
DTTEQMQIY

29455
EACEGAMLE

29456
ECEEPSLMK

29457
EEVESGHFG

29458
EILEEQESC

29459
ELIEKVYRW

29460
FTREDKDRC

29461
GAVEKMVQE

29462
GMEEVEREG

29463
HGEEIHWKD

29464
IWTEAAENC

29465
KGEEMYECG

29466
LREEYKLAG

29467
LTCENQQIP

29468
LWTESVMGN

29469
MALERCDVD

29470
MEEEYMFTA

29471
MMSEQCHLT

29472
MNIERDRNH

29473
NTCEPVQMI

29474
PTLEFLNLM

29475
RPNEPLDYS

29476
SKREIENCH

29477
SLCEMMQPH

29478
SMTEIVVAW

29479
SSNEGGCWM

29480
STKEDFDMF

29481
STVEKMQTL

29482
TTDECHGHP

29483
TTVEMECGM

29484
VEEERPFWS

29485
VRGERLSDL

29486
VTIERNIWQ

29487
VTRELWLPQ

29488
WCTEFSTRQ

29489
WNVETFLGY

29490
YIAEMEFMN

29491
KQHELMVRV

29492
PKHEDMRWW

29493
NQREWHGLA

29494
SECEVLCSL

29495
PHAETHWYT

29496
AAMESCAEI

29497
AEGEIEMWT

29498
AFVEFSDVT

29499
AGCETWNAP

29500
AMHEATDAQ

29501
AQSEWGSAT

29502
ARDESAEEH

29503
ATHEMHLDA

29504
CKKESGSYE

29505
DCDECQDDQ

29506
DKDERAEGM

29507
DQWEYEVQF

29508
EELEQQMCT

29509
EMHEWSRAN

29510
ESEEMHAYE

29511
GKIEARETD

29512
HMFERPRTQ

29513
HPSEECSWR

29514
KLVEMQESK

29515
KWCEVLWQP

29516
LEGEAYAQE

29517
LFSEFFEYE

29518
LSNEMTIDA

29519
LTIENPHDQ

29520
LYKEIGENI

29521
NAEEWWSCD

29522
NCVEAGSFA

29523
NMMEYERFS

29524
NNMEDRWGT

29525
NSNEHGAMW

29526
PTCEVPENC

29527
SFIEFPAMT

29528
SIDEQTFPP

29529
SKEEVQYIF

29530
SWTEISSVK

29531
TAAEMIYEY

29532
TWDEQTAQV

29533
TWRECQSSH

29534
VGNEDKCDP

29535
VMIEKTMMH

29536
VPVEVHKQA

29537
WGAENHGPW

29538
WKDEYTVDD

29539
YAREFGNMY

29540
YTDERAMLN

29541
CDAENSDGG

29542
CLDEPRTDF

29543
GHEEYWEHV

29544
HHREQKDHS

29545
HSAEQVLME

29546
MIQEIKEDN

29547
MWCEVMHCS

29548
SCRECDECP

29549
TPDEQSHYD

29550
TTAEYGMSI

29551
AFHHLGEFQ

29552
ALMPLTRWN

29553
AMHSLHYMF

29554
ASEYLASPM

29555
ATTLLHYKN

29556
AWWWLNAQG

29557
CNIQLHTHY

29558
DRVRLMMVA

29559
DWGYLRIWC

29560
DYSKLDVHY

29561
ERSTLPMSR

29562
FADYLAAAS

29563
FPYHLWVGG

29564
GAEKLHLVH

29565
GGIDLTLMM

29566
HADCLELSC

29567
HGEDLNEFL

29568
HHDDLAPWQ

29569
IICTLSLDS

29570
ILNVLESWE

29571
KERRLWSGV

29572
LFDRLKLNE

29573
LRSMLASYD

29574
MHGHLTQSI

29575
MSISLWGNA

29576
PENPLVTMS

29577
RNTCLNTGD

29578
SLSRLYWSN

29579
TIRSLEMVW

29580
VGIPLQAHC

29581
VMDGLGLVL

29582
VIGFLMIPT

29583
VTRDLQKEQ

29584
VYDYLINGA

29585
WHGRLMITC

29586
WYQFLDNQL

29587
YACLLDTLG

29588
YSYSLMSGW

29589
DKINLAVSY

29590
HMEWLEEQA

29591
ITVALNQGW

29592
KCADLIIRS

29593
HATGLVNFM

29594
ATMNLSWGL

29595
TYESLQNSW

29596
NLSNLVSCR

29597
ALDALHYCS

29598
ANIGLETGV

29599
ARVDLMSHQ

29600
AWTGLHVCN

29601
CRDGLPHTV

29602
CTFWLYFFR

29603
DFNYLNWFC

29604
DILDLVNSF

29605
DIYQLHPSY

29606
DLFCLNYDF

29607
DQHKLTVEN

29608
DTDTLMNSG

29609
DTELLLVQE

29610
DYMLLQGVY

29611
ECHLLEEQR

29612
ECRSLCFQH

29613
EDRDLSWTF

29614
EGMILDHTP

29615
EKANLTYGV

29616
EKRSLPACM

29617
EPHSLDICL

29618
EYFVLHVYD

29619
EYNSLLCHF

29620
FRKLLTRRF

29621
FWCYLVLMV

29622
GEDTLAAWG

29623
GGEFLAVAY

29624
HHMQLLRSG

29625
HPGKLFETH

29626
IGTALERQG

29627
KIVVLAKGE

29628
KQTVLHVHH

29629
KSLPLHDHF

29630
LFTCLQEQE

29631
LSGWLSQFG

29632
LTTHLPEWN

29633
MKTVLYVGD

29634
MNLHLPQWG

29635
NFHSLESTP

29636
NTVTLVQHE

29637
NYGKLGKSY

29638
PMDSLTRVY

29639
PWQNLAVNH

29640
RWQDLLNTG

29641
SQMHLFTSW

29642
SSFELLDHQ

29643
TFNKLSCMW

29644
THAMLDRNF

29645
TQEALMDGQ

29646
TQTNLAECA

29647
TSQLLMHGN

29648
TWGLLWKQY

29649
VKLKLSEGQ

29650
YTYALAHAT

29651
YYNNLNIHT

29652
AHAMLDRNF

29653
DYKYLSCQY

29654
EEMHLDNAL

29655
ENHNLENHG

29656
EQTVLHVHH

29657
HNIDLQNWD

29658
IDTLLYHAI

29659
KCEWLQWVP

29660
KPHPLDPFR

29661
LCSELVHWS

29662
MHGPLYCAE

29663
MWGGLDMVY

29664
NRAQLCRCN

29665
TAMWLHNMV

29666
TAVALFEMD

29667
TNVGLIDGS

29668
VQEWLGLYC

29669
CDGHDPFWY

29670
CEEKTPVWC

29671
CIYLKPDYQ

29672
CLQHMPRLQ

29673
DARSSPDCL

29674
DPWHEPSMG

29675
DRHTKPNST

29676
ETMYWPAMD

29677
FFRDCPYEY

29678
GNNRNPTYW

29679
GTSDFPEAV

29680
HNLKIPMME

29681
IASHNPSKQ

29682
IDFRPPNNT

29683
IPNGPPNNQ

29684
LGFRKPFTR

29685
LQYTFPEGQ

29686
LTEATPKWA

29687
MFQYYPYCD

29688
MGEDRPSMT

29689
MHFKYPQGG

29690
MPDMDPAAL

29691
PQHISPDPT

29692
PYDMIPCNR

29693
SAAFQPRAS

29694
SRQPVPLSK

29695
STARQPALE

29696
SWYMPPMYC

29697
TIFQPPFHY

29698
VMDTTPLHF

29699
WAVLGPQKF

29700
WSGFHPPTM

29701
WTVLGPQKF

29702
YANFSPLWS

29703
SNTPMPYHG

29704
TFVRKPSLM

29705
KDGRNPHCL

29706
TCEDDPSLR

29707
CWMIVPAWH

29708
AAEPIPTWP

29709
CHCMWPSGV

29710
CSENEPFHR

29711
DMGHCPHDM

29712
DSDNIPYCP

29713
DSWRFPGHP

29714
DSYVTPYFT

29715
DTGSTPVGT

29716
EINPEPNWK

29717
FKLWNPDSI

29718
FWRSVPEYW

29719
GISSWPSAH

29720
GPNTHPHMA

29721
GSSYVPICF

29722
GTDACPLHL

29723
GTNGVPKVN

29724
HDYPTPKNW

29725
HMTFKPTAQ

29726
HQYNEPHVS

29727
ICTNYPKMD

29728
IHLMKPAHI

29729
IHNKFPLTM

29730
LKDFDPAIH

29731
LKVLWPTEM

29732
MHLRDPHQH

29733
MMHGYPFYP

29734
MNWLVPYSD

29735
NAMAIPKEC

29736
NDAGEPTRE

29737
RAMMRPHGE

29738
RMEYPPEQF

29739
SCTHDPTQL

29740
SSQPVPVPV

29741
STYHTPFNN

29742
TCVMEPCIC

29743
TIQRYPDYG

29744
TNQPFPSSE

29745
TQNLEPHWN

29746
TSDQYPSSY

29747
TTARRPTYD

29748
VAMAHPMTV

29749
VAVDSPHEM

29750
VLGRSPQVM

29751
WTYAPPLNY

29752
AHSHMPVCL

29753
CKDLSPYKF

29754
EEMQYPMFM

29755
ETCKMPLSD

29756
HHELDPDWE

29757
IPHNEPKAG

29758
KECFIPCAF

29759
MTRIRPDAN

29760
WKAPGPITR

29761
AEHSPHHDF

29762
ASALHCHNQ

29763
ATADNHHPR

29764
CSSLYSHMP

29765
DMCKYSHFV

29766
ETHTRAHDC

29767
ETIFVDHLN

29768
FNIHICHFI

29769
GEASMEHMQ

29770
GRIFSKHPD

29771
GSNQEQHGN

29772
GYQPDQHCP

29773
HRLIEQHDT

29774
IMMLMSHLE

29775
ITCKFTHNG

29776
ITQHMNHFQ

29777
ITSWMAHPQ

29778
KEAVMNHWT

29779
KLFPWEHFG

29780
KLMFWIHHT

29781
KRMQPEHCA

29782
KTDHERHAY

29783
KTWLPYHTW

29784
KWLWNWHDW

29785
LCLSACHFE

29786
LHDNYCHET

29787
LSQDVHHDT

29788
MAFFSMHQG

29789
MCTDYGHEG

29790
MLKCNFHHQ

29791
MPEPREHTR

29792
MSCPHFHVG

29793
NFEVCQHCE

29794
NHQNWMHMD

29795
RFILELHDS

29796
RPVNGVHLH

29797
SAGWIMHNS

29798
SWTWDWHVA

29799
THQADEHDA

29800
TTIQSNHCK

29801
TTMQQGHDL

29802
WSANQMHSH

29803
YIERTNHTP

29804
FRDVCMHDV

29805
FANDFTHNM

29806
GQLSTQHHM

29807
ASKQGTHDY

29808
AAEPYKHSK

29809
ASNTYDHSL

29810
CAMPGSHQI

29811
CDVKDAHHD

29812
DGCDMTHTR

29813
DGWLYQHTY

29814
DTMSKNHCT

29815
DTYAAGHSA

29816
EDTLTLHEN

29817
EFTLFQHEN

29818
ENVDRQHHM

29819
EPNMCCHHR

29820
EYTDVMHDA

29821
FAAVCHHPI

29822
FDTFSVHRL

29823
FTDCMGHTM

29824
GAYQPCHYP

29825
GSWHCVHCP

29826
HAAPQCHYS

29827
KMEFQLHPG

29828
LDGAMSHMC

29829
MHVTPRHPP

29830
MQLIDHHGP

29831
MSKVDRHCA

29832
MTMLGGHCI

29833
NCMYEMHWQ

29834
NFYDSLHCF

29835
NGNLTAHPN

29836
NIELRDHGQ

29837
PCHRKKHPK

29838
RCGRTAHNH

29839
REWYHCHYM

29840
SKYPINHVQ

29841
SSEFAVHCG

29842
TINVKHHFG

29843
WDWFSYHAE

29844
WEAKCAHSV

29845
WPVLNDHEH

29846
WPVLNDHGH

29847
YHEKSCHFG

29848
CQVSYQHYP

29849
DGTVCVHMG

29850
GAESCCHFG

29851
GELMHRHMD

29852
HTQHHSHHL

29853
ISSNFDHFE

29854
KMQNCAHQH

29855
LSSLHDHPE

29856
NTQWEAHSP

29857
SFSSNAHLQ

29858
STECYYHGI

29859
TSTHIEHDT

29860
YRVNCGHPN

29861
AKDLISKPH

29862
AMCNDIAPW

29863
DILPKVYPE

29864
DLEWQHCPP

29865
DMEPQYWPA

29866
DTLGPEAPH

29867
EAMMMYQPN

29868
EIMAAWTPP

29869
EQGSVWTPP

29870
EWFHQMSPH

29871
FMEDNTKPR

29872
HTIWSHDPW

29873
LATQICFPT

29874
LRMDCVPPY

29875
LTGPNLSPA

29876
MGNRSGSPS

29877
MPSHWYRPI

29878
MTHKDIGPS

29879
NFCKGMNPY

29880
PATKTHDPF

29881
SALNAAPPP

29882
SWCRYHEPL

29883
TFMHESEPH

29884
WQYTQNCPA

29885
YLGWICAPC

29886
YCECGHWPN

29887
MPWLDKPPC

29888
CQMLCRPPV

29889
PIWQGYWPW

29890
YDYWMQAPT

29891
EWVVQQIPQ

29892
ADHWEKGPF

29893
AIDPINRPN

29894
AMEKPTLPI

29895
CTNFVNRPG

29896
DIVNHESPG

29897
GDESGLVPE

29898
GNESGLVPE

29899
GPKWGRSPL

29900
HDAPMFTPG

29901
ICMICATPE

29902
KWEHGFMPF

29903
LCKVGMEPE

29904
MEETVTRPG

29905
MEKRKGQPV

29906
NDLSPAEPT

29907
NMAMAKQPG

29908
RCSKVKWPA

29909
RFVVASYPY

29910
RHVSIMPPV

29911
SAVNKCVPG

29912
SETRNNFPY

29913
SIMRCKFPN

29914
STGPELDPP

29915
TDCCVGPPQ

29916
VGNVIARPV

29917
YCVCIQRPF

29918
GLIKIVKPW

29919
HEHTAFRPE

29920
IQIRANCPN

29921
MEGHTGNPC

29922
TPCIWHSPF

29923
AMEIQYMSM

29924
ASTNEEKSM

29925
CIEQKHLCM

29926
EAGIRAEDM

29927
EMSSSNMAM

29928
FFCKYNBIM

29929
FHHNSRLDM

29930
GGHSMTKAM

29931
GIVRQCCIM

29932
GIYYEQDGM

29933
GRRAELGLM

29934
GTMQVTGAM

29935
GYSSTHVQM

29936
IAVQTGSDM

29937
KCMPNVGWM

29938
KLKPICMKM

29939
LCWMAGAQM

29940
NAICTLEEM

29941
NCEFWKMFM

29942
NCVQTNSEM

29943
SADSEEGHM

29944
SKMSNQDAM

29945
SKSDVQSMM

29946
SPLNRNCLM

29947
VWLQSLDRM

29948
WMGHKHKQM

29949
MLTPVQATM

29950
KTSNVMDQM

29951
AKCDVTGGM

29952
AYMAYHTEM

29953
CTTASMSYM

29954
EIGSRNRRM

29955
ERSFHNSIM

29956
ESRIGQECM

29957
EYFNILRNM

29958
FATWHLYDM

29959
FDCWANNNM

29960
FIWDCGDIM

29961
FKTFGSREM

29962
GDDHRVLTM

29963
GGYHKFPVM

29964
GNDIFMVIM

29965
GNTPCRGLM

29966
LIACRENEM

29967
LKATYEYVM

29968
LNDIKWYVM

29969
LQIRAWEYM

29970
MLKVQCTHM

29971
PQDFKHNYM

29972
REFCGALMM

29973
RFGREAGGM

29974
RFTGHMGAM

29975
RHESINRYM

29976
RSDLKVCAM

29977
RYNLQIMWM

29978
SFMLKWWAM

29979
SMFGDRQAM

29980
SNCDNWSAM

29981
SPLHYLVQM

29982
SYRPFRKMM

29983
TLMQPGYQM

29984
VPDHIVGMM

29985
VSSSDGWNM

29986
WTAHMYTNM

29987
WTYHSMAQM

29988
WWWGNFMFM

29989
YCNQQQWTM

29990
YEQVITVMM

Example 10
AAV5 Variants with Tissue Tropism in Spinal Cord

This example describes engineered AAV5 variants with tissue tropism in spinal cord that were discovered using the methods and systems described in EXAMPLE 1-EXAMPLE 2.

CNS, liver, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord tissues were analyzed to identify amino acid residues in the AAV5 VP1 581-589 region that drive spinal cord tropism. The results are shown in FIG. 18O which shows the positional frequency of each amino acid in the 581 to 589 region of AAV5 VP1 in spinal cord tissue as compared to all other analyzed tissues, wherein the capsid polypeptide has the sequence of SEQ ID NO: 2. The position (“Xaan”, n=1-9) is denoted on the x-axis and amino acid residues are denoted on the y-axis. The frequency of a particular amino acid residue in a particular position is indicated in the heatmap as intensity. Darker squares indicate a higher frequency of a particular amino acid in a particular position and lighter squares indicate a lower frequency of a particular amino acid in a particular position.

The frequency of a given amino acid residue occurring at a specified position corresponding to position 581 to position 589 of SEQ ID NO: 1 (generalized in SEQ ID NO: 2) in variants identified in spinal cord over the frequency of that given amino acid residue occurring at the specified position in variants identified in all other harvested tissues was analyzed to identify a set of sequence rules for capsids that preferentially target spinal cord tissue. With reference to TABLE 19 below, and SEQ ID NO: 2 (AAV5 VP1), the following amino acids can, thus, be independently mutated, in any combination, at any one or more positions Xaa1-Xaa9, to provide a VP1 capsid with enhanced spinal cord tropism as compared wildtype AAV5 VP1 capsid (SEQ ID NO: 1), where spinal cord tropism here refers to properties that are preferred for spinal cord transduction over properties that are preferred for transduction of all other harvested tissues.

TABLE 19

Spinal Cord Tropism Rules

Xaa1 is selected from A, C, K, Q, R, S, or W

Xaa1 is selected from K, R, or W

Xaa1 is K

Xaa2 is selected from H, I, K, L, T, V, or W

Xaa2 is selected from H, I, or T

Xaa2 is I

Xaa3 is selected from C, F, G, H, I, K, N, or R

Xaa8 is selected from F, I, or R

Xaa3 is I

Xaa4 is selected from I, M, Q, S, or V

Xaa4 is selected from I, M, or V

Xaa4 is V

Xaa5 is selected from H, K, Q, T, W, or Y

Xaa5 is selected from T, W, or Y

Xaa5 is Y

Xaa6 is selected from H, L, N, Q, R, W, or Y

Xaa6 is selected from L, N, R, or Y

Xaa6 is Y

Xaa7 is selected from D, H, P, Q, or R

Xaa7 is R

Xaa8 is selected from D, F, L, S, T, or Y

Xaa8 is selected from S, T, or Y

Xaa8 is T

Xaa9 is selected from C, I, N, P, R, S, or Y

Xaa9 is selected from I, P, or R

Xaa9 is I

TABLE 20 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that were found in spinal cord tissue and comport to one or more of the rules provided in TABLE 19. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 34991-SEQ ID NO: 35437, as disclosed in TABLE 20. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 20

Sequences of the 581 to 589 Region

in AAV5 VP1 Capsid Polypeptide that

Drive Spinal Cord Tissue Tropism

SEQ

ID
581-589

NO
Sequence

34991
KYNMLYHMD

34992
KGESQCTSQ

34993
KGQHKENRS

34994
KWHGGPGVV

34995
KRMLYPMCF

34996
KATNADGET

34997
KQSDKWGDD

34998
KIMHSWYND

34999
KKEQDHWDK

35000
KASLGACVE

35001
KGGFYGESA

35002
KQNPSMRQD

35003
KSFSWDSSA

35004
KFLIAFEVT

35005
KEFMPCCHR

35006
KEGITGLEL

35007
KTCTEWPGE

35008
KVNRSNRIW

35009
KVTNYYHSG

35010
KYSPTPFMY

35011
KCEGEEVGR

35012
KQEYIMRQR

35013
KWDDMHTQM

35014
KNETFIAST

35015
KAVTRADAS

35016
KVAEGQNML

35017
KHLDMTSCH

35018
KATRDPKFP

35019
KTLPCSNSE

35020
KFNTKDQLM

35021
KGGLSTTNA

35022
KIQPTNHHE

35023
KTGWDSHWS

35024
KGCVPWSKI

35025
KMEVNMSEL

35026
KTLHMLESK

35027
KTGATNLAW

35028
KCRYKDCQC

35029
KSIMMSQLD

35030
KSGMIQEET

35031
KAIPQGSSC

35032
KGQRRDSVS

35033
KIEYGHWIH

35034
KAHDITWNL

35035
KSAQVYWHP

35036
KIQPMMVKA

35037
KEFQLQMEP

35038
KSTMLEHHD

35039
KRFVDMPEM

35040
KYSPFQEHN

35041
KNCIMLASN

35042
KSFQEQGGA

35043
KTMGIGGGP

35044
KVEGYGWTR

35045
KPLTYTVGS

35046
KQATNNNGW

35047
KVEIINWHP

35048
KSEHHWINA

35049
KGNPLDGDT

35050
KCDDRNTQH

35051
KLMYKVMSC

35052
KQKTQNRNA

35053
KCADLIIRS

35054
KHEPMCVQL

35055
KIGCMYSSQ

35056
LIAAQKNFK

35057
QIMQKYQYS

35058
WIPAMTYWC

35059
WIPAMTYWY

35060
WINMPHDRH

35061
DIVCLMNST

35062
MIRPLTHCK

35063
FIIRCMLIW

35064
TISSVFLLD

35065
EICQHMGMA

35066
SIVNHNEAR

35067
GIELPVKHH

35068
EICSRGRTI

35069
GIKYKDVLS

35070
GIKFFVDNC

35071
GIIERYGDC

35072
NICAKHRMS

35073
RIQKLKWCQ

35074
SIDVPPTAN

35075
TIVWSECAW

35076
NIDAVERIW

35077
TIGWKELNE

35078
TIMFGCLAE

35079
GIFMEVAWC

35080
RICPLFCIK

35081
LIQSYPHQC

35082
MISAPPDTC

35083
QIGPKAFGW

35084
DIACLDQWQ

35085
GILVQGGSM

35086
TIRPHHAKS

35087
AIFHACNSG

35088
RIVDGNACG

35089
CIHSQGMVC

35090
QILQWDCPS

35091
QIYSLASTW

35092
SIKFHERQH

35093
TILQQMEEI

35094
AIGPDRSNN

35095
TIHPAWVTL

35096
QISMKGEHG

35097
EIIQAHLIN

35098
CIIGRQEWL

35099
TIWVHEYGM

35100
SIAQDQVWS

35101
PIWQGYWPW

35102
IIKFLWRPT

35103
DITSLGCIM

35104
WINEANCLM

35105
GIKKPNEDN

35106
EIRQFVKGA

35107
TILPKKTLC

35108
DIGRFTAYY

35109
TISQIPWVR

35110
TIVVIPIKS

35111
LINSIVCGD

35112
GIPLWVFID

35113
FIDEKQCHE

35114
DICQKQSPN

35115
NIGVIPHIQ

35116
CIDVLFKHL

35117
QIIYRCRST

35118
EIHMFGQSE

35119
QIHSEHCAT

35120
HIHMSDRGA

35121
AIGCFQNGG

35122
HIKVEHEEV

35123
LIEKQRWML

35124
GIWVRDTNY

35125
EICMRSGFI

35126
CIQPTWDSY

35127
QISKPSVQC

35128
SGITFPGSN

35129
DFIPDWSST

35130
EMISNNCHR

35131
EMINACFQA

35132
APIFVCDCC

35133
YSIIWRVYR

35134
RKIESYCIT

35135
EKIMVKRAF

35136
ATISKLECQ

35137
LKICFSPTV

35138
ELIYKIKTD

35139
ARIWNDSMA

35140
DQILPMNGV

35141
IHIHFNWFY

35142
RGIYCYYVC

35143
WRIYEEDPT

35144
YKIGAAECD

35145
DPILQTYLS

35146
DQILSMNGV

35147
GFIERYGDC

35148
DSIIKDQSD

35149
LFIKVKDTS

35150
MAIQVFMSD

35151
ISIENEIMG

35152
APIQITKAY

35153
EKIRQNAAP

35154
ISICAQDID

35155
VGIMRNDTG

35156
GTIHMYCGW

35157
TRILNEFFC

35158
QNIRLGVPT

35159
DVIHTTHSG

35160
SHIQLNAWR

35161
ILIFIQNCP

35162
TRIVEPDAN

35163
ATIAQWCVH

35164
HVIGMGAGE

35165
RAIYNDMTE

35166
SAICQNYDA

35167
SQIQIRQAD

35168
NRIQMIDFQ

35169
FTIFPGNAG

35170
YVIGQKDEM

35171
IWITCPYAA

35172
NAIPYFVQA

35173
AVIHEKPHG

35174
ACIPQMHSA

35175
TSIEGLKTM

35176
ASIATMDRI

35177
PVICVWHDN

35178
CWILHHRCS

35179
VTWVHYDMY

35180
YTMVAPDYE

35181
SMPVGTKID

35182
MPKVHCASC

35183
WCLVNELPT

35184
VTSVAGGGE

35185
QQTVVQFCL

35186
ADVVMQTCC

35187
GVHVTESGA

35188
NYCVNKRAV

35189
MTSVCNWDS

35190
REKVWSDLK

35191
ICFVNLDHV

35192
MKHVSTMTK

35193
SGFVILEGI

35194
VFTVAHCVH

35195
WFNVRQNGQ

35196
QAPVIPIMA

35197
TNGVQIMHV

35198
INMVSMYAY

35199
QGFVVHSPS

35200
QFYVAEHLE

35201
RQCVIGDYD

35202
HTRVVKSTG

35203
GTAVSRQSK

35204
QTDVKVNFD

35205
NWTVHLIYG

35206
TAMVSECKD

35207
GHLVPFDCG

35208
HYVVRHEDP

35209
PTDVITTDQ

35210
CMEVTCFVV

35211
IQAVMASGW

35212
LESVKMDCN

35213
EMYVQRHSL

35214
AAFVMNGDG

35215
HCWVNEKCE

35216
SKDVYGMYC

35217
EDLVVYMGS

35218
VGNVMIKMC

35219
IHRVVCRDH

35220
MFVVNQNWA

35221
HEWVLDFFP

35222
RVEAYAPTT

35223
THCHYQGND

35224
WAGCYTYAN

35225
WDHNYFTYL

35226
IVMEYAFIP

35227
NSCTYNKDA

35228
FKNWYINMH

35229
ETNSYNSHH

35230
LSSCYYMTY

35231
GSTQYPCCK

35232
QAEGYRDCY

35233
IGQWYECMT

35234
GNAGYQADL

35235
GCMPYQINV

35236
SLCIYDHRH

35237
ETNHYLISD

35238
EGSDYVGYN

35239
NMMQYQWAI

35240
LPKIYLSFC

35241
QNLMYCSSV

35242
RPVTYPHTA

35243
FDKTYFHDC

35244
LMEHYLREG

35245
NVWYYFRTW

35246
WPSFYANYR

35247
IACKYERVC

35248
LMEHYLREW

35249
SMHDYNFNV

35250
GYRNYFIWP

35251
FAMSYQRQN

35252
EVYCYQLQS

35253
YKNSYSQWP

35254
SHMSYGQVQ

35255
IQCKYIHFT

35256
ANHLYHAAN

35257
NYHNYMFGM

35258
CFNTYDQCF

35259
SDNMYHTIE

35260
RPVHYQGTN

35261
MEHFYDRES

35262
PNGHYWMIP

35263
ATNKYANAI

35264
DKTCYLTCP

35265
TMSNYGCLQ

35266
MGMAYSDSE

35267
ELPRYYGQM

35268
DKYLYMYSD

35269
LLAKYMVHT

35270
MTERYYWCQ

35271
DFMWYHICS

35272
HGSSYNTVQ

35273
SLTQYQWRG

35274
TVNWYSGFG

35275
MEPHYWEWP

35276
STTAYHQDY

35277
VVTQYVDAP

35278
AVDHYSNGA

35279
NGNHYNAMR

35280
LHKAYVTDS

35281
SGNMYESGE

35282
CSESYRGPD

35283
YLEMYSKDW

35284
DPGWYGLAP

35285
MNSFYRAEW

35286
NEDQYMHVE

35287
WHGFYKIMF

35288
YTWPYPETH

35289
PGGCYRMME

35290
MYLNYRHFA

35291
WVHPYSYCA

35292
DYFQYNEAP

35293
RQTTYDCLD

35294
AVEDYWRYF

35295
TSCCYESSS

35296
EPRLYCAQE

35297
HTLLWYGIC

35298
MDHPTYGIY

35299
DGCQKYQEQ

35300
FKFISYVGD

35301
IYLKFYFSL

35302
DCDMKYLYE

35303
LGSPFYECG

35304
LLLNSYKWY

35305
RLSLWYFCH

35306
MDRMMYMLG

35307
AMGYQYLYE

35308
SVMLDYHDM

35309
IVKQCYDCH

35310
QVEDGYWSY

35311
EWLSLYCHW

35312
CTEIRYYGT

35313
CLHHHYPVT

35314
QYVGMYTQQ

35315
QHSRAYMWY

35316
TSSDHYALP

35317
YAMPLYFCE

35318
ECVSDYACC

35319
GHHQCYKAD

35320
VSLQKYDMT

35321
NQQRTYNNG

35322
QEYHIYFPF

35323
AEDTPYTIY

35324
YSVQAYMAN

35325
QTAEWYCGA

35326
ECGCMYWGY

35327
RKWSCHRGH

35328
RQTDTHRAN

35329
NVDAHERYR

35330
SRENLGRYN

35331
WMPNQSRGW

35332
SATARFRDN

35333
EKRLFDRTY

35334
LGHHKGRWH

35335
YEWPWQRGL

35336
YEWPWQRWL

35337
YEWPWRRWL

35338
GCLGTKRQI

35339
HDSLFKRTE

35340
HEWPWQRWL

35341
SYLRQQRDT

35342
ALANKRRNH

35343
HEGLIMRLE

35344
AQRCFHRWG

35345
LFCPTCRYA

35346
HLYSDGRGS

35347
QCKTHPRSF

35348
LFTDTKRPE

35349
EFTHAIRQC

35350
GATWEIRFP

35351
ASWLCGRNV

35352
AMLQKKRQD

35353
DWYLSSRDQ

35354
LHSYQWRLG

35355
CGHLLRRAE

35356
IEFTHSRKV

35357
PWYNWDRRC

35358
LCTSTERQG

35359
LVDRSGRTA

35360
NLDMSRRDF

35361
NAMCFGRAK

35362
VHHKTNRSY

35363
TVNRAMREE

35364
EAGPRKRDV

35365
CSVWMQRDR

35366
TFVYGERDR

35367
LREEELMTR

35368
WHRDEMLIT

35369
AMKPMILTN

35370
RAAGIQGTA

35371
EQELKNSTQ

35372
MATYSDPTS

35373
NPPHRNCTI

35374
SVTPDSITT

35375
EACHPDLTP

35376
ASEHKANTT

35377
AYPLENDTD

35378
TYYNMWHTK

35379
DDYQQHWTG

35380
DTLDMPETN

35381
NEQCHAKTS

35382
NEQCHVKTS

35383
PYKIVMITR

35384
IKHITHNTY

35385
STNPNTETR

35386
HMALVMQTA

35387
YDLWGWSTR

35388
IYMSTGNTP

35389
QQTKNNSTD

35390
DTFSNQNTV

35391
SAPRNEITR

35392
GSKIIMHTE

35393
MTKMMCETI

35394
WCHITWPTN

35395
CVGYKSNTP

35396
NCDHRFGTL

35397
SSQHASHTD

35398
IMTMQMDTA

35399
IKHITRNTY

35400
QFMMGQQTP

35401
MAMWIADTH

35402
RKVAVESTW

35403
IGQHNVQTT

35404
MLMHADMTQ

35405
MQPMESATW

35406
HRVHSEKTD

35407
RFDQFWDTY

35408
QSRCVDNTV

35409
FNESGWPTW

35410
ESSQEIKTC

35411
DQEAQVVTG

35412
TCKNLTYYI

35413
ECLLFLVGI

35414
EMKCIGAGI

35415
QPHEAEFKI

35416
YTNFEDVLI

35417
PPHIGTQFI

35418
WDNWSNTQI

35419
WVTLCADLI

35420
RSLDIQYDI

35421
RSFYHPKLI

35422
DHFYMSYGI

35423
VEQMVWADI

35424
CPWFIKWGI

35425
PHVSGEGKI

35426
QAWPKTEYI

35427
FNWFVRAYI

35428
DMGMDNKGI

35429
GTMCREQMI

35430
TSVIRLFWI

35431
YMHFRQYPI

35432
EFVLGCQGI

35433
WPNIKHQPI

35434
AAMESCAEI

35435
HQCQSRSYI

35436
YSDPKNMSI

35437
FFRICGSYI

Example 11
AAV5 Variants with Tissue Tropism in Mammary Gland

This example describes engineered AAV5 variants with tissue tropism in mammary gland that were discovered using the methods and systems described in EXAMPLE 1-EXAMPLE 2.

CNS, liver, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord tissues were analyzed to identify amino acid residues in the AAV5 VP1 581-589 region that drive mammary gland tropism. The results are shown in FIG. 18I which shows the positional frequency of each amino acid in the 581 to 589 region of AAV5 VP1 in mammary gland tissue as compared to all other analyzed tissues, wherein the capsid polypeptide has the sequence of SEQ ID NO: 2. The position (“Xaan”, n=1-9) is denoted on the x-axis and amino acid residues are denoted on the y-axis. The frequency of a particular amino acid residue in a particular position is indicated in the heatmap as intensity. Darker squares indicate a higher frequency of a particular amino acid in a particular position and lighter squares indicate a lower frequency of a particular amino acid in a particular position.

The frequency of a given amino acid residue occurring at a specified position corresponding to position 581 to position 589 of SEQ ID NO: 1 (generalized in SEQ ID NO: 2) in variants identified in mammary gland over the frequency of that given amino acid residue occurring at the specified position in variants identified in all other harvested tissues was analyzed to identify a set of sequence rules for capsids that preferentially target mammary gland tissue. With reference to TABLE 21 below, and SEQ ID NO: 2 (AAV5 VP1), the following amino acids can, thus, be independently mutated, in any combination, at any one or more positions Xaa1-Xaa9, to provide a VP1 capsid with enhanced mammary gland tropism as compared wildtype AAV5 VP1 capsid (SEQ ID NO: 1), where mammary gland tropism here refers to properties that are preferred for mammary gland transduction over properties that are preferred for transduction of all other harvested tissues.

TABLE 21

Mammary Gland Tropism Rules

Xaa1 is selected from C, K, M, Q, R, or Y

Xaa1 is selected from C, Q, or R

Xaa1 is C

Xaa2 is selected from A, F, I, K, S, T, or V

Xaa2 is selected from A, S, or V

Xaa2 is V

Xaa3 is selected from A, F, G, I, K, L, R, T, or Y

Xaa3 is selected from F, G, K, R, or Y

Xaa3 is selected from F, K, or Y

Xaa3 is F

Xaa4 is selected from A, I, K, Q, R, or T

Xaa4 is selected from A, I, or R

Xaa4 is I

Xaa5 is selected from I, L, M, Q, R, T, V, or Y

Xaa5 is selected from I, M, or Y

Xaa5 is Y

Xaa6 is selected from H, N, S, or V

Xaa6 is H

Xaa7 is selected from A, H, I, N, S or Y

Xaa7 is N or S

Xaa7 is N

Xaa8 is selected from A, C, D, G, H, M, Q, or S

Xaa8 is selected from G, M, or Q

Xaa8 is G

Xaa9 is selected from A, E, L, W, or Y

Xaa9 is selected from A, L, or W

Xaa9 is A

TABLE 22 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that were found in mammary gland tissue and comport to one or more of the rules provided in TABLE 21. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 22118-SEQ ID NO: 23117, as disclosed in TABLE 22. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 22

Sequences of the 581 to 589 Region

in AAV5 VP1 Capsid Polypeptide that Drive

Mammary Gland Tissue Tropism

SEQ

ID
581-589

NO
Sequence

22118
CIKTDWQRI

22119
CAVSGPECY

22120
CCEIWLASS

22121
CDYWLVVPW

22122
CFTQKESAY

22123
CGYWLAVPW

22124
CIPTYHHFG

22125
CIRGQTHYN

22126
CNTPYEVWA

22127
CQSTGAHME

22128
CQSTGSHMA

22129
CSKATGGAE

22130
CSKTTWGAE

22131
CYNSGDCGN

22132
CIMSHCRQH

22133
CSNKTCCKA

22134
CGYWLVVPW

22135
CHMLNDCMS

22136
CSKATWGAE

22137
CKNPSWYMD

22138
CFEMYYAMW

22139
CDPCMECGS

22140
CTDGKHASW

22141
CFCTYKIEH

22142
CIGPHWHIP

22143
CAHQQHNED

22144
CAHSYQMSR

22145
CAIHIFLQD

22146
CAIQISDTS

22147
CALSALTME

22148
CALVSRSQD

22149
CAMIHGKYD

22150
CANFAMQQF

22151
CASRSMLTE

22152
CAVRTFMGS

22153
CCDDCPPNR

22154
CCGIFNGIY

22155
CCHMQYRES

22156
CEMYREYFI

22157
CFAKMEKLQ

22158
CFAPYMNVD

22159
CFMHYGRIA

22160
CFSQFASGS

22161
CFYQKGAAY

22162
CGFWLDQNE

22163
CHRLVMGYH

22164
CIAPRPEWP

22165
CIKLKCSHS

22166
CILWKHAMP

22167
CISNTRFGV

22168
CITMPMNFG

22169
CIYEWFTRD

22170
CKSGMVHHA

22171
CKSMGLFLV

22172
CKYDYHQNT

22173
CLGYMWQNT

22174
CMKADGMQR

22175
CMSDRGLQS

22176
CMYMQNAMG

22177
CNNLSVCDE

22178
CNSLFTNQQ

22179
CPTMYKMQA

22180
CQKCYDHYQ

22181
CQYRTRMIT

22182
CSAPTLSGN

22183
CSFHIIMGE

22184
CSFHNNEHA

22185
CSICEHCMD

22186
CSMAEQKSQ

22187
CSMRYMAVH

22188
CSRPLHDSY

22189
CSTQYMKHM

22190
CSVNFYTSF

22191
CSWEWFRAE

22192
CSWQLWQSC

22193
CSYSTQIGH

22194
CTAITLRYP

22195
CTERTESNR

22196
CTHDIWAQA

22197
CTHQIHLPA

22198
CTKTYMHDF

22199
CTLNVQSDG

22200
CTLQWSAGP

22201
CTMRVQMKP

22202
CTVLRFDHQ

22203
CVAYSYNST

22204
CVCGKKQMG

22205
CVFAQSGSY

22206
CVFYQRKVL

22207
CVIAKYYQE

22208
CVIHTPYQY

22209
CVIRDGHYD

22210
CVKMKQDFN

22211
CVMEQQKER

22212
CVNTTHDYQ

22213
CVNVMAADR

22214
CVQIILDGF

22215
CVRMCVDHA

22216
CVSMEVPMV

22217
CVICKFDEQ

22218
CVTLTHRDK

22219
CVTYSSAPK

22220
CVYPTMKAD

22221
CWICMCVSP

22222
CWLHTRTHW

22223
CWLHTRTHW

22224
CWTTANTSN

22225
CYAKVTANT

22226
CYEQWNGLA

22227
CYFLHNNHE

22228
CYYTIEKAE

22229
CAVGGPECY

22230
CKTQERANS

22231
CYNSGDCDN

22232
CTSTDPIAP

22233
CRGKSHDHT

22234
CPALNYDCV

22235
CMLTHIMEF

22236
CTFPSADMC

22237
CQSTGSHME

22238
CSFHMNITS

22239
CIPTYRHFG

22240
CYGRVAHHE

22241
CTEIRYYGT

22242
CVKPFLTGW

22243
CAWYMKQGN

22244
CAEPVYMWK

22245
CAGTSMAKE

22246
CASHVKWGP

22247
CAVAYMSCT

22248
CAYKPHVAP

22249
CCHNEHRHM

22250
CCVSFAYSC

22251
CDAVHNDAS

22252
CEYLPDMHK

22253
CFHWVNHMT

22254
CFQTANIGG

22255
CHDAQFKTD

22256
CHKSWWRLD

22257
CHYWVSTDG

22258
CIAHRHAQP

22259
CIFRLSALS

22260
CIGSRQAIA

22261
CIHIASRLG

22262
CISFYTSAG

22263
CITKLMVNA

22264
CIVTDSMAP

22265
CIYYPNFYN

22266
CKQMYLATE

22267
CKVPVFSLE

22268
CLRVCNCSP

22269
CLRVYNCSP

22270
CMLRYMLGR

22271
CMLRYMLGR

22272
CMPQKYHKD

22273
CMSAMCMWS

22274
CMSKIVWSS

22275
CNDQAQYDW

22276
CNTSMAIMG

22277
CPWWSDNCM

22278
CRVTITYPT

22279
CSFQMQFMG

22280
CSFVLDNAF

22281
CSGPVSASA

22282
CSKYFNDWG

22283
CSKYSAEYP

22284
CSTPHHLQL

22285
CSVNIGQHA

22286
CSYKDQSQQ

22287
CSYTHWRSA

22288
CTAHPMLQA

22289
CTATKILHA

22290
CTFVALSMQ

22291
CTGSIVKQM

22292
CTHPRITGA

22293
CTIKQLCNG

22294
CVANCQQCI

22295
CVARNINAN

22296
CVCMYQNMQ

22297
CVFLAQKMK

22298
CVFMRSHDK

22299
CVFPVHSSN

22300
CVHKICMHH

22301
CVHQNSRAK

22302
CVHRGLTYT

22303
CVHVAMHYP

22304
CVHYANMML

22305
CVLSKMWCH

22306
CVLRSNVQL

22307
CVNEFFTCF

22308
CVNMKTDGN

22309
CVNQLCDSQ

22310
CVNVENLTA

22311
CVPDTIDTE

22312
CVQMIMKDW

22313
CVQTDNRRP

22314
CVRHYGEYW

22315
CVVADQGRG

22316
CVWHAPIQT

22317
CVWHASIQT

22318
CVYTKIIGH

22319
CWEFMFWHG

22320
CWGVISKAP

22321
CWIRGTQDR

22322
CYLITFREH

22323
TVADSSRLM

22324
TVCMFQVLP

22325
TVFPMTVVP

22326
TVGFSEQDA

22327
TVIQWYWHH

22328
TVLSWGLCN

22329
TVNQECNCQ

22330
TVRPQDNKN

22331
VVDHSCSRV

22332
VVDPFGHEQ

22333
VVHENQLKP

22334
WVTCRGTVF

22335
YVEIHHAQK

22336
YVHEGGENP

22337
YVLESNYCQ

22338
YVSASWMTP

22339
YVTKFAECH

22340
YVVTHVPWH

22341
YVYDEFLWS

22342
FVMCYNDCK

22343
AVHWHRKWE

22344
AVINPPCHP

22345
AVSLQPQFG

22346
DVKAFMGCV

22347
GVEAYRNFK

22348
GVIHRQYCP

22349
KVEANPRVC

22350
LVHAVSYIC

22351
MVAPPNCCV

22352
NVNVRESEW

22353
PVHAVSYIC

22354
RVTIMFTDT

22355
SVHTADVCC

22356
SVPQQKIDE

22357
TVKINSISP

22358
YVQCTHYAQ

22359
GVIHRQYCQ

22360
QVPRPSMWW

22361
PVSPLRSYV

22362
QVTSDGKII

22363
VVGWDNYVA

22364
NVHHEWCDW

22365
MVDPMSVPF

22366
DVDYRSEAW

22367
IVPQQKIDE

22368
VVHPMAYCF

22369
LVVTKFTYV

22370
AVKSRTTWV

22371
AVQGDPGVA

22372
AVSVETWWT

22373
AVVDCPLNI

22374
AVWNVVSDG

22375
AVYQYVETP

22376
DVDKLEHGQ

22377
DVDNCYKGD

22378
DVICFGKWE

22379
DVQTRNLSK

22380
DVYRDAMTP

22381
EVALQWAST

22382
EVDVRVLFD

22383
EVESWMWVC

22384
EVHRVASCP

22385
EVSKFNHYF

22386
EVSTITFWS

22387
EVTPHWTGG

22388
EVYYAMASW

22389
FVNVMKVTC

22390
GVKPGASCG

22391
GVQQFYRYD

22392
GVRLSSPWE

22393
GVYPMHEAC

22394
HVCMMASCD

22395
IVCAYSNAS

22396
IVEPAIVCA

22397
IVNKWDGFA

22398
IVYYCGKRK

22399
KVICHEFTS

22400
KVMHTGWHP

22401
KVVQRIDHD

22402
LVCRYWDDE

22403
LVFMQEYAA

22404
LVKCNTCME

22405
CVNHCGQMG

22406
LVTEQVNSP

22407
MVDRMKWHK

22408
MVYPIEWGA

22409
NVGIITFQD

22410
NVLQNSIMC

22411
NVYTHHVLS

22412
QVDNYFNRL

22413
QVNFGLHAQ

22414
RVNEATHDD

22415
SVCPFSYNL

22416
SVFSNQDGV

22417
SVGFEQSWY

22418
SVIPLPHGD

22419
SVITTEFQD

22420
SVVNHAGAC

22421
TVAHERSFQ

22422
TVDNTNWLK

22423
TVHYSSQDP

22424
TVSGLWQEL

22425
VVGTYALTY

22426
VVHTCAMCT

22427
VVSAHMPGV

22428
VVTVAWTPG

22429
WVEKYLISY

22430
YVLSAIPFG

22431
YVWYRRIDL

22432
KVCNSEYWD

22433
QVHIKVTDH

22434
KVDRNTNWC

22435
GVEAYCNFK

22436
HVWNDVFHM

22437
EVQRNIYSP

22438
GVAQYLHHC

22439
DVPIHRVQL

22440
SVNLNCTEH

22441
FVFAMIPGF

22442
AVHWHRKCE

22443
GVMIYEADE

22444
KVAGGHETT

22445
FVPWCHSFG

22446
NVTPGRTHS

22447
LVSQWDYDM

22448
RVDIGGFAQ

22449
HVDQHRVQA

22450
AVEACYHSP

22451
AVFTFATCG

22452
AVQWSAENM

22453
DVDYRPDAQ

22454
DVELGGSNF

22455
DVFQYEMLA

22456
DVGYASDPK

22457
DVKVTEDAE

22458
DVRLDDGAQ

22459
DVRPDEFHN

22460
DVRPDKFHN

22461
DVVSTESTT

22462
EVFREWMWM

22463
EVGSDCHQG

22464
EVKYMRHIC

22465
EVRPSVGSD

22466
EVTSAGFDF

22467
FVIWRQESR

22468
FVRCLSTHN

22469
FVSEQMTAQ

22470
GVDSSRPVQ

22471
HVCQKTHSI

22472
HVNTSCKQF

22473
HVQHHDAAT

22474
IVAYGTCGA

22475
IVHQKQLAA

22476
LVDEMFSQW

22477
LVKGSFAPD

22478
LVKPQSGEL

22479
LVRQDSQIG

22480
LVVCSCPFP

22481
MVEFYCSDE

22482
MVEFYYSDE

22483
NVAYHMVNN

22484
NVENAYDAM

22485
NVFLNLDLA

22486
NVHTIENSA

22487
NVVDILAGS

22488
NVYYKHAHP

22489
PVIRTWVDI

22490
QVASHTQPE

22491
QVATWSMYH

22492
QVFMASTGE

22493
RVGVQEAVC

22494
RVHISILKG

22495
RVNMKTDGN

22496
SVCLHNLVG

22497
SVHGFDQSM

22498
SVIKVAFFF

22499
SVRLHNLVG

22500
TVDPNVRVD

22501
TVVPQGFDR

22502
YVCIPAHDR

22503
YVCMIHMDD

22504
YVDLIMSVD

22505
YVISKCAGQ

22506
TSFCQIASR

22507
VCFKLFHVE

22508
AFFKFMTQM

22509
DKFPMPTHS

22510
ETFGIRTFA

22511
GIFWASYGF

22512
GKFPMPTHS

22513
GMFCASYGF

22514
KDFWGHQCF

22515
LLFTLMSSS

22516
QTFYEGHAT

22517
RDFRMNAGA

22518
RTFYEGNAT

22519
TTFQSLQQM

22520
NQFHWVRYD

22521
LWFFMKFAK

22522
YTFWDYHRS

22523
KDFWEHQCF

22524
VHFTVLNWG

22525
DYFQYNEAP

22526
QEFLEERYV

22527
GTFHCCAWE

22528
ETFGIRTFD

22529
EQFADRAPR

22530
ARFMWNHTR

22531
DTFLEMSMG

22532
DTFPIAGSF

22533
EAFRHDHFH

22534
EGFFYHHGG

22535
EGFQFSNQG

22536
ERFMTASMQ

22537
FEFTVHCTH

22538
GKFEPPVNA

22539
HAFPTVRWG

22540
IMFDPNIPN

22541
KAFHIQREK

22542
LDFQIRFER

22543
LPFHLQSDT

22544
LPFTQMHGS

22545
LSFPVVLTE

22546
NAFEYEKIM

22547
NAFTKTQDQ

22548
NSFSTNWSY

22549
NTFCETVTF

22550
QKFEGEWFT

22551
RAFVGCSPE

22552
RQFAHRFKM

22553
SCFAILSAN

22554
TSFELNDER

22555
TSFSPELFN

22556
WNFNCIILH

22557
STFISSGIQ

22558
FKFWDMETR

22559
KRFVDMPEM

22560
VSFQRMNDM

22561
ERFKTFCEA

22562
LLFPLMSSS

22563
EKRNFKTMN

22564
NWFQAIEIG

22565
GIFCASYGF

22566
IWFWWEKAE

22567
RTFYEGHAT

22568
TTFPQVIMN

22569
DAFINRLSG

22570
DIFDRPSSD

22571
ETFLPWVHT

22572
ETFSKTWFG

22573
KSFGSPFER

22574
KSFGSPFEW

22575
MAFYKSQFN

22576
NAFGMEYNT

22577
NSFKPSGVT

22578
PWFMVPNWP

22579
QCFFNHRCE

22580
RAFQDHYAY

22581
TIFSASDSK

22582
VAFYQENTT

22583
WHFSSKWMD

22584
TPGITQQQW

22585
TSHICVSSL

22586
TWLIDADMA

22587
VAMIMEGQF

22588
VFKIREDDN

22589
WADIEYSHD

22590
WTGIKCQSS

22591
YFCILEKMT

22592
YMCIQMAGW

22593
ESPIMLNQA

22594
RTPIELMHG

22595
AGQITTSVI

22596
ESDIFDGRQ

22597
HPSICKGGD

22598
QCHIIVARE

22599
QCHIIVARG

22600
TAKIIMLAS

22601
DMSIATWYP

22602
ESDIFDGHQ

22603
TCVIGKGRR

22604
VACIEHAKC

22605
QFDIDQCCT

22606
HPSICQGGD

22607
AWIISQEGR

22608
DMWILKDAS

22609
DYYEIDCFM

22610
EHMIEFPHR

22611
FREIKIYDM

22612
GLYITHSME

22613
HCNIFSREQ

22614
IICICNRGH

22615
KEVIMKRCV

22616
KLEIMCQAK

22617
KPGIMKFYN

22618
LYNIPMGYT

22619
MKDIRVTQR

22620
MYTIGTCYG

22621
NINIYWMID

22622
NSAIMSNYM

22623
QEDILQTQY

22624
QFEICHSQE

22625
QHLIMPEPH

22626
QPLIQHAGG

22627
QQTIDSYGH

22628
QSGINDLYG

22629
RCMIYHQNA

22630
RGGIFPQRP

22631
RMCIDYLRP

22632
TACIQQKLL

22633
TNRIVTRII

22634
TQQILHDAF

22635
TRIINVILG

22636
TSLITGQLY

22637
VEPIMFQPA

22638
VNYIICKMD

22639
VNYIICKMG

22640
IKHITHNTY

22641
MYAIADELD

22642
NPSIQPKTM

22643
PENIAEDHE

22644
MRKIKIACM

22645
LRAILDCGW

22646
ELAIDQDIH

22647
SRTICVHGN

22648
IANITACAN

22649
EESIRLLDE

22650
EIYELPKFP

22651
GSAIFVKEE

22652
GYTIKNGYN

22653
HHEIANLHQ

22654
KHQINAVER

22655
NGEIFSCMS

22656
NGEIFSYMS

22657
QFKIYDCVG

22658
QIGITYYLP

22659
QMGINHDDR

22660
QPAINTQCA

22661
SFKILNRGG

22662
SPDIDLINH

22663
STSITELAM

22664
TTLIKMHVE

22665
TWEIKHNQV

22666
VACIANLWT

22667
VFAIMTRTQ

22668
WSNIYWRQD

22669
THGHYTMSA

22670
TRAGYNEQD

22671
TSVKYSAWG

22672
TYWNYAEAK

22673
VFTDYWEQG

22674
VFTDYWERG

22675
VIDWYPHQQ

22676
VSDEYVMEQ

22677
WELTYDCLA

22678
WIICYEALQ

22679
WLIVYDHGN

22680
YAHSYGHFD

22681
MAEQYKYGH

22682
TGAKYHDWL

22683
EAENYFPRH

22684
GLMDYSLCA

22685
KEIQYHSYE

22686
MDTCYMYAD

22687
QPPKYFVHQ

22688
RNTPYEFWA

22689
SGNMYESGA

22690
TEPHYWEWP

22691
TTERYYWCQ

22692
KEIPYHSYE

22693
LMAWYENWY

22694
ITRTYQEMQ

22695
VREYYPKFH

22696
QQEAYILRI

22697
DMTFYLSRD

22698
MMPEYFNKD

22699
FITAYHPCL

22700
GQKSYDGCH

22701
MFEKYFQTM

22702
NEAPYSTWQ

22703
SMSEYHYDE

22704
AAVQYSLMG

22705
AEEVYGYHQ

22706
AQMEYDCAM

22707
DDALYAVFY

22708
DSWMYHVQL

22709
DYIHYKNGM

22710
EFSYYHTHG

22711
ESEKYTWSG

22712
FGATYNQTI

22713
FGLWYRDHK

22714
FHITYLMDK

22715
GAQQYADSI

22716
GFCHYWKQG

22717
GNCKYHVTQ

22718
HIVTYEHHE

22719
HPCYYSWAP

22720
HWDNYWKMD

22721
HWREYPSKG

22722
ICHQYTRHG

22723
IPLAYPEYQ

22724
ISKAYVDWW

22725
IYHPYDMLG

22726
KQMEYDNFH

22727
LMIQYVNKH

22728
LMREYMSVP

22729
MCQWYATNL

22730
MSDYYKEDS

22731
MYGGYAAGF

22732
NFCGYQAMH

22733
NGLQYMSSA

22734
NYHYYDHAG

22735
PTVAYENLD

22736
QAIGYNYFW

22737
QGQYYYECG

22738
QWPLYVMDR

22739
REELYQCAP

22740
REKNYELSE

22741
RWYWYESCN

22742
SMTSYYRQQ

22743
SWVWYPNLC

22744
WATAYWQGI

22745
WCGFYFLAA

22746
WNSPYKKRY

22747
YQVHYASHG

22748
KCTEYVHTC

22749
PIHTYKRHP

22750
RMKWYLPCV

22751
FAMSYQRQN

22752
EASWYYPAI

22753
MEPHYWEWP

22754
MSTDYCKVN

22755
TSKDYNEEI

22756
MTERYYWCQ

22757
QTPKYFVHQ

22758
YSKQYAEYY

22759
HSDWYMKYP

22760
AEGYYVFQP

22761
AWGCYQKGY

22762
DIKMYWEGA

22763
FGILYNNNE

22764
FNQLYMITH

22765
GAEGYSKDQ

22766
GAEGYSKGQ

22767
GINDYKSGN

22768
GMDFYFNLE

22769
HKMAYGRDQ

22770
EAYEYWDYW

22771
IKEGYQLGG

22772
ISQFYLLDQ

22773
KECNYQARG

22774
KPLQYEAIQ

22775
LPNPYKTVV

22776
LPVQYEDRD

22777
MCDNYKSLA

22778
NHGNYEEAW

22779
NTMPYGHSY

22780
NYIPYGRLA

22781
QTQTYAYMD

22782
RFDHYNNEQ

22783
RKEYYPNVQ

22784
SAYQYLGCH

22785
SCGPYLWGT

22786
SHGNYEIAW

22787
SMNLYNRGV

22788
SMPPYRMLS

22789
SYQPYSKTV

22790
TANCYPILG

22791
TSWMYYDYG

22792
TYCLYGISE

22793
VALHYDHPN

22794
YFQLYMKEG

22795
YWAGYHMWP

22796
YYTCYPPSM

22797
TTHSDHHHA

22798
TYNLEHQCS

22799
TYRLEHECI

22800
VCQKVHGLF

22801
VLDNIHWHW

22802
VSEETHNFR

22803
VWNWEHKCT

22804
WYQCLHYQN

22805
YAWCQHCKN

22806
YHMDVHDSM

22807
YNMLTHVSM

22808
ETTLAHRWE

22809
HSAARHMFV

22810
LAECKHSSA

22811
MFMMMHDIW

22812
MFMTMHDIW

22813
NLNNWHTDM

22814
QNNWGHYNK

22815
QTLGGHMCT

22816
SSNFNHHIY

22817
MTAKEHAYL

22818
HQVLDHQLQ

22819
DMRRAHQEK

22820
VNQMMHTQA

22821
MFSFKHVHW

22822
AQASAHIMS

22823
DAIPRHICR

22824
DCSQQHMTL

22825
DEGVEHRPM

22826
DEMLQHSIM

22827
DFHNAHFGS

22828
DHDTNHWSH

22829
EASTFHRQG

22830
ECAETHEMK

22831
ECHVGHSGC

22832
EMCSVHQAW

22833
FCKFMHETH

22834
GFCQLHTQE

22835
GPSLIHQAA

22836
HYETEHDGC

22837
IAKASHKDR

22838
IASGTHHCQ

22839
IIEQSHTLE

22840
KLDLTHRMQ

22841
LFTKQHVYS

22842
LKHFVHAAS

22843
LTLGNHGEY

22844
NAGCQHEYE

22845
NQRVEHAFS

22846
NTEREHRYN

22847
QIVHDHRHR

22848
RYDCMHCYA

22849
SAGEIHEHF

22850
SLDGVHDGF

22851
SLHYWHSAN

22852
SSNYWHASS

22853
STKAHHWCQ

22854
TEAATHWHR

22855
TEDLSHLWW

22856
TFQQSHAAV

22857
TISLVHLSE

22858
TMDVLHCDY

22859
TWCYRHNVE

22860
VSSQGHRFY

22861
VWMTWHASY

22862
YHAPQHHMG

22863
TQKGNHCRW

22864
APRNVHCGT

22865
LMEYCHDRF

22866
NYDFLHHVE

22867
NCIAKHEVT

22868
NSAHVHYFH

22869
NGYLVHACQ

22870
SDGMVHNCW

22871
KCGVEHEHR

22872
QATNEHIFW

22873
DTHHTHWQY

22874
FTQAIHHPD

22875
LEIVMHSLP

22876
GAKYRHYLQ

22877
APNFCHCSF

22878
ATHDMHRLC

22879
DDKCDHLQH

22880
DTKWMHQHC

22881
DTKWMHQHC

22882
EGVPLHRAF

22883
VSMFNQNSC

22884
WAHYTINQQ

22885
YHRHIANNG

22886
YLHNQGNGL

22887
YRGAACNPH

22888
DLGMINNME

22889
DEKAEKNES

22890
FASMQANDR

22891
GGHLEGNWQ

22892
HITMWYNGD

22893
HRQESPNDK

22894
NEIFNQNSY

22895
NGVALENRG

22896
NPWFLDNQF

22897
QMTSSTNCV

22898
TKQQLINSN

22899
NEISNQNSY

22900
VECTDTNDE

22901
HITVWYNGD

22902
RNNWKVNSQ

22903
YCCSDTNTE

22904
QMTSSTNCA

22905
PACQKGNHA

22906
ACLWQRNDE

22907
ATVQFANDG

22908
DALTHGNFE

22909
DHAKLNNPS

22910
DITGHCNPA

22911
EAINRVNCQ

22912
EHEHRINQC

22913
EHRQLVNNN

22914
EKIPHYNME

22915
EKWDMFNTE

22916
FTYCVNNYQ

22917
GKICAENWI

22918
GYRPELNSA

22919
HFDVICNCY

22920
IMALDQNTN

22921
IMEMINNAF

22922
IRMYAYNWL

22923
LESQQSNYT

22924
LIGPQANLY

22925
LTSMQENHS

22926
MIQYNKNMG

22927
MSEPRYNDL

22928
NCEWEMNDR

22929
NRSMFKNVR

22930
PTDFNENDC

22931
QFYSMDNHD

22932
QQEVKKNNA

22933
RNCMDTNGM

22934
RWADVTNMS

22935
STIYAFNWA

22936
TCMQCSNMP

22937
TWAPHNNGM

22938
VAKTSWNHC

22939
WICPKINAP

22940
WSILGTNWG

22941
YDPHNNNGV

22942
YSVARCNMD

22943
YSVCHMANY

22944
YIRVGGNNE

22945
FQIRQDNLA

22946
KIDAHSNME

22947
NSVALENRG

22948
KNTMCDNDA

22949
ALKSWMNGS

22950
ILIFIQNCP

22951
QMKPLINDE

22952
TISHHLNRE

22953
DHLFNNNEW

22954
ETGYRENPC

22955
TSDTMWEGD

22956
TTVRKQFGY

22957
VGGSVNRGG

22958
VLYYSNYGH

22959
WAMFMYEGA

22960
YAQCFGQGI

22961
YAQCFSQGI

22962
YLRSLKCGN

22963
NGCTTTKGE

22964
DPKWEQYGM

22965
EFVMGCQGI

22966
FCQAINIGH

22967
GCMTRLDGL

22968
HFHEVAGGS

22969
HHMHIWLGL

22970
HTVGMAEGD

22971
IQAVMVSGW

22972
KSMPPGQGY

22973
SRDTWQIGR

22974
TEGVKRWGQ

22975
VLMFNRGGS

22976
VLMFNRWGS

22977
DPKWEKYGM

22978
TNHCRDAGT

22979
KWTDCIEGY

22980
DTDKQGQGE

22981
MTRQPEDGF

22982
ASDKMDVGS

22983
QPEQGGWGT

22984
KSMPPGQGD

22985
AACHFTYGS

22986
AATCSLKGH

22987
AHADMTTGF

22988
AMQRSVAGE

22989
AYEHCEHGP

22990
DIADRNVGH

22991
DRMCMMCGV

22992
DRSEFRQGT

22993
DTPDFNTGI

22994
ETEGQVAGA

22995
ETNLHTKGH

22996
HALSLTWGD

22997
HSLPHTEGL

22998
IHSRVNAGH

22999
IQKVTKRGA

23000
KEGPTPVGH

23001
KFDFWVAGH

23002
KHTQQDSGS

23003
LATHQKDGR

23004
LMELTNEGW

23005
LSPKNDPGV

23006
LSYSIVTGE

23007
MAVTLMHGC

23008
MCVRWSVGD

23009
MDTHLQHGA

23010
MGGSNVWGN

23011
MKWDCLIGP

23012
NFVCSDIGK

23013
NLMRRDHGG

23014
QAMYTTAGE

23015
QLKNAFEGS

23016
QQYAPVIGP

23017
QSREPRGGK

23018
SAVGWGVGQ

23019
SYIFSEIGT

23020
SYMVIGWGG

23021
TFTYAYDGQ

23022
TKKRNCFGM

23023
TSIHDNQGM

23024
VATSKDVGP

23025
VNRPTNFGT

23026
WPQRVPLGM

23027
YFPFHMAGQ

23028
EIRMIISGC

23029
GRHYEAAGY

23030
MLAFDPMGR

23031
HHMPIWLGL

23032
HFHEVAWGS

23033
AGQFKSEGS

23034
TGACMVEGL

23035
TLVSCDQGN

23036
DGGLRVQGM

23037
WINSPIQGF

23038
EGGKNIFGM

23039
LDGPIGMGT

23040
EPLYSWVGT

23041
TNIKERTGS

23042
TEDVKRWGQ

23043
TSHLAWCEA

23044
VAMMGCMQA

23045
VCENEAGVA

23046
VMCQTLVKA

23047
VSLVRYVDA

23048
YPCFGTSLA

23049
AHTWLQHWA

23050
DAGWQQFAA

23051
DTEQWEATA

23052
HQYELQMHA

23053
LGYHCCQAA

23054
NSTPMACMA

23055
QIEYCMFYA

23056
SDDDREREA

23057
SHDLVAAWA

23058
THAWLQHWA

23059
SAQCAGHNA

23060
VLPEGTMTA

23061
LGMQADRFA

23062
LGYYCCQAA

23063
KYAYSLDCA

23064
GTEQWEATA

23065
WSKGTMLEA

23066
KGIWQGMDA

23067
NERTGACSA

23068
KSEHHWENA

23069
FSEQNVSAA

23070
ASYYLVRNA

23071
DGERFMMQA

23072
DIGLRNMNA

23073
DTIQFVQNA

23074
KWKWKTMMA

23075
DYRGQDHVA

23076
EFTTCVELA

23077
EICFKEYWA

23078
EQIGVWWHA

23079
ETMWNMWDA

23080
IMITEVRSA

23081
KQETRGSVA

23082
KSCITSHCA

23083
LSHQECAAA

23084
LTNDKSVTA

23085
MCRKNKILA

23086
MEGSDNLSA

23087
MHEGVAGIA

23088
MNEYMIKCA

23089
MYGFVMWEA

23090
NIYATGSQA

23091
NQVHSVDLA

23092
NYTPFYKRA

23093
PEYWGQKIA

23094
QFNFTESDA

23095
QTDWKVVVA

23096
SAHDTIRDA

23097
SFMVMACYA

23098
SECGTCQRA

23099
SSSCVVIAA

23100
SSTTVWEYA

23101
STGTNAEHA

23102
SYQWLCADA

23103
TSEGITREA

23104
TSSHQTVKA

23105
VCNMTDLQA

23106
VKESSDACA

23107
VMEQLQHTA

23108
WCTPMNRVA

23109
YKQSHMVAA

23110
MSWDMVEAA

23111
QFRCMMDDA

23112
EIVQPQCYA

23113
ALQFVSSTA

23114
TERRMFRTA

23115
QTPKSLLDA

23116
AEVMQWQAA

23117
EADTFTLIA

Example 12
AAV5 Variants with Tissue Tropism in Lung

This example describes engineered AAV5 variants with tissue tropism in lung that were discovered using the methods and systems described in EXAMPLE 1-EXAMPLE 2.

CNS, liver, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord tissues were analyzed to identify amino acid residues in the AAV5 VP1 581-589 region that drive lung tropism. The results are shown in FIG. 18E which shows the positional frequency of each amino acid in the 581 to 589 region of AAV5 VP1 in lung tissue as compared to all other analyzed tissues, wherein the capsid polypeptide has the sequence of SEQ ID NO: 2. The position (“Xaan”, n=1-9) is denoted on the x-axis and amino acid residues are denoted on the y-axis. The frequency of a particular amino acid residue in a particular position is indicated in the heatmap as intensity. Darker squares indicate a higher frequency of a particular amino acid in a particular position and lighter squares indicate a lower frequency of a particular amino acid in a particular position.

The frequency of a given amino acid residue occurring at a specified position corresponding to position 581 to position 589 of SEQ ID NO: 1 (generalized in SEQ ID NO: 2) in variants identified in lung over the frequency of that given amino acid residue occurring at the specified position in variants identified in all other harvested tissues was analyzed to identify a set of sequence rules for capsids that preferentially target lung tissue. With reference to TABLE 23 below, and SEQ ID NO: 2 (AAV5 VP1), the following amino acids can, thus, be independently mutated, in any combination, at any one or more positions Xaa1-Xaa9, to provide a VP1 capsid with enhanced lung tropism as compared wildtype AAV5 VP1 capsid (SEQ ID NO: 1), where lung tropism here refers to properties that are preferred for lung transduction over properties that are preferred for transduction of all other harvested tissues.

TABLE 23

Lung Tropism Rules

Xaa1 is selected from A, E, K, M, Q, R, S, or T

Xaa1 is selected from A, E, or Q

Xaa1 is E

Xaa2 is selected from A, I, K, S, T, or V

Xaa2 is selected from S, T, or V

Xaa2 is T

Xaa3 is selected from A, E, K, M, Q, R, S, T, or V

Xaa3 is selected from A, K, R, or S

Xaa3 is R

Xaa4 is selected from M, P, R, S, or T

Xaa4 is selected from P, Q, or T

Xaa4 is Q

Xaa5 is selected from I, K, L, M, T, V, or Y

Xaa5 is selected from L, M, or Y

Xaa5 is L

Xaa6 is selected from D, G, H, M, N, R, or S

Xaa6 is selected from H or N

Xaa6 is N

Xaa7 is selected from A, K, M, Q, or R

Xaa7 is selected from A, K or R

Xaa7 is R

Xaa8 is selected from A, F, G, S, W, or Y

Xaa8 is selected from A, F, or G

Xaa8 is F

Xaa9 is selected from A, E, G, P, R, or Y

Xaa9 is selected from G, P, or R

Xaa9 is G

TABLE 24 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that were found in lung tissue and comport to one or more of the rules provided in TABLE 23. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 16118-SEQ ID NO: 17117, as disclosed in TABLE 24. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region

TABLE 24

Sequences of the 581 to 589 Region in AAV5 VP1

Capsid Polypeptide that Drive Lung Tissue

Tropism

SEQ

ID
581-589

NO
Sequence

16118
ETDYCYQCL

16119
EHVFGLLRE

16120
EDHISDSQL

16121
ENTEGIGPS

16122
EVTIHFVPQ

16123
EPRMSDMGD

16124
EPFCTQQLM

16125
EHPRQQDSA

16126
EVTRNHHFN

16127
ETRSGIAWQ

16128
EANLSHSWG

16129
EFLGTRLPD

16130
EGETVISQW

16131
EGRLKRHDF

16132
EGTEEGPTH

16133
ELRFVKTQR

16134
ENLARDAHY

16135
ERTIQHGEA

16136
ESTKAKKFP

16137
EVKLFKRNH

16138
EYKVYRHID

16139
EQKIKNGGH

16140
ERTCIATMW

16141
EVQRNIYSP

16142
EFSAMYSWY

16143
EHITEQDLR

16144
EACYAPSWA

16145
EAEYWNCGR

16146
EAGILMPAL

16147
EANSCEGFQ

16148
EANTWLNSA

16149
EARYSCWWG

16150
ECEQPNRWS

16151
ECFGKRFYD

16152
ECLPVLHND

16153
ECMGPGVCG

16154
ECQLCNWWP

16155
EDFWVFHYK

16156
EEEGQTAAG

16157
EEEQWGHMY

16158
EEVPCEKCF

16159
EEVRPKFGV

16160
EFCEMQNLC

16161
EFCHAMVCQ

16162
EFEDYGSWE

16163
EFEWECVQA

16164
EFFIIGAID

16165
EFKMSVFSN

16166
EFKQEFEAF

16167
EFTRKSCKF

16168
EGEVVDCDK

16169
EGGSQTYQG

16170
EGHQCQEGL

16171
EGRIHAQWM

16172
EGSPQESMS

16173
EGTGMQQAF

16174
EHADSWACP

16175
EHEGLNNHL

16176
EHFFKREDR

16177
EHFTKITVE

16178
EHFYICVHD

16179
EHNKEVYYG

16180
EHRQMHDSY

16181
EHSCTHMGN

16182
EHTRKDVCD

16183
EIHSNIAAG

16184
EILQGTVVQ

16185
EISMRPVTQ

16186
EIVYVDYPH

16187
EKCFCEVCG

16188
EKDHFDVMQ

16189
EKLNHEAGY

16190
EKMHQLATP

16191
EKRMMIDTH

16192
EKWMYDKDL

16193
ELDDICGIL

16194
ELEHYMSYQ

16195
ELKWWWRIF

16196
ELPTILKHR

16197
ELQKWFRKS

16198
EMEGYRAIQ

16199
EMFWQVAGA

16200
EMKFFAYAA

16201
EMKNLKRTG

16202
EMKVIGVDF

16203
EMRQQKMSR

16204
EMRTVMAGV

16205
EMTTSWYMN

16206
EMWSVKFGM

16207
ENAQFLVWA

16208
ENDSVSAIG

16209
ENGHCVSCT

16210
ENHRITGQS

16211
ENRKWTLAF

16212
ENSQTSAFG

16213
EPGELSVHV

16214
EPLTMDTGD

16215
EPRSCNFWC

16216
EQASSGERG

16217
EQMGYVHKG

16218
EQNTANLCD

16219
EQQFWNRLS

16220
EQQTGGYGP

16221
EQRFWNRLS

16222
EQVLMTYWF

16223
ERAEFEMKE

16224
ERAFSHNQS

16225
ERDHIKGCI

16226
ERSDFHWYM

16227
ESDWMNVSM

16228
ESFNYAHFK

16229
ESHPFTNFS

16230
ESISRDQSW

16231
ESKMSHKEA

16232
ESLHALAHM

16233
ESMFWDSDK

16234
ESMRRLGDT

16235
ESNSNRWAY

16236
ESRIMAPLW

16237
ESWNILLTH

16238
ETADRPSLD

16239
ETAHCPLNA

16240
ETCIANAWL

16241
ETDDQMMPP

16242
ETHTSEANM

16243
ETKLLPDLG

16244
ETMAQSCYG

16245
ETMGICHNS

16246
ETMHPVVDG

16247
ETMSTVYGQ

16248
ETNSCEGFQ

16249
ETRQAHLYH

16250
ETTAQKLKI

16251
ETVEYLSYE

16252
EVASKFFEH

16253
EVCRIGWMS

16254
EVCTIMEHP

16255
EVGTSCRFE

16256
EVNDQYREL

16257
EVNMPTGQH

16258
EVPHFDVSQ

16259
EVQCCMAAM

16260
EVQPVRRFT

16261
EVSFKARCA

16262
EVSMPEKTP

16263
EVSMTGHIA

16264
EVSPMRHIM

16265
EVTNEHCCW

16266
EVTQFDNPV

16267
EVTQGECFY

16268
EVVHDPGYA

16269
EVVKCPNSI

16270
EVVKIDMIP

16271
EVYAPRHMC

16272
EWDKMQKVP

16273
EWGRRANKD

16274
EWHAFMSHC

16275
EWWRYMADY

16276
EYCWANKNG

16277
EYIREWAQD

16278
EYKQQGVRR

16279
EYQGNYMAT

16280
EMMERQIHE

16281
EFITSGQCS

16282
EVVFMSGDI

16283
EPGHPGYEM

16284
EVMGLGCDG

16285
EAQWGMDDT

16286
ECKTHRDVY

16287
ENIPQSIWT

16288
ETCWTACDR

16289
EAYTDMPKT

16290
NTRPMRLHM

16291
NTRPMRPHM

16292
QTMQWMCKG

16293
QTMWLKEHR

16294
RTEIRYYGT

16295
STRPAIQKS

16296
STTVWAQCS

16297
STVMESAMY

16298
TTGTFPALP

16299
TTMPKGGNL

16300
VTESWSSWQ

16260
EVQPVRRFT

16302
YTEQEDNAA

16303
KTYCQDGWT

16304
LTTVYNWPH

16305
ITPPYCDMV

16306
LTMMAMCND

16307
ATQTDPVAR

16308
HTLHSHQNT

16309
STERGTPQY

16310
TTGTFPALQ

16311
ATVMFIEIS

16312
FTNQLPEAN

16313
CTLRVEYVQ

16314
VTWMVKTSN

16315
ATQFSGENA

16316
ATQLYAPIA

16317
FTVYVGARE

16318
KTKCCKTHD

16319
KTQIHMGDE

16320
RTEAHLPAP

16321
TTERYEITP

16322
VTVYVGARE

16323
ITASIKDQW

16324
DTAPNESDR

16325
HTRKEGFNS

16326
GTWMEENLH

16327
RTDVMMKVM

16328
ATEMFDCDV

16329
ATNVLYLDC

16330
ATQQVFWNL

16331
ATSHYAKGL

16332
ATSIFCVFR

16333
ATTHWAGHH

16334
ATTMSAFYG

16335
ATTSYNLTR

16336
ATVDISNVK

16337
CIPPIVMDR

16338
CTSFACKYF

16339
DTAMMHKIA

16340
DTGEIMTRS

16341
DTIPYCRGC

16342
DTKFWMQWN

16343
DTKIQTIHT

16344
DTLLPANWS

16345
DTNKLNDLS

16346
DTRQCLDTQ

16347
DTTTSTMAL

16348
DTVNTCGQM

16349
FTELHCNGN

16350
GTEYDRLSP

16351
GTFMDQRPN

16352
GTMPMDAGG

16353
GTMTTLYAN

16354
GTPHVVSQE

16355
GTQVNRATE

16356
GTSVNMKDQ

16357
GTSYWPQWN

16358
HTAICQDAS

16359
HTAYPYQCP

16360
HTDCLFTDD

16361
HTIPFFEAE

16362
HTKMTHGTS

16363
HTMQREYME

16364
HTNCLFTDD

16365
HTTTQVKHP

16366
HTVKIDQTC

16367
ITDQRRRAC

16368
ITELRPLIL

16369
ITNKQQLAR

16370
ITNQFMSKH

16371
ITRVDCSHD

16372
KTEPSVRIA

16373
KTLMWDCQT

16374
KTTILEDKA

16375
KTYICASCF

16376
LTAVSVFRR

16377
LTHDGWFYS

16378
LTKCNEKGN

16379
LTMLHMHCQ

16380
LTSGEGFEV

16381
LTTGIWKTW

16382
LTWMGLSME

16383
LTYVQGDWR

16384
MTEQSGRAY

16385
MTETTDMAH

16386
MTFFVNQAH

16387
MTIGFHVNG

16388
MTKQNKTHR

16389
MTTSSHWAK

16390
MTV0WAQYG

16391
NTAPKMSCT

16392
NTDVPQFMP

16393
NTEPMWSCS

16394
NTEVMDTGQ

16395
NTLRPQHTF

16396
NTMEDWECY

16397
NTNFWNWSL

16398
NTRSKDFWN

16399
NTSAHGYNQ

16400
NTTTFCHDE

16401
NTVFMHEQT

16402
NTVSHFAYD

16403
QTAANFCFD

16404
QTDAQGIGH

16405
QTELDTGLS

16406
QTFLAMDGK

16407
QTHDEYECA

16408
QTIRDMSGF

16409
QTLLSNCCA

16410
QTLWVHGTR

16411
QTMPCMYVE

16412
QTNVELHQE

16413
QTPQYHVGG

16414
QTQLNCAAC

16415
QTSFAETDY

16416
QTTVFQLEN

16417
QTVQLGTPV

16418
QTWAAHCSR

16419
QTYHVYTCC

16420
RTDRNTDFL

16421
RTGFEVPTS

16422
RTHLMEVTQ

16423
STDLKFCAK

16424
STDNKFWLG

16425
STEAAYENP

16426
STERMFKEA

16427
STFTAGATM

16428
STHGHISHN

16429
STMNFLPCE

16430
STMSCEILR

16431
STMTCGEFA

16432
STSDHHCHS

16433
STSSLDKLI

16434
STTNPYQLA

16435
TTAMYPVHT

16436
TTDCMWENH

16437
TTDQKRHGS

16438
TTEQWDFCR

16439
TTERTRICN

16440
TTMPHHEWQ

16441
TTMQCNSYD

16442
TTNPVEMFN

16443
TTTHMLTRC

16444
TTVPFQAWC

16445
TTYCPPTVE

16446
VTDLVRHFA

16447
VTEHQQRSV

16448
VTENLMMYH

16449
VTEQDDGQW

16450
VTKEMVWQC

16451
VTNQCCKAM

16452
VTTQSCKWM

16453
WTDNYFHES

16454
WTLHNQSLK

16455
WTVHNKDEA

16456
YTKAEDSIC

16457
YTKQIDNIE

16458
STMMVGRQC

16459
ITNSERATY

16460
PTDVITTDQ

16461
DTITFKQQH

16462
MTTEHITWW

16463
ITAIMCRSS

16464
CTESCQTGP

16465
MTESVENPG

16466
STTAYHQDY

16467
CTEIRYYGT

16468
QERGSLWYG

16469
WWRAMEKDM

16470
LDRNQSYHA

16471
SLRTPECDC

16472
CYRCALWTV

16473
ANRFYDCSH

16474
DRREKQLMK

16475
DVRAMHSTC

16476
FCRMYANAD

16477
IGRLNASTP

16478
IRRSMQQQN

16479
PSRWYKLGI

16480
TIRYIGMSG

16481
ASRSCMTQE

16482
AERQACTMG

16483
VGRERVWDS

16484
ALRPYMDDR

16485
ASRYDYNHG

16486
ASRYDYSHG

16487
CERGALKMI

16488
CLRQDRIIF

16489
DARPSPISQ

16490
DARVSNIQQ

16491
DCRCYGNNS

16492
DKRDHHYFA

16493
DSRFYEVQL

16494
DSRQLHHIG

16495
DSRYTQCYI

16496
DVRMNQDLY

16497
FGRNYTDHL

16498
FGRYRLWKG

16499
GERWPHNML

16500
GFRVGSLRH

16501
GIRLKGNIM

16502
GSRPRMKQP

16503
HCRLKEDEG

16504
HERTAKFRY

16505
HRRLFNKMN

16506
IERNSGQGS

16507
IGRDIDTSF

16508
INRENRRGG

16509
ISRMCESAL

16510
KLREVGRPC

16511
KQRDYMRKR

16512
LMRWPWDDT

16513
LNRILDRMG

16514
MGRTCADWT

16515
MIRNETQWA

16516
MVRFAPAYP

16517
NMRYTEMGV

16518
QCRNQDKSQ

16519
QERWQIQTW

16520
QSRLEHYGL

16521
SHREMNRAR

16522
SPREKNHYW

16523
TAREDCQHA

16524
TARLIEKYN

16525
TVRWQADTM

16526
VERHKMKWQ

16527
VSRSLERQA

16528
WCRQTMMDW

16529
WFRRHNYFY

16530
WNRDHNDWT

16531
DVRSHDEMT

16532
TERRMFRTA

16533
QVRFEQNPA

16534
MDRMHKSGI

16535
KNRDFEGFD

16536
NQDQARTEH

16537
QAFQFECHD

16538
QATQAGANQ

16539
RMAQFFRAH

16540
RNLQGGNHG

16541
SCLQYKATE

16542
SIEQKLTFT

16543
VGWQYCEEG

16544
TRKQGEGPY

16545
TGHQLPYNK

16546
LIDQNTKGN

16547
CGYQFSETM

16548
SGCQYEPRN

16549
IQAQMDQQQ

16550
CFNQRIQDS

16551
MGLQNDPTS

16552
TATQREAFV

16553
IAEQRLVEV

16554
QPSQQGRPG

16555
AVDQLRMEL

16556
CDHQRHQQP

16557
FRKQGHGEK

16558
IIGQWGQSD

16559
IIGQWGRSD

16560
NELQQGTSE

16561
NVSQWMITY

16562
QYVQCAAKD

16563
SLTQYQWRG

16564
VDMQKQTES

16565
VNVQMPSAP

16566
WAMQWWAKP

16567
YESQKGRRA

16568
MGMQQDTQV

16569
AAQQGLNPC

16570
ADIQISLCD

16571
AFAQYSRGN

16572
AFNQQEMWA

16573
ANHQVTSHP

16574
ANMQPNAIM

16575
ASGQFPKEG

16576
ASLQATCNH

16577
AYAQKYRTA

16578
CFSQTDCMS

16579
CIWQHNITQ

16580
CSQQVNCLS

16581
CWMQSQENW

16582
DCAQITYGG

16583
DCAQLRQDG

16584
DFVQSDTEF

16585
DGLQIQIQE

16586
DKMQCSSGG

16587
DMAQQQKSH

16588
DVMQGNRAF

16589
DWAQFCNHT

16590
DYAQFSVVP

16591
DYAQGAAMP

16592
FGMQRRHKE

16593
FHKQFHSKF

16594
GDEQTHCNG

16595
GFTQFYHHE

16596
GIHQETYCP

16597
GITQMSQHA

16598
GMVQDYVWL

16599
GSMQVMKKC

16600
GVCQMSEMR

16601
GVFQEASHK

16602
GVTQGHEIT

16603
GWMQWHRQR

16604
GYEQPVKCE

16605
GYIQCQMHA

16606
HASQEYTSL

16607
HASQLGAMV

16608
HDAQFLPDQ

16609
HGLQVGQFY

16610
HPEQFVACE

16611
HSNQMWYCY

16612
HWAWLKVTY

16613
IHKQFEMES

16614
IHQQTHYQF

16615
IMMQRALMG

16616
INQQYNSCW

16617
ISTQMENTG

16618
IVEQKPTVQ

16619
KDAQQWNCA

16620
KGNQQITSL

16621
KMIQSTHDF

16622
KPNQSNRQE

16623
KSAQVYWNP

16624
LAVQSIHSH

16625
LHCQIWFQD

16526
LIAQQMHIN

16627
LICQSDCFP

16628
LMTQEAQRA

16629
LSVQAQWDV

16630
LWMQHMCLD

16631
LYCQSPKDC

16632
MAHQSDIMH

16633
MAQQYHDEM

16634
MGHQWSGNK

16635
NFEQYVHNY

16636
MFKQPNHWD

16637
NHQQQNLIG

16638
QAEQENEDH

16639
QMAQREDDY

16640
QPAQMEDMG

16641
QSAQQPAGS

16642
QVFQLTMVK

16643
QVGQYTLPE

16644
RCWQADISG

16645
RPAQIDPWY

16646
SAVQIQVLH

16647
SLAQDIACF

16648
SNDQNIHNG

16649
SNEQIPRVP

16650
HGLQVGQFY

16651
SQTQWDHMD

16652
SSEQAGPHM

16653
SVQQIQKQP

16654
TFHQCERSD

16655
THQQVALGN

16656
THTQYWKYA

16657
TISQMVYSK

16658
TMGQICVHA

16659
TQCQWNVAK

16660
VEAQFMDNA

16661
VLSQYTQHA

16662
VVSQTYQNR

16663
VWQQYILVI

16664
WYIQVDRKG

16665
YIKQIDNIE

16666
YQTQYSFWH

16667
LVGQCTSHW

16668
GIKQTLVRQ

16669
GCQQIQLQT

15670
KYMQENVLS

15671
NCHQLGDNM

16672
MMVQWGAMA

16673
FSEQNVSAA

16674
TSGQVDHRT

16675
YYEQMDGEF

16676
QPVQAPHCA

16677
SEAQDQVWS

16678
PLKALEWEY

16679
QMEALMDCE

16680
QSKTLCYQC

16681
QVSTLENVQ

15682
RQAKLATLH

15683
SKESLPQWQ

16684
TGVSLVNYS

16685
MLHNLWMQC

16686
AAQTLECHK

16687
YWFVLNPCV

16688
CHVNLKAWW

16689
DFGMLEWQW

16690
KYEILNCNH

16691
MQYTLPTWP

16692
TMANLCWRT

16693
TNIPLGTNM

15694
TWMPLAPCH

15695
TWMPLAPCY

16696
HQNLLECNE

16697
TGKWLVYQA

16698
EPADLDVDH

16699
FFLKLNDVP

16700
AHYFLLEYP

16701
AIIPLPMPT

16702
AIVNLHTQQ

16703
ANLPLQQQD

16704
AVEFLAHRY

16705
AVYPLFAPD

15706
CHNLLHCVY

16707
CNSTLQYGH

16708
DFCMLQMGM

16709
DHCILPQTC

16710
DMFMLDEAP

16711
DYFPLVVAP

16712
GANFLVSQT

16713
GKNPLMPSP

16714
GLDKLMMLA

16715
GVMMLGTAN

16716
HIDHLNAAT

16717
HPLPLSVMS

16718
HSMLLQEYM

16719
ENTALFVNA

16720
KEYDLSLTM

16721
KNFELDQFE

16722
KPYSLAMTE

16723
LGTPLEAGL

16724
LIDRLPACY

16725
LRDCLHVLG

16726
LVDELNQIK

16727
MFDDLMVGR

16728
MINNLDECC

16729
MVDGLTKEY

16730
NCQSLATNV

16731
NEVHLEHNY

16732
NLEMLAMFQ

16733
NVMPLIQYR

16734
PITILCCSQ

16735
PROILHFSQ

16736
QASLLDHYP

16737
QGQPLMRYS

16738
QQNRLNMNW

16739
QQSPLLWER

16740
QRHDLEVDH

16741
QVVDLVDVY

16742
RGEILQLQQ

16743
SAQCLEMWP

16744
SGLILQCNP

16745
SENYLGEGH

16746
SSKPLTNEC

16747
SVAFLNQEW

16748
SWNRLNWCL

16749
TCESLYSRG

16750
THDSLIAHW

16751
TIGNLTKPI

16752
TLMGLWNQG

16753
TNDPLMRSD

16754
TQCGLKMYP

16755
TRMLLEWTY

16756
TSEPLEGED

16757
VDSWLSMRD

16758
VIHYLTHEC

16759
YCIHLKDDI

16760
YCIPLQPMN

16761
YSVPLHHCF

16762
YVGNLCRWK

16763
YVVKLEDYK

16764
YYADLNAHD

16765
RNMALHHPT

16766
DGCSLYAFN

16767
ACDRLMFQG

16768
SLVDLYKTA

16769
FCNKLTSVR

16770
IDAFLANRG

16771
NMMTLWSHL

16772
QIVSKNSRQ

16773
RGVHSNCAE

16774
SAFLPNTVC

16775
SEYDNNPCQ

16776
SFTIYNHKW

16777
SSMTRNNCK

16778
THDYFNGDY

16779
TWANGNGGP

16780
VNHHENWKK

16781
IEWPTNSDH

16782
NAYVDNLRY

16783
QQCMGNDMT

16784
AFLHENMEG

16785
ANGTYNLQE

16786
ANMFRNGMQ

16787
DSYFINELY

16788
FFEMVNTEQ

16789
HSW1NNQEH

16790
KSVTQNRTR

16791
NSYFINELY

16792
TFAAVNCCQ

16793
TMKTFNYVF

16794
YCQLGNHNA

16795
YHENWNMPM

16796
GWMRHNREQ

16797
QSGYMNKLW

16798
VSMSSNYDK

15799
KCAMFNAGE

16800
AEYMTNASM

16801
AGYASNWAD

16802
AESPTNAER

16803
AKGTYNQGN

16804
ASETNNGGY

16805
AYQMFNNVQ

16806
DAEHKNAIR

16807
DAHYTNECR

16808
DCQPVNRIL

16809
DENFDNHAG

16810
DMNSRNVQD

15811
DMVPCNLMH

15812
DWAANNWWP

15813
FEAPINLSF

16814
FVHVENRCD

16815
GAMPGNGCT

16816
GHKVCNCWE

16817
GVWPDNCPQ

16818
GYCGCNHMG

16819
HHVWPNRQV

16820
HIEMRNAHM

16821
HMYVTNHDG

16822
ICSYYNQTQ

15823
IFYATNHSW

16824
IKHDFNSAF

16825
1NAPPNLRT

16826
KGDEQNCYC

16827
KEDVENCCC

16828
KPGTVNPAA

16829
KQFPCNSGH

16830
LCAGTNLTP

16831
LEADNNDAD

16832
LKIPFNMTC

16833
LMAYANTDD

16834
LRTPYNFWW

16835
LVKVINERQ

16836
MCTPRNEQE

16837
MKGFDNYSP

16838
MLSYHNESW

16839
MQNEKNDNP

16840
MRKDYNDTW

16841
MYQDANQIT

16842
NVHTTNYAF

16843
QFSGYNYHY

16844
QLHWPNNFG

16845
QMHVANINL

16846
QSNEFNPVA

16847
QVAPKNCWQ

16848
QYICFNCYH

16849
SACEPNVSP

16850
SANKMNAER

16851
SEGTCNTCG

16852
SIAMYNNMH

16853
SREAQNRET

16854
SVKDNNRRL

16855
SWTASNWRD

16856
SWTSRNAHH

16857
TAQYVNREI

16858
TQDVQNMYV

16859
TSMLHNEEN

16860
TVKEENAAC

16861
VFIAQNEGN

16862
VGVGVNRFD

16863
VHANANMSH

16864
VPELGNGAY

16865
VQLPTNKQW

16866
VSINTNEPG

16867
VVQYKNHAT

16868
YGTEVNSMI

16869
YQVANNQQG

16870
YYADMNAHD

16871
HMWMYNKCT

16872
ARTIANDTQ

16873
FREDMNATM

16874
SLWVGYRTD

16875
TPKNDQRAE

16876
LNARTMRSA

16877
MEDFSDRWP

16878
RMVIMERDR

16879
DGQLVERYD

16880
APKNDQRAE

16881
TYYWDCRHV

16882
LMEAASRSY

16883
TQQTADRWA

16884
IVTCNGRWD

16885
YDYYKMREG

16886
ASKAKQRVA

16887
FQATTKRLP

16888
NAQLGKRGH

16889
YLWMIARPD

16890
YLWMITRPD

16891
KQNPSMRQD

16892
SATARERDN

16893
AAEMHQRGE

16894
AAEMHQRGV

16895
AFKATRRGV

16896
ASGSHQREN

16897
ASGSHQRKN

16898
ASHIYCRHE

16899
DIKMQKRGV

16900
DVGLALRNH

16901
FQADYKRHE

16902
FVKMKQRYG

16903
GAGYRMRVN

16901
GVSVEGRDE

16905
GYKNCHRHG

16906
HIDLQTRKE

16907
HVEWTMREH

16908
IGGMFSRHA

16909
IQKLMRRMH

16910
KAISHERGQ

16911
LAYTCHRGD

16912
LHWYWCRNW

16913
LKKKRGRAC

16914
LQEPRMRYE

16915
MCQSYERGA

16916
MFNRIPRFS

16917
MICRMDRMK

16918
MQKAHDRSA

16919
MSGFHIRCA

16920
MSSFHIRCA

16921
NLIYQPRKS

16922
QATFTTRLV

16923
QGDHFDRNS

16924
QGGMMKRGQ

16925
QIDEVTRNL

16926
QVEPVEREY

16927
RCDGVMRSQ

16928
SAELVQREA

16929
SGSHCGRMA

16930
SLKMRPRQI

16931
SRGDYLRVG

16932
SWEWQMRYP

16933
TAIHEHRGL

16934
TIEPFWRSW

16935
TKYVDDRPG

16936
TSIPITRQE

16937
VINMNRRWG

16938
VSINMWRAQ

16939
WDNLMQRTC

16940
WIQPISRTG

16941
YGSYYMRGW

16942
YMHEWMRLT

16943
YSAPRIRDA

16944
SMNLWCRGN

16945
CDLFPIRWP

16946
TEGPMHRAW

16947
QGTGHSLFL

16948
RHWRHTMFL

16949
RMETWCGFS

16950
PMWTQMPFP

16951
DINASEWFC

16952
MIVMGPHFT

16953
SLYNCMFFL

16954
WMKVRQGFG

16955
ILEAIVNFW

16956
KISPWDGFY

16957
AALENGWFN

16958
ACMVNPGFW

16959
ACSTVMDFN

16960
ASIWRKLFD

16961
AVHAEDIFM

16962
DASVPGVFE

16963
DCNWHWPFT

16964
DKHFYQKFH

16965
DSSLHGAFM

16966
DSVNTSAFH

16967
DVIRDDSFQ

16968
FGMVHHGFF

16969
FKYYIPEFC

16970
FPGWCASFP

16971
HICDMDMFL

16972
HYFYTQQFG

16973
IAIPCASFY

16974
KNVMHSEFR

16975
KYDREWPFL

16976
LCCDVCMFL

16977
LGDTGLMFG

16978
LMSICAAFP

16979
LQGWRWWFA

16980
LVSDHIEFG

16981
MASSEHCFF

16982
MHIEGVMFQ

16983
MIWYNRKFM

16984
MNEIFVEFP

16985
MWTAQQTFM

16986
MYIPVYITN

16987
NVQFESTFF

16988
PAMPFYWFE

16989
QHQPFSMFS

16990
QNSHGPVFG

16991
QNVMHSEFR

16992
QQAPATIFP

16993
QQVEWSVFQ

16994
QSISDMPFP

16995
QVTPSMIFK

16996
RDSCYINFP

16997
SATCWLWFG

16998
SCMGFTSFP

16999
SDEWEDVFG

17000
SGCHWGIFH

17001
SMYSTHCFC

17002
SYPGHHWFY

17003
TASGNTYFV

17004
TILEGDNFA

17005
TITFIDLFN

17006
TKNAMHNFF

17007
TVSRASYFD

17008
VAVTVTDFD

17009
VMEKYAHFP

17010
WGDRYGLFV

17011
YSQWFTFFE

17012
QVHAKLGFM

17013
SATIRFYFN

17014
QHQAFFVFT

17015
QMKGATEFC

17016
LVNMFQCFW

17017
ASTERAKFG

17018
NVKWHCDGG

17019
PYFRFFPHG

17020
QCGAIYQTG

17021
QQNLADQAG

17022
RQGKAMSMG

17023
SCTHIKVPG

17024
SCTMKDWWG

17025
SCTPIKVPG

17026
FLNNEELKG

17027
WDNMDQTQG

17028
AMLRQEKTG

17029
DSQNNWPTG

17030
KCGWHGENG

17031
VQQNNLKAG

17032
VAANTYCYG

17033
GEPWFQTHG

17034
GEPWFRTHG

17035
GMASMEHCG

17036
GQGTSRLMG

17037
ICHLRHIAG

17038
MMLGYMEQG

17039
NAKPKQVQG

17040
PCYNAMNRG

17041
QIHPYLEAG

17042
SRDCVVHDG

17043
VCPSCRDYG

17044
WIKWKCSEG

17045
CLNNSTCRG

17046
QHSKCHMHG

17047
AADFSGYYG

17048
AAGATMLRG

17049
AEIPIMFMG

17050
AFVNWPIHG

17051
ANQPKHGAG

17052
APL1KASWG

17053
AWDWCDMRG

17054
AYNSRANGG

17055
CHGHFQHIG

17056
DAKTCHSRG

17057
DCKLMRHQG

17058
DIMLRTCCG

17059
DRCMQTKHG

17060
DSATQHMEG

17061
KCGWHGDAG

17062
FEGSRQLWG

17063
FREWRAMDG

17064
GCTPSVCSG

17065
GVMLMIAGG

17066
GYVPWLMEG

17067
HCHMIFKDG

17068
HKDDYVQTG

17069
ISEDIKENG

17070
ISWLCEMNG

17071
KNEWTYHQG

17072
LEDMMSVEG

17073
LIMEQPLAG

17074
LNAECTSTG

17075
LSTCWAIAG

17076
LVVNQYTLG

17077
MAAPTALNG

17078
MIGTIQSDG

17079
MKASTMELG

17080
MKVYHEMVG

17081
MQVSADHHG

17082
NAAWQSITG

17083
NCSHYDSRG

17084
NGMVSGQYG

17085
NHPYQHIEG

17086
NKMHCDTMG

17087
NKVMCLWQG

17088
NNAPVWLHG

17089
NSVSCGGQG

17090
PGDFTSGYG

17091
QCDEQQEAG

17092
QFGSFWPTG

17093
QNARSSMVG

17094
QYDTIQLNG

17095
SCHHVGMCG

17096
SFKWTIVNG

17097
SIMKYDQMG

17098
SMHHTQFTG

17099
SVGFPYESG

17100
TAQYDVWHG

17101
TGYPKLKDG

17102
TQEPWGCAG

17103
TQGLPRGRG

17104
TRLYNWPMG

17105
TSHHWAAQG

17106
TWWKDPQEG

17107
VAHYYCWEG

17108
VLMNWQPRG

17109
VLNHWEPIG

17110
VVMHPYAAG

17111
VVSHYDMIG

17112
WHTVHYMGG

17113
WLTFDRFEG

17114
AKCPWHHTG

17115
VNPWYVKHG

17116
DVASRCWVG

17117
SGTPHQQSG

Example 13
AAV5 Variants with Tissue Tropism in Heart

This example describes engineered AAV5 variants with tissue tropism in heart that were discovered using the methods and systems described in EXAMPLE 1-EXAMPLE 2.

CNS, liver, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord tissues were analyzed to identify amino acid residues in the AAV5 VP1 581-589 region that drive heart tropism. The results are shown in FIG. 18D which shows the positional frequency of each amino acid in the 581 to 589 region of AAV5 VP1 in heart tissue as compared to all other analyzed tissues, wherein the capsid polypeptide has the sequence of SEQ ID NO: 2. The position (“Xaan”, n=1-9) is denoted on the x-axis and amino acid residues are denoted on the y-axis. The frequency of a particular amino acid residue in a particular position is indicated in the heatmap as intensity. Darker squares indicate a higher frequency of a particular amino acid in a particular position and lighter squares indicate a lower frequency of a particular amino acid in a particular position.

The frequency of a given amino acid residue occurring at a specified position corresponding to position 581 to position 589 of SEQ ID NO: 1 (generalized in SEQ ID NO: 2) in variants identified in heart over the frequency of that given amino acid residue occurring at the specified position in variants identified in all other harvested tissues was analyzed to identify a set of sequence rules for capsids that preferentially target heart tissue. With reference to TABLE 25 below, and SEQ ID NO: 2 (AAV5 VP1), the following amino acids can, thus, be independently mutated, in any combination, at any one or more positions Xaa1-Xaa9, to provide a VP1 capsid with enhanced heart tropism as compared wildtype AAV5 VP1 capsid (SEQ ID NO: 1), where heart tropism here refers to properties that are preferred for heart transduction over properties that are preferred for transduction of all other harvested tissues.

TABLE 25

Heart Tropism Rules

Xaa1 is selected from I, K, L, M, T, or V

Xaa1 is selected from K or L

Xaa1 is K

Xaa2 is selected from A, C, G, I, K, or S

Xaa2 is selected from A, C, or S

Xaa2 is A

Xaa3 is selected from A, D, E, G, K, M, or V

Xaa3 is selected from E or V

Xaa3 is E

Xaa4 is selected from F, H, R, T, W, or Y

Xaa4 is selected from F, R, or T

Xaa4 is R

Xaa5 is selected from F, L, M, or R

Xaa5 is L

Xaa6 is selected from A, H, N, W, or Y

Xaa6 is selected from H, N, or Y

Xaa6 is H

Xaa7 is selected from A, C, E, F, K, or T

Xaa7 is selected from C, F, or T

Xaa7 is F

Xaa8 is selected from A, C, M, S, or T

Xaa8 is selected from C, M, or S

Xaa8 is C

Xaa9 is selected from A, D, G, or P

Xaa9 is selected from A or G

Xaa9 is A

TABLE 26 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that were found in heart tissue and comport to one or more of the rules provided in TABLE 25. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 13118-SEQ ID NO: 14117, as disclosed in TABLE 26. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 26

Sequences of the 581 to 589 Region in AAV5 VP1

Capsid Polypeptide that Drive Heart Tissue

Tropism

SEQ

ID
581-589

NO
Sequence

13118
KAAPMDISI

13119
KADCPAYKA

13120
KADVWMAIA

13121
KAELSDHVL

13122
KATDALPSC

13123
KCSKQASYP

13124
KDGYALLSC

13125
KDNEAQYQP

13126
KDYMFNLPF

13127
KEAMMYNAN

13128
KEGLIFSTE

13129
KEHTNECRH

13130
KEIVEVDGN

13131
KEMAYQHMW

13132
KEMLAMWLQ

13133
KEWFVTWKA

13134
KFLPCITSY

13135
KFPYLPHRN

13136
KGCMSWEEG

13137
KGDRFCGEV

13138
KGEPGYMYY

13139
KGMFVANSH

13140
KGTKGPQMQ

13141
KHAGTRYKG

13142
KHFTSCDRT

13143
KHHDAAREM

13144
KH1FNECDC

13145
KHMQKYVHY

13146
KHPRFTGWH

13147
KIESDWHAQ

13148
KKHQFCEGT

13149
KLELHGCQN

13150
KLMSFRELE

13151
KLQLEKPLN

13152
KMDCHKYGV

13153
KMNETTNPR

13154
KMVDQSLAT

13155
KMVMEDTDV

13156
KNFMNHEPQ

13157
KNNRTIRWG

13158
KNNTWKWWY

13159
KNQYFETSQ

13160
KNWEKQNEV

13161
KNYNPVNYE

13162
KPCKACLNS

13163
KPCWDQAMQ

13164
KPDQKLWWD

13165
KPDWWIKVN

13166
KPSNSKQTV

13167
KPTQHGVGP

13168
KQAEGDDVG

13169
KQH1FQWHH

13170
KQ1HEWEGQ

13171
KQLPEQOPL

13172
KQMLEFAPY

13173
KQNEINHSC

13174
KREDHYGWG

13175
KRSFEWLER

13176
KSDYNSYDA

13177
KSLSSEMGV

13178
KSMDKDLTS

13179
KSSCVDDSS

13180
KSVAEGAND

13181
KTAQLSVME

13182
KTAYELVDG

13183
KTCDSQTMT

13184
KTEMYSDQC

13185
KTFEWLCPY

13186
KTMPVGAWD

13187
KTTWVDDFD

13188
KTYYVMAEV

13189
KVLDTLTEH

13190
KVMHACNAN

13191
KWDETMFMS

13192
KWEDFGWWK

13193
KWEQPNPQA

13194
KWEYVRCAS

13195
KWLEARMYL

13196
KWLKTLMCL

13197
KWQDAHYIL

13198
KWTLDAWLT

13199
KWWVDRLCQ

13200
KYAVSPCDP

13201
KYDWLDAGG

13202
KTDPEAEYC

13203
KGNPLDGDT

13204
VAHICDPET

13205
VASEHSCLP

13206
VAVEDRPAL

13207
WAREELGFA

13208
YAMCHRWMR

13209
YAPNNSHWP

13210
AACPHQFKD

13211
AADLGQSMC

13212
AAESAKFHE

13213
AAGIAVACT

13214
AAIRCMQTS

13215
AAKLYMDLS

13216
AAMKGEV1G

13217
AAMLRLLGK

13218
AANMLYCST

13219
AANWTHCSL

13220
AAPMHLHEI

13221
AAQFMEKNQ

13222
AASGLEVVA

13223
AATNWGVET

13224
AAVKWDQPL

13225
AAVPELHHS

13226
AAVQQPEHT

13227
AAYSQSAGS

13228
CADSCYWYP

13229
CAEAWFPQR

13230
CAFLDWDWC

13231
CAFRWVEWD

13232
CAHHVMWYP

13233
CAHQPDGYA

13234
CANAENATC

13235
CANCVAISP

13236
CAQPNTHNS

13237
CAVAWKGDY

13238
DAAQCTWAR

13239
DAHPEGPQR

13240
DALEYHSPF

13241
DAMRTHENT

13242
DAQKQQVMH

13243
DAQWIGCLE

13244
DARWRSQCP

13245
DARYSWPGA

13246
DATFNTSSL

13247
DAVLFTAMQ

13248
EAFGNGIDP

13249
EAGEHQKVE

13250
EAGRNYMDL

13251
EANQCHVHI

13252
EATWTAQGA

13253
EAYLWENQL

13254
FAEWKTEQQ

13255
FAHCAMNES

13256
FAHWRDCWY

13257
FAKIRERHV

13258
FASCMRGRA

13259
FASPAGPWS

13260
FAVNEHDVF

13261
FAYATIRRD

13262
GAEIMVNGW

13263
GAEPYVKGG

13264
GAERQWFTP

13265
GAHVNSMNA

13266
GANSSELQQ

13267
GAVMWERDG

13268
HADCYVADF

13269
HADRMEGPW

13270
HADRPMSAC

13271
HAEWTEPVD

13272
HAIPACKRD

13273
HAKLETMWG

13274
HALAWCTKH

13275
HATYCQMAM

13276
HAVSQERAG

13277
IACVHMKQW

13278
IADERWCMA

13279
IADYEQCSS

13280
IAFETYWTN

13281
IAGQWAAFG

13282
IAKCIDVMD

13283
IAKWIEKIG

13284
IAMLSQHTV

13285
IAQPYDNWG

13286
IAINHSAGP

13287
IAVTGFAMS

13288
LADGMGKVP

13289
LAFSDITWS

13290
LAHSEDRTS

13291
LALQUAGAH

13292
LAMWFNESC

13293
LAMWKMRQH

13294
LATMEDQSN

13295
MAAPYORYN

13296
MAAVMIDQA

13297
MACRAWIFV

13298
MADKCANNA

13299
MAEQAVNSK

13300
MAGNVCLCV

13301
MAHMNTGVP

13302
MAKWDDSFS

13303
MALMVEECD

13304
MAMLCSICS

13305
MANPEMYMQ

13306
MAQFVWCNG

13307
MASMMNACN

13308
MASMSKDDS

13309
MASQEHQES

13310
MAVHRPTFC

13311
MAWDIMFWP

13312
MAYQQVIAG

13313
MAYQRHAAF

13314
NACIRRTTW

13315
NACIRWAFD

13316
NADALGALQ

13317
NADREMFEH

13318
NAEHRGMTN

13319
NAGAQWLGI

13320
NAGLEWKLQ

13321
NAHCGAACC

13322
NAMAYENDY

13323
NAMLTLGAG

13324
NAMQRANIR

13325
NAPCFRYYN

13326
NARDDSIFN

13327
NASHYEKDG

13328
NATSYHKFM

13329
PAEIKMPSQ

13330
PARLMMWEF

13331
QADQYYSLE

13332
QAMLKQGPR

13333
QASCISWSQ

13334
QASQENRLT

13335
QAVECTQAP

13336
QAYTGDKDI

13337
RADDYQPVS

13338
RAELFNNGK

13339
RAEQADWHE

13340
RAILNEKRD

13341
RAINFDKAS

13342
RAPELVMNE

13343
RAPYEADSN

13344
SAAYIVKGP

13345
SADGQQAAT

13346
SADLSLTYD

13347
SADQVPECP

13348
SA1YRNWFE

13349
SAKLSEQNV

13350
SAYALQSPR

13351
TAA1VGKES

13352
TACMDHHIR

13353
TACNMMCAY

13354
TADWYNGGP

13355
TAEVWWVFH

13356
TAGFAKHFP

13357
TANMCDEGI

13358
TANSEAHYT

13359
TARWLDEHL

13360
TARYITSRA

13361
TASEFPCGV

13362
TAWKADSGM

13363
TAWWNDCWF

13364
VAAGPLFYG

13365
VACAPEAMG

13366
VADGRQPME

13367
VAFYNKMRG

13368
VAIQSVGYH

13369
VAIVWMCSE

13370
VALPRCQLV

13371
VAMYTRCHT

13372
VAPKDNQQM

13373
VAPQCTNEH

13374
VAQPVQDMG

13375
VASKTQFAM

13376
VAYYFDQAG

13377
WAPPCYYDV

13378
YAANKDCFL

13379
YAKYGQRWM

13380
YATFEVWFR

13381
YATMEYNNN

13382
EAFQKCNWQ

13383
HATGLVNFM

13384
AADIFAIND

13385
EAELENVNM

13386
IAYHYDQQN

13387
TSEPLLAML

13388
TTEDCPNRV

13389
TTEYTCEDQ

13390
VFENCSRTF

13391
VGEWEDEHD

13392
VIEHAHHVN

13393
VSEGQVNMD

13394
VSEHDDCGL

13395
VSEVYLAER

13396
WYEKLMDAF

13397
YCEVPQNPV

13398
YDEYCESKS

13399
YIEIVRGPM

13400
YSEDATPKG

13401
AFEFTDGCC

13402
AFESAADHW

13403
AIEVEGMWD

13404
ALELEAHRP

13405
AMERIKMRN

13406
ATEAYSKLQ

13407
ATEKDHRVC

13408
AVEFQHGHC

13409
AVEIHKCID

13410
AVEPGISPF

13411
AYEHDFKCS

13412
CCEIKRHYY

13413
CIEKSISST

13414
CLEQMCRTN

13415
CPEPFQKVD

13416
CREQNNCDR

13417
CSEHGCDIP

13418
CSEWIRAVR

13419
CTEQYLVDK

13420
CTEVRNWYD

13421
DCEFPMGAN

13422
DDEGYIFNQ

13423
DGEFRYVVS

13424
DGEQESGQT

13425
DHEVSVAGS

13426
DIEMCPCYN

13427
DKEIVNMVW

13428
DLEYVEFDW

13429
DQEVFQTHF

13430
EEEFWMHNA

13431
EIEPTIGPE

13432
EKETMWAEP

13433
ELEKYKLTN

13434
ELEVRHSND

13435
EVEEETDEK

13436
EWEYERGAE

13437
FCETEPSGA

13438
FKEHVQNHM

13439
FQEACMACH

13440
FSEPSNNFC

13441
FWESTSPFT

13442
GEEWVEPTA

13443
GHEHGWQLS

13444
GNERLSVCI

13445
GNETLEMHG

13446
GRECKQGGH

13447
GTEHWLSAC

13448
GYENYNKRD

13449
HCEYCQMMN

13450
HFEAGYACS

13451
HIEGYFRWP

13452
HIEYTNKCL

13453
HPEKARPLA

13454
HPEMKHQNH

13455
HVENALVFT

13456
ICEEQFLMG

13457
IHEEQECGS

13458
IHEGYKTAP

13459
IMEQSAQHT

13460
IRECGQHGS

13461
ITETWDIFF

13462
LDEFGFWTH

13463
LGESEHCMA

13464
LKEAWICKG

13465
LKEGINHIC

13466
LMEGSAKAM

13467
LYEGRPWYN

13468
MDEQMGKMS

13469
MIEKQYPTM

13470
MKEESKSWP

13471
MLENYEHMG

13472
MNEPQKMRQ

13473
MPEYMKTMQ

13474
MPEYQARLM

13475
MREQMPVQY

13476
MRETVFHEP

13477
MTESGMKDK

13478
MVEVSRGEA

13479
MWECGYGGN

13480
MWERIGMGG

13481
MYEDTHCVC

13482
MYEHINSQR

13483
MYEKQDGVL

13484
NGEGSQTCE

13485
NHEQKSYIF

13486
NLEIYLQMN

13487
NVEFMRCNS

13488
NVEWWAPHV

13489
NWEQEFVMI

13490
NWEYMRRKC

13491
QDERMDLVQ

13492
QIESGCPHC

13493
QRELITMGC

13494
QSEVTHLKW

13495
QTEQWMCSH

13496
QWEHIWCWH

13497
QYENINKVD

13498
RDEVVFAHC

13499
RNEPNVWVN

13500
RTEQSRVWE

13501
RVEVWYGEL

13502
SEESVVTHF

13503
SHEQLAGCP

13504
SMENRWEYL

13505
SMEYYYGME

13506
SNEQMMTMQ

13507
STECSGLSG

13508
SYEVIYHMD

13509
TCEKLISVC

13510
TKELTRSEA

13511
TNEAIMQVC

13512
TNEIGEWWF

13513
TSELLVNGD

13514
TSENREYMQ

13515
TWEKQSRNG

13516
TWELIQTKF

13517
VCEEAGELG

13518
VHEYYRSCA

13519
VLEFCKDEE

13520
VMEKQGNGG

13521
VMEYMLIAW

13522
VNENNELSA

13523
VNEQWQIMC

13524
VNEWSADCS

13525
VSEFHDLHL

13526
VSELCTNMA

13527
VTERWIGPM

13528
VYEKFSYIT

13529
VYEREQMDV

13530
YHECKEGHI

13531
YSENRIFFR

13532
YYECLGQDL

13533
WIEGTLIGA

13534
NWEFNARHD

13535
YWEKVPCMK

13536
GNEDCNLAC

13537
MNECFSGAQ

13538
TYESLQNSW

13539
YCECGHWPN

13540
CWECCTDSG

13541
VCPRDMTWE

13542
VPCRPIYQH

13543
VRWRGDVAN

13544
WVLRCTYEY

13545
ADARVMTMM

13546
ADLRPSVDY

13547
AGDRSQILG

13548
AMMRAWSTF

13549
AQDRFMLGC

13550
ATNRIMAAW

13551
AVVREHVES

13552
CIWRKYPEF

13553
CQCRFCHPL

13554
CTIRVQNGE

13555
DEFRFLPAD

13556
DGGRFDDYY

13557
DHWRKLHKA

13558
DIARLQVLG

13559
DKYRQQENS

13560
DQIRTLQMT

13561
DRLRDDYPC

13562
DRPRPMWGH

13563
DVIRIVHDY

13564
DYARNHMMW

13565
DYTRYHSYF

13566
EGNRPEGLA

13567
ERVRPLSTD

13568
ESDREEQSR

13569
ESWRECMCR

13570
ETMRCQTMA

13571
EVARGQDWA

13572
EVNREETIQ

13573
EYLRESMMP

13574
FDDREPFNG

13575
FDGRPRMFE

13576
FGPRKNRHE

13577
FPRRLDHHQ

13578
FPYRIEGED

13579
FPYRKVECG

13580
FTARSYQDA

13581
FVDRVDKHH

13582
GNDRAEPDR

13583
GTCREYGRN

13584
HGARCCWGV

13585
HLSRNEYKS

13586
HSLRDWFKH

13587
HSPRRATMV

13588
IHGRMINSI

13589
IPPRYRHMA

13590
IRLRDPELH

13591
IWIRWPSMG

13592
LFDRVGIEA

13593
LGCRETRHD

13594
LIMRNASWC

13595
LW1RWGSCT

13596
MFIRIRWLV

13597
MKCRIEKVN

13598
MVYRIKMTY

13599
NFHRWDMNT

13600
NRDRSWCQH

13601
NSDRMHMIE

13602
NYPRDHRQH

13603
NYYRALKWD

13604
PVTRWEAGS

13605
QIDRAEKQE

13606
QNVRSNAYT

13607
SDLRNLCAR

13608
SHRVLRSN

13609
SIYRMVMQC

13610
SNDRFPIMC

13611
SQGRMASGA

13612
SQTRDTLNF

13613
SVKRFQHNN

13614
TQQRVYSIE

13615
TYDRMQKCN

13616
VPCRGLVVL

13617
VPLRHLTDH

13618
VTDRAFSMN

13619
VTPRHKTLF

13620
VITRVEHFP

13621
VVMRDEGGR

13622
WCHRVGCTG

13623
WDGRFQAAY

13624
WNLRKANAP

13625
YHDRFERAH

13626
YHYREPSAD

13627
YPGRQHVAN

13628
YRIRKTCAF

13629
YVNRYGAGW

13630
NNVRKVGNT

13631
TSDWLKHFC

13632
TWSLLQQGD

13633
VRAYLMGPN

13634
VSHALIDWH

13635
YPKQLHVWT

13636
ACHTLDQQD

13637
AHQVLHVRS

13638
AKRELLDHD

13639
ANCYLNHTT

13640
APKKLEHVD

13641
AQNALWERI

13642
CFAALDYAS

13643
CGNALPLIH

13644
CNDCLMFCD

13645
CNTMLVSAG

13646
CSMYLMEFG

13647
CWYMLKAYT

13648
DFVQLINRA

13649
DGYHLEGQC

13650
DIKALMDGW

13651
DKSKLDVSY

13652
DTCKLGGWF

13653
EHMLLVEQL

13654
EYNGLGQAY

13655
FCCLLYARR

13656
FFSGLEGRH

13657
FTDQIFTNP

13658
GGNILDIGP

13659
GIIPLHWQP

13660
GSDLLVYSA

13661
GSMLLGGME

13662
GTMLLWRQN

13663
GVANLIGLT

13664
HELYLTTNT

13665
HSACLMTMS

13666
IDTALNVHC

13667
IENWLFVWL

13668
IMTMLTSET

13669
INAVLWWRL

13670
ITHGLQTWD

13671
ITTDLQMDH

13672
IWNELILGC

13673
LDCELYQTR

13674
LGMYLQLGQ

13675
LGVQLSSAP

13676
LHLKLYGQL

13677
LLTLLMYWV

13678
LMYMLPPRH

13679
LRPWLPDST

13680
LTHALGTWE

13681
LWTHLMANH

13682
MDQKLLTNQ

13683
MEDMLMEGH

13684
MHMLLHDVK

13685
MITWLTDQV

13686
MNVGLWALW

13687
MYACLAQPP

13688
NHCMLHHMG

13689
NHDHLFERY

13690
NQPHLWSCG

13691
NTRDLHTKR

13692
PNGLLQMVI

13693
Q1AALHQNH

13694
QLGMLPRAM

13695
QMVMLKDQY

13696
OQGQLKTWS

13697
QVSCLVRDD

13698
RTWTLECDF

13699
RVNILEKIA

13700
SFAHLIDDR

13701
S1AVLCHGN

13702
SIDALKSAF

13703
SIVKLNRSV

13704
SLNKLFHQE

13705
SLQFLYHGS

13706
TERLLQQFY

13707
TFATLCDQQ

13708
TIMGLLGNV

13709
TMYPLMHYS

13710
TPAQLNGYG

13711
TQCMLHGPY

13712
TQGWLTLCY

13713
TSDWLQNYI

13714
TTQCLDRWQ

13715
TTWILPHHD

13716
TYVPLWAYW

13717
VEVQLKALQ

13718
VMHCLRQFM

13719
VRPWLVMDN

13720
VTDGLGFRQ

13721
WVRDLKCPV

13722
YRIFLMHLC

13723
CIDVLFKHL

13724
VFKPLDKRY

13725
TQGSVHVMN

13726
VEPTWHCDW

13727
VISNRHHLP

13728
WRVSAHYYI

13729
YDHTIHRNP

13730
YLLYPHSSG

13731
ACDLQHNIA

13732
ADQQQHKVI

13733
ADWHCHRND

13734
AEAQFHTGE

13735
ANHWKHEWE

13736
ASGWGHCWR

13737
AVIKAHCAA

13738
AYCHQHQHA

13739
AYMPYHKEQ

13740
AYQAEHVGM

13741
CINWKHVAS

13742
DDHWHHRAK

13743
DFLTAHCIE

13744
DFLYMHRMG

13745
DGDAAHTKP

13746
DHQLCHVAL

13747
DHVDNHEEA

13748
DNAYFHIKG

13749
DPCNHHWCI

13750
DVVYTHCIN

13751
EENHSHDER

13752
EINFQHRHV

13753
EIQWNHWYV

13754
EKDMHHVAS

13755
ELLKVHGPK

13756
EMNGRHWWP

13757
EPAVTHDSA

13758
EQHIMHEQW

13759
ERMVCHWDA

13760
EWPINHQKV

13761
FDKAFHVHM

13762
FKPIVHHCG

13763
GEDQIHVWH

13764
GHQSEHRCS

13765
GINSHHNDL

13766
GKQNNHRIE

13767
GNKYMHHHA

13768
GPKPVHFDV

13769
GSVMGHCGS

13770
GWGCGHGVH

13771
HGYNWHHLG

13772
HTLHRHWFW

13773
HTQFAHGVD

13774
HVAIGHGGF

13775
IERHEHNPY

13776
IIPHFHLCY

13777
IKCIQHVHY

13778
IKDITHNHP

13779
LCRQTHKWV

13780
LFGVYHFSG

13781
LICKYHYCQ

13782
LQPMMHTHS

13783
LTLCMHEAD

13784
MCGLYHTEG

13785
MCHGCHRSY

13786
MFDNCHVFP

13787
MNDTYHRRA

13788
MNLTIHEQF

13789
MQMFKHNWI

13790
MTGVEHVFY

13791
NCFWTHVSG

13792
NDYPIHCYS

13793
NFTSFHTYS

13794
NHMLTHHYH

13795
NYAFNHQGV

13796
NYQQHHFFS

13797
QHSWKHFDS

13798
QILHYHYHG

13799
QMNWKHVWN

13800
QNSERHDEP

13801
QRMFTHQEY

13802
QSFSPHFGV

13803
QSVHTHANL

13804
QTGNAHWQE

13805
RCNEDHEQD

13806
RIIEQHAHG

13807
RQCEMHTVD

13808
SCGWKHDNP

13809
SCTEMHAGR

13810
SLYVHHKDG

13811
STGLQHDFN

13812
TEGQTHHTA

13813
TERSAHVTY

13814
TFKWMHEGW

13815
TFNNCHMQS

13816
THTQQHVVN

13817
TMSDRHICH

13818
TNATSHWCG

13819
TNGWMHHWD

13820
TTMIRHHGH

13821
TVFGTHRAL

13822
TVGMFHCQG

13823
TYTPRHRFS

13824
VRKWQHHGG

13825
VTTHCHHTY

13826
WEAKWHAWE

13827
WGFNDHKRG

13828
WRGIGHADL

13829
WTTAFHKRT

13830
YIVQTHCGG

13831
YKCHDHCHD

13832
YNNFHHQMY

13833
YRNEPHQGF

13834
YTNYKHCCF

13835
TGCNHHGRF

13836
DVCDDHKPN

13837
DVAPMHQVE

13838
VFKPNSFQT

13839
VPIQFSFAS

13840
VPLWDVFQT

13841
VTGVTIFEL

13842
YTPTRYFGK

13843
YVGWHMFLD

13844
IWKIQEFGT

13845
AGQPVTFHF

13846
AHASHMFGG

13847
ARQMICFGG

13848
ATYYNQFHD

13849
CCTWKCFCQ

13850
DFPLFTFGN

13851
DIRHMSFEF

13852
DPSQPGFNA

13853
DVAHKCFIS

13854
EGGKERFQQ

13855
EKMTWVFAG

13856
ENTTCNFQG

13857
ESLGCEFML

13858
FGCCGRFVI

13859
FTGKRDFMK

13860
FIMWHRFAW

13861
GDNAKDFRG

13862
GHANALFAD

13863
GIRSTDFYQ

13864
GMIVDVFWV

13865
GMTIMAFQG

13866
HGAKAPFTA

13867
HNDYPQFQP

13868
HVNAGSFGS

13869
IFILRTFFA

13870
IFPWFKFHW

13871
IQVHSLFGC

13872
IRPFIEFLQ

13873
LHQKPNFDC

13874
LIHHCTFEQ

13875
LQLEDRFNG

13876
MCSHYQFMS

13877
MFYLEDFRQ

13878
MITQFIFNI

13879
MTVMQGFLS

13880
NPQWGLFNQ

13881
NTGARPFAG

13882
PCLQTRFAQ

13883
PMYQDMFTL

13884
QGDLMGFHY

13885
QIGMNTFMS

13886
QMHVMIFGS

13887
REWDHYFQI

13888
RLRISPFNI

13889
RMGAWSFPS

13890
RVTTAEFLC

13891
RWSQNKFQR

13892
SDHDNVFED

13893
SHTTYDFFK

13894
SNVCNSFVN

13895
SPHAGPFGC

13896
SQFMMDFGP

13897
SQVPGRFIF

13898
TCCLYEFIH

13899
TGGTAEFGL

13900
TSNWHEFTN

13901
TSPCCTFGI

13902
VHDAQGFCA

13903
VKMMWDFAD

13904
VTHITEFMA

13905
VVSTDMFIH

13906
WCAPAVFNE

13907
YFREKPFDV

13908
YQCKMAFHV

13909
GVWMQGFHP

13910
MWGCKLFVC

13911
YLDSSECCW

13912
YVKPCYDCI

13913
AGLQHDTCV

13914
AITHVMHCH

13915
ASVLQQLCV

13916
ATTGMLACH

13917
AVGATMMCH

13918
AVHDNFWCL

13919
CCKYNQLCH

13920
CDKIDPYCD

13921
CDTDESGCY

13922
CHRCKVNCQ

13923
CITPYRPCD

13924
CMYSWVPCG

13925
CVHYCFGCS

13926
CVKSYEKCF

13927
DEHAHWSCS

13928
DEHQMMECG

13929
DIGFRIDCD

13930
DMCEFMYCD

13931
DMFYWQPCE

13932
DRRDHEACN

13933
EFDSPNQCY

13934
EFHPHDWCL

13935
ERGFMEHCH

13936
ESMSTRACG

13937
ETDPYKSCH

13938
ETVSPVGCH

13939
EVAQTMRCA

13940
EYQLSETCT

13941
FHKCYSACR

13942
FKDMIARCP

13943
GDDCKFDCL

13944
GTGSTWECT

13945
GVAVTTCCR

13946
HSNMRSACE

13947
IKRTYPACW

13948
IMCECQHCE

13949
IPFPMCGCC

13950
IRAPFNRCE

13951
ITCTYRNCE

13952
ITDYQSDCY

13953
LCALHEQCQ

13954
LSVSSDACY

13955
LVVAHCGCV

13956
MDAYFNSCQ

13957
MGTFVAHCY

13958
MIALNQNCT

13959
MMPSYTLCW

13960
MPAWMMNCH

13961
MQQCDGGCG

13962
MSWSVYWCI

13963
MTNCIEYCA

13964
MTTLYQSCL

13965
MVHGALMCR

13966
MVRSREQCI

13967
NGGKGQQCD

13968
NLGYCEHCY

13969
NLMTPWPCQ

13970
NSNMMAHCE

13971
NWVPVYDCH

13972
PFILSNCCA

13973
QNGVWENCE

13974
QSDGFEDCS

13975
QTTNNQRCD

13976
RDTKRSCCH

13977
RFVDFGSCQ

13978
RHYYWVLCP

13979
RNMYFVDCM

13980
RYDKVCYCV

13981
SGFSEKCCK

13982
SGRPKLVCN

13983
SHDPHNTCG

13984
SIGYYCHCM

13985
SILPTWGCD

13986
SKHDFDHCF

13987
THILYSWCI

13988
TIFYCRWCN

13989
TKNEYGLCF

13990
TVQYGRGCS

13991
TWGLKWMCP

13992
VDMVVDNCC

13993
VFGDGAHCI

13994
VHDQRLVCD

13995
VNPCAQGCH

13996
VNPQFSQCT

13997
VPMYTQVCN

13998
VSALIDHCC

13999
VTGPMPTCL

14000
VTMQVRQCA

14001
VVMQKIGCI

14002
VVNTVGCCV

14003
WHLFSACCS

14004
WHMLCDGCS

14005
WMRTQCWCP

11006
YGGLSCGCL

14007
YGPSTEPCQ

14008
LSAQRLVCD

14009
WVHPYSYCA

14010
TMHDIIVTA

14011
VPNVQNMKA

14012
VWAWQNGYA

14013
AEAIWLPWA

14014
AECLGYVDA

14015
AHVVHTSGA

14016
ALLFAPARA

14017
AMPNGLDVA

11018
ARLCTWPYA

14019
ATNKNCAHA

14020
CCFQNNTQA

14021
CLNGWKQAA

14022
CSQSYAADA

14023
CTNHNDHDA

14024
DFIMMQDQA

14025
DPLVHTMIA

14026
DRDNVDSQA

14027
DRSEMGAYA

14028
DRSVVDVDA

14029
DTDMIAQFA

14030
DTWKVMMAA

14031
EGQSMMYQA

14032
EIVGQKGWA

14033
EKQLYMYEA

14034
ELTIIMVVA

14035
ERDGWNMAA

14036
ESSLFEHYA

14037
ETQWWDVIA

14038
FVIPSGAGA

14039
GGHNYGMHA

14040
GIAASCHLA

14041
GSCSTLISA

14042
GSLPYSMYA

14043
GTMIYSHMA

14044
HHGQCGGMA

14045
HTFLCNITA

14046
IISAKQHDA

14047
IKCIPGWYA

14048
ILAQANIHA

14049
ILHSVTVRA

14050
INRQCDMIA

14051
1QKYGAEVA

14052
IQMLVQGSA

14053
IRMHMPRDA

14054
ISKPYQLTA

14055
ITKCDGNTA

14056
IVTASVCQA

14057
LNSLVCSFA

14058
LTKIVTTGA

14059
LWCVSNSFA

14060
MEIGIMKIA

14061
MERIMLSLA

14062
MGWYIVRHA

14063
MHNWGGKIP

14064
MHWLKYNYA

14065
MMAHSGHMA

14066
MMDPCNPPA

14067
MNGMPTSFA

14068
MPDSWVRVA

14069
MRIAQCNGA

14070
MSMQHLEHA

14071
MSTHEDHVA

14072
MVNTGGNTA

14073
MVTCVQSGA

14074
MWMMQGMNA

14075
NCDQMSISA

14076
NCKLESDLA

14077
NDMYVKGTA

14078
NINGIINQA

14079
NRTDRMMDA

14080
NTCQHVNFA

14081
PCSTWLLAA

14082
QKMFGDSYA

14083
QNMSNWLDA

14084
QQVCDIPSA

14085
QSTKEYWQA

14086
QVMCMCAAA

14087
RTNLQQVDA

14088
SCAQERHDA

14089
SILTGGGFA

14090
SLVCHRNFA

14091
SMDVHCNDA

14092
SMKLLSEAA

14093
SMNATRWKA

14094
SQACSNHYA

14095
SYMLKSDYA

14096
THVNRARWA

14097
TRCTCERYA

14098
TTDACWQHA

14099
TTHHAQAGA

14100
TTSMGKHYA

14101
TVGLDAHSA

14102
TYWPGCANA

14103
VCTAINDNA

14104
VIDTIAGHA

14105
VLSLFKEPA

14106
VNQQIESDA

14107
VTQLVTRLA

14108
VTTQIKRIA

14109
VWHVPRDQA

14110
VYCGNKTKA

14111
YDAKSSVNA

14112
YGREKQAYA

14113
YVHADPEFA

14114
YVKIKEAIA

14115
YWLEVDSKA

14116
DVQGSGQWA

14117
SVLHVQALA

Example 14
AAV5 Variants with Tissue Tropism in Colon

This example describes engineered AAV5 variants with tissue tropism in colon that were discovered using the methods and systems described in EXAMPLE 1-EXAMPLE 2.

CNS, liver, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord tissues were analyzed to identify amino acid residues in the AAV5 VP1 581-589 region that drive colon tropism. The results are shown in FIG. 18M which shows the positional frequency of each amino acid in the 581 to 589 region of AAV5 VP1 in colon tissue as compared to all other analyzed tissues, wherein the capsid polypeptide has the sequence of SEQ ID NO: 2. The position (“Xaan”, n=1-9) is denoted on the x-axis and amino acid residues are denoted on the y-axis. The frequency of a particular amino acid residue in a particular position is indicated in the heatmap as intensity. Darker squares indicate a higher frequency of a particular amino acid in a particular position and lighter squares indicate a lower frequency of a particular amino acid in a particular position.

The frequency of a given amino acid residue occurring at a specified position corresponding to position 581 to position 589 of SEQ ID NO: 1 (generalized in SEQ ID NO: 2) in variants identified in colon over the frequency of that given amino acid residue occurring at the specified position in variants identified in all other harvested tissues was analyzed to identify a set of sequence rules for capsids that preferentially target colon tissue. With reference to TABLE 27 below, and SEQ ID NO: 2 (AAV5 VP1), the following amino acids can, thus, be independently mutated, in any combination, at any one or more positions Xaa1-Xaa9, to provide a VP1 capsid with enhanced colon tropism as compared wildtype AAV5 VP1 capsid (SEQ ID NO: 1), where colon tropism here refers to properties that are preferred for colon transduction over properties that are preferred for transduction of all other harvested tissues.

TABLE 27

Colon Tropism Rules

Xaa1 is selected from C, F, H, N, P, W, or Y

Xaa1 is selected from F, P, or W

Xaa1 is P

Xaa2 is selected from D, E, F, L, or P

Xaa2 is selected from D, E, L, or P

Xaa2 is P

Xaa3 is selected from C, F, H, I, L, P, or Y

Xaa3 is selected from C, H, or P

Xaa3 is P

Xaa4 is selected from C, D, E, or P

Xaa4 is selected from C, D, or E

Xaa4 is C

Xaa5 is selected from D, E, G, P, or W

Xaa5 is selected from G, P, or W

Xaa5 is P

Xaa6 is selected from C, K, R, or V

Xaa6 is selected from K or R

Xaa6 is R

Xaa7 is selected from D, M, P, or V

Xaa7 is P

Xaa8 is selected from D, I, K, L, P, R, or V

Xaa8 is selected from K, P, or R

Xaa8 is P

Xaa9 is selected from C, H, I, K, L, M, or W

Xaa9 is selected from I, L, or M

Xaa9 is I

TABLE 28 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that were found in colon tissue and comport to one or more of the rules provided in TABLE 27. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 10118-SEQ ID NO: 11117, as disclosed in TABLE 28. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 28

Sequences of the 581 to 589 Region in AAV5 VP1

Capsid Polypeptide that Drive Colon

Tissue Tropism

SEQ

ID
581-589

NO
Sequence

10118
PENIAEDHE

10119
PYALAEGNP

10120
PLAGRFVAQ

10121
PAEAKQRFD

10122
PAAYDGRLL

10123
PACHCYESL

10124
PACRFEANS

10125
PACSPYGWW

10126
PADFVNSNY

10127
PAECWRMTF

10128
PAEFDYLFT

10129
PAHEIKNLE

10130
PAIDHLLHR

10131
PAMQGHMVD

10132
PAMRTVHMW

10133
PAQWAWADG

10134
PARCAVSEH

10135
PASLKEHCV

10136
PATCSQKKY

10137
PATPPTIVG

10138
PAYHTWQYN

10139
PCAPYHPNQ

10140
PCDCSYYFR

10141
PCGSDIGCM

10142
PCHMWMNFY

10143
PCIWNWETD

10144
PCKPYPKDI

10145
PCNNCGHIR

10146
PCPMSPGEW

10147
PCQLKSDGE

10148
PCRDTLEGI

10149
PCRPLPHLL

10150
PCSMPKAGG

10151
PCTVEGRQY

10152
PCYMKSWVM

10153
PDCQEFWKQ

10154
PDGFSCRFA

10155
PDHCLQSTK

10156
PDHRDEMEW

10157
PDHTNYHIV

10158
PDIIIEGNT

10159
PDINYSCAI

10160
PDLRTQLFS

10161
PDMCGRMDM

10162
PDMQTEAGA

10163
PDPRRTNWQ

10164
PDPVDHRGD

10165
PDPWSTSHK

10166
PDQQYELSC

10167
PDRNLSQGP

10168
PDRWCQLYK

10169
PDSHVPLRD

10170
PDTKYLHRC

10171
PDVQNRWEM

10172
PDVTTNNVI

10173
PDWLMWSHH

10174
PDWYHCGFQ

10175
PEAEFGQKS

10176
PEAEYEQMK

10177
PECHNYNQI

10178
PEDENPLYE

10179
PEDQHNTWC

10180
PEDWKRHAN

10181
PEEQTCEGR

10182
PEFPTWGHY

10183
PEGLWSVAC

10184
PEHKGYAGW

10185
PEISHKNVY

10186
PEKESTVIS

10187
PEKYCHEGY

10188
PELAMPWEM

10189
PELTYAKSI

10190
PEMCKMGSP

10191
PENWFWPHQ

10192
PEPSAMTIP

10193
PEQLLYSMK

10194
PEREDCHNP

10195
PESEGRMTK

10196
PESKLMWGA

10197
PETKTFCQQ

10198
PEWTTTTVF

10199
PFACGMSTW

10200
PFAEFVGRT

10201
PFAGGEYDG

10202
PFCIPWTEG

10203
PFCQPPEFN

10204
PFDRMDHLA

10205
PFGLSQCKG

10206
PFGQLKFNM

10207
PFHVPPPTH

10208
PFKCQDIDC

10209
PFMSTYQVE

10210
PFPVQNVDS

10211
PFSKWVYEW

10212
PFVSQWEHM

10213
PFWWVCKVI

10214
PGGYKEHTW

10215
PGIPIVWMH

10216
PGKQTLTIQ

10217
PGMMPGCTA

10218
PGPFRGPER

10219
PGVTCFSQT

10220
PHCLKAGPL

10221
PHDARVSRM

10222
PHEDNNHQT

10223
PHELCHISH

10224
PHENKFLDC

10225
PHHGANGFC

10226
PHHWCIVES

10227
PHKCMFSWN

10228
PHNYRVFWE

10229
PHPPNPGEN

10230
PHQHPQIPE

10231
PHRDVIWLG

10232
PHRVNVERS

10233
PHSNSVGKC

10234
PHSQKLNSG

10235
PHTDAGYIH

10236
PHTQALICP

10237
PIASEMYED

10238
PICEFCTMN

10239
PIEFYTIKH

10240
PIEGYHIAD

10241
PIESKPRSS

10242
PIEPPGQKP

10243
PIHHQLMCF

10244
PIIPCGGWQ

10245
PIISTFNNQ

10246
PILCTMPFV

10247
PILIPSLQP

10248
PILPNTHTG

10249
PIMCKEPGV

10250
PIMMHQTLG

10251
PIMPKYNAL

10252
PINTSHSYF

10253
PIPHVMNEP

10254
PIRQNAMMW

10255
PITLRQPWH

10256
PIVAWIWYD

10257
PIVHQNPMN

10258
PIVVDGRKF

10259
PlYSIVIKG

10260
PKCEILTGA

10261
PKGADRYNY

10262
PKGVAMGGT

10263
PKIEPNLCK

10264
PKIMPKSCS

10265
PKLEDDEHC

10266
PKLTCETWF

10267
PKSDLCEWV

10268
PKSELQKSF

10269
PKSIDELFE

10270
PKTWMHGVP

10271
PLCLKYEGC

10272
PLECNRGTD

10273
PLEMSNTPW

10274
PLGFAPEIT

10275
PLHQTSEYK

10276
PLLMTKTGS

10277
PLMMGPQRQ

10278
PLNHACYCC

10279
PLNWIIACR

10280
PLQTSKWQA

10281
PLVKLEEIH

10282
PMCICWASE

10283
PMFYDEPKH

10284
PMGCNGYVT

10285
PMGYSNLNC

10286
PMHPTCFVI

10287
PMIVPDVAY

10288
PMLKTEAMS

10289
PMLPEKFPD

10290
PMMISCNSV

10291
PMMWSTNVK

10292
PMNWAHSCA

10293
PMPNYYHPQ

10294
PMRPHRDAE

10295
PMSPFPFEH

10296
PMTSTEPME

10297
PMWYYYHRM

10298
PMYSQVHLG

10299
PMYVLEAHY

10300
PNAKPCCFE

10301
PNCLKSIHG

10302
PNCPFYPHP

10303
PNDLNRRTK

10304
PNEPMTPGQ

10305
PNESGHQSS

10306
PNFYSDISR

10307
PNIKYVYNG

10308
PNIPLIDMC

10309
PNMPTCNAP

10310
PNQYTANEI

10311
PNRHGPDAE

10312
PNSSTPYNW

10313
PNWQNLESD

10314
PPDAFRWCP

10315
PPDIQPNCS

10316
PPELEKCTP

10317
PPFCKWYIV

10318
PPILLIQMR

10319
PPKDKYLRE

10320
PPKPAEEGH

10321
PPKYPQQQI

10322
PPPSVRMMM

10323
PPQQCEWIA

10324
PPRHSNSQG

10325
PPTIGCSCN

10326
PQAEVMLGE

10327
PQCRMLIME

10328
PQDDKGVIP

10329
PQECSHVHP

10330
PQKLNFTYC

10331
PQKTLAVGN

10332
PQLVSHIGH

10333
PQNPKMIPA

10334
PQNTFVRKQ

10335
PQPCSYDSG

10336
PQRYIFGAW

10337
PQSCENNCR

10338
PQSHCHMET

10339
PQTFWGETC

10340
PQVMQLAEA

10341
PQYIQAIED

10342
PQYRCIDEH

10343
PQYTNRVIG

10344
PQYVPDRGT

10345
PRCLKCRPH

10346
PRGANSMMT

10347
PRISMREID

10348
PRLNVELDW

10349
PRLQWMEPG

10350
PRMCMSENE

10351
PRMWDKLDQ

10352
PRRDKDVYM

10353
PRVCVTWVT

10354
PRWMQTYQV

10355
PSACMVGPC

10356
PSADCNPWA

10357
PSAGDLSQY

10358
PSAQGNFSM

10359
PSCVIFKPW

10360
PSDHTWKNV

10361
PSEGDSWLC

10362
PSESLELCP

10363
PSFLCGKYI

10364
PSGCYKCMK

10365
PSKHKDLNE

10366
PSLYCCDGV

10367
PSMYEDMTN

10368
PSNPEQHLV

10369
PSPRYPVCM

10370
PSVLQIITN

10371
PSVTVDVGS

10372
PTCLGKIKW

10373
PTCQTPPLH

10374
PTEFDQEQT

10375
PTEPREDAA

10376
PTETRWHTG

10377
PTFMFRRDL

10378
PTGIYAMQC

10379
PTHLFNFHW

10380
PTINRYFNE

10381
PTKEMMSMF

10382
PTKSSGGAA

10383
PTMLAVKNA

10384
PTMPWKKAE

10385
PTMYRPMHV

10386
PTNGIASAH

10387
PTPSIVLCQ

10388
PTQYIGDHR

10389
PTRIQMTVD

10390
PTVWANVAE

10391
PTVWRTFEF

10392
PTWYVLAHF

10393
PTYPFISYS

10394
PVAGDRYGG

10395
PVAVPTESP

10396
PVDRLVHTN

10397
PVEMPDSCH

10398
PVGSHMTFW

10399
MVIHGMHID

10400
PVIKPNTEV

10401
PVIQQRRDQ

10402
PVIRKIYEL

10403
PVKCEDHPE

10404
PVKSPWFNP

10405
PVLCEMKQY

10406
PVNWPPFFC

10407
PVPRRKCSS

10408
PVPTFCSTC

10409
PVPYETHMY

10410
PVRKAGWQK

10411
PVSPPCDAQ

10412
PVVMIGHMS

10413
PWDLRNQYH

10414
PWEAMYMTA

10415
PWERRHAER

10416
PWETCRWKQ

10417
PWIAMVAQE

10418
PWNQKCNYP

10419
PWNWDGHMR

10420
PWQCLRSPD

10421
PWVRWRTCQ

10422
PYAANHQQG

10423
PYACKIIRG

10424
PYCNIWMES

10425
PYCPMQIDC

10426
PYCWMVDDA

10427
PYEHGKEHH

10428
PYGYNCVID

10429
PYHRIMEYA

10430
PYIRCKKDG

10431
PYMRCEHQH

10432
PYMVTCYDF

10433
PYNPQYCCC

10434
PYPWDRCWI

10435
PYRKPWNAE

10436
PYRVEGEHN

10437
PYSFKDLLE

10438
PYTIQKGEY

10439
PYTNEQQYT

10440
PYVQAQAGR

10441
PYVRRVTEI

10442
MPTRQRHEK

10443
APILLQASC

10444
DPRRSMWYL

10445
IPRIVYMCD

10446
SPCNMDTHH

10447
RPVTYPHTA

10448
NPGWILSMA

10449
IPSHVHFGE

10450
APDNEAGMY

10451
APDPSTANR

10452
APFNANCCN

10453
APGCYFYRN

10454
APICEGEPK

10455
APITIGAMS

10456
APKWFNSLD

10457
APLHNFPVT

10458
APMDMGRMT

10459
APMHSFDAE

10460
APNDCMMQN

10461
APPHQDHNP

10462
APQCVIVFT

10463
APRQJWMHH

10464
APVYWDHQM

10465
APWWEFPQY

10466
APYDPKYVF

10467
APYQAVMES

10468
CPALNQLGR

10469
CPAMPGYCH

10470
CPDDSRLDQ

10471
CPDLPDWSG

10472
CPFFNYMIQ

10473
CPGYYETEI

10474
CPHAHYVWR

10475
CPIYYGPVS

10476
CPKGHEYEN

10477
CPKQWLPAC

10478
CPLIDWCMY

10479
CPLPARWGD

10480
CPNCWYDEV

10481
CPPQSENKV

10482
CPPRRYVNG

10483
CPQCMQGAS

10484
CPQMPWLHE

10485
CPRQLYHWQ

10486
CPSCEISVP

10487
CPSECDNLK

10488
CPSKQTRNP

10489
CPTQLYLHN

10490
CPVCEDMPL

10491
CPVLFTHME

10492
CPWLELCYH

10493
CPYCDQEGH

10494
DPAGDLMQG

10495
DPCNVDPGF

10496
DPCTCHTAK

10497
DPDPVDMYA

10498
DPGAPGPKA

10499
DPHCIPTWK

10500
DPHCVKNNC

10501
DPICVVWTG

10502
DPIHAHVEC

10503
DPKHYNETM

10504
DPKICWQER

10505
DPKSCPRRE

10506
DPKSGFNGT

10507
DPMCFHAYQ

10508
DPMQMHRSG

10509
DPPKCYRYE

10510
DPPNTSFFH

10511
DPPRCDMQK

10512
DPPSPVMCG

10513
DPQLFCSDL

10514
DPRMGGSVN

10515
DPSYNERHR

10516
EPCHWMHEF

10517
EPDQMQERL

10518
EPDQPRWDS

10519
EPECTEFKV

10520
EPFPHVMWG

10521
EPGIVHWYD

10522
EPGYYDGGN

10523
EPHKYNQPS

10524
EPHLQPHQK

10525
EPHLTAVPT

10526
EPKNYSERH

10527
EPLKYNHES

10528
EPLQFMELM

10529
EMIRLGEWS

10530
EPPVCQGNE

10531
EPRRVDDQQ

10532
EPVMVRMQF

10533
FPALIKPHE

10534
FPASPDILI

10535
FPCGIVWTH

10536
FPCQNGPHN

10537
FPDRMDYVE

10538
FPGGQKPCL

10539
FPHFTCWCD

10540
EPHGYAMIW

10541
FPIQGNHFP

10542
FPKAPKYYL

10543
FPKNNLTKC

10544
FPLCVDPGP

10545
FPNIDNPKA

10546
FPPPIHLAI

10547
FPPQQEFIH

10548
FPQWGKEPI

10549
FPRTTNTRV

10550
FPSHDDFLG

10551
FPVTCGTVF

10552
FPWYSLILT

10553
FPYMAEQYR

10554
FPYWDIDDT

10555
GPDPWSQWW

10556
GPDPWSQWW

10557
GPDTEYRGC

10558
GPFADPPAT

10559
GPFLSVPAH

10560
GPFRFEWEW

10561
GPGTLRHLL

10562
GPHWYWHDM

10563
GPINPYAHG

10564
GPLSKHFDV

10565
GPMHNTGPP

10566
GPPPSDLRS

10567
GPPWKHCDA

10568
GPSCYLDYW

10569
GPTIWIHTY

10570
GPTIYQYMQ

10571
GPTKLNYNN

10572
GPVVPCNSW

10573
GPVWEFEWM

10574
GPWHACKIN

10575
HPCCDRYLA

10576
HPDAPGPIQ

10577
HPDVRNYEN

10578
HPEVDSSGA

10579
HPGCEYSWE

10580
HPGIEVDRG

10581
HPGWTHAVF

10582
HPHDVFHET

10583
HPILPAQYF

10584
HPISPWQWL

10585
HPLSRGSES

10586
HPMFEAYQN

10587
HPNFIWCNH

10588
HPNGNEREH

10589
HPNPWGVFW

10590
HPRFNRLQV

10591
HPSFIKCQV

10592
HPYLCTIRI

10593
IPDAWWANM

10594
IPHRNIHPS

10595
IPLWETMNM

10596
IPNDRDRCA

10597
IPRVHWKAE

10598
IPTFRTMKI

10599
IPWFDFAME

10600
KPAPWMRME

10601
KPCPPHAYV

10602
KPEAWAVCG

10603
KPESKMLSE

10604
KPFVSECTE

10605
KPGFRYFAE

10606
KPGTVKDCD

10607
KPILMSEEA

10608
KPITEGRRD

10609
KPKCCQYNW

10610
KPMCDGVAY

10611
KPMCLYVLT

10612
KPNDCLREG

10613
KPPMPTNTG

10614
KPPYSCREQ

10615
KPQHENKQL

10616
KPSMYPRNS

10617
KPTDMVGAW

10618
KPTQIGGEH

10619
KPVEPTTNP

10620
LPAAYFWGA

10621
LPAWYRARC

10622
LPIEQQKSW

10623
LPIESEGEI

10624
LPLGQADEA

10625
LPLVYAKNW

10626
LPLYFHEHY

10627
LPMDEKEGA

10628
LPMQCDETY

10629
LPMVWGNPK

10630
LPMYIEHMC

10631
LPNPVNATQ

10632
LPNWFCRNY

10633
LPPAHMFYC

10634
LPQHYGIWH

10635
LPQPYPDNY

10636
LPQRGTPIV

10637
LPRHIYKST

10638
LPSHELWFW

10639
LPTSDCRAE

10640
LPTYNKYVQ

10641
MPAHANSTC

10642
MPCTCSYPH

10643
MPDNFGNGP

10644
MPIMMNVSP

10645
MPMDLNYIQ

10646
MPMFCQIEQ

10647
MPPFHENES

10648
MPPPVQDWN

10649
MPPVRLYSY

10650
MPQDCGWNL

10651
MPRDGTWWC

10652
MPSHGYWYQ

10653
MPSPYGEAY

10654
MPSQGVIWH

10655
MPSVWGPDM

10656
MPTEYEAHG

10657
MPTQAYMPQ

10658
MPVDIANQM

10659
MPWWYGCIC

10660
NPCFMIWYT

10661
NPCMLQDHH

10662
NPCTFWNNE

10663
NPGHAWATM

10664
NPGWWCRMW

10665
NPIMYPCLS

10666
NPLIRDMWT

10667
NPLKLDYPE

10668
NPPWLQPFS

10669
NPQFLHFQI

10670
NPRAMDAAH

10671
NPRHDAGHS

10672
NPSWNACNV

10673
NPTMPGIRG

10674
NPTQDFKVM

10675
NPVTGEYSL

10676
NPWHYTYAF

10677
NPYLHMFNQ

10678
NPYNQHC1C

10679
NPYRTFTDG

10680
QPAFEHAQV

10681
QPAHMEVMY

10682
QPAHPGTGL

10683
QPAQYAVMH

10684
QPCDITYQS

10685
QPDCEWPNN

10686
QPDMYDCYA

10687
QPEFTYLQF

10688
OPHSSVILD

10689
QPPAAAKQD

10690
QPTDDVIQG

10691
QPTQRMYHC

10692
QPVTFYEKH

10693
QPVWFPKCW

10694
QPWMVWNSH

10695
QPWMYVMGS

10696
QPWITWWNR

10697
QPYCERTGI

10698
RPAVDDEEC

10699
RPCPNMTDG

10700
RPDVRNELC

10701
RPELGGQQT

10702
RPETTAERP

10703
RPHHFWVPF

10704
RPKECEQMW

10705
RPLDTSQED

10706
RPMFGAVAG

10707
RPMHGDYDD

10708
RPNCVMYDK

10709
RPPDEIWGH

10710
RPRDNGVEG

10711
RPWSSEFRV

10712
SPAEIPSIF

10713
SPCPNGVRQ

10714
SPDVGWTES

10715
SPEVTEYGF

10716
SPFMWWRWW

10717
SPGFILGGY

10718
SPGKFWHPC

10719
SPHFQKHPS

10720
SPIRIDWQG

10721
SPISDLRNC

10722
SPIWEGERT

10723
SPKKAEQAI

10724
SPLHVHWQS

10725
SPLISVLEG

10726
SPLQNYEER

10727
SPMDTTEPK

10728
SPMQKQKVS

10729
SPQTCSEPT

10730
SPRCMTNGG

10731
SPSGIMEGN

10732
SPYQCPQAS

10733
TPDHKTRAH

10734
TPDPDLNNR

10735
TPGSDMSMV

10736
TPHSSMKVG

10737
TPKLEFPVD

10738
TPLLTSGHE

10739
TPLMAYWIP

10740
TPMDYEDQE

10741
TPMFDRHSL

10742
TPMLGMLWW

10743
TPMWQASWQ

10744
TPNCMGQRM

10745
TPNVWEQQM

10746
TPPHKCSVG

10747
TPQCAMWNE

10748
TPSQVASDD

10749
TPSRNMVDL

10750
TPYIRHHFE

10751
TPYTWCWPD

10752
VPCPKWSMI

10753
VPEQGRWEA

10754
VPKLIDTDQ

10755
VPLLILNNN

10756
VPLVSYSQQ

10757
VPMMCQWSQ

10758
VPQHKNEFQ

10759
VPQWIDERW

10760
VPRHWAGVR

10761
VPRYNSDEQ

10762
VPSCQCSES

10763
VPTIKVIRA

10764
VPTQSVKGC

10765
VPTSYWCTN

10766
VPWFVMLMM

10767
VPWMVLDNN

10768
WPALFVADA

10769
WPDPFIHAT

10770
WPEATHRFA

10771
WPKYDMPLH

10772
WPMCPVCYF

10773
WPMTRTNGH

10774
WPNYHSWYE

10775
WPPNGKWFM

10776
WPQRGTMDM

10777
WPTGHCAYW

10778
WPYYCCIEH

10779
YPANKCEWE

10780
YPASYDCHP

10781
YPDTVQKSP

10782
YPDWWEMIT

10783
YPEGEMNYE

10784
YPFQPIEEP

10785
YPFWAGWGD

10786
YPGMKICGH

10787
YPIIMLPCR

10788
YPMQGGIRW

10789
YPNDAKHGN

10790
YPNLHHSMM

10791
YPSDGLFEQ

10792
YPSGHKPYW

10793
YPS1NCPTN

10794
YPSPPERLE

10795
YPVNCTQKW

10796
YPWENNAHL

10797
YPWMTACHD

10798
YPYICEFVW

10799
YPYPRLWEC

10800
NEPYCIWAY

10801
GRPVMHRHS

10802
DSPSNHPAV

10803
YYPFPGPTD

10804
AAPIPGKAF

10805
AFPNVMYQN

10806
AGPKMIEPW

10807
AHPQQPHVT

10808
AIPMIGSYC

10809
AIPQEGVCC

10810
AKPYKFSGE

10811
AMPIVMWHT

10812
ANPADWHWY

10813
ANPFVLAPW

10814
ARPWWMLEQ

10815
ASPPDYKAQ

10816
AVPQMTGWC

10817
AYPHCEAVM

10818
AYPSGIRAH

10819
CCPVDNEFW

10820
CDPPIDCYI

10821
CDPSYELNI

10822
CDPWHKYNG

10823
CGPFRHQNG

10824
CHPQEWHGQ

10825
CIPDFISEY

10826
CLPWVMSIA

10827
CMPPDCKCP

10828
CNPGWPKPG

10829
CTPYPPRDQ

10830
CVPNSHIWQ

10831
CYPTLEKHA

10832
DFPLNPVQT

10833
DFPLQVTGY

10834
DFPRNNAIM

10835
DGPGTQCIK

10836
DLPAGQRMF

10837
DLPLPKLMQ

10838
DNPGYLKEE

10839
DSPNYVWTH

10840
DTPEYAPSM

10841
DTPLQYKAQ

10842
DYPKGEKCD

10843
EEPELQRLN

10844
EHPPHAFAH

10845
EKPLYYHHA

10846
ELPATLFQF

10847
ENPQRIRSI

10848
ESPCHQNQG

10849
EYPSYWSDG

10850
EYPYDVPNF

10851
FAPEICGCC

10852
FDPVCKCMK

10853
FEPQNPTFQ

10854
FFPTMHARF

10855
FIPCVISNQ

10856
FLPEWKAFH

10857
FLPTSKLHS

10858
FLPTYDGMH

10859
FNPPIRQLW

10860
FQPFTSNLL

10861
FQPYMTSTS

10862
FRPIISMKR

10863
FRPMITCWW

10864
FSPPMVMKY

10865
FSPYFMVPH

10866
FTPDSLIVH

10867
FTPECTRGM

10868
FVPHDCNRD

10869
GAPFDELPH

10870
GCPLIVPDL

10871
GDPLVIWLC

10872
GFPREMYTE

10873
GGPEKEAHR

10874
GGPPIDEGN

10875
GHPFKDYHS

10876
GMPWEDISN

10877
GNPCSPKIN

10878
GNPKLLESF

10879
GNPPTWLCP

10880
GNPYMYPWH

10881
GRPWDPYFE

10882
GTPQWWNKG

10883
GTPSMHMGI

10884
GVPQDRIQE

10885
HCPNQGRRS

10886
HFPRSEAHR

10887
HGPMEVMHE

10888
HHPFAKPII

10889
HIPQHSRMC

10890
HKPEQMEAY

10891
HKPKMWMYG

10892
HNPLIRTMV

10893
HNPLQENDC

10894
HQPELMTDN

10895
HRPDWNDGK

10896
HRPPCEMGD

10897
HYPAYQKAC

10898
HYPWTPWTP

10899
ICPTKGMEA

10900
ICPVCSWDC

10901
IDPEIYAWC

10902
IDPLDGNHF

10903
IHPOQWWDH

10904
IIPHQNIMW

10905
IKPEPMWMT

10906
ILPPEKHQA

10907
IMPNNLEMM

10908
INPRKLSSK

10909
IQPMYVIYQ

10910
IQPWDYVRP

10911
ISPLNICWF

10912
ITPEVLKTT

10913
KCPLGKPAS

10914
KCPRGGMNC

10915
KDPCCGCHN

10916
KGPPHKDDQ

10917
KHPNIWCAN

10918
KLPDTYKNF

10919
KMPPPAKDI

10920
KTPHCIKHY

10921
KVPVLTAGG

10922
KWPTFKWPK

10923
KYPEPGRGT

10924
LAPQLGWRN

10925
LCPGEWLVI

10926
LEPKDLAAP

10927
LEPPSTVNV

10928
LFPNQCNFV

10929
LFPQHIAQP

10930
LGPHRWYGH

10931
LHPWMPAGR

10932
LKPSWQDKA

10933
LLPMDEMKT

10934
LMPCHGDIA

10935
LMPELGCTD

10936
LSPLIAGIY

10937
LTPDVRINA

10938
LTPSCRHKW

10939
LVPTLINCL

10940
LWPSSCQMK

10941
MAPQPMGGG

10942
MCPKTFRFE

10943
MEPKKMKVW

10944
MEPMSRDHL

10945
MEPWEQCHI

10946
MGPRYDKEW

10947
MHPDSYNVT

10948
MKPDQNHHK

10949
MLPLPNEFQ

10950
MLPMAWYQI

10951
MLPMRQLTQ

10952
MLPWSSRSN

10953
MMPDILRSY

10954
MQPCWQPQC

10955
MRPQQVWNG

10956
MSPQPELSG

10957
MVPAAGPGE

10958
MVPKMMRSQ

10959
MWPAMKVCW

10960
NCPQPNCGW

10961
NMPERKLTF

10962
NMPIGGYCC

10963
NNPQKQQSR

10964
NNPSNSLIW

10965
NTPEFHWQQ

10966
NTPLDFTGE

10967
NTPNNMEAR

10968
NTPQQGQKG

10969
NVPTEPMDH

10970
NWPVSDEDM

10971
NYPCTDKNH

10972
NYPVGYHEL

10973
QAPMVQDTQ

10974
QAPWKHEGP

10975
QCPDEKAEF

10976
QCPIQSVVC

10977
QDPFWRGLH

10978
QHPDQGQHC

10979
QIPYEMWWS

10980
QLPCIKKCP

10981
QLPGCRTDQ

10982
QLPKMWLPN

10983
QMPNGEIVT

10984
QNPDDVINL

10985
QNPSPMMHR

10986
QNPSTYCHY

10987
QQPWPHQRG

10988
QRPTMLQGY

10989
QSPPIEPTI

10990
QSPWKLWSH

10991
QWPDFMVTI

10992
QYPQLPHWA

10993
REPFLHVNS

10994
RFPVKLGAM

10995
RHPQLGLPH

10996
R1PWAELMK

10997
RKPWREHNI

10998
RLPYYVQFC

10999
RNPCSKHEV

11000
RNPRDYPMF

11001
ROPANREKE

11002
RTPFPHDSL

11003
RWPSEKEKF

11004
SAPPSFAQK

11005
SAPVEWMRK

11006
SCPNRESGR

11007
SGPRTDCPL

11008
SGPVVMIQS

11009
SMPHACYNG

11010
SMPTTSSSC

11011
SSPDNWKHM

11012
STPLGNSAP

11013
STPMVCEMG

11014
SVPHEVHCE

11015
SVPLKFWTL

11016
SWPDEPSVA

11017
SWPHCEIEI

11018
SYPLTRWPA

11019
SYPYMHMQD

11020
TAPQCVSDA

11021
TCPNLTAYC

11022
TCPRNLCNY

11023
TDPLCKHQT

11024
TDPLNPLCD

11025
TEPPCPGQT

11026
TEPSIMNEV

11027
TFPCNREGI

11028
TFPSIQWSR

11029
TGPNFQPAL

11030
TIPECKKCE

11031
TIPYEANCA

11032
TKPQITEIS

11033
TQPCMPNRT

11034
TRPPSSLDE

11035
TIPTWQNSP

11036
TVPHFYFYG

11037
TWPMSCDCD

11038
TYPFGWDKH

11039
VAPAILYRC

11040
VAPIHDHRI

11041
VDPAMMPDN

11042
VEPTASQLC

11043
VHPEQTLIG

11044
VHPFHKYEQ

11045
VIPVHNMCG

11046
VKPYNGLVQ

11047
VLPHEWNQG

11048
VLPKSHFFP

11049
VMPEESVRN

11050
VMPYAWGIS

11051
VMPYAWGIS

11052
VQPCFMMDH

11053
VQPFPYCCE

11054
VQPHPFNPH

11055
VQPPILEQH

11056
VQPWWDAMG

11057
VRPLHNFID

11058
VSPANADND

11059
VTPNLQVQS

11060
VTPYDWKFQ

11061
WAPSPRQDR

11062
WDPVYGERN

11063
WEPDGIVPQ

11064
WHPKCYDVF

11065
WHPMDKNER

11066
WIPTEIYal

11067
WKPEGQNFG

11068
WLPRMPSCD

11069
WMPCCPHGC

11070
WMPGMHWRC

11071
WMPSTTRWN

11072
WQPDWMYEH

11073
WRPPVRVQD

11074
WSPERERFE

11075
WWPIWWIPQ

11076
WYPLDVLAS

11077
WYPMLWELE

11078
LS1CQPVMP

11079
TTRCFRHAK

11080
WATCDAPPV

11081
NYVCSTSGY

11082
LTKCMNKGW

11083
SCECPCCLA

11084
VCECCRQNI

11085
ALCCFCCQG

11086
TSYCIQTVD

11087
DQECCQTLF

11088
QTQVPIWNT

11089
LATHPPYTY

11090
LTYNPTMIG

11091
TGIIPIRVC

11092
TCQEPHNIT

11093
QVYAPEIEG

11094
DYQSERLWS

11095
KEQEQRPVR

11096
ISMQNRCAD

11097
IFQLSRPAW

11098
ECKTHRDVY

11099
YHNQDRHYL

11100
KVDTVRNPK

11101
TVSRGRPVA

11102
NQAYGRGHV

11103
SVRLNEPHV

11104
CLVNDMPRK

11105
AHTFEAPKW

11106
IWNYCVPKH

11107
HWEQHNPIW

11108
CIDWAEMPQ

11109
ACVMYDCPP

11110
WVGNANRPA

11111
SVEVGCSPG

11112
CVFSTNHPA

11113
AMRQIIAPC

11114
TVVFSENEI

11115
WKGRWTTTI

11116
NSHYTEGVI

11117
AWTPTEKQI

Example 15
AAV5 Variants with Tissue Tropism in Thyroid Gland

This example describes engineered AAV5 variants with tissue tropism in thyroid gland that were discovered using the methods and systems described in EXAMPLE 1-EXAMPLE 2.

CNS, liver, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord tissues were analyzed to identify amino acid residues in the AAV5 VP1 581-589 region that drive thyroid gland tropism. The results are shown in FIG. 18L which shows the positional frequency of each amino acid in the 581 to 589 region of AAV5 VP1 in thyroid gland tissue as compared to all other analyzed tissues, wherein the capsid polypeptide has the sequence of SEQ ID NO: 2. The position (“Xaan”, n=1-9) is denoted on the x-axis and amino acid residues are denoted on the y-axis. The frequency of a particular amino acid residue in a particular position is indicated in the heatmap as intensity. Darker squares indicate a higher frequency of a particular amino acid in a particular position and lighter squares indicate a lower frequency of a particular amino acid in a particular position.

The frequency of a given amino acid residue occurring at a specified position corresponding to position 581 to position 589 of SEQ ID NO: 1 (generalized in SEQ ID NO: 2) in variants identified in thyroid gland over the frequency of that given amino acid residue occurring at the specified position in variants identified in all other harvested tissues was analyzed to identify a set of sequence rules for capsids that preferentially target thyroid gland tissue. With reference to TABLE 29 below, and SEQ ID NO: 2 (AAV5 VP1), the following amino acids can, thus, be independently mutated, in any combination, at any one or more positions Xaa1-Xaa9, to provide a VP1 capsid with enhanced thyroid gland tropism as compared wildtype AAV5 VP1 capsid (SEQ ID NO: 1), where thyroid gland tropism here refers to properties that are preferred for thyroid gland transduction over properties that are preferred for transduction of all other harvested tissues.

TABLE 29

Thyroid Gland Tropism Rules

Xaa1 is selected from A, K, M, N, Q, or R

Xaa1 is selected from K, N or Q

Xaa1 is K

Xaa2 is selected from A, F, K, L, M, T, V, or W

Xaa2 is selected from F, V, or W

Xaa2 is W

Xaa3 is selected from A, I, K, R, S, T, V, or W

Xaa3 is selected from A, R or T

Xaa3 is R

Xaa4 is selected from A, D, E, I, P, or V

Xaa4 is selected from A, E, or I

Xaa4 is A

Xaa5 is selected from F, I, M, Q, V, or Y

Xaa5 is M, V, Y

Xaa5 is M

Xaa6 is selected from H, M, N, or Y

Xaa6 is N

Xaa7 is selected from H, I, N, Q, S, or W

Xaa7 is selected from H, I, or N

Xaa7 is H

Xaa8 is selected from A, D, F, Q, S, or Y

Xaa8 is selected from A, F, or S

Xaa8 is F

Xaa9 is selected from A, Q, S, or Y

Xaa9 is selected from A or S

Xaa9 is A

TABLE 30 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that were found in thyroid gland tissue and comport to one or more of the rules provided in TABLE 29. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 40438-SEQ ID NO: 41437, as disclosed in TABLE 30. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 30

Sequences of the 581 to 589 Region in AAV5 VP1

Capsid Polypeptide that Drive Thyroid

Gland Tissue Tropism

SEQ

ID
581-589

NO
Sequence

40438
KAEDSQPDC

40439
KAMQVLENC

40440
KASWIPSNK

40441
KCEDNTFAS

40442
KCTDLGEYF

40443
KDHEEHGPK

40444
KERTAWSRE

40445
KFMQDPCNS

40446
KGELMCLDV

40447
KGPKECAAT

40448
KHILWGSLA

40449
KIGISEQDF

40450
KKYTDQDYG

40451
KMTPVEADA

40452
KNPTVFWKM

40453
KQAIYLDLD

40454
KSAMQEGVA

40455
KSFGSEVFF

40456
KSKWGHYWD

40457
KSMDLGQFM

40458
KSNQTYCAA

40459
KSYCLHMKQ

40460
KTGMACDIN

40461
KTLVKQMDY

40462
KTVWVYRVH

40463
KTYPHCFPV

40464
KVEDGLQWH

40465
KVEGMVTQS

40466
KVIEFCGEE

40467
KWLEANMTG

40468
KYAFRYEAS

40469
KCLQMCPNG

40470
KAIGWFHFD

40471
KAYTVEMDP

40472
KCCLPWQNI

40473
KCEQWQKVC

40474
KCHVGGNVY

40475
KCNPWSEPT

40476
KDDGDDNIP

40477
KDRDQVHWM

40478
KEGNMDACS

40479
KEYDPQHWW

40480
KFIYGQREV

40481
KFKQEDAQM

40482
KGIWQDHSC

40483
KHCENCEYF

40484
KIAPVAASP

40485
KIEMLCCQG

40486
KILNPNDSY

40487
KKRRDTQIY

40488
KLHIWNVPC

40489
KMQPNGEWP

40490
KNDQTTYPE

40491
KNEPDNLCE

40492
KPCTFAPFM

40493
KPEWAMMYD

40494
KPYQDIARM

40495
KQEIQGVCR

40496
KQETRTTDH

40497
KQFWMKVEV

40498
KQMLQQCFG

40499
KQWELYQID

40500
KRVVKFTST

40501
KRYTHGKNW

40502
KSADMPTTL

40503
KSFLHPAAH

40504
KSFYMCRCG

40505
KSYVGSYES

40506
KTCELTAMR

40507
KTEMSDMHW

40508
KTLVCVNWF

40509
KTYLWEHES

40510
KVSEAHYDL

40511
KVTTETGVP

40512
KVVAVNQSL

40513
KWQQAEYDE

40514
KYCHWWYKL

40515
KYQNAKPVA

40516
KKNSMDCQV

40517
KPYFKDCPR

40518
KSIFQATLQ

40519
KTAHDKDID

40520
KHSSWCCYT

40521
KSEHHWINA

40522
KVGMDQTAV

40523
KFFYTELMH

40524
KDFNFEAQG

40525
KDQQAERHV

40526
KDYANFGMN

40527
KHIHCTIQC

40528
KNPCLVWQK

40529
KSYEMVNQW

40530
KTGWDSHWS

40531
KVAYMWYNW

40532
KHGVLPMPA

40533
KTLHMLESK

40534
KCGLTPGVE

40535
KVDTVRNPK

40536
KAHDMAINL

40537
KAEMKMGCN

40538
KTTIVPAWN

40539
KAIPQGSSC

40540
KMTNSNENS

40541
KWYQFGMVA

40542
KFDVIRQAM

40543
KQLTFPMLA

40544
KQEWHVHTE

40545
KIDQTQWQR

40546
KRFVDMPEM

40547
KLYGLNHSC

40548
KIQPMMVKA

40549
KMLHKVWID

40550
KGPIFHSAY

40551
KGMQMWNCC

40552
IWKFKNAHL

40553
IWNDQWDCP

40554
IWNWMTEDC

40555
IWTLNWAQL

40556
LWEQWSTLS

40557
LWQFCSICE

40558
LWVECHCNQ

40559
MWVELPSDD

40560
NWIHWNACF

40561
NWMERDHNH

40562
NWSYCNVSA

40563
NWIHQDMFD

40564
NWYFMSQTD

40565
QWAKWDWSS

40566
QWHQFNRMH

40567
QWTTITKEE

40568
RWKSQEEIQ

40569
SWAFANHTH

40570
SWEQNVVST

40571
TWTAMEIYG

40572
SWSPTGVAN

40573
AWDHTMPYG

40574
AWGNRYHYC

40575
AWHDYFRDN

40576
AWMDCSIMC

40577
CWQAALHSA

40578
CWWCALLST

40579
DWIMDTPKP

40580
DWIVDTPKP

40581
DWNYYACYT

40582
EWICVQSHN

40583
GWMIAMDGK

40584
GWNTFIWGQ

40585
GWTLNPSEG

40586
HWIPQWARS

40587
HWISFERHQ

40588
IWDEKCTQC

40589
IWKDYLHVG

40590
LWCNENAMD

40591
LWQQIPWGK

40592
MWAAMPVGH

40593
MWEQEPNRN

40594
MWFAENNGL

40595
MWGINGVAY

40596
MWRPARYKS

40597
NWEDSPWCY

40598
NWEPCWWAH

40599
NWGPQQSER

40600
NWNKTRFTC

40601
NWVSHDKEA

40602
PWAETEKPN

40603
PWIPNQIKQ

40604
PWTCNENQW

40605
QWPEMTCCD

40606
QWTRTGSSF

40607
RWHNGENCS

40608
RWNARWELG

40609
RWRCRMQGN

40610
SWIDMANNQ

40611
SWTALGSVP

40612
SWTLRMWAS

40613
TWLPVGAAH

40614
TWVNFPTHC

40615
VWWFAPMEA

40616
WWAQPWHAL

40617
WWTDMSSWT

40618
YWWLDHANF

40619
GWIINLHYH

40620
NWVVWPMEN

40621
RWVCANAQD

40622
CWECYQFKR

40623
CWSAIMTGC

40624
DWMQGGIYN

40625
EWFEDEVLW

40626
EWGMHTSNR

40627
FWSGYNAEK

40628
GWTHFHWQE

40629
LWKTILEGV

40630
RWGMYDSYY

40631
RWTDTNFRG

40632
TWAMMGQWW

40633
LWKLGNKVF

40634
TWTSRCELL

40635
VWNPDLKMN

40636
EWIKKIADH

40637
IWNYCVPKH

40638
YWAFPCCGF

40639
CWGPIMHPF

40640
TWKSPRGYM

40641
AWMCCARHD

40642
CWWNFGKDC

40643
SWCAMIYEM

40644
IVRFYFKSW

40645
LIRQGDCFI

40646
LTRDRFTEW

40647
LYRTSIALV

40648
MERYGHCHP

40649
MLRAESGLT

40650
NNRTYDWWN

40651
PVRMFTESG

40652
QGRQILKAI

40653
QHRYYTCQS

40654
QLRKHYTLL

40655
QYRPTWVHV

40656
RFRWVHFHW

40657
SGRCNALNT

40658
TIRDSVFGC

40659
VAREDEWQN

40660
VCRTQESSA

40661
VHRHRAHKF

40662
VPRVWERAH

40663
VVRSYNFGE

40664
NDRHVPCWD

40665
AFRNKEFYV

40666
AHRDYACTN

40667
CMRGDKRNR

40668
CMRGDKRNR

40669
CNRPKCPPM

40670
DARFGLNFG

40671
DARFYITEG

40672
DHRETDAEE

40673
DYRNYFIWP

40674
EARCTDMCY

40675
EFRPVLECC

40676
EIRYYYPES

40677
ELRWDARPH

40678
EMRITNARC

40679
ERRMEQAQT

40680
ESRAGYMHY

40681
EYRCVYRKC

40682
EYRLCPQTS

40683
FHREQAAHQ

40684
HSRQSEETI

40685
IAREPLMAN

40686
IDRKDRRES

40687
IIRNRCAEP

40688
IYRLMHWDN

40689
LMRSTVKNG

40690
LPRCVIAAC

40691
LTREDFTQM

40692
MLRAMDIDA

40693
MVRDKNNDH

40694
NRRMVVAQS

40695
NTREYPEYI

40696
PMRMAIEFD

40697
PSRPCTYHS

40698
QGRWMYHWW

40699
QLREIQMIQ

40700
SNRCLTYEF

40701
SQRDEMFAL

40702
SQRHIECIG

40703
STRDWMSCM

40704
TDRMLAGQR

40705
TFRMSVTGV

40706
TIRSFNLEA

40707
TMRTHSSWW

40708
TYRVFMVHD

40709
VFRMVMEQY

40710
VYRLREKQE

40711
IGRHQMTDK

40712
ANRADWQHL

40713
QCRHTDQVK

40714
ETRGFVKCH

40715
ETRVREDAV

40716
QDRRPENRF

40717
CERSKCPKS

40718
DIRSYKGFQ

40719
QVRDIANMC

40720
VKRLENEWH

40721
YGRLREAMH

40722
DDRVKMFHY

40723
ANRMIWYPY

40724
GHRPKLCDQ

40725
VNRGDLSLF

40726
QSRCDPVSP

40727
EHRDNTTRP

40728
GVRNALNLS

40729
GIRCISKAG

40730
LTRSGPIGQ

40731
MTRVSMTWR

40732
CCRQIWVQY

40733
STRPIACHS

40734
NNRALMSVE

40735
TVRHQMDAY

40736
NNRRQFGYV

40737
SSRMQMQGP

40738
MFEAIVTGL

40739
MHGAFVDTQ

40740
MNVAYHGFS

40741
MTQADPCIG

40742
NAYAAGNIG

40743
NHSANPHGY

40744
NIMAIHHDS

40745
NMVAMNEPR

40746
NVWAWKEVE

40747
QLIAIHDIN

40748
QTVAHENCD

40749
QTVAPGGHC

40750
RADATGVPN

40751
REDAYLAGH

40752
RSEAGIRQI

40753
SGLAFDEDD

40754
SHVAYKLLD

40755
TMVAFSSCL

40756
TSYAMWSDV

40757
VASAMRHHT

40758
VTEAFPTSV

40759
VVDAIAQAD

40760
WGLANNCHE

40761
GIWAYGCTS

40762
AACAIWKGE

40763
CGDAYDEHC

40764
DDSAQYWQV

40765
DHTACKKLC

40766
DSKADHCVN

40767
DSMADDQHL

40768
EASAFPHDQ

40769
ECHAIDQSD

40770
ECKARNSCG

40771
ECTAHGVHC

40772
EFDARVAHE

40773
EFFALDSQS

40774
EHGAVHNAN

40775
EVAASSAIH

40776
EVIAGHVGA

40777
FPSAPTEPG

40778
GKEAAYAVW

40779
GMWAQWCSN

40780
GVEAYKFPH

40781
HEQAVQKHI

40782
HNCAGWDDG

40783
HQVATQLIK

40784
HTKAYNHCV

40785
HVHAVPNFA

40786
IADAQTMQP

40787
IFVAPMPGM

40788
IHVALDQSL

40789
IRVAKKGAR

40790
ITGAMMYMG

40791
ITLACVYHD

40792
IVMADVYGN

40793
IVNACMVEK

40794
LEIALHACW

40795
LFTAFRQFT

40796
LTSAVGAGM

40797
MISAFTGSH

40798
MISAPPDTW

40799
MTFAHDTTN

40800
NEFAIQNAE

40801
NKEARGALD

40802
NTTASTDVP

40803
PCKAKEKAV

40804
QQDAMEQYF

40805
RAAAQGPPQ

40806
RFFAAELQW

40807
RLEAMYNWG

40808
SAIATHQYI

40809
SEYAAMRDS

40810
SGCAMLNMG

40811
SKCAHSFEH

40812
SNWARSSAE

40813
SRFASWEGM

40814
SRMAIDVIG

40815
STSADPHCG

40816
TQMAHMIAH

40817
TTCAHSKMG

40818
ITEADDPTT

40819
VGEAPGHWQ

40820
VVGADRCQA

40821
WEEACKLDT

40822
WHSAVQHLL

40823
WVDAFCCQC

40824
YEQAMEYET

40825
YNNADCHYH

40826
YTQAMCITA

40827
TQHALSWWS

40828
NVLARCDDI

40829
VRAAQRSEP

40830
TQNACKVRT

40831
AAKALMSCH

40832
DFHACYKCS

40833
ICAAPNSAK

40834
LSMAMQVGC

40835
MNKAMTTWA

40836
QCNALNSFV

40837
RTCAYESGT

40838
TKNAMEIQE

40839
TPNAFVFAS

40840
TQEACHHDP

40841
CMEAQGECY

40842
QMQAWYCQE

40843
GTNAYMAHK

40844
AVKAPSQMS

40845
QSDAVHEPA

40846
QGVACLDQL

40847
NVYATPVVC

40848
TNAAYEMWF

40849
DQLALTFED

40850
RTEAHLPAP

40851
LDGYMNHDC

40852
LILPMGVNN

40853
LMSVMFAAH

40854
LQDHMCHNG

40855
LQEDMSPYA

40856
LVIKMHQLW

40857
LYHEMTHAF

40858
MCAVMDIFG

40859
MMQRMVYQC

40860
MNSLMQVAW

40861
MVQRMVYQC

40862
MVTTMGSTE

40863
NCCNMQLTF

40864
NCHVMSINL

40865
NGCGMVWQR

40866
NLAGMLPKN

40867
NMTGMPNGT

40868
QALKMENDQ

40869
QFYMMPFSL

40870
QGFFMMALM

40871
QKNHMYHQM

40872
QSDMMCHCM

40873
QTCSMLKDR

40874
RATEMAWSW

40875
SAGFMQEAA

40876
SHTRMGAFS

40877
SIAWMHLTQ

40878
SNMGMKHYE

40879
STDFMIHHF

40880
THDSMFFAC

40881
VAGNMDDHA

40882
VPYWMNTQQ

40883
VSVDMNCAS

40884
VTSLMQMGC

40885
YDTRMQTLG

40886
YRGWMNMED

40887
MSLWMATMQ

40888
AFHHMMNTS

40889
AHTIMMRDF

40890
AMDLMGDCE

40891
AMMQMMKAN

40892
ATAPMCFYQ

40893
CCCPMILFN

40894
CGEDMKICQ

40895
CMFGMVNYT

40896
CVATMHCHQ

40897
DELKMGHNI

40898
DGTPMHEYE

40899
DKMVMVFKR

40900
DKWTMQHWE

40901
DMVKMADCY

40902
DYEDMDQYY

40903
EGNIMMTQH

40904
ETSPMMTKP

40905
FQSQMPKMA

40906
FSQTMIFGA

40907
GMIGMARLA

40908
GVDPMVTHQ

40909
HKDTMHSFS

40910
HKFWMPNQH

40911
HMCWMATDK

40912
HMYGMQKHE

40913
INPKMHYAS

40914
LATTMQAFN

40915
LCEVMGCDA

40916
LENTMWKCA

40917
LGAVMTCHY

40918
LHQTMACQL

40919
LNGTMLAMS

40920
NANDMSSDR

40921
NCEPMNEWP

40922
NEPPMGLKY

40923
NESPMNDTC

40924
NITEMGAYP

40925
NTGEMAEYV

40926
PNILMMFEP

40927
PVGWMVQWP

40928
QCNHMDHVA

40929
QDNMMVRDY

40930
QISTMLARC

40931
QMATMFTCR

40932
QPISMQGQM

40933
QQVRMDRLD

40934
RDEDMHIAE

40935
RHQHMKYLD

40936
RTVDMNLKG

40937
SGLGMPNWT

40938
SGSCMWGYA

40939
SMIPMDSMP

40940
TPNNMWWME

40941
TQVLMHATW

40942
VAQTMHYHH

40943
VKDEMHYMR

40944
VLDQMHAPP

40945
VVNSMWLMH

40946
WFHNMLATN

40947
WGMCMQRPI

40948
YRTEMNEAH

40949
YVSQMWMQC

40950
YHAPMLMCN

40951
DPVEMCSYA

40952
DRTTMVGWY

40953
LPWHMMCMP

40954
AFCRMKFGG

40955
GTEMMGDHA

40956
PCWPMMIPG

40957
GTIRMPEHR

40958
FCSPMFKQE

40959
AKAGMSYSG

40960
ACAIMKMWY

40961
AHFQMNMSP

40962
CESKMCQEP

40963
CLYNMWQCE

40964
DTLPMAMDP

40965
DYGSMAASW

40966
EDFSMPFNH

40967
ESSSMNEEK

40968
IEVGMTFPE

40969
IHQYMTAHG

40970
LDKQMWQFT

40971
MEAMMKHME

40972
PQGIMWWTL

40973
TMWMMHSSP

40974
VGLSMHSMD

40975
VKGRMCNNF

40976
GSTHMNQEG

40977
YDYWMQAPT

40978
HNIHMWCNI

40979
MCLNMAKQG

40980
QINTMQMLS

40981
NNCNMNVMK

40982
VLTTMWKNN

40983
FHDRMAWAR

40984
AQWYMWCWD

40985
YTELMKKAC

40986
CISPMRVHQ

40987
YASTMHSIP

40988
CEFPMGPSD

40989
LLPIMHTHW

40990
FQHRMEEDA

40991
STALMDANM

40992
EFGEMLGVD

40993
VTSHMATAC

40994
PAYRMECCE

40995
GAQSMQFSC

40996
YVIPMVPSS

40997
AAFVMNGDG

40998
FTEPMCMQG

40999
LHDSRNKSP

41000
LRDFENLIL

41001
LTEDGNWWS

41002
LVITFNQGC

41003
MGLSFNHTR

41004
MSTERNRAC

41005
PQOHINFSK

41006
QCTVQNQFG

41007
QMELSNQWN

41008
QTVDHNITL

41009
SAKYDNVQI

41010
SAYSTNGDA

41011
SCQDVNNYH

41012
SHGYGNRFA

41013
SHYQDNIGF

41014
SSKPSNEMW

41015
TKKEINDCC

41016
TMVMNNWVA

41017
TQHSNNQAT

41018
WHKLHNRHV

41019
AHAFNNVVD

41020
AKHWFNESG

41021
AVVGANQVL

41022
AYPHRNVGL

41023
CKQQFNGQN

41024
CMNEWNKLG

41025
CMVDANHLW

41026
CNFNNNDHA

41027
CTLFSNFQY

41028
DSCKANTQS

41029
EAWMHNFNW

41030
EIYITNCTS

41031
ERHGFNHCN

41032
ESDPRNLGP

41033
ETEERNGFG

41034
GFGHSNGFL

41035
GISTFNLWK

41036
HMTEKNMGE

41037
HYMMNNYLV

41038
ICAIVNTCN

41039
ICSGVNADN

41040
IQVRWNAEL

41041
IRYILNPEF

41042
ISDCFNCAD

41043
LCAHYNPHN

41044
LHGNYNYSV

41045
LNSHINGHM

41046
LSCCTNVFC

41047
LSCMRNMSL

41048
LSDYVNDEG

41049
LVMPYNSAQ

41050
LYEERNAQM

41051
MCNGYNVSF

41052
MHVFSNVCG

41053
MQYTINYQD

41054
NETSLNQHM

41055
NFVYWNYLP

41056
NGVNINQCF

41057
NHVNANLRA

41058
NKQYFNQKD

41059
NSHDYNCFH

41060
NSLHNNEDP

41061
NTGMDNGQG

41062
NVLPFNCNF

41063
QHEYLNQKS

41064
QSQRLNGQI

41065
RTEYANHMH

41066
SFYSTNFQL

41067
SSGWLNSEN

41068
STCIDNGRT

41069
SVWWGNSAA

41070
TPMWVNMVC

41071
TVGVINLVR

41072
TVELLNPGT

41073
TYHCQNHDH

41074
VHQWTNMQF

41075
VPQTYNVEA

41076
VQDLFNQFC

41077
VVHCNNTSW

41078
WDCLLNKWR

41079
YGMYENFHC

41080
YRCHNNEKT

41081
IEEGKNALY

41082
GSATPNETP

41083
YVSLCNDGN

41084
ADHLHNVWI

41085
CHCHYNPFC

41086
CVMPSNCQD

41087
CYDMWNMTT

41088
DIQYKNHMN

41089
DKLLSNSWR

41090
DTESHNETP

41091
FKMDTNFAP

41092
ITYEGNISP

41093
LHLWRNSGE

41094
MTTHSNCGP

41095
PTKYHNMIG

41096
QLSKVNVYE

41097
RHSDHNYIG

41098
SNQHNNQSM

41099
SPGTENRGG

41100
SQMQVNPLH

41101
TLTEVNRLA

41102
TQEHINIWN

41103
TSHFKNCIM

41104
TTWEQNVMD

41105
TVCFENLGG

41106
YALLHNAMW

41107
MSARINTPD

41108
ERNMFNICH

41109
GRQSINEIA

41110
STGCYNWPQ

41111
WGQSYNYWP

41112
AVKSCNTCK

41113
QESMCNYWQ

41114
DKCLNNKEY

41115
HMAYCNKVC

41116
GAMLNNNAS

41117
GKTQYNHHN

41118
TYDTRNHCL

41119
NGNHYNAMR

41120
DYSQVNCEK

41121
GTMKHNGRA

41122
EKIRQNAAP

41123
NLERCNTMM

41124
SRYVNNGTV

41125
LRHEGYHSE

41126
MEYWPIHCQ

41127
MFLPTDHQF

41128
MQHEETHDY

41129
MREQKSHGY

41130
MTHGHSHSF

41131
NLDMVTHAE

41132
NTQGTVHIQ

41133
NVDNHDHFE

41134
PQPSEYHWN

41135
QEHQYEHQS

41136
QGPECTHNP

41137
QSKWQDHDY

41138
QSVWLEHTL

41139
QSVYDMHTP

41140
QVDGQCHTT

41141
RDSRPGHQY

41142
RQFKQTHFQ

41143
RSPHVHHQL

41144
SAHTDRHNS

41145
TSPCDMHFS

41146
TYNPWQHFE

41147
VHCQTDHDG

41148
WAHYKAHYP

41149
WQDFYIHYA

41150
WRYMWPHRQ

41151
YAVMYQHHA

41152
YCLQYAHKG

41153
AFWNPTHFD

41154
AMIPNLHME

41155
AMLGYQHQP

41156
AQDIYEHMQ

41157
ASYHWTHAG

41158
AVVIEKHDG

41159
CMKSYDHGS

41160
CNVWPCHQE

41161
DADLAAHWP

41162
DAQLGMHFI

41163
DKANCYHWL

41164
DKIPIDHFA

41165
DKIPIDHFV

41166
DVHHLTHSP

41167
EANKAFHLA

41168
EHGDEMHPP

41169
EHMNCMHCE

41170
GAGMRMHDT

41171
HKASWCHDQ

41172
HMMLRDHNN

41173
ICSIYGHHQ

41174
IDSQIYHML

41175
LCAEFPHGR

41176
LITQPVHQQ

41177
LQAGGEHNW

41178
LTLESPHPA

41179
LYKWIVHNS

41180
LYYITEHST

41181
MAVSQSHQF

41182
MAVWVKHQH

41183
MSSMLEHSW

41184
MYGFYGHDM

41185
NAYQNSHYP

41186
QLYGRRHWH

41187
QMMDFQHPA

41188
QNYWGRHPA

41189
QSHYRMHSA

41190
QVNHVRHWS

41191
RHICFQHCK

41192
RIEIGQHYV

41193
RSEVSSHLD

41194
SANWFTHDK

41195
SVSLKVHHE

41196
SYVTTQHST

41197
TASGIDHPK

41198
TQAEYSHKS

41199
TTWGTGHMA

41200
VFNLYSHSH

41201
VIKPFAHHG

41202
YASYKCHGD

41203
YVQSAQHQQ

41204
YYASLDHYL

41205
NFWNQVHCV

41206
MATNKYHIQ

41207
QYYNYCHMN

41208
GCMCIRHAY

41209
YIYFALHTH

41210
WFDRSWHIL

41211
NIAMPAHIP

41212
HDPLAGHWP

41213
QRQGQLHYW

41214
IACISMHNH

41215
AQEHLDHCL

41216
ECTMRMHSH

41217
EDSFSFHTI

41218
EYCETMFIN

41219
FMPYCVHSG

41220
HSKNTVHKR

41221
IMGIHLHPA

41222
IRNIAKHAL

41223
LDKRQDHNF

41224
MQAEACHRW

41225
NTHLFHHYL

41226
QHTSAFHTM

41227
WHKCLSHQD

41228
WRKCLSHQD

41229
NAYYDSHCE

41230
HDPPPMHWL

41231
RCQLFDHDE

41232
LGKVQTHIR

41233
DVIHTTHSG

41234
TKGRNDHME

41235
NSTRWYHTG

41236
DTDNKPHFA

41237
QGYSHLHAL

41238
CQAGTDHFP

41239
CASSKSHYL

41240
ALGNRHHQC

41241
VQYYYGHMQ

41242
GVFYVGHWQ

41243
LVQQDWREW

41244
MCAYLHMFH

41245
MEKVKCSFA

41246
PKNWAPMFT

41247
QQDIYIDFF

41248
QTEHLWMFH

41249
SGHWNDYFH

41250
STIPIWNFL

41251
TEQFSRLFA

41252
TQDSKMGFG

41253
TVAWATAFE

41254
VKKLAKYFH

41255
VKVDKSQFD

41256
VVNIRGSFQ

41257
AFEVSESFE

41258
AGEMDEAFG

41259
AMELADDFA

41260
AVDHCFKFE

41261
DFKKPSLFR

41262
DMQTICRFP

41263
GMGWQVSFN

41264
GPIPSEVFP

41265
HCYIKEGFQ

41266
HNMPSGQFT

41267
HYESRLDFY

41268
ICKSDPVFD

41269
LEPCTDEFG

41270
LIMRISPFD

41271
LSGSIMEFN

41272
MVSENSTFW

41273
MYSRTIPFH

41274
NKHEVGCFG

41275
NKYEVGCFG

41276
NMHWRYQFE

41277
PYPTHSMFS

41278
QDSHVHSFF

41279
QGQYWHYFK

41280
SCGHQTRFG

41281
SCGHQTWFG

41282
SDHYCGTFY

41283
SHMMSHAFW

41284
SNSLYRAFL

41285
SSVCTQIFT

41286
TGEMDEAFG

41287
TNDIWWQFS

41288
TRTELANFF

41289
VCWLWFYFG

41290
VGICHPEFW

41291
WVTNQSDFA

41292
YPQCAQWFW

41293
HQKMTYAFT

41294
VRWGHTCFY

41295
NSAECREFQ

41296
WKNNQKWFK

41297
DIGSYKGFQ

41298
DSAHYCTFT

41299
EKSTILPFA

41300
EKSTILPFT

41301
EVEYGCWFE

41302
GNMRQSRFP

41303
HGLHEDSFY

41304
TNNYPMPFD

41305
DMPPLHSFC

41306
ASTMKSIFE

41307
SLASEVQFQ

41308
DLTTDTIFQ

41309
SADRTAMFH

41310
PTYPYEWFQ

41311
NMKPKDVFR

41312
SATIRFYFN

41313
FVPWCHSFG

41314
ITEQYDAFA

41315
NTMCQLNFC

41316
LYQKDQRFS

41317
LFGTHPGQA

41318
LQELSSQYA

41319
LTVNLMKIA

41320
NAIDCMSGA

41321
NAWIDWSLA

41322
NQGTSAWGA

41323
NSLPFGMVA

41324
PFNKSKGLA

41325
PNONGTQGA

41326
QDQQVHEGA

41327
QLMIQGSYA

41328
QSAYLIMVA

41329
QTTFAERHA

41330
SDHGRIMTA

41331
SLAHWDGRA

41332
SSGWTQQMA

41333
TFIMLGADA

41334
VDYYSPYLA

41335
VGCIAQAAA

41336
VNDCNWCEA

41337
VRTPWQTVA

41338
WFNVKKTAA

41339
YAPQQHMWA

41340
YSQKQCPTA

41341
ACYQQCVSA

41342
AKSSIEYSA

41343
ALWDKFEAA

41344
CDYHTLSPA

41345
CFENLQVMA

41346
CRDWDGFNA

41347
CTTDTPQNA

41348
CYHECQGCA

41349
DIFFKCEWA

41350
DVGGRAVQA

41351
EATDLCLNA

41352
EATDLCPNA

41353
EDNHYQMIA

41354
EFYNYEIMA

41355
EGGSNMMNA

41356
EKAMLLNGA

41357
EKTCVTREA

41358
EQAMWMYPA

41359
EQGHCEVAA

41360
ESKWYYVMA

41361
EVERDKVGA

41362
FDEKRSESA

41363
FPANYHSNA

41364
FVSTQMPIA

41365
GPNDWMFDA

41366
GYVAWMYVA

41367
HCDDVLWCA

41368
HSAYFGVGA

41369
IEELGKPRA

41370
INGMKSAMA

41371
INVPDLLGA

41372
ITSIRMYQA

41373
IVQPYHVEA

41374
LHQPRMDCA

41375
LIYHTPVMA

41376
LSSCRGLIA

41377
LTIRDYGEA

41378
LVNVRYMHA

41379
MHEPILKYA

41380
MICWWKQPA

41381
MMNHSQLYA

41382
MQHPYMQMA

41383
MSTVSGSYA

41384
NGFMAAVMA

41385
NHADYMRNA

41386
NMWPTFRAA

41387
NTDEESGMA

41388
NTMMLQSAA

41389
QAPLKRAAA

41390
QCNRCDQYA

41391
QFIQYGTCA

41392
QIGLKMTPA

41393
QLNPWFYYA

41394
REISVEAVA

41395
SAKITWASA

41396
SIVEFESAA

41397
STHMATQLA

41398
TKSSIEYSA

41399
TNLRQHINA

41400
TVSDGDWIA

41401
TVVLTLKEA

41402
VTSWFHEIA

41403
VITVSGGCA

41404
VVMCKSRDA

41405
WVMHYYQYA

41406
YSSPDLRPA

41407
TVLVPVQAA

41408
EVGIKYRAA

41409
EVNWQCQYA

41410
GSMFNPGAA

41411
MTNWFPACA

41412
MVKTSCKPA

41413
MVKTVYPDA

41414
PPEEQCGMA

41415
QFTETWPCA

41416
RIIWPDVVA

41417
SSDTPANQA

41418
TFVECIQYA

41419
TGHFFTTAA

41420
VGYGCKGYA

41421
VSWSRMEQA

41422
YRLHCCNRA

41423
MRVQVFSDA

41424
RTESWSSLA

41425
QKWMQMDGA

41426
TAAGYIVDA

41427
TQQTADRWA

41428
FQIRQDNLA

41429
NTGMEGQMA

41430
IMTMQMDTA

41431
EYSSQHPYA

41432
QPFCFSIGA

41433
MGVPPLMQA

41434
VFGSTRSAA

41435
ATNYTMEPA

41436
HIHMSDRGA

41437
GSHDYYQSA

Example 16
AAV5 Variants with Tissue Tropism in Lymph Node

This example describes engineered AAV5 variants with tissue tropism in lymph node that were discovered using the methods and systems described in EXAMPLE 1-EXAMPLE 2.

CNS, liver, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord tissues were analyzed to identify amino acid residues in the AAV5 VP1 581-589 region that drive lymph node tropism. The results are shown in FIG. 18G which shows the positional frequency of each amino acid in the 581 to 589 region of AAV5 VP1 in lymph node tissue as compared to all other analyzed tissues, wherein the capsid polypeptide has the sequence of SEQ ID NO: 2. The position (“Xaan”, n=1-9) is denoted on the x-axis and amino acid residues are denoted on the y-axis. The frequency of a particular amino acid residue in a particular position is indicated in the heatmap as intensity. Darker squares indicate a higher frequency of a particular amino acid in a particular position and lighter squares indicate a lower frequency of a particular amino acid in a particular position.

The frequency of a given amino acid residue occurring at a specified position corresponding to position 581 to position 589 of SEQ ID NO: 1 (generalized in SEQ ID NO: 2) in variants identified in lymph node over the frequency of that given amino acid residue occurring at the specified position in variants identified in all other harvested tissues was analyzed to identify a set of sequence rules for capsids that preferentially target lymph node tissue. With reference to TABLE 31 below, and SEQ ID NO: 2 (AAV5 VP1), the following amino acids can, thus, be independently mutated, in any combination, at any one or more positions Xaa1-Xaa9, to provide a VP1 capsid with enhanced lymph node tropism as compared wildtype AAV5 VP1 capsid (SEQ ID NO: 1), where lymph node tropism here refers to properties that are preferred for lymph node transduction over properties that are preferred for transduction of all other harvested tissues.

TABLE 31

Lymph Node Tropism Rules

Xaa1 is selected from A, D, E, Q, S, or T

Xaa1 is selected from D, E, or T

Xaa1 is E

Xaa2 is selected from A, H, I, S, T, or V

Xaa2 is selected from I, T, or V

Xaa2 is V

Xaa3 is selected from A, E, H, I, T, or V

Xaa3 is selected from A, I, T, or V

Xaa3 is T

Xaa4 is selected from A, D, E, or P

Xaa4 is selected from D, or E

Xaa4 is E

Xaa5 is selected from I, L, M, V, or Y

Xaa5 is selected from I, L, V, or Y

Xaa5 is L

Xaa6 is selected from D, E, I, N, or Q

Xaa6 is selected from D, E, or I

Xaa6 is D

Xaa7 is selected from A, E, G, Q, or V

Xaa7 is A, Q, or V

Xaa7 is V

Xaa8 is selected from F, G, M, or W

Xaa8 is selected from F or W

Xaa8 is W

Xaa9 is selected from I, P, T, or Y

Xaa9 is I or P

TABLE 32 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that were found in lymph node tissue and comport to one or more of the rules provided in TABLE 31. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 19118-SEQ ID NO: 20, as disclosed in TABLE 32. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 32

Sequences of the 581 to 589 Region in AAV5

VP1 Capsid Polypeptide that Drive Lymph

Node Tissue Tropism

SEQ
581-589

ID NO
Sequence

19118
ETTMWNSWA

19119
ENTMWQSSQ

19120
ERQAHNHYA

19121
EAKSRPQYA

19122
FCQQDDTDI

19123
EDICHWHHT

19124
EECIMQRSK

19125
EEERTCKHC

19126
EEFARIPGH

19127
EEPRCVNPC

19128
EFFCQADKA

19129
EGAGLYEFA

19130
EGPELTCDK

19131
EGTLRYTMS

19132
EHDTQDISV

19133
EHHPLGVMG

19134
EHHYCVPWS

19135
EHILTAERI

19136
EIAAKIFCR

19137
EIILIWKQC

19138
EIMSYPHTN

19139
EIQHVSHEH

19140
EKHGCDKAE

19141
ELNHKWHSV

19142
EMALVVSNQ

19143
EMHQFMECQ

19144
EMRFTNFCY

19145
ENIHNNDCT

19146
ENKEKEGQW

19147
ENTMAIIEH

19148
EPILYHEHK

19149
EPIQIFNDA

19150
EQHRDTMRP

19151
EQNACDHYG

19152
EQVFGFWNY

19153
ERMETFIXP

19154
ESCEDEGIL

19155
ESKASGANG

19156
ESMLPIMGR

19157
ESVPLEYAN

19158
ETCCFTNMG

19159
ETCGQGGYT

19160
ETDHCESWY

19161
ETPIEWDEQ

19162
EVEVHSTAL

19163
EVHIAPSAC

19164
EVKRITEQS

19165
EVKRWMMVD

19166
EVMAMVDVA

19167
EVPQNELNV

19168
EVRSIEYWN

19169
EVWHTKILI

19170
EWAFNYNQN

19171
EYKMFEKYE

19172
EDHWHGPGD

19173
FVALKAVID

19174
FVAPVDSFY

19175
FVFEGPMHQ

19176
FVKVRIWTQ

19177
FVQPTFIRH

19178
FVWYGPRNH

19179
GVACVQETD

19180
GVANFMLNN

19181
GVCEENEWM

19182
GVCQMCCWP

19183
GVDARHAQR

19184
GVEFCLING

19185
GVEWPPFMQ

19186
GVFMTWQWL

19187
GVHGINVMM

19188
GVHIMQCQF

19189
GVHIYACHY

19190
GVHQVPTWN

19191
GVLQMDWQE

19192
GVMQHWTCD

19193
GVMWDGVAP

19194
GVQMTVRQA

19195
GVTYAQRIC

19196
GVVFCDQKD

19197
GVVLFQGGA

19198
GVVNSMQTK

19199
GVVQQGAMI

19200
HVFVMDGPI

19201
HVIWDSMKL

19202
HVMDHVATS

19203
IVDFENRHP

19204
IVDPIFRQT

19205
IVFEGGIAS

19206
IVGAPCLVT

19207
IVGDCQFHV

19208
IVGSNTYGY

19209
IVHNVEYSN

19210
IVHSQPNSV

19211
IVLQLQQAK

19212
IVNALEFCK

19213
IVNNAQMHE

19214
IVQMHYINQ

19215
IVQVESCQN

19216
IVSARDTHN

19217
KVDGMDIEG

19218
KVDPPGIFW

19219
KVDQGLCQP

19220
KVDWDGNID

19221
KVEPHHPAD

19222
KVNPKEFQD

19223
KVPICAHKQ

19224
KVQICDFWC

19225
LVCLCHSCA

19226
LVDQYMWQN

19227
LVDRVKHVD

19228
LVDYWAWNG

19229
LVEQERMGV

19230
LVFAKLDQN

19231
LVGICDLCP

19232
LVGSTSHVT

19233
LVMHQQRNY

19234
LVYQHDTAP

19235
LVYTELSGA

19236
MVATESHDK

19237
MVDDPGQQF

19238
MVDHWGDWN

19239
MVDHYICMY

19240
MVDSFGKYL

19241
MVECEGEQY

19242
MVEDSTCHW

19243
MVEEANVPC

19244
MVEEGSWNI

19245
MVEGRQDLT

19246
MVENCDLPP

19247
MVEWGGSNQ

19248
MVGCFYTWT

19249
MVGYQYPAA

19250
MVHEAELYA

19251
MVKEDITEH

19252
MVKEQWQCT

19253
MVLPANGFQ

19254
MVNALTDYG

19255
MVNPWVVFY

19256
MVNQAWQTD

19257
MVQPMPQHH

19258
MVSFNYEIG

19259
MVTKEYMCC

19260
MVTMGYWSS

19261
MVTPYWNQY

19262
MVYPKRGEN

19263
MVYSGQQEF

19264
NVAFAHCGG

19265
NVDDHPTMC

19266
NVDMRNVES

19267
NVDTPSGNQ

19268
NVDWDSECS

19269
NVECCLNRP

19270
NVEEKWEWW

19271
NVEFRYFYN

19272
NVESFPHEQ

19273
NVEWWGHCA

19274
NVFATNCWV

19275
NVGAHFAPF

19276
NVGGPCYNL

19277
NVGGQNMEH

19278
NVGIGHGGN

19279
NVGIHFAQQ

19280
NVGNEHIAY

19281
NVHIALEQH

19282
NVIQHAIGG

19283
NVKFSIEHT

19284
NVLGTLVHV

19285
NVMDCESCC

19286
NVSYEQNFM

19287
PVATALRMP

19288
PVNLNCTVS

19289
PVSLCRFGG

19290
PVTGENHMN

19291
QVALRGATC

19292
QVDDCGHLF

19293
QVDMELPCW

19294
QVETTMIGH

19295
QVIQNVWER

19296
QVIQRRNVM

19297
QVMIECRQS

19298
QVMNGSPKT

19299
QVQIGFEDG

19300
QVVTWGYEY

19301
QVYFWGTHS

19302
RVCDKNLDL

19303
RVDEKLDHD

19304
RVDTNMRDR

19305
RVEHWVNMS

19306
RVNEWGAYG

19307
RVPLYPYWT

19308
RVSPWDQWF

19309
RVVMPLGSL

19310
SVAYKWDAF

19311
SVDACDYDD

19312
SVDDCMHSA

19313
SVDNDRAWP

19314
SVEAKSATC

19315
SVEFHEDFS

19316
SVEGANAEV

19317
SVEGVIIDR

19318
SVEYYSMPQ

19319
SVFQTHCDW

19320
SVGPLQYWK

19321
SVGWGWMMG

19322
SVHEWDPCT

19323
SVHGKVSWF

19324
SVHTIWPMA

19325
SVICWTHMA

19326
SVIYNEHVT

19327
SVMKQELSV

19328
SVNMYMLPD

19329
SVSMASAHG

19330
SVTACPNWN

19331
SVTASYGFS

19332
SVVEDAMRG

19333
SVVPTNNDM

19334
SVYAPTKWG

19335
SVYWPTTEF

19336
TVADMFFDG

19337
TVAHVADCY

19338
TVAPCNQGL

19339
TVATHCFIH

19340
TVAVWWRNA

19341
TVDAIQGWQ

19342
TVDMRHFFT

19343
TVDSEHRLC

19344
TVEAMHLGP

19345
TVEDGQWSW

19346
TVEFTYGWC

19347
TVEPTNYDG

19348
TVFHSTGVG

19349
TVGICFCPV

19350
TVGLLEMTR

19351
TVGTGCDTC

19352
TVGWYQHDW

19353
TVHNCLTMG

19354
TVLSTTQKD

19355
TVMHHDGRL

19356
TVMSWQQHE

19357
TVMWQTQIG

19358
TVNIILQGS

19359
TVQMYQDLE

19360
TVSESRKHN

19361
TVTDEDDCN

19362
TVTIAVQNM

19363
TVVQWHAMI

19364
TVWTEVSPP

19365
VVDPRPDWF

19366
VVDRWDHWM

19367
VVDSNFCSM

19368
VVGCDKSDA

19369
VVHEAWCDT

19370
VVHNFLDQS

19371
VVKEFTGHY

19372
VVKICGCCR

19373
VVMLQSGMT

19374
VVMREWRCF

19375
VVNQGVACT

19376
VVQHMANGI

19377
VVRWPGSEW

19378
VVSRNEVGA

19379
VVTFTNILQ

19380
VVVCRPVAP

19381
VVYEKQDHM

19382
VVYPLAPYR

19383
VVYSNICVE

19384
WVASQGAQC

19385
YVANVPVLW

19386
YVLGRHCVF

19387
YVQIKGLQL

19388
YVTALATYD

19389
SVTNGSTKH

19390
LNCERAGVE

19391
KVNCRMIIM

19392
GVWMQGFHP

19393
AVHTADVCC

19394
DVKSWAVCD

19395
SVLHVQALA

19396
AVSPHWGNY

19397
CVVKRMAEF

19398
CVVNMEYCL

19399
CVWSRHQIC

19400
DVRLIGHEA

19401
DVVRQWQAP

19402
FVEEDLYEE

19403
FVHMWSRLC

19404
FVRPLEEFY

19405
HVFQGDYIA

19406
HVGNLTAFN

19407
IVANAQIPG

19408
EVGYQNCQS

19409
EVLGQMTCT

19410
EVSEWEDWQ

19411
EVSNSVCRV

19412
KVCVVAGPM

19413
KVEVKRDYW

19414
KVVDRETYS

19415
KVYEIGEAP

19416
MVEWEEEWD

19417
MVLIRGMSF

19418
MVLPTKLWD

19419
MVSVVQEPN

19420
NVLSEVYGH

19421
PVKDCILWS

19422
QVCSDWCEC

19423
QVDTRCYGG

19424
QVMMDDSNV

19425
QVSCNVEPK

19426
FPTQMKMCN

19427
FPTWDKPPT

19428
FTTCFSIMV

19429
FTTFWVPQQ

19430
FTTLHNMEN

19431
EYTCELACV

19432
GETTIWPDH

19433
GLTADLWHY

19434
GMTYVSGHH

19435
GNTLIVTGP

19436
GSTTQHDGT

19437
GWTPHTLER

19438
GYTANFYTN

19439
HGTTEASGP

19440
HKTIDDIFT

19441
HNTEPPGMP

19442
HPTNMSACE

19443
HSTGSGTFH

19444
HSTPFVFNS

19445
IATNQSAIL

19446
IGTAYPQWR

19447
IGTIHPQAQ

14948
IGTVTVGAY

19449
IITMMCRSD

19450
IMTSVDSQR

19451
IRTRELDQY

19452
ISTEKNESY

19453
ISTLFSSGD

19454
ITTKYDCMG

19455
ITTPFYKVG

19456
IWTRADLGR

19457
KMTVRQSDS

19458
KTTDDTVGE

19459
KWTQFGWMD

19460
KWTRIGDSA

19461
LGTQEQGMP

19462
LHTRTQCEP

19463
LNTGYMWNQ

19464
LPTSVNMMH

19465
LQTAIPEWP

19466
LSTTEDEMP

19467
LTTAVAIRE

19468
LTTQLFLQE

19469
LTTQYECGP

19470
LTTWNTYVN

19471
LWTQISEFA

19472
MATEGACNP

19473
MATMFTNWC

19474
MITCIDGIF

19475
MPTLCECGL

19476
MSTMSMDSY

19477
MSTTEVLAE

19478
MWTGQLRQC

19479
MYTQWSKAP

19480
NATNSQNMY

19481
NATPYKAEY

19482
NFTVTNLQM

19483
NITLGAACE

19484
NNTNQCQNC

19485
NQTEMSSNG

19486
NRTCNAGEW

19487
NRTGPAAWA

19488
NSTAFFRDQ

19489
NSTIQGGAW

19490
NTTEISRLP

19491
NYTERVQDL

19492
NYTNCEFGN

19493
PATTADMHY

19494
PPTHQMWYC

19495
QATSAMIPS

19496
QCTCEGEAQ

19497
QDTELYCCM

19498
QITQMGRQE

19499
QLTWQAHSW

19500
QMTPGWWKT

19501
QNTCLPVFY

19502
QTTSAMING

19503
QYTHGTCFS

19504
RYIGTWQDC

19505
SATYTEMQS

19506
SETHNDHGW

19507
SKTCEIGKT

19508
SNTAYFNGS

19509
SSTAEPNGR

19510
STTAIDCMR

19511
SWTACTPCH

19512
SYTYRLTDQ

19513
THTEEDVFN

19514
TKTAFGDMG

19515
TNTAFGCYG

19516
TSTEYKMGL

19517
TTTMLHRIN

19518
TTTTQDSQR

19519
TTTYQALNA

19520
TYTCNKWFR

19521
VATKFETNT

19522
VATMAEDNP

19523
VATTDGPVY

19524
VCTNLAHGA

19525
VDTIFTDEH

19526
VGTPLDTSV

19527
VETVFMEQN

19528
VKTPGATDF

19529
VPTNTFDTH

19530
VSTAVMPEG

19531
VSTGLQDAC

19532
VSTTGPEDG

19533
VITEMPMME

19534
VITGYQCAS

19535
VTTTTKVSN

19536
VWTQEMWDA

19537
WDTVPKIRD

19538
WFTGPWAMH

19539
WRTQCQSEK

19540
WYTYNMCER

19541
YATFQDCNA

19542
YATNQCYNG

19543
YCTDQSETS

19544
YETKGNLAR

19545
YMTFKGYHG

19546
YPTKDPFKN

19547
YSTQCNTMK

19548
YTTDDGMSP

19549
YTTHRDTVT

19550
YTTVDNSLQ

19551
KQTVEVDQE

19552
RITQGESGM

19553
ESTKWDYEA

19554
QQTKNNSTD

19555
WCTQFDEYT

19556
KITVHDRIP

19557
DMTLQSVSS

19558
PLTIEVNCT

19559
QCTAHNTCM

19560
FNLEFSPTL

19561
FPQESNHDG

19562
GAVEIDCLY

19563
GHAEQWFES

19564
GHFEFMRDA

19565
GHVEVCTLS

19566
GIGEEMFWD

19567
GSPEVSVAS

19568
GWSEAHKSD

19569
GYYEVVKMY

19570
HGYEIQFFQ

19571
HHDEKLKML

19572
HMDELEYCG

19573
HNKEKWPTE

19574
IACESDESW

19575
IHYEHGMQC

19576
ILLEFQAPM

19577
IQQERNRYH

19578
ISAEHIFAG

19579
ISFEWLELN

19580
ISSEAIFYP

19581
TIMEILGQN

19582
KACESIQDL

19583
KAEEIENMC

19584
KAIESMLEM

19585
KAMESHDCC

19586
KFEEVHASG

19587
KHPEQYRKY

19588
KHVEALKAY

19589
KKYEECPLA

19590
KMVEDEPSA

19591
KNEEDYALY

19592
KPVEIDMEG

19593
KQYELNNWC

19594
KSSECGWAC

19595
KTIEQTIPH

19596
KTYERYQCA

19597
LAKELCCIQ

19598
LANEKDSMC

19599
LFREKMFNP

19600
LIVEHDCWF

19501
LKCEYPPMS

19602
LLDEVRQAG

19603
LLWECEKGV

19604
LTLEQSVPM

19605
LWYEVMIWY

19606
LYKENWQFP

19607
LYQENQHEW

19608
MFAEYSHIG

19609
MFEEKNLHT

19610
MHKECEACR

19611
MHMEMNYSE

19612
MIAEFVSNY

19513
MNEEQEHGA

19614
MPDEQQQMW

19615
MQAELTMSA

19616
MRHEEQRLK

19617
MRHEIVAMY

19618
MSAEVLLMT

19619
MSCELTGMH

19620
MYVEYTVGM

19621
NAVEILLCG

19622
NCNEPKYAA

19623
NEYECMCWY

19624
NGDEFTVHG

19525
NHAEFLSGC

19626
NHGEYFDWF

19627
NHYEMQDWE

19628
NKLENCEGM

19629
NNHEQEWRN

19630
NSREAEMLR

19631
NTHEFKFQF

19632
NTHEFMMQD

19633
NYDEKHVCH

19634
PFFERNIRE

19635
PKFEFYEMI

19636
PMEEMMHFC

19637
PTNELYDNE

19638
PYPEASVKM

19639
PYREGFRSP

19640
QALECTCQR

19641
QARECEAMD

19642
QASEHDTVV

19643
QHNEKPMDA

19644
QHYEFQKHC

19645
QIDEGMYSE

19646
QINEFAEHE

19647
QMREIQAEQ

19648
QNPEVHFSK

19649
QSEECMECR

19650
QSNEDLADI

19651
RAMEMQDQG

19652
RCNEYTGEK

19653
RIIEQDEIT

19654
RINEQDECM

19655
RWMELENAT

19656
SAGEWGQMW

19657
SIDEFQENW

19658
SPDEFGTFR

19659
SPHEIFVWN

19660
SSEEWCQCE

19661
SSVEGIRER

19662
SWAENKQSF

19663
TAEEIWYER

19664
TCCEPSWND

19665
TGNELYCWT

19566
THAEFEHMH

19667
TKMEYQGAT

19668
TMQEFVVKG

19669
TMYETLLMG

19670
TQNEHGKYD

19671
TREEDDKVN

19672
TRMEQPSGR

19673
TSNEMKKHW

19674
TTEEGQTFR

19675
TWGENGAWC

19676
VCEEYYQMR

19677
VFEEDAYGL

19678
VFLEGMHFP

19679
VKNEQTVFF

19680
VLGELGDMG

19681
VSPECPSFT

19682
WGAEKQWAF

19683
WGHEIMTEW

19684
WHDEKSRYL

19685
WENWENAGQ

19686
WMIEMWNCM

19687
YADECAYGR

19688
YAGEHDMGQ

19689
YANEPQHHF

19690
YMVEYAKYG

19691
YNEEEWFFA

19692
YQVERGASA

19693
MGKEYSQCF

19694
TYEEENGFL

19695
NTCEGWSHF

19696
GLFEGSPGA

19697
MWHELIADQ

19698
NQREWHGLA

19699
GDYQLLNSP

19700
GFIDLNSRR

19701
GIDWEDQVG

19702
GIMLLPNGH

19703
GSLTLLEQT

19704
GYVSLWSMY

19705
HCVALMMDR

19706
HEEVLIAGS

19707
HGLTLKWDC

19708
HHESLGMHM

19709
HMEALNTNV

19710
HTGFLTKGM

19711
HTICLAVTP

19712
HTWQLHNWN

19713
IDLNLNIQI

19714
IHDTLGAVA

19715
KCEVLEVQD

19716
KEVYLSTAA

19717
KHVDLSNIP

19718
LGNQLAYGG

19719
LHYPLVNDG

19720
LMDHLCTPN

19721
LRGCLHLEH

19722
LTNFLQNDN

19723
MFLPLHYKD

19724
MHELLQSVH

19725
MIAQLHTWA

19726
MNRWLLTDG

19727
MPQSLRVCS

19728
MYEWLAHCW

19729
NANLLTEQY

19730
NEMRLECSY

19731
NHKWLKCSL

19732
NHVSLNCDP

19733
NISWLHSFH

19734
NTCYLYAFF

19735
PDPVLSIKD

19736
PHCTLSFIG

19737
PRKSLWMEP

19738
PWAVLNQEP

19739
PYEQLGTII

19740
QFGSLVNSW

19741
QGYKLDNQM

19742
QKEYLMNSG

19743
QMWTLPSHN

19744
QPMVLNQDC

19745
QQHRLTFMP

19746
QSQSLTEEA

19747
QTNALLNGE

19748
QWNGLNSCL

19749
RCLDLVNEP

19750
SADQLHQFA

19751
SAEDLYYHY

19752
SCVCLEAHQ

19753
SDWPLMRYQ

19754
SEVGLPCYQ

19755
SFNQLICGL

19756
SHDDLRMQC

19757
SPGILQMIG

19758
SREILDVVA

19759
SSQHLWHRD

19760
STVTLHTHQ

19761
SYDGLDQVA

19762
SYEILGDWC

19763
TCLNLSWYG

19764
TFGFLSDPQ

19765
THEHLANHN

19766
TMEPLGWSF

19767
TSEVLDHFC

19768
TYMQLSKYC

19769
VFHTLHTKG

19770
VGMPLRKEE

19771
VGNCLHNYW

19772
VIYQLGEMV

19773
VREFLNGLN

19774
WCWQLHDQF

19775
WDDLLSFWC

19776
WTLMLHNFM

19777
NMPSLRTRI

19778
ASFFLNPLE

19779
ATMNLSWGL

19780
FQDPKDTVG

19781
FTNSDDDVS

19782
GCEACDFMG

19783
GDNKIDMHN

19784
GHMWTDSMA

19785
GIHVTDLQE

19786
GMQWPDSME

19787
GNDIWDFMA

19788
GQNVMDYHA

19789
GSSVDDGVS

19790
GTECIDWTA

19791
GTNAKDGTP

19792
GWDRHDGFV

19793
HDKDFDWAN

19794
HHDTSDMSC

19795
HHGPMDWGA

19796
HIMCYDEWY

19797
HLYIKDMTD

19798
HSMVHDCFN

19799
HSWTRDRLE

19800
HWVMGDVKL

19801
HYDQRDEHQ

19802
HYHQSDWMH

19803
IAIPEDKQY

19804
IASLEDMNG

19805
IEMVRDCGY

19806
IHLKSDYVW

19807
IKDWVDGIQ

19808
IQQTIDMVA

19809
ITHMIDQDN

19810
ITWPEDGSI

19811
KDYASDIHK

19812
KEFRMDSGS

19813
KHHTSDTAV

19814
KIYDHDAWT

19815
KSLCYDDWM

19816
KWVNVDDMR

19817
LADSDDPMD

19818
LIASRDEAH

19819
LINMWDFWP

19820
LQHFIDEGA

19821
LQKVCDVYT

19822
LTAKPDCCN

19823
LTVVNDTVF

19824
LWLARDLMC

19825
MASFTDHPQ

19826
MEVAQDVIN

19827
MHCAVDAQC

19828
MHRHMDHEG

19829
MNCQQDCVR

19830
MQHVRDQFN

19831
MSHTTDGDE

19832
MTGYWDVEY

19833
MTIPIDEMA

19834
MTVHRDAWY

19835
MTYKWDCHV

19836
MWNIIDRHN

19837
NAPHADTPS

19838
NCRHCDPNC

19839
NCSYHDASP

19840
NDGKDDTYQ

19841
NEDMWDYNH

19842
NFRTFDHSE

19843
NHLTRDQWG

19844
NMLNADAMY

19845
NNSVCDKMS

19846
NQVGRDVMD

19847
NSDTGDMNP

19848
NTMHTDTME

19849
NTMLTDTGS

19850
QCMHDDWSS

19851
QCMWPDIAG

19852
QEKLQDMYG

19853
QFMDFDFHY

19854
QLMPRDGLC

19855
QQEHADWWR

19856
QSENMDYWD

19857
QTFDADGMH

19858
QTQPHDRHH

19859
QYHHIDYGK

19860
QYQNDDRAS

19861
QYYRADGAK

19862
RQVMTDRIN

19863
RSEFADHWW

19864
SAEYWDMQD

19865
SAIWGDNSM

19866
SDETRDKHD

19867
SFSFCDQRT

19868
SIEDFDEDK

19869
SMEPYDTFF

19870
SNRWQDTYY

19871
SPEIWDMGA

19872
SSSQCDVYE

19873
STLQSDPFE

19874
STMFKDHFA

19875
TARNHDHTA

19876
TCHDVDFTM

19877
TGGLWDWFE

19878
THVGIDNIP

19879
TIRYFDGCG

19880
TLHDNDSAC

19881
TMEVSDQHN

19882
TMGIYDMNE

19883
TMRTSDVYE

19884
TNYQVDWDS

19885
TQMMYDCMQ

19886
TSKSYDGQC

19887
TSSVVDLNA

19888
VCALYDEQS

19889
VCCYIDCVQ

19890
VCSMNDSCF

19891
VDCIIDPQA

19892
VEKGQDMDN

19893
VGDQPDAGP

19894
VGVQDDFMY

19895
VHYNVDQCA

19896
VMVKRDVAR

19897
VNGVTDMDD

19898
VPPSADDPH

19899
VPWVSDWRC

19900
VQLIKDWDQ

19901
VSMRCDADY

19902
VSQPFDPYP

19903
VSRIDDAAC

19904
VTHWCDHSY

19905
WDSCSDSWM

19906
WFRMRDCCG

19907
WLVIKDVNL

19908
WMMQVDGYP

19909
WRKMEDNCM

19910
YEQGVDRTF

19911
YGCKSDEGF

19912
YQIQFDQII

19913
YSEMGDFYG

19914
YSSFTDQNV

19915
YTFYKDCCC

19916
DNILRDVMA

19917
QISVYDTKY

19918
STALMDANM

19919
MSGPNDTEF

19920
GIWVRDTYN

19921
DHLSNDAEM

19922
ATGLTDHQW

19923
QSRCVDNTV

19924
GCAVIEVSA

19925
GKWYNGVEL

19926
GRNNAWVHV

19927
GSMMFGVAQ

19928
GTSWVLVQY

19929
GTYLHVVAD

19930
GWSDMTVVF

19931
HHSIKNVCH

19932
IDFITTVHY

19933
IDHHFVVIE

19934
IHEHHPVIF

19935
ILGVHNVEA

19936
IPHHIFVQD

19937
IYVMPVVER

19938
KDENNPVAP

19939
KEDMTSVGQ

19940
KEVTSQVSY

19941
KFVHTQVEC

19942
KNGCITVQP

19943
KQFNHMVTH

19944
KSVDKYVVD

19945
LCHDKIVHV

19946
LEVVQVVGG

19947
MLPTCMVVN

19948
MMASKFVGR

19949
MRMPDGVNT

19950
MWEATCVRS

19951
NAELYPVEW

19952
NGEQAFVMV

19953
NHCYVEVGG

19954
NPQYGCVME

19955
NYHDYQVCY

19956
PGIFGCVGW

19957
PSNNKCVWC

19953
QKHCQCVSV

19959
QTGTGWVGN

19960
QWGTTNVIT

19961
RADNTNVSW

19962
RINHYHVWV

19963
RSCGEVVET

19964
RWDGQEVWK

19965
SGYYPWVLT

19966
SHNFKQVMR

19967
SLHPVFVNS

19968
SQFWAHVCP

19969
SRVNCFVIV

19970
STILMPVHI

19971
TAVAWCVSS

19972
TAVLREVFM

19973
TCYTCCVWY

19974
THCTKVVDT

19975
TIEVMSVWN

19976
TMRTQCVAL

19977
TPLTCMVCA

19978
TQFHCPVVK

19979
TQSSQTVAY

19980
TSQQGIVSL

19981
TYADMWVGC

19982
TYHTVSVCD

19983
VGSWVEVQN

19984
VKQYWHVEE

19985
VMETHWVWA

19986
VNPTNKVHG

19987
VTELRYVSH

19938
YAEVNPVIG

19989
YHQWPFVDM

19990
YKKAWMVMQ

19991
YNPSEEVDG

19992
YQGTMIVGS

19993
YSCKERVGV

19994
SFSYDVVMN

19995
MFSHGHVMF

19996
DCVLTMVDK

19997
QWFTKGVGE

19998
FPGVFKNWV

19999
GASWYCSWM

20000
GFQWDAHWN

20001
GFVSYHSWD

20002
GIAVRSAWF

20003
GISDDAYWW

20004
GMMLMPSWC

20005
GNAACNAWP

20006
GTPRDEQWY

20007
GYNTANSWQ

20008
HLDWFNMWQ

20009
ICEHMEWED

20010
IDQNFCMWG

20011
IHHGCKMWI

20012
IPMLCRIWM

20013
IREPMPAWA

20014
ISAIAGMWG

20015
ITAATTTWK

20016
IWYVRCWWA

20017
KKKNCILWR

20018
LADSQVFWT

20019
LPDKAAHWS

20020
LQPRTGPWK

20021
LRPHIHAWE

20022
LSENITGWW

20023
LSPWNFGWD

20024
LTVPHCSWQ

20025
MASCKVAWS

20026
MCDWREKWT

20027
MFVCRQFWK

20028
MGAANYSWY

20029
MGSIWLEWH

20030
MIESVSHWD

20031
MIFPIYSWA

20032
MPMCHCGWN

20033
MREVHCTWM

20034
MTAADVCWG

20035
MTLDFSTWG

20036
NGMQICCWY

20037
NHVIYHDWE

20038
NIDHQFHWW

20039
NLKWHKRWT

20040
NMEQDPYWN

20041
NMVNAGGWS

20042
NQMFWNIWG

20043
NSGPFYCWT

20044
PLRWDESWG

20045
QANWTMMWC

20046
QIKEVEYWG

20047
QNRYSELWA

20048
QSYWFQQWA

20049
QTSSCEEWG

20050
QYKQVLDWY

20051
RIDAQQQWF

20052
RSADSWEWT

20053
RTIAWEKWQ

20054
RWDAGRTWF

20055
SEDQFHHWC

20056
SIDNKWPWR

20057
SKFVRYDWG

20058
SLCADPDWP

20059
SMWGPRAWT

20060
SPYNECHWL

20061
SQGITGRWV

20062
STEDCVSWY

20063
TADVEPPWN

20064
TALRQMAWH

20065
TFEPYSWWL

20066
THGCMLTWT

20067
THHQMFEWT

20068
TIASFMQWD

20069
TKAMIWHWG

20070
TMRHWCRWP

20071
TRMMMQEWP

20072
VAVKEAQWH

20073
VCHWSLCWL

20074
VFHSTHWWY

20075
VSEYYWEWN

20076
VTSTILHWQ

20077
WELNAPEWQ

20078
WQCAMVMWI

20079
YPWCYMDWV

20080
YTHDTGLWM

20081
LERTIPIWQ

20082
YWCPSQWWR

20083
KMAGIGCWY

20084
MYDTMGAWC

20085
FTPMNRDRI

20086
GHYWAHEAI

20087
GIFTTNFEI

20088
GMDVNHCGI

20089
GSCITMGSI

20090
HSDLMVCQI

20091
HYCYGEFGI

20092
EACTSLPII

20093
EAENFQGCI

20094
ECHAKATNI

20095
ELKLPWLNI

20096
MNMTYYTEI

20097
MSEAEGHKI

20098
MTDLARTYI

20099
NCPSDQPSI

20100
NEVHYEMAI

20101
NRSKASGDI

20102
NSWFKPEFI

20103
NWYKDVIRI

20104
PAESCNQKI

20105
RYPHEVNGI

20106
SCQAVWCGI

20107
SRETFTKHI

20108
TCADGMDHI

20109
TKECSHEVI

20110
TSRFEMHTI

20111
VTAMFGQEI

20112
VTNPKQEAI

20113
WMGAMVHDI

20114
YGPKFWIAI

20115
YMDMRNLVI

20116
WPNIKHQPI

20117
YSDPKNMSI

Example 17
AAV5 Variants with Tissue Tropism in Skin

This example describes engineered AAV5 variants with tissue tropism in skin that were discovered using the methods and systems described in EXAMPLE 1-EXAMPLE 2.

CNS, liver, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord tissues were analyzed to identify amino acid residues in the AAV5 VP1 581-589 region that drive skin tropism. The results are shown in FIG. 18J which shows the positional frequency of each amino acid in the 581 to 589 region of AAV5 VP1 in skin tissue as compared to all other analyzed tissues, wherein the capsid polypeptide has the sequence of SEQ ID NO: 2. The position (“Xaan”, n=1-9) is denoted on the x-axis and amino acid residues are denoted on the y-axis. The frequency of a particular amino acid residue in a particular position is indicated in the heatmap as intensity. Darker squares indicate a higher frequency of a particular amino acid in a particular position and lighter squares indicate a lower frequency of a particular amino acid in a particular position.

The frequency of a given amino acid residue occurring at a specified position corresponding to position 581 to position 589 of SEQ ID NO: 1 (generalized in SEQ ID NO: 2) in variants identified in skin over the frequency of that given amino acid residue occurring at the specified position in variants identified in all other harvested tissues was analyzed to identify a set of sequence rules for capsids that preferentially target skin tissue. With reference to TABLE 33 below, and SEQ ID NO: 2 (AAV5 VP1), the following amino acids can, thus, be independently mutated, in any combination, at any one or more positions Xaa1-Xaa9, to provide a VP1 capsid with enhanced skin tropism as compared wildtype AAV5 VP1 capsid (SEQ ID NO: 1), where skin tropism here refers to properties that are preferred for skin transduction over properties that are preferred for transduction of all other harvested tissues.

TABLE 33

Skin Tropism Rules

Xaa1 is selected from A, C, K, Q, R, or T

Xaa1 is selected from C, K, or R

Xaa1 is C

Xaa2 is selected from A, C, I, S, T, or V

Xaa2 is selected from A, S, T, or V

Xaa2 is V

Xaa3 is selected from A, C, F, G, M, Q, S, or V

Xaa3 is selected from A, C, F, M, or Q

Xaa3 is C

Xaa4 is selected from C, K, L, P, R, or W

Xaa4 is selected from L, P, or R

Xaa4 is R

Xaa5 is selected from F, H, I, M, V, or Y

Xaa5 is selected from M, V, or Y

Xaa5 is Y

Xaa6 is selected from F, H, I, M, N, Q, or S

Xaa6 is selected from M, N, or Q

Xaa6 is N

Xaa7 is selected from A, H, K, M, N, R, or V

Xaa7 is A, H, K, or R

Xaa7 is K

Xaa8 is selected from A, F, G, H, S, or Y

Xaa8 is selected from A, F, or S

Xaa8 is S

Xaa9 is selected from A, E, G, P, Q, R, or S

Xaa9 is selected from A, Q, or S

Xaa9 is A

TABLE 34 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that were found in skin tissue and comport to one or more of the rules provided in TABLE 33. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 31991-SEQ ID NO: 32990, as disclosed in TABLE 34. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 34

Sequences of the 581 to 589 Region in AAV5

VP1 Capsid Polypeptide that Drive Skin

Tissue Tropism

SEQ
581-589

ID NO
Sequence

31991
CAFAYTIQS

31992
CAFMTHCYA

31993
CAGPWCHHE

31994
CAIVCFDSF

31995
CAMWHQKGM

31996
CANCTWVAE

31997
CASWYATDR

31998
CAVWANWLC

31999
CAWLATDFK

32000
CCDHVQSSW

32001
CCMPMFRNA

32002
CFAAYRMQG

32003
CFAKWWGYE

32004
CFALKINYS

32005
CFCIMVNGY

32006
CFCQRPLWP

32007
CFLRNNAEY

32008
CFRYAEIGP

32009
CGAIHSKRD

32010
CHARGYIMP

32011
CHCMPLVIF

32012
CITHTLKDG

32013
CKVMKMEGE

32014
CLVWERRFS

32015
CMSMNHWWR

32016
CNAKMTDLA

32017
CPNIAPIPG

32018
CQLTVHSFG

32019
CQSSYSMYA

32020
CQTRFYVMP

32021
CRGTIAVGI

32022
CRMFKIDAI

32023
CRSRYEAQC

32024
CSIKCAFGC

32025
CSITHQRGC

32026
CSQKIAYAK

32027
CSSPQYFSM

32028
CSTYYCNGL

32029
CSVKCAFGC

32030
CSYPWDNLG

32031
CTATYTIRD

32032
CTCQYGWTV

32033
CTFYYFKHH

32034
CTKGWMVIG

32035
CTLRTWSGM

32036
CTMFYEQSK

32037
CTMKYQEHK

32038
CTMQALRQA

32039
CTTKFEKCW

32040
CITTFACPT

32041
CTWMAFTAP

32042
CTWPRCDSF

32043
CTWVTHGQQ

32044
CVCGKMGHG

32045
CVDPDDYGY

32046
CVEIMYMNE

32047
CVEYWGQCN

32048
CVGSRPEAA

32049
CVIWDQAQN

32050
CVLTQQFNA

32051
CVMHSTDHH

32052
CVQRKDRGT

32053
CVQRKMQAV

32054
CVQTKINCG

32055
CVQYNKCRS

32056
CVRYYPRPA

32057
CVTMALMDQ

32058
CVTTSNYTM

32059
CVTVTYDGR

32060
CVVHKQKEY

32061
CVVHTMRWC

32062
CVVKHLFGH

32063
CWSYGQTAA

32064
CAAQHCAQV

32065
CAAQHYAQV

32066
CAGIQATGD

32067
CAIHTRSHS

32068
CAIMFSAGP

32069
CAIMLQHMT

32070
CALPVLVAM

32071
CALPYAWSR

32072
CALSMYSQE

32073
CALTYAWSR

32074
CAVYFGKCA

32075
CCRYYVNVE

32076
CEKLTQNVD

32077
CEYFKRHPD

32078
CFHQYHQAH

32079
CFHRYHQAH

32080
CFKIWDLGN

32081
CFMGLSFHW

32082
CFVEPKTSC

32083
CFVWEFNYP

32084
CGDCYGWCQ

32085
CGGFCTALE

32086
CHAQIQHGG

32087
CHAQSIMGP

32088
CHFMINAGR

32089
CHHFATESE

32090
CHHYTNTSQ

32091
CHVPLNNCD

32092
CIKACYDHE

32093
CIKSYNVMM

32094
CILHKHSFP

32095
CIMLNARVG

32096
CIQLNKMGP

32097
CIVTVPQQA

32098
CIWMQHFSL

32099
CKFDMPSFF

32100
CKNFHWMND

32101
CKQQINDAY

32102
CKSHFDQVE

32103
CKYDITKMA

32104
CLELVKVYE

32105
CLNTYHYQG

32106
CLSSSYNSY

32107
CMHLVNMNY

32108
CMQFIRIGS

32109
CMQFIRVGS

32110
CMVHNHKGF

32111
CMYWLTNQE

32112
CNCHIRHTG

32113
CNFQTNVHL

32114
CNTQVYGNY

32115
CNVDRYNRP

32116
CPKPFLIQE

32117
CQELRTQYA

32118
CQEVYAHMS

32119
CQSVHYRAP

32120
CRAAHSVCA

32121
CRATHSVCA

32122
CSGCYMKMP

32123
CSGTMLSQP

32124
CSHHVDRVM

32125
CSHQKLMES

32126
CSMIQNSFQ

32127
CSTDHALVE

32128
CSTMVRSQT

32129
CSTWLNQHM

32130
CTCNLPYAI

32131
CTEMLQRKG

32132
CTGQLMHKE

32133
CTHGFLTGP

32134
CTHKVNSWI

32135
CTIFNTVDQ

32136
CTIRANWHF

32137
CTMLANHYQ

32138
CTPGFWSER

32139
CTQFTKHQE

32140
CYTAWQGAW

32141
CTIVKMWHT

32142
CTTYRNLLS

32143
CTYFSMMDR

32144
CTYMQMAYG

32145
CVFQCYRSP

32146
CVFVQQNFQ

32147
CVHAASLMP

32148
CVHLGHWQW

32149
CVLPVLVAM

32150
CVLWQVHSE

32151
CVNQHSLAM

32152
CVPISAAPV

32153
CVRLIDWCC

32154
CVVMKFSEH

32155
CVYWQQNHE

32156
CWGPKFVDF

32157
CWMPQQVAT

32158
CYPVYKDWS

32159
CYTIKQMDI

32160
CANHLNQTI

32161
CWAVISMSS

32162
CTCWYNRGD

32163
CTHPVHDNE

32164
CYQVTIQHN

32165
CTLMHSHTR

32166
CIIGRQEWL

32167
CSRNRCWSG

32168
CSIQINNCR

32169
CGHRCEMNM

32170
CNSYNKPWM

32171
CMHLSCDWT

32172
CYAEEPVWP

32173
CNSWSTGSL

32174
CMNHRWSTT

32175
CWHFFGADR

32176
CARLEGRGN

32177
CMEAQGECY

32178
CTKPIPCWM

32179
CDCLHHCCM

32180
CIWSSPDQP

32181
CADPFSIRG

32182
CSMPQQCME

32183
CTMCHDCSP

32184
CTEISWGGV

32185
CVGYKSNTP

32186
CSESYRGPD

32187
CWECCTDSG

32188
EVMPVNRKH

32189
EVTRFSEPG

32190
GVAYEEDYY

32191
HVFSHWNCN

32192
IVADLNMFY

32193
IVGMQMTGE

32194
IVNSFQKMS

32195
IVTIVSRTA

32196
KVEQYMQCH

32197
KVTMESFDL

32198
LVDHMSTGC

32199
LVGPTDQLP

32200
MVGQLMKES

32201
NVFYVGEWP

32202
NVMLGAGTI

32203
NVQKQQTDA

32204
NVQRLCCLQ

32205
PVSLFGLIH

32206
QVCAMVQCR

32207
SVKHKEMVG

32208
SVNYCPRGP

32209
SVTCLYTSF

32210
TVDRCFQYA

32211
TVFYCTKMH

32212
TVGIEWLNW

32213
TVGSWAPGS

32214
TVMRALVYN

32215
VVVDLWWHT

32216
WVGCTNHGQ

32217
YVVDLKNFG

32218
AVETEFRER

32219
AVNCTIYGP

32220
AVNTQDDLF

32221
AVRNVEVSR

32222
AVSTWAMQS

32223
DVCKVMTRV

32224
DVGSWLNKC

32225
DVNHTKEEN

32226
DVRVDMNHA

32227
DVYNNVKLG

32228
EVCRIERYM

32229
EVDIVTNNY

32230
EVIFMNCPL

32231
EVNEEEVTY

32232
EVWWEESKS

32233
EVYPCMSYG

32234
FVHCTNALF

32235
FVTCNQYAP

32236
GVFYTMASY

32237
GVKCINSPV

32238
GVMNHIAWT

32239
HVCMLEFGT

32240
HVMQGAPTD

32241
HVVMKHGES

32242
IVAKISW5E

32243
IVCPAGQYH

32244
IVEPSGHFL

32245
IVVSMDQWL

32246
LVGPEGAWN

32247
LVMQLYCAG

32248
LVMRMCADG

32249
LVTQDMKRY

32250
MVTQPCAWQ

32251
NVEREQSLG

32252
NVFIATAQT

32253
NVGIFWYCG

32254
PVAWCDKLG

32255
QVAYAFEAN

32256
QVHYMMCYE

32257
QVLSQWFDN

32258
QVVQKAMAE

32259
SVMTTFCHN

32260
TVGITMLPM

32261
TVHEIMENN

32262
VVMPKESNE

32263
YVCGSGMEQ

32264
YVDKRSTME

32265
AVETGESAM

32266
AVATRAMYS

32267
AVSCGQQRE

32268
DVAIMQGKG

32269
DVLMDKRHC

32270
DVNEPIQCQ

32271
DVNLDPDVC

32272
DVYKSPCTT

32273
EVGINSEGL

32274
EVGLCQSGR

32275
EVLKSFMCQ

32276
EVMFMKKHL

32277
EVQVIKAQE

32278
EVSHRVSAA

32279
EVTQLSCQV

32280
EVVCSMYYG

32281
FVASVPDMS

32282
FVDFNHKQW

32283
FVDTYEAPG

32284
FVEPYTDHQ

32285
FVTMLHWQS

32286
GVAVIRKMQ

32287
GVEEFNAQD

32288
GVFPNMFAP

32289
GVGMKCYSH

32290
GVLPVQQGF

32291
GVQPWESCT

32292
GVSAYCTSA

32293
GVWIMRSKR

32294
GVYYYTKQP

32295
HVAFQWSGQ

32296
HVALQNDWG

32297
IVAHQCQYC

32298
IVAKYGMTS

32299
IVDQRSVWV

32300
IVQPCQ5NW

32301
IVRSALTNT

32302
KVCFKEDAF

32303
LVNEMWYHS

32304
LVQHSVVWM

32305
LVTGMMSMG

32306
MVAQHGISL

32307
MVAQYGISL

32308
MVEHNRLAN

32309
MVGMLYKTK

32310
MVVYRENMG

32311
MVWEVKFND

32312
MVWEVKFNG

32313
NVCAMPMTH

32314
NVCTMPMTH

32315
NVMPSPMDR

32316
NVQAYERAN

32317
NVQRHHIFQ

32318
NVVQSFTAK

32319
PVQHIYWGL

32320
QVEVDHCWA

32321
QVGVCDQCQ

32322
QVMENTELK

32323
QVMQNQCYT

32324
QVTMQPYSK

32325
QVTRDGKGG

32326
QVVQYLTNH

32327
RVDVTNHRY

32328
RVEPTTVPD

32329
RVQSQWDGC

32330
SVCPTRYAD

32331
SVVCTTEFS

32332
SVWASTVFH

32333
TVARGPVPG

32334
TVGNSKMQN

32335
TVHWDITGC

32336
TVMSAFDWG

32337
TVTTKVRDS

32338
VVCMHHCDM

32339
VVGQKMTQT

32340
VVQQIRDFH

32341
WVEWPQMGS

32342
WVVCRCAWE

32343
YVPDRDGCD

32344
YVQLKQEWY

32345
YVVGMGKFC

32346
PVLHIYWGL

32347
YVHPWFGVS

32348
KVNACGNLT

32349
PVNLAQRDK

32350
EVAGCATAV

32351
SVKLAIDQK

32352
LVGKYIDSM

32353
DVMQNQCKS

32354
HVVPCCPFH

32355
DVISRNGRC

32356
LVDCSNTCQ

32357
AVEDYWRYF

32358
GVAQYLHHC

32359
QVEDGYWSY

32360
MVFLEQHGR

32361
LVQKWMRTC

32362
VVISTKHTH

32363
DVDVVALPM

32364
RVTIMFTGT

32365
AVNACSNSH

32366
TVWIQSYSN

32367
PVNESVHIC

32368
SVRTCHQDE

32369
DVTQHTSCT

32370
PVEMKFAVQ

32371
GVWMQGFHP

32372
QVHMQRYYG

32373
KVAHEMMSA

32374
DVCKATTTE

32375
AVHNVKDVH

32376
NVYATPVVC

32377
NVMGTQAKE

32378
DVKAFMGGV

32379
RVNGSTCRH

32380
SVYMMTEGW

32381
HVTKLMTSH

32382
ELCTNWTAN

32383
EPCQSHEFY

32384
ETCPYFDFD

32385
GGCVYWQMD

32386
GTCTDMGIC

32387
HTCHFGGFV

32388
ISCQSMCCE

32389
ITCQKMAMN

32390
KDCFNGECE

32391
KPCMFWMSR

32392
KTCAVNVYL

32393
LACQIQWMT

32394
MTCEYRTSM

32395
MWCDYGTGW

32396
QACFIAGQQ

32397
QDCCKQSVA

32398
QGCKQMGSM

32399
SSCYHSTGA

32400
STCTIRGNP

32401
TACTMFECQ

32402
VACWYTKHS

32403
VICQVQGRA

32404
VYCLTHAGP

32405
YICRLWTQC

32406
YTCVMEHMH

32407
AMCPTHWHQ

32408
DRCNFNCSE

32409
DSCPQIFRD

32410
EWCLADGMT

32411
GACKFNGHK

32412
GRCQYFQMA

32413
GWCWYMRMG

32414
IGCKVQSTN

32415
ISCKAHWQG

32416
ITCVDNKMC

32417
KSCEWMLQQ

32418
KTCEWGQDK

32419
LCCPFDTDA

32420
LKCMNHSAW

32421
LKCVFMRAE

32422
LSCLATYQS

32423
MLCEPEGAR

32424
NYCTYHSTH

32425
QSCATIIRE

32426
QSCLQQIQA

32427
RSCTKEDYW

32428
RTCWFMGVG

32429
SGCPQRVAT

32430
SHCWYWYWN

32431
SNCPYHTFY

32432
SNCVTDATA

32433
TNCAMQMYY

32434
TTCRTWEAF

32435
VACRENMLT

32436
VLCHEADGP

32437
VYCPCDGPD

32438
YACHMGHYD

32439
YPCLMEKEY

32440
YSCMVNTHQ

32441
YSCSQAMHG

32442
AACENKVCF

32443
AMCLFETMQ

32444
AQCKLTTVE

32445
ATCWQPAAK

32446
DNCKIMFMC

32447
DPCPNMIWT

32448
DWCTYRAQD

32449
EACYMYHNT

32450
EGCYFWKQV

32451
EICTASPQA

32452
ESCLTGGNT

32453
ETCQKYVAK

32454
FICQIKSAS

32455
HACPKHYCR

32456
HCCFKNGDM

32457
HMCPFMDQA

32458
IACVSWFTC

32459
ILCWHHYSV

32460
ISCRTQSAC

32461
ITCITWVNG

32462
KACDYFMHG

32463
KHCHSMQFE

32464
KSCLYFDAY

32465
MACDYFMHG

32466
MACPKNISG

32467
MACSMNVAL

32468
MMCFNYFDR

32469
NACSHQHLG

32470
NPCPNMIWT

32471
NYCRRTFHH

32472
QACRLTYWY

32473
QACWLTYWY

32474
QMCTLTKAD

32475
RACHKQLWV

32476
RPCWPWDHL

32477
RSCFEMKHI

32478
RTCNLPYAI

32479
RYCSTIYYS

32480
SACPWNFYC

32481
SGCFWRNTS

32482
SHCLYWMKH

32483
SICQTNFAV

32484
SKCGFDVAA

32485
SLCTIHGAQ

32486
TACHISHYD

32487
TACYKNTCE

32488
TACYPHSFG

32489
TICSMGQKD

32490
TICYMSTSS

32491
TYCRSGDVQ

32492
VMCTWQACR

32493
WHCNFHWHL

32494
WYCLTQRSD

32495
WYCMCIIRE

32496
YACATQQHL

32497
YACPKHYCR

32498
YHCKFNWLG

32499
YHCQNYAAS

32500
YKCAHDPTG

32501
IYCWMPMSP

32502
EHCNHDQGV

32503
KACMSHQGR

32504
MSCPAKIVV

32505
QNCIMLASN

32506
GICLFKTHK

32507
KPCNPVGQE

32508
RLCDTHKCL

32509
EMCGGGLSC

32510
EACMPWAHH

32511
IMCHARMMA

32512
AQCHDMWGH

32513
ESCHLQHGE

32514
WACIIYMWN

32515
GRPRTPLGN

32516
GTIRKEKYY

32517
IPRRLQLPI

32518
ITQRDNIAM

32519
IYERHHCQA

32520
KDNRRIYKQ

32521
MALRSPFNL

32522
MCDRAIRSD

32523
QMYRLAYSR

32524
RLSRAHLPQ

32525
SIERHCRME

32526
YDWRTMAEV

32527
YSERYINAY

32528
AASRSHQSE

32529
DGVRMHKDW

32530
ETKRSDHEV

32531
LCARGHKSG

32532
MCLREIQMQ

32533
MPQRYNRET

32534
NCARGWMII

32535
TGERTHMCL

32536
VSRRSDWSE

32537
YFPRPFKMT

32538
YMDRMSECY

32539
AFDRHSDLA

32540
AGGRIGDSQ

32541
DFVRTNIGV

32542
EFVRNPCFE

32543
ESARYIDFT

32544
FAQRTTDHR

32545
FHDRPFTFT

32546
FQDREQFWY

32547
GWMRGEYNV

32548
ICVRWHCGP

32549
IRVRWHCGP

32550
KCQRMDTWK

32551
KGDRFMVDQ

32552
LAARIGEVE

32553
LIARYGKQW

32554
LMARIQNIT

32555
MADREMAAR

32556
MCIRSGFIV

32557
MEDRLAHYH

32558
NCIRNWQHA

32559
NGIRIFLFG

32560
PMSRQLYSD

32561
QGVRILTAA

32562
QWMRWLIVT

32563
TADRWCDAS

32564
TCTRKDDDM

32565
TMARFPCIS

32566
VSERAHCGE

32567
WEHRFSASM

32568
WGIRNMELR

32569
YADRFACYQ

32570
YANRNFSPE

32571
YENRGREHN

32572
AFDRHSDWA

32573
MEVRECGWD

32574
MGQRYDIQG

32575
QAVRRWWFF

32576
HHLRNVHHC

32577
KATRDPKFP

32578
NSTRNTSYY

32579
MTERYYWCQ

32580
MTQRVVHWG

32581
NNDRHQHLK

32582
NILRYLICD

32583
MPTRMQKGA

32584
DIGRFTAYY

32585
EAWRNSSSG

32586
AQARTIGLG

32587
TKGRNDHME

32588
GQFRSGAYR

32589
LNARTMRSA

32590
MMKRNRFIC

32591
EYSRMNKAS

32592
SYARKVYDC

32593
ICIRGARAD

32594
ERMCYEDSY

32595
ESGFYATWN

32596
FGGHYWPCP

32597
GHGSYLAED

32598
GNQPYGDNQ

32599
GQVGYQMHG

32600
GYHMYQKTS

32601
ITMSYAHMC

32602
IWRDYDQCY

32603
QKRDYQFMP

32604
QMNSYPMYE

32605
RADAYTQIA

32606
SCELYMARD

32607
ACSCYMHTG

32608
DISMYMFQD

32609
DMNMYLGSQ

32610
ECIWYDNYF

32611
GFYWYEMAL

32612
HYAAYEQAM

32613
IKMLYHWKD

32614
ISYDYAQAP

32615
ITMPYAHMC

32616
KTPWYTWHD

32617
MASLYPFGQ

32618
MFTPYRMQA

32619
NGASYNQIE

32620
QEMDYRMST

32621
QHQHYELHQ

32622
QWSMYNMYD

32623
RLTCYRMHK

32624
SAAFYMDVD

32625
SGLEYMGQQ

32626
SIQMYRYQA

32627
TPLWYHRTE

32628
WCGCYHIFN

32629
WWFMYWADI

32630
YPHKYWQMK

32631
YRHEYAHYS

32632
TAVLYETEF

32633
ADHVYGTEQ

32634
AINEYWDVP

32635
ASDGYFMEL

32636
ASGCYATMW

32637
DIKTYMGLQ

32638
DKGGYGCHE

32639
DLVNYGNHG

32640
DYQWYQVAR

32641
EMRFYESGN

32642
EQLQYEPSH

32643
GILPYVKFQ

32644
GSTHYTDAN

32645
ICISYEGGT

32646
ICQVYSKTG

32647
IPMQYAHFK

32648
IPVQYAHFK

32649
IQDDYQLMT

32650
LADLYDLSH

32651
LEQFYEGNI

32652
LGGDYFRSD

32653
LKKWYETHG

32654
MAEPYGLAT

32655
MCDDYETSQ

32656
MTKPYMTQC

32657
MYLGYDMDC

32658
NQSAYRVAE

32659
QHEGYVRNG

32660
QQPSYMVSC

32661
QTPAYCHHQ

32662
QYQWYAKIE

32663
SAHWYCHRT

32664
THYTYAVAQ

32665
TNGHYHCKY

32566
TTTIYRRNG

32667
TTTIYSRNG

32668
TYAKYALRG

32669
VLNTYPCNH

32670
YIRMYDIAS

32671
YTMCYSHQS

32672
YTQNYVFPP

32673
SIDTYDHRN

32674
EYSPYQVME

32675
QHFWYEMES

32676
QIQLYQNDE

32677
MCFPYSCAA

32678
QSEHYVLPC

32679
RNLNYNTAV

32680
ATNKYANAI

32681
LLAKYMVHT

32682
ITEQYDAFA

32683
FRLDYDYKN

32684
LPTQYQQQN

32685
GQNGYGQCM

32686
HSDWYMKYP

32687
NMPAYQIQC

32688
YISEYVMMV

32689
NGNHYNAMR

32690
DPGWYGLAP

32691
VQMQYERLF

32692
AYHCYMSWM

32693
GDVSYDAEA

32694
MMLGYMEQD

32695
ESNKWNPYG

32696
GIHAHNSTY

32697
KFASANGPL

32698
LCKTENGSD

32699
LGNMENPPP

32700
LRAGNNITC

32701
MFQLPNHFC

32702
NATQKNGMY

32703
NPKPNNYQY

32704
SSDAENRCG

32705
DAGCSNDNG

32706
GCLYANIAQ

32707
GTATTNFDQ

32708
LLFPRNVGG

32709
MHRCRNTDQ

32710
QAMTNNVRC

32711
TLDVQNVQN

32712
TSGYCNIGA

32713
VAAQVNQSQ

32714
VSNWENVAG

32715
AASGLNAFK

32716
AKSCFNGFE

32717
ASNIMNISF

32718
ASQWLNDSF

32719
AWVMNNWWA

32720
DKSEENRQE

32721
ETSCVNLFN

32722
FKESRNQGA

32723
GAFYINNTF

32724
GLTLANIGA

32725
HEPLTNQVS

32726
HPVDTNAGA

32727
ITSLCNEFH

32728
ITSLWNEFH

32729
MHAERNANM

32730
MTDQFNKMQ

32731
MYAERNANM

32732
MYVWRNDCA

32733
NFRTINDNM

32734
NIDIHNHEE

32735
PLKGTNQIF

32736
PMVIFNTDY

32737
QCDYRNMHF

32738
QGTHHNHVQ

32739
QKVPVNDHQ

32740
QMLNHNHHA

32741
QRMITNWGC

32742
QTDIRNVSG

32743
QWQTTNGGA

32744
RADQSNPCQ

32745
RHFMINAGR

32746
RTECCNVAA

32747
RTSQENKWC

32748
SAVEGNRHN

32749
SCFEVNQAS

32750
SLRNRNADY

32751
SMVIFNTDY

32752
SWICCNAST

32753
SYVHWNVWS

32754
TAYIHNSEQ

32755
THNEVNYAQ

32756
TTDQFNKMQ

32757
WSHTANKYP

32758
YILSMNGGE

32759
YQVKFNVMC

32760
YRPPCNRKD

32761
YSTAHNEMP

32762
DHWQFNTHM

32763
EAMVINTES

32764
WHEQSNHIW

32765
FTSNRNWQA

32766
RIVDGNACG

32767
QPQSRNGAR

32768
KISGLNDAS

32769
QESMCNYWQ

32770
TRETMNEWH

32771
GAMLNNNAS

32772
RMTPNNCQR

32773
KIAAWNDFL

32774
KTGATNLAW

32775
SQAHQNRTC

32776
VHHKTNRSY

32777
STVGQNHCG

32778
KMTNSNENS

32779
LPSHCNLGS

32780
TPQPMNVAT

32781
KWTPREKDY

32782
MSQKGFKAG

32783
RKWMKAKAT

32784
SGNPTMKNW

32785
SSVCFPKAG

32786
TWTMNEKDS

32787
AHKPAKKWL

32788
AIWQPSKMD

32789
ALYYPEKYA

32790
ETEPLPKQD

32791
IMTPHWKAC

32792
ITTEFMKAC

32793
MIGISMKGC

32794
MSSALEKCG

32795
RQIEVTKWG

32796
TERKMHKAA

32797
TRASVMKYQ

32798
AEALHIKAY

32799
ANLKHAKFG

32800
DFFYMDKLS

32801
DIIWAQKAA

32802
GFNPVIKYS

32803
GLEWHTKDW

32804
HQVTIFKDR

32805
ICWLITKTE

32806
KMYSSAKSY

32807
LKHPTWKEE

32808
LPGYVEKHQ

32809
MQNACFKDP

32810
NIYETLKRI

32811
QKLNIQKQS

32812
QLMMEEKVS

32813
RASFNDKAG

32814
RCVQPQKEG

32815
SAEHSSKWK

32816
SLGCFHKWE

32817
SQWMIWKCR

32818
TFLCTLKEE

32819
TFLYTLKEE

32820
WSAKSTKEQ

32821
YYSPMVKAH

32822
ETHQRCKMS

32823
RTDVMMKVM

32824
SCSFCPKDC

32825
ATVHMPKHE

32826
DWAWTMKIA

32827
PDLCQDKLN

32828
LFWQFGKQG

32829
PRWQSEKCQ

32830
TFTYEAKFW

32831
NHDPSIKDQ

32832
RKWMNAKAT

32833
IKTTMVKED

32834
FEEFITISR

32835
GGIFEQSSS

32836
GSFWFTDSK

32837
IAGPAWFSM

32838
KQHYAYLSP

32839
KSTIEPFSI

32840
MSAECLHSD

32841
NCIDMWWSS

32842
PKSEDITSD

32843
QHNHMWSSW

32844
QTMKFTSSW

32845
RHAPTDISP

32846
THAVLIRSK

32847
TQESSRVSS

32848
TSLKIYHSP

32849
VEPISSQSY

32850
WLMKWLESD

32851
DCIDMWWSS

32852
DFYTVSMSC

32853
MIKFIEGSQ

32854
KTGLPDISA

32855
LGTFTCSSP

32856
LSNTCCMSN

32857
QWQLSWCSC

32858
QWTQWAMSC

32859
RCWHIPNSS

32860
SHGWFHQSQ

32861
TGAQAANSC

32862
TWSYNVTSF

32863
YIVIRCVSN

32864
ALRVGWTSE

32865
DKQAMSHSQ

32866
DTVYNVLSW

32867
EATDNLTSN

32868
EDESLHMSW

32869
EDESRHMSW

32870
EHVNVTYSP

32871
ESKSEQPSA

32872
FLEAWANSM

32873
FRDGMIVSC

32874
GIGQPEDSP

32875
ICATILRSA

32876
KSGISEMSE

32877
LGRDCSRSH

32878
LNMATSLSR

32879
LSKVQQGSY

32880
LYQDLLYSE

32881
NIVESTESP

32882
NNPVLVASC

32883
NNTYVMSSG

32884
QAEYFDASA

32885
RSMWPPSSG

32886
TAHCFTV5P

32887
TASLLWYSN

32888
TEEKCYISP

32889
TQLPTMQSL

32890
TSSPHGASG

32891
VDRFICASW

32892
VDRFICASW

32893
DHDFAQFSG

32894
NNGPIWISS

32895
PDAMWMASS

32896
SDRMVSISA

32897
AAMSSSHSY

32898
VHSFHVESY

32899
LRVSQSFSN

32900
ATMEKCRSI

32901
LLFPLMSSS

32902
NTECRQNSD

32903
YGENISPSP

32904
TWQSMVNSE

32905
TSQYSSVST

32906
GAQSMQFSC

32907
KTYSAMYSP

32908
ENTMWQSSQ

32909
KLKESMESA

32910
QWMHCQLSR

32911
MQQHMKYSL

32912
ECRMAVQSC

32913
GAANRIQSW

32914
QRNEMSASR

32915
DMTLQSVSS

32916
ENGAISEPA

32917
HGNLMYTGA

32918
ISVCKQMIA

32919
IWQYKDSDA

32920
LSAFMGRNA

32921
MTNIAWQKA

32922
QSAFMGRNA

32923
QSSLEAGAA

32924
SEKFIIRTA

32925
SIGFWMRQA

32926
TGQWTFATA

32927
WEFSLADVA

32928
AIVKMGNPA

32929
ATNDKVHCA

32930
DASESHFEA

32931
DQHEVQNVA

32932
EWLPCGDLA

32933
FGRLHQWLA

32934
GCHSFCEKA

32935
HCSQMDTYA

32936
KAESFTYNA

32937
LITTGEFLA

32938
MSNSIRWMA

32939
NAELSCYGA

32940
NFAYTMLNA

32941
QNSHVIHEA

32942
SGTYLYDNA

32943
TQTKMMDMA

32944
YMTEWCSWA

32945
EIVQPQCYA

32946
AGGQKPAFA

32947
AISCGKWYA

32948
ALAWMEFCA

32949
FFGTPYHMA

32950
GFELVKAYA

32951
HTMNGVLYA

32952
IDSWQPCDA

32953
KCALIQDIA

32954
KHAQMSSDA

32955
KKGDSTREA

32956
LMMVILDTA

32957
MAANCEVFA

32958
MAEYTDAAA

32959
MWNYGHNYA

32960
MYGQNWNVA

32961
NCIHNWQHA

32962
NHFLASVLA

32963
NSKIVDYCA

32964
QCWFRGDYA

32965
QCWLRGDYA

32966
RATCGISMA

32967
RKHGLLFMA

32968
RMAHTFWDA

32969
SSFPMWWIA

32970
SSGLNWRDA

32971
STGSHGVCA

32972
WTKWECCVA

32973
KMLPTCSDA

32974
KPNPFVAWA

32975
FGQFCCQVA

32976
MPEKDGCPA

32977
DCDWQSAMA

32978
ALQFVSSTA

32979
AWDANLAFA

32980
QKGMMLDCA

32981
VFGSTRSAA

32982
RGGWCMRKA

32983
INENMQWLA

32984
PPMPVSHFA

32985
KRTIEHPCA

32986
RQMCMAHHA

32987
TSEPHCSYA

32988
ICNMDFRCA

32989
AHAWLQHWA

32990
HIHMSDRGA

Example 18
AAV5 Variants with Tissue Tropism in Bone Marrow

This example describes engineered AAV5 variants with tissue tropism in bone marrow that were discovered using the methods and systems described in EXAMPLE 1-EXAMPLE 2.

CNS, liver, skeletal muscle, heart, lung, spleen, lymph node, bone marrow, mammary gland, skin, adrenal gland, thyroid, colon, sciatic nerve, and spinal cord tissues were analyzed to identify amino acid residues in the AAV5 VP1 581-589 region that drive bone marrow tropism. The results are shown in FIG. 18H which shows the positional frequency of each amino acid in the 581 to 589 region of AAV5 VP1 in bone marrow tissue as compared to all other analyzed tissues, wherein the capsid polypeptide has the sequence of SEQ ID NO: 2. The position (“Xaan”, n=1-9) is denoted on the x-axis and amino acid residues are denoted on the y-axis. The frequency of a particular amino acid residue in a particular position is indicated in the heatmap as intensity. Darker squares indicate a higher frequency of a particular amino acid in a particular position and lighter squares indicate a lower frequency of a particular amino acid in a particular position.

The frequency of a given amino acid residue occurring at a specified position corresponding to position 581 to position 589 of SEQ ID NO: 1 (generalized in SEQ ID NO: 2) in variants identified in bone marrow over the frequency of that given amino acid residue occurring at the specified position in variants identified in all other harvested tissues was analyzed to identify a set of sequence rules for capsids that preferentially target bone marrow tissue. With reference to TABLE 35 below, and SEQ ID NO: 2 (AAV5 VP1), the following amino acids can, thus, be independently mutated, in any combination, at any one or more positions Xaa1-Xaa9, to provide a VP1 capsid with enhanced bone marrow tropism as compared wildtype AAV5 VP1 capsid (SEQ ID NO: 1), where bone marrow tropism here refers to properties that are preferred for bone marrow transduction over properties that are preferred for transduction of all other harvested tissues.

TABLE 35

Bone Marrow Tropism Rules

Xaa1 is selected from A, E, G, Q, S, or T

Xaa1 is selected from A, E, or T

Xaa1 is E

Xaa2 is selected from A, I, Q, S, T, V, or Y

Xaa2 is selected from A, S, T

Xaa2 is A

Xaa3 is selected from A, G, I, M, Q, S, or T

Xaa3 is selected from A, Q, or T

Xaa3 is Q

Xaa4 is selected from A, E, P, Q, T, or V

Xaa4 is selected from A, P, or Q

Xaa4 is Q

Xaa5 is selected from F, I, L, M, Q, V, or Y

Xaa5 is selected from F, V, or Y

Xaa5 is V

Xaa6 is selected from F, I, N, Q, S, or V

Xaa6 is selected from I, N, Q, or S

Xaa6 is S

Xaa7 is selected from A, C, M, S, or V

Xaa7 is A, C, or V

Xaa7 is C

Xaa8 is selected front A, C, D, G, M, S, or Y

Xaa8 is selected from A, M, S, or Y

Xaa8 is M

Xaa9 is selected from D, E, G, L, P, S, or Y

Xaa9 is selected from D, E, or P

Xaa9 is P

TABLE 36 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that were found in bone marrow tissue and comport to one or more of the rules provided in TABLE 35. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 4118-SEQ ID NO: 5117, as disclosed in TABLE 36. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 36

Sequences of the 581 to 589 Region in AAV5

VP1 Capsid Polypeptide that Drive Bone

Marrow Tissue Tropism

SEQ
581-589

ID NO
sequence

4118
EAEDEIQKA

4119
EASWPMDWD

4120
ECIHDQCSS

4121
EDIDLKGAQ

4122
EEGPQRMNH

4123
EGCCYWSHA

4124
EHWPMAMFN

4125
EHWQKDYNH

4126
EIYWMMSNV

4127
EKKSCQCWH

4128
ELDSWWRIA

4129
ELKESMESA

4130
EMAQTSETP

4131
ENGHKFNEE

4132
ENQTRMTSW

4133
EPTSEVHHD

4134
EQFNHTWHG

4135
EQHDWYQKY

4136
EQIALHCAW

4137
ERFAFVADH

4138
ESSRYRMMM

4139
ETHEFEKLC

4140
ETRTTAWSA

4141
EVITWGHMQ

4142
EVKDKPSIF

4143
EVWNAHEEG

4144
EWESSSILS

4145
EYTHESHWA

4146
EIEGMYWPL

4147
ETAWKWEWS

4148
EQLLSLCPS

4149
EKNQSTCET

4150
EYARGELEN

4151
EHPEWKCMY

4152
EMEHYPCYN

4153
EYSMSEQTN

4154
EKGGQMCAN

4155
ECDRLKNER

4156
EWCPGEPDG

4157
EEPRILYAG

4158
EHKLFMKPE

4159
EKIMVKRAF

4160
EAKEDWGVR

4161
EDKYMTKRH

4162
EGDLSGFQC

4163
EGKPRCFDW

4164
EHFQYQNHI

4165
EHKYFNSGH

4166
EHWYNRKGL

4167
EIWMWTQGC

4168
EKTAGMQTM

4169
ELPWQRWEN

4170
ELVPMKLRS

4171
ENHEEKMAE

4172
ENLSVNNLF

4173
ENMDDHYDT

4174
ENWCERYAK

4175
EQHLKPFEA

4176
EQKHCWSRD

4177
ERMLVHMRS

4178
EVQWNNPKP

4179
HAAGNEFCY

4180
HACDTQEHP

4181
HAENGAVWR

4182
HAISRQQVE

4183
HALTTDYYN

4184
HANQEGWCL

4185
HATSSTGDD

4186
HAVMHSAVS

4187
HAYNTFQFG

4188
IADHTEADP

4189
IADNIMQAC

4190
IAGMTADDG

4191
IAHDGQADS

4192
IAHEKKVSL

4193
IAELLHNFD

4194
IANSKMWEA

4195
IAYCTDAYE

4196
KACQNSNFW

4197
KACTDESGF

4198
KARDLEYEQ

4199
KAYDTMKCA

4200
KAYSEELGY

4201
LAANSVLYF

4202
LADWYSARE

4203
LAEIMKCPP

4204
LAFHLNGTS

4205
LAGCMGLQE

4206
LAMAAHEEA

4207
LAMNQSKAI

4208
LANLHEWEG

4209
LASITTPPS

4210
MAANNECLV

4211
MAGMMEEGS

4212
MAGPCSSDT

4213
MAIERAEVN

4214
MAERWHEVY

4215
MAEYKCAVN

4216
MAKYPVNWG

4217
MANDKHDPQ

4218
MARQEGWCK

4219
MASGSVEKM

4220
MASHIWNQW

4221
MAVGNLSEA

4222
NAALHSADW

4223
NAANQDHTW

4224
NAEAAWMTF

4225
NAELMDAHG

4226
NAFCGNHYL

4227
NAFNYRLVE

4228
NAGHYMEYK

4229
NAGPCWSEE

4230
NAMPREQFP

4231
NAMSNAAML

4232
NANPHHAVP

4233
NANQSCANN

4234
NASHDMHDL

4235
NAVQTLQLY

4236
NAWFPMTNT

4237
PACFRVMFV

4238
PAMNFFIEF

4239
PAPKNQSTD

4240
QAACTHPWS

4241
QADQMPTQE

4242
QAEGGPNMP

4243
QAKTSYGGE

4244
QALVHGAPL

4245
QAMMAQRAG

4246
QASQYTVKY

4247
QATHWPMWR

4248
RACHPPHLK

4249
RAEPMIYNF

4250
RAEQAPVCN

4251
RAQDTEMML

4252
RAQLYGMDS

4253
SAATFKGSP

4254
SAAYCSYLR

4255
SAAYFPARG

4256
SACGAQYPP

4257
SAEAVVRLW

4258
SAEVQATTY

4259
SAGLRVCDR

4260
SAEFCLKKH

4261
SAEEVDCHY

4262
SALDVQRPS

4263
SALQISMLY

4264
SANWVQRDM

4265
SAPSMWWAS

4266
SAQNHDCWL

4267
SARPFEFME

4268
SASNEFDGL

4269
SAVAHMSET

4270
TACIAYVDN

4271
TADFNVRVE

4272
TAFWMKGPQ

4273
TAGLSHFKS

4274
TAGLVTGLG

4275
TAHCHITYL

4276
TAHGCPTQH

4277
TAKTHLFPW

4278
TALHEDWEP

4279
TALMSSLQA

4280
TANLHTIEG

4281
TANQAYSTE

4282
TAPTEDYGG

4283
TATGMKHVY

4284
TAVHLLDPD

4285
TAVRYEVCG

4286
TAWKQNVSD

4287
TAYYEEFWR

4288
VAAHEGRKS

4289
VADRQNEHK

4290
VAFSLMEMD

4291
VAGFVSPTG

4292
VAGNGIPMP

4293
VAGWATPQL

4294
VARDHRAGT

4295
VASLGLYCM

4296
WAPVHWMKD

4297
WAYNSVASE

4298
YAASHMEMP

4299
YAHEVYRHG

4300
YAMDCMGGV

4301
YAQTDGCLE

4302
YATQNTFSY

4303
YAVEYGWNQ

4304
AAHNYEGVA

4305
CATHMERTR

4306
DADDQAVSG

4307
DADFCPRMP

4308
DAGLDSAGY

4309
DAHALQNVI

4310
DATGKFNFN

4311
FADKWANQF

4312
FANLFANTW

4313
FANLEINTW

4314
GANNQNSCE

4315
GANQHDVGH

4316
GAQCLFFQN

4317
HAIHDCYPP

4318
HAILYFYFP

4319
LAIQPCDER

4320
MAGDYACAM

4321
MASSVVHSC

4322
MATNREWWG

4323
MAVWAKAAE

4324
NARDPACDN

4325
PADRNNLWQ

4326
PAECISDSQ

4327
PATVKPMYW

4328
QALNRVQFN

4329
QATGMLYEE

4330
QATVTKEYW

4331
RAIHDCYPP

4332
SARDGISMH

4333
SAVYIQPDY

4334
TADVVPCHH

4335
VADYLNEFT

4336
VANNGMAVK

4337
VAWSQEFSQ

4338
YAQQFTMPT

4339
QAAGETFYG

4340
AACWTNMQW

4341
HAPMDNAPF

4342
NACDWKNDY

4343
IAMLESWWG

4344
DAGRFTGTS

4345
TAPRIDQQE

4346
VATDSVHES

4347
MAEQMGCQA

4348
DARFDQWHH

4349
PACFDDGEF

4350
LAEPWATSH

4351
PAVGFKDFQ

4352
HANHLMSHE

4353
LAHCDMMMC

4354
GADDHHLMT

4355
YAPKMWAEA

4356
WANTYSENV

4357
RAIYNDMTE

4358
AAMGYAKMS

4359
AAPERCYFE

4360
CAHPEEYHQ

4361
DAHFPGLAQ

4362
DAQCFSFGH

4363
HAVGPVMLE

4364
GIQDEAWVA

4365
GQQSFQQVD

4366
GSQMMQDQA

4367
GVQLTIQQA

4368
GVQPALYED

4369
GYQPDMGMS

4370
HCQIDVLRV

4371
HTQFEPGYH

4372
HVQVDQSKS

4373
HWQHETMAQ

4374
INQLKHSAY

4375
ISQCDTAII

4376
ESQWCANNK

4377
KDQCIKQAP

4378
KEQPNDRTQ

4379
KVQENEVLG

4380
LKQQFAEGM

4381
LVQLNMAGA

4382
LVQSEWGEE

4383
LWQLFTMQQ

4384
LYQQLESSG

4385
MLQSHTCLG

4386
MNQFGNMKQ

4387
MQQPFYRSM

4388
MYQVWCTDT

4389
NCQHNEESP

4390
NDQFRVVDA

4391
NFQRMHWST

4392
NIQGVIDYR

4393
NPQKWNPCI

4394
NQQCFDYYS

4395
NVQEATTDS

4396
NYQPFYVWA

4397
PHQLHENWG

4398
PLQLDHECT

4399
PSQIDHAEQ

4400
PWQYCELGA

4401
QCQSEHRDE

4402
QCQSFGVTQ

4403
QDQLYGRDE

4404
QDQQKQYDW

4405
QHQHLCRTS

4406
QKQFNAQMG

4407
QLQWFVKDS

4408
QNQIEHWCA

4409
QPQFMVGQI

4410
QVQFDRCYL

4411
RFQCEGHLD

4412
RHQLLEWYD

4413
RTQPDGQTR

4414
SDQQWKPTH

4415
SGQPFSYSK

4416
SHQQCVMDP

4417
SKIMMQDHV

4418
SIQTTFNDD

4419
STQKYWNYE

4420
TGQLNTGCV

4421
THQTARQNV

4422
TLQWEPKGV

4423
TNQCICAAP

4424
TNQNLGNDY

4425
TASNIVCCE

4426
TTQWSQLQS

4427
TVQSMTSFA

4428
TVQTPPQYV

4429
TWQSLCDFE

4430
VCQLMKYQP

4431
VDQVFVEDR

4432
VPQTDLDLC

4433
VQQIIGTIG

4434
VSQPCLIVD

4435
VVQWFAQQV

4436
VWQQQFVWL

4437
WDQFCRLMM

4438
WNQIMEYDY

4439
WNQPLMWCF

4440
YCQQTYTEY

4441
YEQKENQSY

4442
YGQWFDMSY

4443
YEQHMHIQR

4444
YEQTKQIMQ

4445
YNQAVCQEP

4446
YPQPFMLMF

4447
YTQLCHHGV

4448
AEQSRRSVG

4449
AKQCLQDYP

4450
CFQQKHNLF

4451
CGQSVSHAG

4452
CQQYTLVQA

4453
DDQFQDSTC

4454
FDQPTPDHP

4455
GEQHQMPMV

4456
IGQHDVQTT

4457
IMQQWPAMF

4458
ISQHTCSKD

4459
KEQPSQMHG

4460
MQQANSDYA

4461
MTQCRDCWL

4462
MTQDFNCDY

4463
QRQQMSHDC

4464
QRQRMSHDC

4465
RHQLLEWDS

4466
RVQHCYGDM

4467
SKQPNADSD

4468
VCQKYDEWG

4469
VTQQYWACD

4470
VTQQYWACN

4471
WDQPNMHTF

4472
WGQHKHTVA

4473
YCQEFEQYP

4474
YDQWECDEH

4475
YDQWERDEH

4476
HDQMMGLER

4477
TNQCIFKTM

4478
CHQCEGQDN

4479
GDQFQDSTC

4480
THQYFASLA

4481
IGQHNVQTT

4482
LKQCQSTMF

4483
ALQQNPSMT

4484
MMQCRDCWL

4485
SCQEWGIQW

4486
ANQFWKIHC

4487
GHIQYMRST

4488
GIIQTGCYK

4489
GIVQGYEGS

4490
GKTQFRKCS

4491
GMYQKCYFA

4492
GNCQQVQMG

4493
GQDQCCREW

4494
GSSQGRARA

4495
GSVQEGYDA

4496
GTRQFEGNC

4497
GTTQDWRVP

4498
GVIQDQGMF

4499
GVVQRATIE

4500
GVYQKQSMQ

4501
HDLQYPMSF

4502
HFHQMIEGG

4503
ICAQANIAR

4504
ICDQPGTTG

4505
IGVQTSFQD

4506
IHLQWQSMM

4507
IIDQKYSWC

4508
INLQGHHWP

4509
ISCQEEISM

4510
ISDQNVAHR

4511
ITLQTANAP

4512
IVLQYKFME

4513
IVMQQWFPF

4514
KCEQIENNP

4515
KEIQFMTMH

4516
KLVQNMKRD

4517
KQDQCMCWY

4518
KYEQLLGCY

4519
KYYQAFSKY

4520
LNWQFVECQ

4521
LNYQIALCR

4522
LPHQYNVVY

4523
LQTQRHWIP

4524
LSVQGLCSY

4525
LTSQSNHWG

4526
LWMQHQHGS

4527
LWSQYGLWV

4528
LYLQNMHCE

4529
MDVQTMNDG

4530
MSFQETTSI

4531
MTVQWFDIN

4532
MWDQTEMFN

4533
MYSQRNCLY

4534
NDRQHHHKC

4535
NNYQKLSFD

4536
NSIQFSSWR

4537
NTAQNEGWM

4538
NTEQMPPLS

4539
NTVQDMGVC

4540
NVLQLDAFL

4541
NWAQAHWYG

4542
NWDQTMSQD

4543
PEYQVERSE

4544
PMIQTDCRW

4545
PPPQIYLMA

4546
PWVQTNIGL

4547
QCFQEGWQD

4548
QDIQLSHHT

4549
QEVQFFATD

4550
QHVQRNYAS

4551
QQCQTGTPG

4552
QSCQAIVED

4553
QTTQQWCMQ

4554
QYSQIHMQK

4555
RGYQHEMMG

4556
RQHQGHYDY

4557
SNEQLDDDR

4558
SNIQDDPKG

4559
SQAQCEMMD

4560
SSDQQGWVT

4561
SSLQCFDME

4562
SSMQMPSGN

4563
SYVQTEWVY

4564
TIA0THWFW

4565
TIKQCSFCL

4566
TISQTQGYM

4567
TQTQINQGR

4568
TSMQSTQGK

4569
TTMQRLHFE

4570
TYAQRHVMG

4571
VDKQMQIYL

4572
VHCQADTAN

4573
VINQNIQCQ

4574
VMKQGNQEE

4575
VPAQQSTFV

4576
VPHQLCHIG

4577
VQVQGLAIM

4578
VTAQAESWF

4579
VTAQQMNLG

4580
VTVQKQEDH

4581
VVIQDIRKD

4582
VVLQQVDPA

4583
WERQYVPFV

4584
WIHQQFAEI

4585
WPCQQQSTW

4586
WSGQNYNGA

4587
YEKQEMNCY

4588
YMNQQNKDE

4589
YSLQTEVND

4590
YVSQSSYNY

4591
YYGQNYMGC

4592
ARAQIQDEW

4593
CVMQERQAN

4594
DLEQAMHAG

4595
DPCQEWDSE

4596
FEAQIVLPG

4597
IHLQWIWAN

4598
KHDQSGIAN

4599
LSGQIDEMS

4600
LYMQAPSEG

4601
NCLQKMQIW

4602
NDLQDKSHQ

4603
NVSQITQDC

4604
PNNQARGQC

4605
PVIQPCDER

4606
RWKQWCWEF

4607
VSYQTQDML

4608
YDLQTIKQL

4609
NITQLQCYC

4610
FHPQTSDND

4611
MEWQSEYIS

4612
DHPQMHFYW

4613
LVAQQDSCE

4614
LWKQWCWEF

4615
LVIQPCDER

4616
KPVQCACQN

4617
LIIQHPNMA

4618
SFLQYHERT

4619
GLPQSADVI

4620
DQVQGAHLF

4621
SGDQHLSEG

4622
LKSQGCEAY

4623
SCCQHDNNA

4624
YHEQCQWHS

4625
NMMQYQWAI

4626
MIPQHARGP

4627
TGRQYYSCC

4628
GNLYVPSWG

4629
GVDLVMMLK

4630
GVMLVGHKY

4631
HDFHVKRRC

4632
HNDWVIMAP

4633
HNVHVTTKW

4634
ICKLVMKED

4635
IEIYVNFCT

4636
IIDYVAHLL

4637
INELVEWCP

4638
ISCSVESLS

4639
KDKEVIVNC

4640
KDMCVHKYT

4641
KEWLVDESQ

4642
KMKMVKEVC

4643
KVEEVNVQQ

4644
KWTEVDGSW

4645
LCEHVQNCM

4646
LDDKVADHW

4647
LDKIVGMWS

4648
LGDNVDCWQ

4649
LMGDVTHMF

4650
LRVWVMYLS

4651
LSCHVQMNS

4652
LTSDVHIHM

4653
LYEAVAQME

4654
MFPLVGMIV

4655
MMTMVCNAL

4656
MPASVGSYT

4657
MWSPVFNWL

4658
NCEWVTKLA

4659
NETGVFRQA

4660
NFCAVEQGV

4661
NFGYVRAVG

4662
NIWGVDATE

4663
NPAPVDQHY

4664
NQNAVECIQ

4665
NRDVVRAWH

4666
NTEHVLVTP

4667
NVFLVPDMD

4668
NVVSVPVKS

4669
PESHVVSVW

4670
PNYVVQWNF

4671
PTTIVGWRD

4672
QCDDVMISE

4673
QCTFVPFET

4674
QGKYVGMIM

4675
QGYYVNSVD

4676
QMKMVNTSK

4677
QWCPVQMQC

4678
QYNVVKCVA

4679
RCDNVEACF

4680
RDGPVVPPP

4681
RIASVQWEE

4682
RPTLVNAHL

4683
RQIPVEDSN

4684
RTDLVAVCG

4685
RWSHVRVLW

4686
SFAHVGLPI

4687
SGHMVVCCR

4688
SNAPVHTCD

4689
SPATVAAGC

4690
SSIGVMGLY

4691
SSLCVNDQW

4692
STFRVGDCI

4693
STTIVGWRD

4694
TCDYVCAQV

4695
THTSVGGQC

4696
TRCDVEGYA

4697
TTYWVREMS

4698
TVMTVAAMQ

4699
TYRLVQQCM

4700
VFESVVITE

4701
VFTPVEKSG

4702
VGDRVQKNS

4703
VGTTVNVRS

4704
VHNMVLAFD

4705
VIGGVISYW

4706
VRELVMWGY

4707
VSGFVFVWR

4708
VTDYVPMNQ

4709
VTEIVCESG

4710
VVGVVIYTR

4711
VVVAVNADG

4712
VWELVSHVQ

4713
YDSGVETIS

4714
YYHGVVDSE

4715
CHAAVTNLY

4716
DLIIVPWSN

4717
LEILVAINE

4718
LMTWVEKGA

4719
LMTWVEKGD

4720
NTYGVAFDT

4721
QYHWVTRII

4722
RPPHVGPAL

4723
RQEFVFDCY

4724
SGKLVMCGY

4725
SSGPVIILR

4726
TCFHVAEMF

4727
PQCHVMADR

4728
CSDLVGTNC

4729
IRMNVVLTP

4730
ISMEVNRGE

4731
ALDRVSMAC

4732
FFEGVNINY

4733
PMYVVPEMC

4734
GKLDISNLC

4735
GLCNPSIVT

4736
GQIYCSRYD

4737
GSNFGSIDK

4738
GSNPFSRHG

4739
GYAKESAKR

4740
HHMRMSTLP

4741
HMDDSSICA

4742
ICREESETP

4743
IGEKISAMP

4744
INEGESTYD

4745
IVNTCSQSW

4746
KRKEASTRW

4747
LFEHSSVAN

4748
LGGLESCPG

4749
LHISISKSC

4750
LISEFSHDK

4751
LLTHISYME

4752
LPGIQSTMR

4753
LVSAYSEKF

4754
MDIDLSSHA

4755
MGAFTSINI

4756
MNVAFSMED

4757
MTPAMSFSP

4758
MTIRESICE

4759
NFDHISAST

4760
NLRYASNWP

4761
NMMSKSLDS

4762
NMSPFSSHS

4763
NMSPMSESY

4764
NQYTASNIC

4765
NTYSWSREW

4766
QTGAASANR

4767
QVECPSVDA

4768
RGVGMSTDN

4769
RVAEMSNTP

4770
SKNCLSFSW

4771
SMVTDSRWT

4772
SSGKDSNAA

4773
STDDESMPG

4774
TECPKSLTS

4775
TGKDESHFY

4776
THFHTSREP

4777
TMEFTSPCS

4778
TNHGFSNWV

4779
TQDWNSTAA

4780
TTHWQSWIQ

4781
TYGMCSTFQ

4782
VCLTHSFSN

4783
VIEMESGDF

4784
VNLVDSIRD

4785
VVNLFSFRS

4786
WFMEPSPQR

4787
WMHEDSSQY

4788
WTFMDSTTD

4789
YCSPFSGDA

4790
YLNKRSVNA

4791
CISMYSEGD

4792
ISGLMSDYQ

4793
IWMLPSVTT

4794
KQHNYSGRA

4795
KSGLQSITR

4796
LSEVFSAQP

4797
LSMVFSKGE

4798
MHFPWSGEA

4799
QVPWESKLC

4800
SKEEDSTML

4801
VEMCYSWNY

4802
YYTIASANV

4803
IPSADSWPM

4804
LPRTSSQIG

4805
QWGFGSDET

4806
AVLLASDQR

4807
DWNWKSCEW

4808
TSSNDSTAN

4809
IHGYHSCYF

4810
HVMLLSPNQ

4811
LSELFSAQR

4812
DIKMFSFHT

4813
GHTGSWCDR

4814
GKYRIACDT

4815
GMADLKCPS

4816
GMVEAECMD

4817
GQICILCSE

4818
GQYFKECYD

4819
GVNILTCEC

4820
GWKYEQCTD

4821
GWTPQLCSS

4822
HCSSMLCQS

4823
HNIPFYCMH

4824
HRLMYDCHK

4825
HSCPLACMN

4826
HTIGYQCHD

4827
HVAEYCCKH

4828
HYDTHQCWF

4829
IEWDHLCSH

4830
IHHGSICCT

4831
ITETRACAA

4832
KEEMMQCEE

4833
KGEPENCST

4834
KQDMARCWW

4835
KTDSINCQQ

4836
LFEPINCYH

4837
LSEVCLCSE

4838
MCDANHCNS

4839
MKGHNDCIS

4840
MMGVRNCHM

4841
MSTRCHCRY

4842
MTYKADCPK

4843
NDDTMHCAY

4844
NHTHITCTE

4845
NMMIMHCHT

4846
NNTTTRCLM

4847
NPLMQCCYL

4848
NRADMICAK

4849
NSAPSMCST

4850
NTEPNNCWC

4851
NTVYDNCGE

4852
NVKHCHCEI

4853
NWTYTFCQQ

4854
PFPMEQCWQ

4855
QCKWGDCAC

4856
QVHRPECRH

4857
RFWLQCCTE

4858
SFENSECTK

4859
SFYERACES

4860
SGFLTVCDR

4861
SHCMLNCWA

4862
SKILFDCTS

4863
SPLITNCEA

4864
SQVRTVCGF

4865
SRNPMMCHP

4866
STKNDICEG

4867
STLPSKCGM

4868
STMGTKCSM

4869
TDREIHCMN

4870
TFFPGHCYQ

4871
TLIEKLCED

4872
TPMRCYCFG

4873
TTDTATCKS

4874
TTHKMTCCP

4875
ITYKQECTI

4876
VDEWYVCMW

4877
VDRHGGCQF

4878
VFSKFVCQL

4879
VGSLYNCSG

4880
VNTLRHCQS

4881
VPTNLNCCQ

4882
VREHYKCGW

4883
YENAMICEM

4884
YTENKECCP

4885
AKNYFWCKP

4886
CISILGCAH

4887
FTEEGACGM

4888
GGMDLICHG

4889
HWVMQMCIG

4890
IMEPLVCDD

4891
KGCMDWCDS

4892
MKYDRPCCT

4893
NNDHFVCYL

4894
NTNMRHCGI

4895
RSIPGGCVA

4896
SFYSHICRE

4897
TCHWIQCFN

4898
TGEAQDCNP

4899
TVPMDHCWA

4900
VMKIKRCEG

4901
VNALFDCCY

4902
VTMNFHCGC

4903
VNDYDQCCM

4904
DYAENWCIE

4905
AWRPPNCEL

4906
NPIKFQCAH

4907
VYANENCAP

4908
PHEDCECLR

4909
TSVMWACPG

4910
GICGTRSMK

4911
GIMLSQDML

4912
GIVADRSME

4913
GLGYGHHMM

4914
GMVFTNYMR

4915
GSELGGVMG

4916
GVLPSYMMM

4917
HSPPCQIMP

4918
HWTTEEQMM

4919
IMCTKARMD

4920
IMIFEQMMH

4921
IQGCQMGME

4922
ISHNNAHMQ

4923
ITTPYGHMY

4924
KDLRSMEMC

4925
KFMGYMVMP

4926
KFMGYTVMP

4927
KHWFDPGMW

4928
KIMEPTFMC

4929
LCGLNTKMY

4930
LFHPDMMMP

4931
LGDDKYIMS

4932
LNANMKEMF

4933
LQTSTINMQ

4934
LSFDFPTMC

4935
MCALNPGMP

4936
MCFMQNDMR

4937
MDWSEGKMG

4938
MHDGGMEMP

4939
MILGEMSMY

4940
MNLNKIMMY

4941
MPYCEHWMI

4942
MSIPFCWMQ

4943
MVKPCGSMF

4944
NFHRWDRMT

4945
NIEWFKTMG

4946
NKEDNHMMM

4947
NKNEMAMMG

4948
NNCHYNIMA

4949
NQFPATEMQ

4950
NTFLCQKMA

4951
NTRFNVDME

4952
PLTLADKMQ

4953
PWYWPFEMY

4954
QGNWTMLMQ

4955
QKECYPNMV

4956
QSGVDFAMA

4957
QSLREENMQ

4958
QVLKGEEMP

4959
RHDHMMNMA

4960
RSVVNEKMF

4961
RVCNNESMF

4962
SCVVTDNMR

4963
SDHVLHEMC

4964
SEREMRHMP

4965
SFASKLAME

4966
SLDHMPMMG

4967
SLTFFQYMD

4968
SPSSHCVMI

4969
STSPYEHME

4970
SVGTRINMA

4971
THSSLASMV

4972
TLCSQNWMM

4973
TNHIQNEMD

4974
TNNEDTQMH

4975
TRHYNCTML

4976
TVVTKGTMP

4977
TWIEMDYME

4978
TYPKRMNMM

4979
VCDPWFAMP

4980
VDAPQPNMP

4981
VMYMRDQMW

4982
VPKWGTGMF

4983
VSAYEQNMK

4984
VTWHPLSMH

4985
VVEPQMRMC

4986
WITLNQKMV

4987
WWCHPMHMM

4988
YDKTYMAMG

4989
YQFNWGNMT

4990
YTKETGGMV

4991
DKIPINGMS

4992
DVEKTMQMN

4993
FHFIRHEMV

4994
FQMAMCSMG

4995
GRFEEGAMY

4996
EGVMLNAMQ

4997
KDRCQYEMA

4998
MCGFETTMP

4999
MHDDYRGMQ

5000
MSRRKEAMP

5001
QRIWTTYMH

5002
RRFEEGAMY

5003
SELFRANMD

5004
VMCNMMQMT

5005
WNAPKVMMA

5006
DKIPINGMP

5007
SFHPSWWMY

5008
NCFDSEDMG

5009
VTTNELGMH

5010
TTFPQVIMN

5011
DENWNHNMF

5012
TSGEMLRML

5013
LIEVLNHMN

5014
LFDRCNNMW

5015
KFHSKERMN

5016
YMNSLWHMR

5017
GIVTCAGMC

5018
GIHICKACP

5019
GNWSKVSKP

5020
GWENLFMTP

5021
HHPTLFHFP

5022
HPDNSQMSP

5023
HQTRSELAP

5024
HSAAPTEGP

5025
HVFSIFHVP

5026
IMRLDARCP

5027
INTGFMNKP

5028
ISHPSVGEP

5029
ITMPRMDNP

5030
KDVEFFHGP

5031
KSEENYNHP

5032
KVDDHAFEP

5033
KVDNEFYVP

5034
KVWPCNPDP

5035
LCAIWTLCP

5036
LFRFDMSPP

5037
LHCSPEIGP

5038
LHESLDDFP

5039
LPCHAIVSP

5040
LVTLNHNKP

5041
LYGDAQGGP

5042
MGGHMHEYP

5043
MMVCGDRSP

5044
MPNMWQEWP

5045
MQEELVHCP

5046
MRGIEVEQP

5047
MYIRREGNP

5048
NCDERPHWP

5049
NEGPIMYSP

5050
NEWNPLPKP

5051
NILCKHKVP

5052
NISGQLHTP

5053
NMSGEYVFP

5054
NQCEIIDFP

5055
NSTNNGWQP

5056
NSVKQNAAP

5057
NTGLSDEPP

5058
NVTTAWNCP

5059
NWHWPPVFP

5060
NYDFYGMIP

5061
NYGEAYNFP

5062
PLIIQQFWP

5063
PSACMAGKP

5064
QFEDMYKSP

5065
QHANFAVQP

5066
QNSPCTAFP

5067
QSCLFMQIP

5068
QSTWAERWP

5069
QTMLYWYLP

5070
QTWKHMLGP

5071
RCIWKCEGP

5072
REDPSALEP

5073
RHDLYVWSP

5074
RYDYAKYHP

5075
SECTHDHFP

5076
SGVPYWAHP

5077
SSIYEPEYP

5078
STSCSPKWP

5079
STTSQHHPP

5080
SVTSQWMPP

5081
SVVCQQDEP

5082
TESDFAIGP

5083
TMTRIVHEP

5084
TNSSGEFPP

5085
TSALELQKP

5086
TWPRGAMHP

5087
TWSSIDMWP

5088
VFLVSGMLP

5089
VGLTMTGVP

5090
VRNEGGEWP

5091
WDCTMYHFP

5092
YPHTPHNCP

5093
YPHTPYNCP

5094
AQPNGNEHP

5095
CISEIAITP

5096
CISITAITP

5097
CVLSWCVRP

5098
DFNDSNWLP

5099
DMMVSGHVP

5100
DQLTWHTGP

5101
DQMDEFLYP

5102
GIDRPHFVP

5103
HNNNHAIPP

5104
LGNWHHNSP

5105
LHYNNERDP

5106
MSEVMDFLP

5107
STAHRPWTP

5108
TELVRQVAP

5109
TITALMVDP

5110
VKLTHRVRP

5111
VQPVQLFCP

5112
YTCCENHWP

5113
YPNDMTFEP

5114
AQPSMDYVP

5115
FKVRKFWDP

5116
QTEEPDDYP

5117
INEGLLYPP

Example 19
AAV5 Variants with Tissue Tropism in CNS as Identified by Machine Learning

This example describes preferred properties of engineered AAV5 variants that display CNS tissue tropism as identified, by machine learning methods.

First, sequencing data from CNS tissues was subject to bioinformatics pre-processing. An overview of the bioinformatics pre-processing steps are as follows. First, raw NGS paired end reads were merged, quality-filtered (>Q30 avg), and base-corrected (fastp). Next, reads <150 nt were removed (readlength). Accurate alignment of reads was ensured using fuzzy matching of constant sequences on both 5′ and 3′ of variant region (align, fuzzy Align). Reads with stop codons were removed (stop). Additional relevant filters were used to select variants for subsequent analysis (e.g., “deduped”-shows number of unique UMI-var pairs detected in a given sample). Finally, filtered variants were moved into the machine learning pipeline of the present disclosure. FIG. 19A shows the results at various steps of bioinformatics pre-processing of the NGS data. Stepwise read count for 48 individual samples through filters is shown. FIG. 19B shows a positional comparison of CNS/non-CNS variant sequences. Position 1-9 refers to the 581-589 positions of AAV5 VP1 (SEQ ID NO: 2).

Additional filters were developed taking into consideration the experimental design. First, as the libraries were designed such that a single amino acid residue is encoded by a single codon, each variant amino acid sequence should be represented by roughly one nucleotide sequence. Consequently, the number of sequencing reads for a given nucleotide sequence should approximately match the number of reads of the encoded amino acid sequence (the ratio of sequencing reads for a given nucleotide sequence and its encoded amino acid sequence should be approximately equal to 1). As such, in some cases. DNA variants were eliminated for which this ratio was <0.9. Furthermore, as many errors can arise during library preparation and sequencing. DNA variants were eliminated that were only present once after deduplication of the sequencing reads.

Finally, as variant sequences isolated from tissues must be amplified during library preparation, the abundance of a variant nucleotide sequence in the starting material should be proportional to its abundance after amplification. Thus, variant sequences in the starting material should show greater amplification as compared to variant sequences that arise due to PCR or sequencing errors. Thus, comparing the ratio of sequencing reads of a nucleotide sequence before and after deduplication produced a bimodal distribution, with a higher-ratio group likely representing variants in the starting population, and a lower-ratio group of variants presumably resulting from PCR/sequencing errors. Therefore, as an optional stringent filter, in some embodiments the low-ratio group variants were removed.

FIG. 20A shows an example of peripheral and CNS tissue samples analyzed by the methods described herein. FIG. 20B shows amino acid positional abundance in the top 1000 predicted/filtered variants recovered from the CNS compared to non-CNS variants. An overview of the machine learning process is as follows. Filtered variants were binary classified as groups of Present(1)/Absent(0) for the targeted tissue(s): those found in both target/non-target tissue(s) (“shared” variants) were assigned as Absent(0). Each 9 aa variant sequence is represented as a 144-feature vector, composed of 16 biophysical properties per position (TABLE 37 below). Featurized variants were input into 2 separate ML models: Random Forest (RF) and Histogram-based Gradient Boosting Tree (HGB). Training and testing was done with cross-validation and all predictions of tissue Presence/Absence were made on hold-out data not used in model training (FIG. 22). Weights were assigned to each training sample to correct for class-imbalance. Machine learning performance was assessed through concordance between two models (FIG. 21A-FIG. 21B) and the average predicted probability of the two models was taken as the prediction. The top predicted variants were outputted. Important features contributing to tissue targeting/de-targeting were extracted (FIG. 23 and FIGS. 24A-C) as Shapley Additive Explanations (SHAP) values, representing the contribution of individual features to model output.

To accommodate potential sparse sampling of recovered variants from tissues, we focus the model on biophysical properties through featurization of the variant sequence. In this embodiment, the biophysical properties listed in TABLE 37 were used to construct a 144-feature vector for each variant as input for the ML. In TABLE 37, amino acid biophysical features are derived from the following amino acid properties. “Charge” refers to the electrostatic property of the amino acid side chain as an acid or base, having a positive or negative charge in an aqueous solvent at neutral pH. “Phosphorylation” refers to whether the functional group of an amino acid residue can have a phosphate group added as a post-translational modification. “Ionic_bond” refers to the capacity of an amino acid residue side chain to participate in electrostatic interactions. “Hydrogen_bond” refers to the capacity of an amino acid residue side chain to participate in hydrogen bond(s), “hbond_donors” refers to the number of amino acid residue side chain atoms that can donate a hydrogen atom to a hydrogen bond under neutral pH conditions, whereas “h-bond_acceptors” refers to the number of amino acid residue side chain atoms that can accept a donor hydrogen atom in a hydrogen bond under neutral pH conditions and “total_potential_h_bonds” refers to the number of amino acid residue side chain atoms that can participate in a hydrogen bond. “Mol_mass”, is the predicted molecular weight of an amino acid residue in unit Daltons. “Volume” refers to the predicted volume of a given amino acid residue in aqueous solution, and is adapted from Zamyatnin, A. A., Protein volume in solution, Prog. Biophys. Mol. Biol., 24:107-123 (1972). “Hydropathy” represents the hydrophobic (repels water) or hydrophilic (attracts water) properties of the side chain of a given amino acid residue, and the values are adapted from Kyte, J. and Doolittle, R. F., J. Mol. Biol., 157:105-132 (1982). “Goldman_engelman_steitz” is a particular measurement of hydrophobicity, as it refers to the free energy transfer from amino acid residues in an alpha-helix from non-aqueous condition to water, and the values for individual amino acids (kcal/residue) are adapted from Engelman, D. M., Steitz, T. A. and Goldman A., Annu. Rev. Biophys. Chem., 15:321-353 (1986). “Flexibility” refers to the symmetric/asymmetric distribution of amino acid residues in polypeptides and is adapted from Bhaskaran, R. & Ponnuswamy, P. R., Int. J. Peptide and Protein Res., 32:4:241-255 (1988). “Mutability” refers to the probability that a given amino acid residue would change in across an evolutionary interval, and is calculated by the relative frequency at which a residue is replaced with another, and in this case Alanine is arbitrarily set at “100” as adapted from Dayhoff. M. O., Schwartz, R. M., & Orcutt, B. C. Atlas of Protein Sequence and Structure, Vol. 5, Suppl. 3 (1978). See also references on the world wide web at:imgt.org/IMGTeducation/Aide-memoire/_UK/aminoacids/charge/and imgt.org/lMGTeducation/Aide-memoire/_UK/aminoacids/abbreviation.html#refs, each of which are incorporated herein by reference in their entirety.

TABLE 37

Biophysical properties used to featurize variant amino acids

Amino

phosphor-

Acid
category
charge
hydropathy
solubility
ylation
average_flexibility_idx
ionic_bond
hydrogen_bond
hydrophilicity

Y
amphipathic
0
−1.3
63
1
0.42
0
1
−2.3

W
amphipathic
0
−0.9
97
0
0.31
0
1
−3.4

D
charged
−1
−3.5
−55
0
0.51
1
1
3

E
charged
−1
−3.5
−31
0
0.5
1
1
3

R
charged
1
−4.5
−14
0
0.53
1
1
3

K
charged
1
−3.9
−23
0
0.47
1
1
3

V
hydrophobic
0
4.2
76
0
0.39
0
0
−1.5

P
hydrophobic
0
−1.6
−46
0
0.51
0
0
0

M
hydrophobic
0
1.9
74
0
0.3
0
0
−1.3

A
hydrophobic
0
1.8
41
0
0.36
0
0
−0.5

G
hydrophobic
0
−0.4
0
0
0.54
0
0
0

F
hydrophobic
0
2.8
100
0
0.31
0
0
−2.5

I
hydrophobic
0
4.5
99
0
0.46
0
0
−1.8

L
hydrophobic
0
3.8
97
0
0.37
0
0
−1.8

H
polar
1
−3.2
8
0
0.32
1
1
−0.5

N
polar
0
−3.5
−28
0
0.46
0
1
3

Q
polar
0
−3.5
−10
0
0.49
0
1
0.2

S
polar
0
−0.8
−5
1
0.51
0
1
0.3

T
polar
0
−0.7
13
1
0.44
0
1
−0.4

C
polar
0
2.5
49
0
0.35
0
0
−1

Amino

muta-

vol-

Acid
Surface_accessibility
bility
hbond_donors
hbond_aceptors
total_potential_hbonds
mol_mass
ume
goldman_engelman_steitz

Y
1.089
41
1
1
2
181
193.6
−0.7

W
0.808
18
1
0
1
204
227.8
1.2

D
1.283
106
0
4
4
133
111.1
−9.2

E
1.445
102
0
4
4
147
138.4
−8.2

R
1.475
65
5
0
5
174
173.4
−12.3

K
1.545
56
3
0
3
146
168.6
−8.8

V
0.606
74
0
0
0
117
140
2.6

P
1.236
56
0
0
0
115
112.7
−0.2

M
0.714
94
0
0
0
149
162.9
3.4

A
0.815
100
0
0
0
89
88.6
1.6

G
0.714
49
0
0
0
75
60.1
1

F
0.695
41
0
0
0
165
189.9
3.7

I
0.603
96
0
0
0
131
166.7
3.1

L
0.603
40
0
0
0
131
166.7
2.8

H
1.18
66
2
2
4
155
153.2
−3

N
1.296
134
2
2
4
132
114.1
−4.8

Q
1.348
93
2
2
4
146
143.8
−4.1

S
1.115
120
1
2
3
105
89
0.6

T
1.184
97
1
2
3
119
116.1
1.2

C
0.394
20
0
0
0
121
108.5
2

Favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an AAV5 VP1 capsid polypeptide, which are associated with a higher probability of CNS tissue tropism as predicted by machine learning are described below. Any of the below machine learning-derived positional amino acid preferences described in TABLE 38 can be present alone or in combination with each other or in combination with any of the amino acid preferences observed and described in EXAMPLE 5, TABLE 9.

TABLE 38

Machine Learning-Derived CNS Tissue Tropism Rules

Low amino acid solubility at position Xaa1

Xaa1 is selected from K, R, or Q

Low amino acid hydropathy at position Xaa1

Xaa1 is selected from K or R

High average amino acid flexibility index at position Xaa1

Xaa1 is selected from D, E, R, K, G, I, N, Q, or S

High hydrogen bond donors at position Xaa1

Xaa1 is selected from K or R

Amino acid mutability at position Xaa1

Xaa1 is selected from K, R, P, or H

Low amino acid solubility at position Xaa2

Xaa2 is selected from R, K, Q, or S

Low amino acid hydropathy at position Xaa2

Xaa2 is selected from R, K, D, E, N, Q, H, P, Y, W, S, or T

High amino acid charge at position Xaa2

Xaa2 is selected from R, K, or H

High amino acid solubility at position Xaa3

Xaa3 is selected from A, M, V, W, L, or I

High amino acid solubility at position Xaa5

Xaa5 is selected from C, M, V, W, L, or I

High hydropathy at position Xaa5

Xaa5 is selected from M, V, or I

Low average amino acid flexibility index at position Xaa5

Xaa5 is selected from M, W, F, or C

High amino acid solubility at position Xaa8

Xaa8 is selected from H, V, or I

TABLE 39 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that exhibited CNS tissue tropism and comport to one or more of the rules provided in TABLE 38. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 8118-SEQ ID NO: 9117, as disclosed in TABLE 39. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 39

Sequences of the 581 to 589 Region in AAV5

VP1 Capsid Polypeptide that Drive CNS

Tissue Tropism

SEQ
581-589

ID NO
Sequence

8118
HWELVRCHD

8119
SQSRTIEQA

8120
KMAGIGCWD

8121
KSEMMSKLD

8122
ICIRGARTD

8123
CCKSKEACA

8124
RVAGEGIQS

8125
DNYWIIPFA

8126
TERDHKTIY

8127
VKHTAVDVL

8128
RGLSQDCCR

8129
KMEGNMSEL

8130
KNPPWICVW

8131
GVFRPDSFR

8132
AKCTIQNFQ

8133
KTVTKEVTP

8134
RIFYVMGIG

8135
HWEMIRCHD

8136
GKVQWSAQA

8137
VAANSRMMI

8138
KSPCYMSVQ

8139
KSGKVWYVL

8140
PWRARVGAR

8141
RGLTQDCCR

8142
RQYFEDSAG

8143
KSVQVYWHP

8144
IPINDREWG

8145
QIRSLITHA

8146
RMTPHNCQR

8147
EACYWGHCS

8148
MMAVIRQKS

8149
LSAKQRFFC

8150
KFNTKYQLM

8151
QNKHNWWIT

8152
SRLPVHKCP

8153
IRNRYAQAP

8154
KVNCRMTIM

8155
KSYLIANRH

8156
KTTFDVTCY

8157
MEDRVFACC

8158
NLNNLVSCR

8159
DAIWEHIRL

8160
KWAIVGYAL

8161
KFGFMYSEF

8162
KSTMLEHHA

8163
GRWMCNEAA

8164
KDHQLWGVW

8165
ASSFCKFTS

8166
PKRIRYRTT

8167
TKITMPGYQ

8168
TFHQSDYWA

8169
GKRVGDTSC

8170
FAQRDKQDR

8171
KTYSAMHSP

8172
TRAYYDSYS

8173
RMTPNNFQR

8174
KWGNEGRMS

8175
TVRHRMDAY

8176
RQQRNHASS

8177
TISQIPLVR

8178
FNWLVRAYI

8179
MYTGRRCWR

8180
QCHRGYIYE

8181
NYFAFWVFI

8182
AIDCCYSCG

8183
KIYMQCDRE

8184
DRKQAGQIY

8185
RDVGEENSA

8186
IAKKNFMKT

8187
RKAYLNYVA

8188
EMHAFWQRG

8189
FCKCFVKHC

8190
RKKVEVVVA

8191
NTRTGNDRL

8192
LADCMINHH

8193
LDTKTLASS

8194
EVDSKVICV

8195
RTFMKDMCE

8196
DAHSPWQVI

8197
TRHPTITMH

8198
SSRHRGGCQ

8199
RCTDHGESI

8200
SSMCSGSLL

8201
GKLTFDTER

8202
RKDQSDLWS

8203
RSMWSAFKH

8204
FRLDYNYKN

8205
RKKVEVVGE

8206
GYNYYWQHV

8207
TKLMYNTVD

8208
KTLHMPESK

8209
GRPVWCVCM

8210
MVHRVQGDD

8211
ATRYQICGE

8212
DIRWLPYDL

8213
HMHNLSVKP

8214
THRTMDDYF

8215
NYFTFWVFI

8216
HGCKCQYSF

8217
VSANEDRAG

8218
YPVNMMPPL

8219
GFRRTHVDQ

8220
CWMLHHRCS

8221
CVNLMPVNN

8222
TMRAGLENS

8223
QGFSTDAKP

8224
QIVSCDTET

8225
QKGMMLDYA

8226
DSHRCLKAA

8227
NTSEMLMAQ

8228
GKIARDQLN

8229
QYWWPTTHD

8230
KTNFGDAIE

8231
YIYMMLNLH

8232
QWFAKGVGE

8233
QLMSCANVE

8234
DIEFCMKQY

8235
DYEKFVKWM

8236
FCNYAAMSP

8237
CYWSMTNCT

8238
KWPVTANLH

8239
DTRHKWCLE

8240
FCVKSPRVD

8241
DAIQWGSWQ

8242
SALYRRGHV

8243
FMTVRNWAG

8244
KISTTWADT

8245
MHNAHVIIE

8246
MTGCRETVD

8247
QYKWHDVGV

8248
VACDMCLVQ

8249
KSEWTCCPH

8250
ARKPCNGWD

8251
TRYPTIAMH

8252
TFMFNSKWS

8253
RAMAYHFYW

8254
LYNMRMKDR

8255
DIWWTLTRD

8256
YHVHMMPPL

8257
KSGSCAMRV

8258
TRWTEKRQA

8259
QLMPCTNVE

8260
MGAMKSDDF

8261
AMKSHGCLT

8262
QNGPILTSR

8263
YCCNQEPFT

8264
NWCTMYRYC

8265
KDPDFYDWP

8266
LWIKQHVKC

8267
TRLEIEYMG

8268
DMTHTGIMS

8269
SMHMRVCYT

8270
IANKSNCHK

8271
VGRFNAYGE

8272
GLVANVTPR

8273
NTHTGNARL

8274
MVKPCLDEW

8275
QIKAHSNVF

8276
LVDRSGRAA

8277
TSKGKPTCC

8278
NCYCAYGWP

8279
IWQSLKKAT

8280
ARSGINGHA

8281
NWTRNMQWA

8282
NCLSRKKFA

8283
CIMLNGCFP

8284
KNGATNLAW

8285
NNCKLAMVH

8286
KATHCDKYA

8287
VFNPMLQDT

8288
ARPEIEMIG

8289
NSVHVHSSN

8290
SCPWKCMIV

8291
CSNDINGRT

8292
KMGMHDMAT

8293
KKLVNEGTW

8294
QLNQWTFAG

8295
FERRIGWDH

8296
VWNVMIKMC

8297
TGGCYRMME

8298
KPDPRQADQ

8299
LSCKFGWDG

8300
KYPFKYYVT

8301
TMLQKKRQD

8302
GYAQRSFFM

8303
ATDGYVAQD

8304
KMSTYHTYD

8305
HVADNQVVQ

8306
AKTFVMSFD

8307
TCNVSWEHE

8308
RRTDHCEPI

8309
RTDYSRLSD

8310
TDLRKRSNL

8311
IKAWWHDHQ

8312
KFKWEACCP

8313
VNVDCDVQQ

8314
QYQSTGVVR

8315
LIHEERKEQ

8316
ETNVVEDWY

8317
LACTPRYIF

8318
HGGCYRMME

8319
QSVNERHFH

8320
DWIKQHIKG

8321
QLMPCANVA

8322
TQREGETSY

8323
HNDWQQVFS

8324
YEDNDVMQI

8325
ARRAGLENS

8326
GATWLFRSP

8327
RTLWALYAI

8328
DCIICSSKG

8329
HMALVMQTV

8330
HSNMSLVHQ

8331
TNGAQIMHV

8332
STRPIACQS

8333
CMYVPADQE

8334
SPHYFADQW

8335
GFDLMRQIN

8336
GHHQCYKAN

8337
EMNSDYSGI

8338
SYWHVLLLV

8339
SADRTVMFH

8340
RFTVKDGDA

8341
QCKPGAEGF

8342
KAIQAFEHD

8343
ATRYQICDE

8344
MWWREGAMR

8345
LFMTYSTRN

8346
QPPNREAWC

8347
TPKSAGICC

8348
KPTYEHRGF

8349
CNYPIIWDR

8350
FTGTPPVGM

8351
QGQTGKYVM

8352
AHLHNQVIF

8353
KATVEMQSL

8354
CVRIGMRGP

8355
HLSFMMSSF

8356
TTMWIWFPF

8357
TTMMKWCAG

8358
FIQPTWDSY

8359
RC1WCHMQG

8360
LVHRVQGGD

8361
NVRSPAVRD

8362
LKENWMEWG

8363
YSASFGCQS

8364
GLKYWMHGI

8365
QTGTGHGFE

8366
QMNYQHEHR

8367
AHLHNKFIF

8368
STKQMTKSF

8369
QEENGKSVT

8370
RVADEGIQP

8371
CGLSTHEYL

8372
RANTESDQP

8373
QSAEWYLNG

8374
CWQLFNEQN

8375
AADGYVIQD

8376
SANQRHHDW

8377
KCVEWNAPA

8378
MHDTMGAWC

8379
GDNMVQWGS

8380
DVWCRKLMP

8381
SYRTLFNGW

8382
MRQHMKYSL

8383
PDQYLVVGD

8384
RELDKENCV

8385
MTGQCRNGQ

8386
TSLWQQMAS

8387
HSMFRHPCP

8388
GNTKCPSWH

8389
HECDTVAVN

8390
ERTGEKGIR

8391
GSGPVQSEE

8392
DGWCQKLMP

8393
QSVNEHHFP

8394
RAAKYQNSH

8395
DYERADCWM

8396
IRNYCRMTA

8397
LAVWQQKME

8398
INMSVSKDQ

8399
ACDKWDCCC

8400
SQPVCCVCM

8401
GRALANCAE

8402
ITTHMNCAD

8403
MKTTMQDYF

8404
HCIKVESVG

8405
HKGTHAFAM

8406
HYTEMSCDD

8407
DFWWTLTRD

8408
SMQYFFKTA

8409
LNCPSKDGT

8410
CTHMKNVRT

8411
ANADNSFCQ

8412
EERPWEWET

8413
CSHGTGAMC

8414
FAIWDYESK

8415
HLRFIMHEP

8416
HLMPRSQCF

8417
QLMVAFLCP

8418
AHSPFLDHM

8419
KQAGYDLFH

8420
DTMQWGSWQ

8421
SGFFVHTSS

8422
AKIIAQLCV

8423
NWTGNMQWE

8424
QREGNNYLT

8425
NAFWTQNME

8426
IKQVFNQIV

8427
HAVSGPTTG

8428
NTSGLLMAQ

8429
MYLSFPVHA

8430
STLSRHGWG

8431
QWMWQRLID

8432
TQGTMDSWY

8433
RPIHTRQNY

8434
EWLRQPRMT

8435
NQQRTNNNG

8436
QPRTVGVAG

8437
ASWPCGRNV

8438
LNSMIMWDN

8439
ESRSPNNPA

8440
PTSWLNDLT

8441
MIRKRFEGQ

8442
AMWAGLENS

8443
KLTDIWAHM

8444
TSYNCGRDP

8445
HCVKVESVG

8446
TACGSFHGC

8447
MSVPKRQGP

8448
LKQAKQMEP

8449
DCDVWHANV

8450
MGDWLGVPT

8451
FCKHNAQTQ

8452
QTKPMIGNE

8453
LSAQFNNQS

8454
PALMRRNIF

8455
ISTICPKIC

8456
TGSFMGCVD

8457
MTGCREAVA

8458
ERGLDNKPR

8459
NYCINKRAV

8460
SGQPVDRMF

8461
QIMSAKVFG

8462
MFVVNRNWA

8463
WPDPESEEV

8464
MNTMLMSNS

8465
HFVDNQVVQ

8466
NTRQRYHTW

8467
TSLVMPSLP

8468
DYRQCIDWE

8469
VLRYKCYPS

8470
HWEIEWVYW

8471
QVMMGQQTP

8472
RGCQVKCHT

8473
EQWKTHQQN

8474
YSKWARALS

8475
QCMITISCP

8476
DVWWQKLMP

8477
VANTWQVHC

8478
RMMWQSHCG

8479
VANITACAN

8480
PWAVEYSLV

8481
SYMWPHFPH

8482
FFEQEAHNR

8483
LCTGTERQG

8484
MAVGTVSAK

8485
QSSSPVMWV

8486
SSVVMVKQP

8487
EPRLYCAQA

8488
YANIHNRHG

8489
EAYIYLGEA

8490
PVETARCSG

8491
NAIPYFIQA

8492
FCKCFMKHF

8493
EKLVMYHKC

8494
YEDCMDMYF

8495
GIWVWDTYN

8496
AEWDHKTIY

8497
TQIQIRQAD

8498
RCNWQFAQH

8499
ESRHCKYPY

8500
SSNCMRRAS

8501
GVGQFRLLS

8502
KIDQTQWRR

8503
HLSFLVSSF

8504
NAGMSFRNR

8505
VMNRMEWSV

8506
LFMGMSDST

8507
ANVVMRKCQ

8508
GIYYQWRGQ

8509
DVHAEHSPQ

8510
QHPMTFSHE

8511
EPRLYCTQE

8512
NAKEQVVNT

8513
EFCHDDILM

8514
PLFRSEWLA

8515
GNMLRNFGS

8516
HAETDHAGL

8517
MKMHQLNGC

8518
SSERFRAEG

8519
QGGCAPMDS

8520
REVIMTGCG

8521
SDNGWCAHS

8522
YCCNQELFA

8523
TFVYGEWDR

8524
IFPSKIPCQ

8525
YVTQMQHLE

8526
NSRLLIKHP

8527
KTFPAADMK

8528
VGHVMIKMC

8529
GTDHVIVGY

8530
GAWSMQSSC

8531
YAVQTNNIM

8532
WGGMSNNQG

8533
QMVTQLNWG

8534
VPKQGAKQE

8535
DEYQQESFR

8536
GGTRSGKVV

8537
CQNTAIRAI

8538
TIHPPWVTL

8539
CMVSTPKLA

8540
RIWWALYAI

8541
SNYTDVETW

8542
WTSSKAVYD

8543
ESSFQDNVI

8544
YAHAFHCQP

8545
RRPPKPGTT

8546
QMYATMDTY

8547
EGGQCEKCK

8548
MGDWLGTPT

8549
GVKYATLHD

8550
PWEIEWGYW

8551
MYLLFPVHE

8552
HVLNGTGGP

8553
NIGQSPTQL

8554
VSCMWCQEN

8555
TMSVHYGQV

8556
NAAMITTVC

8557
HAVNSDCKK

8558
NIRPCVNQE

8559
QMECRGGAM

8560
VGVQCPNGG

8561
MQGMQTFWT

8562
QTMGIGGGP

8563
RSQQDRVMA

8564
QQGMCVTAI

8565
AIGPRLIDD

8566
HGINVELGS

8567
NVDGRHPMI

8568
DAARERHWE

8569
QTEIGHWVE

8570
TCGQCVNGV

8571
EGSMFAERQ

8572
HSCKYLMRV

8573
MCDWLGAPT

8574
EVSMFAERP

8575
ESAITSCAW

8576
RVCEMRTVD

8577
AHQGCRPGQ

8578
ANMISPYGL

8579
EKHRCAKDR

8580
TQQRSQATA

8581
IPMSREFPE

8582
GNQLITMSG

8583
YEMTSKCAY

8584
TIGQIPWVW

8585
HHGASLGKY

8586
QMMPQMDCS

8587
DSWQYYNGL

8588
VTYYGHYSV

8589
TIVHMPKHE

8590
QSHHFLNSA

8591
TNTMIWNDC

8592
KAIQASERD

8593
CRSTGMMRQ

8594
TSSLTDVIW

8595
TQAKIECWS

8596
KYAHSSCED

8597
NYENNFRAQ

8598
SPSQKGYGP

8599
QSHGGVVVQ

8600
WIQLAILVC

8601
NTHTGNYRL

8602
EACYLGHCH

8603
DEYQQESFP

8604
YRQQMFERQ

8605
AQQSTMISL

8606
AHQTPWAWI

8607
SISRADLQG

8608
LNVMSIHRC

8609
AGRFHKHYA

8610
GEMTRCFRN

8611
QVKWTANPD

8612
KTISPHVFA

8613
MGRPGACQA

8614
FMQPYGHGE

8615
DCWCYTTKC

8616
LIRRHCHYP

8617
QMGVTVVQR

8618
VCYNQEKAA

8619
EMNSDRSGI

8620
HLWMCPPRY

8621
TASLTDVIW

8622
LETTMRHGE

8623
FIGENNAEA

8624
QQQRTVFGM

8625
CYNQFHTYN

8626
LTISHPKSQ

8627
AVNRIQQVH

8628
GWFMIQSMK

8629
WCNWKFAQH

8630
MSYLVKGVG

8631
NTSRELIMP

8632
TVNPCVNGE

8633
QMGHQLCVQ

8634
DERHKWCLE

8635
MWHELITDQ

8636
LTRSGPVGQ

8637
AQFHIHRWQ

8638
VCTPPLNMQ

8639
SWSQLQHTA

8640
VATYIHAMH

8641
STPQGKQRQ

8642
TTTITNRLR

8643
SSLCRTYDN

8644
MLEKRNHWG

8645
NSELLRYTF

8646
CGVGLTQCK

8647
TGQQIHYKG

8648
DLEPIMTQA

8649
PIAQDKVWS

8650
YSTIQGSGP

8651
ATTTIQGKA

8652
ISICAQDIG

8653
STNPNLTGD

8654
THELHGHCV

8655
QEMMIVNCD

8656
EADTFTLIV

8657
GMNAYPAHS

8658
NCRQLGDNM

8659
ARCSMLDHG

8660
ASTWMQSYS

8661
HYCSCHNKY

8662
VSNVTIGCC

8663
DNAFMHHHV

8664
SKVHWDLPC

8665
NPQHQIDIG

8666
MWWCESAMR

8667
FIQCCQDTP

8668
RMVCDGRID

8669
ELPRYYGQV

8670
VTAFTTCHY

8671
TCSSTEGSG

8672
EWRTISYGN

8673
QSTPTLGPK

8674
GWALIVNDL

8675
VTILGLNPT

8676
VTCHWHQGG

8677
QGSVVHSPS

8678
NQARENWFH

8679
HIKVCTLGD

8680
QYYWFAQFL

8681
LKVDHKIGD

8682
DSPAFHQSG

8683
WSCRSICPP

8684
QHATTCTHP

8685
QYQHAVWAP

8686
LVCSNVDDR

8687
GKEHMPHHM

8688
SMSDCEVSF

8689
DHIWLHWME

8690
WSTLYLMTS

8691
MANDQRVDY

8692
SYWDVLLLV

8693
KMGTLHNME

8694
DHHRTSAVR

8695
MNTPCQQAP

8696
NSTHNTSYY

8697
AVSMFAERQ

8698
STDLSNHFQ

8699
DSIIKDQSA

8700
FCGKGANQI

8701
TQRSTQWSK

8702
CHYQINKFA

8703
QATPACPAV

8704
DVFWPDSFR

8705
FVIKPNQAT

8706
CRKSKEVCA

8707
ASGFGHWWG

8708
RKRTAGVHF

8709
TYMYRSDCS

8710
MSVKKECGG

8711
WHFLMTFYI

8712
IADDYFMYH

8713
RIHDTMSDY

8714
CNMPQQCME

8715
TVGMRQAMM

8716
KKIFCSRDY

8717
NHDEKMWAP

8718
LCAVCGGCD

8719
VHPQTHVTD

8720
QHEQSNRIW

8721
EAYATVHMC

8722
LMHTYVVQQ

8723
DACAAHDHW

8724
DIMMKHCRR

8725
LTQYKALVQ

8726
MDKHKIGYA

8727
ESDVALGTI

8728
NVGYYKWQS

8729
IECFPRESI

8730
LSMPYAPTT

8731
AVKILMHDH

8732
THELHGHCS

8733
NSQTIMLHQ

8734
HIHMSDRGT

8735
TGRAWMWHS

8736
NSHAHGHMK

8737
DYTIMPYIA

8738
SVTYHERHP

8739
CTAMGWVHN

8740
SVTYHKWHP

8741
VSCEFGQEY

8742
KDLEYQRSN

8743
AKGPHWGQG

8744
RYHHSCDSH

8745
ACTWLHGTN

8746
SIHNVLVQN

8747
KVDQNTNWC

8748
FGEFTTDEN

8749
VWMRFDYVD

8750
MAKKREVGV

8751
MDEHFSVGC

8752
EFRCRLIDS

8753
NHCHCIAHP

8754
IIMLGCTGR

8755
VTLTSKMCQ

8756
RAECRYQII

8757
FDVHKMCIY

8758
VSADTPYMN

8759
THRTRHTDC

8760
DYYLNRASP

8761
SVSTKPHAE

8762
VADWIGRTP

8763
KLYIVMNQA

8764
NPIPGYFEQ

8765
SCKWIQSNR

8766
QWGVISGHA

8767
ESEFGHWWG

8768
KKAGAKDAL

8769
CTCEPRSAW

8770
YLYHSMMKQ

8771
ILRTIPNRT

8772
IEESIQNAD

8773
AMMPKTETH

8774
VVTLHSKAR

8775
VCCSHADSA

8776
DGSSHFTMQ

8777
VAYPIQTDA

8778
YAQQSPHCE

8779
YMLHAIMES

8780
AAVITINGL

8781
LNMVWGQKF

8782
NALPAQQHH

8783
QKPTIWDHF

8784
YQDIRQCIF

8785
TCTHFEFVD

8786
TLEPAYWHE

8787
PVHFWTQSK

8788
RCKRAVQVV

8789
WCESWGPQG

8790
AVVYKMNYH

8791
KNHMMLYVN

8792
ANMCGCDSF

8793
LFSDHGVSY

8794
TWSCCLHQY

8795
WTDSDSDCT

8796
VIDCCYSCD

8797
PIAQDQVLS

8798
LCRWYRYAN

8799
QNIRPGVPT

8800
RTTDHRADE

8801
QIHSEHCTT

8802
QSHDGVMVQ

8803
DLLANTICT

8804
SANIVFSVC

8805
LSMTRELWH

8806
MLHKYLHMW

8807
QYAITYQKG

8808
VQMYWSSKD

8809
GFNLMHMAG

8810
LFYSPMHTH

8811
VKAKKMANP

8812
NCMPTIAGS

8813
SHWQMQVMA

8814
HIMQEGRSS

8815
HFAMKCHNN

8816
EIIQAHLVN

8817
VKNALCQHM

8818
KTWDAHRSS

8819
IRGFMQNIY

8820
VSYAPMDAQ

8821
SGSMTQHHT

8822
AICTTASFD

8823
YPTECNHLT

8824
QSQHLAYKG

8825
LKAQAVTVT

8826
CTVQFELGC

8827
GTSQLQHMA

8828
PIYCNALCS

8829
VSKPDMHVS

8830
GPLVCISAG

8831
HKVKFGALH

8832
TQGWLAAGH

8833
GPRLIVHKL

8834
MFGFSYSDG

8835
TMGMANCGM

8836
QSAIMSART

8837
VWGMAYTAC

8838
DATFMKNAF

8839
TEGPMHRVW

8840
FFRICDSYI

8841
CTALVNEDL

8842
SQAHQNRAC

8843
AMIGHGAYA

8844
HAETHHVGL

8845
ATVDVKGSH

8846
SSDGESHYI

8847
RTLLPNANS

8848
ITCNWCVMP

8849
GSLLQGVSD

8850
FCIMASDMA

8851
QGAFQMMLP

8852
FMWTTHRWH

8853
MFAQQMGVD

8854
ALVMIILQG

8855
ATISTESEV

8856
LTCNWTHHG

8857
TFKEYQGYH

8858
IRQPIHMTP

8859
QTAMWCWQE

8860
HIEVCTLGA

8861
QTHLKLECE

8862
GRNEQGYCH

8863
TFMQNTYQG

8864
LTNYHIAQE

8865
ATTEKCRSI

8866
ENAQMKCLM

8867
HKLWYWAFF

8868
AHAFRHKHF

8869
KFICTTITC

8870
QPGIVICKK

8871
IMGYKCKWL

8872
QPGYMNKLW

8873
QCDAALNWS

8874
ISTVCPKIF

8875
NDQHAPCTE

8876
ASSHRHWVY

8877
AMMLFQTSY

8878
SKEDLCHQQ

8879
AKIHGAWNA

8880
ASAPRVNEQ

8881
SCPWKCPIV

8882
PNRYRPFCC

8883
DHLRDHPLY

8884
GNGTVGKAN

8885
EQAGAASMW

8886
VQKLTLNAK

8887
GHNHMYTQP

8888
VSKTPWERD

8889
QSCHVMNMP

8890
IVLWFVNTT

8891
WCQHIPHNG

8892
CIAREYTGY

8893
YFVVNHFWS

8894
STLEDEKPY

8895
ARCHDMWGH

8896
MAAIKWYMS

8897
WMLHGWATL

8898
MVDCMKWHE

8899
AGMYSFSGE

8900
VALQTKIEQ

8901
GTQNLLHGY

8902
MTQYKALAQ

8903
GCATYSCYV

8904
QGDMGTWVG

8905
SAVVSMRND

8906
YPYGKCSNY

8907
RVINTEWGR

8908
TIYHHGNTL

8909
QQTTYDCLD

8910
LTHLQHSAT

8911
YMPLAMQYC

8912
TREPCQQCD

8913
IQDLQIAIK

8914
ATTNHDMQQ

8915
FCNKLTSVW

8916
EAVCMDTFP

8917
SAALVADGP

8918
QPEIFPVMA

8919
HTESGQDET

8920
TRNTWQQDF

8921
GVNTNMNGS

8922
CQAELIDVR

8923
TSTMCSWSM

8924
NAEPQQADI

8925
IRQMAGGHC

8926
LNQCAQAGC

8927
TATCGIEAE

8928
TNLHNNHYW

8929
AYSYYSVLP

8930
STMLLLARD

8931
VAYYSGHMM

8932
IQQLSMVGS

8933
ARVKETCEW

8934
TFMFNSEGS

8935
CVPKTIACC

8936
ACYEFVNAC

8937
TRMQFNKCH

8938
SRDIAKSST

8939
YESCNLTRD

8940
MVGRNMWAV

8941
MVGRNMWAV

8942
CQMQVLNNI

8943
YLVHEGHSR

8944
TGTQFMRWA

8945
MCMQEDRLA

8946
LATNHREIM

8947
GRWPYFLAC

8948
IPSFYWGDY

8949
GDDWIIWQA

8950
MECIHMRND

8951
SIWDCNHRS

8952
NTVFQLYHT

8953
GYGHTFWDA

8954
VHEEYSNSH

8955
GTLACDQKE

8956
FCCMYCQHP

8957
ESSMWCTDC

8958
QTADWCHPV

8959
MPHGRCNTI

8960
GITQHFVKH

8961
AMDCGQSFG

8962
LAYSQYVGC

8963
NRNCTDAWH

8964
RSHFQEGVQ

8965
MFMAKQNAM

8966
PIWSVYQDE

8967
DKRIICQDN

8968
MFYAMVGPE

8969
ETCCSEWHV

8970
AIHAHTAAN

8971
AIHAHTAAN

8972
NVSSHQHCW

8973
CRCCWYQDQ

8974
DMSDCEVSF

8975
SMCWFNQVA

8976
AAPMVGEFY

8977
SPMSFLNVL

8978
TKGPKELMA

8979
EGWTQFRPE

8980
QAGDSMWAK

8981
SADTNVVVE

8982
LKSGQYRFT

8983
YIHDYSSPN

8984
QTEEAMGFT

8985
QLCIPLTWE

8986
EANCNHVGV

8987
SGMKGRTQI

8988
AEAIINDIW

8989
LTEFTPYIG

8990
FCAHNFTSD

8991
SFLMMMNCT

8992
LCMPADRQS

8993
MTNSQHQHW

8994
PKHHVTWED

8995
VSQPAWQQK

8996
QAQSHIHYT

8997
EVLMAMSLF

8998
GQALDKQYQ

8999
SNEPLDTCY

9000
SQAVRKCCD

9001
QGSPLDSFQ

9002
LLFPLMGSS

9003
PNSPGVCSC

9004
MSTHVLKWV

9005
FCHAFPDAY

9006
VEGWHGTNA

9007
TQGVSTIGV

9008
NDEQRNTLY

9009
TSQHKYAEE

9010
HDCIWMECR

9011
DFAYRSTGV

9012
DSATSKSAD

9013
EPASCGSTR

9014
SMGWDHMMS

9015
MASTESDMC

9016
QMEVHWHMR

9017
AQWISPWPT

9018
QENDANRPQ

9019
LVNQLFHSW

9020
ACDQVNTFQ

9021
MKSVFMHEW

9022
WGEKCLGTS

9023
VSRSDWTMP

9024
TSTEPGVGT

9025
YHTFVECAQ

9026
ASCYVWGPA

9027
QHQSVMGFA

9028
HMQAQIHYW

9029
KQHELIVRV

9030
AYCCFNRFQ

9031
TYMQCMVSG

9032
CAVNTYQKH

9033
RNINEYEHY

9034
SARVNHGRV

9035
HSHMYDCQQ

9036
NTECWQNSD

9037
LNCQKANQN

9038
TVMMLMDNG

9039
MTTGLMAYE

9040
LLYRNMILS

9041
QSHLSDKMY

9042
TMAMLGWMN

9043
GLWHGFYAV

9044
GGCCGGELN

9045
LQYSPGQQT

9046
TNNCYWCAP

9047
AVLGSNYLK

9048
MDTKVVYGT

9049
WADRQGALA

9050
VITLKNNCH

9051
AIAEIKEQS

9052
VTMPMATWH

9053
VHFEAWRGM

9054
VEGVEMKCM

9055
TDAGMFAHA

9056
ITKFKMQDH

9057
AMCEVCIAP

9058
KSGYQCDSM

9059
LALMIQQAH

9060
LSAQDTMFH

9061
YQAQLNNCW

9062
SCEATMHAI

9063
TELGEPSSK

9064
DMTHTDEMP

9065
LEGPNTKWG

9066
EARMPWAHH

9067
HVHMCTPSV

9068
LNTYYRLED

9069
INCQTKKEM

9070
VPEPKGTLN

9071
STHMAGHYA

9072
APGMAFGGQ

9073
SVSAQQTMQ

9074
RTVYDVQDD

9075
GCYGNQPPT

9076
TATWGIEVE

9077
ECATNVGDR

9078
WDQKNLMDT

9079
ERTLTKTGS

9080
IVNMNHSAM

9081
FWMKWESYG

9082
AMRDNSVAH

9083
CWSYTNKNC

9084
QANPYNHGI

9085
IHVKFPPFD

9086
EYIQHSSAR

9087
TTTTPCNMP

9088
CCTGLGSMG

9089
TVTHMSTWG

9090
NNMLHGAYR

9091
WGQHLPMID

9092
MMKMCEYDG

9093
GANNMTAMN

9094
THAAYGTNL

9095
LSSLPMHYC

9096
ETEQYDAFV

9097
YTGIKVMGR

9098
DVEHVSCPK

9099
WHCEFTQHH

9100
LSTQSAKEY

9101
ISDPNVGFP

9102
SVCSKWDGT

9103
NNMWDTIFM

9104
NHKMVRDAL

9105
HVGIYEHCH

9106
SPKMMVSMT

9107
WMVRQKYMA

9108
LQWWERMGK

9109
CCGEREEYG

9110
FAGFTPGWM

9111
PHKDSMQMT

9112
DKTVQSLSL

9113
PWACEHVWH

9114
EYCEFMSEA

9115
YGVMCPSHY

9116
PIIRGRSDS

9117
EPMEANQCR

Example 20
Engineered AAV5 Variants with Tissue Tropism in Liver as Identified by Machine Learning

This example describes preferred properties of engineered AAV5 variants that display liver tissue tropism as identified by machine learning methods. The same bioinformatics pre-process, machine learning methods, and analyses as described in EXAMPLE 19 and FIGS. 19-FIG. 22 were carried out to identify preferred properties of engineered AAV5 variants that display liver tissue tropism. FIG. 25 shows a set of top 20 positional features contributing to model output probability. Shapley Additive Explanations (SHAP) values can be used to interrogate the relative contribution of features to model predictions. As shown in FIGS. 26A-C, these features were further compared between tissue targeting and non-targeting variants. FIGS. 26A-C show a comparison of top predictive features positionally. Features were selected if they were found to be important to both the HGB & RF models. Summaries are shown of the features in the top 1000 ML-predicted liver variants compared to random 2% liver variants.

Favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an AAV5 VP capsid polypeptide, which are associated with a higher probability of liver tissue tropism are described below. Any of the below machine learning-derived positional amino acid preferences described in TABLE 40 can be present alone or in combination with each other or in combination with any of the amino acid preferences observed and described in EXAMPLE 4, TABLE 6. Listed below in TABLE 40 are a summary of positional features shared between the top 30 important features for liver tropism extracted from two ML models (HGB & RF).

TABLE 40

Machine Learning-Derived Liver Tissue Tropism Rules

High surface accessibility at position Xaa1

Xaa1 is selected from K, R, or E

Low hydropathy (<−3.5) at position Xaa1

Xaa1 is selected from K or R

Low amino acid mutability at position Xaa1

Xaa1 is selected from H, P, K, or R

Low amino acid solubility at position Xaa1

Xaa1 is selected from Q, K, or R

High surface accessibility at position Xaa2

Xaa2 is selected from E, R, or K

Low hydropathy at position Xaa2

Xaa2 is selected from K orR

High amino acid volume at position Xaa2

Xaa2 is selected from S, L, I, A, R, or K

High mutability at position Xaa3

Xaa3 is selected from N, I, A, M, E, or D

Low solubility at position Xaa3

Xaa3 is selected from N, K, R, or E

Low hydropathy at position Xaa4

Xaa4 is selected from K or R

High amino acid volume at position Xaa4

Xaa4 is selected from K, R, I, or L

Medium amino acid solubility at position

Xaa5 Xaa5 is selected from H or T

Low surface accessibility at position Xaa8

Xaa8 is selected from V or C

Low average flexibility index at position Xaa8

Xaa8 is selected from W, V, M, A, F, L, H, or C

TABLE 41 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid poly peptides that exhibited liver tissue tropism and comport to one or more of the rules provided in TABLE 40. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 44438-SEQ ID NO: 45437, as disclosed in TABLE 41. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region

TABLE 41

Sequences of the 581 to 589 Region in AAV5 VP1

Capsid Polypeptide that Drive Liver Tissue Tropism

SEQ

ID
581-589

NO
Sequence

44438
KAVAGCDFW

44439
KVQSAVEVT

44440
TTKPKQSCM

44441
QATKLQDVA

44442
RARHTEECA

44443
TSHKQSDCT

44444
KSGHTWDMS

44445
RVVITSNCT

44446
VAFRMKWVC

44447
KDTFTKQWQ

44448
KAQSDCDFY

44449
KAVSSLDGV

44450
LVGRCYDLH

44451
KDGKCVSCG

44452
KCEVKQCDH

44453
SQVIRLACY

44454
ACNVCAH1R

44455
TTYKEMVSG

44456
AAMKLGNLA

44457
GSNRHMIYA

44458
A1SKFIEGM

44459
ECGRLYEHE

44460
KADQFSRDR

44461
AAYKAQVLR

44462
ASWLDMCMR

44463
KYEHTTSPG

44464
QGLKCEQFE

44465
TKNTIVKAE

44466
RSNLLLQHM

44467
EKRMRAECE

44468
AIKMTNRCR

44469
TMAKYNEME

44470
RAERWGGCV

44471
NAKLEHAVN

44472
IMTKVHHVT

44473
QKRHCQTCT

44474
TKRDCNTQI

44475
RGEQKVYCS

44476
SRRLTTAHY

44477
SKKLTQQHM

44478
MGFRTSQPV

44479
AASRGNDCN

44480
KYETENGIN

44481
KAKLETWWD

44482
GMTKNTMGH

44483
ARSFVEKCR

44484
FGKYFWNSF

44485
VMVKWMRCN

44486
IVHKCIDIG

44487
IVHKCIDIG

44488
KAQYGFAVP

44489
KSEFVQSNF

44490
FTKIHWWFT

44491
TKMQVTSST

44492
MRATSHIVS

44493
KSTFAVLWA

44494
GATFCWNCG

44495
VKTQYCDMD

44496
KVGPAADFA

44497
AASFHAMDR

44498
EKVMRKPAD

44499
KVTIGTLAN

44500
AKMNYANVD

44501
SKQVCLAHE

44502
VAKMSFDCC

44503
RPSYKVKCQ

44504
ATHKIIGVE

44505
KSMHTVNEP

44506
VCARTNNLC

44507
TKVIYEKAV

44508
RSGVTYAWA

44509
RTQTFTMIP

44510
KCEMNISCH

44511
KPLQAFAFN

44512
AELRYQFYM

44513
DKRVIVKCE

44514
SNCLLGQCT

44515
QKQGFAINQ

44516
SKLLYSTSG

44517
KITFCWPCH

44518
RMQYTQSCS

44519
GTRTSFAMD

44520
VVKQTTGMD

44521
SATRAFEVQ

44522
SNGLQNIAF

44523
KMQSRLYCY

44524
TIKPSWLCA

44525
VSVRMHEMH

44526
KAMMDHAYD

44527
LASRAENHS

44528
KMQVDNANE

44529
MCMKAQRSG

44530
TAMTDVRDR

44531
RINRNNCRI

44532
MAQLLYSCQ

44533
QCRRMEGVE

44534
AMQRALINA

44535
DRKTQMIST

44536
NGFIQYDSK

44537
SKNAANSWD

44538
QKKFMECAQ

44539
SLFLEKSCG

44540
KVLECHGAA

44541
KIESTSSHC

44542
KHQFTGLHS

44543
QRKMNPDCM

44544
KMSTCMEHN

44545
AIGRNDQTT

44546
NAVVMMSNK

44547
RNIQGISWA

44548
AKSPITNGM

44549
SSNRYGDVT

44550
MRSGFQMST

44551
IAAEFKACR

44552
KLNSTMYVG

44553
LAVRTKQFH

44554
KQTSPLHIG

44555
KVANFFDEP

44556
TKQHFPLGE

44557
MIVKIERGV

44558
ACSKLTCCY

44559
LMYKMHQIL

44560
TAGKTWEGK

44561
KFHNSIVVT

44562
AACRISEVP

44563
ISTKAHQME

44564
SQKSHTVWA

44565
VKFLVQCHD

44566
SVTIWENFV

44567
KANPSNGFQ

44568
TTGWQVQCM

44569
SYVFAQJGC

44570
ATRQLFDCR

44571
GTHRTEMGR

44572
AQNRIGNQN

44573
MSNTAVQFV

44574
QKASYVEFD

44575
FGMKSKQMS

44576
IQREVTWCT

44577
NVNREHRFA

44578
MDPLLIKMA

44579
TRFMTDCMF

44580
VKGQMYSCE

44581
KCEEQKPCP

44582
VCLQFYQGR

44583
MTRASHVMC

44584
RADQQVRWA

44585
RADRCSDLY

44586
TSVTIVECV

44587
SNRTCAVSV

44588
VRTYVQACE

44589
ALMKLWCAM

44590
QKNPTLDSA

44591
SMRRNKYWC

44592
TKAEHVVMH

44593
VAVKHHTTC

44594
FCKTSTSFN

44595
SANLLSDFH

44596
KSVMDDVVD

44597
KGETTQNYG

44598
KAMDGAALT

44599
KMEVRMDHG

44600
TKTTTNISI

44601
TIAREKGYP

44602
AIKPSTWQQ

44603
MKFLLELWS

44604
KGMMKDTEQ

44605
IMKPFMEMW

44606
ACVTCTTFK

44607
AMKSRCRCE

44608
MCKFSVWAL

44609
YNKAYGYMA

44610
QRAHYIDCD

44611
LAAVWVVDG

44612
MAHGFQRWC

44613
STKIQYFCE

44614
EYVAWMKQW

44615
KSVQKECYF

44616
MCAQWCQCN

44617
RAEAHWHNF

44618
NKLIQEGCP

44619
EKAGSGAWN

44620
SAGMKPMCC

44621
RKSQKRMIN

44622
SAAIDRLQR

44623
AQAYYGVCD

44624
MAMRVGSHQ

44625
QKTPTMNEW

44626
NCTMIQRNY

44627
KCNSQPSSD

44628
IKQPYALTA

44629
GQQLIARCG

44630
EERPWYCKE

44631
NRELRMHHE

44632
TEQIFMKGC

44633
QKRLGEEVE

42634
TTYKAMSGD

44635
SHMFQYSHT

44636
TLKVHGIGG

44637
KIQQNMETI

44638
KAMQPECGA

44639
ASYRMGDHP

44640
AFAKLIHTE

44641
KAEPNMALE

44642
FRKSYWYTG

44643
MGGGQASWY

44644
VLHKAEICL

44645
YCTQCRGGV

44646
MCSQCAVLD

44647
RVNTSSRAT

44648
IRYAYDGNE

44649
VATQLIHSY

44650
IWKQLVEMQ

44651
KGGTENGWN

44652
MSQRIWTVG

44653
NKQMFCKGP

44654
MTYRSEFMR

44655
APGKGNRAC

44656
MFTRFMRCG

44657
RCMQSFTFH

44658
TIFRLRKSC

44659
MMSSQMKCG

44660
HTARNVRMR

44661
TGKSWTMLG

44662
EAAKESRLA

44663
GVKLQSYMT

44664
DRRYFTYAG

44665
LCYMLFEMT

44666
TIKCWNWSN

44667
STGMWQCAQ

44668
KLTECPLCG

44669
KTFFYHAPC

44670
RISDAYQCP

44671
SSKYAHYCA

44672
IATVWARCP

44673
TIGRTQFCH

44674
SSHMEFSAK

44675
MIQRVPTVA

44676
SKTDKMEFY

44677
TTAIRMYCV

44678
KCQYCSDMY

44679
TCKPIYHIE

44680
RVNRKQINI

44681
SGGKSQIVP

44682
MFRINTECC

44683
NAATIYREA

44684
ASTKQEYWR

44685
TSALKPACN

44686
TVNLNGESR

44687
SGTFAPFGW

44688
ACAKDRYAS

44689
KCMSALEQQ

44690
ALTMLIKGC

44691
SKTLFHMIQ

44692
VSVIKFKGE

44693
KPAEAHSVW

44694
TRQLYGLHD

44695
KKVMGMLYH

44696
MKTPLAEAH

44697
KVEGCKSHY

44698
HVAMSCNLM

44699
KKDTVNENA

44700
LSGRMAQQA

44701
ATLVKAFCA

44702
ACTHLMCRP

44703
QVTKLPPFS

44704
SCHMIAEFA

44705
SGYFRIQLS

44706
STKIGAKFL

44707
TSNFNCKMW

44708
GSQRIGIIP

44709
QRAGSFGWD

44710
NMRMWEWTT

44711
IAVKAWFDH

44712
KSNKRCGHL

44713
MGLQRMEFC

44714
KPSHYSCAF

44715
KGATDVPSG

44716
LAIKCMDEW

44717
ADNFLVKHG

44718
SRVIRSDSC

44719
MKQFCTCMS

44720
NCLLQWGQC

44721
GAAVLSALH

44722
KKNTAFAST

44723
QRTATLEAW

44724
SKLTVAAVE

44725
SMYRVMPCV

44726
HGILQNCFI

44727
LTVQLLECA

44728
MMMRQHLYG

44729
TAHFSFDTA

44730
QKKLWYGPV

44731
TKLQFTSPH

44732
TKLQFTSPH

44733
VGNFLYRAY

44734
TSGTSYQCQ

44735
RWQNAYEVS

44736
ARVMFDMSR

44737
SGQFFNVSL

44738
VSMMLCASL

44739
KGGLQENFG

44740
TALAKLHVG

44741
ACQPSFAGR

44742
ADKPLYQNE

44743
DWVLWTRIC

44744
VANAHWCVG

44745
SCAKTFAYS

44746
KIQQDIDMG

44747
KLKTQELWS

44748
IVMREAQAS

44749
TKNFTKGAW

44750
LCAKEGSHE

44751
TCLHVWQCG

44752
TSHVMYQYC

44753
AAVMQFMGV

44754
KMDWPNVLN

44755
TNQRIPNCE

44756
RSKDGGNCW

44757
SHTIFLHSF

44758
TGRLQEAYA

44759
SFCFCSANF

44760
TASFFMSVM

44761
ACQRVEKYT

44762
RAIAGGECV

44763
VAVKGWWGT

44764
KRCVTHECR

44765
TAYMTFDQA

44766
SSYRGVACE

44767
FCGSCVAFE

44768
AMGINMSGF

44769
KVVPNFPAQ

44770
TCSKDGSWW

44771
ERSPTFESQ

44772
LKASYHYQR

44773
FANTLANNA

44774
ATRQDGMAY

44775
SPASCQAAA

44776
AMGFSPHER

44777
SRYVVNSAH

44778
MANIWHKGA

44779
TKTPCWVWS

44780
AWGSFIHMR

44781
VYNFSARER

44782
VEAKMCEQN

44783
NCNLYYDHL

44784
TIFTTYSMK

44785
LAGKAQMWQ

44786
KMNGCQSTE

44787
MKQRTWSGK

44788
MMVVWQNMC

44789
FCVYFVDGK

44790
SAMIAGLGK

44791
DRKLFQFSG

44792
TSWTCPRFG

44793
MMKILQHSA

44794
QEVVWFKEP

44795
VSRITTIMA

44796
SAGSCAHMQ

44797
LSLRLERMC

44798
KGTPNLVSE

44799
TMFSCKYEG

44800
VTNMRLEGV

44801
AVKLPMCVD

44802
KENLTQSAT

44803
SAQTHEKFM

44804
TAVIRSNTA

44805
VFNRLIQSI

44806
QKAHIFDVT

44807
NSSKLEKYT

44808
GAGKSNCNP

44809
RGDTQHCMY

44810
KKEYNNGWW

44811
AAYQMTGIY

44812
TAMHTHKLG

44813
NHNMCAVVV

44814
DNRFMFVDT

44815
KWAENTVSG

44816
HALLKMCEG

44817
ESSTCKFGQ

44818
TRLKLDVWS

44819
SAYQLTGTA

44820
ACVASIGMK

44821
FCRIHMMCW

44822
QCRKDWGHY

44823
FCNTLVISA

44824
AWYRTEQMW

44825
TMKSWVHGC

44826
SAAGMNDCL

44827
KITTCWADP

44828
ARRIEEGYG

44829
LCMPLLTFD

44830
NGNVVMKEY

44831
SSLTCSHSV

44832
QGLKEYCHP

44833
AGVGSWQNE

44834
KGAIQQEWW

44835
AKNPYLWYQ

44836
ACSELYRHL

44837
NTWMNLESC

44838
TGGQLFQCW

44839
MNVKKSGSL

44840
MGLPVADWG

44841
FSTSHFHLW

44842
GSVKGMSVS

44843
YSGMFHEGR

44844
GWKPYAYAE

44845
ACCLTMCSY

44846
STRTTSSSM

44847
AAVPCSMQQ

44848
STQMQTVGL

44849
NFFLTAVEH

44850
YAQSKVRMG

44851
AGGKAQAVE

44852
TATLCANFC

44853
TGTVKMGME

44854
TTYIRLGGP

44855
SVFLSQSCR

44856
AKYEKVGMA

44857
ITQIWARVI

44858
QMNRLGNWN

44859
QSSWHSKVW

44860
KSDQSVCDQ

44861
LAASCGHVG

44862
GATTCKQID

44863
YKNHFMHMW

44864
VALVVVMSH

44865
ACVTNLWGD

44866
MMMRQEFCE

44867
IRKYYHNGV

44868
KFAIAQPVS

44869
TAMKQPTLP

44870
SVIIHPSYE

44871
VGAQFAVQF

44872
SKNELFVMH

44873
TACFNALSC

44874
DCYKSEVGV

44875
VKFLVQQGQ

44876
GTQTRFDNS

44877
KSGPVVCKE

44878
VAHISSGAL

44879
ARNLAQGGY

44880
VFGINGKCY

44881
FFTQCARVP

44882
TSWFKDCVG

44883
STQIWSKMH

44884
ISGTNRMVP

44885
GAHTCAKAG

44886
TIVFSLHRD

44887
MCAQMMGVE

44888
TSASCCQLT

44889
KGRWVDPMA

44890
MWKEITGQS

44891
SALETNADQ

44892
TCTTLTNQA

44893
SSVHQMLTC

44894
AGKITWQND

44895
SGKTHSGEQ

44896
MGHMCPGCF

44897
TSTLHFSAD

44898
KYETENGIK

44899
RASVTMGSA

44900
RASVTMGSA

44901
FTTVCAGCV

44902
MRVTYPWAD

44903
AGAFMMPNW

44904
SKSDKLECR

44905
GGTYKLNGC

44906
KTIPVCAHS

44907
KADQGVKDW

44908
KSTPLAEGV

44909
VGKGMNRFN

44910
KINTECMHD

44911
MAIIMSELG

44912
TPKHQQGYY

44913
FREEYRRGG

44914
ACNYFSREC

44915
ATLKVMGHH

44916
KCYTEGTNA

44917
AMLMNTACG

44918
TINRWMVAF

44919
NGNMIWRGK

44920
IVNTFWNYC

44921
SVYLKPTVS

44922
KHALAVSQP

44923
VGNQIPRGE

44924
STHILEGWD

44925
KSVGGTEHP

44926
QKNATAVNN

44927
FCSQHLTGP

44928
QCRVCQFHA

44929
VAAQFSRFE

44930
RSSSETLWS

44931
HKKEFVMNS

44932
TKNPTQLER

44933
TGMVLNGAM

44934
MKSHQMWVM

44935
VPKYWQQAC

44936
LVWRLHQAP

44937
KSTAPEMMN

44938
SFVIHSRTA

44939
SMFIFLHIC

44940
AMASFWECG

44941
QSFIMYGDC

44942
ASLKSMEYR

44943
VGMANMTCQ

44944
MCGHHAQLG

44945
ISQLVWKCV

44946
TAYRAIGPY

44947
QRGTYNQMM

44948
ATVGKVDIQ

44949
KTATNMNSF

44950
GCGLWVYSS

44951
VTTSLMDSA

44952
KSMMNESAV

44953
TIIACPRGS

44954
AAFGMNTFP

44955
VLYKIWQEK

44956
TCRRYECSV

44957
EKAMKSCTF

44958
AQNLHFYAP

44959
AKSQCSKGL

44960
VGTPTCQGV

44961
GSHSHMLDR

44962
SAAYRYCEP

44963
KVNPLNGHW

44964
GRIQFIMCG

44965
KATQEVWHK

44966
KKEAVGIMN

44967
GASSSYQNS

44968
EREAHCNQA

44969
VPHISFKCC

44970
LAHMAYPSV

44971
TALVQYNYP

44972
ESMRGQQWW

44973
TVAFSMCRP

44974
RCAMDIPHG

44975
LAMPCSGLC

44976
GKQFNMGDP

44977
NKTLVGIQS

44978
TANIASVPG

44979
MIVKTVMDV

44980
VTWSCSELA

44981
IGHGVQTAF

44982
KCNNHVNSS

44983
TRVPTQSIF

44984
TCHVSQDCT

44985
NTFMSTFLE

44986
TYQFRNRSE

44987
SAAWQKRCP

44988
RKEEFHACN

44989
GSMFTCYLA

44990
GESRLNEHW

44991
IKTPYCSSG

44992
FANSTQGCA

44993
AACLNTQYR

44994
FISFHEGYR

44995
HQMKAMNLG

44996
ACMGIKHVH

44997
IAQQTWQFE

44998
ERYMFLNME

44999
HKTPYPHLR

45000
RAEMLHEAV

45001
SGQHMPGCM

45002
GYMLKSDVQ

45003
YCVQTAFMH

45004
FRAYFQSAQ

45005
VSFTWNRCI

45006
ACASLINQM

45007
TNRHAEAYI

45008
SCQPAYGGG

45009
YTNLVIGVW

45010
MIFVISVGN

45011
TFYVQVNDK

45012
SANKVVHTD

45013
KCAMALIHK

45014
MLFWFRCLL

45015
DIEKLHHVF

45016
RYVCPLDHY

45017
SCCQEANGA

45018
THYFRNMHG

45019
AFVPFGNDL

45020
LGKVVYQAY

45021
ASKLDTALT

45022
MCKLLATSQ

45023
SLKAEYSGN

45024
NTQTVNRMF

45025
QVHKGWACV

45026
SQRVWEQVC

45027
THTKAQIGH

45028
MGSRHFHFP

45029
GHYVCSMAY

45030
DKHGNANWN

45031
GMSRQSIAD

45032
KAEHFNVDA

45033
IKMNHHIQG

45034
IFGKQHRLH

45035
ATGTKISVE

45036
MAKINPLHG

45037
TTRHCAYAQ

45038
MIQFDMMHG

45039
KTKSGAVFT

45040
AQKLMMGQG

45041
VSNLTWPGP

45042
ESKIVRYNN

45043
SAGRWCQTY

45044
NGALQMVQW

45045
GMFIQAVEA

45046
AGNTTIKFE

45047
AYFSCMHVN

45048
LGMTCVAHG

45049
VAILALRCH

45050
TKQQVGDCW

45051
TGLMLQQIC

45052
NVHFLFNCA

45053
NYRVAEWVA

45054
AARIACFDS

45055
VCVTKQMAC

45056
DRRPVCCYT

45057
KNDKGQCYF

45058
LG1KSQDAA

45059
QSALMSKYW

45060
TVGIKFQEM

45061
YTKFMSGGM

45062
AFRSPNVMY

45063
YSSLLALRA

45064
TQAKLQCAE

45065
TANHATMCD

45066
SPQPILDLQ

45067
TTVLIQICE

45068
SAASLIPTL

45069
MKVDSTFIH

45070
LGAANQMCH

45071
MSTADMIRI

45072
KAALMGWEK

45073
NCVVLQKLH

45074
DRAPKLSWE

45075
VCNQFQHWH

45076
KISFEVTEY

45077
TFYIWNHCD

45078
MCKQIIQGG

45079
SCSVWRCMS

45080
VSILMYQNQ

45081
TSMFYEMCN

45082
SYKHCSQVG

45083
FAAQWFWTG

45084
TWATWLEGQ

45085
SAWVCQFIQ

45086
NAMSQVSLA

45087
SNQFFPQSG

45088
TGTLMGSQG

45089
MCNYSNSGE

45090
TCSQPPCCG

45091
GHYIFNCSQ

45092
IGHRYYWSM

45093
GCHTIQFYR

45094
NCNMYHYFA

45095
GAQPRVVLS

45096
KATCYMDAN

45097
SAKQWTTVE

45098
NANTNGHMF

45099
TAGQPGILG

45100
WGTYWFNMP

45101
VSSKSVHDV

45102
SPQSFPQFE

45103
SKAANNQCC

45104
MGYQPAVGN

45105
AEMRTMIQA

45106
GGTFIQGQG

45107
KGTDDGQWQ

45108
GCNIQNMAA

45109
AGMGIHAMP

45110
AGQLVQACH

45111
TRDTFFKMG

45112
STYVQMDGG

45113
TSTTKMLSC

45114
TANATVFHS

45115
TCEQEWKAR

45116
SAMPGASMN

45117
GASYANNVV

45118
AAFTHMGTF

45119
AVKNWEVHE

45120
VASFWHCGA

45121
AIYPTNFVK

45122
NPALHNQCQ

45123
QKMAVCDMP

45124
KLHQRTWAA

45125
SAKYHMDVE

45126
QCVANYIAR

45127
GWHVQSCAM

45128
VPITCYNHG

45129
DKQPHNTGF

45130
AQNGMAICE

45131
VLFQTMVED

45132
TRKAVCQPV

45133
VALKMFCTY

45134
GANTEVLAG

45135
MGAVTMVGS

45136
MSYLKTTCH

45137
TMGLHQDHP

45138
LMHPLGTCA

45139
TMGLAGKSA

45140
YGQHWNPLT

45141
ACYQLPSAN

45142
ATKEKHPFY

45143
MMKNTQFCE

45144
KTNFRCEYY

45145
MVAKEIDGP

45146
MGSLQNMEF

45147
RAQPQERVV

45148
TKAECMPYQ

45149
VAMFNRLDR

45150
KAHDVLSNP

45151
VTNLLMRAS

45152
VGWACQAFY

45153
MTNSSHKWS

45154
KAAWSEDGR

45155
ASGLYYSHG

45156
MTGLQRVAV

45157
KPSTKMTLR

45158
IATFIRGFY

45159
ARGVLNLED

45160
FMNMMGSCS

45161
FAHQHPGCP

45162
NCVSLAAHW

45163
DSMRCTPVP

45164
FDRLITQGE

45165
TGLADQDWW

45166
NSIASKKSM

45167
MFYYAFTCG

45168
NSMVGGMSE

45169
LV1PWARVE

45170
FAAMACDMT

45171
NSTSIFDAC

45172
IAAHVVMCD

45173
SAVPVCSCI

45174
NKNSTNQWE

45175
TRFEYWSNS

45176
QMTMWANFY

45177
SPNPLPWCD

45178
KAINPLKAT

45179
SVSIFKGSN

45180
GNYISSMAC

45181
YATTTMSML

45182
KVHTCGMNY

45183
NWNMWLKSL

45184
FAAITRFQS

45185
SVTAKNLCL

45186
VCTICPVQM

45187
NIKLTHFEM

45188
VTQLSCVDT

45189
GAGMEASMD

45190
AAVSVHKCL

45191
TAALWK1EC

45192
TGQPYYNTA

45193
AILPCSTPK

45194
EKQVYIVRN

45195
MAKPAHAED

45196
KVHEASYFP

45197
VMIKTWASL

45198
HSATAVREG

45199
GQIFMCQHI

45200
AVSLHEHLQ

45201
AAMPVMKCW

45202
LGAMMALHC

45203
RCGLSSDAT

45204
LALQIQENR

45205
TARTPNDFE

45206
MGTFKLDFN

45207
TSTRANMLC

45208
AFKLVSCLE

45209
ISLPCHEVK

45210
EKGYIFVID

45211
TTSVLTGTG

45212
FARQFMEYW

45213
FGHFVGLTG

45214
STMSIVRTP

45215
ASVTNVKGM

45216
MSKLHAGIH

45217
GIAFAVHGI

45218
MFQTTILES

45219
MAHSYQSFC

45220
VTQMIANVQ

45221
DRFTQQSAK

45222
VQKYVTQAF

45223
SASTTRFIG

45224
VAATAYMDE

45225
RGEHFKCSY

45226
HSNMWLDQY

45227
NKTSNIMYH

45228
AAYSPMFWR

45229
SVMPCVDAY

45230
TAVPILDWT

45231
SGMLFIYSG

45232
NRLTIHTWY

45233
VAVITQDYP

45234
NSRLIRPHG

45235
RTIHPMRFI

45236
SKFEFQNAA

45237
MGRPVPSHS

45238
MAHNLVSLS

45239
KMMCNPTLL

45240
NIKLTHCNF

45241
AASSWVLME

45242
MCLQPYNCY

45243
VTHSTPQCV

45244
TSKVVSDAP

45245
ASSIVETAA

45246
KLHSGGYLC

45247
TAGIIYFYH

45248
SGKQLQNTY

45249
RFTMYSEWL

45250
TSRPISHEI

45251
SGMLKPMYS

45252
RTTMGVKME

45253
TCLTRHSQG

45254
MADKKWRYC

45255
SFYIHQEAN

45256
ASYMKSELA

45257
SYTRAVECN

45258
SVQMEGLGW

45259
GTKHTMQQC

45260
TSQHLQVLC

45261
MHNRIQKGG

45262
VIKSYVSTA

45263
RLQYAGQFL

45264
DKQFIAEWP

45265
SHEHCMKMC

45266
KTEDYQHHQ

45267
AVYMAGRYM

45268
GATPLCDAN

45269
GAYHVTNQT

45270
SHQHHPWGA

45271
GATIVMTGH

45272
TAVAQVDPV

45273
TARNLMGSS

45274
KGNHKMKHE

45275
RIEDTRHNN

45276
MCHQWMSCT

45277
STHYKIWAG

45278
TMKAMMEVP

45279
ALAHEAGKR

45280
SSNIIQYHE

45281
TQKSGSFNN

45282
ALKLAQSHY

45283
VCSHFCNEQ

45284
MSAWKMRDR

45285
TVTVWPVCC

45286
NSKHLNTYV

45287
VSHWITKVR

45288
TSMPYKAYG

45289
VYLQHNCCW

45290
AANVGMLGS

45291
KLQMNYEEK

45292
LGGFHVSAS

45293
GATFLQFDG

45294
SQMLSEHLE

45295
TPGWVNQIA

45296
TGTFVQMDK

45297
ATKQTPWAI

45298
TMCHEMCSL

45299
VGAACKMVA

45300
GQVTMQRCC

45301
TWTIKQIGE

45302
SARQVLDPD

45303
VSKSWLNSF

45304
GGMFLAEGS

45305
NYVTLGNHM

45306
ISKHCHDQG

45307
SAQSALDAL

45308
FAHGHSKAR

45309
AYFYLGAEF

45310
QKTQHPVWN

45311
NLCVSLESY

45312
MSQRVSVPS

45313
KEKLQMNHM

45314
HKGFTALFQ

45315
GTRTRIQQE

45316
ACTNMFAYD

45317
QSMHCANLK

45318
LGSHYCNYA

45319
VAMLEFQSA

45320
SSHMLELHI

45321
HGVIPMNW1

45322
SGAKMAWVA

45323
SQWMLPCGE

45324
MPTHMWDAP

45325
NVMMQTYSR

45326
TCOGIGSVT

45327
TCRVAEFVP

45328
MGVMLTSEW

45329
AQNLYSAFW

45330
NHQLTYCSG

45331
MIKLRHWDV

45332
HCGPCLKGD

45333
AAQYQAIGV

45334
TNKMVGHGH

45335
RCGVQQGSN

45336
TFMRNEWTY

45337
AKYICONYC

45338
GCARNNGAI

45339
GCGKLEIHF

45340
TAVQTHTGA

45341
ARMPRESAD

45342
VMIPRMGCG

45343
YGYMANKMK

45344
NSYTNFKGG

45345
VLYSWWHFD

45346
KCFEDLCST

45347
VSNPMSCKK

45348
SCQHFMDDP

45349
VAAHWKMSN

45350
AAVAMQMCE

45351
ISQHCHCKL

45352
QWKQAQCCT

45353
GHRSRLSGA

45354
TCASLQSNQ

45355
KSAWMYGES

45356
RSATDYGNH

45357
ASGTKWAHK

45358
SPVTLHNGE

45359
RKERVIKEK

45360
VAARYTAPN

45361
AVKQTIYMP

45362
FKILKPGLG

45363
AAIPFACMG

45364
MCRLWHDAW

45365
AQKYILVGQ

45366
IQIFCFWCG

45367
NRKGQHVQL

45368
TKKTCVTWG

45369
TVNMWGADD

45370
GFKPEMRMA

45371
HRKHKFVPI

45372
KLSWEEYVR

45373
VIAQQGNAR

45374
TTAHVYDGM

45375
VKQMKLEQH

45376
MCVMSQEHL

45377
GRMQTSRCG

45378
TMRGTGAQH

45379
HYRPFWSAC

45380
ELFMDHRSW

45381
FSQPCVNGQ

45382
RPFTNEGLC

45383
VGMSPFKFC

45384
TVKIYGEAS

45385
AYRYEYCAI

45386
IGITWYGDT

45387
ERARSHECT

45388
KIESIVCNT

45389
MGASNMRFV

45390
VANYCNRPE

45391
AGQTFIYVH

45392
LAMMCHADQ

45393
MHVMSGHLF

45394
AMRQANAGV

45395
QYSIWNFDH

45396
TTHLGMNHF

45397
MSLHKCDVR

45398
NFAMSMRDH

45399
MKYEKNSCW

45400
GSSYLNQPT

45401
MSRHCSMHD

45402
MANATSREN

45403
EAYRNKNVC

45404
KNNTKKQWP

45405
RVAFTLESF

45406
AYKQIHDST

45407
NATGAPRAW

45408
MKHDVKEWN

45409
FSTQQCCVE

45410
EKKFHLKYQ

45411
ALGQFATCD

45412
GGFRFTAQA

45413
IALSCQSHL

45414
MSQFGMLAG

45415
AMKFVNYFY

45416
SMHLHMKDL

45417
ISTLLSYCG

45418
TEMSLPGME

45419
NAKLGDACG

45420
AMKKEGEWG

45421
SAHSTAVYR

45422
VTYSCVCTH

45423
GSNWIYEEA

45424
VSVIFPQQG

45425
MVIREWCQC

45426
MVMRQLFYN

45427
VSYYMYEAP

45428
CVIRAHQLC

45429
YGAAKQAMA

45430
AVTSSLRDL

45431
TGQVQGFHY

45432
DKLMFGEWC

45433
AASLKPWMY

45434
TRGLYMWSY

45435
LSSRSEHPN

45436
NASMYGAQC

45437
SIFFFAASP

Example 21
AAV5 Variants with Liver-Detargeting Tissue Tropism as Identified by Machine Learning

This example describes preferred properties of engineered AAV5 variants that display liver-detargeting tissue tropism as identified by machine learning methods. The same bioinformatics pre-process, machine learning methods, and analyses as described in EXAMPLE 19 and FIGS. 19-FIG. 22 were carried out to identify preferred properties of engineered AAV5 variants that display liver-detargeting tissue tropism. FIG. 27 shows a set of top 20 positional features contributing to model output probability. Shapley Additive Explanations (SHAP) values can be used to interrogate the relative contribution of features to model predictions. As shown in FIGS. 28A-B, these features were further compared between tissue targeting and non-targeting variants. FIGS. 28A-B show a comparison of top predictive features positionally. Features were selected if they were found to be important to both the HGB & RF models. Summaries are shown of the features in the top 1000 ML-predicted liver-detargeting variants compared to random 2% liver-detargeting variants.

Favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an AAV5 VP1 capsid polypeptide, which are associated with a higher probability of liver-detargeting tissue tropism are described below. Any of the below machine learning-derived positional amino acid preferences described in TABLE 42. Listed below in TABLE 42 are a summary of positional features shared between the top 30 important features for liver tropism extracted from two ML models (HGB & RF).

TABLE 42

Machine Learning-Derived Liver-

Detargeting Tissue Tropism Rules

Low solubility at position Xaa1

Xaa1 is selected from D, or P

Low mutability at position Xaa1

Xaa1 is selected from C, K, or L

Low solubility at position Xaa2

Xaa2 is selected from N, K, P, E, or D

Low hydropathy at position Xaa2

Xaa2 is selected from D, E, R, K, H, N, or Q

Low charge at position Xaa2

Xaa2 is selected from D or E

High number of total potential hydrogen bonds at position Xaa2

Xaa2 is selected from H, N, Q, D, E, or R

Medium volume at position Xaa2

Xaa2 is selected from D, E, V, P, N, or T

Low solubility at position Xaa3

Xaa3 is selected from P or D

Medium volume at position Xaa4

Xaa4 is selected from D, E, V, P, N, or T

Low solubility at position Xaa5

Xaa5 is selected from N, P, E, or D

Low solubility at position Xaa8

Xaa8 is selected from K or Q

Low hydropathy at position Xaa8

Xaa8 is selected from K or R

High surface accessibility at position Xaa8

Xaa8 is selected from E, R, or K

TABLE 43 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that exhibited liver-detargeting tissue tropism and comport to one or more of the rules provided in TABLE 42. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 46438-SEQ ID NO: 47437, as disclosed in TABLE 43. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 43

Sequences of the 581 to 589 Region in AAV5 VP1

Capsid Polypeptide that Drive Liver-Detargeting

Tissue Tropism

SEQ

ID
581-589

NO
Sequence

46438
IDGDDEPRA

46439
PRSDDDQRF

46440
WDCEDERRH

46441
DDPDDLECC

46442
PESDPERTN

46443
REWWWCLRI

46444
DDHEEVFKA

46445
DEDECEGRM

46446
AEIEKDPRW

46447
PEPLPEPSK

46448
YECVERDRF

46449
PHPQATGPS

46450
QEHHPRDRW

46451
CDTEAEFRR

46452
MEPGEKSRP

46453
LEMGGKTRT

46454
CHPHDEHKT

46455
MDLFPPHRH

46456
YEDNDVMQI

46457
PDMKSVVKA

46458
KWPIEWGHY

46459
VDRLPPNKS

46460
MDWDDAIRG

46461
WDRPPVMRP

46462
YDWGRVTPN

46463
REPWQTTEH

46464
PWKHEPSEF

46465
FDSHPKIER

46466
LRDRDGNRM

46467
RRGMEPTKH

40468
GDERPGQRN

40469
PLDRDQATQ

46470
PKNCRWYRT

46471
PECGDRGDR

46472
WDHEPETKH

46473
EGPDPCNSA

46474
PECFPKGRN

46475
CDHDTKHPT

46476
PADFRDTRV

46477
CEYDDESQF

46478
DDEDVSEEI

46479
HDKPGKPPY

40480
KNPPWICVW

40481
PIEKDDNVD

46482
PKMDKVEPC

46483
PDPCTEGTC

46484
ADNCPKNDN

46485
PDHITKCDE

46486
DDHDDKLVA

46487
WEDEPPWKI

46488
PQPDENYPW

46489
FDREKRDGN

46490
GNPHEGQKF

46491
PEGSMDRRF

40492
GDWSERSIT

40493
PERWERPVW

46494
NDLCQDQRV

46495
TRPDESPRQ

46496
WECADRRPT

46497
YESCNLTRD

46498
PPPVHNDRS

46499
VHPEACMRN

46500
PDQVDRNTP

46501
REPNSGLVY

46502
EEPEFTGRG

46503
TDHNGEIRP

46504
CDRDPSWRD

46505
QDRNRVHRQ

46506
IDPYDCDPT

46507
PDQDDMMHG

46508
YEYRPELRA

46509
CVPEPWDYR

46510
MDPKHHCES

46511
QDQCDDKIE

46512
MHYCDRERK

46513
PPDRCTAEK

46514
RNPSQDQKG

46515
CDTHDVSKD

46516
DVADVPDTT

46517
DEKPMDGRG

46518
PDTSPYCKS

46519
QLPDKWPKS

46520
YRCCDDSKT

46521
PAECISDSQ

46522
PDPLYWIMN

46523
LEEGDHMHM

46524
GDHEPKHEH

46525
WEVEQRPRH

46526
CECWPEYKR

46527
PFPLNPERH

46528
DVISRNGRC

46529
PNEDKIKPD

46530
YPCDDTKPE

46531
DTDEENSPH

46532
IDPGMMFRG

46533
PEQRDRLGF

46534
DEPSDDDEE

46535
HHPPKQPPN

46536
CRWEDYDSR

46537
CPPQADRIQ

46538
PQPQFDMMQ

46539
EFPDGGHKM

46540
CNCPDQHPD

46541
WPKCDECRM

46542
ADNEDARPK

46543
PICRPDTDV

46544
APCHDKDRE

46545
LDPPDSACG

46546
VETEEKMIL

46547
MEDTDERSN

46548
CDRDKRHDH

46549
GEEADEYHM

46550
RDIWECIRR

46551
AECQEDRRG

46552
TEPDTVDES

46553
PDPGIRWPM

46554
PRDVDDSAY

46555
VEEDQSCKT

46556
RQYFEDNAG

46557
YDQWERDIH

46558
RKPEWSCMV

46559
LNPQSPEPK

46560
PPCKMKIDT

46561
PNDWITDPH

46562
VERMENPKH

46563
DAPEPHAQK

46564
PQEPEEGKT

46565
HDMDDVLNH

46566
VDCTPDRRN

46567
PHQARKNKP

46568
CEDEGKTWL

46569
PFPQMNSNA

46570
ADLRPSVDY

46571
GEPWDTRIV

46572
YKPKFDAMG

46573
PDPQQGGTA

45574
MDGRDKVDP

45575
CEDQRQPME

46576
PDFHHKRDF

46577
NEGCRDEKS

46578
LQPNAVGKT

46579
PKLCSHOTE

46580
QVPDEGYSW

46581
NEPDKGYTM

46582
DLDDVCNKC

46583
QDTDEGDQC

46584
FDNPDEYVT

46585
YPDCEGGKM

45586
DVNKPDEKW

45587
EVNDDHAKP

46588
PHQRNRMDP

46589
QEPDSNRGV

46590
SLPQPDTRY

46591
FEPCPRSCW

46592
KEPKMEKRG

46593
PGDHRFYWP

46594
ADTANILRH

46595
KPCRDNQDV

46596
LPPCYCQQY

46597
PECIEGPNH

46598
HDNKDVNRN

46599
GEMTRCFRN

46600
PDKEKVMPA

46601
WNSPDDERV

46602
VDRDDGKDY

46603
DFEDELSKV

46604
YEDGDH1VE

46605
VEGEEILRI

46606
CEWLDLMRT

46607
HEAEDWRPF

46608
KMPCEGEFT

46609
GDPWEEWRK

46610
HERWDANPY

46611
PGCCDLTNI

46612
PIIKGEPAD

46613
PDTDDIIHW

46614
EDLNEKSKS

46615
EVDDGNVGE

46616
DDWNEAKRN

46617
PPDMPMSAD

46618
QNPWDTDNH

46619
PDTWAYAPH

46620
TEPEIRNWF

46621
FPSDPDGEH

46622
PPPVVSPKP

46623
FDPHNPCFR

46624
ANAEPVHRC

46625
LVPESQMHG

46626
WERCPELNQ

46627
MDEHREQRS

46628
SDPEKYPWD

46629
ERDDPRFSN

46630
FDSSQEWKL

46631
CRKNPT1HM

46632
EAEEMGLKH

46633
CKYLSHERL

46634
IDPCDELFW

46635
CKPTSDMKE

46636
RPWEWWIRK

46637
FEPQVEANT

46638
LETAEKSKS

46639
FDSCSRHRW

46640
DNTDPDGSW

46641
ADPOSVCCT

46642
PHCQTVSDI

46643
IEEPDTCLV

46644
PEMQNDMDS

46645
DHLPEDKRY

46646
QENDANRPQ

46647
KQGCEKNPM

46648
HRCDPDMGR

46649
DVPRDRKEN

46650
EDNNNDVKH

46651
IDFRPPNNT

46652
DFPDKGELG

46653
CIPDVAIMM

46654
MEDHTAPRW

46655
PENQQNHRD

46656
CRKSETPWD

46657
RIPPMMIML

46658
PQPWNQQAF

46659
CEIQGGQRY

46660
VDFHDGFND

46661
WDDHKLWRW

46662
RDPFSPDPM

46663
HLPRDDPVM

46664
SD1PPNCRK

46665
DPPRSKDDS

46666
NDPDTSRTA

46667
IDWPDNQKV

46668
YDCENILRT

46669
VEKPSEEPQ

46670
LKPLPHFRE

46671
HDEEEKNKA

46672
PHGWEENDP

46673
PLVGMDHRE

46674
HRPEQLHEG

46675
PGPHDQGPY

46676
EVEDRSGWC

46677
PEPDWVQEC

45678
CRSEGWIHT

46679
PETTSRWKW

46680
RDYQDFICY

46681
CDFQSSWDQ

46682
IECFPRESI

46683
PERASEMFW

46684
NYPRDHRQH

46685
FEAHDRPSN

46686
WEMNDMODY

46687
LDTCEMLKQ

46688
QFPEFDRET

46689
PVPHPEKEF

46690
EFDEFNTRG

45691
GDQAGATRF

45692
EPPTNDART

46693
AEVGTEHKV

46694
DDPSGQLRL

46695
EDPHSKKEH

46696
CRKEQGQRT

46597
PEAGGKSWD

46598
PWDPEEDGD

46699
ENPEMMGFE

46700
MNPNEKGGL

46701
PDAAERKGV

46702
EPPDGQVDA

45703
FDANDCRPS

45704
FRSHDREEN

46705
IKCEEPMRL

46706
FIPDTDNWG

46707
IDCRSRLEE

46708
CKEQPTERF

46709
DDMEDQQQY

46710
DKPEQLKEH

46711
PDSHKQPDC

46712
CDPSVEDRV

46713
PEGCHHQEE

46714
FDPCQGGEQ

45715
NDEDNMSRH

45716
PECLPKNSC

46717
AEDASRDLF

46718
CNPRPQMVG

46719
CDDHAKRNA

46720
MRNYPDARF

46721
DNPPGRRKP

46722
PDNSDATHQ

46723
MDCKEWNKQ

46724
GNQCELNPH

46725
PNYKKRSTV

45726
LRCCGMETE

45727
PPDMQSAKW

45728
DHPSNDAEM

46729
RIPGSRPPQ

46730
WDPNQGGAT

46731
PSCAPCLEK

46732
IDSHCKMRE

46733
CEYDGTMDT

46734
EVADENGNY

46735
PQPNIPVRW

46736
WDYNSIFPG

46737
IHSNDOPKG

45738
PHDLKHQRS

45739
PIPLMMHDE

45740
FDCLPDREH

46741
PNDWHIAYG

46742
LDTKRTFKK

46743
CDRASDEGK

46744
DDSSQEKRW

46745
PEHQDEQSC

46746
TEPETYPAT

46747
FKDWDHHTV

46748
WKTCPQEAY

46749
EEKDEQERA

45750
WPNTPWEPH

45751
CPTGDQQKW

45752
RPCMQQDPH

46753
CDAGLISKM

46754
EASDMSIEK

46755
AEPRAIWRH

46756
FHISPKEKW

46757
PIPPIKTHS

46758
PDPQNHQFV

46759
TEVEQLPRA

46760
IDPNNLTSM

46761
LGDEPWAKF

46762
LNHCDKEDH

46763
WEDHDLHRN

46764
PALYRRKTD

46765
NEDIPDPPH

46766
WEIQDVIRV

46767
PENWNTGKH

46768
PDRYPAFEM

46769
HNPWFSCRW

46770
INQKETWKF

46771
PVPYSGPKQ

46772
YEREPRMFH

46773
LDEEPVFVG

46774
PVDADHQLW

46775
RPPHVGPAL

46776
ADPTTETPH

46777
PDWWPRCKP

46778
YQPRSTPPG

46779
NNGDDRRQH

46780
REWKDNCKQ

46781
PDDWSGTLY

46782
PEVYPGGTT

46783
PKCCCKCDW

46784
PDCRWGEDN

46785
CDLPMSWRV

46786
PEDCDDRIS

46787
QDCEPEYDF

46788
DVSEDHFND

46789
PKQLDQPTN

46790
EFHDMDMVI

46791
CEHPESKDV

46792
TEHKDHFET

46793
PSNEPSVKT

46794
GEVTGHERN

46795
PNPQSHLHI

46796
CPRCDSRPA

46797
MEDNTVCSV

46798
QPPRRAMKM

46799
SLPEEHPFE

46800
ANPLFKCIS

46801
DQSEDLDQG

46802
DDPEYTLCL

46803
KFPYLPHRN

46804
PVGNKDSRY

46805
WDVDHKLRM

46806
PQHGAETPH

46807
PHREEDDHC

46808
CVPNPLGHI

46809
NDTGPHGRH

46810
DDTENLRE1

46811
VDPWDPAQP

46812
CDPWDWEIE

46813
EQPHDPSST

46814
PVPCNTVKF

46815
KMPNELGYY

46816
KEREERRFW

46817
DFPEPYYGF

46818
QDQRRQKRM

46819
PRHTMLGPS

46820
DDREKCNWM

46821
FEQWKDQKV

46822
LRPCEGEQD

46823
CPTKPRYPS

46824
PSESKLMNY

46825
CDKNWILQQ

46826
PSPPPGHNA

46827
IKPEPNWSS

46828
PEETRHNDC

46829
DDVQVEPKW

46830
PQNGMLHYS

46831
FDSRHIEKP

46832
HEKHSLDRY

46833
ETPDDWNCY

46834
CDAEAGFRR

46835
EEFGPTKPM

46836
DVPQKAHND

46837
IHPCEGSQT

46838
VDLQQEDIY

46839
AFPDNSKTE

46840
EELASSDKY

46841
EEYTGECRG

45842
NIDDKPSMQ

45843
WPPEPWCCC

45844
IDPFCAVRI

46845
DEYCQWIRS

46846
PODEWQNRV

46847
LNPHVSQIW

46848
MDPYEVTSC

46849
WDAFEPMRY

46850
YDFSERKCF

46851
MDPNHVHAG

46852
FDCFDRNLC

46853
YDTDDAQPN

46854
WDPHKENTM

45855
RDPNAQGWT

45856
EWPHIDAGK

45857
EMDEKLDGK

46858
CQPPEEGMV

46859
PPANDVPHQ

46860
IECSPRETI

46861
VDIHKEMRV

46862
PAEHDPNQV

46863
PKPPPHYEQ

46864
PGNPENDPN

46865
DWNLEDDRA

46866
CEYWHQVRP

45857
HDPCTQIKP

45858
CQPPEEGMV

45859
PRVDNGNQL

46870
PHESCECSG

46871
ADTKRKRRT

46872
DEPRDWGAQ

46873
INEDAQEKM

46874
LNSEPKCRA

46875
PMPNTGDEE

46876
TVDEQGIQH

46877
FDGCSKMPM

46878
AEPDTIDWD

45879
DIWDTEVQE

45880
PQDLKPGNS

45881
IDRCDVLQH

46882
HDPWNVLDW

46883
EDPSCPAAQ

46884
MNTNENCRK

46885
CKHVDWDSK

46886
SEGMDRDRC

46887
YKELDRDPQ

46888
PRLQSRKYL

46889
PGLWIAPDH

46890
PYLHSKQDK

46891
PQCHDHRQD

46892
EQPTQTPFS

46893
HPPKRHIPC

46894
VDTQKTERA

46895
DVDDLVEIM

46896
CELNGKIDL

46897
EEENCEQGN

46898
DEDERTLDI

46899
YPPSQEIKT

46900
PHSEEGREI

46901
EQPECETNV

46902
PHCNDGSEP

46903
CECWGCTRG

46904
PNDFTTHGA

46905
CDTNPQDQV

46906
PRWQSEECQ

46907
VEATPKSPN

46908
PEEGSPAEY

46909
DTFQADKRY

46910
PYEKEKKKD

46911
DFYHHDDRG

46912
DTEEPVVNP

46913
PNEPNEEHK

46914
VRIYDRCEM

46915
LRCDPWLPQ

46916
LEWSEKINP

46917
VTPEETVVI

46918
PDPPKQYED

46919
HPGHDRPDF

46920
EHGHDDQHR

46921
DVENQDWAM

46922
PHSEKYAKH

46923
PDCLDEGAA

46924
QDCSDWTKF

46925
QDHENNLKT

46926
ADSQDPLQQ

46927
PLESHESEF

46928
HDDTHRIVN

46929
CNEFKHERR

46930
NEARDGEPF

46931
PQNPSEWPP

46932
CRCDTKIRE

46933
QEPLSKHGV

46934
INPEIRQIE

46935
YDLSPKMYY

46936
EVSDYQQWV

46937
RPPHKMSKE

46938
EDRCPGNPG

46939
LMPDPVQGI

46940
SIDRDHPKE

46941
ADPQVEWAM

46942
CQRQEPDRE

46943
DAMEDSKTC

46944
PDPKAWSNY

46945
PQPKQCMCW

46946
DMSDEGQKT

46947
QEPFTESKS

46948
YECDSEHEV

46949
PHGTEDWDY

46950
VRPHDVFYM

46951
FHPDKLMPM

46952
QDRRDDHGG

46953
LKLRDEFNE

46954
CEVQDEWTG

46955
EVPNNHHYT

46956
PEPSWKYRP

46957
CPWCDTQTY

46958
KDKCDYMTV

46959
CATDPIHRV

46960
CESFPGPVM

46961
PHVCGEGKI

46962
LREEELMTR

46963
WNPANMFNI

46964
CDPCMECGS

46965
LHPPPHQCP

46966
NDFSDEPCM

46967
IDPPGTESM

46968
PYPQQSHLE

46969
VRPFDDGPM

46970
TEPSKMPIW

46971
PTMKFKDRD

46972
CHSGDILDT

46973
PPKMMDNEF

46974
PSWGGREEE

46975
LDQNNGPET

46976
NETPDAWNV

46977
WEVPRWPPI

46978
FDTDQRQSE

46979
WIPCQFSKI

46980
PLDHRPHTF

46981
IWEDMKGKF

46982
WRIRPQQEI

46983
PGHFEPTNY

46984
MEPPDMQCP

46985
DVPFVNASD

46986
LRTCPVWRQ

46987
PIECMGQHW

46988
VDPSNAAFK

46989
DPPIDMSKD

46990
WKPGYGAKP

46991
CPPSETRVP

46992
MQDRDTPTI

46993
RHPEGRRVD

46994
PFKQGRENP

46995
LDVTPQPEG

46996
EVGDMAHQS

46997
PGHHPLEDH

46998
HLPVYAPKD

46999
DVEAMDFGT

47000
CTPGFWSER

47001
IDHIGDDRF

47002
PQSHKLIES

47003
CNPPCTPFN

47004
DVCDDHKPT

47005
PPPLAGWMW

47006
YDPNMFPYT

47007
IKPKDQAPL

47008
ERCHDTEKY

47009
CKKESTCAL

47010
PKPVQPHND

47011
DPELDIPEE

47012
HEDTHVLRQ

47013
ESEEHRVMS

47014
PMEEMHSHE

47015
DEEVEGMSV

47016
DEPFFAIRP

47017
YEMMPNIKY

47018
EYPENSSHF

47019
WNPGPVLTH

47020
GERNGGWRL

47021
PADIDDDGN

47022
QRWSDGDKC

47023
AKICDADQL

47024
VEPRYDDPC

47025
PSEHWGCDG

47026
RNCCGPNPY

47027
PEQMDRAHG

47028
MVPDQYQMS

47029
WVPDLEDVQ

47030
IDKITWQKH

47031
DDWTPYLNE

47032
PDQSEMTKA

47033
FHPDRRNPT

47034
QDTNETNDV

47035
WNEIDEGRW

47036
DDNQSRGCP

47037
EIQQHDNVI

47038
SPPDWWPFD

47039
MEPDMRQYS

47040
DIEFLDCET

47041
LEMQQLTKV

47042
IEELEEESH

47043
EDWRKDNNF

47044
PDMKDFKQE

47045
WPWKPPYSD

47046
LQPDRECIG

47047
WDMMFPCKW

47048
PHDPGECYP

47049
PQDHMVNMQ

47050
DNDEQLGGC

47051
CKPHQQRIH

47052
GDMNVWVRY

47053
CEGAAWPQI

47054
SVDDFSSHV

47055
VIPESMNHA

47056
DDPCMRCNV

47057
KKPWDDHTR

47058
SWPQGDYRF

47059
PVCPMKQRS

47060
DGEEDWMDP

47061
MESEESVKP

47062
QDTESKFRD

47063
DDMDLEIKH

47064
ENPSNWHDW

47065
HHVSMDNKH

47066
NRDWFEWKM

47067
CQPESVWHP

47068
PPCGKGTLC

47069
PYMLCNSRS

47070
GEPEAKGNG

47071
VNLDEMPKA

47072
ADRCEQEVL

47073
VNPGQESGE

47074
CMPDEMTWS

47075
ENHENCDQD

47076
MDDMDRRNS

47077
MDWCEYHRS

47078
PDHERVEYC

47079
IRPIICRKG

47080
PPEHCEHTQ

47081
LEPPGLDSM

47082
LKTCPEGNT

47083
WDKCAWAWD

47084
MRVCDELEP

47085
MRHDPFHRR

47086
LVPDDRLWD

47087
VWDDKSEAE

47088
VNPCSIMKA

47089
GPDEEWNIQ

47090
PMDEFGHYV

47091
PGEENEPPA

47092
ENDEASICM

47093
PSRTEGDKM

47094
PDRQQPMQH

47095
PNQLDKPAM

47096
EHPNDKIAI

47097
CNMATSNRS

47098
PWPDHQRAC

47099
WPDSRNSPN

47100
FDDRKRDAR

47101
LYCEADRKI

47102
PNENKGQGF

47103
TYPDVCIGW

47104
PQQQEGGTE

47105
PPNFKIYKE

47106
LPAGDEKGE

47107
REPLVTEGG

47108
LCDEGDMPS

47109
TEDFENHDG

47110
NEPQSPVGY

47111
WDGIDRQNT

47112
MKVTDKLDD

47113
QDETDEWQR

47114
QEFIGEERT

47115
QDDIDLASW

47116
PTDAALHGN

47117
PTINDWKTK

47118
PMDHEKRYH

47119
YKCDDNFIP

47120
RDSNCHFDK

47121
YNDFNRGEW

47122
PCDQTLIWQ

47123
DFGCKDEST

47124
CNLCTHAKC

47125
ANPDSSVMH

47126
FRDGDAAEE

47127
PFPCQQDIF

47128
VKNAERPRE

47129
CDFCRQFIK

47130
PDTSNKTFH

47131
HERCSELEY

47132
TDANRLCRK

47133
PNGCDRSGG

47134
LESDSFDRD

47135
PDLQKRQGA

47136
YEYWPDVSW

47137
CPWWDFAEC

47138
DQKDVKGKM

47139
YELCPLPQH

47140
DVEDCLICC

47141
IMPEKMHTD

47142
PHAEDRDNN

47143
TVPDVRDCQ

47144
EQHRDTMRP

47145
CVMPHNHRL

47146
PQEDEGVRQ

47147
SQPNPEENW

47148
PIDGNSHKD

47149
PGEPADEKT

47150
PDHWECIHD

47151
QQPPLAPDV

47152
YNPQNHVEG

47153
GDNAKDFRG

47154
FNVSWENKP

47155
PERLNGPTP

47156
FDPHRCSCH

47157
DRLRDDYPC

47158
VSEDDESHQ

47159
FESTQEPIA

47160
CKNFPMKNN

47161
PVPMSHEMQ

47162
HNGIEKTRE

47163
KDPCADRWA

47164
EVVDNMRSQ

47165
ADYKDWAEH

47166
PRNLPAYHS

47167
PMSQICCDM

47168
YDCWAKEAT

47169
ENPEVWIEQ

47170
IERWGVIDV

47171
DADDRVEAT

47172
CKDRPEIDC

47173
NGDDHVFRE

47174
DYIRIFDRH

47175
QDHEREEYK

47176
AEAKERFEM

47177
PDDEKIEHP

47178
MNPAVEKGM

47179
CDPQYDQED

47180
YEAHSDDRK

47181
MDPCEEPGR

47182
CODEDEAMQ

47183
PEAQDKVWS

47184
PRLTDIREV

47185
VPPFLDIRD

47186
PEHPDPFSQ

47187
YWCLDDPRD

47188
ERVEDGMDH

47189
RDYAPLLID

47190
QVPEIQSMP

47191
PRNGPYCDE

47192
CDNGEDDHH

47193
PFDTMDYRS

47194
PNTHPEYSK

47195
WNREDKRAQ

47196
PDWCQDPGM

47197
PQQPHWCDG

47198
PPDPPGDNV

47199
SPPNLRAFP

47200
GEEDEQSNH

47201
CHPTHQIRC

47202
PFPYHLEVC

47203
PNNQARGQC

47204
KNPTWVMSD

47205
YRHEEYWHN

47206
LEHENGTPQ

47207
QDPHAHRDD

47208
VDLCRKQVK

47209
PYWCDWPNG

47210
YPCQNHWRT

47211
MIPCHDRTY

47212
QPADPPNDS

47213
PKHADALRM

47214
LEPCTDEFG

47215
PDCDPAHHD

47216
CRPQAIWLH

47217
HEEIPQQPK

47218
CEDLNKPMM

47219
HNPSGPEFS

47220
GDFIPKQEV

47221
CKTQERANS

47222
SDDDREREA

47223
FEPVSGETP

47224
PHSDSWCDR

47225
WHKRDDKTQ

47226
PYPPMWRGM

47227
IEDVWECPY

47228
DYDWSDMKC

47229
IEMHEKKEE

47230
CEPRDCNFY

47231
PTMRESMES

47232
EVPFKTSTT

47233
VSPHKTSRN

47234
ADHCNWSRT

47235
SDHDNVFED

47236
SRPGDRGVQ

47237
QEPDSYACM

47238
DEECQRVPM

47239
VDNMSEGRQ

47240
EQEHDEEKP

47241
PEFGWWIAM

47242
PDELMGWQQ

47243
CECAGWSYE

47244
MEPFHVNKV

47245
HKDWHDADL

47246
PCNCDSWQN

47247
NEDWEAPEG

47248
PIEAMHDGW

47249
MIPQSEERW

47250
YKTGDEYGL

47251
PLDKEPMER

47252
HEDHDPAQE

47253
GNQDRTNRK

47254
CQTRDTLEL

47255
EADEQKTRW

47256
TEADEEWDT

47257
GDKWIGCEL

47258
ECDCRDTPC

47259
VQPQQMHKT

47260
PKCCSGMTT

47261
YNPFYGADM

47262
CPWCDMKPW

47263
KDRDTCRAD

47264
SELTNWVRW

47265
YNPAACVEL

47266
CVMPTIYRG

47267
EIKPYDARN

47268
EFSGHDEPD

47269
VDIKPKGAI

47270
MEQWDPDEW

47271
VEVTPGFKF

47272
PPNLCWNQH

47273
SRVKDCHKA

47274
ENEKPRTRC

47275
PHDWSPEFA

47276
PHQGDREGF

47277
IESDDIRTQ

47278
HDDPPGGGF

47279
EIQDLLCKA

47280
CVEDHWLHI

47281
HIAEPKNKP

47282
HEWKCEKRV

47283
YEATPHGRQ

47284
YPPNEHWQC

47285
LNPAGKIWD

47286
PEVGSPYPS

47287
IERHEHNPY

47288
HDPPKLGCM

47289
NDDCEESQK

47290
PVVERFNDP

47291
PFEEYEQTD

47292
VRPEKQRVG

47293
YSDEQGICQ

47294
HNVPDVRRN

47295
EADERSKEQ

47296
PYIWQAMHR

47297
KDHIAEHRL

47298
AKPKCPPEG

47299
DDDIGADSV

47300
DDPFENIGV

47301
WDGRTNDKA

47302
EGDDCMWCR

47303
VNRRRRDEM

47304
TEVDDKICY

47305
EHGDEMHPP

47306
LEKFPYQPL

47307
FDEMHTCYD

47308
KCPDEMNNI

47309
IDTGDWLVG

47310
PVPCCHYDP

47311
PGTEDYSRW

47312
LNYQPCCRI

47313
PAEEYHSRE

47314
PEELYLYRK

47315
MDVSDQWPE

47316
WFPDHMSGI

47317
CPPPSHHID

47318
LPPMKQDEE

47319
WRPCAYCQT

47320
CDDNGCTRD

47321
RYPEPDAQV

47322
QVPIMDDFF

47323
PKLPEYTDS

47324
MNSEPVTET

47325
HVPENFVQC

47326
PDRCHSGDW

47327
YPPPPNQHH

47328
IFPYTDKFI

47329
PGDCYWLER

47330
HEHRPYFRV

47331
DLFNHDSPH

47332
YDDNNWGSA

47333
HDSGPTFDM

47334
EIDLEDAQK

47335
PQEDKQCKP

47336
DVVHLDWKE

47337
IVPCLWPIP

47338
WEPVNAAMA

47339
QEPLRC1PG

47340
GNCEPSEPE

47341
DNVNNKQKG

47342
PDWFKADDT

47343
KHPAELDAA

47344
PTGHQRGTF

47345
WENEEDKVT

47346
QNWYDKLPT

47347
SDARECDIM

47348
QDSQDQQQW

47349
DNCDAGKWD

47350
VKDLADPPG

47351
PSAPNMWKQ

47352
ENEETWIFY

47353
WDNMDQTQG

47354
EYPCSKMRY

47355
QCDNESDSM

47356
SDPDIACVH

47357
HDDQMHCSS

47358
CEEETIEQS

47359
HRAQDPQQF

47350
HRTDEPTQN

47351
DVGTGDDRQ

47362
VDACSHNRS

47363
FDHQNWFEQ

47364
VNCPDQHDV

47365
CDMAHMLPV

47365
CDMAHMLPV

47367
YENKTDMKF

47368
EYPDNQWSQ

47369
CDWPIENRA

47370
VDHVKRCPV

47371
WRTSDKYER

47372
PK1QRVTET

47373
LDDWMMPVS

47374
HYPRNLVEE

47375
NEFQEDKSV

47376
DRGQDNHRI

47377
PFNDGGIIE

47378
CYPCLQDKG

47379
CTPNHCGGK

47380
SDTEKMHQQ

47381
GPCCPKYNC

47382
TLPADKPKG

47383
FTEEDEIRE

47384
PDFQKVVSH

47385
CNPDHMYCQ

47386
HETCGENVC

47387
WELAQKHKP

47388
YYNEPKAKG

47389
CKTDEFDNH

47390
KRPEICEWG

47391
LDPPNSHVN

47392
PHDNHRHGL

47393
PIVDWKRQD

47394
YRDMPGQHN

47395
YYPMADPSC

47396
PVNHKDQMT

47397
EYEWVDEPQ

47398
PYPGGAVRA

47399
RKPWVASRV

47400
NDPWTPISY

47401
YRFGPFEQE

47402
RRLPDFDDV

47403
QPPESMVQL

47404
DEWTNKRNS

47405
CRFTDDSSV

47406
PWNLPWRPG

47407
YESNTKSDV

47408
IEQEDQGYH

47409
EVLDDMKWN

47410
FNPMVYAHK

47411
QVDEWGGHE

47412
PMLQSRGWN

47413
PYACHHPDF

47414
PHTNRECDC

47415
WYDDDLLAN

47416
PPGCGWYDN

47417
WQPTHPARF

47418
EWDDPYGWE

47419
DCWWFGWQT

47420
KHPDQWTMT

47421
YIDRDDSEE

47422
VDYRTTVKS

47423
CDDERMDQP

47424
GVPIIEPRV

47425
WPKGTKDEA

47426
DRVHPDHLY

47427
NEQHNRDQQ

47428
QEPKETLMI

47429
KRWMEVDTY

47430
LNNKDPPPV

47431
FPHPDSSKH

47432
PDHSRLEYP

47433
LEPEQLDMF

47434
WPPCHETQL

47435
CPLRDDDPA

47436
EDENMKSTN

47437
PTEPDEQYA

Example 22
AAV5 Variants with Skeletal or Cardiac Muscle Tissue Tropism as Identified by Machine Learning

This example describes preferred properties of engineered AAV5 variants that display skeletal muscle tissue tropism or cardiac muscle tissue tropism as identified by machine learning methods. The same bioinformatics pre-process, machine learning methods, and analyses as described in EXAMPLE 19 and FIGS. 19-FIG. 22 were carried out to identify preferred properties of engineered AAV5 variants that display skeletal muscle tissue tropism or cardiac muscle tissue tropism. FIG. 29 shows a set of top 20 positional features contributing to model output probability. Shapley Additive Explanations (SHAP) values can be used to interrogate the relative contribution of features to model predictions. As shown in FIGS. 30A-B, these features were further compared between tissue targeting and non-targeting variants. FIGS. 30A-B show a comparison of top predictive features positionally. Features were selected if they were found to be important to both the HGB & RF models. Summaries are shown of the features in the top 1000 ML-predicted skeletal muscle or cardiac muscle variants compared to random 2% muscle variants.

Favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an AAV5 VP1 capsid polypeptide, which are associated with a higher probability of skeletal muscle tissue tropism or cardiac muscle tissue tropism are described below. Any of the below machine learning-derived positional amino acid preferences described in TABLE 44. Listed below in TABLE 44 are a summary of positional features shared between the top 30 important features for skeletal muscle tissue tropism or cardiac muscle tissue tropism extracted from two ML models (HGB & RF).

TABLE 44

Machine Learning-Derived Skeletal Muscle Tissue

Tropism or Cardiac Muscle Tissue Tropism Rules

Low solubility at position Xaa1

Xaa1 is selected from D, E, R, K, P, N, or Q

Low hydropathy at position Xaa1

Xaa1 is selected from D, E, R, K, Q, N, Y, or P

High surface accessibility at position Xaa1

Xaa1 is selected from E, R, or K

High hydropathy at position Xaa2

Xaa2 is selected from V, I, F, L, or C

Low mutability at position Xaa2

Xaa2 is selected from R, V, I, H, or C

Medium volume at position Xaa2

Xaa2 is selected from E, V, or Q

Low solubility at position Xaa3

Xaa3 is selected from D, R, or Q

Low solubility at position Xaa4

Xaa4 is selected from D, E, P, or N

Low charge at position Xaa4

Xaa4 is selected from D, or E

Low amino acid solubility at position Xaa5

Xaa5 is selected from D, E, R, K, N, or Q

Low solubility at position Xaa8

Xaa8 is selected from D, E, K, P, or N

High flexibility index at position Xaa8

Xaa8 is selected from Q, S, P, E, or D

High surface accessibility at position Xaa8

Xaa8 is selected from S, D, P, N, E, R, or K

TABLE 45 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that exhibited skeletal tissue tropism or muscle tissue tropism and comport to one or more of the rules provided in TABLE 44. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 25118-SEQ ID NO: 26117, as disclosed in TABLE 45. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 45

Sequences of the 581 to 589 Region in AAV5 VP1

Capsid Polypeptide that Drive Skeletal Tissue

Tropism or Muscle Tissue Tropism

SEQ

ID
581-589

NO
Sequence

25118
RDQCRMEAG

25119
EEREFRASN

25120
DVCDDHKPT

25121
EVREQCEID

25122
WERCLELNP

25123
PVMCVWHDN

25124
CWECCPDSG

25125
TGCNHNGRF

25126
PVICVWHDS

25127
RDQCRMEYG

25128
KGNPLDGDK

25129
EINEFRASN

25130
YSDAKNMSI

25131
DLLVATEWF

25132
PYGTIEYDE

25133
FPKVSECNW

25134
TQHEDHWLA

25135
NGVGRPGDN

25136
AADIFALND

25137
EDNIQIMME

25138
AVCDDHKPN

25139
TQPEDHWLS

25140
FPKGSEGNW

25141
WEQCLELNQ

25142
TYEALQNSW

25143
WERCMELNQ

25144
RYQCRMESG

25145
SGDWDFGAW

25146
DLEVATAWF

25147
VFKPLDNRY

25148
DSYTDSWSI

25149
PFACDRYEV

25150
SGDLDFGEW

25151
KDGRNPDCL

25152
DLSVATEWF

25153
SADIFAIND

25154
CWACCTDSG

25155
CWECYTDSG

25156
YKGMEKDPV

25157
TYESLOTSW

25158
QTNEEEYGN

25159
YYNHKIVEQ

25160
CWEGCTDSG

25161
AADEFAINA

25162
PYGPEEYDA

25163
EAGCKVWPT

25164
EFGCMYWGY

25165
DVAPMHQVA

25166
EVLIQCKED

25167
NKMEIFDSW

25168
DVADVPDTT

25169
HTIMEWSPT

25170
CGECCTDSG

25171
DVTTTSWIE

25172
ECGCMYWCY

25173
GNEDCNLAF

25174
HHFNVNLDE

25175
TQTEDHWLA

25176
ECGCMYWGC

25177
YSDPKNMYI

25178
EIHMFGPSE

25179
EAMPFRENQ

25180
YSDPMNMSE

25181
DLEVATELF

25182
KEFVQWAAA

25183
ASDMQFWSM

25184
DTSDLQVMK

25185
EVYKNDDGN

25186
NIDEMNCNA

25187
YKGMEKTPV

25188
NTVEKMQTL

25189
ESYPKLTQF

25190
RDAWKWKRY

25191
TIDDKQSWA

25192
ATMDGHMSM

25193
WGRCLELNQ

25194
CQMLCRTPV

25195
VFKTLDKRY

25196
EMAELWDTG

25197
EISSSNLDP

25198
DLIVATEGF

25199
FCDPAHHYE

25200
VMDEYEANP

25201
ETSEHNRMP

25202
ECGCMYWGS

25203
PYGLIEYDE

25204
EIAGNDTYM

25205
VDRLFRVEY

25206
PTEPDEQYA

25207
SVDQDQPGN

25208
GWECCTDSG

25209
SGDWDFGEG

25210
DMENPNAGP

25211
VFKPLDKRS

25212
DVIPRDGAE

25213
PVICVWHGN

25214
ITAEVTWQE

25215
IRTWGCKKQ

25216
GSHWYDCNP

25217
RLTEDMSDM

25218
FDWMANVSV

25219
THESLQNSW

25220
FQMLCRPPV

25221
EIHMVGQSE

25222
ETADRIHWI

25223
PYGPIEYYE

25224
CDHVTKNPT

25225
EVHTKDEHM

25226
TVCNHHGRF

25227
QGDYSWWRY

25228
GTLDVPTMF

25229
SVICVWHDN

25230
HVDITDNDM

25231
CDHATKHPT

25232
TYEYLQNSW

25233
ATSEIQPFV

25234
EVIEGCPMG

25235
DNVDLDIQH

25236
NVLGFDDGG

25237
SNVMATQNW

25238
QVTDNAAGM

25239
AKYLPGLYY

25240
HNCPWDQQY

25241
CWECCTYSG

25242
WNTIMTVES

25243
PQYVHNWRQ

25244
HTGMEWSPT

25245
WDYSMAKKD

25246
WKHCEREIN

25247
SIFDYDTFQ

25248
QTNGEEDGN

25249
EAALENVNM

25250
WQKPINCEC

25251
EIRMFGQSE

25252
DALEWPMPP

25253
DIAPDWKGE

25254
TKMNIFDSW

25255
MVRPTDQPA

25256
ITEDQARSC

25257
IEYWEHETK

25258
TIVEDAPLA

25259
YKEMPTILC

25260
TVLEDSMWA

25261
AVCDWLIAE

25262
PSSCDWIPV

25263
LSTELHHFG

25264
CTEGENDEH

25265
PMPDGMFGI

25266
CTVDGLHFH

25267
DNYMQLNGV

25268
DVDGEHEVY

25269
EVQHELASG

25270
FTGDFATFA

25271
TTDQQCTGP

25272
ESVEKSISN

25273
AMGEEINYQ

25274
VAGDIEWYG

25275
DTQPDTLNP

25276
AITDDNRMQ

25277
TNDQQCTGH

25278
VCEELDREW

25279
GNGYWMCMS

25280
HQYVHTWRQ

25281
SYHMFDYGE

25282
CMHDEMQTS

25283
DFNSIDHSG

25284
DASERNRWQ

25285
RHNPHDACE

25286
QYVDQENCN

25287
DASPQCDRD

25288
EIKPAAEIT

25289
DALTRVDGD

25290
DYQWPDVEE

25291
DIVENDSLQ

25292
VIVESEEYM

25293
GFVDKMVMD

25294
IVHEWEVNH

25295
EVMQPEASV

25296
QTLDKWDGP

25297
DTFMMQSAH

25298
DAIPWWWSY

25299
WERCLVLNQ

25300
NVEEGMAAD

25301
EAGPWKRDV

25302
TIHDVAMMD

25303
MNLQIGGKG

25304
NNVRQVGNT

25305
QFPGGDDEG

25306
VYVEMIGWY

25307
TVEHTNHDA

25308
HQYVHDWRQ

25309
NLIVATEWF

25310
MLMHADMTP

25311
DSSESLNQN

25312
KCEFYQFCQ

25313
EVCSQWTQF

25314
MIDEVWHHW

25315
PNMPHAEDQ

25316
TFDDFDMCA

25317
PWQQHRKMS

25318
GDPWEEWRK

25319
EQLEDNDMQ

25320
MIHSQDMFE

25321
QVFHQDDVN

25322
HYKYDANRK

25323
DMMNIEKMG

25324
WEHCYRVSW

25325
DILFWEDDD

25326
MTDEHLQYT

25327
LSVDRDYFL

25328
DVQQIHNKE

25329
MITNNCNIP

25330
NISWNESHD

25331
ETVHPSASN

25332
TKMIIFDAW

25333
IIAQWDQVN

25334
TADDFQGDE

25335
SQERMAYDG

25336
EAMQDHTDC

25337
CDWMANVAV

25338
DVKPVQKDW

25339
ANEYQYDTM

25340
MVVDRMLGW

25341
MVVEYLTAG

25342
MPAYSKLKW

25343
AHEEDEAMQ

25344
ETRPTVAIQ

25345
VDNMSEGRQ

25346
SQESMDYDG

25347
CVDDLASSF

25348
EVKPSMRDD

25349
LCSQPIPEP

25350
ETRPGMSDS

25351
ETSFKIHEH

25352
DTVEGGYIQ

25353
YYEHPINKK

25354
SIDWINQQG

25355
EITPYAKWP

25356
EMCERNAGI

25357
DVCINPWYT

25358
NDYFRVYRK

25359
SQEHMDYDG

25360
LGPWPQTDQ

25361
AGEFVQANW

25362
TVKPHDNTE

25363
QDEMMMACI

25364
VTWEDVYIN

25365
NVHESDTMF

25366
NIAGYEWWM

25367
WLLGMWNTG

25368
VINPHDRSV

25369
TVIPYEMFD

25370
PSTNLTDSA

25371
SVYDDFTST

25372
NIHGVNLQG

25373
DWEKVPCMK

25374
TQQVADYTK

25375
FIGSSDKQS

25376
CLDELVEED

25377
ATLDLDSHA

25378
DTQQEQSQC

25379
TAEIFAVND

25380
DVVFNLWAP

25381
GSMSSTVPH

25382
NVVMLDQSY

25383
QQDMIVTKQ

25384
DFMFLISTH

25385
FQVDTFMFE

25386
IFEWGDATY

25387
LPPGIEWEH

25388
MDRLAEFKD

25389
DTVVKNMYH

25390
EQIPVTWTT

25391
WHMMSRDGN

25392
STEEGKGWF

25393
DVMMALATQ

25394
SAEPQDANV

25395
SVTMDEEAH

25396
AVPTSQAEY

25397
PCAMHNWCC

25398
EVYVESQNE

25399
CIDTYDIDS

25400
SNMELYCRV

25401
GTHEHEYGA

25402
DTLPVQVYQ

25403
NIFDTNVLT

25404
VSEDKEAIS

25405
VMHDIPNFF

25406
ETMSRHEGM

25407
ATEIDHWQW

25408
LSMDVTMAG

25409
AIHWMDWCQ

25410
TVHFESEET

25411
EMAGPFVTT

25412
DIHVNNFWN

25413
EKQMDGINH

25414
QEDGDFSTF

25415
EITQTYDHL

25416
AHSEIMCDQ

25417
VTKLAEDVY

25418
PEFMWCVPW

25419
DTTTDMWVA

25420
DVFIENQHW

25421
NVSLQSIHK

25422
ESGMEQISQ

25423
PNGCYTDCE

25424
AVKEFHDCF

25425
WVADHGDGS

25426
EIGMTQPCT

25427
WLYTSNFKD

25428
DDMEDQQQY

25429
EATTTSEKD

25430
QTQDTMNLH

25431
QVHHNNVDM

25432
SVVLCNLCQ

25433
DAPDFMTFC

25434
FPRIEGQGE

25435
POLFKGPVE

25436
TTEWLDFDA

25437
FEWVDWGYV

25438
MRNYPDARF

25439
EIGSCPEWC

25440
DNPPGRRKP

25441
TTTDSPMHF

25442
SCDESEAYY

25443
WEHVLDYAK

25444
DASFVDODE

25445
GVNEAEFTI

25446
MIEVTSHST

25447
AVQHTEFNL

25448
DIQGMLEAA

25449
DVVMTAMDY

25450
FKDWDHHTV

25451
NVEHNNMGA

25452
HVHEIMEEE

25453
SVMTNDDID

25454
GPIEFGWQI

25455
EFANTEHTS

25456
VTEDMIMNT

25457
SQISVEFNC

25458
EFWSRAWSD

25459
DADELRSAK

25460
EMRTYNKDY

25461
SSTELFQAG

25462
TEPETYPAT

25463
VMDDESHRG

25464
DACEEGSSM

25465
WYTQDIEKE

25466
IGDHDCQEQ

25467
DVGNTYNHW

25468
QSYTTMYQC

25469
ISGEIDYME

25470
ACCEFELTQ

25471
TANEWETME

25472
AEKMDEIWK

25473
VTKPKWSNF

25474
VVVLSDFGE

25475
GSHWYDYNQ

25476
GMYESNQGS

25477
DVDCVWHEE

25478
LVYDPHFTS

25479
NDVDLWHVD

25480
AVEYAGWEK

25481
FYQADIGWA

25482
NTEAEDHQD

25483
RCDCKVHEF

25484
HLPCGWPAV

25485
IPVHFDWFF

25486
NVEGIVTAC

25487
DVGNMAAIE

25488
ETLQAQHCD

25489
NTADIHMKE

25490
SPLEVWTPA

25491
KFVWDTWAW

25492
KCFLCGSTI

25493
WECADRRPT

25494
VIDDYFQMC

25495
GATCTIVRY

25496
TIDCYAAMD

25497
PYCQQAANW

25498
MVYEATELD

25499
MTVDLYEVL

25500
EVDKGLAQA

25501
DSELLPEAH

25502
AVYDHAPQN

25503
WHHTHSHNC

25504
DIMPQWAFM

25505
DIRHYNISR

25506
VTLDGNRMG

25507
IIETQCQAI

25508
EASHIFTGE

25509
QIAEACAQH

25510
ETTCNMGAK

25511
VFDMTNLRV

25512
LSPWHQTDQ

25513
ECSHQDRSN

25514
TMMDAEVGL

25515
YLTELIQKT

25516
EIVELRQTR

25517
FPMVWFVLF

25518
SNEWPTMIH

25519
QISEHAGRS

25520
VVQEKSINW

25521
DAQPQRNGS

25522
EVWTSCMTP

25523
EATNFTIGR

25524
VTEPHSFTV

25525
LHEVQTILH

25526
EVVTQMEHL

25527
WRVWQNLPQ

25528
TSEYMATGE

25529
KDWRFEDEK

25530
IQAEQMSGC

25531
DAKNWLDKN

25532
MPNFAWEAQ

25533
DLLNYGMFH

25534
EQPFDEFKA

25535
HATGLVNVM

25536
KEEFNQEPA

25537
EAYTRVGYE

25538
MQEEMGPRK

25539
DFTNQDHDN

25540
EVRANCESY

25541
DNTDPDGSW

25542
TEGPSNFST

25543
DNANFSGAD

25544
EYMERDMSI

25545
FDKLGTMWG

25546
CVDQLDDVT

25547
ESGGYMRMC

25548
VQTMPKPIY

25549
TYVDNNHMV

25550
EIVVEFEDQ

25551
IPDCMNLSQ

25552
HTVMEGSPT

25553
PIIPIAEHV

25554
DVQGSGQWV

25555
HTNQQANSP

25556
STDDEYSLA

25557
TAVEWHYFD

25558
MAPQSLWNM

25559
SSGEMDHVY

25560
TEDDYGPIS

25561
TTEAGTALE

25562
TTEAGTALE

25563
ETDLKCHSY

25564
ACVEQEKAD

25565
SMNEMYQYP

25566
GEDTTTQAF

25567
FPQMNEWEC

25568
DVHTTCYHY

25569
SQPNPEENW

25570
EVMLSKMEY

25571
EVDHCNMYP

25572
EFHQFGNHM

25573
TVQMGEWGY

25574
NMSDGHEVC

25575
DDNEYHVWS

25576
DNWMGQDDK

25577
DEMDAQMGA

25578
TNDWMRLRV

25579
IHYDYNTIC

25580
DTGSFTSTP

25581
LTEATPKWT

25582
GDWSERSIT

25583
TTCPMEKLN

25584
DVAQYGLGN

25585
DVAQYGLGN

25586
TADMMPMNP

25587
TDESEFESS

25588
HDMSYHDPI

25589
TVVERHEHG

25590
CVETQWRAD

25591
SEDIESARQ

25592
EVEVVKEFQ

25593
NTOTTDEHG

25594
PQKGDDWLY

25595
PCEQMPYIM

25596
DEYDNVWQA

25597
DMRDSHAWP

25598
YTCHEEWPL

25599
ESGLAPMNL

25600
PMDHEKRYH

25601
VPEPDDKPY

25602
AIIWTTDQP

25603
QSMPECEGH

25604
DAGPLSMRS

25605
VTEVQHMAK

25606
WEGFYMLTL

25607
MIMCMDCFP

25608
DIAGMPHIS

25609
GVDAGLVSE

25610
TIDVPMQYA

25611
MELEGHNLP

25612
DEWYRSKED

25613
VWDCPKWRH

25614
SYDDTVICW

25615
EVDNKCYYN

25616
DSTPIMQRC

25617
YWDCQHSYM

25618
TKMIIFGSW

25619
SVVERQLVD

25620
EVCEHEQCM

25621
DCSHEAHRY

25622
INPELSEGC

25623
DTSSWGVEP

25624
HYRDNKSLP

25625
MNSEPVTET

25626
QMEWLAAGP

25627
EFTTADGEF

25628
MFPSEAKEI

25629
DVKPGLFYT

25630
ETDQPQLAA

25631
DVLLYMSAT

25632
DIIFKLEAG

25633
CDAGLISKM

25634
MFTGPDVRH

25635
GVMESQYIS

25636
DCTLQPAMY

25637
PEAQRVAGE

25638
DVEMCQKMS

25639
WHGCYKIMF

25640
VHMPYTEET

25641
ENGTHQYND

25642
QCAYSWWRY

25643
CEWLGTCNH

25644
DTCYNNWKD

25645
LSAQRLVGD

25646
NLDMYSESQ

25647
SELFYSETH

25648
MDYDGMAVY

25649
LCSDEVCWT

25650
TIEGKSLYC

25651
ETSMPMEHT

25652
VE1NAKWGN

25653
EEPWDCQIP

25654
EEPEFTGRG

25655
ENDCRERMA

25656
CVMHVQCDH

25657
AIATGWQDM

25658
EYHGYLEHT

25659
IQWVHGCLF

25660
NDCSDCOIL

25661
AGMEIIVQC

25662
DHEMPESTF

25663
VKCFPCQLE

25664
NEPELKLQP

25665
DTCQAMHGD

25666
EWHFVNEGT

25667
MVQQYNGSE

25668
D1ERFSTVC

25669
NEAAHSPGP

25670
HNPCWPFMI

25671
IVDMPVRHP

25672
SADEDKCGW

25673
SFPEAWPIA

25674
MIVSQQWVW

25675
HIEYWDQRD

25676
SLMYEDDYP

25677
DAAIYVMLD

25678
MQKNPPKRV

25679
TMVDWELGK

25680
QTGLNEYAP

25681
DDMWCGGLY

25682
VIYFVGLIA

25683
HHACGTREH

25684
DISSNEYDI

25685
AIECYKMGS

25686
EESNRGYQH

25687
MTEVVVSSM

25688
HEGHPHLSC

25689
ESCTGEWWA

25690
MTDTVQEYA

25691
PRNGPYCDE

25692
NKMTEARKH

25693
DNGQVEETA

25694
DAYVQQPSF

25695
FVYEWSNLV

25696
EVRGQGSHY

25697
WFDCPPCCA

25698
WREFSEWTF

25699
QLFAPDMDW

25700
TYNDFYTAF

25701
EVGSMPQDW

25702
SKELGSVET

25703
DVTISMQAG

25704
KYNFNLNKY

25705
VSDWPQIKA

25706
DTKSRQWDC

25707
AYEGECIHY

25708
MAEFEQWLE

25709
CELHELFDG

25710
TAEYIDISY

25711
EIAQFSGVC

25712
DWNEEMAHL

25713
WQEYRMTDQ

25714
SNENICPTC

25715
LPVDVLVKT

25716
SADWRHKSP

25717
EDGQSMCVS

25718
MHHEYAMAG

25719
DDHDINGGP

25720
MTEDQTWWQ

25721
WNEPWYVRP

25722
ECQQDSWHY

25723
EVOROMSNK

25724
DMHSFKWWS

25725
DHGEIPAFT

25726
EYDKMMYAE

25727
DADREDFWS

25728
QQEAEHKGV

25729
AHEVITVRP

25730
QPIDVSAAI

25731
DIYLKCFIK

25732
EIFLENRFP

25733
FTKTNDHNT

25734
WQALWCLVH

25735
AFKALDVQC

25736
DIHSTNWLA

25737
NLTDKCTAD

25738
NSEWWSSVL

25739
TNEIMWCSP

25740
MVAPSQALQ

25741
APGEYMDVE

25742
ECAKTEWNF

25743
KDLHDAIHN

25744
LAPSCKIYC

25745
FMWPPQIVV

25746
HMQEPSIAS

25747
MEPVMHYLN

25748
YCKCPTCNT

25749
ADGPVLQMF

25750
EVSSCDDFT

25751
CNEEMEFSC

25752
MLTESQQSM

25753
WPGFEKLPS

25754
SPYCDDRGN

25755
QYCLTEASH

25756
TSEWLWVDS

25757
DNMVWRGAH

25758
CDNDYLTAT

25759
VIEYNSNIY

25760
HVSAIEKAQ

25761
QVATFECKQ

25762
LHIHRDVTI

25763
NHEETEFGY

25764
MDSCYDYQS

25765
EVLGCNYWP

25766
STEDWATLH

25767
CQTDKNATE

25768
ALEQEMVKF

25769
MITFASASK

25770
DDEVYVWGQ

25771
QVMPRSIER

25772
AVHTFQMWD

25773
LVGNADHGQ

25774
NVGLPSYFA

25775
EDYSGTKWG

25776
TVPLKTYGG

25777
TYMPMDCKW

25778
FIVQQSIWK

25779
NEQHYEYWK

25780
LDVTPQPEG

25781
ETSLPHWQG

25782
DSKPVMMAN

25783
VYRHFDMTP

25784
DSLAAWSDK

25785
STIPTPIWL

25786
VQVQDWLPE

25787
DWNCAALKP

25788
EVGPSWEQR

25789
MHDFLTSAQ

25790
NLPCSQQAM

25791
DQRCNKAPG

25792
PLAFSPVQR

25793
ETDLAMKAE

25794
NVEWRQACN

25795
ETTTCWQYF

25796
AMPYEEVPA

25797
ESAQFVAYS

25798
VLDCAMCNP

25799
AEPHFQNKS

25800
MNESFPTWN

25801
EVQCDQIHP

25802
MDQCFSYNQ

25803
EVGSQRTAY

25804
DHSPVKSAY

25805
YICQQDLIM

25806
KIDHEQDCY

25807
LPVEPIDCE

25808
GLMVGQWPH

25809
AHIWDTLIP

25810
ATIVGDSGN

25811
TANDYGPTE

25812
NVELPKIHE

25813
MLPDHWVIT

25814
HPFNEAVKH

25815
IISSTTSTG

25816
IKEHECRRI

25817
AVHQINVAA

25818
LVMEALPMA

25819
GQIWAAMRR

25820
MWVCKLFVC

25821
PIHHGEHVY

25822
NHQCNGLAQ

25823
TCEEPYNFA

25824
LVHEKWEQF

25825
ALAKFRCWS

25826
ITSVFDNKA

25827
DVITHWIGQ

25828
QQASSSGGF

25829
EGDDLDRHY

25830
MPELKFSYC

25831
HQTHQHNIW

25832
AFGTLEHEP

25833
TKPTHDVGS

25834
QVMQNESER

25835
SDESNWFGQ

25836
FSADVNNYD

25837
LWCCIDTEC

25838
SMEEWGCES

25839
MHFCIWMRM

25840
AHNESDDCG

25841
SHQFVQPNN

25842
AIHMFGQSE

25843
PRKCSNEEV

25844
HFPLGWLWL

25845
QQLELDGSL

25846
INHGWEKVA

25847
HQHCAEFCH

25848
DCSHNGMRE

25849
WINQQTLQC

25850
QERMFEMTA

25851
QINMSMHES

25852
MVTARCWLS

25853
PLWFEPALN

25854
EIGYRKVYN

25855
ESVMEKSMF

25856
NVNHFGMGD

25857
SVNVDNEAM

25858
NEWFQCDVH

25859
NPNCTKQQY

25860
QMGPHERHA

25861
ETRMHMDAL

25862
LNMYEASLS

25863
MKVTDKLDD

25864
ADETCAKKF

25865
INQWLTCRW

25866
HNVGDMFCN

25867
SNQPQGQYC

25868
LEKVINVLH

25869
SFVHMFQGP

25870
DASYYSPVP

25871
VNDDIYQQG

25872
LEMMGWPIG

25873
MEHALCTRT

25874
ECSCVKRGY

25875
ESGFPNPYH

25876
STDWSPSLG

25877
ETDYIQMGA

25878
DIVTVIKRT

25879
LAWDNQQFN

25880
DTDHRNKFP

25881
MSNDRCSMG

25882
FRQSVPHWA

25883
QVGQEAKNC

25884
YLEWDSWRC

25885
MSDFPISTP

25886
MQNLSSVTE

25887
STEYRNALN

25888
VIVGNNVGI

25889
DGPCDQMTN

25890
MDEYFCGFT

25891
EPAQSQHGF

25892
SFDYVMWGC

25893
DVFKEHYHQ

25894
HFVQPACEH

25895
DVANLKHHV

25896
VSEQIWNAD

25897
IVPCLWPIP

25898
GHQEFKGSE

25899
ANCSSEIYV

25900
DSTHDIAHS

25901
TTDRQCTGH

25902
V1SAMNVYP

25903
AHQFIQWLH

25904
ETYRYGSQR

25905
KGNPLAGDT

25906
NTMQMPMIG

25907
MYEVLNNDQ

25908
ICLHDHFRN

25909
EMKGTLYAY

25910
APDFRMITR

25911
PQWMPVNQI

25912
EFKTQQFDP

25913
VNEWLDSIY

25914
GSIQSVMWE

25915
DFMCDQFVC

25916
PCSDSTELD

25917
DQECWQSSQ

25918
EEPLPSDQK

25919
QTMKLEHLK

25920
PGTHWWWDC

25921
MTVSWDSWK

25922
DAWKPHQFY

25923
THEEYQLGS

25924
TVIQTECNT

25925
SNTFIDWQA

25926
LYDWSCAQS

25927
WREPNCHGL

25928
TMAEMCGSP

25929
SETDWQRKC

25930
QVIIGYCES

25931
GFDECMFWY

25932
PCTQIGEIY

25933
FMDFFGMAS

25934
DQMLDNTAY

25935
SQDCQEVGG

25936
SDHMSERVD

25937
DVLQREEIC

25938
VIALYSLAY

25939
EWNYGGLKC

25940
EVQYGKYWA

25941
DMTMYSNAV

25944
MTQGYPSLM

25943
SIPCNHCFE

25944
MTQGYPSLM

25945
DIASVRQTE

25946
IVDSASVCQ

25947
AIEAKMTQV

25948
ALAKFQGWS

25949
PAFLGACGF

25950
ESEEHRVMS

25951
CQGCVPFKG

25952
FIEFDAVTW

25953
TTCSSNREW

25954
VCNELQKYF

25955
VPCYHKGRW

25956
DVKCEPDRG

25957
LICWYQQGN

25958
EMQTHYAAR

25959
AETDHYIPT

25960
PQSDDWAMP

25961
NHQWTQRRY

25962
FEGNYTATK

25963
CNCPDQHPD

25964
IFQCLYTGT

25965
VETYQINDT

25966
FFEWLLDTG

25967
SDALGPEWW

25968
MTNCKLHEQ

25969
IYEACFWWL

25970
WCMQEGTMS

25971
EIRLEHDGG

25972
HVELSLWNQ

25973
CWMEFMAGN

25974
AWDYWNSQW

25975
NNITTPDAY

25976
SVTMHSMEL

25977
TYDHIKFAP

25978
LQQMENASP

25979
CTDDKMCPV

25980
IDSCNTSLN

25981
CKEMRMDQW

25982
EPEHGAMDK

25983
VINCHTENN

25984
ETHLINPGI

25985
CDHDTKHPT

25986
CSAEIIDTC

25987
EVGFYMVPR

25988
AIWSIEHES

25989
MVNLGYAEC

25990
ELEGMSRRE

25991
VDATTGDYQ

25992
TVGFGLKAS

25993
WMEASITGP

25994
WTDGWHTMC

25995
SVTLGMKPN

25996
GHVQESWNC

25997
STVAADWDH

25998
ESEIAISWN

25999
NETLDSKHF

26000
TFMWLDRVG

26001
TTDSWMDVE

26002
DEAEEHGCT

26003
FTPPKLHYK

26004
QYLTAEFTS

26005
MSTNNCNIS

26006
EAWSCGIYD

26007
NYDTLLETH

26008
NVEYVNWHW

26009
FVMPISAQG

26010
SQESHMTWH

26011
GIVALDNAD

26012
TVMWKDYID

26013
PWCEDPGCS

26014
TVEYVTDDA

26015
TQDCSTMPS

26016
SMCWLENGN

26017
MVDLETTYV

26018
DNTQKQNDP

26019
NVDRCDSWQ

26020
TYEPCSTSH

26021
DFHCQTERQ

26022
TTAPMEMSL

26023
NAVEGQSHK

26024
DNYSIKVGQ

26025
FMMFHMGLD

26026
ETRFDDRRA

26027
NTGWQEAER

26028
WVFMSWCDV

26029
PCLITVAPH

26030
DIAVIGAGA

26031
EATLITYCS

26032
TTEDMQCDM

26033
ITMPRVRPT

26034
LFVMPAAIE

26035
ALEHTLFGT

26036
MGCDVMKAS

26037
DSWPTLMHD

26038
CYHEECDGN

26039
ITTPQTQSH

26040
IHDWYEVGS

26041
SMQMTSHYH

26042
TVAVFSIAM

26043
QGDEINEQA

26044
PCSQYYPSC

26045
PFAAVSMVF

26046
MQLTNMEIE

26047
VTESIDYVH

26048
TIAPYTYMD

26049
DIQLTHIHG

26050
MYMTVQQNV

26051
PFHERGLMQ

26052
TYVNQGMGP

26053
FVCELQSAN

26054
ETKSHPVMV

26055
ENDEASICM

26056
VVHMENDQM

26057
NTFILETFD

26058
HDENWVIKE

26059
WFVTQGAYM

26060
FWATSHWGY

26061
IPYTEKADI

26062
HDEEEKNKA

26063
WLHYVEDTL

26064
NWAFNARHD

26065
CSGYWMCMS

26066
DEVLTGTIN

26067
DVNQNAKYD

26068
ITDIFHWSC

26069
HMRTMDAFE

26070
ESIQKHSVC

26071
MSEMEKQQF

26072
LMDEQPMWC

26073
HWDVQVNNA

26074
EQNHAQLMP

26075
DTAKVVELC

26076
ETSCGTLMC

26077
EQQW1APKH

26078
EVKOATIWC

26079
TYDPFWMQH

26080
THDTTIFAF

26081
THDTTTFAF

26082
WVTPQCIHL

26083
LGHDVNGHH

26084
CHELIKKQW

26085
SNWSKCAPT

26086
LNDYTHHQA

26087
ACEQENIQH

26088
LYDYMHMGS

26089
EVPLHTVCQ

26090
VHGCGKQYH

26091
MHMDQPTMC

26092
VKFSNEMKW

26093
MELHKCNSP

26094
ELHGTYNSC

26095
SEAEMDWIG

26096
AFRTMDEVH

26097
IIDNVLCNG

26098
MKMCNCPYQ

26099
MILVQQLQN

26100
PSTGNIQFM

26101
QHPGFGLVI

26102
IWWEHMGPA

26103
QWRWHKSAC

26104
DSESRLCED

26105
ETSAEFEQV

26106
FLGSMWPRD

26107
CVTLEQSSF

26108
HMTAMCLAG

26109
NVELRHHGY

26110
MALDQRSKP

26111
DFIYDGEDM

26112
FDRHDKSCC

26113
TIGQHCADF

26114
HVANGTDAC

26115
DSHTRRYDT

26116
MIPQPVEPF

26117
TVAHHFVET

Example 23
Treatment of a Neurological Disease or Condition with an AAV5-Derived Virion Encapsidating a Therapeutic Payload

This example describes treatment of a neurological disease or condition with a variant AAV5-derived virion having any one of the engineered CNS tropic variant AAV5 VP capsid polypeptides disclosed herein, wherein the variant AAV5 virion encapsidates a therapeutic payload. Polynucleotide sequence encoding for AAV Rep, an AAV5-derived variant Cap and helper proteins and a therapeutic payload are transfected in cells to produce variant AAV5 virions, where the polynucleotide sequence encoding for the variant AAV5 Cap comprises at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 in the VP1 capsid polypeptide and where the polynucleotide sequence encoding for the variant AAV5 Cap encodes for an amino acid sequence of any one of SEQ ID NO: 7118-SEQ ID NO: 10117. The therapeutic payload is a guide RNA targeting an mRNA encoded for by a gene implicated in the neurological disease or condition or a transgene. The variant AAV5 virion encapsidating the payload is administered to a subject. The subject is a human or non-human animal. The route of administration is a systemic route of administration. The systemic route of administration is intravenous administration. Upon administration to the subject, the variant AAV5 virions encapsidating the therapeutic payload exhibit enhanced CNS tissue tropism as compared wildtype AAV5 virions encapsidating the therapeutic payload. Upon administration to the subject, a lower dose of the variant AAV5 virions encapsidating the therapeutic payload is administered as compared to wildtype AAV5 virions encapsidating the therapeutic payload. Upon administration to the subject, at least one symptom of neurological disease or condition is alleviated or the subject is cured.

Example 24
Treatment of Alzheimer's Disease with an AAV5-Derived Virion Encapsidating a Therapeutic Payload

This example describes treatment of Alzheimer's disease with a variant AAV5-derived virion having any one of the engineered CNS tropic variant AAV5 VP capsid polypeptides disclosed herein, wherein the variant AAV5 virion encapsidates a therapeutic payload. Polynucleotide sequence encoding for AAV Rep, an AAV5-derived variant Cap and helper proteins and a therapeutic payload are transfected in cells to produce variant AAV5 virions, where the polynucleotide sequence encoding for the variant AAV5 Cap comprises at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 in the VP1 capsid polypeptide and where the polynucleotide sequence encoding for the variant AAV5 Cap encodes for an amino acid sequence of any one of SEQ ID NO: 7118-SEQ ID NO: 10117. Each such variant is detargeted for all non-CNS tissues, including being detargeted for cardiac tissue. The therapeutic payload is a guide RNA targeting an mRNA encoded for by a gene implicated in Alzheimer's disease or a transgene. The mRNA targeted by the guide RNA is an mRNA encoded for by a gene encoding for amyloid precursor protein (APP), alpha-synuclein (SNCA), Tau protein (MAPT), or Apolipoprotein E (ApoE). The transgene is a gene encoding for amyloid precursor protein (APP), alpha-synuclein (SNCA), Tau protein (MAPT), or Apolipoprotein E (ApoE). The variant AAV5 virion encapsidating the payload is administered to a subject. The subject is a human or non-human animal. The route of administration is a systemic route of administration. The systemic route of administration is intravenous administration. Upon administration to the subject, the variant AAV5 virions encapsidating the therapeutic payload exhibit enhanced CNS tissue tropism as compared wildtype AAV5 virions encapsidating the therapeutic payload. Upon administration to the subject, a lower dose of the variant AAV5 virions encapsidating the therapeutic payload is administered as compared to wildtype AAV5 virions encapsidating the therapeutic payload. Upon administration to the subject, at least one symptom of Alzheimer's disease is alleviated or the subject is cured.

Example 25
Treatment of Parkinson's Disease with an AAV5-Derived Virion Encapsidating a Therapeutic Payload

This example describes treatment of Parkinson's disease with a variant AAV5-derived virion having any one of the engineered CNS tropic variant AAV5 VP capsid polypeptides disclosed herein, wherein the variant AAV5 virion encapsidates a therapeutic payload. Polynucleotide sequence encoding for AAV Rep, an AAV5-derived variant Cap and helper proteins and a therapeutic payload are transfected in cells to produce variant AAV5 virions, where the polynucleotide sequence encoding for the variant AAV5 Cap comprises at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 in the VP1 capsid polypeptide and where the polynucleotide sequence encoding for the variant AAV5 Cap encodes for an amino acid sequence of any one of SEQ ID NO: 7118-SEQ ID NO: 10117. The therapeutic payload is a guide RNA targeting an mRNA encoded for by a gene implicated in Parkinson's disease or a transgene. The mRNA targeted by the guide RNA is an mRNA encoded for by a gene encoding for leucine-rich repeat kinase 2 (LRRK2). The transgene is LRRK2; GBA1; GBA1+alpha-synuclein; AADC; GDNF; Neurturin; GAD; NTN; hFOXG1; hKCNQ2; hFMR1; anti-Tau/miRNA; EPM2A or EPM2B. The variant AAV5 virion encapsidating the payload is administered to a subject. The subject is a human or non-human animal. The route of administration is a systemic route of administration. The systemic route of administration is intravenous administration. Upon administration to the subject, the variant AAV5 virions encapsidating the therapeutic payload exhibit enhanced CNS tissue tropism as compared wildtype AAV5 virions encapsidating the therapeutic payload. Upon administration to the subject, a lower dose of the variant AAV5 virions encapsidating the therapeutic payload is administered as compared to wildtype AAV5 virions encapsidating the therapeutic payload. Upon administration to the subject, at least one symptom of Parkinson's disease is alleviated or the subject is cured.

Example 26
Treatment of a Tauopathy with an AAV5-Derived Virion Encapsidating a Therapeutic Payload

This example describes treatment of a Tauopathy with a variant AAV5-derived virion having any one of the engineered CNS tropic variant AAV5 VP capsid polypeptides disclosed herein, wherein the variant AAV5 virion encapsidates a therapeutic payload. Polynucleotide sequence encoding for AAV Rep, an AAV5-derived variant Cap and helper proteins and a therapeutic payload are transfected in cells to produce variant AAV5 virions, where the polynucleotide sequence encoding for the variant AAV5 Cap comprises at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 in the VP1 capsid polypeptide and where the polynucleotide sequence encoding for the variant AAV5 Cap encodes for an amino acid sequence of any one of SEQ ID NO: 7118-SEQ ID NO: 10117. The therapeutic payload is a guide RNA targeting an mRNA encoded for by a gene implicated in the Tauopathy or a transgene. The mRNA targeted by the guide RNA is an mRNA encoded for by a gene (MAPT) encoding for Tau protein. The transgene is MAPT. The variant AAV5 virion encapsidating the payload is administered to a subject. The subject is a human or non-human animal. The route of administration is a systemic route of administration. The systemic route of administration is intravenous administration. Upon administration to the subject, the variant AAV5 virions encapsidating the therapeutic payload exhibit enhanced CNS tissue tropism as compared wildtype AAV5 virions encapsidating the therapeutic payload. Upon administration to the subject, a lower dose of the variant AAV5 virions encapsidating the therapeutic payload is administered as compared to wildtype AAV5 virions encapsidating the therapeutic payload. Upon administration to the subject, at least one symptom of Tauopathy disease is alleviated or the subject is cured.

Example 27
Treatment of Alpha-1 Antitrypsin Deficiency (AATD) with an AAV5-Derived Virion Encapsidating a Therapeutic Payload

This example describes treatment of AATD with a variant AAV5-derived virion having any one of the engineered CNS tropic variant AAV5 VP capsid polypeptides disclosed herein, wherein the variant AAV5 virion encapsidates a therapeutic payload. Polynucleotide sequence encoding for AAV Rep, an AAV5-derived variant Cap and helper proteins and a therapeutic payload are transfected in cells to produce variant AAV5 virions, where the polynucleotide sequence encoding for the variant AAV5 Cap comprises at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 in the VP1 capsid polypeptide and where the polynucleotide sequence encoding for the variant AAV5 Cap encodes for an amino acid sequence of any one of SEQ ID NO: 43438-SEQ ID NO: 46437. The therapeutic payload is a guide RNA targeting an mRNA encoded for by a gene implicated in AATD or a transgene. The mRNA targeted by the guide RNA is an mRNA encoded for by a gene encoding for SERPINA1. The transgene is SERPINA1. The variant AAV5 virion encapsidating the payload is administered to a subject. The subject is a human or non-human animal. The route of administration is a systemic route of administration. The systemic route of administration is intravenous administration. Upon administration to the subject, the variant AAV5 virions encapsidating the therapeutic payload exhibit enhanced CNS tissue tropism as compared wildtype AAV5 virions encapsidating the therapeutic payload. Upon administration to the subject, a lower dose of the variant AAV5 virions encapsidating the therapeutic payload is administered as compared to wildtype AAV5 virions encapsidating the therapeutic payload. Upon administration to the subject, at least one symptom of AATD is alleviated or the subject is cured.

Example 28
Treatment of Cystic Fibrosis with an AAV5-Derived Virion Encapsidating a Therapeutic Payload

This example describes treatment of cystic fibrosis with a variant AAV5-derived virion having any one of the engineered CNS tropic variant AAV5 VP capsid polypeptides disclosed herein, wherein the variant AAV5 virion encapsidates a therapeutic payload. Polynucleotide sequence encoding for AAV Rep, an AAV5-derived variant Cap and helper proteins and a therapeutic payload are transfected in cells to produce variant AAV5 virions, where the polynucleotide sequence encoding for the variant AAV5 Cap comprises at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 in the VP1 capsid polypeptide and where the polynucleotide sequence encoding for the variant AAV5 Cap encodes for an amino acid sequence of any one of SEQ ID NO: 16118-SEQ ID NO: 19117. The therapeutic payload is a guide RNA targeting an mRNA encoded for by a gene implicated in cystic fibrosis or a transgene. The mRNA targeted by the guide RNA is an mRNA encoded for by a gene encoding for CFTR. The transgene is CFTR. The variant AAV5 virion encapsidating the payload is administered to a subject. The subject is a human or non-human animal. The route of administration is a systemic route of administration. The systemic route of administration is intravenous administration. Upon administration to the subject, the variant AAV5 virions encapsidating the therapeutic payload exhibit enhanced CNS tissue tropism as compared wildtype AAV5 virions encapsidating the therapeutic payload. Upon administration to the subject, a lower dose of the variant AAV5 virions encapsidating the therapeutic payload is administered as compared to wildtype AAV5 virions encapsidating the therapeutic payload. Upon administration to the subject, at least one symptom of cystic fibrosis disease is alleviated or the subject is cured.

Example 29
Treatment of Duchenne Muscular Dystrophy with an AAV5-Derived Virion Encapsidating a Therapeutic Payload

This example describes treatment of Duchenne muscular dystrophy with a variant AAV5-derived virion having any one of the engineered CNS tropic variant AAV5 VP capsid polypeptides disclosed herein, wherein the variant AAV5 virion encapsidates a therapeutic payload. Polynucleotide sequence encoding for AAV Rep, an AAV5-derived variant Cap and helper proteins and a therapeutic payload are transfected in cells to produce variant AAV5 virions, where the polynucleotide sequence encoding for the variant AAV5 Cap comprises at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 in the VP1 capsid poly peptide and where the polynucleotide sequence encoding for the variant AAV5 Cap encodes for an amino acid sequence of any one of SEQ ID NO: 25118-SEQ ID NO: 26117, SEQ ID NO: 28991-SEQ ID NO: 31990, or SEQ ID NO: 13118-SEQ ID NO: 16117. The therapeutic payload is a guide RNA targeting an mRNA encoded for by a gene implicated in Duchenne muscular dystrophy or a transgene. The mRNA targeted by the guide RNA is an mRNA encoded for by a gene encoding for DMD. The transgene is DMD. The variant AAV5 virion encapsidating the payload is administered to a subject. The subject is a human or non-human animal. The route of administration is a systemic route of administration. The systemic route of administration is intravenous administration. Upon administration to the subject, the variant AAV5 virions encapsidating the therapeutic payload exhibit enhanced CNS tissue tropism as compared wildtype AAV5 virions encapsidating the therapeutic payload. Upon administration to the subject, a lower dose of the variant AAV5 virions encapsidating the therapeutic payload is administered as compared to wildtype AAV5 virions encapsidating the therapeutic payload. Upon administration to the subject, at least one symptom of Duchenne muscular dystrophy disease is alleviated or the subject is cured.

Example 30
Treatment of Frontotemporal Dementia with an AAV5-Derived Virion Encapsidating a Therapeutic Payload

This example describes treatment of frontotemporal dementia (FTD) with a variant AAV5-derived virion having any one of the engineered CNS tropic variant AAV5 VP capsid polypeptides disclosed herein, wherein the variant AAV5 virion encapsidates a therapeutic payload. Polynucleotide sequence encoding for AAV Rep, an AAV5-derived variant Cap and helper proteins and a therapeutic payload are transfected in cells to produce variant AAV5 virions, where the polynucleotide sequence encoding for the variant AAV5 Cap comprises at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 in the VP1 capsid polypeptide and where the polynucleotide sequence encoding for the variant AAV5 Cap encodes for an amino acid sequence of any one of SEQ ID NO: 7118-SEQ ID NO: 10117. The therapeutic payload is a guide RNA targeting an mRNA encoded for by a gene implicated in FTD or a transgene. The mRNA targeted by the guide RNA is an mRNA encoded for by a gene encoding for progranulin. The transgene is progranulin. The variant AAV5 virion encapsidating the payload is administered to a subject. The subject is a human or non-human animal. The route of administration is a systemic route of administration. The systemic route of administration is intravenous administration. Upon administration to the subject, the variant AAV5 virions encapsidating the therapeutic payload exhibit enhanced CNS tissue tropism as compared to wildtype AAV5 virions encapsidating the therapeutic payload. Upon administration to the subject, a lower dose of the variant AAV5 virions encapsidating the therapeutic payload is administered as compared to wildtype AAV5 virions encapsidating the therapeutic payload. Upon administration to the subject, at least one symptom of FTD is alleviated or the subject is cured.

Example 31
AAV5 Variants with Adrenal Gland Tissue Tropism as Identified by Machine Learning

This example describes preferred properties of engineered AAV5 variants that display adrenal gland tissue tropism as identified by machine learning methods. The same bioinformatics pre-process, machine learning methods, and analyses as described in EXAMPLE 19 and FIGS. 19-FIG. 22 were carried out to identify preferred properties of engineered AAV5 variants that display adrenal gland tissue tropism.

Favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an AAV5 VP capsid polypeptide, which are associated with a higher probability of adrenal gland tissue tropism are described below. Any of the below machine learning-derived positional amino acid preferences described in TABLE 46 can be present alone or in combination with each other or in combination with any of the amino acid preferences observed and described in EXAMPLE 7, TABLE 13. Listed below in TABLE 46 are a summary of positional features shared between the top important features for adrenal gland tropism extracted from the ML models.

TABLE 46

Machine Learning-Derived Adrenal Gland Tissue Tropism Rules

Low mol mass at Xaa1

Xaa1 is selected from V, P, S, or C

Low hydropathy at Xaa1

Xaa1 is selected from T, S, W, or Y

Low hydropathy at Xaa2

Xaa2 is selected from R

Low mutability at Xaa2

Xaa2 is selected from C

Low solubility at Xaa2

Xaa2 is selected from K

Low average flexibility at Xaa3

Xaa3 is selected from W, M, or F

High solubility at Xaa3

Xaa3 is selected from M

High surface accessibility at Xaa4

Xaa4 is selected from K or R

High average flexibility at Xaa4

Xaa4 is selected from K, I, or N

Medium mutability at Xaa5

Xaa5 is selected from R or H

High goldman engelman steitz at Xaa5

Xaa5 is selected from V or L

Low hydropathy at Xaa5

Xaa5 is selected from R

High volume at Xaa5

Xaa5 is selected from Y, R or F

High solubility at Xaa6

Xaa6 is selected from Y, V, M, A, or C

Medium mutability at Xaa7

Xaa7 is selected from V, H, or R

Low solubility at Xaa7

Xaa7 is selected from R

High average flexibility at Xaa8

Xaa8 is selected from K, I, or N

High mol mass at Xaa8

Xaa8 is selected from R, or Y

High mutability at Xaa9

Xaa9 is selected from N

TABLE 47 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that exhibited adrenal gland tissue tropism and comport to one or more of the rules provided in TABLE 46. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 2118-SEQ ID NO: 3117 as disclosed in TABLE 47. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 47

Sequences of the 581 to 589 Region in AAV5 VP1

Capsid Polypeptide that Drive Adrenal Gland

Tissue Tropism

SEQ

ID
581-589

NO
Sequence

2118
AAGGFLSGC

2119
AAVQKERYN

2120
ACYFMMRQN

2121
AFGHAMKQF

2122
AIGCIQNGG

2123
AIGCVQNGG

2124
AIGGFPNGG

2125
AKSYFQNGG

2126
AILKLTKVL

2127
AILKMTKVM

2128
AILKTTKVL

2129
AIMKMTKVL

2130
ALGHRHHQC

2131
ALNKTLAEG

2132
ANMFRNGML

2133
ANMFRNGMR

2134
ANYIQYVWE

2135
AQMQYERLF

2136
ARAPNEGVI

2137
ARQWMCVHC

2138
ARTLYMINR

2139
ARVIEHKAG

2140
ASMFWHYQV

2141
ATCKHTDHL

2142
ATCYFVSSQ

2143
ATFKHTDHF

2144
ATFKHTDHW

2145
ATLKHTDHL

2146
ATNKYANAT

2147
ATVKHTDHL

2148
AVGLNERNG

2149
AVGNRHHQC

2150
AWGMQDQRY

2151
AWMCRGVYC

2152
AWMCRHVYC

2153
AWMCRNFYC

2154
AWMCRNVDC

2155
AWMCRNVHC

2156
AWMCRNVNC

2157
AWMCRNVYY

2158
AWMCRSVYC

2159
AWMCRTVYC

2160
AWMFRNVYC

2161
AWRCRNVYC

2162
AWRCRNVYC

2163
AWVCRNVYC

2164
AWVMKDQQY

2165
AWVMKNQRY

2166
AYCHRAHNG

2167
CAGIHLNEG

2168
CAGIHMNEA

2169
CAGIHMNEA

2170
CARFVARWP

2171
CARFVEGWP

2172
CARFVERWH

2173
CARFVERWR

2174
CAVIHLNEA

2175
CAVPPSRYP

2176
CAVPTARYP

2177
CAVPTSRYL

2178
CAVPTSRYT

2179
CAVQKERYN

2180
CAVVMMRQP

2181
CCAKFKMCT

2182
CCEDMKIWQ

2183
CCVFRHHYC

2184
CDIDYDLGF

2185
CECIVQMAT

2186
CECLVQMGT

2187
CFDIIQRYD

2188
CFKFKFRLV

2189
CFKFKVRLA

2190
CFKSKFSWQ

2191
CFNIIERYD

2192
CGEDMKMWQ

2193
CGKDIKIWQ

2194
CGKDMQIWQ

2195
CHGRVAHHE

2196
CHLRDCSPI

2197
CHMDYDLGF

2198
CHRFIQPSY

2199
CHRFIQRSY

2200
CHRYIQQSY

2201
CHSRDCSPL

2202
CIKSMFSWQ

2203
CILQTRKGN

2204
CKIGTQKGQ

2205
CKIVAWKGQ

2206
CKIVIQKGQ

2207
CKIVIQKSQ

2208
CKLVTQKGQ

2209
CKMVTQKGQ

2210
CKMYMHEAR

2211
CKMYMQESR

2212
CKMYMRESR

2213
CLMALNRSG

2214
CLMAMKRSG

2215
CLMGLKRSG

2216
CLSPFARAM

2217
CLVALKRSG

2218
CMKRYMMLP

2219
CPDEPKWFP

2220
CRKQGHGEK

2221
CSIVRWMSP

2222
CSKFIVAWE

2223
CSKFIVGWE

2224
CSKFIVVWE

2225
CSSFRWMSP

2226
CSSVRWMGP

2227
CSSVRWMST

2228
CTFVMMRQP

2229
CTGQRGFTN

2230
CTGVMMRQP

2231
CTIRGNSSC

2232
CTLRGNASC

2233
CTVGMMRQP

2234
CTVLMMRQP

2235
CTVLMTRQP

2236
CTVVLMRQP

2237
CTVVMMREP

2238
CTVVMVRQP

2239
CVDTYRMWQ

2240
CVHQRMHVT

2241
CVIQTSRGT

2242
CVKFKVRLV

2243
CVLEFSRQW

2244
CVLKFSREW

2245
CVLKFSRHW

2246
CVLKFSRQG

2247
CVLKFSRQS

2248
CVLKFSRRW

2249
CVLKVSRQW

2250
CVLKYSRQW

2251
CVLQTSRGI

2252
CVMQTSRGI

2253
CVNKLSYEN

2254
CVRHTENWR

2255
CVSLFARAM

2256
CVSPFARAK

2257
CVSPFARAT

2258
CVSPFARGM

2259
CVSPFARSM

2260
CVSPFARVM

2261
CVSPFGRAM

2262
CVSPFVRAM

2263
CVSQFARAM

2264
CVSSFARAM

2265
CVTTRHKQG

2266
CVWPFKFDF

2267
CVWPMKFDF

2268
CWDVTCQEN

2269
CYGRVAHLE

2270
CYRFIQQSY

2271
CYVFRHHDC

2272
DASSNDRNT

2273
DCESPEKQN

2274
DCESSEKPN

2275
DCQSPEKPN

2276
DEDVCKFVF

2277
DEVACKFVF

2278
DEVDCKFVF

2279
DEVVCKFMF

2280
DEVVCKIVF

2281
DFKLFHNRE

2282
DFKLFHNRP

2283
DFKLFNNRA

2284
DFKVFHNRA

2285
DFQLFHNRA

2286
DFRLFHNRA

2287
DFRWFIRMN

2288
DMKRPTRYN

2289
DMQRPPRYN

2290
DMQRPSRYN

2291
KNNMIVRCV

2292
DNQWMNSLE

2293
DNSMIVRCA

2294
DQPHLHRIG

2295
DRCLVEWVS

2296
DRCLVKWVA

2297
DRMDFHFFF

2298
DSKQIAKAM

2299
DVCNVLGWK

2300
DVDVVARPM

2301
DVKLFHNRA

2302
DVRGLNREY

2303
DVRNFFDHC

2304
DWHQYDGCE

2305
DWNQHDGCE

2306
DWNQYAGCE

2307
DWNQYDGGE

2308
DWNQYNGCE

2309
DWSQYDGCE

2310
DYKQTAKAM

2311
ECAHVIKFN

2312
ECHKMRCLE

2313
ECHKMRCVE

2314
ECNKMRCIE

2315
EETKKVKHW

2316
EHKKNINMH

2317
EKRHISRAP

2318
EQRHISRAR

2319
EQRHISRTP

2320
ERVIEHRAG

2321
ERVLEHKAG

2322
EYDVNMRQM

2323
FGKDAFVSS

2324
FGKDTFASS

2325
FGKDTFLSS

2326
FGKDTFVSA

2327
FGKDTVVSS

2328
FGMDTFVSS

2329
FMMHRVMSD

2330
FMMHRVMSG

2331
FMMMAKTQA

2332
FQKLRDVCF

2333
FREQGHGEK

2334
FRKEGHGEK

2335
FRKPGHGEK

2336
FRKQAHGEK

2337
FRKQGHGAK

2338
FRKQGHGEE

2339
FRKQGHGQK

2340
FRKQGNGEK

2341
FRKQGPGEK

2342
FRKQVHGEK

2343
FRNQVHGEK

2344
FSSCCRQKG

2345
FTSNRNWEA

2346
FTSNRNWQD

2347
FTSNRNWQP

2348
GAHHRCRMT

2349
GAWMQGFHP

2350
GCAKFQMCT

2351
GCAKTRFEN

2352
GCDKNRFEN

2353
GCDKTRFAN

2354
GCDKTRFES

2355
GCDKTRFEY

2356
GCHKMRCIE

2357
GEGIRKMNN

2358
GEGIRQMNT

2359
GEGVRQMNN

2360
GEVVCKFVF

2361
GFDRQDAHR

2362
GFKLFHNRA

2363
GMKTRCDDD

2364
GMKTRCDYA

2365
GMKTRCDYV

2366
GMKTRCHYD

2367
GMKTRCNYD

2368
GMKTRCVYD

2369
GMQTRCDYD

2370
GMQTRCDYD

2371
GRAFVKIDT

2372
GRICMQGMK

2373
GRIFMLGMK

2374
GRIFVKIDT

2375
GRLFVKIDT

2376
GRWFQIGTE

2377
GTFKHTDHL

2378
GVAYRESQA

2379
GVDKLSYEN

2380
GVHHRWRMT

2381
GVKPRWGLF

2382
GVKTRCDYD

2383
GVWMQGIHP

2384
GWEKKINFQ

2385
GWEKKINGQ

2386
GWEKKLNVQ

2387
GWEKKVNVQ

2388
GWEKTINVQ

2389
GWFFKNYYC

2390
GWKKKINVQ

2391
GWMCRNVYC

2392
GWQKKINVQ

2393
HACCLLQQY

2394
HACSLLKQY

2395
HACSLLQQF

2396
HIEWVSMQR

2397
HIQWVSMHR

2398
HMEWVSMHR

2399
HRADIMCWN

2400
HREAIMCWN

2401
HREDIMCGN

2402
HREDIMCWD

2403
HREDIMCWI

2404
HREDIRCWN

2405
HRGDIMCWN

2406
HRKDIMCWN

2407
HRQDIMCWN

2408
HRTETECWC

2409
HSCSLLQQY

2410
HVGALTRSD

2411
IARHIDHDL

2412
IAWHVDHDL

2413
ICHLRHIAA

2414
ICHLRHIPV

2415
ICHLRHISG

2416
ICHLRHNAG

2417
ICHLRHVAG

2418
ICTTLIQGM

2419
ICTTQIKGM

2420
ICYKWWVMF

2421
IDAALDMSW

2422
IEVKYWNRH

2423
IEYVRSCPW

2424
IFMAFDHLE

2425
IGMDKPVDA

2426
IHMKPQESP

2427
IHMKQQDSP

2428
IHMKQQESA

2429
IHMKQQEST

2430
IHMKQQQSP

2431
IHMKQRESP

2432
IHMKTIVGE

2433
IHMRQQESP

2434
IILHNMKSW

2435
IMLALMHED

2436
IMLHRVMSD

2437
IMMAHWFAH

2438
IMMAQWFAQ

2439
IMMAQWFGH

2440
IMMAQWFSH

2441
IMMAWEIAH

2442
IMMHRMMSD

2443
IMMHRVMSE

2444
IMMHRVMSV

2445
IMMHRVMSY

2446
IMMHRVMYD

2447
IMMLRVMSD

2448
IMMNRVMSD

2449
IMMQRVMSD

2450
IMMVQWFAH

2451
IMMYRVMSD

2452
IMVALMHEG

2453
IVCRMWHQP

2454
IWHKWWVMF

2455
IWYKWWAMF

2456
IWYKWWLMF

2457
IWYKWWVMC

2458
IWYKWWVMV

2459
IWYKWWVMY

2460
KAIPENSNR

2461
KALEVHFQK

2462
KAMEFHFQK

2463
KAMPEPVTE

2464
KAQDFATEN

2465
KAQEFATED

2466
KAQEFATEH

2467
KAQKFATEN

2468
KAOQFATEN

2469
KAREFATEN

2470
KASTEAGDK

2471
KATTEASDK

2472
KCDKPMLEP

2473
KCDKPMLQP

2474
KCDTFRGME

2475
KCEPFRGME

2476
KCETFLGME

2477
KCGILYYQG

2478
KCGNLYYQG

2479
KCGSLNYQG

2480
KCGSLYCQG

2481
KCGSLYHQG

2482
KCGSLYSQG

2483
KCGSLYYQV

2484
KCGSMYYQG

2485
KCGTLYYQG

2486
KCYFMMRQN

2487
KDCQYEGAC

2488
KDCQYEGSW

2489
KDCQYLGAW

2490
KFELNQTAV

2491
KFELNRKAV

2492
KFETFRGME

2493
KFGLNQKAV

2494
KFIDVWDAR

2495
KGEYQADAP

2496
KHDPMCGQL

2497
KHEQMCVQL

2498
KHERMCVQL

2499
KHETMCVQL

2500
KHKENAVER

2501
KHKPMCVQL

2502
KHKYEDPMV

2503
KHKYEDTMG

2504
KHQINTVER

2505
KHQPMCVQL

2506
KHSSHMEAC

2507
KIHYHFEDK

2508
KINYHFEDE

2509
KINYHFEDQ

2510
KISGLNDAT

2511
KKDEANGQT

2512
KKDPMQTYP

2513
KMGTEVSQQ

2514
KMRMIFRKD

2515
KNEPMCVQL

2516
KPVEYASAP

2517
KQDFHHGLA

2518
KQLYHIPTC

2519
KQQYVTMNA

2520
KQRDESRAP

2521
KQRHISRAP

2522
KQSAHMEAC

2523
KQSPHMEAC

2524
KQSSHMEAF

2525
KRDLANGQF

2526
KREEEHPCA

2527
KRMDFHFFF

2528
KRSSHMEAC

2529
KRVIEHKAG

2530
KRYFWMSYY

2531
KSMLEDRIC

2532
KSMWEARIC

2533
KSMWEDREG

2534
KSMWEDRTC

2535
KSMWEERIC

2536
KSMWENRIC

2537
KSMWEYRIC

2538
KSVWEDRIC

2539
KTFFWMSYY

2540
KTFSESTCR

2541
KTMEVHFQK

2542
KTQWDEAIV

2543
KTYEADTIC

2544
KTYFADTES

2545
KTYFADTMG

2546
KTYFADTVG

2547
KTYFAETEG

2548
KTYFANTIG

2549
KTYFSDTEG

2550
KTYFWMSYC

2551
KTYFWMSYS

2552
KTYFWTSYY

2553
KTYFWVSYY

2554
KVDVFTVKD

2555
KVDVETFKD

2556
KVDVETVOD

2557
KVDVVTVKD

2558
KVILDSFRR

2559
KVMPEPVPE

2560
KVMPEPVTA

2561
KVMPEPVTV

2562
KVMPVPVTE

2563
KVMSEPVTE

2564
KVTWMDLED

2565
KVVLDSFRQ

2566
KVYMEPANW

2567
KYEELIKYI

2568
KYEFLLKHI

2569
KYEFLLKYL

2570
KYEFLLRYI

2571
KYEELNCNP

2572
KYEVLLKYI

2573
KYEYLLKYI

2574
KYEAVWDAR

2575
KYIDFWDAR

2576
KYIDIWDAR

2577
KYEDVWEAR

2578
KYKAPTENP

2579
KYKHPTINP

2580
KYKPLTENP

2581
KYKPPTMNP

2582
KYKRPTINP

2583
KYQPPTINP

2584
LAVDKSRLL

2585
LCGLMPRPA

2586
LCGVMPRPG

2587
LFKPEDEDR

2588
LFKPEDPDR

2589
LFKPEEQDR

2590
LHRDPMPIM

2591
LHRDQMPFM

2592
LHRDQMPLM

2593
LIMAHWLHM

2594
LIMAYWLDM

2595
LIMSHWLDM

2596
LKRPQGEAE

2597
LLMAHWLDM

2598
LQKYYMNQP

2599
LQNYRDEDG

2600
LQPWWLGMI

2601
LQTYHHVDG

2602
LQTYQPVDG

2603
LRFLSDADP

2604
LRKEGHGEK

2605
LRYLSDTDP

2606
LSTNRLVNA

2607
LSTNRMVSA

2608
LSTNRMVTA

2609
LYRDQMPIM

2610
MCGHASKAV

2611
MCGHASRAE

2612
MCGPASKAE

2613
MCHLRHAIG

2614
MCRANGNMP

2615
MCSLRHWQP

2616
MEGMWNRSN

2617
MISMLNRSN

2618
MRSEYMNES

2619
MRWILTEME

2620
MSAQRHVQT

2621
MSAQRYAQT

2622
MSAQRYVQA

2623
MSMLRMHEA

2624
NAMCFGRAR

2625
NAMCFGREK

2626
NAMCFGRTK

2627
NAMCIGRAK

2628
NAMGFGRAK

2629
NATCFGRAK

2630
NCNKCINRT

2631
NCNKCKHRT

2632
NCNKCKYRT

2633
NCRHTILYV

2634
NCSKCAKYD

2635
NCVKLECQS

2636
NELQDQCGM

2637
NEVVCKFVF

2638
NFSKCAKYV

2639
NFSKCTKYD

2640
NFSKGAKYD

2641
NGMDKAVDA

2642
NGMDKPFDA

2643
NGMDKPIDA

2644
NGMDKPVDG

2645
NGMDKPVDS

2646
NGMDKPVNA

2647
NGMDQPVDA

2648
NGMDRPVDA

2649
NGMDTPVDA

2650
NGVDKPVDA

2651
NGWESEQHM

2652
NHMKQQESP

2653
NLDMSRRAF

2654
NLDMSRRDC

2655
NMIKRCCQG

2656
NMIKRCSKG

2657
NMIKRCSQA

2658
NMIKRCSQC

2659
NMIKRCSQS

2660
NMIKRCSQV

2661
NMIKWCSQG

2662
NMVKRCSQG

2663
NNQWKNSLE

2664
NNQWTNSLE

2665
NPCKINVGT

2666
NQCKINFGT

2667
NQCKINVGN

2668
NQCKINVGP

2669
NQCKLNVGT

2670
NQLKDVAAS

2671
NQWKINVGT

2672
NRAETECWC

2673
NRCLVEWVA

2674
NRMDFHFFV

2675
NRMDFNFFF

2676
NRNKCKNRT

2677
NRPFNYFQR

2678
NRTLYMINR

2679
NSSYFEFSN

2680
NSVKLGCQN

2681
NTIKRCSQG

2682
NVDMSRRDF

2683
NVMCFGRAK

2684
NVRNFFAHC

2685
NVRNFFEHC

2686
NVSKCAKYD

2687
NVWPNIAEM

2688
NVWTNISEM

2689
NWNGRWNQC

2690
NWNKCKNRT

2691
NWPCHDNRH

2692
NWSYHDNRH

2693
NWTGRWHQC

2694
NYCDRAHNG

2695
PAVQKERYN

2696
PCFSIARYY

2697
PCYFMMRQN

2698
PKVTCRYRM

2699
PPLHSRCGM

2700
PQLHSRCRM

2701
PQLLSRCGM

2702
PQLNSRCGM

2703
PQLRSRCGM

2704
PWVMKDQRY

2705
QAYCCMHQD

2706
QCDEFKMRM

2707
OCHKMRCIE

2708
QESNRCWCD

2709
QHKERMLFD

2710
QHKERMVFD

2711
QHQKNINMH

2712
QLDEFKMRM

2713
QMSWVQNWE

2714
QMYHNHHTA

2715
QMYWAQNWE

2716
QMYWGQNWE

2717
QMYWIQNWE

2718
QMYWVQNLE

2719
OMYWVQSWE

2720
QPVTRMTNE

2721
QQLHSRCGM

2722
QQRHISRAP

2723
QRDLAHGQF

2724
QRDLANGKS

2725
QRDLANGLF

2726
QRDLANGQI

2727
QRDLANGQS

2728
QRDLANGQV

2729
QRDLANGQY

2730
QRDMANGQF

2731
ORDPANGQF

2732
QSAFRQSDS

2733
QSTFRQCDS

2734
QSTMRQYYA

2735
QSYCCMHQY

2736
QTYWVQNWE

2737
QVSGMFHPA

2738
QVYCCMHQY

2739
QVYWVQNWE

2740
QWDEFKKRM

2741
QWDEFKMRR

2742
QWDEFKVRM

2743
QWDVFKMRM

2744
RACSLLQQY

2745
RCDKTRFEN

2746
RCEAFNWRE

2747
RCELFNWRE

2748
RCEPFHWRE

2749
RCEPFNLRE

2750
RCEPFNWHE

2751
RCGHASKAE

2752
RCGPFNWRE

2753
RCGQVRNEM

2754
RCKLFDHDE

2755
RCVEDHFHK

2756
RCVLDHFHK

2757
RCVQDHFHE

2758
RCVQDHFHR

2759
RCVQDHFHT

2760
RCVQDHFNK

2761
RCVQDHIHK

2762
RCVQDHSHK

2763
RCVQDNFHK

2764
RCVQHHFHK

2765
RCVRDHFHK

2766
RCYFMMRQN

2767
RDLSLHLMR

2768
RECLIERCG

2769
RECLIERCI

2770
RECLIKRCS

2771
RECLVERCS

2772
RIERVDVFN

2773
RKDEANGQA

2774
RMDEANGQT

2775
RQCLIERCS

2776
RQMWMAHHA

2777
RRDAKVANG

2778
RRDEANGQT

2779
RREDIMCWN

2780
RRKDIMKER

2781
RRYRELTEE

2782
RSEPFNWRE

2783
RTDAIHHQV

2784
RTDAIHPQI

2785
RWVQDHFHK

2786
RYCQVWGCA

2787
RYDLFSYHQ

2788
RYDLVSHHQ

2789
RYDLVSYHR

2790
RYDLVSYYQ

2791
RYNLVSYHQ

2792
RYRQEWGCA

2793
SAGQKERYN

2794
SA1QKERYN

2795
SALYGSKAI

2796
SAVQQERYN

2797
SAVQRERYN

2798
SAVRNERYN

2799
SAWYGSKSI

2800
SCKGFHLQN

2801
SCKSHQMHE

2802
SCRHIYREA

2803
SCRHLYHEA

2804
SCRHNYHEA

2805
SCRRIYHEA

2806
SFDRQDVHR

2807
SFDRQGAHR

2808
SFGHAMKQC

2809
SFKGFHLEN

2810
SFKGFHVQN

2811
SFMGFHLQN

2812
SFVRQDAHR

2813
SGQKFIFCP

2814
SIGCFQNGG

2815
SIIFSNTCM

2816
SILLPGIQG

2817
SIMHEGVQS

2818
SIMLPGIQA

2819
SLNQAIHLE

2820
SMGPTVHEM

2821
SMNQAVHLE

2822
SMNQPIHLE

2823
SMRSQHADY

2824
SNCALHNAW

2825
SNCAQHNSW

2826
SNCAQYNAW

2827
SNCSQHNAW

2828
SNCVQHNAW

2829
SNGAQHNAW

2830
SNGFEWACE

2831
SNGFEWVCA

2832
SNGFEWVGE

2833
SNGFEWVSE

2834
SNGFEWVYE

2835
SNMFRNGMQ

2836
SNVFEWVCE

2837
SRDEAKSWV

2838
SRICMLGMK

2839
SRIILDYYS

2840
SRPFNYIQR

2841
SRSFNYFQR

2842
SRWCQIGTE

2843
SRWFKIGTE

2844
SRWFQIGTK

2845
SSGDRRSDS

2846
SSKGFHLQN

2847
SSKQDFYGL

2848
SSKSHRMHE

2849
SSMLEGVQS

2850
SSVDRRSDG

2851
SSVHALYFP

2852
SSVHTLYYP

2853
SSVQKERYN

2854
STCYFVASL

2855
STDPYTSHH

2856
STFKHTDHL

2857
STGFEWVCE

2858
STSYFVASQ

2859
SVGPTVHQM

2860
SVSKFQADK

2861
SVVQKERYN

2862
SWEKKINVQ

2863
SWGLIHGWN

2864
SWGLIHWWN

2865
SWGLIHYWN

2866
SWGLVHCWN

2867
SWKWDKYCN

2868
SWMCRNVYC

2869
SWRHIYHEA

2870
SWRTRGNFE

2871
SWSKCNMEN

2872
SWVLIHCWN

2873
SYDRQDAHR

2874
SYVFRHHYC

2875
TACATYRSF

2876
TACMTYRSF

2877
TACVAYRSF

2878
TACVNYRSF

2879
TACVTYRGL

2880
TACVTYR1F

2881
TAFVTYRSF

2882
TAFVTYRSL

2883
TAGHEGHSC

2884
TAGHQGHIC

2885
TAGVTYRSF

2886
TAITMWQYM

2887
TAMCFGRAK

2888
TAMSMCMYG

2889
TAMTICMYG

2890
TANDHRVDA

2891
TATDRRVDA

2892
TAVQKERYN

2893
TAVSICMYG

2894
TAVTMWQSM

2895
TAWVTYRSF

2896
TAYVTYRSF

2897
TCCFMMRQN

2898
TCCSIARYY

2899
TCDFMMRQN

2900
TCFFMMRQN

2901
TCFGIARYY

2902
TCFNIARYY

2903
TCFSIARCY

2904
TCFSIARYD

2905
TCFSIARYF

2906
TCFSIARYS

2907
TCFSIDRYY

2908
TCFSIGRYY

2909
TCFSITRYY

2910
TCFSIVRYY

2911
TCFSNARYY

2912
TCFSTARYY

2913
TCFSVARYY

2914
FCFTIARYY

2915
TCFTVARYY

2916
TCGSLYYQG

2917
TCHFALHGC

2918
TCHFMMRQN

2919
TCHYALHAC

2920
TCNKCKNRT

2921
TCRYALHGC

2922
TCSFMMRQN

2923
TCSRFDNQA

2924
TCVSLARYY

2925
TCWALFRGE

2926
TCWDLFRGG

2927
TCYFMMGQN

2928
TCYFMMREN

2929
TCYFMMRKN

2930
TCYFMMRPN

2931
TCYFMMRQD

2932
TCYFMMRQK

2933
TCYFMMRQT

2934
TCYFMMRRN

2935
TCYFMVRQN

2936
TCYSIARYY

2937
TCYSMMRQN

2938
TCYVMMRQN

2939
TCYYMMRQN

2940
TDCVTYRSF

2941
TFDLSNKVT

2942
TFGWFMRIP

2943
TFKGFHLQN

2944
TFSKCAKYD

2945
TFYFMMRQN

2946
TGCVTYRSF

2947
TGGLNERNG

2948
TGGTFVRTM

2949
TGMDKPVDA

2950
TGQATERAL

2951
TGTDHRVDA

2952
TGWSTCMIP

2953
TITKPINLF

2954
TITKPSHLF

2955
TLNKTLAES

2956
TNTCECVDR

2957
TNYIQYVWD

2958
TPLMMCRAS

2959
TPMMMCQAS

2960
TRCYFVASQ

2961
TRDAKVSNG

2962
TREWMCVHC

2963
TRHMGVSAG

2964
TRKWMCVHC

2965
TRLWMCVHC

2966
TRMDFHFFF

2967
TRPWMCVHC

2968
TRQRMCVHC

2969
TRQWMCAHC

2970
TRQWMCIHC

2971
TROWMCLHC

2972
TRQWMCVHG

2973
TRQWMCVHW

2974
TRQWMCVYC

2975
TRQWMGVHC

2976
TRQWMWVHC

2977
TRTLYMIDR

2978
TRTLYRINR

2979
TRTPYMINR

2980
TRWFQIGTE

2981
TRYFMMRQN

2982
TSCDRAHNA

2983
TSCVTYRSF

2984
TTCFEVASQ

2985
TTCFIVASQ

2986
TTCYFVASE

2987
TTCYFVASP

2988
TTCYFVASW

2989
TTCYFVGSQ

2990
TTCYFVSSQ

2991
TTCYFVVSQ

2992
TICYIVASE

2993
TFCYLVASP

2994
TTFKHTDHL

2995
TTGYFVASQ

2996
TTRYFVASQ

2997
TTWYFVASQ

2998
TTYYFVASQ

2999
TVGLNERNA

3000
TVGLNERNS

3001
TVGLNKRNG

3002
TVGVNERNG

3003
TVKMHFAAP

3004
TVKTNFTSK

3005
TVOSNEFSK

3006
TWALAAKYE

3007
TWFSIARYY

3008
TWKWDEYCN

3009
TWKWDKYCH

3010
TWKWDKYYN

3011
TWKWDRYCN

3012
TWNKCNMHV

3013
TWPYHDNRH

3014
TWQTKDFYN

3015
TWSKCNLHV

3016
TWSKCNMRV

3017
TWSKCNVHV

3018
TWSKCSMHV

3019
TWSKCTMHV

3020
TWSKFNMHV

3021
TWSRCNMHV

3022
TWTYCEQRH

3023
TYCDRADNG

3024
TYCDRAHDV

3025
TYCDRAHNA

3026
TYCDRTHNG

3027
TYFDRAHTG

3028
TYWDRAHNG

3029
TYYDRAHNG

3030
VACRMWHQP

3031
VAEKRMDRS

3032
VARHVDHDL

3033
VAWSNEDQK

3034
VCHTFLRLG

3035
VFCRMWHQP

3036
VGEKRMDRS

3037
VHEQLEGCK

3038
VILKMTKVL

3039
VLGNRHHQC

3040
VLKKFGHSF

3041
VLMQYERLF

3042
VLNKTLAES

3043
VLPKFGHSF

3044
VLQKFGHSV

3045
VLRKFGHSF

3046
VMKTRCDYD

3047
VMMHRVMSD

3048
VPIEYMTEK

3049
VPIPYMTEK

3050
VPIQYMAEK

3051
VPMQYMTEK

3052
VQEQLESCK

3053
VQKLRDLCC

3054
VQMQYARLF

3055
VQMQYERLY

3056
VQMQYQRLF

3057
VQMQYVRLF

3058
VQTYQHVDG

3059
VQVQYERLF

3060
VRIQYMTEK

3061
VRMQYERLF

3062
VSAECYFDM

3063
VSDEFYFDM

3064
VSDVCYFDM

3065
VSGECYFDM

3066
VTWMVRTSN

3067
VVCRMWYQP

3068
VVEKRMERS

3069
VVFRMWHQP

3070
VVWMQGFHP

3071
VVYRMWHQP

3072
VWRYFCVSL

3073
VWYKWWVMF

3074
WATKWDVHR

3075
WCNTRLAMA

3076
WCTSRLAMA

3077
WCTTRLTMA

3078
WCTTRLVMA

3079
WWMHGRGGR

3080
YACSLLQQY

3081
YAVQKERYN

3082
YCHKTAVAA

3083
YCHQQDRIT

3084
YCNKTAVAA

3085
YCYKNAVAA

3086
YCYKTAVAV

3087
YCYRTAVAA

3088
YGEAOITSP

3089
YIKQICADA

3090
YIKQICSDS

3091
YINKNPHQM

3092
YLHKNPHQM

3093
YLNKNHHQM

3094
YLNKNLHQM

3095
YLNKNPHQR

3096
YLNKNPHQV

3097
YLNQNPHQM

3098
YLTKNPHQM

3099
YMIKRCSQG

3100
YPTQRLHHL

3101
YRDEAKCWV

3102
YRDEAKSWA

3103
YRDEAQSWV

3104
YRDEDKSWV

3105
YRGEAKSWV

3106
YRHEAKSWV

3107
YRKQGHGEK

3108
YRMEYGTCK

3109
YRMEYGTWE

3110
YSAQLRQQM

3111
YSVDRRSDS

3112
YWCPNQWWR

3113
YWCPSKWWR

3114
YWCPSQLWR

3115
YWCPSQWLR

3116
YWGLIHCWN

3117
YWGPSQWWR

Example 32
AAV5 Variants with Bone Marrow Tissue Tropism as Identified by Machine Learning

This example describes preferred properties of engineered AAV5 variants that display bone marrow tissue tropism as identified by machine learning methods. The same bioinformatics pre-process, machine learning methods, and analyses as described in EXAMPLE 19 and FIGS. 19-FIG. 22 were carried out to identify preferred properties of engineered AAV5 variants that display bone marrow tissue tropism.

Favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an AAV5 VP capsid polypeptide, which are associated with a higher probability of bone marrow tissue tropism are described below. Any of the below machine learning-derived positional amino acid preferences described in TABLE 48 can be present alone or in combination with each other or in combination with any of the amino acid preferences observed and described in EXAMPLE 18, TABLE 35. Listed below in TABLE 48 are a summary of positional features shared between the top important features for bone marrow tropism extracted from the ML models.

TABLE 48

Machine Learning-Derived Bone Marrow Tissue Tropism Rules

High hydropathy at Xaa1

Xaa1 is selected from V, I, or L

Low mutability at Xaa1

Xaa1 is selected from Y, L, F, or C

Low hydropathy at Xaa2

Xaa2 is selected from Y or W

High mol mass at Xaa2

Xaa2 is selected from W

Low surface accessibility at Xaa2

Xaa2 is selected from W or A

Low hydrophilicity at Xaa2

Xaa2 is selected from W

Low mutability at Xaa2

Xaa2 is selected from C

Low average flexibility at Xaa2

Xaa2 is selected from W, M, or F

Low average flexibility at Xaa5

Xaa5 is selected from W, M, or F

Low average flexibility at Xaa6

Xaa6 is selected from W, M, or F

Low mutability at Xaa6

Xaa6 is selected from Y, F, L, or C

High solubility at Xaa6

Xaa6 is selected from W, F, I, or L

Low surface accessibility at Xaa7

Xaa7 is selected from C

Or

High surface accessibility at Xaa7

Xaa7 is selected from D or N

Low mutability at Xaa7

Xaa7 is selected from C

High solubility at Xaa7

Xaa7 is selected from C

Or

Low solubility at Xaa7

Xaa7 is selected from D

Low charge at Xaa8

Xaa8 is selected from D or E

High mutability at Xaa8

Xaa8 is selected from D, E, A, or T

High mol mass at Xaa9

Xaa9 is selected from H or F

Low mutability at Xaa9

Xaa9 is selected from Y, F, or L

TABLE 49 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that exhibited bone marrow tissue tropism and comport to one or more of the rules provided in TABLE 48. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 5118-SEQ ID NO: 6117, as disclosed in TABLE 49. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 49

Sequences of the 581 to 589 Region in AAV5

VP1 Capsid Polypeptide that Drive Bone Marrow

Tissue Tropism

SEQ

ID
581-589

NO
Sequence

5118
AADVVPCHH

5119
ACFHVAEMF

5120
AEMHMVADF

5121
AESLCNELC

5122
AKKSCQCWH

5123
ANDYDQCCM

5124
ANGHKFNEE

5125
ANNEFIQPF

5126
ANQCIFKTM

5127
AQCLCNELC

5128
AQGLCNELC

5129
AQHDWYQKY

5130
AQPNMDYVP

5131
AQPSMDCVP

5132
AQPSMDYAP

5133
AQPSMDYVA

5134
AQPSMEYVP

5135
AQSLCNELF

5136
AQSLCNELG

5137
AQSLCNQLC

5138
AQSLFNELC

5139
AQSVCNELC

5140
AQTLCNELC

5141
ARAKIQDEW

5142
ARALIQDEW

5143
ARARIQDEW

5144
ARGQIQDEW

5145
ARPSMDYVP

5146
ARVQIQDEW

5147
AWESSSIIS

5148
AYAENWCIE

5149
CANHMIRTR

5150
CATHMIRTK

5151
CCEHKEYLN

5152
CCEPKGYLN

5153
CCRKDNKKS

5154
CCRMANKKS

5155
CCRMDDKKS

5156
CCRMDHKKS

5157
CCRMDIKKS

5158
CCRMDNKES

5159
CCRMDNKIS

5160
CCRMDNKKA

5161
CCRMDNKRS

5162
CCRMDNQKS

5163
CCRRDNKKS

5164
CDAEAEFQR

5165
CDAEDEFRR

5166
CDAKAEFRR

5167
CDAVAEFRR

5168
CDDEAEFRR

5169
CDGEAEFLR

5170
CDGEAEFRR

5171
CDGVAEFRR

5172
CFKQKHNLF

5173
CFLQKHNLF

5174
CFQEKHNLF

5175
CFQPKHNLF

5176
CFQQKHNIF

5177
CFQQKHNLI

5178
CFQQNHNLF

5179
CFQRKHNLF

5180
CGYJEFMVG

5181
CHCDAVECF

5182
CHFDAMECF

5183
CHKCIGQDN

5184
CHQCIGPDN

5185
CHQCIGQDS

5186
CHQCIGQDT

5187
CHQCVGQDN

5188
CHWDAMECF

5189
CIKIQLIDQ

5190
CIKIQMIDK

5191
CIKIRMIDQ

5192
CIKVQMIDQ

5193
CINITAITP

5194
CISITVITP

5195
CMAHIWEAH

5196
CMAHIWEGD

5197
CMDHIWEGH

5198
CMSHIWEGH

5199
CPQCIGQDN

5200
CRCDAMECF

5201
CWYDEFMVG

5202
CWYHEFMVA

5203
CWYHEFMVD

5204
CWYHEFMVR

5205
CWYHEFMVS

5206
CWYHEFMVV

5207
CWYHEFVVG

5208
CWYHEHMVG

5209
CWYHEVMVG

5210
CWYHKFMVG

5211
CWYLEFMVG

5212
CWYNEFMVG

5213
CWYPEFMVG

5214
CWYREFMVG

5215
CWYYEFMVG

5216
DACDWKNDY

5217
DACQEWDSE

5218
DADFCARMP

5219
DADFCPRMA

5220
DADFCPRMS

5221
DADFCPRMT

5222
DARFDQWHY

5223
DCAENWCIE

5224
DCIHDQCSS

5225
DDDCWVFYH

5226
DDDFWVFYH

5227
DDDHWVFYH

5228
DDDYWAFYH

5229
DDDYWGFYH

5230
DDGSWVFYH

5231
DFAENWCIE

5232
DGDYWVFYH

5233
DGGCWPREC

5234
DHAENWCIE

5235
DHELTGSEA

5236
DITQLQCYC

5237
DKNEMFMKW

5238
DLDCDWLHS

5239
DMDCEWLHS

5240
DPCQDWDSE

5241
DPCQEWNSE

5242
DPCQKWDSE

5243
DPCQVWDSE

5244
DPCREWDSE

5245
DPGQEWDSE

5246
DQCQEWDSE

5247
DRCQEWDSE

5248
DSCQEWDSE

5249
DSRFDQWHH

5250
DTCQEWDSE

5251
DYAEHWCIE

5252
DYAEIWCIE

5253
DYAEKWCIA

5254
DYAENGCIE

5255
DYAENWCIA

5256
DYAENWCIG

5257
DYAENWCME

5258
DYAENWCNE

5259
DYAENWCVE

5260
DYAENWWIE

5261
DYAETWCIE

5262
DYAQNWCIE

5263
DYAVNWCIE

5264
DYDENWCIE

5265
DYGENWCIE

5266
DYPENWCIE

5267
DYTENWCIE

5268
DYVENWCIE

5269
EAEDE1OKS

5270
EAEDE1QRA

5271
EAGDE/QKA

5272
EATGMLYEE

5273
EAWNAHEEG

5274
ECESSS1IS

5275
EC1HAQCSS

5276
ECIHDKCSS

5277
ECIHDQCCS

5278
EC1HDQCSA

5279
EC1HDQCSP

5280
EC1HDQCST

5281
EC1HDQCTS

5282
ECIHHQCSS

5283
ECIHVQCSS

5284
EDFHEFAQT

5285
EDGHKFNEE

5286
EEKSCQCWH

5287
EGIHDOCSS

5288
EHAEWKCMY

5289
EHHEWKCMY

5290
EHPEGKCMY

5291
EHPELKCMY

5292
EHPEWKCLC

5293
EHPEWKGMY

5294
EHPEWTCMY

5295
EHREWKCMY

5296
EHSEWKCMY

5297
EHTEWKCMY

5298
EINQSTCET

5299
EKDQSTCET

5300
EKESCQCWH

5301
EKHQSTCET

5302
EKINWKRPF

5303
EKIMVKRTF

5304
EKIMVKWAF

5305
EKKACQCWH

5306
EKKPCQCWH

5307
EKKSCKCWH

5308
EKKSCLCWH

5309
EKKSCPCWH

5310
EKKSCQCWD

5311
EKKSCQCWN

5312
EKKSCRCWH

5313
EKKSFQCWH

5314
EKKSWQCWH

5315
EKKSYQCWH

5316
EKKTCQCWH

5317
EKLLSLCPS

5318
EKNESTCET

5319
EKNQCTCET

5320
EKNQGTCET

5321
EKNQSTCEA

5322
EKNQSTCEP

5323
EKNQSTCES

5324
EKNQSTCGT

5325
EKNVWNNKW

5326
EKQSCQCWH

5327
EKRSCQCWH

5328
EKTQSTCET

5329
EKTSCQCWH

5330
ELPEWKCMY

5331
ENAHKFNEE

5332
ENGDKFNEE

5333
ENGHEFNEE

5334
ENGHKFDEE

5335
ENGHKFNAE

5336
ENGHKFNEA

5337
ENGHKFNEK

5338
ENGHKFNKE

5339
ENGHKFNQE

5340
ENGHKESEE

5341
ENGHKFTEE

5342
ENGHQFNEE

5343
ENGHRFNEE

5344
ENPEWKCMY

5345
ENVHKFNEE

5346
EQHDWCQKY

5347
EQHDWYKKY

5348
EQHDWYLKY

5349
EQHHWYQKY

5350
EQHNWYQKY

5351
EQKSCQCWH

5352
EQLLSLCHS

5353
EQLLSLCLS

5354
EQLLSLCPN

5355
EQLLSLCSS

5356
EQLLTLCPS

5357
EQLSSLCPS

5358
EQNDWYQKY

5359
EQPLSLCPS

5360
EQVLSLCPS

5361
ERIHDQCSS

5362
ERKSCQCWH

5363
ERNQSTCET

5364
ESEDEIQKA

5365
ESGHKFNEE

5366
ESIGMVCSD

5367
ETALKWEWS

5368
ETAWKWDWS

5369
ETGHKFNEE

5370
ETSWKWEWS

5371
EVPWESKLC

5372
EVWDAHEEG

5373
EVWNSHEEG

5374
EWEGSSEIS

5375
EWESNSIIS

5376
EWESSSIIA

5377
EWESSSTIS

5378
EWETSSIIS

5379
EWGFGSDET

5380
EWIHDQCSS

5381
EWLPWINER

5382
FEMAMCSMG

5383
FKIRKFWDP

5384
FKMAMCSMG

5385
FKVGKFWDP

5386
FKVIKFWDP

5387
FKVRKFLDP

5388
FKVRKFWDS

5389
FQMAMGSMG

5390
FQMSMCSMG

5391
FQVRKFWDP

5392
FRIIWGADM

5393
FRILWQCEG

5394
FRMAMCSMG

5395
FRPIWGADM

5396
FRPIWGGDM

5397
FRTIWGADL

5398
FRTIWGADV

5399
FRTIWGPDM

5400
FRTIWGTDM

5401
FRVRKFWDP

5402
FVDSMLHKW

5403
FWDRQWDDF

5404
FWLGLFPTK

5405
FWLGVFPAK

5406
FWMGLFPAK

5407
GATHMIRTR

5408
GCEAKEYLN

5409
GCLTWAFRG

5410
GCVTWAFRC

5411
GCVTWDFRG

5412
GDHDMLNSL

5413
GEQEARNCS

5414
GEREGRNCS

5415
GEYLRHTVT

5416
GEYLRNTIT

5417
GEYLRSTVT

5418
GEYQRNTVT

5419
GGAAMWGRY

5420
GGAAMWRKY

5421
GGAGMWRRY

5422
GGASMWRRY

5423
GGATMWRRY

5424
GGDAMWRRY

5425
GGEERVGEA

5426
GGGAMWRRY

5427
GIEGMYWPL

5428
GILIMAYSW

5429
GLDCEWLHS

5430
GLLYDFNPE

5431
GLLYDFNQG

5432
GLPHSADVI

5433
GLPQSADGE

5434
GLPRSADVE

5435
GLSVMTYTS

5436
GMHHAVWAS

5437
GMNHAVGAS

5438
GMNHAVWAN

5439
GMNHAVWTS

5440
GMNHSVWAS

5441
GMNNAVWAS

5442
GMNQAVWAS

5443
GMNYAVWAS

5444
GMPQSADVI

5445
GMSHAVWAS

5446
GMTHAVWAS

5447
GNGHKFNEE

5448
GQSLCNELC

5449
GVPQSADVE

5450
GWMCEVPWT

5451
GWMWFVHWT

5452
GWMWEVPWR

5453
GWMWEVTWT

5454
HDEMMGLER

5455
HDIMFAHQM

5456
HDIMFSHQM

5457
HDEMFTHKM

5458
HDIMFTHKR

5459
HDIMFTHQR

5460
HDIMFTYQM

5461
HDKMMGLER

5462
HQLMFTHQM

5463
HDQMMDLER

5464
HDQMMGMER

5465
HNIMFTHQM

5466
IADVVPCHH

5467
IAMLESWWS

5468
ECFHVAEMF

5469
EDMLESWWG

5470
EGEAQDCNP

5471
EGMIRATRE

5472
IGMIRDARE

5473
IGMIRDTHE

5474
IGMIWDTRE

5475
IGMLRDTRE

5476
IGVMLNAMK

5477
IGVMLNDMQ

5478
ILHMMHRGA

5479
IMAPLVCDD

5480
IMAPENCDG

5481
IMEALVCDD

5482
IMEHLVCDD

5483
IMEHMVCDD

5484
IMEPLACDD

5485
IMEPLVCAD

5486
IMEPLVCDA

5487
IMEPLVCDN

5488
IMEPLVCDV

5489
IMEPLVCGD

5490
IMEPLVCND

5491
IMEPPVCDD

5492
IMEPRVCDD

5493
IMEPVVCDD

5494
IMERLVCDD

5495
IMERMVCDD

5496
IMKPLVCDD

5497
IMKQWPAMF

5498
IMPQWPAMF

5499
IMQEWPAMF

5500
IMQPLVCDD

5501
IMQPWPAMF

5502
IMQQWLAMF

5503
IMQQWPAKF

5504
IMQQWPAMY

5505
IMQQWPDMF

5506
IMQQWPSMF

5507
IMQQWPTMF

5508
IMRQWPAMF

5509
IMVPLVCDD

5510
INNEFIQPF

5511
IPDLMHNVH

5512
IPGLMSDYQ

5513
IRQQWPAMF

5514
IRTIWGADM

5515
ISALMSDYQ

5516
ISDLMHDVH

5517
ISDLMHNAH

5518
ISDLMHNGH

5519
ISDLMHNVN

5520
ISDLMHSVH

5521
ISDLMHYVH

5522
ISDLMNNVH

5523
ISDLVHNVH

5524
ISDMMHNVH

5525
ISELMSDYQ

5526
ISGLMSDDQ

5527
ISGLMSDHQ

5528
ISGLMSDYE

5529
ISGLMSDYK

5530
ISGLTSDYQ

5531
ISGLVSDYQ

5532
ISNLMHNVH

5533
ISQHTCNKD

5534
ISQHTCSED

5535
ISQQTCSKD

5536
ISRHTCSKD

5537
ISSADSWPM

5538
ISVLMSDYQ

5539
ITCIEAQEN

5540
ITCIEGLEN

5541
ITCIEGQED

5542
ITCIEGREN

5543
ITCLEGQEN

5544
ITEPLVCDD

5545
ITQHTCSKD

5546
IWARQWDDF

5547
IWDHOWDDF

5548
IWDMMHRGA

5549
IWDRKWDDF

5550
IWDRLWDDF

5551
IWDRPWDDF

5552
IWDRQLDDF

5553
IWDRQWADF

5554
IWDRQWDAF

5555
IWDRQWDDY

5556
IWDRQWGDF

5557
IWDRQWNDF

5558
IWDRQWYDF

5559
IWDSQWDDF

5560
IWHRQWDDF

5561
IWKLPSVIT

5562
IWKMPPIDM

5563
IWMLPSETT

5564
IWMLPSVAT

5565
IWMLPSV1T

5566
IWVRQWDDF

5567
IWYRQWDDF

5568
KCCLGKNSR

5569
KFHSKERMS

5570
KENEFAYTY

5571
KINIFEYTC

5572
KENVFEYTY

5573
KETIFEYTY

5574
KKDVWNNKW

5575
KKHVWNNKW

5576
KKNAWNNKW

5577
KKNEMFMKW

5578
KKNEWNNKW

5579
KKNGWNNKW

5580
KKNVWDNKW

5581
KKWAIHNKW

5582
KKWAINNEW

5583
KKNVWNSKW

5584
KKNVWNTKW

5585
KKNVWNYKW

5586
KKNVWSNKW

5587
KKNVWYNKW

5588
KKSVWNNKW

5589
KKYVWNNMI

5590
KLLPWINER

5591
KLNIFEYTY

5592
KNDHFVCYL

5593
KNGHKENEE

5594
KNNEFIQPF

5595
KPVQCACEN

5596
KQHDWYQKY

5597
KRLPWINER

5598
KTNVWNNKW

5599
KWGFGSDET

5600
KWLPCINER

5601
KWLPGINER

5602
KWLPWIHER

5603
KWLPWIKER

5604
KWLPWISER

5605
KWLPWVNER

5606
KWLQWINER

5607
KWLRWINER

5608
KWMPWINER

5609
KWPPWINER

5610
KWQPWINER

5611
KWRPWINER

5612
LADSMLHKW

5613
LAGQMDEMS

5614
LAIQPCDER

5615
LATGMLYEE

5616
LDDSMLHKW

5617
LDQMMGLER

5618
LGIQPCDER

5619
LIDSMLHKW

5620
LILIMAYSW

5621
LPAWQQHGR

5622
LQGGFDHLK

5623
LQGGFDHLP

5624
LQGGFDHLS

5625
LSASMEYEE

5626
LSGLIDEMS

5627
LSGQIAEMS

5628
LSGQIDEMA

5629
LSGQIDEMP

5630
LSGQIDEVI

5631
LSGQIDEMY

5632
LSMVFSRGE

5633
LSVQIDEMS

5634
LSVVFSKGE

5635
LVDIMLHKW

5636
LVDSMLHEW

5637
LVDSMLHRW

5638
LVDSMLRKW

5639
LVDSMPHKW

5640
LVDTMLHKW

5641
LVHSMLHKW

5642
LVIEPCDER

5643
LVIKPCDER

5644
LVIQACDER

5645
LVIQHCDER

5646
LVIQPCAER

5647
LVIQPCDEG

5648
LVIQPCDEH

5649
LVIQPCDEP

5650
LVIQPCDKR

5651
LVIQPCNER

5652
LVIORYDER

5653
LVIQSCDER

5654
LVIRPCDER

5655
LVLQPCDER

5656
LVMQPCDER

5657
LVNSMLHKW

5658
LVTQPCDER

5659
LVVQPCDER

5660
LWAAMEYEE

5661
LWAFLEYEE

5662
LWAFMEYEE

5663
LWASMAYEE

5664
LWASMEFEE

5665
LWASMEHEE

5666
LWASMESEE

5667
LWASMEYDK

5668
LWASMEYEA

5669
LWASMEYEQ

5670
LWASMEYEV

5671
LWASMGYEE

5672
LWASMKYEE

5673
LWASVEYEE

5674
LWENYHSEF

5675
LWEQWCWEF

5676
LWESKENKS

5677
LWGSMEYEE

5678
LWKEWCWEF

5679
LWKIMCWEF

5680
LWKMPPIDM

5681
LWKPRHYDF

5682
LWKPWCWEF

5683
LWKQGCWEF

5684
LWKQLCWEF

5685
LWKQSCWEF

5686
LWKQWCGEF

5687
LWKQWCLEF

5688
LWKQWCSEF

5689
LWKQWCWEC

5690
LWKQWCWEI

5691
LWKQWCWES

5692
LWKQWCWEV

5693
LWKQWCWEY

5694
LWKOWCWGF

5695
LWKQWCWKF

5696
LWKQWFWEF

5697
LWKQWGWEC

5698
LWKRWCWEF

5699
LWMQWCWEF

5700
LWPSMEYEE

5701
LWQQWCWEF

5702
LWRQWCWEF

5703
LWTQWCWEF

5704
LWTSMEYEV

5705
LWVRQWDDF

5706
LWVSMEYEE

5707
LYEIRFSDM

5708
LYEIRFVDM

5709
LYKIRFADM

5710
LYKQAPSEG

5711
LYMPAPSEG

5712
LYMQATSEG

5713
LYVIRFADM

5714
LYVQAPSEG

5715
MAEPMGCQA

5716
MAEQMGCRA

5717
MAEQMVCQA

5718
MAGDCACAM

5719
MDHPNDHFF

5720
MDHTNDLFF

5721
MEWQSEYVS

5722
MGEQMGCQA

5723
MGEVMDFLP

5724
MKNVWNNKW

5725
MLGHANDGL

5726
MLGHASDAL

5727
MMEPLVCDD

5728
MSAVMDFLP

5729
MSEIMDFLP

5730
MSEVMDFMP

5731
MSEVMDFPP

5732
MSEVMDFQP

5733
MSEVMDFRP

5734
MSGVMDFLP

5735
MSQVMDFLP

5736
MSWNWWALY

5737
MSWNWWNLY

5738
MSWNWWTVY

5739
MSWTWWTLY

5740
MTEQMGCQA

5741
MWASMEYEE

5742
MWDRQWDDF

5743
MWKQWQNEF

5744
NACDLKNDY

5745
NACDWKSDY

5746
NACDWKTDY

5747
NACDWKYDY

5748
NACDWRNDY

5749
NADVVPCHH

5750
NAYDWKNDY

5751
NCFHVAEMF

5752
NFFDSEDMG

5753
NGGCWAREC

5754
NGGCWLREC

5755
NGGCWPWEC

5756
NGGCWSREC

5757
NGGCWTREC

5758
NGGFWPREC

5759
NGGGWPREC

5760
NINQLQCYC

5761
NITQLQCCC

5762
NKHEMFMKW

5763
NKNAMFMKW

5764
NKNGMFMKW

5765
NKNVWNNKW

5766
NLDCEWLHS

5767
NMEPLVCDD

5768
NMTQLQCYC

5769
NNDDFVCYL

5770
NNDHCVCYL

5771
NNDHFICYL

5772
NNDHFLCYL

5773
NNDHFVCFL

5774
NNDHFVCHL

5775
NNDHFVCNL

5776
NNDHFVCYM

5777
NNDHIVCYL

5778
NNDHSVCYL

5779
NNDHVVCYL

5780
NNDHYVCYL

5781
NNDRFVCYL

5782
NNHHFVCYL

5783
NPCQENDSE

5784
NRGMNAWTA

5785
NSCDWKNDY

5786
NTCIEGQEN

5787
NVGQITQDC

5788
NWFDSEDMG

5789
NYAENWCIE

5790
NYRVHGGAG

5791
NYRVHGSAA

5792
NYRVRGSAG

5793
PATGMLYEE

5794
PKELFPTCM

5795
PLCHVMADR

5796
PMDWVAECR

5797
PNNEFIQPF

5798
PQCHVMDDR

5799
PQCHVMSDR

5800
PQCHVTADR

5801
PQCLVMADR

5802
PQCRVMADR

5803
PQSLCNELC

5804
PQYHVMADR

5805
PVIQPCDER

5806
PWGFGSDET

5807
PWKIDFWYH

5808
PWKVDFWDH

5809
PWLSEFMHW

5810
QAAGMLYEE

5811
QANGMLYEE

5812
QATRMLYEE

5813
QATVMLYEE

5814
QDFHEFAET

5815
QDFHEFAPT

5816
QDFHEFAQA

5817
QDFHEFDQT

5818
QDFHEFPQT

5819
QDFPEFAQT

5820
QDIHFFAQT

5821
QFMCEIECR

5822
QFMFEIECK

5823
QHPEWKCMY

5824
QKGISEFQR

5825
QKNVWNNKW

5826
QMDWVAGCR

5827
QMDWVGECR

5828
QNCFFMHWW

5829
QNGHKFNEE

5830
QNWFFLHWW

5831
QPTGMLYEE

5832
QQCHVMADR

5833
QQHDWYQKY

5834
QVPWESRLC

5835
QVTGMLYEE

5836
QVWNAHEEG

5837
QWAFGSDET

5838
QWESKEMGS

5839
QWESKEMYS

5840
QWGCGSDET

5841
QWGFASDET

5842
QWGFESDET

5843
QWGFGADET

5844
QWGFGSDEA

5845
QWGFGSDEN

5846
QWGFGSDEP

5847
QWGFGSGET

5848
QWGFGSHET

5849
QWGFGSNET

5850
QWGFGSVET

5851
QWGFVSDET

5852
QWGIGSDET

5853
QWGSGSDET

5854
QWGVGSDET

5855
QWGYGSDET

5856
QWKQWCWEF

5857
QWVFGSDET

5858
RDFHEFAQT

5859
RDIMFTHQM

5860
RHELLEWDS

5861
RHQLLEWAS

5862
RHQLMEWDS

5863
RHQLVEWDS

5864
RHRLLEWDS

5865
RKNVWNNKW

5866
RMDWVAECR

5867
RNNEFIQPF

5868
RRQLLEWDS

5869
RSSTFEHHK

5870
RSTIFERHK

5871
RVIQPCDER

5872
RVTRDERWG

5873
RWGFGSDEA

5874
RWKQWCWEF

5875
SCFHVAEMF

5876
SCILRLISS

5877
SCYSHICRI

5878
SFDPSWWMY

5879
SFFLRLISS

5880
SFHPIWWMY

5881
SFHPSWWMS

5882
SFHPSWWVY

5883
SFHPTWWMY

5884
SFHRSWWMY

5885
SFHSSWWMY

5886
SFHTSWWMY

5887
SFILRLICS

5888
SFILRLISF

5889
SFILRPISS

5890
SFNPSWWMY

5891
SFNSHICRI

5892
SFSSHICRI

5893
SFYNHICRI

5894
SFYPSWWMY

5895
SFYSHICHI

5896
SFYSHICPI

5897
SFYSHICRM

5898
SFYSLICRI

5899
SFYSNICRI

5900
SFYSPICRI

5901
SFYSYICRI

5902
SGFPMWDYW

5903
SGIPMWDHW

5904
SGIPMWDYR

5905
SGIPMWEYW

5906
SGIPMWNYW

5907
SGIPVWDYW

5908
SGIQMWDYW

5909
SGIRMWDYW

5910
SGNPMWDYW

5911
SILIMAYSW

5912
SIYSHICRI

5913
SMNSLWHMR

5914
SQSLCNELC

5915
SRAQIQDEW

5916
SRTIWGADM

5917
SSILRLISS

5918
SSIPMWDYW

5919
SVPMDHCWA

5920
SWDRQWDDF

5921
SWKIDFWDH

5922
SYAVLQRTM

5923
SYILRLISS

5924
SYYSHICRI

5925
TADAVPCHH

5926
TADVVLCHH

5927
TADVVPCDH

5928
TAEAQDCNP

5929
TAEVVPCHH

5930
TANVVPCHH

5931
TCEAQDCNP

5932
TCEGREVDS

5933
TCEGRKLDS

5934
TCFDSEDMG

5935
TCFHAAEMF

5936
TCFHFAEMF

5937
TCFHGAEMF

5938
TCFHIAEMF

5939
TCFHVAAMF

5940
TCFHVDEMF

5941
TCFHVTEMF

5942
TCFNVAEMF

5943
TCFPVAEMF

5944
TCFRVAEMF

5945
TCGRDEFAF

5946
TCGRDLFAF

5947
TCGRDQCAF

5948
TCGRDQFAY

5949
TCGRDQFSF

5950
TCIHVAEMF

5951
TFFHVAEMF

5952
TFHPSWWMY

5953
TGDVVPCHH

5954
TGEAEDCNP

5955
TGEAQDCNA

5956
TGEAQDCSP

5957
TGFHVAEMF

5958
TGGCWPREC

5959
TGIPMWDYW

5960
TGKAQDCNP

5961
TGMSELADE

5962
TGPMDHCWA

5963
THFPWSGEA

5964
THQLLEWDS

5965
TMEPLVCDD

5966
TMSVVQCER

5967
TNDEFIQPF

5968
TNDHFVCYL

5969
TNHCIFKTM

5970
TNHEFIQPF

5971
TNIEFIQPF

5972
TNMSELADE

5973
TNNAVIQPF

5974
TNNECIQPF

5975
TNNEFFQPF

5976
TNNEFIEPF

5977
TNNEFIQLF

5978
TNNEFIQQF

5979
TNNEFIQRF

5980
TNNEFIQSF

5981
TNNEFIRPF

5982
TNNEFLQPF

5983
TNNEFVQPF

5984
TNNEIIQPF

5985
TNNEVIQPF

5986
TNNQFIQPF

5987
TNQCLFKIM

5988
TNSEFIQPF

5989
TNTEFIQPF

5990
TNYEFIQPF

5991
TRAQIQDEW

5992
TSDVVPCHH

5993
TSMNELADE

5994
TSNEFIQPF

5995
TVPMDHCWP

5996
TVPMDHCWS

5997
TWFHVAEMF

5998
TWLPWINER

5999
TYAVLKRTM

6000
VATDSVNES

6001
VGEERVGEP

6002
VGGERVGEA

6003
VGVMLNAMQ

6004
VKKIKRCEG

6005
VMEPLVCDD

6006
VMYPWISCL

6007
VNDDDQCCM

6008
VNDHDQCCM

6009
VNDNDQCCM

6010
VNDYAQCCM

6011
VNDYDKCCM

6012
VNDYDQCCR

6013
VNDYDQCCV

6014
VNDYDQCFM

6015
VNDYDQCWM

6016
VNDYDQCYM

6017
VNDYEQCCM

6018
VNDYVQCCM

6019
VNGHKFNEE

6020
VNGYDQCCM

6021
VNHYDQCCM

6022
VNNYDQCCM

6023
VQKIINSTV

6024
VQLLSLCPS

6025
VSDLMHNVH

6026
VSGQIDEMS

6027
VVIQPCDER

6028
VWAAMEYEE

6029
VWDCQWDDF

6030
VWEMPPIDM

6031
VWESSSIIS

6032
VWKKPPIDM

6033
VWKMLPIDM

6034
VWKMPPIDT

6035
VWKMPPIDV

6036
VWKMPPLDM

6037
VWKMPPMDM

6038
VWKMPPVDM

6039
VWKMPQIDM

6040
VWKMPSIDM

6041
VWKMSPIDM

6042
VWKQWCWEF

6043
VWKRPPIDM

6044
VWKTPPIDM

6045
WVKVPPIDM

6046
VWQMPPIDM

6047
VWTMPPIDM

6048
WAKCQFGAT

6049
WCRMDNKKS

6050
WCRWPFYSE

6051
WHCDAMECF

6052
WILIMASSW

6053
WILIMAYAW

6054
WILIMAYSL

6055
WILIMAYTW

6056
WILIMEYSW

6057
WILIMSYSW

6058
WILIMTYSW

6059
WILIMVYSW

6060
WILSMAYSW

6061
WIMIMAYSW

6062
WIPIMAYSW

6063
WIQIMAYSW

6064
WIRIMAYSW

6065
WIVIMAYSW

6066
WLIHLWTQF

6067
WLIPLWAQF

6068
WLIPLWPQF

6069
WLIPLWTEF

6070
WLIPMWTQF

6071
WLIPPWTQF

6072
WLIPVWTQF

6073
WLIRLWTQF

6074
WLLIMAYSW

6075
WLLPLWTQF

6076
WPSIWKNEQ

6077
WPSIWNNER

6078
WQMRMQNKF

6079
WSKCQFGAA

6080
WSKCQFGAM

6081
WSKCRFGAT

6082
WSKFQFGAT

6083
WSKWQFGAT

6084
WSLIMAYSW

6085
WSQCQFGAT

6086
WSRCQFGAT

6087
WTIACIQRG

6088
WTNDCIQRG

6089
WVDSMLHKW

6090
WVLIMAYSW

6091
YASYKAMED

6092
YCFDSEDMG

6093
YDIMFTHQM

6094
YDQMMGLER

6095
YFAADERWG

6096
YFQQKHNLF

6097
YFTADEQWG

6098
YGGCWPREC

6099
YISDSVSHG

6100
YKVRKFWDP

6101
YPNGMTFEP

6102
YQEHFHNLI

6103
YQENFHNWI

6104
YQNDMTFEP

6105
YRSHEKFQF

6106
YRSPEKFEF

6107
YRSPEQFQF

6108
YRTPEKFQF

6109
YSDEMDWSK

6110
YSNEMDWNK

6111
YSSSKAMED

6112
YVGDSVSHG

6113
YVSDCVSHG

6114
YVSDGVSHG

6115
YVSDSVNHG

6116
YVSDSVSHS

6117
YVSDSVSRG

Example 33
AAV5 Variants with Colon Tissue Tropism as Identified by Machine Learning

This example describes preferred properties of engineered AAV5 variants that display colon tissue tropism as identified by machine learning methods. The same bioinformatics pre-process, machine learning methods, and analyses as described in EXAMPLE 19 and FIGS. 19-FIG. 22 were carried out to identify preferred properties of engineered AAV5 variants that display colon tissue tropism.

Favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an AAV5 VP capsid polypeptide, which are associated with a higher probability of colon tissue tropism are described below. Any of the below machine learning-derived positional amino acid preferences described in TABLE 50 can be present alone or in combination with each other or in combination with any of the amino acid preferences observed and described in EXAMPLE 14, TABLE 27. Listed below in TABLE 50 are a summary of positional features shared between the top important features for colon tropism extracted from the ML models.

TABLE 50

Machine Learning-Derived Colon Tissue Tropism Rules

High mol mass at Xaa1

Xaa1 is selected from Y or W

High solubility at Xaa1

Xaa1 is selected from W, F, I, or L

Low solubility at Xaa2

Xaa2 is selected from D

Low mutability at Xaa2

Xaa2 is selected from P or K

Medium mol mass at Xaa2

Xaa2 is selected from D, E, N, K, M, Q, I, or L

Low hydropathy at Xaa2

Xaa2 is selected from D, E, R, K, H, N, or Q

Low mutability at Xaa3

Xaa3 is selected from K, V, P, or C

High solubility at Xaa3

Xaa3 is selected from W, F, I, or L

High average flexibility at Xaa5

Xaa5 is selected from S, P, G, R, E, or D

High surface accessibility at Xaa5

Xaa5 is selected from D or N

Low hydropathy at Xaa6

Xaa6 is selected from R

Low mutability at Xaa6

Xaa6 is selected from Y, R, F, or L

Low solubility at Xaa6

Xaa6 is selected from R or Q

High surface accessibility at Xaa6

Xaa6 is selected from E, R, or K

High average flexibility at Xaa6

Xaa6 is selected from G or R

Low solubility at Xaa8

Xaa8 is selected from D

TABLE 51 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that exhibited colon tissue tropism and comport to one or more of the rules provided in TABLE 50. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 11118-SEQ ID NO: 12117, as disclosed in TABLE 51. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 51

Sequences of the 581 to 589 Region in AAV5

VP1 Capsid Polypeptide that Drive Colon

Tissue Tropism

SEQ

ID
581-589

NO
Sequence

11118
ACCKVRRDC

11119
ACPVEFVPW

11120
ACYIDAAPN

11121
ACYLRNEDR

11122
ADASRTELD

11123
ADASRTEPD

11124
ADASRTKRD

11125
ADERPNATD

11126
ADIMFEIIF

11127
ADIMKESIV

11128
ADIMKVSIF

11129
ADIMMESIF

11130
ADIMQESIF

11131
ADIMRESIF

11132
AD1NRWSYV

11133
ADITKESIF

11134
ADLHNNHEV

11135
ADLNNNHED

11136
ADRDGPHWY

11137
ADRGGPHWD

11138
AECCLEPKH

11139
AEDGADMII

11140
AEGCKFYNG

11141
AELMMGSII

11142
AEQQHRLIE

11143
AFCRGHYKA

11144
AFCRGHYPA

11145
AFCRGHYRA

11146
AFCRGLYQA

11147
AFCRGRYQA

11148
AFDTFYAFS

11149
AFHHDRKYY

11150
AFKEFSLCC

11151
AFLYAAGWC

11152
AFNLDRKYY

11153
AFNQDRKYY

11154
AFPQNEHQR

11155
AFVTFYAFR

11156
AGPVEFFPW

11157
AGRQEKPDG

11158
AGRQEKPYV

11159
AGWQKCYDS

11160
AGWQMCDDS

11161
AGWQTCYDS

11162
AHDLATEWE

11163
AIDYLIDIL

11164
AINHEKTDW

11165
AKDPKPIRK

11166
ALNHDRKYY

11167
ALPDTNHAD

11168
ALSNDNAIL

11169
ALSNHNAVL

11170
AMHMTRGKC

11171
AMIWPQVLD

11172
AMQKTRGKC

11173
ANCDMKDPW

11174
ANCDMTDRW

11175
ANCNKRHNW

11176
ANNHEKTDL

11177
ANNHEKTEW

11178
ANNHVKTDW

11179
APPPHEVKI

11180
APPPHEVKS

11181
APPPREVKF

11182
APPQMCDFI

11183
AQFKDRSSF

11184
ARPKNLPLS

11185
AVTNHCIDR

11186
AVWQMCYDS

11187
AWDGSIEFL

11188
AYDAPRWFM

11189
AYFQARLLL

11190
AYIMKESIF

11191
AYNHDRKYY

11192
CDKMCAPQA

11193
CDPHGMVGC

11194
CDRKGAGNC

11195
CDRMYAPQA

11196
CERKGAGHC

11197
CERKGAGNG

11198
CERKGSGNC

11199
CERRGAGNC

11200
CFCKCGMPS

11201
CFDCSRAHP

11202
CFDRGKAFW

11203
CGFPGFSPV

11204
CGRWNFQYL

11205
CGSSGHLIY

11206
CHPLTAPSV

11207
CIHTCESII

11208
CINPCESII

11209
CLLDGKSDL

11210
CMCPPRLEV

11211
CMKFAFWEC

11212
CMKIAFWGC

11213
CMKMAFWEC

11214
CMPTARICM

11215
CQIWAKNEY

11216
CQIWGKNEF

11217
CQLWAKNEF

11218
CQPHWEKGD

11219
CRKMMORIS

11220
CSGSGHLIY

11221
CWDERNVFC

11222
CWFMGEGCH

11223
CWLIWKLDR

11224
CWVLWKLDR

11225
DCKCDMFNC

11226
DCPLSNSYE

11227
DDKIAVGWP

11228
DDKIAVRWH

11229
DDVDCCDRK

11230
DFIMKYHAM

11231
DFNHDRKYY

11232
DFPQNEHQR

11233
DEQEFSLCC

11234
DGFMDFKHH

11235
DGMRDFKPG

11236
DGVVPVTNL

11237
DGYMDFKHL

11238
DGYMDFKRH

11239
DHALATEWE

11240
DHDLATGWE

11241
DHDLTIEWE

11242
DHDLVTEWE

11243
DHVLATEWE

11244
DKDCDLTSR

11245
DKDGDRFYI

11246
DKDHSRIDG

11247
DKHDSRIDG

11248
DKHHGRIDG

11249
DKHHSRFDG

11250
DKHHSRIDC

11251
DKHHSRIDD

11252
DKHHSRIDR

11253
DKHHSRIDV

11254
DKHHSRIHG

11255
DKHHSRITG

11256
DKHHSRIYG

11257
DKHHSRMDG

11258
DKHHSRNDG

11259
DKHHSRVDG

11260
DKHLSRIDG

11261
DKHNSRIDG

11262
DKHPSRIDG

11263
DKHRSRIDG

11264
DKLHSRIDG

11265
DKNHSRIDG

11266
DKPHSRIDG

11267
DKYHSRIDG

11268
DMHHSRIDG

11269
DNGFKLQKS

11270
DNQCSDKGL

11271
DPPNDMGAV

11272
DQHHSRIDG

11273
DRDCNLTSR

11274
DRHHSRIDG

11275
DSRCPAAYF

11276
DTFDRIFCS

11277
DWDHMFEME

11278
DWSHIFGRG

11279
DYNWHACSI

11280
EAIHRQACT

11281
EDIMKESIF

11282
EDNKWHGWH

11283
EEAPGFFCF

11284
EEAPGFICY

11285
EEATGFICF

11286
EEDPGFICF

11287
EEGPGFICF

11288
EEGPGFIFF

11289
EETPGFICF

11290
EFKEFSLCC

11291
EFPIDRRFP

11292
EGDEHEFAG

11293
EGDEPEFAG

11294
EGDEQEFVG

11295
EGFPFEGIS

11296
EGFRFEGII

11297
EGHNYEAML

11298
EGIYFVKHD

11299
EGQNYEALL

11300
EGQNYEAVL

11301
EGQNYESML

11302
EGQNYETML

11303
EGRNYEAML

11304
EIAYDYFPM

11305
EIILKDMCH

11306
EIILKDMWN

11307
EIIVKDMWH

11308
EKAPGFICF

11309
EKCGPCARY

11310
EKCMCHDIS

11311
EKHDGLMRR

11312
EKPDGLMRQ

11313
EKPEPRYKE

11314
EKPQPWYKE

11315
EKPSITCRE

11316
EMCMDVPSV

11317
EQCLAAAGE

11318
EQCLAAAVE

11319
EQCLAAGAE

11320
EQCLAAPAE

11321
EQCLAASAE

11322
EQCVAAAAE

11323
EQDIYGAIV

11324
EQPVGQSHR

11325
EQTEGQSHR

11326
ESINRQACT

11327
EVAPGFICF

11328
EVEYDYFPM

11329
EVIYFVQHD

11330
EVLYFVKHD

11331
EVWCQVEPN

11332
EWACAHAMR

11333
EWDQPNYIE

11334
EWVCAHAMR

11335
EYFQARLWL

11336
EYRQPQSIA

11337
FACPVLWGQ

11338
FANPVLWGQ

11339
FASP1LWGQ

11340
FASPVLWGR

11341
FCWRDNEPE

11342
FDGPSVRGE

11343
FDKMYAPQA

11344
FDMCIEYSN

11345
FDMWGVGIP

11346
FDNHANTWA

11347
FDSPVLWGQ

11348
FEQTRFPMQ

11349
FEREFHYYD

11350
FEREFKYYD

11351
FERPPKLGI

11352
FEWEFQYYD

11353
FHDNARLFL

11354
FHLREKDDC

11355
FHYDCQDKV

11356
FHYDCRDNV

11357
FIAHKGQQW

11358
FIEFHVILV

11359
FIGKGYAIF

11360
FIGMGYAIC

11361
FIVHKGRQW

11362
FIVLKGQQW

11363
FIVMGYAIF

11364
FIVNKGQQW

11365
FLERYQHMD

11366
FLFMEPRNG

11367
FLGCYCHYA

11368
FLGCYCLYA

11369
FLGCYCPDA

11370
FLGCYCPSA

11371
FLGCYCPYG

11372
FLGCYCPYS

11373
FLGCYCRYA

11374
FLGCYFPYA

11375
FLGCYGPYA

11376
FLGCYRPYA

11377
FLGFYCPYA

11378
FLGMGYAIF

11379
FLGNTWLGP

11380
FLGWYCPYA

11381
FLGYYCPYA

11382
FLRNAWLGP

11383
FLRNTWLDP

11384
FLRNTWLGA

11385
FLRNTWLGS

11386
FLRNTWLSP

11387
FLRNTWLVP

11388
FMCNIGCFT

11389
FMGCYCPYA

11390
FNVCTMWAQ

11391
FPDCKNKAF

11392
FPLRENDDC

11393
FPTCWRYAK

11394
FPTIGLMAC

11395
FPYCKNKPF

11396
FQRNTWLGP

11397
FRIRCRVNK

11398
FRIRRRVNK

11399
FRRNTWLGP

11400
FSEQHAVND

11401
FSEQHAVNT

11402
FSEQHVVNN

11403
FSERHAVNN

11404
FSGWFRVCD

11405
FSPVERHNG

11406
FTSPVLWGQ

11407
FVCMEPRNG

11408
FVFMDPRNG

11409
FVFVEPRNG

11410
FVKGEFHFH

11411
FVQPFLHDG

11412
FWCRPRLHY

11413
FWPGHNPHN

11414
FWSRMIPLV

11415
FWTGHNPHI

11416
FWTGHNPPN

11417
FWTGRNPHN

11418
FYPGERHNG

11419
GCDGYALAI

11420
GCDPYNMVD

11421
GDDDCCDRK

11422
GDHHGMVGC

11423
GDKIKESDR

11424
GDKMGICHC

11425
GDKMGIHHC

11426
GDKMYAPQA

11427
GDPHGMGGC

11428
GDPHGMLGC

11429
GDPPGMVGC

11430
GDPQGMVGC

11431
GDVDCCDRT

11432
GDYHYRVNW

11433
GEAPGFICF

11434
GENKVNDPA

11435
GEYHCRVNW

11436
GEYKLNDPA

11437
GEYKVNDPP

11438
GEYRVNDPA

11439
GFCRGHYQA

11440
GFMCNLCGP

11441
GFQTKFHLC

11442
GGDGYAIAI

11443
GGRHGYLHD

11444
GGWQMCYDS

11445
GGYKVNDPA

11446
GIIHHCVSE

11447
GIIRHCFSE

11448
GIIRHCLSE

11449
GIIRHCVIE

11450
GIIRNCVSE

11451
GKHHSRIDG

11452
GLRDGNSIF

11453
GMIPYNWSW

11454
GMPATELWL

11455
GMRRDGLDV

11456
GMRWDRDQL

11457
GMSSHTPIY

11458
GNCNKRHNW

11459
GPKKGGQAM

11460
GPKQGGOSM

11461
GPVLPDVGG

11462
GPVMPDVGA

11463
GRSSHPP1Y

11464
GSKSGGADD

11465
GSKYGGDDD

11466
GSNSGGDDD

11467
GVRRVKIEW

11468
GWIEDKREV

11469
GWSEEKREV

11470
HDDWKKQPN

11471
HGAERFDRF

11472
HGAERVDRV

11473
HGLVDFDPH

11474
HKHHSRIDG

11475
HNHTRKDPV

11476
HVANEEVGG

11477
HVGNEEVSG

11478
HWRVQLPWN

11479
ICHANNVRV

11480
ICKARIEKY

11481
ICKPRLEKY

11482
ICPANNVRG

11483
ICPANNVRL

11484
ICPVNNVRV

11485
ICRPRIEKY

11486
ICSANNVRV

11487
ICSIDNNNY

11488
ICSMDNKNY

11489
IDHDNWDMR

11490
IEKDEMPVC

11491
IERPPKLGT

11492
IFLNIFELR

11493
IFLNIFEML

11494
IGANGWQCC

11495
IGLVDFDPH

11496
IHAPKLLWL

11497
IHCPDSWSQ

11498
IHCPEAWSQ

11499
IHCPESWSL

11500
IHGPCTPWD

11501
IHHEGRSVC

11502
IHPPKLLWL

11503
IHSPKLLWL

11504
IHTNMRVIS

11505
IHTNMRVIV

11506
IHTPRLLWL

11507
IIKFEWQEV

11508
IIKFEWQKF

11509
IIKFEWQQV

11510
IIKFEWQRV

11511
IKDQNKEWN

11512
ILRNTWLGP

11513
IMNEAHYRF

11514
IMNKDHYRF

11515
1MNRAHYRF

11516
1NGSNTLTN

11517
IPECWWRWH

11518
IPEFWLRWH

11519
IPEIWWRWH

11520
IPKDEMPVC

11521
IPPLKTEDN

11522
IPRNVYMCD

11523
IPRTVYMCD

11524
IQKDEMPFC

11525
IQKDEMPVW

11526
IQKYEMPVC

11527
IQRDEMPVC

11528
IRPNDSSFH

11529
IRPNEGSFH

11530
IRPNESSFP

11531
ISGKEMNST

11532
ISNEMSKIL

11533
IVADNTVCA

11534
IVDDHTVCA

11535
IVDTNTATK

11536
IVPRFLVEA

11537
IWPANNVRV

11538
KDDWAGYQM

11539
KDIREIWDI

11540
KDERGEWDI

11541
KDIRVIWDI

11542
KDMRDIWDI

11543
KDSWAGYQM

11544
KDWMPSYAL

11545
KDYWAEYQM

11546
KDYWAGCQM

11547
KDYWAGYKM

11548
KDYWAGYRM

11549
KDYWAVYQM

11550
KEGCHGRMG

11551
KEGCHVRVG

11552
KELSCDQNW

11553
KFLLAFEVT

11554
KFLYAAGWC

11555
KGIVIDRIL

11556
KHENKKDVL

11557
KIMQFCWDF

11558
KKFLCWEWS

11559
KLCNPVVFV

11560
KMCAYQPDT

11561
KMCHDERQV

11562
KMCPDERKV

11563
KMCPDERQL

11564
KMCTDERQV

11565
KMFPDERQV

11556
KMGPDERQV

11567
KMLIDEPTL

11568
KMLIDGATL

11569
KMLMDEATL

11570
KMLNDEATL

11571
KMRIDEATL

11572
KMWPDERQV

11573
KNFEELDND

11574
KNFEKLDNE

11575
KNFEKLDYD

11576
KNFEKLNND

11577
KNKC1ENRW

11578
KPCPVRPPH

11579
KPDDEFDCD

11580
KPVYLYCSD

11581
KQCLAAAAE

11582
KSEWQLMYG

11583
KTADEFDCD

11584
KWERYRDFI

11585
KYKREEYEK

11586
LCKLPEIRK

11587
LDAKMVQPS

11588
LDCDSVWSP

11589
LDLKMVQPS

11590
LEGEGMWAF

11591
LERKPRNDC

11592
LERKPRNYF

11593
LERKPRSYC

11594
LERQPRNYC

11595
LESEGMWSF

11596
LHHEGRSVC

11597
LKHPPQPKV

11598
LKQINRHPS

11599
LMDAIDSIW

11600
LMVAIDSIW

11501
LNYCDQYGD

11602
LNYCDQYVG

11603
LNYERRDNY

11604
LPAYTTPIV

11605
LPFGNKFPD

11606
LPGPVTVSW

11607
LPKPHFDKQ

11608
LPKPYFDKK

11609
LPLLKTEDN

11610
LPLTDAPMV

11611
LPPLETEDN

11612
LPPLKTEAN

11613
LPPLKTEDD

11614
LPPLKTEDT

11615
LPPLKTKDN

11616
LPPLKTVDN

11617
LPPLRTEDN

11618
LQRLGYSPD

11619
LQTYTTPIV

11620
LR1DPKDSF

11621
LSICHPVMP

11622
LSICKPVMP

11623
LWDHGASYK

11624
LWHHGASYM

11625
LYDQSRSQL

11626
LYPCKMPMN

11627
LYYPSRSQL

11628
LYYQSRSKL

11629
MCKPRIEKY

11630
MCLEDGKDW

11631
MCSIDNKNY

11632
MDKHGCCCL

11633
MDKLGCCCW

11634
MDMQGGKVC

11635
MDQHGCCON

11635
MDWCVQQSS

11637
MDWCVQQST

11638
MEADHKSDP

11639
MKGCFVHHS

11640
MKGYHHFIS

11641
MKGYFVHHY

11642
MMPCMVNFG

11643
MNKCIENRG

11644
MNKWIENRW

11645
MQFVDRLAM

11646
MQFVDRWAL

11647
MRCCNVPWS

11648
MRDRMKDEE

11649
MVIHPAISR

11650
MWDHRNETM

11651
MWDNRNESM

11652
NAMKRSPDV

11653
NAMQRSPDF

11654
NCENATDGD

11655
NCLVDFDPH

11656
NCMNATDWD

11657
NCNNELTDR

11658
NDMAQSYRE

11659
NDMEQSDRE

11660
NDMEQSYHE

11661
NELAYQDWS

11662
NELEYQHWS

11663
NFCPENAWQ

11664
NFEDQESHV

11665
NFGDLESHV

11666
NFITQEMYS

11667
NFNTQEMYS

11668
NFTTKEMYS

11669
NFTTMNMTC

11670
NFTTQEMYY

11671
NFWMDMHRI

11672
NGGTWANVE

11673
NGHNELTDR

11674
NGLDDFDPH

11675
NGLVDCDPH

11676
NGLVDFAPH

11677
NGLVDFDPL

11678
NGLVDFDRH

11679
NGLVDFDTH

11680
NGLVDFVPH

11681
NGLVHFDPH

11682
NGNNELADR

11683
NGNNELTDK

11684
NGNNGLTDR

11685
NGRVDFDPH

11686
NGWEGSEDD

11687
NGWSDVDRQ

11688
NHNVPACII

11689
NIEQMLAIG

11690
NKHHSRIDG

11691
NMHSEQTPL

11692
NMKFSVGMN

11693
NMKISVDMN

11694
NMKTESAHK

11695
NMKTKSADK

11696
NMKTKSAHE

11697
NMKTKSAHT

11698
NMKTKSAPK

11699
NMKTKSARK

11700
NMKTKSDHK

11701
NMKTKTAHK

11702
NMKTRSAHK

11703
NMPWNMEWN

11704
NMPWNMPWN

11705
NMPWNMQLN

11706
NMPWNMQWK

11707
NMPWNMQWS

11708
NMPWNVQWN

11709
NMQTKSAHK

11710
NNMIVAKIA

11711
NPCHGYLKM

11712
NPPCARECR

11713
NPPCARGGR

11714
NPPCARKWR

11715
NPPCARQWR

11716
NPRCGIFTL

11717
NQGLPCCQI

11718
NQGMHCCQI

11719
NQGMPCCRI

11720
NQNNLEYPI

11721
NRHPPAAIS

11722
NRNPPAAII

11723
NRNPPEALS

11724
NRNSFHQEE

11725
NRPWQMRWA

11726
NRTSFHKFE

11727
NSITANCSK

11728
NSITENCAK

11729
NSITENCSN

11730
NTKVKDEDN

11731
NVLGPPSFN

11732
NVNNELTDR

11733
NVTANYCCF

11734
NYDHFIAAS

11735
NYGKDMHKL

11736
NYNNFIAAS

11737
NYRESRSDL

11738
PAIHNHMCT

11739
PAIRNHMCP

11740
PAYKNSGGD

11741
PDFVAEQQP

11742
PETLWANIW

11743
PETLWTDIW

11744
PETPWTNIW

11745
PFCRGHYQA

11746
PFISPKQVS

11747
PF1SPREVS

11748
PHYTDLGP

11749
PFLYTDWGP

11750
PFNNWFAAG

11751
PGWQMCYDS

11752
PINFPYEWL

11753
PKFAFNQCY

11754
PKGMINWPS

11755
PKVRINWPS

11756
PKVVINWPS

11757
PMCMEPFPM

11758
PMNRDTLIS

11759
PPPLNRKWD

11760
PQAYTTPIV

11761
PQLYFTTDK

11762
PQTKKDCPA

11763
PRKMMQRIS

11764
PSDLKLVNN

11765
PSTIKHVDP

11766
PSTIKRIDP

11767
PWDGSIEFL

11768
QCLLWNDCK

11769
QELFHGKIA

11770
QFPRHEPIQ

11771
QFPRQEPLQ

11772
QFYNRSDDL

11773
NVTANYCCF

11774
QINPFYMWF

11775
QINQFCQIV

11776
QMCPDERQV

11777
QPCLVRPPH

11778
QPCTVRPPH

11779
QSMWETDRN

11780
QWCWMAVSI

11781
QWNMFQVIF

11782
RAAKVIPPT

11783
RADKFIPPT

11784
RADKVITPT

11785
RAYKVIPPT

11786
RCDLKEMWK

11787
RCYLKEMRK

11788
RDCHMRITD

11789
RDCWMVFII

11790
RDFVAGQQP

11791
RDFVGEQQP

11792
RDGKWALEN

11793
RDGKWVREN

11794
RDYHMLITD

11795
RGFHHADNV

11796
RGFNHADNG

11797
RHIYRKGWG

11798
RLRDGNSIF

11799
RMCPYEAWK

11800
RMCTYEPWK

11801
RPEFNAFQA

11802
RSFIDIVGT

11803
RTQPIGVDP

11804
RYCWMVVII

11805
SADYNFGFS

11806
SCGWDHGNL

11807
SCPANNVRV

11808
SCPVEFFPW

11809
SDASRTERD

11810
SDKMGIYHC

11811
SDLHNNHED

11812
SDNWGAQCC

11813
SDPHGMVGC

11814
SFGPSQVCC

11815
SFIGNAQWC

11816
SFIGNARWC

11817
SFLAHNNFH

11818
SFLANNHFH

11819
SFLANNTFH

11820
SFNHDRKYY

11821
SFYEWPHCE

11822
SGICCCDNA

11823
SGIGCCDNS

11824
SGWKMCYDS

11825
SGWQMCYDS

11826
SHDQALMFE

11827
SHFYEDARS

11828
SIIRHCVSE

11829
SIQPFLHDG

11830
SLNGYTLVC

11831
SLPEYSNVK

11832
SNPWPMGKV

11833
SPHHDKDPF

11834
SPMINLWPS

11835
SRKMMKRIS

11836
SRKMMPRIS

11837
SRQCPLEIG

11838
SSDLKLVNS

11839
SSGGDHGNL

11840
SSIHHQCCT

11841
SSIQHHCCT

11842
SSVWDHGNL

11843
SVQLFLHDG

11844
SVQPFLHDA

11845
SVQPFLYDG

11846
SVQPVLHDG

11847
SYDNWDFCC

11848
TCSIDNKNY

11849
TCTWIGAGS

11850
TDASRTERD

11851
TDIMKESIF

11852
TDYWAGYQM

11853
TFPDGEMYQ

11854
TGLVDFDPH

11855
TGTWIGTGS

11856
TGWQMCYDS

11857
TIDDHSFDQ

11858
TINCKWCFC

11859
TINPHKSDW

11860
TISCKWCVC

11861
TNICYDQIK

11862
TNICYDQIR

11863
TNNHEKTDW

11864
TPCCLHHPA

11865
TPPPHEVKF

11866
TQMFAMPPS

11867
TQPPLMNIL

11868
TRNPPAAIS

11869
TRPSAASIR

11870
TRPSAASVG

11871
TRPSASSIG

11872
TSICYDQIQ

11873
TSPPDQVPV

11874
TWDGSIEFL

11875
VADHRPMYS

11876
VAIVDGWIE

11877
VAPSESSWE

11878
VCLVDGKDW

11879
VCPANNVRV

11880
VCTIYYMCG

11881
VDCNKRHNW

11882
VDHCTAIGA

11883
VDIFDGWIE

11884
VDPFCAVRI

11885
VDPMGEAGT

11886
VDVDCCDRK

11887
VEFADNPWN

11888
VERGNKIAS

11889
VERSNKISS

11890
VFCRGHYQA

11891
VGTKEGPDP

11892
VHHEGRSFC

11893
VHHEGRSVF

11894
VHHKGRSVC

11895
VHLRENDDC

11896
VHPEGRSVC

11897
VHQEGRSVC

11898
VHREGRSVC

11899
VHYEGRSVC

11900
VISRECYEE

11901
VKCKDKYQS

11902
VKCQDKYQY

11903
VKHHSRIDG

11904
VLGLPPYPE

11905
VLLYMRNNQ

11906
VMAAEQLNQ

11907
VMIWPQILD

11908
VMPTDCILV

11909
VNCDKRHNW

11910
VNCNKRHNL

11911
VNCNKRHNM

11912
VNCNKRRNW

11913
VNCNKRYNW

11914
VNCNNRHNC

11915
VNCNRRHNW

11916
VNCSKRHNW

11917
VNCYKRHNW

11918
VNGNKRHNW

11919
VNLQDLSDR

11920
VNPSDQTNH

11921
VNSSDQNNH

11922
VPDFDPFWS

11923
VPIFDGWIE

11924
VPNFAAFCM

11925
VPPACRHKK

11926
VPPGCRHQK

11927
VPTFDPFWS

11928
VQFVDRWAM

11929
VQTKKDCPA

11930
VRFDMCKWT

11931
VRFEMMSWT

11932
VIDFHTDML

11933
VTDFNTDMQ

11934
VVFINYNDQ

11935
VVFTNYNDK

11936
VVPPGNVIL

11937
VVPPGNVLP

11938
VVYRDTCNG

11939
VWDHRNESM

11940
VYNWHACAI

11941
WAEDAYRRF

11942
WCACVFEYW

11943
WCERVFEYW

11944
WCEWVFEYW

11945
WCFKKELCN

11946
WCKLPEIRR

11947
WCQCVFEYW

11948
WCQLPEIRK

11949
WCYLMNLES

11950
WDHNCQATI

11951
WDHNYQDTI

11952
WDNNCQDTI

11953
WDQNCQDTI

11954
WERMGQIEC

11955
WERMGQIED

11956
WERMGQIEH

11957
WERMGQIES

11958
WERMGQIQY

11959
WERMGQMEY

11960
WEWDRPFMK

11961
WFFHYKPWM

11962
WFMKCNWWT

11963
WGCCEEMDS

11964
WINTGPLRH

11965
WKIKPDHRV

11966
WKLKNCKSD

11967
WLFEGKLDT

11968
WLFYGCEPN

11969
WLPHGPRME

11970
WLPNRRIDD

11971
WLQDGNSIF

11972
WLRDGDSIF

11973
WLRDGNSIS

11974
WLRDGNSIY

11975
WLREGNSIF

11976
WLRGGNSIF

11977
WLRVGNSIF

11978
WMGWDRDQL

11979
WMPNRRIDA

11980
WMPNRRIDG

11981
WMRCDRDQL

11982
WMRLDRDQL

11983
WMRWARDQL

11984
WMRWDQDQL

11985
WMRWDRAQL

11986
WMRWDREQL

11987
WMSDMLKAV

11988
WMWWDRDQL

11989
WNIPNRSQA

11990
WNNEHLKVW

11991
WNNFYCGCN

11992
WPFPVQFCD

11993
WPHHAKDPF

11994
WPHHDKDPV

11995
WPHHNKDPF

11996
WPRDGNSIF

11997
WPSEGLIGA

11998
WPVPVQVCD

11999
WQIWAKNEF

12000
WRFDGCEPN

12001
WRFYGCEPT

12002
WRFYGCERN

12003
WRFYGCESN

12004
WRGCMFMSK

12005
WRNCMFMSK

12006
WRSYGCEPN

12007
WSSEGLIGA

12008
WVWNGCRDM

12009
WYAQCWKAF

12010
WYCWMVFII

12011
WYDAGHKCK

12012
WYDAGNNCK

12013
WYDAVNKCK

12014
WYDDGNKCK

12015
WYECVFEYW

12016
WYYAGNKCK

12017
YAHYCWDPI

12018
YAVAVGISS

12019
YDFGGRVQT

12020
YDHHKIIWA

12021
YDHHKIIWV

12022
YDINYYPEE

12023
YDINYYPKD

12024
YDINYYPNE

12025
YDIPKSTCS

12026
YDNSKYCKN

12027
YDTNYYPKE

12028
YDYGGRVQK

12029
YEAKSLQPN

12030
YEENVLFMG

12031
YEINYYPKE

12032
YETGMWLNR

12033
YFDCSRAHS

12034
YFDCSRANP

12035
YFDCSRGHP

12036
YFDCSRTHP

12037
YFDRSRAHP

12038
YFDWSRAHP

12039
YGCNRSYQD

12040
YHANNMEPW

12041
YHDLATEWE

12042
YHDNALLFL

12043
YHENVLFMG

12044
YHINNMEPW

12045
YHPHHMYWR

12046
YHPRTALMN

12047
YHQHNACMR

12048
YHT1DMEPW

12049
YHT1NMEPW

12050
YHTNNMERW

12051
YHTNNMERW

12052
YHTNNMETW

12053
YHTNNMVPW

12054
YHTNNREPW

12055
YHTNNVEPW

12056
YHTSNMEPW

12057
YHTTNMEPW

12058
YHTYNMEPW

12059
YIVHKGQQW

12060
YIWLVMFFS

12061
YKAGDRFYI

12062
YKDCDRFYI

12063
YKDDDRFYI

12064
YKDGDRIYI

12065
YKDSDRFYI

12066
YKHHSRIDG

12067
YKYGDRFYI

12068
YLPSAELIK

12069
YMWICPPDQ

12070
YMWIHPPDQ

12071
YMWISPPDQ

12072
YMWIYAPDQ

12073
YMWEYPPDK

12074
YMWEYPPDL

12075
YMWIYPPDP

12076
YMWIYPPGQ

12077
YMWIYPTDQ

12078
YMWIYQPDQ

12079
YMWNYPPDQ

12080
YMWSYPPDQ

12081
YMWTYPPDQ

12082
YNMAHGDEP

12083
YNMVDGDEP

12084
YNMVHGDEA

12085
YNMVHGDES

12086
YNMVHGDET

12087
YMNVHGDGP

12088
YNMVHSDEP

12089
YNPRIALMN

12090
YNPRSALMN

12091
YNTVHGDEP

12092
YNVCILWAQ

12093
YNVCTMLAQ

12094
YNVVHGDEP

12095
YPEDRLNMR

12096
YPLGSAGSD

12097
YPQHNACMQ

12098
YPTNNMEPW

12099
YQAAARQNA

12100
YQADPRQNA

12101
YQATPRONA

12102
YQAYGRGKV

12103
YQDAPRQNA

12104
YQIFQYPLH

12105
YQLFQYPWH

12106
YQLLHTIEN

12107
YQLLHTIFQ

12108
YRTLQCKWE

12109
YRWIYPPDQ

12110
YRYLWPIVR

12111
YTGLNELQN

12112
YTICDVPDP

12113
YTPNFRIGD

12114
YVIPFEMDD

12115
YVIPVEMDN

12116
YYDCSRAHP

12117
YYPCVQYRQ

Example 34
AAV5 Variants with Heart Tissue Tropism as Identified by Machine Learning

This example describes preferred properties of engineered AAV5 variants that display heart tissue tropism as identified by machine learning methods. The same bioinformatics pre-process, machine learning methods, and analyses as described in EXAMPLE 19 and FIGS. 19-FIG. 22 were carried out to identify preferred properties of engineered AAV5 variants that display heart tissue tropism.

Favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an AAV5 VP capsid polypeptide, which are associated with a higher probability of heart tissue tropism are described below. Any of the below machine learning-derived positional amino acid preferences described in TABLE 52 can be present alone or in combination with each other or in combination with any of the amino acid preferences observed and described in EXAMPLE 13, TABLE 25. Listed below in TABLE 52 are a summary of positional features shared between the top important features for heart tropism extracted from the ML models.

TABLE 52

Machine Learning-Derived Heart Tissue Tropism Rules

Low solubility at Xaa1

Xaa1 is selected from N or E

Low hydropathy at Xaa1

Xaa1 is selected from H, N, Q, P, Y, D, or E

High mutability at Xaa1

Xaa1 is selected from A or E

High hydropathy at Xaa2

Xaa2 is selected from V or I

Medium mutability at Xaa2

Xaa2 is selected from V

Medium volume at Xaa2

Xaa2 is selected from V, E, or Q

High solubility at Xaa2

Xaa2 is selected from V or M

Low solubility at Xaa3

Xaa3 is selected from R or Q

Low surface accessibility at Xaa4

Xaa4 is selected from C

High solubility at Xaa4

Xaa4 is selected from C

Low charge at Xaa4

Xaa4 is selected from D, E, Y, W, V, P,

M, A, G, F, I, L, N, Q, S, T, or C

High hydropathy at Xaa4

Xaa4 is selected from C

High surface accessibility at Xaa5

Xaa5 is selected from D, E, R, K, N, or Q

Low solubility at Xaa5

Xaa5 is selected from D

Low mutability at Xaa6

Xaa6 is selected from C

Low solubility at Xaa6

Xaa6 is selected from D

High surface accessibility at Xaa8

Xaa8 is selected from D or N

High average flexibility at Xaa8

Xaa8 is selected from D, R, P, G, or S

Medium mol mass at Xaa9

Xaa9 is selected from N, D, L, or I

TABLE 53 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that exhibited heart tissue tropism and comport to one or more of the rules provided in TABLE 52. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 14118-SEQ ID NO: 15117 as disclosed in TABLE 53. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 53

Sequences of the 581 to 589 Region in AAV5

VP1 Capsid Polypeptide that Drive Heart

Tissue Tropism

SEQ

ID
581-589

NO
Sequence

14118
AADFFAIND

14119
AADIFAISD

14120
AADIFGIND

14121
AADIFVIND

14122
AADIYAIND

14123
AADMFAIND

14124
AAYIFAIND

14125
ACDCHHWDQ

14126
ACIEWNWGE

14127
AGDCCLAER

14128
AHECCLPNR

14129
AIQPRDMYS

14130
AKECTGLKN

14131
AKGMEKLRT

14132
AKNGDVLEM

14133
ALPCDNWRG

14134
APRCESCNM

14135
AQEDCITGL

14136
ASDYCYTER

14137
ATMNLSWAL

14138
AVCDDHKPN

14139
AVDWHCEVL

14140
AVESKDIPQ

14141
AVGDEQPID

14142
CCECGHWPN

14143
CDDCYLINR

14144
CDICKHEEY

14145
CDNCYAEPH

14146
CDWMASVSV

14147
CDWMPNVSV

14148
CDWMSNVSV

14149
CEYGWILKR

14150
CGDWDFGEW

14151
CKCCRMEKC

14152
CKFRDFAES

14153
CKMLCRPPV

14154
CKPGDFVRK

14155
CLECCTDSA

14156
CPDCGPTGP

14157
CPECSVDPW

14158
CQMLCRSPM

14159
CRDCCLRGW

14160
CRECSNERV

14161
CRECVHNRC

14162
CRICRATIW

14163
CSDPKNMSI

14164
CWECCNDSG

14165
CWECCPDSG

14166
CWECCTDGG

14167
CWECCTDNG

14168
CWECCTDRV

14169
CWECCTDSV

14170
CWECCTDSW

14171
CWECFTDSG

14172
CWECGTDSG

14173
CWECYTDSG

14174
CWEWCTDSG

14175
CWGCCTDSG

14176
CWVCCTDSG

14177
CYWCGWDNR

14178
DAMDQREQY

14179
DAVDEVKDE

14180
DCNQFNWSE

14181
DDDDFAEKG

14182
DDEILKEDG

14183
DDEVGKPKW

14184
DDVEKDETN

14185
DERERKTKD

14186
DFKALDVPC

14187
DFKVLDVQC

14188
DGCDDHKPN

14189
DGKIEPIRF

14190
DGMRCKEKK

14191
DGVGRPGDN

14192
DHREDNIDA

14193
DIDDLNFAT

14194
DIDFMECAF

14195
DIEEWEMAE

14196
DIEGQNNGM

14197
DIEHIRDPP

14198
DIEMFNENG

14199
DIEMNITAL

14200
DIGDIWMSQ

14201
DIEEYQKIE

14202
DIIPKDAAP

14203
DIKVLDSQP

14204
DILCICETS

14205
DINNNNCGY

14206
DIQPFDYYL

14207
DIQPNDGCS

14208
DIRDEDNIG

14209
DIRDSWTES

14210
DITEEDVTL

14211
DITHHNDGK

14212
DIVDHDHAG

14213
DIVECIQYN

14214
DIYNDSWST

14215
DLCRPNPKN

14216
DLIVATELF

14217
DLIVVTEWF

14218
DLPNSQPPH

14219
DLSEPTEWF

14220
DLTVATEWF

14221
DMDCPEDQQ

14222
DMFQLKPQN

14223
DMVDLDVNP

14224
DNVNMNDAI

14225
DQRCNKAPG

14226
DRCDSIILA

14227
DSDAKNMSI

14228
DSDPKNMSI

14229
DSDSKNMSI

14230
DSDTKNMSI

14231
DSNDQDTPE

14232
DSSEVDMNF

14233
DSYDDSWSI

14234
DSYNDGWST

14235
DSYNDSWSP

14236
DTAPKQEAE

14237
DTIQKKMDH

14238
DTIWADINN

14239
DVADWNGYD

14240
DVAERVEQI

14241
DVAGLDKNA

14242
DVAPMDQVE

14243
DVAPMHEVE

14244
DVAPMHQDE

14245
DVAPMHQIE

14246
DVAPMHRVE

14247
DVAPMNQVE

14248
DVCDDHKAN

14249
DVCDDHKPH

14250
DVCDDHKPI

14251
DVCDDHKPS

14252
DVCDDHKPT

14253
DVCDDHKQN

14254
DVCDDHKRN

14255
DVCDDHKSN

14256
DVCDDHKTN

14257
DVCDDHQPN

14258
DVCDDHTPN

14259
DVCDDPKPN

14260
DVCPFPDGK

14261
DVCPVDVQN

14262
DVDDIHVSF

14263
DVDEISDGV

14264
DVDIHEWPP

14265
DVDMIDTCP

14266
DVDPWDVTE

14267
DVDQMHHND

14268
DVEEKDNKA

14269
DVEEKDNKA

14270
DVEETEAAY

14271
DVEHCNTNA

14272
DVEHQDWGG

14273
DVEIHKWGG

14274
DVETIDNLK

14275
DVEWDNMKQ

14276
DVGDDHKPN

14277
DVGEDEFQP

14278
DVGEWTMNN

14279
DVGILDFNG

14280
DVGPMDCQP

14281
DVHEYDHPE

14282
DVHFFENSR

14283
DVIPRDGAE

14284
DVKDQSIML

14285
DVKEIDELC

14286
DVMPMPEAQ

14287
DVNEDGFNM

14288
DVNENEWTD

14289
DVRDSMKRD

14290
DVSDFIQHF

14291
DVSEGDDST

14292
DVSQIPITF

14293
DVTEYQWST

14294
DVTPNSWIE

14295
DVFPTSWSE

14296
DVTTLNESQ

14297
DVVDFSIVK

14298
DVVEQRECP

14299
DVYDDHKPN

14300
DVYEKMWAN

14301
DYFDMNLTD

14302
DYGPEDFSM

14303
EAAPRDSDD

14304
EAECKMWPT

14305
EAELGNVNM

14306
EAGCRMWPT

14307
EARIQCKID

14308
EARMDIWPI

14309
EAVNRPNMK

14310
ECGCMYWCY

14311
ECGCMYWGC

14312
ECGCMYWVY

14313
EDDWQRMLY

14314
EDRIQCKID

14315
EDRMDKIFT

14316
EDTGEREAW

14317
EDYAACFEF

14318
EEGMQMISQ

14319
EERCYDGSW

14320
EFHEFRASN

14321
EFRIQCKID

14322
EIAIYNFAN

14323
EIAPNMCPP

14324
EIAPREVVN

14325
EICPVDRTQ

14326
EICSFEEGH

14327
EIDDDMLEE

14328
EIDDTDQRW

14329
EIDDVGEPG

14330
EIDEFSASN

14331
EIDQKDISQ

14332
EIDRGDSYK

14333
EIEALMKER

14334
EIEEDDEWY

14335
EIEEFMKMR

14336
EIEQDNEPG

14337
EIEQSFLDS

14338
EIEVTAWAC

14339
EIEYIDAGG

14340
EIGCVDSAK

14341
EIGMQIAQE

14342
EIGTFDNSE

14343
EIHDQQIVF

14344
EIHDVDFGM

14345
EIHEFFHNR

14346
EIHEFGASN

14347
EIHEFPASN

14348
EIHEFRAAN

14349
EIHEFRAPN

14350
EIHEFRASI

14351
EIHEFRAST

14352
EIHEFRAYN

14353
EIHEFRDSN

14354
EIHEFRGSN

14355
EIHEFRSSN

14356
EIHEFSASN

14357
EIHEISTHD

14358
EIHGMDTCP

14359
EIHMFFQSF

14360
EIHPWHRTN

14361
EIIDTSCNC

14362
EIISMNDFM

14363
EILDYCSYN

14364
EILVRCEVD

14365
EIMPDSIMC

14366
EIMPNHNMI

14367
EINDQRPWP

14368
EINEFRASN

14369
EINPKSEDE

14370
EIPEFRASN

14371
EIQDIVKID

14372
EIQPLNAVN

14373
EIRCHHKFA

14374
EIRDMAHHG

14375
EIRDTHEMQ

14376
EIREFRASN

14377
EIREQCIGP

14378
EIRFEGNME

14379
EIRIQCKID

14380
EIRPWDCKW

14381
EIRTIDWQW

14382
EISDIHDLR

14383
EISEYCORS

14384
EISPYNDVE

14385
EISQIPPGW

14386
EITCLCVHS

14387
EITDFQYEN

14388
EITDKDVCT

14389
EITEFWGNY

14390
EITPKDSHT

14391
EITPKDSHT

14392
EIVDFDNYF

14393
EIVEQCGNR

14394
EIVHQDDVN

14395
EIYEFQMWD

14396
EIYEFRASN

14397
EKQMDGINH

14398
EKYFKMFDN

14399
ELRCTKLQN

14400
ELRIQCKID

14401
ELSDSMDSN

14402
EMHECPMAW

14403
EMKCIDVED

14404
EMRTYNKDY

14405
ENYPWEQLE

14406
EPRCENYME

14407
EQCRGEPDE

14408
EQKDNMMMN

14409
ERRCAWFNA

14410
ESDEREEAD

14411
ESDPINEAN

14412
ESGCKMWPT

14413
ESHEIRDHN

14414
ESQEAEWGW

14415
ESRCGDKGC

14416
ESRCVDNTV

14417
ESTDFDELN

14418
ESTELRTGN

14419
ESTNIPMAF

14420
ESVPKHEMI

14421
ETAGKSIHH

14422
ETEDYRTSN

14423
ETEEIQKGH

14424
ETHEFRASN

14425
EMIDKLWAN

14426
ETMNQSWGL

14427
ETNEEEDGN

14428
ETQMFEIGE

14429
EVACFDDKN

14430
EVACLGVIN

14431
EVAETDNFF

14432
EVAHENFRM

14433
EVAHLDATK

14434
EVAPECKFE

14435
EVCDLIHDN

14436
EVCEEEFDP

14437
EVDDKYHED

14438
EVDEGHNNE

14439
EVDGRNRGQ

14440
EVDPLEIAL

14441
EVECRHETC

14442
EVEELWNPF

14443
EVEEMNWGK

14444
EVEEQIELI

14445
EVESLPCSL

14446
EVEWSCRFE

14447
EVGGMGEVK

14448
EVGIQCKID

14449
EVGIQCKVD

14450
EVGPLNEWG

14451
EVGTHDCLD

14452
EVHDFHYIP

14453
EVHDKNTQF

14454
EVHEARPYF

14455
EVHEFRASN

14456
EVHELENIN

14457
EVHMIPEMQ

14458
EVHTEDYGK

14459
EVHVSEIMK

14460
EVIEWCNYE

14461
EVIMQCMMK

14462
EVIPTCHFG

14463
EVKPKDEAH

14464
EVKQWNEVM

14465
EVLIQCKID

14466
EVMHWNIEW

14467
EVMMFSCYE

14468
EVMSLESFE

14469
EVMYRCLMP

14470
EVPEIEHDK

14471
EVPIQCKID

14472
EVPTRDCCD

14473
EVQCDQIHP

14474
EVQDDSKAN

14475
EVQDDYESN

14476
EVQDLDMMD

14477
EVQDTDCSP

14478
EVQEDCMFE

14479
EVQIQCKID

14480
EVRCFHELE

14481
EVREKEPND

14482
EVRESDFRE

14483
EVRFDMRYE

14484
EVRIECKID

14485
EVRIHCKLD

14486
EVRIPCKID

14487
EVRIQCEID

14488
EVRIQCKFD

14489
EVRIQCKFG

14490
EVRIQCKIN

14491
EVRIQCKIV

14492
EVRIQCKIY

14493
EVRIQCKLD

14494
EVRIQCKLV

14495
EVRIQCKMD

14496
EVRIQCQID

14497
EVRIQCTID

14498
EVRIQFKID

14499
EVRIQWKID

14500
EVRIRCKID

14501
EVRIRCKMD

14502
EVRIYCKID

14503
EVRMQCKID

14504
EVRNKCKID

14505
EVRNQCKID

14506
EVRSECKID

14507
EVRSKCKID

14508
EVRSQCKID

14509
EVRTKCKID

14510
EVRTKNEAW

14511
EVRTQCKID

14512
EVRTQIIMD

14513
EVRVQCKID

14514
EVRYNPFHW

14515
EVSELDGAW

14516
EVSEVKENG

14517
EVSGTDELI

14518
EVSLWEIGC

14519
EVIDPNMLD

14520
EVIDYSGMF

14521
EVTEGCPMG

14522
EVTPMQDID

14523
EVTQIWLFP

14524
EVVDCNWLM

14525
EVVEFHWQQ

14526
EVVVRNMPT

14527
EVWIQCKID

14528
EVYDHDTYI

14529
EVYHICGAN

14530
EWIWCYDRG

14531
EWTMFIEAH

14532
FCDCGHWPN

14533
FCECGHWPN

14534
FDCHTFGYR

14535
FDECFPEYW

14536
FDKMFREIL

14537
FDRCRNANW

14538
FDRPGRNIL

14539
FEDCGWDKT

14540
FEIGGKTRT

14541
FENCRDAGR

14542
FERCEGDNH

14543
FHCICAMGH

14544
FHSLCAMGH

14545
FKFEPWEYW

14546
FKHSLRQWT

14547
FKHSQREWT

14548
FKHSRREWT

14549
FKKEDAGQF

14550
FKQDDRSEV

14551
FKVLPWWYH

14552
FLRCKKWHI

14553
FMRPCDMGM

14554
FNRCGRPKN

14555
FNVDCGTWF

14556
FPAFRGGSF

14557
FPIDFVPFR

14558
FPKASECNW

14559
FPKECECNW

14560
FPKGSECTW

14561
FPKHDWPTS

14562
FPPFCWSGM

14563
FPPWPLDLV

14564
FPRFGDDRM

14565
FPRGSECNW

14566
FPSCHQCNI

14567
FQPWPKYWC

14568
FQWRSEMKN

14569
FRHSLREWT

14570
FRMENRNAN

14571
FRQSPWPVV

14572
FRVTPKPEA

14573
FSDPKNMSI

14574
FWECCTDSG

14575
FWGCNWDPK

14576
FWQCPWQED

14577
GGDWDFGEW

14578
GKSADLLKQ

14579
GPDCECQRG

14580
GQPTPWLDH

14581
GRDCKYINW

14582
GRGCQIEKY

14583
GRPCCGYNY

14584
GRPCEPILQ

14585
GRYCECVEH

14586
GSDEFMAYG

14587
GTMNLSWGL

14588
GVCDDHKPN

14589
GWECCTDSG

14590
HARCVDNTV

14591
HCECGHWPN

14592
HDCKLKPIM

14593
HDRCFASYC

14594
HDRCFDSTA

14595
HGVGRSGDN

14596
HHFHVSLDI

14597
HHFQVNLDI

14598
HHFYVNLDI

14599
HHYHVNLDI

14600
HKGMEKNPV

14601
HKYIDWIDG

14602
HPFHVNLDI

14603
HPKCSEQYR

14604
HPRCATDHA

14605
HQRCPDRAD

14606
HRQCNLLDI

14607
HRSCDRMIM

14608
HSDPKNMSI

14609
HTPDRTEFE

14610
HTRCVDHDT

14611
HVDDLNDWA

14612
HVKCLDYQI

14613
IDICGIEHR

14614
IDRMGMILN

14615
IDYISFVDR

14616
IGIGRSGDN

14617
IIAQRDQLN

14618
IIGDFSQLG

14619
IIKEMDIHW

14620
IIPDREDIS

14621
IITQRDQVN

14622
IMDCDTNKY

14623
INDISFVDR

14624
INFISFVDR

14625
INYISFDDR

14626
INYISFIDR

14627
INYISTVAR

14628
INYISFVDP

14629
INYISFVDS

14630
INYISFVNR

14631
INYSSFVDR

14632
ISYISFVDR

14633
ITYISFVDR

14634
IVDQWNMGI

14635
IWDCMVEKF

14636
KDDCGTQER

14637
KDDFDPDQG

14638
KDEIEDFKS

14639
KDKFEWLTH

14640
KDWRGWEER

14641
KDWWEDDKQ

14642
KDWYAWEEW

14643
KEGCETDKS

14644
KEMCQWDAQ

14645
KRDCPWHNL

14646
KSRCADNTV

14647
KVIEFDMDI

14648
LAECMEEMC

14649
LCDYSWWRY

14650
LCPCRLWDW

14651
LCTDKMDGD

14652
LDECCYDTR

14653
LDEYARPQQ

14654
LDRCLVIVQ

14655
LDWAEHFNC

14656
LEFGGKTRT

14657
LEIGAKTRT

14658
LEIGGKTKT

14659
LERCLGLNQ

14660
LESAGKTRT

14661
LHECRKEIH

14662
LKDCHGDGR

14663
LKGCPCDTM

14664
LNCQCWPER

14665
LNGCDSRMR

14666
LTDGWDPER

14667
LWQECDEAW

14668
MDKEIETWP

14669
MDPODEMKF

14670
MDRCPPERT

14671
MDSCYDRVR

14672
MDWFKGERS

14673
MEGSCKQER

14674
MEILGTMKQ

14675
MEQQPGPKR

14676
MGDCQTLSY

14677
MHPRPRPEV

14678
MIDFREDVQ

14679
MIEPYEMHY

14680
MINDMKMFF

14681
MKWINIIDW

14682
MMFENHQRN

14683
MNRQRDHIR

14684
MPKFEDVDH

14685
MQIECTWSH

14686
MQLDCTWHN

14687
MRGCMDKTR

14688
MSDEWMCNN

14689
MTRCVDADY

14690
MVEEQEEND

14691
MVEICCSNK

14692
MVMIWDHDQ

14693
MVNPIDWQC

14694
MVRCVEGDD

14695
MYECGNIGE

14696
NCECGHWPN

14697
NERWDHDQL

14698
NESIDDIEQ

14699
NHWNQELDF

14700
NIALRDSMN

14701
NIDAWDNHN

14702
NIVDDQYGN

14703
NKPFDAFKW

14704
NKWHPMIVI

14705
NLDCRPVNG

14706
NMFPLKPQN

14707
NMFQLKPQH

14708
NMFQLKPQT

14709
NMIQLKPQN

14710
NNWLIRPNE

14711
NSDPKNMPI

14712
NSDPKNMSI

14713
NVNEWEECE

14714
NVREYDRYE

14715
NVVEQHEYN

14716
NWEFNARND

14717
PAEGGNVSD

14718
PAFCQRPEQ

14719
PCAMHNWGF

14720
PCDMNHWGC

14721
PCGMHNWGC

14722
PCGPIEYDE

14723
PCPCDHGGK

14724
PCSQPIHEP

14725
PCTMPYVEN

14726
PCTMTYVED

14727
PDCLRPVMK

14728
PDHCDPMPE

14729
PDHCVKNER

14730
PDICVWHDN

14731
PDIDMDMDY

14732
PDLRPRDAC

14733
PDRFPQIKS

14734
PDTEKEMRF

14735
PDTYVRERY

14736
PEAGDQFKT

14737
PEEMKYPLK

14738
PEVFTDIRP

14739
PEWGTSCNW

14740
PFACDGYEM

14741
PGGCRGIRY

14742
PGICGEPGK

14743
PGICVWHDN

14744
PGKCANERM

14745
PGPCVMEIW

14746
PHFHVNLDI

14747
PHPCMFPTM

14748
PICQGMMEN

14749
PIICVWHDN

14750
PKGSNMFDW

14751
PKKVFCPEE

14752
PKQCKVHEQ

14753
PKSCKFIDT

14754
PKYFECIYY

14755
PMDCGKHKK

14756
PMFHPRQYP

14757
PMMISYIQR

14758
PMRCGDDAK

14759
PNCKQWVDN

14760
PNDLKFETF

14761
PNICPLDEQ

14762
PNKLSGGEE

14763
PNRCHDDRG

14764
PPVRPWEEM

14765
PQFFSWSEY

14766
PQHAKCGEF

14767
PQRVLPMEY

14768
PREFEWYPH

14769
PRGCDCLQS

14770
PRRCDNWKT

14771
PSGPIQYDE

14772
PSKCEWTDN

14773
PSRCFDNTV

14774
PSRCVDNTV

14775
PTMNLSWGL

14776
PTNEEEDGN

14777
PVACVWHDN

14778
PVFCVWHDN

14779
PVGFDPPPF

14780
PVHDFDAAY

14781
PVHSHNIDN

14782
PVICDWHDN

14783
PVICGWHDN

14784
PVICGWHEN

14785
PVICLWHDN

14786
PVICVWHAN

14787
PVICVWHDH

14788
PVICVWHDS

14789
PVICVWHDT

14790
PVICVWHGN

14791
PVICVWHHN

14792
PVICVWHNN

14793
PVICVWPDN

14794
PVICVWYDN

14795
PVIFVWHDN

14796
PVIYIWHDN

14797
PVLCVWHDN

14798
PVLCVWHDN

14799
PVMCVWHDN

14800
PVNCVWHDN

14801
PVQCNWERE

14802
PVRCTKPRD

14803
PVSCVWHDN

14804
PVTCVWHDN

14805
PVVCVWHDN

14806
PWLDDSEME

14807
PWLWHKSAC

14808
PWQCKGPIN

14809
PWQFCYPWT

14810
PWRVQKHGV

14811
PWRWHKSAW

14812
PWRWHKSDC

14813
PWRWHKSPC

14814
PWRWHKSSC

14815
PWRWHKSTC

14816
PWRWHKSVC

14817
PWRWHRSAC

14818
PWRWRKSAC

14819
PWRWYKSAC

14820
PWSCOPEQA

14821
PWTYDKEFY

14822
PWVFTYPSN

14823
PYGPFEYDE

14824
PYGWRNIDQ

14825
PYIMEFHAF

14826
PYIMEFHPC

14827
PYIMEFHPV

14828
PYIMEFHTC

14829
PYSMEFHPF

14830
QANEEEDGN

14831
QCDCSWWRY

14832
QCDYSWWRD

14833
QCDYSWWRH

14834
QCDYSWWRN

14835
QDGTGEPKG

14836
QFRCVDNKV

14837
QIEFIWDAH

14838
QIEMTEISQ

14839
QLSEGEVLN

14840
QKNEEEDGN

14841
QKPCKVEQV

14842
QKRCCWHRC

14843
QLDSWVEKR

14844
QMEEIQIGH

14845
QNLEIIGAG

14846
QPICRRDKR

14847
QPMEWKMVR

14848
QPRCLDNTV

14849
QPRCYRNIS

14850
QSDEYNIAF

14851
QSRCEDNTV

14852
QSRCLDNTL

14853
QSRCVDITV

14854
QSRCVDNPG

14855
QSRCVDNRV

14856
QSRCVDNSV

14857
QSRCVDTTV

14858
QTHEEDDGN

14859
QTHEEEDGN

14860
QTNDKEDGN

14861
QTNEEEAGN

14862
QTNEEEDAN

14863
QTNEEEDGH

14864
QTNEEEDGT

14865
QTNEEEDSN

14866
QTNEEEDVN

14867
QTNEGEDGN

14868
QTNEQEDGN

14869
QTSVEEDGN

14870
QTTEEEDGN

14871
QTYEEEDGN

14872
QVGDDRSFA

14873
QVRIQCKID

14874
QWDDDQWST

14875
QWPDDQWST

14876
QYRCSDSAF

14877
RDCCDAWSR

14878
RDECRMESG

14879
RDKCRMESG

14880
RDPCRMESG

14881
RDPLNDSRY

14882
RDQCRMEAG

14883
RDQCRMECG

14884
RDQCRMEDG

14885
RDQCRMEFG

14886
RDQCRMEHG

14887
RDQCRMESA

14888
RDQCRMESD

14889
RDQCRMESR

14890
RDQCRMESS

14891
RDQCRMESV

14892
RDQCRMETC

14893
RDQCRMEYG

14894
RDQCRMKSG

14895
RDQCRVESG

14896
RDSHDEFEW

14897
REDCWPPEN

14898
REHIGFMKF

14899
REIGGKTRT

14900
REKCHQHQC

14901
RSRCVDNTV

14902
RWGCDLPQI

14903
RYQCRMESG

14904
SADIFAIND

14905
SCECGHWPI

14906
SCECGHWPN

14907
SDPGEILKR

14908
SGDGDFGEW

14909
SGDLDFGEW

14910
SGDWAFGEW

14911
SGDWDFGKW

14912
SGDWNFGEW

14913
SGRCPFSSK

14914
SIEECCEEM

14915
SMFQLKPQN

14916
SNDACIQSF

14917
SNECWNEMA

14918
SNGDCIQAF

14919
SNRCHDDRG

14920
SNVDCIQSF

14921
SPERMDYDG

14922
SPKGSECNW

14923
SQQWCEQPD

14924
SRPCCKDIR

14925
SSDPKNMSI

14926
STMDLSWGL

14927
STMNLSWGF

14928
SVDYEDEVW

14929
SVEPKSMTG

14930
SVETLDOSL

14931
SVICVWHDN

14932
SVLRRDEVN

14933
SVMDIDCHQ

14934
SWLDNILPK

14935
SYFWCFMDC

14936
TADIFAIND

14937
TAEIFAVND

14938
TDKYCDQKR

14939
TFSEIRDQC

14940
THPWEMTKE

14941
THRCHDDRG

14942
TIDSWFQER

14943
TIGEINDAN

14944
TKFHDGDWC

14945
TKMLDRDAE

14946
TNRCHDDRC

14947
TNRCHDDRD

14948
TNRCHDDRV

14949
TNRCHDHRG

14950
TNRCHDYRG

14951
TPPWESIRK

14952
TQPFSKDED

14953
TREHQIISP

14954
TRPCGTLSH

14955
TTHEGHEDN

14956
TVICVWHDN

14957
TVPCMWQTY

14958
TVWEQDLLE

14959
TWAPWCCDC

14960
TWAPWCCYY

14961
TWGPWCCYF

14962
TWPPWCCYF

14963
TWRCGDIGH

14964
TYRCHDDRG

14965
VADCKQPRF

14966
VDCSKILRR

14967
VDDRPKPEH

14968
VDRLFRMEF

14969
VDRQVRMEY

14970
VDSHKIIRR

14971
VEIRGKIRT

14972
VHEGKTFDI

14973
VHQDKIIGE

14974
VICEFEEGN

14975
VITEDDQAE

14976
VNEWEADAE

14977
VQLCDLLRR

14978
VRYSDMFWW

14979
WAEHDFPKG

14980
WCDDDMRRQ

14981
WCTEEWEEK

14982
WDCSMTKKH

14983
WDFSGRIDN

14984
WDHREERTH

14985
WDICSGLRQ

14986
WDKCIPMHW

14987
WDLVDPPKY

14988
WDMNGKDGS

14989
WDPRRPVKK

14990
WDRCLELNK

14991
WDSLEWDRA

14992
WDYSMPKKH

14993
WDYSMSKKH

14994
WDYSMTKKN

14995
WDYSMTKKP

14996
WEFTMAPEV

14997
WEKCGWPPI

14998
WELCMELNQ

14999
WEPCLELNQ

15000
WERCLELDQ

15001
WERCLELHQ

15002
WERCLELNL

15003
WERCLELNP

15004
WERCLEQNH

15005
WERCLEVNQ

15006
WERCLGLNQ

15007
WERCLGLTQ

15008
WERCLQLNQ

15009
WERCLVLNQ

15010
WERCMELNQ

15011
WERCQELNH

15012
WGAFRGVCE

15013
WHGFHKIMF

15014
WIAPRHMGL

15015
WKAFRGGIG

15016
WKDCKQMKM

15017
WKFGDKLIM

15018
WKGAPALER

15019
WKHCEREIN

15020
WKIDPCVLQ

15021
WKLGDGIQD

15022
WKRPPGGEH

15023
WKSHEWDPH

15024
WLECNPPEQ

15025
WLHPYSYCA

15026
WNLQDKPGE

15027
WNPARKDDI

15028
WNSCPWQLP

15029
WPCHDAPIW

15030
WPIGEKMVF

15031
WPKERVPKG

15032
WPNDDMTRM

15033
WQFTGKPTL

15034
WQHIMKPQI

15035
WQMLCRPPV

15036
WQRCLELNQ

15037
WRQFPCAEQ

15038
WRSCPKGLV

15039
WSRCVDNTV

15040
WTIEPEWYF

15041
WTKDDEQLE

15042
WVHPFSYCA

15043
WVHPYTYCA

15044
WVHQYSYCA

15045
WVHTYSYCA

15046
WVLDISHCG

15047
WVRCLELKQ

15048
WVYPYSYCA

15049
WWRVPCESH

15050
WYDWIHERT

15051
WYICQKDSM

15052
WYKCQDVDR

15053
YADPKNMSI

15054
YCECEHWPN

15055
YCECGHCPI

15056
YCECGHSPN

15057
YCECGHWPD

15058
YCECGHWPH

15059
YCECGHWPI

15060
YCECGHWPY

15061
YCECGHWQN

15062
YCECGHWSN

15063
YCECGHWTS

15064
YCECGNWPN

15065
YCECGRWPI

15066
YCECGRWPN

15067
YCECGYWPN

15068
YCEGGHWPN

15069
YCGCGIHQQ

15070
YDESVRISR

15071
YEEFPEIIM

15072
YEETRVEAR

15073
YELYQDIDN

15074
YERCHHQGD

15075
YGPCCHFQF

15076
YHKFGDMDI

15077
YHRCDEFFL

15078
YIRPEQPVC

15079
YKDRDICRL

15080
YKEHEAVQF

15081
YKFYPGLPT

15082
YKGMEKNAG

15083
YKGMEKNRV

15084
YKGMEKTPV

15085
YKMCDHDDC

15086
YKPCMTIDV

15087
YKQHEGEQF

15088
YPHCRLLGP

15089
YPHMDCSEE

15090
YRDCEQMEV

15091
YRDCRQFNR

15092
YRVLPRIKD

15093
YSDAKNMSI

15094
YSDGKNMSI

15095
YSDHQNMSI

15096
YSDPKDMSI

15097
YSDPKNISM

15098
YSDPKNMFI

15099
YSDPKNMPI

15100
YSDPKNMSV

15101
YSDPKNMTL

15102
YSDPKNMTT

15103
YSDPKNMYI

15104
YSDPKTMSI

15105
YSDTKKMSI

15106
YSGPKNMSI

15107
YSHPKHMSI

15108
YSYPKNMTI

15109
YVDPYEMYG

15110
YVFGQKDEM

15111
YVIEQKDEM

15112
YVIGQKDEV

15113
YVLDSWRTS

15114
YWECGHWPN

15115
YWGCPDQVR

15116
YYNHKIFEP

15117
YYNPKIVEP

Example 35
AAV5 Variants with Lymph Node Tissue Tropism as Identified by Machine Learning

This example describes preferred properties of engineered AAV5 variants that display lymph node tissue tropism as identified by machine learning methods. The same bioinformatics pre-process, machine learning methods, and analyses as described in EXAMPLE 19 and FIGS. 19-FIG. 22 were carried out to identify preferred properties of engineered AAV5 variants that display lymph node tissue tropism.

Favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an AAV5 VP capsid polypeptide, which are associated with a higher probability of lymph node tissue tropism are described below. Any of the below machine learning-derived positional amino acid preferences described in TABLE 54 can be present alone or in combination with each other or in combination with any of the amino acid preferences observed and described in EXAMPLE 16, TABLE 31. Listed below in TABLE 54 are a summary of positional features shared between the top important features for lymph node tropism extracted from the ML models.

TABLE 54

Machine Learning-Derived Lymph Node Tissue Tropism Rules

High average flexibility at Xaa1

Xaa1 is selected from D, E, P, G, Q, S, or R

High hbond donors at Xaa1

Xaa1 is selected from R

High mol mass at Xaa1

Xaa1 is selected from Y, W, R, or F

Low solubility at Xaa2

Xaa2 is selected from N or E

Low average flexibility at Xaa3

Xaa3 is selected from W, M, or F

Low mutability at Xaa3

Xaa3 is selected from R, H, K, P, Y, F, L, or C

Low mutability at Xaa4

Xaa4 is selected from C

High mutability at Xaa5

Xaa5 is selected from N

Medium mol mass at Xaa5

Xaa5 is selected from D, I, or N

High mol mass at Xaa6

Xaa6 is selected from Y, W, R, or F

High average flexibility at Xaa6

Xaa6 is selected from G or R

High average flexibility at Xaa7

Xaa7 is selected from D, E, K, P, I, N, Q, or S

Low solubility at Xaa7

Xaa7 is selected from N or E

Low solubility at Xaa8

Xaa8 is selected from N, E, or D

Medium mutability at Xaa8

Xaa8 is selected from R or H

Low mutability at Xaa9

Xaa9 is selected from P or K

High average flexibility at Xaa9

Xaa9 is selected from D, E, P, or S

High solubility at Xaa9

Xaa9 is selected from M or V

TABLE 55 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that exhibited lymph node tissue tropism and comport to one or more of the rules provided in TABLE 54. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 20118-SEQ ID NO: 21117, as disclosed in TABLE 55. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 55

Sequences of the 581 to 589 Region in AAV5

VP1 Capsid Polypeptide that Drive Lymph

Node Tissue Tropism

SEQ

ID
581-589

NO
Sequence

20118
ACKLDHMPP

20119
ADCCSCKAN

20120
ADDCTHKNH

20121
ADSCTHKNH

20122
ADYCAHKNH

20123
ADYCPHKNH

20124
ADYCTHKTH

20125
ADYCTHRNH

20126
ADYGTHKNH

20127
ADYWIHKNH

20128
AECCSIFSS

20129
AERLCIEWD

20130
AESKCDIGT

20131
AESRCDVGT

20132
AEWSWAEHS

20133
AFWGTDVWG

20134
AFWSPDVWG

20135
AGFRIDFRC

20136
AGGWLFEWH

20137
AGNNEDTQL

20138
AGQWGSLNP

20139
AGSNEDKQL

20140
AGTDQTMHM

20141
AHRWSWSES

20142
AISQDMFMQ

20143
AISQDMLMQ

20144
AMLCNRVCE

20145
ANCCSCKAD

20146
ANCCSCKDN

20147
ANCCSCKPN

20148
ANCCSCKTN

20149
ANCCSCKTS

20150
ANCCSGKAN

20151
ANCYSCKAN

20152
ASASHDFMG

20153
ASCEDEGIL

20154
ASGSNDFMG

20155
ATDRVVVNF

20156
ATSRCDVQT

20157
AVPWEWQEG

20158
CCAALIQSG

20159
CCDALIOSS

20160
CCDDLIQSG

20161
CCDPLIQSG

20162
CDMWWYQDP

20163
CEKYNLPKI

20164
CKDRPIIDC

20165
CLREAVVGI

20166
CVLNMEYCL

20167
CVQRLLFKC

20168
CVVNMEYYL

20169
DAAIKMDES

20170
DCIFFHCPK

20171
DDFGPNHRH

20172
DDHQPMQSR

20173
DDRGFTRSM

20174
DDWSWAEHS

20175
DECIMQRSK

20176
DEFDPNHRH

20177
DEFGPNHGH

20178
DEFGPNHIH

20179
DEFGPNHKH

20180
DEFGPNHRR

20181
DEFGPNHRY

20182
DEFGPNYRH

20183
DEFGPSHRH

20184
DEFGTNHRH

20185
DEIGPNHRH

20186
DEMGYDQLD

20187
DERCYDQLD

20188
DEWAWAEHS

20189
DEWPWAEHS

20190
DEWSGAEHS

20191
DEWSLAEHS

20192
DEWSWAAHS

20193
DEWSWAEHA

20194
DEWSWAERS

20195
DEWSWAGHS

20196
DEWSWAQHS

20197
DEWSWSEHS

20198
DEWSWTEHS

20199
DEWSWVEHS

20200
DEWTWAEHS

20201
DEWWSGHAH

20202
DEWWSGHAP

20203
DEYGPNHRH

20204
DFHSRTNIN

20205
DFLDKSTFF

20206
DFLHGTSIM

20207
DFLHVTSTM

20208
DFLPVTSIM

20209
DFLYVTSIM

20210
DGEWGSLNP

20211
DGFRIEFRC

20212
DGLKNDDRK

20213
DGNDQTMHM

20214
DGPDQTMHM

20215
DGQWGSIAL

20216
DGQWGSLNQ

20217
DGQWGSLNS

20218
DGTDPTMHM

20219
DGTDQTMHV

20220
DGTGQTMHM

20221
DGTNQTMHM

20222
DHQGPKLSK

20223
DMYMOQIWR

20224
DNNSPQEAS

20225
DNRCYKYWN

20226
DPICKVEDW

20227
DPIWLIKEC

20228
DPIWRVEDW

20229
DPIWTVEDW

20230
DPKWANCDC

20231
DPMWEREMF

20232
DPSWKVEDW

20233
DPVWKVEDW

20234
DQDAGGYDS

20235
DOWSWAEHS

20236
DRQWGSLNP

20237
DSKNONCEK

20238
DSLHVTSIM

20239
DSNYQGTKH

20240
DSPNQNCFK

20241
DSQNENCFK

20242
DSQNQKCFK

20243
DSQNQNRFK

20244
DSRNQNCFK

20245
DTFGLRFDD

20246
DTFGLRIDD

20247
DTFGQRVDD

20248
DTFWLRVDD

20249
DTQNQNCFK

20250
DTRCDKYWN

20251
DTRCSKYWN

20252
DTRCYKYWT

20253
DVQWGSLNP

20254
DWTDQTMHM

20255
EAHTNSIGK

20256
EANWVYIEA

20257
EARTNSICK

20258
ECEQDDTDI

20259
ECGLWNFSG

20260
ECKITESLT

20261
EDHLHGPGD

20262
EDHWLGPGD

20263
EDHWRGPGD

20264
EDRWHGPGD

20265
EDYWHGPGD

20266
EEAHSVKFV

20267
EEANSVKFA

20268
EEARCVNPC

20269
EEAYSVKFA

20270
EEDHSVKFA

20271
EEFARIPGN

20272
EEFDRIPGH

20273
EEFSRIPGH

20274
EEPETYTQM

20275
EEPRCGNPC

20276
EEPRCVDPC

20277
EEPRCVNPG

20278
EEPRCVNNV

20279
EEPRCVNPY

20280
EEPRCVNTC

20281
EEPRCVSPC

20282
EEPRCVTPC

20283
EFCENLTAI

20284
EFCENMTSI

20285
EFFCEADKA

20286
EFFCPADKA

20287
EFFCQADKT

20288
EFFCQADKV

20289
EFECOANKA

20290
EFFCQVDKA

20291
EFFFQADKA

20292
EFQQDDTDE

20293
EGIKFQTEL

20294
EGKYMDYFM

20295
EGNSNWVFQ

20296
EGPQNELNV

20297
EGVGLYEFA

20298
EHDNQDISV

20299
EHDTQAISV

20300
EHDTQDIIV

20301
EHDTQDSSV

20302
EHHALGVMG

20303
EHHDCVPWS

20304
EHHHSFGCP

20305
EHHNLGYHK

20306
EHHNVGYHT

20307
EHHPLGAMG

20308
EHHPVGVMG

20309
EHHTLGVMG

20310
EHLNLGYHT

20311
EHRNLGYHT

20312
EHVRGMEER

20313
EHVRGMVDR

20314
EHYTODISV

20315
EIAHKLDDI

20316
EIALKLDDL

20317
EINEDYAKI

20318
EKDQRGIGS

20319
EKRCHAIAL

20320
ELAGKAFDH

20321
ELYVILIWW

20322
EMPIEWDEQ

20323
EMRFNNFCY

20324
ENDTQDISV

20325
ENTMSIIEH

20326
ENTMTIIEH

20327
EPIKIENDA

20328
EPIQIFDDA

20329
EPIQIFNDS

20330
EPIQIFSDA

20331
EPIQIVNDA

20332
EPIQMENDA

20333
EPIQSFNDA

20334
EPIWKVEDW

20335
EPLQIENDA

20336
EPNIHAKCV

20337
EPPLSPSAS

20338
EPVEGEWNY

20339
EQHNLGYHT

20340
EQIASYDRV

20341
ERDTQDISV

20342
ESCEDEGLL

20343
ESCEDEGML

20344
ESCEDEGTL

20345
ESCEDEGVL

20346
ESCEDGGIL

20347
ESCEDQGIL

20348
ESCQDEGIL

20349
ESKYVFPNQ

20350
ESKYVEPNL

20351
ESKYVIPYQ

20352
ESKYVLSNQ

20353
ESKYVVPNQ

20354
ESMERWGYD

20355
ESTNDNITI

20356
ESTTDNIAI

20357
ETCGPGGYT

20358
ETHYNEFCK

20359
ETRCYKYWN

20360
ETVETHRDR

20361
EVGGVRIGL

20362
EVKRWMMAD

20363
EVKRWMMDD

20364
EVKRWMMGD

20365
EVKRWMMVG

20366
EVKRWMTVD

20367
EVPKNELNV

20368
EVQRWMMVD

20369
EVRRWMMVD

20370
EWAQPMIMD

20371
EWIQPMIMD

20372
EWTQPLIMD

20373
EWTQPMIMG

20374
FATPKEGFD

20375
FCDGCAPED

20376
FCDRCAAED

20377
FCDRCAPEG

20378
FCDRCAPEV

20379
FCDRCAPEY

20380
FCDRCAPKD

20381
FCDRCATED

20382
FCDRCDPED

20383
FCDRCTPED

20384
FCGRCAPED

20385
FFFFPENED

20386
FFICPENED

20387
FFIFAENED

20388
FFIFHENED

20389
FFIFLENED

20390
FEIFPEDED

20391
FEIFPEHED

20392
FEIFPEIED

20393
FFIFPENAD

20394
FFIFPENEA

20395
FFIFPENEV

20396
FFIEPENGD

20397
FFIEPENQD

20398
FFIEPENVD

20399
FFIFPETED

20400
FFIFPEYED

20401
FFIFRENED

20402
FEIFSENED

20403
FEIFTENED

20404
FEIIPENED

20405
FFIVPENED

20406
FFIYPENED

20407
FFLFPENED

20408
FFNINGSVE

20409
FFVFPENED

20410
FGLKNDDRK

20411
FHALWAECF

20412
FIDAKIWDV

20413
FIIFPENED

20414
FIYKCLFGF

20415
FLVFMIIEG

20416
FLVFMLIEV

20417
FNGNDMLMV

20418
FPEHKDQGI

20419
FQEHKDKGI

20420
FQEHKDQGV

20421
FTNAAPVQD

20422
FWESDRVMM

20423
FWKSDRVMV

20424
FWKTDRVMM

20425
FYGPNWEFH

20426
FYIEPENED

20427
FYLPNWEFH

20428
FYQQFFEGI

20429
FYREFFEGI

20430
FYRKFFEGI

20431
FYRPFFEGE

20432
FYRPNWECH

20433
FYRPNWEFD

20434
FYRPNWEFL

20435
FYRPNWEFY

20436
FYRPNWEEH

20437
FYRPNWEVH

20438
FYRQFFAGE

20439
FYROFFEGF

20440
FYRQFFEGL

20441
FYRQFFEGV

20442
FYRQFFQGE

20443
FYRQYFEGE

20444
GARKNGFMQ

20445
GARMNGFMR

20446
GARMNGFTQ

20447
GARTNGFMQ

20448
GCQQDDTDE

20449
GDCACKFSA

20450
GDCECKFAA

20451
GDCECKFSS

20452
GDCECTFSA

20453
GDCGCKFSA

20454
GDHLGNVWY

20455
GDHWHGPGD

20456
GDEPHRVAC

20457
GDMWWYQDP

20458
GDPADRNTC

20459
GDPGDRNTC

20460
GDPLDRNTC

20461
GDPVDHNTC

20462
GDPVDRNAC

20463
GDRLDREVF

20464
GDYCTHKNH

20465
GDYECKFSA

20466
GDYLGNIWY

20467
GEAQQRVVK

20468
GECECKFSA

20469
GEFGPNHRH

20470
GEFMEQGCG

20471
GEFWNWSRP

20472
GEFWTWSRT

20473
GEIPDRVAC

20474
GEIPHRVSC

20475
GEIPNRVAC

20476
GEIPPRVAC

20477
GEPVDRNTC

20478
GEWSWAEHS

20479
GFLHVISEM

20480
GFPDEGSLQ

20481
GFPDEKSLQ

20482
GFPDERNLQ

20483
GFPDERSLE

20484
GFPDERSLK

20485
GFTDERSLQ

20486
GLELGHVWG

20487
GPFHQDARA

20488
GPFKQDARA

20489
GPFLQDARA

20490
GPFQEDARA

20491
GPFQQDARS

20492
GPFQQDDRA

20493
GPFQQDGRA

20494
GPVQQDARA

20495
GSESNVKAN

20496
GSQCGVPND

20497
GTFSNVKAN

20498
GTISNVKDN

20499
GTESNVKPN

20500
GTESNVKSN

20501
GTESNVKTN

20502
GTESNVNAN

20503
GTESNVRAN

20504
GTKPDTFHG

20505
GTVSNVKAN

20506
GVDMCGLAE

20507
GVESCPDCE

20508
GVISCPECL

20509
GYAGVESLG

20510
GYAGVGELG

20511
HANQDPWDG

20512
HATLDPWDG

20513
HFCANRHIA

20514
HFCDNRHIA

20515
HFCFNRHIA

20516
HFCLNRHIA

20517
HFCVNRHIE

20518
HFCVNRHNA

20519
HFCVNRNEA

20520
HFCVNRPIA

20521
HFFVNRHEA

20522
HFGVNRHEA

20523
HGLDLGCAV

20524
HGLKNDDRK

20525
HGTDQTMHM

20526
HIRLWPFEG

20527
HPMDDACKQ

20528
HSANRDHFV

20529
HSEDRDHFV

20530
HSEHRDHFV

20531
HSENRDHFG

20532
HSENRDHEV

20533
HSENRDPFV

20534
HSENRDYFV

20535
HSGNRDHFV

20536
HSKNRDHFV

20537
HTHYNEFCK

20538
HTKYECYRF

20539
HTSLLHFDH

20540
HWPTHWNIP

20541
HWPTHWNIP

20542
HWWFRKRWV

20543
HWWYRKRWE

20544
HYTKSISHS

20545
IDMWWYQDP

20546
IFDTRWSNV

20547
IFNYSNHND

20548
IFPYSNHND

20549
IINCNGMNW

20550
IINRTGMNW

20551
IIRLWPFEG

20552
ELGQWESYV

20553
EPDWLWKHV

20554
EPYKNQNNE

20555
ESEWNFQDW

20556
EVCQNNANS

20557
IYRPNWEFH

20558
KDHNREHII

20559
KDNECVSHT

20560
KDPETYTQM

20561
KEPEAYTQM

20562
KEPENYTQM

20563
KEPETYIQM

20564
KEPETYTKM

20565
KEPGTYTQM

20566
KEPKTYTQM

20567
KERETYTQM

20568
KESETYTQM

20569
KETETYTQM

20570
KGIDMAMEL

20571
KELTDEFDS

20572
KIWNFTKMM

20573
KIWNVTKVM

20574
KEWSVTKMM

20575
KKPETYTQM

20576
KMQCLTFCD

20577
KNEDYLKWN

20578
KNNEYLKWN

20579
KPHHWNKKT

20580
KPPHWNKET

20581
KPPHWNNKT

20582
KOPETYTQM

20583
KRNEDVLEQ

20584
KSMERWAYD

20585
KSMGRWGYD

20586
KTHYNEFCK

20587
KTEWGWSDC

20588
KTWNVTKMM

20589
KYESEWMEL

20590
LAMDCRPEH

20591
LDDPDTSGQ

20592
LFCVNRHEA

20593
LGECCYRDP

20594
LGMDWGMTD

20595
LGSLNGAMA

20596
LGSSNGAMD

20597
LGSSNGAVA

20598
LHCALGQHS

20599
LHCELGEHS

20600
LHCELGQHP

20601
LHCGLGQHS

20602
LHCKLGQHS

20503
LHCVLGQHS

20604
LHGKVSMHR

20605
LHWELGQHS

20606
LIFPNGDFQ

20607
LIPLWPFEG

20608
LIQLWPFEG

20609
LIRLWQFEG

20610
LIRLWRFEG

20611
LKMDCRPEH

20612
LLICCYRDP

20613
LLINDWISP

20614
LLVYFFVPN

20615
LNVADLPIN

20616
LNVDDMPIN

20617
LNVHDLPIN

20618
LNVVDLPIN

20619
LPADLVDDW

20620
LPADLVQDW

20621
LPADLVYDW

20622
LPDDLVHDW

20623
LPPHWIGHY

20624
LPSDLVHDW

20625
LPVDLVHDW

20626
LQMANRDPF

20527
LQMANRDSC

20628
LQMANRDTF

20629
LQMDNRDSF

20630
LQMPNRDSF

20631
LQMSNRDSF

20632
LOTHSICRD

20633
LOTKSICRD

20634
LRCELGQHS

20635
LRMDCRPEH

20636
LSEWNFHDW

20637
LSICCYRDP

20638
LSMDCRPEH

20639
LTMDCGPEH

20640
LTMDCRAEH

20641
LTMDCRPED

20642
LTMDCRPER

20643
LTMDCRPEY

20644
LTMDCRPGH

20645
LTMDCRPKH

20646
LTMDCRPKR

20647
LTMDGRPEH

20648
LTMGCRPEH

20649
LTMPNSEGK

20650
LTMPNSEQK

20651
LTPDWIGHY

20652
LTPHWIGHS

20653
LTPHWIVHY

20654
LWFCCYRDP

20655
LWICCYQDP

20555
LWICCYRDA

20657
LWICFYRDP

20658
LWMCCYRDP

20659
LWNCCYRDP

20660
LWVCCYRDP

20661
MAGINGMAY

20652
MAGLNGMAN

20663
MAGLNGMAS

20664
MAGLNGMEY

20665
MAGLNGVAY

20666
MAGVNGMAY

20667
MAHNGNWNR

20568
MDMSKAWMI

20669
MELGKDWIQ

20670
MFENWNLGV

20671
MFETKKRVV

20672
MGGLNGMAY

20673
MGIDKAMEL

20674
MGIDMAMEF

20675
MHCELGQHS

20676
MHDDNLNWN

20677
MHSPLIQYG

20678
MIEDRDYFI

20679
MIETKKRVI

20680
MISPLIQYG

20681
MISWWKVDF

20682
MKDAGEWLV

20683
MKFIQMIWV

20684
MLSALIQYG

20585
MLSHLIQYG

20686
MLSPLIQYC

20687
MLSTLIQYG

20688
MNFIQMILV

20689
MNFIQMLWV

20690
MNFISVHMC

20691
MNICVLDGP

20692
MNICVLDGT

20693
MNSAYTMVY

20694
MNSTQTMAY

20695
MNVDDLPIN

20696
MSHCLYVFG

20597
MTCNDMAFY

20698
MTIDSMAPC

20699
MTKWDRLHD

20700
MTKWDRWRD

20701
MWICCYRDP

20702
MWYLWWNCL

20703
MYLDCCDFK

20704
MYLDCRNFK

20705
MYMDCCNFK

20706
MYVDCCNFK

20707
NATEGLGWV

20708
NCVLFECCG

20709
NCVLYEFCG

20710
NCVVYECCG

20711
NEWSWAEHS

20712
NGARADAKK

20713
NGLDLGCGV

20714
NGTDQTMHM

20715
NGTRVDAKK

20716
NKVNWNHSS

20717
NNGFDRYID

20718
NRILDVEHD

20719
NSENRDHFV

20720
NTAEGLGWV

20721
NTIEGLGWV

20722
NTTEGLGWG

20723
NTIQGLGWV

20724
NVMEKYECR

20725
NVMEKYECS

20726
NWSAPQFSD

20727
NWSGPQFTD

20728
NWSTPQFTD

20729
PECCGIFSS

20730
PECCSICSS

20731
PECCSIFAS

20732
PECCSIFCS

20733
PECCSIFSA

20734
PECCSIFST

20735
PECCSIFYS

20736
PECCSIVSS

20737
PECCSIYSS

20738
PECCSLFSS

20739
PECCSSFSS

20740
PECCSTFSS

20741
PECCSVFSS

20742
PECCTIFSS

20743
PECWSIFSS

20744
PECYSIFSS

20745
PEELGYCYV

20746
PEELLYCYV

20747
PEELWYCHV

20748
PEEQWYCYV

20749
PEFCSIFSS

20750
PEKGMVEHQ

20751
PERENLERC

20752
PEWCSIFSS

20753
PFAHMESDA

20754
PFEHMASDA

20755
PFEHMDSDA

20755
PHOWSWSES

20757
PHRWSWIES

20758
PHRWSWSEP

20759
PILWTNHSN

20760
PIPWTNHSN

20761
PIQWTNHIN

20762
PIQWTNHNN

20763
PIQWTNPSN

20764
PISWCTQDL

20765
PISWCTQHV

20766
PLINQCREL

20767
PMLCNRGCE

20768
PMLCNRLCE

20769
PMLCNRVGE

20770
PMLWNRVCE

20771
PMMCNRVCE

20772
PPLGAWWYG

20773
PPPNTFHDS

20774
PQCCSIFSS

20775
PRNNDDWRT

20776
PRQLWEVWQ

20777
PSRDCDSWE

20778
PTIWGWSDC

20779
PTMDCRPEH

20780
PTRCLPLWT

20781
PVEWAYWHM

20782
QAMRCLFDH

20783
QARENLERC

20784
QECCSIFSS

20785
QEGENLERC

20786
QEPETYTQM

20787
QEPHPFSEQ

20788
QERENFERC

20789
QERENLDRC

20790
QERENVERC

20791
QIGCCMLMW

20792
QIIEFEERV

20793
QITCCMLMW

20794
QIWNVTKMM

20795
QLTCPLLDD

20796
QMLCNRVCE

20797
QMPMPPVAD

20798
QNHYNEFCK

20799
QNIEYLKWN

20800
QSDKNWYGM

20801
QSDQNWYSM

20802
OSKRCLEDH

20803
QTDYNEFCK

20804
QTHNNEFCK

20805
QTHYNEFCE

20806
QTHYNEFCR

20807
QTIGGWSDC

20808
QTIWGWGDC

20809
QTIWGWIDC

20810
QTIWGWNDC

20811
QTIWGWSDF

20812
QTIWVWSDC

20813
QTNYNEFCK

20814
QVCSDWCEG

20815
RAHECHQCD

20816
RAHVCDQCD

20817
RAHVCHHCD

20818
RAHVCHPCD

20819
RAHVCHQCA

20820
RAHVCNQCD

20821
RAHVCQQCD

20822
RARPHQGQQ

20823
RDQLDREVF

20824
RDRLDRAVF

20825
RDRLDREAF

20826
RDRLDREGF

20827
RDRLDRELF

20828
RDRLDREVC

20829
RDRLDREVS

20830
RDRLDREVV

20831
RDRLDREVY

20832
RDRLDRQVF

20833
RDRLDWEVF

20834
RDRLGREGF

20835
RDRLGREVF

20836
RDRMDREVF

20837
RDRRDREVF

20838
RDRSYEQMD

20839
RDRVDREVF

20840
RECIPMNSA

20841
REFWTWSRP

20842
REPETYTQM

20843
RERPQQGQQ

20844
RFCVNRHIA

20845
RFPDERSLQ

20846
RGGPQQGQQ

20847
RGHVCHQCD

20848
RGRPEQGQQ

20849
RGRPQEGQQ

20850
RGRPQQGEQ

20851
RGRPQQGKQ

20852
RGRPQQGQR

20853
RGRSQQGQQ

20854
RHFLAWSEI

20855
RHIDFFEGG

20856
RHNDYETHN

20857
RHRWSWSES

20858
RHSDYETHN

20859
RHTDYETHT

20860
RHWWGAACV

20861
RGRPQQGKQ

20862
RIGHNGGGF

20863
RIRLWPFEG

20864
RIWNVTKMM

20865
RKPVEWHWE

20866
RLDLFIGRT

20867
RLGLFMGRT

20868
RLINDWIAP

20869
RLINDWIFP

20870
RLINDWIST

20871
RLNCLENHS

20872
RLNNDWISP

20873
RLNSLDNHS

20874
RLSNDWISP

20875
RLVNDWISP

20876
RMGSKEQED

20877
RMQCLTYCD

20878
RNPISVCSV

20879
RNPVDWHWE

20880
RNPVEWHWK

20881
RNPVEWNWE

20882
RPHVCHQCD

20883
RPWYHENDG

20884
RPWYHEYGG

20885
RRPNAHVDC

20886
RSDVCDNYD

20887
RSHVCHQCD

20888
RSMERWGYD

20889
RTDAQIGWQ

20890
RTDELIGWQ

20891
RTDEQIGWR

20892
RTDQQIGWQ

20893
RVGCLNVWG

20894
RVHVCHQCD

20895
RVMMDWQPD

20896
RWICCYRDP

20897
RYADMMSEY

20898
RYCLAWSEI

20899
RYFFAWSEI

20900
RYFLAWIEI

20901
RYFLAWSAI

20902
RYFLAWSES

20903
RYFLAWTEI

20904
RYFLDWSEI

20905
RYFSAWSEI

20906
RYTWAWSEI

20907
RYILAWSEI

20908
RYVLAWSEI

20909
SAPRLIEDR

20910
SCDRCAPED

20911
SCHTDDAKP

20912
SCKFDHMPP

20913
SCKLAHMPP

20914
SCKLDHMAP

20915
SCKLDHMSP

20916
SCKWDHMPP

20917
SCNPEDSIV

20918
SCNPEGSIG

20919
SDDGDHCTI

20920
SDDGYDIFG

20921
SDHGMHRDG

20922
SDMCWYQDP

20923
SDMGWYQDP

20924
SDMSWYQDP

20925
SDMWCYQDP

20926
SDMWGYQDP

20927
SDMWLYQDP

20928
SDMWWHQDP

20929
SDMWWYEDP

20930
SDMWWYKDP

20931
SDMWWYPDP

20932
SDMWWYQAP

20933
SDMWWYQDA

20934
SDMWWYQDH

20935
SDMWWYQDR

20936
SDMWWYQDT

20937
SDMWWYQHP

20938
SDMWWYQVP

20939
SDMWWYQYP

20940
SDMWWYRDP

20941
SDTWWYQDP

20942
SECCSIFSS

20943
SEGEHQCKA

20944
SEMWNYQDP

20945
SFWGPDVWG

20946
SGLKNDDRK

20947
SGMWWYQDP

20948
SMLCNRVCE

20949
SNCCSCKAN

20950
SNDCNLFDL

20951
SNDCSLCDL

20952
SNMWWYQDP

20953
SNNCNLCDL

20954
SPFQQDARA

20955
SSGSHDFMG

20956
SVFDNAFNN

20957
SVFDNGFDN

20958
SVLPDWGDS

20959
SVPRLIEDG

20960
SWKLDHMPP

20961
TCKLDHMPP

20962
TCNPEDSIG

20963
TCSELPERI

20964
TCVELPERI

20965
TDHNREHIS

20966
TDHNREHMI

20967
TDIDKQRDK

20968
TDVGDYWHA

20969
TDVMLAEWT

20970
TENDIGCAT

20971
TGGWMFFWH

20972
TGIDMAMEL

20973
TGLDFYRWF

20974
THNDIGCAT

20975
TKLNIRDRW

20976
TKTGNNVMH

20977
TKWNNLHIS

20978
TMLCNRVCE

20979
TMPMPPVAD

20980
TNCCSCKAN

20981
TNGFDKYID

20982
TNIDGLWPI

20983
TNNNDRFQV

20984
TNNVSEFMP

20985
TRLNDEDRG

20986
TSGSHDEMG

20987
TSMERWGYD

20988
TSPADHKWR

20989
TSSADHKWR

20990
TSTNDRFQV

20991
TTLNLRDRW

20992
TTMNIRDRW

20993
TTMRGFYHC

20994
TTTGNNVLH

20995
TTWNNLHII

20996
TYITKMDIV

20997
VCDRCAPED

20998
VEQLCIEWD

20999
VERLCIEWV

21000
VESNCIDDV

21001
VFQWINYYM

21002
VGTDQTMHM

21003
VHCELGQHS

21004
VHISNMEGD

21005
VLGQWIGYV

21006
VLGRWISYV

21007
VMEITNLWH

21008
VMGITHLWH

21009
VNCCSCKAN

21010
VNVHNWRCS

21011
VPHNFEFYT

21012
VSADKCNLN

21013
VSAGKCNVN

21014
VSCEDEGIL

21015
VSQNQNCFK

21016
VIKWDRWHD

21017
VWKSDRVMM

21018
VYRPNWEFH

21019
VYRQFFEGI

21020
VYTVNDGCV

21021
WEDSMFCQY

21022
WEHSMFCKY

21023
WEHSMFCRY

21024
WELSMFCQY

21025
WLYIWEKWP

21026
WLYTWEKWT

21027
WLYTWGKWP

21028
WLYTWVKWP

21029
WNPSNMFNI

21030
WRKAIPYVM

21031
WRKGITYVM

21032
WRKGTPYVM

21033
WRKGVPYVM

21034
WRKVIPYVM

21035
WRQGIPYVM

21036
WRRGIPYVM

21037
WVVNFIRQV

21038
WWIIEEHFK

21039
WWIIEEHGK

21040
WWVIEEHCK

21041
WYFLAWSEI

21042
YALKNDDRK

21043
YCHEYDEIV

21044
YCIFEYNGR

21045
YCIGCVDAI

21046
YCLKNDDRK

21047
YCNEFDEIV

21048
YCNEYDEII

21049
YCPAANWNW

21050
YCPEANWNL

21051
YCPEANWSW

21052
YCPEASWNW

21053
YCPETNWNW

21054
YCPGCGLMW

21055
YCPVANWNW

21056
YCREANWNW

21057
YCTEYDEIV

21058
YDHGMHRAG

21059
YDHGMHRDA

21060
YDHGMHRYG

21061
YDHGWKMAC

21062
YDIHPRFFH

21063
YDLCEHRWD

21064
YDPGMHRDG

21065
YEWSWAEHS

21066
YFCVNRHIA

21067
YFIFPENED

21068
YGIIMKIDL

21069
YGIRLKIDL

21070
YGIRNGKIG

21071
YGKISTIIN

21072
YGLENDDRK

21073
YGLKDDDRK

21074
YGLKNDARK

21075
YGLKNDDCK

21076
YGLKNDDRE

21077
YGLKNDDRR

21078
YGLKNDDSK

21079
YGLKNDGRK

21080
YGLKNDNRK

21081
YGLKNDYRK

21082
YGLMNDDRK

21083
YGLQNDDRK

21084
YGMKNDDRK

21085
YGMKNDDRR

21086
YGRKNDDRK

21087
YGVGYLMPA

21088
YGVKNDDRK

21089
YHLIAIGPS

21090
YHLIAIWPP

21091
YHLIAKWPS

21092
YHLIDIWPS

21093
YHPGCDCAC

21094
YLVGYLMPA

21095
YMKHFKNPS

21096
YMKHFRDPS

21097
YMKHFRNAS

21098
YMKHFRNSS

21099
YMKHIRNPS

21100
YMKHVRNPS

21101
YMKNFRNPS

21102
YMKQFRNPS

21103
YMKRFRNPS

21104
YMMHFRNPS

21105
YMRHFRNPS

21106
YNISIMEAG

21107
YQPGCDCSC

21108
YQPGCDCTC

21109
YSENRDHFV

21110
YSERKGEYG

21111
YITGLRVDD

21112
YTNVDGFPI

21113
YTPGNNVTR

21114
YVFDNAFDN

21115
YVKISTVIN

21116
YVLKNDDRK

21117
YWNEYDEIV

Example 36
AAV5 Variants with Mammary Gland Tissue Tropism as Identified by Machine Learning

This example describes preferred properties of engineered AAV5 variants that display mammary gland tissue tropism as identified by machine learning methods. The same bioinformatics pre-process, machine learning methods, and analyses as described in EXAMPLE 19 and FIGS. 19-FIG. 22 were carried out to identify preferred properties of engineered AAV5 variants that display mammary gland tissue tropism.

Favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an AAV5 VP capsid polypeptide, which are associated with a higher probability of mammary gland tissue tropism are described below. Any of the below machine learning-derived positional amino acid preferences described in TABLE 56 can be present alone or in combination with each other or in combination with any of the amino acid preferences observed and described in EXAMPLE 11, TABLE 21. Listed below in TABLE 56 are a summary of positional features shared between the top important features for mammary gland tropism extracted from the ML models.

TABLE 56

Machine Learning-Derived Mammary Gland Tissue Tropism Rules

Low surface accessibility at Xaa1

Xaa1 is selected from C

Medium mol mass at Xaa1

Xaa1 is selected from C

High surface accessibility at Xaa2

Xaa2 is selected from D, N, or Q

Low hydropathy at Xaa2

Xaa2 is selected from D, E, R, K, H, N, or Q

High average flexibility at Xaa3

Xaa3 is selected from D, E, R, P, G, or S

Medium mutability at Xaa3

Xaa3 is selected from R or H

High mutability at Xaa4

Xaa4 is selected from M, I, Q, or T

High solubility at Xaa4

Xaa4 is selected from W, F, I or L

High surface accessibility at Xaa4

Xaa4 is selected from E, R, or K

High solubility at Xaa5

Xaa5 is selected from W, F, I, or L

Low mutability at Xaa5

Xaa5 is selected from Y, F, or L

High hydropathy at Xaa6

Xaa6 is selected from V, I, or L

Medium mol mass at Xaa6

Xaa6 is selected from D, I, L, or N

Low surface accessibility at Xaa8

Xaa8 is selected from C

Low mutability at Xaa8

Xaa8 is selected from C, R, or H

Medium mutability at Xaa9

Xaa9 is selected from R or H

TABLE 57 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that exhibited mammary gland tissue tropism and comport to one or more of the rules provided in TABLE 56. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 23118-SEQ ID NO: 24117, as disclosed in TABLE 57. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV 5 VP1 581 to 589 region.

TABLE 57

Sequences of the 581 to 589 Region in AAV5

VP1 Capsid Polypeptide that Drive

Mammary Gland Tissue Tropism

SEQ
581-589

ID NO
Sequence

23118
AAAQFGKHT

23119
AASKVKLHH

23120
ACDRFLAMY

23121
ACDRFMTMY

23122
ACDRFMVMY

23123
ACDRIMAMY

23124
ACDRVMAMY

23125
ACDRYIAEY

23126
ACDRYMAMY

23127
ADQCKWWCM

23128
ADQCQWWCV

23129
AELMALAPC

23130
AELMVLAQC

23131
AFDRFMAMY

23132
AFTIKDDWW

23133
AGAKYHDWL

23134
AGGKLKTIG

23135
AGGKLQTLV

23136
AIGQFMMHL

23137
AIQQFMMHL

23138
AIRLFMMHL

23139
AIRQFMMHR

23140
AIRQFMMHV

23141
AIRQFTMHL

23142
AIRRFMMHL

23143
AIWPAAWMD

23144
AKLMALAQC

23145
AKVNCVYYG

23146
AKVSTINGM

23147
ALREFWTWW

23148
ALRQFMMHL

23149
ANCKHDHCH

23150
ANCKPDHCH

23151
ANCKQHHCH

23152
ANCRQDHCH

23153
ANRKQDHCH

23154
ANRQFMMHL

23155
ANWKQDHCH

23156
ANYKQDHCH

23157
AQISTLNGM

23158
AQNGLGTVQ

23159
ARGKLQTEG

23160
ARGPMLAVK

23161
ARVSTLNGM

23162
ASARLYYCW

23163
ASDIFDGHQ

23164
ASDKMDGGS

23165
ASDKMDIGS

23166
ASERLHYCW

23167
ASERLYYCL

23168
ASKQWDHKH

23169
ASNKMDVGS

23170
ASQRLYYCW

23171
ASRQFMMHI

23172
ASVRLYYCW

23173
ASYKDQRYY

23174
ATQKITGTS

23175
ATRQFMMHL

23176
ATTQWGMCN

23177
AVHPMAYCF

23178
AWDRFMAMY

23179
CAHKEHVCH

23180
CAHWSYDCY

23181
CAHWTYECY

23182
CAHWTYNCY

23183
CAHWTYYCY

23184
CAHWYYDQR

23185
CANKENVCH

23186
CANKGHVCH

23187
CANWTYDCY

23188
CATKEHVCH

23189
CEGWGTYCL

23190
CEGWVTYCI

23191
CFCELEKMT

23192
CFCTYEIEH

23193
CFCTYKEQH

23194
CFCTYKNEH

23195
CFGFVNSVT

23196
CFGTYKEEH

23197
CFGVVNSVN

23198
CFRTYDLLS

23199
CFWMYDLLS

23200
CEYTYKIEH

23201
CGHWTYDCY

23202
CGLIMEMCC

23203
CGLIMEMCL

23204
CGLIMEMWW

23205
CGLIMEVCW

23206
CGLEMQMCW

23207
CGLEMVMCW

23208
CGLLMEMCW

23209
CGMIMEMCW

23210
CGPIMEMCW

23211
CGVIMEMCW

23212
CHFQYKNQA

23213
CHFRYQNQA

23214
CHMSFNQCY

23215
CHRKWEKQE

23216
CEALRLAWD

23217
CECSSPDHP

23218
CICSSPEQP

23219
CEDLQLAWD

23220
CEDLRLAWG

23221
CIDLRLAWH

23222
CIDLRLDWD

23223
CIDLRLVWD

23224
CIDLWLAWD

23225
CIEWGGMMG

23226
CIEWWDMMG

23227
CIEWWGKMG

23228
CIEWWGMMA

23229
CIEWWGMVG

23230
CIEWWSMMG

23231
CEGAHWHEP

23232
CEGINLDHH

23233
CEGLRLAWD

23234
CEGLRLDWD

23235
CIGRFDHKQ

23236
CIGWWGMMG

23237
CEGYKWFPG

23238
CIHADGKYS

23239
CIIPKLKER

23240
CIIPKMEER

23241
CIIPKMKAR

23242
CIIPKMKEK

23243
CIIPKMKKR

23244
CIIPKMMER

23245
CIIPKMRER

23246
CIIPKVKER

23247
CIIPRMKER

23248
CEIRKMKER

23249
CEISKMKER

23250
CIKNDWQRI

23251
CIKPDWQRI

23252
CIKTDWERI

23253
CIKTDWQGE

23254
CIKTDWQRN

23255
CIKTDWRRI

23256
CILPKMKER

23257
CILSSPEQP

23258
CIMGHCRQH

23259
CIMPKMKER

23260
CIMSDCRQH

23261
CIMSHCREH

23262
CIMSHCRKH

23263
CIMSHCRQD

23264
CIMSHCRQY

23265
CIMSHFRQH

23266
CIMSHGRQH

23267
CIMSHNRQH

23268
CIMSHWRQH

23269
CIMSHYRQH

23270
CIMSRCRQH

23271
CIMTDWQRI

23272
CENADGKYP

23273
CINRFDHKQ

23274
CINSDGKYS

23275
CIPSDRHFG

23276
CIPTYRHLV

23277
CIPTYRHLV

23278
CIRSHCRQH

23279
CIRSSTDQP

23280
CERTDWQRI

23281
CESREDHKL

23282
CISRYDHKQ

23283
CITADGKYS

23284
CETTMNVKT

23285
CITTYSHFG

23286
CIVLRLAWD

23287
CIVPKMKER

23288
CIVWWGMMG

23289
CKHPSWYMD

23290
CKKWTNQAH

23291
CKPWINQAH

23292
CKQWTNQAQ

23293
CKVHKHWDR

23294
CKVHRNWDR

23295
CKVRKNWDR

23296
CLAIFRLQS

23297
CLGATWVAA

23298
CLGSTWAAA

23299
CLGSTWFAA

23300
CLGSTWGAA

23301
CLGSTWVAT

23302
CMCLKKFYY

23303
CMCLKKYYY

23304
CMCLRKCYY

23305
CMCVKKCYY

23306
CMENYHQQY

23307
CMMSHCRQH

23308
CQDFRHDCA

23309
CQDFRHECS

23310
CQGKWEKQE

23311
CQKRIIMQA

23312
CQPKWEKQE

23313
CQQRLIMQA

23314
CQREWEKQE

23315
CQRKWEKQA

23316
CQRRWEKQE

23317
CRGKSHEHT

23318
CRGKSHNHT

23319
CRGKSRDHT

23320
CRGQSHDHT

23321
CRLIMEMCW

23322
CRRKSHDHT

23323
CSIWEDRSG

23324
CSMERRNEL

23325
CSMSHCRQH

23326
CSPNFQDQK

23327
CSRATWGAE

23328
CTDRMVKAA

23329
CTDRMVKAT

23330
CTGNYHQQY

23331
CTHKDIEMD

23332
CTHKQSIAV

23333
CTHRMVKAS

23334
CTLFSGRTA

23335
CTQSFVMYQ

23336
CTVFSGRAA

23337
CTVSSGRTA

23338
CVAIEMSVA

23339
CVCLKKCYY

23340
CVCTYKIEH

23341
CVFREIKTA

23342
CVGINLEHH

23343
CVGINLYHH

23344
CVGLNLDHH

23345
CVHGQYYGQ

23346
CVLIMEMCW

23347
CVLYYNQQC

23348
CVMCYNDCK

23349
CVMSHCRQH

23350
CVQGQYNQE

23351
CVQGQYYQA

23352
CVQGRYYQE

23353
CVQTMNGRS

23354
CVVINLDHH

23355
CVYREIKTS

23356
CVYREIQTA

23357
CVYREITTA

23358
CVYRELKTA

23359
CVYREVKTA

23360
CWAIERLRS

23361
CWAIERMQS

23362
CWAIVRLQS

23363
CWAQLNTDP

23364
CNAQLNTDS

23365
CWDIERLQS

23366
CWDIYNACD

23367
CWGIERLQS

23368
CWGQLNTDT

23369
CWPQLNTDT

23370
CWSIERLQS

23371
CWVMMKTQE

23372
CYCTYKIEH

23373
DAAKFGKHT

23374
DASKVKLHR

23375
DDLGAGRSE

23376
DDNWHAWFM

23377
DDVGAGRSD

23378
DDVMWQLIH

23379
DDWMPQWVD

23380
DEFIWLVMH

23381
DEISNQNSY

23382
DGGVTTYGG

23383
DHVIIKKNY

23384
DIRPSVSDS

23385
DKEWSVHMG

23386
DLDAQGAMW

23387
DLGMINNMA

23388
DMDAKGAMW

23389
DMDAQGSMW

23390
DMSIATWNP

23391
DPDKQGQGI

23392
DPKWEKYAM

23393
DPKWEKYGT

23394
DQAWSVHMG

23395
DQEWSVHMC

23396
DQEWSVNMG

23397
DQKWEKYGL

23398
DQKWEKYGM

23399
DQKWSVHMG

23400
DQVWSVHMG

23401
DREWSVHMG

23402
DSKWLDRPV

23403
DSSKVKLHH

23404
DTDKKGQGI

23405
DTDKQGQGF

23406
DTDKQGQGL

23407
DTDKQGQGM

23408
DTDKQGQGV

23409
DTDKQSQGI

23410
DTNKQGQGI

23411
DVDYRSKAW

23412
DVDYRYEAW

23413
DVFIDWSGR

23414
DVKTDTSRD

23415
DVRQILLCH

23416
DVRQILVCH

23417
DVRQIMMCH

23418
DVRQLLMCH

23419
DYFQYNEAR

23420
DYYQYNEAP

23421
EAISYVENV

23422
ECDIFDGHQ

23423
ECDILDGHQ

23424
ECDRYIAEH

23425
ECDRYIAQY

23426
ECDRYISEY

23427
ECGFIRLNE

23428
ECKPIAGDR

23429
ECYHVLANT

23430
EDIHMIAGH

23431
EDTHMIAGP

23432
EDTHMIVGH

23433
EEFDTIAAP

23434
EEFDTLAAP

23435
EEFDTVAAR

23436
EEFDTVATP

23437
EEFDTVGAP

23438
EEYDTVAAP

23439
EFDIDQCCT

23440
EGDIFDGHQ

23441
EGIWQGMDA

23442
EHVIIEKNY

23443
EHVILKKNY

23444
EHVIMKKNY

23445
EIKVVNSKD

23446
EIKYRRCRA

23447
EIKYRREMA

23448
EIKYRRSMA

23449
EIREGEECC

23450
EIRVVNSRD

23451
EISPAAWMD

23452
EKMESPECL

23453
EKTWLVVWC

23454
EMDAQGAMW

23455
EMRMNIQCE

23456
EPEGALDKT

23457
EPEGELDRT

23458
EPEGELYKT

23459
EPEGEVDKT

23460
EQEAYILRI

23461
ESDIFDAHQ

23462
ESDIFDGHE

23463
ESDIFDGHK

23464
ESDIFDGHP

23465
ESDIFDGHR

23466
ESDIFDGRQ

23467
ESDIFDVHQ

23468
ESDIFHGHQ

23469
ESDIFVGHQ

23470
ESDISDGHQ

23471
ESDIVDGHP

23472
ESDIVDGHQ

23473
ESDPMQLFT

23474
ESDPMQRFT

23475
ESDVFEGHQ

23476
ESKSCGLKG

23477
ESKSCGLRG

23478
ESNIFDGHQ

23479
ESTSCGLTG

23480
ESVDWIKCE

23481
ESWPAAWMD

23482
ETDEFDGHQ

23483
EVEAIRGIR

23484
EVEIFRLEG

23485
EVIAYVENV

23486
FDWMPQWVD

23487
FGGQLRVMY

23488
FGRERVWDS

23489
FGSLYRRRG

23490
FIGLRYKDQ

23491
FKLWKLFQA

23492
FKMDMTKRS

23493
FKMWKLFKA

23494
FKRWKLFQA

23495
FNIPCLFSR

23496
FNIQCLCSR

23497
FNIQCLFTR

23498
FNIQCLISR

23499
FNIQCLYSR

23500
FNIRCLFSR

23501
FNLQCLFSR

23502
FQTSMYCDW

23503
FRGHAMWDA

23504
FRGHGMWYA

23505
FRGHGTWDA

23506
FRGHGVWDA

23507
FRGLGMWDA

23508
FSEQNVSEA

23509
FSIQCLFSR

23510
FTEQIGRQM

23511
FTQQEDRQM

23512
FVCLRYKDQ

23513
FVGWYLNSP

23514
FVGWYWDSP

23515
FVGWYWNPP

23516
FVGWYWNSH

23517
FVGWYWNSR

23518
FVMGYNDCK

23519
GCGLVQRFM

23520
GHFTVLNWG

23521
GNCKQDHCH

23522
GRHTAVATG

23523
GSERLYYCW

23524
GSVMVAITG

23525
GYGRMNKAS

23526
HASICQGGD

23527
HAYIYETWP

23528
HDCDMQASE

23529
HDKWGWWRE

23530
HDRKACMIA

23531
HDVMWQLIH

23532
HDWMPQWVD

23533
HEAKLVPKA

23534
HEFIWLEMH

23535
HEYIYETLP

23536
HEYIYVTWP

23537
HFDFNHQCQ

23538
HFLEVAWGS

23539
HHRHIANNG

23540
HKHMILYHE

23541
HMDIQMVMP

23542
HMDIQMYMP

23543
HMDIQVFMP

23544
HMSIATWYP

23545
HMTFYLSRD

23546
HPSICEGGD

23547
HSEMAMLMQ

23548
HSEMVMWMQ

23549
HSGELYDAF

23550
HSGELYDCF

23551
HSGELYDSF

23552
HSGIHCCCG

23553
HSNFFHLND

23554
HSNFFHMTD

23555
HVDGNEACN

23556
HVDSNIACT

23557
HVDSNISCN

23558
HVDSNLACN

23559
HVDSNVACN

23560
HVNSNIACN

23561
HYCKIIPCH

23562
HYCKIIPSH

23553
HYCKIIPYN

23564
HYCKIITYH

23565
HYCKILPYH

23566
HYCKLIPYH

23567
HYCKMIPYH

23568
HYCKNIPYH

23569
HYCKTIPYH

23570
HYCMIIPYH

23571
HYCQIIPYH

23572
HYCRIIPYH

23573
HYFKIIPYH

23574
HYGKIIPYH

23575
HYYKIIPYH

23576
ICKIRSRLD

23577
ICTIGSRLD

23578
IDRKACMIA

23579
IGVQFLKDH

23580
IIGDKTQDD

23581
IIRDKTEDD

23582
IIRDKTQDG

23583
IIRDKTQND

23584
IIRDKTRDD

23585
IIRDRTQDD

23586
IIRDTTQDD

23537
IIREKTQDD

23588
IIRGKTQDD

23589
IIRYKTQDD

23590
IMRDKTQDD

23591
IPRTYQEMQ

23592
IQHQSRRHM

23593
IQLKAVPSC

23594
IQLKAVICC

23595
IQLKTVTSC

23596
IRGHGMWDA

23597
IRLKAVTSC

23598
IRRHQMTDK

23599
ISRDKTQDD

23600
ITRTYQEME

23601
ITRTYQEMK

23602
ITRTYQERQ

23603
IVRDKTQDD

23604
KANMWLACV

23605
KCGFIRLNE

23606
KCVIGKGRR

23607
KDFWEHECF

23608
KDFWEHLCF

23609
KDFWEHQCV

23610
KDFWERQCF

23611
KDFWEYQCF

23612
KDFWGHQCF

23613
KDFWKHQCF

23614
KDRMGINQC

23615
KDRMGLNQR

23616
KDRMGLNQW

23617
KDRMVLNQC

23618
KDTHMIAGH

23619
KDVTRLRCP

23620
KDVWEHQCF

23621
KEFDTVAAP

23622
KFDEDQCCT

23623
KKFWDTVIY

23624
KKHMILCHF

23625
KQTQIRRHM

23626
KSDIFDGHQ

23627
KSVDWIKCE

23628
KYAYILDCA

23629
KYAYSLYCA

23630
KYGYSLDCA

23631
KYTYSLDCA

23632
KYWNYAEAK

23633
LADIEYSHD

23634
LAGKEGAHF

23635
LCASRGLDA

23636
LCGAIIKAS

23637
LCGLIIKAS

23638
LCGPIIKAA

23639
LCGPIIKAP

23640
LCGPIIKAY

23641
LCGPIIKPS

23642
LCGPIIKSS

23643
LCGPIIRAS

23644
LCGPILKAS

23645
LCGPIVKAS

23646
LCGPLIKAS

23647
LCGQIIKAS

23648
LCTSRGLDP

23649
LDRKACLIA

23650
LDRKACMIG

23651
LDRKACMIS

23652
LDRKACMMA

23653
LDRKACMVA

23654
LDRKACVIA

23655
LDRKAFMIA

23656
LDRKAWMIA

23657
LDRKDCMIA

23658
LDRKPCMIA

23659
LDRKSCMIA

23660
LDRKVCMIA

23661
LDRQACMIA

23662
LDRRACMIA

23663
LFTDYWEQG

23664
LGILYRRRG

23665
LGSPYRRRG

23666
LIRDKTQDD

23667
LKAQFHWGA

23668
LKDQFHWCA

23669
LKDRFHWGA

23670
LKMDMTKHS

23671
LKVDMTKRS

23672
LLFFMKFAK

23673
LQMDMTKRS

23674
LQPQMRRHM

23675
LQPQNRRHM

23676
LQPQSRRHV

23677
LRCQMGDHW

23678
LRMDMTKRS

23679
LRVIQGDNY

23680
LTHKRESEW

23681
LTPKRESEC

23682
LTRKRESEC

23683
LWFYMKFAK

23684
MANKWLACV

23685
MANRWLACV

23686
MASMWLACV

23687
MATMWLACV

23688
MCGLRYYWA

23689
MCGPIIKAS

23690
MCLLIRACE

23691
MCLWILACE

23692
MCLWIRPCE

23693
MCRHIIPDN

23694
MEHQSRRHM

23695
MEPESRRHM

23696
MEPQSRRHI

23697
MEPQSRRHM

23698
MFEKYFQSM

23699
MFMKDELIA

23700
MFMKNELMA

23701
MHKHFEDHK

23702
MHKHFKGHK

23703
MHKHYKDHK

23704
MHKRFKDHK

23705
MHLLKDFCT

23706
MHQHFKDHK

23707
MHRHFKDHK

23708
MIDWGNKCT

23709
MKPQGRRHM

23710
MKPQSRRHI

23711
MLAGWNQDT

23712
MLGEWWPGC

23713
MLNWGNKCT

23714
MNKHFKDHK

23715
MPKHFKDHK

23716
MQHLSRRHM

23717
MQHQCRRHM

23718
MQHRSRRHM

23719
MQLQCRRHM

23720
MQLRSRRHM

23721
MQPDSRRHM

23722
MQPHIRRHM

23723
MQPHSRRDM

23724
MQPLSRRHM

23725
MQPPSRRRM

23726
MQPQFRRHM

23727
MQPQSRRDM

23728
MQPQSRRHK

23729
MQPQSRRNM

23730
MQPRGRRHM

23731
MQPRSRRHM

23732
MQRRSRRHM

23733
MQTESRRHM

23734
MQTQSLRHM

23735
MQTQSRRNM

23736
MRIIQGDNY

23737
MRPQCRRHM

23738
MRPQSRRHK

23739
MRPQSRRHM

23740
MRPQSRRHR

23741
MRPQSRRHR

23742
MSNMWLACV

23743
MSPKPAYWG

23744
MSTKPAYWA

23745
MSTKPFYWG

23746
MTAKMAGRM

23747
MTNMWLACV

23748
MVAIKQYHT

23749
MVGVIRDAS

23750
MVVIKQYQT

23751
MVVIKRYHT

23752
MWFFMKFAK

23753
MWGEWWPGG

23754
MWVIQGDNY

23755
MYKHFKDHK

23756
NDGMVHNCW

23757
NDHWHAWFM

23758
NDKWGWWRQ

23759
NDLMWQLIH

23760
NDNWDAWFM

23761
NDNWHAWFR

23762
NDNWHAWFV

23763
NDNWHAWIM

23764
NDNWHEWFM

23765
NDVMWELIH

23766
NDVMWHLIH

23767
NDVMWQLIR

23768
NDVMWQLIY

23769
NDVMWQLVH

23770
NDWMPQWVD

23771
NEISNQNTY

23772
NFDFNHQCH

23773
NFGIYHLQG

23774
NKHCIAMPC

23775
NKYCIAMAC

23776
NKYCIAMPF

23777
NKYCITMPC

23778
NKYCLAMPC

23779
NMDAQGAMW

23780
NNGEWDVCY

23781
NPKWEKYGM

23782
NQDWLLSVP

23783
NQEWSVHMG

23784
NVDVRESEW

23785
NWHEKRHRG

23786
NYKPSFLTR

23787
PCAIEESHD

23788
PCDRFMAMY

23789
PEDFENHDG

23790
PLRAEMSFH

23791
QCDRVWLNQ

23792
QCEFIRLNE

23793
QCGFIRLDG

23794
QCGFIRLHE

23795
QCGFIRLNG

23796
QCGFIRLNQ

23797
QCGFIRVNE

23798
QCGFLRLNE

23799
QCHIIVAPE

23800
QCHIMVARE

23801
QCHINVARE

23802
QCPIIVARE

23803
QDFWEHQCF

23804
QEFDTVAAP

23805
QEHMILCHF

23806
QETWLVVWC

23807
QFGFIRLNE

23808
QFHIDQCCT

23809
QGGFIRLNA

23810
QGGFIRLNG

23811
QIGCNVMIS

23812
QIGFNVMLS

23813
QIKYRRCMA

23814
QIWPAAWMD

23815
QKDMIICHF

23816
QKDMILCHF

23817
QKDWGRRVT

23818
QKEAYILRI

23819
QKHMIICHF

23820
QKHMILCGF

23821
QKHMILCRF

23822
QKHMILCYF

23823
QKHMLLCHF

23824
QKHMMLCHF

23825
QKHMSLCHF

23826
QKHMTLCHF

23827
QKHRILCHF

23828
QKHRLLCHF

23829
QKHTILCHF

23830
QKHVILCHF

23831
QKIWLVVWC

23832
QKKMILCHF

23833
QKNMILCHF

23834
QKPMILCHF

23835
QKPWLVVWC

23836
QKRMILCHF

23837
QKTWLIVWC

23838
QKTWLVGWC

23839
QKTWLVVWF

23840
QKTWLVVWV

23841
QKTWLVVWY

23842
QNYWCVCGA

23843
QPDPMKQWS

23844
QPKEWVIKD

23845
QPREWVIKE

23846
QQDPMKQWT

23847
QQEACILRI

23848
QQEAFILRI

23849
QQEAYILKI

23850
QQEAYILRL

23851
QQEAYILRV

23852
QQEEYILRI

23853
QQEGYILRI

23854
QQESYILRI

23855
QQEVYILRI

23856
QQHMILCHF

23857
QSDDWIKCE

23858
QSDIFDGHQ

23859
QSDRVWLNP

23860
QSEMAMWMQ

23861
QSGWLLEAK

23862
QSIDWIKCE

23863
QSLDWIKCE

23864
QSVDWIKGE

23865
QSVDWIKYE

23866
QSVDWIQCE

23867
QSVDWIRCE

23868
QSVDWLKCE

23869
QSVDWMKCE

23870
QSVDWVKCE

23871
QSNNWIKCE

23872
QSVYWIKCE

23873
QVKYRTCQW

23874
QVTSDGKIV

23875
QYSRMNKAS

23876
RAKTCIYKY

23877
RAKTCLYKC

23878
RCDGLWCHF

23879
RCGFIRLNE

23880
RCGFIRLNG

23881
RCMKLVGTA

23882
RCRNCGSNG

23883
RDFWEHQCF

23884
RDQCVASCA

23885
REAKVFEIK

23886
REAKVFEFK

23887
REPKVFELK

23888
RESKVFELK

23889
RETKVFELK

23890
RFKLLMTGR

23891
RKTWLVVWC

23892
RLKPSAVQR

23893
RLRNCGSNG

23894
RLVRDSKNT

23895
RLVRDSKTN

23896
RMIQPEHVV

23897
RMKWYLLCV

23898
RMKWYLTCV

23899
RMPIELMHG

23900
RNNWKVDSQ

23901
RNNWKVNSR

23902
RNYWCMCGA

23903
RPATVHASF

23904
RPEMVHASF

23905
RPERVHASF

23906
RQEAYILRI

23907
RQGCLLAWQ

23908
RQPQSRRHM

23909
RRDELLIPD

23910
RRIELLIPD

23911
RRKWYLPCV

23912
RRNELLLPD

23913
RRSELLIPD

23914
RRTELLIPD

23915
RSVDWIKCE

23916
RVADLMYIG

23917
RVGDVMYIG

23918
RVSDVMYIG

23919
RVTDVMYIG

23920
RVVRDSKNN

23921
RWGFIRLNE

23922
RWRHCGSNG

23923
RWRICGSNG

23924
RWRNCGGNG

23925
RWRNCGNNG

23926
RWRNCGSNA

23927
RWRNCSSNG

23928
RWRNGGSNG

23929
RWRSCGSNG

23930
RYCKIIPYH

23931
RYWNYAEAK

23932
SAGIFYAHK

23933
SAYIFYAHK

23934
SCGLVERFM

23935
SDQCQWWCM

23936
SGGLVQRFM

23937
SGNIYESGQ

23938
SIRDKTQDD

23939
SIRQFMMHL

23940
SIRQYMMHL

23941
SKGCIIAMQ

23942
SQVSTLNGM

23943
SSHAANFCG

23944
SSHATTFCG

23945
SSKYEMKFL

23946
STDKDIEMD

23947
STHKDIEMG

23948
STHKDIEMN

23949
STHKDIVMD

23950
STTQWGYCN

23951
SWHEKRHRG

23952
SWSMQSKST

23953
SYGQIEGYQ

23954
TCAVEVIHC

23955
TCDRFMAMY

23956
TCFIGKGRR

23957
TCGIGKGRR

23958
TCIIGKGRR

23959
TCIIGQGRR

23960
TCLIGKGRR

23961
TCVIAKGRR

23962
TCVIEKGRR

23963
TCVIGKGQR

23964
TCVIGQGRR

23965
TCVIGRGRR

23966
TCVIVKGRR

23967
TEDFENHDA

23968
TIRQFMMHL

23969
TKGCIIDMQ

23970
TKLNCVYYG

23971
YKVNCIYYG

23972
TLRAAMSFH

23973
TLRAEMSFR

23974
TLRAEMSIH

23975
TLRAVMSFH

23976
TNCKQDHCH

23977
TRVNCVYYG

23978
TSERLYYCW

23979
TSHATNFCG

23980
TSNKDQRYY

23981
TSSKDQRYY

23982
TSYKDERYY

23983
TSYKNQRYY

23984
TTTQWGMCN

23985
TVLAWGLCN

23986
TVRADMSFH

23987
TYKPSFWTR

23988
TYWNYAESK

23989
VASKTLWET

23990
VCFKLFHVG

23991
VDRKACMIA

23992
VFTDYWEKG

23993
VFTDYWERG

23994
VFVGKNAAW

23995
VGSLYRRRG

23996
VIRDKTQDD

23997
VLDAIHWHW

23998
VQPESRRHM

23999
VRFTVLNWG

24000
VRIQPQHNK

24001
VRLKPQHNK

24002
VRLQPKHNK

24003
VRLQPQHDK

24004
VRLQPQHDR

24005
VRLQPQHSR

24006
VRLQPQLNK

24007
VRLQPQRNK

24008
VSDPMQQFT

24009
VVHQMAYCF

24010
VWDWEHKCT

24011
VWFFMKFAK

24012
VWNGEHKCT

24013
WAAIEYSHD

24014
WADIDYSHD

24015
WADIEYGHD

24016
WADIEYSHA

24017
WAGIEYSHD

24018
WAYIEYSHD

24019
WCGPIIKAS

24020
WDFHSIFPG

24021
WDYHIIFPG

24022
WDYHSICPG

24023
WDYHSIFAG

24024
WDYHSIFPC

24025
WDYHSIYPG

24026
WDYHSVFPG

24027
WDYHTIFPG

24028
WQHGREADH

24029
WSNFALWCQ

24030
WSSFAVWCQ

24031
WSSFILWCQ

24032
YAQSFSQGI

24033
YAWMPQWVD

24034
YCFINSAYP

24035
YCPRLVEAS

24036
YDCDMKASE

24037
YDCDMQASG

24038
YDCDVQASE

24039
YDCVMQASE

24040
YDFDMQASE

24041
YDGMPQWVD

24042
YDLMPKWVD

24043
YDLMPQWVD

24044
YDWDMQASE

24045
YDWMAQWVD

24046
YDWMHQWID

24047
YDWMPEWVD

24048
YDWMPHWGD

24049
YDWMPHWVE

24050
YDWMPKWVD

24051
YDWMPPWVD

24052
YDWMPQLVD

24053
YDWMPQWAD

24054
YDWMPQWDA

24055
YDWMPQWDD

24056
YDWMPQWFD

24057
YDWMPQNGA

24058
YDWMPQWGD

24059
YDWMPQWID

24060
YDWMPQWIG

24061
YDWMPQWIV

24062
YDWMPQWLG

24063
YDWMPQWVA

24064
YDWMPQWVH

24065
YDWMPQWVV

24066
YDWMPQWVY

24067
YDWMRQWVD

24068
YDWMSQWVD

24069
YDWMTQWVD

24070
YDWRPQWVD

24071
YDWVPQWVD

24072
YDYDMQASE

24073
YEAPTILNP

24074
YEWMPQCVD

24075
YFCILDKMT

24076
YFCIMEKNT

24077
YFCIQEKMT

24078
YFCSWNTQE

24079
YFDFNHQCQ

24080
YGHEKRHRG

24081
YGLIMEMCW

24082
YGWMPQWVD

24083
YHGHIANNG

24084
YHMGFNQCY

24085
YHMNFNQCY

24086
YHMSFKQCY

24087
YHMTFNQCY

24088
YHRHIADNG

24089
YHRHTANNG

24090
YHRNIANNG

24091
YIRDGGSHD

24092
YIRDGVSHG

24093
YIRDGVSHH

24094
YIRVGGNNQ

24095
YIRVGVSHD

24096
YLHEKRHRG

24097
YLRSLKCGD

24098
YNWMPQWVD

24099
YQASIYCDW

24100
YQTSIYCDW

24101
YSDVLIYAN

24102
YSDVLVCAN

24103
YSDVMICAN

24104
YVDSNIACN

24105
YVLESNYCE

24106
YVRDGVSHD

24107
YVRQILMCH

24108
YVVESNYCQ

24109
YVWMPQWVD

24110
YWHEQRHRG

24111
YWHERRHRG

24112
YWREKRHRG

24113
YYCKIIPYH

24114
YYFQYNEAP

24115
YYKPSFWTR

24116
YYWHVSAFG

24117
YYWHVSTFW

Example 37
AAV5 Variants with Lung Tissue Tropism as Identified by Machine Learning

This example describes preferred properties of engineered AAV5 variants that display lung tissue tropism as identified by machine learning methods. The same bioinformatics pre-process, machine learning methods, and analyses as described in EXAMPLE 19 and FIGS. 19-FIG. 22 were carried out to identify preferred properties of engineered AAV5 variants that display lung tissue tropism.

Favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an AAV5 VP capsid polypeptide, which are associated with a higher probability of lung tissue tropism are described below. Any of the below machine learning-derived positional amino acid preferences described in TABLE 58 can be present alone or in combination with each other or in combination with any of the amino acid preferences observed and described in EXAMPLE 12, TABLE 23. Listed below in TABLE 58 are a summary of positional features shared between the top important features for lung tropism extracted from the ML models.

TABLE 58

Machine Learning-Derived Lung Tissue Tropism Rules

High mutability at Xaa1

Xaa1 is selected from D, E, M, A, I, Q, or T

High mol mass at Xaa2

Xaa2 is selected from F

Low mutability at Xaa2

Xaa2 is selected from Y, F, or L

Low mutability at Xaa3

Xaa3 is selected from K, V, P, or H

Low hydropathy at Xaa3

Xaa3 is selected from K or R

Low mutability at Xaa4

Xaa4 is selected from K or P

High average flexibility at Xaa4

Xaa4 is selected from D, E, P, or S

Low average flexibility at Xaa5

Xaa5 is selected from W, M, or F

High solubility at Xaa5

Xaa5 is selected from W, F, I, or L

Medium mutability at Xaa6

Xaa6 is selected from R or H

High surface accessibility at Xaa6

Xaa6 is selected from T

Low mutability at Xaa7

Xaa7 is selected from C

High solubility at Xaa7

Xaa7 is selected from W, V, M, F, I, or L

High mutability at Xaa8

Xaa8 is selected from D, E, M, A, I, Q, or T

Low hydropathy at Xaa8

Xaa8 is selected from R or K

High average flexibility at Xaa9

Xaa9 is selected from R or G

TABLE 59 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that exhibited lung tissue tropism and comport to one or more of the rules provided in TABLE 58. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 17118-SEQ ID NO: 18117, as disclosed in TABLE 59. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 59

Sequences of the 581 to 589 Region in AAV5

VP1 Capsid Polypeptide that Drive Lung

Tissue Tropism

SEQ
581-589

ID NO
Sequence

17118
AAKAIHYWM

17119
AAMPFYWFE

17120
AANKMNAER

17121
ACSTVMDFS

17122
ACSTVTDFN

17123
ADADMMWMA

17124
ADCSSSDWS

17125
AEKAIHYWM

17126
AFCCDHNAA

17127
AFEDYGSWE

17128
AFKAPRRGV

17129
AFKATHRGV

17130
AFKATRGGV

17131
AFKATRLGV

17132
AFKATRQGV

17133
AFKATRRAV

17134
AFKATRRGA

17135
AFKATRRGM

17136
AFKATRRSV

17137
AFKSTRRGV

17138
AFKTTRRGV

17139
AFMPKESKE

17140
AFNPQEMWA

17141
AFNQQEVWA

17142
AFQSTDWGW

17143
AFVHWPIHG

17144
AFVNWPLHG

17145
AFVNWTIHG

17146
AFVTWPIHG

17147
AGDFSGYYG

17148
AGETNNGGY

17149
AGSASNWAD

17150
AGVERKITM

17151
AGYASHWAD

17152
AGYASNWGD

17153
AHCHFSNMQ

17154
AHMVENDTP

17155
AHWDFSNMQ

17156
AIHDQLEAM

17157
AKAFIEHTY

17158
AKEQGAVGR

17159
AKGAYNQGN

17160
AKGFLEHTY

17161
AKGSYNQGN

17162
AKGTYNHGN

17163
AKHFYQKFH

17164
AKISAAWDR

17165
AKNAMHNFF

17166
AKNYMHCYW

17167
AKRDHHYFA

17168
AKTDNHVLH

17169
ALTYCHWTD

17170
ALTYCNWAD

17171
AMKNLKRTG

17172
AMVCQAIVF

17173
AMVCQIIVF

17174
AMVCRVIVF

17175
ANSSNHSQE

17176
AQHNSESNI

17177
ARHVNFEMS

17178
ASKMSHKEA

17179
ASKRSGKQP

17180
ASMEHMKKA

17181
ASRIYCRHE

17182
ASRSLERQA

17183
ASYFMEITR

17184
ATEQWDFCR

17185
ATSVFCVFR

17186
ATTSNNLTR

17187
ATTSSNLTR

17188
ATTSYNLAR

17189
AVDPTFKPH

17190
AVEFLAHQY

17191
AVKCTEYSA

17192
AVNPAFKPH

17193
AVYPLFAPE

17194
AWKMMHCDI

17195
AWKMMHCHN

17196
AWKMMHFDN

17197
AWKMMHGDN

17198
AWKMMNCDN

17199
AWQMMHCDN

17200
AWRMMHCDN

17201
AWYASNWAD

17202
AYKNCHRHG

17203
AYRMFNNVQ

17204
CAKACDTSS

17205
CAKDCDASS

17206
CAKNCDTSS

17207
CARDCDTSS

17208
CDYVQREWD

17209
CDYVQRKWD

17210
CDWQRRWID

17211
CERRALKMI

17212
CESIMHPDQ

17213
CFKWTIVNG

17214
CFRQDRIIF

17215
CIESFIENH

17216
CIWQHNVTQ

17217
CKAHFGEME

17218
CKDHFGFMV

17219
CKDHFGVME

17220
CKEAFDVNE

17221
CLRQDRIIS

17222
CPEAFDVNE

17223
CQDHFGFME

17224
CSKDCDTSS

17225
CSPPIVMDR

17226
CSYFMEITR

17227
CTAPIVMDR

17228
CTKDCDTSS

17229
CTPPIAMDR

17230
CTPPIFMDR

17231
CTPPIIMDR

17232
CTPPILMDR

17233
CTPPIVTDR

17234
CTPPTVMDR

17235
CTSFACKSF

17236
CVKDCDTSS

17237
DADNYMIYQ

17238
DADYYMIYR

17239
DARVSNIRQ

17240
DCRWYGNNS

17241
DCTNNELKR

17242
DCTNNKVKR

17243
DCTNTKLKR

17244
DEPCEELAF

17245
DFIQSDTEF

17246
DFVQSDTGF

17247
DGCSLYAFS

17248
DGDYYMIYQ

17249
DGTNNKLKR

17250
DHCGPIAWP

17251
DHPYQHIEG

17252
DICCNYDAQ

17253
DIGMRPSGW

17254
DKHFYEKFH

17255
DKNYMHCHW

17256
DKNYMHWYW

17257
DKNYMHYYW

17258
DKNYTHCYW

17259
DKRDNHYFA

17260
DKRDYHYFA

17261
DKRNHHYFA

17262
DKRYHHYFA

17263
DKSYMHCYW

17264
DKTYMHCYW

17265
DMSHTFVAA

17266
DNAFVHCNQ

17267
DNAIVHCDQ

17268
DNAIVHCNE

17269
DNAIVHCNL

17270
DNAMVHCNQ

17271
DNAVVHCNQ

17272
DNGIVHCNQ

17273
DRCMQAKHG

17274
DRCMQTKHA

17275
DRNYMHCYW

17276
DSAIVHCNQ

17277
DSKEEGLAF

17278
DSMAMSARA

17279
DSMGMTARA

17280
DSREEGLTF

17281
DSRQLHHLG

17282
DSRQLHHTG

17283
DSRQLHRIG

17284
DSRQLHYIG

17285
DSRRLHHIG

17286
DTAIVHCNQ

17287
DTNQINDLS

17288
DTTPNESDR

17289
DVCTVFVTA

17290
DVDYYMIYQ

17291
DVVRVQKMA

17292
DWASNNWWP

17293
DWMTRHKVI

17294
DWMTRNKVI

17295
DWMTRYKLI

17296
DWMTRYKVL

17297
DWSANNWWP

17298
EAEYWSCGR

17299
EAEYWTCGP

17300
EAHYTNECR

17301
EARDSCWWG

17302
EARYCCWWG

17303
EARYSCWSG

17305
ECKQQGVRR

17306
EFCEMQSLC

17307
EFCHALVCQ

17308
EFEDYGGWE

17309
EFEMSVFSN

17310
EFFIFGAID

17311
EFFIIGAIG

17312
EFFIIGAIV

17313
EFFIIGAVD

17314
EFFIIGTID

17315
EFFIIVAID

17316
EFFIVGAID

17317
EFGHAMVCQ

17318
EFITSVQCS

17319
EFKKEFEAF

17320
EFKMSGFSN

17321
EFKMSVFSS

17322
EFKMSVFTN

17323
EFKMSVISN

17324
EFKMSVVSN

17325
EFKMTVFSN

17326
EFKQEFVAF

17327
EFKQEYEAF

17328
EFRMSVFSN

17329
EFTRKSGKF

17330
EFTRRSCKF

17331
EHGCTHMGN

17332
EHKQQGVRR

17333
EHQPFSMFS

17334
EHRQMHDAY

17335
EHRQMHDSF

17336
EHRQMHDSS

17337
EHSCNHMGN

17338
EHSYTHMGN

17339
EHTRQDVCD

17340
EKDHFGVMQ

17341
EKHHFDVMQ

17342
EKLMYDKDL

17343
EKLNHETGY

17344
EKLSHEAGY

17345
EKLTHEAGY

17346
EKMGYVHKG

17347
EMKDLKRTG

17348
EMKNLKRAG

17349
EMKNLKRNG

17350
EMKNLKRTC

17351
EMKNLKRTS

17352
EMKNLRRTG

17353
EMRPQKMSR

17354
ENAQFLVWP

17355
ENAQILVWA

17356
ENAQVLVWA

17357
ENAQYLVWA

17358
ENDSVSAIA

17359
ENEFIYCHP

17360
ENEWTYHQG

17361
ENGQFLVWA

17362
ENHRITGKS

17363
ENHSVSAIG

17364
ENSQASAFG

17365
ENSQFLVWA

17366
ENSQNSAFG

17367
ENSQTSPFG

17368
ENVMHSEFR

17369
ENVQFLVWA

17370
ENYRITGQS

17371
EPRPCNFWC

17372
EPRSCNFWG

17373
EPRSCNFWW

17374
EPRSCNYWC

17375
EQMGYVHKA

17376
ERAEFEMKQ

17377
ERNDFHWYM

17378
ERSCTHMGN

17379
ERSDFHWCM

17380
ERSEFEMKE

17381
ERSGFHWYM

17382
ESKMNHKEA

17383
ESKMSHKQA

17384
ESMFRDSDK

17385
ESMGICHNS

17386
ETIRDMSGF

17387
ETMAQSCYR

17388
ETRSCNFWC

17389
EVASQFFEH

17390
EVFKIDMIP

17391
EVGEMGILR

17392
EVGRIGWMS

17393
EVQKWFRKS

17394
EVRYSCWWG

17395
EVTDEHCCW

17396
EVTNEHCCL

17397
EVTNEHGCW

17398
EVVKIDMIS

17399
EVVKIDMTP

17400
EWKMMHCDN

17401
EWWRYMAYY

17402
EYICFNCYH

17403
EYKEQGVRR

17404
EYKKQGVPR

17405
EYKMSHKEA

17406
EYKPQGVRR

17407
EYKQEGVRR

17408
EYKQKGVRR

17409
EYKQPGVRR

17410
EYKQQCVRR

17411
EYKQQGLRR

17412
EYKQQSVRR

17413
EYQQQGVRR

17414
EYRQQGVRR

17415
EYTKQGVRR

17416
FAPKKCDTW

17417
FCKLQKLKA

17418
FCKLRKLKG

17419
FCKLRKLRA

17420
FCKMRKLKA

17421
FCKSRKLKA

17422
FEAPFNLSF

17423
FEAPINLGF

17424
FEAPINMSF

17425
FEVPINLSF

17426
FFKLRKLKA

17427
FGAPINLSF

17428
FGKLRKLKA

17429
FGRYRLLKG

17430
FGRYRLWEG

17431
FGRYRLWKA

17432
FGRYRLWQV

17433
FGRYRLWRG

17434
FHKKFHSKF

17435
FHKQFHSEF

17436
FHKQFHSRF

17437
FHKQYHSKF

17438
FLKQFHSKF

17439
FNIDCFNAA

17440
FNKQFHSKF

17441
FNVDCFSAA

17442
FRKQFHSKF

17443
GDYVQRQWD

17444
GERWAHNML

17445
GERWPHHML

17446
GERWPHNMW

17447
GERWPNNML

17448
GERWTHNMI

17449
GFDAEHGYQ

17450
GGNMRSSNI

17451
GHIHARDGA

17452
GIKQTLVRR

17453
GIRLKGDIM

17454
GIRLRGNIM

17455
GKRWPHNML

17456
GLRQDRIIF

17457
GNQPTHSPR

17458
GSETNNGGY

17459
GSIIESSAR

17460
GSI1KSSAR

17461
GSIIQGSAR

17462
GSISQSSAR

17463
GSIVQSSAR

17464
GSKMSHKEA

17465
GSMIQSSAR

17466
GSNIQSSAR

17467
GSQPTHSPR

17468
GTPPIVMDR

17469
GTRLKGNIM

17470
GVEFLAHRY

17471
GVKDNNRRL

17472
GWDKLMMLA

17473
GWHCFSFVV

17474
GWKMMHCDN

17475
GWNCFSFVV

17476
GYEPTHSPR

17477
GYKDCHRHG

17478
GYKPTHSPR

17479
GYQMFNNVQ

17480
GYQPTHSHR

17481
GYQPTNSPR

17482
GYRPTHSPR

17483
HATGVGKDR

17484
HAVTHGWHR

17485
HDAQFLPDE

17486
HGEMFDRNS

17487
HIEMRNSHM

17488
HIEMRNTHM

17489
HIEMRNVHM

17490
HIKCTHEDN

17491
HIQCTHEDD

17492
HIQCTHKDN

17493
HIQCTHQDN

17494
HMQCTHEDS

17495
HRIYIMKKM

17496
HRIYIMKQM

17497
HRNYIMKEM

17498
HSAGVGKDP

17499
HSEVKCVER

17500
HSNGVGKDR

17501
HSPTYKWKR

17502
HSTGIGKDR

17503
HSTGLGKDR

17504
HSTVVGKDR

17505
HTDGLFTDD

17506
HTKMTHGTA

17507
HTNCLFTDD

17508
HTRSKDFWN

17509
HVAANYDTF

17510
HVASNYDTS

17511
HVGTHGWHR

17512
HVITHGWHR

17513
HVVHLDDYW

17514
HVVTHGWQR

17515
HVVTLGWHR

17516
HVVTNGWHR

17517
IAIKQRDGA

17518
IANKQQLAR

17519
IAVKQPDGA

17520
IAVPCASFY

17521
IAVPSCWAR

17522
ICHDCMNYR

17523
ICVPSCWAR

17524
IEHDFNSAF

17525
IFHDCMNSR

17526
IFHDFMNYR

17527
IFSPQQLGA

17528
IGEKPVTDR

17529
IGGMFSRNA

17530
IGGMYSRHA

17531
IGRDIDTAF

17532
IGRDVDTSF

17533
IHEWIANVE

17534
IHEWLANVQ

17535
IHGLIPMDS

17536
IHKQFHSKF

17537
IHQWIANVQ

17538
IIANHSDMA

17539
IIANHSHMS

17540
IIANHSHMV

17541
IIKPTRAPA

17542
IIVNHSHMA

17543
IKHDFHSAF

17544
IKHDFNNAF

17545
IKHDFNSAC

17546
IKHDFNSDF

17547
IKHDFNSGF

17548
IKHDFNSPF

17549
IKHDFNSSF

17550
IKHDINSAF

17551
IKHDYNSAF

17552
IKHNFNSAF

17553
IKNAMHNFF

17554
IKNDFNSAF

17555
IKRDFNSAF

17556
IKYDFNSAF

17557
ILLYSCVKD

17558
IMLYSCVQD

17559
INATPNLRT

17560
INVDCFNAA

17561
INVDCFNAA

17562
IQHDFNSAF

17563
IRADLDVDH

17564
IRHDFNSAF

17565
ISAIADLDR

17566
ISAIADMDQ

17567
ISAPPNLRT

17568
ISAPSCWAR

17569
ISDPSCWAR

17570
ISEDIKDNG

17571
ISEDIKENA

17572
ISEDIKENS

17573
ISGPSCWAR

17574
ISIPSCWAR

17575
ISLPSCWAR

17576
ISNKQQLAR

17577
ISVHSCWAR

17578
ISVPGCWAR

17579
ISVPICWAR

17580
ISVPRCWAR

17581
ISVPSCWER

17582
ISVPSCWGR

17583
ISVPSCWSR

17584
ISVPTCWAR

17585
ISVSSCWAR

17586
ISVTSCWAR

17587
ITNKEQLAR

17588
ITNKQELAR

17589
ITNKQLLAR

17590
ITNKQPLAR

17591
ITNKQQLER

17592
ITNKQQLGR

17593
ITNKQQLPR

17594
ITNEMQLSR

17595
ITNKQQQAR

17596
ITNKRQLAR

17597
ITSKQQLAR

17598
ITTKQQLSR

17599
ITTQFMSKH

17600
IVAISCYDN

17601
IVGISCYDD

17602
IVGNSCYDN

17603
IVGTSCYDN

17604
IVKDNNRRL

17605
IWEKPATDR

17606
IWEKPVADR

17607
IWEKPVTAR

17608
IWEKSVTDR

17609
IWEKTVTDR

17610
IYSPNRQDR

17611
IYVPSCWAR

17612
KAEYWNCGR

17613
KCEMISLSR

17614
KEFCCCCKA

17615
KFGSFWPTG

17616
KGDHFDRNS

17617
KHFCCCCKA

17618
KHVMHSEFR

17619
KHWRHTMFL

17620
KKDHFDVMQ

17621
KKEPCNSGH

17622
KLREVGRLC

17623
KMEWFPHNS

17624
KMKNLKRTG

17625
KMRQQKMSR

17626
KNAPTNQPC

17627
KNEWIYHQG

17628
KNEWNYHQG

17629
KNEWTYHQA

17630
KNEWTYHQC

17631
KNVKHSEFR

17632
KNVMHSAFR

17633
KNVMHSQFR

17634
KNVMHSVFR

17635
KPLEMWPDF

17636
KPLLMLPDF

17637
KPMLMWPDF

17638
KPRSCNFWC

17639
KPVLMWPDF

17640
KQAPCNSGH

17641
KQFCCCCRA

17642
KQFCCCRKA

17643
KQFCCCWKA

17644
KQFCFCCKA

17645
KQFCWCCKA

17646
KRFCCCCKA

17647
KSESAASDN

17648
KSEVKCVER

17649
KSKMSHKEA

17650
KSRIMAPLW

17651
KTDYCYQCL

17652
KTERDMSGF

17653
KVGEMGIIR

17654
KVGEMGLLR

17655
KVGEMSILR

17656
KVGNEKSCE

17657
KVHTFFANE

17658
KVQPVRRFT

17659
KWEKPVTDR

17660
KYKQQGVRR

17661
LAAVSVFRR

17662
LACACQDAA

17663
LAETEKLWN

17664
LASANHGGN

17665
LCMHTMWSR

17666
LFDDVMVGR

17667
LFGESMVGT

17668
LGKVENERQ

17669
LGNFEDMPA

17670
LHYAFDQCM

17671
LKEPFNMTF

17672
LKKERGRAC

17673
LKKKRGRSC

17674
LQKKRGRAC

17675
LQSFRSMMD

17676
LQSPLLWER

17677
LRAPYNFWW

17678
LRIYIMKEM

17679
LRKKRGRAC

17680
LRTPFNFWW

17681
LRTPYDFWW

17682
LRTSYNFWW

17683
LRTTYNFWW

17684
LRYSFDQCM

17685
LSTGVGKDR

17686
LSYVQGDWR

17687
LTAVSGFRR

17688
LTFVQGDWR

17689
LTPVSVFRR

17690
LTRCNEKGN

17691
LTTGMIKNW

17692
LTYAQGDWR

17693
LTYVEGDWR

17694
LTYVKGDWR

17695
LTYVQGNWR

17696
LTYVQRDWR

17697
LVEFYSENL

17698
LVHELNQIK

17699
LVHNSPFDF

17700
LVKDEDNNC

17701
LVKMDWKEA

17702
LVNELNQIK

17703
LVRVINERQ

17704
LYCRMHNWA

17705
LYCRMNDWA

17706
MASIFQYPR

17707
MCDDLMVGR

17708
MCKSYERGA

17709
MCMHKMWSR

17710
MCYGFWCEI

17711
MFDDLMVAR

17712
MFDDLMVCR

17713
MFDDLMVGH

17714
MFDDLMVSR

17715
MFDDVMVGR

17716
MFDEVMVGR

17717
MFDNLMVGR

17718
MFDYLMVGR

17719
MFGDLMVGR

17720
MFHDCMNYR

17721
MFNDLMVGR

17722
MGIRDWKRV

17723
MGIREWRRV

17724
MGIWKSQRH

17725
MGTREWKRV

17726
MHNDKLMDE

17727
MHNDKWMDA

17728
MHSPNRQDR

17729
MHYSFDQCM

17730
MIHWDYHMD

17731
MIHWDYNME

17732
MIHWDYWME

17733
MILWDYHME

17734
MINWDYHME

17735
MIRSFTQWA

17736
MIWSNRKFM

17737
MIWYNRKCM

17738
MIWYNRKVM

17739
MIWYNRQFM

17740
MEYWDYHME

17741
MKDEHDDMP

17742
MKDEQDDTP

17743
MKETMDCSY

17744
MKKAHDRSA

17745
MKKKRGRAC

17746
MKLDYDVGC

17747
MKNEKNDNP

17748
MKWDYDVGG

17749
MMKPWACLE

17750
MMKPWVFLE

17751
MMKQWVCLE

17752
MMMMRATMD

17753
MMMMRVAMD

17754
MNTALFVNA

17755
MQIKVSWNR

17756
MQKAHDRAA

17757
MQKEHDRSA

17758
MQKPHDRSA

17759
MQKTHDRSA

17760
MQKWTEEGC

17761
MQLKGSWNR

17762
MQNEKNDHP

17763
MQNEKNDTP

17764
MQNEKNENP

17765
MQTKGSWNR

17766
MRDEQDDMP

17767
MRNDKWMDE

17768
MRNEKNDNP

17769
MRTPYNFWW

17770
MSRTCADWT

17771
MTAVSVFRR

17772
MTKQNKIHR

17773
MTSIFQHPR

17774
MTYVQGDWR

17775
MVDGLMKEY

17776
MVEREWKRV

17777
MVKCTEYSA

17778
MVKVENERQ

17779
MVWYNRKFM

17780
MWKCKSTYA

17781
MYNPNRQDR

17782
MYSANRQDR

17783
MYSPNHQDR

17784
MYSPNREDR

17785
MYSPNRHDR

17786
MYSPNRKDR

17787
MYSPNRQAR

17788
MYSQNRQDR

17789
MYSTNRQDR

17790
MYTPNRQDR

17791
NASNCFWCF

17792
NASTCFWCC

17793
NCRCYGNNS

17794
NESPYHMCL

17795
NFKTYRFCE

17796
NFVQYVHNY

17797
NGSNCFWCC

17798
NHAPAKNKA

17799
NECCNYDAK

17800
NECCNYDAP

17801
NKNAMHNFF

17802
NKNYMHCYW

17803
NLEMLAMFP

17804
NLEMLAMFR

17805
NMRSTEMGV

17806
NNAPEWLHG

17807
NNGVFHWKD

17808
NNPYQHIEG

17809
NNSVFHWKD

17810
NNVKIQYMG

17811
NNWAFHWKD

17812
NNWGFHWKD

17813
NNWVCHWKD

17814
NNWVFHWKA

17815
NNWVFHWKH

17816
NNWVFPWKD

17817
NNWVIHWKD

17818
NPRSKDFWN

17819
NQCGLKMYP

17820
NSAIKEEMA

17821
NSAIRQEMA

17822
NSDMKETGD

17823
NSRQLHHIG

17824
NSRSKDFWN

17825
NSSNCFWCC

17826
NTEPMWSCS

17827
NTRPRRLHM

17828
NTRRMRLHM

17829
NTRSKDFWY

17830
NTSNCFWCC

17831
NMIPLIEYR

17832
NVMTLIQYR

17833
PAEINKMHC

17834
PQAKIDMEF

17835
PQSPLLWER

17836
PQVKVDMEF

17837
PRRLLHFSQ

17838
PRYMGTKDA

17839
PSRLEHYGL

17840
PTKMTHGTS

17841
PTSFAETDY

17842
PVHTFFANE

17843
QAHTFFANE

17844
QAESDMPFP

17845
QFFWSLHAV

17846
QFGTFWPTG

17847
QGEPLMRYS

17848
QHQAFSMFS

17849
QHQPFTMFS

17850
QHSCTHMGN

17851
QICPSFDAD

17852
QINPSFDAD

17853
QEYPSFEAD

17854
QPITSFDAD

17855
QKATTDMYW

17856
QKSPLLWER

17857
QKTPTDMYW

17858
QMKNLKRTG

17859
QMVYAQKNM

17860
QNAWIECQL

17861
QNTWEQCQL

17862
QNVMHSEFR

17863
QPRSCNFWC

17864
QQEPLLWER

17865
QQQPFSMFS

17866
QQSPPLWER

17867
QQSPQLWDR

17868
QQSPQLWER

17869
QQSTLLWER

17870
QRHDLEVDL

17871
QSIRDMSGF

17872
QSQVKCVER

17873
QSREMAPLW

17874
QSSFAETDY

17875
QTERDMGGF

17876
QTIRDMNGF

17877
QTTFAETDY

17878
QTWAAHCTR

17879
QTWSAHCSR

17880
QTWTAHCSR

17881
QYVCFNCYH

17882
RACIPKFKE

17883
RACVPKFKQ

17884
RAEILQLQQ

17885
RARIPKFKQ

17886
RASIFQHPR

17887
RCDEWAEDQ

17888
RCDEWAKDK

17889
RCDEWAKDR

17890
RCDEWAKNQ

17891
RCDEWARDQ

17892
RCDEWVKDQ

17893
RCVPWMNQW

17894
RDAQFLPDQ

17895
RFDDLMVGR

17896
RGDEWAKDQ

17897
RGEILELQQ

17898
RGEILQLQR

17899
RHQPCECYE

17900
RHQPFSMFS

17901
RHWRHAMFL

17902
RHWRLTMFL

17903
RIMVHEWGD

17904
RLDHAETRA

17905
RMLAYKQRH

17906
RVDHAESRA

17907
RVVDLVDVY

17908
RWEKPVTDR

17909
SAEAAYENP

17910
SDCSYSDWS

17911
SEVHLEHNY

17912
SFCCDHNAG

17913
SFCCDHNAS

17914
SFCCDHNGA

17915
SFDERGVWC

17916
SFGSCNTCG

17917
SFHDEEGLF

17918
SFKATRRGV

17919
SFKLTIVNG

17920
SFKWPIVNG

17921
SFKWTILNG

17922
SFKWTIVDG

17923
SFKWTIVNA

17924
SFKWTIVND

17925
SFKWTIVNV

17926
SFKWTIVTG

17927
SGKDNNRRL

17928
SHIHKDSCW

17929
SIMKYDEMG

17930
SIMKYDQMV

17931
SIMLNECNT

17932
SIRLKGNWI

17933
SKISATWDR

17934
SKLHKQYCC

17935
SKYDTDSAC

17936
SLEMRPRQI

17937
SLKDNNRRL

17938
SLKMRPRKI

17939
SLKMRPRQS

17940
SLKMRPRQT

17941
SLQMRPRQI

17942
SLTASNWRD

17943
SMVCQVIVF

17944
SNDKNIHNG

17945
SNWVFHWKD

17946
SPREKNDYW

17947
SPREKNRYW

17948
SRIHKDICW

17949
SRINKDSCW

17950
SRLHKDSCW

17951
SRTMKDWWG

17952
SSAIKQEMA

17953
SSEQTGPHM

17954
SSHFMEITR

17955
SSIIQSSAR

17956
SSKQAGPHM

17957
SSRPRMKQP

17958
SSTASNWRD

17959
SSYFMAITR

17960
SSYFMEIAR

17961
SSYFMEVTR

17962
SSYFMKITR

17963
SSYFTEITR

17964
SSYIMEITR

17965
SSYVMEITR

17966
SSYYMEITR

17967
STDDKFWLG

17968
STDNRFWLG

17969
STDTKFWLG

17970
STEAAHENP

17971
STEASYENP

17972
STESAYENP

17973
STQQVFWNL

17974
STRPMRLHM

17975
STSILDKLI

17976
STSSLDKLV

17977
STTAYNLTR

17978
SVHAKMEGC

17979
SVHSKMQGC

17980
SVHSTMEGC

17981
SVKDINRRL

17982
SVKDNNGRL

17983
SVKDNNQRL

17984
SVKDNNRIL

17985
SVKDNNRKL

17986
SVKDNNRTL

17987
SVKDTNRRL

17988
SVKNNNRRL

17989
SVMQSIDSS

17990
SVNSKMEGC

17991
SVRDNNRRL

17992
SWAASNWRD

17993
SWKMMHCDN

17994
SWTSSNWRD

17995
SWTTSNWRD

17996
SWTVSNWRD

17997
SYQMFNNVQ

17998
TAQYLNREI

17999
TAREDYQHA

18000
TARLLEKYN

18001
TCMGFTSFP

18002
TFFVTASWS

18003
TFHQCERSV

18004
TFKNYRFCE

18005
TFKTYRYCE

18006
TFPNPVHRG

18007
TFRTYRFCE

18008
TFSNPVHRV

18009
TFVNWPIHG

18010
TKIFTENDQ

18011
TKNAMHHFF

18012
TKNDMHNFF

18013
TKNGMHNFF

18014
TKNPMHNFF

18015
TKTAMHNFF

18016
TITTEMIWA

18017
TNWVFHWKD

18018
TPEAEHHKI

18019
TRGMRDFCY

18020
TRMVLEWTY

18021
TSEVFTVNC

18022
TSQYVNREA

18023
TSVPSCWAR

18024
TTEQWDVCR

18025
TIEVEHHKI

18026
TVKCTGYSA

18027
TVKDENAAC

18028
TVKEENAAF

18029
TVKEENATC

18030
TVKEENSAC

18031
TVKEENTAC

18032
TVRCTEYSA

18033
TWKMMHCDN

18034
VARPWAMNS

18035
VATQSCKWM

18036
VAVTITDFD

18037
VAVTVRDFD

18038
VCKWMGRQQ

18039
VCSTVMDFN

18040
VEADNNDAD

18041
VFNQQEMWA

18042
VGKWMGKQQ

18043
VGVTVTDFD

18044
VHALIPMDS

18045
VHGLIPMDT

18046
VHRQMHDSY

18047
VICATQDAA

18048
VICATQDPS

18049
VICPTQDPA

18050
VICTTQDPA

18051
V1HDQLETM

18052
VINMNRRWS

18053
VKKKRGRAC

18054
VKNYMHCYW

18055
VKSFRSMMD

18056
VMRWPWDDT

18057
VPRSCNFWC

18058
VRTPYNFWW

18059
VSETNNGGY

18060
VSINTNQPG

18061
VSKMSHKEA

18062
VSKPIEVGY

18063
VSMSSIYDK

18064
VSMSSNSDK

18065
VSNQCCKAM

18066
VSRALERQA

18067
VSRSLERKA

18068
VSRSLERQG

18069
VSRSLERQT

18070
VSRSIKRQA

18071
VSRSLQRQA

18072
VSRSQERQA

18073
VSVPSCWAR

18074
VSVTVTDFD

18075
VTEHQQRGV

18076
VTKCNEKGN

18077
VINKQQLAR

18078
VINQCCKTM

18079
VTSQCCKAM

18080
VTTGIWKTW

18081
VVGGDKAAY

18082
VVKVINERQ

18083
VVSQTYRNR

18084
VWKQYILVI

18085
VWMQHMCLD

18086
VWQQYILII

18087
WCRQTMMDL

18088
WDAMDDKKA

18089
WETDMDMDA

18090
WFRRHDYFY

18091
WFRRHSYFY

18092
WGDKKATDR

18093
WGDKMAADA

18094
WGDKPATDR

18095
WGDKRATDR

18096
WKNDMDMDA

18097
WKTEMDMDA

18098
WNCVMPDNA

18099
WNTDMDMDA

18100
WVDKQATDR

18101
YCRCYGNNS

18102
YIMVHEWGD

18103
YMILQHDAL

18104
YMTLEHDAL

18105
YQSGGILRM

18106
YQSGGIRRM

18107
YQTGGIHRM

18108
YSHTYKWKR

18109
YSLTYKWKR

18110
YSPTYMWKR

18111
YSQLFTFFE

18112
YSSTYKWKR

18113
YSTTYGGRH

18114
YVTEVNSMI

18115
YWVDFVMPF

18116
YWVNFVVPF

18117
YWVTFVMPF

Example 38
AAV5 Variants with Sciatic Nerve Tissue Tropism as Identified by Machine Learning

This example describes preferred properties of engineered AAV5 variants that display sciatic nerve tissue tropism as identified by machine learning methods. The same bioinformatics pre-process, machine learning methods, and analyses as described in EXAMPLE 19 and FIGS. 19-FIG. 22 were carried out to identify preferred properties of engineered AAV5 variants that display sciatic nerve tissue tropism.

Favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an AAV5 VP capsid polypeptide, which are associated with a higher probability of sciatic nerve tissue tropism are described below. Any of the below machine learning-derived positional amino acid preferences described in TABLE 60 can be present alone or in combination with each other or in combination with any of the amino acid preferences observed and described in EXAMPLE 8, TABLE 15. Listed below in TABLE 60 are a summary of positional features shared between the top important features for sciatic nerve tropism extracted from the ML models.

TABLE 60

Machine Learning-Derived Sciatic Nerve Tissue Tropism Rules

High average flexibility at Xaa1

Xaa1 is selected from G or R

Low solubility at Xaa1

Xaa1 is selected from R or Q

Low mutability at Xaa1

Xaa1 is selected from C, L, F, Y, R, K, P, or H

High volume at Xaa1

Xaa1 is selected from Y or F

High surface accessibility at Xaa2

Xaa2 is selected from E, R, or K

Medium mutability at Xaa3

Xaa3 is selected from H or R

Medium average flexibility at Xaa3

Xaa3 is selected from V or Y

High mutability at Xaa4

Xaa4 is selected from N

High average flexibility at Xaa4

Xaa4 is selected from I, N, G, or R

Low solubility at Xaa4

Xaa4 is selected from N

Low mutability at Xaa6

Xaa6 is selected from C, L, F, or Y

High volume at Xaa6

Xaa6 is selected from K, M, I, or L

Low mutability at Xaa7

Xaa7 is selected from L, F, or Y

Medium mol mass at Xaa7

Xaa7 is selected from D, I, L, or N

High surface accessibility at Xaa8

Xaa8 is selected from S, Y, T, D, P, H, or N

Low mutability at Xaa9

Xaa9 is selected from C, H, or R

Medium solubility at Xaa9

Xaa9 is selected from Q, T, or C

Low surface accessibility at Xaa9

Xaa9 is selected from C

TABLE 61 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that exhibited sciatic nerve tissue tropism and comport to one or more of the rules provided in TABLE 60. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 26991-SEQ ID NO: 27990, as disclosed in TABLE 61. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 61

Sequences of the 581 to 589 Region in AAV5

VP1 Capsid Polypeptide that Drive Sciatic

Nerve Tissue Tropism

HQ
581-589

ID NO
Sequence

26991
AAYLYNVPM

26992
AIRIAMCDW

26993
AIRTAMCDW

26994
AIRTTMCHW

26995
ANGCAIEYH

26996
ANGCTIENH

26997
ANGGAIENH

26998
ANGGAIENN

26999
ARYRMRYHG

27000
ASGCAIENH

27001
ATFGWMNWF

27002
ATSRQAVWA

27003
ATYLYNIPM

27004
AVLLWCEVQ

27005
AYPRNAKYI

27006
AYSRNAKYI

27007
CADWPCMSL

27008
CAGWACMSL

27009
CAKENNAIH

27010
CAKFTNAIY

27011
CAKFTNTIH

27012
CAKFITAIH

27013
CANWACMSL

27014
CCCEALMPK

27015
CCCEALMSQ

27016
CCCEAMMPQ

27017
CCCEAVMPQ

27018
CCCEVLMPQ

27019
CCCPDATNN

27020
CCCSDATNN

27021
CCGQVEFGW

27022
CCSGALMPQ

27023
CDGQVEFGW

27024
CEDPLCQSS

27025
CEDWACMSL

27026
CEHPLCQSS

27027
CEIPLCQSS

27028
CENPLCKSS

27029
CENPLCPSS

27030
CENPLCQSN

27031
CENPLGQSS

27032
CENPLRQSS

27033
CENPLWQ5S

27034
CENPQCQSS

27035
CENSLCQSS

27036
CETPLCQSS

27037
CEVNEEYDW

27038
CFAEPITDC

27039
CFGAEGYAG

27040
CFNEPITDC

27041
CFTAPITDC

27042
CFTEPIADC

27043
CFTEPINDC

27044
CFTEPITDR

27045
CFTEPITNC

27046
CFTESITDC

27047
CFTETITDC

27048
CGGKVEFGW

27049
CGGQAEFGW

27050
CGGQFEFGW

27051
CGGQIEFGW

27052
CGGQIEVGW

27053
CGGQVEAGW

27054
CGGQVECGW

27055
CGGQVEFDW

27056
CGGQVEFSL

27057
CGGQVEFSW

27058
CGGQVEFVW

27059
CGGQVEIGW

27060
CGGQVKFGW

27061
CGGQVQFGW

27062
CGKFTNAIH

27063
CGVQVEFGW

27064
CHPQHHYSH

27065
CHPQHRHSH

27066
CHPQLHHSH

27067
CKDTRKMRE

27068
CKEHRKMRE

27069
CKEPGKMRE

27070
CKEPRKMRA

27071
CKEPRKMRK

27072
CKEPRQMRE

27073
CKEPRRMRE

27074
CKETRKMRE

27075
CKKPRKMRE

27076
CKVNEEYDW

27077
CQIREDRHH

27078
CQNPLCQSS

27079
CQVNAEYDW

27080
CQVNEAYDW

27081
CQVNEEDDW

27082
CQVNEEFDW

27083
CQVNEEYNW

27084
CRCKDAWNM

27085
CRCNAAWNM

27086
CRCNDAGNM

27087
CRCNDAWHR

27088
CRCNDAWNK

27089
CRCNDAWNR

27090
CRCNDAWNV

27091
CRCNDAWIM

27092
CRCNDEWNM

27093
CRCNEAWNM

27094
CRCNGAWNM

27095
CRCSDAWNM

27096
CRFNDAWNM

27097
CRGQVEFGW

27098
CRVNEEYDW

27099
CSSCNHRVF

27100
CVFGWNNYQ

27101
CVGQVEFDW

27102
CVGQVEFGW

27103
CVKFTNAIH

27104
CVWSVGTEW

27105
CWAADSKPC

27106
CYPQHHHSH

27107
DDLDAWQTS

27108
DDLDLWQTS

27109
DDLDVGQTS

27110
DDLDVWETS

27111
DDLDVWKTS

27112
DDLDVWRTS

27113
DDMDVWQTS

27114
DDQDVWQTS

27115
DDRDVWQTS

27116
DFEMWDWVG

27117
DFEVWDWFG

27118
DFNHIGWWQ

27119
DFSHIAWWQ

27120
DFTHIAWWQ

27121
DFYFSGFLI

27122
DFYISGFLI

27123
DFYLSGFLI

27124
DPADKPSNS

27125
DPQDKPSNS

27126
DRIMISIQP

27127
DRVMISIQQ

27128
DRVMVSIQP

27129
DVMMPALKV

27130
DVMMRGLKV

27131
DVMMSGLKV

27132
DWRMVCVHC

27133
EACLYNIPM

27134
EAYFYNIPM

27135
EAYLCMIPM

27136
EAYLYHIPM

27137
EAYLYMIPM

27138
EAYLYNFPM

27139
EAYLYNIRM

27140
EAYLYNISM

27141
EAYLYNVPM

27142
EAYLYSIPM

27143
EAYMYNIPM

27144
EEYLYNIPM

27145
EFKRMTCNR

27146
EGSRKWFAC

27147
EGYLYNIPM

27148
EIGIMRHGH

27149
EIILDDEQR

27150
EIILDDQKR

27151
EIIVDDQQR

27152
EIIVDEQQR

27153
EIMLDDQQR

27154
EIVLDDQQR

27155
EKMIDTCLQ

27156
EKMRHTCLQ

27157
EKMTHPCLQ

27158
EKMTHTCLE

27159
EKMTHTCLK

27160
EKMTHTCLR

27161
EKMTHTCVQ

27162
EKMTHTCVR

27163
EKMTHTWLQ

27164
EPYLYNIPM

27165
EQMTHTCLP

27166
EQMTHTCLQ

27167
ERMTHTCLR

27168
ESHVRPLNM

27169
ETMKHKKRE

27170
ETMQYKKRE

27171
EWTRRMAAP

27172
EWTRRMEAS

27173
EWTRRMEAT

27174
EWTRRMEDP

27175
FAAANQNHM

27176
FAPIVKRHH

27177
FDLSTCYHQ

27178
FEKNSSQMR

27179
FGGQVELGW

27180
FHPYFPTYK

27181
FHVRFDRHH

27182
FIAANQNHM

27183
FKEPRKMRE

27184
FKIRFDRHH

27185
FKMDCGSCG

27186
FKMDCHGCG

27187
FKMDCHSCC

27188
FKMDCHSTG

27189
FKMDCHSGG

27190
FKMDCHTCG

27191
FKMDGHSCG

27192
FKMDWHSCG

27193
FKMGCHSCG

27194
FKMVCHSCG

27195
FKRDCHSCG

27196
FKRICLEAG

27197
FKRICWEAA

27198
FKRICWEGG

27199
FKRICWEPG

27200
FKRICWEVG

27201
FKRICWGAG

27202
FKRICWQAG

27203
FKRMCWEAG

27204
FKRTCWEAG

27205
FKRVCWEAG

27206
FKVDCHSCG

27207
FKVNRDDSW

27208
FKVRFDRHH

27209
FLIRFDRHH

27210
FNAGPLAYN

27211
FNLPFCYHQ

27212
FNLSFCYHL

27213
FNLSICYHQ

27214
FPGVMEAEI

27215
FPGVMQGEI

27216
FPHIVKRHH

27217
FPIRFDRHH

27218
FPIRFDRHL

27219
FPIRFDRHR

27220
FPIRFDRPH

27221
FPPIVKQHH

27222
FPPEVKRHY

27223
FPPTVKRHH

27224
FQIGFDRHH

27225
FQIKFDRHH

27226
FQILFDRHH

27227
FQIPFDRHH

27228
FQIQFDRHH

27229
FQIQIDRHH

27230
FQIRFARHH

27231
FQIRFDGHH

27232
FQIRFDRDH

27233
FQIRFDRHC

27234
FQIRFDRHD

27235
FQIRFDRHR

27236
FQIRFDRNR

27237
FQIRFDRPH

27238
FQIRFDRRQ

27239
FQIRFDRYH

27240
FQIRFGLHH

27241
FQIRFHRHH

27242
FQIRFNRHH

27243
FQIRFYRHH

27244
FQLRFDRHH

27245
FQMRFDRHQ

27246
FQSRFDRHH

27247
FQSRFDRHR

27248
FQTRFDRHH

27249
FQVRFDRHH

27250
FRIRFDRHH

27251
FRIRFDRHY

27252
FSAGALAYN

27253
FSAGGLAYN

27254
FSAGPLAYC

27255
FSAGPLAYY

27256
FSAGPLDYN

27257
FSAGPLSYI

27258
FSAGPLVYN

27259
FSAGPMDYN

27260
FSAGSLAYN

27261
FSAGTLAYS

27262
FSDGPLAYN

27263
FSIRFDRHH

27264
FSPGPLAYN

27265
FSTGPLSYN

27266
FSVGPLAYN

27267
FTPIVKRHH

27268
FTPQKDNSC

27269
FTPQKDTSC

27270
FTQEKDTSC

27271
FTQQKDASC

27272
FVAANQNNM

27273
GAHNELVGY

27274
GASRQAVWA

27275
GAYLYNIPM

27276
GAYLYNLPM

27277
GCCSDATNN

27278
GEVVVCILR

27279
GFAADSKPC

27280
GFNHAIWWQ

27281
GFTGMSDQD

27282
GFYLTREKK

27283
GGGQVEIGW

27284
GIANDDDRE

27285
GIANDDDWA

27286
GIANDDDWE

27287
GIANLDDWE

27288
GIANVADWA

27289
GIANVADWE

27290
GIANVDAWV

27291
GIANVDDLE

27292
GIANVDDWK

27293
GIANVDDWR

27294
GIANVDDWT

27295
GIANVDEWK

27296
GIANVDHWE

27297
GIANVDNWE

27298
GIANVDYWE

27299
GIANVEAWE

27300
GIANVGDWG

27301
GIDNFDDWE

27302
GIDNVDDLE

27303
GIDNVDDWG

27304
GIDNVDEWE

27305
GIDNVDGWE

27306
GIDNVDYWE

27307
GIGNVDDWE

27308
GIPNVDGWE

27309
GISNVDDWE

27310
GISNVDDWG

27311
GISRQAVWA

27312
GITNVDDWE

27313
GITNVDGWE

27314
GIVNVDDWE

27315
GIWRSREHY

27316
GIWRTREHH

27317
GKANDDDWE

27318
GKANVDDRE

27319
GLPNVDDWE

27320
GNWRTREHY

27321
GRANVDDCE

27322
GRCNDAWNM

27323
GSHARPLNM

27324
GSHVGPLNM

27325
GSHVGTLNM

27326
GSHVQPLNM

27327
GSHVRPLHM

27328
GSHVRPLSM

27329
GSHVRSLNM

27330
GTANVDDLE

27331
GTHNELVGY

27332
GTSRQALWA

27333
GTSRQAVWS

27334
GTSRRAVWA

27335
GVTDLQNIC

27336
GVVNVDDWE

27337
GVYRSMFQE

27338
GWAADAKPC

27339
GWAADSEHC

27340
GWAADSEPC

27341
GWAADSKAC

27342
GWAADSKHC

27343
GWAADSKLC

27344
GWAADSKRC

27345
GWAADSKSC

27346
GWAADSKTC

27347
GWAADSRPC

27348
GWAADSTPC

27349
GWAADTKPC

27350
GWAAEAKPC

27351
GWAAGSTPC

27352
GWAANSKPC

27353
GWADDSKPC

27354
GWADDSKTC

27355
GWAGDSKPC

27356
GWASDSKPC

27357
GWATDSKPC

27358
GWAVDSKPC

27359
GWAVNSKPC

27360
GWDDDSKPC

27361
GWGVDSKPC

27362
GWHQKCMGD

27363
GWSADSKPC

27364
GWTADSKPC

27365
GWYKKCMGD

27366
GWYQKCMGN

27367
GWYQKCVGD

27368
GWYQKYMGD

27369
HFNINIYNC

27370
HFYVSGFLI

27371
HGSRNWFAC

27372
HLACLKMEM

27373
HLDCLKMQM

27374
HLDCLQMEM

27375
HLDCLRMEM

27376
HLNCLKMEM

27377
HMDCLKMEM

27378
HQDCLKMEM

27379
HWRMACVHC

27380
HWRMGCVHC

27381
HWRMVCVHG

27382
HWRMVCVYC

27383
IRDWQAYWL

27384
IRDWQCYWL

27385
IRDWQGHWL

27386
IWGIYVHYC

27387
IWGIYVLYF

27388
IWGIYVYYF

27389
KAGCFVDVW

27390
KARFHRERS

27391
KDLDVWQTS

27392
KDRQIDQYH

27393
KENNTNRID

27394
KGAENIHWG

27395
KGAPWAYNC

27396
KGDAYIHWG

27397
KGDENIHWC

27398
KGDEYIHWA

27399
KGDEYIHWC

27400
KGDEYIQGG

27401
KGDEYIRWG

27402
KGDEYIYWG

27403
KGDEYVHWG

27404
KGDGYIHWG

27405
KGDVHIHWG

27406
KGDVYIHWG

27407
KGGAYIHWG

27408
KGGCFVDVW

27409
KGGEYIHWC

27410
KGGLWAYNC

27411
KGGPLAYNC

27412
KGGPWAHNC

27413
KGGPWGYNC

27414
KGGQWAYNC

27415
KGGRWAYNC

27416
KGGVYIHWG

27417
KGHEYIHWG

27418
KGNEYIHWG

27419
KGPEYIHWG

27420
KGTEYIHWG

27421
KGVAYIHWG

27422
KPRFHRERS

27423
KSGRGKLTW

27424
KSRLGKLTW

27425
KSRRAKLTW

27426
KSRRGKLAW

27427
KSRRGKLKW

27428
KSRRGKLPL

27429
KSRRGKLPW

27430
KSRRGKLRW

27431
KSRRGKLSW

27432
KSRRGKLTL

27433
KSRRGKMTW

27434
KSRRGKRTW

27435
KSRRGKVTW

27436
KSRRVKLTW

27437
KSWGWVDTA

27438
KTGHFNQRG

27439
KTRFHREPS

27440
KTRFHRERN

27441
KTRFHRERT

27442
KTRRGRLTW

27443
KTWFHRERS

27444
KVGCSVDVW

27445
KVGCVVDVW

27446
LEKNGSQMR

27447
LIKNSAQMR

27448
LEKNSSEMR

27449
LEKNSSKMR

27450
LEKNSSLMR

27451
LEKNSSPMR

27452
LEKNSSQMK

27453
LEKNSSQTR

27454
LEKNSSQVR

27455
LEKNTSQMR

27456
LEQNSSQMR

27457
LERNSSQMR

27458
LLDCLKMEM

27459
LMENHEFQD

27460
LQDNINYSY

27461
LYLPLCHRI

27462
MASIDKQNV

27463
MCTGELDGT

27464
MCTGELSGT

27465
MCTGEWNGT

27466
MCTGKLNGT

27467
MCTRELNGT

27468
MEKNSSQMR

27469
MEMTQEWMA

27470
MEQNLQLTM

27471
MFTGELNGT

27472
MICGTHDNV

27473
MIFGTHDHV

27474
MEFGTHDNF

27475
MEFGTHDNI

27476
MIIGTHDNC

27477
MMVTKEFNI

27478
MMVTKEFNL

27479
MNLCYSSMM

27480
MQIRFDRHH

27481
MQMAQEWMA

27482
MQMMQMCVP

27483
MQMTQEWMG

27484
MQMTQKWMA

27485
MTAYFKALC

27486
MWTGELNGT

27487
NDLDVWQTS

27488
NELLWCEVQ

27489
NEMLWCEVQ

27490
NFCHSGFLI

27491
NFDSLGFLI

27492
NFEMWDWFG

27493
NFHENEYNC

27494
NFHLSGFLI

27495
NFNEHEYNC

27496
NFNINIFNC

27497
NENINISNC

27498
NFNINTYNC

27499
NFNENVYNC

27500
NFNETEYNC

27501
NFNLSGFLI

27502
NFNMNIYNC

27503
NFSINIYNC

27504
NFSTNIYNC

27505
NFTINIYNC

27506
NFYCSGFLI

27507
NFYFSGFLL

27508
NFYHSGFLI

27509
NFYISGFLI

27510
NFYLGGFLI

27511
NFYLIGFLI

27512
NFYLSGFLM

27513
NFYLSGFLI

27514
NFYLTGFLI

27515
NFYLTGYLI

27516
NFYVSGFLI

27517
NKIDVQWLQ

27518
NMECEKFLT

27519
NMECERFLT

27520
NMECQQFLT

27521
NQEDVQWIQ

27522
NQEDVQWLK

27523
NQLDVQWLQ

27524
NQPCINTYW

27525
NQPCVNTCW

27526
NSRRGRLTW

27527
NVMMPGLKV

27528
PELLWCEVQ

27529
PEVMVCILR

27530
PIFGWMNWF

27531
PKYAWSTSP

27532
PKYHWHMYH

27533
PRLQIRKYL

27534
PSCVTQESE

27535
PSTTVMNVM

27536
PTFGWMDWF

27537
PTFGWMNLF

27538
PTFGWMMAN

27539
PTFGWMMAN

27540
PTFVWMNWF

27541
PTYGWMNWF

27542
PWGDMKDPQ

27543
PWGDMKDSH

27544
PWGVMKDPH

27545
PWGYMKDPH

27546
QAYLYNIPM

27547
QCRLHMQCP

27548
QCRQSHEQE

27549
QCSRNWFAC

27550
QDSRNWFAC

27551
QGGRNWFAC

27552
QGSGNWFAC

27553
QGSRNWFGC

27554
QGSRNWFSC

27555
QGSRNWYAC

27556
QGTRNWFAC

27557
QIGRFAMAH

27558
QIGRFAMPH

27559
QIGRFAMYH

27560
QIGRFTMSH

27561
QNSRHRYAW

27562
QSRRGKLTW

27563
QSSRNWFAC

27564
QTLGWAMGT

27565
RAEGWFKYD

27566
RAHNGLVGY

27567
RANMCDCFE

27568
RANNWTRRH

27569
RASECDYFE

27570
RASMCDFFE

27571
RASMCDGFE

27572
RASMCDYFE

27573
RASMWDCFE

27574
RCHNELVGY

27575
RDSMCDCFE

27576
RDVIVCILR

27577
REAMVCILR

27578
REDMACILR

27579
REDMGCILR

27580
REDMLCELR

27581
REDMVCMLR

27582
REDMVGILR

27583
REDTVCILR

27584
REFMVCILR

27585
REGMLCILR

27586
REGMVCILR

27587
REGVVCILR

27588
REIMVCILR

27589
RELMVCILR

27590
RETMVCILR

27591
REVKVCILR

27592
REVLVCILK

27593
REVMACILR

27594
REVMCCILR

27595
REVMFCILR

27596
REVMGCILR

27597
REVMGCMLR

27598
REVMICILR

27599
REVMLCILR

27600
REVMVCIII

27601
REVMVCIIK

27602
REVMVCIIR

27603
REVMVCIIT

27604
REVMVCILG

27605
REVMVCILK

27606
REVMVCIPR

27607
REVMVCIRR

27608
REVMVCIVI

27609
REVMVCIVR

27610
REVMVCLLR

27611
REVMVCTLR

27612
REVMVCVLK

27613
REVMVCVLR

27614
RFIGMSDQD

27615
RFNGMSDQD

27616
RFPGMSDQD

27617
RFTGMGDQD

27618
RFTGMNDQD

27619
RFTGMSAQD

27620
RFTGMSDED

27621
RFTGMSDKD

27622
RFTGMSDKV

27623
RFTGMSDQY

27624
RFTGMSDRD

27625
RFTGMSGQD

27626
RFTGMSHQD

27627
RFTGMSYQD

27628
RFTGMTDQD

27629
RFTGVSDQD

27630
RGHNGKGPM

27631
RGSMCDCFE

27632
RGVMQQMNH

27633
RIANVDEWE

27634
RIHNELVGY

27635
RIPNVDDWE

27636
RLDCLKMEM

27637
RMSGMLCTV

27638
RMSGMLFTL

27639
RMSGMVCTL

27640
RNHNEIVGY

27641
RNHNELVGC

27642
RNHNELVGY

27643
RNSRHKYAW

27644
RPDLNEEVC

27645
RPDLNEVEC

27646
RPGLNEEEC

27647
RPHNELVGY

27648
RPHNGLVGY

27649
RSHNGLVGY

27650
RSHVQPLNM

27651
RSRRGKLTW

27652
RSRRGKMTW

27653
RSSMCDCFE

27654
RSWRGKLTW

27655
RTDNEEVGY

27656
RTDNELVGD

27657
RTDNGLVGY

27658
RTEGWFEYD

27659
RTEGWFKSD

27660
RTEGWFKYA

27661
RTEGWFKYE

27662
RTEGWVKYD

27663
RTHEELVGY

27664
RTHNALVCY

27665
RTHNALVSY

27666
RTHNALVVY

27667
RTHNDLVGH

27668
RTHNDLVVY

27669
RTHNEFVGY

27670
RTHNEIFGY

27671
RTHNEIIGY

27672
RTHNEEVGF

27673
RTHNEIVGS

27674
RTHNEIVGY

27675
RTHNELFGH

27676
RTHNELFGS

27677
RTHNELEGY

27678
RTHNELGGY

27679
RTHNELHGY

27680
RTHNELEGY

27681
RTHNELLCY

27682
RTHNELLGF

27683
RTHNELVAH

27684
RTHNELVCD

27685
RTHNELVCH

27686
RTHNELVCS

27687
RTHNELVCY

27688
RTHNELVFY

27689
RTHNELVGE

27690
RTHNELVGF

27691
RTHNELVGG

27692
RTHNELVGH

27693
RTHNELVGQ

27694
RTHNELVGV

27695
RTHNELVIY

27696
RTHNELVRH

27697
RTHNELVRY

27698
RTHNELVSF

27699
RTHNELVSY

27700
RTHNELVVF

27701
RTHNELVVY

27702
RTHNEPVGY

27703
RTHNEVVGC

27704
RTHNEVVGD

27705
RTHNEVVGS

27706
RTHNGLVGF

27707
RTHNGLVGS

27708
RTHNKLGGY

27709
RTHNKLVGN

27710
RTHNKLVGY

27711
RTHNQLVGC

27712
RTHNQLVGY

27713
RTHNVLVGS

27714
RTHNWLVGY

27715
RTHSELVGY

27716
RTHSQLVGY

27717
RTHTKLVGY

27718
RTMGQFYYE

27719
RTNNEIVGY

27720
RTNNELAGY

27721
RTNNELIGY

27722
RTNNELVGS

27723
RTPNALVGY

27724
RTQGWFKYD

27725
RTQNELVGS

27726
RTQNELVVY

27727
RTQNGLVGY

27728
RTRNELEGY

27729
RTRNELLGY

27730
RTRNELVGY

27731
RTRNELVVY

27732
RTRNKLVGY

27733
RTRSELVGY

27734
RTYNELVGY

27735
RVGMQQMNH

27736
RVMAEMVWE

27737
RVSGMLCTL

27738
RVVMQPMNH

27739
RVVMVCILR

27740
RWAADSKPC

27741
RWGDMKDPH

27742
RWGDMKDPH

27743
RWHNGKAPM

27744
RWHNGKEPM

27745
RWHNGKGHM

27746
RWHNGKGPK

27747
RWHNGKGRM

27748
RWHNGKGSM

27749
RWHNGKVPM

27750
RWHNGQGPM

27751
RWHNGRGPM

27752
RWHTGKGPM

27753
RWNNGKGPM

27754
RWRMVCVHC

27755
RYGNVAHHA

27756
RYSNVAHYA

27757
RYTGMSDQD

27758
SATNWECSK

27759
SCCADATNN

27760
SCCSDAANK

27761
SCCSDAANN

27762
SCCSDANNN

27763
SCCSDATDN

27764
SCCSDATNH

27765
SCCSDATQN

27766
SCCSDATSN

27767
SCCYDATNN

27768
SCTTRVWEC

27769
SCWSDATNN

27770
SCYSDATNN

27771
SEVMVCVLR

27772
SEYHWHMYH

27773
SFCSDATNN

27774
SFCSDATNS

27775
SFYLSGFLI

27776
SKCHWHMYH

27777
SKFAWSTSP

27778
SKFHWHMYH

27779
SKYAWSTAP

27780
SKYAWSTSQ

27781
SKYAWSTSS

27782
SKYAWSTTP

27783
SKYDWSTSP

27784
SKYGWSTSP

27785
SKYHWDMYH

27786
SKYHWHEYP

27787
SKYHWHMYL

27788
SKYHWHMYR

27789
SKYHWNMYH

27790
SKYHWPMYH

27791
SKYHWQIYH

27792
SKYHWRMYH

27793
SKYLWHMYH

27794
SKYNWHMYH

27795
SKYNWHMYQ

27796
SKYPWSTSP

27797
SKYQWHMYH

27798
SKYRWHMYH

27799
SKYSWSTSP

27800
SKYTWSTSP

27801
SKYVWSTSP

27802
SKYYWHMYH

27803
SPTNWECSN

27804
SRYAWSTSQ

27805
SRYHWHMYH

27806
STFGWMNWF

27807
STMTDAPCG

27808
STTNWECSN

27809
SVCSDATNN

27810
SVMMSPHLM

27811
SWAADSKPC

27812
SYARKAYDC

27813
SYPRRVYDC

27814
TALLWCEVL

27815
TAYMTAMYC

27816
TCCSDATNN

27817
TDLLWCELQ

27818
TDLLWCEVH

27819
TDRQIAQYH

27820
TDRQIDQYL

27821
TDRQIDQYR

27822
TDRQIGQYH

27823
TDRQIHQYH

27824
TDRQVDQYH

27825
TELFWCDVQ

27826
TELFWCEVL

27827
TELLLCEVH

27828
TELLWCAVQ

27829
TELLWCEIK

27830
TELLWCEIQ

27831
TELLWCELQ

27832
TELLWCELR

27833
TELLWCEVA

27834
TELLWCEVE

27835
TELLWCEVG

27836
TELLWCGVQ

27837
TELLWCKVQ

27838
TELLWCQVQ

27839
TELLWCVVQ

27840
TELLWFEVQ

27841
TELLWGEVQ

27842
TELLWREVQ

27843
TELLWSEIQ

27844
TELSWCEVQ

27845
TELVWCEVK

27846
TEMLWCEVK

27847
TEMLWCEVL

27848
TEMLWCEVQ

27849
TERLWCEVL

27850
TFNINIYNC

27851
TRAMPDDNC

27852
TFWMPGNNC

27853
TFYLSGFLI

27854
TGLLWCEVQ

27855
TISHDEAHK

27856
TKLLWCEVK

27857
TKYAWSTSP

27858
TKYHWHMYH

27859
TMGEVVINL

27860
TMSEVVINL

27861
TNGCAIENH

27862
TPGIHRHCC

27863
TPGISRHCC

27864
TPGITRHCC

27865
TPGIYRHCC

27866
TPGLNRHCC

27867
TQLLWCEVQ

27868
TRRSWECSP

27869
TRYRMPYHG

27870
TRYRMRYHC

27871
TRYRMRYHS

27872
TRYRMRYNG

27873
TSCGYHDNN

27874
TSCGYNDNS

27875
TSCRYNDNN

27876
TSRSAAADL

27877
TSRTEAADL

27878
TSSHDEAHR

27879
TTFGWMNWF

27880
TTGINRHCC

27881
TTMPDAPCR

27882
TTSHDEAHK

27883
TVLLWCDVQ

27884
TVLLWCEVH

27885
TVLLWCEVQ

27886
TWEIQIDCI

27887
TWETQIDCI

27888
TWGSQIDCI

27889
TWQSQIDCI

27890
TYPRNAKYT

27891
VEDKINYSY

27892
VEKNSSQMR

27893
VGYRSMFQE

27894
VIGIIRHGH

27895
VISNVDDWE

27896
VLYRSMFQE

27897
VQAQINYSY

27898
VQDEINYSY

27899
VQDKFNYSY

27900
VQDKINFSY

27901
VQDKINYGY

27902
VQDKINYNY

27903
VQDKINYSH

27904
VQDKISYSY

27905
VQDKITYSY

27906
VQDKLNYSH

27907
VQDKLNYSY

27908
VQDKRNYSY

27909
VQDKSNYSY

27910
VQDKVNYSY

27911
VQDMLNYSY

27912
VQDMVNYSY

27913
VQDNINYSH

27914
VQDQINYSY

27915
VQDRINYSY

27916
VQDRVNYSY

27917
VQDTINFSY

27918
VQDTINYSF

27919
VQDTINYSY

27920
VQDTVNYSY

27921
VQENINYSY

27922
VQGRINYSY

27923
VQGTINYSY

27924
VQNKINYSY

27925
VQVKINYSY

27926
VQVRINYSY

27927
VQVTINYSY

27928
VRDKINYSY

27929
VRDWQGYWL

27930
VSRRGKLTW

27931
VTMPDAPCG

27932
VWEANPPFR

27933
VWEDNAPFR

27934
VWEDNPPVR

27935
VWENNPPFR

27936
WEKNSSQMR

27937
WEVMVCILR

27938
WHAMVNSGP

27939
WHDMGNSGP

27940
WMCDFSCIQ

27941
WMCEFSCIK

27942
WRDMVNSGP

27943
YAEVAVQKE

27944
YANSKIMSF

27945
YAQVAVQQE

27946
YGGQVEFGW

27947
YHGVICKEL

27948
YHPHFPTYK

27949
YHPYFPAYK

27950
YHPYFPTHK

27951
YHPYFPTNK

27952
YHPYFPTYQ

27953
YHPYFPTYR

27954
YHPYFSTYK

27955
YHPYVPTYR

27956
YIANPCANC

27957
YIDNPCGNC

27958
YIDNPCTNC

27959
YIDTPCANC

27960
YIHNRCANC

27961
YKLNRDDSW

27962
YKMDCHSCG

27963
YKVNGDDSW

27964
YKVNLDDSW

27965
YKVNQDDSW

27966
YKVNRDASW

27967
YKVNRDGSW

27968
YKVNRDVSW

27969
YKVNRDYSW

27970
YKVNRNDSW

27971
YNGGICKEL

27972
YNGVFCKEL

27973
YNGVICKVL

27974
YNGVICQEL

27975
YNGVICREL

27976
YNGVIGKEL

27977
YNGVLCKEL

27978
YNGVMCKEL

27979
YNGVTCKEL

27980
YNGVVCKEL

27981
YQIRFDRHH

27982
YRVNRDDSW

27983
YSAGPLAYN

27984
YSNSIKMSL

27985
YSNSVQMSF

27986
YSNTIQMSL

27987
YIGVICQEL

27988
YWRMVCVHC

27989
YYMHWINSG

27990
YYMHWVNAG

Example 39
AAV5 Variants with Skeletal Muscle Tissue Tropism as Identified by Machine Learning

This example describes preferred properties of engineered AAV5 variants that display skeletal muscle tissue tropism as identified by machine learning methods. The same bioinformatics pre-process, machine learning methods, and analyses as described in EXAMPLE 19 and FIGS. 19-FIG. 22 were carried out to identify preferred properties of engineered AAV5 variants that display skeletal muscle tissue tropism.

Favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an AAV5 VP capsid polypeptide, which are associated with a higher probability of skeletal muscle tissue tropism are described below. Any of the below machine learning-derived positional amino acid preferences described in TABLE 62 can be present alone or in combination with each other or in combination with any of the amino acid preferences observed and described in EXAMPLE 9, TABLE 17. Listed below in TABLE 62 are a summary of positional features shared between the top important features for skeletal muscle tropism extracted from the ML models.

TABLE 62

Machine Learning-Derived Skeletal Muscle Tissue Tropism Rules

High average flexibility at Xaa1

Xaa1 is selected from G or R

Or

Low average flexibility at Xaa1

Xaa1 is selected from W, M, F, or H

High mol mass at Xaa1

Xaa1 is selected from R, F, or W

Low hydropathy at Xaa2

Xaa2 is selected from K or R

Low mutability at Xaa2

Xaa2 is selected from C, R, or H

High average flexibility at Xaa2

Xaa2 is selected from G or R

High average flexibility at Xaa3

Xaa3 is selected from G or R

High hydrophilicity at Xaa4

Xaa4 is selected from D, E, R, K, or N

Low mutability at Xaa4

Xaa4 is selected from C, R, or H

Low mol mass at Xaa5

Xaa5 is selected from A

Low average flexibility at Xaa5

Xaa5 is selected from A or L

High mutability at Xaa5

Xaa5 is selected from D, A, or E

Low average flexibility at Xaa6

Xaa6 is selected from W, M, or F

Low mutability at Xaa6

Xaa6 is selected from C

High mol mass at Xaa6

Xaa6 is selected from W

Low goldman engelman steitz at Xaa7

Xaa7 is selected from R

High average flexibility at Xaa7

Xaa7 is selected from D, R, P, G, or S

High mutability at Xaa7

Xaa7 is selected from R, H, or N

Low solubility at Xaa7

Xaa7 is selected from R or Q

High hydrophilicity at Xaa8

Xaa8 is selected from D, E, R, K, or N

Low mutability at Xaa9

Xaa9 is selected from Y, F, or L

TABLE 63 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that exhibited skeletal muscle tissue tropism and comport to one or more of the rules provided in TABLE 62. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 29991-SEQ ID NO: 30990, as disclosed in TABLE 63. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 63

Sequences of the 581 to 589 Region in AAV5

VP1 Capsid Polypeptide that Drive

Skeletal Muscle Tissue Tropism

SEQ
581-589

ID NO
sequence

29991
AAGDIGSVM

29992
AAGNIGSVM

29993
AANLLWRLW

29994
AANLLWRLY

29995
ACAWIESFT

29996
ACPDHHRWL

29997
ACPNVWWHM

29998
ACQCKVIGQ

29999
ACQCRVIWQ

30000
ACTWPGVGQ

30001
ACWDTHQSI

30002
ACWDTKQSI

30003
ACWDTLQSI

30004
ACWDTQKSI

30005
ACWDTQQGI

30006
ACWRHVQWN

30007
AFGEFDNHS

30008
AFIHIDREC

30009
AGGHCTYVA

30010
AGWIGACMA

30011
AHWNAIMRW

30012
AIFEPHRWH

30013
AIFEPRRWP

30014
AIIEPRRWH

30015
AIKWRSQDK

30016
AIRVDDSTI

30017
AIVSMDDWH

30018
AKREIENCH

30019
AMHKRWTND

30020
ANNVYCVKK

30021
APANTPWRK

30022
APRPPRSAR

30023
ARDNESNLI

30024
ARSGAQSMM

30025
ARSGFASNT

30026
ARSSFASNN

30027
ARSSFASNS

30028
ARSSFPSNT

30029
ARSSFSSNT

30030
ARSSFTSNT

30031
ARSSVASNT

30032
ASTEVFYMI

30033
ATGIDNNAT

30034
AWSCVHADV

30035
AWWARCKQQ

30036
AWWTPCKQQ

30037
AWWTRCKQL

30038
CADVRENAN

30039
CCEFWLASA

30040
CCELWLASS

30041
CCRHWRPTT

30042
CCRYWRPTP

30043
CGGKIDIGL

30044
CHQENFIFR

30045
CHQENFVFH

30046
CHQENFVFP

30047
CIETGLLNG

30048
CIRQYDAIH

30049
CITYEGWAD

30050
CKVSESAKN

30051
CPPICEGGT

30052
CTETTVRWP

30053
CTKTTVRWP

30054
CTPTTVRWP

30055
CTQSSVRWP

30056
CTQTAVRWP

30057
CTQTTIRWP

30058
CTQTTVRWR

30059
CTQTTVRYP

30060
CTRSTVRWP

30061
CTRTIVRWP

30062
CVGMDSYNG

30063
CYGHLTAMC

30064
DARASPDCL

30065
DARPQWDIP

30066
DARSSPDCI

30067
DCEEFDRIT

30068
DCLVQKAAL

30069
DCNVNPACF

30070
DCWAIHSSW

30071
DCWDTQQSI

30072
DEVHGFLSG

30073
DEVYSFLSG

30074
DFHSYCMSF

30075
DFIHVGDWE

30076
DFTHVGDWD

30077
DGDPNFRIQ

30078
DHGYRNNSM

30079
DILIDHDFF

30080
DKQHDCRAS

30081
DKQQDCRAY

30082
DQQHDCRAY

30083
DRSSLCADE

30084
DRYSLCADA

30085
DSGRIYAWI

30086
DSNVQFCID

30087
DSRSSPDCL

30088
DSTLQFCID

30089
DTRKWMKWH

30090
DVPIDHDFF

30091
DVVGDAACP

30092
DWWTRCKQQ

30093
EARLQWDIP

30094
EARPHWDIP

30095
EARPQWDIL

30096
EARPQWDIS

30097
EARPQWDIT

30098
EARPQWDLP

30099
EARPQWDSP

30100
EARPQWDVP

30101
ECWQHVQWN

30102
ECWRHVKWN

30103
ECWRHVQLN

30104
ECWRHVQWI

30105
ECWRHVQWK

30106
ECWRHVQWS

30107
ECWRQVQWN

30108
ECYRKVVFR

30109
ECYSIYCEQ

30110
EDNIRMMMI

30111
EFAMDWVSD

30112
EFWGSMRNG

30113
EFWSSMRTG

30114
EGRPQWDIP

30115
EILAAWTPP

30116
EKCFQWTSP

30117
EKCFQNTSS

30118
EKCYQWTSA

30119
EKDLSHATF

30120
EKDLSQDTF

30121
EKTSRHWKT

30122
EKVHCMWLH

30123
EQEGDYVDM

30124
EQGAVWTPP

30125
EQGTVWTPP

30126
EQVRAFRQI

30127
ERAGFKMMD

30128
ERAGFQMMH

30129
ERAGVKMMH

30130
ERAGYKMMH

30131
ERASFKMMH

30132
ERDGFKMMH

30133
ERGGEKMMH

30134
ERVGFKMMH

30135
ESRPQWDIP

30136
ETCEMETPG

30137
ETGNDNNAT

30138
ETPINSEKG

30139
EVGTDPSKN

30140
EVRPQWDIP

30141
FAGKIDIGL

30142
FAGPNSYWP

30143
FAMEASPDD

30144
FAVMIHYHN

30145
FAVMVHYPN

30146
FAVMVRYHN

30147
FCAAEWGCL

30148
FCPAEWACL

30149
FCPAEWGCV

30150
FCPAEWGGL

30151
FCPAEWGYL

30152
FCPAKWGCL

30153
FCPEHWFDI

30154
FCPEHWFNM

30155
FCPEHWFNN

30156
FCPEHWFNS

30157
FCPEHWLNI

30158
FCPEHWVNI

30159
FCPQHWFNI

30160
FCSAEWGCL

30161
FCTAEWGCL

30162
FCTEHWFNI

30163
FDLFASPDD

30164
FDMFASPAD

30165
FDMFASPDA

30166
FDMFASPGD

30167
FDVFASPDD

30168
FFELSKTVF

30169
FFELSQTEF

30170
FFRDCPYES

30171
FERDYPYEY

30172
EGGKEDIGL

30173
FGGKIAIGL

30174
FGGKIDIGV

30175
FGGKIDIVL

30176
FGGKIDVGL

30177
FGGKIHIGL

30178
FGGKINIGL

30179
FGGKLDIGL

30180
FGGTIDIGL

30181
FGLVHQKNA

30182
FGRKIDIGL

30183
EGVVHQKDA

30184
EHHLTEYHG

30185
FHHQPFYHG

30186
FHHQTEYRG

30187
FHHQTGYHG

30188
FHHQTQYHG

30189
FHKWGINLG

30190
FHKWWIHLG

30191
FHKWWINLA

30192
FHKWWINLS

30193
FHKWWINQG

30194
FHKWWISLG

30195
FHKWWITLG

30196
FHKWWLNLG

30197
FHRQTEYHG

30198
FLKWWINLG

30199
FMGATIPED

30200
FMGVTIPAD

30201
FMGVTIPEG

30202
FMGVTIPKD

30203
FMGVTIPVD

30204
FMKCENFNS

30205
FMKCENVNT

30206
FMKCESVNS

30207
FMKGENVNS

30208
FMTCENVNS

30209
FMWHANSCG

30210
FMWHANSGA

30211
FMWYANSCA

30212
FNINAWDWD

30213
ENVTAQDWD

30214
EPHHLWVGG

30215
FPYHLWIGG

30216
FPYHLWVAG

30217
FPYHVWVGG

30218
FPYRLWVGG

30219
FRECLQIVN

30220
FRECLQLVT

30221
FRECMQIVT

30222
FREGLQIVT

30223
FRGCLQIVT

30224
FRGIETEQG

30225
FRGIETGEG

30226
FRGIETGQV

30227
FRGIETVQG

30228
FRGTETGQG

30229
FRGVETGQG

30230
FRKCLQIVT

30231
FRRWQSNTT

30232
FTETTVRWP

30233
FTQPTVRWP

30234
FVLVHQKDA

30235
FWKFALLSF

30236
FWKLALHSF

30237
FWPQIQYRM

30233
FYGKSAPVG

30239
FYGKTALVG

30240
EYGKTAPVA

30241
FYGPVTKSC

30242
FYGQLTKSC

30243
FYGQTAPVG

30244
FYGQVTESC

30245
FYGQVTMSC

30246
FYGQVTQSC

30247
FYGQVTTSC

30248
FYKWWINLG

30249
GARPQWDIP

30250
GARSSPDCL

30251
GASWHDREH

30252
GATEVFYMI

30253
GCDMDAYSN

30254
GCDVDAYSN

30255
GDCGASNCY

30256
GFHNTELPI

30257
GGAKALRFN

30258
GGARALRFI

30259
GGARALRFS

30260
GGGCEDWPM

30261
GGPEIKHYI

30262
GHWNAIMRW

30263
GIDWEDVWE

30264
GIKWRSQDK

30265
GIRDDDSTI

30266
GIRGDDSTI

30267
GIRVDDNTI

30268
GIRVDDSTK

30269
GIRVDDSTV

30270
GMDEVEREG

30271
GMEEGEREG

30272
GMEELEREG

30273
GNIYHFNNV

30274
GPDGAKLNK

30275
GPDGAKRHK

30276
GPDGARRNK

30277
GPDKCTVNL

30278
GPGLKAWPC

30279
GPLATTDPV

30280
GPTEVEYMI

30281
GRDWAFCDY

30282
GRHWACCDY

30283
GRHWAFCDD

30284
GRHWAFCDF

30285
GRHWAFCDH

30286
GRHWAFCDN

30287
GRHWAFCGY

30288
GRHWAFCHY

30289
CRHWAFFDY

30290
GRHWAFGDY

30291
GRHWAVCDY

30292
GRHWDFCDY

30293
GRHWSFCDY

30294
GRHWTFCDY

30295
GRPWAFCDY

30296
GRQWAVCDY

30297
GRRWAFCDY

30298
GRVSGHVHW

30299
GRYSLCADE

30300
GRYWAFCDY

30301
GSAEVFYMI

30302
GSNEVFYMI

30303
GSPEVFYMI

30304
GSTELFYMI

30305
GSTEVFCMI

30306
GSTEVFYMV

30307
GSTEVFYVI

30308
GSTKVFYMI

30309
GVEPMSRAA

30310
GVRDAVHWC

30311
GWSARSVNM

30312
HALSVGEWD

30313
HARGEWDFH

30314
HCAHYWMQD

30315
HCIRHFCEV

30316
HCLLQKAAL

30317
HCSCHFCEV

30318
HCSHHFCEV

30319
HCSRHFCEA

30320
HCSRHFCKV

30321
HCSRHFCQV

30322
HCSRHFGEV

30323
HCSRHYCEV

30324
HCTRHFCEV

30325
HCTRYWMQD

30326
HCYRKVVFR

30327
HDIDGWQMA

30328
HDIWQWSIK

30329
HDMLQWSEK

30330
HDMWQNTEK

30331
HDQVKCLRK

30332
HDTWQWSEK

30333
HEIDGWQTA

30334
HEIDVWQMA

30335
HEVDGWQMA

30336
HEVHSFLSG

30337
HKRNLYALQ

30338
HLHFHWGLW

30339
HLWRTFEAQ

30340
HLYSDYWFG

30341
HLYSEYWFD

30342
HRDAHLLME

30343
HRYSLCADE

30344
HSHFHWGLW

30345
FETCHEVDG

30346
HTCSELVEG

30347
HTELSHDPW

30348
HTEWSHDQW

30349
HTLLLSREP

30350
IACMKCPNI

30351
IANNDLRFG

30352
ICEMGPYHG

30353
ICMSQEKRC

30354
IDVGAYVWS

30355
IFEQCQRMR

30356
IFRDCPYEY

30357
IGIVFQRDF

30358
IGLVFQRDF

30359
IGVVFPRDF

30360
IGVVFQRDC

30361
IGVVFQRDE

30362
IGVVFQRDV

30363
IGVVFQRDY

30364
IGVVFQRVF

30365
IHVEYFDAW

30366
IHWNAEMRW

30367
IIKWRSQDK

30368
IENKDHDKF

30369
IENKHHDRF

30370
EINKRHDKF

30371
INQMARHPK

30372
INVEYFDTW

30373
INVEYFEAW

30374
INVEYFVAW

30375
IRTSFHVNA

30376
ISNNDLRFV

30377
ISNNDLRVG

30373
ISNNDVRFG

30379
ISTNDLRFG

30380
ITCKFTQNG

30381
IICKMDFWQ

30382
ITTGSRPTK

30383
IVFPSLARG

30334
IWPQSQYRM

30385
KAHKTVQDK

30386
KAMSVPWGK

30387
KANGSMDHH

30388
KANYACDWD

30389
KCFSMDFSG

30390
KCMPGCNDW

30391
KCMPNVGWR

30392
KCNVNPACF

30393
KCQWVGREN

30394
KCVSMAFSG

30395
KCVSMDFAG

30396
KCVSMDFTG

30397
KCVSMGFSG

30398
KCVSMVFSG

30399
KCVSMYFSG

30400
KCVSVDFSG

30401
KCWRHVQWN

30402
KDHNRWNRV

30403
KDPNRWNRL

30404
KDVGDDAHG

30405
KDYNRWNRL

30406
KEHNRWNRL

30407
KFAKDWVSD

30408
KFAMDWDSD

30409
KFAMDWVSV

30410
KFVSMDFSG

30411
KHEADYVDM

30412
KHEPRQSPR

30413
KHKARQSPR

30414
KHKLRQSPR

30415
KHMPRQSPR

30416
KHSELWREN

30417
KKCFQWTSA

30418
KKVRAFRQI

30419
KMKDVEENG

30420
KMKPVQRVG

30421
KMMSFSEVD

30422
KQEGDDVDM

30423
KQEGDSVDM

30424
KQVRAFREE

30425
KQVRAFRPE

30426
KQVRAFRQL

30427
KQVRAFRQM

30428
KQVRAFRQS

30429
KQVRAFRQV

30430
KQVRAFRRE

30431
KQVRDFRQI

30432
KQVRPFRQE

30433
KQVRSFRQE

30434
KQVRVFRQI

30435
KRVHFEGGD

30436
KSMTELDQP

30437
KTCELFTPG

30438
KTCEMFTPA

30439
KTCEMFTPS

30440
KTCERFTPG

30441
KTCGMFTPG

30442
KTELAMVID

30443
KTELDRDSM

30444
KTEWAMAID

30445
KTEWAMVLD

30446
KTEWAVWVD

30447
KTESGFQDK

30448
KTKWAMVID

30449
KTMSGPWGK

30450
KVNYACDWH

30451
KVTYACDWD

30452
KWDSIFVEH

30453
KWDSIFVQR

30454
KYAANWLKD

30455
KYADNWLKA

30456
KYADNWVKD

30457
KYADYWLKD

30458
KYSDNWLKD

30459
KYSEVWREN

30460
KYSKLWREN

30461
KYWMRHvym

30462
LATSMCYQS

30463
LCHFLRTCK

30464
LDKIEHDWN

30465
LFCEVQSRE

30466
LIDREWFNW

30467
LSGPEHYEP

30468
LSGSIHYES

30469
LSVKMDRAK

30470
LSVQMDRSK

30471
LSVRMDRAK

30472
LTGQEAGKG

30473
LVCIRADTD

30474
LVGKYWPRD

30475
LVGPYWPRD

30476
MADHCCANS

30477
MADHCCTDS

30478
MAFSSMHQG

30479
MASLRGSDE

30480
MCYQNIAFQ

30481
MFGECYNNG

30482
MFKYYPYCD

30483
MFLECYNNG

30484
MFQSYPYCD

30485
MFQYYQYCD

30486
MFRECYDNG

30487
MFRECYKNG

30488
MFRECYNSG

30489
MFREWYNNG

30490
MFRKCYNNG

30491
MGDGLWVKV

30492
MHFKYPQRG

30493
MNINKIRNS

30494
MPVKKEYGG

30495
MRHWAFCDY

30496
MRIHFEGGD

30497
MRVHFEGGG

30498
MRVHFQGGD

30499
MRVRFEGGD

30500
MSSLKGSDE

30501
MSWLGAAGE

30502
MTLLMGHES

30503
MVCIHADTD

30504
MWLAKARQG

30505
MWVGKARQG

30506
MYKYGQDWY

30507
NAINVGEWD

30508
NAISVGKWD

30509
NALSVGEWD

30510
NARSSPDCL

30511
NCSRHFCEV

30512
NCWAIHGSW

30513
NCWAILSSW

30514
NDLGDDAHG

30515
NDVADDAHG

30516
NDVGDDDHG

30517
NDVGDEAHG

30518
NDVGGDAHG

30519
NDVGNDAHG

30520
NEIDGWQMA

30521
NEIMMWRAN

30522
NETMMWRTN

30523
NFLDYHDMV

30524
NFMDYNDMV

30525
NFMEYHDMV

30526
NHQADEHDA

30527
NIRPSDASG

30528
NIRPSDASS

30529
NIRPSDTSA

30530
NIRQSDASA

30531
NIRRSDASA

30532
NKRNLYAMQ

30533
NKRNLYAVQ

30534
NMGIASPWH

30535
NMVDENRFW

30536
NMVHEHRFW

30537
NMVHESRFW

30538
NMVHQNRFW

30539
NMVRENRFW

30540
NSISVGEWD

30541
NTSCWKLEN

30542
NTVIRYRHS

30543
NVNEDVQWQ

30544
NWVNGFVDH

30545
NWWAIHSSW

30546
NYVLFVDHE

30547
PAWDLVEML

30548
PAWDLVGMF

30549
PAWDLVGMH

30550
PAWDLVGMI

30551
PAWDPVGML

30552
PAWELVGMV

30553
PAWHLVGML

30554
PCDFDNQPH

30555
PCHFDNQPH

30556
PCLDHHRWL

30557
PCNFDSQPH

30558
PCNVDNQPH

30559
PCPDHHRLL

30560
PCPDHHRRL

30561
PCPDHHRWV

30562
PCPDHNRWL

30563
PCPDNHRWL

30564
PCPDPHRWL

30565
PCPEHHRWL

30566
PCPNVWWHM

30567
PCQCKVIWQ

30568
PCTDHHRWL

30569
PCTFDNQPH

30570
PCYRKVVFR

30571
PDGERCERF

30572
PDWRKEKKF

30573
PFMKDGECE

30574
PFMKDVKCE

30575
PGDPQEDTV

30576
PGIDLWHMA

30577
PGIEWWHMA

30578
PGWEGACMA

30579
PHFDKCQHF

30580
PEDWEDVWE

30581
PKAMAKICE

30582
PKGMAKICA

30583
PKGMAKIFE

30584
PKGMAKIWE

30585
PKGMARICE

30586
PKGMSKICE

30587
PKGMVKICE

30588
PKGNRTNQI

30589
PKVMAKICE

30590
PMHQRWTND

30591
PMMKKLCCG

30592
PNGEKCERF

30593
PQHISADPT

30594
PQHISSDPT

30595
PQLESPDPT

30596
PQRESPDPT

30597
PRDDESNLI

30598
PRDKIWFMN

30599
PRDNVSNLI

30600
PRFCISLKC

30601
PRGNESNLI

30602
PRMSYEEKG

30603
PRTSYEFKG

30604
PSKPFSKEE

30605
PSQPFSEEE

30606
PTWDLVGML

30607
PWCECHRLF

30608
PWSWCEFNK

30609
PWSWCEYNQ

30610
PWWTRCKQQ

30611
PYGEKCERF

30612
PYTHWKLNN

30613
QARIEDMAK

30614
QCEWQKMEH

30615
QCFLCGSAT

30616
QCHFLRNCK

30617
QCMPGCNDL

30618
QCPVECDHV

30619
QCVSMDFSG

30620
QCYREVVFR

30621
QFAMDWVSD

30622
QIRIEDMAK

30623
QKGNRTDQI

30624
QKGNRTTQI

30625
QKRGIEAVD

30626
QKRIEDMAK

30627
QLNICQATF

30628
QMHNTFCGW

30629
QMHSTFCSW

30630
QRDKVWFMN

30631
QRDRIWFMN

30632
QSVDVGLMH

30633
QTGDWWQYG

30634
QTGNWWEYG

30635
QTGNWWKYG

30636
QTGNWWQYA

30637
QTGTWWQYG

30638
QTISGFKDK

30639
QTISGFQYK

30640
QTIVAWHHS

30641
QTLSGFQDK

30642
QTQIEDMAK

30643
QTRIDDMAK

30644
QTRIEDMTK

30645
QTRIQDMAK

30646
QTRRLEPYH

30647
QTRVEDMAK

30648
QVCIHADTD

30649
RADVDMTSI

30650
RCEIWLASS

30651
RCISNEGMQ

30652
RCISNEWME

30653
RCISNEWMP

30654
RDHNRWNRL

30655
REEIARPWY

30656
REKIARPWY

30657
REQIACPWY

30658
REQIARPGY

30659
REQISRPWY

30660
REQLARPWY

30661
RETLEQVAY

30662
RFNLPNFYD

30663
RFNWPIFYD

30664
RFNWPNVYD

30665
RFPFRAQVW

30666
RFRFLAQVW

30667
RFRFRAKVW

30668
RFRFRAQGW

30669
RFRFRAQIW

30670
RFRFRTQVW

30671
RFRFRVQVW

30672
RFRIRAQVW

30673
RFRVRAQVW

30674
RFTWPNFYD

30675
RHDWQHRTV

30676
RHELTMSHH

30677
RHELTMSQY

30678
RHQPLHYFH

30679
RHVPLHYFH

30680
RIDFNVDCY

30681
RIDINVDCH

30682
RIDINVDWY

30683
RIDINVNCY

30684
RIDINVYCY

30685
RIDISVDCY

30686
RIDLNVDCY

30687
RIDVNVDCY

30688
RIGENVDCY

30689
RIMFFDKFT

30690
RIMYFDEFT

30691
RIMYFDKCT

30692
RIMYFDRFT

30693
RINWPNFYD

30694
RITYFDKFT

30695
RKDTPKAGH

30696
RLARDNNES

30697
RLARENNEA

30698
RLELDNNEA

30699
RLEQYNNEA

30700
RLERANNEA

30701
RLERDDNEA

30702
RLERDHNEA

30703
RLERDINEA

30704
RLERDNIEA

30705
RLERDNNAA

30706
RLERDNNAV

30707
RLERDNNED

30708
RLERDNNEP

30709
RLERDNNES

30710
RLERDNNET

30711
RLERDNTEA

30712
RLERDTNEA

30713
RLERHNNEA

30714
RLERNNNEA

30715
RLGRDNNEA

30716
RLKRDNNEA

30717
RLKRVNNEA

30718
RLQRDNNEA

30719
RLYSDYWFD

30720
RMGYEVDWQ

30721
RMMYFDKFT

30722
RNLDYNTAV

30723
RNLNYNTGV

30724
RNPNYNTAV

30725
RNVNYNTAV

30726
RPERDNNEA

30727
RQQIARPWY

30728
RRCCISLKC

30729
RRDSDSVCK

30730
RRFCVSLKC

30731
RTECVPDTK

30732
RTLDSEYFF

30733
RTLDSEYWF

30734
RTQCAPDTK

30735
RTQCVPATK

30736
RTQCVPDAK

30737
RTQWVPDTK

30738
RTVDSEYLF

30739
RVCPFCCCF

30740
RVCTFCGCF

30741
RVDINVDCY

30742
RVDNDMADG

30743
RVERDNNEA

30744
RVKRDNNEA

30745
RWEKMAYSW

30746
RYCDAWPDV

30747
SAWDLVGML

30748
SCENWGRQH

30749
SCETGGRQH

30750
SCETWGRHH

30751
SCETWGRPH

30752
SCETWSRQH

30753
SCETWVRQH

30754
SCEWSKQCK

30755
SCMDGTTGV

30756
SCMHGTIGV

30757
SCMNGATGV

30758
SCMVGPTGV

30759
SCMNGTAGV

30760
SCMNGTTVV

30761
SCMNVTTGV

30762
SCPWPTGEE

30763
SCQTWGRQH

30764
SCRHWRPTP

30765
SCVTWGRQH

30766
SCWDTQQSI

30767
SDKWYRVSF

30768
SFQHAQWMG

30769
SGWIAACMA

30770
SGWIRACMA

30771
SGWNGACMA

30772
SHFESHQWC

30773
SHGESHQWC

30774
SIMMPDYSC

30775
SITDANLGF

30776
SKIYAVDQS

30777
SKRDIENCH

30778
SKRDVQSMM

30779
SKREIENCQ

30780
SKREIETCH

30781
SKRELENCH

30782
SKRKIENCH

30783
SKWACHPKM

30784
SKWNCHPKM

30785
SLNELKRDQ

30786
SMAKHFTSS

30787
SMHKIWYRA

30788
SQEFVCAQE

30789
SRDNESNLI

30790
SREQVNCDA

30791
SRISFVCSD

30792
SRKQVNCDA

30793
SRLQVNCDA

30794
SRLSFVCSD

30795
SRQPVNCDA

30796
SRQQANCDA

30797
SRQQVHCDA

30798
SRQQVNCDV

30799
SRQQVNCEA

30800
SRQQVNGDA

30801
SRQRVNCDA

30802
SRSMSSSME

30803
SRSMYSGME

30804
SRVAFVCSD

30805
SSAIQCDAA

30806
SSGIECDAA

30807
SSGIQCDAT

30808
SSGIQCEAA

30809
SSGLQCDAA

30810
SSILTWTWI

30811
SSRIQCDAA

30812
STDNHWGHG

30813
STQATVRWP

30814
STQEDFDMF

30815
STQTSVRWP

30816
SVQWYRVSF

30817
SWCDCHRLF

30818
SWCECHRLL

30819
SWCECHRLV

30820
SWCECHRMF

30821
SWCECHRSF

30822
SWCECHRVF

30823
SWCGCHRLF

30824
SWCVCHRLF

30825
SWNELKRNQ

30826
SWSWCEFNQ

30827
SYTHLKLNN

30828
SYTPWKLNN

30829
TAWDLVGML

30830
TAWRKIKKE

30831
TCKWVGREN

30832
TCMFRVYDN

30833
TCPNVWWHK

30834
TCPNVWWHL

30835
TCPNVWWRM

30836
TCQWFGREN

30837
TDWPKIKKE

30838
TDWRKFKKF

30839
TDWRKLKKF

30840
TDWRKMKKF

30841
TDWRKVKKE

30842
TDWRRIKKF

30843
TEVKGWESD

30844
TFMDYHDMV

30845
TGEHNTFKD

30846
THQGDEHDA

30847
TISDIPGEP

30848
TISDIPGKP

30849
TLDECCRRG

30850
TLPCYQRYT

30851
TMPCYQRYT

30852
TNVEYFDAW

30853
TNWREMNAR

30854
TNWRKIKKF

30855
TPRPPRSGR

30856
TPRPSRSAR

30857
TPRTPRSAR

30858
TRSGAQAMM

30859
TSGTCMPIK

30860
TWMVVHRNN

30861
TYQWVGREN

30862
VAEMHWMVN

30863
VAIGFHWHA

30864
VAWAVSYVC

30865
VAWGVSYVG

30866
VCDIDAYSN

30867
VCEMGPYDG

30868
VCEMGPYNG

30869
VCEMGPYRG

30870
VCEMGQYHG

30871
VCERGPYHG

30872
VCQMGPYHG

30873
VCVAIMKDQ

30874
VCVAIMKGK

30875
VCVAIMQDK

30876
VCVMGPYHG

30877
VCWDTQQSI

30878
VCWMAGAQM

30879
VDKAWWNLI

30880
VDKVWWNLV

30881
VDRSYGPSG

30882
VEGDFDNHS

30883
VFVACHVNG

30884
VGDLNFRIQ

30885
VHLNADTKF

30886
VHWDAIMRW

30887
VHWNAIMRG

30888
VHWNAIMRL

30889
VHWNALMRW

30890
VHWNASMRW

30891
VHWQQCGHQ

30892
VIDRIWENW

30893
VIKGRSQDK

30894
VIKLRSQDK

30895
VIKWGSQDK

30896
VIKWRGQDK

30897
VIKWRSLDK

30898
VIKWRSRDK

30899
VIQWRSQDK

30900
VIRWRSQDK

30901
VLPCQRKRV

30902
VLWNAIMRW

30903
VMQNVWSCW

30904
VMSCHSRDH

30905
VPKNVNRDM

30906
VPKTANRDM

30907
VPKTVDRDM

30908
VPTIVNRDM

30909
VQDKCTVNL

30910
VRHWAFCDY

30911
VSVQMDRAK

30912
VTGKMDFWQ

30913
VTKNVWCSW

30914
VTKWRSQDQ

30915
VTMIMMDCH

30916
VTRDLKKEQ

30917
VVDMCRKGK

30918
VVRDLPNQY

30919
VWFGKCVNC

30920
VWLKSLDRM

30921
VWPKGMREM

30922
VWSCVHTDV

30923
WDVVAADGP

30924
WDVVATDVP

30925
WERAWLFGG

30926
WERFWHEYW

30927
WHGRLMITF

30928
WHQENFVFR

30929
WKDPFNRKI

30930
WLANGWFRE

30931
WLANSWFRG

30932
WLDNSWFRE

30933
WLPNSWFRE

30934
WLSNSWFRE

30935
WMCRNHHFD

30936
WMCRNHPFN

30937
WMCRNHPVD

30938
WMCRSHPFD

30939
WRCLCARID

30940
WRCMCARIA

30941
WRCMCARLD

30942
WRCMCARMD

30943
WRCMCARTD

30944
WRCMCARVD

30945
WRCMCSRID

30946
WRCMCTRID

30947
WRCWYAPLT

30948
WRELSNTGY

30949
WREWSHTGY

30950
WREWSNAGY

30951
WREWSNTAY

30952
WREWSNTCY

30953
WREWSNTVY

30954
WRGWSNTGY

30955
WRQWSNTGY

30956
WRRFASGWE

30957
WRRFASWLE

30958
WRRFSSWWE

30959
WRRFVSWWE

30960
WRTADEGDA

30961
WRTADESDT

30962
WRTAHESDA

30963
WRTGDESDA

30964
WTETTVRWP

30965
WTQTTVRWP

30966
WVANSWFRE

30967
WVCRNHPFD

30968
WYGHLNAMC

30969
WYSWRVRNR

30970
YARGIWDIH

30971
YARGIWDVH

30972
YARGTWDFH

30973
YCSRHFCEV

30974
YDRQWDFIA

30975
YEVHSFLSG

30976
YHDLCFAWY

30977
YHDLFFDWY

30978
YHDLFFGWY

30979
YKREIENCH

30980
YMGIASPWH

30981
YMWWEEGTL

30982
YMWWQAGTL

30983
YRQQVNCDA

30984
YRRLQSNTI

30985
YRRWESNTT

30986
YRRWQSNTI

30987
YRRWQSNTS

30988
YVKFNVNIN

30989
YVKFSVDIN

30990
YVKISVNIN

Example 40
AAV5 Variants with Skin Tissue Tropism as Identified by Machine Learning

This example describes preferred properties of engineered AAV5 variants that display skin tissue tropism as identified by machine learning methods. The same bioinformatics pre-process, machine learning methods, and analyses as described in EXAMPLE 19 and FIGS. 19-FIG. 22 were carried out to identify preferred properties of engineered AAV5 variants that display skin tissue tropism.

Favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an AAV5 VP capsid polypeptide, which are associated with a higher probability of skin tissue tropism are described below. Any of the below machine learning-derived positional amino acid preferences described in TABLE 64 can be present alone or in combination with each other or in combination with any of the amino acid preferences observed and described in EXAMPLE 17, TABLE 33. Listed below in TABLE 64 are a summary of positional features shared between the top important features for skin tropism extracted from the ML models.

TABLE 64

Machine Learning-Derived Skin Tissue Tropism Rules

Low surface accessibility at Xaa1

Xaa1 is selected from C

Low volume at Xaa1

Xaa1 is selected from C

Low mutability at Xaa1

Xaa1 is selected from C

High surface accessibility at Xaa2

Xaa2 is selected from R or K

High average flexibility at Xaa2

Xaa2 is selected from K, I, or N

Low mutability at Xaa2

Xaa2 is selected from P or K

High hydropathy at Xaa3

Xaa3 is selected from I or V

Low mutability at Xaa4

Xaa4 is selected from L, F, or Y

Low average flexibility at Xaa4

Xaa4 is selected from W, H, F, or M

High average flexibility at Xaa5

Xaa5 is selected from G, R, K, I, or N

High average flexibility at Xaa6

Xaa6 is selected from G, R, K, I, or N

High surface accessibility at Xaa8

Xaa8 is selected from M, G, or F

Low average flexibility at Xaa8

Xaa8 is selected from H, F, M, or W

Low mutability at Xaa8

Xaa8 is selected from L, F, or Y

High average flexibility at Xaa9

Xaa9 is selected from D, E, R, K, P, or G

High mutability at Xaa9

Xaa9 is selected from D, E, R, V, A, or H

TABLE 65 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid poly peptides that exhibited skin tissue tropism and comport to one or more of the rules provided in TABLE 64. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 32991-SEQ ID NO: 33990, as disclosed in TABLE 65. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 65

Sequences of the 581 to 589 Region in AAV5 VP1

Capsid Polypeptide that Drive Skin

Tissue Tropism

SEQ

ID
581-589

NO
Sequence

32991
AADAETPWF

32992
AADPRADCP

32993
AADPRGDCP

32994
AADPRIDCP

32995
AADPRVDCA

32996
AADPRVDCL

32997
AADPRVDCR

32998
AADPRVECP

32999
AADPTKSWC

33000
AADSETPWY

33001
AASPYITYR

33002
AEHCYRHKH

33003
AEPFPCDWS

33004
AFFYMDKLS

33005
AIVCRVNYP

33006
AKRDMDPAQ

33007
AKRDMDPAS

33008
AKRNMDPAP

33009
ANETHDNFH

33010
ANVGCIMIL

33011
ASARYIDFT

33012
ASWCSDERC

33013
ATAGCIMIL

33014
ATDPRVDCP

33015
ATVGFIMIL

33016
AVATRAMYY

33017
AVDPRVDCP

33018
CAFQCYRSP

33019
CAFVQQNFQ

33020
CAGMQATGD

33021
CAHHLNQTI

33022
CAIFNTVDQ

33023
CAIHTQSHS

33024
CAIMCSAGP

33025
CAIMFSAGQ

33026
CAIMFSAGT

33027
CAIMLKHMT

33028
CAIMLQHMA

33029
CAIMLQHMI

33030
CALHTRSHS

33031
CALMFSAGP

33032
CALPVRVAM

33033
CALPYAGSR

33034
CALSMYSHE

33035
CANHLNKTI

33036
CANHLNQTM

33037
CANHLNRTI

33038
CANHLSQTI

33039
CANHVNQTI

33040
CAPISAAPV

33041
CARLIDWCC

33042
CASMFSAGP

33043
CATHLNQTI

33044
CATVKMWHT

33045
CAVMFSAGP

33046
CAVMKFSEH

33047
CAVYFAKCA

33048
CAVYFGKCV

33049
CAVYFGRCA

33050
CAVYYGKCA

33051
CAYFKRHPD

33052
CAYFSMMDR

33053
CAYHLNQTI

33054
CCLYYVNVE

33055
CCTIKQMDI

33056
CCVWEFNYP

33057
CDHQFIDQA

33058
CEHFKRHPD

33059
CERLTQNVD

33060
CEYFKRDPD

33061
CEYFKRHPA

33062
CEYFKRHPG

33063
CEYFKRHTD

33064
CEYFKRRPD

33065
CEYFKRYPD

33066
CEYIKRHPD

33067
CFHPYHQAH

33068
CFHQYNQAH

33069
CFLWEFNYP

33070
CFMALSFHW

33071
CFMGLSTHL

33072
CFMVLSFHW

33073
CFPVYKDWS

33074
CFVGLSFHW

33075
CFVLEFNYP

33076
CFVWAFNYP

33077
CFVWEFDYP

33078
CFVWEFNDP

33079
CFVWEFNHP

33080
CFVWEFNSP

33081
CFVWEFSYP

33082
CFVWGFNYP

33083
CFVWGINYP

33084
CFVWQFNYP

33085
CFVWVFNYP

33086
CFYQYHQAH

33087
CHAPSIMGP

33088
CHCHIRHTG

33089
CHFMFNAGR

33090
CHFMIHAGR

33091
CHFMINAAR

33092
CHFMINACR

33093
CHFMINAGH

33094
CHFMINDGR

33095
CHFMINSGR

33096
CHFMINTGR

33097
CHFMINVGR

33098
CHFMLNAGR

33099
CHFMSNAGR

33100
CHFMTNAGR

33101
CHFMVNAGR

33102
CHFVINAGR

33103
CHHATNTSQ

33104
CHHFATASE

33105
CHHFATETE

33106
CHHFATQSE

33107
CHIMINAGR

33108
CHPVYKDWS

33109
CHSMINAGR

33110
CHTIKQMDI

33111
CHVMINAGR

33112
CHYMINAGR

33113
CIKACNDHE

33114
CIKACYDRE

33115
CIKACYDYE

33116
CIKACYHHE

33117
CIKACYNHE

33118
CIKAYNVMM

33119
CIKAYYDHE

33120
CIKGCYDHE

33121
C1KPCYDHE

33122
CIKSCYDHE

33123
CIKSYNVMV

33124
CIKTYNVMM

33125
CIKYYNVMM

33126
CILHKHNFP

33127
CILHKHSSP

33128
CILHKHSVP

33129
CILHKNSFP

33130
CILQKHSFP

33131
CILYKHSFP

33132
CIMHKHSFP

33133
CIQACYDHE

33134
CIQFTKHQE

33135
CIQLSKMGP

33136
CIQLTKMGP

33137
CIRSYNVMM

33138
CITACYDHE

33139
CITMVRSQT

33140
CKAHCPGQY

33141
CKCHFDQVE

33142
CKDFHWMND

33143
CKFDITKMA

33144
CKFDMASFF

33145
CKFDMPCFF

33146
CKFDMPGFF

33147
CKFDMPSCF

33148
CKFDMPSFI

33149
CKFDMPSFV

33150
CKFDMPSFY

33151
CKFDMPSVF

33152
CKFDMPTFF

33153
CKFDMQGFF

33154
CKFDMQSFF

33155
CKFDMSSFF

33156
CKFDMTSFF

33157
CKFHMPSFF

33158
CKFNMPSFF

33159
CKFYIPSFF

33160
CKGHFDQVE

33161
CKHFHWMND

33162
CKIDMPSFF

33163
CKIHFDQVE

33164
CKNFHLMND

33165
CKNFHRMND

33166
CKNFHWMDD

33167
CKNFHMVHD

33168
CKNFHWMNE

33169
CKNFHWMNH

33170
CKNFHWMSD

33171
CKNFHWRND

33172
CKNFLWMND

33173
CKNFPWMND

33174
CKNFYWMND

33175
CKNHFDQVE

33176
CKNIHWMND

33177
CKPQINDAY

33178
CKQQINDAC

33179
CKQQINDAD

33180
CKQQINDAF

33181
CKQQINDAH

33182
CKQQINDAN

33183
CKGQINDAS

33184
CKQQINDDY

33185
CKQQINDGY

33186
CKOQINDSY

33187
CKQQINHAY

33188
CKQQLNDAY

33189
CKRQINDAY

33190
CKSFHWMND

33191
CKSHFAQVE

33192
CKSHFDKVE

33193
CKSHFNQVE

33194
CKSHIDQVE

33195
CKSHVDQVE

33196
CKTFHWMND

33197
CKTHFDQVE

33198
CKTQCPGQY

33199
CKVDMPSFF

33200
CKYDFTKMA

33201
CKYDIPKMA

33202
CKYDISKMA

33203
CKYDITKMG

33204
CKYDITMIS

33205
CKYDITKMT

33206
CKYDITKMV

33207
CKYDITMMA

33208
CKYDITQMA

33209
CKYDLTKMA

33210
CKYDMPSFF

33211
CKYDNTKMA

33212
CKYDTTKMA

33213
CKYDVTKMA

33214
CKYFHWMND

33215
CLELRTQYA

33216
CLELVKAYE

33217
CLELVKDYE

33218
CMFHNHKGF

33219
CMFQCYRSP

33220
CMFVQQNFQ

33221
CMHLSCDWA

33222
CMHLVNMNS

33223
CMKACYDHE

33224
CMKFIRVGS

33225
CMPFIRVGS

33226
CMQFIRAGS

33227
CMQFIRIGS

33228
CMQFIRLGS

33229
CMQFIRVAS

33230
CMQFIRVGG

33231
CMQFIRVGN

33232
CMQFLRVGS

33233
CMQFNRVGS

33234
CMQFSRVGS

33235
CMQFTRVGS

33236
CMQFVRVGS

33237
CMQVIRVGS

33238
CMQYIRVGS

33239
CMRFIRVGS

33240
CMSWLTNQE

33241
CMVHNHKGS

33242
CMVHNHQGF

33243
CMYWLNNQE

33244
CMYWLTNKE

33245
CNCNLPYAI

33246
CNCPIRHTG

33247
CNCQIRHTG

33248
CNFMINAGR

33249
CNFQTNVHM

33250
CNFQTSVHL

33251
CNHKFIDQA

33252
CNHKVNSWI

33253
CNHPFIDQA

33254
CNHQFIDQG

33255
CNHQFIDQS

33256
CNHQFIDQT

33257
CNHQFIDQV

33258
CNHQFLDQA

33259
CNPQFIDQA

33260
CNYQTNVHL

33261
CPKPFLIKE

33262
CPKPVLIQE

33263
CQELRTQHA

33264
CQELRTQYT

33265
CQEVHAHMS

33266
CQEVYAHMT

33267
CQKWRSIWK

33268
CQQVYAHMS

33269
CQSHFDQVE

33270
CRAAHPVCA

33271
CRAAHSICA

33272
CRAAYSVCA

33273
CRATHSVCA

33274
CRAVHSVCA

33275
CRFMINAGR

33276
CRTAHSVCA

33277
CRVAHSVCA

33278
CSAMVRSQT

33279
CSGCHMKMP

33280
CSGCYMKMS

33281
CSHPVHDNE

33282
CHIAKHSFP

33283
CSLPYAWSR

33284
CSMIQNSFP

33285
CSTMVRSQA

33286
CSVCYMKMP

33287
CSVWEFNYP

33288
CTAMLQRKG

33289
CTAVKMWHT

33290
CTCGYNRGD

33291
CTCHIRHTG

33292
CTCNLPSAI

33293
CTCNVPYAI

33294
CTCWYNRGE

33295
CTEVLQRKG

33296
CTGCYMKMP

33297
CTGIQATGD

33298
CTGLLMHKE

33299
CTGQLMHEE

33300
CTGQLMHQE

33301
CTGQLMRKE

33302
CTGWYNRGD

33303
CTHAVHDNE

33304
CTHGFLTAP

33305
CTHGFLTSP

33306
CTHGFLTVP

33307
CTHGVLTGP

33308
CTHKVNTWI

33309
CTHPVHDHE

33310
CTHPVNDNE

33311
CTHQFIDQA

33312
CTHQVHDNE

33313
CTHTVHDNE

33314
CTHVFLTGP

33315
CTIFHTVDQ

33316
CTIFNNVDQ

33317
CTIFNTVDK

33318
CTIFTTVDQ

33319
CTIMFSAGP

33320
CTIRANLHF

33321
CTIYRNLLS

33322
CTKACYDHE

33323
CTKMLQRKG

33324
CTKSYNVMM

33325
CTLKVNSWI

33326
CTLMHIHTR

33327
CTLMHSHAR

33328
CTLMHSHNR

33329
CTLMHSHTI

33330
CTLMLSHTR

33331
CTMLANHDQ

33332
CTMLANHHQ

33333
CTMLANHYK

33334
CTMLANHYP

33335
CTMLANHYR

33336
CTMLANNYQ

33337
CTMLANYYQ

33338
CTMMANHYQ

33339
CTMVANHYQ

33340
CTMWANHYQ

33341
CTNFHWMND

33342
CTNVKMWHT

33343
CTNYHLKAG

33344
CTNYHVKAV

33345
CTPVKMWHT

33346
CTQFIKHQE

33347
CTQFPKHQE

33348
CTQFTQHQE

33349
CTQFTTHQE

33350
CIQLNKMGP

33351
CTTAWQGTW

33352
CTTFKMWHT

33353
CTTGWQGAW

33354
CTTVKMLHT

33355
CTTVKMWHA

33356
CTIVKMWHI

33357
CTTVKMWNT

33358
CTTVQMWHT

33359
CTTYRNLMS

33360
CTTYRNLPS

33361
CTTYRNVLS

33362
CTVFNTVDQ

33363
CTVLANHYQ

33364
CTYFSMMDP

33365
CTYFSMTDR

33366
CTYKVNSWI

33367
CTYVSMMDR

33368
CVAMKFSEH

33369
CVEPYTDHQ

33370
CVFQCNRSP

33371
CVFQCYGSP

33372
CVFQCYRGP

33373
CVFQCYRNP

33374
CVFQCYRTP

33375
CVFQYYRSP

33376
CVFVPQNFQ

33377
CVFVQENFQ

33378
CVFVQQHFQ

33379
CVFVQQSFQ

33380
CVRIQOTFQ

33381
CVFVRQNFQ

33382
CVGMKFSEH

33383
CVHLVHWQW

33384
CVHQYHQAH

33385
CVLMKFSEH

33386
CVLVQQNFP

33387
CVMGLSFHW

33388
CVRMIDWCC

33389
CVSISAAPV

33390
CVVMKFNEH

33391
CVVMKFSED

33392
CVVMKFSER

33393
CVVMKFSKH

33394
CVVMKFSVH

33395
CVVMKFTEH

33396
CVVMKYSEH

33397
CVVMTFSEH

33398
CVVVQQNFQ

33399
CVVWEFNYP

33400
CVYQCYRSP

33401
CVYWLTNQE

33402
CWAVTSMGS

33403
CWMPQQVAA

33404
CWVVTSMSS

33405
CYAIKQMDI

33406
CYHTTNTSQ

33407
CYHVYKDWS

33408
CYMGLSFHW

33109
CYPFYKDWS

33410
CYPLYKDWS

33411
CYPVSKDWS

33412
CYPVYKAWS

33413
CYPVYKDWT

33414
CYPVYKNWS

33415
CYPVYQDWA

33416
CYPVYTDWS

33417
CYSVYKDWS

33418
CYTIKHMDI

33419
CYTIKKMDI

33420
CYTIKQMAI

33421
CYTIKQMDL

33422
CYTIKQMDM

33423
CYTIKQMDS

33424
CYTIKQMGI

33425
CYTIKQMHI

33426
CYTIKQMNI

33427
CYTIKQVDI

33428
CYTMKQMDI

33429
CYTVYKDWS

33430
DEPCPCDWS

33431
DEPFPCDWA

33432
DEPFPCDWT

33433
DESFPCDWS

33434
DFVRTNLGV

33435
DHANQRVDR

33436
DIFERSMMF

33437
DKAGYGCHE

33438
DKEDPQQWL

33439
DKEEPEQWL

33440
DKGGNGCHE

33441
DKGGYGCNE

33442
DKGGYGGHE

33443
DKGGYSCHE

33414
DKSEESRQE

33445
DKSPWGANH

33446
DKVGYGCHE

33447
DQDGRCYKG

33448
DQDGRCYKQ

33449
DSEPSRGDH

33450
DSQPSRGDY

33451
DYAHQRVDR

33452
DYANERVDR

33453
DYANKRVDR

33454
DYANQRGDR

33455
DYATQRVDR

33456
DYCRRTFHH

33457
DYQPCHVHM

33458
DYSNQRVDR

33459
DYTNQRVDR

33460
EDESRHISW

33461
EDEYRHMSW

33462
EDKSRHMSW

33463
ERITRKAAD

33464
EFVTRKAAT

33465
EGEDRQAFW

33466
EHCNHDQGG

33467
EIFERSMMY

33468
EKAPWGANH

33469
ESARYIDFA

33470
ESARYLDFT

33471
FAEPYTDHQ

33472
FAERTTDHR

33473
FAQRATDHR

33474
FAQRNTDHR

33475
FAQRTTHHR

33476
FAQRTTNHR

33477
FFGTPYHMS

33478
FGHWAQDAS

33479
FGQLIIKHR

33480
FGQRTIDHR

33481
FHDRHFTFT

33482
FHDRPFNFT

33483
FHDRPFTFA

33484
F1CLIKSAS

33485
FICQIKSAA

33486
FKCAHDPTG

33487
FKMQMOYDS

33488
FKVQMQYDN

33489
FRAGMIVSC

33490
FRDAMIVSC

33491
FRDGMIFSC

33492
FRDGMTVSC

33493
FRDRPFTFT

33494
FRHGMIVSC

33495
FRNGMIVSC

33496
FTQQRCAQC

33497
FTQQRCGQC

33498
FTQRTTDHR

33499
FTRQRCVQC

33500
FVEPYTDHR

33501
FVEPYTHHQ

33502
FVQPYTDHQ

33503
FYWCCNREC

33504
FYWCYTREC

33505
FYWFCTREC

33506
GAAVIRKMQ

33507
GFAVIRKMQ

33508
GGWIMRSKR

33509
GIAVIRKMQ

33510
GILPSVKFQ

33511
GILPYGKFQ

33512
GILPYVKVQ

33513
GKCGFDVAA

33514
GKEDMPSFF

33515
GLAVIRKMQ

33516
GLEHCRPGT

33517
GLEWHTKNW

33518
GLQWHTKDW

33519
GNETSDNFH

33520
GSQWLNDSF

33521
GVAAIRKMQ

33522
GVAGIRKMQ

33523
GVAIIRKMQ

33524
GVALIRKMQ

33525
GVAVFRKMQ

33526
GVAV1RKME

33527
GVAVIRKMR

33528
GVEWHTKDW

33529
GVFPNMFAH

33530
GVFPNMFSP

33531
GVTVIRKMQ

33532
GVVVIRKMQ

33533
GVWIVRSKR

33534
GYANQRVDR

33535
HCCEKNGDL

33536
HCCFKNVDM

33537
HCCFKSGDM

33538
HCCFKTGDM

33539
HCCFKYGDM

33540
HCCERNGDM

33541
HKVNLKGDE

33542
HKVTIFKDR

33543
HQLEFDFVG

33544
HQMEFDFVG

33545
HRVT1FKDR

33546
HYCRSGDVQ

33547
IAAQIDDRE

33548
IAAQSDDRQ

33549
IAARSDDRE

33550
IADQSDDRE

33551
IAVCVIRDN

33552
ICARWHCGP

33553
ICLLITKTE

33554
ICLRWHCGP

33555
ICVRWHCGS

33556
ICVRWHCGT

33557
ICWLETKTE

33558
ICWLIAKTE

33559
ICWLIKKTE

33560
ICWLIPKTE

33561
ICWLISKTE

33562
ICWLITKNE

33563
ICWILTKTG

33564
ICWLITKTQ

33565
ICWSITKTE

33566
ICWWITKTE

33567
ISACVIRDN

33568
ISCRTQSDC

33569
ISVCVIRDT

33570
IVQQIRDFH

33571
IWECPRAGS

33572
IWECPRLGS

33573
KADRFMVDQ

33574
KAEHEEMWN

33575
KAEYFDASA

33576
KCAFNGIDE

33577
KCDYRNMHF

33578
KELLCSDDA

33579
KGDRFMVDP

33580
KGKPWGHHN

33581
KHCHSMQFG

33582
KHCRSMQFE

33583
KHCYSMQFE

33584
KIVCRVNYP

33585
KKLNIQKQS

33586
KLCFKEDAF

33587
KMCSSAKSY

33588
KMCTLTKAD

33589
KMFSSAKSY

33590
KMYPSAKSY

33591
KMYSSVKSY

33592
KQCHSMQFE

33593
KSAMPMNDD

33594
KSARCTLHM

33595
KSCLYFDAC

33596
KSCLYFDAF

33597
KSCLYFDSY

33598
KSSWTDSFG

33599
KSVRCKLHM

33600
KVCFKEDAI

33601
KVCFKEDGF

33602
KVCFKEDPF

33603
KVCFKEDSF

33604
KVCYKEDAF

33605
KWCLSIDYR

33606
KWELSIDYP

33607
KWELSIDYS

33608
KWFLSMDYR

33609
KWEPSIDYR

33610
KWFVSIDYR

33611
KWVLSIDYR

33612
KYARCKLHM

33613
LAAQSDDRE

33614
LAARIGEVK

33615
LAARIGKVE

33616
LAARSGEVE

33617
LCVRWHCGP

33618
LCWLITKTE

33619
LGIRNMELR

33620
LKHPAWKEE

33621
LKHPIWKEE

33622
LKHPNWKEE

33623
LKHPPWKEE

33624
LKHPSWKEE

33625
LKHPTWREE

33626
LKHYTWKEE

33627
LKLPTWKEE

33628
LKPPTWKEE

33629
MAANCEVFG

33630
MANENRGIR

33631
MANENRGVH

33632
MANENRVIH

33633
MASENRGIH

33634
MATPERQMP

33635
MAVNCEVFA

33636
MCIRSGFVV

33637
MCIRSGIIV

33638
MCLRSGFIV

33639
MCVRSGFIV

33640
MCWLITKTE

33641
MEDRLAHYY

33642
MEDRVAHYH

33643
MKHPTWKEE

33644
MKIPNIQTI

33645
MKMPRWGMP

33646
MKMTRWGML

33647
MKMTRWGMQ

33648
MKMTRWGMS

33649
MKTPTIQTI

33650
MKVTRWGMP

33651
MQNACFKNP

33652
MSVCVIRDN

33653
MVAHNRLAN

33654
MVANCEVFA

33655
MVEHNRLAD

33656
MVEHNRLAH

33657
MVEHNRLDN

33658
MVEHNRLGN

33659
MVEHNRVAN

33660
MVVYREHMG

33661
MVVYRENMA

33662
MVVYRETMG

33663
MYAERNADM

33664
MYVWRNDFA

33665
MYVVANDGA

33666
NAIRIFLFG

33667
NCCFKNGDM

33668
NCIHNWKHA

33669
NCIHNWQHT

33670
NFFYMDKLS

33671
NFQHVRMLE

33672
NEVETLKRI

33673
NGIRIFLFA

33674
NGIRIFLFV

33675
NGIRNFLFG

33676
NGIRSFLFG

33677
NGLRIFLFG

33678
NGVRIFLFG

33679
NKCAHDPTG

33680
NKCGFDVAA

33681
NSCRRTFHH

33682
NYANQRVDR

33683
NYCRRTFHL

33684
NYCRRTFHP

33685
NYCRRTFHQ

33686
NYCRRTFHR

33687
NYCRRTFHY

33688
NYCRRTFYH

33689
PANHARNPE

33690
PANHARNPQ

33691
PANHARNPR

33692
PANHARTPK

33693
PVCPTRYAD

33694
PVNHARNPK

33695
QAESRDVLD

33696
QAKSRDGLD

33697
QAKSRDVLA

33698
QAKSRDVLN

33699
QAKSRDVMD

33700
QCAYRNMHF

33701
QCCFKNGDM

33702
QCDYRDMHF

33703
QCDYRNMDF

33704
QCDYRNMNF

33705
QCHYRNMHF

33706
QCNYRNMHF

33707
QCVYRNMHF

33708
QFDYRNMHF

33709
QGGRILTSA

33710
QGKSRDVLD

33711
QGNDAIVWQ

33712
QHCNHDQGV

33713
QKIPVNDHQ

33714
QKLDIQKQS

33715
QKLKIQKQS

33716
QKLNIKKQS

33717
QKLNIQKQP

33718
QKLNMQKQS

33719
QKLNVQKQS

33720
QKLPVNDHQ

33721
QKVHVNDHQ

33722
QKVNIQKQS

33723
QKVPLNDHQ

33724
QKVPVSDHQ

33725
QKVPVTDHQ

33726
QKVQVNDHQ

33727
QMCTLTKTD

33728
QQVTIFKDR

33729
QSARYIDFT

33730
QTKGRDGAC

33731
QVKSRDVLD

33732
QWTRWLIVT

33733
RACFEMKHI

33734
RADVFNHRY

33735
RAQSQWDGC

33736
RASCGISMA

33737
RAVTPCHYN

33738
RCCFKNGDM

33739
RCVQPQKEA

33740
RCVQPQKKG

33741
RDLMYTDKA

33742
RHKWRSIWK

33743
RINPDIIAP

33744
RISHDIIAP

33745
RISPDIIAL

33746
RISPDIIAS

33747
RISPDIIGP

33748
RISPDILAP

33749
RISPDTIAP

33750
RISPEIIAP

33751
RISPNIIAP

33752
RISRDIIAP

33753
RIVCRVNYP

33754
RKHGLLFMG

33755
RKHGLLFMT

33756
RKHGLLFMV

33757
RKHGMLFMA

33758
RKHPTWKEE

33759
RKLNIQKQS

33760
RLHGMGDVP

33761
RMAHNFWDA

33762
RMAHTFWNA

33763
RMAPTFWDA

33764
RMARTFWDA

33765
RMHVLVIYT

33766
RMSPDIIAP

33767
RMYSSAKSY

33768
RPCFEMKHI

33769
RQEWRSIWK

33770
RQKLRSIWK

33771
RQKWRIIWK

33772
RQKWRNIWK

33773
RQKWRSFWK

33774
RQKWRSIWE

33775
RQKWRSIWQ

33776
RQKWRSIWR

33777
RQKWRSIWT

33778
RQKWRSMWK

33779
RQKWRSVWK

33780
RQQWRSIWK

33781
RQRWRSIWK

33782
RQTWRSIWK

33783
RSATPCHYN

33784
RSCFEMKHL

33785
RSCFEMRHI

33786
RSCFVMKHI

33787
RSCYEMKHI

33788
RSMWPPSSV

33789
RSVTPCHSN

33790
RSVTPCHYS

33791
RSVTPCHYT

33792
RSVTPCHYY

33793
RTHCHAKAD

33794
RTHGNAKAD

33795
RTHGPAKAD

33796
RTSWKWEYY

33797
RVAVTNHRY

33798
RVCFKEDAF

33799
RVDATNHRY

33800
RVDVINHRF

33801
RVEPTIVAD

33802
RVESQWDGC

33803
RVQSQWHGC

33804
RWFLSIDYR

33805
SADPRVDCP

33806
SADSETPWF

33807
SCWLITKTE

33808
SEHCYRHQH

33809
SEWMIWKCR

33810
SGCFWRHTS

33811
SGCFWRNAS

33812
SGCFWRNTP

33813
SGCFWRSTS

33814
SGCFWRTTS

33815
SGIRIFLFG

33816
SHCLYWMRH

33817
SHCMYWMKH

33818
SHCVYWMKH

33819
SHDVRMHML

33820
SHFLYWMKH

33821
SIVCRVNYP

33822
SKCGFDVAD

33823
SKCGFDVAG

33824
SKCGFDVAP

33825
SKCGFDVAV

33826
SKCGFNVAA

33827
SKCSFDVAA

33828
SKRDMDPAP

33829
SLGNRNADY

33830
SLQNRNADY

33831
SLRDRNADY

33832
SMDTSDWWE

33833
SQWMIWKCQ

33834
SVCPNRYAD

33835
SVCPTRHAD

33836
SVCPTRYAE

33837
SVCPTRYAH

33838
SVCPTRYAN

33839
SVRNRNADY

33840
TAAWRTDDR

33841
TADPRVDCP

33842
TADRWFDAS

33843
TADRWGDAS

33844
TAELRTDDR

33845
TAEPKMDHE

33846
TAEWRNDDR

33847
TAEWRTDAR

33848
TAEWRTDDW

33849
TAEWRTDYR

33850
TAGWRTDDR

33851
TAKWRTDDR

33852
TCILRQSPT

33853
TCLLRQSQT

33854
TCPRKDDDM

33855
TCTRKDDDK

33856
TCTRKDDDT

33857
TCWLITKTE

33858
TFVCRVNYP

33859
TGDRFMVDQ

33860
TGGRIGDSQ

33861
TIVCRINYP

33862
TIVCRVDYP

33863
TIVCRVHYP

33864
TIVCRVNYH

33865
TIVCRVNYR

33866
TIVCRVNYS

33867
TKKKLQCES

33868
TKKKMQCEA

33869
TKKKMQCEY

33870
TKKKMQCQS

33871
TKQKMQCES

33872
TLVCRVNYP

33873
TMKLGDMDG

33874
TSEWRTDDR

33875
TVAMVMDFC

33876
TVEHNRLAN

33877
TVVCRVNYP

33878
VAAQSDDRE

33879
VAQQIRDFH

33880
VAQRTTDHR

33881
VCMHLRQGG

33882
VCMHVRQGA

33883
VCMPVRQGG

33884
VCWLITKTE

33885
VCWMITKTE

33886
VCWVITKTE

33887
VDDKHITGY

33888
VDDKNITCY

33889
VDDKYITCY

33890
VEDKHITCY

33891
VEPFPCDWS

33892
VGGDYFRSD

33893
VKILSEMMT

33894
VKLLSEMMN

33895
VKMLSEMMN

33896
VPVDNLWND

33897
VVAVIRKMQ

33898
VVCMNHCDM

33899
VVEHNRLAN

33900
VVEPYTDHQ

33901
VVQQIRDFN

33902
VVQQIRDFP

33903
VVQQIRDFR

33904
VVQQLRDFH

33905
VVQQSRDFH

33906
VWECPRVGS

33907
WAEYRKTHA

33908
WAEYRKTHH

33909
WAEYRQTHD

33910
WAIRNMELR

33911
WAKYRKTHD

33912
WEHRFPASM

33913
WFAARQHFP

33914
WFIARQHFP

33915
WFVARKHFP

33916
WFVARQHFA

33917
WFVARQHFT

33918
WFVARQNFP

33919
WFVGRQHFP

33920
WGIRNMELG

33921
WGIRNMELQ

33922
WGIRNMEPR

33923
WGIRNMGLR

33924
WGIRNMKLR

33925
WGIRNMVLR

33926
WGIRNTELR

33927
WGNRNMELR

33928
WGSRNMELR

33929
WGVRNMELR

33930
WLVCRCAWE

33931
WSAKSTKKQ

33932
WSAKTTKEQ

33933
WSIRNMELR

33934
WVVCRCDWE

33935
WVVCRCSWE

33936
WVVCRCTWE

33937
YAARFACYQ

33938
YACAIQQHL

33939
YADRFAGYQ

33940
YANLGREHN

33941
YAPDRDGCD

33942
YDEQWNDNS

33943
YDIMNISPF

33944
YDIIVMSPC

33945
YDMMTISPF

33946
YENLAREHN

33947
YFASWIRNH

33948
YFNSWIRNH

33949
YFQSMGHMW

33950
YFRAMGHMW

33951
YFTGWIRNH

33952
YGCFWRNTS

33953
YHCQNYVAS

33954
YIALRATFQ

33955
YIDKIAGWL

33956
YIGLRATFE

33957
YIGLRATFK

33958
YIGLRATFP

33959
YIGVRATFQ

33960
YINKIAGWW

33961
YIPDRDGCD

33962
YIVLRATFQ

33963
YKCAHDPNG

33964
YKCAHDPTE

33965
YKCAYDPTG

33966
YKCGHDPTG

33967
YKCPHDPTG

33968
YKCSHDPTG

33969
YKCTHDPTG

33970
YKCVHDPTG

33971
YKFDMPSFF

33972
YKGAHDPTG

33973
YKGGYGCHE

33974
YLGLRATFQ

33975
YQDGRCYKE

33976
YSGQHFRTL

33977
YTMCKANWN

33978
YVHDRDGCD

33979
YVPDRAGCD

33980
YVPDRDGCA

33981
YVPDRDGCC

33982
YVPDRDGCG

33983
YVPDRDGYD

33984
YVPDRDVCD

33985
YVPDRNGCD

33986
YVPDRVGCD

33987
YVPYRDGCD

33988
YVRDRDGCD

33989
YVTDRDGCD

33990
YYCRRTFHH

Example 41
AAV5 Variants with Spinal Cord Tissue Tropism as Identified by Machine Learning

This example describes preferred properties of engineered AAV5 variants that display spinal cord tissue tropism as identified by machine learning methods. The same bioinformatics pre-process, machine learning methods, and analyses as described in EXAMPLE 19 and FIGS. 19-FIG. 22 were carried out to identify preferred properties of engineered AAV5 variants that display spinal cord tissue tropism.

Favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an AAV5 VP capsid polypeptide, which are associated with a higher probability of spinal cord tissue tropism are described below. Any of the below machine learning-derived positional amino acid preferences described in TABLE 66 can be present alone or in combination with each other or in combination with any of the amino acid preferences observed and described in EXAMPLE 10, TABLE 19. Listed below in TABLE 66 are a summary of positional features shared between the top important features for spinal cord tropism extracted from the ML models.

TABLE 66

Machine Learning-Derived Spinal Cord Tissue Tropism Rules

High volume at Xaa1

Xaa1 is selected from F, W, or Y

Low mutability at Xaa1

Xaa1 is selected from Y, F, L, or C

High solubility at Xaa1

Xaa1 is selected from W, F, I, or L

Low average flexibility at Xaa1

Xaa1 is selected from F, M, or W

Low hydropathy at Xaa2

Xaa2 is selected from P or Y

Low hydrophilicity at Xaa3

Xaa3 is selected from Y, W, V, M, F, I, or L

High solubility at Xaa3

Xaa3 is selected from W, F, I, or L

High volume at Xaa6

Xaa6 is selected from W, R, K, M, I, or L

High mol mass at Xaa6

Xaa6 is selected from W

High mol mass at Xaa8

Xaa8 is selected from W, E, K, M, H, or Q

High volume at Xaa8

Xaa8 is selected from W, K, M, I, or L

High goldman engelman steitz at Xaa8

Xaa8 is selected from V or L

High hydropathy at Xaa9

Xaa9 is selected from V or I

High solubility at Xaa9

Xaa9 is selected from W, F, I, or L

TABLE 67 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid poly peptides that exhibited spinal cord tissue tropism and comport to one or more of the rules provided in TABLE 66. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 35438-SEQ ID NO: 36437, as disclosed in TABLE 67. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 67

Sequences of the 581 to 589 Region in AAV5 VP1

Capsid Polypeptide that Drive Spinal Cord

Tissue Tropism

SEQ

ID
581-589

NO
Sequence

35438
ACLLFLVGI

35439
ACYWLLNWF

35440
ACYWNLNWF

35441
ACYWWMNWF

35442
ADHPENFSD

35443
ADWLRRNIW

35444
AEFRPKISR

35445
AFIQQNNQD

35446
AFIQQSDQD

35447
AFIQQSNED

35448
AFIQQSTQD

35449
AFIQRSNQD

35450
AFVQQSNQD

35451
AFWYSMAFP

35452
AGDRTISMW

35453
AHFRAYIQY

35454
AHKWFCEGS

35455
AHKWVCEDS

35456
AKNVFEFHV

35457
ALFAPNGEV

35458
ALFYMEKMP

35459
AMVLFLVDI

35460
AMWIRGIWG

35461
ANDMCNYYG

35462
AQGNKTYVI

35463
AQKKCLSFG

35464
AQKWFCEDS

35465
AQQKCLSFG

35466
AQVNKTYLI

35467
AQVNKTYVI

35468
AQVIKTYVI

35469
ATMFSVIHQ

35470
ATWVHYDMY

35471
AVIQQSNQD

35472
AWFDMEKMP

35473
AWFHMEKMP

35474
AWFNMEKMP

35475
AWFYKEKMP

35476
AWFYMFEMP

35477
AWFYMFKMS

35478
AWFYMENMP

35479
AWFYMGKMP

35480
AWFYREKMP

35481
AWLQGGIYN

35482
AWSYMEKMP

35483
CAVKQECSR

35484
CCCSEIGVN

35485
CHVMSVPCA

35486
CKNWYINMH

35487
CQWTPEIFQ

35488
CTQCYGDMP

35489
CWAFFRMRD

35490
CWGEFRMRV

35491
CWTEFRMRV

35492
CWVEFRMRV

35493
CWWTPEIFQ

35494
DAKSPCDSV

35495
DAKSPCESF

35496
DAVNDLTWP

35497
DCIPDWSST

35498
DCYWWLNWF

35499
DEANKTYVI

35500
DEDNKTYVI

35501
DEVNKTYVI

35502
DFILDWSST

35503
DFIPDWSSI

35504
DFIRDWSST

35505
DFITDWSST

35506
DFVPDWSST

35507
DFWYGMAFP

35508
DFWYSMACP

35509
DFWYSMAVP

35510
DFWYSMTFP

35511
DFWYSRAFP

35512
DFWYSVAFP

35513
DFWYTMAFP

35514
DHVNKPYVI

35515
DIIPDWSST

35516
DIRIVTDGR

35517
DIWYSMAFP

35518
DKGNKTYVI

35519
DKNVFEEDV

35520
DKNVFEQHV

35521
DKNVYEEHV

35522
DNVNKTYVI

35523
DPGNKTYVI

35524
DPVNKTYVL

35525
DPVNKTYVV

35526
DPVTKTYVI

35527
DQAEVLSDH

35528
DQANKTYVI

35529
DQDNKTYVI

35530
DQDTKTYVI

35531
DQGNKTYVI

35532
DQGSKTYVI

35533
DQINKPYVI

35534
DQINKTYVI

35535
DQINKTYVS

35536
DQLNKTYVI

35537
DQVDKTYVI

35538
DQVNKNYVI

35539
DQVNKPYEI

35540
DQVNKTHVI

35541
DQVNKTNVI

35542
DQVNKTSVI

35543
DQVNKTYAI

35544
DQVNKTYEI

35545
DQVNKTYML

35546
DQVNKTYVF

35547
DQVNKTYVN

35548
DQVNKTYVV

35549
DQVNMTYVI

35550
DQVNNTYMI

35551
DQVNQTYVI

35552
DQVNRTYVE

35553
DQVSKTYVE

35554
DQVTKTYVI

35555
DQVTQTYVE

35556
DRNVFEEHV

35557
DRVNKTYLE

35558
DRVNKTYVS

35559
DVNLKITTH

35560
DVNLQEATH

35561
DVYYCPEAI

35562
DWMQGGIYH

35563
DWMQGGIYY

35564
DYAQYYRTW

35565
DYDPDIKDP

35566
DYDPDLKDQ

35567
DYGPDIKDQ

35568
ECLLFLIGI

35569
ECMLFLVGI

35570
ECVSKWIVF

35571
EDWIRRNIW

35572
EDWLRCNEW

35573
EDWLRRDIW

35574
EFIQQSNQD

35575
EHFRAYAQY

35576
EHFRAYEQY

35577
EHFRDYTQY

35578
EHFRSYTQY

35579
EHWLRRNIW

35580
EIIPQFAYF

35581
EINREVREN

35582
EINRVVRQN

35583
EISREVRQN

35584
EKELKNSTQ

35585
ELIYKFKTD

35586
ELIYKIETD

35587
ELIYKMKTD

35588
ELVNKTYVE

35589
ELVYKIKTD

35590
EMINACFQT

35591
EMKCIGAAI

35592
EMKCMGAGI

35593
EMKCTGAGI

35594
EMKWLGAGI

35595
EMKYIGAGI

35596
EMWIGGIWG

35597
EMWIRGVWG

35598
EMWVRGVWG

35599
EQEEKNSTQ

35600
EQELKDSTQ

35601
EQELKNSTL

35602
EQELKTSTQ

35603
EQEVKNSTQ

35604
EQQLKNSTQ

35605
ERFRAYTQY

35606
ETFHNTATI

35607
ETWVHYDMY

35608
EYGTKVKMD

35609
EYSTKVKMD

35610
EYVTKVKMD

35611
FAEASITRW

35612
FASAFANTD

35613
FASAFSNSD

35614
FASAFVNSD

35615
FASVFANSD

35616
FCMEWEVMD

35617
FFMAWEVMD

35618
FFMDWEVMD

35619
FFMEWEGMD

35620
FFMEWELMD

35621
FFMEWEVKD

35622
FFMEWEVRD

35623
FFMEWEVTD

35624
FFMEWEVVD

35625
FFMEWKVMD

35626
FFMEWQVMD

35627
FFYECYSNK

35628
FIIGCMLIW

35629
FIIHCMLIW

35630
FIIRAMLIW

35631
FIIRCMLIL

35632
FIIRCMLVW

35633
FIIRCMVIW

35634
FIERCVLIW

35635
FIIRCVLVW

35636
FIIRGMLIW

35637
FIIRWMLEW

35638
FILRCLLEW

35639
FILRCMLIC

35640
FILRCMLIW

35641
FIVGGLAFA

35642
FIVRCMLIW

35643
FKFISYEGD

35644
FKHWYINMH

35645
FKICFSPTV

35646
FKNGYINMH

35647
FKNHYEGGP

35648
FKNLYENMH

35649
FKNWFINMH

35650
FKNWHINMH

35651
FKNWSINMH

35652
FKNWYEHMH

35653
FKNWYINMD

35654
FKNWYINMF

35655
FKNWYINML

35655
FKNWYINMN

35657
FKNWYINMP

35658
FKNWYINMR

35659
FKNWYINMY

35660
FKNWYINRH

35661
FKNWYINTH

35662
FKNWYINVH

35663
FKNWYESMH

35664
FKNWYLHMH

35665
FKNWYNNMH

35666
FKNWYVNNM

35667
FKSWYINMH

35668
FKVGGLAFA

35669
FKYWYINMH

35670
FLIRCMLIW

35671
FMIRCMLIW

35672
FPEASLTRW

35673
FQNWYINMH

35674
FRNWYINMH

35675
FRNWYMNMH

35676
FTFGGLAFA

35677
FTIGGLAFA

35678
FTNTKKQAF

35679
FTNWYINMD

35680
FTNWYINMH

35681
FTNWYINMN

35682
FTVAGLAFA

35683
FTVGGLAFG

35684
FTVGGLAFP

35685
FTVGGLAFT

35686
FTVGGLAFV

35687
FTVGGLGFA

35688
FTVGGLSFA

35689
FTVGGLTFA

35690
FTVGGMAFA

35691
FWCHIIGEY

35692
FWCNIIGIY

35693
FWGNIIGIY

35694
FWSNFIGIY

35695
FWSNIEGFY

35696
FWSNIEGIC

35697
FWSNIIGIH

35698
FWSNIIGIN

35699
FWSNIIGLY

35700
FWSNIIGMY

35701
FWSNIIGVY

35702
FWSNIMGIY

35703
FWSNLIGIY

35704
FWSNMIGEY

35705
FWSNVEGIY

35706
FWSSIIGIY

35707
FWSTIIGIY

35708
FWTNIIGIY

35709
FYMEWEVMD

35710
GCYWWLNWF

35711
GDEYYDEMG

35712
GDHTENFSD

35713
GEFRPKISR

35714
GFIQQSNQD

35715
GHVMTVPCA

35716
GIKYKDILS

35717
GINREVRQN

35718
GKNVFEEHV

35719
GQGNKTYVI

35720
GSASRNQRD

35721
GSSPGKGWM

35722
GSSSRNQRD

35723
GTWVHYDMF

35724
GWAEFRMRV

35725
GWAEFRVRV

35726
GWSEFRMRV

35727
HPVNKTYVI

35728
HQGNKTYVI

35729
HRFQEKGSR

35730
HRFQEKGTH

35731
HRFREKGSH

35732
HILLWYGIW

35733
HTLLWYGIY

35734
HVYCCPEAI

35735
ICHCCMAWG

35736
ICHCCMVWG

35737
ICHCCMYWG

35738
ICHWCMDWG

35739
ICYCCMDWG

35740
ICYWWLNCF

35741
ICYWWLNWF

35742
IDRWWVIDV

35743
IHIGTNSYA

35744
IHTATNSYA

35745
IHTGTNSYV

35746
IIYECYSNK

35747
IKHIAAESR

35748
IKHIATETR

35749
IKHISTESR

35750
IKNWYINMH

35751
IKNWYINMP

35752
IKSQTLTDC

35753
IMFHCREMG

35754
IMVHCRQMG

35755
IMVHCRVMG

35756
IRNMFEQYT

35757
IRSHTLTYC

35758
IRSQTITDC

35759
IRSQTLTDW

35760
IRSQTLTDY

35761
IRSQTLTNC

35762
IRSQTVTDC

35763
IRSRTLTDC

35764
IRWLRLHKK

35765
ISWIIRADI

35766
ISWINRADI

35767
ISWITRADV

35768
ISWITRAGI

35769
ISWITRTDI

35770
ITSQTLTDC

35771
ITWLHYDMY

35772
IWFYMEKMP

35773
IWSNIIGIY

35774
IWYWWLNWF

35775
KAVWDPSSM

35776
KCWDCHDAG

35777
KCWDCHEGG

35778
KEDFGWKHV

35779
KESRSQTGD

35780
KEVFGWKHM

35781
KGVWDPGSM

35782
KGVWDPSST

35783
KHIGSYTQF

35784
KHNMLYHMD

35785
KHYKWGHGQ

35786
KINREVRQN

35787
KKWSCHRGH

35788
KKYQDTHTF

35789
KMWIRGIWG

35790
KRIKMPPPR

35791
KRIKMPPTK

35792
KWHAGPGVV

35793
KWHCGPGVV

35794
KWHGGPGAV

35795
KWHGGPGDV

35796
KWHGGPGFV

35797
KWHGGPGVM

35798
KWHGGQGVV

35799
KWHGGRGVV

35800
KWLGGPGVV

35801
KWLYGWPIK

35802
KWYGGPGVV

35803
KYNMMYHMD

35804
KYNMRYHMD

35805
KYNMVYHMD

35806
KYNVLYHMD

35807
KYSMLYHMD

35808
LATMSLTWW

35809
LCYWWLNWF

35810
LEKWGHMGT

35811
LKICFGPTV

35812
LKIGFSPTV

35813
LPEIYLSFC

35814
LPKRYLSFC

35815
LPQIYLSFC

35816
LPTIYLSFC

35817
LQICFSPTV

35818
LRICFSPTV

35819
LRWVRLHKK

35820
LSKIYLSFC

35821
LSSCYYTTY

35822
LSWITRADI

35823
LVFLRGQHY

35824
LWFYMEEMP

35825
LWFYMEKMP

35826
LWSNMIGIY

35827
MFRIEAVCP

35828
MKFQDTHTF

35829
MKRDAAFCA

35830
MKREAAVCA

35831
MKRETAFCP

35832
MKRETAFCS

35833
MKRETAFCT

35834
MKRETAVCA

35835
MKRETGFCA

35836
MKYKDTHTF

35837
MKYQDTHTC

35838
MKYQDTHTI

35839
MKYWDTHTF

35840
MYFFVWRFM

35841
MYIFGWRFM

35842
MYIFVWGFM

35843
MYIFVWRFT

35844
MYIFVWRIM

35845
MYIFVWRVM

35846
MYIFYWRFM

35847
MYIIVWRFM

35848
MYISVWRFM

35849
MYIVVWRFM

35850
MYIYFWRFM

35851
MYIYVWRFM

35852
MYLFVWRFM

35853
NANDVQWYD

35854
NAPDVQWYD

35855
NATDFQWYD

35856
NATDGQWHD

35857
NATDVEWYD

35858
NATDVKWYD

35859
NATDVLWYD

35860
NATDVRWYD

35861
NCDRSLAEY

35862
NCHIDRVQD

35863
NCHVDHVQD

35864
NCKNLTYYI

35865
NFIPDWSST

35866
NFWYSMAFP

35867
NIDAHERYR

35868
NKNVFEEHV

35869
NLCRHLFQC

35870
NLCRHLFQF

35871
NNYKFQQDG

35872
NNYKGQQDG

35873
NNYKLQQDG

35874
NNYKVQEDG

35875
NPAYGRGHI

35876
NPELGKCEI

35877
NPGWRYMWF

35878
NPGWRYMWW

35879
NQAYGRGHI

35880
NQVNKTYVI

35881
NRAYGRGHV

35882
NSWITRADI

35883
NSWRHICDH

35884
NVAAHERYR

35885
NVCRHLFQF

35886
NVDAHERYK

35887
NVDAPERYR

35888
NVDVHERYR

35889
NVGAHERYR

35890
NVYYCPEAI

35891
NWCKHLFQF

35892
NWCRDLFQF

35893
NWCRHLFEC

35894
NWCRHLFEF

35895
NWCRHLFKF

35896
NWCRHLFKY

35897
NWCRHLFQC

35898
NWCRHLFQI

35899
NWCRHLFQV

35900
NWCRHLFQY

35901
NWCRHLFRF

35902
NWCRHLIQF

35903
NWCRHLVQF

35904
NWCRHRFQF

35905
NWCRILFQF

35906
NWCRLLFQF

35907
NWCRNLFQF

35908
NWCRQLFQV

35909
NWCRVLFHF

35910
NWFRHLFQF

35911
NWGRHLFQF

35912
NWWRHLFQF

35913
NYAQHYRTW

35914
NYSQYYRTW

35915
PADRTISMW

35916
PDDRTISMW

35917
PGDRTISKW

35918
PGDRTITMW

35919
PGDRTLSMW

35920
PGNRTISMW

35921
PHKWFCEDS

35922
PPDIGTQFI

35923
PPHIGTEFI

35924
PPHIGTKFI

35925
PPHLGTQFI

35926
PPHMGTQFI

35927
PPHTGTQFI

35928
PPLIGTQFI

35929
PRFQEKGSH

35930
PRHIGTQFI

35931
PWFYLEKMP

35932
PWFYMEEMP

35933
QCLLFLVGI

35934
QEKWGHKHT

35935
QEKWGHMHA

35936
QEKWGHVHT

35937
QTIIGNVWV

35938
QWFYMEKMP

35939
RAHWSDIAD

35940
RAPADWACA

35941
RDGSNWLCI

35942
RDPADWACG

35943
RDPADWACS

35944
RDPADWICA

35945
RDPADWVCA

35946
RDPSDWACA

35947
RDPTDWACA

35948
RDTADWACA

35949
REDFGWKHM

35950
RHIGSYTQF

35951
RHIHRYWEC

35952
RHKWFCEAS

35953
RHPADWACA

35954
RHVHPYWEC

35955
RHVHQYWEC

35956
RHVHRFWEC

35957
RHVHRYWVC

35958
RIMHSWYND

35959
RKEQDHWDK

35960
RKVESYCIT

35961
RKWSCHRAH

35962
RKWSCHRCH

35963
RKWSCHRGL

35964
RKWSCHRGY

35965
RKWSCHRVH

35966
RKWSCNRGH

35967
RKWYCHRGH

35968
RPHIGTQFI

35969
RPVQEDGCG

35970
RQNDTHRAN

35971
RQVQRYWEC

35972
RQWSCHRGH

35973
RRFQEKGSH

35974
RRIESYCIT

35975
RRIKMPPPK

35976
RTVKEDGCG

35977
RVPADWACA

35978
RWSYMKAGK

35979
RWSYMQAGI

35980
RWSYMQTGK

35981
RWSYMQVGK

35982
RWSYMRAGK

35983
RYIFVWRLM

35984
RYNMLYHMD

35985
RYPADWACA

35986
SAAWQFGES

35987
SAAWQFGVS

35988
SDDMCNYYG

35989
SDMIKLAEI

35990
SGADHYHLK

35991
SGFYMEMMP

35992
SGFYMEQMP

35993
SGFYMERMP

35994
SHKWFCEDS

35995
SHYWPMCWS

35996
SLFYMDKMP

35997
SLFYMEKML

35998
SLFYMEKMP

35999
SLLRFQPSI

36000
SLMLFLPSI

36001
SLMRCLPSI

36002
SLMRFLASI

36003
SLMRFLHSI

36004
SLMRFLPAI

36005
SLMRFLPST

36006
SLMRFLPSV

36007
SLMRFLPYI

36008
SLMRFLRSI

36009
SLMRFLSSI

36010
SLMRFLTSI

36011
SLMRFPPSI

36012
SLMFRVPSI

36013
SLMRVLPSI

36014
SLVRFLLSI

36015
SLVRFLPSI

36016
SMHDSNFNV

36017
SMHDYNFNI

36018
SMHDYTFNV

36019
SNDKCNYYG

36020
SNDMCDYYG

36021
SNDMCNHYG

36022
SNDMCNSYG

36023
SNNMCNYYG

36024
SPYWLMRMG

36025
SQDCGRPE1

36026
SRMRFLPSI

36027
SRMRLLPSI

36028
SSASVNHTS

36029
SSDMCNYYG

36030
SSFYMEKMP

36031
SSSPGKGWK

36032
SSWITRADI

36033
STDMCNYYG

36034
SVADHYHRK

36035
SVADHYNLK

36036
SVADYYHLK

36037
SVAHHYHLK

36038
SVARQMHSW

36039
SVMRFLPSI

36040
SVSDHYHLK

36041
SWCFMEKMP

36042
SWCRHLFEF

36043
SWCRHLFQF

36044
SWCYIEKMP

36045
SWCYMEEMP

36046
SWCYMEQMP

36047
SWCYMVKMP

36048
SWFCVEKMP

35049
SWFDMEKMP

35050
SWFFMEKMP

36051
SWFFMVKMP

36052
SWFFVEKMP

36053
SWFHMEKMP

36054
SWFHMEKMQ

36055
SWFNMEQMP

36056
SWFSKEKMP

36057
SWFSVEKMP

36058
SWFYIERMP

36059
SWFYKETMP

36060
SWFYMAQMP

35051
SWFYMDKMT

35052
SWFYMDMMP

36063
SWFYMFAMP

36064
SWFYMEDMP

36065
SWFYMEEVP

36066
SWFYMEGMP

36067
SWFYMEKIR

36068
SWFYMEKMA

36069
SWFYMEKMI

36070
SWFYMEKMR

36071
SWFYMEKMT

36072
SWFYMEKRP

35073
SWFYMEKVP

35074
SWFYMEMMP

36075
SWFYMERMP

36076
SWFYMETMP

36077
SWFYMEWMP

36078
SWFYMKKMP

36079
SWFYMQKMP

36080
SWFYMVKMP

36081
SWFYRDKMP

36082
SWFYREKLP

36083
SWFYVEEMP

36084
SWFYVEKIP

36085
SWFYVEKMP

36086
SWIYMDKMP

36087
SWIYMEMIT

36088
SWIYMEQMP

36089
SWTYMEKMP

36090
SWVCMEKMP

36091
SWVYMEKMQ

36092
SWVYMEKMS

36093
SWVYMEMMP

36094
SWVYMEQmP

36095
SWVYMKKMP

36096
SWYCMEKMP

36097
SWYYDERSN

36098
SWYYMEEMP

36099
SWYYMEKMP

36100
TAEKFICFP

36101
TCARSLAEY

36102
TCDRSLAQY

36103
TCDRSLGEY

36104
TCDRSLPEY

36105
TCDRSLTEY

36106
TCDRSLVEY

36107
TCDRTLAEY

36108
TCKNLTYSI

36109
TCKNMTYYI

36110
TCKSITYYI

36111
TCNRSLAEY

36112
TCRNLTYYI

36113
TCTNLTYYI

36114
TDKWFCVDS

36115
THIASYTQF

36116
THICSYTQF

36117
THIGSYTEF

36118
THIGTYTQF

36119
THIVSYTQF

36120
THKWCCEDS

36121
THKWFCDYS

36122
THKWFCEDC

36123
THKWFCEDI

36124
THKWFCEDN

36125
THKWFCEDT

36126
THKWFCEHS

36127
THKWFCENI

36128
THKWFCENS

36129
THKWFCEVS

36130
THKWFCEYS

36131
THKWFCGDI

36132
THKWFCQDS

36133
THKWFCVDS

36134
THKWICEDI

36135
THKWVCEDS

36136
THRWFCEDS

36137
THRWFCEES

36138
THRWLCEDS

36139
TLMRFLPSI

36140
TPKWFCEDN

36141
TPKWFCEDS

36142
TPKWVCEDS

36143
TRKWFCEDS

36144
TSERFICFP

36145
TTDHFICFP

36146
TTELYICFP

36147
TWCRHLFQF

36148
TWFHMEKMP

36149
TWFYMDKMP

36150
TWFYMEKMP

36151
TWFYMEKMQ

36152
TWFYMEQMP

36153
TWLCGWPIK

36154
TWLFGWPIK

36155
TWLGGCKSV

36156
TWLYGWPFK

36157
TWLYGWPIQ

36158
TWLYGWPIR

36159
TWLYGWPIT

36160
TWLYGWPVK

36161
TWLYGWSIK

36162
TWLYGWTIK

36163
TWMYGWPIK

36164
TWPVGCKSV

35155
TWRGGCKSV

35156
TYAQYYRTW

35157
VAFRPKISR

36168
VCCWWLNWF

36169
VCSWWLNWF

36170
VCYWWLDWF

36171
VCYWWLHWI

36172
VCYMNLIWF

36173
VCYWWLNLF

36174
VCYWWLNWC

36175
VCYWWLNWI

36176
VCYWWLNWY

36177
VCYWWLSWL

35178
VCYWWLTWF

35179
VCYWWMNWF

36180
VEFGPKISR

36181
VEFKPKISR

36182
VEFRHKISR

36183
VEFRPEISR

36184
VEFRPKFSR

36185
VEFRPKLSR

36186
VEFRPKMSR

36187
VEFRPKTSR

36188
VEFRPKVSR

36189
VEFRPRISR

35190
VEFRPTISR

35191
VEFRRKISR

36192
VEFRSKISR

36193
VEFRTKISR

36194
VEFTPKISR

36195
VEYRPKISR

36195
VFIQQSNQD

36197
VFYWWLNWF

36198
VGYWWLNWF

36199
VIWVHYDMY

36200
VKFRPKISR

36201
VKKCHTCGA

35202
VKNWYINMH

35203
VKWVRLHKK

36204
VNWVHYDMY

36205
VQVNKTYVI

36206
VRWLRLHKK

36207
VRWVRIHKK

36208
VRWVRLHEK

36209
VRWVRLHKQ

36210
VRWVRLHKR

36211
VRWVRLHKT

36212
VRWVRLHRK

36213
VRWVRLHTK

36214
VRYWWLNCF

36215
VSWITRADI

36216
VTVGGLAFA

36217
VTWMHYDMY

36218
VTWVHYAMY

36219
VTWVHYDMF

36220
VTWVHYDMH

36221
VTWVHYNMY

36222
VTWVHYVMY

36223
VTWVHYYMY

36224
VVFLRDQHY

36225
VWSNIIGIY

36226
VWYWWLNWF

36227
VWYWWLNWL

36228
VYYWWLNWF

36229
WANFKTLNR

36230
WANFQTLNK

36231
WAPTPWRTH

36232
WCCKMWSWW

36233
WCMSQQYED

36234
WDGKFWETM

36235
WDIKFWETM

36236
WDLKFWETM

36237
WDMSQQYED

36238
WDNLRNTQI

36239
WDASSNTQI

36240
WDNWINTQI

36241
WDNWSDTQI

36242
WDNWSHTQI

36243
WDNWSITQI

36244
WDNWSNTEI

36245
WDNWSNTKI

36246
WDNWSNTKK

36247
WDNWSNTLI

36248
WDNWSNTPI

36249
WDNWSNTQM

36250
WDNWSNTQV

36251
WDNWSNTRI

36252
WDNWSTTQI

36253
WDNWTNTQI

36254
WDNWTNTQM

36255
WDSWSNTQI

36256
WDVKFWESM

36257
WDVKYWETM

36258
WDVMFWETM

36259
WENWSNSQI

36260
WFCKKWSWW

36261
WFCKMLSWW

36262
WFCKMWGWG

36263
WFCKMWGWW

36264
WFCKMWIWW

36265
WFCKMWSGW

36266
WFCKMWSLW

36267
WFCKMWSWG

36268
WFCKMWSWL

36269
WFCKMWTWW

36270
WFCKVWSWW

36271
WFCNLWSWW

36272
WFCQMWSWW

36273
WFCRMWSWW

36274
WFCTLWSWW

36275
WFCTMWSWW

36276
WFFKMWSWW

36277
WFGKMWSWW

36278
WFNMPHDRH

36279
WFWKMWSWW

36280
WFYKMWSWW

36281
WGNWSNTQI

36282
WGVKFWETM

36283
WHCSVYTCS

36284
WHRDEMLTP

36285
WHREEMLTT

36286
WHVKFWETM

36287
WHWDEMLTT

36288
WIAAMTYWC

36289
WICKMWSWW

36290
WIHAMTYWC

36291
WIHMPHDRH

36292
WINMAHDRH

36293
WINMHHDRH

36294
WINMPDDRH

36295
WINMPHARH

36296
WINMPHDGH

36297
WINMPHDKH

36298
WINMPHDRD

36299
WINMPHDRL

36300
WINMPHDRN

36301
WINMPHDRP

36302
WINMPHDRR

36303
WINMPHDRY

36304
WINMPHGRH

36305
WINMPLDRH

36306
WINMPNDRH

36307
WINMPPDRH

36308
WINMPRDRH

36309
WINMTHDRH

36310
WINRPHDRH

36311
WINVPHDRH

36312
WIPAKTYWC

36313
WIPAMAYWC

36314
WIPAMPYWC

36315
WIPAMTYLC

36316
WIPAMTYWF

36317
WIPAMTYWW

36318
WIPAMTYWY

36319
WIPARTYWC

36320
WIPAVTYWC

36321
WIPGMTYWC

36322
WIPSMTYWC

36323
WIPTMTYWC

36324
WIPVVTYWC

36325
WISAMTYWC

36326
WISMPHDRH

36327
WITAMTYWC

36328
WITMPHDRH

35329
WKNVCPSRQ

36330
WKSVCPSRQ

36331
WKTVCPSRH

36332
WLCQMWSWW

36333
WLPAMTYWC

36334
WMLLGLTWF

36335
WMLLGMAWC

36336
WMLLGMHWC

36337
WMLLGMNWC

36338
WMLLGMPWC

36339
WMLLGMTLC

36340
WMLLGMTWF

36341
WMLLGMTWG

36342
WMLLGMTWW

36343
WMLLGMTWY

36344
WMLLGTTWC

36345
WMLLGVTWC

36346
WMLLVMTWC

36347
WMLNQSRGW

36348
WMLQGMTWC

36349
WMMLGMTWC

36350
WMMMGLTWC

36351
WMNMPHDRH

36352
WMPNQSRVW

36353
WMVLGMNWC

36354
WMVLGMTWC

36355
WNCSVYTCI

36356
WNNWSNTQI

36357
WRLLGMTWC

36358
WSCKMWSWW

36359
WTLLGMTWC

36360
WVCKMWGWW

36361
MCKIMWSWW

36362
WVNLCADLI

36363
WVPAMTYWC

36364
WVPAPWRTH

36365
WVPPPWRTH

36366
WVPTHWRTH

36367
WVPTPGRTH

36368
WVPTPWQTH

36369
WVPTPWRSH

36370
WVPTPWRTN

36371
WVPTPWRTQ

36372
WVPTPWRTY

36373
WVPTTWRTH

36374
WVTLCAVLI

36375
WVTTPWRTH

36376
WYMSKQYED

35377
WYMSQEYED

35378
WYMSQQNED

35379
WYMSQQSED

36380
WYMSQQYAD

36381
WYMSQQYKD

36382
WYMSQQYQD

36383
WYMSQRYED

36384
WYMTQQYED

36385
WYTSQQYED

36386
WYVKFWETM

36387
WYVSQQYED

36388
YAQPDWFMT

36389
YAQRDWFMA

36390
YAWFYWADY

36391
YDQPDWFMA

36392
YDYWPMCWS

36393
YFMEWEVMD

36394
YFWYSMAFP

36395
YHYWHMCWS

36396
YHYWPMCWA

36397
YHYWPMCWC

36398
YHYWPMCWP

36399
YHYWPMCWI

36400
YHYWPMCWY

36401
YHYWPMGWS

36402
YIIRCMLIW

36403
YKNWYINMH

36404
YLEAKTIHA

36405
YLESKTIHP

36406
YPELGKGEI

35407
YPELGQCEI

36408
YPQPDWFMA

36409
YPYWPMCWS

36410
YSWFNWADY

36411
YSWFYWADC

36412
YSWFYWADF

36413
YSWFYWADS

36414
YSWFYWAVY

36415
YSWFYWPDY

36416
YSWFYWSDY

36417
YSWFYWTDY

36418
YSWFYWVDY

35419
YSWIYWADY

36420
YSWVYWADY

36421
YSWYYWADY

35422
YTALMWVWA

35423
YTELGKCEI

36424
YTLLWYGIC

36425
YTNFEAVLE

36426
YTNFEDALE

36427
YTNFEDVII

36428
YTNFEGVLI

36429
YTNFENVLI

36430
YTNEEDVLI

36431
YTNLDDVLE

36432
YTWFYWADY

36433
YWSHIIGIY

35434
YWSNIIGIY

35435
YYAQYYRTW

35436
YYWFYWADY

36437
YYWFYWSDY

Example 42
AAV5 Variants with Spleen Tissue Tropism as Identified by Machine Learning

This example describes preferred properties of engineered AAV5 variants that display spleen tissue tropism as identified by machine learning methods. The same bioinformatics pre-process, machine learning methods, and analyses as described in EXAMPLE 19 and FIGS. 19-FIG. 22 were carried out to identify preferred properties of engineered AAV5 variants that display spleen tissue tropism.

Favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an AAV5 VP capsid polypeptide, which are associated with a higher probability of spleen tissue tropism are described below. Any of the below machine learning-derived positional amino acid preferences described in TABLE 68 can be present alone or in combination with each other or in combination with any of the amino acid preferences observed and described in EXAMPLE 6, TABLE 11. Listed below in TABLE 68 are a summary of positional features shared between the top important features for spleen tropism extracted from the ML models.

TABLE 68

Machine Learning-Derived Spleen Tissue Tropism Rules

Low solubility at Xaa1

Xaa1 is selected from D or P

Or

High solubility

Xaa1 is selected from F, I, or L

Low hydropathy at Xaa1

Xaa1 is selected from Y or P

Low mutability at Xaa1

Xaa1 is selected from C, K, or P

Low solubility at Xaa2

Xaa2 is selected from D, Q, or R

Low hydropathy at Xaa2

Xaa2 is selected from D, E, R, K, H, N, or Q

Low charge at Xaa2

Xaa2 is selected from D or E

Low volume at Xaa2

Xaa2 is selected from T, N, P, or D

High average flexibility at Xaa2

Xaa2 is selected from D, E, R, P, G, Q, or S

Low solubility at Xaa3

Xaa3 is selected from D, E, P, or N

Low hydropathy at Xaa3

Xaa3 is selected from D, E, H, N, Q, or P

Low hydropathy at Xaa4

Xaa4 is selected from K or R

Low solubility at Xaa5

Xaa5 is selected from D, E, P, or N

High average flexibility at Xaa5

Xaa5 is selected from D, E, R, P, G, Q, or S

Low mutability at Xaa6

Xaa6 is selected from C

High surface accessibility at Xaa8

Xaa8 is selected from E, R, or K

Low solubility at Xaa8

Xaa8 is selected from E, P, R, K, N, or Q

Medium volume at Xaa8

Xaa8 is selected from E, D, R, K, V, P, M,

I, L, H, N, Q, or T

Medium mol mass at Xaa9

Xaa9 is selected from E, D, K, M, I, L, H, or N

TABLE 69 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that exhibited spleen tissue tropism and comport to one or more of the rules provided in TABLE 68. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 38438-SEQ ID NO: 39437, as disclosed in TABLE 69. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 69

Sequences of the 581 to 589 Region in AAV5 VP1

Capsid Polypeptide that Drive Spleen

Tissue Tropism

SEQ

ID
581-589

NO
Sequence

38438
ADDMCCSPN

38439
ADEDDCDDP

38440
ADEMEEEHG

38441
ADEMWPIRF

38442
ADHCNWSRT

38443
ADHCSWSRT

38444
ADMCECVKG

38445
ADNRTKPLC

38446
ADPDQKAMN

38447
ADQRDCWTT

38448
ADREPWPRN

38449
ADSRVRPKW

38450
AECKCCVKM

38451
AENQDDWQF

38452
AGDMWRHKM

38453
AGDTDMPED

38454
AGWEEAWCM

38455
AHDCDEPAL

38456
AIENNYCPM

38457
AIPHWDDKH

38458
APDFNREKN

38459
APDHWMEKM

38460
ARKCDFPAL

38461
CDAKRTPGM

38462
CDAWGELRG

38463
CDCKEQEED

38464
CDCKGRIPH

38465
CDDFCMCHC

38466
CDDHNHPRA

38467
CDDKQSHKE

38468
CDDKRFSKK

38469
CDDRERQQN

38470
CDDWGWDQM

38471
CDEIEMQGT

38472
CDELPMAGH

38473
CDEPHFWRC

38474
CDERRWDRP

38475
CDIRCENMG

38476
CDMCREWKW

38477
CDNKCWPHD

38478
CDNKPKNLS

38479
CDNRRNWWE

38480
CDPCNEFNK

38481
CDPMDACQE

38482
CDPNDMREH

38483
CDPRFVEAM

38484
CDPRTLPFY

38485
CDPWDWDIE

38486
CDPWDWEIE

38487
CDQFCEWRQ

38488
CDQFHWDKK

38489
CDRRCFDKH

38490
CDSCCGMHV

38491
CDTWPWSKK

38492
CDWCDQFRH

38493
CDYCDPRKF

38494
CDYRCLWYC

38495
CECYEPFEG

38496
CEDMDQWPW

38497
CELPDRCPQ

38498
CERVPVFSF

38499
CFDHRPLPC

38500
CFPCCWAKF

38501
CGAWWWNHM

38502
CGDEWAPRL

38503
CGDIDHPKS

38504
CGDWAWDRC

38505
CGEKCWHDI

38506
CGHDSKPRC

38507
CGHWCCSDY

38508
CGIEDKPEN

38509
CGLCRYYDM

38510
CGPEPCVRQ

38511
CGPEPCVRQ

38512
CGPWNEPEY

38513
CGTHCPLRQ

38514
CGYDCQPRF

38515
CHKFDRELW

38516
CHNHPMPPL

38517
CKDEFWPLM

38518
CKDFRMSPL

38519
CKDYCRDEL

38520
CKPYEWWVM

38521
CKYTDDEEQ

38522
CMTDWSGMM

38523
CNCKDYVEV

38524
CNDQHMEKD

38525
CNPPEEPVR

38526
CNTQPSPKD

38527
CPEYRMARF

38528
CPPPCSYLL

38529
CPQWCEYYE

38530
CQCDDNWEI

38531
CQERERGKW

38532
CQSECAKRG

38533
CRDCRDGEN

38534
CRDEGLEYE

38535
CRDGDEPRG

38536
CRDWGWICA

38537
CRNDNIERK

38538
CRPLDWKPD

38539
CRYWCSYKD

38540
CWDGCHRRL

38541
CWDHGAPEL

38542
CWEDCQYHE

38543
CYDDWQDRI

38544
DCDMFCMFN

38545
DDARRCHRK

38546
DDCFQRCRN

38547
DDDCNYTRN

38548
DDDEDLNYD

38549
DDDHPRPRS

38550
DDDNWHHKN

38551
DDDPGDAYW

38552
DDECEMYYN

38553
DDEDGGHYM

38554
DDEPKGIRM

38555
DDEVDGDHP

38556
DDGCNYTRN

38557
DDGPDTNRK

38558
DDGPDTYRR

38559
DDGPETYRK

38560
DDGVMDSVD

38561
DDHEEGFKA

38562
DDHQENTKT

38563
DDKGPRNSM

38564
DDKHDRIHH

38565
DDLKCWNNY

38566
DDMHDNMVH

38567
DDNCNYTRN

38568
DDNHPTPTL

38569
DDNREPQDT

38570
DDNWDVFRD

38571
DDPCVEFET

38572
DDPFDVTLQ

38573
DDPFWCQRL

38574
DDPQEVHGF

38575
DDPQRKGYC

38576
DDPWGDRHT

38577
DDQLEGWID

38578
DDQPDNNIY

38579
DDQRLLDDI

38580
DDQTDYPPT

38581
DDQYRIEQH

38582
DDRANVPKH

38583
DDSGPKSQI

38584
DDSIEHFKG

38585
DDSWPIPLG

38586
DDTEPMMNK

38587
DDTEPMPVR

38588
DDINEIEEY

38589
DDWKGWLRF

38590
DDWKPCEFA

38591
DDWMRPKRC

38592
DDYGHDDKN

38593
DECSESMRN

38594
DEGEEEPKL

38595
DEHKDPDEA

38596
DENKDEFWM

38597
DEQCRTHRK

38598
DEQHGGEMD

38599
DESCEWMNQ

38600
DESEERIQD

38601
DESSDDDEE

38602
DGCFDAQIN

38603
DGDHDARIH

38604
DGETDELRM

38605
DGFLDGEWW

38606
DGFMDGEWC

38607
DGKTSREVC

38608
DGPKDRIFN

38609
DGPVPWMMC

38610
DGSDGMSTN

38611
DGVFPRCGN

38612
DHCKWWLDV

38613
DHDCKCEEA

38614
DHDGDVPVF

38615
DHHCDRPQG

38616
DHQCEFPMV

38617
DHTCNWPPN

38618
DICFRVSMM

38619
DIEDTQMSW

38620
DIEQWIAAM

38621
DKDFWSAPN

38622
DKHLEWPNH

38623
DKKNECNWH

38624
DKTCDCEMT

38625
DLSDQYPEL

38626
DLWCMDAEW

38627
DLWTHWNPC

38628
DMDEGSSSN

38629
DMDKDGPRE

38630
DMWDWTYRM

38631
DNDTWQPRI

38632
DNEWDCAGN

38633
DNHREAWQD

38634
DNKRRVPEC

38635
DNQCEDPKP

38636
DPEMEYMGM

38637
DPKPRHDND

38638
DQDICGWVW

38639
DQDRRCDQP

38640
DQECDDHGI

38641
DQHGWRRPG

38642
DQKCERGRI

38643
DQNHRVDRH

38644
DQRHDHRQM

38645
DQSSGWPRD

38646
DQWNDECFF

38647
DRERWWPET

38648
DRKFERQTM

38649
DRKKPVPEW

38650
DRNQWLPRH

38651
DRPRPYCRN

38652
DRQYPPPNH

38653
DRSPWCPQF

38654
DRSQRCNRG

38655
DRTGRCMPP

38656
DRYLGWQFC

38657
DVDCCCVYN

38658
DWDCKTSRN

38659
DWKPADNKC

38660
DWNSDWPEY

38661
DWQRHPPKC

38662
DWRPPCFRF

38663
DWSKEIMGM

38664
DWTCNGHRH

38665
DYDWSDMKC

38666
DYENDEDGV

38667
DYKCCRPEY

38668
EDDFKPARD

38669
EDEGDLWFY

38670
EDEPDGKMG

38671
EDEQSGERE

38672
EDGHKVGFM

38673
EDHDIPRRN

38674
EDNRWIPPL

38675
EDPGDPDMM

38676
EDPHEEFDR

38677
EDQCCFELP

38678
EDTQPLFRH

38679
EEAHRGERC

38680
EEDHEQMCN

38681
EFDHWTMRM

38682
EFEHELDIW

38683
EFPVPLDFW

38684
EGAEGWTQD

38685
EGDPPMPVF

38686
EHDCQDDVN

38687
EHEWDAPSW

38688
EHKYRDMKH

38689
EHPMELEFC

38690
EIDFGTPET

38691
EIDYWWCPH

38692
EMDLCHYSN

38693
EMHPCFWIF

38694
EPSRCRPHV

38695
EQKQEPEFW

38696
ERCEGIQFC

38697
ERDAGDTIN

38698
ERHGEEIRD

38699
ERHYDIMLY

38700
ERKFYENRM

38701
ERQKDRLPD

38702
ESRYCMWGM

38703
ETRELITMM

38704
EVDEDPAFW

38705
EVDFCSAIM

38706
EVEHDVAVW

38707
EVTMGCMIN

38708
FDCDDVIER

38709
FDDCDAGRG

38710
FDDDDKAKC

38711
FDDYDLTVE

38712
FDETDFEAM

38713
FDFPDCHGW

38714
FDHDDEHKC

38715
FDHHEKWNN

38716
FDHRPNCHD

38717
FDNPDQRNA

38718
FDNRDSDKG

38719
FDNRRHWWE

38720
FDPFQFCVG

38721
FDPHHAKQQ

38722
FDPIYEMVE

38723
FDSFDEHHN

38724
FDWISKCSW

38725
FDWQWEDKC

38726
FDYKWMNAY

38727
FEDWGWPKG

38728
FEHRDNWKM

38729
FERWDRKEH

38730
FERYDKPYM

38731
FGEMDGGCY

38732
FGGCDGTPW

38733
FGWLDCCNQ

38734
FGWPDEQQW

38735
FHETQTPKT

38736
FKACDSECD

38737
FKCGNWGII

38738
FKDYRCGWC

38739
FKEGDAKKD

38740
FKGCSGELV

38741
FKMCPCVKG

38742
FKMSDILPH

38743
FKPCMCDPM

38744
FKPDPCFDD

38745
FKPTNIEKG

38746
FKWWDTMNQ

38747
FKYTDHGIQ

38748
FLNDWSNWF

38749
FNPFDCDIG

38750
FNTFDCEIG

38751
FRCNERRWN

38752
FRCYPNQRD

38753
FRDRDGNRM

38754
FRGGEMAKN

38755
FRNENEFEP

38756
FRPKCGPIH

38757
FRPKECALQ

38758
FRQCGEEYH

38759
FRQLDRERQ

38760
FRQWQTMET

38761
FRSEEKAEN

38762
FRSRNPPKH

38763
FRSYGIRLI

38764
FRTWPDDET

38765
FWEHADEEG

38766
FYDGPREMW

38767
FYVGPAMPM

38768
GADWDEPKE

38769
GDCIDGKPH

38770
GDDHDIEFM

38771
GDDHKSHRK

38772
GDDLDCWKN

38773
GDEDHGSKM

38774
GDELHFLRM

38775
GDERPGRRN

38776
GDLCRYWRD

38777
GDPDDDEIW

38778
GEAIDNEKW

38779
GEEEDRMIH

38780
GESCDEKSD

38781
GIPCWQDEW

38782
GKPEDGFDD

38783
GPMRRRINI

38784
GRCQPWYFH

38785
GRDCDEAPD

38786
GRDFYWKIE

38787
GRECSAQRM

38788
GRHGDDLEV

38789
GRPQCWYKH

38790
GRTCDYHDM

38791
HDATPTPRY

38792
HDCCEWWRA

38793
HDCWAEWIA

38794
HDDGDGFQF

38795
HDDIEAIRQ

38796
HDEGPEFKD

38797
HDELWVDRM

38798
HDESGEWLE

38799
HDHKWKPPE

38800
HDKWENPGQ

38801
HDLQPGMNN

38802
HDPCTQIKP

38803
HDPWNVLDW

38804
HDYEPKCMG

38805
HEFKDDMPH

38806
HENKEEQKD

38807
HEQWDNIPC

38808
HFECRTMPN

38809
HFEFWNGGM

38810
HGDHRGNRE

38811
HGSMDSVYW

38812
HHQHNEIEL

38813
HKECPRQRH

38814
HKGFEPERQ

38815
HKPCHGTEF

38816
HMEDWMLEM

38817
HNDTWQPRL

38818
HNEGERWPN

38819
HQCMWDWLT

38820
HRCCETTIN

38821
HRCPIDSRI

38822
HRECGVWDF

38823
HRSDDDQRF

38824
HRSDPPQKM

38825
HTCRLDVNM

38826
HVEEWAEKM

38827
HWSIDAPRK

38828
IDAKPYGRE

38829
IDAWEEHYE

38830
IDCCSFSPT

38831
IDCQRWCQQ

38832
IDCTPWIQT

38833
IDDDPEPTR

38834
IDDKKWIHA

38835
IDDRIWNCD

38836
IDDWLMDKP

38837
IDEWDCIDS

38838
IDFHDCFTK

38839
IDFNDWIPW

38840
IDGDDEHRA

38841
IDHKEAFFE

38842
IDIHEREHF

38843
IDKWEWNRD

38844
IDLQDCNPW

38845
IDMTDMVLW

38846
IDPEGESIS

38847
IDPFDWEGM

38848
IDPFWIFKF

38849
IDPPEQDIM

38850
IDPYDCDPT

38851
IECNDDCPE

38852
IEKFDDPPA

38853
IEPWEHMSN

38854
IEQKGWRRD

38855
IESDPCVAG

38856
IEWDPWIHL

38857
IEWFRWMMW

38858
IFDPRVKAW

38859
IGENMWIWH

38860
IHPMRIGPW

38861
IHTVPDWVM

38862
IIVEVEICH

38863
IKDGPEIDP

38864
IKFWDGLRD

38865
IKPCDEPVD

38866
IKPIGFWVN

38867
IKVKPDNIM

38868
IKWPDEHIW

38869
INPYGTPLN

38870
IPWCDYFFA

38871
IQDCNSIRY

38872
IQERMSIED

38873
IQLGEFWKE

38874
IQWMDMDPW

38875
IRDFGERRI

38876
IRDIDYLPW

38877
IRDMSCLPS

38878
IRGDEEDRN

38879
IRGDEMLED

38880
IRGREDMLQ

38881
IRHCDGDCE

38882
IRHEDGCTY

38883
IRPCAPARR

38884
IRPDGEELQ

38885
IRPNSSLEL

38886
IRPTEFTPN

38887
IRSCPMKCP

38888
ITCDWYYQM

38889
IVSPADIVP

38890
IWERDPDGE

38891
IYCMPYIRA

38892
KDCNHRAEN

38893
KDDCAAWNC

38894
KDEDDIWID

38895
KDHGHQLPP

38896
KDHLGGKRN

38897
KDHMGCPPH

38898
KDHWHPPKY

38899
KDKMKWLKE

38900
KDMCYLSKK

38901
KDNFWMYKT

38902
KDNGNKMDE

38903
KDRTQIWKQ

38904
KDSPWFERT

38905
KDSQRHDKE

38906
KDVFWENKF

38907
KDWSELYEY

38908
KDYIPQLRG

38909
KDYNDQEEN

38910
KEKIQDYPE

38911
KEWFDWCIT

38912
KPLKCWEPQ

38913
KRDYWCWPM

38914
LCSFDDSII

38915
LCSFGDSIS

38916
LDCMRWGFN

38917
LDCPPAPKM

38918
LDCRPEHPD

38919
LDDEESLVF

38920
LDDWDFKPW

38921
LDECEIECN

38922
LDEFKMEPQ

38923
LDGFDTNLE

38924
LDGTDAVRL

38925
LDHKWPDYI

38926
LDIRWYEHV

38927
LDLWRNWKI

38928
LDPCAWVKV

38929
LDPFAQPFS

38930
LDPTDPGRW

38931
LDRLEAPTG

38932
LDRNRVHRQ

38933
LDSCDPRLK

38934
LDTCEMLKQ

38935
LDTGEMLKQ

38936
LDTKRWPKY

38937
LDWKDKERG

38938
LDYRDIARD

38939
LEAGEKWNF

38940
LEAHEGEKF

38941
LEDLRLPRM

38942
LENKWCYKM

38943
LENRPELND

38944
LESCSPIVS

38945
LEWIHDWYH

38946
LEWYECVVT

38947
LGDCPGEPH

38948
LGLCDREMQ

38949
LGSFDDSIS

38950
LHDWWMREM

38951
LHQRDRIEF

38952
LKCGSLIDV

38953
LKLADWWTT

38954
LKLRDEVNE

38955
LKMERCMIA

38956
LKPFEDLLN

38957
LKQWDPWLC

38958
LLDCRGPEV

38959
LNDHEMPGM

38960
LNRCDAPKL

38961
LNWPDYHCM

38962
LQCREGLPM

38963
LQDEPEPPH

38964
LQDFRFCKQ

38965
LQDPACDPS

38966
LQHRDSVES

38967
LRCKWWRTV

38968
LRDFWECRL

38969
LRDWDGNRM

38970
LREVDYPSM

38971
LREWKWCPD

38972
LRPCEGGQD

38973
LRPDIAWEC

38974
LRPGGGLKK

38975
LRTIWSPRM

38976
LRTNDCNFH

38977
LRVMDPLEG

38978
LRYNDLKIH

38979
LTGCSDEVQ

38980
LWADGWELK

38981
LYQCTDPEK

38982
LYQPGEPKS

38983
MDACEGQNN

38984
MDARDECYQ

38985
MDCRREHRH

38986
MDDFEAIRW

38987
MDDSEVLKY

38988
MDEEFWPFN

38989
MDERDAHEE

38990
MDERWWIKI

38991
MDFEEMWVW

38992
MDHKDGYRQ

38993
MDHLHKPKT

38994
MDNHKKPRA

38995
MDPDPKCGM

38996
MDPSHGKIN

38997
MEGFDGWRM

38998
MEMYRWCRH

38999
MFDCRFPRY

39000
MFDFWNLTM

39001
MGECDWPMH

39002
MGMEDDWKH

39003
MHPWDALDM

39004
MHYFRYCKN

39005
MKDFWTEKW

39006
MMNHHDTRP

39007
MNPMAYTRN

39008
MQPFQMHRG

39009
MRCDGWIQF

39010
MRCDLKKKE

39011
MRDWEASKM

39012
MRQFGCPTE

39013
NDCMHNDRW

39014
NDDEPGWKQ

39015
NDDEPPSRL

39016
NDDFDCQEA

39017
NDDLDCCVK

39018
NDDRCFIRA

39019
NDEIDAQYH

39020
NDEIECIGH

39021
NDMFPSFDC

39022
NDPGNRPRE

39023
NDPRRPDSN

39024
NDRCEKPDI

39025
NDRPEADEH

39026
NDSMGEPKS

39027
NDSYGQPKV

39028
NDTGPHGRH

39029
NDTNRALKM

39030
NDYIDEFPY

39031
NDYVEEHPM

39032
NECCDRHPM

39033
NEDHENPSN

39034
NEDQEEDPC

39035
NEYRPPPQV

39036
NGYFDAMRH

39037
NHCDENWYW

39038
NHCKDWEEH

39039
NKPIWYVEM

39040
NPENHKCRF

39041
NQEKTEPKC

39042
NQEMEYHPS

39043
NQTCDQEEM

39044
NRDWFEWKM

39045
NRDWRFFRI

39046
NREQDELKW

39047
NREWPDYRD

39048
NRGVDRLKW

39049
NRPEGRGEC

39050
NRVLEIERK

39051
NRYFDAMRH

39052
NTDFECVGN

39053
NVCEQDIYE

39054
PDALDPCDP

39055
PDAYDDRNH

39056
PDCEPGNVE

39057
PDDCPWPNN

39058
PDDFDKFMW

39059
PDDFEGGPL

39060
PDDHESPRD

39061
PDDHETPTN

39062
PDDIHRFPK

39063
PDEQSLPFD

39064
PDEVPRCRA

39065
PDKLDRFVG

39066
PDLFEKCRI

39067
PDLRDHWQM

39068
PDNAQGPQC

39069
PDNEDARPK

39070
PDPADKCTY

39071
PDPEGEDEP

39072
PDPGIRWPM

39073
PDPHEDGNP

39074
PDPLGWLAH

39075
PDPPKKYED

39076
PDPPKQYED

39077
PDPQQMSRR

39078
PDPRYWIMN

39079
PDPSDKGTY

39080
PDPSPAKGV

39081
PDPSPYCKS

39082
PDPWPWMDD

39083
PDQDDMMHG

39084
PDQFENSFK

39085
PDQFIRWPM

39086
PDQHDPSNL

39087
PDQSHIDLM

39088
PDRCDLTDF

39089
PDRCNYVRH

39090
PDSGEQPVA

39091
PDSPGVCRC

39092
PDTMWREHG

39093
PDTYHYCEW

39094
PDVNQGPWL

39095
PDVSERMMD

39096
PECGDRGDR

39097
PEDCPDGKA

39098
PEDEWGFEY

39099
PEEDEHDVC

39100
PEEDSPWIM

39101
PEFGWWIAM

39102
PEHHFRWPH

39103
PEHKDHFET

39104
PEKLDLINY

39105
PEKYDIWPV

39106
PEMEDASLG

39107
PENDHGMEA

39108
PESFPAERC

39109
PESNDKRES

39110
PETGKAPHI

39111
PETIRGGKC

39112
PEVNEGIHL

39113
PEYPKKGKC

39114
PEYYDDDHS

39115
PFPRDWHDA

39116
PGAIDWQIM

39117
PGCCDRLVE

39118
PGDFGTAED

39119
PGGMCWWER

39120
PGGNDCLLF

39121
PGPKPWKEI

39122
PGPNDVNHC

39123
PGSHGPPVC

39124
PGTMDPPVN

39125
PGYNRLEPS

39126
PHCEEGPED

39127
PHDFWVIED

39128
PHDNDAYFW

39129
PHDRRKNDN

39130
PHEIEYYMN

39131
PHEWDMCAE

39132
PHNNHWIEP

39133
PHNQWMPKS

39134
PHRHGYRKD

39135
PHRQDRMIW

39136
PHTCCGPEE

39137
PHTDRLDIN

39138
PHWFNWPMC

39139
PHYNTVPEN

39140
PKCSNWLNM

39141
PKCTDWNGV

39142
PKDNPIEPK

39143
PKDTKGPET

39144
PKEQPCAEY

39145
PKEWDKPPH

39146
PKHEDCCPE

39147
PKKYDRDVM

39148
PKPWETSRC

39149
PKQGPAIFN

39150
PKSCGDGND

39151
PKTCNCKVT

39152
PKYHESAKS

39153
PLQDGCEMH

39154
PMHYWKSEM

39155
PMPLCCWSL

39156
PNALDMCVE

39157
PNEPWCPVV

39158
PNESNEIEH

39159
PNNAWIDKQ

39160
PNNEENDAM

39161
PNNNDQDKT

39162
PNPHDRPRW

39163
PNQLDKPAM

39164
PNQRNEIEH

39165
PNWGIEPGF

39166
PQCKPLVPM

39167
PQGCDKYLW

39168
PQHYDKWEI

39169
PQLPELEKW

39170
PQNGHICKG

39171
PQNYDQDKV

39172
PQPNRTSPH

39173
PQSEPQPYD

39174
PQSFWGSEM

39175
PRCDPCMGV

39176
PREGDWEGM

39177
PREMWECQC

39178
PREQRPSKS

39179
PRGLPTPMN

39180
PRNCPEDWS

39181
PRNYEAAEC

39182
PRRMPLVPH

39183
PRSDDDQRF

39184
PRSGDDQRF

39185
PRTDGCPWA

39186
PRYQDRCMW

39187
PTPDGSDKN

39188
PTVPQCPRH

39189
PVPHSCERR

39190
PWFHKRYKN

39191
PWNISCPKE

39192
PWPKWCWPA

39193
PWQFGHAKQ

39194
PWTYRGEDF

39195
PYDQRTDDA

39196
PYGPMCTMG

39197
PYKMNHPRE

39198
PYNCRRCPD

39199
PYNPNVDPH

39200
QDAMDPWRI

39201
QDCVERIKS

39202
QDEIDPSEH

39203
QDEIRMPFN

39204
QDEMDGYDQ

39205
QDFLPVLKW

39206
QDHCEPFEP

39207
QDHKDECNE

39208
QDHLADWRN

39209
QDKHEPGKW

39210
QDKMPCWQF

39211
QDMGEKLTH

39212
QDMHDKEPA

39213
QDPCSWLKN

39214
QDPHHNHRH

39215
QDPQWWMRC

39216
QDRYGVPEN

39217
QDTDEGDQC

39218
QENTPGYEG

39219
QEPTEITPM

39220
QKPSDEKRC

39221
QKQCDRTAW

39222
QMFMPYCPF

39223
QQPWCWWEK

39224
QRCDDWIHS

39225
QRCKPRCPM

39226
QRDPEDFET

39227
QRMHDCCDL

39228
QVECSWDKY

39229
RDAKDGGRP

39230
RDAYWAEDN

39231
RDCFWCIGE

39232
RDCHKGIKN

39233
RDDCWPFNP

39234
RDDLRLWPF

39235
RDDLWLWPF

39236
RDHNEKINY

39237
RDHNEQINC

39238
RDIKPWFEM

39239
RDLEKGDEG

39240
RDPAWIWDY

39241
RDTMFEWKF

39242
REEFPLCNQ

39243
REEKDWFKH

39244
RELDCMWEN

39245
REPHDAKNN

39246
REPYWCLGY

39247
REQLWVHPG

39248
REYPKAHRE

39249
RHDTWHDEG

39250
RKDYWSDDG

39251
RLEWWIHPM

39252
RNDFWPAGN

39253
RNQDWGDRQ

39254
RPEMRCFRN

39255
RPKCNLIEM

39256
RPNCYKAEH

39257
RPWEWWIRK

39258
RRPEWLKNY

39259
SDCHENDPH

39260
SDDWRDFDN

39261
SDENDVDDC

39262
SDENEMEPH

39263
SDEWPYYRQ

39264
SDNREWMRY

39265
SDPCKVLRH

39266
SDSADDPKF

39267
SDSKGRERV

39268
SDWDPSWDT

39269
SDYSELPQH

39270
SEHEPDITN

39271
SEKEGIPRD

39272
SEPPDWNTK

39273
SGPRTWIKK

39274
SHDIPVPIM

39275
SHHNPGPRT

39276
SHYPNRPKG

39277
SKDWPPFKA

39278
SKQYDPPKP

39279
SMPLWQDSN

39280
SMPMWWPKH

39281
SNERACEEV

39282
SQPTGWEEN

39283
SRDGDEPNH

39284
SREFWWDGT

39285
SREWDCGSH

39286
SRPPDWPRT

39287
TDDCYGPKG

39288
TDDHERGDC

39289
TDDNGEIRP

39290
TDDWYGPKC

39291
TDEIPEMRY

39292
TDFSDRLKW

39293
TDHCNWSRT

39294
TDHKDGFRI

39295
TDHNGEIRT

39296
TDNRWMDKW

39297
TDPPDGFDM

39298
TDSCEDCKI

39299
TEFECFWNT

39300
TESLEKGPM

39301
TREGPQLKC

39302
TRLKELLIW

39303
TWDRECEYD

39304
VDEFNVWKH

39305
VDFFDWLDH

39306
VDHITKWKF

39307
VDPCDDGNA

39308
VDQHERARN

39309
VDQRDMATD

39310
VDWFARWDS

39311
VEFLDEIRH

39312
VEKMREWPF

39313
VEPYEGVRM

39314
VLDCDNPVD

39315
VNPMAYTRN

39316
VRDCRWCIV

39317
VREFEEDKN

39318
VRPYADSRN

39319
VRRDDPPRP

39320
VRSCGGHDI

39321
VRSWDTDFE

39322
VYWREEIRI

39323
WDCEDGWQQ

39324
WDDHERGQV

39325
WDDRPALKG

39326
WDDRQCEQH

39327
WDEKPAGQV

39328
WDFCKCADI

39329
WDFNDGITP

39330
WDNFNPPKQ

39331
WDNWDDREP

39332
WDPGEGNNF

39333
WDPLEHRLH

39334
WDPSKVNIM

39335
WDQCGWHRQ

39336
WDRASWLEQ

39337
WDRCDCEIP

39338
WDRCPMPRN

39339
WDSCEGREP

39340
WDSHDFEIH

39341
WDSKRLNPP

39342
WDWIEVYRC

39343
WDYRNLNRQ

39344
WECQSRDED

39345
WEDEPPWKI

39346
WEEQRIPSE

39347
WEEWDMMRC

39348
WELPCEWTY

39349
WEPHPYQTN

39350
WFDSRKEEW

39351
WFECWTYPM

39352
WGCKENWNM

39353
WGDCDPRVG

39354
WGDCDYCEK

39355
WGHKPCKGY

39356
WGPCAFGQE

39357
WGPKPCRGY

39358
WGSCECHKC

39359
WGTKPCKGY

39360
WHANEVLKK

39361
WHDTNLEPH

39362
WHSWWTNIM

39363
WKACPLRET

39364
WKAYPWQPT

39365
WKCAEFRTC

39366
WKCPDKLDN

39367
WKDAEKKDD

39368
WKDHTVPDH

39369
WKDNESSEF

39370
WKDRDQCLH

39371
WKLVDLGIG

39372
WKSCGMDSL

39373
WKTTEAPRD

39374
WLDDRLEFF

39375
WNDYDEGEC

39376
WNPADGKQN

39377
WNPWDQLEF

39378
WPCPWFDHM

39379
WPPEPWCCC

39380
WPPWEWYED

39381
WPSCGCTVE

39382
WPVDSWIEN

39383
WQANHWDRT

39384
WQEDQVGRQ

39385
WQPWRHERH

39386
WQQFWWNEC

39387
WQYYPCIDK

39388
WRAPGPLEN

39389
WRCATIPIA

39390
WRCCQFWII

39391
WRCDDRDNC

39392
WRCPRLIDW

39393
WRDHPEDIQ

39394
WRDMDFYIN

39395
WRDQGFSTN

39396
WREMPMSEL

39397
WRHWDGLNL

39398
WRNQGDLVP

39399
WRNRPCLNE

39400
WRQCEFGVE

39401
WRSDGPIER

39402
WRSEEVEHC

39403
WRSRDHDHP

39404
WRTCDMPYF

39405
WRYAEWWNQ

39406
WRYTERIVH

39407
WNDCEPDTT

39408
WNDTDGNEG

39409
WYDNAGNKN

39410
WYPDEADHA

39411
WYQDWHPRM

39412
YDCCCGWKD

39413
YDCIPRCKP

39414
YDDAEMRGH

39415
YDEMYYCVG

39416
YDHRCYVRG

39417
YDNKPLPKH

39418
YDSCDCSRF

39419
YDTRWQPRC

39420
YDWKAADGW

39421
YDYRSYPVA

39422
YEAKETERP

39423
YEVDEEIKI

39424
YEVKEEPVS

39425
YGCFDRCMR

39426
YGDFEWCKR

39427
YGPEDLHKW

39428
YHDCRLHRP

39429
YKDARWPLW

39430
YKECHWHKE

39431
YKTVDVGPH

39432
YPNKDGERN

39433
YPWFDGWQW

39434
YQYFELPKW

39435
YRHEEYWHN

39436
YRHEPEDEG

39437
YRPGPTIRT

Example 43
AAV5 Variants with Thyroid Gland Tissue Tropism as Identified by Machine Learning

This example describes preferred properties of engineered AAV5 variants that display thyroid gland tissue tropism as identified by machine learning methods. The same bioinformatics pre-process, machine learning methods, and analyses as described in EXAMPLE 19 and FIGS. 19-FIG. 22 were carried out to identify preferred properties of engineered AAV5 variants that display thyroid gland tissue tropism.

Favored biophysical properties and favored amino acid residues at each position in the 581 to 589 region of an AAV5 VP capsid polypeptide, which are associated with a higher probability of thyroid gland tissue tropism are described below. Any of the below machine learning-derived positional amino acid preferences described in TABLE 70 can be present alone or in combination with each other or in combination with any of the amino acid preferences observed and described in EXAMPLE 15, TABLE 29. Listed below in TABLE 70 are a summary of positional features shared between the top important features for thyroid gland tropism extracted from the ML models.

TABLE 70

Machine Learning-Derived Thyroid Gland Tissue Tropism Rules

High mutability at Xaa1

Xaa1 is selected from N

Low surface accessibility at Xaa2

Xaa2 is selected from F, G, or M

High solubility at Xaa3

Xaa3 is selected from F

Low mutability at Xaa3

Xaa3 is selected from Y, F, L, or C

Medium mol mass at Xaa3

Xaa3 is selected from D, E, R, K, V, P,

M, I, L, N, Q, T, or C

Low surface accessibility at Xaa3

Xaa3 is selected from V, I, L, or C

High goldman engelman steitz at Xaa4

Xaa4 is selected from L or V

Low surface accessibility at Xaa4

Xaa4 is selected from V, M, A, G, F, I, or L

Low mol mass at Xaa4

Xaa4 is selected from D, A, G, I, L, or N

High solubility at Xaa5

Xaa5 is selected from C, L, F, M, V, or Y

Or

Low solubility at Xaa5

Xaa5 is selected from D

Low average flexibility at Xaa5

Xaa5 is selected from F, M, or W

Low average flexibility at Xaa6

Xaa6 is selected from F, M, or W

High mutability at Xaa7

Xaa7 is selected from N

Low volume at Xaa7

Xaa7 is selected from P, N, or T

Low average flexibility at Xaa8

Xaa8 is selected from F, M, or W

Low surface accessibility at Xaa8

Xaa8 is selected from M, G, or F

Low mutability at Xaa9

Xaa9 is selected from R, K, P, H, or C

Low hydropathy at Xaa9

Xaa9 is selected from R

TABLE 71 below provides sequences of the 581 to 589 region for variant AAV VP1 capsid polypeptides that exhibited thyroid gland tissue tropism and comport to one or more of the rules provided in TABLE 70. The present disclosure, thus, provides for rAAVs composed of engineered AAV5 VP1 capsid polypeptides having a SEQ ID NO: 2, wherein the Xaa1 to Xaa9 region is any one of SEQ ID NO: 41438-SEQ ID NO: 42437, as disclosed in TABLE 71. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 71

Sequences of the 581 to 589 Region in AAV5 VP1

Capsid Polypeptide that Drive Thyroid

Gland Tissue Tropism

SEQ

ID
581-589

NO
Sequence

41438
AACAVWKGE

41439
AANWFTHDK

41440
ACWLWFYFG

41441
ADSADRVYN

41442
AFHHMMNTP

41443
AFKKPSLFR

41444
AFYNYEIMA

41445
AHAIMMRDF

41446
AHHALSFWR

41447
AHHEVSFWR

41448
AHNIMMRDF

41449
AHTIMMRDV

41450
AHTIMMRHF

41451
AHTIMMRYF

41452
AIHEPAENR

41453
AIHEPAGDR

41454
AIHKPAEDR

41455
AIHVPAEDR

41456
AKIPIDHFA

41457
AKSSIEFSA

41458
AKSSIEHSA

41459
AKSSIEYSG

41460
AMHEPAEDR

41461
AMMKMMKAN

41462
AMMLMMKAN

41463
AMMQMMKTN

41464
AMMQMMRAN

41465
ANERHWTSF

41466
ANVRHWTYF

41467
ARAMYVESF

41468
ARSSIEYSA

41469
ARTMYGESF

41470
ASDTMNCGS

41471
ASYHWTHAS

41472
ASYHWTHVG

41473
ATERHWTYF

41474
ATRDWMSCM

41475
AVDEVGSSN

41476
AVHEPAEDR

41477
AVILSSMYS

41478
AVTVDGNNR

41479
AWMDCPIMC

41480
AYDEKKNQM

41481
AYDKKKNQM

41482
AYDQRKNQM

41483
AYLINMLWM

41484
AYNQKKNQM

41485
CFENLQIMA

41486
CGGQFWQGI

41487
CKDEFIVGN

41488
CKIPIDHFA

41489
CKQKENGQN

41490
CKQQFIGQN

41491
CKQQFTGQN

41492
CKQRFNGQN

41493
CMESYDHGS

41494
CMFGMVNYP

41495
CMFGTVNYT

41496
CMKSFDHGS

41497
CMQKDGNDR

41498
CMRGDKRKR

41499
CMRGDORNR

41500
CMRGDTRNR

41501
CMRKDGNDR

41502
CMRMDCNDR

41503
CMRMDGNNR

41504
CMRMVGNDR

41505
CMRSDKRNR

41506
CMRVDGNDR

41507
CMVDANHVW

41508
CNFNNNDHV

41509
CTLFTNFQY

41510
CTLISNFQY

41511
CVGQFWEGI

41512
CVGQFWKGI

41513
CVGQFWRGI

41514
CVVQFWQGI

41515
DANDMSSDR

41516
DARFGINFG

41517
DARFGLNFA

41518
DARFYIAEG

41519
DARFYISEG

41520
DCVIDWCQM

41521
DDSARYWQV

41522
DDSSWDNLF

41523
DELKMGHNS

41524
DFEYTCNQH

41525
DFEYTCNQS

41526
DFQKPSLFR

41527
DGAHFRSCF

41528
DGIIDWCQM

41529
DGSQAGPDK

41530
DGVIDWCKM

41531
DGVIDWRQM

41532
DGVVDWCQM

41533
DHIACKKLC

41534
DHTACKKVC

41535
DHTACKRLC

41536
DHTGCKKLC

41537
DIFFKCEWV

41538
DIFFKCWIA

41539
DIHHLCNIF

41540
DIHHWCNIC

41541
DIHPWCNIF

41542
DINAKKNIC

41543
DINGKKNIR

41544
DINGKKNIW

41545
DINGKKNVC

41546
DINGRKNIC

41547
DIQHWCNIF

41548
DIYHFNHKS

41549
DIYHWCNIF

41550
DKEDDHRRA

41551
DKIAIDHFA

41552
DKIPIAHFA

41553
DKIPIDHFT

41554
DKIPMDHFA

41555
DKIPVDHFA

41556
DKISIDHFA

41557
DKLPIDHFA

41558
DKMLMVFKR

41559
DKMPIDHFA

41560
DKVPIDHFA

41561
DLHHWCNIF

41562
DLNGKKNIC

41563
DQLKMGHNI

41564
DRCFRDDFE

41565
DRCFRDDMV

41566
DRWTMQHWE

41567
DSGKANTQS

41568
DSKADHFVN

41569
DSKADHGVN

41570
DSKADNCVN

41571
DSKADPCVN

41572
DSKADRCVN

41573
DSKADYCVN

41574
DSMADDPHL

41575
DSMGDDQHL

41576
DSRFNESLG

41577
DTVPSHLGC

41578
DVLKMGHNI

41579
DVVIDWCQM

41580
DWIVDPPKP

41581
DWIVDTAKP

41582
DWIVDTPKA

41583
DWIVDTPKL

41584
DWIVDTPQP

41585
DWMDCSIMC

41586
ECERAKGEW

41587
ECERPKGEW

41588
ECERTKGGW

41589
ECERTKGKW

41590
ECERTKGQW

41591
ECERTRGEW

41592
ECKRTKGEW

41593
EDDGDDNIP

41594
EEIAGHVGA

41595
EFHQDMFEC

41596
EFSGYTNQE

41597
EFYDYEIMA

41598
EFYNYEIMT

41599
EFYNYGIMA

41600
EFYNYQIMA

41601
EHGAVHNAS

41602
EHGAVLNAN

41603
EHHGENHCN

41604
EHMTCMHCE

41605
EIHQDMFEF

41606
EKAMLLNGT

41607
EKAMLMNGA

41608
EKAMVLNGA

41609
EKEDDHRRP

41610
EKFMVMNSC

41611
ELMWWQWDR

41612
EMSSEMEKL

41613
ERFMVMNSF

41614
ERFMVMNSW

41615
ERFMVMSSC

41616
ERGAVHNAN

41617
ERHAFNHCN

41618
ERHGFNHCK

41619
ERHGFNHCS

41620
ERHGFNHWN

41621
ERHGINHCN

41622
ERHGVNHCN

41623
ERQTLAMDR

41624
ESMWYYVMA

41625
ESSWPLQWN

41626
ESTISPNHK

41627
ESVDHWGDW

41628
ETAMGTNLY

41629
ETAMLLNGA

41630
ETAYRENPC

41631
ETGPSMFSF

41632
ETGYGENPC

41633
ETGYQENPC

41634
ETGYRENSC

41635
EVSKFWRHT

41636
EVTVDGNNR

41637
EVVAGHVGA

41638
EYCHWWYKL

41639
EYERLGDSW

41640
EYMNCMHCE

41641
FAEPCMAIW

41642
FCERNMMAQ

41643
FCNRNVNRP

41644
FKFWTDNAD

41645
FKFWTDNTE

41646
FKFWTDNTN

41647
FKSIQCSWG

41648
FKSVQCSWA

41649
FMVEFAYMD

41650
FNQTMIFGA

41651
FNVPDLLGA

41652
FPANYYSNA

41653
FPGNYHSNA

41654
FPKFCKCIS

41655
FSDTMNCGS

41656
FSQTMIFCA

41657
FTRDWMSCM

41658
FVEPCMAIL

41659
FVEPCMAIS

41660
FVEPCMTIW

41661
FVERCMAIW

41662
FVQPCMAIW

41663
GAYFFDKYK

41664
GAYFFDKYS

41665
GAYFFHKYN

41666
GCQGFFNMK

41667
GCQGFFNMP

41668
GDAALYNWF

41669
GDGSWDNLF

41670
GDSSLDNLF

41671
GDSSWDNVF

41672
GDSVLYNWF

41673
GDTSWDNLF

41674
GELKMGHNI

41675
GFTVDGNNR

41676
GGPKAANDW

41677
GIHHWCNIF

41678
GKMVMVFKR

41679
GMRGDKRNR

41680
GRFMVMNSC

41681
GSSWPLLWN

41682
GSYHWTHAG

41683
GTADCRNSS

41684
GTGEWMHMA

41685
GVAKAANDW

41686
GVNVDGNNR

41687
GVPKAANDL

41688
GVPRAANDW

41689
GVPVDGNNR

41690
GVSVDGNNR

41691
GVTGDGNNR

41692
GVTIDGNNR

41693
GVTLDGNNR

41694
GVTVDANNR

41695
GVTVDGNDR

41696
GWMDCSIMC

41697
GYLINMLLM

41698
GYLISMLWM

41699
GYLIYMLWM

41700
GYLTNMLWM

41701
GYRINMLWM

41702
HAGSGSNVL

41703
HAGTGSNVL

41704
HHICFQHCK

41705
HIHHWCNIF

41706
HKISLFKVN

41707
HNLMGWYGP

41708
HQTLDMERF

41709
HQTLDMKKF

41710
HQTLDMKRV

41711
HRCFRDDME

41712
HSAYFRVGA

41713
HTGPGSNVL

41714
HVEYVIWPG

41715
HVEYVIWPV

41716
HVKYV1WPD

41717
HW1SFERHP

41718
HYEICHISK

41719
IADIVWQGQ

41720
IAHPFKCGG

41721
ICAIANTCN

41722
IDDDNWEQP

41723
IDDGDDNIP

41724
IEIALHACW

41725
IFAAPMPGM

41726
IFHSNPVVR

41727
IFHSTPIVR

41728
IFVAPMPGR

41729
IFVASMPGM

41730
IFWMKCGWR

41731
IGNRNVNRP

41732
IINWLYNEP

41733
IISIRMYQA

41734
IKFWTDNTD

41735
IMIEFAYMD

41736
IMLEFAYMD

41737
IMVEFADMD

41738
IMVEFSYMD

41739
IMVEFTYMD

41740
INDCCTMPD

41741
INDCCTMSD

41742
INVPD1LGA

41743
INVPDLLGG

41744
INVPDLLGS

41745
IPKFCKCSS

41746
IPKVCKCIS

41747
IPQFCKCIS

41748
IPWEYAMND

41749
IPWPYAMND

41750
ISDCFTCAD

41751
ISGMKSAMA

41752
ITETPGNTH

41753
ITGAKMYMG

41754
ITETPGNTH

41755
ITGATMYMG

41756
ITGIRMYQA

41757
ITSIGMYQA

41758
ITSIRMDQA

41759
ITSIRMFQA

41760
ITSIRMYKA

41761
ITSIRMYRA

41762
ITS1WMYQA

41763
1TSLRMYQA

41764
ITSNRMYQA

41765
IVGADRCQA

41766
IVNACMAEK

41767
IVQSFWNIP

41768
IVVAVNQSL

41769
IWDFKCNQC

41770
IWEDYLHVG

41771
IWKDYLHMG

41772
IWKDYLHVA

41773
1WKDYLYVG

41774
IYEERNAQM

41775
IYICGPMIN

41776
IYIREPMIN

41777
IYIRGPMLN

41778
IYIRVPMIN

41779
IYLRGPMIN

41780
IYRLREKQE

41781
IYTRGPMIN

41782
IYVRGPMIN

41783
KAFGWFHFD

41784
KAIGWFHFH

41785
KAIGWFHFN

41786
KAIGWFYFD

41787
KALGWFHFD

41788
KAMGWFHFD

41789
KAVAVNQSL

41790
KDAGDDNIP

41791
KDDADDNIP

41792
KDDGDDNIL

41793
KDDGDDNIQ

41794
KDDGDNNIP

41795
KDDGGDNIP

41796
KDGGDDNIP

41797
KDNGDDNIP

41798
KDRMLAGQR

41799
KDSHVHSFF

41800
KDYGDDNEP

41801
KECDPQHWW

41802
KEMRKQGMW

41803
KEYAPQHWW

41804
KEYDLQHWW

41805
KEYDPEHWW

41806
KEYDPQNWW

41807
KEYDPQYWW

41808
KEYDPRHWW

41809
KFSGYTNQE

41810
KFYNYEEMA

41811
KGRWMYHWW

41812
KHCHWWYKL

41813
KIEMLCCPG

41814
KIEMLCCQR

41815
KEEMLCCQV

41816
KIHQDMFEC

41817
KIKMLCCQG

41818
KIMWWQWDR

41819
KKRRDTHIY

41820
KLEMLCCQG

41821
KLYGRRHWH

41822
KMQNENRQM

41823
KNDGDDNIP

41824
KPEWTMMYD

41825
KPQWAMMYD

41826
KPYEDIARM

41827
KQEMQGVCR

41828
KQYDPQHWW

41829
KRETLAMDR

41830
KRFMVMNSC

41831
KRHGFNHCN

41832
KSFYMCQCG

41833
KSFYMCRCA

41834
KSFYMFRCG

41835
KSSMLEHSW

41836
KSTMCDNDA

41837
KTEMSDMNW

41838
KTEMSDMYW

41839
KTYVWEHES

41840
KVLICQNGQ

41841
KVVAVDQSL

41842
KVVAVNRSL

41843
KWEQEPNRN

41844
KWRQAEYDE

41845
KWTRTGSSF

41846
KYCHWWYRL

41847
KYCLWWYKL

41848
KYCNWWYKL

41849
KYRHWWYKL

41850
KYWHWWYKL

41851
LAETPGNTH

41852
LAIALHACW

41853
LAIPQPVNG

41854
LATIMQGFN

41855
LATIMQPFN

41856
LATTMRAFN

41857
LCAEFPHCR

41858
LCAEYPHGR

41859
LCTEFPHGR

41860
LDALPMTQA

41861
LEIAMHACW

41862
LEINWWYPG

41863
LELALHACW

41864
LELNWWYPG

41865
LEVNWWFPG

41866
LEVNWWYAG

41867
LEVNWWYPA

41868
LEVNWWYPV

41869
LEVNWWYQG

41870
LFAEFPHGR

41871
LFTAFRKFT

41872
LFTAFRQFA

41873
LFTAFRQFN

41874
LFWMKCGWR

41875
LGDYVNDEG

41876
LGERTMDRI

41877
LHTIDANRR

41878
LIMREPPFD

41879
LIMRESSFD

41880
LIMRLSPFD

41881
LIVRISPFD

41882
LKNTMWKCA

41883
LKSCIQFSC

41884
LKVNWWYPG

41885
LLKFIHLNE

41886
LLKIYESNQ

41887
LMEFTEDCN

41888
LMEFTEDGD

41889
LMEFTEDGS

41890
LMEFTEEGN

41891
LMEITEDGN

41892
LMEPYGNCD

41893
LMRGTVKNG

41894
LMRSTVRNG

41895
LMSCIKFSC

41896
LMSCIPFSC

41897
LMSCIQISC

41898
LMSCLQFSC

41899
LNGMKSAMA

41900
LNVPDLLGA

41901
LPKFCKCIS

41902
LPRCFIAAC

41903
LPRCVIAAY

41904
LPRYVIAAC

41905
LSCCANVFC

41906
LSCCNNVFC

41907
LSCCTNGFC

41908
LSCCTNIFC

41909
LSDFVNDEG

41910
LSDHSQAAV

41911
LSDYVNEEG

41912
LSDYVNNEG

41913
LSDYVTDEG

41914
LSLSGETQL

41915
LSSCRGMIA

41916
LSYHTPVMA

41917
LTAREKKCG

41918
LTDYVNDEG

41919
LTEIPGNTH

41920
LTEKPGNTH

41921
LTEPPGNTH

41922
LTERPGNTH

41923
LTETPGNTQ

41924
LTQEDFTQM

41925
LTQTPGNTH

41926
LTRADFTQM

41927
LTREDFAQM

41928
LTREDFNQM

41929
LTREDFSQM

41930
LTREDITQM

41931
LTRQDFTQM

41932
LTRVDFTQM

41933
LTSERMYQA

41934
LVGADRCQA

41935
LWCNENAMA

41936
LWCNENAME

41937
LWCNENAMG

41938
LWCNENAMN

41939
LWCNENPMD

41940
LWCTENAMD

41941
LWDFKCTQC

41942
LWGNENAMD

41943
LYAERNAQM

41944
LYEERHAQM

41945
LYEERNSQM

41946
LYEWRNAQM

41947
LYHHDILQT

41948
LYKERNAQM

41949
LYQERNAQM

41950
LYRWEVHNS

41951
LYTEYITKH

41952
LYYEAEHST

41953
LYYIPEHST

41954
MAGYLEKPF

41955
MAVTQSHQF

41956
MFVAPMPGM

41957
MGAYIMWDC

41958
MHVFSNVFG

41959
MIANYEGEM

41960
MIGNYEGEM

41961
MIHLPFGKR

41962
MIHLPFGQR

41963
MIHMPFGER

41964
MIVIYEGEM

41965
MIVNYEGEV

41966
MIVNYEGKM

41967
MIVRCSNTW

41968
MIVSYEGEM

41969
MLEQEPNRN

41970
MLFAENNGL

41971
MSSMLEHAW

41972
MSSMLERSW

41973
MTARRESAY

41974
MTARRESSY

41975
MTARRESTS

41976
MTARRGSTY

41977
MWEKEPNRN

41978
MWEQEANRN

41979
MWEQEPNQN

41980
MWFAANNGL

41981
MWGWPGDSF

41982
NAALYGGWI

41983
NACTGTWWR

41984
NAELFGGWI

41985
NAELSGGWI

41986
NAELYGGWV

41987
NAEQYGGWI

41988
NAEVYGGWI

41989
NAGLYGGWI

41990
NALVLQNVR

41991
NANDMGSDR

41992
NANDMSGDR

41993
NANDMSSDH

41994
NANDMSSNR

41995
NANDMSSYR

41996
NANVLSSDR

41997
NANVMSSDR

41998
NAQHAWENG

41999
NAVLYGGWI

42000
NAYENSHYP

42001
NAYLVHIPR

42002
NAYQCSHYP

42003
NAYQNSNYP

42004
NAYQTSHYP

42005
NCALERMPS

42006
NCAPARMPS

42007
NCAQERMPS

42008
NCLVNLCTY

42009
NCPPERMPS

42010
NCVVHLCTY

42011
NCVVNLCKY

42012
NCVVNPCTY

42013
NEAPMGLKY

42014
NEHPMGLKY

42015
NERPMGLKY

42016
NESPMGLKY

42017
NFWDQVHCV

42018
NFWNQVHCG

42019
NFWNQVHFV

42020
NFWSQVHCV

42021
NGAPERMPS

42022
NGCFVYWWL

42023
NGFMPAVMA

42024
NGRLLWQRY

42025
NGRLWWERY

42026
NGRLWWKRY

42027
NGRLWWQRC

42028
NGRLWWQRF

42029
NGVIINQCF

42030
NGVINDCEF

42031
NGVNMNQCF

42032
NGVSINQCF

42033
NGWLWWQRY

42034
NGYLVDIPR

42035
NGYLVNIPR

42036
NGYLVYIPR

42037
NHADYMRDA

42038
NHADYMRHA

42039
NHADYMRKA

42040
NHADYMRNV

42041
NHTACKKLC

42042
NHVDANLRA

42043
NHVNANRRA

42044
NHVNDNLRA

42045
NHVNTNLRA

42046
NIMSYSSWN

42047
NKMVMVFKR

42048
NLKWDWQNH

42049
NLKWREHHP

42050
NLKWRETHP

42051
NMCEYANVQ

42052
NMCKCANVQ

42053
NMCKFANVQ

42054
NMCKYANVP

42055
NMCKYANVR

42056
NMCKYPNVQ

42057
NMCKYSNVQ

42058
NMCRYANVQ

42059
NMHWQYQFE

42060
NMHWRYKFE

42061
NMIPMDSMP

42062
NMLPFNCNF

42063
NMPWRYQFE

42064
NMYWRYQFE

42065
NNCNMNVMR

42066
NNMHSDWCV

42067
NNMWCQKCP

42068
NNVNANLRA

42069
NPSQMDMDT

42070
NQAHCILNC

42071
NQLQMWYRV

42072
NQTHCIINC

42073
NRFASWEGM

42074
NSDEESGME

42075
NSLHNNEDA

42076
NSLHNNEHP

42077
NSLWCQKCP

42078
NSMWCEKCP

42079
NSMWCQECP

42080
NSMWCQKCL

42081
NSMWCQKCR

42082
NSMWCQKCT

42083
NSMWCQQCP

42084
NSMWCRKCP

42085
NSRHNNEDP

42086
NSSCIQECW

42087
NSSLIQECW

42088
NSSWIEECW

42089
NSSWIQECL

42090
NSSWIQQCW

42091
NSSWLQECW

42092
NSVHNNEDP

42093
NTADCRNSS

42094
NTDEESGMP

42095
NTKYAERSG

42096
NTQFAERSG

42097
NTQYADRSG

42098
NTQYAERCG

42099
NTQYAERPG

42100
NTQYAERSA

42101
NTQYSERSG

42102
NTRFADQTE

42103
NTRFPDQAE

42104
NTRFPDQTA

42105
NTRFSDQTE

42106
NTSWIQECW

42107
NTYQNSHYP

42108
NVFVAYCKY

42109
NVIVAYCKS

42110
NVLAFNCNF

42111
NVLPFNCHF

42112
NVLPFNGNF

42113
NVLPVNCNF

42114
NVLSHTMME

42115
NVLVHTMME

42116
NVMGHTMME

42117
NVNDMSSDR

42118
NVQCNRANG

42119
NVQCTRADG

42120
NVQCTRSNG

42121
NVYVAYCKS

42122
NWADSPWCY

42123
NWEHCWWAH

42124
NWENSPWCY

42125
NWEPCWWAY

42126
NWEPCWWDH

42127
NWEPCWWSH

42128
NWEPCWWTH

42129
NWERCWWAH

42130
NWVDSPWCY

42131
PDRMLAGQR

42132
PHICFQHCK

42133
PHTIMMRDF

42134
PIAKFANMS

42135
PLDWKLNNR

42136
PLDWPLNNR

42137
PLYGRRHWH

42138
PNILMMFEA

42139
PNIVMMFEP

42140
PNMLMMFEP

42141
PSPHEWMAF

42142
PSRPCIYHT

42143
PWAQTEKPN

42144
PWGETEKPN

42145
PWIPNQIKR

42146
PWTETEKPN

42147
QAWWKIFWH

42148
QEMRKKGMW

42149
QEMRKRGMW

42150
QEMRQQGMW

42151
QFSGYTNQV

42152
QFYNYEIMA

42153
QGQWMYHWW

42154
QGRWMYNWW

42155
QGRWMYYWW

42156
QGRWVYHWW

42157
QIHQDMFEC

42158
QIMRISPFD

42159
QIYGRRHWH

42160
QKAMLLNGA

42161
QLYGRRHLH

42162
QMSSEMEQL

42163
QNSNVMGCM

42164
QRHGFNHCN

42165
QSKWYYVMA

42166
QSRAGYMHY

42167
QSVDHWGDL

42168
QVHEWVFPR

42169
QVLICKNGQ

42170
QVLICQNAQ

42171
QVLICQNGL

42172
QVLICRNGQ

42173
QVINCQNGQ

42174
QVVICQNGQ

42175
QVV1DHPKY

42176
QWIRTGSSF

42177
QWTRAGSSF

42178
QYCHWWYKL

42179
RAIGWFHFD

42180
REISVKAVA

42181
REISVQAVA

42182
REMRKQGMW

42183
RFFAAELQL

42184
RFFAAEPQW

42185
RFFAAQLQW

42186
RFFADELQW

42187
RFFAVELQW

42188
RHICFKHCK

42189
RHMCFQHCK

42190
RMATMFTCR

42191
RMHDACFDN

42192
RMHDACYDT

42193
RMHDGCYDN

42194
RMHDTCYDN

42195
RMNDACYDN

42196
RMRDACYDN

42197
RMYDACYDN

42198
RPEYANHMH

42199
RQILDMKRF

42200
RSFYMCRCG

42201
RTEYADHMH

42202
RTKYANHMH

42203
RTYLWEHES

42204
RWFAENNGL

42205
RWRCRMRGN

42206
RYFAAELQW

42207
SANDMSSDR

42208
SANWCTHDK

42209
SANWFAHDK

42210
SASWFTHDK

42211
SATWFTHDK

42212
SCNIVEQVH

42213
SDAVLYNWF

42214
SIDIDYVHL

42215
SEYAAMRDP

42216
SEYAAMRDT

42217
SEYAAMRDY

42218
SEYAGMRDS

42219
SFTLLVGDY

42220
SGCAMHNMG

42221
SGCAMLDMG

42222
SGCAMLHMG

42223
SGCAMLNFG

42224
SGCAMLNMA

42225
SGCAMLNME

42226
SGCAMLNMV

42227
SGCAMLNVG

42228
SGCAMLYMG

42229
SGCAMPNMG

42230
SGCAMVNMG

42231
SGCGMLNMG

42232
SGCVMLNMG

42233
SGDYYDGMP

42234
SGFASWEGM

42235
SGLGMPNWP

42236
SGWAMLNMG

42237
SHMMNHAFW

42238
SHMMSNAFW

42239
SIKMCQSTV

42240
SIKVCESTV

42241
SIKVCQSNV

42242
SIKVCRSTV

42243
SIPSIGSEE

42244
SIVDFESAA

42245
SKSSIEYSA

42246
SLHSIGSEE

42247
SLPSIGNEE

42248
SLPSLGSEE

42249
SLPSVGSEE

42250
SMCKYANVQ

42251
SMIPMDSMT

42252
SMIPMDSVP

42253
SMSPMDSMP

42254
SMVPMDSMP

42255
SNERHWTYF

42256
SNMMSHAFW

42257
SNWWFMFEA

42258
SPQHCIGMP

42259
SRFASWDGM

42260
SRFASWEGV

42261
SSAYYDGMP

42262
SSDFYDGMP

42263
SSDYYAGMP

42264
SSDYYDGMT

42265
STAACRNSS

42266
STADCRNNS

42267
STADCRSSS

42268
STADGRNSS

42269
STADYRNSS

42270
STAGCRNSS

42271
STGDCRNSS

42272
STPDCRNSS

42273
STSADPNCG

42274
STSADPYCG

42275
STSGDPHCG

42276
STTDCRNSS

42277
STTFASSWE

42278
STWDWMSCM

42279
SVIQNEMRI

42280
SVLGHTMME

42281
SVLQNEMRI

42282
SVNMRFAKH

42283
SVNMRFTEH

42284
SVNWFTHDK

42285
SVVQNEMPI

42286
SVVQTEMRI

42287
SWIDMPMMQ

42288
SWTALGSVT

42289
SWTALSSVP

42290
SWTAVGSVP

42291
SWTIVEQVH

42292
SYLINMLWM

42293
SYMMSHAFW

42294
TACAGTWWR

42295
TASAIDHPK

42296
TASGIDHPQ

42297
TASGIDHPT

42298
TASGMDHPK

42299
TAVINDCEF

42300
TAWIYWNEF

42301
TDDGDDNIP

42302
TDRMMAGQR

42303
TFVAPMPGM

42304
TGVINNCEF

42305
TGVLNDCEF

42306
TGVMNDCEF

42307
TGVTNDCEF

42308
TGWVYWNEF

42309
THHELSFWR

42310
TIHEPAEDR

42311
TKVNHIMRP

42312
TNDIWLQFS

42313
TNDIWWQFA

42314
TNDIWWQFC

42315
TNDIWWQFF

42316
TNDIWWQVS

42317
TNDIMNQYS

42318
TNKMLACSS

42319
TQDIYEHMQ

42320
TRFASWEGM

42321
TSEFVAISM

42322
TTEMSDMHW

42323
TTSADPHCG

42324
TTWVYWNEF

42325
TVSGIDHPK

42326
TVVINDCEF

42327
TVWVYWNEF

42328
TYDQKKNQM

42329
VCAEFPHGR

42330
VCAIVNTCN

42331
VCNRNVNRP

42332
VCWLWFYCG

42333
VCWLWFYFA

42334
VCWLWFYFV

42335
VCWVWFYFG

42336
VCWWWFYFG

42337
VFHSNPIVR

42338
VFVAPMPGM

42339
VFWMKCGWK

42340
VFWMQCGWR

42341
VGIPERAMP

42342
VGWLWFYFG

42343
VHKWDTLRD

42344
VHMGEKMWS

42345
WHMGWKMWS

42346
VIARCSNTW

42347
VIIRCSNTW

42348
VILRCSNTW

42349
VIVNYEGEM

42350
VIVRCANTW

42351
VIVRCPNTW

42352
VIVRCSNTG

42353
VIVRGSNTW

42354
VKFWTDNTD

42355
VLVRCSNTW

42356
VMEFTEDGN

42357
VNDCCTMRD

42358
VPDLFNQFC

42359
VPHPFKCGG

42360
VSDFKIYMG

42361
VSDTLNCGS

42362
VSFLPDEYC

42363
VSFLPNQYC

42364
VSYHWTHAG

42365
VTAFKIYMG

42366
VTAREEKCG

42367
VTDFEIYMG

42368
VTGAMMYMG

42369
VTIHERRCN

42370
VTSEWMHMA

42371
VTVHEHRCN

42372
VVICKRFVG

42373
VVTVDGNNR

42374
VWFAENNGL

42375
VWMDCSIMC

42376
VYEERNAQM

42377
VYRLRERQE

42378
VYRVREKQE

42379
WDCLLHKWR

42380
WDCLLNRWR

42381
WDCLVNKWR

42382
WECLLNKWR

42383
WGEACKLDT

42384
WHCLLNKWR

42385
WHGMRMIRA

42386
WHGMRMIRT

42387
WHGMRMLRP

42388
WHGMRMVRP

42389
WKVIYHQPG

42390
WKVIYHQRG

42391
WMEHYGNCD

42392
WMEPYDNCD

42393
WMEPYGNCE

42394
WMEPYGNCN

42395
WMEPYGNYD

42396
WMESYGNCD

42397
WMGPYGNCD

42398
WMKSYDHGS

42399
WMSKQMFKG

42400
WMSQQMFKG

42401
WNCLLNKWR

42402
WSSVAHPGN

42403
WVGQFWQGI

42404
WWAPPWHAL

42405
WWTDMSSWK

42406
WWTDTSSWT

42407
WWTNMSSWT

42408
WYGMRMIRP

42409
YGYLVHIPR

42410
YHEICHISK

42411
YHTACKKLC

42412
YKFWTDNTD

42413
YLKDVDFDS

42414
YPQCVQWFW

42415
YPRCAQWFW

42416
YQHWAMVQR

42417
YQHWTMDQR

42418
YVEYVIWPD

42419
YVSQMWMRC

42420
YVSQVWMQC

42421
YWLLDHANF

42422
YYAICHISK

42423
YYEFCHISK

42424
YYEICDISK

42425
YYEICHICK

42426
YYEICHIST

42427
YYEICHLSK

42428
YYEICNISK

42429
YYEICPISK

42430
YYEICQISK

42431
YYEICRISK

42432
YYEICYISK

42433
YYELCHISK

42434
YYGICHISK

42435
YYKICHISK

42436
YYQICHISK

42437
YYVICHISK

Example 44
Treatment of Rett Syndrome with an AAV5-Derived Virion Encapsidating a Therapeutic Payload

This example describes treatment of Rett syndrome with a variant AAV5-derived virion having any one of the engineered CNS tropic variant AAV5 VP capsid polypeptides disclosed herein, wherein the variant AAV5 virion encapsidates a therapeutic payload. Polynucleotide sequence encoding for AAV Rep, an AAV5-derived variant Cap and helper proteins and a therapeutic payload are transfected in cells to produce variant AAV5 virions, where the polynucleotide sequence encoding for the variant AAV5 Cap comprises at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 in the VP1 capsid polypeptide and where the polynucleotide sequence encoding for the variant AAV5 Cap encodes for an amino acid sequence of any one of SEQ ID NO: 7118-SEQ ID NO: 10117. The therapeutic payload is a guide RNA targeting an mRNA encoded for by a gene implicated in Rett syndrome, a suppressor tRNA targeting a premature termination codon (PTC) in a gene implicated in Rett syndrome, or a transgene. The mRNA targeted by the guide RNA is an mRNA encoded for by a gene encoding for MECP2. The suppressor tRNA targets a PTC in MECP2. The transgene is MECP2. The variant AAV5 virion encapsidating the payload is administered to a subject. The subject is a human or non-human animal. The route of administration is a systemic route of administration. The systemic route of administration is intravenous administration. Upon administration to the subject, the variant AAV5 virions encapsidating the therapeutic payload exhibit enhanced CNS tissue tropism as compared wildtype AAV5 virions encapsidating the therapeutic payload. Upon administration to the subject, a lower dose of the variant AAV5 virions encapsidating the therapeutic payload is administered as compared to wildtype AAV5 virions encapsidating the therapeutic payload. Upon administration to the subject, at least one symptom of Rett syndrome is alleviated or the subject is cured.

Example 45
Determination of Enriched Variants

This example illustrates determination of enriched variants in each of the following tissues: adrenal gland, bone marrow, CNS tissues, colon, heart, lung, lymph node, mammary gland, skeletal muscle or cardiac muscle, sciatic nerve, skeletal muscle, skin, spinal cord, spleen, and thyroid gland.

Capsid variants may be analyzed with respect to their observed abundance. Following filtering as described in EXAMPLE 19, variants are scored based on the number of repeated observations between animals, or tissue sections, or CNS regions, or sequencing samples. This results in a distribution of variant frequency and a variant may be ranked by its observation relative to the distribution of observational frequencies of all other variants through statistical methods.

Additionally, variant “enrichment” within a tissue may be calculated using the following equation, or variations of this equation:

$Enrichment score = \frac{\begin{matrix} variant 1 UMI count in tissue 1 / sum \\ of varients UMI count in tissue 1 \end{matrix}}{\begin{matrix} variant 1 UMI count in all t issues / sum \\ of varients UMI count in all tissues \end{matrix}}$

$\frac{VARjk / \sum_{j = 1}^{n} VARjk}{\sum_{k = 1}^{m} VARjk / \sum_{k = 1}^{m} \sum_{j = 1}^{n} VARjk}$

where VAR is the UMI count for variant j in tissue k for all n variants in all m tissues.

As with observations, this results in a distribution of variant enrichment frequency within each tissue and a variant may be ranked by its enrichment relative to the distribution of enrichment frequencies of all other variants through statistical methods.

The resulting observational and enrichment relative scores may then be summed, or averaged, or otherwise integrated, and variants are ranked by this integrated observation/enrichment score to identify candidates likely to infect a given tissue.

To generate the list of capsid variants present in each tissue based on the integrated observational and enrichment scores, preference was given to variants observed. Variants were ranked by the integrated score and up to 1000 of the top variants were included.

Enriched sequences for each tissue are described in the tables below. Also encompassed herein are rAAVs composed of engineered AAV5 VP2 capsid polypeptides and engineered AAV5 VP3 capsid polypeptides having the sequences disclosed in the below table at the regions in AAV5 VP2 (amin acid residues 445 to 453) and AAV5 VP3 (amino acid residues 389-397) corresponding to the amino acids in the AAV5 VP1 581 to 589 region.

TABLE 72

Sequences of the 581 to 589 Region in AAV5

VP1 Capsid Polypeptide that Drive

Adrenal Gland Tissue Tropism

SEQ
581-589

ID NO
Sequence

3118
KIEYGHWIH

3119
TIDVKRWGQ

3120
CAIPRHVFP

3121
CSYKNQQHQ

3122
AAQKGQHYS

3123
AEMPTVNNI

3124
AERTKELHS

3125
AGWHNTFWG

3126
AHDAYTKNS

3127
ANVHSGQCQ

3128
AQVFSVGWT

3129
ASHRYILAE

3130
ATMTFHFSP

3131
ATVLEASPD

3132
AVHLKAACP

3133
AVMHKNADY

3134
AVMSIPFQS

3135
AVQSCWTGT

3136
CFHRCLFIL

3137
CHLSTCRWP

3138
CIHMLKLAN

3139
CIHQYGRVH

3140
CILRHIKEN

3141
CKMRQRPFR

3142
CLIRNVAGG

3143
CPHKFLMER

3144
CPRCYHRAQ

3145
CSGSKMRMG

3146
CTANRAMML

3147
CTQRRMTFP

3148
CTSYRLSTP

3149
CVMSYINWH

3150
CVNYNLGND

3151
CYKWIDMCN

3152
DHEWYCSLA

3153
DILRYHTQL

3154
DSFQTENSM

3155
DTWIMSQVH

3156
DVKAPHGCV

3157
EDGRADWHV

3158
EHDCYMAVV

3159
ETALLEFRR

3160
EVCQDYQQL

3161
EWKRRLKRR

3162
FCHLIMRYY

3163
FCNYVATDD

3164
FHLRSHWMP

3165
GECRESCVT

3166
GKSDCEHMY

3167
GMHETSVND

3168
GPPLFTMIG

3169
GTTQARIGS

3170
GTVCNNECR

3171
GYIQVHVFN

3172
GYQRAYVPF

3173
HALTWMRAK

3174
HFPYNFWYY

3175
HKIYGSTPY

3176
HSQPDKTGA

3177
HSYNWDEGG

3178
HVMTRNNVY

3179
IIQKTQHNT

3180
IKVQMECHM

3181
ILYNHWVIG

3182
IMEEARTYC

3183
IMSNYCLTA

3184
INKVQTFGL

3185
IRCCTKPLT

3186
KCATYPSHD

3187
KIGMEEVMS

3188
KILIRDCGV

3189
KKKYWKMED

3190
KMLYSSYTQ

3191
KNEWPCIGF

3192
KNKTSEYAC

3193
KTFFHLSFA

3194
KWADQRNSF

3195
LCMLAWIME

3196
LLQKMWAPT

3197
LMNRRADER

3198
LVHCFLKSA

3199
LYCLCKNWC

3200
LYGLKMMYN

3201
MANMVWQNW

3202
MGAMALELA

3203
MSKIKQRLN

3204
MVNRPATCV

3205
MVTPHDHWH

3206
NCDSKVVGH

3207
NCLKEEVVD

3208
NHHCCTPSP

3209
NHRWNFGKS

3210
NLVDCECRV

3211
NTCEGWSHF

3212
PHMPYSDQQ

3213
PQYSVCGGP

3214
PSARSEQQD

3215
QAGGSGFNE

3216
QESAYGRPL

3217
QQNGSNNFM

3218
QSTPTLGPK

3219
QVGTHNHME

3220
QWVHKQNLD

3221
QYADFKAQQ

3222
QYYMTIHHF

3223
RAETTKQLM

3224
RCWITEVSY

3225
RELSVQQTI

3226
RGKGHGTDL

3227
RGLKCYMRN

3228
TDLEQGMDQ

3229
TFDLWQWGH

3230
SCCWRGSME

3231
SHVCQEYMN

3232
SLPQTDQHV

3233
SNAEMSQLF

3234
SNKLMMDVM

3235
SPRCGGPFP

3236
SVCMHEGSQ

3237
SVGWSFEPK

3238
TDLEQGMDQ

3239
TFDLWQWGH

3240
TIEWKPSEL

3241
TYLHQPEDS

3242
TYSNDGLFN

3243
VAKETMNTW

3244
VCQVGFLND

3245
VGQPTEKSF

3246
VHCKVWGQC

3247
VMEGQRGWT

3248
VSAYRCSFE

3249
VVMHCAKEF

3250
VYHRLCRDT

3251
VYHRNATWV

3252
WPEQCNVNL

3253
YAQSVSESN

3254
YCERCQDFH

3255
YDEYPRNAT

3256
YDNYWAYNP

3257
YGVCLWNNG

3258
YETQQYASH

3259
YNPQQGKSK

3260
YPYAILVDE

3261
YWQWNTSGN

3262
YWVPDCYNF

3263
GVHATCMHA

3264
ADRQCLQGP

3265
ASDPNRWAG

3266
ATGTYNLQK

3267
CEHRYSKSS

3268
CIHWYGKSS

3269
CEHWYSKSA

3270
CEHWYSKSS

3271
CEHWYSKSY

3272
CLVSYSQGQ

3273
CLVSYSRGQ

3274
CMHWYSKSS

3275
CMQTDKLMN

3276
CMQTGKLMN

3277
CMQTVELMN

3278
CMQTVKLMK

3279
CMQTVKLMN

3280
CMQTVKLMS

3281
CMQTVKLMT

3282
CMQTVRLMN

3283
CSMARRYWG

3284
CSMTRGYWG

3285
CSMTRRYWG

3286
CSMTRRYWS

3287
CSPGMHLSP

3288
CSPSMHLSP

3289
CSPSMHPSP

3290
CSPSVHLSP

3291
CSVTRRYWG

3292
CTMYRNVSW

3293
CVGWQSTAP

3294
CVGWRSAAP

3295
CVGWRSTAP

3296
CVGWWSTAP

3297
CVHWYSKSS

3298
CVIPPRGCV

3299
CVIPPRGWV

3300
CVQWELNSA

3301
CVQWERNSA

3302
CVQWERNSV

3303
CYPSMHLSP

3304
DHDEIFVAR

3305
DHDEMFVAR

3306
DHNEIFVAR

3307
DIQCANEGY

3308
EGEHCWKWE

3309
EGIHCWKGM

3310
EIDPNRWAG

3311
ESAPNRWAG

3312
ESDPNRGAG

3313
ESDPNRLAG

3314
ESDPNRWAC

3315
ESDPNRWAD

3316
ESDPNRWAG

3317
ESDPNRWSG

3318
FSLFCGQYS

3319
FSLFCRQYS

3320
GICFALNAP

3321
GIQCANEGY

3322
GLTIDHQLV

3323
GLTIDHQLV

3324
GLTTDPQLV

3325
GLTTNHQLV

3326
GVQCANEGY

3327
IICFALNAP

3328
IMLSCHNMK

3329
IMLSCHNMR

3330
EMLSCRNMK

3331
IMLSYHNMK

3332
KGEAMCQSL

3333
KGEAMYQSL

3334
KGEAVCQSL

3335
KGHIMCQSL

3336
KGGAMCQSL

3337
KSDPNRWAG

3338
KSFAGTTHM

3339
KSVAGTTHM

3340
KVEAMCQSL

3341
LEWYPNHHN

3342
LEWYPNHLN

3343
LEWYPNHRN

3344
LEWYPNHSN

3345
LEWYPYHRN

3346
LECFALNAP

3347
MVYCNRKSN

3348
QFHVPQSDS

3349
QFHVPQSGS

3350
QFRVPQSDS

3351
QNNNLYTQV

3352
SAGMLAQMC

3353
SAGMRAQMC

3354
SAGMRTQMC

3355
SAGMWAWMC

3356
SAGVRAWMC

3357
SAVMRAQMC

3358
SEQYVAYSG

3359
SKQYVAYSE

3360
SKQYVAYSG

3361
SKQYVTYSG

3362
TASKYEGRT

3363
TATEYEGRT

3364
TATKYEGRT

3365
TKPSMEVNC

3366
VEANSACQN

3367
VEANSECQN

3368
VEANSEFQN

3369
VEANSGCQN

3370
VICFALNAP

3371
VICFALNAS

3372
VICFALNTP

3373
VSLFCRQYS

3374
WCVHAYWQV

3375
WRVHAYWQV

3376
WRVHSYWQV

3377
YIHWYSKSS

3378
WGQSYNYWP

3379
SIRSIVKGE

3380
GMLEFMNMT

3381
QMYWVQNWE

3382
STDPHTSHH

3383
TLTEVNRLA

3384
SCASCRFPL

3385
ETHWDIDSN

3386
EAAHQTNIW

3387
MGTEWTQSY

3388
QQSFFFGEQ

3389
CVDTNRMWQ

3390
DVCKATTTE

3391
TTYPHNTHG

3392
KGLITENNY

3393
LHKAYVTDS

3394
KQATNNNGW

3395
CFARTLSYD

3396
MPYPLNVKE

3397
EQTHTWLCR

3398
ELKYVMMTA

3399
LARTTTNSE

3400
TLTTDKTYS

3401
TRSYFDQSC

3402
PYIMEFHPF

3403
KIRDCDGNV

3404
EIRQFVKGA

3405
APFFTFTNH

3406
TQCKQLTVM

3407
CQNTAVRAI

3408
KAVLFLKDC

3409
DVTQHTSCT

3410
VCQYCELGG

3411
AVVCKMNYH

3412
PCQQLMCFM

3413
TVNRAMREE

3414
EVLKFCKEW

3415
SMSHNGQCL

3416
TNMLAMRGD

3417
MLAFDPMGR

3418
ASFFLNPLE

3419
QKAIVAQND

3420
ISMRSECED

3421
ISWISHASP

3422
CVEQYVEQG

3423
STTAYHQDY

3424
GWDMLSVAK

3425
MITNNCNIS

3426
TCHPNMSVP

3427
MEHTMDFGV

3428
KEFQLQMEP

3429
TIVVIPIKS

3430
SSIYKEIQE

3431
GTRIYGKVS

3432
FFVLGCQGI

3433
QSGMLTITA

3434
YVDHWTLGD

3435
SRYVNNGTV

3436
AAQYMGIMK

3437
AEDTPYTIY

3438
DCVLTMVDK

3439
MTIKMEQQY

3440
NSQLSMTFG

3441
NMPSLRTRI

3442
EIHMFGQSE

3443
MVMMREPSN

3444
MNANETHGA

3445
WTTSIAVDT

3446
NDVWQPHPN

3447
GLSMVLHFV

3448
RCADNPYFR

3449
AFLHGQMWQ

3450
NANWVEYYD

3451
AIYTTASFD

3452
EAGCKMWPT

3453
DARFDQWHH

3454
ATGTYNLQE

3455
TTTNQFMFQ

3456
VCRLDNGPQ

3457
EYACMLMTQ

3458
YDYWMQAPT

3459
SHAQPIASH

3460
DFNGSNWLP

3461
RVNGSTCRH

3462
HIHMSDRGA

3463
NAYYDSHCE

3464
THVKRFDYE

3465
KKHKGWCRA

3466
MPWLDKPPC

3467
RRTDHGEPI

3468
GNECFHNVE

3469
YPTMTFPEE

3470
KVNCRMIIM

3471
WPNIKHQPI

3472
QCTAHNTCM

3473
DMTLQSVSS

3474
YFPSIQCYE

3475
RYPCCQSPF

3476
WKATPGGQC

3477
GHLENNNEW

3478
MNSFYRAEW

3479
QWFTKGVGE

3480
INYISFVDR

3481
KMAGIGCWY

3482
LEIGGKTRT

3483
NGVGRSGDN

3484
AGMYKCQED

3485
YSDPKNMSI

3486
DVKSWAVCD

3487
TVNWYSGFG

3488
AAAGFSAPS

3489
AAARDRRQT

3490
AACAQVYHS

3491
AACGVLWSE

3492
AAEREKAHG

3493
AAGPITCLD

3494
AAGHHANLG

3495
AAGYARTYD

3496
AANLNEGAK

3497
AANPHMHMR

3498
AASHLIVHE

3499
AATITYAPS

3500
AATMAPWMA

3501
AAVKKLSDD

3502
AAYLPSLVH

3503
ACCRMTTQQ

3504
ACDFIGQNE

3505
ACELQEMAM

3506
ACGQAIHHS

3507
ACIFRHQFY

3508
ACLEVYCQD

3509
ACLSSPIFE

3510
ACMTVYRSC

3511
ACQPSPSYT

3512
ACTQRQEEM

3513
ACYCYFMFI

3514
ACYRQHATK

3515
ADIWIQFDA

3516
ADRLINSSK

3517
ADYQTQTYN

3518
AECLSTKWV

3519
AEQWKLGSI

3520
AERDCEKGP

3521
AERMFHQHF

3522
AFCSMNATS

3523
AFIEQVMNC

3524
AFMAIMWTL

3525
AFQQPHKAN

3526
AGDRYTSED

3527
AGISLWCHN

3528
AGLTMQKCE

3529
AGSSCNTFH

3530
AHAEFAVDP

3531
AHAQYQSVA

3532
AHMPREWMA

3533
AHTAIGSCP

3534
AHVCLSGQW

3535
AHVDYNTCD

3536
AHVGVELAE

3537
AHYHVHSHG

3538
AIAHGSDYE

3539
AIDHRTMCY

3540
AIEKQPNFE

3541
AIESSMHCC

3542
AIGKSNQGL

3543
AIIHQSTAW

3544
AILDQNSLQ

3545
AINTIPMAQ

3546
AISQIMSNE

3547
AITAHLAHD

3548
AITQMEWIN

3549
AIVPESAAQ

3550
AKANRCEAE

3551
AKMPQCAWA

3552
AKMQNMACA

3553
AKNLTMLCR

3554
AKSYANDWA

3555
ALHHFVKAQ

3556
ALLMQTFAH

3557
ALMVFKSMW

3558
ALMVNQGNA

3559
ALNLSWWPM

3560
ALTAITTSS

3561
ALTNHMICP

3562
ALTYRDSMY

3563
AMLKYAYPG

3564
AMSWTQKGG

3565
AMVGYHCRE

3566
ANFPVNTLE

3567
ANGSVNCEE

3568
ANGTLAECG

3569
ANMYSNVTS

3570
ANTSGWTSG

3571
APDQANVYE

3572
APMSYSDMD

3573
APRVHSGNC

3574
APSPKAVYF

3575
AQEFIPSHP

3576
AQGTQCANL

3577
AQKQADINA

3578
AQRMAKIWA

3579
ARELLHWYG

3580
ARMPVSCLP

3581
ARSRTLSDG

3582
ARTECHHCG

3583
ARYKLMPHH

3584
ASCRLAQCC

3585
ASEGAGSWE

3586
ASEPITFVS

3587
ASFGYMIDD

3588
ASGIIKSNG

3589
ASGPIGTAA

3590
ASHGDGVGT

3591
ASKPVELGH

3592
ASLMKPEAV

3593
ASMAYCAQE

3594
ASMKRQENL

3595
ASMTDFVGV

3596
ASMWQHFVI

3597
ASQGVWQSH

3598
ASSTAWQWP

3599
ASTLCNRGQ

3600
ASTWQGHTD

3601
ASVESRIRI

3602
ATAAENIYE

3603
ATAVIANAN

3604
ATCKPDSCD

3605
ATDQFGHHD

3606
ATGKMMFPS

3607
ATHCINRAH

3608
ATIGRSCWD

3609
ATMPWDGNE

3610
ATMQESDRI

3611
ATSPSEHTN

3612
ATSSQVATI

3613
ATTFTRHNV

3614
ATTIWHNHM

3615
ATVFMQETT

3616
ATVHMERDC

3617
ATVMTVHQG

3618
ATYNPGVNW

3619
AVCSMHGTI

3620
AVDIIAWDT

3621
AVEEMVAKG

3622
AVEKCMTLH

3623
AVEPHCFGH

3624
AVGCYTMSG

3625
AVKMFIDRN

3626
AVMAQKIAD

3627
AVQKEYQYL

3628
AVSQFQCSQ

3629
AVVEHAVNR

3630
AVVRSVTRD

3631
AWDSKSNWD

3632
AWEMYHVID

3633
AWGTMNHWL

3634
AWHMYEMSE

3635
AWTEYSLSC

3636
AWYHHCHYD

3637
AYAFVGRDN

3638
AYDRMGGTY

3639
AYEQMPESC

3640
AYHNVKEWN

3641
AYSSTLHAE

3642
AYYEPRGWN

3643
CAAQEQRSR

3644
CAAWRQTQE

3645
CAAWYYGRT

3646
CAFRTAQMA

3647
CALIKHQQP

3648
CAMFRNVAQ

3649
CAMKGNVAQ

3650
CAMLKMKYD

3651
CAMPFRQSV

3652
CAMSRMVFS

3653
CAMWTSASP

3654
CAMYYHRAC

3655
CAQWIKEQQ

3656
CASAFVSYQ

3657
CAVGRHMAL

3658
CAYAYANLC

3659
CCEFKMWRG

3660
CCMWKNSWE

3661
CCNSRNVFP

3662
CCTKLQVLA

3663
CCTLKRCYD

3664
CDGQLDFHW

3665
CFFRVRLCI

3666
CFHIRNIDR

3667
CFHPTDQGW

3668
CFKYTFMQH

3669
CGAKMSRAA

3670
CGTIVHWST

3671
CGTTIAYCP

3672
CHFTQHKSD

3673
CHHWFNHAT

3674
CHLTTNVNE

3675
CIAIITRGP

3676
CIARKMVDD

3677
CIFPRNMAT

3678
CIGSIPMAA

3679
CIHAIPRER

3680
CIHARMVGM

3681
CIHDHKECV

3682
CINWTRKAP

3683
CIRLLCRDK

3684
CIRYEMGAA

3685
CITQRQAMW

3686
CIVLRSMGQ

3687
CIVVYSRSQ

3688
CIWKHSHFV

3689
CKAHYHDNF

3690
CKDAMNTGG

3691
CKFLLNQAR

3692
CKIASNEAF

3693
CKMEFMRVE

3694
CLCLTWAFG

3695
CLGYEKQCP

3696
CLHYTIVNN

3697
CLMEVQWGT

3698
CMDEVHMCT

3699
CMMPYSRTN

3700
CMYQSRAAY

3701
CNTPYWQQA

3702
CPFLMNRAF

3703
CQQRYNVGF

3704
CSAYYMKCS

3705
CSCWYQVQF

3706
CSDNLYCAM

3707
CSECWCKYW

3708
CSHDFTQYQ

3709
CSIQWWQWG

3710
CSKLHDHIN

3711
CSLQQMSIR

3712
CSNMYVASE

3713
CSQPMSMPN

3714
CSQSFQWSK

3715
CSSMWFLWG

3716
CSSPMSLQD

3717
CSTSRMASY

3718
CSVNRHHYW

3719
CSYELITNG

3720
CTFHTMRGH

3721
CTFIRHAMQ

3722
CTFLRSTFA

3723
CTFNQIRGP

3724
CTFQPNRVC

3725
CTGFMGWCM

3726
CTHPNALSP

3727
CTIAIRDSS

3728
CTIMAMQSH

3729
CTIRKQQMT

3730
CTKFEHRSL

3731
CTKHMLQGE

3732
CTMHMDAKY

3733
CTMWCYGCP

3734
CTNVPMKYE

3735
CTRQTTFHG

3736
CTVSKNYGQ

3737
CTYQYQAGM

3738
CTYTPLRAP

3739
CVCYHNSSQ

3740
CVDQHHLTP

3741
CVFAFGMKH

3742
CVFAHNRGF

3743
CVFMDRMAY

3744
CVFPRHHMF

3745
CVGLVMRPQ

3746
CVGPFGSMM

3747
CVHCYNRGY

3748
CVHIRNIGW

3749
CVHTVIRGW

3750
CVIMIGELS

3751
CVIQRMHVY

3752
CVKWCNRCS

3753
CVLASRADR

3754
CVLMRWAWP

3755
CVMPRHRFP

3756
CVMSHLSGK

3757
CVMTYWEAH

3758
CVMVWAEKG

3759
CVNHARQMS

3760
CVNPTTTFG

3761
CVPSYYNNR

3762
CVSYMFSQC

3763
CVISLYGQY

3764
CVVEYPWTW

3765
CVVGKAKLA

3766
CWCCWNNPG

3767
CWEWTMINC

3768
CWNDEPEKC

3769
CWNYVHFAQ

3770
CWRVDRNHS

3771
CWSNVHRGS

3772
CWSVRHAWQ

3773
CWWLWNARE

3774
CYCCLRCHW

3775
CYCTNEHPS

3776
CYPACECTQ

3777
CYVYRHVIC

3778
CYYLSIVVC

3779
DAASDMFDP

3780
DAGYFTLHA

3781
DAIRWNKMV

3782
DALMREAGH

3783
DAMYYNFIT

3784
DAQSSPTAF

3785
DARFKPMMC

3786
DASMYYECS

3787
DATHMMCMH

3788
DCEPREGYW

3789
DFGAEKASE

3790
DFHYNSDWW

3791
DFLIHGYSG

3792
DFMKMQQHG

3793
DFYTMAEHF

3794
DGFKMQFDR

3795
DGTDWDQFS

3796
DHSFHWAYD

3797
DHSHNHNFF

3798
DHVPISTYF

3799
DIERSSRSM

3800
DIKCTMTES

3801
DIMCGQEHK

3802
DITINSFGS

3803
DKCRKDWHF

3804
DKKFLECVI

3805
DKLLMPFDR

3806
DKLYVDVMA

3807
DKMLAMVAD

3808
DKNDKWHAM

3809
DKYLCYTTP

3810
DLYVSQKAH

3811
DMAQPQAAE

3812
DMEWEHCHL

3813
DMFMIHAVQ

3814
DMHMWVHGM

3815
DMMYVRKDA

3816
DMYQFPMHG

3817
DNQMKNEMR

3818
DNRCIIGHN

3819
DNSTQIIGE

3820
DPCWIWKTI

3821
DPGRFQIGN

3822
DPPKEPKEG

3823
DPRDYDHHS

3824
DPTGEGDII

3825
DQGVMRQNE

3826
DQMGHTWQG

3827
DQMIQVEFT

3828
DQQPYVSGL

3829
DQRYGTSFS

3830
DRCYFGMGH

3831
DRNWYRHPG

3832
DRRPVDCMN

3833
DSASIPNYE

3834
DSFRKGTFA

3835
DSFTYTHGW

3836
DSIFPPHTI

3837
DSMLNGMGW

3838
DSQAAPTCP

3839
DSRTVFVDN

3840
DSVDWFTSG

3841
DTFGVACYL

3842
DTGMDAKYP

3843
DTKSYQAPA

3844
DTKWLHWMH

3845
DTTNRNMYS

3846
DVAEAWKHA

3847
DVAWHHWTF

3848
DVDRGRSMS

3849
DVDRIMPQP

3850
DVDWTYMTL

3851
DVHFKMTMA

3852
DVLCLHNHF

3853
DVLFTDVGF

3854
DVTFRWGIC

3855
DVTLMENAR

3856
DVWFNHTVS

3857
DVYPPQGLM

3858
DVYQLHAMQ

3859
DWAKSDRIT

3860
DWFIQYHRG

3861
DWSWYFQGS

3862
DYAAISSMA

3863
DYAKQDSSC

3864
DYNLAGLLP

3865
DYRQWDQQY

3866
DYVQVNQQP

3867
EAASADCQD

3868
EAAYGCKIG

3869
EADKYPYCP

3870
EADWNIGDT

3871
EAERIVRGP

3872
EAFCLHGWA

3873
EAGFWTQQA

3874
EAGSFQHCA

3875
EAHNVEDNP

3876
EAHPHWWVT

3877
EANDSLKAN

3878
EANLSNVRA

3879
ECFPSIQNL

3880
ECGLSHDCG

3881
ECHVRDSHL

3882
ECIKQSECA

3883
ECIQVVRGS

3884
ECKDVEDIF

3885
ECMLCPEAS

3886
ECQTSPVAY

3887
ECRAGDPVN

3888
ECSLKDHHS

3889
ECTANTHYY

3890
ECWMQNEHA

3891
EDCALHQSE

3892
EDHWLFWKQ

3893
EETILIDCH

3894
EEVMLGCLQ

3895
EFAADAHGM

3896
EFAGITDMG

3897
EFCEYAQAT

3898
EFHQQVNTW

3899
EFMHAYEAT

3900
EFSECHAFD

3901
EFSNPCSFG

3902
EFVKAFMQI

3903
EGHGAVLNP

3904
EGKWIGLQT

3905
EGMKSWNST

3906
EGRSSENND

3907
EGSPYFMAK

3908
EHALEHTQL

3909
EHELLKQYE

3910
EHGMILRLG

3911
EHLFLIHEA

3912
EHRQKMAST

3913
EHRTAFFYS

3914
EHVFEEVHS

3915
EIDNVCRQA

3916
EIGCAHAEE

3917
EIGRNCALF

3918
EILLSMMEQ

3919
EILQPQVMI

3920
EIMCKGTMG

3921
EIQQVGRDF

3922
EIRHWGWLV

3923
EISNVRCAP

3924
EIVPTMHCK

3925
EIVYQHISQ

3926
EKHNFTQMV

3927
EKKFNHQIN

3928
EKLYSVSVA

3929
EKRLESAGW

3930
EKSNLFTVR

3931
EKYMYHDVQ

3932
ELDFIIMPG

3933
ELNGAWWKA

3934
EMGAVGSFK

3935
EMHRNELAM

3936
EMMVKPSYC

3937
EMRIQTAQQ

3938
EMVLCVTHE

3939
ENDQIAHEQ

3940
ENGYMHQYL

3941
ENIKTPFGT

3942
ENTYQDTIP

3943
EPKDVSHMT

3944
EPQSFNHWS

3945
EQARIDTEG

3946
EQDQTKTMH

3947
EQELPVTLK

3948
EQPQKPSAW

3949
EQTKVMHCC

3950
EQVSKGVYA

3951
ERGPTDNLH

3952
ERHESHQEF

3953
ERINKFNAD

3954
ERRSAESLQ

3955
ESAQSEGEF

3956
ESDIGHPYK

3957
ESDIMRSPM

3958
ESEQSQDNH

3959
ESGPKWRFP

3960
ESNGFVNGV

3961
ESSRTGSHA

3962
ESTESHFCE

3963
ESTQMCTCC

3964
ESVKFGKMP

3965
ESYTLYQHL

3966
ETACNHTAW

3967
ETASQQHDA

3968
ETCNRGRLF

3969
ETFAMNAAW

3970
ETHFYEVKF

3971
ETLAKGYDW

3972
ETLRMFGVY

3973
ETMPNKLQY

3974
ETNIDNRAM

3975
ETRINIEAH

3976
ETTNAAWFR

3977
ETVSGMHER

3978
EVCFDVCGE

3979
EVCMAVVDN

3980
EVDHQFHFC

3981
EVGPGSHCE

3982
EVNHMMCDM

3983
EVQKCMFVE

3984
EVRWIPWHY

3985
EVSLFSCEF

3986
EVYITGHEI

3987
EWKFCWDAI

3988
EWQHVGATC

3989
EYCRSNFQH

3990
EYESCICAS

3991
EYLRFIAHD

3992
FACMRQMAY

3993
FATGKHHYL

3994
FATHTYDYW

3995
FAVPSKVTM

3996
FCYRMTDDR

3997
FDMEGFREL

3998
FERTVHTDK

3999
FGDWKHGDG

4000
FGPGSHGNN

4001
FKGPYHYQS

4002
FLFYLNMHE

4003
FLMRRNGAD

4004
FLTERCQTE

4005
FMNHFMMWT

4006
FMQKECVEE

4007
FNADIAHCD

4008
FNMLVLETW

4009
FNSSFMGQA

4010
FPHKWNAMS

4011
FPVDMQWMY

4012
FQGPIAPME

4013
FSAMFRRHC

4014
FSGNLSTYA

4015
FSKLTGQDF

4016
FSLKTESRI

4017
FTDDDVNRF

4018
FTGLCDIWQ

4019
FTRALLDMG

4020
FVRTKEDLV

4021
GADDKGTHC

4022
GADIYEETS

4023
GADKLFMYG

4024
GADNVQKIC

4025
GADPFEGQN

4026
GAEIEQLAF

4027
GAEYDQRVW

4028
GAHFIMSSL

4029
GAQFCTQAR

4030
GARHNGIFT

4031
GARQFNCLE

4032
GASKLSHER

4033
GASRYQWGN

4034
GATDCGSCP

4035
GAVECSEYK

4036
GAVPHPVCA

4037
GAVQKLIQC

4038
GAVSANWQS

4039
GCASIGEHE

4040
GCDWLMCEL

4041
GCERMERVW

4042
GCFFSQRGY

4043
GCFYTQRAY

4044
GCGKLGVDR

4045
GCHQLMNFG

4046
GCIRSISWP

4047
GCIRVDSGE

4048
GCMHYVLTE

4049
GCMQSIHMQ

4050
GCRFVILDH

4051
GCTKYGRHN

4052
GCVVKNTHS

4053
GDKWISIQD

4054
GEMCNVSGN

4055
GEDQRGMEG

4056
GGTEIAMCF

4057
GHAMFNSRD

4058
GHFQTMMAQ

4059
GHITCATMS

4060
GHKQCFHQT

4061
GHNCCHMIE

4062
GHVQESWMW

4063
GIAQKNRNP

4064
GIEHLQNTS

4065
GIMCSGMCP

4066
GIMSVMGIG

4067
GINHRHVSA

4068
GKKDQVKGG

4069
GKTELHVEV

4070
GLGFKHSMG

4071
GLVEIHYTE

4072
GLYPHQAMF

4073
GMAVINEVE

4074
GMGKYNQWW

4075
GMGMVEIVG

4076
GMWVNPMAL

4077
GNQHWDVCG

4078
GQCYCYVQP

4079
GQSGLHVCY

4080
GRIQRMESD

4081
GSAQEPIWA

4082
GSILSIGFH

4083
GSSWIDQQG

4084
GSTRCVKWV

4085
GTEHHPFFD

4086
GTERLPAHQ

4087
GTEYERHTF

4088
GTFWRDSQL

4089
GTGKHAWCY

4090
GTHYDASAG

4091
GTIRLDTQY

4092
GTLEKRAIF

4093
GTLSYMMWW

4094
GTSTIRAEH

4095
GTYIELACF

4096
GVCFKLTDN

4097
GVFQCPQSV

4098
GVHQKNCPE

4099
GVKAREIEI

4100
GVNDLRHVA

4101
GVNHRMDHF

4102
GVRVFMSDV

4103
GVSMCYNWN

4104
GVSSTHQHR

4105
GVVANLNST

4106
GVVKMENMA

4107
GVYTTEQCV

4108
GWEYHPAFG

4109
GWTNYALMT

4110
GYHWTNTAS

4111
GYLPLEIIA

4112
GYMPCMCQD

4113
GYVQDFHGP

4114
HAEQALSVA

4115
HAGSCTLND

4116
HAGSWQYGP

4117
HAMPFCGRS

TABLE 73

Sequences of the 581 to 589 Region in AAV5

VP1 Capsid Polypeptide that Drive Bone

Marrow Tissue Tropism

SEQ
581-589

ID NO
Sequence

6118
DVDVVFRWV

6119
GNMHNFSAW

6120
QVVKFTGYH

6121
VQMNLDAAP

6122
YSFATGNAA

6123
MVDPYGCEG

6124
CEMQVQLCV

6125
AVDHYSNGA

6126
AAQTLECHK

6127
ADHLACALQ

6128
ADSDVHSEP

6129
AHENYRYHQ

6130
AHLNCDKSM

6131
AHRCGCFKC

6132
AILKMTKVL

6133
AKTGGEPDS

6134
AKVNMMKQF

6135
ANNTYPVCA

6136
AQWISPWPT

6137
ASKFEIACF

6138
ATCLIVCEA

6139
ATNNADCGV

6140
CARANCVLS

6141
CEDGPLYID

6142
CMRHGDRCA

6143
CRIDHMETY

6144
CVMEDNEAP

6145
CWEEGMPFY

6146
CWILHHRCG

6147
CYEHDHGQA

6148
CYNDNWVDV

6149
DAHKSMKSQ

6150
DCLHCWDFC

6151
DENWRDTCG

6152
DFLEEHSHM

6153
DFNFLRQQA

6154
DFTQLRMSG

6155
DGSKNQHSY

6156
DIAQLILTT

6157
DIGRNVGEC

6158
DILGKGEPC

6159
DIMSYATGI

6160
DKNYPTHKQ

6161
DLGCVQENF

6162
DQECNEEGF

6163
DTDCHSYMD

6164
DTNDYTSCD

6165
DVERSESRR

6166
DVSYHKKMH

6167
DVVSMKWGV

6168
ELASSWLVI

6169
ELLEFNAMQ

6170
ENAYKTYPL

6171
ERNRLHHRY

6172
ESHALHMCN

6173
ESSYWDKPK

6174
EVVRQXVIC

6175
EYSWSAQYN

6176
FAAMCNKWD

6177
FCSTYNAGT

6178
FEQSEHKPT

6179
FSHAHNWMT

6180
FSHRFDAIF

6181
FSTPARENN

6182
FTAQLAVWM

6183
GDMCEDMCT

6184
GHDFQKHAE

6185
GLHQFSICM

6186
GPAVPEQYG

6187
GRMLVDRHA

6188
GTGPMHHKW

6189
GYDPPEGMV

6190
GYYSMHHEE

6191
HATTVEFCE

6192
HESKTAHRP

6193
HFHQQRKTY

6194
HGEPFCMAP

6195
HHQQCKLDW

6196
HKMECAQSM

6197
HPHQIPIKI

6198
HRPEQLHEG

6199
HSWMRKSQC

6200
HWCIMKGMR

6201
ICEYFRVQA

6202
ECEYRYCEF

6203
IDAWGHFSE

6204
EGYSTWPRE

6205
ELNTCLVDQ

6206
EMDVRVKRS

6207
IMVMINTPY

6208
ENTNYPCIN

6209
EPCILELVV

6210
EPVMPCNCE

6211
IWILRARGD

6212
KDDTQSHMF

6213
KEQRPPNAG

6214
KFVQYQHYV

6215
KGRWIFEYA

6216
KHDGEIRVL

6217
KHEVGQDGG

6218
KEYLNQELR

6219
KLQMGQDNS

6220
KLSDHFMMY

6221
KQNDTVKIP

6222
KTDRMAMMM

6223
LAFTMMWPW

6224
LATFWGHQD

6225
LCAPTQWYM

6226
LCGKAGLYH

6227
LDLCMLFLF

6228
LEMQRGMET

6229
LEDKDEARK

6230
LEKEHSGIQ

6231
LGLTEGQSM

6232
LNGWWDCTR

6233
LPCTSIMWY

6234
LPEHEQQMH

6235
LPENRNKRH

6236
LPKSKYMFQ

6237
LQANEWNMC

6238
LQQPIVHIE

6239
LVMCPPFFG

6240
LYDVVHREW

6241
MGHYCHARR

6242
MHQFRCHYD

6243
MMAPLKVFD

6244
MNMEDRCTF

6245
MNNRWSDCD

6246
MTEPVLLDT

6247
MVQHEEQWE

6248
NCWLPDNPE

6249
NFKQHDTFC

6250
NGFYWVFVM

6251
NLCPNTVDV

6252
NMGCPMMTE

6253
NNSHLYHDN

6254
NWDSPSNKS

6255
NYFPTHAAW

6256
PDQVDRNTP

6257
PEKAKMVWH

6258
PPADPATES

6259
PQTMMCKRW

6260
PSMLWSVVV

6261
QEGNGFHNT

6262
QEKLWEQGS

6263
QEVRSEHYS

6264
QFGPGCFKP

6265
QQIWLCQEI

6266
QQLCVDESH

6267
QRPTCKNTA

6268
QSMVSCYEG

6269
QTRKNEIGI

6270
QWMLRKEVQ

6271
QWTACQATN

6272
QYMLAEDWR

6273
RADGLATVV

6274
RCNMMGKGA

6275
RDRYCDPHQ

6276
RDRYCDPLQ

6277
RKFWLAGKF

6278
RLQIIMYRV

6279
RMKGAKRWY

6280
RNEYQSACW

6281
RRNMPIPSW

6282
RSMYANNDV

6283
RYNNCPNLW

6284
SAMSRAVYY

6285
SEKTQEVCL

6286
SGEGETKMY

6287
SELETPYAN

6288
SLCWEVDLM

6289
SLYETPYHT

6290
SMYTNCTFF

6291
SQKQECGDT

6292
SWLPENPVH

6293
SYAYSHSAT

6294
TNVYQNVEA

6295
TPPCCVIQF

6296
TQEACHHDP

6297
TRSYFDQSC

6298
TSNTFQRHG

6299
TVKYHQDQD

6300
TVNNDPNGE

6301
VCHAFTCNL

6302
VCTIRAECP

6303
VDTIRFPRM

6304
VEKTHNKWM

6305
VESMNHASM

6306
VGECRLKAV

6307
VHYYACHRF

6308
VKEPATYKY

6309
VNAGEVYRS

6310
VPSHMPHHP

6311
VTDAAMFMG

6312
VTNPLEGRE

6313
VVEMNGDDD

6314
VVGVPARAQ

6315
VVQYRLDPP

6316
VYTSTQPQE

6317
WTHTVISDW

6318
WWDHLPELM

6319
YAFNTPWMT

6320
YVEFEDQNG

6321
YVYDAQRQD

6322
YWRYHYYYM

6323
QNKAFLDAV

6324
FDHNAKVLD

6325
YNNIMQIMV

6326
FWLGLFPAK

6327
NNDHYDSES

6328
MCQNVEKEY

6329
QMDWVAECR

6330
MSSRKEFLT

6331
SCRMVYGRA

6332
AVHKQTAAW

6333
DCAWMQKEW

6334
QKNVSTKYD

6335
DYVDQCQVS

6336
TAVLYETEF

6337
VPDYVPMNW

6338
TSRDGGWAY

6339
LPLPMQSMC

6140
DGGRANTLH

6341
QADNGTMRV

6342
MFSHGHVMG

6343
PYHNPWECR

6344
TTYPHNTHG

6345
EQTYYKITW

6346
TSLCAKPHF

6347
FCHEVFSHT

6348
CKVDIHDYM

6349
ETSTHCCKV

6350
PAYRMECCE

6351
GMVELGSCW

6352
EAMTCTKNH

6353
DNVFSNSPN

6354
HILIMQGVS

6355
MHGIARMQQ

6356
HMDHCQCHE

6357
EKFWFKTMN

6358
QLDTYMEGR

6359
NMKPKDVFR

6360
QPGVRTDDY

6361
WCNWQFAQH

6362
GILVQGGSM

6363
ILKQASCKG

6364
AIIQWACWA

6365
WPTMLSAHM

6366
DFNGSNWLP

6367
FTEPMCMQG

6368
KTDKAGLME

6369
KWEQRSKMS

6370
TLHYFHGYQ

6371
MNHQCMLWD

6372
IYCLCQDSD

6373
TLNSRHPEC

6374
ACTKQKTVQ

6375
KTCLCIFTK

6376
SSNFNHHID

6377
QFMMGQQTP

6378
IAVASHAHG

6379
KTGMGDLFL

6380
RITQGESGM

6381
STVGQNHCG

6382
KVNCRMIIM

6383
YVDHWTLGD

6384
TMPDRDPTY

6385
QKAIVAQND

6386
CMEVTCFVV

6387
LESVKMDCN

6388
VLKSNLTGM

6389
AWMCCARHD

6390
RNDNTCHAY

6391
MYDTMGAWC

6392
ILEAIVNFW

6393
AHAWLQHWA

6394
ATGLIDHQW

6395
DVKSWAVCD

6396
PDLWYEKSS

6397
SECEVLCSL

6398
RYPCCQSPF

6399
MNSFYRAEW

6400
RTRISQMFG

6401
RDVGEDNSA

6402
ATGTYNLQE

6403
DQEAQVVTG

6404
PMELWRTWY

6405
ASMHEETCL

6406
DMTLQSVSS

6407
EVAFTTSSD

6408
SPQQPSVFH

6409
KSTYYNLKW

6410
SQERMDYDG

6411
MNLQIGSKG

6412
EIHMFGQSE

6413
MPWLDKPPC

6414
QCTAHNTCM

6415
DQRHGNVSP

6416
EAGCKMWPT

6417
ATMNLSWGL

6418
QAKMILDGT

6419
INYISFVDR

6420
WKATPGGQC

6421
YCECGHWPN

6422
SNVACIQAF

6423
QSRCVDNTV

6424
YSDPKNMSI

6425
ASKQGTHDY

6426
NHSLCWDSK

6427
HSMMPGWPS

6428
EWMQNWISM

6429
AAADVEGYT

6430
AAAPWCEFR

6431
AAAWPEWTM

6432
AACNEDHHW

6433
AACSRHYES

6434
AAELYKCEN

6435
AAFEGEACF

6436
AAFEWMRAP

6437
AAGWRDGRR

6438
AAHCEYSTI

6439
AAKTFYWAR

6440
AALASDHDF

6441
AAQFLHIVK

6442
AASCGYHGW

6443
AASQAQSLT

6444
AAVCTQFSL

6445
AAVGSAQIE

6446
AAWDIELNE

6447
AAWVSGKLY

6448
ACALDGASN

6449
ACEFGSLNQ

6450
ACFHANNWH

6451
ACHVQLSDA

6452
ACKHEISHA

6453
ACLNMWEER

6454
ACPSAELCY

6455
ACTSSQVKG

6456
ACVFKCWWP

6457
ADCQPQSFN

6458
ADWLYMQSC

6459
ADYDMLTAY

6460
AEAMCSQDH

6461
AEIQNWFAK

6462
AEQHTLGMG

6463
AFDTMVCGT

6464
AFFTNMSPS

6465
AFGPYWMEK

6466
AFHLWKEDQ

6467
AFKSFDSHI

6468
AFRAICAHG

6469
AFWIEQMDY

6470
AGDLWEIGS

6471
AGEHSRSER

6472
AGFPFEKFA

6473
AGGHIHADH

6474
AGQFRKLPY

6475
AGWPTSREM

6476
AHMGQVWWF

6477
AHSQLVSDR

6478
AHVLNTFGN

6479
AIEFWDDAL

6480
AIFHACNSG

6481
AIFRSEITY

6482
AIGNLTGAQ

6483
AIQASMDAF

6484
AIRCESDAA

6485
AISVGEVTT

6486
AISVRHQHH

6487
AITDALLYR

6488
AITSASAKN

6489
AIVGWKTPP

6490
AIWIEEFFH

6491
AKAHVCTIP

6492
AKDQLNITR

6493
AKEVTTCNW

6494
AKHIGVTFG

6495
AKKMWHCDQ

6496
AKYGPWENN

6497
AKYQWQSYP

6498
ALAWKNYDK

6499
ALCTENAER

6500
ALGPLHQGD

6501
ALMPSMFLS

6502
ALQQNSLGE

6503
ALVYAHWWG

6504
ALYEADQFD

6505
ALYQIKWGF

6506
AMDVIIEGA

6507
AMEYLRLQH

6508
AMGTTEVQV

6509
AMHLQERMP

6510
AMHNREGAI

6511
AMIQCAMPP

6512
AMWWGQAMG

6513
ANAAIREHL

6514
ANAERCNLA

6515
ANCRKDPMP

6516
ANENCSKGS

6517
ANGPHTDCI

6518
ANILWYHAS

6519
ANKCLDMRD

6520
ANLMSAKME

6521
ANPLCELQQ

6522
APRLTEACL

6523
APSWRNHMC

6524
APVLITNQQ

6525
AQDHRHSDW

6526
AQDWSATQG

6527
AQEKRLYGQ

6528
AQFSHLHSY

6529
AQHSHHECY

6530
AQSMPYSRH

6531
AQWNQLYGS

6532
ARATWPRTA

6533
ARAVDGLNN

6534
ARFIREHEW

6535
ARLWIPGEP

6536
ARRMDDDWI

6537
ASCAFAEQY

6538
ASCTYYHKW

6539
ASEHYMEVF

6540
ASGIEGEQG

6541
ASGWIQAKT

6542
ASIEKAEMH

6543
ASMDHNLDD

6544
ASMTKHNNF

6545
ASNHRLEKY

6546
ASQPFNPWV

6547
ASSARSIFE

6548
ASTAPVADQ

6549
ASTNSVWHV

6550
ASVMQLLCP

6551
ASWLYDNNC

6552
ATAMPQVRY

6553
ATCCYTHFD

6554
ATDDQMIKG

6555
ATDHSHPWP

6556
ATDPCACNI

6557
ATDVVDECH

6558
ATELYMNGH

6559
ATERWLEGN

6560
ATHDAMVTN

6561
ATKCYEYTW

6562
ATMPQECGA

6563
ATNHSHPWP

6564
ATQFGMTED

6565
ATQSVWMLN

6566
ATQTDYNIP

6567
ATSLPAHYD

6568
ATTLHERTH

6569
ATTYWHRGG

6570
ATYYWQFVW

6571
AVELAPCTN

6572
AVFWTHQEE

6573
AVMPNHWQW

6574
AVMYPMRLH

6575
AVNLKQFKT

6576
AVNPAWKHD

6577
AVNWKQDGY

6578
AVRDHELNN

6579
AVSMACLNT

6580
AVVDWQALE

6581
AVVPLMESA

6582
AVYEALRCL

6583
AVYFVPCRP

6584
AWEMEMEYE

6585
AWNPSGFMG

6586
AWIRCIVCA

6587
AWVMSVMLA

6588
AWVNCISGS

6589
AYAETSTHP

6590
AYDNHYLAP

6591
AYQHHEGVV

6592
AYIESQHNL

6593
CAADCGKGW

6594
CADPQERCD

6595
CAQNLNLWL

6596
CASAVQEDC

6597
CASEHSCWH

6598
CATFQNDVL

6599
CCELLIENT

6600
CDGRRLSMH

6601
CDVQQPTMM

6602
CDYHFKNWG

6603
CEPLVEIHG

6604
CFDYMELSP

6605
CFEDCFYCH

6606
CFEPQCVIN

6607
CFVPDTEWG

6608
CGQQNFRIP

6509
CHADLFWEQ

6610
CHNTANHIQ

6611
CHSYFWMPD

6612
CIATQAFMY

6613
CIHQDAPPH

6614
CISQAVMPP

6615
CKERCGYAN

6616
CKLQKCYYM

6617
CLCYFCCDA

6618
CLHCPIRRR

6619
CLSACDLLV

6620
CMMMYNTCR

6621
CMMQCMDMM

6622
CMNPRSYYF

6623
CNAKQQAAW

6624
CNCASRTEF

6625
CNDLRNQWE

6626
CNNPTTRRW

6627
CNWSKSSIA

6628
CPGVLQSCD

6629
CPQQCIWYG

6630
CQVEKVENE

6631
CQVPAWYDP

6632
CRVNDVMQN

6633
CSCTTDTDP

6634
CSDWYVECK

6635
CSEVLEDNA

6636
CSLPHVVHY

6637
CSQRIHSHC

6638
CSSDYSTLS

6639
CTCQAKIFS

6640
CTDRFAIGA

6641
CTFPNVHEA

6642
CTHSAFHYV

6643
CTIWKQCGH

6644
CTMCYYQNN

6645
CTQMVIHQE

6646
CTTAIGPSG

6647
CTVILPKVG

6648
CTWFDCSQE

6649
CVDTHHANV

6650
CVHATFDYT

6651
CVTTDHCAW

6652
CWENTNVTC

6653
CWEVVNTFR

6654
CWFIKRACA

6655
CWYVWDLWP

6656
DACFVNQPN

6657
DACSHTWAD

6658
DAEDTPATV

6659
DAGKFRKYV

6660
DAHQENLNW

6661
DAKDMMMTC

6662
DALENHAAV

6663
DALSYMQME

6664
DANEFILMH

6665
DANNTLQHS

6666
DANYYFVWQ

6667
DAWPNQQMW

6668
DCAQNWSGS

6669
DCEWMSTGA

6670
DCGFQTAHT

6671
DCHKWDTNQ

6672
DCKMHQAAM

6673
DCKWLALYP

6674
DCMQDPAYR

6675
DCMVMSQGH

6676
DCNFRHQDP

6677
DCPEWGVGC

6678
DCQMILGDY

6679
DCQPMEWWE

6680
DCRHFQSQE

6681
DCTEIVSGM

6682
DCTMYCKQS

6683
DCTPYQGGH

6684
DCYSKRFTP

6685
DCYYLKYSA

6686
DDEYNLTGL

6687
DDPNYAWEA

6688
DDQLYAWGM

6689
DDSLLFHMD

6690
DEASDLQYA

6691
DEHPHPEHC

6692
DEIQIGGSF

6693
DEMTFEQLF

6694
DEQHICMQT

6695
DESEMTSGG

6696
DEWWWSREE

6697
DFASRCNYV

6698
DFCFNESYW

6699
DFHCYSTER

6700
DFMTLVTHQ

6701
DFQAYSWNH

6702
DFRHVDKHA

6703
DFTQGMCTP

6704
DGDAQSCAY

6705
DGDCLQRHL

6706
DGFNYELQP

6707
DGGHMMNQD

6708
DGHNFDGHN

6709
DGIWTHKAD

6710
DGQFIFSQN

6711
DGYNMLWDP

6712
DHCEMHDCW

6713
DHFHMELTK

6714
DHQSYYTQK

6715
DIEFYCAYP

6716
DIEMFGSEM

6717
DIEYQSMGS

6718
DIGWLAPTA

6719
DIICDCDMF

6720
DILLDSRHS

6721
DITLIMSVH

6722
DKGQLCSLQ

6723
DKIWIWQAS

6724
DKLARSVSC

6725
DKMELNAHW

6726
DKQQAKDQL

6727
DKQRPMCVC

6728
DKQWFWEGH

6729
DKTSCMSFR

6730
DKTYLTRME

6731
DLAVMGKCT

6732
DLMWYEGAS

6733
DLQTSYHDQ

6734
DMARALKFF

6735
DMCTMAYTG

6736
DMEYSVCHS

6737
DMKMYEFGP

6738
DMLMNLMEP

6739
DMNFTEQYA

6740
DMTWTMGRQ

6741
DNACHTPFW

6742
DNAVLDGVP

6743
DNMPQPVCH

6744
DNRFIQDAR

6745
DNSSEGSWF

6746
DNVYKGPWA

6747
DPCPYIDHQ

6748
DPYPQGRQK

6749
DQADCNWTW

6750
DQASYDIHG

6751
DQCPLRSGT

6752
DQKSQMQTR

6753
DQLEFICWP

6754
DQNSYGCIP

6755
DQYVPATGY

6756
DRAVKFSIL

6757
DRGDQVMQT

6758
DRGFAHLCQ

6759
DRSTQDKFD

6760
DRTVQVRVD

6761
DRTYTYCWS

6762
DRVMMQRND

6763
DRWGMEEHQ

6764
DRWQIPSKL

6765
DSATFKSAD

6766
DSECKNCMC

6767
DSGGNMQVY

6768
DSGKQPADQ

6769
DSHCEVRFE

6770
DSIQRLSAG

6771
DSNTGQKAY

6772
DSVEWTVHE

6773
DSVRMLGKP

6774
DSWTGWNFA

6775
DTCKYLWTP

6776
DTCSRKHWM

6777
DTDEYQKAQ

6778
DTEWKYKHN

6779
DTKMQHTCT

6780
DTKWYPSTG

6781
DTMLGPQMQ

6782
DTMTCNRHQ

6783
DTNHATMAC

6784
DTNITDQWR

6785
DTQYTRDFV

6786
DTSWDRIFV

6787
DTYMEADSI

6788
DVAEFTGVV

6789
DVETTKAIN

6790
DVFKHLQCT

6791
DVGACFIQD

6792
DVGMINGPR

6793
DVLPAVECP

6794
DVLTSQELY

6795
DVMHDSRCS

6796
DVMPIECQT

6797
DVNIVCMDD

6798
DVRSMYVMG

6799
DVTHGDPAK

6800
DVTKHALAP

6801
DVVDGEASQ

6802
DVVHGGTQL

6803
DVVKNQGIG

6804
DVVQNDCEF

6805
DVYWDECVF

6806
DWACQEDAG

6807
DWCWYTCSW

6808
DWNYRHWFN

6809
DWVETAQES

6810
DWYMHDQPM

6811
DYEQSQKWW

6812
DYGTAGYPR

6813
DYIMHHHCG

6814
DYLCINGFS

6815
DYNSKMDFH

6816
DYRPENTGC

6817
DYTTAIYTD

6818
DYVRRAWMP

6819
EAAWVVRKR

6820
EADNTQRSS

6821
EADSKFVDQ

6822
EAFAYDHKD

6823
EAGFDAPGR

6824
EAGIDKCCH

6825
EAHMIGRHT

6826
EAICVEREY

6827
EAKLQQEIE

6828
EAMGCILRG

6829
EAMHGDAIW

6830
EARAHQNEI

6831
EASANWLKC

6832
EASGLHKIF

6833
EASNRMIDN

6834
EATLGGWCD

6835
EATPFVRFR

6836
EATVWTWNP

6837
EAWHFYEWP

6838
EAYVNKDQW

6839
ECDFPMGTE

6840
ECEVSHFID

6841
ECHHLWCDY

6842
ECHQMNEDQ

6843
ECMQRYIGV

6844
ECNQNWDEF

6845
ECQNQFGWM

6846
ECRCIPTVW

6847
ECRSEMGLP

6848
ECTRGELTH

6849
ECVHKHSGS

6850
ECYHMELMT

6851
EDTRFHQWT

6852
EEAEQEEWN

6853
EEMHDDGDA

6854
EESQRDMIT

6855
EFAAEMTPA

6856
EFCKANGPL

6857
EFDGCQYFA

6858
EFEQFTRYA

6859
EFHQITGSS

6860
EFMMPMIYD

6861
EFMPTVDTK

6862
EFQWWNMSK

6863
EFTHWMQMQ

6864
EFTQLAYTY

6865
EGCWHGHTF

6866
EGHAFNTFN

6867
EGKPRLHDA

6868
EGKTDVMSY

6869
EGWTVMVDQ

6870
EHCNHDMNT

6871
EHCQEMRWH

6872
EHEAMLVND

6873
EHEFLGAKS

6874
EHFRACVVF

6875
EHFSTYMRE

6876
EHGYSHTLA

6877
EHQEQQLVH

6878
EHSYIMANP

6879
EHTNLEKYA

6880
EHTTWHQYT

6881
EIATHWQKG

6882
EICHYNEQY

6883
EIDAMIPMT

6884
EIDLVTRWE

6885
EIEEQGWSG

6886
EIFLNPQYY

6887
EIGYRHRDC

6888
EIGYWCGCA

6889
EIHQVTHYE

6890
EKHLLNCDS

6891
EKMVAEFQC

6892
EKMVAEFQC

6893
EKQFGVKNR

6894
EKTWHQRNM

6895
ELATLVDWN

6896
ELAWFMNTE

6897
ELCKTVFLA

6898
ELGAAVMVC

6899
ELGFWHGKG

6900
ELLNPDNPI

6901
ELMLGNMFW

6902
ELMMNLIGM

6903
ELRCRDSDQ

6904
ELRPITWVD

6905
ELTKKVKAS

6906
EMIHRHQVD

6907
EMKKLCDAC

6908
EMKPWCQAL

6909
EMMLVHQPS

6910
EMNAVFSTD

6911
EMTKNDSFM

6912
EMYFMSEGT

6913
ENIQSIADV

6914
ENKEYPDSN

6915
ENMCNQGIG

6916
ENRLCVGTV

6917
ENVVAIQTI

6918
ENYKEIPYG

6919
EPTASKHDR

6920
EPTNIPDGF

6921
EPVRRTVMG

6922
EPWMPSIAL

6923
EPWSIGKAA

6924
EPYCEGVAW

6925
EQETHEALG

6926
EQHVIAMDH

6927
EQIKQLNWA

6928
EQMKYYWFE

6929
EQNEQKDMW

6930
EQVPFCFTE

6931
ERACSMADR

6932
ERCHYPYIK

6933
ERETLGDQR

6934
ERGMQVWPP

6935
ERKYIMQFD

6936
ERRSHTETA

6937
ERYRGVIAC

6938
ESCILEEHM

6939
ESCIMQCMI

6940
ESCLFHMRQ

6941
ESENNKFGH

6942
ESEPNKHNP

6943
ESFPNKIQG

6944
ESGVEDMQN

6945
ESHHKGDHM

6946
ESKSHWTIG

6947
ESKVGYELG

6948
ESLTHEDGY

6949
ESRAVLSMP

6950
ESRLTMTCP

6951
ESVNRYFKQ

6952
ETAYTTRTE

6953
ETDHRKHYC

6954
ETECWHKSH

6955
ETEMICEFQ

6956
ETFACPSCR

6957
ETGMFKFSG

6958
ETGWKQNIE

6959
ETHFCIRNS

6960
ETHQMGMDR

6961
ETILDQHGG

6962
ETISRVGMQ

6963
ETLFNSYGR

6964
ETLPLYWQC

6965
ETNKQLNLW

6966
ETQQNIWSW

6967
ETQSPMCCL

6968
ETSCTCEVC

6969
ETSGQGVLW

6970
ETTWNVICP

6971
EVADPMTWV

6972
EVCPGSIRS

6973
EVERKVNQE

6974
EVGNPTQYD

6975
EVHYCGNCG

6976
EVIMSHLIT

6977
EVKYFWHCA

6978
EVLMDWSWE

6979
EVLQLLKAD

6980
EVLYGGQQW

6981
EVNSGRDWW

6982
EVQGFIDDR

6983
EVQHIHFPY

6984
EVQKILFLQ

6985
EVQRNGIGM

6986
EVSHFKVDE

6987
EVSIQPQMN

6988
EVSYTIGDI

6989
EVVHLMMRG

6990
EVWSPMHNG

6991
EWGWAWLVG

6992
EWKMANVMQ

6993
EWRHAWNCW

6994
EWRMFINTA

6995
EWTPEMNEG

6996
EWYLRHHMI

6997
EWYQIRHID

6998
EYAIKICNH

6999
EYASTACYD

7000
EYDHIDVWL

7001
EYEGYKSAA

7002
EYEPIASGP

7003
EYEWLMAAA

7004
EYFDNRCAQ

7005
EYFHVVGVD

7006
EYFPTARTF

7007
EYKSRGREV

7008
EYMERWFND

7009
EYSDSMVWP

7010
EYVRWNKQC

7011
EYYLLGCAD

7012
EYYRKQCAI

7013
FAASSEIYG

7014
FAAYDMVSE

7015
FACPEALGT

7016
FADPHTWFS

7017
FAEDIHYGA

7018
FAIEPGLCT

7019
FATIVVHEE

7020
FATVSPGWE

7021
FAYENRLCD

7022
FCFIEDLAM

7023
FCFMYPFHW

7024
FCFYNRCWQ

7025
FCHLEQCMI

7026
FCSCCHCRT

7027
FCVINELGT

7028
FCYQKLNYN

7029
FDMQLTAYA

7030
FDQPIQQCP

7031
FEHVIRTMW

7032
FEKWRHPRH

7033
FGAKYHDGQ

7034
FGEPANVNA

7035
FGGLCWLVV

7036
FGGLVIDAS

7037
FGKQRNEDY

7038
FGLWQQAGY

7039
FHAEWKGPP

7040
FHTRVNACD

7041
FICQQGDSL

7042
FIRGSPMNM

7043
FIRMIIVDD

7044
FKAEYQSGH

7045
FLINPAWQG

7046
FLWGIRVLA

7047
FMGKTDVNM

7048
FMLTMDFWA

7049
FMNVYFEHA

7050
FMQVGGINT

7051
FMTSLRDRF

7052
FNCEGQLYE

7053
FNDEYPCSQ

7054
FNDSEPEVI

7055
FNFQKTMDC

7056
FNLEEWWSP

7057
FNLQYAGLQ

7058
FNVPFAKDD

7059
FPRESSNLL

7060
FQAGLANCA

7061
FQFTAMWYM

7062
FQGHSRDGM

7063
FQLYQNHGA

7064
FQMTKQTQS

7065
FQMVEQQPQ

7066
FQNLLDGCW

7067
FQTRPSCSF

7068
FRVRWWHYR

7069
FRWNIDVTY

7070
FSANMNADI

7071
FSDGLNHSD

7072
FSQHFNRYP

7073
FSQLMNTPP

7074
FTFDQPSGY

7075
FTHPSTDQQ

7076
FTILLFVHN

7077
FTLLNSEGF

7078
FTMRYQKLH

7079
FVETTQVMN

7080
FVKSFNNDV

7081
FVQSSCQQS

7082
FVVDKYIYA

7083
FWNNNLFQS

7084
FWRGITCNF

7085
FYRLDRCEM

7086
GACDVLSGE

7087
GADEPQATL

7088
GADYFTECH

7089
GAGHYITAW

7090
GANAMITEE

7091
GAYEYASKM

7092
GCASEHGRS

7093
GCCQGSPFA

7094
GCELIPIRP

7095
GCEMKYQSF

7096
GCLELEHYE

7097
GCTIAEGAC

7098
GCTQGEQAT

7099
GCVGAGCDM

7100
GDCFKQTFH

7101
GDGNRPTDI

7102
GDHCSLSEE

7103
GDHKTIVMF

7104
GDKYCGVHT

7105
GDLFWKKIH

7106
GDYTKMVNQ

7107
GECTYVGGD

7108
GFAQYGMAL

7109
GFEQYDCEM

7110
GFFCFVTQW

7111
GFMEHPNSA

7112
GFMEKCYDG

7113
GFNALQQAP

7114
GFNCEKPDA

7115
GGGHCWQNA

7116
GGHMKMQME

7117
GGRAFAGYK

TABLE 74

Sequences of the 581 to 589 Region in AAV5

VP1 Capsid Polypeptide that Drive CNS

Tissue Tropism

SEQ
581-589

ID NO
Sequence

9118
AAMESCAEI

9119
EKYQIHWDR

9120
VTGTYNLQE

9121
ATGTYSLQE

9122
ATGTYNLRE

9123
ATDTYNLQE

9124
MWTKFNEYG

9125
QTLGGHMWT

9126
TLCHTDNMV

9127
GIAYEVLWD

9128
SRLPVHECP

9129
TIGTYNLQE

9130
MWTEFNEDG

9131
TFVYGERDR

9132
GQQTIEMSY

9133
NYAMRRDSY

9134
KPLASEGQY

9135
AAFVMNGDG

9136
RVAGEGEQP

9137
TCGHTRFFI

9138
ESAEPRMIP

9139
TTSQNSEAM

9140
NTQVISRWT

9141
AGCNTQNGH

9142
VGPLYAYYS

9143
QHSRAYMWY

9144
DIREVSDGK

9145
TMAQKGLWF

9146
QCEYSNCCY

9147
NGTQCLNMD

9148
MTDNNGPLF

9149
CPEHNQVGV

9150
EMYVQRHSL

9151
LINSIVCGD

9152
ACKPCNGWD

9153
PLTIEVNCT

9154
SKGWADLCP

9155
DMTLQSVSS

9156
RVIEMFTGT

9157
RMFWDTSDR

9158
SMQYFFKTE

9159
SFKGEHLQN

9160
TTYPHNTHG

9161
KSTMLEHHD

9162
TCKWYEESE

9163
DICQKQSPN

9164
ILEAIVNFW

9165
DYEVCGNWS

9166
EWNCWWWHD

9167
SPNGRGLCG

9168
TSMEFTQHQ

9169
ASFFLNPLE

9170
METNNCNIS

9171
MAEQMERDF

9172
VLTIMWKNN

9173
IATHNHILN

9174
MYDTMGAWC

9175
CKQLFCELG

9176
MWTEFNENG

9177
ECVSDYACC

9178
AGESQFDCF

9179
RGKPILNQQ

9180
KSAQVYWHP

9181
AIGPALCLT

9182
QNKHNWWIM

9183
MASQGGQVR

9184
CGHLLRRAE

9185
WGDNKNFMG

9186
FVFAMEPGF

9187
TFMFNSEWS

9188
MAQPYGRTA

9189
CKNHYMMAA

9190
NSGVLIGME

9191
LAESKKRNK

9192
AFNGANSWT

9193
YTELMKKAC

9194
ETNDNMILH

9195
QFMMGQQTP

9196
VKAWWHDHQ

9197
GKEAHDQLN

9198
QVHMQRYYG

9199
MFVVNQNWA

9200
ELINDREWG

9201
DVTQTRYCY

9202
QTGVMAAFG

9203
ESRQVQDCY

9204
IMNPTAQNN

9205
CCCVPRSEY

9206
NRIQMTDFQ

9207
QPYWVAVDP

9208
KMLHKVWID

9209
QWMHCQLSR

9210
GHMFETFWG

9211
NCPMYDMIS

9212
CCCQFSFWN

9213
TLEPAYRHE

9214
NLSNLVSCR

9215
DRKQADQIY

9216
QPKMWMMAS

9217
KEFQLQMEP

9218
QMKGATEFC

9219
LETMMRHGE

9220
LCKFANGAA

9221
FAQQIINAS

9222
EMYLKRCCT

9223
EKRLEGWVY

9224
NVWSPAVRD

9225
DEYQQESFQ

9226
EICMRSGFI

9227
AVKAPSQMS

9228
QTDILRDVQ

9229
DNYWEVPFA

9230
INMPVSKDQ

9231
ENQSTMSHT

9232
ENTPYAHIG

9233
LESVKMDCN

9234
WCEAVGDYA

9235
SMSHNGQCL

9236
HAETDHVGL

9237
QCTLEISYV

9238
SGQPVDCMF

9239
AAAWSCMQW

9240
RTVPTQEMS

9241
LTPRSEGLP

9242
GDVSYDAEA

9243
QHKWHDVGV

9244
TAFQSLQQM

9245
REFAISGGN

9246
PHAETHWYT

9247
NLAPMIQVC

9248
STGSSKDQD

9249
HFEVRSVQR

9250
TMRLFWDMG

9251
NFVNHQPLP

9252
GKGQWTAQE

9253
VHGQEDLSL

9254
YESEYVMMV

9255
GVRNALNLS

9256
HMEQCWCNE

9257
MFFPGFVAH

9258
QCFLWNDCQ

9259
LIHWLKHDL

9260
KCAWIEVTP

9261
YPVHMMPPL

9262
PCCWEQSTN

9263
CSMPQQCME

9264
RRTDHGEPE

9265
GSKEEMHTE

9266
SQMKYAHNQ

9267
EFNWLLWSG

9268
AEDTRYTIY

9269
RTAEDCPFQ

9270
AKCTILNFQ

9271
WFSIIMSKL

9272
QTAEWYCGA

9273
MHGIARMQQ

9274
VTKYCDHEQ

9275
DFNGSNWLP

9276
IHFTSQSCL

9277
QVKWTADPD

9278
CIQPTWDSY

9279
DVKAFMGGV

9280
DPGWYGLAP

9281
DHLSNDAEM

9282
NSIGWDSAS

9283
TNFPTSMHG

9284
RQTTYDCLD

9285
SAKCCINAV

9286
VHHKTNRSY

9287
FGSMQANDR

9288
AVETGESAM

9289
MVENGLRLQ

9290
AIEHGAWKC

9291
SMMEQKEQQ

9292
AFLHGQMWQ

9293
IEFTHSRKV

9294
NWKLPTCMT

9295
EHKGCMQAI

9296
ATGTYNLQE

9297
NGNHYNAMR

9298
QHEQSNHIW

9299
IGRLNASTP

9300
MSNLEYYEA

9301
TSCCYESSS

9302
WINEANCLM

9303
VESAMINQA

9304
EPRLYCAQE

9305
WPNIKHQPI

9306
FTHSCCCLS

9307
SFSDTHLGW

9308
SDSKVQWGM

9309
WDGWVITHF

9310
QTNGHPYFP

9311
QTSHQDQDC

9312
HMGIAYNDH

9313
TFVRKPSLM

9314
QQLCYMSDE

9315
KVAEGQNML

9316
LHYRPTEQC

9317
KTGMGDLFL

9318
AQCHDMWGH

9319
ATGLTDHQW

9320
QWAPLQTWA

9321
SPHNCQHMG

9322
ISMRSECED

9323
KIAAWNDFL

9324
RFDQFWDTY

9325
HQCNCLLSQ

9326
MAMWIADTH

9327
AVHTADVCC

9328
SCAPDTQFG

9329
CWILHHRCS

9330
QVGIEFVVC

9331
TIVVIPIKS

9332
MPWLDKPPC

9333
YMSWYQIRN

9334
AVYLDHVES

9335
KACMSHQGR

9336
TSIEGLKTM

9337
AWSYTGACY

9338
QCVDCPPYC

9339
VTCNWCVMP

9340
KCADLIIRS

9341
NVCDNGHEI

9342
MMKRNRFIC

9343
SRVMIHVYV

9344
WKATPGGQC

9345
WYTSIAVDT

9346
DTHHVHGWQ

9347
MGYAKHDSH

9348
GCEEVGRCQ

9349
KTMGIGGGP

9350
RVWSLEWHL

9351
EQPNMGDYG

9352
FGEFTMDEN

9353
KFGFMYNEF

9354
ERFHSYPLG

9355
FLYTEMFQH

9356
GSVMLMVIR

9357
KMAGIGQNY

9358
RTRISQMFG

9359
AQARTIGLG

9360
MIRPSINGS

9361
TKGRNDHME

9362
ATIAQWCVH

9363
RYNNCPNLW

9364
FKIEWSQDI

9365
KKHKGWCRA

9366
GAANRIQSW

9367
NWIGNMQWA

9368
CMEVTCFVV

9369
RVEMGCEPR

9370
SQIQIRQAD

937/
KIGCMYSSQ

9372
MWHELIADQ

9373
PQEAQIGAP

9374
KTQIHMGDE

9375
LPSHCNLGS

9376
KVESTTQLC

9377
AYAMRNYTS

9378
IIRQSEQYH

9379
AHAWLQHWA

9380
HAVMMCPAP

9381
EVAFTTSSD

9382
PLNQEQIPQ

9383
TSVIRLFWI

9384
QFAQLWCAK

9385
GIWVRDTYN

9386
LCTSTERQG

9387
DRVNLMWKD

9388
CYMNFIVDG

9389
QLSKPSVQC

9390
GITMHPMHH

9391
LPPPEEKWL

9392
TTTNQFMFQ

9393
RCGEIMGLD

9394
MQLRIHTNS

9395
ICHIRTRGN

9396
MSGPNDTEF

9397
NTQYSAKND

9398
KQKTQNRNA

9399
TRETMNEWH

9400
ERQAHNHYA

9401
SKGWADPCP

9402
RTEAHLPAP

9403
GNECFHNVE

9404
ECIFAPRMS

9405
YFPSIQCYE

9406
SSIYKEIQE

9407
THVKRFDYE

9408
LYMMALLPT

9409
SGALKNREE

9410
EASIQASWN

9411
ACIPQMHSA

9412
LHSYQWRLG

9413
ACVERLHHK

9414
TVNWYSGFG

9415
IAKQNFMKT

9416
FTQAIHHPD

9417
YHCCFAMQS

9418
NLERCNTMM

9419
HMHNLPVKP

9420
HCWVNEKCE

9421
FADMITNQF

9422
RQAWPKDGM

9423
CSESYRGPD

9424
VLKSNLTGM

9425
NAYYDSHCE

9426
YPAQYTAFS

9427
GLFEGSPGA

9428
VTMIHNAFH

9429
IYMSTGNTP

9430
YIGMKSCSG

9431
MFMTNVNSF

9432
IAQSEADYW

9433
AVHNVKDVH

9434
HIHMSDRGA

9435
GLFHPQFGQ

9436
GPRHIEEDG

9437
AYHCYMSWM

9438
ITASIKDQW

9439
PAHINDQYM

9440
AAMSSSHSY

9441
MNSFYRAEW

9442
QATGMHRWQ

9443
SSRMQMQGP

9444
MEHTMDFGV

9445
YLEMYSKDW

9446
QWFTKGVGE

9447
KNCIMLASN

9448
AYAPSKNAM

9449
GLSMVLHFV

9450
SPQQPSVFH

9451
PGGCYRMMF

9452
KTCLCIFTK

9453
ACHTGQSGM

9454
DIEFLDCEN

9455
WTLWVMKIN

9456
DVKSWAVCD

9457
LCAVCGDCD

9458
DQEAQVVTG

9459
HFQPVFMQP

9460
GHLFNNNEW

9461
YDYWMQAPT

9462
ASMHEETCL

9463
DCVLTMVDK

9464
MLPLEMKNL

9465
IMTIIPYGP

9466
LIEKQRWML

9467
VSVQSRAQL

9468
THSSCWYQD

9469
VFNLMLQDK

9470
HIKVEHEEV

9471
MMLGYMEQD

9472
TQCKQLTVM

9473
QSSHCKQMV

9474
WGGMPNNQG

9475
FLTCDQFEW

9476
RVNGSTCRH

9477
ETHAAMNSM

9478
DAKLTMHSA

9479
RYPCCQSPF

9480
WGENAWHHN

9481
TSNHNMALE

9482
NPGWQTIGQ

9483
CAWHMKQGN

9484
NVMGTQAKE

9485
TYIQYPTNA

9486
DGMHRELYS

9487
QSAHPEPMC

9488
QAKMILDGT

9489
VHVHCMIGS

9490
TAALGHSFC

9491
DTRRNLCCD

9492
ENTMWQSSQ

9493
NQREWHGLA

9494
PDLWYEKSS

9495
ESSQEIKTC

9496
YVDHWTLGD

9497
NHSLCWDSK

9498
SECEVLCSL

9499
DQRHGNVSP

9500
MCFPYSCAA

9501
ASKQGTHDY

9502
GNKHCMLGT

9503
GTKLIWPYQ

9504
QCTAHNTCM

9505
ACDQVNTVQ

9506
IECSPRESI

9507
RCADNPYFR

9508
QESMCNYWQ

9509
AGMYKCQED

9510
SQAHQNRTC

9511
MGRQKDNVY

9512
NANYWFYYD

9513
EFVLGCQGI

9514
QRSVFHQRS

9515
PIWQGYWPW

9516
HSMMPGWPS

9517
GVHHRCRMT

9518
IQAVMASGW

9519
IAKIGNCVW

9520
CVDTNRMWQ

9521
ARWFSTVEQ

9522
VFKPLDKRY

9523
EVRIQCKID

9524
NGVGRSGDN

9525
CVEQYVEQG

9526
LSAQRLVCD

9527
KGNPLDGDT

9528
PVICVWHDN

9529
EIHMFGQSE

9530
YCECGHWPN

9531
TYESLQNSW

9532
INYISFVDR

9533
MWGCKLEVC

9534
CQMLCRPPV

9535
RDQCRMESG

9536
WVHPYSYCA

9537
QSRCVDNTV

9538
GRALGKKQG

9539
SNVACIQAF

9540
HQYVHNWRQ

9541
SVLHVQALA

9542
ATMNLSWGL

9543
WERCLELNQ

9544
TKMIIFDSW

9545
CWECCTDSG

9546
YSDPKNMSI

9547
FPKGSECNW

9548
EAGCKMWPT

9549
KDGRNPHCL

9550
LEIGGKTRT

9551
YKGMEKNPV

9552
DLIVATEWF

9553
WHTIMTYES

9554
DTAFMHHHV

9555
FQWRTEMNN

9556
SGDWDFGEW

9557
MNLQIGSKG

9558
CNGYWMCMS

9559
SQERMDYDG

9560
SVGYIYDQS

9561
GTWSDPWPN

9562
YPTMTFPEE

9563
AAQYMGIMK

9564
HTVMEWSPT

9565
GQHTISHAG

9566
RYQSHCWLH

9567
LSTMFPEQT

9568
NIGVIPHIQ

9569
PWTTPENYT

9570
YNCPWDQQY

9571
QWHDDQWST

9572
DYAQPTGPD

9573
FKHSLREWT

9574
AARMVGEFY

9575
GSMPSTVPH

9576
ALAKFQCWS

9577
AADIFAIND

9578
QHHWKWSTE

9579
AAGAFLSGC

9580
FDVQFMRVD

9581
ALQYCQGKD

9582
NERDQCQWP

9583
CRMITMMNE

9584
EWHARHHYW

9585
STSWLNDLT

9586
DEGYIMFAP

9587
QMVTQLNWE

9588
VVTQYVDAP

9589
TVIRDSRSY

9590
DEAKKSCWQ

9591
AVNEAQDNR

9592
NERSGSVLE

9593
TYASTRNFH

9594
IRRSMQQQN

9595
GFQRTHVDQ

9596
HKHRMCKAR

9597
TAGQQPTFM

9598
VQMNLDAAP

9599
KDHQLWEVW

9600
GDCIIVISW

9601
RDVGEDNSA

9602
PWMHTMLNP

9603
ETSTHCCKV

9604
QVNMFMMPC

9605
NADMSFRNR

9606
FRLDYDYKN

9607
TNAAYEMWF

9608
TTTITNRLC

9609
KIQPMMVKA

9610
LFRAYAWFQ

9611
DVIRDMIAE

9612
ALTHDDGRT

9613
KTYSAMYSP

9614
NCMWISSTQ

9615
TILPKKTLC

9616
VTTHMNCAD

9617
PPRFLQTTG

9618
VSVPKRQGP

9619
VFGSTRSAA

9620
TSDQCGNED

9621
GAKYIQNTD

9622
MRYHWDEAG

9623
SSEYSQFAI

9624
GVFWPDSFR

9625
GAYSLHDWR

9626
MTALPPSKN

9627
HGFKCQYSF

9628
YGGSEHWGP

9629
TTAYHRSDH

9630
NTHTGNDRL

9631
YPQPCSCHS

9632
VSGFDRVAF

9633
SRLYKKRLF

9634
CCKSKEVCA

9635
TYDTRNHCL

9636
SFKSVSQYN

9637
QAAVTKEYW

9638
EIIQAHLIN

9639
KTGATNLAW

9640
GQFRSGAYR

9641
PMMIIPYVD

9642
EPLVCISAG

9643
KGGLSTTNA

9644
KSTYYNLKW

9645
DVGIGVDQG

9646
MCDNFPSAH

9647
IKHITRNTY

9648
GNMHNFSAW

9649
CSIQINNCR

9650
MREKLVHDC

9651
EQPHISIKG

9652
EMSISHGCH

9653
RSDEMYVTP

9654
SFSYDVVMN

9655
GDMCEDMCT

9656
TVTQGSIGH

9657
RKDQSDLWP

9658
MAAITQNVA

9659
NSTPMACME

9660
PLSSEWRWA

9661
CHLKICMHS

9662
DVWWTLTRD

9663
DATTSTECH

9664
AMLQKKRQD

9665
TIAMPSWTY

9666
NGYVVDNHS

9667
GHHQCYKAD

9668
SANQRYHDW

9669
DCAWMQKEW

9670
KVNCRMIIM

9671
KCRYKDCQC

9672
NTSELLMAQ

9673
QGKQLYHMC

9674
NWYTTWIDE

9675
QAGNSMWAK

9676
RMYATEHCS

9677
NDMQMNWVH

9678
MHDGYRGMQ

9679
WVEPEQCFG

9680
TRCCEPYFQ

9681
LSAKQQFFC

9682
ECTKSMQSG

9683
NCHQLGDNM

9684
PCQQLMCFM

9685
YTKYNWKAA

9686
GTIRMPEHR

9687
VCHNPDPFT

9688
SLTQYQWRG

9689
TLNSRHPEC

9690
IMTMQMDTA

9691
YMHFRQYPI

9692
QTMCITIHP

9693
QEAGYTRHN

9694
MNHQCMLWD

9695
PWYNWDRRC

9696
DASQIPITF

9697
IHRVVCRDH

9698
HHMPIWLGL

9699
AEVPYVMQQ

9700
CVLSRSTMR

9701
AKRECTHIP

9702
TESQIPWVR

9703
STRPIACHS

9704
AWKMAHNSF

9705
RNDNTCHAY

9706
IEHCSNRYA

9707
AVEDYWRYF

9708
KHIVGQDGG

9709
ERFKTFCEE

9710
LVQKWMRTC

9711
DARDQQMGT

9712
NMPSLRTRI

9713
GSQQTTLSC

9714
GYFYALFFM

9715
GFRILDACL

9716
ETDCMEGGH

9717
YPDTADIFW

9718
QFMGVSSHS

9719
VHPGFEKLA

9720
NTFEQCTEH

9721
NNRRQFGYV

9722
QQSFFFGEQ

9723
QSMVACYTI

9724
DSMVYNRPQ

9725
TRDAKVANG

9726
SVKDPMDWG

9727
TMTVRMEQP

9728
ACNTLEHMA

9729
KIRDCDGNV

9730
IAYHYDQQN

9731
QTNEEEDGN

9732
NHILTPVVF

9733
QSYPTMYQC

9734
ECGCMYWGY

9735
EGVNRPNMK

9736
HATGLVNFM

9737
NMFQLKPQN

9738
NDVWQPHPN

9739
TQPEDHWLA

9740
IWITCPYAA

9741
TCHPNMSVP

9742
SNVMATKNW

9743
VDRLFRMEY

9744
FNIQCLFSR

9745
PYGPIEYDE

9746
DYFQYNEAP

9747
TTCYFVASQ

9748
AVVCKMNYH

9749
QVSFVGCNH

9750
ATGTYNLQA

9751
RKWMNAKAT

9752
ENHPTWEKS

9753
KAVLFLKDC

9754
ARYMYEKDA

9755
SHAQPIASH

9756
ATGTHNLQE

9757
IPRPFFTHM

9758
SCQLLNDGG

9759
DGEFLHEVL

9760
LFMTYSARN

9761
QVSFNARYA

9762
DILAFDGSC

9763
NTGVKFCAC

9764
IGQYFNFHF

9765
AEHMVVDHE

9766
MQHLPPAMT

9767
RAEQKFVLL

9768
QGKEFICGP

9769
KCNDIVLYH

9770
STWSANRMT

9771
STWSANMIT

9772
QCGCAPMDS

9773
CCLWSYWAF

9774
QFCICCDGM

9775
CVFSRNNYQ

9776
LTCMHASTP

9777
GYAQRGFFM

9778
HLMPRNQCF

9779
DIATMRNTW

9780
GKVQWSAQE

9781
AHQTAWAWI

9782
GCQCTGHPI

9783
EPLYSWVGI

9784
EIYSQLTDL

9785
KMEVNMSEL

9786
NVTTMFGQK

9787
CKVDIHDYM

9788
ENCMSRMKP

9789
HPLPNTEIC

9790
KIGVNSHYD

9791
GGDGRVHNK

9792
SIRYDFYVP

9793
SNTPMPYHG

9794
HVTMNQLYS

9795
QHLMTFSHE

9796
GREITQVMS

9797
GRWMCNEAE

9798
TWALAMRGD

9799
MYTGMRCWR

9800
LAAIQHCAM

9801
LQIGTGNME

9802
AQQQILNSN

9803
CASSKSHYL

9804
CLYHSMMKQ

9805
FKPFAQQWN

9806
KAVSYHHDG

9807
ICHTEVMFN

9808
QCQSQEWFH

9809
WCTQFDEYT

9810
PACKEWCFC

9811
IHNRYAQAP

9812
ASSRFGSYG

9813
EFGEMLGVD

9814
CEMQVQLCV

9815
DFMWYHICS

9816
EFDWLLGTI

9817
IATKSNCHK

9818
QPFCFSIGA

9819
QFHVWFEQK

9820
EMSPIDWHY

9821
ARAYYDSYS

9822
TVRHQMDAY

9823
VKFRITTEC

9824
SDHQYRGCL

9825
TSSDHYALP

9826
GTDLSNHFQ

9827
RAERYFKDE

9828
SKGDLCHQQ

9829
EAQPSEING

9830
TVCGSFHGC

9831
QTPKYFVHQ

9832
KTDKAGLME

9833
VPTGRRVKL

9834
GAQSMQFSC

9835
TQEACHHEW

9836
MKTIMQDYF

9837
MFSHGHVMF

9838
SVSTKPHVE

9839
RAIYNMITE

9840
MFTPCQEGL

9841
KNAVLNASD

9842
DVYMAVVGS

9843
CIMLNGCFQ

9844
RHWYIALYA

9845
CWQLFNKQN

9846
KPCNPVGQE

9847
SSVVMAKQP

9848
NAKYSMLYG

9849
GMWAQDSSG

9850
YPKRCAEGD

9851
VKHTAVDVP

9852
GEGVEQMDG

9853
MCRPVMMIE

9854
GTNAYMAHK

9855
NIQLLHSYC

9856
MWAETADPA

9857
QIKTHSNVF

9858
RLETTRYIW

9859
VTMTTKSVW

9860
TMPDRDPTY

9861
TVWIQSYSN

9862
CVKPFLTGW

9863
LIWSVYQDE

9864
MPHHSPDCW

9865
HGYQMAYAQ

9866
QVYMMEDNM

9867
VTNWLSLQS

9868
MVFLEQHGR

9869
KRTDLHAAD

9870
QTEIGHWEE

9871
TMGLGMNAI

9872
VLKSCDRHG

9873
TYWCPLMMN

9874
ANGTRECQS

9875
DSQYHVKCG

9876
MALPLHVLS

9877
TRNVHEYGM

9878
QTQICHSWM

9879
TEEVQFVVH

9880
DSATFKSAD

9881
CGCQVKCHT

9882
EAYITYNKA

9883
CTHHNLKAA

9884
AWSDLMNCW

9885
MTIKMEQQY

9886
KATHCDKYV

9887
AHEDPNTWC

9888
AWMCCARHD

9889
QWEEVWWEA

9890
NAIPYFVQA

9891
ITSHWLSTN

9892
KSPCCMSVQ

9893
AGYQCGYNE

9894
ETNGVEDWY

9895
SMKQMTKSF

9896
YACWLQDAL

9897
KWEQRSKMS

9898
KTQTHTSEP

9899
GNVLLGCQP

9900
HAKQANGMV

9901
KCEGEEVGR

9902
HSTNLFEAK

9903
GWADFRELQ

9904
ETEIRQMDG

9905
KITVHDRIP

9906
VCGWWHQYW

9907
IADDYFMCH

9908
GVWMPHGWR

9909
MSLQHHFCT

9910
YNNDMKQRR

9911
QGVACLDQL

9912
RITQGESGM

9913
ASGHMTMCM

9914
NTNRAHYWH

9915
TIKQQMEMM

9916
CQNTAVRAI

9917
AVAEIRPEP

9918
YSFATGNAA

9919
SYLWRSVYC

9920
AVDHYSNGA

9921
ASIATMDRI

9922
NHKMVQDAL

9923
QRALPHPIK

9924
LTHNTPFPK

9925
SIAQDQVWS

9926
CCRQIWVQY

9927
CWGPIMHPF

9928
GNKSPDECF

9929
GHRPKLCDQ

9930
QVACHMTNP

9931
QYVQCAAKD

9932
QDEPMNLSC

9933
GMRTHSVMN

9934
GTNHRDCLV

9935
LMKYQWIGN

9936
KIDQTQWQR

9937
TLHYFHGYQ

9938
QSQWWDTCN

9939
WKPEQRHTP

9940
IYCLCQDSD

9941
DMKQKAEYT

9942
CIPTYRHFG

9943
VRQFGTWAT

9944
ICIRGARAD

9945
TSSKHLVFD

9946
MIKLINCVN

9947
IESNTNKFP

9948
GIKKPNEDN

9949
AMTRCKAGH

9950
HGNMVLVIG

9951
LIQSYPHQC

9952
GGALARSSI

9953
QKAIVAQND

9954
KTLPFAIYH

9955
KIQPTNHHE

9956
KMGMHDMAM

9957
QENDERIWY

9958
APFFTFTNH

9959
YSDMFHKDH

9960
RMVLAYGNF

9961
GQDCWGLAM

9962
PRWQSEKCQ

9963
NTAKLGWGL

9964
MVMMREPSN

9965
MDGRDEIIH

9966
VCRLDNGPQ

9967
MTGIGKCWN

9968
EIHTGRPGS

9969
QIIYRCRST

9970
KCLPYMEMF

9971
PNGYRPFCC

9972
QGQSGTQYA

9973
LVSQWDYDM

9974
ESCHLQHGE

9975
TSEPHCSYA

9976
LSVQAQWDS

9977
FVPWCHSFG

9978
MKEDNKFAV

9979
KTGWDSHWS

9980
TGMTFNSFR

9981
NGYLVHACQ

9982
QVEWFMAQY

9983
YTWPYPETH

9984
IQCKYIHFT

9985
SNVNLMNSR

9986
ATKFFCSSE

9987
MREMPMVGA

9988
SMQEYQKHI

9989
KSMPPGQGD

9990
ECRMAVQSC

9991
VRWGHTCFY

9992
VKRLENEWH

9993
TNRCHDDRG

9994
TTKNAISFE

9995
EAELENVNM

9996
IQNPAMWGT

9997
CDWMANVSV

9998
EQTHTWLCR

9999
AKYLPELYY

10000
EIHEFRASN

10001
WLIGMWNTG

10002
WHVQSKRWT

10003
TTDQQCTGH

10004
NISPYEMAV

10005
YYNHKIVEP

10006
AKPTHDVGS

10007
HTNQQANAP

10008
VTKLAENVY

10009
DVCDDHKPN

10010
IAHWKCHYP

10011
YWEKVPCMK

10012
NWEFNARHD

10013
VRLQPQHNK

10014
KCTEYVHTC

10015
VPTPWMGIS

10016
HHFHVNLDI

10017
DSRTQGGLC

10018
VRADNMPVM

10019
LLPIMHTHW

10020
ISLWVMHYP

10021
MWTEFNEHG

10022
MWAEFNEYG

10023
QVHIGSWPP

10024
YEMTSKCTY

10025
HMSQAGVVS

10026
DQSMMTIGQ

10027
MPSWYIAQA

10028
GAGPVQSEE

10029
FGGMTPYGV

10030
VCSNVRSYC

10031
HGNSFLVHW

10032
MWTEFNEYW

10033
DITSLGCIM

10034
NRPLMSCCF

10035
EFWWMNYMG

10036
TSQYSSVST

10037
MGTEWTQSY

10038
MWTEFDEYG

10039
KQLFMKEWN

10040
EFKCVSDCL

10041
KTIAQTYYG

10042
VQMQYERLF

10043
MDVQQIISG

10044
NPIKFQCAH

10045
NSVHVHASN

10046
CAEGQRALT

10047
QAIDMEFPL

10048
FNESGWPTW

10049
NARVMAKRE

10050
LNDPHCSQN

10051
YFADAWLKT

10052
NIHYHMRRN

10053
QMEVHWHMW

10054
REEWKNASN

10055
ISTKFGKEI

10056
AIHQESMIQ

10057
QEYHIYFPF

10058
TLTHCYSKD

10059
GCLTSECAY

10060
AWFNTYSVH

10061
GYGHTFRDA

10062
FIDEKQCHE

10063
HMSFCNFKV

10064
SVETARCSG

10065
SSEHYSASF

10066
LMKTSPSYW

10067
YGNSVTCTS

10068
KHITWNHCL

10069
DTSKTGKHV

10070
LTQYKALAQ

10071
TRIVEPDAN

10072
LADCMLNHH

10073
VPMWAHWQN

10074
SVTPDSITT

10075
QRNNLFVHD

10076
IHGIDCYEI

10077
GIDLMRQIN

10078
MKNQPTGAS

10079
QKSMLNSIQ

10080
ICNMDFRCA

10081
KTVLKMQDM

10082
RLGMNWKQI

10083
TPKSEGICC

10084
GVITVDAKG

10085
AIGPDRSNN

10086
EQEISDQCR

10087
MQYRGMSAS

10088
MISQMGLKN

10089
AFARQEYAN

10090
TYLHQPEDS

10091
TKLSMMGCE

10092
EELRYTCHT

10093
GSCCLNNET

10094
KKTLYEMAC

10095
RWGVMDLCN

10096
NVEDVGMIE

10097
QSWWHSDTC

10098
AADGYVAQD

10099
ITTPLHHHN

10100
NAQRLAVGM

10101
HVIGMGAGE

10102
RQYFEDNAG

10103
KSELNRFTY

10104
SALYRWGHV

10105
HRAGMSNIP

10106
MARQCHSFH

10107
ICKHLNVWP

10108
TNGKSSSYS

10109
HAVNFDCKK

10110
QISAWNEFF

10111
MMTPMAVEP

10112
KISTTWADK

10113
EVDQYYSLA

10114
GTMSFPTGC

10115
MIMANNRCW

10116
GIGHIGNAT

10117
GGSYVSAPD

TABLE 75

Sequences of the 581 to 589 Region in AAV5

VP1 Capsid Polypeptide that Drive Colon

Tissue Tropism

SEQ
581-589

ID NO
Sequence

12118
AQWYMWCWD

12119
ASHYHEKVW

12120
IANWRWGPD

12121
RVNGSTCRH

12122
YNRLMHQLW

12123
ETTMMIVSP

12124
STAKTGTTT

12125
LPPLKTEDN

12126
QTQICHSWM

12127
QSLWYWQQE

12128
KMGMHDMAM

12129
IRPQYAKMD

12130
MNANETHGA

12131
MSVAAGRGM

12132
QTMCITIHP

12133
CMEAQGECY

12134
FPQTMHAQA

12135
KPLASIGQY

12136
CCCQFSFWN

12137
DAELMWQIE

12138
ENTMWQSSQ

12139
KIDQTQWQR

12140
EQEISDQCR

12141
GKIAHDQLN

12142
MCFPYSCAA

12143
LSVQAQWDS

12144
MMNQCQVWN

12145
TQEACHHDP

12146
DFNGSNWLP

12147
HFEVRSVQR

12148
MAEQMERDF

12149
TGMAMSEQS

12150
PCCWEQSTN

12151
NWYTTWIDE

12152
CCKSKEVCA

12153
LESVKMDCN

12154
PHVSGEGKI

12155
TYDTRNHCL

12156
MAAITQNVA

12157
PDLWYEKSs

12158
AHAWLQHWA

12159
MAMWIADTH

12160
LCAVCGDCD

12161
RITQGESGM

12162
GNKSPDECF

12163
KNCIMLASN

12164
EKRLEGWVY

12165
LETMMRHGE

12166
EQPNMGDYG

12167
QSQWWDTCN

12168
LMKYQWIGN

12169
QISKPSVQC

12170
AIEHGAWKC

12171
SLTQYQWRG

12172
TIVVIPIKS

12173
CSMPQQCME

12174
QTAEWYCGA

12175
GNECFHNVE

12176
VGPLYAYYS

12177
WTLQVMKIN

12178
TSEPHCSYA

12179
AVHTADVCC

12180
QHSRAYMWY

12181
HMEQCWCNE

12182
AYAPSKNAM

12183
DNYWIVPFA

12184
QRSVFHQRS

12185
HEVMMCPAP

12186
KTMGIGGGP

12187
TLNSRHPEC

12188
PLNQEQIPQ

12189
AQCHDMWGH

12190
KACMSHQGR

12191
TTTNQFMFQ

12192
VLKSNLTGM

12193
MFMTNVNSF

12194
SSRMQMQGP

12195
RYPCCQSPF

12196
NARVMAKRE

12197
GLSMVLHFV

12198
VSVQSRAQL

12199
DMTLQSVSS

12200
MTDNNGPLF

12201
PLTIEVNCT

12202
YVDHWILGD

12203
NQREWHGLA

12204
SECEVLCSL

12205
CAWHMKQGN

12206
ATGTYNLQE

12207
KCADLIIRS

12208
QSRCVDNTV

12209
ATMNLSWGL

12210
SVLHVQALA

12211
KMAGIGCWY

12212
YDYWMQAPT

12213
HSMMPGWPS

12214
MFWTTPSHW

12215
TKHCFYGGC

12216
AMNWWAAPQ

12217
AMNWWDAPH

12218
AMNWWDAPP

12219
AMNWWGAPQ

12220
CDRPDVNWF

12221
CDRPDVNWV

12222
CHSLHQVWA

12223
CHSLHQVWD

12224
CHSLHQVWG

12225
CHSLRQVWD

12226
CKTMKDIER

12227
DMQQNDPCC

12228
DVDFRAWAT

12229
DVHLFPTEQ

12230
ECHYRMFQI

12231
ECPYRMFQI

12232
EHHWGDIID

12233
ESAHECEHD

12234
ESAHECGHD

12235
ESAHEWEHD

12236
EYHYRMFQI

12237
FAVECVDCT

12238
FEVECVDCT

12239
FEVGCVDCT

12240
FLGFKRYGG

12241
FLSFERYGG

12242
FLSFKRYGC

12243
FLSFKRYGG

12244
FLSFKRYGS

12245
FLSFQRYGG

12246
FLSFRRYGG

12247
FNLTAHEDN

12248
GLEINMLNC

12249
HCAKFECMF

12250
IANTNPRGR

12251
IEAPKVNEY

12252
IEDPEVNEY

12253
IEDPKVNAY

12254
IEDPKVNEC

12255
IEDPKVNEH

12256
IEDPKVSEY

12257
IEDPRVNEY

12258
IEDPTVNEY

12259
IEDRKVNEY

12260
IGDPKVNEY

12261
IYTYQWDWI

12262
IYTYQWDWT

12263
IYTYQWGWT

12264
KCHYRMFQI

12265
KDTEGLEAP

12266
KDTEGLGAP

12267
LASQKRDCD

12268
LEDPKVNEY

12269
LINHWMHDN

12270
LINKWMHDN

12271
LINQWMHNN

12272
LLNQWMHDN

12273
LSSQKRDCA

12274
LSSQKRDCG

12275
LSSQKRDCN

12276
LSSQKRDGD

122T7
LSSQRRDCD

12278
MCATQHCEA

12279
MCEQYWTED

12280
MCEQYWTGD

12281
MGEQYWTED

12282
MGRMELDGV

12283
MHMQQNQMM

12284
MHMRQNQMM

12285
MHMWQNQMM

12286
MRATQHCEA

12287
MSRMELDGV

12288
MSRMEQDGV

12289
MSRMGLDGV

12290
MSWMELDGV

12291
NDDPGQAVG

12292
NVHLFPNEQ

12293
NVHLFPTAQ

12294
NVHLFPTEQ

12295
NVHLFPTGQ

12296
NVHLFPTKQ

12297
PACYSDVLN

12298
PEFLGEVDA

12299
PEFPGAVDA

12300
PEFPGEGDA

12301
PEFPGEVDA

12302
PEFPGEVDS

12303
PEFPGEVDV

12304
PEFPGEVYA

12305
PGFPGEVDA

12306
PKFPGEVDA

12307
PQCLHHHWY

12308
PQCLHNHWY

12309
PQCLHSHWY

12310
PQCLRNHWY

12311
PQCLYNHWY

12312
QPVGLFAFA

12313
QSKSEGIHI

12314
QTDILDYAW

12315
QTDILDYDW

12316
QTDILDYYW

12317
RDRPDVNWV

12318
RLEINMLHC

12319
RLEINMLNC

12320
RLEINMRNC

12321
RLGINMLNC

12322
SCHQCDLCD

12323
SCHQCNLCD

12324
SCHQCSLCD

12325
SCIFRHWLT

12326
SCSPDAWET

12327
SCSPDDWGT

12328
SCSPDGWET

12329
SCSPGDWET

12330
SLQRWFCRT

12331
SLQRWVCRT

12332
STIHAIEQG

12333
SYHQCNLCD

12334
TMNWWDAPQ

12335
TSVQSVDRG

12336
TSVRSVDRD

12337
TSVRSVDRG

12338
VQCIHHDYH

12339
VQCIHHDYR

12340
VQCIHHGYR

12341
HQCIHHNYR

12342
VQCIHPDYR

12343
VQCIHRDYR

12344
VQCIRHDYR

12345
WKPPLSDAP

12346
WPSMQCAAH

12347
WPSMQCAAN

12348
WPSMQCAAR

12349
LSSQKRDCD

12350
IEDPKVNEY

12351
SCSPDDWET

12352
AMNWWDAPQ

12353
LINQWMHDN

12354
AAEQRFNPE

12355
AAILMSNAG

12356
ACMMMWPYV

12357
ADTGSTAVI

12358
AESVEDYHT

12359
AFELHYHHQ

12360
AFKFKMLGQ

12361
AFKWSHIHL

12362
AGVQYTWCG

12363
AIDQFSGCK

12364
AIEYRKYDR

12365
AIFMMSGPG

12366
AKWMITIGC

12367
ALDWCTRQA

12368
ALICVNVFG

12369
AMHTQGSLG

12370
AMTQEQMAD

12371
ANNSCYRPY

12372
APLPWWVVP

12373
ARRYNPELQ

12374
ARSPWMHMS

12375
ASDAKPAQL

12376
ASDDQRDSK

12377
ASHPCYSMH

12378
ATDIWCAFC

12379
ATNDSPHLA

12380
ATSWNTAIG

12381
ATYVYERFE

12382
AWCITYGLG

12383
AYERYDCYD

12384
AYTKMAAQG

12385
AYVAVASQW

12386
AYVPEFRDT

12387
CATRKDEMF

12388
CEECQEMLT

12389
CFRICVSEY

12390
CGEDMKIWQ

12391
CGGCLNLGT

12392
CGGTHGMMG

12393
CHVDRAMHH

12394
CHVSVPVDC

12395
CLANVDNFD

12396
CIEYTHRLA

12397
CILNTCSFC

12398
CEPPLSDKE

12399
CKAPAQDFR

12400
CKAPTRSYR

12401
CKSIGCMDE

12402
CNDMTAQYV

12403
CPDQYRAYP

12404
CQGFVKGQS

12405
CRENMERVS

12406
CTKKDNMDN

12407
CTNRHTPFI

12408
CWATEWQMP

12409
CWEQEWPFW

12410
CWQNKINHE

12411
CYVMRRIYC

12412
DADQAAKAC

12413
DAGMCEKED

12414
DANEASSGR

12415
DCMVFINYL

12416
DDCTNGSME

12417
DDVNNVAEA

12418
DDWQKLGVP

12419
DELYTGPCA

12420
DFNTKTSSL

12421
DGEFQAYWE

12422
DGGMERGVS

12423
DGNKREWLS

12424
DGSPKNTSS

12425
DHIQMHRFN

12426
DINANMDRK

12427
DEQSNCSEA

12428
DESEQNRMC

12429
DESQHPEGS

12430
DEYLSNGTY

12431
DKCMLMDAS

12432
DNENCVQKT

12433
DQNEEGTKP

12434
DREGYCNNW

12435
DRQGCNAPR

12436
DSMWRTKEL

12437
DSMYASSMP

12438
DSRQVKKEN

12439
DSTWLEAQQ

12440
DTGVYSSWE

12441
DTSADKSGW

12442
DTWISHWWC

12443
DVKSENHDW

12444
DVNVAASWV

12445
DVVQEFWAK

12446
DWHHEHNPN

12447
DWQRVKTMW

12448
DWSTEEWTA

12449
DYWSYPNYF

12450
DYYESWWMH

12451
EAHQSTLST

12452
EAKFLGGCW

12453
EANHYIEEQ

12454
EAPKVTVWG

12455
EASMKWWSN

12456
ECDHHEEHY

12457
ECQCCECQD

12458
ECRMRLDNS

12459
EDRNCSECL

12460
EEKPNYHTA

12461
EFDYYCLKT

12462
EFETKNWEH

12463
EHECPNSGM

12464
EHETTMAEA

12465
EICSHLCWA

12466
EIILGGYHN

12467
EKGWSEPGN

12468
EKQQIEIAS

12469
EMMSQSACA

12470
ENESSHTNP

12471
ENKVCVYMM

12472
EPYQKDCEL

12473
EQQAPDHVK

12474
ERGVMCHFW

12475
ESENPMGPY

12476
ESHMMRISE

12477
ETQTHYTSC

12478
ETWLMHHHV

12479
ETYEGGCCM

12480
EVGWEPPYM

12481
EVNYVKCND

12482
EVINRTIPR

12483
EWKCSQNLT

12484
EWKYSQNLT

12485
EYNQCTEGM

12486
EYYQSVKMS

12487
FAQLHAVGN

12488
FASAMDVTL

12489
FDRHRWQPL

12490
FEHFDLSAS

12491
FLGNEKSYE

12492
FLQNVGFEN

12493
FMMAFKLDC

12494
FMQPIKQMD

12495
FNPWDKYGW

12496
FQPHIRDVG

12497
FQYRPAEHV

12498
FRVLQGNSV

12499
FTETTHCGH

12500
FTSGHGYME

12501
FVHKTEGED

12502
FVHTVTVNP

12503
FYFPEGMRP

12504
GDGRAETWQ

12505
GEDCPSAEE

12506
GELVRRLSQ

12507
GFCYPNKMI

12508
GFTQDPVGH

12509
GGDTGEYLF

12510
GHEPNKHFQ

12511
GHQVIEYMF

12512
GHYVIHKTN

12513
GIITPSMGS

12514
GMNPQLECP

12515
GQPENGKQE

12516
GSARQDGKP

12517
GSCLGISDM

12518
GSMQIAQEG

12519
GSMQSPTRR

12520
GSNSHLPLQ

12521
GSNSHLPLR

12522
GSTSKDSSM

12523
GTHPCGCPE

12524
GTSFNVHGA

12525
GVDHFFWVS

12526
GVHSAFYQT

12527
GVEWEHDEN

12528
GYNHVDSWS

12529
HCHLCSLRG

12530
HDERMWAMD

12531
HEPRTECTF

12532
HEQIGPEYR

12533
HFGYMHCSQ

12534
HNEHKVGYG

12535
HPLKEPEGK

12535
HPLKIPIGK

12537
HQNHPPAFD

12538
HSFEEHDEG

12539
HSNPRCVFF

12540
HTDQDRHAE

12541
HTSPCWGSY

12542
HVEMPAPGC

12543
HWIHMVNQQ

12544
ECEWTIAQW

12545
ECKPEGGEK

12546
EDNMPKCRP

12547
IDVHNGFDQ

12548
IDVRPHFAD

12549
IEMSEQPNC

12550
IETRMESMG

12551
IFSHWFAGW

12552
IGCLYTSNW

12553
IGDQAMECS

12554
IKCEPWVSG

12555
ILAQENRWA

12556
IMEWKNTMV

12557
IMMHRVMSD

12558
IMRDMVLPV

12559
IMTNYHYLD

12560
IMVKRTVNA

12561
INCKPSVQF

12562
INVDKDQFL

12563
IQCSTNSWE

12564
IQFVTWRFE

12565
IQGPSWHKN

12566
IQQVDMMCP

12567
IRWVEPMCQ

12568
ISPCPTVVA

12569
ISPNMQWYP

12570
IVFQGDTGT

12571
IVTVNCAGE

12572
IWCNGQMDD

12573
IYTYFIEWF

12574
KAFPIDLIA

12575
KDDHCAFWN

12576
KEGVDQEFF

12577
KEKQNAKCL

12578
KHHPTDADN

12579
KICMYHLID

12580
KIFWTTEFH

12581
KKTEEHHCQ

12582
KMMHLPQEE

12583
KMVQDEMWC

12584
KQEPMHCSA

12585
KSDQVWQGE

12586
KSTIFKMRH

12587
KTECFFSMH

12588
KTFPAADVK

12589
KVEFTATVM

12590
KVNIYSDTL

12591
LAERWEQNN

12592
LAHEPYWSP

12593
LALCDAVND

12594
LALHSQFCE

12595
LASPKQMVA

12596
LCAPYNREV

12597
LCGCCMMHY

12598
LCGNDKKCD

12599
LCHFIKGVY

12600
LDQGMYFHK

12601
LECWNKIAR

12602
LGYHVKDFM

12603
LIAFLVDRI

12604
LIMPLTTQM

12605
LMKMWISMG

12606
LPDFWRTRN

12607
LPIFVQGDF

12608
LRAIIDCGW

12609
LSATMVWAQ

12610
LSIQDNRGQ

12611
LSTSSQLTP

12612
LSVAVQESN

12613
LSWMACAQA

12614
LSYVMDTAA

12615
LTQFSCVSH

12616
LWVWWMKDH

12617
LYEEHHACW

12618
MAASGMLCY

12619
MADGRPRMP

12620
MATCYHDHW

12621
MCAQPCNLP

12622
MCVHKFLTG

12623
MDGPWMGKL

12624
MEHVPTRDQ

12625
MEMPLMDVI

12626
MGTQYQFTG

12627
MHAETDIQM

12628
MHFTSWPVD

12629
MKIMCENVR

12630
MNMEKLQID

12631
MNRIMMVAD

12632
MNSPRRECI

12633
MQIILEFPR

12634
MQTAHPVYL

12635
MRNHGDFDF

12636
MSILASVRV

12637
MSIRGNWGY

12638
MSKPCRVCE

12639
MTDPGMKRR

12640
MWVQRETGG

12641
NAAIWEHVH

12642
NACTKVKMT

12643
NAYNFTKDC

12644
NCENKQNVI

12645
NCPCHTDAY

12646
NCREHPEEG

12647
NDIECEMAS

12648
NEYEKLFPR

12649
NFMSMYGFE

12650
NHFPILIDH

12651
NHKLHTGPG

12652
NHWMGEKEC

12653
NIGFAIVDP

12654
NIGKERMEN

12655
NIGTHMGHT

12656
NIYVTRAHC

12657
NKSPTWTES

12658
NLHTPAGCM

12659
NLVCRESSC

12660
NPCIKASYF

12661
NPNGPLHYD

12662
NQMPKEKWD

12663
NRTGVTPST

12664
NRVDAMIHY

12665
NRWMMGMKP

12666
NSTEQEIIQ

12667
NTMTSAPVC

12668
NTNMAWKQF

12659
NVTELHNPK

12670
NVTETGPDA

12671
NYASEEFLT

12672
NYMGIDEQK

12673
NYSMPVDWS

12674
PCVEPNAAM

12675
PDARGMHFL

12676
PEHPRDWLM

12677
PHAGQGNKG

12678
PHGPITNPE

12679
PIAIKDQGW

12680
PLASMENGP

12681
PKILFMFSF

12682
PIMIARYDP

12683
PMAPDWRNS

12684
PMTLPLECW

12685
PNEWYLWAC

12686
PPNMTSCDW

12687
PQDQSCAGQ

12688
PQHWKSVGA

12689
PTAMWLTHM

12690
PVATYNVSR

12691
PVEPSCQMK

12692
PVEVMQWKP

12693
PYEPQTINC

12694
PYVDKWANH

12695
QAAPVHAKG

12696
QAEATHAEF

12697
QATCTRYGH

12698
QCGCCKMKW

12699
QCIQIDMER

12700
QDDEYHMTP

12701
QEIAKKWSN

12702
QETGPKSST

12703
QFKDSRGAG

12704
QGDPVKFMS

12705
QGKAKQIQH

12706
QGTHSQNDD

12707
QIAQQPCMW

12708
QIDNVNQDP

12709
QITKKDNLM

12710
QKLVHKESF

12711
QNEKYGKTL

12712
QQGPLQRIM

12713
QQKRVCDQM

12714
QRPWHDLTY

12715
QSAQSHSHV

12716
WSNHEFRWF

12717
QVAVDQSPF

12718
WVYALSKHW

12719
QWDRCLVQV

12720
QWFVQGITL

12721
QYKVKERGM

12722
QYNSHFLYG

12723
RCDQHDDQW

12724
RCTDCSYFY

12725
RDQQIHHED

12726
REVKPMHVH

12727
RNDVCCEMQ

12728
RNLYENQGC

12729
RPCSKSHEF

12730
RPESEHPEG

12731
RPREGEWPG

12732
RSLHYGNSV

12733
RSPPCDSFP

12734
RTNDHPDKH

12735
RVDCETPYH

12736
RVGNERLHH

12737
RWYHYLLRN

12738
RYCLLDEIE

12739
SCKHDDMGK

12740
SCKWIQSNR

12741
SCLDHQDID

12742
SCNCKKNKI

12743
SDKGRQHYN

12744
SDNNAPEFQ

12745
SEGVYNQVY

12746
SFHVAQGDY

12747
SFIAPCDLS

12748
SGCTWDMSP

12749
SGPDTVFFT

12750
SHASGIECT

12751
SHESTNAGA

12752
SHMQDKNYT

12753
SHVEPVMGF

12754
SIRWFYTDC

12755
SIVNHNEAR

12756
SIWGCNVGG

12757
SKHEWDMHV

12758
SLDRFNQSG

12759
SLILMHEQT

12760
SLNWWFWGG

12761
SMGPLWDPL

12762
SMHTVINKG

12763
SNDCRNEFT

12764
SNSCEYQIM

12765
SPCFYPDCS

12766
SPICLQHDR

12767
SQKNSRHLS

12768
SSALITRKE

12769
SSSFWENKP

12770
SSSSHLAAD

12771
SSVCKTACW

12772
STARDLYLM

12773
STSHPDNSI

12774
SVFDVWCDQ

12775
SVMQGSSIQ

12776
SVWSTRMQK

12777
SWEILHDCH

12778
SWHWMTTQN

12779
SWYGNEEDQ

12780
SYHDHEQGI

12781
SYQLPILHS

12782
SYSYVNDEA

12783
TCKGMDNAI

12784
TCTEPIVEC

12785
TCVQMEHVK

12786
TCYWDNCWY

12787
TDRWTRVMS

12788
TEDQYNNIH

12789
TERWGWLPH

12790
TFWCAQLGV

12791
TFWIRMAAG

12792
TGLYYEMQE

12793
TGQNGWSKT

12794
TGVVDTAYS

12795
THSKSDCAY

12796
THTVRDSHI

12797
TIDRCNWHY

12798
TILHGMLLE

12799
TITHEQPGP

12800
TIVPYEYNQ

12801
TMDPCRKSC

12802
TMGVYSFDW

12803
TMMLVVLRG

12804
TNGIGGKKE

12805
TQLHWNDGR

12806
TQQALPLAC

12807
TSAPDRLSS

12808
TTEQIRDID

12809
TTVSCVCKG

12810
TVLRRCNYE

12811
TVYAWLLSM

12812
TWFVGQSTR

12813
TWQRTENNK

12814
TYLMNQFHM

12815
VCKMPMYQH

12816
VCRCTKYTA

12817
VDPNLQYPW

12818
VDWFTAPAP

12819
VFCNYQKKA

12820
VGSKWHVLG

12821
VGYSTVGEW

12822
VHHDLGESK

12823
VHHHGYPTS

12824
VHKEWHWQC

12825
VHMYPMTQV

12826
VKPNETDLH

12827
VLGFDHDPQ

12828
VLLMDGRTF

12829
VLRYVNSAG

12830
VMELMMHNH

12831
VPLVNDGRC

12832
VPTICMATY

12833
VQNEADMNP

12834
VRAMPFDPD

12835
VRNTEMDWF

12836
VSAPECHIY

12837
VSMWYDRDD

12838
VTFMITMFH

12839
VVDHFFWVS

12840
VVITDGGQV

12841
VYMIFSGWQ

12842
WALAPELWY

12843
WAMDNMNRC

12844
WDHRYQTGH

12845
WDQCTMINQ

12846
WDSPQTNFT

12847
WFHTRHPQE

12848
WGPCKHPWQ

12849
WGVVKMVLC

12850
WHDKLRNNK

12851
WIDGIVDIC

12852
WINGPASRY

12853
WIQLNHQEV

12854
WITAFDSIG

12855
WITVKEFST

12856
WLISNSPQW

12857
WRCPPEQWW

12858
WVSADTYEH

12859
YASLWESGW

12860
YCMTVICSE

12861
YDECQKIFV

12862
YDGCMWMPF

12863
YDQKGGQCW

12864
YEDIIWNMH

12865
YENVGSKAW

12866
YGHRRDEAR

12867
YHSHLDAGQ

12868
YKLNCFARP

12869
YLDCPHTQF

12870
YPMAKNRGD

12871
YPRHWTLQH

12872
YQHVWSQER

12873
YQNPDAPAL

12874
YQQQLMMNE

12875
YSDAVYCSM

12876
YTDALGMEP

12877
YTPLPAVNG

12878
YWLCYPKLC

12879
EKLTADAAI

12880
INSESEWNP

12881
QTTEFALVQ

12882
SGLYSQCGG

12883
SDVHMFCML

12884
QSNLMEFHS

12885
EAQNIHDPH

12886
HTTLNYMSN

12887
AATPQKNQE

12888
AFCRGHYQA

12889
AGAPPLAVH

12890
AGWQMCFDS

12891
AGWQMCYGS

12892
AHPILQGWD

12893
AHYQQSTDR

12894
AIGHGAWKC

12895
AMCMLSHIH

12896
ANELSWHPR

12897
AQGCKFYNG

12898
ATVTRARSM

12899
CACHMKQGN

12900
CDSLSLLQN

12901
CFCQCGMPS

12902
CHHSGESHQ

12903
CKGRANWYM

12904
CKWLAIPCR

12905
CQVGLQNCW

12906
CTKMNTMIE

12907
CWALNGQLG

12908
DALPERRYF

12909
DAMQKISCV

12910
DFIYHHQAP

12911
DGVVTVTNL

12912
DHDLATEWE

12913
DKHHSRIDG

12914
DMNMYLGSQ

12915
DSKITQFNH

12916
DVETNNWRC

12917
DVHDSQMCY

12918
DYNDFNWYF

12919
ELGTYDGMG

12920
EMQMEERLP

12921
EPGRMHHSW

12922
EQCLAAAAE

12923
ESRPMIQPF

12924
EVCGHQKIQ

12925
EVHVTACGN

12926
EVVRIENPM

12927
EWFCYCGTC

12928
EYHKTQWGH

12929
FALIYHKDE

12930
FASPVLWGQ

12931
FEGFIERQH

12932
FGHRYDSAY

12933
FGNPMQICP

12934
FGVNDMDSA

12935
FHCICWTHM

12936
FKYDCYQRN

12937
FLGCYCPYA

12938
FLRNTWLGP

12939
FMPYCVHSG

12940
FPSWMMKAP

12941
FSTAHMFGW

12942
FWCRPRLHD

12943
FYHHLPEEP

12944
GAGRMNCNV

12945
GANMMHMMN

12946
GCLISECAY

12947
GDKMGIYHC

12948
GEMPYCNRQ

12949
GFKVEQPYE

12950
GHTNCFAMS

12951
GIDEAKADY

12952
GKYWLQRSV

12953
GLFHPQFVQ

12954
GNISTFCWQ

12955
GQHHEETIY

12956
GQILFRSHT

12957
GQNPKMSAE

12958
GSCCLHIHH

12959
GTQHEKMYQ

12960
GTTQCTVHT

12961
GYDSSEAFE

12962
GYVIEGFDE

12963
HCPCKCTWL

12964
HERHATLMR

12965
HLLGNLEYM

12966
HNVCTMWAQ

12967
HQIQFCPCP

12968
HTQEELITP

12969
HTYCYWQSC

12970
IAAKVQNHD

12971
ICAAPNSAK

12972
IDHDNWDML

12973
IIGQTSIQD

12974
IKEHIPYWD

12975
IMNKAHYRF

12976
IQAGHQATM

12977
IVDMNTATK

12978
KCFTHAIVG

12979
KDWIPSYAL

12980
KEGCHVRMG

12981
KMQGGNGYG

12982
KNAYNMTIQ

12983
KPEILHFHD

12984
KTAENVWAM

12985
KTESIVKLQ

12986
KTIQHGTAD

12987
KYSDFCCPV

12988
KYTEALSCG

12989
LAMCALDNF

12990
LAMRALDNF

12991
LCSTEGMHT

12992
LCSTKGMHT

12993
LDTKRTFKK

12994
LGEATWQAV

12995
LMMKCCAMH

12996
LPDRDWHKW

12997
LQTKKDCPA

12998
LVVEKPSES

12999
MACKKAQDV

13000
MAKRVELVT

13001
MCHSFPTRP

13002
MEVEAEKPN

13003
MMKYYDAIG

13004
MMMPQMWSG

13005
MNRNVEMDH

13006
MTDKLYQED

13007
MTNWLMRGR

13008
MVNNWDWFY

13009
NCDFMVASI

13010
NCQPYGEPT

13011
NCSHYTQHY

13012
NGLLDFDPH

13013
NGLVDFDPH

13014
NIAPGGTIF

13015
NLEAMDCNY

13016
NMCTTFRLL

13017
NMKTKSAHK

13018
NNHRLNDAN

13019
NTMQSPAWV

13020
NVTPTDREG

13021
PCGRTCFNG

13022
PMMSFIFHH

13023
PQTAAQKSD

13024
PTEVTVSDM

13025
PTQWAMGDY

13026
QAAYCRCTF

13027
QATHDLMHQ

13028
QATLHHCTN

13029
QCLLWNDCQ

13030
QGQEKEWYY

13031
QHMELQEHD

13032
QPCIPVQIA

13033
QQLYFTIDK

13034
QVIPVTYRK

13035
QVKMGVIDQ

13036
QVLATMNNK

13037
QWRQMDRMQ

13038
RDDQPPMVW

13039
RDEVAEQQP

13040
REERYFKDE

13041
RIEMHQSVT

13042
RKAEQEDVY

13043
RLVTYPHTA

13044
RNATKIKLT

13045
RNCLTDAWD

13046
RSDQECSKA

13047
RTCKTIGYY

13048
RTEPTFVDP

13049
RTYQYEYEV

13050
SCWRETMKE

13051
SLENEARRC

13052
SLHIHGNGP

13053
SMGMCELPI

13054
SNAHMQVGC

13055
SNAYHAWKN

13056
SQDLECNAA

13057
SSGWDHGNL

13058
SSTWQNVDK

13059
STHPLNLYH

13060
SVQPFLHDG

13061
TADRQLVHS

13062
TCGKMHGPN

13063
TLWDNTNTP

13064
TPSWENVGA

13065
TRGHHCAED

13066
TVTCPEVRF

13067
VNQLHIHVQ

13068
VPWYEGPYE

13069
VRFEMMSCT

13070
VTKCGWMLQ

13071
WDNSKGYNV

13072
WNNEDLKVW

13073
YATQMRHLE

13074
YCLMDKEDT

13075
YDGKTFLHW

13076
YFDCSRAHP

13077
YGTKCERCY

13078
YGTKSERCY

13079
YHDLFFAWY

13080
YHTNNMEPW

13081
YMHVPPSIH

13082
YMWIYPPDQ

13083
YNVCTMWAQ

13084
YIQLYCSHL

13085
YVPCERCEW

13086
YVQSYWNES

13087
YWERHCMTI

13088
YYGWLLVMV

13089
AGWQMCYDS

13090
VNCNKRHNW

13091
PYCSIKQAS

13092
CQTHWEKGD

13093
EMISNNCHP

13094
RQRYMGMQF

13095
SYLFSKHSE

13096
KAISLHAHG

13097
GIGEMQCTE

13098
ILERYQHMD

13099
IQKDEMPVC

13100
GTQFWMSVE

13101
QMSWGKYHQ

13102
KDYWAGYQM

13103
ICDLYSQER

13104
EEAPGFICF

13105
DLPNDMGAV

13106
GMHATELWL

13107
LMHPQMTMC

13108
AAKKGRDLH

13109
RHIYRKGWR

13110
GEFADNPWN

13111
ERQMKLRCA

13112
TDSSDQLQI

13113
YNMVHGDEP

13114
NIKMLVHCG

13115
MKGYFVHHS

13116
MLRTQIHCA

13117
CKDNTFVSY

TABLE 76

Sequences of the 581 to 589 Region in AAV5

VP1 Capsid Polypeptide that Drive Heart

Tissue Tropism

SEQ
581-589

ID NO
Sequence

15118
ADIDQIMCQ

15119
ADIDQIVCQ

15120
ADIDQVMCQ

15121
AHKYDWEYI

15122
ARSDWRCMC

15123
AWCEGNCLE

15124
CCCIMSVMC

15125
CCCIMSVMG

15126
CSRIMALSN

15127
DNRTEGYFT

15128
ECHLLSDYD

15129
EDWHGWNEH

15130
ELYMQMHWP

15131
ENRLRLAHE

15132
EQTMFNSPP

15133
FAMVVITEY

15134
FFPHDGTIV

15135
FYQMFHASG

15136
GFPPRWKIP

15137
GHCCSNGPT

15138
GKQQCPAQI

15139
GKQQCPTQT

15140
GNRTEGYFT

15141
HDCALYGRF

15142
HLNPMTGSN

15143
HNVMCDACC

15144
ICKYPMIPH

15145
ICKYPMISH

15146
ICMYPMIPH

15147
IDEFWYGPF

15148
IDPRRNNDL

15149
KINPSMPMD

15150
KPDTSVGIG

15151
KPDTSVGIV

15152
KWPCHWHHE

15153
KWPCHWHHG

15154
LCINKYFNY

15155
LEPKMKLAD

15156
LEPKMKLAE

15157
LVCEVACKD

15158
MFTQNAGKI

15159
MSCPKFHIV

15160
NEDSLWNMH

15161
NEDSPWNMH

15162
NEDSPWSMH

15163
NNRCKDQTE

15164
PFTQHVAYI

15165
PFTQHVAYS

15166
PQQFAWNND

15167
PTAIQAYMR

15168
QHVSTEPPY

15169
QIADGWGWE

15170
QMCATMNHE

15171
QMQHCMDYL

15172
QVGWKECVM

15173
RCQQFHGQI

15174
RDINIDPCC

15175
SAMVVTTEY

15176
SFTQHVAYS

15177
SKQQCPAQI

15178
SNRTEGYFT

15179
TDIDQIMCQ

15180
TDRHDGVFA

15181
TEYQNARTM

15182
TITCEADNW

15183
VFRNRWLEW

15184
VIEVEQACT

15185
VIKVEQACT

15186
VPNPKNFGW

15187
VSVVQNEAL

15188
YCSIIGRSC

15189
YFALCPPYT

15190
YHAFYWQPY

15191
AADHWVCQF

15192
AAFQCAKQG

15193
AAGMHWNAA

15194
AAHMGHGWP

15195
AANQEQECL

15196
AAPCPDRFV

15197
AASDSNRIH

15198
ACCDKTRPE

15199
ACMFWHRCG

15200
ADGRGMDCP

15201
AEENWRQCA

15202
AEHDLKIVS

15203
AEVEQYAEA

15204
AFDPTFIDT

15205
AFEAFDYDH

15206
AFFLTPAEP

15207
AFGNLEVHM

15208
AGAPLIHQQ

15209
AGAQQYWAD

15210
AGECQLLMT

15211
AHNCNGHVP

15212
AHNWPAMYT

15213
AHNYCGYYV

15214
AHQPDQYCK

15215
AIAHCVQIE

15216
AIEGLTGRD

15217
AIEYMNMGF

15218
AIKDMRLAL

15219
AISSHWQWP

15220
AKNEECDCN

15221
AKVEMEEWN

15222
ALAWQMRAQ

15223
AMNDELKMQ

15224
AMVIIYTDF

15225
AMVLMFLYQ

15226
ANNQHRARA

15227
APFMSEDCS

15228
APLMTAFDP

15229
APMSQWHTV

15230
APNGYTMFY

15231
APSAHHTHL

15232
APYWDCFDR

15233
AQGPMSVQL

15234
ARPIFPWIV

15235
ASTVVYAEM

15236
ASWLADNLC

15237
ATAFRHRIN

15238
ATARLNWDA

15239
ATDQYWFPT

15240
ATEIKGSHS

15241
ATESKWDQD

15242
ATHIHYHGL

15243
ATIEGDGVL

15244
ATIRVITIG

15245
ATNYVVRHT

15246
ATSVMAWWD

15247
ATWRICSSG

15248
AVDFCQTYD

15249
AVHAAYCYW

15250
AVHAGPQAI

15251
AVKPREAWM

15252
AVMYNDTYD

15253
AVPCQLSGA

15254
AVQERFLQN

15255
AVQSQTKNM

15256
AVQWVEHEK

15257
AVRCDPYRF

15258
AVSTTVDWY

15259
ALTTPDVYV

15260
AVVAMNHHT

15261
AWGEMNFSV

15262
AWMPFIDSG

15263
AWPDIMWAS

15264
AWRGSKMSN

15265
AWSVQGPYG

15266
AWVDQEIPA

15267
AWVGAEHAG

15268
AYEHITDGR

15269
AYTDTNAGY

15270
AYVEKMRFD

15271
AYYHEMAVP

15272
CAAMQQGRD

15273
CAEAIIKNF

15274
CAHSNKGGA

15275
CAMQPCLAV

15276
CAVDKCMPH

15277
CCSSFRDMV

15278
CCWLANGGG

15279
CDMLTRMQY

15280
CDPWSWTQI

15281
CDSTRFCCV

15282
CEVARWLDY

15283
CEWKHHAKF

15284
CFHDPYEKQ

15285
CGCYSPFLE

15286
CGSDCNPAY

15287
CGYQLFECF

15288
CHNEDLCVW

15289
CHTMQDIKD

15290
CIGCPNEWC

15291
CIQHIAESE

15292
CISEMDDQR

15293
CISFSNKWV

15294
CKAAGPWTD

15295
CKDSFASVW

15296
CKPRLCEVP

15297
CKYMIHPDC

15298
CLYPLQLEW

15299
CMDQYRIQF

15300
CNACQAIEC

15301
CNKAVQKTS

15302
CNMPQITGA

15303
CNPAKSKDG

15304
CPCKGRRLG

15305
CPCLYCNIA

15306
CPEFGFICE

15307
CQIPKKIGM

15308
CQTAFYVGH

15309
CRDRDMAHQ

15310
CRLKKYYLK

15311
CSPRKSHFC

15312
CSRFMDSQW

15313
CSVDKEDMD

15314
CTKLSGLNA

15315
CTKWQRTLG

15316
CTLEFPSFL

15317
CVGIMTNGK

15318
CVGWAMGLG

15319
CVHKQGENE

15320
CVIQSNTHV

15321
CVQPTYADH

15322
CVTLEMQQG

15323
CWAMDWGDT

15324
CWQFTRCAS

15325
CYYHTCLGG

15326
DACFYGNFE

15327
DAENCMLGW

15328
DAGYCQSGV

15329
DALSFQSSA

15330
DANRKDLRG

15331
DAQAGWRFN

15332
DASSGFTWT

15333
DATDWKCAW

15334
DAVKAEVMW

15335
DAWAKGYEW

15336
DAWSANEAW

15337
DAYWMQWWL

15338
DCASFPAVD

15339
DCQGWTAWD

15340
DCVKAIQGW

15341
DCVQMNVNV

15342
DDCWQDRHV

15343
DDVWKELDV

15344
DEAYRMPLQ

15345
DEGAAAKAI

15346
DEGTSQTKT

15347
DEHDGCKKS

15348
DEMYVSSQF

15349
DFAQRNSAC

15350
DFGMKNYLA

15351
DFKFEMTDR

15352
DFLYSPEGI

15353
DFSTPYQFG

15354
DGCTSKDNQ

15355
DGIHSLTGW

15356
DGNQLCRLT

15357
DGNYERVDV

15358
DGPCAQYRG

15359
DGPMFATDI

15360
DHHGAGYNR

15361
DHISDYQHY

15362
DIILRIRIK

15363
DIKAILSKD

15364
DILGVHLGS

15365
DIQWQHEWM

15366
DISCNLQFL

15367
DKMLKTDLM

15368
DKNSCCLNP

15369
DKPVGRYGH

15370
DKQFVEDNQ

15371
DKRFQKSYS

15372
DLMMHYEHK

15373
DLQMSVNTG

15374
DLSQMNKNY

15375
DMGEVQVGI

15376
DMNPDACVR

15377
DMQVLNTDD

15378
DMSEVDKVF

15379
DMTIAAWAC

15380
DMTKVTPFL

15381
DNACVHCAE

15382
DNCWLNDSA

15383
DNEPNNRKS

15384
DNESWSAWD

15385
DPHQGIAGE

15386
DPPIVQQWH

15387
DQKWDTLKP

15388
DQRRKTNIR

15389
DQTTLRLMG

15390
DQYWAWADS

15391
DRLEDKESQ

15392
DRMQFFVRN

15393
DRMTCDTDR

15394
DSEWTIDEH

15395
DSGEVRTAS

15396
DSKVFEWKC

15397
DSQLTNACG

15398
DTELVHNGW

15399
DTLSSDAFL

15400
DTMCYAWCK

15401
DTQPWVPCG

15402
DTRESGFWV

15403
DTRIKMMLP

15404
DVAKKNMDG

15405
DVASEGHEI

15406
DVCMLVDLP

15407
DVCMSNELT

15408
DVDCCEFCT

15409
DVDFHWQGM

15410
DVDHKWQYD

15411
DVDPMPCFY

15412
DVGIMDYWS

15413
DVLQHQGAL

15414
DVMMCFHWR

15415
DVPDKPWYH

15416
DVQLDQEWD

15417
DVQRKTSGE

15418
DVRFESEQG

15419
DVRPPAVNE

15420
DWPCDDFRK

15421
DYLSFCWKD

15422
DYNKDPKLV

15423
EACMHRTVL

15424
EADFVGPNS

15425
EAEEHCTFW

15426
EAEEYEPWK

15427
EAFQEDAHP

15428
EANSEYNVY

15429
EATKSVETR

15430
EAVTARVQH

15431
ECDDYDCHY

15432
ECTRSGKWG

15433
EDHVMTKKD

15434
EDIEKNTFE

15435
EDKQAQTCQ

15436
EDRSSCVWM

15437
EEEPLSEMA

15438
EEKMSVDRK

15439
EETVSKRKK

15440
EFIFHIHDQ

15441
EFMQSFFIE

15442
EGEDYMDIE

15443
EGNQFWMFS

15444
EGQTSNIWE

15445
EGVTTLQSS

15446
EHDTMEGFG

15447
EHGQEEMIM

15448
EHTAMWMHG

15449
EICNKGQPD

15450
EIEGVKANV

15451
EIGGNPEYD

15452
EIGYCMTCD

15453
EEKLPMEQD

15454
EIKPMVASH

15455
EILNHTRLW

15456
EIMDTKQCS

15457
EIMTMEFQD

15458
EINTPFEMS

15459
EINVTQTHQ

15460
ELQQMDDAP

15461
EIVATYGQN

15462
EIVELATQY

15463
EKHAFSYSF

15464
EKHMTMAGI

15465
EKMAREWQT

15466
EKMWPLNWG

15467
EKVMQMHMQ

15468
ELWKAMPPE

15469
EMEINPTVE

15470
EMHMTNTGW

15471
EMTKYLNHA

15472
ENHCWSAKE

15473
ENNAERWMF

15474
ENVQCELMQ

15475
ENWTWKPEE

15476
EPMYFIVCS

15477
EPVNFNEQV

15478
EQDGHNINE

15479
EQEQFHICG

15480
EQMLWNQIS

15481
EQQWNSVNE

15482
ERIMHLHES

15483
ERMPLTCMS

15484
ERSEHPCMF

15485
ESCWPQVTG

15486
ESEPEVYGW

15487
ESHNTTEGH

15488
ESLENPNDR

15489
ETDNFSDCA

15490
ETFVTCTGA

15491
ETHHGQMFC

15492
ETHMYERRE

15493
ETHVVQNDN

15494
ETELAPSGT

15495
ETLMTVKKC

15496
ETVRQGNPE

15497
EVARKVQNE

15498
EVAWMGRQN

15499
EVDPWGHME

15500
EVHVNWKDE

15501
EVIDHGHME

15502
EVLEWRHYQ

15503
EVNESGEYV

15504
EVTEPVKAI

15505
EVTKHLVAC

15506
EVWTPCFPI

15507
EWCNEWCGP

15508
EWDEWLISF

15509
EWIHLYCCF

15510
EWIPYAVGN

15511
EYDYIFNHH

15512
EYEADTITC

15513
EYMGFRCDR

15514
EYMSIMMNH

15515
EYNPTNHIL

15516
EYPAEFAQC

15517
EYQQLARHW

15518
FACMPNEFL

15519
FAFDTQMVF

15520
FAPPTHTLE

15521
FCHNIFTEC

15522
FDHSERQYH

15523
FEKTRIAAW

15524
FEMGQKSLS

15525
FFEFRPSTK

15526
FGMVMPMWA

15527
FGPDSTHLC

15528
EGQIIMTDG

15529
FGWYYFWDP

15530
FHSCSRMGL

15531
FIDTACSHG

15532
FIGDIYAGG

15533
FIGMQSALE

15534
FITMRQEGP

15535
FIVILNNSD

15536
FNPDTDQDH

15537
FPFGTMHGS

15538
FPFLTKSPK

15539
FPQLPQNEA

15540
FSMDKQQFQ

15541
FTCEYTYAY

15542
FTELSMVQD

15543
FTKQADELP

15544
FTNFEYSME

15545
FTQGGEVYA

15546
FTSWMDVQD

15547
FTTQPSVAW

15548
FVPQWVQIP

15549
FVSSYYAIH

15550
FVSTESVWN

15551
FYIMMEKML

15552
GACCKQIGE

15553
GAKWQAQPD

15554
GATQHECWC

15555
GAYDTVWGW

15556
GCATWHNLY

15557
GCMLRSVTG

15558
GCNLLSVSN

15559
GDAFSCVFP

15560
GDDPMFFEK

15561
GEHTGMMLY

15562
GFMAVVQSW

15563
GGRCYFWED

15564
GGRLYKPEW

15565
GHASWAMWY

15566
GHGHKWIYS

15567
GHMYLLDME

15568
GHTCPCKWD

15569
GIDTDDVGH

15570
GIGMFCVTN

15571
GKARRCATW

15572
GKDVKQCCD

15573
GKLGLDCQD

15574
GKVVDAHMG

15575
GLQNKGYPT

15576
GLSIEISTP

15577
GLSLHKSDH

15578
GMADMVHGP

15579
GMASHSHYI

15580
GNEFIECRQ

15581
GNEFIECRQ

15582
GNHWGWSIS

15583
GPEMTVAQT

15584
GQIPCSSWG

15585
GQVDNMMAT

15586
GRELEIEPQ

15587
GSDRATGKE

15588
GSEFCILNE

15589
GSKPSSKQS

15590
GSSHFNDLG

15591
GTCHGPVLM

15592
GTCQVQLHS

15593
GTEIKTLHG

15594
GTEVIGLNT

15595
GVFLGYAQR

15596
GVISDGKYA

15597
GVKDFDVCT

15598
GVMVNPCNM

15599
GVSAEGKAQ

15600
GVSGMQITG

15601
GVTMNTECE

15602
GVVATDCEI

15603
GVYMESAVR

15604
GWETPLVWF

15605
GWWNHYLFW

15606
GYIYHKMYT

15607
HAAETEIHQ

15608
HCDTHVNLE

15609
HCKSGQQML

15610
HCTCYDAIF

15611
HDKWKWSDR

15612
HENYYREGA

15613
HFLQMEHGH

15614
HFMYDMNFQ

15615
HILKHVHTP

15616
HISFHTIPG

15617
HITGEYIAP

15618
HKESSEGAT

15619
HLNNCHWRR

15620
HMGQMYWHM

15621
HMVDINLIA

15622
HNGVMMHSH

15623
HNKACITAL

15624
HPNNFNRAH

15625
HPVHYNMCS

15626
HSCTMMGGD

15627
HSMGIFSNL

15628
HSMVQWNST

15629
HVEMANAGK

15630
HVIPHNRSE

15631
HVMPWLCQW

15632
HVMTLVDYD

15633
HVNTEQKYY

15634
HWNCNCRNA

15635
HWPQRLNEH

15636
HWSWFASFV

15637
HWTNQDSNK

15638
HYHQYTRLP

15639
IADVIHSKW

15640
IAETLIGVL

15641
IAHGASAYL

15642
IAYSLLANY

15643
ICMFEEEQW

15644
ICPPFQRQV

15645
ICPYAEVSD

15646
ICSNPWNKE

15647
IDCQGKCDW

15648
IEDDPLHAE

15649
IEHRFMIAG

15650
IEPIHNWKK

15651
IESVTHLCD

15652
IGELDEACD

15653
IGFMDNTSC

15654
IGIPIHMWD

15655
IGQSIVTMQ

15656
IHMFRMWPE

15657
IHPQIAEGM

15658
IIDVFPRHT

15659
IIFFLCRDP

15660
IKQPANDTD

15661
IKYGMRHTN

15662
ILVMVWLGQ

15663
IMRWHYGMV

15664
INASSCDYW

15665
INRNYWDFI

15666
INSELPVDV

15667
INYSSVATV

15668
IPQPTMMQD

15669
IQEYIQDAD

15670
IRESWIGQF

15671
IRYVTAYYK

15672
ISEIMPSNP

15673
ISMYQSTQV

15674
ISNWRCSQK

15675
ITCSNEKDF

15676
ITENRGYYH

15677
ITSNELPME

15678
IVEFMHCDD

15679
IVEMYAKNN

15680
IVHDEPRHN

15681
IVHDIWGQA

15682
IVKYRAYPG

15683
IVNKNCSQP

15684
IVPFGCGGF

15685
IWKCLNTQN

15686
IWTCCKGGG

15687
IVWDYEITR

15688
IYEARTIVR

15689
KALPGMSAG

15690
KAPGVCTRS

15691
KCDMKRGQF

15692
KCELMSNQT

15693
KCWYKFITN

15694
KDALFDFPC

15695
KDDGMCYGR

15696
KDEDWNWCD

15697
KDFWAMSMV

15698
KDIGKAAGY

15699
KDVMFREMH

15700
KECEQLMFP

15701
KFDHDSEFY

15702
KFEPRYGDC

15703
KGIHEPISC

15704
KHDSDWMWY

15705
KHIQDARWF

15706
KHWREMITV

15707
KIDDDSRVY

15708
KIQYDDFDG

15709
KIVDIDWWA

15710
KLMWFASTD

15711
KLSNDRFIY

15712
KMQTVMRAL

15713
KNDMLKKKG

15714
KPIPQTVWL

15715
KPQAHYYTI

15716
KQLLTRMYM

15717
KQPEPTPLA

15718
KRMQEMTCR

15719
KSASPTQPY

15720
KSPAIHTEE

15721
KSSGVWMCC

15722
KTEWYDMKN

15723
KVEENLRSM

15724
KVIPEDQNC

15725
KWDSTCYRW

15726
KWTEFDRCD

15727
KYEEQYHFH

15728
KYQPDGWCY

15729
LAAGVWDQE

15730
LACKREAMW

15731
LAVCKTHEP

15732
LCHPGEHWQ

15733
LCVTGEDTH

15734
LDESWSVAG

15735
LDPNRCMDQ

15736
LGFWETQIM

15737
LGHMQASTP

15738
LGNQYWWHF

15739
LHEIHLETN

15740
LHLPQLLHD

15741
LHSAWMHLA

15742
LIAHWIHSF

15743
LIAPYFGQL

15744
LICTMSDMN

15745
LIDHIQNQH

15746
LKEMTWINW

15747
LKQFRHMDG

15748
LLYMAHCPP

15749
LMDGCTPFM

15750
LMEVYRSKA

15751
LMHYGMVWP

15752
LMSKENNCN

15753
LNGECVVGY

15754
LNTDVAEAF

15755
LNVGKKGYG

15756
LQQCEPMKS

15757
LQSEHNEDE

15758
LSICDQYDV

15759
LSMRCMDQW

15760
LSNFSWQYG

15761
LSVKHVHYS

15762
LTCMWGVYP

15763
LTTLVEAIP

15764
LTTQWNAKG

15765
LTVFSNGGV

15766
LTVWFGPFS

15767
LVDTWNNHR

15768
LWCKTWHNY

15769
LWDGEAWCW

15770
LWNMIKPLY

15771
LYELQQFGA

15772
LYHWWGYIE

15773
LYQSTFDWR

15774
MAADRWPLG

15775
MAGQSAAYW

15776
MAHIEGVAN

15777
MASDTDYSF

15778
MAVMIANAA

15779
MCGLTKFRP

15780
MCPPYQQAE

15781
MCSSQHSYS

15782
MDDDNDPVF

15783
MDKDSEIMG

15784
MEEYNTTTC

15785
MEGGHPYRT

15786
MESKYWHSD

15787
MFHDTLGMF

15788
MFMRLGTLP

15789
MFNMYEGSQ

15790
MFNWAEMED

15791
MFTDNITKC

15792
MGIQTNYEA

15793
MGNMPASMA

15794
MGRTFFEEF

15795
MGRWENIWC

15796
MHDMLMGQM

15797
MHFQAWSGA

15798
MHIGVFTQA

15799
MIAFWSTWR

15800
MIAWWGSPG

15801
MIKPLICNK

15802
MIKQTGAEW

15803
MINNSKTPY

15804
MIRTQCCWI

15805
MISQMMSNA

15806
MIYWTVDQQ

15807
MKASKDGGK

15808
MKEEWGHIP

15809
MLSWVYPCM

15810
MMAMKTDSH

15811
MMCQIMHSC

15812
MMNGMMGGN

15813
MMQEIGAEA

15814
MMRGPESDR

15815
MMVQQMFFI

15816
MNESTHIHP

15817
MPCCWWVVT

15818
MPCPLPKYR

15819
MPGYQIHNA

15820
MPKDWVPDA

15821
MPLTHSMHN

15822
MPNAFYQAC

15823
MPSQNGTKH

15824
MQLGEHSWY

15825
MQLLGDSEY

15826
MQSQEDHSP

15827
MRVWRYCDR

15828
MSSCKKMDA

15829
MTGEKTGNA

15830
MTLSWWGIG

15831
MTMLKMAGR

15832
MTNGNTVGA

15833
MTNIMQQGK

15834
MVDGDLPMK

15835
MVDKGFAGQ

15836
MVESELMMC

15837
MVGCCCHFG

15838
MVGPAQIEL

15839
MVHFTSHPI

15840
MVLTDAHTA

15841
MVQCMGQLA

15842
MVQPGHFES

15843
MVVASSASD

15844
MWDSVYYPF

15845
MYAPRCQTE

15846
MYGYTMNSG

15847
MYSCWTVNT

15848
MYSNLVHWS

15849
MYSYPHRRS

15850
MYTTHCHGM

15851
NAGLVNLGK

15852
NAQEEPESW

15853
NARYDKDNL

15854
NAVEQYVHH

15855
NCKEKEDCQ

15856
NCMASGWRG

15857
NCWINTRLP

15858
NDAKNLEGW

15859
NEKEYSWAA

15860
NEKHMSYCF

15861
NERHTFCVD

15862
NEWLMYLLT

15863
NFALATTTE

15864
NFTHREKEC

15865
NGKCCHPGP

15866
NHALMHGFQ

15867
NHCINSTEL

15868
NHESKDASW

15869
NIGFQMTMG

15870
NIGYANLWC

15871
NIQFQVGNY

15872
NIQWITDVG

15873
NIVHYCNPS

15874
NKSDEQLVQ

15875
NLTFCLFMG

15876
NMGVYQFLG

15877
NMVEYKQTA

15878
NNDCENPPL

15879
NNEFDVQHS

15880
NNMEIMKTC

15881
NPKQTHWDY

15882
NPRGEFMAH

15883
NQLQPICKP

15884
NSAHELHSP

15885
NSGTCPAPH

15886
NSSTVCNNR

15887
NSVMQMCVD

15888
NTFMPNGAN

15889
NTHMAHKSE

15890
NTQTLMTNG

15891
NTSLVRESE

15892
NVECVQQQW

15893
NVHDDHGAR

15894
NVMAGHDHE

15895
NVMFEQDDR

15896
NVQMQTLSW

15897
NVWTCDVCS

15898
NWFKMQAGM

15899
NYAETPCYS

15900
NYNFYATHV

15901
NYQNNMTFN

15902
PDVLHPNND

15903
PFRTDVRGS

15904
PGGTPRPEY

15905
PGKTWLWEL

15906
PGLDSTMHY

15907
PHPKSHKQH

15908
PIKYKHRQP

15909
PIRREWHLW

15910
PKEQYEWKD

15911
PKVDKRHKL

15912
PLASMWQHD

15913
PLMLLSKAM

15914
PLTFRGHNH

15915
PMEKFYCGC

15916
PNTERTYEC

15917
PQHAGRHHE

15918
PTDFIWPMM

15919
PTDQDQMLE

15920
PTKMHNHPW

15921
PVKLETHES

15922
PWGRSLVHC

15923
PWPSMACQK

15924
PYAPKNCCV

15925
QAIGRDQSF

15926
QAMWGNGQG

15927
QASVHVMAD

15928
QDHALVYRN

15929
QDYYSKPEW

15930
QEGIRTDVS

15931
QEMPLPMKS

15932
QFAEWWTMA

15933
QFILSHRRD

15934
QFLPCNFRC

15935
QFMTCQNYP

15936
QFVSWYTYR

15937
QGEMPDMFK

15938
QGNSPYHEA

15939
QGQPEVVVW

15940
QGQPTDASG

15941
QGYHFAWND

15942
QHALNQAVA

15943
QHIHEIGKK

15944
QIAMATMFR

15945
QIDNVRHHP

15946
QIMANACQN

15947
QITDMQDMQ

15948
QLVEAQLPP

15949
QLVEKEDMN

15950
QMSPAGCTE

15951
QNKVTQREC

15952
QPSPGHAYP

15953
QQDAAFHWT

15954
QQGFIVYWH

15955
QQIHMGHQA

15956
QQTLFRTGS

15957
QQYRASNFA

15958
QSCVTLQTN

15959
QSEFLVKHI

15960
QSMDCNWDG

15961
QSPSDIQCT

15962
QTADHRKGM

15963
QTATMAVWN

15964
QTDHFDSCR

15965
QTRAIVLLD

15966
QTYAQTHRT

15967
QVDWLKSSF

15968
QVEDELANY

15969
QVEMKDCDQ

15970
QVFPCDYEN

15971
QVMRCDFDD

15972
QVNLENHSC

15973
QVSVDCSQC

15974
QVVVIVWQA

15975
QVWINNVNI

15976
QWGTMMKNQ

15977
QWMTGMCWE

15978
QYAHISAHS

15979
QYQSVSNFS

15980
RCAQYEVAG

15981
RCLKRKTCF

15982
RCTGFDDFE

15983
RDETSFRHC

15984
RDSDKERCK

15985
RFHTDYRRH

15986
RFVMRPKES

15987
RGEQGVLTL

15988
RGLFDNWIF

15989
RGTYSYSAR

15990
RHLDQIPWP

15991
RHSWHPHFE

15992
RHYFYQDDD

15993
RIHEGSISG

15994
RKKGELCSR

15995
RKWCEFDAI

15996
RMCCSDSGE

15997
RMEQQGVMN

15998
RMESHIAQT

15999
RMHMPWCHG

16000
RNPHMDQSM

16001
RPIHDFGVY

16002
RTDQMELAT

16003
RTETKSTYF

16004
RTYWLDDGK

16005
RVDQEGGRD

16006
RYECYRSPF

16007
SAESRQGES

16008
SATPVGKEY

16009
SAVHRNDVA

16010
SAVHYTYMD

16011
SCAFVDEWS

16012
SCDAVDGLW

16013
SCDLKHDSG

16014
SCTDFNSMG

16015
SDHYNPEPY

16016
SEEESKEGG

16017
SEFCAWKCD

16018
SELVRWKHS

16019
SEQNRGSHT

16020
SERKDQRMW

16021
SEWQEFFFN

16022
SFRIANQTN

16023
SFYSTGIWG

16024
SGPDMTAGN

16025
SHRWVMKEH

16026
SHSHQPSAS

16027
SECGAVQNF

16028
SIKGRDWGD

16029
SIMVSTHNN

16030
SIQPLAATT

16031
SKMEEALRM

16032
SLRLELRPQ

16033
SMKHRMQAF

16034
SMLQFLSDD

16035
SMYFKPTQD

16036
SNEMCGFAM

16037
SNLQIQDAK

16038
SNSPTCYAY

16039
SNTMRLNAE

16040
SNYAPFYCD

16041
SPHCYESTP

16042
SPMVKKSGT

16043
SPMYEMSAF

16044
SPNHDAGQN

16045
SQDTEHCWA

16046
SQGCQVKQC

16047
SQMPHSKAA

16048
SQSNFEIME

16049
SRHWRPQSC

16050
SSGLYWKKN

16051
SSLNYSPGA

16052
STAAYQFEE

16053
STAEDQADH

16054
STAPDNTAF

16055
STDALWQEN

16056
STDCIFFQH

16057
STDSMDAEA

16058
STEHLGRND

16059
STEPSGIHQ

16060
STGQDQEHT

16061
STHGQASAI

16062
STKQVEALG

16063
STVNTSSPE

16064
SVCIDIPCI

16065
SVDYTNALT

16066
SVENYERCQ

16067
SVRFDGNDW

16068
SVTVNNVHC

16069
SWAVLHQAQ

16070
SWDMEWDGP

16071
SWDVPLYNP

16072
SYCNLQFMR

16073
SYEKSVNMG

16074
SYQAVFGSI

16075
SYVQRHCVF

16076
TAFSGWKEQ

16077
TAGLGIESV

16078
TAMDCMCGT

16079
TAMKFPGGE

16080
TASPAFDES

16081
TASSWLMHQ

16082
TATGMNRCE

16083
TATSWWAFK

16084
TAVWMDKTS

16085
TCFMGWRCL

16086
TCRMGMEIG

16087
TDQTCSKCE

16088
TEEICQLRE

16089
TEKDNEEHQ

16090
TEKMSHCSE

16091
TESQSGGQP

16092
TFARRADYA

16093
TFPVGYNYT

16094
TGEWQQGKP

16095
THMTIDDAS

16096
TIGVDPSCQ

16097
TIKHILNHK

16098
TIKNYCQGR

16099
TIKYDLHFA

16100
TEMPRHRHA

16101
TITTFWDFE

16102
TKRPFKGQW

16103
TLHHNSDAK

16104
TLIYNAQVT

16105
TMDQKNDMD

16106
TMGTAAKSH

16107
TMPVDVMIP

16108
TMPWQGNHC

16109
TMQNTEPGI

16110
TMRFFGEGD

16111
TNADSWDQE

16112
TNGVPPWWS

16113
TNHINFGWI

16114
TNHSFWEQG

16115
TPDCYSMGA

16116
TPNCGINHE

16117
TQCNTDQRT

TABLE 77

Sequences of the 581 to 589 Region in AAV5

VP1 Capsid Polypeptide that Drive Lung

Tissue Tropism

SEQ
581-589

ID NO
Sequence

18118
QTNTMQMLS

18119
GPRHIEEDG

18120
RKADVEYQG

18121
SCDMQQAHQ

18122
DILMFEWFD

18123
DIRMFAWFD

18124
DIRMFELFD

18125
DIRMFEWFD

18126
DIRMFEWFG

18127
DIRMFEWFN

18128
DIRMFEWFY

18129
DIRMFEWVD

18130
DIRMFKWFD

18131
DIRMFVWFD

18132
DIRMIEWFD

18133
DIRMVEWFD

18134
DIRVFEWFD

18135
DMRMFEWFD

18136
EGKQAYFFP

18137
ELKQAYFFP

18138
EPKRLLRTV

18139
EPKRLLWIV

18140
EWEQAYFFP

18141
EWKPAYFFP

18142
EWKQACFFP

18143
EWKQASFFP

18144
EWKQAYFFA

18145
EWKQAYFFP

18146
EWKRAYFFP

18147
EWRQAYFFP

18148
GIRMFEWED

18149
GMAHRFRGR

18150
GMAHWFRGR

18151
GMTHRFRGR

18152
HIESFTYCN

18153
HTESFAYCN

18154
HTESFTYCN

18155
HTESFTYCS

18155
HTESFTYCT

18157
HTKSFTYCN

18158
HTQFINGRG

18159
HTQFTDGRG

18160
HTQFTNGRG

18151
GMTHRFRGR

18152
NERMFEWFD

18153
NESFPNQSE

18154
NTESFTYCN

18155
HTESFTYCS

18166
QLNARWYRC

18167
QLNARWYRW

18168
QLNAWWYRC

18169
QLSARWYRC

18170
RLDYRHYSK

18171
RLNYRHCSK

18172
RLNYRHYSK

18173
RLNYRHYSR

18174
RLNYRHYST

18175
RLNYRPYSK

18176
RLNYWHYSK

18177
SKNLCTMAG

18178
SKNLYTMAC

18179
SKNLYTMAD

18180
SKNLYTMAG

18181
SKNLYTMAV

18182
SKNLYTMSG

18183
SKNLYTMVG

18184
SKNLYTVAG

18185
TMTSTLQST

18186
VWKQAYFFP

18187
GTKLIWPYQ

18188
CSTLAKHDE

18189
DVIHTTHSG

18190
DFMWYHICS

18191
DVEFCMKQY

18192
MMLGYMEQD

18193
DFNGSNWLP

18194
AGRFKFPQP

18195
AMYQCPSMD

18196
ANWKHWQWR

18197
ATVNSDTLT

18198
AVGHFEYVL

18199
AVWTMLEHC

18200
CQVWWENRD

18201
CQYAVTKMG

18202
CSGVDCTGA

18203
DCSSCMKDL

18204
DRKLGDAHG

18205
DTARWDVMT

18206
DVVNCTVHT

18207
EICSRGRTI

18208
ELGHCLLYE

18209
ELQSFMQYG

18210
ERFKTFCEE

18211
FASSHWYQG

18212
FYLWDTRET

18213
GEANMAPSR

18214
GMSEQIAIT

18215
GWSTDWTET

18216
IASKFNICC

18217
IFVYCEKMP

18218
ITSSPNHIF

18219
IVDTSARYQ

18220
KANWVESME

18221
KTGWGGETG

18222
KWKVETTCY

18223
LLCTCGWCE

18224
LSYENDAFQ

18225
MCSHNLPKS

18226
MHTMTNAMK

18227
MNTHEQWGG

18228
MSLMPHGWH

18229
MTDLMYIQP

18230
MVECLYYHG

18231
MVFLQCQQN

18232
MWNTSNLFH

18233
NVNLQMMMT

18234
NYNGYSDAQ

18235
PSTKPPKCK

18236
QDWSDFNDC

18237
QNVQLENSQ

18238
SKTKKRFAW

18239
SRYDTNCWW

18240
STHNRLDWQ

18241
TGDEIVGTY

18242
TSIPPRHNP

18243
VAAYNFPTC

18244
VMEGVRNME

18245
WTHMYENED

18246
WYNQGAEYH

18247
WYWRLDCLW

18248
YATEPGVFP

18249
YCHRSDCDR

18250
YMDEMITEL

18251
AAMPTHHTM

18252
AANIMISQA

18253
AANVTQDTS

18254
AAPKYSAHQ

18255
AAQPHFVVA

18256
AASDWCRYS

18257
AASWQEFFP

18258
AAYFVGENC

18259
ACEWKQHWA

18260
ACGKMHGPN

18261
ACGVVLKAA

18262
ACKQFKKHA

18263
ACLQQDIAM

18264
ADAHQRTVS

18265
ADGQMLQAH

18266
ADRQSQMHL

18267
AFLELAMYP

18268
AFTEGQLNE

18269
AGNRWWWSI

18270
AHASWKCWF

18271
AIGTLAPNL

18272
AILMQQGAG

18273
AIMHPPYCE

18274
AIQSGNWSS

18275
AKLEQMDSA

18276
AKTHCHCWW

18277
ALHHCIKKA

18278
ALIQFMMDQ

18279
AMAYSWTMA

18280
AMCFNKCWQ

18281
AMSQQHDHQ

18282
AMSTLGECP

18283
APQFYVSYD

18284
APTKVHCQG

18285
AQNPFDIDA

18285
AQNREGAKL

18287
AQTFCSFDR

18288
ARSGENGHE

18289
ASGCLQQIS

18290
ASEDVASWG

18291
ASTPWCWFE

18292
ATDNSDTLT

18293
ATGIDPLGA

18294
ATMIKFECT

18295
ATYISAHDA

18295
ATYISAHDA

18297
AVESPERYH

18298
AVNDWNNDW

18299
AVTRPSNKM

18300
AWDPMTSTF

18301
AWGGFNAEM

18302
AWILFWWFV

18303
AYCSEERSP

18304
AYENVQHPN

18305
AYEPFASSF

18306
AYNRMMLTC

18307
CAFPVNITA

18308
CAGVTSHGE

18309
CAICNAQCP

18310
CAQSVHFHK

18311
CAYEVYDGK

18312
CFCKNGTYL

18313
CGEWLNECS

18314
CHHFNMPSQ

18315
CILHVTDNG

18316
CKSNWHIFK

18317
CLMSNITHI

18318
SPFQHLGPK

18319
CQQYWNAAW

18320
CSAVYLANH

18321
CVDEYYNSS

18322
CVIDWKAMS

18323
CVTKTMWTG

18324
CWRSGSMNL

18325
CWSITNQDH

18326
CYASMSASY

18327
CYDWYEGAE

18328
CYSENDHQV

18329
DAAHMGYEP

18330
DAGHSFQIF

18331
DAGPIWYSS

18332
DANQRATNS

18333
DANSLNSWP

18334
DARSMAPVE

18335
DAYSVHHYV

18336
DCNSLINVA

18337
DCRHTILYV

18338
DDMLMQFQH

18339
DFGCMDYGP

18340
DFHSYLKQQ

18341
DGFQQGEGI

18342
DINRSGNQC

18343
DKWCCEMTA

18344
DLEY1WHWG

18345
DLIGSGPWF

18346
DMHECIGCS

18347
DMHRQHLIE

18348
DMLKFHFNY

18349
DMMDNQIAS

18350
DNIRYPCHS

18351
DNQFFNGDY

18352
DPVWLPFMF

18353
DQFARHKKY

18354
DRPKKNLCC

18355
DRPSTGWYL

18356
DSDIVNDYC

18357
DTKVKKEFG

18358
DTMLRLCYP

18359
DTNWMQWNH

18360
DTSLNDGWM

18361
DVCCQFSAY

18362
DVLVATQVQ

18363
DYQRKTRYS

18364
EACKCSVVP

18365
EAFYLQKPG

18366
EAIQDAHQM

18367
EANKGASQD

18368
ECCNMSPPG

18369
ECGRFAPME

18370
ECHKCEVCV

18371
EDRPMAMME

18372
EEDKYNVVQ

18373
EEKKLTVIM

18374
EFGTYDNIG

18375
EFRYGPQYG

18376
EFVIAEETS

18377
EGANYHWDH

18378
EGAQQESVG

18379
EGARYTKFH

18380
EGGMGPQQS

18381
EGKWMSVHH

18382
EGRPQYTDP

18383
EGSFVLSTE

18384
EHAPMVHAL

18385
EHQRMKFRN

18386
EHVQKVNYP

18387
EHYCVTAWY

18388
EIDLWRTPT

18389
EIHDEASCY

18390
EIHKEMGFG

18391
EIMFYMRKQ

18392
EINIKKKQM

18393
EINKMAALG

18394
EIRTVESAG

18395
EISWIPREN

18396
EKHSVENYG

18397
EKIDWQSCV

18398
EMAICAEHS

18399
EMGRLPNHN

18400
EMVDQIVDF

18401
EMYPEAAAK

18402
ENYPPSQCA

18403
EPNHPESLA

18404
EQKVGNQLS

18405
ERCMSEWSF

18406
ERWSGTRRG

18407
ERYHFMTHQ

18408
ESKYNRKYN

18409
ESRDAVCPD

18410
ETCHTYAQH

18411
ETHGDRKSR

18412
ETICCDSAH

18413
ETMGVHYCE

18414
ETRMQRRSQ

18415
ETWPLIDDA

18416
EVADCLKHF

18417
EVCNEMPEP

18418
EVELDERLS

18419
EVELYGCIC

18420
EVESQEHLH

18421
EVTRETDYM

18422
EYCMLEDYQ

18423
EYNRRYLLE

18424
EYQNYQLVT

18425
EYTRTQWAH

18426
FHHLNTPEM

18427
FIFEEQRWA

18428
FKETWLFPT

18429
FLCCPKECP

18430
FQRLEMKQH

18431
FQRWEMKQH

18432
FSGKYDQRG

18433
FSQYPSAFA

18434
FSRPFEQYH

18435
FVAQMWDDH

18436
FYRQLDEAG

18437
GASSKSLDA

18438
GDIYYKCWV

18439
GDLWNNHLE

18440
GFEDRGNQE

18441
GFMLRMSTD

18442
GFSLFMMNR

18443
GGMTVLIEY

18444
GHQLMFFKF

18445
GIKEQRRFV

18446
GIRMDLCTF

18447
GITHDFMAG

18448
GIVIKPHGE

18449
GIYPQFTWQ

18450
GKEFALSAF

18451
GLATLGLSG

18452
GLGMVHRMH

18453
GMMWRSKWI

18454
GMRCWRHRL

18455
GMRYSEWHW

18456
GMVDVCDCC

18457
GQHTIHKNS

18458
GQTVQPCST

18459
GSCCLNNET

18460
GSLTMPDAH

18461
GSYYVNWDQ

18462
GTCLQDNYN

18463
GTGWEASAD

18464
GTGWGASAD

18465
GTLSSSIVE

18466
GTNQVCLFT

18467
GVDWRHQQN

18468
GVISEGISA

18469
GVMVIAGMG

18470
GVNQYDEMI

18471
GVVQMVIGD

18472
GWMDLMSAR

18473
GYDHHGTEH

18474
HCMEYIHHA

18475
HDCEFLKDE

18476
HEVERRRAK

18477
HGMQLEAFF

18478
HHNIKEAWM

18479
HIESYMCRD

18480
HIRHRDENG

18481
HKNRLLRSV

18482
HKSRATHTG

18483
HNHILIQTS

18484
HPMDGLIQV

18485
HSEHQGRMV

18486
HSFWIAQMQ

18487
HSGIGLTMQ

18488
HSWLENSAC

18489
HVEYVQKYE

18490
HVFQVSDED

18491
HWHNVISKD

18492
IADTPDALQ

18493
IEDISKKWA

18494
IFQPCFRDW

18495
IGQSTQYCA

18496
IHQSVIQKG

18497
IISRYDNDD

18498
IKYSIDHFN

18499
IMELTHHNA

18500
INAVPWQLL

18501
IPAGMTPWC

18502
IQAPTENNN

18503
IRPMYETEE

18504
ISAPGCVFC

18505
ISDRMNYMN

18506
ISERVHQTE

18507
ISQFGANVH

18508
ISSDKKHMG

18509
ITAPRMVET

18510
ITGCIQKAK

18511
ITKWSLGNP

18512
IVRTMDNEQ

18513
IYATNIAWP

18514
IYMHKIALF

18515
IYSQIGRED

18516
KCAGWMPSS

18517
KDVEECNSL

18518
KEYPRIGRM

18519
KGSNDGADK

18520
KHQSTCGQD

18521
KMDFWAVPI

18522
KQSELGIGC

18523
KSSNYHSWP

18524
KTEQGGDAE

18525
KTTDECWHV

18526
KVEDVGDYA

18527
KVTEAAKTM

18528
KWCMFFDQH

18529
KYELSQCFG

18530
KYHQAGQLC

18531
LADSSMLWE

18532
LADVQFLYV

18533
LADWSYEAS

18534
LATDVPVSE

18535
LCFQSVHEE

18536
LCTAVFRDQ

18537
LFMHMIWGA

18538
LFNCTVWLE

18539
LFRYQRKTC

18540
LGDWAPASI

18541
LGNSFKRAR

18542
LGQQAMPLK

18543
LHFKIEQVD

18544
LHNNMNAFG

18545
LIALDMCHP

18546
LIQSYVNTG

18547
LIRAKHRTM

18548
LKKEVQRYQ

18549
LKSEVSIME

18550
LMYWKNGMS

18551
LNAWVNRHS

18552
LNHKYERMT

18553
LNKYNPHKT

18554
LQDLTQLCK

18555
LRQCPLNKL

18556
LSDQCLDLA

18557
LSNQQWYAG

18558
LSRSGHDQE

18559
LSTDNNIQF

18560
LSTMEWKTN

18561
LSTNPQHCG

18562
LTAMKKQNA

18563
LTEVKWCNC

18564
LTGYLNDMT

18565
LVDKNMFIN

18566
LVTQWVPAY

18567
LWAWSGEPK

18568
LWNDMHFHG

18569
MAGRDDGRA

18570
MAIIRLAEM

18571
MAKSSRNDQ

18572
MAPKFSSYG

18573
MASFPMFNK

18574
MCQHEQATF

18575
MCTSVTNMG

18576
MEHSTHDDD

18577
MENYQWLAP

18578
MFGCIIGDF

18579
MGSKRKPWT

18580
MGVKVDGTN

18581
MHDGYRGMQ

18582
MHHCYQTFA

18583
MHIAQMLGP

18584
MHSNLHWYG

18585
MHSPCSGPW

18586
MIDVLSWLE

18587
MIESQGDLQ

18588
MILPSMAWC

18589
MISCGGDDH

18590
MKDHWDFYG

18591
MMIGTYMGE

18592
MMNQTCCHM

18593
MMVVRGKVE

18594
MNSCNNYCM

18595
MPVERNVTP

18596
MQEDCMSGE

18597
MQNEDVPHT

18598
MQQECVDMW

18599
MQQGYGIFA

18600
MRFILLADW

18601
MSQCIVMHR

18602
MTMQDINGQ

18603
MVWNTSMDP

18604
MYTMQQSGI

18605
NAAPISHIF

18606
NCENEWRLN

18607
NCERRKSAG

18608
NCGSLPVTE

18609
NERPYEGMG

18610
NFFSLHMMS

18611
NGCECHECW

18612
NGFQTRMRV

18613
NGWMNQSAG

18614
NIQWQWMTG

18615
NLNVMFSME

18616
NQEGQGMFT

18617
NSAHIMKDG

18618
NSMHGDCQW

18619
NTCPVESAH

18620
NTDNNWTSG

18621
NTNLLEITH

18622
NTTYRTEYF

18623
NVVFKQSAH

18624
NVWVIKDIC

18625
NVYFSWMWT

18626
NWTCLMSIP

18627
NYPLQWGVE

18628
PATDQIDEC

18629
PATRVFADI

18630
PDWMDWNHY

18631
PFVKSNIAL

18632
PKQLFWYWW

18633
PLFAITTIE

18634
PLTMGHGEC

18635
PQHNCGYNN

18636
PSENNKRKK

18637
PVDWIIAAE

18638
QAIFVRQLN

18639
QCGHDHQYT

18640
QCKYSERGT

18641
QFAEKPGSW

18642
QFEGGDSMG

18643
QGTGRYAWG

18644
QGYRKLMNQ

18645
QHHTYMCIS

18646
QIMFNHQNL

18647
QIVGNQKRE

18648
QKMEMEEGS

18649
QLETFTVHN

18650
QNDVMMGNH

18651
QQSFFFGEQ

18652
QQSSMATEY

18653
QQWDVAMMC

18654
QRVDQQSLG

18655
QSEWWRRHA

18656
QSHSMMYFG

18657
QSHSTDYCS

18658
QSQPDNKGM

18659
QSQYKAECK

18660
QSSGEERGT

18661
QSSWDGQDD

18662
QSTQTSYIS

18663
QSTSDECNL

18664
QTDIWMMWA

18665
QTDSVMECT

18666
QTTHFRYFH

18667
QTTQKSEER

18668
QVDTRYMNE

18669
QVGMVAISQ

18670
QVLWANNHD

18671
QVSFEWYKC

18672
QWPLNPYGK

18673
QWRILDVVK

18674
QYAMSYAGE

18675
QYYEHEFLD

18676
RADHTIDFS

18677
RCDTSQWWQ

18678
REGWPGHKY

18679
RENLCNVPP

18680
RGGLIHFST

18681
RHLDIQNHD

18682
RHMESLNSA

18683
RIHLWENNP

18684
RLCWMTPCC

18685
RLENFHEAH

18686
RMELRMIFP

18687
RMEPLHCCE

18688
RPYPAMFKG

18689
RQHHVGNEV

18690
RQPPLKWKR

18691
RTDFLACHC

18692
RTDFLACHC

18693
RTINFSLMI

18694
RTMYWPSVN

18695
RVDFTPIVH

18696
RVGMDPRDG

18697
RWAEMHFID

18698
RWAYLEEFR

18699
SAGWDGKVC

18700
SAVNYTMFG

18701
SAYVVCSHD

18702
SCDPRHVDE

18703
SEANMAPSR

18704
SELDQYTMA

18705
SFLITAYEA

18706
SGHPRHMCY

18707
SHMPFSDMC

18708
SHMQMNWVD

18709
SHTECHTWG

18710
SIISDEYFT

18711
SITPRDEKP

18712
SKIKPADPN

18713
SLSVKFKTN

18714
SMQQAHGWK

18715
SPGSCQAFE

18716
SQNQYIKAA

18717
SREICQLEY

18718
SSPWKDFRQ

18719
SSQIKTSSF

18720
STACAESGW

18721
STHPYMLSK

18722
STHQPDCFR

18723
STMCMVRDS

18724
STTHLQYYC

18725
SVGPQLYLT

18726
SVGTWQNMG

18727
SVHKAENWW

18728
SVLMQMGSE

18729
SVTTCHMPL

18730
TADHNLFDY

18731
TADWEDQTP

18732
TATEGVWGC

18733
TCCYDQRGP

18734
TCDSNATWY

18735
TCEHRLMAT

18736
TCGTTANFE

18737
TEVATGHWG

18738
TGEYTNSWD

18739
THGCGQIMP

18740
THLSCRMWG

18741
TIGIPYSHD

18742
TKKDLDRQW

18743
TLALDCKWP

18744
TLCIREIMM

18745
TLTMMQGCS

18746
TMFAVTVKN

18747
TMGFLWYEQ

18748
TNKMLGRAP

18749
TPGRNNQGM

18750
TPIGVQKLA

18751
TPSQQAEQC

18752
TQAPIWENL

18753
TQEHSHEVS

18754
TSDVRLTEE

18755
TSGHMRNMF

18756
TSIDETELF

18757
TSIWINAVK

18758
TSQQFITAY

18759
TSSWLQCYI

18760
TSTWPGVIA

18761
TTAISLIGE

18762
TTAYSPNDR

18763
TTNLEQMIP

18764
TTQDGQMYS

18765
TTQSMQSGS

18766
TVCMFWVWC

18767
TVDPFKRIW

18768
TVEEAVWAN

18769
TVMMPMRLD

18770
TVNICLQHQ

18771
TWNCTFTGG

18772
VAHQSPNMM

18773
VAQPCGQYF

18774
VCIDHRAYS

18775
VCQQWLTQN

18776
VDYPRKCDK

18777
VGEPLAMFM

18778
VGQWFFKAG

18779
VIDCCYSCG

18780
VIEQHIQQF

18781
VKWDKKTKQ

18782
VISTIGKDS

18783
VLHRTSYKH

18784
VMEEDKRSA

18785
VMKGVDKFH

18786
VNRCCYPNY

18787
VPEHMDRDQ

18788
VQGLLIFVS

18789
VQHVRYWHS

18790
VSFPTNTMQ

18791
VSLPHSKHL

18792
VSNHWMYHL

18793
VTAWWDTAE

18794
VTEVYERST

18795
VTGESETPN

18796
VTGTMLKDA

18797
VTKMPDFAP

18798
VTQYNPKMG

18799
VTRQVIESQ

18800
VTWSTHQWL

18801
VVWDVDYYF

18802
VWDALQNPM

18803
VYVQEGNCS

18804
WAHTNMVYT

18805
WFCFEDSPS

18806
WFMCDYNGD

18807
WGWSPWKKI

18808
WKTKVENPE

18809
WWCWMAGPV

18810
WWTHKYFYL

18811
YAHEEDQNP

18812
YAWISEYEA

18813
YDVTKLGEQ

18814
YEAWKVNID

18815
YHCCWTSHP

18816
YHNIATPIP

18817
YPNTWCVVV

18818
YQGPAVLCA

18819
YRTHCFKGH

18820
YSCHFNQKT

18821
YWNGRLKWD

18822
ITISNVEND

18823
CCFPHYFVG

18824
MFELHDHQF

18825
RLCDTHKCL

18826
NNRALMSVE

18827
DSCIKGLAC

18828
MREKLVHDC

18829
TTYHHRNTL

18830
TWNTWQQDF

18831
DSASPIMGM

18832
MMAMKQGFY

18833
KACMSHQGR

18834
TIGEMVNPS

18835
NWNHCHQWR

18836
SEDKDQGAQ

18837
CAVAGGEWM

18838
NTMLSWGSP

18839
CSIRGESAF

18840
TIDWCEPHC

18841
SFSHTDLEA

18842
AIEHGAWKC

18843
TLTTDKTYS

18844
KDGRNPHCL

18845
WHCVAPNSF

18846
ACIPQMHSA

18847
NYAMRRDSY

18848
IYMSTGNTP

18849
GYRNYFIWP

18850
MFTPCQEGL

18851
GSTWGASWH

18852
GQFRSGAYR

18853
VYILEGGIH

18854
NTQVISRWT

18855
CYMNFIVDG

18856
NHKMVQDAL

18857
DVYMAVVGS

18858
GITMHPMHH

18859
QMTSSTNCA

18860
MHGIARMQQ

18861
RVWSLEWHL

18862
EVYCYQLQS

18863
QENDERIWY

18864
AMTRCKAGH

18865
KPLASIGQY

18866
GAANRIQSW

18867
GHMFETFWG

18868
EAQPSEING

18869
ECLEVKRTC

18870
FGDHLRVMY

18871
WTHCTNNQI

18872
NAYYDSHCE

18873
EKYQIHWDR

18874
QALVGNAAE

18875
QSSSSYLWG

18876
LESVKMDCN

18877
QRSVFHQRS

18878
NDVWQPHPN

18879
EDQWISNPF

18880
DHLSNDAEM

18881
LHKAYVTDS

18882
AVDHYSNGA

18883
DMKQKAEYT

18884
IMYLKRCCT

18885
DQRHGNVSP

18886
RVTIMFTGT

18887
TTYPHNTHG

18888
PPRFLQTTG

18889
YPTMTFPEE

18890
GCEEVGRCQ

18891
MAAITQNVA

18892
WPTILSAHM

18893
KCLPYMEMF

18894
AHAWLQHWA

18895
PDLWYEKSS

18896
DTRRNLCCD

18897
LIEKQRWML

18898
ENQSTMSHT

18899
QWMHCQLSR

18900
ICHIRTRGN

18901
YISEYVMMV

18902
QATGMHRWQ

18903
MFMTNVNSF

18904
KTVETYAAH

18905
HIKVEHEEV

18906
KMAGIGCWY

18907
QCTAHNTCM

18908
RTRISQMFG

18909
MGYAKHDSH

18910
PIWQGYWPW

18911
RCGEIMGLD

18912
LPSHCNLGS

18913
PAHINDQYM

18914
DRVNLMWKD

18915
QCVDCPPYC

18916
AVHTADVCC

18917
NQREWHGLA

18918
HIHMSDRGA

18919
PCCWEQSTN

18920
AFLHGQMWQ

18921
PLTIEVNCT

18922
IAKIGNCVW

18923
YFPSIQCYE

18924
NMPSLRTRI

18925
CKNHYMMAA

18926
SECEVLCSL

18927
ERQAHNHYA

18928
ERFHSYPLG

18929
MAEQMERDF

18930
DQEAWVVTG

18931
ACVERLHHK

18932
NHSLCWDSK

18933
MYDTMGAWC

18934
QRNEMSASR

18935
GHLENNNEW

18936
MLPLEMKNL

18937
ATGTYNLQE

18938
YVDHWTLGD

18939
IAQSEADYW

18940
LINSIVCGD

18941
LVQKWMRTC

18942
AAMESCAFI

18943
DMTLQSVSS

18944
AEDTPYTIY

18945
HFQPVFMQP

18946
LSPWPQTDQ

18947
AGMYKCQED

18948
PHAETHWYT

18949
YPDTADIFW

18950
VTMIHNAFH

18951
DLIVATEWF

18952
HSMMPGWPS

18953
IWITCPYAA

18954
PYGPIEYDE

18955
ATMNLSWGL

18956
NGVGRSGDN

18957
RCADNPYFR

18958
WERCLELNQ

18959
SNVACIQAF

18960
INYISFVDR

18961
LEIGGKTRT

18962
YDYWMQAPT

18963
QSAHPEPMC

18964
ASMHEETCL

18965
DMIDPVRHG

18966
ADRSNSSCF

18967
AEYSTNLHS

18968
AFSYCMAHM

18969
AIDYKTDWT

18970
AKACHLWGK

18971
ALQMAYEWS

18972
ANDIAEHWG

18973
ANVCIITHC

18974
APFMMQCFS

18975
APYPSWPTY

18976
AQWRIMFEA

18977
ARDCWWSDG

18978
ARDRPHRGS

18979
ASCRCLNWH

18980
ASQRSGRQP

18981
ATEHVRSWW

18982
ATIMFIEIS

18983
ATKCVLGLG

18984
ATMYMERAD

18985
ATYEHSCTK

18986
AVNHAQCGF

18987
CARLGGRGN

18988
CCTELNMSG

18989
CFEFLWEHE

18990
CLVRIQKDH

18991
CQAIINADT

18992
CSTMPKLPE

18993
CTMSPGYDQ

18994
CVVMAVDNT

18995
CWGWAPPDD

18996
CWPQSCKPE

18997
DGENMDEMA

18998
DIGRFEAYY

18999
DIKLVEAYA

19000
DKDEFQMAP

19001
DNDMYHSMQ

19002
DNNFKDKEE

19003
DSFYPQLQN

19004
DSQLESTLA

19005
DTINWIASQ

19006
DTLKSDNYI

19007
DTRGKAFFP

19008
DVITVYWDS

19009
DVTDDQIHM

19010
DVVKSWQMG

19011
DYSLLWMYE

19012
ECAFEHVDE

19013
ECMQYFPEW

19014
EDRLIVRMD

19015
EHITEQDLW

19016
EIGMEYGHS

19017
EINNAIASR

19018
EIQQMCVKW

19019
EIYMCKEGN

19020
EMSWKMKPT

19021
ENHPPWLHC

19022
ERDSLWFGK

19023
ESPQIHIYS

19024
ETVQDVGLH

19025
EVSWVPTTK

19026
EVVNSQFNL

19027
EYEWVDEPQ

19028
EYFKLRQFG

19029
EYSNETHQE

19030
FEADNTMVH

19031
FGHFEIKKM

19032
FIKLTTLPS

19033
FLEAMQRTP

19034
FLQYSMSGN

19035
FMDECLRVD

19036
FNSSEQQQM

19037
FSHKESVWC

19038
FTHWVNEQL

19039
FVKVVWGHK

19040
GDNMDQTQG

19041
GFKKFHFTF

19042
GKERFHRID

19043
GMSLKNYEE

19044
GNYHLDQDE

19045
GPNGDVARY

19046
GQQTMEMSY

19047
GTMHALKDE

19048
GTQSERDIQ

19049
GVEFVPIGN

19050
HCIFKDPKI

19051
HEVDRPAPC

19052
HITNYGYCG

19053
HNPATECRC

19054
HPLTQMVLE

19055
HPLTQMVLG

19056
HRRRAKVWV

19057
HTRYWGFAL

19058
IAGMHEFQQ

19059
IFILIEWHH

19060
IKTITYHST

19061
ILWSQHEVL

19062
ENISTPVNC

19063
ISHQCHQIH

19064
ISTQMSWDG

19065
ITFCCRAAK

19066
ITQYTTYEG

19057
IVAEHDPGF

19068
IVMPALMCK

19069
IYKLFQHGM

19070
IYRLFQHGM

19071
KADNRNWSC

19072
KAQPSEENG

19073
KCLPYMKMF

19074
KCTAEKAQC

19075
KCTVEKAQC

19076
KEREERRFW

19077
KKYFPTGIG

19078
KLDYHGVRQ

19079
KMVQVEHYT

19080
KVMTAEIAD

19081
LACACQDAP

19082
LDGSCYHWG

19083
LKATCTAMD

19084
LTTVYNWQH

19085
LVEMGCEPR

19086
LWTMDTVRP

19087
LYVGWMNCL

19088
MCYHTGHFA

19089
MGGMIVMCQ

19090
MHCRTFSHN

19091
MHMRTNEGS

19092
MILTEGIIE

19093
MLHMMPWID

19094
MMAMKQRFY

19095
MMGGKFGTT

19096
MNNEVWKAS

19097
MNQHWPPQI

19098
MPLSMKAKK

19099
MSDTMDMGE

19100
MSIMCCSDC

19101
MSNVCICTG

19102
MTGHNESFT

19103
MTVAGQSRD

19104
MTWTRNSGT

19105
MVDREKKKE

19106
MVSSETGSH

19107
MWKRHCGMG

19108
MWMQMLSAM

19109
MYCLEFMND

19110
MYTGMRCWH

19111
NAPNDLKNY

19112
NASCQYRGA

19113
NASCQYRGV

19114
NATHMYNAH

19115
NGCIVAEAA

19116
NGYIVHACQ

19117
NGYQARTKS

TABLE 78

Sequences of the 581 to 589 Region in AAV5

VP1 Capsid Polypeptide that Drive Lymph

Node Tissue Tropism

SEQ
581-589

ID NO
sequence

21118
AVDCHNSWI

21119
VQQYHITQE

21120
AAAKWDDVW

21121
AACKEKRNN

21122
AACTMAVAR

21123
AACYDVNKA

21124
AADLKGYIG

21125
AAELNSSQS

21126
AAENCQCPF

21127
AAEWKVGQA

21128
AAGEEWSAL

21129
AAMSCADAS

21130
AANESTHDC

21131
AAPYLTGRD

21132
AASPITVLK

21133
AASQMMNHG

21134
AATMNGQCM

21135
AAVGYFHTS

21136
AAVMESFWR

21137
AAVMSLCTV

21138
ACATTNFYD

21139
ACDPSHKHQ

21140
ACDPYSINF

21141
ACETWQWRE

21142
ACEWWFNKE

21143
ACIMQMTQD

21144
ACKLLDSHL

21145
ACMPICMSA

21146
ACTYRMTWH

21147
ACVDLVVDK

21148
ACVKHLYDW

21149
ADAWETPYP

21150
ADCHTQNVA

21151
ADCRYNLAC

21152
ADCSDQMSQ

21153
ADFDGILWY

21154
ADIQFARMP

21155
ADLDINQYW

21156
ADLHKRHNW

21157
ADNLTPHQW

21158
AEHKWPEEL

21159
AEQFVDVAG

21160
AESPVAYQQ

21161
AEWVEQAVH

21162
AFEHQYCHL

21163
AFEIVHGAN

21164
AFETLRNVK

21165
AFGEFVILA

21166
AFGHFPGIG

21167
AFHMMEKWE

21168
AFISKWVHP

21169
AFITQVPWY

21170
AFMNTYVAD

21171
AFMPWMHTY

21172
AFMQTICDM

21173
AFQYSNQAH

21174
AFRDISKMI

21175
AFRLLGYSY

21176
AFRNFTHCP

21177
AFSLNNCTH

21178
AFTCCGYEN

21179
AGDQKYVGC

21180
AGEEYTTDF

21181
AGEITKSMF

21182
AGHRDRAET

21183
AGLYLTNED

21184
AGNAMDFCF

21185
AGNEVQTMP

21186
AGQYKLIFW

21187
AGYCSQWRF

21188
AHAECKFWS

21189
AHAKWWDYQ

21190
AHALNEWYA

21191
AHAMVVNQG

21192
AHCYVLRHE

21193
AHELSYSLP

21194
AHFDYDIFW

21195
AHGLFMCMN

21196
AHHEVCWQH

21197
AHHLQICHT

21198
AHLWVQGSH

21199
AHMHTHAIM

21200
AHNQINDNL

21201
AHPHFEENG

21202
AHSWWHRAE

21203
AIAECIGFT

21204
AICQLLGDF

21205
AIEPKEDWM

21206
AIGHGENYR

21207
AIHETIAGT

21208
AIHYLWQDQ

21209
AIMPVRDGQ

21210
AIPCEMNAN

21211
AIQLSQDLM

21212
AISVMHYDM

21213
AITDTSCCN

21214
AITTQGDFY

21215
AITYQGQGV

21216
AIVPITFNM

21217
AIVYKNWAK

21218
AIWRANEST

21219
AIYSLDFEQ

21220
AKFFPHTNT

21221
AKGLNLDIM

21222
AKHLFDAAP

21223
AKHRCMSEW

21224
AKMWECNPK

21225
AKNSLYNYH

21226
AKYWNPELQ

21227
ALHWWQCLT

21228
ALINQPDSA

21229
ALMMYFENQ

21230
ALTEQCVYG

21231
ALVLWDMLH

21232
AMAGPQFSA

21233
AMAHKGMDA

21234
AMAQPVKCA

21235
AMHITQYKT

21236
AMHQNIFGI

21237
AMQQWVSGC

21238
AMQQWVSGC

21239
AMSLQYKNE

21240
AMTSRGSYP

21241
AMVFNMQLG

21242
AMVSSWLVG

21243
AMYVMYHGM

21244
ANDCRGMSE

21245
ANDLLTMQF

21246
ANFGSHAAM

21247
ANHFESSRS

21248
ANHPNMLIP

21249
ANMIRKGYG

21250
ANSFEVCGM

21251
ANSYYHQYC

21252
ANVEQAMCA

21253
ANVPMFRMD

21254
ANYFPTKFP

21255
APGCVNQLH

21256
APLTSYPTL

21257
APPVFYMPQ

21258
APTVINMLK

21259
APWQFNHQW

21260
AQCPMTWSL

21261
AQEVTITHE

21262
AWLLLIVMA

21263
AQRADWTYL

21264
AQWPLHRDT

21265
AQYGRNFAG

21266
ARMQEPIQV

21267
ARNANCPRM

21268
ARQINEHQE

21269
ARVDDGQMT

21270
ASANNPVFF

21271
ASASYEGEQ

21272
ASAVAKEWD

21273
ASDGEEKPC

21274
ASDHLNPFV

21275
ASDPWNWWP

21276
ASGWIQLMC

21277
ASMAYLKGE

21278
ASNFHSGLY

21279
ASNFVMEFP

21280
ASNKICNFK

21281
ASREEVKHP

21282
ASTHASESQ

21283
ASTSQCQFQ

21284
ASTTIGQEM

21285
ASVELGGYS

21286
ASVKDFTVQ

21287
ASYGDGPMW

21288
ATAEGEDCT

21289
ATECAWYHG

21290
ATFICLTSI

21291
ATHEYASWD

21292
ATHPVMLDA

21293
ATICSGWDR

21294
ATEGWERYN

21295
ATEMVCRFE

21296
ATKELAVQW

21297
ATKLDFHPT

21298
ATLTFQQHH

21299
ATMLRKEVW

21300
ATMSPECMR

21301
ATNWEGEDS

21302
ATNYVTNHD

21303
ATRVGGSTW

21304
ATSAHSCSS

21305
ATSENCNAW

21306
ATVDYWQAW

21307
ATWTDSWKV

21308
ATYCEQYMY

21309
AVAGVVWDN

21310
AVAMGGQDD

21311
AVANWFQHM

21312
AVCEGEITP

21313
AVCSKELDH

21314
AVDHMAMES

21315
AVDQINMEQ

21316
AVELSMRAV

21317
AVEPYCSML

21318
AVESTAIES

21319
AVEHQGSGA

21320
AVITWSQYH

21321
AVKPSVEPW

21322
AVMPQGHYG

21323
AVMTAQGWV

21324
AVNPFPCQT

21325
AVQSGIETC

21326
AVSAAMQMD

21327
AVSDKPYQH

21328
AVSFYQYGC

21329
AVTHKWCEE

21330
AVTILLYPI

21331
AWAKDVMQC

21332
AWVISQNEN

21333
AYASSQDSL

21334
AYENTFPNP

21335
AYEPFWCWG

21336
AYHHVDYHW

21337
AYNQNIGTG

21338
AYYDLFNQC

21339
CADFMVSWG

21340
CADWCNTQP

21341
CADYQYQQD

21342
CAIGYWEHD

21343
CAMMDDRGD

21344
CAQLFWLPQ

21345
CASDLMENR

21346
CASDSLVNG

21347
CAVNWNVKE

21348
CAVQFNYCV

21349
CAYQAMDFF

21350
CCCNRCHEY

21351
CCFSTFECY

21352
CCGDTRCET

21353
CCGLPARLL

21354
CCERSFVCA

21355
CCKTYQRAG

21356
CCMTYNNQG

21357
CCPGNGDEW

21358
CCPMRHANF

21359
CCSGNTGMY

21360
CDHGCWCKG

21361
CDHVVCMAP

21362
CDWAAYHCW

21363
CEGIQVVDF

21364
CEVTINGAE

21365
CFAELSNVA

21366
CGDHYRKGH

21367
CGHLQQFIL

21368
CGIGVNMHH

21369
CGIWNNNAF

21370
CGRQMEDAA

21371
CGTQSFIWG

21372
CGWCGLSKI

21373
CHAIFCRCA

21374
CHDGSFNPH

21375
CHGQYVTWG

21376
CHHRMHRAQ

21377
CHMGINRLN

21378
CHSPFNHWG

21379
CHWYCSKVP

21380
CIDEGVSPD

21381
CIETSDQER

21382
CIEYTPMGR

21383
CIGFYMCAG

21384
CIGVINGMN

21385
CIHYVQHCQ

21386
CIILRSNDG

21387
CILCQPHKW

21388
CIWKNVPPW

21389
CKDAYRSHA

21390
CKFMYLMKE

21391
CKLTMKRQN

21392
CKSTHDMAW

21393
CLHCGTFST

21394
CLHCHSLQC

21395
CLMLQSHHT

21396
CLRAVDNCF

21397
CMQSESMYC

21398
CNATWGVFA

21399
CNAVKNNDM

21400
CNEVYTHIH

21401
CNNYSMVHR

21402
CNNYSMVHR

21403
CNRLHDTWT

21404
CNRSEIQFR

21405
CNVERDPYL

21406
CPCFYINYP

21407
CPETRHVVH

21408
CPKDCRIVI

21409
CPLSWIQQE

21410
CPQPSNRTW

21411
CPQQIQHYN

21412
CPTEVREKK

21413
CQGFGVPWD

21414
CQHLHTSYW

21415
CQKALMQLG

21416
CQTCLQFCD

21417
CQVEFQMTT

21418
CRDHYDRSC

21419
CRFEHQYNC

21420
CRKLQSIEA

21421
CRTNGHYYM

21422
CRVEPVDSD

21423
CSACTPFSK

21424
CSADMNSMP

21425
CSADMQQDI

21426
CSDASHHFG

21427
CSEAEGDWY

21428
CSFNVMNTA

21429
CSKPDCQWD

21430
CSMQNNRMY

21431
CSMYYCWTG

21432
CSNSQDFGA

21433
CSTEMQMNC

21434
CSYQRTEYF

21435
CTATPEIIH

21436
CTEININQG

21437
CTFVRELQC

21438
CTGDVDQSE

21439
CTGLAQHWW

21440
CTGQRHASP

21441
CTHHYMEQH

21442
CTKHMDGQA

21443
CTNDWCRSD

21444
CTNLISSYC

21445
CTQVIEECP

21446
CTTYQDEMN

21447
CTWWIPLVY

21448
CTYTELQYM

21449
CVDQLSLGH

21450
CVHAISCVG

21451
CVIGRWLYM

21452
CVISMAYQW

21453
CVLKYNHHY

21454
CVMMQAGGE

21455
CVSYSGQHD

21456
CVVNAHQMR

21457
CVVRAQSAM

21458
CVVTMSMSD

21459
CWMFPDYWN

21460
CWMSFSAYE

21461
CYESTEGEH

21462
CYHEYCCNM

21463
CYKYGPPSQ

21464
CYMVFMFKA

21465
CYQDIEAGY

21466
CYQFKKLTN

21467
CYQSEKNGY

21468
CYSGVAMAS

21469
CYVTKSWDH

21470
CYWFMYKPK

21471
DAAQTEQYP

21472
DAAYSNWFF

21473
DACKESSNI

21474
DADEPWLDR

21475
DADRAIMHE

21476
DAEDCHTHK

21477
DAEGQVCYM

21478
DAETVDSQF

21479
DAFLEQELC

21480
DAGKPQYSI

21481
DAHDYDSNF

21482
DAHNYWGWQ

21483
DAHWVEKTN

21484
DAIPYKQPT

21485
DAKFCGQGH

21486
DALYEMGTT

21487
DAMPSDRWW

21488
DANFTMMTD

21489
DANNKKYHV

21490
DARHYFGLQ

21491
DARPDYEMQ

21492
DARTIVLTH

21493
DASDCCYSY

21494
DASLWSVPM

21495
DASMHGTVP

21496
DASPTRLVY

21497
DASVNRVIE

21498
DATKIPMQQ

21499
DATQNHFQY

21500
DAVDVAMAM

21501
DAVYYLFQA

21502
DAWHSTCNS

21503
DAYYDHRGC

21504
DCGWLAYEW

21505
DCKCPQPDL

21506
DCPWKAFHA

21507
DCRGLSSIF

21508
DCSVFWIGA

21509
DCTFTQQQW

21510
DDKPWVIMA

21511
DDLQPMERT

21512
DDVNGAVIM

21513
DDVNKMRKE

21514
DDVWMWQWV

21515
DDYKKWRHP

21516
DDYKPHCKF

21517
DECYNLFTV

21518
DEDDYNMPY

21519
DEHLITMQT

21520
DEQWSPYTI

21521
DEYVPEAYF

21522
DFAACQDYL

21523
DEAFENWMG

21524
DFAWQECWC

21525
DFEKVSEQA

21526
DFGCQEQMT

21527
DFGPRCCFG

21528
DFGVQKSNE

21529
DFHDQYVID

21530
DFEWQQCSN

21531
DFIEGTREG

21532
DFSRMAGGP

21533
DFTAFKNDM

21534
DGCPSGNWP

21535
DGELGKEDI

21536
DGEDFEKIG

21537
DGGFQQAEW

21538
DGHSDLLLS

21539
DGKHLDTDP

21540
DGKKTMHET

21541
DGQLHMFYA

21542
DGSGSEWHR

21543
DGTADDKRY

21544
DGTETIYNM

21545
DGTYANHMG

21546
DGTYKCPEP

21547
DHCNFMAPY

21548
DHDWLQNMG

21549
DHEVVQNMP

21550
DHKLIQMHT

21551
DHLICQNGP

21552
DHMWVNWSQ

21553
DHPFNVKQE

21554
DHQLTMEGM

21555
DHSMYATQE

21556
DECDKNHEG

21557
DIEFDITCD

21558
DEGEFWTAH

21559
DIHRFAEQF

21560
DIHVITKGI

21561
DIKQHQLWG

21562
DILDVVKQG

21563
DIMINDLYS

21564
DIQHLTESG

21565
DIQQHNIAA

21566
DEQVPLAYR

21567
DIRRMVASL

21568
DISMESWMP

21569
DITETQCGS

21570
DIYYSMRGR

21571
DKACMLWDS

21572
DKCAIEFAS

21573
DKGALEFWI

21574
DKGEWQCYK

21575
DKGFIQHVA

21576
DKMTLPFSP

21577
DKPCSICGC

21578
DKRWSANQW

21579
DKWPPLEPH

21580
DLAWQYQPA

21581
DLEDAHVGL

21582
DLHGEGVLA

21583
DLEAGNWPA

21584
DLPSPRRED

21585
DLRVNHWHH

21586
DLVCAQWRE

21587
DMAAFHYAG

21588
DMAANHTFP

21589
DMALYHSNV

21590
DMDCYTMME

21591
DMDSPFPWD

21592
DMDTYYGHG

21593
DMFTFAHNS

21594
DMGFVNRHE

21595
DMIHYPLYG

21596
DMLMLVTHD

21597
DMMGFWDKP

21598
DMNELAWAY

21599
DMRVGNGYF

21600
DMSVIQDGA

21601
DMTRNPEWE

21602
DNAACIEMS

21603
DNAEMNSFD

21604
DNCYSCEAT

21605
DNEGWLMAE

21606
DNEVWLSAV

21607
DNHALEFDC

21608
DNMQRKWAG

21609
DNQIRSIRS

21610
DNQSKESQA

21611
DNSDFKWHW

21612
DNVPYCYYQ

21613
DNYINQEAA

21614
DPAHTTKIC

21615
DPDRTPSQH

21616
DPEEHNNYG

21617
DPEFQQWTH

21618
DPIDEVNAQ

21619
DPNRIDNQY

21620
DPSEVRARV

21621
DQCFMTYDT

21622
DQDNPSCWL

21623
DQEPNWNGV

21624
DQFDSHSQG

21625
DQGCKLRRC

21626
DQGMGSCKD

21627
DQHGASTVF

21628
DQIPMKSMM

21629
DQKMYVDED

21630
DQKSVQCYE

21631
DQLEGGESN

21632
DQLGLRDNT

21633
DQQPALDAQ

21634
DQWSILTES

21635
DRAKECQCE

21636
DRLQCDECS

21637
DRNTAMTGG

21638
DRQINWQQE

21639
DRSDKGCLY

21640
DRSVHVRAS

21641
DRVEERAYG

21642
DRYLFTHWH

21643
DRYQWAKRH

21644
DSALQFIDR

21645
DSAMFGQYK

21646
DSANVGWCT

21647
DSAPGKMWP

21648
DSAPLQSQL

21649
DSAPPPNVI

21650
DSDEGSEHI

21651
DSDLYEGWM

21652
DSEWLHWHA

21653
DSGFAMWTP

21654
DSHFYDQWQ

21655
DSHPIAHGE

21656
DSMKGTMFQ

21657
DSMMHDCHF

21658
DSMNGVEAS

21659
DSNEEQRQG

21660
DSPEQFGLD

21661
DSQHTAGWP

21662
DSQLIKQWM

21663
DSQPMFACV

21664
DSREAVAVD

21665
DSRQDHWFE

21666
DSVHSYEQK

21667
DSVPNLKAY

21668
DSYPNQACT

21669
DTCTTCVSL

21670
DTCVTAANH

21671
DTDTRCIPC

21672
DTEGDHREG

21673
DTELSMVEP

21674
DTFFHGAFE

21675
DTGIWAYVG

21676
DTHRIIMDD

21677
DTILEQYGP

21678
DTIQWDQGI

21679
DTKMVPYMP

21680
DTKPENNWI

21681
DTMHCLSCD

21682
DTMPTMDLM

21683
DTQWQEWCE

21684
DTRDNWLIR

21685
DTRTPAVWP

21686
DTSQTSNNT

21687
DTSRCLAYW

21688
DTWILDFEV

21689
DTYAEIMYG

21690
DTYPLVIGS

21691
DVAMTSLIA

21692
DVCDSIGYC

21693
DVCEQRNWL

21694
DVDDSQCEY

21695
DVDDWFHQS

21696
DVDHICEGH

21697
DVDMDTNRG

21698
DVENHYMGS

21699
DVEPMKNDF

21700
DVESRYDCG

21701
DVFCMLASH

21702
DVFIQQAYS

21703
DVFMGARFE

21704
DVFPCQKTE

21705
DVFQCEELS

21706
DVGFNRISR

21707
DVGIQHRPL

21708
DVGLFHGGF

21709
DVGPQKNIV

21710
DVHMADGNN

21711
DVILTYDRA

21712
DVIRTHDGV

21713
DVKCMQNTF

21714
DVKGNNGGG

21715
DVKIVWSGE

21716
DVKLAGMIH

21717
DVKPAMNQA

21718
DVKPGWMGD

21719
DVKWMESWL

21720
DVKYLTHQW

21721
DVLLLEHER

21722
DVLTNLNGF

21723
DVNFVGEPE

21724
DVNSYKLYG

21725
DVQTIRMEH

21726
DVRERDSDD

21727
DVRTEPTIT

21728
DVRTVTQHG

21729
DVSQSDWQP

21730
DVTAQRAWK

21731
DVTAYSESI

21732
DVTEDVVSP

21733
DVTENREVW

21734
DVTIYWNDY

21735
DVTLNEGVV

21736
DVTPEDACV

21737
DVVALQASK

21738
DVVANLTSS

21739
SVVEQFHWT

21740
DVVIVDCAN

21741
DVVVSQMAM

21742
DVWAQSQVL

21743
DVWLQIDGQ

21744
DVWWVEGEY

21745
DWASPKSYV

21746
DWCMSMVWT

21747
DWDQQRQYF

21748
DWGAMWVCG

21749
DWGHRIFEH

21750
DWHHPPVDH

21751
DWKPIYEAS

21752
DWPIRSAPI

21753
DYCTGACDC

21754
DYDVASFYH

21755
DYEGPIRPN

21756
DYEHYWHET

21757
DYKLHPIGY

21758
DYKMDVMPC

21759
DYNMMHWAA

21760
DYQRCTFLP

21761
EACSNSHEK

21762
EADCVHRVW

21763
EADVSLMHC

21764
EAEAEVIIR

21765
EAECTWASL

21766
EAELKTNFM

21767
EAESGGAYK

21768
EAEYRNWNP

21769
EAGMFVHRA

21770
EAGQWLGSP

21771
EAHAWEKTE

21772
EAHEEWSFD

21773
EAHVNVYKY

21774
EAKCMHDFS

21775
EAMIWNNTM

21776
EANQILSAH

21777
EANSSQSQF

21778
EANWEHSCI

21779
EAQTTGSTW

21780
EASLIKEDK

21781
EASSLYSQQ

21782
EASWSLHGQ

21783
EATGESFWF

21784
EAWHPGCCQ

21785
EAWMCVEAQ

21786
EAWSMNAPY

21787
EAYFDTAEC

21788
ECAPWVVLS

21789
ECCIGDNMM

21790
ECEMMDYSK

21791
ECFATGEFM

21792
ECHKCSKLN

21793
ECKLSVCGY

21794
ECMHAEWQA

21795
ECMLIGTDG

21796
ECPDYMRKD

21797
ECQMHRTFA

21798
ECRPWGMHE

21799
ECSESAEAK

21800
ECTVMQRGA

21801
ECWFCDIWR

21802
EDLHQWAQH

21803
EDMFCPTKA

21804
EEDFRNEAM

21805
EEDMQRTVQ

21806
EEEAGKWMC

21807
EELEQIWHG

21808
EESLDVVMK

21809
EFCYEPDWD

21810
EFETFWEVG

21811
EFGTMHYAP

21812
EFGTSQKDA

21813
EFGYSPGVS

21814
EFIHAENWY

21815
EFKARMVTP

21816
EFLVNTSWL

21817
EFMCSCMLN

21818
EFMTYETNV

21819
EFNSIMHWW

21820
EFQWRACFQ

21821
EFRMASSDA

21822
EFSDVQRTA

21823
EFSITMAPW

21824
EFSKMSGTA

21825
EFVEKWLWG

21826
EGAELEWDH

21827
EGAQLDVMS

21828
EGDMTTNQY

21829
EGEELRVAY

21830
EGEEYTLPS

21831
EGFLGFGQN

21832
EGFYLEGDP

21833
EGKEWIFYE

21834
EGMKTQQTF

21835
EGMWPHSAF

21836
EGQRLTWAQ

21837
EGRQHFVYA

21838
EGSQWRTSI

21839
EGTMSNGWQ

21840
EGVRLNLYG

21841
EGWFMKKQD

21842
EGYEYHTQR

21843
EHATESWAG

21844
EHATIENWP

21845
EHDGVWMYP

21846
EHEDTIAHD

21847
EHFPQQSGD

21848
EHGPCMSGC

21849
EHIGCGYWQ

21850
EHIRFGHYW

21851
EHKPDGANY

21852
EHLKLLIAC

21853
EHMSICDAP

21854
EHNSEDCIC

21855
EHPMENTYQ

21856
EHQFMWQDH

21857
EHRDHDAWA

21858
EHVKCNHFF

21859
EIAGSEMHL

21860
EIAKWNCEL

21861
EIAQL0FDQ

21862
EICAFDVGW

21863
EICVIECEQ

21864
EIDYHICSG

21865
EIEMAVAYA

21866
EIETSAREQ

21867
EIFQYMHHH

21868
EIGIGAPDY

21869
EIHHGHAVP

21870
EIIAQKWTQ

21871
EIIMYEQHM

21872
EIIPTSMFG

21873
EIITVRDDE

21874
EIIYREVDA

21875
EILSDDDMP

21876
EIMHRCHFW

21877
EIMIQVIRD

21878
EIMMEHDNK

21879
EIMWPLAGN

21880
EINLGHQHN

21881
EIQMILTAV

21882
EIRSHDTGH

21883
ELSWQDRDL

21884
EITFNQYGH

21885
EITWESNIA

21886
EIVDLTVME

21887
EIVEHDYRE

21888
EIVYMMCPC

21889
EKAGNLERK

21890
EKDQYKMGS

21891
EKEQFSHKG

21892
EKGMIDLVY

21893
EKGMNMHAG

21894
EKHQLSVQQ

21895
EKQWLSRQD

21896
EKSGYHFSC

21897
EKVICLYRG

21898
EKVTLNMDM

21899
EKYMNFMCP

21900
ELADQNQIQ

21901
ELAWIWQIH

21902
ELCCNDMFK

21903
ELERTVKEF

21904
ELGYSEKVC

21905
ELLTCIECG

21906
ELNFCWQQR

21907
ELYMNNHTE

21908
EMAELNQHV

21909
EMDQVPWGF

21910
EMEFPDMCM

21911
EMEWQDSWW

21912
EMGFGKQGM

21913
EMGMPSYMK

21914
EMKEAWLSA

21915
EMMVLCENE

21916
EMPTICKNC

21917
EMQCYQTYH

21918
EMSECSITA

21919
EMTPQQFWH

21920
EMWICHEGG

21921
EMWSRGMVN

21922
EMYPFSKQL

21923
ENAEEKDHM

21924
ENAIKHHDS

21925
ENAQRHNYA

21926
ENDAEVPDH

21927
ENEAMAKWF

21928
ENFMLRPCN

21929
ENHASAHVR

21930
ENHIEFWWP

21931
ENLESRLQH

21932
ENLHCDCLY

21933
ENLRMGKWE

21934
ENVKLDQCD

21935
ENWHLVAHR

21935
ENYCQCCLE

21937
ENYRHHTGP

21938
EPEWLGHEE

21939
EPGPMQCWD

21940
EPGQFFHDC

21941
EPLDNVGCG

21942
EPMNGKVME

21943
EPNAQMTAA

21944
EPQEVENMT

21945
EPTLVGEVP

21946
EPTNIWAVH

21947
EPTSATWVY

21948
EPVHLPEWM

21949
EQFGCGQWL

21950
EQKMKQIAQ

21951
EQKSTQTFH

21952
EQLDSHWWK

21953
EQLDVKYWH

21954
EQLIEWSSH

21955
EQNMLDADC

21956
EQQPMASFT

21957
EQQYLAQGK

21958
EQTATMRWD

21959
EQVYYWNTP

21960
EQWYDEEPH

21961
EQYYFNQLR

21962
ERFYQHAAK

21963
ERGYNMAQH

21964
ERIQWTHMG

21965
ERTAPSSRG

21966
ERTMVHQYW

21967
ERVDQENCH

21968
ERVTDEHEE

21969
ERYHMRWEP

21970
ESANYMNLF

21971
ESCYMMMNW

21972
ESGMLFYFM

21973
ESKAHAQTP

21974
ESKTTSCAF

21975
ESLRAENMP

21976
ESMGFEECG

21977
ESMKLIQCD

21978
ESMNEMTYK

21979
ESNDNGNQW

21980
ESNPVCVLK

21981
ESRCAWLSD

21982
ESRKEIGYA

21983
ESRRHHWAH

21984
ESSDESNYL

21985
ESTLLLCHG

21986
ESTNLQCVL

21987
ESVISLDQQ

21988
ESYETQLAC

21989
ESYFSETQP

21990
ESYEVEDNT

21991
ESYKCIRDC

21992
ETAHYSMRE

21993
ETANQEWER

21994
ETASKNVQT

21995
ETCILAVDG

21996
ETDEEYEVG

21997
ETDQQYDQY

21998
ETEEIVNMA

21999
ETEGYLFSP

22000
ETEPPDTQG

22001
ETESMMFCN

22002
ETHTYDDCS

22003
ETEAMQYRC

22004
ETKCYAMLG

22005
ETKEYDCNG

22006
ETKLAIDEF

22007
ETKSVLDEH

22008
ETMCPKGPL

22009
ETMNQIGSC

22010
ETNLFEHST

22011
ETPGTKQCS

22012
ETQRDRHTN

22013
ETRDYLEAD

22014
ETRQMDGNN

22015
ETSMRFESL

22016
ETSRMNVDA

22017
ETTCYAYME

22018
ETVERQCEQ

22019
EVAETSHSY

22020
EVANGKGDH

22021
EVCLGQRYA

22022
EVDRHLGMQ

22023
EVEACKYEY

22024
EVECTHRGE

22025
EVEVWSVYH

22026
EVFPYGCQN

22027
EVGMEVHPD

22028
EVGTCYFSK

22029
EVHKNDKMH

22030
EVHSMDTQW

22031
EVIQTSDNR

22032
EVKFEMLCQ

22033
EVKHKQDWA

22034
EVKNKGGFC

22035
EVKPCLVVQ

22036
EVKTRMCAG

22037
EVLKAQREV

22038
EVLLSTDEA

22039
EVLPYEWCE

22040
EVMIRKLDM

22041
EVMYSERTW

22042
EVNMEMQLA

22043
EVQGEKCLN

22044
EVQQFHRDQ

22045
EVQRWSGYH

22046
EVRGAEANH

22047
EVSKLATTY

22048
EVSQFGIMN

22049
EVTFWTCEG

22050
EVTIRSHYQ

22051
EVILEPDSW

22052
EVVHSMSDG

22053
EVVNVPMRT

22054
EVYRNTMKC

22055
EWAGKQTDM

22056
EWAYMGSYP

22057
EWDDTSNSG

22058
EWHFPKAMN

22059
EWNDYPSWY

22050
EWRHVELHY

22061
EYAGHHCFY

22062
EYCMCMWGA

22063
EYCNVMIPC

22064
EYDDVPEPQ

22065
EYEQDCRFQ

22065
EYHDKHWAG

22067
EYHTEWIGM

22068
EYEGEELHK

22069
EYILWHWCT

22070
EYISTQADA

22071
EYKYYTYIF

22072
EYMATEHGN

22073
EYMHYYGSE

22074
EYPLESTYK

22075
EYPLPVTLQ

22076
EYSQLHPSP

22077
EYTCRLLYD

22078
EYWEGQQSC

22079
EYWQESAWS

22080
FACEIQYSD

22081
FAEDCQRFD

22082
FALNGEEKW

22083
FASRYDGHH

22084
FAVFCWAER

22085
FAYCESEED

22086
FCDACLGGY

22087
FCELYFHCA

22088
FCLDPNFYY

22089
FCTGASHAH

22090
FDDCTWQGP

22091
FDHATMQFG

22092
FDRCNESYC

22093
FEPHGCQIE

22094
FEQADMMQM

22095
FEQYIRNIK

22096
FERLFMGWT

22097
FEVMRAIRC

22098
FFTKISWSW

22099
FGTAVMVWE

22100
FGWDAYCRV

22101
FHQYHYQAM

22102
FHWWGAVVK

22103
FICDVQMGD

22104
FIEFYMGEQ

22105
FIEGEMHIR

22106
FIGLQWSYP

22107
FIPISHQHP

22108
FIRSYNVEC

22109
FIRVSWWLA

22110
FIYTDFRMT

22111
FKEELMKYQ

22112
FKFEQENNS

22113
FKKPMVCHV

22114
FKLVSGDQW

22115
FKQIDHYPR

22116
FMAELKTDD

22117
FMKSKLETV

TABLE 79

Sequences of the 581 to 589 Region in AAV5

VP1 Capsid Polypeptide that Drive

Mammary Gland Tissue Tropism

SEQ
581-589

ID NO
Sequence

24118
FGDHLRVMY

24119
LPTQYQQQN

24120
TKKIIESSS

24121
IMYLKRCCT

24122
TSDQCGNED

24123
GQFPHYGIF

24124
YTELMKKAC

24125
LGMHYKDNC

24126
KMGTMDFNH

24127
QWDDEWDMD

24128
THACMYHEY

24129
LEQCHDHCA

24130
TKVNCVYYG

24131
QELGVNCAA

24132
QKDWGPRVT

24133
APMTCKEFL

24134
ASWFPVLFP

24135
CTYAAMMSY

24136
CVQCECNQN

24137
DMFRPDVRC

24138
DTDNKPHFA

24139
DTHIFIRHA

24140
EFHFKCRTD

24141
EHHAMMLHE

24142
FSHMPTSGG

24143
FVCPRAFNM

24144
FWMNCWVTV

24145
GQYIAVNYS

24146
HYEMPQAMQ

24147
KCGNEGRMS

24148
LQKGWANCE

24149
LSTLITMMD

24150
LTDFHPKML

24151
MAMKLIEGN

24152
NFCKQGNTC

24153
NIRCGNDAS

24154
NQQPGQHSQ

24155
NVVCVRYKS

24156
QATMDAWVH

24157
QIPKAWNRY

24158
QTTINDSSQ

24159
SADRTAMFH

24160
SARTDCQCK

24161
SDSKVQWGM

24162
SEDHFGFWV

24163
SHWFFHFDL

24164
STSWLNDLT

24165
TEFSQQYKC

24166
VAFEYGERA

24167
VCGWWHQYW

24168
VHTEMSNID

24169
VTQRPLHHS

24170
WLYLAFAQN

24171
WWKPCNCQY

24172
YATQMQHLE

24173
YMGSTNIWM

24174
GVGPSHGER

24175
GVMHHGIFD

24176
AISMYWSSW

24177
AVQCYIHPS

24178
CSTIHAGHP

24179
DIGMYWSSW

24180
DISMCWSSW

24181
DISMDWSSW

24182
DISMYWGSW

24183
DISMYWSGW

24184
DISMYWSSL

24185
DISMYWSSW

24186
DMSMYWSSW

24187
GISMYWSSW

24188
KPMINEIVY

24189
LHALINQGS

24190
LHALVNQGS

24191
LHALVNQGY

24192
LRALVNQGS

24193
MCLRDHNGL

24194
MPMINEIVY

24195
MYLCDHNGL

24196
MYLRDDNGL

24197
MYLRDHDGL

24198
MYLRDHNCL

24199
MYLRDHNDL

24200
MYLRDHNGL

24201
MYLRDHNSL

24202
MYLRDHNVL

24203
MYLRDHSGL

24204
MYLRDHTGL

24205
MYLRDRNGL

24206
MYLRGHNGL

24207
MYLRNHNGL

24208
NISMYWSSW

24209
QCYAIAHLI

24210
RCCAIAHLI

24211
RCYAIADLI

24212
RCYAIAHLI

24213
RCYAIAHLL

24214
RCYAIAHLV

24215
RCYAIAHVI

24216
RCYAIAHWI

24217
RCYAIALLI

24218
RCYAIAPLI

24219
RCYAIARLI

24220
RCYAIGHLI

24221
RCYAISHLI

24222
RCYAITHLI

24223
RCYAIVHLI

24224
RCYAVAHLI

24225
RCYDIAHLI

24226
RCYSIAHLI

24227
RCYTIAHLI

24228
RGYAIAHLI

24229
RWYAIAHLI

24230
SEQCYIHPS

24231
SVQCCEHPS

24232
SVQCYIHPS

24233
SVQCYIRPS

24234
SVQCYMHPS

24235
SVQCYVHPS

24236
SVQFYIHPS

24237
TPMIDEIVY

24238
TPMINEIFY

24239
TPMINEIVC

24240
TPMINEIVY

24241
TPMINEMVY

24242
TPMINEVVY

24243
TPMINGIVY

24244
TPMISEIVY

24245
TPMVNEIVY

24246
VYLRDHNGL

24247
YSTIHAGHP

24248
RVTIMFTGT

24249
GTRENTCNG

24250
AVGSLRTFQ

24251
CNTPYEFWA

24252
DLRLMACCG

24253
RRREGISGL

24254
VTYYSMTQQ

24255
MASRKSMED

24256
GMVELGSCW

24257
TVTITKWRD

24258
QHTQQLVQN

24259
SGNMYESGE

24260
QDFRMNAGA

24261
GLSMVLHFV

24262
AMTRCKAGH

24263
GSNRNPIMM

24264
ASAGRNWTA

24265
TYWTKQSEV

24266
CIWSSPDQP

24267
NYLQMCWFN

24268
TTYPHNTHG

24269
TCTNSEPHP

24270
ECQYEMNDD

24271
THAQMSLRD

24272
KYASMDIYS

24273
AQSKSLMRC

24274
TVMQCGTMP

24275
TAFQSLQQM

24276
HMDEVCSKD

24277
YVSLCNDGN

24278
RVEYVTDSH

24279
MCQNVEKEY

24280
NSTPMACME

24281
TSEPHCSYA

24282
DPGWYGLAP

24283
CWGPIMHPF

24284
LSVQAQWDS

24285
IAKIGNCVW

24286
KAVSYHHDG

24287
SLTQYQWRG

24288
MATCYMYAD

24289
APFFTFTNH

24290
TRMQAVDHE

24291
FWMQTEHGT

24292
GCEEVGRCQ

24293
IKHITRNTY

24294
WKWYGLPHH

24295
ESTFWQHTF

24296
SQMKYAHNQ

24297
AEVPYVMQQ

24298
TCGHTRFFI

24299
NTSGELIMP

24300
DRVNLMWKD

24301
AVHTADVCC

24302
EFVLGCQGE

24303
DVKAFMGGV

24304
TTSQNSEAM

24305
YASSCQNAM

24306
QSMVACYTE

24307
MGYAKHDSH

24308
SSRMQMQGP

24309
SQIQIRQAD

24310
VHHKTNRSY

24311
RVNGSTCRH

24312
NGNHYNAMR

24313
WITSEAVDT

24314
FTEPMCMQG

24315
DVKSWAVCD

24316
AYHCYMSWM

24317
RYPCCQSPF

24318
AHAWLQHWA

24319
HQCNCLLSW

24320
TNFPTSMHG

24321
QRNEMSASR

24322
QRNEMSASR

24323
WDGWVTTHF

24324
PAHINDQYM

24325
VLKSNLTGM

24326
ATGTYNLQE

24327
LINSIVCGD

24328
ACHTGQSGM

24329
DHLSNDAEM

24330
SECEVLCSL

24331
VFNLMLQDK

24332
ILINDREWG

24333
QWETKGVGE

24334
NVMGTQAKE

24335
YSDPKNMSE

24336
LEIGGKTRT

24337
ASMHEETCL

24338
WERCLELNQ

24339
MPWLDKPPC

24340
ATMNLSWGL

24341
WVHPYSYCA

24342
QSRCVDNTV

24343
NRIQMTDFQ

24344
SNVACEQAF

24345
INYISEVDR

24346
EVRIQCKID

24347
SVLHVQALA

24348
CWILHHRCS

24349
KMAGIGCWY

24350
LCAVCGDCD

24351
HSMMPGWPS

24352
DQEAQVVTG

24353
ISLVWMHYP

24354
AACNGLEYQ

24355
AADCYHWFR

24356
AAEWDRFAE

24357
AAGLQVDSW

24358
AAMASAFVY

24359
AAMLGWESE

24360
AASPKTMNE

24361
ACDRFMAMY

24362
ACSVSECTE

24363
ACYDYYQQV

24364
ACYSYEFEE

24365
ADCFIFAEY

24366
AELEIMWDD

24367
AFGVYPSVS

24368
AFNHVHEGT

24369
AFTIEDDWW

24370
AGVQENVSV

24371
AILTVCIGA

24372
AINMSGSGN

24373
AKNDNLCAD

24374
AKRQKVVLS

24375
AKWRGTLIN

24376
ALCDYMSWH

24377
AMEYRSWHT

24378
ANCIRPFTS

24379
ANCKQDHCH

24380
ANICTIESD

24381
ANIRGRSMK

24382
ANQVTGSWK

24383
ANTQYPAHM

24384
ANWPMACDQ

24385
APFRKQMPG

24386
APWHSNDMR

24387
AQNVLDTVQ

24388
AQNVLGTVQ

24389
AQVSTLNGM

24390
AREDPCAPR

24391
ARGWDYKCP

24392
ASERLYYCW

24393
ASILQFTVD

24394
ASKPTRKVT

24395
ASQYQQSLS

24396
ATAINQRQA

24397
ATECKLNYL

24398
ATERQTLLN

24399
ATTCITSDD

24400
ATTKGMSGW

24401
ATTLINQEW

24402
AVGHFYTNS

24403
AVHTADVFC

24404
AVEFMSVCE

24405
AVTQQVDEN

24406
AWCNIDMST

24407
AWSCIMSYD

24408
AYENFHWYD

24409
AYHDSGHEW

24410
AYYQLQMSS

24411
CADVPITMA

24412
CAEQWAAMI

24413
CAHWTYDCY

24414
CAHWYYEQR

24415
CANKEHVCH

24416
CANWKDHGY

24417
CAQHRDTAF

24418
CASLKVEDS

24419
CASTYTGIN

24420
CATAMFRYQ

24421
CATKSMMYG

24422
CAWLSQTAS

24423
CCMPLQSFG

24424
CEGFEMRVE

24425
CEGWGTYCI

24426
CEHGNENWP

24427
CFGFVNSVN

24428
CFMWNWHHE

24429
CFRMYDLLS

24430
CFTNNVVWT

24431
CGCSEVTPH

24432
CGLIMEMCW

24433
CGWWFNAAE

24434
CHAEYSHEQ

24435
CHFQYQNQA

24436
CHLLLTDCY

24437
CHMMDWQCG

24438
CHMQFMEAG

24439
CHTEYSHEQ

24440
CIDLRLAWD

24441
CIEWWGMMG

24442
CIIPKMKER

24443
CILHRYVIE

24444
CILLKKCMA

24445
CINADGKYS

24446
CIQAVIGGT

24447
CIRLTEYLG

24448
CIRTMNGSW

24449
CISREDHKQ

24450
CKQWTNQAH

24451
CKVHKNWDR

24452
CLGSTWVAA

24453
CMCLKKCYY

24454
CMQQYDNGQ

24455
CMREQMNAC

24456
CNKDINQSC

24457
CNWLFMTQW

24458
CPGPMMQMA

24459
CPKPIGCGG

24460
CQDFRHECA

24461
CQQRIIMQA

24462
CQRKWEKQE

24463
CRTELGSMY

24464
CSAKVPMPT

24465
CSAQIKAAC

24466
CSCAMYNFH

24467
CSDQYQTQN

24468
CSGRLGSLY

24469
CSHHLMTFG

24470
CSHNFQDQK

24471
CSIWEDKSG

24472
CSMECRNEL

24473
CSTTMNVKT

24474
CSVITQTTG

24475
CTDKLLNAG

24476
CTDRMVKAS

24477
CTENYHQQY

24478
CTGQYMGYI

24479
CTHQQSLAV

24480
CTLKQNMPA

24481
CTQLSVNMQ

24482
CTQSFVLYQ

24483
CTVFSGRTA

24484
CTVRQNHKP

24485
CVAIDMSVA

24486
CVEYLKQLY

24487
CVGDREPWG

24488
CVGINLDHH

24489
CVGKQIMAE

24490
CVIHPPNFD

24491
CVLYYNQHC

24492
CVQGQYYQE

24493
CVQHNLVGY

24494
CVQTMNGKS

24495
CVYREIKTA

24496
CWAIERLQS

24497
CWAQLNTDT

24498
SWEMMKTQE

24499
CWNIYNACD

24500
CWSSWQKRD

24501
CYDGWWCPS

24502
CYKFVGAAD

24503
CYRDVRARG

24504
CYVYWTTCP

24505
DAAQFGKHT

24506
DACWGHEYA

24507
DAGTNSFCG

24508
DAGVTTYGG

24509
DASKVKLHH

24510
DASSYLHLC

24511
DCAKFPFMI

24512
DCEDIKKWE

24513
DCGQFSIIE

24514
DDHWDWLCY

24515
DDLGAGRSD

24516
DDLSMTNDC

24517
DEEFYPPAW

24518
DEFIWLEMH

24519
DETVFRNPV

24520
DFEYFSTKD

24521
DGFQYDAHC

24522
DGVQVDWGR

24523
DHKEYMDKG

24524
DHRQWKVLA

24525
DICHYIRNE

24526
DIMRYINTA

24527
DIRTSVSDS

24528
DITQMVVGC

24529
DIVIKIGMG

24530
DKGCKCQWG

24531
DKKPVEDDM

24532
DKVVQQCKD

24533
DLEHKHQFF

24534
DLRCDPQMW

24535
DMDAQGAMW

24536
DMYFTRHTW

24537
DMYSMVKHN

24538
DNFPWCTGN

24539
DQCVLSRGH

24540
DQEWSVHMG

24541
DQFCTYDYT

24542
DQGPSESDL

24543
DQKVMFECY

24544
DQQCAQMGV

24545
DRQCCKLTF

24546
DSHRLPGNW

24547
DSMHYYMMH

24548
DSNYEEKVI

24549
DSQYMCVHR

24550
DSRQFENQC

24551
DSTGRPMMG

24552
DSVAHSSQK

24553
DTDRGLVYE

24554
DTHAVEFAA

24555
DTHWWNILC

24556
DTYGIQDFQ

24557
DVDFEWSQW

24558
DVEPEYCAG

24559
DVFIDWSGQ

24560
DVIMGPIAN

24561
DVINRYECG

24562
DVKQQPLAI

24563
DVKTDTSPD

24564
DVLLADTEP

24565
DVRDNEPER

24566
DVRQILMCH

24567
DVWQNSGAL

24568
DWNQVDNYY

24569
DWPAQKGWA

24570
DWQPMLHMN

24571
DYLAMNLHF

24572
DYMHMLTAQ

24573
DYMQNQYGT

24574
EAEIFRLEG

24575
EALMPMLGA

24576
EASTQWLMQ

24577
EATCDPPYN

24578
EATIMDQVV

24579
EATKQSWTH

24580
EAYLTIDKL

24581
ECDRYIAEY

24582
ECGLWWGSE

24583
ECKPIAGDK

24584
ECPNVNTAQ

24585
ECVANPFQP

24586
ECYHVLANS

24587
EDTHMIAGH

24588
EEFDTVAAP

24589
EFMLSSKDI

24590
EFYHFMYKD

24591
EGVESMLYA

24592
EHDWITVYE

24593
EHVIIKKNY

24594
EICVVTGEQ

24595
EIEMMGRGE

24596
EIKVVNSRD

24597
EIKYRRCMA

24598
EIRECEECC

24599
EITDWGVVP

24600
EITYRRCMA

24601
EIYMRHNFP

24602
EKAHSWQDP

24603
EKISICSQY

24604
EKMESPECI

24605
EKPYHEEKE

24606
ELLMINAMS

24607
EMARNIEQN

24608
EMRMNIQWE

24609
ENCEPMHCE

24610
ENCHITKMD

24611
ENSYTHDTI

24612
ENTLFMRYH

24613
EPKQEEWYV

24614
EPVTFVGQA

24615
EPYFCCCVP

24616
ERDHVNESC

24617
ERSWYYDQY

24618
ESCEGILTF

24619
ESDPMQQFT

24620
ESEYNSSYN

24621
ESKSCGLTG

24622
ESNLTPWSQ

24623
ESRWMDEHT

24624
ESTHKFMQD

24625
ESYSTAEMA

24626
ETEWAWVTQ

24627
ETLPIEIRS

24628
ETSKWHAVC

24629
ETVLHQWAA

24630
EVAMLEEDK

24631
EVCLRHYWQ

24632
EVDCKVSHV

24633
EVDVHDEWV

24634
EVEAIRGIQ

24635
EVEFMSYRW

24636
EVISYVENV

24637
EVLHRDLHM

24638
EVMCSDVPK

24639
EVNCTHLYN

24640
EYARKTKGW

2464/
EYPYRYEFS

24642
EYRWMPNHL

24643
FACQCKGCS

24644
FAKYMNHGG

24645
FDEPMIHQW

24646
FEYIWVYWY

24647
FFCPKHKWP

24648
FICLRYKDQ

24649
FIWVCHTYK

24650
FKMWKLFQA

24651
FLELMQIGL

24652
FLRYMVCAH

24653
FMEQDIGHA

24654
FNGSICDTY

24655
FNNNDVHPI

24656
FPSAYQHQH

24657
FQSMGEIWC

24658
FRGHGMWDA

24659
FRLCSDVRC

24660
ESNLVEVSK

24661
FSTCTSCCA

24662
FTCPYKFGM

24663
FTEQEDRQM

24664
FTMASAQAD

24665
FVCLGRKGP

24666
FVGWYWNSP

24667
FWKIKSGGA

24668
FWQMKSCLP

24669
FWVLDMCQA

24670
FWYPKDRMV

24671
GACERRDAM

24672
GADQCMLWE

24673
GAIYENKLG

24674
GCAPLVQDD

24675
GCAYQAVNP

24676
GCISTLNGM

24677
GCMKLVGTA

24678
GEVVPHISW

24679
GFHLEEKYD

24680
GEKQYATYI

24681
GGHFDGWQR

24682
GGMWLQNVS

24683
GGWFMGVMG

24684
GHREFQMPY

24685
GHVAYPPDQ

24686
GIARMFRWV

24687
GIGYKWFPG

24688
GLMPVMNYT

24689
GNATKIKLN

24690
GNRRCWVPF

24691
GQGGNCKSH

24692
GSHEYWEMA

24693
GSLFQNYQW

24694
GSYMACRMD

24695
GTCNEQMDS

24696
GTDCFWWHE

24697
GTSFVGNPL

24698
GTYVCQNHD

24699
GVAHPEAVC

24700
GVTEHNEVC

24701
GVVDESYDP

24702
GVVGLNTCV

24703
GWCNRYFWD

24704
GWIRYKNSE

24705
GYFGKCKRI

24706
HADMEQGNS

24707
HADSNIACN

24708
HAESMVCHH

24709
HASLNLNPS

24710
HCIWNWSDA

24711
HEAKQVPKA

24712
HERCSELEY

24713
HEYIYETWP

24714
HFYVGSPED

24715
HHQYIDCFV

24716
HIDPDHWHG

24717
HKLYEDRIN

24718
HKNDLAAVQ

24719
HKSDMWWTA

24720
HLPAPELYL

24721
HMDIQMFMP

24722
HNFCMYWPQ

24723
HNHSADSFA

24724
HPELLDRFE

24725
HPSSIWSIL

24726
HQNKFEWSP

24727
HRNEQLIRA

24728
HSGELYDGF

24729
HSMLETTGK

24730
HSNFFHMND

24731
HSYQQCAVG

24732
HTPQVLKGE

24733
HTTAHQNWA

24734
HTTFALVMH

24735
HVDSNIACN

24736
HVKSPTFQT

24737
HVKVHEMMI

24738
HVMPKQGFN

24739
HYCKIIPYH

24740
HYEYCFGIF

24741
IAGTYTNWY

24742
IAMDNQNTD

24743
IAQDTGTLH

24744
ICKIGSRLD

24745
IEGHWAQFP

24746
IEIATVNVH

24747
IFMVHDSDY

24748
IFVGKNAAW

24749
IGIQFLKDH

24750
IGTYGWIEA

24751
IHDASTKDQ

24752
IHMHEVGVY

24753
IIKDVGDFG

24754
IIPQVDQYY

24755
IIRDKTQDD

24756
IKCDKCQDQ

24757
IKGRFDWDC

24758
IMEYVHKPG

24759
IQAATNLWQ

24760
IQGEVTVTA

24761
IQGPEAIMK

24762
IQGVQWMNH

24763
IREEMHEQF

24764
ISGLAQMGM

24765
ISICYHMFP

24766
ISMQICAGW

24767
ITCQSNEQH

24768
ITGTRFAEV

24769
ITMPAEAFQ

24770
ITTKYQIDH

24771
ITTPQLNSV

24772
IVDHLPRHG

24773
IVGLATTAM

24774
IVHPTLNTP

24775
IVMREHGAC

24776
IWADLECGR

24777
IWDLAGDAM

24778
IYNQRQMCP

24779
KACDRQIYE

24780
KACEMYASF

24781
KADAKVPCC

24782
KADVRSAQH

24783
KASNDYLCR

24784
KCDGLWCHF

24785
KDRMGLNQC

24786
KDVTKLRCP

24787
KEPITAEHK

24788
KFAGAGIGW

24789
KGKLEYRIR

24790
KGLDVPHWG

24791
KHCEILCNW

24792
KIAEWEDWM

24793
KICSQLDSP

24794
KKFWDTVIC

24795
KKNCREPEQ

24796
KMQAMGVNW

24797
KPTKYEWLH

24798
KREAWMVGN

24799
KSLIQQSWD

24800
KSMVIRHAQ

24801
KTCHPEGMP

24802
KTNMHAKSV

24803
KTNRTYMWR

24804
KVEACQSVE

24805
KVGAHVAQE

24806
KVGIHDSLV

24807
KVKWKDKDL

24808
KVYYIISHQ

24809
KWIMWIEYN

24310
KYHNVPSKI

24811
KYQYYQEGP

24812
LAGCLCYTN

24813
LAGCLWYTN

24814
LAICHSNLA

24815
LANMPNMGF

24816
LATQACSCN

24817
LCASRGLDP

24818
LCGPIIKAS

24819
LCIRLWTSA

24820
LCPNWWRYQ

24321
LDRKACMIA

24322
LEAELMGHC

24823
LEVDMHASY

24824
LFEQKMVWS

24825
LFSPYEACC

24826
LFVVTTTNN

24827
LGSLYRRRG

24828
LGYHVQEAG

24829
LHCQMGDHW

24830
LHEIQKPNH

24831
LHSQADGLY

24332
LHTPQHDCH

24333
LKDQFHWGA

24334
LKELIQFGN

24835
LKMDMTKRS

24836
LLPPMMMGQ

24837
LNLYWFWSA

24838
LNPMTQNHE

24839
LPLQVSNMA

24840
LPRLNLVDR

24841
LQDVIVDCE

24842
LQTPQHHGA

24843
LRYYIEMDR

24844
LSQMLAHTA

24845
LSVWAIRND

24846
LTHKRESEC

24847
LTMGFEIGW

24848
LTNPWHYFA

24849
LTVIIFDDA

24850
LVAMVPQSY

24851
LVDQGKCMQ

24852
LWHWLWLKT

24853
LWTRMVYFG

24854
LYAHGNCYP

24855
LYASVMQSE

24856
LYKMVTKLH

24857
MAPTVVRKK

24858
MATPKEVMQ

24859
MCAGWCDDQ

24860
MCGLRYYWP

24861
MCLWIRACE

24862
MCPNHKPCW

24863
MESLCVYFY

24864
MFMKNELIA

24865
MGCQSQKPK

24866
MGMFNTMAE

24867
MHETHTWGA

24868
MHMSTLDAK

24869
MHVQQQRYG

24870
MIYKDYNHA

24871
MIYPCMTSC

24872
MKHESDGGF

24873
MLAGWWQET

24874
MMIVQCSNK

24875
MMKWTEYMV

24876
MMTLKNTMD

24877
MNFNSLAQW

24878
MNLPKSHDC

24879
MNMYYTGEV

24880
MNNIYECIP

24881
MNSHNHTST

24882
MNTPTFHQI

24883
MPTMMGSYM

24884
MQFRTNNAN

24885
MQNISCPQC

24886
MQPRYNCEG

24887
MQSFVGVMW

24888
MRALSVSQG

24889
MRTMFYTSP

24890
MRVIQGDNY

24891
MSFTYVGGS

24892
MSTKPAYWG

24893
MSTKPVYWG

24894
MTDLQLIER

24895
MTEILQITG

24896
MTEPFMQDL

24897
MTIGYFVQA

24898
MTNRMQALQ

24899
MTRFKIYHE

24900
MTTHCNTAI

24901
MVNCAVTYQ

24902
MVSDTQHGK

24903
MVSYDVLGK

24904
MVSYNVLGK

24905
MVTTQQIVC

24906
MVVIKQYHT

24907
MVVVIRDAS

24908
MWGEWWPGC

24909
MWQEFQWQY

24910
MYIPRNGEP

24911
NACMVTGCG

24912
NAEWQRDTF

24913
NASPCQIFG

24914
NCFVMYCGR

24915
NDKWGWWRE

24916
NDVMWQLIH

24917
NFDFNHQCQ

24918
NFEIYHLQC

24919
NEGIYHLQC

24920
NFNWTRADC

24921
NFRTINAHP

24922
NGDPVFRMA

24923
NIEGIEFNA

24924
NIVGATEWD

24925
NKDDTCFQF

24926
NKYCIAMPC

24927
NLGETITNA

24928
NMIHLWTTE

24929
NMNQNPTMS

24930
NMYWCMQIH

24931
NNRETMIHG

24932
NNREWDVCY

24933
NPPDVGCPS

24934
NPQSLCYSC

24935
NQHWLLSVP

24936
NQMPCWVFF

24937
NSCFIAENC

24938
NSGIATADH

24939
NSIQYGQAP

24940
NSKSPSEGC

24941
NSQPWYRHS

24942
NSTAHAFAY

24943
NSTAHPMMW

24944
NTHVCNLHE

24945
NTLFWDGNP

24946
NVEERNRHF

24947
NVEKQVAMG

24948
NVHHTTQIP

24949
NVMSALDLS

24950
NVTQWDEDL

24951
NVVQCNGPH

24952
NWFGINSWH

24953
PANYSMPNC

24954
PDGYRRSHG

24955
PDWCQDPGM

24956
PEELYLYRK

24957
PEYRQWMLI

24958
PFLFTVIHN

24959
PFVHDDMAR

24960
PGEENEPPA

24961
PIWTQHHTM

24962
PLQQYDGCS

24963
PQHCNTMWG

24964
PQIDSLQDY

24965
PRNDDNCLP

24966
PVIWLPVDS

24967
PWDKEEPQG

24968
PYAIEESHD

24969
PYIRRVHVS

24970
QAACAMATT

24971
QADKQFYGN

24972
QAGYQSALY

24973
QANGENLDH

24974
QCDRVWLNP

24975
QCIPKQKDY

24976
QCQTLDCCD

24977
QCWYCELCP

24978
QESEVHVFG

24979
QFQLVTIQG

24980
QGAFHTAWT

24981
QGEKCVPDF

24982
QGIMTSCQT

24983
QGVRMVGNS

24984
QHCMHSMVE

24985
QHQHDLPTG

24986
QIAVVSCQR

24987
QIAVVCSQW

24988
QIFKALVGE

24989
QIGWDDAIT

24990
QIISETGWE

24991
QKFCNIHEY

24992
QKQTSISFS

24993
QKTWLVVWC

24994
QMATPVDMT

24995
QMGCTFQTP

24996
QMGLLDTYP

24997
QMYMDFTQP

24998
QNYWCMCGA

24999
QPKEWVIKE

25000
QPRNDCNEH

25001
QQAMYTSSG

25002
QQDPMKQWS

25003
QQEVEQTMG

25004
QQEYFDDMQ

25005
QQSAVGAHY

25006
QQVYEMKWH

25007
QRVMSGKPL

25008
QSDPQPVIW

25009
QSGIHCCCG

25010
QSGWFLEAK

25011
QSHFCFENA

25012
QSIPEWSCA

25013
QSNGQKDSQ

25014
QSVDWIKCE

25015
QTDMANKMG

25016
QTGWLHSYG

25017
QTKTGEIQK

25018
QIKTGKSAT

25019
QVHTKWDGM

25020
QVQYRTCDW

25021
QWTEKTFEG

25022
QYEPLGRAM

25023
QYKSTQRRN

25024
QYYMLSESA

25025
RAKTCIYKC

25026
RAMCILHNQ

25027
RAMPTAAQI

25028
RDLFIQFGP

25029
RDMDMQMWC

25030
REAKVFELK

25031
REGCYNTQE

25032
RFELLMTGR

25033
RGFWVEKSG

25034
RGSAKDAWG

25035
RISSEGMWC

25036
RLHIDDEDH

25037
RLVRDSKNN

25038
RMEELINDK

25039
RMVTWDQSE

25040
RNMCRHNDI

25041
RPETVHASF

25042
RQGCLLAWP

25043
RQICNEPWN

25044
RRNELLIPD

25045
RSEMVNTQM

25046
RSQWHNSID

25047
RSSEYLTQQ

25048
RTLVKGAQC

25049
RTQGCGPSR

25050
RVADVMYIG

25051
RVECRPWGE

25052
RVMDNACDM

25053
RWDYYWTGM

25054
RWRNCGSNG

25055
SACIFYAHK

25056
SAEMWVKDE

25057
SALPCNTAQ

25058
SARTEERES

25059
SATTMPRTD

25060
SCFINSAYP

25061
SCGLVQRFM

25062
SCGQVEHYC

25063
SCQMYTYGI

25064
SCRLRFNYA

25065
SCTTCHSWQ

25066
SFASMYNIK

25067
SFMYKTQAL

25068
SFYKNGMSN

25069
SGGFCNYAT

25070
SIGPASNAL

25071
SIHMAEGVE

25072
SIHRWNWMG

25073
SINPESQGS

25074
SISPSMGEK

25075
SIVHPPTTS

25076
SKGCIIDMQ

25077
SKLLVTRNH

25078
SKRLVTRNH

25079
SLKDVDLWH

25080
SLYKHEIWW

25081
SMFPFNMGP

25082
SMTDCDFHQ

25083
SNHPNMHTY

25084
SQGNTRYSY

25085
SRCLRHQEC

25086
SRMDMNHGL

25087
SRTMYKWMP

25088
SRTNIEWGG

25089
SRYKCQHRN

25090
SSDQLIMWG

25091
SSHATNFCG

25092
SSHTTNFCG

25093
SSTGMQSST

25094
SSTTHSRMQ

25095
STFPKWTSE

25096
STHFRTEDR

25097
STLVYPCGP

25098
STSAQPNFD

25099
STTQWGMCN

25100
STVSQMNYF

25101
SVCPSHMWM

25102
SVFSFHSDT

25103
SWDVRVTAQ

25104
SWSMQNKST

25105
SWVSADEVP

25106
SYGQIECYQ

25107
SYLRAHHKD

25108
TAALMGNMA

25109
TALDNSTGI

25110
TAQLMQTAC

25111
TAYLNNWVP

25112
TAYPEYCGN

25113
TCAPQGKQE

25114
TCAVFVFHC

25115
TCEEYYNPF

25116
TCELMQVMH

25117
TCQNIAMQL

TABLE 80

Sequences of the 581 to 589 Region in AAV5 VP1

Capsid Polypeptide that Drive Sciatic Nerve

Tissue Tropism

SEQ

ID
581-589

NO
Sequence

27991
EIILDDQQR

27992
LDDNAWRHR

27993
LGGKPYALD

27994
GSQQTTLSC

27995
TWESQIDCI

27996
VVGLKPMHM

27997
NFNINIYNC

27998
ANHATGGQS

27999
EFKRMTCNK

28000
NKHQGMHRM

28001
KGGPWAYNC

28002
VLIPIGQHY

28003
DPFECTVWP

28004
TVQWNFAHM

28005
TAYMTDMYC

28006
SATNWECSN

28007
KVDAEGFHF

28008
VWEDNPPFR

28009
SVLMSPHLM

28010
LEKNSSQMR

28011
CRCNDAWNM

28012
AAPFMWRDL

28013
ACKYPPHVR

28014
ANKMFMEQA

28015
ANKMFMGQA

28016
ANKMVMEQA

28017
AWSEFQWFD

28018
CYRCLPPEA

28019
DIDRHAHFD

28020
DKGYSLYQP

28021
DRDWSDVRR

28022
DRGWSDVRR

28023
EHGYMRCTK

28024
EHGYMRCTR

28025
EHGYMRCTT

28026
EHGYMRYTK

28027
EIECLQTVA

28028
EIEFLGTVA

28029
EIEFLQTVS

28030
EIEFLQTVV

28031
EIEILQTVA

28032
EIEVLQTVA

28033
EIFLDDQQR

28034
EIILDAQQR

28035
EIILDDEQR

28036
EIILDDQQC

28037
EIILDDQQH

28038
EIILDDQRR

28039
EIILDGQQR

28040
EIILDVQQR

28041
EIILGDQQR

28042
ELALGVLWK

28043
FPPFCALCT

28044
FPPFYALCT

28045
FPPFYAPCT

28046
FPPFYTLCT

28047
FTAYPQAVM

28048
GAIVFDLCD

28049
GAWDYMYEC

28050
GCFWAANSC

28051
GCFWAANSG

28052
GCFWATNSG

28053
GIILDDQQR

28054
GMPTNHQYI

28055
GMPTNRQYI

28056
GMSTNRQYI

28057
GNLIDWQHI

28058
GNLIDWRHI

28059
HADFERHSQ

28060
HADFGRHSQ

28061
HLSFMDLGS

28062
HLSFMNLCS

28063
HLSFMNLDS

28064
HLSFMNLGI

28065
HLSFMNLGS

28066
HLSFMSLGS

28067
HLSFVNLGS

28068
IALTVTRFP

28069
IQHWMSHSH

28070
IQHWMSHSR

28071
ISHCHEDHK

28072
ISHSHEDHK

28073
ISHYHFDHE

28074
ISHYHFDHK

28075
ISHYHEDHR

28076
ISHYHFDQK

28077
ISHYHFGHK

28078
ITQCTMMGV

28079
KLPGSHAER

28080
KLPGSHTER

28081
KLPGSRAER

28082
KYASVLEQA

28083
KYASVPEQA

28084
LDLKWWCTF

28085
MFFQKPKWS

28086
MIALRMEGN

28087
MIDLRMEGN

28088
MIDLRMEGT

28089
MIDLRMGGN

28090
MIDLRVEGN

28091
MIDLWMEGN

28092
MQEQQKAYE

28093
MQEQQKTYA

28094
MQEQQKTYE

28095
MQEQQRTYE

28096
MQQSEQCYP

28097
MQQSEQCYQ

28098
MQQSEQFYQ

28099
MQQSGQCYQ

28100
MREQQKTYE

28101
MSFQEPKWS

28102
MSFQKPKWS

28103
MSFQKPMWS

28104
MSFQKPRWS

28105
MSFQKTKWS

28106
NCLPWGQFC

28107
NCLPWSQFC

28108
NCLSWSQFC

28109
NLRECTIMN

28110
NLREGTIMN

28111
NLWECTIMN

28112
NRGCSDRIS

28113
NWLPWSQFC

28114
NYIEACAMN

28115
PREICWSHS

28116
PVCSTHFWQ

28117
PVSMGTKRE

28118
PVSMGTRRE

28119
QCRHEGSNG

28120
QCRHEWSNG

28121
QCYTFNDTK

28122
QHNYVSVVE

28123
QHNYVSVVG

28124
QHVGLQAEK

28125
RENTEDQIR

28126
RHGHGHHYD

28127
RHSHGHHYD

28128
RHSHGHHYG

28129
RHSHGHHYY

28130
RHSHGPHYD

28131
RLSFMNLGS

28132
SIEVRPPLC

28133
SIEVWPPLC

28134
SKAKYKTQM

28135
SMEFEDQCV

28136
SMEFGDQCV

28137
SMEVEDQCV

28138
SVEVRPPLC

28139
TFCFVQTLV

28140
TFCFVQTRV

28141
TEKENWMDN

28142
TEKENWMDS

28143
TFKENWMDT

28144
TFKFNWMGN

28145
TFKFNWMYN

28146
TFKVNWMDN

28147
TFRFNWMDN

28148
TFTENWMDN

28149
THCPFQHDI

28150
THCQFPHDI

28151
THCQFQHDI

28152
THCQFQHDM

28153
TNKMFMEQA

28154
VAIVFDLCD

28155
VAIVFGLCD

28156
VAWDCMYEC

28157
VAWDDMYEC

28158
VAWDYMYEC

28159
VAWDYMYEG

28160
VAWGYMYEC

28161
WANLRQMDL

28162
WGNLRQMDL

28163
WSSREYLHK

28164
WVNLRQMDL

28165
WVNLRQMGL

28166
WNVLWQMDL

28167
YYYSSKSSV

28168
YYYSSTSSV

28169
WSSREYWHR

28170
WSSRGYWHK

28171
WSSREDWHK

28172
MHNPLWFSS

28173
ACVDHMEER

28174
AGYTIHTVA

28175
AGYTIHTVE

28176
AIHLPSACW

28177
AITMDAQPN

28178
ATIDHNSPG

28179
AVGSEDSSD

28180
CAHIMDELG

28181
CDINVLIMM

28182
CDINVLIMV

28183
CDKLDKPAV

28184
CDKLDKPTV

28185
CFAHMQIAN

28186
CISNHNSTY

28187
CLSMFVGNT

28188
CVKLYVNED

28189
CVKLYVNGD

28190
CVRKCDLCG

28191
DMPWVDYMG

28192
DMTFEAYPT

28193
NATEEPYPT

28194
NATEETYPT

28195
DNDVEMAAK

28196
DSDNGPNIF

28197
EAHPHMMSQ

28198
EAQPHMMSQ

28199
ECGRYGVFS

28200
EFMEVMDQV

28201
EGGFQDFCE

28202
EGGFRDFWE

28203
EMLRCCDCT

28204
EQLDVQEVY

28205
EVDVSNDNC

28206
EVDVSNNNC

28207
EVSDGTPTV

28208
FALWSNCAE

28209
FALWSNCGE

28210
FALWSNCVE

28211
FDCHNASKP

28212
FLTEMONHA

28213
FLTGMQWHA

28214
FVADWIWPA

28215
FVADWIWPT

28216
EVADWIWSA

28217
GCFTFWDHQ

28218
GCLQVNQME

28219
GCNMIPKHD

28220
GDMPERETG

28221
GHVSTSNQG

28222
GQHSLVTMY

28223
GVQFRYSMP

28224
GVRVQDSHH

28225
GVTHKAKHE

28226
GVWVQDSHH

28227
HSDYEMNDD

28228
HYTEGYRQT

28229
IADHFLAHF

28230
IADLCNCEN

28231
IGAPVDTIN

28232
IGTAFRNSG

28233
KCESDYLPE

28234
KTNWWDGQC

28235
LAIRKIANH

28236
LAIRKITNH

28237
LARYCEKTP

28238
LCMESIMMI

28239
LCMESLMMI

28240
LCTESLMMI

28241
LDRRSSAYS

28242
LFSAWNDFA

28243
LFSAWNDFV

28244
LMCYMQKIT

28245
LMGYMQKIT

28246
LMSTMFNTF

28247
LQVHWWLLL

28248
LTIPLHAWS

28249
LYGQVVAID

28250
MAIFDSAAV

28251
MFWSGNKQG

28252
MQCDFLHDT

28253
MQKDVEQMW

28254
MSMCYGIKG

28255
NAMTQGNHQ

28256
NAMTQINHQ

28257
NAMTQVNHQ

28258
NAMTQVNHR

28259
NHNPTNRNC

28260
NKPCKYMYK

28261
NPTTVCLKM

28262
NTHEGTDGN

28263
NTTPNCNSW

28264
NTWPYPENS

28265
NYLQFGHKQ

28266
PGTQPYMGW

28267
PQEIKSTSC

28268
PRECHQSHQ

28269
QASFKQMGD

28270
QASFKQMGG

28271
QATCYDTSP

28272
QFTHNQAAN

28273
QGIQIGKCQ

28274
QHVAFNRWH

28275
QILDQQEPP

28276
QINNVVDNR

28277
QISMNVAFR

28278
QMAEFTYYG

28279
QMFDTWKCW

28280
QTSSYDAHC

28281
QTSSYDTHC

28282
QVTGIPWFY

28283
QWEDRTQMS

28284
RATCYDTSP

28285
REMEYQGER

28286
RPPNFVATG

28287
SAMPETTIF

28288
SAMPGTTIF

28289
SQPMLKVNA

28290
SSSDLCQNR

28291
TCMPFGTSQ

28292
TFKLTTGGE

28293
THDDCQWWE

28294
TIHLPSACW

28295
TIKLTTGGE

28296
VIMWNYEHY

28297
VSVSIYSGW

28298
VTMVKRKGT

28299
YDDWYSNVY

28300
YPMPMCCQP

28301
YHAPMCCQS

28302
WCRTPQREN

28303
GMTNEVQAA

28304
EGGFRDFCE

28305
RYEISERRC

28306
SAISCQCAK

28307
EVQRNIYSP

28308
HCTEGYRQT

28309
SAPEFSVIC

28310
MFRQFYIQG

28311
STVGQNHCG

28312
EFCHQHSAI

28313
TGWNFPVSC

28314
GCLTSECAY

28315
TMFYCHGPN

28316
HITPMITMW

28317
AITRHVADT

28318
QRSVFHQRS

28319
VKRLENEWH

28320
MMAMKQGFY

28321
EGRTISYGN

28322
STCYPFAMG

28323
QIIYRCRST

28324
GTAVSRQSK

28325
GCEEVGRCQ

28326
QHCFRAMED

28327
MNSFYRAEW

28328
VCHNPDPFT

28329
NGYLVHACQ

28330
NPGWQTIGQ

28331
LCAVCGDCD

28332
ATLMSDGGG

28333
YFPSIQCYE

28334
GAANRIQSW

28335
MFMTNVNSF

28336
VSVQSRAQL

28337
PLTIEVNCT

28338
KTMGIGGGP

28339
NRIQMTDFQ

28340
DFNGSNWLP

28341
FAIWKRNES

28342
HFQPVFMQP

28343
DQEAQVVTG

28344
SECEVLCSL

28345
AAMESCAEI

28346
NQREWHGLA

28347
NYVNQGMGP

28348
NISPYEMAV

28349
ATGTYNLQE

28350
HATGLVNFM

28351
NAIPYFVQA

28352
CVEQYVEQG

28353
AVAEIRPEP

28354
ASKQGTHDY

28355
SSEYSQFAI

28356
TMTVRMEQP

28357
MWGCKLFVC

28358
AGMYKCQED

28359
TYESLQNSW

28360
TKMIIFSDW

28361
LCKFANGAA

28362
INYISFVDR

28363
RQAWPKDQM

28364
QSAHPEPMC

28365
YCECGHWPN

28366
SQERMDYDG

28367
WINEANCLM

28368
SNVACIQAF

28369
YTKYNWKAA

28370
NYAMRRDSY

28371
PVICVWHDN

28372
SFKSVSQYN

28373
ENTMWQSSQ

28374
NAYYDSHCE

28375
KCADLIIRS

28376
EKRLEGWVY

28377
KDGRNPHCL

28378
QSRCVDNTV

28379
ATMNLSWGL

28380
YSDPKNMSI

28381
PDLWYEKSS

28382
TFVRKPSLM

28383
PYGPIEYDE

28384
EPRLYCAQE

28385
DRKQADQIY

28386
KGNPLDGDT

28387
GLFEGSPGA

28388
DIEFLDCEN

28389
EIHMFGQSE

28390
WGGMPNNQG

28391
SVLHVQALA

28392
ASMHEETCL

28393
MHGIARMQQ

28394
EAGCKMWPT

28395
KMAGIGCWY

28396
AVYLDHVFS

28397
GHLFNNNEW

28398
SNSHPFQSH

28399
LSAQRLVCD

28400
TVNWYSGFG

28401
YIGMKSCSG

28402
NHSLCWDSK

28403
PIWQGYWPW

28404
AELFYDAMY

28405
CQVNEEYDW

28406
DTEAIMVVD

28407
ESWRTLCQI

28408
GHHNTPDMQ

28409
GWDKKQNCG

28410
IKADVDAME

28411
LTTTVPTKQ

28412
MAEPVKRCK

28413
MCHKVMDII

28414
MSECKNPAY

28415
TEVEISASF

28416
TTVAGPHTT

28417
VSFWYDCHH

28418
AACYDYKDH

28419
AAKLPVAKG

28420
AEFLTYEQK

28421
AFWQDLYYV

28422
AGGGSVQCN

28423
AHLNCDKSM

28424
AMATCQMQH

28425
AMGRLHDSK

28426
AMMLTDLML

28427
APHWMEISS

28428
AVMWTVLHF

28429
CADWACMSL

28430
CAVKQASAM

28431
CCWFVYEST

28432
CFTESITDC

28433
CIIVRSTAA

28434
CIKYNSSIG

28435
CQSNKQQFC

28436
CVFSWNNYQ

28437
CYEITMNQG

28438
DCLISECAY

28439
DFETWDWFG

28440
DFNHIAWWQ

28441
DHHGIDTTL

28442
DIWLWDTFA

28443
DRPNHSTDG

28444
DSMFIIMNF

28445
DSMFTIMNF

28446
DVASWCQCP

28447
DYENCFESY

28448
DYGYVSINF

28449
DYTTHWQYA

28550
EEKYQPCLK

28451
EFKRMTRNK

28452
EGTQTHSAT

28453
EIKFPACGL

28454
EKLNKTWAG

28455
ENDHRDYNI

28456
ENYMCYHSL

28457
EQAPLDVMF

28458
ETMEHKKRE

28459
ETMQHKKRA

28460
ETMQHKKRE

28461
EVADENGNY

28462
EVCMAVVDN

28463
FKRICWEAG

28464
FNLSSCYHQ

28465
FTEYEHRAS

28466
FVAANQNHM

28467
GAAVSRKGY

28468
GAEQHTNWP

28469
GDIRTYPAE

28470
GFTQNYAQN

28471
GLIHSTHTV

28472
GMTNEDQAA

28473
GNPHLIEHC

28474
GRAHGNVRI

28475
GRVLLMAQD

28476
GSFPCYSLG

28477
GSFPCYSLS

28478
GSTEVEYMI

28479
GTAVSKQSK

28480
GTNHSAEVY

28481
GTRATKHHV

28482
HAPPHGTWA

28483
HAVNDHSWM

28484
HCKVKECQY

28485
HDEWLGHGM

28486
HIDRPPCCM

28487
HWMEQPPYG

28488
IQCNLCEAW

28489
IQYNLCEAW

28490
IWGICVHYF

28491
KENTINRID

28492
KHHMFVDKH

28493
KKDEWHCYP

28494
KMCWWPEAM

28495
KONASDQVH

28496
KQVDPLDFW

28497
KSDFRVLHQ

28498
KSEYVVGVP

28499
KTREHRERS

28500
KVDAEGSHF

28501
KVMVVDSVY

28502
LATMWFCAD

28503
LCYHFWKVN

28504
LEMHYKDNC

28505
LFRSHWDAQ

28506
LKASLREQW

28507
LMENYEIQD

28508
LMQKWMRTC

28509
LYLPHCHRI

28510
LYLSHCHRI

28511
LYMWQNQHV

28512
MATCYMYVD

28513
MCSYFNNRE

28514
MEFYGRAGN

28515
MESGTLLGV

28516
MFCEFPKCG

28517
MFGTKMSQY

28518
NWREFPKCG

28519
NIDLGNRGF

28520
MIDWGGEWG

28521
MMLHKQKDY

28522
MMWKENAGW

28523
MSECKNHAY

28524
MSNYHITHE

28525
MTAHKENHP

28526
MTAQSPLIP

28527
MTDYFKALC

28528
NCDHWFGTL

28529
NDKSWLKHC

28530
NETDVGTDV

28531
NQREWHGLS

28532
NRGYANVAW

28533
NTFVKNEFW

28534
NTLTMQSYL

28535
NTLTMRSYL

28536
NTNYWFYYD

28537
NVEGTRMRP

28538
NVEHSHCVG

28539
PKLCSHQTE

28540
PRLQSRKYL

28541
PSCITQESI

28542
PSTTIMNVM

28543
PTFGWMNWF

28544
QCERYHAAD

28545
QCRLDMQCP

28546
QFEDYWAIN

28547
QGQKQNTTM

28548
QHFVQGTYP

28549
QICPQSMMP

28550
QKDERHCYP

28551
QMFMQMSGH

28552
QNSRHKYAW

28553
QTLGCEMGT

28554
QTLGWEMGT

28555
QTVETYAAH

28556
QVGQYQNTI

28557
RANDGYYIN

28558
RDGFQEFTP

28559
RFTGMSDQD

28560
RNMSRQEGM

28561
RRFVQGTYP

28562
RTMGMDWFT

28563
RVAMQQMNH

28564
SAWQWAQMN

28565
SCAVCGDCD

28566
SCTARAWIC

28567
SESQEGVYG

28568
SGALKNREG

28569
SKYHWHMYH

28570
SRDAAAYSA

28571
STEWAALGV

28572
STFISSGER

28573
SYIPFDTQK

28574
TAAGYIVDV

28575
TAYMIDMYC

28576
TCNVSWKHA

28577
TCSYFNNRE

28578
TGKVDGMQY

28579
TLPKMYMWP

28580
TMRTTMCIK

28581
TMRTTMCIT

28582
TPGINRHCC

28583
TTFITETMD

28584
TTHQTAFEW

28585
TVNEALTCH

28586
TVRWCHWCL

28587
TWAMHPSHE

28588
VAWSCDKFA

28589
VEDMGQVEA

28590
VGQNFNTGR

28591
VGTDCATSL

28592
VPSLNTSAG

28593
VQKETDEVC

28594
VSDDYECGI

28595
VTQWKYGDT

28596
VIRNEEWFP

28597
VTYTNGMAE

28598
VVTVHVRGH

28599
WQIGGCPEE

28600
WTHTYATGR

28601
YAEVAVQQE

28602
YAMPLYFWE

28603
YAPQNEKPE

28604
DFKFHSTGS

28605
MMVIKEVNL

28606
RTEGWFKYD

28607
TSMWGLETS

28608
FPGVMEGEI

28609
TAKQHFMVW

28610
FKMDCHSCG

28611
ATWPERWQN

28612
LMENYEFQD

28613
ESYHYKLMT

28614
QPIHFQQRD

28615
VVNQQEFYY

28616
AQKITDEVC

28617
DVRSSMDHC

28618
FNLSFCYHQ

28619
RWHNGKGPM

28620
RYSNVAHHA

28621
FAMEQVKCD

28622
DRVMESIQP

28623
AGSGSVQCN

28624
LPTLRVADK

28625
DTDDWTCVF

28626
SWHNNWTMD

28627
WHDMVNSGP

28628
RQCTDEECW

28629
RVVMQQMNH

28630
DRLNHSTDG

28631
FLIQGDLTF

28632
FPPIVKRHH

28633
LCEGQQWHA

28634
SGSIYHHMC

28635
VVMWQFDSQ

28636
RPDLNEIEC

28637
SADNKIGGP

28638
HAYQTMSEQ

28639
CFTEPITDC

28640
MTFVCNFGT

28641
TSCGYNDNN

28642
HPMFPNTED

28643
AFAAYCSRE

28644
CAKFTNAIH

28645
NHQDNSAYP

28646
IWGIYVHYF

28647
YWWGFCWGL

28648
KVEGYGWTR

28649
YDQYGDRTM

28650
EIGIIRHGH

28651
VDHTLDFYH

28652
TVVINACLE

28653
LRHWFFSNM

28654
AMTRQNGWM

28655
WMCEFSCIQ

28656
LTTKDDKEQ

28657
PECKGAVDP

28658
FQTPVANVC

28659
QVGQYQNMI

28660
QNADQAMSG

28661
RFVLCDYME

28662
HSEMHNHHA

28663
TAFANAHTF

28664
KVGCFVDVW

28665
NWRMVHGRW

28666
GVNDLQNIC

28667
CVVNRGSIT

28668
DTHTCSGFS

28669
DFEMWDWFG

28670
TRRSWECAP

28671
SCTTRAWEC

28672
MIEPNDHGK

28673
KKDERHCYP

28674
CHCQMEAEW

28675
RVEMQYWEN

28676
DWTINDGWS

28677
TVREDCCEN

28678
SSQKYKDWE

28679
CVWSVGAEW

28680
DTAATMEPK

28681
EPRPETEGC

28682
MVTERKPWS

28683
SYDKEHAWV

28684
RANTWTRRH

28685
HLTILHCPE

28686
YCGWKEFRR

28687
MASFVEHCI

28688
KAMGMLEWA

28689
EEKDPHMDS

28690
MDNNYHQFG

28691
W1QWVRKMG

28692
NEWSRWLTG

28693
FQNVEIEKF

28694
QNMFGNQPY

28695
CKQRNCDGN

28696
LCNNADMRC

28697
VKQSMPMKG

28698
MANLKHQLA

28699
EDFVTTSEL

28700
LKRTVGVCQ

28701
ALILYARKH

28702
ASIKFEGQE

28703
SQKQFSMQH

28704
CDNAGEFGQ

28705
NRCQFGKDW

28706
VTNEFNGPR

28707
GYKCGCVCC

28708
ACEQIFEGI

28709
AHGWSQGAS

28710
AHGWSQGES

28711
AHRWSQGES

28712
AEHEVQSSF

28713
AELKITKVL

28714
AMINPKNDH

28715
AMINPNNDH

28716
AMTNPNNDH

28717
ANEEMMAAD

28718
APAPTLREW

28719
AQSPGIHQE

28720
AQSPGMHQE

28721
ATDQPANLC

28722
ATDQPVNLC

28723
ATGTQLNQD

28724
AVHNVKDVH

28725
CASSYPRDL

28726
CASYYPRAL

28727
CASYYPRDL

28728
CEFEKMDYG

28729
CEFKKMDYG

28730
CEFKKVDYG

28731
CFCKYDMQM

28732
CFCKYEMQM

28733
CFCTYDMQM

28734
CGMCHECTL

28735
CGTAYWTAY

28736
CGTTYWTAY

28737
CPKAWEKAA

28738
CPKDWAKAA

28739
CPKDWEKAA

28740
CPKDWGKAA

28741
CPKGWEKAA

28742
CQMLLHAFK

28743
CQMLLRAFK

28744
COTETILCG

28745
COTKTILCG

28746
CQTKTMLCG

28747
CTKECMLWP

28748
CTKEYMLWP

28749
CVECNQPVR

28750
CVMCHACTL

28751
CVMCHECTL

28752
DAGCMGGWF

28753
DCARQHYYA

28754
DICLEPAQT

28755
DICLETAQT

28756
DICLETTQT

28757
DICSETAQT

28758
DKSHVDQDK

28759
DLYTYIMEN

28760
DLYTYIMGN

28761
DLYTYLMEN

28762
DLYTYMMEN

28763
DLYTYVMEN

28764
DMEEHTHSC

28765
DNLCIGAFG

28766
DNLCIGASG

28767
DNPCIGAFG

28768
DQRMLLKEL

28769
DQRMLLKGL

28770
DSCSIKCDE

28771
DSCSITCDE

28772
DSCTHHEGF

28773
DSHTANFRG

28774
DSHTVNFRG

28775
DSYTHHEGF

28776
DYASLAGQA

28777
DYASLEGQA

28778
DYASLEGRA

28779
DYASLGGQA

28780
DYASLVGQA

28781
DYGPKDFFA

28782
DYGPKEFFA

28783
EAIFAMTST

28784
EAYAMKQNT

28785
EDQVASHWI

28786
EFMCVDQLE

28787
EFMCVDQSE

28788
EGVIISLLE

28789
EMMQTCVQH

28790
EMMRTCVQH

28791
EQICSEIDQ

28792
EQVMDTWQL

28793
ESLSGGEHP

28794
ESLSNGEHP

28795
ESLSSGEHP

28796
ESLYSGEHP

28797
ESQSSGEHP

28798
ETQYWYARQ

28799
EVFLESQDE

28800
EVERESQDE

28801
EVKYKGNYN

28802
FADLILSDG

28803
FGDKMTVTS

28804
FSPNTPRSD

28805
FSPNTQRSD

28806
FWSHVGLQE

28807
FWSPVGLQE

28808
GASYFSQPV

28809
GCTWEQHAE

28810
GCTWEQHGE

28811
GCTWEQHVE

28812
GKSHVDQDK

28813
GKSHVEQDK

28814
GKSHVNQDK

28815
GKSPVDQDK

28816
GQDCWGLAM

28817
GRRYPECTT

28818
GRWYPECTT

28819
GSSYFGQPV

28820
GSSYFIQPV

28821
GSSYFNQPV

28822
GSSYFSQPV

28823
GTSYFSQPV

28824
GTYGSSFYM

28825
GVAHYHHHP

28826
GVAHYHHHQ

28827
GVAHYHHRQ

28828
GVAHYQHHQ

28829
GVAHYRHHQ

28830
GVAPYHHHQ

28831
GVFECSTCQ

28832
GYTWEQHVE

28833
HAVWRQSSH

28834
HCWGHAFFF

28835
HCWGHEFFF

28836
HCWGHGEFF

28837
HCYSTSEME

28838
HCYSTSETE

28839
HTDDLKKVI

28840
HTIITNQPY

28841
HVFSANVDK

28842
HVFSANVNK

28843
HVVWRQSSH

28844
IAHAAMHDG

28845
IAHEAMHDG

28846
IAHEAMRDG

28847
IAHEAMYDG

28848
IAHGAMHDG

28849
ICFQHQIRK

28850
ICFQYQIRK

28851
ICMAKNGWW

28852
ICMDKHGWW

28853
ICMDKNGWW

28854
ICMDKSGWW

28855
IIGFMPPTG

28856
ISNPLMTQM

28857
ITTAYPECD

28858
IVAIAHGEH

28859
IVASAHGEH

28860
KAIPQGSSC

28861
KARQDDVNT

28862
KHDEYHTYT

28863
KHNPSRHGI

28864
KQVNFNAKN

28865
KSDTQCRFF

28866
KSDTQWRFF

28867
KTVPECESS

28868
KTVQECESS

28869
KTVQEGESS

28870
KTVQEWESS

28871
LAAMQQGPW

28872
LAMRRFDNF

28873
LCFQHQIRK

28874
LENKNDCSM

28875
LFNKNGCSM

28876
LFNKNGCSV

28877
LFRAYAWFQ

28878
LGRKPYALD

28879
LGSDFNAND

28880
LLTGQNSSH

28881
LMWIALEAW

28882
LPWIHTNAN

28883
LSHKPADPT

28884
LSMQWAVIH

28885
LSMQWAVLH

28886
LSMQWVVLH

28887
LTCYHNELS

28888
LVMRRFDNF

28889
MADLIVEHD

28890
MAEHRKQEH

28891
MCHRASDQN

28892
MCQRASDQN

28893
MDGQAWKEA

28894
MGHKPVLSS

28895
MIRADIRLI

28896
MIRAVIRLE

28897
MIREAIRLI

28898
MIREDIRLI

28899
MIREGIRLI

28900
MIREVIRLI

28901
MIREVMRLI

28902
MERGVIRLI

28903
MERVVIRLI

28904
MLTGQNSSH

28905
MLYICMMAT

28906
MNMCFGQST

28907
MSMQWAVLH

28908
MTTVIYDSC

28909
MVEMDHWAK

28910
MVETDHWAK

28911
MYQRASDQN

28912
NATSIYDID

28913
NATSIYDVD

28914
NFEMQPRAD

28915
NFMLVNGPL

28916
NNEPYQKFN

28917
NNEPYRKFN

28918
NNFNDAAGL

28919
NNFNDEAGL

28920
NPDYRNEMP

28921
NTRPFEKDI

28922
PERRIGQWY

28923
PGICKANWT

28924
PGIRKANWT

28925
PMFPDSMSS

28926
PMSPDSMSS

28927
QCSYCQQVY

28928
QHVPVGPPA

28929
QHVQVGPPA

28930
QIMWFPPEE

28931
QLCSPPMLD

28932
QLCYPPMLD

28933
QLERTGVHP

28934
QLHYYNVGH

28935
QMFQGLWSH

28936
QPERTGVHP

28937
QQERFPHSW

28938
QRVQVGPPA

28939
QSDTQCRFF

28940
RHVQVGPPA

28941
RIHYELYIC

28942
RPEAVICWN

28943
RPEDVICWN

28944
SAAAVPSWH

28945
SAADVPSWH

28946
SAAVKSEPP

28947
SADVKSEPP

28948
SAHMYHAGY

28949
SATQNWNEH

28950
SATVKSEPP

28951
SDTWAMYHH

28952
SGTWAMYHH

28953
SHATFMMGN

28954
SKSHVDQDK

28955
SRDWPASMN

28956
SRDWPESMN

28957
SRDWSESMN

28958
SSPNTQRSD

28959
STHIYWICP

28960
STHIYWICS

28961
STHVYWICP

28962
STVPQMRWT

28963
SVSPHTPMH

28964
SVSQHTPMH

28965
SYCPVVQWD

28966
SYCQVVQWD

28967
TDPLVMMEH

28968
TECLCVYNF

28969
TLSSGARDM

28970
TMMFEQNSK

28971
TTDQPANLC

28972
TVETDHWAK

28973
VAEHRKQEH

28974
VAEHRQQEH

28975
VDGQAWKEA

28976
VKEQTYAAN

28977
VLAGLECQY

28978
VNEEMMAAD

28979
VTDRNAWVG

28980
VTEHRKQEH

28981
VTGTQLNQD

28982
VTNRWKRKS

28983
VTNRWKRMS

28984
VVECASHSG

28985
VVNQFEHWM

28986
WIHSELYIC

28987
WIHYELYIC

28988
WSFSERPGS

28989
WTGYSWSVA

28990
YAVMSHPAG

TABLE 81

Sequences of the 581 to 589 Region in AAV5 VP1

Capsid Polypeptide that Drive Skeletal Muscle

Tissue Tropism

SEQ

ID
581-589

NO
Sequence

30991
ACIFVVLPL

30992
ACIFVVLPL

30993
AGKEVSAGM

30994
AKKGMRHDQ

30995
AKKGMWHDQ

30996
APSKPWLHF

30997
ARLCKRKQA

30998
AWVEHAAFM

30999
AYIFVVLPL

31000
CAWDKHHKM

31001
CAWDKHHKV

31002
CDPHWAWCE

31003
CDPHWDWCE

31004
CDPHWDWWE

31005
CDPHWGWCE

31006
CFVHTSCQN

31007
CPWFGLCNW

31008
CPWFSLCNW

31009
CPWFSLWNW

31010
CRTCDEDGA

31011
CRTCDEDVA

31012
DDCQFDNEN

31013
DDFGTLPRM

31014
DENIKIIYG

31015
DENIKVIYG

31016
DIDPLRNSG

31017
DIDQLRNSC

31018
DIDQLRNSG

31019
DKLKFVKQW

31020
DKPKFVKQW

31021
DLRSPGFYA

31022
DMTPKNMQM

31023
DRAMYHKVV

31024
DRAMYRKVG

31025
DRAMYRKVV

31026
DRTMYRKVV

31027
DVYQFWTSF

31028
EACYMCRNV

31029
EAYDVAAVN

31030
EAYDVATVN

31031
EFECFHNGP

31032
EFGCFHNGP

31033
ELNLPVRWY

31034
ERRMEVLNA

31035
ETYDVAAVN

31036
EVECFHNGP

31037
FTDTHEMVL

31038
FYPRPELTG

31039
GCIQFESTG

31040
GDGYLEDSW

31041
GMAPKNMQM

31042
GMTPKNMQM

31043
GNKDCYGSM

31044
GRNFQESLQ

31045
GTMLTTEDL

31046
GVWQIHARE

31047
GVWQIPARE

31048
GVWRIHARE

31049
GTTTMQTLH

31050
HLDAIWPLW

31051
HLDEEWPLW

31052
HLDEIWPSW

31053
HLDESWPLW

31054
HMQWMGFRG

31055
HSRCSHYDW

31056
HTPHDDYRS

31057
HYMSMTTYE

31058
IAYTLQFQW

31059
ICNEQEFVA

31060
ICNEQEVVA

31061
ICNKQEFVA

31062
IDHCGCTQP

31063
IFRCFGVEY

31064
ILARWSEKE

31065
IPAFPLRMV

31066
IPEFPLRMV

31067
EPEFPLRRV

31068
EVEMPFCST

31069
KENLKTNWW

31070
KQEGDYVDM

31071
KVKACEGSK

31072
KVKSCAGSK

31073
KVKSCEESK

31074
KVKSCEGAK

31075
KVKSCEGSK

31076
KVKSWEGSK

31077
KVRSCEGSK

31078
KWMQDITYF

31079
LALNAPNFC

31080
LELPDSEKV

31081
LELPVSEKV

31082
LLPLFTPYV

31083
LMWIGQVVE

31084
LMWTGQVVE

31085
LQAQEQFHE

31086
LRNCECNDH

31087
LRTMVQNDH

31088
LRTMVRNDH

31089
LTGWRIVVV

31090
LVLNAPNFC

31091
LWTMVQNDH

31092
MAGYNDSCM

31093
MAGYNDSWM

31094
MAGYNGSCM

31095
MAGYSDSCM

31096
MIEWQFCTS

31097
MMIGNQMHC

31098
MPGYNDSCM

31099
MRGNCEIKR

31100
MTGYNDSCM

31101
NAPCMWMPD

31102
NAPCMWMPY

31103
NAPCMWMSY

31104
NDFHIEPIH

31105
NEDPGAGCN

31106
NEDPGAGCY

31107
NGFHIEPIH

31108
NGFRIEPIH

31109
NLQSMFCEP

31110
NQPSSDDRQ

31111
NVAQPTIHR

31112
NVTQPTEHR

31113
PFECEANEQ

31114
PFGCEANEQ

31115
PHEQKMLDR

31116
PPCGLKCMN

31117
PPCGLKCMS

31118
PQAQEQFHI

31119
QFFHTLWKY

31120
QFSHTLWKY

31121
QNWHMKNKM

31122
QNWHMKNNM

31123
QSTQICHNR

31124
RFWPRKVQP

31125
RFWQRKVQP

31126
RKDDQMYAL

31127
RKEGCTSHG

31128
RSGMARYAY

31129
RTMLTTEDL

31130
SHYFCTMST

31131
SPPCCKKEL

31132
SWKLKLCQT

31133
SWRLKLCQT

31134
TCCLPWQRI

31135
TGKEASAGM

31136
TGKEDSAGM

31137
TGKEGSAGM

31138
TGKEVSAGM

31139
TKKGMRHDQ

31140
TSFSFPDCM

31141
TVSCGPADM

31142
TVSCGPTDM

31143
TWVEHAAFM

31144
VDLHHPEYR

31145
VVKFSWKPW

31146
VVPQPPLET

31147
VVQFSWKPW

31148
WNPHHDFEA

31149
WNPHHDFED

31150
WNPHHDFEG

31151
WNPHRDFED

31152
YGNCRGHRL

31153
YGNCWGHRL

31154
YIDQLRNSG

31155
YNPYRFINK

31156
YNPYRFMNK

31157
AENLPRWGM

31158
CIETGILNG

31159
CTQTTVRWP

31160
EKVHCMWWH

31161
ETHEMNVCM

31162
FCPAEWGCL

31163
GRHWAFCDY

31164
HCSRHFCEV

31165
HQQSEAFVS

31166
EACMGEVQG

31167
KAHQTVQDK

31168
KCVSMDFSG

31159
CTQTTVRQP

31170
NAISVGEWD

31171
NKRNLYALQ

31172
NMVHENRFW

31173
PAWDLVGML

31174
PKGMAKICE

31175
QKKGIEAVD

31175
RFRFRAQVW

31177
RLERDNNEA

31178
VIKWRSQDK

31179
WLANSWFRE

31180
YMWWQEGTL

31181
YVVGDAACP

31182
KFAMDWVSD

31183
LNELPRAEY

31184
ISLQEMMGK

31185
VGECVASAP

31186
MREMPMVGA

31187
AAANYDQIE

31188
AACCPFAIM

31189
AAGMSGQFE

31190
AANEKPTHQ

31191
AAVQFVLSC

31192
ACMSRAMFD

31193
ACQCKVIWQ

31194
ACTWPGVGP

31195
ADQSWYIKG

31196
AEIYNCEFK

31197
AEVEADQRM

31198
AFANALACY

31199
AFEYFGNPI

31200
AFNRIGRYT

31201
AFYYLGHDS

31202
AGSTAMQSQ

31203
AGTSEDMMP

31204
AHMENELEM

31205
AHNYSLAEQ

31206
AHRQLDAEW

31207
AIAWWHEPM

31208
AIDWEDVWE

31209
AIEMQRQTD

31210
AIFEPRRWH

31211
AIQLHFEWG

31212
AKGNTNNNA

31213
ALVHKDLYG

31214
AMMELMWTD

31215
ANHHCGLAC

31216
ANHLFVIEH

31217
APDDENCDF

31218
AQATESSQP

31219
AQVEYTYET

31220
AQYPTLEYA

31221
ASFFSSVAQ

31222
ATDALSTSL

31223
ATDNHWGHG

31224
ATENVMWTC

31225
ATGLHITTC

31226
ATHERDFFV

31227
ATHEVITPA

31228
ATNWGPMSP

31229
ATQEFLGLN

31230
ATVATYINT

31231
AVGEVDAYS

31232
AVIYSSYNW

31233
AVMYYGYES

31234
AVWIEPRQW

31235
AVYLNEFNM

31236
AWFCPLLYN

31237
AWSARSVNM

31238
AYGWENQFE

31239
AYHELTSCA

31240
CADSKMTST

31241
CAHFAMNNH

31242
CAVCPALSA

31243
CCRHWRPTP

31244
CEHDMCQPT

31245
CEQVQIGDE

31246
CFICCHCFQ

31247
CGEHDKFKS

31248
CGGCEDWPM

31249
CHMYHRREG

31250
CHPHDEHKT

31251
CHQENFVFR

31252
CIASFTDGK

31253
CKVNESAKN

31254
CNPRPQMVG

31255
CNTKFLSQS

31256
CQKTFELHG

31257
CQMPERWTN

31258
CSGRGEIEF

31259
CTCDYTAQY

31260
CTCEGGSTA

31261
CVLGHIHHG

31262
CVMQRAADL

31263
CVQPHVDPM

31264
CVTCTNVYG

31265
CWSQEANRG

31266
CWTDNDDMD

31267
CYGHLNAMC

31268
DAEHSPNIM

31269
DAKHHQMVH

31270
DAKPYQRNI

31271
DATATGQWS

31272
DAWTMASAM

31273
DCEEFDRTP

31274
DCKWEHHSD

31275
DCPQDEQMN

31276
DDEPKTDWC

31277
DDMWQRDWF

31278
DDPAGHCYV

31279
DEHKRLDIC

31280
DEVPNKCDC

31281
DFKPKQHDH

31282
DFNWQDSGY

31283
DFSSINCQS

31284
DGMCNGGHP

31285
DGVSDHQRP

31286
DHTDMLCFG

31287
DIFPNCKTD

31288
DIFPSMNMS

31289
DIFQLTDDS

31290
DIHEWQVMS

31291
DIHGSQEFA

31292
DINENMSVE

31293
DIVCKFASP

31294
DLWRTFEAQ

31295
DNHDCGRSH

31296
DNHGYDRHY

31297
DNRCCWHMN

31298
DNRWWVAFS

31299
DNVIMTTVG

31300
DPSNCMRIM

31301
DQEVCTAMD

31302
DQLNVNESG

31303
DQNIFLSID

31304
DRSDGGFVG

31305
DSGEGLHNK

31306
DSLLMREAY

31307
DSRRIYAWI

31308
DSYCVNHEH

31309
DTAYVWCEM

31310
DTNRCMMQS

31311
DTQLDDIAG

31312
DTYPMQSAD

31313
DVDDLVEIM

31314
DVEDHDSQQ

31315
DVIANIHSR

31316
DVMHNHVKA

31317
DYRQYLEVQ

31318
EADKQYDDQ

31319
EAGMKQLCA

31320
EAEQFPSRG

31321
EAQDKWAGA

31322
EAQSIACPT

31323
EAYMMVCSH

31324
ECLFLRNVE

31325
ECNRMAMAQ

31326
ECNVNPACE

31327
ECSDWTHMQ

31328
ECWRHVQWN

31329
ECYSIYCEE

31330
EDNIRIMMI

31331
EDYHYTHTE

31332
EEAYVNCSV

31333
EFDEFNTRG

31334
EFESGLLDH

31335
EFHSYCMSF

31336
EFIHIDREC

31337
EFKCVDATE

31338
EFMEAHCGC

31339
EFMPEISTG

31340
EFRHLKHGG

31341
EGLENNDMF

31342
EHAGCQLWS

31343
EHEQLPSCY

31344
EFIFFDDNG

31345
EHKLVMCFP

31346
EHKWYTTLG

31347
EIHHFEYWS

31348
EIIHVVGND

31349
EILKNYTEA

31350
EIRPQDTEM

31351
EKAKRISGW

31352
EKPMMQNEC

31353
EKYSIIAHS

31354
ELAYLWPPH

31355
ELHSVNYML

31356
EMCVQHNTS

31357
EMEAKAFQD

31358
EMFYCGAYH

31359
EMMASAQSN

31360
EMMPLHQQP

31361
ENFSFYKCT

31362
ENIHYACSN

31363
ENLVINHIS

31364
ENNKWYSAW

31365
ENRCFSDEQ

31366
EPHWFFAES

31367
EPQATQKAI

31368
EQRIVQQAT

31369
ERCESHRAS

31370
ERQTCWRGS

31371
ESADMACSQ

31372
ESHSYVIAA

31373
ESRCVLEQG

31374
ESTNSHWGA

31375
ESVDVGLMH

31376
ETEPHTFFS

31377
ETGIDNNAT

31378
ETNLECGQK

31379
ETQYCRGQQ

31380
EVAHKLKHA

31381
EVDEVSPSA

31382
EVGTNPSKN

31383
EVRVDPCCL

31384
EVSHDAQCT

31385
EVTEVQTKA

31386
EWAMWKPYA

31387
EWSDEHLKA

31388
EWVREDGLP

31389
EYGVVKTPP

31390
EYMNYEHES

31391
EYSQNHSEV

31392
FAAHYGQND

31393
FAAPNSYWP

31394
FAKTDVWCG

31395
FANQGAIHP

31396
FCPEHWFNI

31397
FDMFASPDD

31398
FDNSCPKRW

31399
FFALNPNYP

31400
FFELSQTVF

31401
FGGKIDIGL

31402
FHCRGYLGQ

31403
FHHAMTEWQ

31404
FKPVNAPQL

31405
FLESWEKDQ

31406
FLPPNCIEW

31407
FLWLMQAIN

31408
FMEQYMMVD

31409
FMGQQNVAC

31410
FPQHMNVCV

31411
FRAQLPTCS

31412
FRGIETGQG

31413
FSGCYCMRI

31414
FYGQVTKSC

31415
FYQTIGQQE

31416
GAFMDMLIS

31417
GAIDYHTQM

31418
GAMLAQLSC

31419
GASWNDREH

31420
GEPELPNNA

31421
GEFIYQNRI

31422
GGARALRFN

31423
GGIQLDREW

31424
GHRKAFLAT

31425
GIFQYMSKH

31426
GIGPIMSAM

31427
GIIQVRMEW

31428
GINQEYTKC

31429
GITLTELQF

31430
GLRHDPAQF

31431
GMAYLQNEG

31432
GMQWMDRQA

31433
GMVQKQHGR

31434
GNVVWLARP

31435
GPDGAKRNK

31436
GPIGMDWED

31437
GPISGRYSV

31438
GPVVSPCKP

31439
GQFEDLLVL

31440
GQGLKAWPC

31441
GQVAFDKDY

31442
GRHDWMDKE

31443
GSLFSHFWV

31444
GSYNRLHDN

31445
GTDRNYAQC

31446
GTEQVSAEA

31447
GVHHNEHCR

31448
GVKFFEYDQ

31449
GVMVWNQLR

31450
GVYMEHNFE

31451
GVYSNCHDG

31452
GWYCMRQKI

31453
GYCGLMQYP

31454
GVTDDGVGY

31455
HADHAMAYF

31456
HASLQCMKT

31457
HCEGGQTIV

31458
HDLSYIDMA

31459
HEDRKHAGE

31460
HEDVGWCSR

31461
HEEFEYWNL

31462
HEHKRSSTE

31463
HEIDGWQMA

31464
HFDEITAGT

31465
HFERSPYWI

31466
HFQPLFEYN

31467
HGTEVCHWS

31468
HHEEHDIFA

31469
HHSFTIPYQ

31470
HILHLYICW

31471
HNFYPVTGG

31472
HPEEFRGFT

31473
HPGTYAIHA

31474
HPHAGGINK

31475
HPVWVRSDH

31476
HQETFFHGP

31477
HRDPHLLME

31478
HSFLDNNAD

31479
HSGMDHAQY

31480
HSMQGAPLY

31481
HSRWFEDYE

31482
HSTCPCVVR

31483
HSYPTAFYY

31484
HTGMEHMLY

31485
HTKQMDVDM

31486
HVIDHTGIL

31487
HVKINGTGK

31488
HWYMMEDYV

31489
HYFTKYKVD

31490
IAGLFLKTY

31491
IAMISLYNG

31492
IANCNTKHL

31493
IAVQQFYAA

31494
IAYQLTEIT

31495
ICFTPMHDS

31496
ICLPKMYFG

31497
IDIDMCYDY

31498
IDPPHKMLD

31499
IFGCTSAAE

31500
IGCHRMLNP

31501
IGNCDTFTK

31502
IGVVFQRDF

31503
IHHALQCHK

31504
IHVTSKTVK

31505
IIQGPVKAN

31506
ILMSMYING

31507
ILVNQQNGQ

31508
IMEITNVHE

31509
IMELTHHNA

31510
INEMARHPK

31511
INYTYLWAE

31512
IPGYAEFCE

31513
IPGYLEIHR

31514
IPTYRNRDH

31515
IQGVLPAMA

31516
IQMDFMLWR

31517
IRPTQRPCA

31518
ISGKSHGDR

31519
ISQWEHSCW

31520
ISTDCAEFK

31521
ITETEAALY

31522
ITKHHMRNC

31523
ITSEGFSFT

31524
IVANGHWLQ

31525
IVEQCQRMR

31526
IVFQSLARG

31527
IVTMGITEI

31528
IYEIMDYLA

31529
IYFMFYCSV

31530
IYPQCALQW

31531
KATYMEFDT

31532
KCEEFAHPQ

31533
KCFLCGSAT

31534
KCFWAELWA

31535
KEAVLNHWT

31536
KHELVTRMY

31537
KHKPRQSPR

31538
KHVCLVSWY

31539
KMEWLAAGP

31540
KMMAFSEVD

31541
KNNEPPMFF

31542
KSYCFMCPT

31543
KTCEMFTPG

31544
KTTILEDCC

31545
KTVEWWGRW

31546
KVVDCGRYQ

31547
KWADAEYGH

31548
KWCYHLFCQ

31549
LANVWVCME

31550
LAPEIECRH

31551
LATSKCSAM

31552
LATSMCYQA

31553
LATYGEWHN

31554
LCDYMHMGS

31555
LCEENSVDP

31556
LCFHYMLAQ

31557
LCWGPLHIS

31558
LCYFGSRWY

31559
LCYRTEKWD

31560
LDSQWMYYN

31561
LEEAVAAGN

31562
LESTSFCTY

31563
LEYDWNGWW

31564
LFCIVQSRQ

31565
LGWSNYECA

31566
LHIRYQSAP

31567
LHNSYWSLA

31568
LHWCSQHGH

31569
LIGFRTMFG

31570
LILGAKFMG

31571
LITWDGPTF

31572
LKFWASEKG

31573
LKYQFWTYD

31574
LMMPYMGTH

31575
LNITLVMGC

31576
LNPHVSQIW

31577
LPCKEVCCR

31578
LPGTPPDNC

31579
LPKWKEEIY

31580
LPPMKQDEE

31581
LQHHKYSSM

31582
LRCIDPPCQ

31583
LRNNNCSDI

31584
LRRWDDFQT

31585
LRSIEEKLS

31586
LRSLVAEDF

31587
LSDRCDVSG

31588
LSEVMMQPG

31589
LSHIKVEQD

31590
LSKNCEYWE

31591
LSTGNIQFM

31592
LTKPMPIIL

31593
LTMISMFSD

31594
LTRPRVQKL

31595
LTSRSDFDL

31596
LVDMCRKGK

31597
LVGQYWPRD

31598
LVGSYQHFQ

31599
LVTRGVSPL

31600
LWDQTQIAI

31601
LYAVENQHW

31602
LYCQQAANW

31603
LYDHYAAAS

31604
LYNWMKLIH

31605
MADLKWEDS

31606
MAFVSKYIM

31607
MAHDIPVPV

31608
MAHQLNHNS

31609
MATDISGWF

31610
MAWTHCHKP

31611
MCLCNMVIQ

31612
MCSARRQDT

31613
MCTQPSVKP

31614
MCYQNIAFP

31615
MDEQRQHEE

31616
MDFPSCEMC

31617
MDHQLCIQY

31618
MDPILRCLH

31619
MDPRAHGKV

31620
MEQWDPDEW

31621
MFELFQEGW

31622
MFGGFVKMK

31623
MFIKWRRYW

31624
MFMWWNYFP

31625
MFRISRFMA

31626
MGILKGEFS

31627
MGTSCCNYE

31628
MHLQKHIGC

31629
MHYLVMRDA

31630
MIDPKMGDR

31631
MIHKKDGMY

31632
MIMQCNCSM

31633
MKQLSNDGE

31634
MMDLAYHWQ

31635
MMEYTHSFE

31636
MNLPDPSVG

31637
MNVPWYKDR

31638
MPCFSVAAA

31639
MPHPEIRPG

31640
MQEPMRCAI

31641
MSFLFFVFM

31642
MSLLMGHES

31543
MTEVMREIR

31544
MTIILPMTS

31645
MTKPWSIIE

31646
MTKQTLCHL

31647
MTQLCEPWL

31648
MVIQAIWPV

31649
MVIQDGHEY

31650
MVMVMEAHN

31651
MVQWEWCDN

31652
MYLEKDLSG

31653
MYQLCHAFA

31654
NAEFDNNRG

31555
NAIPWINMD

31555
NAVAQIVSD

31557
NCCPGFAIA

31658
NCPWPTGEE

31659
NCRMHNAWF

31660
NDDRCSIEL

31651
NEQWWVWYR

31652
NETMMWRAN

31653
NFHYIGTLG

31664
NFIFMQDMT

31565
NFMDYHDMV

31666
NGMLETDHV

31567
NIYMPHTCL

31568
NMATANTCV

31569
NMDGKNCRT

31670
NNEMPGWRA

31671
NNMPMDAFH

31672
NNMSAHHSK

31673
NNYHYPHIG

31674
NNYLELHRY

31675
NPSSHNQLN

31676
NQNIWKHNV

31677
NQRPYDQWA

31678
NRGCYEMEN

31679
NRTFKAECH

31680
NSIFCLENQ

31681
NSNQDQWRI

31682
NTCCWKLEN

31683
NTVERYRQS

31684
NVCLEPMDP

31685
NVEWQVRGP

31686
NVQWADIGG

31687
NYKQPVQAP

31688
NYNETKFFG

31689
NYWQAQEFY

31690
PAEVWTHSE

31691
PCCQEHCQR

31692
PCNFDNQPH

31693
PDAAYWFDG

31694
PDCPYKCYC

31695
PDGEKCERF

31696
PDNYIQNHH

31697
PDQWDFEQY

31698
PEFMTEMCT

31699
PFHMALAEE

31700
PFYHMCAAP

31701
PGCMIVSQN

31702
PGELGDENF

31703
PHNWRSVSW

31704
PMMQKLCCG

31705
PNNEGGMKP

31706
PPGSHKGEN

31707
PPPDRTFAA

31708
PQEFREMCP

31709
PQLKEGQLY

31710
PQNGMLHYS

31711
PRAENDESR

31712
PTETDRFFQ

31713
PTTNWHYVQ

31714
PTYHWDSKT

31715
PVEHVEKEM

31716
PWHEYAGFT

31717
PWLAEPRGG

31718
PWQSGSDGD

31719
PYASQECIS

31720
PYPAIELGK

31721
PYSPHKQSA

31722
QANVNGHDF

31723
QCAFIPNSV

31724
QCEHDRQIM

31725
QCPVECEHV

31726
QCYRKVVFR

31727
QDIWPVCRE

31728
QEDDFRNKS

31729
QEKIAMRTN

31730
QFTSQNEGP

31731
QFVGRERQV

31732
QGFYTHTGY

31733
QHMPRHIWH

31734
QHWQFCTNA

31735
QIADQTGIV

31736
QIGPTISEY

31737
QIMNYCYFN

31738
QISCLEHGL

31739
QISYHIQHH

31740
QITQHPCEQ

31741
QITSMKCAN

31742
QIVQSRSAV

31743
QIYNWEKMT

31744
QKGNRTNQI

31745
QLIVCDTGH

31746
QMGSRMEMC

31747
QMHSTFCGW

31748
QMPIGGCSE

31749
QNSHKHWDH

31750
QPCVDWPYF

31751
QQAPGQMFY

31752
QQDRLWTTG

31753
QQKDQAWRH

31754
QQMIRQLSE

31755
QQMWGCACC

31756
QQNVLEHHF

31757
QQSPVEQCY

31758
QRQNVQQTW

31759
QSDAELKYY

31760
QSDLQHWWY

31761
QSKELSCMK

31762
QSKSGQHIA

31763
QTCTSQDYS

31764
QTLESMIYH

31765
QTMEYEIGG

31766
QTTQTDCTK

31767
QTVEMSATV

31768
QVDEWGGHE

31769
QVDNREGDV

31770
QVKCMQMWP

31771
QVVHNEFIH

31772
QVWITDAPI

31773
QVWITAKEE

31774
QYDAFAVLA

31775
QYMDQDVAI

31776
RADEDMTSI

31777
RADHWGSWP

31778
RENIHCEGE

31779
RHELTMSHY

31780
RHEPLHYFH

31781
RHWLNGCSN

31782
RIDINVDCY

31783
RIELMACTT

31784
RIKYEHCQL

31785
RIMYFDKFT

31786
RITYFDDQC

31787
RNGFRWTVL

31788
RPEAYWVPV

31789
RRDSDSVFK

31790
RRFCISLKC

31791
RSAEGISHW

31792
RSHHHQRMM

31793
RTIEEENTS

31794
RTVEIESWV

31795
RVDDENGGN

31796
RVDHDMADG

31797
RVESQACFQ

31798
RVSVDESDK

31799
RWEQMAYSW

31800
SAGVVVQSI

31801
SATMVADKE

31802
SAYMALHPL

31803
SCEWPKQCK

31804
SCTHWNTWR

31805
SDAPEHDGQ

31806
SDLKEETAY

31807
SDQWYRVSF

31808
SHMVVTSQG

31809
SHVESHQWC

31810
SIDVTSCAF

31811
SIHPLNQEE

31812
SIRVYPELS

31813
SISPERDMD

31814
SIVSEVCPN

31815
SKMAGFPHW

31816
SKWTCHPKM

31817
SLKMVDVAV

31818
SMFKYEARD

31819
SMHEKQHFE

31820
SNLDFPNQE

31821
SNPIMAKGV

31822
SNPQSVCRG

31823
SNPRTQTMV

31824
SNSPHPAVF

31825
SNVILDFEN

31826
SNVVKQASD

31827
SNYRALYGH

31828
SRDGWPNCI

31829
SREPIIKGH

31830
SSGGIWYFD

31831
SSMEMWTCN

31832
SSRFAPPPE

31833
SSRPPSGWV

31834
SSSQCCTPI

31835
STHHFSYGE

31836
SVEAGEKEG

31837
SVKEEMFSP

31838
SVLESDECK

31839
SVNDTVDYF

31840
SVWDMVKMQ

31841
SWCPEYWEI

31842
SWHYLAMQC

31843
SYHAHQPHC

31844
SYKQVTAPR

31845
SYLSFGAPW

31846
TAAEQKADN

31847
TAHCTDWQH

31848
TASMMECAD

31849
TATTHQRCP

31850
TCPNVWWHM

31851
TCVIRNMMI

31852
TEAPWWSGH

31853
TEEVDSMAC

31854
TEGNWDSCW

31855
TEKAIVYYP

31856
TFGYMATQV

31857
TGFHNTFRD

31858
THCHRHHID

31859
THEHSGVYG

31860
THKWDADAK

31861
TIATVRTCT

31862
TIDLVYLDK

31863
TIIPIDWTE

31864
TILPTCKQA

31865
TIPCYQRYT

31866
TISDEPGQP

31867
TKFMPRGYI

31868
TLFYHDWYM

31869
TLSSHWKER

31870
TMTKRDDFW

31871
TNTAFMNGT

31872
TNTSMPAAW

31873
TNWREMNPR

31874
TPWFWQETA

31875
TQTELYCCP

31876
TRHCHECLG

31877
TSDEAEHYT

31878
TSFMCQKFP

31879
TSGPCMPIK

31880
TSQHAIWNA

31881
TSTLVSNNV

31882
TTEFVRIKG

31883
TTNDDNCNN

31884
TISEKNFDD

31885
TTTRDEAFM

31886
TVCTTKSMG

31887
TVRCMIQLG

31888
TVSDDWCWE

31889
TWETTGQFV

31890
TWMFVHRNN

31891
TWSDNHAAQ

31892
TWTNKLCDM

31893
TYEDHVDRF

31894
TYEWSMLDR

31895
TYQNELACY

31896
VATMAHYGY

31897
VCIGYDHTP

31898
VCPLPNQLH

31899
VCSOTTLWH

31900
VDKVWWNLI

31901
VETCRDCEG

31902
VFGEFDNHS

31903
VFHFQTKGM

31904
VFKFDFSYP

31905
VGDPNFRIQ

31906
VHAQTIQRE

31907
VHHTEPERQ

31908
VHWPQCGHQ

31909
VIHAPTHKT

31910
VIPPSIFNN

31911
VEYHTSYYW

31912
VKECRRTDQ

31913
VKSEWYDEQ

31914
VLGDWSNIC

31915
VLIPILVAQ

31916
VLVNVDQLG

31917
VMPNWKEQT

31918
VMSCYSRDH

31919
VMTFRQKAC

31920
VPGPMIYEF

31921
VPKTVNRDM

31922
VQSVRRITG

31923
VRMICTSSC

31924
VRVWDYKGT

31925
VSALQQTQQ

31926
VSCHWGLAN

31927
VSRLRQNHW

31928
VTDHAWTMP

31929
VTMLMAGMQ

31930
VTSYWEFQE

31931
VTYHHHNFL

31932
VVNFVGAEG

31933
VVWMNSNHD

31934
VWFYPQWDS

31935
VWSCVHADV

31936
VYWFPDQMM

31937
WCNGHGSIF

31938
WDLYLKTMG

31939
WEAGECDLM

31940
WEKFWHEYW

31941
WEWPMTWTT

31942
WHIFVVIGI

31943
WIPHCKCPA

31944
WKAHTEHLG

31945
WKCWYAPLT

31946
WLIWGIIEQ

31947
WMCRNHPFD

31948
WQMTTRYYM

31949
WSSTQDQTN

31950
WTVGDEQPM

31951
WVEPQTTIY

31952
WVEPSSKPP

31953
WWQTFCYGE

31954
YADMDQAVQ

31955
YAMSRKQTI

31956
YATVEEDLQ

31957
YEHWYEHDD

31958
YERLFDEQT

31959
YFRDQEEGQ

31960
YFSLLMENC

31961
YHSDYHHNN

31962
YIEPNHHSG

31963
YIQRGTDSM

31964
YLVRQWAIM

31965
YMSKHEADI

31966
YNNNHLMMC

31967
YQQQMTFHN

31968
YQYFPDNKK

31969
YRHGSVDGP

31970
YSCNRLTRF

31971
YTQLNASQW

31972
YTTFQGYWA

31973
YVAEPYYHG

31974
YVCNICPTP

31975
YVGQTSSSE

31976
YVGTIPTFS

31977
AAEENEAAD

31978
AAGPDALRP

31979
AAGTVCLPM

31980
AAGTVEFGH

31981
AAGWGGATS

31982
AAMEVMVKC

31983
AAMHESCNW

31984
AAPCCLYYT

31985
ACAWIENFT

31986
ACHATTLRY

31987
ACKQDSELV

31988
ACMSRAMFN

31989
ACTRPMVLA

31990
ACVEHYVAQ

TABLE 82

Sequences of the 581 to 589 Region in AAV5 VP1

Capsid Polypeptide that Drive Skin Tissue

Tropism

SEQ

ID
581-589

NO
Sequence

33991
NMWYSFVMI

33992
FWLGISPAK

33993
DSQYHVKCG

33994
AIEWKSTVT

33995
AIEWRSTVT

33996
ANAHQAESF

33997
ANAHQAGSF

33998
ANLEQSLMA

33999
ANLEQSLMT

34000
ANLKQSLMA

34001
ANTHQAESF

34002
ATKNRWPIA

34003
ATKNRWPIV

34004
CCHGWHNGA

34005
CCHGWHNGS

34006
CCHGWYNGS

34007
CECEKLGTS

34008
CEGEKLGTS

34009
CEYEKLGTS

34010
CFHGWHNGS

34011
CIALLEQYM

34012
CNATYQQND

34013
CSCQWYISV

34014
CSCRWYISV

34015
CSYRWYISV

34016
CYLWCAYQH

34017
CYLWCDYQH

34018
CYLWCDYQR

34019
CYLWCDYRH

34020
CYLWCVYQH

34021
CYLWYDYQH

34022
DHPPVKEVS

34023
DNLNFNSPT

34024
DRPPVKEVS

34025
DTYQNYMFM

34026
DVHPSNNVQ

34027
DVHPSNTVQ

34028
EGPWMFAQG

34029
EGPWMFTQG

34030
EYQCWGRFT

34031
EYQCWGWFT

34032
GYELQLQMN

34033
HMYDNELNP

34034
IAEPQWHKQ

34035
ITEPQWHKQ

34036
MCFMAFRFD

34037
MDYCCEHFP

34038
MHYCCEHFP

34039
MLFLKRNIA

34040
MRFMAFRFD

34041
MRFMAFRFG

34042
MRFMAFWFD

34043
PTPPMQHTA

34044
QAAHEPQIC

34045
RHIVHKRTE

34046
RMYDNELNP

34047
RRIVHKRTA

34048
RRIVHKRTE

34049
RRIVHKRTG

34050
RSKCPVEIH

34051
RSKCPVEIR

34052
TIEWKSTVT

34053
TNAHQAESF

34054
TNLEQSLMA

34055
TTHPMQHTA

34056
TTPPMQHTA

34057
WCHGWHNGS

34058
WPAQGRITT

34059
YSCRWYISV

34060
AAVQMSSLS

34061
ACANTDQRG

34062
ACMSILEKE

34063
AEHQNNCAF

34064
AELPEGWMC

34065
AGFYEHRSN

34066
AHGCTNAMS

34067
AIRISSLVT

34068
AIVHHGKEP

34069
AKNYQFQVG

34070
ALDSCMVSE

34071
AMIHRNDHC

34072
ARGMCQDLQ

34073
ASEVHHAEF

34074
ASHCMILHT

34075
ASVDNSGSP

34076
ASWCLNHMA

34077
ATSTRQQHA

34078
AVDPAWHGC

34079
AVSAFQQDN

34080
AVSMKAFAI

34081
CAAHVVLTF

34082
CAKAKKMTA

34083
CAQQIYSQT

34084
CCARIAVAE

34085
CDTESLAML

34086
CEHCCITQF

34087
CEHCCITRF

34088
CFAFQNMDD

34089
CFQSNITAN

34090
CFRSADNWW

34091
CGAHIMSGH

34092
CIQWWEMWN

34093
CKQYIVTDV

34094
CLNRIQTAS

34095
CMMPWQSQS

34096
CMTLILAQE

34097
CMVWTNVSP

34098
CNRSIQCMV

34099
CPGRYWKVG

34100
CPKLIRQFA

34101
CPNVWARPW

34102
CQCRYFDWC

34103
CSAFTAHQS

34104
CTFDCNGYW

34105
CTFSTSEQP

34106
CTSWVWMQG

34107
CVAWHAHSA

34108
CVKQTFTDS

34109
CVLPLFQET

34110
CVMAGAIHR

34111
CVNPMAVAC

34112
CVQYTRIGR

34113
CVSPENGMM

34114
CVWLKQMYE

34115
CWMNTHIGE

34116
CWQYQMVTA

34117
DASPVSRSV

34118
DFVLRSDPA

34119
DGTHMDTWL

34120
DIVGPNTGF

34121
DKGFKTNSL

34122
DKHLFDMCR

34123
DLYTISTRD

34124
DMFMWNTQE

34125
DMQCYKRPG

34126
DMTNCQRCR

34127
DQRWENQHA

34128
DTYLNVMFE

34129
DVAKKRTAG

34130
DVCRKGEQF

34131
DVPLNECDT

34132
DWRRNTEKQ

34133
DYAVMKIAM

34134
EAEEGMGSW

34135
EARMWHHQW

34136
ECLESGIQT

34137
EGWLMMKTG

34138
EHIHGKFFL

34139
EIERDTSHE

34140
EIHTSEMDL

34141
EIKSWHQVY

34142
EKNSPWQFA

34143
ELCWHHGVA

34144
ENKHHEAFY

34145
ERMNMIRYA

34146
ESKIEEAME

34147
ESVHDNGGH

34148
ETHHNACSP

34149
ETHRMTWTT

34150
ETHTVDADM

34151
EVNIECCNG

34152
EVWTWQDAH

34153
FFIGRHSGG

34154
FPCMSWKQA

34155
FVFPKHVTL

34156
FVVPISAYQ

34157
GACNYMAHM

34158
GACVMFVGR

34159
GAQCFIMTP

34160
GHCACWVAG

34161
GHFLTTCSF

34162
GIGFVDMQL

34163
GIVDCIAAM

34164
GNDKFLVHN

34165
GQVATISQF

34166
GSSQIVKAE

34167
GTCPYRAIS

34168
GTFMSSRQS

34169
GTTAKTEQI

34170
GTVNFEKMS

34171
GVNDYRDAG

34172
GYAMKYLNA

34173
HAYWCQMFM

34174
HCDVIVGDW

34175
HCVPHVGYQ

34176
HFMCCWFKR

34177
HHAASATCF

34178
HIQGMNMWG

34179
HKMRAIWFA

34180
HMGMMQCQE

34181
HNMCPCVDY

34182
HSGPFCRHG

34183
HSSPFCRHG

34184
HSSSKEACF

34185
HTNWKDGYG

34186
HTYPAHWLQ

34187
HWCMLINVE

34188
IDIDRGHYG

34189
IFGCIQSAR

34190
IGFQYGSVE

34191
IGVSLLNDP

34192
IIKESVYCQ

34193
IISMTNRGP

34194
IMFYTNERV

34195
IMSKTPSFL

34196
INVEGGFEY

34197
IPMEWEHAG

34198
IRVHDNRQQ

34199
ISVDCMQRI

34200
ITAPKWQEK

34201
ITVEAVHHC

34202
IWDVRQDMD

34203
KFDTFIGYQ

34204
KFWCGHMEE

34205
KMTSVSWSV

34206
KNDLEAFSA

34207
KNGDTMAVG

34208
KPKCTYPLA

34209
KTDILDTIV

34210
KTQEFEVQP

34211
KVESTTQLC

34212
KWSYNHFRV

34213
LAKCGRWSV

34214
LATLLAQIY

34215
LCWFLLGVP

34216
LENCMWGGN

34217
LFWPDCHFP

34218
LGLQEWNTP

34219
LGMPCAWCP

34220
LIYPWKPGA

34221
LPEKWNIWK

34222
LQMNYKAQN

34223
LSAYLTEAG

34224
LSIPKEQCW

34225
LSMWRWESE

34226
LSNFFGAMQ

34227
LSSTFPHWH

34228
LTAKPCTAP

34229
LTGLQEMAA

34230
LVQFIYPFE

34231
LVQFTYPFE

34232
LWSLRSANF

34233
LYLMDIAMC

34234
LYLMDITMC

34235
LYNLFMHTE

34236
MAIVDPRSC

34237
MAQKKCKCP

34238
MAYCYLMHN

34239
MDGSYHWDQ

34240
MFMCYINDN

34241
MGVEVTQAM

34242
MIDRHWTAS

34243
MIRPSINGS

34244
MITLLPNWP

34245
MLFKAQAFF

34246
MLFMAQAFF

34247
MMCTFDFAG

34248
MMEIQESCE

34249
MPEHGMDNW

34250
MPPWYRHHL

34251
MQLEANKYH

34252
MRCIEVKGF

34253
MSDHLSNGA

34254
MTHDRAMQC

34255
MTQCNTEHL

34256
MTTVHWHSS

34257
MVLHEMFVY

34258
MVNPCLLGN

34259
MVTCAGPTV

34260
NAGSSGQGA

34261
NCKYRMELE

34262
NCVRYKNMD

34263
NFQMEMDAF

34264
NFSLTNNGQ

34265
NILVLCQNN

34266
NKLDQDNWK

34267
NKYCFPTSW

34268
NLMAHGVAP

34269
NMPNTHCNG

34270
NQVCWPRIV

34271
NRLSFMHYG

34272
NTEFSHLFP

34273
NVNVNKHNH

34274
NVNVNKHNR

34275
NVRDTMRGF

34276
NVRVEFIQC

34277
NVVHVGDVK

34278
PACNCPKTH

34279
PIWPGVLQH

34280
PTIKTQFSC

34281
PTMSLFNEQ

34282
QAKYVTESA

34283
QAMKHAFYQ

34284
QATIWNNRE

34285
QCCQQPCHN

34286
QCCQTLFQS

34287
QCFDNQDEH

34288
QEAGECKWC

34289
QHQCYYNQQ

34290
QIMCNTTGH

34291
QKSWIVGEK

34292
QPIFYDRSQ

34293
QQDGDQQPL

34294
QQKAISSLP

34295
QSNNALMGN

34296
QTFSFAPGY

34297
QVMDLVYPT

34298
RAEPFPEEK

34299
RCEHDKLCL

34300
RCWFSELHE

34301
REVYGLFMD

34302
RGLMCEMMN

34303
RHMFHEAGW

34304
RKPPFGPPY

34305
RLMSGAIVE

34306
RNKHDMIGC

34307
RRDCMDSAF

34308
RVAAPWLPE

34309
RVILAMGPY

34310
SAHQFQDYQ

34311
SAPQFQDYQ

34312
SDMPHPPLS

34313
SGCRYNQEQ

34314
SGLQLMWHH

34315
SHKFMDLMM

34316
SHQLIRREN

34317
SIRYDFYVP

34318
SLCSMLCTR

34319
SLCWMHAHH

34320
SMASVSEIK

34321
SMEQYWTSN

34322
SMGVPQAAN

34323
SMMPKCKAS

34324
SMQCEVVGP

34325
SMQCEVVGS

34326
SMQEYQKHI

34327
SMYQFADPM

34328
SQDIESLWP

34329
SRMQFHNQE

34330
SRWCPKVPF

34331
SSGEGKAED

34332
SSRPKYFCV

34333
STCHVGKMQ

34334
STCSIDAQM

34335
SVCWSNKVP

34336
SVDRQFVHS

34337
SVQLWGYQP

34338
SVRYECSHG

34339
SYQSCESMQ

34340
SYSWKQYGV

34341
TALGIYAHK

34342
TCIRAMTSC

34343
TCRPSDWAW

34344
TCYQNQRSH

34345
TEWEQNWTP

34346
TFGTTAIHN

34347
TGNASNDCW

34348
THFSKMSDQ

34349
TKCSWNHIS

34350
TLCIVPPLI

34351
TMSTCQQML

34352
TNQLYRGFW

34353
TSFMPPSEA

34354
TSGFFHMYY

34355
TSHCIPEVE

34356
TTIGWHVCQ

34357
TYCKSYRTR

34358
TYKCWIADN

34359
VAVDLSWCG

34360
VFASEWCGS

34361
VGTKHDVWY

34362
VKKRDNQST

34363
VKKRVNQST

34364
VMKHQHSGR

34365
VQGYKDSNE

34326
VSCYTHRQD

34367
VTFCVHRYQ

34368
VTTSRWELI

34369
VYILEGGIH

34370
WFRITDTCG

34371
WIITKSTEF

34372
WMTCFPMQN

34373
WRDFFNFSH

34374
WTKMQQNDT

34375
WVAVLNLNA

34376
WWQLMNLLP

34377
YCGIYWNSF

34378
YEHIDWTFC

34379
YLHKRCWPS

34380
YRLLTCASA

34381
YSALNSYAC

34382
YSQLFQMTN

34383
YTFWINNHA

34384
DTSPVEYAH

34385
GMVDDKWLY

34386
QKNVSTKYD

34387
YCHQEDRIT

34388
SQETFQKMD

34389
SRYVNNGTV

34390
DGDFLQNHH

34391
ICHTEVMFN

34392
QTANVKAMY

34393
SAICQNYDA

34394
MALTVYWKG

34395
DTSKTGKHV

34396
TQCKQLTVM

34397
KTMGIGGGP

34398
RVMMGNNQA

34399
MNSFYRAEW

34400
AAGHIGSVM

34401
AAILTDYDE

34402
AATGFTFFE

34403
ACDQQQRES

34404
AHAPQFGPC

34405
APEIHDYCA

34406
AYMWEEQAT

34407
CSMPSVNSD

34408
CSSCNCHCS

34409
CTKAYRKPG

34410
DCMLVPVSG

34411
DEPFPCDWS

34412
DGRQVWFFQ

34413
DHSQNTEPT

34414
DKFSLPSQD

34415
DNVTIKKVQ

34416
DQCPGHISV

34417
DYCNKKRPV

34418
EAYANMFHG

34419
EDEQHLWND

34420
EEGTVPEVL

34421
ELPRNYPDP

34422
EQMPLTNVP

34423
EQPSVNYLR

34424
EQTGDFVHH

34425
ERQARNHYA

34426
EVNCDWHWQ

34427
FGPCLMNER

34428
FGQLIIKHP

34429
FKMFNWPAY

34430
FSGVYKQYE

34431
GDSYYMWKA

34432
GERGHGERY

34433
GFHDCSMMI

34434
GIHEIVVNQ

34435
GKCIMQMCM

34436
GLSMVMHFV

34437
GSMQHRRDC

34438
GSMRHQRDC

34439
HGYEHMMSG

34440
HTERWHMLG

34441
HTHDWYSAG

34442
HVYEHMMSG

34443
IAGISMHNH

34444
IDDPTMFEV

34445
IHWHCWPKW

34446
IIQQYGCST

34447
IMQPPWHTG

34448
KEPMPRGFL

34449
KICHEDISV

34450
KQALYDTFK

34451
KQMPLTNVP

34452
LAYPHERDG

34453
LHYDTMKNE

34454
LLANYMVHT

34455
LPIEQVKWF

34456
LWSWYDQPS

34457
MCWSWQDAY

34458
MQTPQAVMS

34459
MSMKYLEQT

34460
MTFQDAATY

34461
MTIDIDQWC

34462
MTVIWIWSH

34463
NNVEQHCQV

34464
PEYQFECIF

34465
PGWSVANCS

34466
PKVNLKGDE

34467
PQVMWPGWF

34468
PSTHHTQGM

34469
PVEELYGTF

34470
QFFACGVYD

34471
QPTSHMNMN

34472
QTIWFIDDN

34473
FDLMYTDQA

34474
RIEWEMISD

34475
SCATAHARE

34476
SHEKYWYFS

34477
SKCIMQMCM

34478
SMFMGRRTE

34479
SPYSCWMQE

34480
SRYLYYNYH

34481
SSYYFMGTP

34482
SVNRMDQDH

34483
TCELTMCYL

34484
THSSCWYQA

34485
TIRPVMWGH

34486
TKPPLGCSS

34487
TQTNKTFMK

34488
TVCEPKCAA

34489
TVICIPSGS

34490
VIWYCFCTL

34491
VMGIVEPSK

34492
VMWNQNKTE

34493
VVHNVKDVH

34494
WTSILDHSS

34495
WVASSYKPG

34496
WYQPRFGRQ

34497
YDAVWRKYT

34498
YDEHWNDNS

34499
YDIMTISPF

34500
YHKDYMCLG

34501
YTGRFTCMS

34502
AATTYNQHA

34503
ACEQQQRES

34504
AGDWYIALI

34505
AGHLQFTCP

34506
AGVRTQCDN

34507
ALCSCNQLF

34508
AMRKIGVWH

34509
AQGHKCYCS

34510
AQTNKTFMK

34511
ASMPVWKHY

34512
ATIMMHQFG

34513
AYVEHNASR

34514
CKTHCPGQY

34515
CNHQFIDQA

34516
CSCSYQDMM

34517
CVNMEMMTA

34518
CYEQVNSND

34519
DCADMGYRR

34520
DCEWMIRLG

34521
DDEYPAHWD

34522
DHLPMNQER

34523
DHQWTDAWV

34524
DIGYRSTTF

34525
DRAYRPWND

34526
DSKSYWMDH

34527
DVGCLQNMY

34528
EAYTNMFHG

34529
ECRMYWFED

34530
EMMHKMDAG

34531
ESRQVHVGN

34532
ESSYVNGIG

34533
FASYDNTHL

34534
FGEDGGFKG

34535
FGPRENNHN

34536
FHGNYQHDS

34537
FLCLPQGHI

34538
FQKVVEWQA

34539
FQRPNFSLT

34540
FQSLNKNQS

34541
FVTQQHSHE

34542
FWNHHDVAR

34543
GLCSLNNMP

34544
GSMQHQRDC

34545
HEWWGPVSH

34546
HKELILQPT

34547
HVQHWMNDR

34548
IADIDLTSF

34549
ICQPDMKQS

34550
IPKYKQFFD

34551
ISAQCPHVH

34552
ISHWQQRAE

34553
ISKAQQHNP

34554
KHDLTQQRQ

34555
KHVITGGWW

34556
KKPWDDHTR

34557
KLESMAMSM

34558
KLVCVDAYQ

34559
KMNTGDNYH

34560
KQPTPVNGI

34561
KSARCKLHM

34562
KSLSGEKNY

34563
LDMWGDFLL

34564
LDQSMDIAP

34565
LQSHSQCFR

34566
LTWGANAYT

34567
LVCFADIMQ

34568
LYRLVAQMN

34569
MDQLVINSK

34570
MIKTVAKKE

34571
MILYFWKSH

34572
MLAPIDCNT

34573
MLGPLWKDQ

34574
MNTRKDPPL

34575
MTGWSSKQS

34576
MTYQHVAGA

34577
MVCGNQRNN

34578
MVLADNEFI

34579
NNDRWAIPQ

34580
NPGWYGLAP

34581
NQKGHDYSQ

34582
NQVEVIYQS

34583
NTSYVTMCT

34584
PAFVCESPQ

34585
PKPPLGCSS

34586
QCEMKHHLA

34587
QDPHKCRRS

34588
QPAAKNKWN

34589
QQNTEVKWE

34590
QVVVRVQDH

34591
SCEHCNKPY

34592
SCHKYTDGN

34593
SDPDIACVH

34594
SFCTMQHSV

34595
SHQWSRQMM

34596
SNTYAGFQW

34597
SRMGLELVN

34598
SSMIMCGYV

34599
TGTNLHQCQ

34600
TMHIKRNSH

34601
TMQCISHLS

34602
TMRGSNHEP

34603
TSMKTHFDR

34604
TSNHENRAD

34605
TWNEPHQNQ

34606
VDDQHPTQK

34607
VIFMPIVHC

34608
VLVDHPVNE

34609
WYQSYLNWS

34610
YAQHEFRLD

34611
YERQMATPG

34612
YHPLQSDFG

34613
YPHHEQHAT

34614
ATQFYQACI

34615
GFHDRSMMI

34616
NCWVNEKCE

34617
QGSKQMYQA

34618
RTHGHAKAD

34619
TSKSISHMD

34620
YTQFCAAMW

34621
AQDINRLQA

34622
IAQSEADYW

34623
DAVIHTGFA

34624
DHLNEEQTK

34625
LHQFEAYYK

34626
KQQMMIVGN

34627
YKTSWGQHV

34628
VDNTDPWIM

34629
ANIPIEMVC

34630
TGNHEDPDN

34631
MMAMKQGFY

34632
ALTHDDGRT

34633
AEHCYRHQH

34634
IEFGVKISG

34635
KELLCSHDA

34636
LGDQKLVDE

34637
HCTETHHTN

34638
TVGFRFDRA

34639
GGAEIVSAD

34640
YIQFEAYAN

34641
KVHFAKVRI

34642
LIAYKSHAH

34643
SPQQPSVFH

34644
PAEAKQRFD

34645
MKVSNWVFP

34646
TIAFNGAVC

34647
MNTNVMQKD

34648
PANHARNPK

34649
YYTKHEQDP

34650
RQAWPKDGM

34651
QTAEWYCGA

34652
THVKRFDYE

34653
EISICTAKF

34654
CSECQAIHS

34655
KERMQVSKA

34656
FHNAPEHHE

34657
EPRLYCAQE

34658
DIEFLDCEN

34659
DATKEMSWK

34660
QQAMWLHSS

34661
CADPQERCD

34662
TTYPHNTHG

34663
AVNQWMHSL

34664
CTNYHVKAG

34665
WPNIKHQPI

34666
KNCIMLASN

34667
PLTIEVNCT

34668
ATGTYNLQE

34669
WATWPEPLQ

34670
YVDHWTLGD

34671
GLFEGSPGA

34672
SVLHVQALA

34673
NGVGRSGDN

34674
MNLQIGSKG

34675
NWTGNMQWA

34676
EVRIQCKID

34677
SNVACIQAF

34678
ATMNLSWGL

34679
QSAHPEPMC

34680
DHSQNPEPT

34681
EECPNCEAP

34682
GWNYMFGSA

34683
HHTMPIDWN

34684
GTMPMTGQC

34685
AQAAAVICW

34686
DQLAHNAQE

34687
INMPCPNTK

34688
RMWMEQYQA

34689
IMQMHPASG

34690
IPSHMDAHQ

34691
HTDSDCRHS

34692
SYLPRHHKG

34693
ASDCDGKTW

34694
QIIMLQHEN

34695
EICHEDISV

34696
KTTPAPDQG

34697
SWYLYYNYH

34698
LSIRNGMCD

34699
QFMYCAKGN

34700
HSSCKNCPW

34701
DGDYACFKN

34702
QFLGGPKIQ

34703
RAVHARQYQ

34704
EHTQRMDMA

34705
IVTMQMDSW

34706
QEINDLKMH

34707
YTYLHQSSG

34708
DDRRNTKWE

34709
VCCSIGPEH

34710
ALKSFNWCS

34711
TNDWGEKNA

34712
CWPEAMIDD

34713
GAETEPKDS

34714
CPTYKNKDQ

34715
HVGAGGMPG

34716
VTHMTCVEG

34717
AYMWEGQAT

34718
KCGNVLCEV

34719
EHDWRMFHQ

34720
GPKEHNLCD

34721
ANSLEGGFE

34722
YLHERTDKD

34723
VAEAGYCYK

34724
KECCFPPCE

34725
IGEVFDAFS

34726
AFDGEMSWE

34727
IEMPTESKH

34728
YRLHCCNRA

34729
DAMAYGQFM

34730
ITELMGCLH

34731
RGMAHHHLC

34732
TGGWQRGAF

34733
FCWSMCEAC

34734
FCIKDQHVR

34735
PVTHPQTSK

34736
RSGVQCCAE

34737
IWQQCNQKN

34738
KQSDKWGDD

34739
MIVHQVSQI

34740
TSNKKGEAA

34741
TMFYCHGPN

34742
LCMNDDRLN

34743
DCLKKNHSL

34744
RMDCKWEDL

34745
TGWNFPVSC

34746
GSQQMQASQ

34747
NSKACRGCT

34748
QCLWVDSNV

34749
EFCHQHSAI

34750
HQNLLICNE

34751
FAGFTPVWM

34752
NSPHSGVGD

34753
DKTCYLTCP

34754
CHGFSGCQT

34755
NSRLLIKHL

34756
NCKPDQIAM

34757
TAMVSECKD

34758
YDYYKMREG

34759
SAPEFSVIC

34760
REAWGIRDS

34761
KIETRPTCT

34762
HCTEGYRQT

34763
MCLNMAKQG

34764
QNPNCKVMW

34765
YFHWHDCIG

34766
KYINFDHHQ

34767
SFFHQTINP

34768
KAGYQEGSC

34769
IACISMHNH

34770
QPGVRTDDY

34771
QCQNWRHLS

34772
LIEMCCSSL

34773
PTEFRVCPV

34774
DVCNQSGMS

34775
RYEISERRC

34776
NCERWFQIE

34777
DRREKQLMK

34778
ENQARCIFE

34779
ESTHMLYEM

34780
TWHQGWAEY

34781
EGGFRDFCE

34782
AFARQEYAN

34783
ASSFCKFAS

34784
ASSRFGSYG

34785
CHQPKNSDR

34786
CIAKQIARC

34787
CMYHEQPGR

34788
CPQCDVMTS

34789
CSHETGAMC

34790
CSNCRQIMG

34791
CSVCLKYEQ

34792
CTLKTQAET

34793
CVKFSHHHA

34794
DANHCMWWQ

34795
DCVKSSDGI

34796
DGCPMTRDL

34797
DGLFEGHIP

34798
DIKWQHGAH

34799
DMYDHEWLM

34800
DPKCFWVSY

34801
DWIVMSQKA

34802
EFNWLLWSG

34803
EIAHDTSMS

34804
EIGHKQMAI

34805
EIVQPQYYA

34806
EMHKVPSGM

34807
EVTAVEHIW

34808
EWTKKTADH

34809
FLHISSWYQ

34810
FNMNTNDSG

34811
GCKKSEWIT

34812
GGSMLGNGY

34813
GNKLKNDQM

34814
HCLAGESYV

34815
HFHEVAWGS

34816
HPETMIYGW

34817
ICNAMLGSE

34818
IFNHLLPTE

34819
IRVMFGKHN

34820
KAIWQGTEN

34821
KFICTTIAC

34822
KICPTEYAS

34823
KIVNQENMK

34824
KNTMCDNDA

34825
KSDKVWYVL

34826
KTQTHTSEP

34827
KWEQRSKMS

34828
LMAQWEQTG

34829
LMAYCYANN

34830
LMHGVMDHV

34831
LTQKNLSWQ

34832
LWPQVMEQR

34833
MAGHYFNSR

34834
MAIDNLECK

34835
MAIISQIEF

34836
MATISQIEF

34837
MIHTPDAGC

34838
MIKFPAKQQ

34839
MMGSNHCSS

34840
MNSFYRAEL

34841
MSPFFWEQG

34842
MTSTQIRAG

34843
MYMEQACYD

34844
NADEKYNNT

34845
NAPMMACQI

34846
NGLFEGHIP

34847
NWDRGDKCI

34848
QCMVMTEAD

34849
QDFRMNAGA

34850
QDFWMNAGA

34851
QILPHMYAY

34852
QNNVITVDW

34853
QPFLHRPCA

34854
QPFLHRPCV

34855
QQPRKINLC

34856
QTMCITIHP

34857
QWDEFKMRM

34858
QYQTHPMMH

34859
QTWWPATHD

34860
RCEPFNWRE

34861
REVHVNQNN

34862
RGSQQMETH

34863
RKKVEVVVE

34864
RLKLSAVQR

34865
RSTWGHIQR

34866
RVNWEQRMN

34867
RYQPFSRQW

34868
SALAHHANF

34869
SALAHRANF

34870
SAMCSGSLL

34871
SANQAHGEF

34872
SANQRYHDW

34873
SAVQEERYN

34874
SAVQKERYN

34875
SDHQYRGCL

34876
SMTTWWMFP

34877
SNSHPFQSH

34878
SWRWHHWHW

34879
SYVMNLLGQ

34880
TAHIMTDVI

34881
TIDGMMGDH

34882
TLPKMHMWP

34883
TQQYTWRDA

34884
TSYLRKIHS

34885
TTAGKFAAQ

34886
TWARYKVGE

34887
TYDTFGCME

34888
TYRTRHTDC

34889
VKHLFYWLP

34890
VLKSCDRHG

34891
VVKHGVTTN

34892
WGATSEFMG

34893
WHCVAPNSF

34894
WNDLVRCEE

34895
WWQPVEYAW

34896
YHAPMLMCN

34897
YLAMMMSGD

34898
YLAMTMSGD

34899
YQNHYQKKW

34900
YSVVMRTSY

34901
YWDACYDQK

34902
DADCYVPAD

34903
EGKVPGQFA

34904
ACDLVKKDQ

34905
ACEAYQDEK

34906
AFSQQDYSF

34907
AGQLNFCSI

34908
AHNVRCATA

34909
ALCWKQMDH

34910
AMCWYYHMQ

34911
ANVISPQSN

34912
ASCNTHHEV

34913
ASGTKYRCN

34914
ASTMFIDVE

34915
ATHNVHMFH

34916
ATMAYRHVN

34917
AVDCSDHSY

34918
AVEQWVNHD

34919
AVVIAQKVV

34920
AWELVVHQA

34921
AWNFIRVGN

34922
CAANVTKMG

34923
CAAPTFADK

34924
CAEHLTPSE

34925
CAGLLGFHW

34926
CAGVQTTWW

34927
CAHYFAHAG

34928
CAKWFDVQI

34929
CAQSWCSFE

34930
CCFLINWDR

34931
CCGWYHEFG

34932
CCMEVFMYN

34933
CFTMPVHWG

34934
CHIQLHAGI

34935
CIHYKEQYA

34936
CIHYKEQYE

34937
CIIHQSKYG

34938
CIMSRQQDC

34939
CIRRDDHHH

34940
CIRWFDWLG

34941
CIWYTQMGT

34942
CKETVETVY

34943
CKMQLFSHG

34944
CLGWCTACP

34945
CLTALMKLS

34946
CLVRYFRDG

34947
CMTKTSKMS

34948
CQCSIMSIE

34949
CRGHIWSMD

34950
CSACIASAT

34951
CSLKGPPTH

34952
CSSSLFSAC

34953
CSTPITAAV

34954
CSTPITTAV

34955
CTCAGVVTC

34956
CTNLYAMAC

34957
CTQHMYVKD

34958
CTWKMMWGG

34959
CTWTTGEAS

34960
CVCLAEYVV

34961
CVFYLPHSP

34962
CVMSRWNYD

34963
CVQHFHWDK

34964
CVRFLNRVA

34965
CVTRCMARD

34966
CVVYHNHHT

34967
CWAQMNHVF

34968
CWMGIKRQD

34969
CWMLQHKMM

34970
CWSINYNYY

34971
CWTIRHGGP

34972
CWYLWYNMP

34973
CYYMLQAAD

34974
DDDTENDFM

34975
DGGQTHVSY

34976
DIHNYLMSE

34977
DISMNQDVN

34978
DKRCNKWDP

34979
DMRLQDHVR

34980
DNFFTQYLA

34981
DQESLNAHY

34982
DREQNEMLG

34983
DSSSSIGHW

34984
DTCRRLINE

34985
DVDWANSLL

34986
DVRLIGHEA

34987
DYARCQSVW

34988
EAEPNGFGL

34989
EAGDVRRFG

34990
EAGPHSDTW

TABLE 83

Sequences of the 581 to 589 Region in AAV5 VP1

Capsid Polypeptide that Drive Spinal Cord

Tissue Tropism

SEQ

ID
581-589

NO
Sequence

36438
GCLQIERRH

36439
EVTRNHHEN

36440
CCKPQCKCQ

36441
MVFLEQHGR

36442
YITCQYSHH

36443
ELVPYYTGS

36444
AWSDLMNCW

36445
MEEPYTSYN

36446
NWYTTWIDE

36447
MMKRNRFIC

36448
ACVERLHHK

36449
ECRMAVQSC

36450
APFFTFTNH

36451
ASLVKWDFE

36452
ATMWGDCDY

36453
AWKMAHNSF

36454
CHHDVNVPI

36455
CEMLNGCFQ

36456
DLCAGNKSQ

36457
DLCEGNKSQ

36458
DLCGGNKSQ

36459
DPMKEFVSH

36460
EKEWRPLQW

36461
ESNGMYFDR

36462
GNCVYHCYE

36463
HITCQYSHH

36464
KHMPTLQWQ

36465
KSTEQMHRA

36466
KVNCRMIIM

36467
LNLARYEAP

36468
MGMWKVWAC

36469
MTGDSGKFR

36470
NIHAVFVQS

36471
NIHDVFVQS

36472
NITLVMGQH

36473
QCCVTYVNS

36474
QEFYCWIDG

36475
QNEGRGSIV

36476
RTAMFESRL

36477
SHARKVYDC

36478
SIYQKWVTN

36479
SKCPWFHNG

36480
SKNERNYDI

36481
SQCVMQFPL

36482
SYARKVYDC

36483
TAALGHSFC

36484
TQGMYVESY

36485
TTLLVQAEW

36486
VIIYPQFKC

36487
YGGMEFEDC

36488
YGGVEFEDC

36489
YQTLCMHQD

36490
YSKVFYFFS

36491
YTQAMCITA

36492
ADQHTSPTS

36493
AIILLCHAE

36494
ATRCWWREI

36495
CIIARSTAA

36496
CNIHWTMAE

36497
CNNRLDVFN

36498
CVHERANAN

36499
DERPNMFWS

36500
DNAVNSHVF

36501
DNYYYWGSL

36502
EDFVTTSEL

36503
ETANGMCST

36504
EVKQGFTQG

36505
EWHYRNKDA

36506
GEPDHSPAN

36507
GIKSCWQNS

36508
GITEQPLAR

36509
GSKGMAFCS

36510
GVNEFLDTN

36511
HMGYVNNQS

36512
HQHEAPMDE

36513
HTTPCARSR

36514
KSDGQMRQS

36515
KTYSAMYSP

36516
LFYMKEDMR

36517
LPTLRVADK

36518
MHMSYHWHE

36519
MHMSYHWRE

36520
MMLSDDRTP

36521
NIDQWKHOQ

36522
QISMAHFLP

36523
QMEQWHRGG

36524
QNWAALACP

36525
QVSVSVIAW

36526
SITEQPLAR

36527
SMAICMHDN

36528
SMQSAEPSC

36529
STHQSEFTT

36530
STVYRWDMP

36531
SVMLKANEV

36532
SVYHCWRSQ

36533
TAEVRWPCP

36534
TASFAQRRS

36535
TDDPPLLTY

36536
TFCESMPWV

36537
TIRYEGMSG

36538
TKKQMEWQM

36539
TQQTGWVYN

36540
TTYIPASAA

36541
VCHNPDPFT

36542
VSSDKQVLE

36543
WECQWDMSM

36544
WFCYLSPMI

36545
WFEPSNLIK

36546
LGMHYKDNC

36547
ATGTYDLQE

36548
APKAPKMYH

36549
DMVTTRSPF

36550
DQVNKTYVI

36551
DWSVWIMET

36552
FLTTVLRVC

36553
HLTTIPGDR

36554
IFYECYSNK

36555
MYIFVWRFM

36556
PEKVWSDLK

36557
SWFYMEKMP

36558
TWLYGWPIK

36559
VMPIAWQQD

36560
VQKCHTCGA

36561
VQKCRTCGA

36562
WMLLGMTWC

36563
FTNTKEQAF

36564
YSWFYWADY

36565
VRWVRLHKK

36566
VCYWWLNWF

36567
THKWFCEDS

36568
DPYSSKYES

36569
EDWLRRNIW

36570
KHYKWGHCQ

36571
ESVPLRETT

36572
SSSPGKGWM

36573
GSTSRNQRD

36574
AHTQYQGQN

36575
KFDVIRQAM

36576
GTIPKOHEF

36577
QKAIVTQND

36578
MDYHCQVED

36579
ERQSTHGQY

36580
KCWDCHEAG

36581
NHDEKLWAP

36582
DGMHRELYS

36583
AKMWQLENP

36584
LTNRTQESW

36585
STALMDANM

36586
KQAGYDFFH

36587
SPQQPSVFH

36588
TTYPHNTHG

36589
CMEAQGECY

36590
ISLVWMHYP

36591
NNGPIWISS

36592
AHLHNQFMP

36593
RTEAHLPAP

36594
EGASPLACT

36595
DANHCMWWQ

36596
EKYQIHWDR

36597
KQEWHVHTE

36598
KLYGLNHSC

36599
QSSSSYLWG

36600
DIRIVSDGK

36601
MALPLHVLS

36602
QTMCITIHP

36603
RHWYIALYA

36604
LSAKQQFFC

36605
GEGVEQMDG

36606
RQIGTGNME

36607
IMTIIPYGP

36608
VNDDYECGI

36609
NTQYSAKND

36610
SRTICVHGN

36611
KHIVGQDGG

36612
KCLPYMEMF

36613
KAYMMMEGP

36614
SVTYHEWHP

36615
YSDMFHKDH

36616
GNECFHNVE

36617
IAKIGNCVW

36618
SGNQIVQAP

36619
ASFFLNPLE

36620
DMKQKAEYT

36621
MMEATHKLS

36622
GCEEVGRCQ

36623
CIHPVLVTM

36624
YDWGRVTLN

36625
QKAIVAQND

36626
TFTYEAKFW

36627
ISWISHASP

36628
PCQQLMCFM

36629
VSVQSRAQL

36630
NVCDNGHEI

36631
HFQPVFMQP

36632
HMEQCWCNE

36633
LPPPEEKWL

36634
ESAEPRMIP

36635
ICIRGARAD

36636
KKHKGWCRA

36637
AEVPYVMQQ

36638
MAEQMERDF

36639
ITASIKDQW

36640
ETDCMEGGH

36641
QFAQLWCAK

36642
KACMSHQGR

36643
GLFEGSPGA

36644
AYHCYMSWM

36645
GQQTIEMSY

36646
ATGTYNLQE

36647
GHLFNNNEW

36648
FAQQIINAS

36649
AGMYKCQED

36650
LEIGGKTRT

36651
NWTGNMQWA

36652
DIEFLDCEN

36653
KMAGIGCWY

36654
QCTAHNTCM

36655
LHGIINQDN

36656
AAGREYWLY

36657
AAVPVNKLT

36658
ADQSYNEYY

36659
AELCNNIIK

36660
AEYTHDCAM

36661
AGHVFWDGE

36662
AIGPSWMGV

36663
AIRPHHAKS

36664
AQKMEDQSL

36665
AQKMKDQSL

36666
ATEMFVQHD

36667
ATIAQWCIH

35668
AYQWFMSQP

36669
CDSIYHHMP

36670
CHFKYPRIE

36671
CSYIESWQI

36672
CIGHFSCQQ

36673
CTYTMATNR

36674
DAHKFARYP

36675
DAKSPCDSF

36676
DAVNDLTCP

36677
DDTEMSYCY

36678
DFWYSMAFP

36679
DGAWVKMMY

36680
DQAELLSDH

36681
DQFRALDLR

36682
DVASYCQLQ

36683
EAGMWPSKA

36684
EAWMVMNDQ

36685
ECRSNDYDR

36686
EDVFYQCNH

36687
EFRRNELGA

36688
EFWVCTRNH

36689
EGEARCHWH

36690
EHFRAYTQY

36691
EIKCSGGYA

36692
EKRESFGCG

36693
ENEMYMKQD

36694
EQDRGPMQC

36695
ESMKFRTFS

36696
FASAFANSD

36697
FMHSYEKFD

36698
FNIAESTPM

36699
FNPNRITKT

36700
FNVFIDRSN

36701
FQRRTQLIT

36702
FTVFYRLQW

36703
FTVGGLAFA

36704
GDEYYDTMG

36705
GDLSRDQED

36706
GFRTQDMIE

36707
GPCMETQHH

36708
GSEQKIHIN

36709
GTQSYLVKN

36710
HATFTWRCL

36711
HFWITEQMD

36712
HGCVETNEH

36713
HMLKMEMFN

36714
HNDQYEHDE

36715
HVDNDNMHE

36716
HVYYCPEAI

36717
IACPMGIHD

36718
IAESYRMMH

36719
IDSWMMIPS

36720
IGEPQAYMT

36721
INHGVNHQW

36722
ISMRSECKD

36723
ISWITRADI

36724
IVNLQHYHG

36725
IVSTSGNND

36726
IWEDMKGKF

36727
KAHLVLAGD

36728
KAMTDAANW

36729
KCAITETSD

36730
KFTTAMTEF

36731
KIVESHGTN

36732
KKTEEMPVT

36733
KPCTQAAEP

36734
KSYSCEQTS

36735
KTHVMHWEV

36736
KTYSVMYSP

36737
KVCNHELFG

36738
LAAMSLTWW

36739
LDGVFFEFF

36740
LFRSHTCHH

36741
LRTQLLLQD

36742
LSDSYVRKF

36743
LVKGREQFN

36744
LVQCHHVHC

36745
MAMWARTEP

36746
MDSQHNKWQ

36747
MEQFDHCP1

36748
MFRIEAACP

36749
MGAHFMNPP

36750
MKTFWDATF

36751
MVGPSWDTT

36752
MYYLMQKTG

36753
NAMLWRMHN

36754
NCCQEHNVG

36755
NEGKWFWAS

36756
NHDEKLWVP

36757
NHPVGWDND

36758
NHVTMCVLR

36759
NIVDCDDVR

36760
NLGWQEWDS

36761
NNPFSRYMP

36762
NSMWLCTNQ

36763
NSVMMTMIA

36764
NSVVMTMIA

36765
NTIKVEDLG

36766
NWGDQRRHV

36767
NYAQYYRAW

36768
NYCTYHSTH

36769
PADHKQTSV

36770
PFCESACHI

36771
PGDRTISMW

36772
PGMMYQFAW

36773
PHVCGEGKI

36774
PKGWADPCP

36775
PNWGLDMAP

36776
PTCIIMKGN

36777
PITHRIGSY

36778
QAARMQKYQ

36779
QAVSAVDKC

36780
QDLEWIAGC

36781
QHHPINCAT

36782
QHMVTSHNP

36783
QHNTSYPSF

36784
QPTLPDEHE

36785
QTVPPMVQE

36786
QWIVFTCII

36787
QWSDIMNCD

36788
QYAHHYNIE

36789
QYVDMYTQQ

36790
RCDTHRSCT

36791
RDSSNWLCI

36792
RNYLPLQFD

36793
RSMLGCHTN

36794
RTEACTSLP

36795
RTQWCIADP

36796
RTYYEENGR

36797
SATDMTMLT

36798
SDYEKFLAY

36799
SEHWHEDAW

36800
SGNSQNAFM

36801
SHQHMWHQT

36802
SIETMDTNA

36803
SILNAMSLQ

36804
SNYFNNFGF

36805
SNYFNNFWF

36806
SPPNLRAFP

36807
SPSIDRFWV

36808
SPYWVMRMG

35809
SQKVHIDNN

35810
STHYASTYL

36811
SVADHYHLK

36812
SVAMQMHSW

36813
SVCWIWWTF

36814
SVKPHYSCW

36815
SVVYEAIWA

36816
SWHAEVVNQ

36817
TAIHMNWRQ

36818
TAPLVYPHC

36819
TASDAENYQ

36820
TCDRSLAEY

35821
TCDWSLAEY

35822
TCGYTHIFS

36823
TCQHNQHVA

36824
TCSPDEQLD

36825
TEALAQFMA

36825
TEKEFMYIC

36827
THIGSYTQF

36828
THQYWYGQE

36829
TIKGQHGMT

36830
TISDHGAEW

36831
TLLFAHDYA

36832
TRATSLTPC

35833
TTIAQWCVH

35834
TTNQNSEAM

36835
TWHNPRNDK

36836
VCQYCELRG

36837
VRMVPESEW

36838
VINWLSLQY

36839
VVLPWGLRE

36840
WKFMFEKWR

36841
WKTVCPSRQ

36842
WLAPCAKCG

36843
WNCSVYTCS

36844
WSPLYQTVR

36845
YIHTVENND

36846
YQNHYQKKW

36847
IKHIATESR

36848
VMGTGRMQH

36849
NWGDQRQHV

36850
LVFLRDQHY

36851
CRWTPEIFQ

36852
KRIKMPPPK

36853
SWYYDEQSN

36854
NNYKVQQDG

36855
HHQLHLKDS

36856
QDDVLDVTE

36857
VKNHYEGGP

36858
KPSQCLPHN

36859
DYDPDIKDQ

36860
CDSIYRHMP

36861
GAMMGNSLP

36862
CTQGYGDMP

36863
MIYTGWDMM

36864
IGKEFIVMP

36865
TVNTYEHCW

36866
ECASVDISY

36867
VNSCRHAWY

36868
AVFAPNGEV

36869
KIESRNHYS

36870
NSQYAHEPP

36871
AIGHMTQLP

36872
QIIPQFAYF

36873
YFMTWGKCD

36874
IKHGGWGFH

36875
ECVSKWVVF

36876
SPPNWRAFP

36877
AFIQQSNQD

36878
VSNQFKYND

36879
QMHRLFNDE

36880
SKMEEVMDP

36881
MEQPTCMME

36882
QPGTQEVLQ

36883
GCQG1PRMQ

36884
YSDNLERCG

36885
YANQAMDYM

36886
TWPGGCKSV

36887
AVDDCESER

36888
KGVWDPSSM

36889
MTGIVLQNN

36890
QVNYYTKGM

36891
QIGSHSFSC

36892
EINREVRQN

36893
QTIIGNVWG

36894
EIGLNGAGT

36895
RTVQEDGCG

36896
NCHIDHVQD

36897
EIKCSVKLE

36898
WVPTPWRTH

36899
CTSTMHAFQ

36900
MSANMMTKG

36901
LFNYEGRTQ

36902
CPNLPGMIN

36903
SAAWQFGGS

36904
MMATAPELQ

36905
NYAQYYRTW

36906
SQDCGRPES

36907
HEAQKEDCN

36908
EMDSITQFH

36909
FHMNSWLSR

36910
YWIPTHVLN

36911
WYEMMVGYH

36912
WDVKFWETM

36913
SGTSIHTYL

36914
ECVEWNAPD

36915
MIEGHAAPR

36916
PILLCHAPE

36917
VFLPIARDQ

36918
FPEASITRW

36919
PKGVSCPNL

36920
KHDGEIRVL

36921
DSDPITVDL

36922
SDCEKFLAY

36923
DNVVMWWNQ

36924
KDFNFEAQG

36925
PYGQCMRTF

36926
TNKPKNVLM

36927
TLGMLLENI

36928
DVNLQITTH

36929
ENDYVHGCM

36930
IDLSHCRCT

36931
NRTAGSCAC

36932
LAKPKCKLT

36933
ATGLPARFP

36934
NHAWTKPWQ

36935
CCTQFKCVS

36936
AHVFEPQRV

36937
ALFCYNDHV

36938
AMMPCVGWQ

36939
AQLCCHNET

36940
ASCEWSWRL

36941
AVHWTQKRE

36942
AVTKCMKTP

36943
AWAEWCEYS

36944
AWKDIPSAA

36945
AWMHAQLSR

36946
AWMHTQLSR

36947
AWTEWCEYS

36948
AYYQTICAA

36949
CCDLWDDMS

36950
CCGPSATAQ

36951
CFCMKEWPN

36952
CIPPPGYKD

36953
CKEAWADCT

36954
CKNVETQMA

36955
CSAAFQLQP

36956
CSTKAHRQY

36957
CTEQIHRHV

36958
CTQQEASMM

36959
CVLQENANA

36960
CVTYLDAWA

36961
CWGFACLLI

36962
CWWFACLLI

36963
DADSSIQLG

36964
DADSTIQLG

36965
DAGIGMKFY

36966
DELWWIANG

36967
DFAYQCYLN

36968
DFRTFNQES

36969
DHEHYTWFG

36970
DHHTCLEQY

36971
DKRAWDWLR

36972
DLMKIDKWD

36973
DMHMLNCCE

36974
DMHMLNCCG

36975
DPLKHCYDN

36976
DRHNHDVAI

36977
DRHNHDVTI

36978
DSRQQMSLE

36979
DTEYQTNAN

36980
DVAHKDDML

36981
DVQQANGRY

36982
DVTMAEVIG

36983
DVWTLPECS

36984
DYEVCGNWS

36985
DYEYVMSIK

36986
DYNESDMQN

36987
EEMLVTRAS

36988
EHETVWHNP

36989
EHNIYNNGL

36990
EIMQPMQGM

36991
ELNCLSRYV

36992
ENGPVRRIG

36993
ENQRMRQIN

36994
ERTDTNLWS

36995
ERTDTNVWS

35996
ESGSTRNDG

36997
ESYEHQMHR

36998
ETHRTSNNC

36999
ETHYPKCQE

37000
ETHYPKYQE

37001
ETKFRLFPK

37002
EVMIAAALP

37003
EVMIAAALS

37004
EVVGYPIIC

37005
EWDDWYSQS

37006
FAQLKWMMS

37007
FICQIPETK

37008
FWHCINACP

37009
FWPCINACP

37010
GAPRYHESR

37011
GATRQEHAF

37012
GATWQEHAF

37013
GELWWIANG

37014
GFVQKVNDT

37015
GICWPNWYH

37016
GQNIVEMSW

37017
GVYKTNQPQ

37018
GYEGFEIVD

37019
GYEGSEIVD

37020
GYEYVMSIK

37021
HCSPDDMKC

37022
HDFPFWSKG

37023
HIHFDEWNC

37024
HKGFLQEWI

37025
HPAGYEKWW

37026
HQKSEWPNT

37027
HSDRDEIEE

37028
HSDRDEIEG

37029
HVLHQDFFI

37030
HVLHQDFFT

37031
HWNTGMLTN

37032
HWYVVLRDW

37033
IAKDMHEMS

37034
IAVLHSSQR

37035
IFALIPDAN

37036
ILDNDQVTY

37037
ILQCHEAIG

37038
IMLMCTPRK

37039
IPGFKHSDW

37040
ISTTFQWYS

37041
ITGAVOPFL

37042
KGGACDACI

37043
KKGWEFMYG

37044
KKGWEVMYG

37045
KVDPIRHGQ

37046
KVPTDGANS

37047
KYVRWFQAW

37048
LAESHYPFE

37049
LDTKWDQHI

37050
LDTKWDQHV

37051
LGGMHPCNN

37052
LGGMHPYNN

37053
LINKFPQQE

37054
LISKFPQQE

37055
LMEMTLRMG

37056
LNNHMKKYP

37057
LRISKWMTG

37058
LVCDSGYAA

37059
LVRIWHETM

37060
LWECYGTHG

37061
MAFMQGLGW

37062
MDGQACLNQ

37063
MDNQEDRCE

37064
MFALIPDAN

37065
MHAQRMKYP

37066
MHTQRMKYP

37067
MLMYLGHLT

37068
MLMYLGHRT

37069
MNEDCMQTS

37070
MNHEHGPIT

37071
MPRHKHSQT

37072
MSDDVVLAL

37073
MTVQNVPEM

37074
MVDGPAASG

37075
MYYVTQMCN

37076
NAWMYVNMS

37077
NAYPWHLFC

37078
NAYSWHLFC

37079
NIIKCVECC

37080
NIIKCVEGC

37081
NLRIHAHRT

37082
NPMRPSVEQ

37083
NVPIRALQK

37084
PIALYANCS

37085
PIELYANCS

37086
PIGLYANCS

37087
PLTTDCSPN

37088
QGFADGRPT

37089
QHLEINCVS

37090
QLVYNMWDC

37091
QMCMNISHY

37092
QMCPSCCVT

37093
QPDYEEPPS

37094
QPWVPSILH

37095
QVETVPYLS

37096
QYGHFQFEY

37097
QYRVDSYPF

37098
RCQDKVMPY

37099
RCSHDDMKC

37100
RCSPDVMKC

37101
RFDMWWLEI

37102
RHKDTEWFI

37103
RIEMFWCNN

37104
RIEMFWCTN

37105
RIKMFWCNN

37106
RLDCNQFWE

37107
SALTEPPFP

37108
SAPTELPFP

37109
SCGQFEWDN

37110
SGSWEAFGF

37111
SGSWEDFGF

37112
SGSWEGFGF

37113
SHVFEPQRV

37114
SIEMEWCNN

37115
SMHTWQTAK

37116
SMHTWQTTK

37117
SNERRHMCA

37118
SPMGVAMNG

37119
SSIPWNYYT

37120
SVTYLDAWA

37121
TCAGFCAPE

37122
TCLHDYGYG

37123
TLFCYNDHV

37124
TNAYQYHVG

37125
TPEHGLMVG

37126
TQCEIFEAH

37127
TRKVAEEMW

37128
TRPVEMQPG

37129
TVEPEYFQA

37130
TVEPEYFQD

37131
TVEPEYFRD

37132
TVFMNAHWA

37133
TVNEGEYGP

37134
TWSYDPASS

37135
VAEPSWQNN

37136
VCTLNNHTM

37137
VGNETHLGD

37138
VGNKTHLGD

37139
VHITVCRYG

37140
VIYCHPMVP

37141
VMDWWLDEN

37142
VPHTTCHTH

37143
VPHTTYHTH

37144
VQKWGLMQQ

37145
VSGSTWNDG

37146
VTEFIMHCG

37147
VTEVIMHCG

37148
VTHTVHQET

37149
VVVVNGVTE

37150
VYQRFCGKR

37151
WINPTEMGA

37152
WEVLSDTIG

37153
WKHCWMLMQ

37154
WLRILRAPH

37155
WLRILRAPY

37156
WSQEQITFD

37157
YNGKWPRLT

37158
YNMCSAGSE

37159
YNMYSAGSE

37160
YSGCWWQHG

37161
YSNRIPIGF

37162
YVPPNYTEQ

37163
LHWPITVQP

37164
VLNKNWTMC

37165
ISDCMDNYW

37166
YCVNMQYDV

37167
NSEREYAIR

37168
RAQSHMEDY

37169
EKFSANKFQ

37170
RSGQGEGHM

37171
GSHQLMWIG

37172
YMWGTELGC

37173
VTNLYEGSG

37174
EMWHHLSTP

37175
WWRATFGQA

37176
NWQWIAVQD

37177
HNLYGVYWG

37178
NQSANWLQN

37179
TGGQNYAHE

37180
CSEPDPAWN

37181
AFSSERVYA

37182
CCILWESGT

37183
CPQCDVMTS

37184
QSSFWPAPT

37185
SKWHFYLRQ

37186
VRFIRTSLQ

37187
KAEMKMGCN

37188
MANHSISAD

37189
IDRCCWDYA

37190
EPGLLWHTQ

37191
HTHDVRVHK

37192
WKPAAYSNC

37193
CGHTACSQC

37194
DICLTHDHC

37195
REIRDDYIY

37196
VYTPKYQDL

37197
KCESWKDFF

37198
QNEPEKQVK

37199
LASYESVEY

37200
HMHFDEWNC

37201
KPFHVEMSL

37202
GRHYIAAGY

37203
TTMTHSTLD

37204
AEICHKRED

37205
DAAEELRNA

37206
LIAHDHKCF

37207
YTKIESVHL

37208
ALMMDDITP

37209
TQSHWNKMS

37210
KTFQYMGPT

37211
SNHESMHMD

37212
SCGQFEWGN

37213
VEPYMFAPM

37214
TTETRNWVL

37215
TAQQFAHSG

37216
EANPYTATK

37217
FQGMCQLRE

37218
IIIQRSRTY

37219
DFTEEGEAY

37220
MVKLDAQDA

37221
EIMQPLQGM

37222
VPNRNCQFS

37223
CIHSVHFAA

37224
MFAKWMSGS

37225
ESESWIHMP

37226
SALTELPFP

37227
FGLQMMYIV

37228
PAPMSEYNF

37229
ECALAQYGS

37230
SHMGKHHHM

37231
ITAKQEVTS

37232
AADCGSFDH

37233
AADCGSFYH

37234
ADHTENFSD

37235
AEDCGSFDH

37236
AEDTPYNIY

37237
AESPQRWAL

37238
AEWSEGECA

37239
AEWSEWECA

37240
AEWSEWEGA

37241
AGQKHWHYS

37242
AIDHHTEVV

37243
ATETFNEDN

37244
AVCMEEREP

37245
AWCWQVILM

37246
CAPKQECSR

37247
CDVWWFPNL

37248
CNRQKYTAK

37249
CNSSESHYY

37250
CWTMLMFST

37251
CYQHLNTMN

37252
DANHCMWWR

37253
DDHCCTIPT

37254
DEPTLEEDM

37255
DGTLAILCT

37256
DGTLAIMCA

37257
DGTLAIMCT

37258
DGYQKYQEQ

37259
DIACLMNST

37260
DIEGYGHSW

37261
DISQTTITK

37262
DKLPMAMDP

37263
DKNVFEEHV

37264
DNCQVENHL

37265
DPPHRNCTI

37266
DQEKCLSFG

37267
EARMPDDWS

37268
EARMPNDWP

37269
EFGFKIHRY

37270
EIEHACNSG

37271
ELQYYKEHT

37272
ELYGLNHSC

37273
EMaRGRIVG

37274
ESCHLQHEE

37275
ESCHPQHGE

37276
EVCTDDGNV

37277
EYATKVKMD

37278
FHCAMEGRT

37279
FSRHWECVQ

37280
FSRRWECVQ

37281
FWSNIIGIY

37282
GFNMNNMVA

37283
GHLDPFDCG

37284
GLCIYDHRH

37285
GMSMSQSHK

37286
GNECFHNVA

37287
GNNTFNQDQ

37288
GQNINRGVC

37289
GTAGPCRSS

37290
GWCWQVILM

37291
GYNKWGRNE

37292
HEGLIMRLA

37293
HIGNKHIEC

37294
HMEQCRCNE

37295
HMYATEHCS

37296
IACRTHQTE

37297
IARRTHQTE

37298
ICHCCMDWG

37299
ICIRGARAA

37300
ICIRGVRAD

37301
ICTFYQPSP

37302
IHSPFMPYH

37303
ITTARNQYN

37304
IVSPFQTGT

37305
IYRLCQDSD

37306
KAEVKSPAP

37307
KATMTNCCF

37308
KCEGEEAGR

37309
KEDFGWKHM

37310
KKEQDHRDK

37311
KQAGYGFEH

37312
KYEVPGKYG

37313
LGSPFYERG

37314
LHSYQWPLG

37315
LKVVQAYEC

37316
LNNRTQESW

37317
LQLIVVSQT

37318
LQYPTACGC

37319
LSAKRQFFC

37320
LTNRTQESR

37321
MAIQVFMYD

37322
MAIQVIMSD

37323
MGTILQEVA

37324
MHAEDVTSV

37325
MISAPPDTY

37326
MKRGTAFCA

37327
MKYQDTHTF

37328
MTIQVFMSD

37329
MVASVAAEE

37330
MYMHHALSP

37331
NGKTNPCQK

37332
NGTQCLNMG

37333
NGTQCPNMD

37334
NISWIQDYM

37335
NMMTLWSHI

37336
NPGWRYMWC

37337
NQDEKLWAP

37338
NSEKVYGMF

37339
NSNHYNAMR

37340
NTEPTLVTR

37341
NTEYSAKND

37342
NVDAHEQYR

37343
NVDRQHAQG

37344
NVRNGTNTC

37345
NVYVTPVVC

37346
NYRCFDPMH

37347
PMMIIPYVA

37348
PNGHHWMIP

37349
QAPVIPIMT

37350
QHNSCKDAW

37351
QMNCLKLGP

37352
QNIATMRGF

37353
QNICLGVPT

37354
QSFPCMFKD

37355
QSPVPNRYA

37356
QTESEGWRL

37357
QVNKGAPQE

37358
RADWSDIAD

37359
RCNLYLEYS

37360
RDGDANCSL

37361
REKWGHMHT

37362
REQHIAGAD

37363
RGLVQDCCR

37364
RHVHRYWEC

37365
RVKAYAPTT

37366
RWSYMQADK

37367
SAMYGSTWA

37368
SAVYTAACN

37369
SGMTWNFHA

37370
SHIQLNAWW

37371
SIFMEVAWC

37372
SIQRNCAFW

37373
SLMRFLPSI

37374
SMGWAILEG

37375
SMPVGTEID

37376
SSASVNHTN

37377
SSDMQVCWT

37378
SSDNSHQGR

37379
SSKFHERQH

37380
STDNTGTAC

37381
STGSCAPLY

37382
TAVKSEAST

37383
TDHTENFSD

37384
TFANPPENA

37385
THGIYGTHD

37386
TISWLDTEV

37387
TISWPDTEV

37388
TKDAWNGLC

37389
TMSNYGCPQ

37390
TQIMRVISV

37391
TTQRDGQWY

37392
TVGPAGISE

37393
VDGLQCCCP

37394
VDIMRNDTG

37395
VEFRPKISR

37396
VGIMWNDTG

37397
VNWEVRAYI

37398
VSYWFQWGS

37399
VTATSPGIE

37400
VVTKPAYFV

37401
WADLWQYQW

37402
WATPPLQRC

37403
WRIRPQQEI

37404
WRMFESAPG

37405
YAQPDWVMA

37406
YCMWYATPP

37407
YHVPYQHLP

37408
YHWHINSFD

37409
YPELGKCEI

37410
YTGHTNNGR

37411
YTGLMWVWA

37412
AMVLFLVDN

37413
CALKQECSR

37414
CHNQMWQIC

37415
CISQTFRYY

37416
CWAEFRMRV

37417
CWMLGRDCC

37418
DGAAPLTFC

37419
DIAMNPRMC

37420
DITLWMATE

37421
DLMWHVNAI

37422
EMCHSMGDF

37423
EMCVMSFGY

37424
FFGWKKSNY

37425
FGREGCPTE

37426
FVELHCHTE

37427
GDTVMLVPS

37428
GGPFKVASS

37429
GTEHNTATI

37430
HTRVVESTG

37431
IAKIGNCVC

37432
IHTGTNSYA

37433
IMVHCREMG

37434
IPQVCQFGQ

37435
IRSQTLTDC

37436
IVHKLESVN

37437
KEARSQTGD

TABLE 84

Sequences of the 581 to 589 Region in AAV5 VP1

Capsid Polypeptide that Drive Spleen Tissue Tropism

SEQ

ID
581-589

NO
Sequence

39438
CTTYYCWQE

39439
TMMYYESGQ

39440
YNMVHGDEP

39441
WGSLEWDRA

39442
DSGTKKRMH

39443
QHELEGNRF

39444
TEESLVRTY

39445
DDYYQLHES

39446
LLEHPEKDL

39447
LWQHTCIHG

39448
SDFTLGYHH

39449
MIEQWDACS

39450
PQYDCWKGS

39451
NHYPDQEAE

39452
EQPHCKWTA

39453
SHNNNPSGE

39454
EVADIVSLA

39455
LVELSIKQQ

39456
LWEHTFDCD

39457
VQSFENQAC

39458
VFRVKCAAD

39459
QSDWISIPC

39460
RANTWTRRH

39461
KVPSQLVFE

39462
ENCMSRMKP

39463
RLTEDMPDM

39464
NNGVHIFCE

39465
SHMSYGQVQ

39466
DVCDDHKPN

39467
GDNMVQWVS

39468
GYASIVKRD

39469
DNYMQLNCV

39470
DVAPMHQVE

39471
QLRCEYQHT

39472
EASIQASWN

39473
PCAMHNWGC

39474
MNANETHGA

39475
CECHWAYPS

39476
GFQRTHVDQ

39477
MEPHYWEWP

39478
RYMNAAKFF

39479
MNEYIMSMH

39480
IYMSTGNTP

39481
QQTKNNSTD

39482
EIYSQLTDL

39483
VTKLAENVY

39484
WDYSMTKKH

39485
FKHSLREWT

39486
ALAKFQCWS

39487
FCDLAHHYE

39488
PWTTPENYT

39489
NLAPMIQVC

39490
EAELENVNM

39491
YMALDLIPY

39492
TTKNAISFE

39493
STLMPDEMA

39494
LSAQRLVCD

39495
EPMPMPVWN

39496
HYVFGMSST

39497
GHNVTAMAI

39498
LWSRFISMD

39499
VPTPWMGIS

39500
IAYHYDQQN

39501
IEHCSNRYA

39502
FAQQIINAS

39503
YVLYSWRTS

39504
VINWLSLQS

39505
QMVTQLNWE

39506
IQNPAMWGT

39507
EFAHEWYVN

39508
PYGQCMRTF

39509
INYISFVDR

39510
GVNDLQNIC

39511
CSKATWGAE

39512
DYVDQCQVS

39513
MCGCGWAKG

39514
NAELQQADI

39515
ESSQEIKTC

39516
AIRQFMMHL

39517
QTQICHSWM

39518
WFDRSWHIL

39519
FRAVYYYHS

39520
TFHQSDYWE

39521
CGCQVKCHT

39522
WNWLKVMHE

39523
MASQGGQVR

39524
DVEFCMKQY

39525
INMPVSKDQ

39526
QMCVWMTRQ

39527
SSEYSQFAI

39528
DSIIKDQSD

39529
EHPEWKCMY

39530
WKATPGGQC

39531
STARHNMQL

39532
RYQSHCWLH

39533
TFAKSDWLA

39534
NSQLSMTFG

39535
RFDQFWDTY

39536
QTSHQDQDC

39537
DHELTRSEA

39538
MITNNCNIS

39539
FNESGWPTW

39540
MWGCKLFVC

39541
SKGWADLCP

39542
HIHDTMSDY

39543
WHTIMTYES

39544
SFVHMFQGH

39545
WVHPYSYCA

39546
QVSFNARYA

39547
FSTSYCHED

39548
WLIGMWNTG

39549
NSTMEHSRW

39550
KYEFLLKYI

39551
HCDSPAATP

39552
DIEFLDCEN

39553
YSDPKNMSI

39554
ACIPQMHSA

39555
NCMHLQGCQ

39556
SRVMIHVYV

39557
SMFTHQLAQ

39558
DVTPTSWIE

39559
WCEAVGDYA

39560
GDMYYWKIQ

39561
GWDMLSVAK

39562
NMEPLKLQA

39563
CWECCTDSG

39564
GNECFHNVE

39565
LSTMFPEQT

39566
TAFQSLQQM

39567
NGVGRSGDN

39568
QAWPKTEYI

39569
YNNDMKQRR

39570
MLHTSECVQ

39571
GSHWYDCNQ

39572
PTNYGWMWR

39573
AHEDPNTWC

39574
QSMVACYTI

39575
PWRWHKSAC

39576
WINEANCLM

39577
LSPWPQTDQ

39578
LETMMRHGE

39579
LGSPFYECG

39580
ACQFYISRP

39581
VFKPLDKRY

39582
TFVRKPSLM

39583
NDMQMNWVH

39584
HAKQANGMV

39585
VKFRITTEC

39586
CVEQPVEQG

39587
MQPMESATW

39588
VEDRVFACC

39589
EVRIQCKID

39590
KQKTQNRNA

39591
KVAEGQNML

39592
YPTMTFPEE

39593
ASKQGTHDY

39594
YVIGQKDEM

39595
PHASGYTCI

39596
EKRLEGWVY

39597
NLSNLVSCR

39598
YTELMKKAC

39599
QCTAHNTCM

39600
TWAPWCCYF

39601
ICHIRTRGN

39602
CICRTWDSY

39603
EQELKNSTQ

39604
NGYLVHACQ

39605
NADMSFRNR

39606
DQRHGNVSP

39607
TWYMNWMWH

39608
KGNPLDGDT

39609
VTMIHNAFH

39610
SFKSVSQYN

39611
AFGLTTGHT

39612
CVLSRSTMR

39613
LCKFANGAA

39614
QSYPTMYQC

39615
QPYWVAVDP

39616
CKYKTKSIR

39617
QCTLEISYV

39618
EICMRSGFI

39619
WVDQNVAQD

39620
YYNHKIVEP

39621
QHEQSNHIW

39622
YVDHWTLGD

39623
NIQLLHSYC

39624
IMNPTAQNN

39625
ATMNLSWGL

39626
HTNYTVGYE

39627
PWMHIMLNP

39628
EAGPRKRDV

39629
LEIGGKTRT

39630
ARWFSTVEQ

39631
KSDAQRNVA

39632
WERCLELNQ

39633
RCADNPYFR

39634
QTDILRDVQ

39635
MDGRDEIIH

39636
ASDMQFWST

39637
CSIQINNCR

39638
P1WQGYWPW

39639
GAANRIQSW

39640
SGDWDFGEW

39641
TQCKQLTVM

39642
MHGIARMQQ

39643
LMKTSPSYW

39644
KMHGNERCF

39645
RYPCCQSPF

39646
AVYLDHVFS

39647
ITAARNQYN

39648
VGNVMIKMC

39649
HHFHVNLDI

39650
NRIQMTDFQ

39651
QISKPSVQC

39652
DVMQYQNHM

39653
MQHTDWTSG

39654
CWILHHRCS

39655
FSWPIHEPD

39656
RDQCRMESG

39657
QIKTHSNVF

39658
DSAKQWHHQ

39659
SVLHVQALA

39660
ETNDNMILH

39661
FQWRTEMNN

39662
FQTCDFWAT

39663
TVNWYSGFG

39664
YNFDYMKDG

39665
KCADLIIRS

39666
CCLWSYWAF

39667
FQTLILSGL

39668
ILINDREWG

39669
EPEGELDKT

39670
MYAGWLNHG

39671
FLTCDQFEW

39672
NMFQLKPQN

39673
PYGPIEYDE

39674
GGDGRVHNK

39675
VDRLFRMEY

39676
QWFTKGVGE

39677
QSRCVDNTV

39678
QIIYRCRST

39679
KDGRNPHCL

39680
AIGPALCLT

39681
TQPEDHWLA

39682
TGMTFNSFR

39683
NAIPYFVQA

39684
DVKSWAVCD

39685
VLPMGPVDP

39686
CDHVTKHPT

39687
SNVACIQAF

39688
THVKRFDYE

39689
VFNLMLQDK

39690
ECGCMYWGY

39691
MYTGMRCWR

39692
MDNCPPMWN

39693
ATMWMQWWD

39694
TNSCECVDR

39695
GHLFNNNEW

39696
ASMHEETCL

39697
VCDGHTAFP

39698
TISQIPWVR

39699
KNCIMLASN

39700
QGKQLYHMC

39701
YFPSIQCYE

39702
KMAGIGCWY

39703
GGALARSSI

39704
PDLWYEKSS

39705
NVWTNIAEM

39706
HIKVEHEEV

39707
EHVQCSADM

39708
LYMMALLPT

39709
ASIATMDRI

39710
YKGMEKNPV

39711
QMKGATEFC

39712
AADIFAIND

39713
NSGVLIGME

39714
TSNHNMALE

39715
LIEKQRWML

39716
SGFFVHTSN

39717
QSSHCKQMV

39718
RDYIERREP

39719
RQAWPKDGM

39720
QHHWKWSTE

39721
YCECGHWPN

39722
HMGIAYNDH

39723
LCAVCGDCD

39724
TPLMMCQAS

39725
TYESLQNSW

39726
GRALGKKQG

39727
YTKYNWKAA

39728
TATCGIEVE

39729
MPWLDKPPC

39730
TRQWMCVHC

39731
DQEAQVVTG

39732
MVENGLRLQ

39733
EAGCKMWPT

39734
AIGCFQNGG

39735
GLFEGSPGA

39736
YLNKNPHQM

39737
CQMLCRPPV

39738
VNLPYGYAK

39739
LMTPQVKQA

39740
SQERMDYDG

39741
NTHTGNDRL

39742
QGHKTMHVF

39743
FGGMTPYGV

39744
SECEVLCSL

39745
EIHMFGQSE

39746
QAKMILDGT

39747
VSDECYFDM

39748
REWMFTLMA

39749
CNGYWMCMS

39750
QIRSLITHD

39751
QFAQLWCAK

39752
RYNNCPNLW

39753
EVAFTTSSD

39754
YVKMCSMRN

39755
HTVMEWSPT

39756
TFMFNSEWS

39757
AGMYKCQED

39758
FCILASDMA

39759
DLIVATEWF

39760
QGFSTDVKP

39761
FMIMAKTQA

39762
QSAHPEPMC

39763
YDYWMQAPT

39764
VVCRMWHQP

39765
GSKIIMHTE

39766
NHSLCWDSK

39767
TSSKHLVFD

39768
TKMIIFDSW

39769
ENTMWQSSQ

39770
PVICVWHDN

39771
NWTGNMQWA

39772
MNLQIGSKG

39773
DRKQADQIY

39774
GGNAGRDTT

39775
WGGMPNNQG

39776
HSMMPGWPS

39777
DSDTWAMHE

39778
EGIMNHAPT

39779
FQVMDHANP

39780
NSNSMNCVS

39781
QQVLGLAGN

39782
QSSPDQRAL

39783
AAPIAHSMR

39784
ADPTTETPH

39785
AFNEMEQGD

39786
AHAPWCWAV

39787
AMTESVSCS

39788
ANYALHYDR

39789
AREFWHCRD

39790
ATNIKESVN

39791
AVHAFPWSF

39792
AVKDMKKWD

39793
CDEDLYVTP

39794
CHIQVDHHF

39795
CMDGQLKWF

39796
CPWNSQVTL

39797
CQQYAAWAV

39798
CRIASFKAI

39799
CSAYCHACE

39800
CSSDRDGLP

39801
DDGHHVPIM

39802
DDMFAQYFS

39803
DHSMWTWAE

39804
DICNATCWV

39805
DITQTDMWG

39806
DIWTAKENV

39807
DLAMTMVNP

39808
DSGPQTQMV

39809
DTAINQCWI

39810
DVEPSECSW

39811
DVQNANQMF

39812
DVRYELPYR

39813
EAVGMPLCN

39814
EDWYWMNLC

39815
EFKVYQNCH

39816
EKYTFCIFL

39817
ELLYTLMFH

39818
EMEAQLLGE

39819
EPCLVNISH

39820
EQNASMTMH

39821
ERDHIPCYA

39822
ESSNTQPWH

39323
ETASDPQTI

39824
ETKYSYEVG

39825
ETTDGYGQV

39826
ETVWNDEQS

39827
EVALQMFSQ

39828
EVCQNINEA

39829
EVMPITGYG

39830
FHELRHVDT

39831
FKDWNTVGM

39332
FKTRETTEI

39333
FMHHTVRSF

39834
FMPAKSVAD

39835
FMRSVQWNA

39836
FSGMETRPP

39837
FTLVATNIH

39838
FVSSVTQVA

39839
GAGDMPMLD

39840
GDQGPRQQL

39841
GFHPIPWFP

39842
GIEMDLCNG

39843
GIGPQLNWA

39844
GIRFFEKGI

39845
GMHMPCDKN

39846
GQAKYWMGT

39847
GYKPDRLHD

39848
HMVINQMDM

39849
HPRQGWMHS

39850
IAGYRHNTD

39851
IFHNYDLTG

39852
INDARQHFG

39853
INVAMQKEG

39854
IRGDEMLED

39855
ISNYVGFHG

39856
IVLHMSEGY

39857
KAMPEQLWP

39858
KAVSYQNPF

39859
KMHRRIWGA

39860
KNTETMYSE

39861
KNWPMDKHL

39862
KVNEMVAEV

39863
KVTEYCDCG

39864
LIICTWKAG

39865
LIMEAMLFS

39866
LLRRNDELK

39867
LMDCHSVIP

39868
LMQMDKPNG

39869
LPHMCKCKG

39870
LTMADGLMR

39871
LTNEVREEN

39872
LTWISHGVN

39873
MAMEYIEMY

39874
MARIYPGTC

39875
MAYMEHHVG

39876
MEMEYEEFM

39877
MKQDSTQGM

39878
MPLCGYNVR

39879
MTDQPPSKV

39880
MVWNVAMMD

39881
NDELCVYCW

39882
NGVEQIKHE

39883
NMVMFVMDN

39884
NQICHQYDA

39885
NSTVHWQQN

39886
NTQWYMMNA

39887
NVEDNRKVF

39888
NYRNQKNCH

39889
QRATTKRHA

39890
QVHPVNHWG

39891
QVQVVDEWS

39892
QYQHERGWL

39893
RGGYIVAGI

39894
RGYWDVREG

39895
SAECMTVSS

39896
SIWHYQHMQ

39897
SIYKYTALN

39898
SMLLSNQNQ

39899
SNQCLPYDC

39900
SSAVIMKFP

39901
STHTVSWDM

39902
STQPSAKQV

39903
SVDYVEGYC

39904
TAMAYTSCE

39905
TCQHAPSPM

39906
TFEQIPCWT

39907
TFHVCQTMG

39908
TMQVMEVIS

39909
TNLRPMTER

39910
TNNTMRHEF

39911
TRFSSQGKM

39912
TSHAQTQMD

39913
TSRTRNCTH

39914
TYALDMVST

39915
TYGLYDDSY

39916
VAMYYESEM

39917
VDNKFMEHK

39918
VDPCANEGH

39919
VIEEMTMVC

39920
VIMPMQPRC

39921
VMLHLIHYP

39922
VPCIWVLKD

39923
VSTCCQANG

39924
VVECSGRAN

39925
VWVVRNHAD

39926
WCAGATMQW

39927
WEKLSQRNR

39928
WEMNDMQDY

39929
WINIPCHCI

39930
WKCIVPGQD

39931
YMVWSMMMI

39932
AAAMCGYQV

39933
AAAPSDDEP

39934
AADREKSWM

39935
AADTQSYIR

39936
AADTYVNHM

39937
AADYDDQQW

39938
AAEEGHTWS

39939
AAFQPDVNF

39940
AAHEKQCAA

39941
AAHTWHVAW

39942
AAHYNDGCT

39943
AAISMLWVE

39944
AAKEATSEG

39945
AAKWKAKCA

39946
AANPIESVD

39947
AAPLKRGLP

39948
AASHTEGFL

39949
AASPCNDFW

39950
AASQCFCMP

39951
AATFKSRHA

39952
AAWHANLSR

39953
ACDVKKHFG

39954
ACHAYPSAW

39955
ACMEAIMTT

39956
ACTDFMSAC

39957
ACVEYIDSA

39958
ADAVQLMCP

39959
ADCDYGRKF

39960
ADCIFESDG

39961
ADMYANENE

39962
ADPKYVPTT

39963
ADQCNPLCA

39964
ADQCTMLRF

39965
AEMCKTLIT

39966
AEPCKLKHF

39967
AEPRHDKAC

39968
AFKGHYMAA

39969
AFNSGMAKI

39970
AFVENMWNF

39971
AGALPQAWA

39972
AGCFFTGIK

39973
AGDHFLLVG

39974
AGRMRFAKM

39975
AGSYYGVIN

39976
AHCVPWSWT

39977
AHEATSCMV

39978
AHGCGFAWP

39979
AHKQMDWHT

39980
AHLPLMLGT

39981
AHQGVAGTY

39982
AHVDNNLLF

39983
AIAAENDCT

39984
AIAEHWEGR

39985
AIAEYAQQG

39986
AIAWYMGEA

39987
AIECSWGWI

39988
AIKMDGDFQ

39989
AILPMLMQS

39990
AINIMDQEA

39991
AIPDYQIYE

39992
AISHNMKTP

39993
AISSCARQI

39994
AITMSMPTW

39995
AITVGCGWA

39996
AIVEYWSSC

39997
AIYYCLSDW

39998
AKCTWRSYH

39999
AKEDQHQFQ

40000
AKEYMMLSY

40001
AKLKQAWSG

40002
AKMDWKLPM

40003
AKQQPEKQY

40004
AKSCWPSVW

40005
AKTHNPNTD

40006
ALCSRDSSN

40007
ALLAMQLAT

40008
AMLHTMGHW

40009
AMMNWQMMN

40010
AMMTVEYFN

40011
AMREWRVFP

40012
AMTSMIQSG

40013
AMYGKAAGD

40014
AMYNPYTQK

40015
ANCQPSRVC

40016
ANHTKLKMY

40017
ANMENWVQS

40018
ANMSATYYE

40019
APAPNVRYS

40020
APNMMNWIA

40021
APVIGWGAY

40022
AQDNNNCAA

40023
AQEPVQGNY

40024
AQHVAIFNL

40025
AQMYRWVMT

40026
AQPLEHHYN

40027
AQQEVKCGY

40028
AQSHGTEMF

40029
AQTWGGAGS

40030
ARDHGKWNW

40031
ARDLYHRSE

40032
ARITWNVCT

40033
ARKQIVGMD

40034
ARQHMRRQP

40035
ASADVHYAQ

40036
ASANRKQMD

40037
ASAWQMMMP

40038
ASCEMTQTN

40039
ASDQFMADE

40040
ASHKDGWAT

40041
ASHNFSDEG

40042
ASNMHANAH

40043
ASRDYDLAP

40044
ASSSCNGGH

40045
ASTLGCDCC

40046
ASVTRVAVL

40047
ASVVSDECN

40048
ATAFGGEYY

40049
ATCYLMKLD

40050
ATDDECCVG

40051
ATDDVRCLW

40052
ATDGYSKWD

40053
ATDPERSQW

40054
ATDTWDRAS

40055
ATEYQPNTW

40056
ATFSCDPWN

40057
ATGEEDVNW

40058
ATGFGTEDS

40059
ATHEMKGMN

40060
ATIQHEAGM

40061
ATKFFRMET

40062
ATLTWNYSQ

40063
ATNFDVGHE

40064
ATNSNYEHR

40065
ATNWMRDQT

40066
ATSEQRIVV

40067
ATSLAHCVC

40068
ATTDNYHDR

40069
ATTHETVLY

40070
ATTNSSGSH

40071
AVAMLVDVW

40072
AVAMLVDVW

40073
AVCNDDHSA

40074
AVCRPSTSY

40075
AVDCYGLFS

40076
AVELETQWW

40077
AVFYPYITD

40078
AVGHNGSHR

40079
AVLNSWPAC

40080
AVMPYCELE

40081
AVQDKVCTC

40082
AVVLTDWNE

40083
AWAFHHAVP

40084
AWANYIRHE

40085
AWEFVNYPP

40086
AWEVNGHYH

40087
AWWFEAPMV

40088
AYAPEPMCP

40089
AYLNFYDGP

40090
AYLQFQIQG

40091
AYMTSNSFG

40092
AYSRATTHI

40093
AYVAKKNMN

40094
CANEHNHTA

40095
CAPVSTRKA

40096
CAQSIPEAW

40097
CATCHILHM

40098
CCCEENYRR

40099
CCEQFELGS

40100
CCLPGWGSL

40101
CCNLSDEHH

40102
CCRSGDCMP

40103
CCTVTQAPE

40104
CCYHTQLLH

40105
CDMALVGGD

40106
CDNNWPIKQ

40107
CDWSNIYDV

40108
CDYLLAQCC

40109
CEDFCMVNR

40110
CEFRPVVDH

40111
CEHTGSFQS

40112
CFDRSIGYE

40113
CFHYLMNTW

40114
CFMDGWPFV

40115
CGAICDAVV

40116
CGEPPHIQD

40117
CGRHWDPWF

40118
CGTGMNGAF

40119
CHTLKSGEH

40120
CHYPYCVPL

40121
CIGFAMESG

40122
CIQRYTTEQ

40123
CIRVQEFGG

40124
CIVEEMAHW

40125
CIYEQERKM

40126
CKNGPAYVP

40127
CLFYDEAMG

40128
CLHVPHNSL

40129
CLLVAECLQ

40130
CLQFKCRLS

40131
CLQHTSMKC

40132
CLSDNYHNQ

40133
CLVSIKDCH

40134
CMACTDIYM

40135
CMGNEDMST

40136
CMIEKNMDY

40137
CMPLSMGTR

40138
CMQNSIKEC

40139
CMRDCNHYG

40140
CNASKFWQD

40141
CNQQIQFFG

40142
CNRQGCMHE

40143
CNVYRFHEA

40144
CPCHEIHPQ

40145
CPFFHNKLA

40146
CPQCLPVTE

40147
CPYCARQHE

40148
CPYQDVFQH

40149
CQFEYQIFQ

40150
CQHLVRQGL

40151
CQLSDSFLF

40152
CQMGFCCHS

40153
CQQFYERVD

40154
CQQGSPNDW

40155
CQVESIGPH

40156
CRCWSQGHQ

40157
CREDKLEAR

40158
CRIPMPARC

40159
CRVWEPLRM

40160
CSEMMQIHC

40161
CSESGLWCS

40162
CSKGDCQPH

40163
CSLLFVSCV

40164
CSMYWDWLF

40165
CSTHIAQSE

40166
CSTSFSQRG

40167
CSYNDMDRC

40168
CTCMTKCVD

40169
CTELVQFIG

40170
CTFSIACDH

40171
CTGIPKCVK

40172
CTNQHKLSS

40173
CVCQYHQTN

40174
CVDPGVSWP

40175
CVIHMKTKI

40176
CVKQYMFAQ

40177
CVMDAMAIP

40178
CVMTEDFQF

40179
CVSESNIWH

40180
CVYRHQREY

40181
CWDSTLYGY

40182
CYEPYHRHV

40183
CYFFARNEM

40184
CYGHRQIKD

40185
CYKLSNYIV

40186
CYQWHHDGT

40187
DACKTINQF

40188
DACLYWAPW

40189
DADAWQMVT

40190
DAGMPWFMN

40191
DAGPSVVVQ

40192
DAKCGRMVA

40193
DAMPSCEFS

40194
DAQLLVMRM

40195
DAQWPLQRD

40196
DARFTFVES

40197
DCAEWLVRD

40198
DCGRAQMMA

40199
DCHGAFDRE

40200
DCSLMGSDP

40201
DCSRHHMAQ

40202
DCSTQRTIW

40203
DCWSEQQPM

40204
DCYHIDMGC

40205
DDAPLKITA

40206
DDDKKHACR

40207
DDGQCDRKG

40203
DEHCDSPAI

40209
DEMKWLNCS

40210
DFCKQCNGL

40211
DFDWEVNPT

40212
DFEPNEEHY

40213
DFLVFSVQG

40214
DFSENGHTF

40215
DFSERRIVV

40216
DFSQFNQGH

40217
DFTLSFDAH

40213
DGPLTCEQA

40219
DGPMSKQEL

40220
DGQCPNAWV

40221
DGTHNHLQG

40222
DGTYLMFHA

40223
DGVDKEWYE

40224
DHQANRTCC

40225
DHTWQHEGH

40226
DICAQSPGA

40227
DIDSVQEVP

40228
DIDVGHSFT

40229
DIFLNKCTY

40230
DIGTRVCWH

40231
DIHNDIQHG

40232
DILPVQAPG

40233
DIMPYASQG

40234
DIQIQTYQS

40235
DIQLYNNAF

40236
DISAMAGWY

40237
D1WQASTRA

40238
DKESWCEVT

40239
DKHTYEHGM

40240
DKRCFQQSN

40241
DKSSTARGM

40242
DKYDTDWAF

40243
DKYSNKCLF

40244
DLEGTEQSL

40245
DLEYIWRQD

40246
DLIMGRKKQ

40247
DLMLNFHLD

40248
DLQHFMSYP

40249
DLTQRISMS

40250
DLVNCNNAN

40251
DMAFMHSIC

40252
DMCKKMLSW

40253
DMIGSQAEL

40254
DMVYLAGNA

40255
DNEDVLHMG

40256
DNGWFENMA

40257
DNQCNEITL

40258
DNRDQHWRT

40259
DNRNERRQS

40260
DNTVNTIIW

40261
DNVGRVNSV

40262
DNWKPTRIG

40263
DPEQAYTKH

40264
DPLLKSEWG

40265
DPNNYKGYY

40266
DPPLHWVTM

40267
DQAFEWFIY

40268
DQCKTLFYT

40269
DQGQVGCFV

40270
DQGYMPTKT

40271
DQLNQQAMS

40272
DQPFMNRQH

40273
DQRICGEKC

40274
DQWLMQCKN

40275
DRCNKIWPD

40276
DRGWQNWTY

40277
DRKQAQRNG

40278
DRNTIRKMA

40279
DRPHGQMMT

40280
DSKLLCCSC

40281
DSKMAGQAN

40282
DSKQAFVMT

40283
DSKTIQVYW

40284
DSLIQDFLH

40285
DSLLANQDG

40286
DSQYQAIQF

40287
DSRHLACYT

40288
DSRILQAGD

40289
DSRMCPPYN

40290
DSRQPNAGL

40291
DSRVDSDDY

40292
DSVNSCAFP

40293
DTDWIDCMG

40294
DTEFRVMDW

40295
DTISEQMRT

40296
DTMLKFVST

40297
DTNIDLKPP

40298
DTRIDSVYE

40299
DTRQWDDGL

40300
DTSQYGNHV

40301
DTVQLMGSQ

40302
DTYELESQL

40303
DVAQWFEAS

40304
DVARCMDCQ

40305
DVASKDDWP

40306
DVCKYQHNP

40307
DVCYAGLTE

40308
DVDDRMEMC

40309
DVFNAGPPF

40310
DVHVPCVIG

40311
DVIMCNCPK

40312
DVMSMFTDV

40313
DVRSGNFYP

40314
DVSICVTCT

40315
DVSWHYAQG

40316
DVTDNGTCC

40317
DVVNEDTGP

40318
DVWEHSHPS

40319
DVWTPWNAS

40320
DWAKYWHFP

40321
DWESDYYQN

40322
DWFQCHCAA

40323
DWILELMST

40324
DWKLFNAWK

40325
DWMEGSENG

40326
DYELTPSWY

40327
DYGESKLEP

40328
DYGMPECVD

40329
DYILLTTWE

40330
DYRWIKGLA

40331
DYSQFGGGQ

40332
EAAFIDAHC

40333
EADWDSCWM

40334
EAEWVLCCK

40335
EAGPLKDCC

40336
EAHMRRYSK

40337
EAHSVRYLM

40338
EALACQDPG

40339
EAMRRSNLH

40340
EAMRRYRSP

40341
EANDYNQNC

40342
EANRIDAVS

40343
EANRVLCMN

40344
EAPLPKWQC

40345
EASCYWEGR

40346
EASMGRNGH

40347
EAYPQKECA

40348
EAYTVSGYA

40349
ECANHEKTY

40350
ECDFRMIWD

40351
ECEFPDVYA

40352
ECGEFTCML

40353
ECKVYSGGK

40354
ECQRPHPIQ

40355
ECRMIWNHK

40356
ECRPWVWPG

40357
ECSAFHWQP

40358
ECSCEEIPC

40359
ECSQMISSE

40360
ECTMTNASN

40361
ECWNWESFI

40362
ECWNYLMSW

40363
EDCLFPACF

40364
EDDPVSVAD

40365
EDFHCFPLS

40366
EDIAPAHGT

40367
EDIKQQMVC

40368
EDKRDAWAR

40369
EDMTVNWSS

40370
EDQLCHRTH

40371
EDQWVQMCA

40372
EDWHTFYWK

40373
EECVTGLHA

40374
EEEPLYVDN

40375
EFDQIGLFN

40376
EFLLHHCAP

40377
EFLPKSDQH

40378
EFMSHSTGA

40379
EFNPAMWIV

40380
EFPGKHWVW

40381
EFRQMGACG

40382
EFVVLPEAI

40383
EFYQQAYQH

40384
EGEQPSMAI

40385
EGHGCDPWF

40386
EGIKKEGMP

40387
EGLNDDSRA

40388
EGLSMNEYW

40389
EGQTSSECF

40390
EGRLTMKHH

40391
EGYQHPNEF

40392
EHDPYMKVF

40393
EHDWPADRI

40394
EHEQDCEQI

40395
EHPIDWRKE

40396
EHQLAHESA

40397
EHQLPQVDF

40398
EHRFRVKLA

40399
EHRQEQTRL

40400
EIASGSMYW

40401
EIATNYYSQ

40402
EICQINHSG

40403
EIEMDGAHW

40404
EIEVFWKMH

40405
EIGIRLGQD

40406
EIGWHMNYV

40407
EIGYRSMMT

40408
EIHEGQLAQ

40409
EILKPGAHM

40410
EIMISIGHT

40411
EIMTRYGGH

40412
EISCKKWNQ

40413
EIVYSVFGL

40414
EIWASQQVN

40415
EKEFVHSCD

40416
EKEPLHCDG

40417
EKFMLWFWG

40418
EKGCVGPVS

40419
EKHTMTWHN

40420
EKKFRTADA

40421
EKMCEWLTD

40422
EKMFTLSDS

40423
EKYGTQREN

40424
EKYHVDHHA

40425
EKYTYNMGT

40426
ELCKGNCFL

40427
ELFSFGHRE

40428
ELTLRMNWG

40429
EMAIRWDYT

40430
EMAMNGRTG

40431
EMCGSSKVY

40432
EMESWDYCQ

40433
EMGEHHSTL

40434
EMHMIKKHY

40435
EMIKTAWAS

40436
EMTNAETFT

40437
ENESCMHIE

TABLE 85

Sequences of the 581 to 589 Region in

AAV5 VP1 Capsid Polypeptide

that Drive Thyroid Gland Tissue Tropism

SEQ

ID
581-589

NO
Sequence

42438
KGVWCKIDV

42439
QQFPHNRQC

42440
DLKERMRGF

42441
ISTKWDYIA

42442
ISWISHASP

42443
DKTVQTSAR

42444
RMYATEHCS

42445
GLWLNVAPR

42446
SMHYYDTFE

42447
FNIHGDLLC

42448
WMSEQMFKG

42449
ILNMKACDV

42450
VFKRVVGFA

42451
QMQNENRQM

42452
RTREPFWWW

42453
MQYRGMSAS

42454
VSDTMNCGS

42455
EAEPWDYWA

42456
IAWADGLEF

42457
IAWCDGLEF

42458
IAWSDGLEF

42459
IAWSDGMEF

42460
IAWYDGLEF

42461
IVWSDGLEF

42462
MAWSDGLEF

42463
MWIEFNEYG

42464
MWPEFNEYG

42465
MWTECNEYG

42466
MWTEFDEYG

42467
MWTEFNAYG

42468
MWTEFNECG

42469
MWTEFNESG

42470
MWTEFNEYE

42471
MWTEFNEYV

42472
MWTEFNGYG

42473
MWTEFNKYG

42474
MWTEFNVYG

42475
MWTEFSEYG

42476
MWTEFTEYG

42477
MWTESNEYG

42478
MWTEVNEYG

42479
MWTGFNEYG

42480
SAWSDGLEF

42481
VWTEFNEYG

42482
MWAEFNEYG

42483
MWTEFNEYW

42484
MWTEFNENG

42485
MWTEFNEHG

42486
LFLQLYRHC

42487
LWRSLQDGN

42488
QISVYDTKY

42489
AMHCQPAFA

42490
NAIPYFVQA

42491
GLNICKNQQ

42492
MMVQWGAMA

42493
MTSTSSRGE

42494
DGQNRDQMC

42495
DKIDTCCFL

42496
NTQVISRWT

42497
GTMCREQMI

42498
KTCLCIFTK

42499
EASIQASWN

42500
HACSLLQQY

42501
RVTIMFTGT

42502
LWGWPLHKL

42503
TKQFVHNED

42504
AQIQPAMFP

42505
KGNSVTHVQ

42506
LIHWLKHDL

42507
ICHTEVMFN

42508
RVMMGNNQA

42509
SPQQPSVFH

42510
LTLRQGYIT

42511
EFVLGCQGI

42512
YSSHHSMDY

42513
SVTYHEWHP

42514
GCLQMPCHE

42515
MVENGLRLQ

42516
SRCTVMDPG

42517
IVQPMCYQQ

42518
DVKAFMGGV

42519
AYAPSKNAM

42520
NVYMRTCNM

42521
NAAYQMAAY

42522
LVQKWMRTC

42523
EKHFMQVDR

42524
LEIVMHSLP

42525
NQREWHGLA

42526
LADGCCCVN

42527
AVETGESAM

42528
RCADNPYFR

42529
ETHTVSREC

42530
QAAWIPFAG

42531
AVEDYWRYF

42532
QYVQCAAKD

42533
ACIPQMHSA

42534
AVTGHVIGA

42535
WGDARMIGP

42536
TGMAMSEQS

42537
VFNLMLQDK

42538
VTCNWCVMP

42539
DANHCMWWQ

42540
QISKPSVQC

42541
WTTSIAVDT

42542
QTSHQDQDC

42543
QAKMILDGT

42544
IHNRYAQAP

42545
NWYTTWIDE

42546
CMEVTCFVV

42547
AYRVKEVAY

42548
APVHPTQSM

42549
QKAIVAQND

42550
AQGFKPSDG

42551
NANYWFYYD

42552
TRETMNEWH

42553
GVWMPHGWR

42554
AVHNVKDVH

42555
ETSTHCCKV

42556
FQDERTTHH

42557
KSNLCYHVQ

42558
DTRRNLCCD

42559
AEHMVVDHE

42560
DVYTMHELH

42561
MKEDNKFAV

42562
MFFPGFVAH

42563
RFDQFWDTY

42564
HFQPVFMQP

42565
DIEFLDCEN

42566
QCLQTNPYN

42567
RTMDCEYTG

42568
MSGPNDTEF

42569
IGDEYATHT

42570
SFSDTHLGW

42571
YFPSIQCYE

42572
IRLMTPDEN

42573
ICHIRTRGN

42574
MCDNFPSAH

42575
SSIYKEIQE

42576
PMMIIPYVD

42577
PLTIEVNCT

42578
NVCDNGHEI

42579
KIAAWNDFL

42580
HQCNCLLSW

42581
GAANRIQSW

42582
DVWWTLTRD

42583
CSESYRGPD

42584
TISQIPWVR

42585
MFMTNVNSF

42586
GKGQWTAQE

42587
SGALKNREE

42588
LHSYQWRLG

42589
DVKSWAVCD

42590
LPSHCNLGS

42591
GQQTIEMSY

42592
QWFTKGVGE

42593
QRNEMSASF

42594
YPAQYTAFS

42595
DHLSNDAEM

42596
DQEAQVVTG

42597
ASMHEETCL

42598
TTYPHNTHG

42599
QFAQLWCAK

42600
MTDNNGPLF

42601
RYPCCQSPF

42602
CAWHMKQGN

42603
TSVIRLFWI

42604
TTTNQFMFQ

42605
MMLGYMEQD

42606
GLFEGSPGA

42607
AEDTPYTIY

42608
QHKWHDVGV

42609
NRIQMTDFQ

42610
YVDHWTLGD

42611
ACVERLHHK

42612
DQRHGNVSP

42613
ATGTYNLQE

42614
EAGCKMWPT

42615
KQKTQNRNA

42616
ASKQGTHDY

42617
TYESLQNSW

42618
MNLQIGSKG

42619
GRALGKKQG

42620
EVRIQCKID

42621
PHAETHWYT

42622
WERCLELNQ

42623
AGMYKCQED

42624
TVNWYSGFG

42625
ATMNLSWGL

42626
QSSHCKQMV

42627
YSDPKWMSI

42628
GHLFNNNEW

42629
DINTCRVCY

42630
MSDQKEHMS

42631
AGCSISWTP

42632
AVIVTADMQ

42633
AWQQNTFWY

42634
AWTQASRGN

42635
AWYKAWSHV

42636
DFFLMKIAD

42637
DLDFWNGQS

42638
DVQEAMNLV

42639
DVRLATAWH

42640
DVYATPVVC

42641
ECVEGQSCW

42642
ETKYWWVNF

42643
EVTQCDFFV

42644
FTGASFHIA

42645
GCMYIRHAY

42646
GIDAHAIIY

42647
GTMCRERMI

42648
GYMCIRHAY

42649
HAYSLLQQY

42650
HLAPYCQQA

42651
HLTMWLVMT

42652
IHDLVPAYD

42653
IKQFWTKKM

42654
ITHRYGWNL

42655
KGMHSMPGI

42656
KLTIDTACR

42657
KQAGMLLGW

42658
KSNDSDNEF

42659
LMWTRNTAH

42660
LNICKTNQA

42661
LRHANRRTY

42662
LVKAHCAWS

42663
MAELCMEGH

42664
MYMPATMAV

42665
NAEDVDQGE

42666
NCCFVYWWL

42667
NDRHVPCRD

42668
NLMTDWWDI

42669
PPEQFQSGK

42670
PVQKWMRTC

42671
QACLQIMWN

42672
QGYRSSEMQ

42673
QKALNVHHA

42674
QMHMVHKCE

42675
QNMLMQAGV

42676
QYIDTYHWT

42677
QYQTHPMMH

42678
RNKSPDECF

42679
RTAHRDDFA

42680
RVEYPPFTS

42681
SFLVMNRMG

42682
TDLWYEKSS

42683
TEAKRTIDW

42684
TGCYHVHMD

42685
TICKCKISN

42686
TIQQCMHLG

42687
TPQNNAMSM

42688
TRRTCPIAM

42689
VRWGYTCFY

42690
YFEHMMITT

42691
YFHMIDQIP

42692
YNDFNRGEW

42693
YSNCQPTHM

42694
ADGWWMDSL

42695
AHLEMMNQF

42696
ALLMFSLSN

42697
AMLDSHSAF

42698
AQSRMHTMD

42699
ARTMYVESF

42700
ASEYFPVFK

42701
ASIWPEGKD

42702
AVICKRFVG

42703
AVIMTADMQ

42704
AVKANDTVH

42705
AWDANLAFV

42706
CAHTDKVHH

42707
CEIVVSHVE

42708
CLRSDIGKQ

42709
CNEKGYHWH

42710
CTLSFMEDK

42711
CVRFTPTHN

42712
DCAWQTNTS

42713
DCMCNFAGM

42714
DFFPMKIAD

42715
DHRYMMSSV

42716
DISLKLCHQ

42717
DITWPNHSY

42718
EEGATSGWP

42719
EIQPMMRKP

42720
EMTLQSVSS

42721
EPKYWWVNF

42722
ESILPGKIL

42723
FASLCNNGL

42724
FASLSGPPN

42725
FTCNHLAFY

42726
GEEADEYHM

42727
GGMAHAIIN

42728
GLENVVVGL

42729
GNDFFVNIY

42730
HASSEHVQD

42731
HMGPYMESE

42732
IAMRTLNQK

42733
IAMTAILCD

42734
IATWGEQQM

42735
IECEWPQDE

42736
IECKWPQDK

42737
ITPKFMFPS

42738
KASEHLPGF

42739
KGLTIQKYA

42740
KMCTHDLAC

42741
KPQPTGIFT

42742
KTDFVFLPQ

42743
KVVPAEHQY

42744
LAGPKLWEQ

42745
LAHMIEITA

42746
LCGHCGESC

42747
LCKYCFTAD

42748
LRSLWEAWH

42749
LTKFLHKGD

42750
MGYKRAASR

42751
MMFRPNSAH

42752
MNKFREEYG

42753
MPQGAVNSD

42754
MSSFMASRE

42755
MTNNKMMYE

42756
NCVVNLCTY

42757
NEPLFMMGS

42758
NGVWCKIDV

42759
NLSPINWHL

42760
NQLPEMIET

42761
NRDGVPQSY

42762
NSSMIEWDC

42763
NTAHLQDGW

42764
NTRLVFGIT

42765
NTSIWERDQ

42766
NTWVPHEFY

42767
NVFVAYCKS

42768
NVMRVPTGQ

42769
PRNQLGPGI

42770
PSSQFKCDQ

42771
PTIACGNMV

42772
PWKYCCMDQ

42773
PYVDKWANH

42774
QIHRAGAKQ

42775
QNRLVNRQH

42776
QRDFAWQFC

42777
QRQGQPHYW

42778
QSFSYDAKE

42779
RFKTCRYMY

42780
RKVRIDGDT

42781
RNEKGYHWH

42782
RPEDVPVAP

42783
RSFMCGLIG

42784
RYKSRGALT

42785
SATILANEV

42786
SIVDCQGAF

42787
SLVPVSKTR

42788
SWTCCATQC

42789
SYDEICSTV

42790
TAILNQEIH

42791
TFSPARHPK

42792
TGHPLSNCL

42793
TLGYVWSVG

42794
TNERFTIQC

42795
TQDIWYNYV

42796
VCYCSSTAK

42797
VDQPTTHWG

42798
VHGQYPCQE

42799
VHNWFLTWW

42800
VLTTIWKNN

42801
VNAQQNWGV

42802
WANTQWQDD

42803
WGYNYQCAF

42804
WLKYKKTWI

42805
WPDQNMFAV

42806
WYTFLQFIT

42807
YCNAYMIGM

42808
MEHVVSVQP

42809
NTRFPDQTE

42810
ACLSPDHDR

42811
AMCMLSHIH

42812
ANRNCVTPH

42813
ARMINDNDP

42814
ATWSADVTA

42815
CNFTVCNMS

42816
CTAEMAPMT

42817
CTDGKHASW

42818
DEISDPEEF

42819
DGAQDKWAC

42820
DLIYKGQNG

42821
DRPKRQVLY

42822
DSHIDQFGF

42823
DTEKGQAWW

42824
EDILEITKH

42825
EMAGMREFQ

42826
ENNPSHTQL

42827
EPYKQMGAG

42828
ERTLTKAGS

42829
ETVMMEVRD

42830
EVTKCEPPS

42831
EVTKCGPPS

42832
GQKSYDGCH

42833
GVDNQDFSW

42834
HAKCNRNIK

42835
HQHDDRPDM

42836
HVMEWLQCP

42837
IAGPIEIGM

42838
IKQYPQNKP

42839
IQAVMASGW

42840
ITDEYFMNM

42841
KACSMQQEW

42842
KGVALCDYQ

42843
KIYMQCDQE

42844
KPEWNHTSA

42845
LANHAKMGL

42846
LFSKEKHHR

42847
LMSPYNFLT

42848
LMYQDDMFE

42849
LQHQLHVYE

42850
LSQCQSYVA

42851
LTKPCWPWA

42852
MAAGNYTCA

42853
MCSSDLEGC

42854
MDCCARQAP

42855
MPSCINGPY

42856
MTNRCAKYK

42857
MVTFRENPA

42858
NASVGAWAQ

42859
NEQVATPMA

42860
NIGYMNVLF

42861
NLRECTIMN

42862
NSLLRHGEQ

42863
QLHQCPAVG

42864
QSQPDHVSW

42865
QYECESDNS

42866
RNDYSDQYD

42867
RSDQHSPHM

42868
SCQRNQQTN

42869
SFHNCMLDP

42870
SSTNNQTVD

42871
STCVTEMFY

42872
TCFNQNNTW

42873
TFFQAVVIA

42874
TMASWECYQ

42875
TMDSIQWFE

42876
TQKAHKNPG

42877
TTDSKLLGH

42878
TVEDRRKQG

42879
VADCPHHFD

42880
VHPYAESVD

42881
VVMVQHRTA

42882
VVPHPFPAY

42883
WWEACPAAS

42884
YCTTLSMES

42885
YFVMQWFHY

42886
YMVQDRDEW

42887
YMYSNMQTM

42888
GTSEWMHMA

42889
TVGTAKSQI

42890
DEKFGQVED

42891
GTPDMRLTL

42892
STAFASSWE

42893
HIQCQGDSW

42894
NNDIRIEGN

42895
WAPLPYNWC

42896
LVDGYRMRW

42897
DLCVHANVR

42898
TLLPHWQAT

42899
SIPMGNNWG

42900
FNTLGLLFD

42901
ENMHKMGML

42902
RNSHGWRLD

42903
QCARPWYRS

42904
MWGWPGDAF

42905
KVIRKDSCV

42906
MSPNTKDCT

42907
NGRLWWQRY

42908
TYSQPPVSG

42909
HFTHTESAH

42910
SRFWNRWDF

42911
CLGPWNVKS

42912
CCMSKMPDE

42913
MTHPYWEDS

42914
THNVKTDYP

42915
LDYHFPFEA

42916
WETLEHWRG

42917
DANWRTWQH

42918
CVVPYYRFT

42919
HHEPYGSNC

42920
THHHVLEVP

42921
AAMILESHK

42922
CMRGKWDRL

42923
VTMIYDLHQ

42924
LDHHAEVLQ

42925
MFYPFQCGN

42926
FASHQMNMM

42927
PPESQESST

42928
PDPKAWSNY

42929
IRDLVPAYD

42930
QCPCSRLIC

42931
TICKCTISN

42932
DEEFCEMSA

42933
LPRPGILME

42934
PYGKAAEKL

42935
EKMRYPPMI

42936
SFRCCWDWH

42937
KPPGSRWIT

42938
LACVAHWPS

42939
MSHEMFWVD

42940
KNTMCDNDA

42941
AVKMPVEQS

42942
FTANEDQCH

42943
CRRGQLLYE

42944
DKIMMAEMD

42945
PIAFGCERA

42946
ETGYRENPC

42947
RFADYCKEE

42948
NWVFYTNFP

42949
WECQIQDMK

42950
DIYHFNHRS

42951
NSFANNFVM

42952
DAQWSFEGA

42953
RINCNYGQM

42954
CHPPMTNMG

42955
WPDSNMTGV

42956
QTIMRHAQT

42957
VERLCACQA

42958
VMMYNDSIS

42959
AWDANLAFA

42960
DAGKFGVDI

42961
NQSVCYSSV

42962
LRHDVFSQA

42963
DFAAQQRAQ

42964
SAGPFFDHY

42965
CMASVNYFD

42966
CWARMQGYM

42967
CFVDSVFMY

42968
APRNVHCGT

42969
GRCFMPPGR

42970
KNHAHEFIE

42971
IIAVSMRGR

42972
LPRMPECSV

42973
TTMVQMGAW

42974
TLVSCDQGN

42975
SADRMISHE

42976
MPHHSPDCW

42977
TPSQMDMDT

42978
SCNFEFLWM

42979
TTVDYAGTG

42980
WKCCQDQLL

42981
SWFAGQWEW

42982
EITDKYWGE

42983
CRGSLSNAN

42984
KAYMMMEGP

42985
VPLGFNFAG

42986
AVSNWLETS

42987
MWTVARMFP

42988
ITHKYGWNL

42989
QGRWACWYS

42990
NHASFKRFM

42991
RNMALHHPT

42992
MWTKFNEYG

42993
HLLKKDASD

42994
GQAGVHNLS

42995
SFFREDYSF

42996
RCTDCSYFY

42997
IKRAYDNNG

42998
SQQHCQHDQ

42999
QGDFFNICP

43000
CMQKYIPDV

43001
PSLWKEKVW

43002
YGTAMDSLY

43003
IHQCIVCAP

43004
HLAPYCQQE

43005
MIQSGQKDF

43006
EVDQYYSLA

43007
KIQMAVSLD

43008
LINQWMHDN

43009
HTHDVRVHK

43010
MPASNWVAE

43011
RTVHRDDFA

43012
TSRDGGWAY

43013
KNETFIAST

43014
EVAGCATAV

43015
MWTEFNEDG

43016
QPGHTYNAP

43017
MESHHHAHA

43018
DCVCCSELV

43019
TKQPVHFLG

43020
AMNWWDAPQ

43021
YFMTCSTYP

43022
GSFMSCEKN

43023
VDQHGWNDM

43024
HHSWQPQKY

43025
QFCTSCASN

43026
KMLLVMDGD

43027
ERTCIATMW

43028
GVDNRQWLF

43029
NEAPYSTWQ

43030
SGNIDVHTM

43031
AADNYSWVV

43032
AAHRGECMA

43033
AAVPRMWAM

43034
AAWGYNQFF

43035
AAWVDKWWV

43036
AAYMSSECL

43037
ACCCHENVL

43038
ACEHPLNYD

43039
ACGVTQMNP

43040
ACQYDVETL

43041
AEDVDEGCD

43042
AEIRNVACW

43043
AFFMDHMWH

43044
AGINCQWCP

43045
AHFPFFVYW

43046
AIFSREENG

43047
AIGIDHQAF

43048
AIGNFHMAY

43049
AIIWGAIQE

43050
AIQMPDVTY

43051
AKDMHVANE

43052
AKLSNCAIP

43053
AKMGFDRFT

43054
AKSHHDDSA

43055
ALEYIRQGT

43056
ALHHYPVYH

43057
ALHVVMDVG

43058
AMDYLMEHV

43059
AMGNQRNYY

43060
AMHAQQNRE

43061
AMIASHHCQ

43062
AMKFAGECA

43063
AMKFAGEGA

43064
AMMHRNNET

43065
ANAYQINGQ

43066
ANCMDFHIT

43067
ANFYNEEAT

43068
ANHHQTYDT

43069
ANQISVRGV

43070
APCWEGSMR

43071
APESARPAN

43072
APIWNDSLM

43073
APQVSKRFA

43074
APSTFANWG

43075
APVNMENPI

43076
AQQCSRLMF

43077
AQRPCDDWQ

43078
AQTMSHCFW

43079
ARAAQLSYW

43080
ARQCNPNCT

43081
ASLDQAEQN

43082
ATENEHAMP

43083
ATNQRKAYT

43084
ATTNMFVDT

43085
ATVKYNCHW

43086
AVAFGSQED

43087
AVGFCQYSS

43088
AVNHTPIWA

43089
AVNYGHEGG

43090
AVIDKCDNP

43091
AVTSYDNMQ

43092
AWTLNNNHW

43093
AYTDIWNFW

43094
CAAMAYEKG

43095
CAEVQGETS

43096
CASGFGRLA

43097
CAWVMQWHH

43098
CCHFDEFYM

43099
CDNSENFLT

43100
CDTRWYCLC

43101
CEAEKVEQP

43102
CEGVANPWE

43103
CFPLCSFST

43104
CFTPHESMQ

43105
CGESWAGFN

43106
CGPNVSMMP

43107
CGVVESGMN

43108
CHKPYHLGI

43109
CHYEYSKFG

43110
CIETVHFHF

43111
CIWPYQAIQ

43112
CKPLECQLV

43113
CLCLIVDCE

43114
CLLGEVMFD

43115
CLTDSQDLP

43116
CMHWGWIPM

43117
CMKIFKFNS

43118
CMMTYMHLG

43119
CMNCRHQCL

43120
CMQKIDHSW

43121
CMSVSTRWY

43122
CMVCPGREE

43123
CNDRYMFAH

43124
CPCTFADEQ

43125
CPKPSDGSF

43126
CPLTSWHFE

43127
CQHRTHCCM

43128
CQIPEGHEG

43129
CQVKRNATG

43130
CSLRLIAES

43131
CTALCYSRQ

43132
CTVIDINKN

43133
CTWLTQHQF

43134
CVAIENVNF

43135
CVRQTVTLC

43136
CVSHYASQM

43137
CWPFAINHI

43138
CWTMFCPPT

43139
CWTQMKFWG

43140
CYHQYETRC

43141
CYKSTQGAE

43142
DAGDENAPE

43143
DAKMGAEFS

43144
DAKNECASW

43145
DALLHRHYE

43146
DAREYWRYQ

43147
DASPKGGAS

43148
DAWLHEQGT

43149
DCNAADPGR

43150
DCQYVVNDC

43151
DCRMCERHV

43152
DCRWMSIMG

43153
DDGKTYCLY

43154
DDRNVWSAD

43155
DDVRQGDCW

43156
DEIDIGREG

43157
DEMLKQFNG

43158
DFNVTAFRD

43159
DFVGCSHWG

43160
DGTPISCVG

43161
DHDDTRLHR

43162
DHDKREIVT

43163
DHFYIAKQY

43164
DHGFEGGWP

43165
DIWHTQDME

43166
DKAQIKMYG

43167
DKDEHDTQD

43168
DKKQSCDTS

43169
DKQLMMLHT

43170
DKRWQKMVR

43171
DKSQMHKWG

43172
DLCFMEQLH

43173
DLEVTSASQ

43174
DLHDVVFVV

43175
DMHGIENAR

43176
DMPEDSIHW

43177
DNKSIVMSQ

43178
DNNVQKCGQ

43179
DNQPTVCHT

43180
DNRPPLFWQ

43181
DPEYTVVLT

43182
DQHPSFLGC

43183
DQNESGFGC

43184
DQWQATIAE

43185
DRCQTVIDW

43186
DRKCMIPNK

43187
DRQQMDCYQ

43188
DRTPLHQQQ

43189
DSAELHWPH

43190
DSFTFLKKA

43191
DSGILETCD

43192
DSHAYWMYP

43193
DSNFSYLGH

43194
DSQFKMAAW

43195
DTAAQKWEA

43196
DTDTVLQSS

43197
DTEWNHHNT

43198
DTFQPHWYS

43199
DTKHSSYDS

43200
DTMTKEKRH

43201
DTMVDEYDE

43202
DVCSTFGWM

43203
DVDQEQDPI

43204
DVGIHEWCT

43205
DVGLSGFER

43206
DVHFLGEFF

43207
DVLFEGQGS

43208
DVNPASHFC

43209
DVQSWCRQG

43210
DWKPEMDGS

43211
DWRMKKSSY

43212
DWSHTQYGA

43213
DWVCMDWCQ

43214
EADDHDQNR

43215
EAHISAEFK

43216
EAIFHDVEM

43217
EAKTSYRMW

43218
EARETAIMC

43219
EATQHETRV

43220
ECMCVQFCQ

43221
ECTMIEYMG

43222
EDCNTNERS

43223
EDIQFDGDS

43224
EDKTQEMWI

43225
EDLLMNSNT

43226
EEPAMKIFM

43227
EFEHFNYQD

43228
EFRWQQSTP

43229
EFTDQADWE

43230
EFTQDNWGQ

43231
EFYPSTPCS

43232
EGNWWHEYA

43233
EGTDGDMFS

43234
EHESMCCGW

43235
EHKLWEGFA

43236
EHYQKHCYL

43237
EIFQAFLLS

43238
EIQRNHAIQ

43239
EIWMMNGEY

43240
EKAELVRDL

43241
EKCLLMNAR

43242
EKLYSTLQP

43243
ELAEYGNGW

43244
ELEVSKLWW

43245
ELEWMMHCA

43246
EMMYKKLDQ

43247
EMTMEGQDI

43248
EMVQGDTHE

43249
ENDFLCAWT

43250
ENMWGYANH

43251
ENNEYWHVE

43252
EPEMQNQNQ

43253
EPRVHTWRK

43254
ERDYCAYTS

43255
ERKMCFCHP

43256
ESAWKHFVP

43257
ESKWNNPDA

43258
ESMLAIMST

43259
ESNRMMGCG

43260
ESRANATLR

43261
ESVHEGTSS

43262
ESYPNNHDL

43263
ESYYKPDFQ

43264
ETFFLPNCN

43265
ETMCGNELG

43266
ETMDREEDK

43267
ETMFYHIDH

43268
ETQAGAPCN

43269
ETQSAAYDE

43270
ETTWQCIDP

43271
EVAQNTPHQ

43272
EVGDCMPQE

43273
EVGQHEATI

43274
EVSTQTKLS

43275
EVTYWQEWT

43276
EVYQGMLYF

43277
EVYQQQKDH

43278
EWKMWYLQG

43279
EWTDWRDEG

43280
EYEFEGNAP

43281
EYRQAQSFS

43282
FAEQEHMAS

43283
FAIQEYHIT

43284
FATLQAAQD

43285
FATTTYADC

43286
FCHYLVYDD

43287
FFRCEKITA

43288
FISTFERHI

43289
FITEIQDYT

43290
FITRHPVYD

43291
FKYMWNFSN

43292
FLKNKDCLL

43293
FLLQYMQPS

43294
FLVQLSDGV

43295
FMGFCGPYQ

43296
FMNPIGVNG

43297
FMSSRVINF

43298
FPAFPKEAC

43299
FQHDRHTYL

43300
FQIPFCRSF

43301
FQMHHKWDL

43302
FSAMYPHMN

43303
FTALLHYDI

43304
FTAYMQEFA

43305
FTCPVSHFE

43306
FTPWPQCCQ

43307
FTTQFIDGK

43308
FTWTQCVVP

43309
FVEVRMMCH

43310
FVKMRLSDD

43311
FVTVWRTTG

43312
FVWNTMSTE

43313
FYLPMEHMD

43314
FYTETNHAP

43315
GAGLYMVMG

43316
GATCNMFSY

43317
GAVWQQGSD

43318
GDSQLNHWM

43319
GEVEFVPRN

43320
GFTTMYKHG

43321
GHEVQNWPM

43322
GHHQRTTIN

43323
GHKHGIFAG

43324
GHKTPSGPG

43325
GHMCDCTSM

43326
GIDENHAGL

43327
GIGCPQQWE

43328
GIIPWMHFM

43329
GIKANIGTK

43330
GKCENKLST

43331
GKHKTYEPS

43332
GLQAFCFQS

43333
GLQHYGEID

43334
GMCENVSSF

43335
GMPEDSIHW

43336
GMQVIHVTG

43337
GPPMYKDCM

43338
GPYKNITFQ

43339
GQGFRRQHR

43340
GQVEMFADR

43341
GQVSSTWWH

43342
GRNNNTTEQ

43343
GSAPFQCIG

43344
GSHQFSGPH

43345
GSNPPMMQY

43346
GTGTCMWAE

43347
GTQPVNWLI

43348
GTVMRGMAL

43349
GVAPLMDQA

43350
GVEFQHNQA

43351
GVGCEILGA

43352
GVHPDNWRN

43353
GVQLAMSDC

43354
GVQSLHFNS

43355
GVTCKDMYH

43356
GWSQEGHFA

43357
GWWGLVHNQ

43358
GWWYYGQCR

43359
GYASHQHWQ

43360
HAGCSTRTG

43361
HAIHKWPRW

43362
HALGRVANR

43363
HAMLNEICD

43364
HATQRSADA

43365
HAYLAGWCN

43366
HCQDLQIGM

43367
HCWAQYLRF

43368
HGARGEQFA

43369
HGCYVAHCG

43370
HGEMCAFFQ

43371
HGRCIWPMW

43372
HGYADCKAH

43373
HHCNTSEFV

43374
HHEIKVIFE

43375
HHILNVWTE

43376
HHNFRMAMN

43377
HIAIVCLMD

43378
HLLHVCLEK

43379
HMIRGYCDY

43380
HNGMDTQWH

43381
HNHHECRQM

43382
HPPPLPWQS

43383
HQDQPEWAK

43384
HRIAEICRP

43385
HSEMLDLQE

43386
HSQGFDVTQ

43387
HTADMRAYD

43388
HTLFHRDCA

43389
HVATKEECT

43390
HVEPLEETM

43391
HYDPYNRTL

43392
HYYTPDRMT

43393
IACMENLSL

43394
IAFWIDGQC

43395
IAHWCHKCW

43396
IATGRESAI

43397
IATGWESAI

43398
IAWEKCDAW

43399
ICHSGGWST

43400
ICRREWNPV

43401
IDWVGHMGA

43402
IECNYCKFY

43403
IECYKGMFC

43404
IEPEGHLVT

43405
IEYPNKWRW

43406
IFAVMADMS

43407
IFHPQWKQG

43408
IFKHCKHDE

43409
IGAGFYYKP

43410
IGEAFASTG

43411
IGLLTSLHD

43412
IGRVKCART

43413
IGVMPGIVA

43414
IHSAHHWAS

43415
IHSQMIEAP

43416
IIGRFESMS

43417
IINPLRHGD

43418
ILDPQHWSM

43419
ILTHVSDAC

43420
IMGNYIQAN

43421
IMMDWEGNW

43422
IMWVLNELE

43423
INEGIGRHF

43424
INHGTTATF

43425
INMPAVMKF

43426
IPGTRIAWA

43427
IQCYYKGEQ

43428
IRFGSREAK

43429
IRGQANSCV

43430
ISEVENWYD

43431
ISNKLTMPV

43432
ISTMIEYSN

43433
ITEYAQQDC

43434
ITHHRDNWF

43435
ITKIMRQAE

43436
IVMPVAAFH

43437
IVQCFVDSQ

TABLE 86

Sequences of the 581 to 589 Region in AAV5 VP1

Capsid Polypeptide that Drive Liver Tissue Tropism

SEQ

ID
581-589

NO
Sequence

45438
AFEKVNCPD

45439
ILIFIQNCP

45440
ASQRTHHVD

45441
ATNLINKLF

45442
EVLLGCQNY

45443
FYMLSNQGS

45444
GSVGMLDGQ

45445
GVNIMFRQQ

45446
KGEPGMNYY

45447
KIAVYADWA

45448
KNAWMDCVN

45449
KNGAGDAHY

45450
LAKMYWDMG

45451
LTQSFQTYH

45452
MIVQQSTGC

45453
NRDYYRGNY

45454
QAVQLWSHV

45455
QCGSMQGVY

45456
REAEYWRDQ

45457
SSFYAQREG

45458
VIISRHMDS

45459
VTHLFGSQA

45460
AAEAGIHQA

45461
ACQSYWNEA

45462
DATFCTYMR

45463
DKTSRLADQ

45464
KANMCEKIC

45465
KPNNFDTQF

45466
NINKFTCHE

45467
NSAIEYQNE

45468
QATPSEWEN

45469
QSLMCWNMG

45470
SARYQIEGC

45471
SIVHSGLSF

45472
TFYPTQNQH

45473
TSNWLNFFP

45474
VAAQSDTVN

45475
VFKLISNQA

45476
VVVRSNECA

45477
YHNQDRHYL

45478
HIKVEHEEV

45479
ACASTSPNA

45480
AHSRMDWTP

45481
AIGRRRLTH

45482
AKGHDKACC

45483
AMGSLMRAS

45484
ARNSMPFGM

45485
CGTGEDCED

45486
DKCNVTRAD

45487
DMSDCYEHA

45488
DNILRDVMA

45489
DVANHITQM

45490
DVDWNMGVE

45491
DVNQYTMRH

45492
DWRNKRNHA

45493
DYVFWGFPC

45494
EAAKLVEPT

45495
ECHVCTHKA

45496
ELPRYYGQM

45497
ETHRRCKMS

45498
FAKHCVMTA

45499
FDEALWKNN

45500
FTMRMASGF

45501
GTHTQQWHH

45502
HVFFAPLEG

45503
HYMGLLCGC

45504
IFDVYARWT

45505
IHIGKNDNC

45506
IHKFCFKGY

45507
ISGFTKHEW

45508
IVKQCYDCH

45509
KAANDHHVA

45510
KHVILRQLP

45511
KIVSNESMT

45512
KVDGCQKLC

45513
LMYGVGTCF

45514
LNNHHGTGS

45515
MFQWRWIDG

45516
MGFPKMSAW

45517
MQVVLQHTS

45518
MYYQIENDP

45519
NSICMGGWK

45520
NVCTKRENE

45521
NYCKYNKMR

45522
PESVKMDCN

45523
QVYYVHTER

45524
RASHDGCCD

45525
RIFWDFHAT

45526
RVPRCVYEC

45527
TDALTDSMH

45528
TEDDRWRHA

45529
TIWERQSFH

45530
TQGMYVESY

45531
TSNIFQRHG

45532
TVNVDSQYN

45533
VASSHRESQ

45534
VIKVNTVNP

45535
VSGMLAACN

45536
VVNVWGCAP

45537
WNIVLMFYC

45538
WWVVVPCFM

45539
YCTGLGSMD

45540
YFALNDITH

45541
YNLQWFKHY

45542
YSDNGRRFD

45543
YSIIWRVYR

45544
YSMDVMMSI

45545
YVHKPPDQG

45546
EVRSTRKNH

45547
LDGPIGMGT

45548
GAYSLHDWR

45549
IMVERWQHA

45550
MCDNFPSAH

45551
NGMDKPVDA

45552
AAWQSEIAN

45553
ACILDGFTL

45554
CAYNAPDWP

45555
EFTPEHRCW

45556
ETLKNIMEK

45557
GFSPISSPC

45558
GIDTISTSI

45559
GSEWKTFGD

45560
IMMCVWRSF

45561
KAFDNELHN

45562
KGEMEMSMA

45563
KKCDTMIGQ

45564
KMEHMWKMP

45565
MNGFCFGEA

45566
MVKQAQTMC

45567
NGKQFESGP

45568
NSASYCNPT

45569
NTLVNMESP

45570
TNSVMQNME

45571
TTGPKHCSV

45572
YSTYSEKSC

45573
YVETTKVWP

45574
TSERHSCMV

45575
ATAFKMLNT

45576
SPNGRGLCG

45577
AACWTVPQH

45578
AAESQGHRH

45579
AAHCIPNHD

45580
AAISSGFPA

45581
AAKVFPDGM

45582
AAMCCHLGV

45583
AAQCCTFGT

45584
AATASGYAS

45585
AAVFNDVEK

45586
AAVGINTRE

45587
AAVGSNEFA

45588
AAWMGNAVM

45589
ACAYSVASE

45590
ACCKAGFLV

45591
ACEYKVVNQ

45592
ACHNNVYHG

45593
ACKANTAQM

45594
ACKFTELSM

45595
ACLTKNWTR

45596
ADNVMHWWT

45597
ADSQSRVMP

45598
ADYHEGMFP

45599
AEHQHLACV

45600
AEMRTMGSQ

45601
AFAGTLQHP

45602
AFAWAHCPV

45603
AFMPQSWTQ

45604
AFMRNFDYP

45605
AFMSTGMLA

45606
AFMVTASNL

45607
AFNHWHYRN

45608
AFRSVVIKW

45609
AGCNRCWKQ

45610
AGDKWMASF

45611
AGGFWCEPF

45612
AGKWWWQYH

45613
AGVTLPVTL

45614
AHIAVFDCD

45615
AHLDMWWSN

45616
AHMCTQQSG

45617
AHMTEIAFA

45618
AHNMDVWCI

45619
AHPPTQYNH

45620
AHTLRQFWE

45621
AIACAHMGG

45622
AIEFHEAFT

45623
AILFLKGND

45624
AINGTGTFT

45625
AINWPMHIE

45626
AIQHPNQWC

45627
AIQNEENPG

45628
AIQTTETEE

45629
AISDKQPWR

45630
AITPCLEFK

45631
AIVEGVKHY

45632
AIVPIWTTN

45633
AIVQEDVEE

45634
AIVTASCPP

45635
AKCCSQLDY

45636
AKCYKEWWP

45637
AKDQYGLLS

45638
AKGLTHLTC

45639
AKLWRYRWI

45640
AKMLKDRAA

45641
AKMPYMSTH

45642
AKQMTAAQM

45643
AKSCEESRP

45644
AKSEFSVSI

45645
AKSGWTLME

45646
ALAHNFNGM

45647
ALDTFCRWP

45648
ALGLNSNHG

45649
ALMTWVRDC

45650
ALWYSKKEC

45651
AMASVRGGD

45652
AMENWYSYQ

45653
AMLDGMTSG

45654
AMLMSKEVA

45655
AMMQGVSNQ

45656
AMNVIASHF

45657
AMSCYTLWC

45658
AMSFTQFCS

45659
AMSPIVDAP

45660
AMSRLMMFS

45661
AMVEPQCQG

45662
AMWFCSVGD

45663
AMWRLNNVF

45664
ANCTQENQT

45665
ANECKQNVE

45666
ANHVTDTSC

45667
ANKMIVLNH

45668
ANKSVGEWR

45669
ANLFYSDRD

45670
ANLLGHYQA

45671
ANMLTNCDT

45672
ANMNCDKFS

45673
ANNVHCVKK

45674
ANNVTQAER

45675
ANQCHMGYT

45676
ANTRFPMKD

45677
APDTWVPVY

45678
APGNHNNWW

45679
APHCEKENG

45680
APISWQMAL

45681
APSMRKIIP

45682
APSNPRKVA

45683
AQAEKPCSY

45684
AQAKEWKTK

45685
AQCYKFNGH

45686
AQHTVWRFA

45687
AQLPFCQQD

45688
AQQKGGITF

45689
AQSAISVMV

45690
AQSAWHSWP

45691
ARLACCSAD

45692
ARMYYDDQP

45693
ARRLWQNDQ

45694
ARVQENHVG

45695
ASGMQGSWH

45696
ASIMQTKGI

45697
ASKPGIIFD

45698
ASMPFLSYT

45699
ASNILDLPE

45700
ASQWKIHGA

45701
ASSPFSQVR

45702
ASTCKKLAM

45703
ASYSILKQR

45704
ATAHIAPPS

45705
ATDTMQFIW

45706
ATIAQWCVH

45707
ATKLGCSRI

45708
ATLGHVMMC

45709
ATLTNQHAE

45710
ATMMKWDAN

45711
ATMWLEDQC

45712
ATRAMQVLE

45713
ATRLAFHQG

45714
ATRLITYAV

45715
ATSFTQFCS

45716
ATSGQTYSV

45717
ATSMCMHPR

45718
ATSPHLAWG

45719
ATVKFSDCV

45720
ATYKIDTSC

45721
AVAIRSWCG

45722
AVAMEFNVV

45723
AVDPMCFDE

45724
AVESICSTE

45725
AVFIPGQFW

45726
AVGMCQMDL

45727
AVLIAGAGH

45728
AVLKAEVVK

45729
AVPFPWGCC

45730
AVQLWWWHN

45731
AVRFYSTPI

45732
AVRWSQTPG

45733
AVTVVGTQI

45734
AVVAQCQSW

45735
AVVGWTAGQ

45736
AVYIMTQLQ

45737
AWAEPDMRW

45738
AWDAESCYK

45739
AWEKIHRSP

45740
AWITCLIAP

45741
AWMHDTCTK

45742
AWRSVASNH

45743
AWTHHFTLD

45744
AWWRVTHEQ

45745
AYAPSLHNN

45746
AYCEIPTNP

45747
AYDHVLESR

45748
AYEYRNHEF

45749
AYFPNSWGQ

45750
AYNRVWSCG

45751
AYPQCFQNP

45752
AYQAKYDLA

45753
AYTETSYLH

45754
AYTLCLCNE

45755
CANCPMFPT

45756
CAQFKMTYF

45757
CARQPMPPA

45758
CAYAIMPCE

45759
CCFQFELEC

45760
CCIFIWWLY

45761
CCSNKSRFV

45762
CCSPLCQQP

45763
CCVPMMNVC

45764
CCVTAWKNQ

45765
CDAIYVSCG

45766
CDCLFMLHQ

45767
CDNGYETNE

45768
CFCQGFRKR

45769
CFEHWADFE

45770
CFEKKQTDG

45771
CGEQKDQIG

45772
CGHLYNRDF

45773
CGLDYAMCQ

45774
CGRHWKEWE

45775
CGRQCKACF

45776
CHCQWMCQN

45777
CHGAHQKLE

45778
CHMPWHIHE

45779
CHMSTVMKV

45780
CHQETFLDA

45781
CHTGLMDAM

45782
CHVQFDGFG

45783
CICREGKFY

45784
CIMACQDRT

45785
CIMGMQWQF

45786
CIRHQNHYW

45787
CITAEQCGE

45788
CITPQFTSS

45789
CIVKRMECF

45790
CKAMTLAEM

45791
CKCYEMEVS

45792
CKEPHTMHE

45793
CKKPLGTRI

45794
CKNEYRIHT

45795
CKSVNPRMM

45796
CKVEWWHGN

45797
CKYEISSGL

45798
CLVSCPALN

45799
CLYVPLFAD

45800
CMEGGLMDW

45801
CMEMMKWAN

45802
CMFWANCSQ

45803
CMGSGFAEL

45804
CMKMWDCGK

45805
CMMPVLEEC

45806
CMRWFLLGS

45807
CNEVPSVMP

45808
CNSQKSYAH

45809
CNSWNCAYA

45810
CPDVGFQSG

45811
CPHLFVFDW

45812
CPPSNNPCE

45813
CPSRTWAIE

45814
CPTIIVKDW

45815
CPVANTTHP

45816
CQEFNSHTD

45817
CQNFSSLPW

45818
CQSVSANHI

45819
CRARRNPRL

45820
CRIRKRHIT

45821
CSCDEMEID

45822
CSDACQAMV

45823
CSDMLTKCN

45824
CSEHENLGI

45825
CSEKFNLHC

45826
CSETQVSTG

45827
CSQPHQMAS

45828
CSQRCMSFG

45829
CSRSDLLCG

45830
CSVWCIYYR

45831
CSYPAHRFH

45832
CTEVMAMGT

45833
CTFSSDKSD

45834
CTKLVNWYT

45835
CTLRCIMHT

45836
CTMRSGVGT

45837
CTPTILRGV

45838
CTQSQCHLC

45839
CTTEVRGTP

45840
CTTMKMNQA

45841
CTWFIQGPK

45842
CTYLAKLKC

45843
CVAAWGENG

45844
CVDNPFVLV

45845
CVFMGMMWD

45846
CVGRWWNFY

45847
CVHISDFGD

45848
CVIQGHSIE

45849
CVMAIESRG

45850
CVNRQHRIN

45851
CVPASGGWD

45852
CVTMKEFAM

45853
CWRQYVAGQ

45854
CYCTCEQPD

45855
CYELTMPLD

45856
DADRNNMSV

45857
DAEMLCYDY

45858
DAFWSNLHL

45859
DAHWGALSV

45860
DAIPKKWYG

45861
DALANNESR

45862
DALSMLRAP

45863
DALTNRASH

45864
DASMMWSRG

45865
DASYEYQSL

45866
DCAQTGEYG

45867
DCKNHYQSS

45868
DCMWYNRID

45869
DCPRSKMMT

45870
DCPTYYWRD

45871
DCSCHELAL

45872
DCYHICVIP

45873
DDELCPQCN

45874
DDHQSTWMN

45875
DDRCVQAAK

45876
DFAVSTSFD

45877
DFCMKEKAV

45878
DGCVHNQAI

45879
DGPICKGIY

45880
DGTHPCWNG

45881
DHCKTALPN

45882
DHERQRWLH

45883
DHIRQHTCS

45884
DHLKIRGAY

45885
DHQKRNVIQ

45886
DHQLVLVSG

45887
DHTVLFEGQ

45888
DHWITGPCR

45889
DHYLGHASY

45890
DICWKGWCF

45891
DIDRFTDVQ

45892
DIECKEIAI

45893
DIEITMQEN

45894
DIGPQDHWP

45895
DIHWMDHDF

45896
DIIEKVNEM

45897
DILERMNRA

45898
DINMNAQER

45899
DIRKDQHKN

45900
DIVIKKCNA

45901
DKASNQHNG

45902
DKFAAHALA

45903
DKGHHNSMH

45904
DKIDFSSVM

45905
DKKFGQFMS

45906
DKKWEEICA

45907
DKLELIGLD

45908
DKNFWVHWF

45909
DKTEMVIID

45910
DLASVGRWN

45911
DLCKMKEDK

45912
DLFRQERLF

45913
DLINWWHLQ

45914
DLKPWNPAW

45915
DLRLKVISR

45916
DLSRSFLAQ

45917
DLVIIYMFH

45918
DMELQFMYA

45919
DMGNYPCFE

45920
DMMLRHCVF

45921
DMRSNLPWH

45922
DMSHECVDA

45923
DMSHECVGA

45924
DMVDLEMYP

45925
DMWDEGDGD

45926
DNADMFNIG

45927
DNAKRPMTI

45928
DNENAIPMG

45929
DNGNMFQQH

45930
DNKLTEPCN

45931
DNKYYMSKC

45932
DNMEFESPG

45933
DNNAMINCT

45934
DNPVTRCTD

45935
DNSWFVQGG

45936
DNVYTMHQW

45937
DPMYIPVNH

45938
DQCLCHTQA

45939
DQGLWENIP

45940
DQYFEGWRK

45941
DRCVMTDVL

45942
DRFMEFTGA

45943
DRFWSKYKD

45944
DRGMPTNWC

45945
DRGQSFQMF

45946
DRGTVNRDC

45947
DRSILHAHP

45948
DRTIIQNIP

45949
DRTSYPLHL

45950
DSAWYYWHP

45951
DSCHSRIIL

45952
DSELVRDYI

45953
DSIGLERGG

45954
DSKIAHGRQ

45955
DSLRKYQCR

45956
DSLTVNMAH

45957
DSMAAHCAV

45958
DSNGIAHVP

45959
DSNKWAGLG

45960
DSNTHLPIG

45961
DSPQIVDVY

45962
DSQGINKLH

45963
DSRHSTFCL

45964
DSRHYANFM

45965
DSTPDRSIW

45966
DSTRIITQY

45967
DSYTMNWFY

45968
DTAAEVNMQ

45969
DTAEYRAGY

45970
DTDCARAIG

45971
DTDRQVKQS

45972
DTFEYTNAF

45973
DTKEYKIAY

45974
DTLQRACML

45975
DTLWYVNFA

45976
DTMMMPSSE

45977
DTMTTCYNS

45978
DTQNMKSSK

45979
DTRWWGMYA

45980
DTSSHECCD

45981
DTVHSGWQD

45982
DTVNQQKTW

45983
DVCKKGQCV

45984
DVCSQGQVP

45985
DVEMMIQRA

45986
DVEYYDHCF

45987
DVGYSAVWA

45988
DVKWGTSNL

45989
DVLPMEWCV

45990
DVLWGNRDS

45991
DVMAMAPSC

45992
DVNAGTRFP

45993
DVTDCKWGF

45994
DVWYPEFIG

45995
DWAHWAAYP

45996
DWCTDMTRK

45997
DWHNLVEYP

45998
DWLNLGRYM

45999
DYFREHMCP

46000
DYKLRARNA

46001
DYNGLSVPI

46002
DYYQWDGQI

46003
EADRWSSPN

46004
EAGRQAWTD

46005
EAHLDWIYP

46006
EAHWFTNYN

46007
EAKPWSGYL

46008
EAKRYHVDG

46009
EANMKEGLI

46010
EATIKALVF

46011
EATWTHADE

46012
ECDPRQMMC

46013
ECHQGSNME

46014
ECMDQQATW

46015
ECMQQCFVR

46016
ECQPFATVT

46017
ECVYNPANN

46018
EEKYKDAQW

46019
EEPGSQMTV

46020
EERPWKRDL

46021
EETEGIEPH

46022
EFDAAVIEK

46023
EFGYEHIVP

46024
EFRNSSCTN

46025
EGAHNKFDR

46026
EGCIPGGWS

46027
EGGSCNTQM

46028
EGNFIMGCS

46029
EGNQEGLNM

46030
EGNREGLNM

46031
EGNYMAGLG

46032
EGWHFPAME

46033
EHDGTPFNE

46034
EHDHQPSYA

46035
EHERFCQGR

46036
EHHRRGAFK

46037
EHMFNKDCT

46038
EHQRGHACN

46039
EHSYLLHDT

46040
EICSEINMK

46041
EIEQEEGGC

46042
EIGLYWIKC

46043
EIGYKQMYF

46044
EIKASHNYD

46045
EINQGDEDG

46046
EIQSVMPQP

46047
EISNLRQGF

46048
EISPHKILP

46049
EITGGHASW

46050
EIVWYFQDT

46051
EKAMRNGQA

46052
EKCCCLQWE

46053
EKEIKMLFG

46054
EKGPYWHQH

46055
EKHAMECVS

46056
EKIAPNHQA

46057
EKKMWVYTG

46058
EKLMAYIML

46059
EKLPIQMFN

46060
EKMPWLISN

46061
EKNWYQHIW

46062
EKQTRNQDG

46063
ELHHPIRRS

46064
ELMASNRCH

46065
ELMFVMHGT

46066
ELPMMNMAQ

46067
ELSFKTAQG

46068
ELTMANCMN

46069
EMERKNMFE

46070
EMGCDNKSF

46071
EMHLYHICH

46072
EMTSTMKSH

46073
EMYRALGHG

46074
ENGPMCDSN

46075
ENHSPNVDP

46076
ENILAIPAL

46077
ENKWQPKQA

46078
ENNSRQLIN

46079
EPCMRNLFG

46080
EPIRTEGAE

46081
EPLCHQIMP

46082
EPLVDEFGG

46083
EPRECFHIE

46084
EPWVMKLNK

46085
EQADYRAVQ

46086
EQERYEWDP

46087
EQGCQLVTT

46088
EQHPRMNFH

46089
EQIHQLRFG

46090
EQKWQWCQY

46091
EQQMLLQLS

46092
EQYMVNTRD

46093
ERCPWSVGE

46094
ERENSLWMS

46095
ERFLLEAVE

46096
ERGKWHALS

46097
ERQPFTMQW

46098
ERSWTHAHW

46099
ERTTKAEGY

46100
ERVARLQFT

46101
ESAKFVCEY

46102
ESCEDGIRT

46103
ESGFRKLRV

46104
ESHMHDHWI

46105
ESILVCATP

46106
ESIQLRSSQ

46107
ESSFVHEEQ

46108
ESTEPMSAT

46109
ESTLKTTYA

46110
ETCTEPKYW

46111
ETEMTSWQN

46112
ETKAVISAQ

46113
ETLERDWDY

46114
ETMKDVRFM

46115
ETNQLSGYC

46116
ETPKFNWSN

46117
ETRTDVPML

46118
ETTAAFKPG

46119
ETYDNNLQA

46120
ETYVCWFGS

46121
EVATSYARV

46122
EVDQAPMMI

46123
EVHRSELQR

46124
EVIGFLYHA

46125
EVMAMCDRY

46126
EVMPMCDRY

46127
EVQLTYLHE

46128
EVVTKHDSR

45129
EWCKQWMRF

46130
EWMTCGVSG

46131
EWSMLQYNH

46132
EWYIANISD

46133
EYELTQPAP

46134
EYIFENEMC

46135
EYLLCRGEA

46136
FADHFEAMF

46137
FAEHEIEHS

46138
FAIIRNDSH

46139
FAIMSSDHP

46140
FALNTEISA

46141
FAPFLKAGH

46142
FAQMNKDRC

46143
FARHPLKCA

46144
FASQHAVQA

46145
FCFWMPTTN

46146
FCGKLTHVG

46147
FDWRCGCWS

46148
FEQPPMHHQ

46149
FEVTMTVPE

46150
FFQRFLSAW

46151
FGAMTSDTI

46152
FGAYYNDRG

46153
FGETYNHSH

46154
FGEWITFYF

46155
FGHKIEAPH

46156
FGQHVWQGF

46157
FGRYFQFDL

46158
FGWWYKTFT

46159
FHAPTECMA

46160
FHFQFTMDK

46161
FHHLFDCNL

46162
FHLACIHAQ

46163
FHMTRYKMT

46164
FHRPSEGQE

46165
FIRLFMKLN

46166
FKECTNGMH

46167
FKMATAECP

46168
FKPCKDDDV

46169
FKVTEYELP

46170
FMDEIAVTM

46171
FMGFMGVMG

46172
FMQIWNKYS

46173
FMRMEMMGQ

46174
FMRPFLEPD

46175
FNFCFNHQF

46176
FNGIDQMRY

46177
FNMLLQVNR

46178
FNTTCQMSI

46179
FPICNQERE

46180
FPQRPCTYA

46181
FPTNQNDPH

46182
FPTQMHYNA

46183
FQAKMHTLA

46184
FQDWPVTWC

46185
FQQQTGTRC

46186
FRFSTQHEA

46187
FRLNLLKTN

46188
FSEVTLQES

46189
FSKTGHCKF

46190
FSQCPETCC

46191
FSQGPGMHF

46192
FSQHQEHMR

46193
FSQRVEKTW

46194
FSSEAADYD

46195
FSTFTKPSP

46196
FSVPLGATT

46197
FSVSAQHSC

46198
FTFRVGEVN

46199
FTHGSVLHD

46200
FTMLGMEAI

46201
FTNPHAFYP

46202
FTQHIVGET

46203
FYIKYVYFD

46204
FVDHYYGHQ

46205
FVNHTRCNP

46206
FVRHDVIGQ

46207
FVTHWEYYP

46208
FVTMMQIAF

46209
FVTMRYEGH

46210
FWTSYTTNP

46211
FYGQNYWWY

46212
FYPRSWPQA

46213
GAARYGYNI

46214
GADSSGGWN

46215
GAFIRDFDI

46216
GAGSVQEIH

46217
GAGTTMRPD

46218
GAHWWGGCV

46219
GAIPLLNWL

46220
GAKMQIAFW

46221
GAKQHMPCY

46222
GAVNSEVCE

46223
GCCWSYYCG

46224
GCDTTKFDA

46225
GCFIYWKDS

46226
GCVKDRCWM

46227
GCWNERCQN

46228
GCYWACWLT

46229
GDNYTAWVG

46230
GEFMMFDRG

46231
GEGWNDDLG

46232
GEREYWNWE

46233
GFACLDHFD

46234
GFDIVLKDF

46235
GFELNMRIS

46236
GFGNTFCIT

46237
GFGTVSMER

46238
GFGYMWCKG

46239
GFLLTYGED

46240
GFMIGNQYL

46241
GFRVIISNW

46242
GFSCLICFD

46243
GFTQTCMTN

46244
GGLLYIECY

46245
GGTIDGHVG

46246
GGVKFNNWW

46247
GHMFVNIDQ

46248
GHMMMMECW

46249
GHSLTMRAH

46250
GHTARDIHA

46251
GHTHSQPAA

46252
GHTPGGSFY

46253
GIANTSDSY

46254
GICCILWDR

46255
GIDSMVIQR

46256
GIHEIMVQR

46257
GIHMVTAMS

46258
GIKFTQQDL

46259
GILRQIEKP

46260
GISICGQNW

46261
GKDTPNSFP

46262
GKGEVLAVA

46263
GKGTMEFGN

46264
GKGYGQHDG

46265
GKILWMSAR

46266
GKMQVLHSA

46267
GKNMLIDGG

46268
GKPYTQIIR

46269
GKSTCKRDV

46270
GLEMRNQAM

46271
GLFQQAMQG

46272
GLGCLMQGN

46273
GLSVGQRIE

46274
GLYERLACC

46275
GMHYSQCQH

46276
GMKAVFIGD

46277
GMWVNHGAK

46278
GNEHCVKYA

46279
GNEKIERYD

46280
GNEMPNYGG

46281
GNHVNYNMH

46282
GPEKLTWGP

46283
GPPHYRNPE

46284
GQFIFDEFV

46285
GQGFMLWNG

46286
GQGKNWVSW

46287
GQGMCDRKT

46288
GQLNCMKGS

46289
GQQENPVFT

46290
GQTSPLMYP

46291
GRAPMCAAA

46292
GRLVVHQFK

45293
GRNMFLWWW

46294
GRSLSQSEW

46295
GRYQSSQFC

46296
GSASHNVWY

46297
GSDYWLCDK

46298
GSFYAQYEE

46299
GSFYCQWNN

46300
GSGKMWQED

46301
GSLLFNPEE

46302
GSMKEMMMR

46303
GSMVASSWC

46304
GSNQRDCSR

46305
GSRPTPADW

46306
GSSHYMEFY

46307
GSYTKPTNV

46308
GTAHNVFMG

46309
GTCLDQTGC

46310
GTETHCDFE

46311
GTGAFTQVP

46312
GTIAFSDNN

46313
GTIPMNCFW

46314
GTKMFQESI

46315
GTMFWGWTG

46316
GTNNIINVR

46317
GTQHFWMDH

46318
GTRWIHDMA

46319
GTSQEKRGQ

46320
GVAAVGLYK

46321
GVIVPISRS

46322
GVLKQANQQ

46323
GVQNTALPR

46324
GVTSPVGTC

46325
GVVDNQQCL

46326
GVWPHYENK

46327
GWCITKARN

46328
GWDHFMQNT

46329
GWMDYIAAH

46330
GWYNWMIQY

46331
GYESKNTWY

46332
GYEWVQEIE

46333
GYFPNHWCK

46334
GYHCTGAHS

46335
GYICEGTCH

46336
GYNERFEHK

46337
GYNPMMSAA

46338
GYSQKPRTP

46339
GYTYNQQVC

46340
HADAENQVM

46341
HAIPHTMWH

46342
HAIQEGTVP

46343
HAIYWIPQV

46344
HAMATECGP

46345
HAMLSHGCC

46346
HANESISMT

46347
HASYQSSFG

46348
HATSQELCQ

46349
HATTFEWDS

46350
HAYALAVEH

46351
HCCSANNSP

46352
HCELISRIA

46353
HCELLPVNE

46354
HCFMAHCSY

46355
HCMNWMING

46356
HCQILPHWA

46357
HCVNEHAWE

46358
HDGFPWADE

46359
HERNWPFAC

46360
HESCLKLAG

46361
HEWAGELQT

46362
HEYGFGEAR

46363
HFANICVRG

46364
HFEHLWCQS

46365
HFGHYLGMF

46366
HFISHDYHN

46367
HFMPNNFLA

46368
HFVNSYCWW

46369
HGAQFARCS

46370
HGNEVDHSM

46371
HHAVMNKMD

46372
HHEVTTKFQ

46373
HHFHSGCCW

46374
HHPPELDRM

46375
HHTDRNGGD

46376
HHVLKGDMA

46377
HIDGATVSG

46378
HIKWDCWCP

46379
HINAFDNIR

46380
HISWWPACG

46381
HITTLYMEA

46382
HKLVAHERH

46383
HKPMCDRVF

46384
HLMGLCAMS

46385
HLQSISHCH

46386
HLVGMSLTQ

46387
HMEMPYKLG

46388
HMKCDVWQP

46389
HMLKKEDMN

46390
HMTTVNLQY

46391
HNGKSVLEW

46392
HNKMTYNYE

46393
HNNMFFWDR

46394
HNRAHENDK

46395
HNSVVNDHE

46396
HNTQNGFAD

46397
HNWYKRIVQ

46398
HPGIDLVCG

46399
HQCFWIYGS

46400
HQFQFYQWM

46401
HQPPIMNQG

46402
HRCTMQPDI

46403
HRCTPYNRD

46404
HRDHDITDF

46405
HRFSPPMET

46406
HRHFEMSVI

46407
HRMVIRKSQ

46408
HRVQCDEAP

46409
HSCTSGNPG

46410
HSESIDVWS

46411
HSETQRHQM

46412
HSGPWSNCE

46413
HSMQTKEDW

46414
HSSALFDME

46415
HSTQAGGMY

46416
HSTTACVHN

46417
HSVMWGKEN

46418
HTDKWWRDI

46419
HTEKGGEQA

46420
HTIHGAGLI

46421
HTIPVPMAA

46422
HTNTCLFEE

46423
HTQIIGGSD

46424
HTQWFNMGE

46425
HVDQYYRSA

46426
HVDTEKGRE

46427
HVELSQLHS

46428
HVEWVNAEK

46429
HVLPVCYRM

46430
HVMFPWGMW

46431
HVMYQHWMM

46432
HVNIEHQGT

46433
HVSDCCKSQ

46434
HVVKYPLSG

46435
HVYRGNGME

46436
HWDECNYVG

46437
HWDQFNGQF

Example 46
Treatment of Spinal Muscular Atrophy (SMA) with an AAV5-Derived Virion Encapsidating a Therapeutic Payload

This example describes treatment of spinal muscular atrophy (SMA) with a variant AAV5-derived virion having any one of the engineered CNS tropic variant AAV5 VP capsid polypeptides disclosed herein, wherein the variant AAV5 virion encapsidates a therapeutic payload. Polynucleotide sequence encoding for AAV Rep, an AAV5-derived variant Cap and helper proteins and a therapeutic payload are transfected in cells to produce variant AAV5 virions, where the polynucleotide sequence encoding for the variant AAV5 Cap comprises at least one mutation in a region from a position corresponding to 581 to a position corresponding to 589 in the VP1 capsid polypeptide and where the polynucleotide sequence encoding for the variant AAV5 Cap encodes for an amino acid sequence of any one of SEQ ID NO: 34991-SEQ ID NO: 37437. The therapeutic payload is a guide RNA targeting an mRNA encoded for by a gene implicated in SMA or a transgene. The mRNA targeted by the guide RNA is an mRNA encoded for by a gene encoding for survival motor neuron (SMN) protein. The transgene is SMN. The variant AAV5 virion encapsidating the payload is administered to a subject. The subject is a human or non-human animal. The route of administration is a systemic route of administration. The systemic route of administration is intravenous administration. Upon administration to the subject, the variant AAV5 virions encapsidating the therapeutic payload exhibit enhanced spinal cord tropism as compared to wildtype AAV5 virions encapsidating the therapeutic payload. Upon administration to the subject, a lower dose of the variant AAV5 virions encapsidating the therapeutic payload is administered as compared to wildtype AAV5 virions encapsidating the therapeutic payload. Upon administration to the subject, at least one symptom of SMA is alleviated or the subject is cured.

2. Certain Sequences

>AAV5

VP1, from Chiorini et al., J. Virol. 73(2): 1309-1319 (1999)

[SEQ ID NO: 1]

MSFVDHPPDWLEEVGEGLREFLGLEAGPPKPKPNQQHQDQARGLVLPGYN

YLGPGNGLDRGEPVNRADEVAREHDISYNEQLEAGDNPYLKYNHADAEFQ

EKLADDTSFGGNLGKAVFQAKKRVLEPFGLVEEGAKTAPTGKRIDDHFPK

RKKLRTEEDSKPSTSSDAEAGPSGSQQLQIPAQPASSLGADTMSAGGGGP

LGDNNQGADGVGNASGDWHCDSTWMGDRVVTKSTRTWVLPSYNNHQYREI

KSGSVDGSNANAYFGYSTPWGYFDFNREHSHWSPRDWQRLINNYWGFRPR

SLRVKIFNIQVFEVTVQDSTTTIANNLTSTVQVFTDDDYQLPYVVGNGTE

GCLPAFPPQVFTLPQYGYATLNRDNTENPTERSSFFCLEYFPSKMLRTGN

NFEFTYNFEEVPFHSSFAPSQNLFKLANPLVDQYLYRFVSTNNTGGVQFN

KNLAGRYANTYKNWFPGPMGRTQGWNLGSGVNRASVSAFATTNRMELEGA

SYQVPPQPNGMTNNLQGSNTYALENTMIFNSQPANPGTTATYLEGNMLIT

SESETQPVNRVAYNVGGQMATNNQSSTTAPATGTYNLQEIVPGSVWMERD

VYLQGPIWAKLPETGAEFHPSPAMGGEGLKHPPPMMLIKNIPVPGNITSF

SDVPVSSFITQYSTGQVTVEMEWELKKENSKRWNPEIQYTNNYNDPQFVD

FAPDSTGEYRTTRPIGTRYLTRPL

>AAV5 Library scaffold

[SEQ ID NO: 2]

MSFVDHPPDWLEEVGEGLREFLGLEAGPPKPKPNQQHQDQARGLVLPGYN

YLGPGNGLDRGEPVNRADEVAREHDISYNEQLEAGDNPYLKYNHADAEFQ

EKLADDTSFGGNLGKAVFQAKKRVLEPFGLVEEGAKTAPTGKRIDDHFPK

RKKARTEEDSKPSTSSDAEAGPSGSQQLQIPAQPASSLGADTMSAGGGGP

LGDNNQGADGVGNASGDWHCDSTWMGDRVVTKSTRTWVLPSYNNHQYREI

KSGSVDGSNANAYEGYSTPWGYFDFNRFHSHWSPRDWQRLINNYWGFRPR

SLRVKIFNIQVKEVTVQDSTTTIANNLTSTVQVFTDDDYQLPYVVGNGTE

GCLPAFPPQVFTLPQYGYATLNRDNTENPTERSSFFCLEYFPSKMLRTGN

NFEFTYNFEEVPFHSSFAPSQNLFKLANPLVDQYLYRFVSTNNTGGVQFN

KNLAGRYANTYKNWFPGPMGRTQGWNLGSGVNRASVSAFATTNRMELEGA

SYQVPPQPNGMTNNLQGSNTYALENTMIFNSQPANPGTTATYLEGNMLIT

SESETQPVNRVAYNVGGQMATNNQSSTTAP

Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, Xaa9

IVPGSVWMERDVYLQGPIWAKIPETGAHFHPSPAMGGFGLKEPPPMMLIK

NTPVPGNITSFSDVPVSSFITQYSTGQVTVEMEWELKKENSKRWNPEIQY

TNNYNDPQFVDFAPDSTGEYRTTRPIGTRYLTRPL

Xaa1, Xaa2, Xaa3, Xaa4, Xaa5, Xaa6, Xaa7, Xaa8, and Xaa9 are

each independently selected from A, R, N, D, C, E, Q, G, H, I,

L, K, M, F, P, S, T, W, Y, and V

SEQ ID NO: 3

MSFVDHPPDWLEEVGEGLREFLGLEAGPPKPKPNQQHQDQARGLVLPGYN

YLGPGNGLDRGEPVNPADEVAREHDISYNEQLEAGDNPYLKYNHADAEFQ

EKLADDTSFGGNLGKAVFQAKKRVLEPFGLVEEGAKTAPTGKRIDDHFPK

RKKLRTEEDSKPSTSSDAEAGPSGSQQLQIPAQPASSLGADTMSAGGGGP

LGDNNQGADGVGNASGDWHCDSTWMGDRVVTKSTRTWVLPSYNNHQYREI

KSGSVDGSNANAYFGYSTPWGYFDFNRFHSHWSPRDWQRLINNYWGFRPR

SLRVKIFNIQVKEVTVQDSTTTIANNLTSTVQVFTDDDYQLPYVVGNGTE

GCLPAFPPQVFTTYQYGYATLNRDNTENPTERSSFFCLEYFPSKMLRTGN

NFEFTYNFEEVPFHSSFAPSQNLFKLANPLVDQYLYRFVSTNNTGGVQFN

KNLAGRYANTYKNWFPGPMGRTQGWNLGSGVNRASVSAFATTNRMELEGA

SYQVPPQPNGMTNNLQGSNTYALENTMIFNSQPANPGTTATYLEGNMLIT

SESETQPVNRVAYNVGGQMATNNQSSTTAPATGT

Xaa5

NLQE
IVPGSVWMERDVYLQGPIWAKIPETGAHFHPSPAMGGFGLKHPPPMM

LIKNTPVPGNITSFSDVPVSSFITQYSTGQVTVEMEWELKKENSKRWNPEI

QYTNNYNDPQFVDFAPDSTGEYRTTRPIGTRYLTRPL

Wherein Xaa5 is V (Y585V)

SEQ ID NO: 4

MSFVDHPPDWLEEVGEGLREFLGLEAGPPKPKPNQQHQDQARGLVLPGYN

YLGPGNGLDRGEPVNRADEVAREHDISYNEQLEAGDNPYLKYNHADAESQ

EKLADDTSFGGNLGKAVFQAKKRVLEPFGLVEEGAKTAPTGKRIDDHFPK

RKKARTEEDSKPSTSSDAEAGPSGSQQLQIPAQPASSLGADTMSAGGGGP

LGDNNQGADGVGNASGDWHCDSTWMGDRVVTKSTRTWVLPSYNNHQYREI

KSGSVDGSNANAYFGYSTPWGYFDFNRFHSHWSPRDWQRLINNYWGFRPR

SLRVKIFNIQVKEVTVQDSTTTIANNLTSTVQVFTDDDYQLPYVVGNGTE

GCLPAFPPQVFTLPQYGYATLNRDNTENPTERSSFFCLEYFPSKMLRTGN

NFEFTYNFEEVPFHSSFAPSQNLFKLANPLVDQYLYRFVSTNNTGGVQFN

KNLAGRYANTYKNWFPGPMGRTQGWNLGSGVNRASVSAFATTNRMELEGA

SYQVPPQPNGMTNNLQGSNTYALENTMIFNSQPANPGTTATYLEGNMLIT

SESETQPVNRVAYNVGGQMATNNQSSTTAPATGTYN

Xaa7

QE
IVPGSVWMERDVYLQGPIWAKIPETGAHFHPSPAMGGFGLKHPPPMMLI

KNTPVPGNITSFSDVPVSSFITQYSTGQVTVEMEWELKKENSKRWNPEIQY

TNNYNDPQFVDFAPDSTGEYRTTRPIGTRYLTRPL

Wherein Xaa7 is T (L587T)

SEQ ID NO: 5

MSFVDHPPDWLEEVGEGLREFLGLEAGPPKPKPNQQHQDQARGLVLPGYN

YLGPGNGLDRGEPVNRADEVAREHDISYNEQLEAGDNPYLKYNHADAEFQ

EKLADDTSFGGNLGKAVFQAKKRVLEPFGLVEEGAKTAPTGKRIDDHFPK

RKKARTEEDSKPSTSSDAEAGPSGSQQLQIPAQPASSLGADTMSAGGGGP

LGDNNQGADGVGNASGDWHCDSTWMGDRVVTKSTRTWVLPSYNNHQYREI

KSGSVDGSNANAYFGYSTPWGYFDFNRFHSHWSPRDWQRLINNYWGFRPR

SLRVKIFNIQVKEVTVQDSTTTIANNLTSTVQVFTDDDYQLPYVVGNGTE

GCLPAFPPQVFTLPQYGYATLNRDNTENPTERSSFFCLEYFPSKMLRTGN

NFEFTYNFEEVPFHSSFAPSQNLFKLANPLVDQYLYRFVSTNNTGGVQFN

KNLAGRYANTYKNWFPGPMGRTQGWNLGSGVNRASVSAFATTNRMELEGA

SYQVPPQPNGMTNNLQGSNTYALENTMIFNSQPANPGTTATYLEGNMLIT

SESETQPVNRVAYNVGGQMATNNQSSTTAPXaa1TGT

YNLQE
IVPGSVWMERDVYLQGPIWAKIPETGAHFHPSPAMGGFGLKHPPPMM

LIKNTPVPGNITSFSDVPVSSFITQYSTGQVTVEMEWELKKENSKRWNPEI

QYTNNYNDPQFVDFAPDSTGEYRTTRPIGTRYLTRPL

Wherein Xaa1 is T (A581T)

SEQ ID NO: 6

MSFVDHPPDWLEEVGEGLREFLGLEAGPPKPKPNQQHQDQARGLVLPGYN

YLGPGNGLDRGEPVNRADEVAREHDISYNEQLEAGDNPYLKYNHADAEFQ

EKLADDTSFGGNLGKAVFQAKKRVLEPFGLVEEGAKTAPTGKRIDDHFPK

RKKLRTEEDSKPSTSSDAEAGPSGSQQLQIPAQPASSLGADTMSAGGGGP

LGDNNQGADGVGNASGDWHCDSTWMGDRVVTKSTRTWVLPSYNNHQYREI

KSGSVDGSNANAYFGYSTPWGYFDFNRFHSHWSPRDWQRLINNYWGFRPR

SLRVKIFNIQVKEVTVQDSTTTIANNLTSTVQVFTDDDYQLPYVVGNGTE

GCLPAFPPQVFTLPQYGYATLNRDNTENPTERSSFFCLEYFPSKMLRTGN

NFEFTYNFEEVPFHSSFAPSQNLFKLANPLVDQYLYRFVSTNNTGGVQFN

KNLAGRYANTYKNWFPGPMGRTQGWNLGSGVNRASVSAFATTNRMELEGA

SYQVPPQPNGMTNNLQGSNTYALENTMIFNSQPANPGTTATYLEGNMLIT

SESETQPVNRVAYNVGGQMATNNQSSTTAPA

Xaa

GTYNLQE
IVPGSVWMERDVYLQGPIWAKIPETGAHFHPSPAMGGFGLKHPP

PMMLIKNTPVPGNITSFSDVFVSSFITQYSTGQVTVEMEWELKKENSKRWN

PEIQYTNNYNDPQFVDFAPDSTGEYRTTRPIGTRYLTRPL

Wherein Xaa2 is A (T582A)

SEQ ID NO: 7

MSFVDHPPDWLEEVGEGLREFLGLEAGPPKPKPNQQHQDQARGLVLPGYN

YLGPGNGLDRGEPVNRADEVAREHDISYNEQLEAGDNPYLKYNHADAEFQ

EKLADDTSFGGNLGKAVFQAKKRVLEPFGLVEEGAKTAPTGKRIDDHFPK

RKKARTEEDSKPSTSSDAEAGPSGSQQLQIPAQPASSLGADTMSAGGGGP

LGDNNQGADGVGNASGDWHCDSTWMGDRVVTKSTRTWVIPSYNNHQYREI

KSGSVDGSNANAYFGYSTPWGYFDFNRFHSHWSPRDWQRLINNYWGFRPR

SLRVKIFNIQVKEVTVQDSTTTIANNLTSTVQVFTDDDYQLPYVVGNGTE

GCLPAFPPQVFTLPQYGYATLNRDNTENPTERSSFFCLEYFPSKMLRTGN

NFEFTYNFEEVPFHSSFAPSQNLFKLANPLVDQYLYRFVSTNNTGGVQFN

KNLAGRYANTYKNWFPGPMGRIQGWNLGSGVNHASVSAFATTNRMELEGA

SYQVPPQPNGMTNNLQGSNTYALENTMIFNSQPANPGTTATYLEGNMLIT

SESETQPVNRVAYNVGGQMATNNQSSTTAPATG

Xaa4

YNLQE
IVPGSVWMERDVYLQGPIWAKIPETGAHFHPSPAMGGFGLKHPPP

MMLIKNTPVPGNITSFSDVPVSSFITQYSTGQVTVEMEWELKKENSKRWN

PEIQYTNNYNDPQFVDFAPDSTGEYRTTRPIGTRYLTRPL

Wherein Xaa4 is A (T584A)

SEQ ID NO: 8

MSFVDHPPDWLEEVGEGLREFLGLEAGPPKPKPNQQHQDQARGLVLPGYN

YLGPGNGLDRGEPVNRADEVAREHDISYNEQLEAGDNPYLKYNHADAEFQ

EKLADDTSFGGNLGKAVFQAKKRVLEPFGLVEEGAKTAPTGKRIDDHFPK

RKKARTEEDSKPSTSSDAEAGPSGSQQLQTPAQPASSLGADTMSAGGGGP

LGDNNQGADGVGNASGDWHCDSTWMGDRVVTKSTRTWVLPSYNNHQYREI

KSGSVDGSNANAYFGYSTPWGYFDFNRFHSHWSPRDWQRLINNYWGFRPR

SLRVKIFNIQVKEVTVQDSTTTIANNLTSTVQVFTDDDYQLPYVVGNGTE

GCLPAFPPQVFTLPQYGYATLNRDNTENPTERSSFFCLEYFPSKMLRTGN

NFEFTYNFEEVPFHSSFAPSQNLFKLANPLVDQYLYREVSINNTGGVQFN

HNLAGRYANTYKNWFPGPMGRTQGWNLGSGVNRASVSAFATTNRMELEGA

SYQVPPQPNGMTNNLQGSNTYALENTMIFNSQPANPGTTATYLEGNMLIT

SESETQPVNRVAYNVGGQMAINNQSSTAPATGT

Xaa5

NXaa7QE
IVPGSVWMERDVYLQGPIWAKIPETGAHFHPSPAMGGFGLKHPPPMM

LIKNTPVPGNITSFSDVPVSSFITQYSTGQVTVEMEWELKKENSKRWNPEI

QYTNNYNDPQFVDFAPDSTGEYRTTRPIGTRYLTRPL

Wherein Xaa5 is V (Y585V) and Xaa7 is T (L587T)

(581-589 sialic acid binding region of AAV5 VP1 capsid and

reference for Xaa1-Xaa9)

SEQ ID NO: 9

ATGTYNLQE

(FIG. 10, Middle Variant DNA)

SEQ ID NO: 111

AATCCAGCTATGTTCAATTGCGATTAC

(FIG. 10, Middle Variant; amino acid)

SEQ ID NO: 112

NPAMFNCDY

(FIG. 10, Right Variant, DNA)

SEQ ID NO: 113

AAGTTGACCGCCCACATCTACGCCTTG

(FIG. 10, Right Variant; amino acid)

SEQ ID NO: 114

KLTAHIYAL

SEQ ID NO: 115-1114 (TABLE 8, wherein the listed AAs represent

Xaa1-Xaa9 of SEQ ID NO: 2)

AAV5 VP2 sequence

(SEQ ID NO: 1115)

TAPTGKRIDDHFTKRKKARTEEDSKPSTSSDAEAGPSGSQQLQIPAQPASSLGADTMSAGGG

GPLGDNNQGADGVGNASGDWHCDSTWMGDRVVTKSTRTWVLPSYNNHQYREIKSGSVDGSNA

NAYFGYSTPWGYFDFNRFHSHWSPRDWQRLINNYWGFRPRSLRVKIFNIQVKEVTVQDSTTT

IANNLTSTVQVFTDDDYQLPYVVGNGTEGCLPAFPPQVFTLPQYGYATLNRDNTENPTERSS

FFCLEYFPSKMLRTGNNFEFTYNFEEVPFHSSFAPSQNLFKLANPLVDQYLYRFVSTNNTGG

VQFNKNLAGRYANTYKNWFPGPMGRTQGWNLGSGVNRASVSAFATTNRMELEGASYQVPPQP

NGMTNNLQGSNTYALENTMIFNSQPANPGTTATYLEGNMLITSESETQPVNRVAYNVGGQMA

TNNQSSTTAPATGTYNLQEIVPGSVWMERDVYLQGPIWAKIPETGAHFHPSPAMGGFGLKHP

PPMMLIKNTPVPGNITSFSDVPVSSFITQYSTGQVTVEMEWELKKENSKRWNPEIQYTNNYN

DPQFVDFAPDSTGEYPTTRPIGTRYLTRPL

AAV5 VP3 sequence

(SEQ ID NO: 1116)

MSAGGGGPLGDNNQGADGVGNASGDWHCDSTWMGDRVVTKSTRTWVLPSYNNH

QYREIKSGSVDGSNANAYFGYSTPWGYFDFNRFHSHWSPRDWQRLINNYWGFRPRS

LRVKIFNIQVKEVTVQDSTTTIANNLTSTVQVFTDDDYQLPYVVGNGTEGCLPAFPPQ

VFTLPQYGYATLNRDNTENPTERSSFFCLEYFPSKMLRTGNNFEFTYNFEEVPFHSSF

APSQNLFKLANPLVDQYLYRFVSTNNTGGVQFNKNLAGRYANTYKNWFPGPMGRT

QGWNLGSGVNRASVSAFATTNRMELEGASYQVPPQPNGMTNNLQGSNTYALENTM

IFNSQPANPGTTATYLEGNMLITSESETQPVNRVAYNVGGQMATNNQSSTTAPATGT

YNILQE
IVPGSVWMERDVYLQGPIWAKIPETGAHFHPSPAMGGFGLKHPPPMMLIKN

TPVPGNITSFSDVPVSSFITQYSTGQVTVEMEWELKKENSKRWNPEIQYTNNYNDPQF

VDFAPDSTGEYRTTRPIGTRYLTRPL

3. Equivalents and Incorporation by Reference

All references cited herein are incorporated by reference to the same extent as if each individual publication, database entry (e.g., Genbank sequences or GeneID entries), patent application, or patent, was specifically and individually indicated incorporated by reference in its entirety, for all purposes. This statement of incorporation by reference is intended by Applicants, pursuant to 37 C.F.R. § 1.57(b)(1), to relate to each and every individual publication, database entry (e.g., Genbank sequences or GeneID entries), patent application, or patent, each of which is clearly identified in compliance with 37 C.F.R. § 1.57(b)(2), even if such citation is not immediately adjacent to a dedicated statement of incorporation by reference. The inclusion of dedicated statements of incorporation by reference, if any, within the specification does not in any way weaken this general statement of incorporation by reference. Citation of the references herein is not intended as an admission that the reference is pertinent prior art, nor does it constitute any admission as to the contents or date of these publications or documents.

While the invention has been particularly shown and described with reference to a preferred embodiment and various alternate embodiments, it is understood by persons skilled in the relevant art that various changes in form and details can be made therein without departing from the spirit and scope of the invention.

Number	Date	Country
63030038	May 2020	US
63119554	Nov 2020	US
63134885	Jan 2021	US
63181037	Apr 2021	US

HIGH THROUGHPUT ENGINEERING OF FUNCTIONAL AAV CAPSIDS

Information

Publication Number

Date Filed

Date Published

Inventors

Original Assignees

CPC

International Classifications

Abstract

Description

Claims

1. CROSS-REFERENCE TO RELATED APPLICATIONS

Provisional Applications (4)