Nucleic acids, proteins, and antibodies

STATEMENT UNDER 37 C.F.R. §1.77(b)(4)

[0001] This application refers to a “Sequence Listing” listed below, which is provided as an electronic document on two identical compact discs (CD-R), labeled “Copy 1” and “Copy 2.” These compact discs each contain the following files, which are hereby incorporated in their entirety herein:

1FileSize inDate ofDocumentNamebytesCreationSequence ListingPJZO2_seqList.txt2,365,610Jan. 15, 2001V Viewer Setup FileSetupDLL.exe695,808Dec. 19, 2000V Viewer Helpv.cnt7,984Jan. 05, 2001File ControllerV Viewer Programv.exe753,664Dec. 19, 2000FileV Viewer Help Filev.hlp447,766Jan. 05, 2001

[0002] The Sequence Listing may be viewed on an IBM-PC machine running the MS-Windows operating system by using the V viewer software, licensed by HGS, Inc., included on the compact discs (see World Wide Web URL: http://www.fileviewer.com).

FIELD OF THE INVENTION

[0003] The present invention relates to novel proteins. More specifically, isolated nucleic acid molecules are provided encoding novel polypeptides. Novel polypeptides and antibodies that bind to these polypeptides are provided. Also provided are vectors, host cells, and recombinant and synthetic methods for producing human polynucleotides and/or polypeptides, and antibodies. The invention further relates to diagnostic and therapeutic methods useful for diagnosing, treating, preventing and/or prognosing disorders related to these novel polypeptides. The invention further relates to screening methods for identifying agonists and antagonists of polynucleotides and polypeptides of the invention. The present invention further relates to methods and/or compositions for inhibiting or enhancing the production and function of the polypeptides of the present invention.

BACKGROUND OF THE INVENTION

[0004] The brain regulates every aspect of human behavior and physiology, from movement, heart rate, blood pressure, and body temperature, to language, emotion, and memories. The prominence of the nervous system among other bodily systems is evidenced by the disproportionate amount of resources consumed by the brain. While comprising only 2% of the body's weight, the brain consumes 20% of the body's oxygen, 25% of the body's glucose, and receives 15% of the cardiac output (see Circulation and energy metabolism of the brain. In: Siegel G, Agranoff B, Albers R W, and Molinoff P, eds. Basic Neurochemistry: molecular, cellular, and medical aspects. 4th ed. New York: Raven Press (1989)).

[0005] The nervous system is organized into the central nervous system (CNS; comprising the brain and spinal cord), and the peripheral nervous system (PNS; comprising the network of nerves that connects the brain and spinal cord to the rest of the body). The basic functional units of the CNS and PNS are neurons, usually composed of dendrites (branching specializations which receive input from other neurons), a cell body (containing the machinery to sustain cellular functions), and an axon (which transmits electrical signals to other neurons or muscle cells). Electrical impulses, propagated along axons by voltage-gated ion channels, are converted to chemical signals at junctions between neurons called synapses. Calcium-mediated exocytosis of storage vesicles in the axon terminal leads to neurotransmitter release into the synaptic cleft. The signaling molecules passively diffuse to the postsynaptic membrane and bind to neurotransmitter-specific receptor proteins. Depending on the type of receptor activated, neurotransmitter binding can have a variety of effects on the postsynaptic cell, including activation of second messenger biocehmical cascades and modulation of ion channel permeability. These biochemical and biophysical changes influence the subsequent behavior of the neuron, for example making the cell more or less excitable to incoming signals.

[0006] The elaborate circuitry of the adult nervous system arises through an interaction between genetically programmed growth patterns and environmental influences. During embryonic development, neural connections are formed via the programmed extension of axons under the influence of local molecular cues. Through post-natal development, this coarse pattern of connections is refined based on specific interactions between the child and the environment. It is believed that there are critical periods of neural development in childhood, during which environmental stimulation has a more profound effect on nervous system organization than during adulthood. For example, it is known that sensory deprivation in early childhood (such as blindness or deafness), leads to measurable differences in brain organization (see, for examples, Roder et al., Nature 400(6740):162-6 (1999); Buchel et al., Brain 121 (Pt 3):409-19 (1998)).

[0007] Because of its integral role in human behavior and physiology, disorders of the nervous system are among the most debilitating diseases known. Since the adult nervous system has very limited ability to regenerate, neural injury due to illness or trauma can produce life-long impairments. About half of all spinal cord injuries result in permenant loss of movement and sensation in the arms and legs (quadriplegia). Similarly, up to 30% of stroke survivors are left permanently disabled (American Heart Association 2000 Heart and Stroke Statistical Update, Dallas, Tex. (1999)). Methods of promoting neural tissue regeneration—for example, to repair spinal cord injuries or brain tissue damage—is a major focus of modern neurobiology. However, currently there is no effective way to repair damaged adult neural tissues.

[0008] A number of neurological conditions, including schizophrenia, depression, and myesthenia gravis, involve impaired or inappropriate synaptic communication between neurons. Drug therapies designed to correct the synaptic chemical imbalances underlying these disorders, such as dopamine receptor antagonists for schizophrenia and serotonin uptake inhibitors for depression, have had varying degrees of success, at the cost of sometimes serious side-effects.

[0009] The immune system is suspected to play a role in some neurological disorders and conditions. For example, multiple sclerosis, which is characterized by sensory impairments (tingling, numbness, dizziness, loss of vision) and motor impairments (tremor, weakness, loss of coordination), is thought to be an autoimmune disorder in which immune cells destroy the insulating myelin sheath covering axons. In addition, the inflammatory immune response can be a serious complication of brain injury (e.g. trauma and stroke), spinal cord damage, and infection (e.g. encephalitis and meningitis), and may be a common pathological mechanism in many other neurological disorders (Hays, Curr. Pharm. Des. 4:335-48 (1998); Halliday et al., Clin. Exp. Pharmacol. Physiol. 27:1-8 (2000)).

[0010] The field of neurobiology is only beginning to uncover the biological basis of neurological diseases. In fact, in most cases the underlying cause or causes remain poorly understood. Thus, the discovery of new human nervous system-associated polynucleotides, the polypeptides encoded by them, and antibodies that immunospecifically bind these polypeptides, satisfies a need in the art by providing new compositions which are useful in the diagnosis, treatment, prevention and/or prognosis of neurological diseases, disorders, and/or conditions, including, but not limited to, neuropsychiatric disorders, neurodegenerative diseases, vascular disorders, developmental disorders, infections, and neoplastic disorders.

SUMMARY OF THE INVENTION

[0011] The present invention relates to novel proteins. More specifically, isolated nucleic acid molecules are provided encoding novel polypeptides. Novel polypeptides and antibodies that bind to these polypeptides are provided. Also provided are vectors, host cells, and recombinant and synthetic methods for producing human polynucleotides and/or polypeptides, and antibodies. The invention further relates to diagnostic and therapeutic methods useful for diagnosing, treating, preventing and/or prognosing disorders related to these novel polypeptides. The invention further relates to screening methods for identifying agonists and antagonists of polynucleotides and polypeptides of the invention. The present invention further relates to methods and/or compositions for inhibiting or enhancing the production and function of the polypeptides of the present invention.

DETAILED DESCRIPTION

[0012] Tables

[0013] Table 1A summarizes some of the polynucleotides encompassed by the invention (including cDNA clones related to the sequences (Clone ID NO:Z), contig sequences (contig identifier (Contig ID:) and contig nucleotide sequence identifier (SEQ ID NO:X)) and further summarizes certain characteristics of these polynucleotides and the polypeptides encoded thereby. The first column provides the gene number in the application for each clone identifier. The second column provides a unique clone identifier, “Clone ID NO:Z”, for a cDNA clone related to each contig sequence disclosed in Table 1A. The third column provides a unique contig identifier, “Contig ID:” for each of the contig sequences disclosed in Table 1A. The fourth column provides the sequence identifier, “SEQ ID NO:X”, for each of the contig sequences disclosed in Table 1A. The fifth column, “ORF (From-To)”, provides the location (i.e., nucleotide position numbers) within the polynucleotide sequence of SEQ ID NO:X that delineate the preferred open reading frame (ORF) that encodes the amino acid sequence shown in the sequence listing and referenced in Table 1A as SEQ ID NO:Y (column 6). Column 7 lists residues comprising predicted epitopes contained in the polypeptides encoded by each of the preferred ORFs (SEQ ID NO:Y). Identification of potential immunogenic regions was performed according to the method of Jameson and Wolf (CABIOS, 4; 181-186 (1988)); specifically, the Genetics Computer Group (GCG) implementation of this algorithm, embodied in the program PEPTIDESTRUCTURE (Wisconsin Package v10.0, Genetics Computer Group (GCG), Madison, Wis.). This method returns a measure of the probability that a given residue is found on the surface of the protein. Regions where the antigenic index score is greater than 0.9 over at least 6 amino acids are indicated in Table 1A as “Predicted Epitopes”. In particular embodiments, polypeptides of the invention comprise, or alternatively consist of, one, two, three, four, five or more of the predicted epitopes described in Table 1A. It will be appreciated that depending on the analytical criteria used to predict antigenic determinants, the exact address of the determinant may vary slightly. Column 8, “Tissue Distribution” shows the expression profile of tissue, cells, and/or cell line libraries which express the polynucleotides of the invention. The first number in column 8 (preceding the colon), represents the tissue/cell source identifier code corresponding to the key provided in Table 4. Expression of these polynucleotides was not observed in the other tissues and/or cell libraries tested. For those identifier codes in which the first two letters are not “AR”, the second number in column 8 (following the colon), represents the number of times a sequence corresponding to the reference polynucleotide sequence (e.g., SEQ ID NO:X) was identified in the tissue/cell source. Those tissue/cell source identifier codes in which the first two letters are “AR” designate information generated using DNA array technology. Utilizing this technology, cDNAs were amplified by PCR and then transferred, in duplicate, onto the array. Gene expression was assayed through hybridization of first strand cDNA probes to the DNA array. cDNA probes were generated from total RNA extracted from a variety of different tissues and cell lines. Probe synthesis was performed in the presence of 33P dCTP, using oligo(dT) to prime reverse transcription. After hybridization, high stringency washing conditions were employed to remove non-specific hybrids from the array. The remaining signal, emanating from each gene target, was measured using a Phosphorimager. Gene expression was reported as Phosphor Stimulating Luminescence (PSL) which reflects the level of phosphor signal generated from the probe hybridized to each of the gene targets represented on the array. A local background signal subtraction was performed before the total signal generated from each array was used to normalize gene expression between the different hybridizations. The value presented after “[array code]:” represents the mean of the duplicate values, following background subtraction and probe normalization. One of skill in the art could routinely use this information to identify normal and/or diseased tissue(s) which show a predominant expression pattern of the corresponding polynucleotide of the invention or to identify polynucleotides which show predominant and/or specific tissue and/or cell expression. Column 9 provides the chromosomal location of polynucleotides corresponding to SEQ ID NO:X. Chromosomal location was determined by finding exact matches to EST and cDNA sequences contained in the NCBI (National Center for Biotechnology Information) UniGene database. Given a presumptive chromosomal location, disease locus association was determined by comparison with the Morbid Map, derived from Online Mendelian Inheritance in Man (Online Mendelian Inheritance in Man, OMIM™. McKusick-Nathans Institute for Genetic Medicine, Johns Hopkins University (Baltimore, Md.) and National Center for Biotechnology Information, National Library of Medicine (Bethesda, Md.) 2000. World Wide Web URL: http://www.ncbi.nlm.nih.gov/omim/). If the putative chromosomal location of the Query overlaps with the chromosomal location of a Morbid Map entry, an OMIM identification number is disclosed in column 10 labeled “OMIM Disease Reference(s)”. A key to the OMIM reference identification numbers is provided in Table 5.

[0014] Table 1B summarizes additional polynucleotides encompassed by the invention (including cDNA clones related to the sequences (Clone ID NO:Z), contig sequences (contig identifier (Contig ID:) contig nucleotide sequence identifiers (SEQ ID NO:X)), and genomic sequences (SEQ ID NO:B). The first column provides a unique clone identifier, “Clone ID NO:Z”, for a cDNA clone related to each contig sequence. The second column provides the sequence identifier, “SEQ ID NO:X”, for each contig sequence. The third column provides a unique contig identifier, “Contig ID:” for each contig sequence. The fourth column, provides a BAC identifier “BAC ID NO:A” for the BAC clone referenced in the corresponding row of the table. The fifth column provides the nucleotide sequence identifier, “SEQ ID NO:B” for a fragment of the BAC clone identified in column four of the corresponding row of the table. The sixth column, “Exon From-To”, provides the location (i.e., nucleotide position numbers) within the polynucleotide sequence of SEQ ID NO:B which delineate certain polynucleotides of the invention that are also exemplary members of polynucleotide sequences that encode polypeptides of the invention (e.g., polypeptides containing amino acid sequences encoded by the polynucleotide sequences delineated in column six, and fragments and variants thereof).

[0015] Table 2 summarizes homology and features of some of the polypeptides of the invention. The first column provides a unique clone identifier, “Clone ID NO:Z”, corresponding to a cDNA clone disclosed in Table 1A. The second column provides the unique contig identifier, “Contig ID:” corresponding to contigs in Table 1A and allowing for correlation with the information in Table 1A. The third column provides the sequence identifier, “SEQ ID NO:X”, for the contig polynucleotide sequence. The fourth column provides the analysis method by which the homology/identity disclosed in the Table was determined. Comparisons were made between polypeptides encoded by the polynucleotides of the invention and either a non-redundant protein database (herein referred to as “NR”), or a database of protein families (herein referred to as “PFAM”) as further described below. The fifth column provides a description of the PFAM/NR hit having a significant match to a polypeptide of the invention. Column six provides the accession number of the PFAM/NR hit disclosed in the fifth column. Column seven, “Score/Percent Identity”, provides a quality score or the percent identity, of the hit disclosed in columns five and six. Columns 8 and 9, “NT From” and “NT To” respectively, delineate the polynucleotides in “SEQ ID NO:X” that encode a polypeptide having a significant match to the PFAM/NR database as disclosed in the fifth and sixth columns. In specific embodiments polypeptides of the invention comprise, or alternatively consist of, an amino acid sequence encoded by a polynucleotide in SEQ ID NO:X as delineated in columns 8 and 9, or fragments or variants thereof.

[0016] Table 3 provides polynucleotide sequences that may be disclaimed according to certain embodiments of the invention. The first column provides a unique clone identifier, “Clone ID”, for a cDNA clone related to contig sequences disclosed in Table 1A. The second column provides the sequence identifier, “SEQ ID NO:X”, for contig sequences disclosed in Table 1A. The third column provides the unique contig identifier, “Contig ID:”, for contigs disclosed in Table 1A. The fourth column provides a unique integer ‘a’ where ‘a’ is any integer between 1 and the final nucleotide minus 15 of SEQ ID NO:X, and the fifth column provides a unique integer ‘b’ where ‘b’ is any integer between 15 and the final nucleotide of SEQ ID NO:X, where both a and b correspond to the positions of nucleotide residues shown in SEQ ID NO:X, and where b is greater than or equal to a +14. For each of the polynucleotides shown as SEQ ID NO:X, the uniquely defined integers can be substituted into the general formula of a-b, and used to describe polynucleotides which may be preferably excluded from the invention. In certain embodiments, preferably excluded from the invention are at least one, two, three, four, five, ten, or more of the polynucleotide sequence(s) having the accession number(s) disclosed in the sixth column of this Table (including for example, published sequence in connection with a particular BAC clone). In further embodiments, preferably excluded from the invention are the specific polynucleotide sequence(s) contained in the clones corresponding to at least one, two, three, four, five, ten, or more of the available material having the accession numbers identified in the sixth column of this Table (including for example, the actual sequence contained in an identified BAC clone).

[0017] Table 4 provides a key to the tissue/cell source identifier code disclosed in Table 1A, column 8. Column 1 provides the tissue/cell source identifier code disclosed in Table 1A, Column 8. Columns 2-5 provide a description of the tissue or cell source. Codes corresponding to diseased tissues are indicated in column 6 with the word “disease”. The use of the word “disease” in column 6 is non-limiting. The tissue or cell source may be specific (e.g. a neoplasm), or may be disease-associated (e.g., a tissue sample from a normal portion of a diseased organ). Furthermore, tissues and/or cells lacking the “disease” designation may still be derived from sources directly or indirectly involved in a disease state or disorder, and therefore may have a further utility in that disease state or disorder. In numerous cases where the tissue/cell source is a library, column 7 identifies the vector used to generate the library.

[0018] Table 5 provides a key to the OMIM reference identification numbers disclosed in Table 1A, column 10. OMIM reference identification numbers (Column 1) were derived from Online Mendelian Inheritance in Man (Online Mendelian Inheritance in Man, OMIM. McKusick-Nathans Institute for Genetic Medicine, Johns Hopkins University (Baltimore, Md.) and National Center for Biotechnology Information, National Library of Medicine, (Bethesda, Md.) 2000. World Wide Web URL: http://www.ncbi.nlm.nih.gov/omim/). Column 2 provides diseases associated with the cytologic band disclosed in Table 1A, column 9, as determined using the Morbid Map database.

[0019] Table 6 summarizes ATCC Deposits, Deposit dates, and ATCC designation numbers of deposits made with the ATCC in connection with the present application.

[0020] Table 7 shows the cDNA libraries sequenced, and ATCC designation numbers and vector information relating to these cDNA libraries.

[0021] Table 8 provides a physical characterization of clones encompassed by the invention. The first column provides the unique clone identifier, “Clone ID NO:Z”, for certain cDNA clones of the invention, as described in Table 1A. The second column provides the size of the cDNA insert contained in the corresponding cDNA clone.

[0022] Definitions

[0023] The following definitions are provided to facilitate understanding of certain terms used throughout this specification.

[0024] In the present invention, “isolated” refers to material removed from its original environment (e.g., the natural environment if it is naturally occurring), and thus is altered “by the hand of man” from its natural state. For example, an isolated polynucleotide could be part of a vector or a composition of matter, or could be contained within a cell, and still be “isolated” because that vector, composition of matter, or particular cell is not the original environment of the polynucleotide. The term “isolated” does not refer to genomic or cDNA libraries, whole cell total or mRNA preparations, genomic DNA preparations (including those separated by electrophoresis and transferred onto blots), sheared whole cell genomic DNA preparations or other compositions where the art demonstrates no distinguishing features of the polynucleotide/sequences of the present invention.

[0025] As used herein, a “polynucleotide” refers to a molecule having a nucleic acid sequence encoding SEQ ID NO:Y or a fragment or variant thereof, a nucleic acid sequence contained in SEQ ID NO:X (as described in column 3 of Table 1A) or the complement thereof, a cDNA sequence contained in Clone ID NO:Z (as described in column 2 of Table 1A and contained within a library deposited with the ATCC); a nucleotide sequence encoding the polypeptide encoded by a nucleotide sequence in SEQ ID NO:B as defined in column 6 of Table 1B or a fragment or variant thereof; or a nucleotide coding sequence in SEQ ID NO:B as defined in column 6 of Table 1B or the complement thereof. For example, the polynucleotide can contain the nucleotide sequence of the full length cDNA sequence, including the 5′ and 3′ untranslated sequences, the coding region, as well as fragments, epitopes, domains, and variants of the nucleic acid sequence. Moreover, as used herein, a “polypeptide” refers to a molecule having an amino acid sequence encoded by a polynucleotide of the invention as broadly defined (obviously excluding poly-Phenylalanine or poly-Lysine peptide sequences which result from translation of a polyA tail of a sequence corresponding to a cDNA).

[0026] In the present invention, “SEQ ID NO:X” was often generated by overlapping sequences contained in multiple clones (contig analysis). A representative clone containing all or most of the sequence for SEQ ID NO:X is deposited at Human Genome Sciences, Inc. (HGS) in a catalogued and archived library. As shown, for example, in column 2 of Table 1A, each clone is identified by a cDNA Clone ID (identifier generally referred to herein as Clone ID NO:Z). Each Clone ID is unique to an individual clone and the Clone ID is all the information needed to retrieve a given clone from the HGS library. Furthermore, certain clones disclosed in this application have been deposited with the ATCC on Oct. 5, 2000, having the ATCC designation numbers PTA 2574 and PTA 2575; and on Jan. 5, 2001, having the depositor reference numbers TS-1, TS-2, AC-1, and AC-2. In addition to the individual cDNA clone deposits, most of the cDNA libraries from which the clones were derived were deposited at the American Type Culture Collection (hereinafter “ATCC”). Table 7 provides a list of the deposited cDNA libraries. One can use the Clone ID NO:Z to determine the library source by reference to Tables 6 and 7. Table 7 lists the deposited cDNA libraries by name and links each library to an ATCC Deposit. Library names contain four characters, for example, “HTWE.” The name of a cDNA clone (Clone ID) isolated from that library begins with the same four characters, for example “HTWEP07”. As mentioned below, Table 1A correlates the Clone ID names with SEQ ID NO:X. Thus, starting with an SEQ ID NO:X, one can use Tables 1, 6 and 7 to determine the corresponding Clone ID, which library it came from and which ATCC deposit the library is contained in. Furthermore, it is possible to retrieve a given cDNA clone from the source library by techniques known in the art and described elsewhere herein. The ATCC is located at 10801 University Boulevard, Manassas, Va. 20110-2209, USA. The ATCC deposits were made pursuant to the terms of the Budapest Treaty on the international recognition of the deposit of microorganisms for the purposes of patent procedure.

[0027] In specific embodiments, the polynucleotides of the invention are at least 15, at least 30, at least 50, at least 100, at least 125, at least 500, or at least 1000 continuous nucleotides but are less than or equal to 300 kb, 200 kb, 100 kb, 50 kb, 15 kb, 10 kb, 7.5 kb, 5 kb, 2.5 kb, 2.0 kb, or 1 kb, in length. In a further embodiment, polynucleotides of the invention comprise a portion of the coding sequences, as disclosed herein, but do not comprise all or a portion of any intron. In another embodiment, the polynucleotides comprising coding sequences do not contain coding sequences of a genomic flanking gene (i.e., 5′ or 3′ to the gene of interest in the genome). In other embodiments, the polynucleotides of the invention do not contain the coding sequence of more than 1000, 500, 250, 100, 50, 25, 20, 15, 10, 5, 4, 3, 2, or 1 genomic flanking gene(s).

[0028] A “polynucleotide” of the present invention also includes those polynucleotides capable of hybridizing, under stringent hybridization conditions, to sequences contained in SEQ ID NO:X, or the complement thereof (e.g., the complement of any one, two, three, four, or more of the polynucleotide fragments described herein), the polynucleotide sequence delineated in columns 8 and 9 of Table 2 or the complement thereof, and/or cDNA sequences contained in Clone ID NO:Z (e.g., the complement of any one, two, three, four, or more of the polynucleotide fragments, or the cDNA clone within the pool of cDNA clones deposited with the ATCC, described herein), and/or the polynucleotide sequence delineated in column 6 of Table 1B or the complement thereof. “Stringent hybridization conditions” refers to an overnight incubation at 42 degree C. in a solution comprising 50% formamide, 5×SSC (750 mM NaCl, 75 mM trisodium citrate), 50 mM sodium phosphate (pH 7.6), 5× Denhardt's solution, 10% dextran sulfate, and 20 μg/ml denatured, sheared salmon sperm DNA, followed by washing the filters in 0.1×SSC at about 65 degree C.

[0029] Also contemplated are nucleic acid molecules that hybridize to the polynucleotides of the present invention at lower stringency hybridization conditions. Changes in the stringency of hybridization and signal detection are primarily accomplished through the manipulation of formamide concentration (lower percentages of formamide result in lowered stringency); salt conditions, or temperature. For example, lower stringency conditions include an overnight incubation at 37 degree C. in a solution comprising 6×SSPE (20×SSPE=3M NaCl; 0.2M NaH2PO4; 0.02M EDTA, pH 7.4), 0.5% SDS, 30% formamide, 100 ug/ml salmon sperm blocking DNA; followed by washes at 50 degree C. with 1×SSPE, 0.1% SDS. In addition, to achieve even lower stringency, washes performed following stringent hybridization can be done at higher salt concentrations (e.g. 5×SSC).

[0030] Note that variations in the above conditions may be accomplished through the inclusion and/or substitution of alternate blocking reagents used to suppress background in hybridization experiments. Typical blocking reagents include Denhardt's reagent, BLOTTO, heparin, denatured salmon sperm DNA, and commercially available proprietary formulations. The inclusion of specific blocking reagents may require modification of the hybridization conditions described above, due to problems with compatibility.

[0031] Of course, a polynucleotide which hybridizes only to polyA+ sequences (such as any 3′ terminal polyA+ tract of a cDNA shown in the sequence listing), or to a complementary stretch of T (or U) residues, would not be included in the definition of “polynucleotide,” since such a polynucleotide would hybridize to any nucleic acid molecule containing a poly (A) stretch or the complement thereof (e.g., practically any double-stranded cDNA clone generated using oligo dT as a primer).

[0032] The polynucleotide of the present invention can be composed of any polyribonucleotide or polydeoxribonucleotide, which may be unmodified RNA or DNA or modified RNA or DNA. For example, polynucleotides can be composed of single- and double-stranded DNA, DNA that is a mixture of single- and double-stranded regions, single- and double-stranded RNA, and RNA that is mixture of single- and double-stranded regions, hybrid molecules comprising DNA and RNA that may be single-stranded or, more typically, double-stranded or a mixture of single- and double-stranded regions. In addition, the polynucleotide can be composed of triple-stranded regions comprising RNA or DNA or both RNA and DNA. A polynucleotide may also contain one or more modified bases or DNA or RNA backbones modified for stability or for other reasons. “Modified” bases include, for example, tritylated bases and unusual bases such as inosine. A variety of modifications can be made to DNA and RNA; thus, “polynucleotide” embraces chemically, enzymatically, or metabolically modified forms.

[0033] The polypeptide of the present invention can be composed of amino acids joined to each other by peptide bonds or modified peptide bonds, i.e., peptide isosteres, and may contain amino acids other than the 20 gene-encoded amino acids. The polypeptides may be modified by either natural processes, such as posttranslational processing, or by chemical modification techniques which are well known in the art. Such modifications are well described in basic texts and in more detailed monographs, as well as in a voluminous research literature. Modifications can occur anywhere in a polypeptide, including the peptide backbone, the amino acid side-chains and the amino or carboxyl termini. It will be appreciated that the same type of modification may be present in the same or varying degrees at several sites in a given polypeptide. Also, a given polypeptide may contain many types of modifications. Polypeptides may be branched, for example, as a result of ubiquitination, and they may be cyclic, with or without branching. Cyclic, branched, and branched cyclic polypeptides may result from posttranslation natural processes or may be made by synthetic methods. Modifications include acetylation, acylation, ADP-ribosylation, amidation, covalent attachment of flavin, covalent attachment of a heme moiety, covalent attachment of a nucleotide or nucleotide derivative, covalent attachment of a lipid or lipid derivative, covalent attachment of phosphotidylinositol, cross-linking, cyclization, disulfide bond formation, demethylation, formation of covalent cross-links, formation of cysteine, formation of pyroglutamate, formylation, gamma-carboxylation, glycosylation, GPI anchor formation, hydroxylation, iodination, methylation, myristoylation, oxidation, pegylation, proteolytic processing, phosphorylation, prenylation, racemization, selenoylation, sulfation, transfer-RNA mediated addition of amino acids to proteins such as arginylation, and ubiquitination. (See, for instance, PROTEINS—STRUCTURE AND MOLECULAR PROPERTIES, 2nd Ed., T. E. Creighton, W. H. Freeman and Company, New York (1993); POSTTRANSLATIONAL COVALENT MODIFICATION OF PROTEINS, B. C. Johnson, Ed., Academic Press, New York, pgs. 1-12 (1983); Seifter et al., Meth. Enzymol. 182:626-646 (1990); Rattan et al., Ann. N.Y. Acad. Sci. 663:48-62 (1992)).

[0034] “SEQ ID NO:X” refers to a polynucleotide sequence described, for example, in Tables 1A or 2, while “SEQ ID NO:Y” refers to a polypeptide sequence described in column 6 of Table 1A. SEQ ID NO:X is identified by an integer specified in column 4 of Table 1A. The polypeptide sequence SEQ ID NO:Y is a translated open reading frame (ORF) encoded by polynucleotide SEQ ID NO:X. “Clone ID NO:Z” refers to a cDNA clone described in column 2 of Table 1A.

[0035] “A polypeptide having functional activity” refers to a polypeptide capable of displaying one or more known functional activities associated with a full-length (complete) protein. Such functional activities include, but are not limited to, biological activity, antigenicity [ability to bind (or compete with a polypeptide for binding) to an anti-polypeptide antibody], immunogenicity (ability to generate antibody which binds to a specific polypeptide of the invention), ability to form multimers with polypeptides of the invention, and ability to bind to a receptor or ligand for a polypeptide.

[0036] The polypeptides of the invention can be assayed for functional activity (e.g. biological activity) using or routinely modifying assays known in the art, as well as assays described herein. Specifically, one of skill in the art may routinely assay nervous system polypeptides (including fragments and variants) of the invention for activity using assays as described in Examples 24, 34, 37, 48, and 59.

[0037] “A polypeptide having biological activity” refers to a polypeptide exhibiting activity similar to, but not necessarily identical to, an activity of a polypeptide of the present invention, including mature forms, as measured in a particular biological assay, with or without dose dependency. In the case where dose dependency does exist, it need not be identical to that of the polypeptide, but rather substantially similar to the dose-dependence in a given activity as compared to the polypeptide of the present invention (i.e., the candidate polypeptide will exhibit greater activity or not more than about 25-fold less and, preferably, not more than about tenfold less activity, and most preferably, not more than about three-fold less activity relative to the polypeptide of the present invention).

[0038] Table 1A summarizes some of the polynucleotides encompassed by the invention (including contig sequences (SEQ ID NO:X) and clones (Clone ID NO:Z) and further summarizes certain characteristics of these polynucleotides and the polypeptides encoded thereby.

[0039] Polynucleotides and Polypeptides of the Invention

2TABLE 1AAATissue DistributionSEQLibrary code: countOMIMGeneContigSEQ IDORFID(see Table IV for LibraryCytologicDiseaseNo:Clone ID NO: ZID:NO: X(From-To)NO: YPredicted EpitopesCodes)BandReference(s):1HTPAD461048901111567-1148609AR061: 0, AR089: 0L0794: 4, H0039: 2,S0358: 1, H0013: 1,H0575: 1, L0770: 1,L0769: 1 and L0749: 1.503313335103-309933His-50 to Leu-69.2HCWFF88109256612628-227610AR089: 15, AR061: 6H0305: 2506577336 41-187934Pro-1 to Gly-6,Ala-41 to Leu-47.3HSSAX53119888913209-361611H0135: 1 and H0063:1.507509337209-3619354HCEPH7152273914 3-410612Val-1 to Lys-8,AR089: 1, AR061: 1Pro-36 to Lys-41,H0052: 1 and T0067:Gln-49 to Lys-57,1.Ser-63 to Ser-70,Asp-79 to Gln-92,Asn-103 to Thr-122.5HTEDF74108340515 2-970613Pro-37 to Trp-53,AR061: 3, AR089: 2Arg-56 to Pro-62.L0731: 2, H0341: 1,H0392: 1, H0038: 1,H0641: 1, L0803: 1 andH0445: 1.522982338 3-575936Pro-20 to Trp-36,Arg-39 to Pro-45,Gly-62 to Glu-69,Asp-77 to Lys-82,Pro-87 to Ala-93.6HTTEK47113453416 2-1009614Val-21 to Pro-27.AR061: 8, AR089: 6L0439: 5, H0622: 3,H0040: 2, L0794: 2,L0805: 2, L0758: 2,L0803: 1, L0375: 1,L0659: 1, L0789: 1,L0665: 1, H0579: 1,L0750: 1, L0779: 1,L0777: 1, L0752: 1 andL0755: 1.573649339 2-349937Leu-13 to Val-18,Thr-37 to Lys-46.7HTOBE75116388317 149-1075615Arg-36 to Gln-44,AR061: 2, AR089: 1Ser-49 to Gln-57,H0038: 3, L0748: 3,Lys-276 to Cys-286.L0659: 2, L0743: 2,L0744: 2, H0486: 1,H0421: 1, H0024: 1,H0031: 1, H0272: 1,L0662: 1, L0384: 1,L0809: 1 and L0779: 1.591896340 1-4149388HCFAT05115631018 1-747616Leu-21 to Gln-29,AR061: 1, AR089: 1Ala-95 to Gly-101,H0556: 2, H0634: 1,Arg-163 to Gln-172,L0766: 1 and H0422: 1.Ser-183 to Glu-202,Thr-219 to Ser-226,Thr-231 to Ser-238.592118341 2-490939Arg-1 to His-11,Ser-18 to Gly-27,Gly-36 to Gly-44,Asp-97 to Phe-103,Pro-127 to Gly-132.9HFIAH37118900119 69-872617Lys-65 to Thr-71,AR089: 7, AR061: 1Lys-104 to Gly-109,H0457: 2, H0529: 2,Lys-116 to His-122,H0561: 1, H0521: 1,Asn-140 to Asp-146,S0192: 1 and L0600: 1.Lys-184 to Lys-203,Glu-205 to Asn-239,Ala-256 to Phe-267.615597342 70-375940Lys-65 to Thr-71.10HFTDF15108488720475-672618AR089: 3, AR061: 2H0563: 1 and H0123:1.657020343129-25494111HPFCU80101759321341-3 619AR089: 16, AR061: 13H0163: 3 and H0169:1.685294344162-34194212HSVAW49115391622767-501620Pro-19 to Thr-24,AR061: 9, AR089: 7Thr-78 to Lys-89.H0309: 1689674345 44-208943Glu-21 to Glu-27.13HWHQC94111646323 2-916621AR089: 2, AR061: 1L0665: 4, S0132: 2,L0438: 2, L0439: 2,L0596: 2, H0542: 2,H0543: 2, S0114: 1,H0614: 1, H0592: 1,H0587: 1, S0280: 1,H0253: 1, H0581: 1,H0457: 1, H0012: 1,H0083: 1, H0687: 1,H0290: 1, H0622: 1,H0135: 1, S0150: 1,L0796: 1, L0646: 1,L0643: 1, L0764: 1,L0773: 1, L0649: 1,L0659: 1, L0663: 1,H0658: 1, H0555: 1,H0478: 1, L0752: 1,L0599: 1 and H0506: 1.715096346 1-627944Trp-62 to Pro-67.14HRSMD49106545824166-342622Phe-16 to Trp-24,AR089: 3, AR061: 2Leu-30 to Val-37,H0394: 1 and L0589:Phe-41 to Ile-49.1.723025347190-456945Gln-36 to Ile-46,Ser-55 to Phe-65,Ser-67 to Lys-78.15HFTDY671151220251527-1 623Asn-64 to Pro-73,AR089: 89, AR061: 75Asp-83 to Glu-94,S0002: 2, H0521: 2,Leu-144 to Pro-153,S0360: 1, H0123: 1,Glu-162 to Thr-167,S0250: 1, L0654: 1,Asp-178 to Ser-189,S0152: 1, L0740: 1 andGly-197 to Leu-210,L0749: 1.Pro-217 to Pro-222,Arg-234 to Asp-251,Gly-279 to Phe-293,Asp-357 to Gly-367,Gly-379 to Val-396,Glu-421 to Met-426,Asn-441 to Leu-447,Glu-467 to Trp-474.745221348 1-228946Gly-23 to Phe-37.16HYABL89109073326247-417624AR089: 1, AR061: 1S0114: 1, H0583: 1 andH0013: 1.786157349246-41694717HCUEV29113779127 3-569625Glu-18 to Val-28,AR089: 1, AR061: 1Pro-31 to Glu-47,H0457: 15, H0271: 11,Glu-88 to Asp-94,H0494: 7, H0521: 7,Ser-154 to Lys-178.H0141: 6, H0255: 6,S0434: 6, L0758: 5,S0354: 4, S0358: 4,S0278: 4, H0179: 4,L0771: 4, L0783: 4,H0436: 4, H0556: 3,H0069: 3, H0618: 3,L0776: 3, L0659: 3,H0435: 3, H0661: 2,S0418: 2, S0420: 2,H0580: 2, S0222: 2,H0486: 2, H0013: 2,H0581: 2, H0083: 2,H0266: 2, S0003: 2,H0424: 2, S0036: 2,H0090: 2, H0038: 2,H0634: 2, H0616: 2,S0344: 2, S0002: 2,L0770: 2, L0646: 2,L0662: 2, L0381: 2,L0655: 2, L0809: 2,L0666: 2, L0665: 2,S0216: 2, H0703: 2,H0547: 2, H0593: 2,H0670: 2, H0539: 2,S0027: 2, L0748: 2,L0439: 2, L0751: 2,L0591: 2, H0543: 2,H0624: 1, H0650: 1,H0656: 1, S0116: 1,H0484: 1, H0402: 1,S0376: 1, S0444: 1,S0360: 1, S0045: 1,S0046: 1, H0619: 1,S6026: 1, H0261: 1,H0438: 1, H0586: 1,H0559: 1, H0101: 1,H0427: 1, H0036: 1,T0048: 1, H0318: 1,S0474: 1, H0421: 1,H0052: 1, H0205: 1,H0231: 1, L0738: 1,H0150: 1, H0081: 1,T0010: 1, H0416: 1,T0006: 1, H0213: 1,H0598: 1, H0135: 1,H0040: 1, H0087: 1,H0264: 1, H0488: 1,H0623: 1, H0334: 1,H0561: 1, S0440: 1,L0369: 1, L0769: 1,L0667: 1, L0773: 1,L0648: 1, L0364: 1,L0766: 1, L0649: 1,L0375: 1, L0378: 1,L0806: 1, L0653: 1,L0657: 1, L0636: 1,S0052: 1, S0428: 1,H0702: 1, S0374: 1,L0438: 1, S0328: 1,S0330: 1, H0710: 1,S0146: 1, S0406: 1,H0576: 1, H0631: 1,S3014: 1, L0779: 1,L0759: 1, H0445: 1,S0436: 1, S0011: 1,S0026: 1, S0242: 1 andH0506: 1.816065350 2-29894818HCESP56112175128452-72 626Glu-1 to Leu-6,AR061: 8, AR089: 5Asp-90 to Asp-107.H0052: 3, S0282: 1,H0194: 1, H0009: 1,L0789: 1, L0602: 1 andL0439: 1.827671351147-512949Asp-15 to Thr-21,Gln-83 to Ile-91.19HLQDT35115406429230-502627Ile-1 to Ser-6,AR089: 3, AR061: 3Leu-81 to Glu-91.S0358: 8, L0766: 7,L0777: 7, L0731: 7,L0659: 4, L0748: 4,L0751: 4, L0783: 3,L0663: 3, S0418: 2,S0360: 2, H0486: 2,S0010: 2, S0250: 2,S0422: 2, L0763: 2,L0803: 2, L0775: 2,L0789: 2, H0520: 2,L0756: 2, L0752: 2,H0656: 1, S0376: 1,H0208: 1, H0574: 1,H0632: 1, S0414: 1,H0581: 1, H0052: 1,H0024: 1, H0014: 1,H0355: 1, H0688: 1,H0090: 1, H0623: 1,H0509: 1, H0529: 1,L0520: 1, L0761: 1,L0650: 1, L0809: 1,L0666: 1, L0665: 1,S0126: 1, H0684: 1,H0648: 1, S0390: 1,L0740: 1, L0745: 1,L0749: 1, L0750: 1,L0755: 1, L0591: 1,L0362: 1 and S0242: 1.839777352222-49495020HDPBS6484662430 2-685628Phe-7 to Pro-15,AR089: 1, AR061: 0Trp-34 to Gly-40.S0222: 1, S0002: 1,L0804: 1, L0663: 1 andH0521: 1.21HTBAB41105238831619-798629Met-1 to Ser-8.AR089: 30, AR061: 8S0044: 2, L0748: 2,H0392: 1, H0075: 1,L0770: 1 and H0519: 1.867287353 11-178951Lys-1 to Asp-7,Gln-47 to Arg-53.22HTLGE3187024732 2-307630Val-47 to Gly-65.AR089: 163, AR061:32H0618: 1, L0368: 1and S0053: 1.23HWLHK29115227933 3-608631Gln-24 to Thr-32,AR089: 15, AR061: 5Ser-154 to Phe-163.L0754: 5, L0755: 5,S0354: 3, L0483: 3,H0648: 3, L0777: 3,S0374: 2, L0751: 2,L0758: 2, L0605: 2,L0362: 2, H0543: 2,S0114: 1, S0358: 1,H0411: 1, H0575: 1,L0105: 1, H0263: 1,H0596: 1, H0510: 1,S0003: 1, H0166: 1,H0169: 1, H0090: 1,H0059: 1, S0440: 1,L0373: 1, L0372: 1,L0800: 1, L0662: 1,L0794: 1, L0649: 1,L0803: 1, L0804: 1,L0659: 1, L0783: 1,L0809: 1, L0789: 1,L0790: 1, L0666: 1,L0665: 1, S0378: 1,L0602: 1, H0436: 1,L0779: 1, L0780: 1,S0434: 1, S0196: 1 andS0446: 1.876064354 3-491952Gln-18 to Thr-26.24HHEGG20110681634 3-971632Arg-4 to Glu-12,AR089: 2, AR061: 1Glu-121 to Gly-126,S0360: 1, H0013: 1,Ala-141 to Pro-146,L0664: 1 and H0542: 1.Gln-161 to Phe-176,Lys-186 to Ser-194.894409355 26-82095325HDPRU43121703535 1-2460633Pro-1 to Gly-7,AR089: 1, AR061: 1Val-127 to Val-133,H0046: 34, L0731: 5,Leu-162 to Ser-171,L0534: 4, L0769: 4,Arg-178 to Glu-185,H0521: 4, S0356: 3,Pro-195 to Thr-200,L0800: 3, L0794: 3,Gln-243 to Trp-248,L0439: 3, L0749: 3,Gln-252 to Asn-265,L0752: 3, L0759: 3,Glu-313 to Cys-319,L0562: 2, H0486: 2,His-417 to Glu-422,L0803: 2, L0805: 2,Arg-464 to Ala-473,L0809: 2, L0789: 2,Met-530 to Lys-538,L0744: 2, L0485: 2,Arg-594 to Gly-599,H0556: 1, H0657: 1,Glu-641 to Gly-649,H0637: 1, H0580: 1,Asp-660 to Ala-668,H0208: 1, H0261: 1,Arg-705 to Ser-727,H0609: 1, H0455: 1,Ser-777 to Glu-783,H0600: 1, H0586: 1,Leu-796 to Gly-806.H0331: 1, H0635: 1,H0618: 1, H0544: 1,H0009: 1, H0050: 1,H0620: 1, H0288: 1,S0312: 1, S0314: 1,H0252: 1, H0688: 1,H0644: 1, S0366: 1,H0135: 1, H0063: 1,H0087: 1, H0551: 1,H0264: 1, S0002: 1,L0639: 1, L0771: 1,L0648: 1, L0766: 1,L0650: 1, L0378: 1,L0655: 1, H0699: 1,H0660: 1, L0743: 1,L0750: 1, L0777: 1,L0758: 1, L0097: 1,S0194: 1 and H0543: 1.909841356 1-342954Pro-8 to Gln-16.26HE8PK12122764736 2-367634Val-30 to Ser-37.AR089: 6, AR061: 4L0754: 6, L0777: 6,L0740: 5, L0731: 4,L0758: 4, L0759: 4,S0001: 3, S0280: 3,L0770: 3, L0764: 3,L0747: 3, L0749: 3,L0366: 3, S0412: 3,S0007: 2, H0411: 2,H0013: 2, L0471: 2,T0004: 2, L0598: 2,L0638: 2, L0662: 2,L0783: 2, L0438: 2,H0696: 2, L0744: 2,L0748: 2, L0751: 2,L0745: 2, L0779: 2,L0752: 2, H0170: 1,S0282: 1, H0662: 1,H0574: 1, T0060: 1,H0427: 1, H0590: 1,S0010: 1, L0105: 1,S0049: 1, H0194: 1,H0373: 1, L0163: 1,H0201: 1, H0031: 1,H0553: 1, S0306: 1,L0776: 1, L0659: 1,L0526: 1, L0809: 1,L0663: 1, H0144: 1,H0547: 1, H0648: 1,H0672: 1, L0743: 1,L0780: 1, S0031: 1,H0343: 1, L0604: 1 andH0653: 1.909884357 2-367955Val-30 to Ser-37,Gln-43 to Asp-62,Pro-74 to Glu-79,Thr-102 to Phe-109.27HE9HV92122751937 1-1050635Asn-2 to Gly-10,AR061: 1, AR089: 0Asp-86 to Ile-110,L0439: 3, H0616: 2,Gly-116 to Gln-121,L0749: 2, S0420: 1,Ala-135 to Arg-140,H0415: 1, H0013: 1,Ala-167 to Ser-172,H0590: 1, S0010: 1,Leu-176 to Lys-183.H0046: 1, H0050: 1,H0375: 1, H0615: 1,S0366: 1, H0529: 1,H0144: 1, S0126: 1,S0152: 1, S3014: 1 andL0779: 1.911510358 2-1048956Thr-1 to Gly-9.28HOHCE47121705938 578-2143636Tyr-83 to Ser-92,AR061: 1, AR089: 0Leu-118 to Tyr-123,S0040: 1, H0580: 1,Leu-137 to Ser-143,S0222: 1, H0355: 1,Gln-148 to Ser-158,S0250: 1, L0565: 1 andThr-258 to Pro-266,S0152: 1.Gln-274 to His-283,Asp-325 to Ser-334,Gln-343 to Thr-349,Ser-366 to Val-378,Arg-381 to Asp-388,Pro-426 to Asn-431,Cys-446 to Ser-457,Leu-469 to Lys-486,Cys-501 to Arg-510.911566359 1-429957Gly-1 to Trp-6.29HSDII69115406739 74-412637His-13 to Gly-21,AR061: 6, AR089: 5Tyr-61 to Asp-66,H0328: 4, H0031: 3,Ala-105 to Thr-110.L0519: 3, L0748: 2,L0777: 2, L0731: 2,S0260: 2, H0624: 1,S6024: 1, H0650: 1,S0116: 1, H0254: 1,S0007: 1, H0393: 1,H0441: 1, H0438: 1,H0574: 1, H0156: 1,H0599: 1, S0051: 1,H0615: 1, H0039: 1,L0564: 1, L0763: 1,L0766: 1, L0774: 1,L0776: 1, L0659: 1,L0518: 1, L0792: 1,L0666: 1, L0663: 1,S0242: 1 and H0423: 1.917180360202-540958His-13 to Gly-21,Tyr-61 to Asp-66,Ala-105 to Thr-110.30HKAKM10122763940 3-2840638Leu-15 to Ser-21,AR089: 1, AR061: 1Leu-89 to Tyr-94,L0794: 4, L0438: 4,Gly-130 to Gln-136,L0761: 3, L0766: 3,Asn-163 to Leu-168,L0748: 3, L0439: 3,Lys-176 to His-181,H0556: 2, L0602: 2,Ile-187 to Arg-193,L0754: 2, L0779: 2,Ala-239 to Thr-244,H0580: 1, H0208: 1,Pro-263 to Val-268,H0013: 1, T0082: 1,Ala-401 to Ser-406.S0010: 1, H0428: 1,H0553: 1, H0038: 1,H0616: 1, H0494: 1,L0796: 1, L0800: 1,L0773: 1, L0533: 1,L0803: 1, L0776: 1,L0657: 1, L0791: 1,H0520: 1, H0519: 1,H0521: 1, H0187: 1,L0731: 1, S0031: 1 andL0366: 1.918685361 2-547959Gly-25 to Gln-31,Asn-58 to Leu-63,Lys-71 to His-76,Ile-82 to Arg-88,Ala-134 to Thr-139.31HCEPU561226120411037-1840639Lys-65 to Thr-71,AR061: 6, AR089: 5Lys-104 to Gly-109,Lys-116 to His-122,Asn-140 to Asp-146,Lys-184 to Lys-203,Glu-205 to Asn-239,Ala-256 to Phe-267.920347362219-593960Lys-16 to Thr-22,Lys-55 to Gly-60,Lys-67 to His-73,Asn-91 to Asp-97.32HUSHB5492805442125-355640AR089: 1, AR061: 0H0056: 3, H0437: 1,H0050: 1 and S0002: 1.33HLMDO9592834443 88-435641AR089: 27, AR061: 11H0271: 3, H0250: 2,H0635: 2, S0216: 2,H0254: 1, H0638: 1,H0069: 1, H0416: 1,H0090: 1, L0761: 1,L0800: 1, L0776: 1,L0789: 1 and S0052: 1.34HHASQ32119890244 50-892642Gly-1 to Gly-6,AR089: 21, AR061: 7Arg-12 to Arg-17,H0510: 3, S0438: 2,Gln-56 to Ser-71,L0803: 2, L0615: 1,Glu-82 to Glu-89,S0418: 1, H0393: 1,Phe-94 to Glu-104,H0632: 1, H0355: 1,Ala-126 to Asn-131,L0774: 1, H0144: 1,Pro-167 to Gly-177,L0749: 1, L0750: 1,Thr-224 to Ala-233,L0605: 1 and L0581: 1.Leu-237 to Lys-281.928730363 40-855961Gly-1 to Gly-6,Arg-12 to Arg-17.35HARAB87116434045 3-548643AR051: 29, AR050:24, AR054: 18, AR089:1, AR061: 0T0082: 1, T0023: 1 andL0596: 1.933441364181-76896236HTNGF69115226846501-803644Ser-1 to Gly-23,AR061: 3, AR089: 2Gly-85 to Leu-91.L0758: 3, L0666: 2,L0751: 2, L0777: 2,H0663: 1, L0021: 1,H0309: 1, H0688: 1,H0617: 1, H0477: 1,L0766: 1, L0775: 1,L0367: 1, L0789: 1,L0663: 1, L0438: 1,L0749: 1, L0779: 1,L0757: 1 and S0456: 1.933614365483-785963Ser-1 to Gly-23,Gly-85 to Leu-91.37HMSJL96115478847 1-1461645Thr-15 to Arg-22,AR054: 16, AR051:Ala-38 to Met-43,15, AR050: 12, AR089:Gln-49 to Lys-64,0, AR061: 0Thr-97 to Gln-108,L0777: 6, L0758: 5,Tyr-132 to Ser-139,L0779: 4, L0803: 3,His-219 to Gln-265,S0358: 2, H0004: 2,Ala-354 to Gly-362,L0662: 2, L0775: 2,Ser-379 to Ala-386,H0144: 2, S0126: 2,Asp-402 to Ser-417,S0328: 2, S3014: 2,Leu-424 to Glu-433,S0027: 2, L0743: 2,Arg-443 to Gly-459,L0748: 2, H0265: 1,Ser-463 to Ser-472.H0656: 1, S0212: 1,H0663: 1, H0638: 1,H0580: 1, H0632: 1,H0486: 1, H0599: 1,H0618: 1, L0105: 1,H0251: 1, H0309: 1,H0544: 1, H0123: 1,H0050: 1, L0471: 1,H0024: 1, H0399: 1,S0003: 1, H0364: 1,H0553: 1, H0038: 1,H0412: 1, H0413: 1,T0041: 1, S0344: 1,S0002; 1, L0598: 1,H0529: 1, L0645: 1,L0363: 1, L0649: 1,L0804: 1, L0805: 1,L0558: 1, L0659: 1,L0528: 1, L0789: 1,L0792: 1, L0666: 1,S0374: 1, H0555: 1,S3012: 1, S0028: 1,S0206: 1, S0032: 1,L0439: 1, L0757: 1,S0031: 1, H0707: 1,S0192: 1, H0423: 1,S0042: 1 and H0008: 1.934483366 1-426964Thr-15 to Arg-22,Ala-38 to Met-43,Gln-49 to Lys-64,Thr-97 to Gln-108,Thr-131 to Lys-137.38HDTBT06120526148 3-905646AR089: 2, AR061: 1L0754: 6, H0318: 3,H0486: 2, H0014: 2,L0777: 2, H0543: 2,H0171: 1, S6024: 1,H0650: 1, S0354: 1,H0455: 1, H0013: 1,L0483: 1, H0494: 1,S0450: 1, L0520: 1,L0763: 1, L0769: 1,L0641: 1, L0521: 1,L0662: 1, L0774: 1,L0776: 1, L0783: 1,L0663: 1, S0136: 1,H0478: 1, L0742: 1,L0439: 1, L0780: 1,L0592: 1, S0192: 1 andS0424: 1.935404367 1-90696539HTTIE47116536349 2-574647Thr-8 to Ser-16,AR061: 4, AR089: 4Arg-34 to Leu-42,L0615: 1, S0420: 1,Thr-46 to Glu-51,H0333: 1, H0286: 1,Thr-57 to Arg-66,H0634: 1 and H0144: 1.Gln-94 to Ala-100,Ser-127 to Glu-134.941834368 1-567966Thr-6 to Ser-14,Arg-32 to Leu-40,Thr-44 to Glu-49,Thr-55 to Arg-64,Gln-92 to Ala-98,Ser-125 to Glu-132.40HHFBP4794666850825-253648Cys-9 to Arg-14,AR089: 5, AR061: 5Arg-21 to Gly-28.L0748: 2, L0749: 2,H0085: 1, H0050: 1,H0090: 1 and L0758: 1.41HCCCC81108355351562-2 649AR089: 12, AR061: 7H0583: 1, H0675: 1and H0457: 1.949062369139-921967Gly-8 to Ile-13,Glu-141 to His-146,Glu-186 to Glu-195,Asp-213 to Asn-218.42HPJEV71119784152976-164650Gln-49 to Pro-66,AR089: 7, AR061: 6Ser-96 to Thr-108,H0521: 2, H0039: 1,Glu-116 to Glu-135,H0641: 1, H0529: 1,Arg-140 to Pro-152,L0654: 1, H0701: 1,Ser-167 to Arg-172,H0518: 1, S0152: 1 andPro-175 to Leu-185,H0522: 1.Ala-199 to Lys-215,Pro-228 to Leu-237,Pro-247 to Ser-253.949153370 269-2029968Ser-5 to Gly-20.43HTEIL07113612153 43-1143651Glu-13 to Asp-29,AR061: 1, AR089: 0Glu-50 to Lys-58,L0758: 4, L0617: 2,Thr-61 to Glu-66,L0794: 2, H0253: 1,Ala-94 toTyr-100,H0038: 1, H0616: 1,Gln-146 to Ser-156,L0789: 1 and L0779: 1.Pro-171 to Asp-177,Ile-179 to Trp-191,Glu-197 to Val-203,Asp-238 to Lys-244,Pro-304 to Ala-315.953803371 42-443969Glu-13 to Thr-27.44HTEAG4995461454510-208652AR089: 1, AR061: 0L0759: 4, L0770: 2,S0040: 1, S0318: 1,S0334: 1, S0316: 1,S0340: 1, H0038: 1,L0598: 1, L0800: 1 andS0276: 1.45HSLCF9663767055 355-1248653Val-2 to Trp-7,AR054: 38, AR050:Lys-9 to Trp-18,26, AR051: 25, AR061:Gln-20 to Gly-25,2, AR089: 1Gln-79 to His-85,S0028: 1Pro-134 to Asp-139,Asp-164 to Thr-171,Pro-223 to Arg-228.9547773721332-430 970Thr-1 to Cys-6,Ser-52 to Gly-57,Gln-111 to His-117.46HNHCI3286167356183-593654Lys-17 to Thr-23,AR051: 23, AR050:His-95 to Thr-101.14, AR061: 10, AR054:4, AR089: 3S0053: 1956105373963-553971Lys-17 to Thr-23,His-95 to Thr-101.47HPMFL08105068457191-391655AR089: 1, AR061: 1H0031: 2959569374191-346972Met-43 to Trp-52.48HTXRA1395962258 3-1091656Ser-1 to Ser-6,AR061: 1, AR089: 0Thr-14 to Gly-28.H0556: 2, L0756: 2,H0423: 2, S0134: 1,H0580: 1, H0271: 1,T0006: 1, H0264: 1,H0560: 1, H0641: 1,S0142: 1, L0805: 1,L0809: 1, L0789: 1,H0555: 1, L0780: 1 andS0031: 1.49HCE3H71119789859410-988657His-14 to Gly-19,AR089: 14, AR061: 10Pro-21 to Pro-28,L0439: 12, L0438: 5,Arg-49 to Gln-54,L0741: 4, H0052: 2,Pro-82 to Pro-91,H0009: 2, L0769: 2,Gly-102 to Ser-108,L0794: 2, H0229: 1,Arg-150 to Ser-155,H0572: 1, L0770: 1,Pro-160 to Asn-168,L0796: 1, L0789: 1 andAla-175 to Glu-188.L0786: 1.961681375275-82697350HUTSF11113491460 3-323658Pro-4 to Glu-13,AR089: 0, AR061: 0Asn-23 to Arg-29,S0464: 1 and L0356: 1.Gln-91 to Arg-100.966029376 3-302974Glu-1 to Glu-6,Asn-16 to Arg-22.51HTEGI48102123561 1-549659Arg-1 to Ser-12,AR089: 4, AR061: 1Leu-33 to Leu-40,H0038: 2, H0556: 1,His-42 to Phe-49,H0341: 1 and L0596: 1.Glu-51 to Met-57,Gly-72 to Phe-78.530595377 32-523975Val-3 to Tyr-15,Leu-17 to Thr-27,Ser-34 to Ser-61,Leu-82 to Leu-89,His-91 to Phe-98,Glu-100 to Met-106,Gly-121 to Phe-127.52HSFAM09115391362152-295660Arg-1 to Ser-8,AR061: 5, AR089: 2Lys-42 to Lys-48.H0154: 2573345378147-332976Arg-1 to Ser-8,Lys-42 to Lys-48.53HNFHK77118228663165-662661Pro-38 to Lys-43,AR061: 1, AR089: 0Glu-126 to Tyr-132,H0271: 2Trp-161 to Arg-166.576186379165-422977Pro-38 to Lys-43.54HFXDO83101260264 12-608662AR089: 1, AR061: 1S0001: 3578847380 16-207978Arg-1 to Cys-10.55HSDIW73110440665 1-1068663Leu-4 to Pro-9,AR051: 4, AR054: 1,Ile-64 to Arg-69,AR089: 1, AR061: 0,Asn-142 to Pro-147,AR050: 0Pro-349 to Asp-356.S0046: 1, S0028: 1,S0031: 1 and S0260: 1.587311381 2-1075979Leu-6 to Pro-11,Ile-66 to Arg-71,Asn-144 to Pro-149,Pro-351 to Asp-358.9548213821595-549 980Ile-57 to Arg-62,Asn-135 to Pro-140,Pro-342 to Asp-349.56HFVGD23119964566 2-1003664Ser-4 to Arg-9,AR089: 5, AR061: 3Glu-41 to Ser-53,S0250: 4, L0745: 2,Arg-67 to Ser-72,H0393: 1, H0587: 1,Asn-111 to Arg-122,L0744: 1, L0748: 1,Gly-212 to Arg-218,L0439: 1 and L0752: 1.Gln-229 to Ser-235,Arg-239 to Lys-244.676214383 2-478981Ser-4 to Arg-9,Glu-41 to Ser-53,Arg-67 to Ser-72,Asn-111 to Arg-122.57HMSBZ24108236767 2-718665Lys-79 to ASp-87,AR061: 4, AR089: 2Lys-100 to Asp-106.H0331: 1, S0002: 1,H0519: 1 and L0741: 1.678707384 2-32298258HWHHB69121704268 3-2186666Thr-1 to Ser-10,AR089: 1, AR061: 1Ala-73 to Tyr-80,L0803: 3, S0354: 2,Arg-133 to Ser-143,H0052: 2, H0617: 2,Gly-174 to His-179,L0770: 2, L0646: 2,Ser-201 to Arg-224,S0028: 2, L0753: 2,Asn-236 to Gly-241,H0445: 2, H0556: 1,Tyr-260 to Cys-272,S6024: 1, H0657: 1,Pro-274 to Thr-284,S0418: 1, S0420: 1,Gln-292 to Glu-306,H0351: 1, H0441: 1,Cys-409 to Arg-414,H0586: 1, H0013: 1,Arg-424 to Arg-432,S0280: 1, H0156: 1,Asp-523 to His-531,L0021: 1, H0122: 1,Thr-552 to Pro-557,S0010: 1, H0571: 1,Asn-601 to Pro-606,L0163: 1, H0135: 1,His-612 to His-618,H0412: 1, H0100: 1,Pro-678 to His-684,L0351: 1, L0769: 1,Asn-698 to Gln-705.L0639: 1, L0764: 1,L0649: 1, L0659: 1,L0809: 1, L0530: 1,H0520: 1, H0547: 1,H0519: 1, H0690: 1,H0539: 1, S0136: 1,H0696: 1, L0748: 1,L0747: 1, L0756: 1,L0779: 1, L0757: 1,S0434: 1, S0436: 1,S0011: 1 and H0136: 1.690442385 1-261983Gly-1 to Ser-7.59HFXLC69116254369 16-939667Gly-49 to Gly-60,AR061: 1, AR089: 1Arg-84 to Cys-97,H0052: 2, H0135: 2,Pro-100 to Gln-106,S0282: 1, H0254: 1,Ala-113 to Ala-137,H0051: 1, H0634: 1,Ala-145 to Trp-156,S0152: 1, H0436: 1 andAla-172 to Tyr-182,H0677: 1.Asn-218 to Tyr-225.692773386 3-23098460HBXBW40115676570124-588668Ser-1 to Ser-12,AR089: 16, AR061: 8Arg-33 to Arg-50,S0038: 2, H0438: 1,Tyr-117 to Leu-125.S0049: 1 and H0547: 1.706115387124-456985Gln-3 to Ser-12,Arg-33 to Arg-50,Ser-93 to Glu-98.61HCE1L51114049871 3-434669AR089: 4, AR061: 3S0206: 2, H0052: 1,T0010: 1 and H0604: 1.715899388 3-43498662HRADM45114804672 2-472670Lys-1 to Leu-6,AR089: 14, AR061: 6Asp-25 to Pro-30.H0555: 1 and L0777:1.717358389 2-472987Lys-1 to Leu-6,Asp-25 to Pro-30.63HTEFO45115391873 61-597671Leu-59 to Thr-82,AR061: 6, AR089: 3Lys-89 to Gly-94,L0758: 2, S0222: 1,Gln-155 to Val-161,H0038: 1 and H0539: 1.Lys-169 to Ala-179.723446390 3-482988Gln-6 to Lys-14,Leu-68 to Glu-90.64HOHBN82115227174 2-1465672Asn-41 to Pro-50,AR089: 1, AR061: 0Asp-60 to Glu-71,S0002: 2, H0521: 2,Leu-121 to Pro-130,S0360: 1, H0123: 1,Glu-139 to Thr-144,S0250: 1, L0654: 1,Asp-155 to Ser-166,S0152: 1, L0740: 1 andGly-174 to Leu-187,L0749: 1.Pro-194 to Pro-199,Arg-211 to Asp-228,Gly-256 to Phe-270,Asp-334 to Gly-344,Gly-356 to Val-373,Glu-398 to Met-403,Asn-418 to Leu-424,Glu-444 to Trp-451,Cys-465 to Tyr-474.724322391 3-629989Asn-41 to Pro-50,Asp-60 to Glu-71,Leu-121 to Pro-130,Glu-139 to Thr-144,Asp-155 to Ser-166,Gly-174 to Asp-188.65HWHGF52121702675834-1 673Lys-1 to Ala-6,AR089: 1, AR061: 0Ser-38 to Gln-43,L0776: 5, L0764: 4,Pro-88 to Ala-112,L0743: 4, L0740: 3,Pro-141 to Asp-148,L0750: 3, L0777: 3,Gly-186 to Thr-200,L0731: 3, S0001: 2,Pro-231 to Ala-238,H0438: 2, H0052: 2,Leu-248 to Ser-254.H0194: 2, H0201: 2,L0526: 2, H0144: 2,L0742: 2, H0662: 1,H0619: 1, H0261: 1,H0392: 1, H0455: 1,H0586: 1, H0587: 1,H0574: 1, H0486: 1,H0013: 1, H0427: 1,S0010: 1, S0346: 1,T0110: 1, H0009: 1,L0157: 1, H0320: 1,H0051: 1, T0006: 1,H0604: 1, H0163: 1,H0646: 1, L0763: 1,L0638: 1, L0630: 1,L0646: 1, L0773: 1,L0651: 1, L0523: 1,L0805: 1, L0666: 1,L0663: 1, L0664: 1,H0547: 1, H0660: 1,S0404: 1, L0744: 1,L0439: 1, L0752: 1,S0434: 1 and L0595: 1.726102392 1-453990Gln-1 to Lys-8,Gly-10 to Trp-17,Val-28 to Gly-43,Thr-54 to Glu-63.66HBKDI301223861761115-321 674AR089: 1, AR061: 0S0364: 3, S0366: 3,L0604: 3, H0624: 1,L0622: 1, L0623: 1,H0041: 1, L0791: 1,S0380: 1 and L0748: 1.729048393 1-381991Gly-15 to Thr-21,Glu-76 to Lys-86.67HSQFR54118514377 1-1170675Ser-116 to Asp-125,AR061: 4, AR089: 2Glu-183 to Ser-188,H0650: 2, H0052: 2,Arg-228 to Gln-234,H0547: 2, H0542: 2,Leu-280 to Lys-303.S0212: 1, S0222: 1,T0114: 1, L0483: 1,H0628: 1, L0455: 1,H0413: 1, S0344: 1,L0766: 1, L0775: 1,L0805: 1, L0665: 1,H0520: 1, H0519: 1,S0126: 1, H0521: 1,S0044: 1, S0390: 1,L0592: 1 and S0026: 1.730964394 2-319992Glu-41 to Ser-46.68HAGBA56110259378 1-735676Pro-2 to Gly-10.AR061: 2, AR089: 1S0010: 1, H0135: 1,L0766: 1, L0745: 1,L0779: 1 and L0758: 1.732597395115-633993Asp-52 to Leu-57,Lys-82 to Thr-87,Ser-90 to Trp-98,Ser-118 to Leu-123.69HHSAE29122085179563-985677His-13 to Asn-24.AR061: 2, AR089: 1S0282: 1 and S0051: 1.743166396 2-32599470HMSHO6474658280 1-411678Ser-11 to Ser-21,AR089: 2, AR061: 2Ser-84 to Ala-89,S0002: 2Pro-98 to Arg-107.71HFPBW22115478681762-376679Lys-1 to Ala-6,AR061: 4, AR089: 2Ala-17 to Leu-25,S0222: 1, S0280: 1,Arg-54 to Ala-59,L0774: 1, L0376: 1 andVal-61 to Arg-66,S0378: 1.Ser-90 to Gly-95.750631397224-61999572HTLBH671224371822886-2005680Lys-31 to Ala-48,AR061: 2, AR089: 1Gln-51 to Thr-62,L0752: 3, L0747: 2,Gln-105 to Ser-110,H0294: 1, H0253: 1,Cys-126 to Leu-134,H0046: 1, H0040: 1,Gln-137 to Gly-150,H0063: 1, H0494: 1,His-174 to Ala-205,S0352: 1, L0769: 1,Arg-212 to Pro-220,L0766: 1, L0804: 1,Pro-227 to Gly-232,L0805: 1, L0791: 1,Gly-245 to Ala-251,H0521: 1, L0779: 1,Ala-257 to Ser-263,L0780: 1, L0731: 1 andLeu-266 to His-283.L0758: 1.751985398 1-28299673HNTMH70114352383 2-679681Pro-1 to Glu-6,AR089: 0, AR061: 0His-17 to Lys-22,H0520: 1Pro-52 to Gln-58,Gly-123 to Arg-130,His-205 to Ala-210.757184399 2-688997Pro-1 to Glu-6,His-17 to Lys-22,Pro-52 to Gln-58.74HCETC59118333484105-326682AR089: 1, AR061: 1H0052: 1 and H0194:1.761881400105-32699875HE8UX76116122385 2-1237683Asp-8 to Ala-13,AR061: 1, AR089: 1Ala-26 to Arg-33,H0539: 4, L0439: 4,Pro-38 to Ala-50,L0438: 2, H0013: 1,Pro-60 to Asn-65,L0758: 1 and L0592: 1.Asp-68 to Ser-74,Arg-109 to Arg-132,Asp-140 to Leu-145,Ala-149 to Ser-154,Ile-158 to Asp-169,Glu-171 to Ala-177,Cys-213 to Pro-218,Pro-226 to Lys-231,Thr-244 to Phe-249,Arg-361 to Ile-370.767871401214-798999Arg-39 to Arg-62,Asp-70 to Leu-75,Ala-79 to Ser-84,Ile-88 to Asp-99,Glu-101 to Ala-107.76HTLEN77113612486 3-947684His-1 to Thr-6,AR089: 33, AR061: 18Arg-30 to Thr-35,L0748: 2 and H0253:Lys-40 to Ala-71,1.Pro-209 to Glu-222,Arg-231 to Tyr-237,Pro-239 to Tyr-245,Arg-263 to Ala-271,Gln-290 to Trp-306.77236340290-4221000Ala-1 to Ala-35.77HBGDI80112469587 2-523685Asp-44 to Ile-50,AR061: 13, AR089: 5Arg-121 to Leu-132,L0805: 2, H0436: 2,Lys-148 to Ser-160.L0439: 2, L0362: 2,S0358: 1, L0483: 1,H0181: 1, S0422: 1,L0369: 1, L0804: 1,L0787: 1 and L0663: 1.781600403 1-4291001Asp-13 to Ile-19,Pro-37 to Arg-42.78HELHB88122563288 138-3230686Gln-1 to Thr-7,AR061: 2, AR089: 2Glu-28 to Gln-35,L0777: 3, L0794: 2,Lys-188 to Lys-207,S0027: 2, L0748: 2,Ser-238 to Gly-245,L0747: 2, L0601: 2,Asp-278 to Gly-283,S0342: 1, S0212: 1,Pro-317 to Ser-324,S0282: 1, L0004: 1,Ser-335 to Glu-342,S0045: 1, H0581: 1,Pro-344 to Lys-355,T0110: 1, L0471: 1,Glu-362 to Asn-373,S6028: 1, H0551: 1,Glu-385 to Arg-393,H0494: 1, H0509: 1,Arg-399 to Gln-417,L0646: 1, L0665: 1,Lys-422 to Gln-457,H0520: 1, H0547: 1,Glu-461 to Glu-477,S0390: 1, L0591: 1,Leu-514 to Glu-529,L0366: 1 and H0653: 1.Leu-538 to Met-548,Gln-562 to Gln-567,Asn-569 to Asp-574,Arg-594 to Gln-609,Asn-626 to Met-636,Ala-638 to Lys-649,Glu-654 to Gln-670,Gln-676 to Leu-716,Ser-736 to Gly-741,Phe-762 to Asp-768,Glu-808 to Val-815,Ser-847 to Trp-856,Asp-858 to Trp-876,Gln-892 to Ala-898,Pro-964 to Ile-976.811935404136-5671002Arg-1 to His-10.79HTEMV66115226189 1-669687Ile-33 to Ser-40,AR061: 5, AR089: 1Val-63 to Gln-69,H0616: 1 and L0758:Phe-84 to Ser-94,1.Lys-205 to Lys-212.813038405 1-3181003Ser-38 to Pro-45.80HMTAJ73106398990704-3 688Gly-5 to Gln-15,AR061: 24, AR089: 14Gly-104 to Gly-111,L0806: 3, L0772: 2,Gly-136 to Asp-141,L0648: 2, H0255: 1,His-228 to Pro-234.L0717: 1, H0586: 1,H0599: 1, H0618: 1,H0581: 1, H0052: 1,H0123: 1, L0629: 1,L0659: 1, L0663: 1,S0330: 1, H0518: 1 andH0555: 1.813296406 1-4381004Pro-23 to Lys-28,Gln-39 to Thr-51,Lys-93 to Ala-106,Gln-112 to Pro-129,Pro-132 to Pro-143.81HE9TD3181584591 3-674689Ser-119 to Thr-127,AR089: 0, AR061: 0Gln-134 to Ser-152.S0354: 1, L0657: 1,H0144: 1 and S0330: 1.82HGBDG55114136392 26-535690Gly-1 to Ala-8,AR089: 1, AR061: 1Phe-31 to Leu-36,S0040: 1, H0014: 1,Glu-54 to Lys-62,H0030: 1, H0063: 1,Gly-69 to Gly-75,L0803: 1, H0521: 1 andLeu-100 to Gly-106,S0028: 1.Ser-125 to Lys-131.815858407 26-4001005Gly-1 to Ala-8,Phe-31 to Leu-36,Glu-54 to Lys-62,Gly-69 to Gly-75,Leu-100 to Gly-106.83HOUHL51112591493 2-508691Ala-12 to Trp-19,AR061: 6, AR089: 2Ala-21 to Arg-27,L0758: 6, L0794: 3,Glu-38 to Phe-49,H0038: 2, L0768: 2,Arg-90 to Val-102,L0790: 2, L0731: 2,Glu-144 to Gly-151,S0342: 1, H0664: 1,Tyr-164 to Ala-169.H0616: 1, S0210: 1,L0773: 1 and L0608: 1.815891408 3-527100684HEOPP67102011994 1-972692Asp-27 to Val-32,AR061: 1, AR089: 0Asp-66 to Gly-71.H0457: 2, H0650: 1and H0622: 1.827630409 2-4481007Arg-8 to Arg-17,Asp-47 to Val-52,Asp-86 to Gly-91.85HKAOV71116542395 1-732693AR089: 6, AR061: 2H0013: 2, H0046: 2,H0036: 1, H0590: 1,H0581: 1, H0551: 1 andH0494: 1.827679410 1-732100886HDQID90113775296 2-361694Leu-12 to Gln-21.AR089: 7, AR061: 3L0766: 14, H0521: 4,L0748: 4, L0804: 3,L0776: 3, L0749: 3,L0731: 3, L0485: 3,S0376: 2, H0580: 2,L0483: 2, H0316: 2,S0002: 2, L0803: 2,L0775: 2, L0805: 2,L0659: 2, L0438: 2,H0265: 1, H0686: 1,H0656: 1, H0341: 1,S0212: 1, H0638: 1,H0125: 1, S0360: 1,H0411: 1, S0222: 1,H0409: 1, H0587: 1,H0014: 1, S0003: 1,H0163: 1, H0591: 1,H0488: 1, H0494: 1,H0641: 1, L0598: 1,H0529: 1, L0772: 1,L0764: 1, L0768: 1,L0774: 1, L0655: 1,L0783: 1, L0809: 1,L0792: 1, L0663: 1,L0665: 1, H0702: 1,H0519: 1, S0126: 1,H0682: 1, H0435: 1,H0672: 1, H0704: 1,S3012: 1, L0751: 1,L0750: 1, L0777: 1,L0752: 1, L0757: 1,L0758: 1, L0759: 1,L0362: 1, H0423: 1 andH0506: 1.831976411221-724100987HFRBN811182552972331-598 695AR050: 10, AR051: 3,AR061: 1, AR089: 1,AR054: 0S0028: 4, S0001: 2,S0278: 2, S0050: 2,S0282: 1, H0632: 1,H0271: 1, H0416: 1,H0027: 1, S0038: 1,S0052: 1, S0053: 1,H0684: 1 and S0044: 1.833061412 1-11161010Gly-113 to Gly-119,Gln-173 to Thr-181,Ala-362 to Pro-368.973206413 1-4351011973208414138-5871012Leu-51 to Ser-62.88HFKJW01118713498487-8 696AR089: 1, AR061: 0H0620: 2, H0012: 1,S0152: 1 and S0260: 1.836491415 3-440101389HSDFL631219300992590-1844697Ser-83 to Lys-88,AR061: 402, AR089:Pro-95 to Asn-112,142Arg-180 to Asp-185,H0038: 7, L0758: 5,Met-231 to Arg-240.H0616: 4, L0731: 4,S0002: 3, L0637: 3,H0623: 2, L0794: 2,L0809: 2, L0663: 2,H0522: 2, L0779: 2,L0777: 2, S0046: 1,H0431: 1, T0060: 1,H0013: 1, S0010: 1,H0545: 1, H0050: 1,S0023: 1, S0003: 1,H0328: 1, H0135: 1,H0163: 1, H0412: 1,H0102: 1, H0100: 1,T0042: 1, L0768: 1,L0803: 1, L0375: 1,L0542: 1, L0647: 1,L0367: 1, L0791: 1,L0664: 1, H0693: 1,S0328: 1, S0168: 1,S0031: 1 and H0008: 1.836498416 1-2491014Glu-1 to Asp-7,Met-53 to Met-60,Ile-78 to Ser-83.90HPJET90836503100 39-377698His-8 to Gly-18,AR061: 0, AR089: 0Leu-40 to Ile-45,S0152: 2Asn-100 to Asp-105.91HEMFC61836514101 1-714699Glu-4 to Ser-9,AR061: 4, AR089: 3Ser-58 to Arg-65.H0038: 7, L0758: 5,H0616: 4, L0731: 4,S0002: 3, L0637: 3,H0623: 2, L0794: 2,L0809: 2, L0663: 2,H0522: 2, L0779: 2,L0777: 2, S0046: 1,H0431: 1, T0060: 1,H0013: 1, S0010: 1,H0545: 1, H0050: 1,S0023: 1, S0003: 1,H0328: 1, H0135: 1,H0163: 1, H0412: 1,H0102: 1, H0100: 1,T0042: 1, L0768: 1,L0803: 1, L0375: 1,L0542: 1, L0647: 1,L0367: 1, L0791: 1,L0664: 1, H0693: 1,S0328: 1, S0168: 1,S0031: 1 and H0008: 1.92HDTBR501174351102467-234700Thr-26 to Arg-31,AR089: 41, AR061: 4Gly-73 to Trp-78.H0486: 2846630417130-3421015Ala-2 to Glu-7,Arg-50 to Glu-58.93HACCH94847143103 1-897701Gly-1 to Ser-6,AR061: 4, AR089: 2Arg-76 to Gln-88,L0754: 6, L0766: 3,Lys-113 to Ser-119,L0731: 2, H0624: 1,Tyr-125 to Lys-132,H0170: 1, S0116: 1,Ser-167 to Tyr-179,S0280: 1, H0545: 1,Arg-263 to Tyr-281,T0006: 1, S0344: 1,Ser-294 to Thr-299.S0426: 1, L0770: 1,L0790: 1, L0748: 1,L0756: 1, L0779: 1,L0589: 1 and L0462: 1.94HE8T1391223481104 441-1166702Thr-21 to Trp-26,AR061: 2, AR089: 2Thr-72 to Val-88,L0438: 4, L0746: 4,Arg-115 to Tyr-127.H0581: 2, H0656: 1,H0013: 1, L0471: 1,H0266: 1, H0328: 1,H0553: 1, S0438: 1,H0529: 1, L0766: 1,L0805: 1, H0520: 1,H0521: 1, L0752: 1 andS0192: 1.849161418 3-4701016Ser-8 to Thr-15,Arg-73 to Thr-79,Phe-86 to Leu-92.95HEGAP321140393105525-1 703AR061: 6, AR089: 3H0550: 1, H0494: 1and L0659: 1.851207419 2-3971017Arg-1 to Thr-14.96HCWFU66853005106 3-311704AR061: 1, AR089: 1H0305: 1 and H0589:1.97HUSY129853149107 3-470705Gln-57 to Ile-67,AR061: 5, AR089: 2Asp-77 to Asp-85.H0650: 2, H0052: 2,H0547: 2, H0542: 2,S0212: 1, S0222: 1,T0114: 1, L0483: 1,H0628: 1, L0455: 1,H0413: 1, S0344: 1,L0766: 1, L0775: 1,L0805: 1, L0665: 1,H0520: 1, H0519: 1,S0126: 1, H0521: 1,S0044: 1, S0390: 1,L0592: 1 and S0026: 1.98HMEFT661134131108 82-933706Gly-61 to Glu-67,AR061: 1, AR089: 1Ala-88 to Gly-96,H0175: 1, H0266: 1,Gly-127 to Trp-137.H0292: 1, H0628: 1 andL0779: 1.856149420 2-349101899HKAAR71863023109 54-977707Gln-1 to Gln-6,AR061: 1, AR089: 0Ser-24 to Thr-31,H0494: 2, H0693: 2,Pro-57 to Gln-63,H0521: 2, H0580: 1,Ala-96 to Met-104,H0253: 1, H0628: 1,Asn-124 to Lys-133,H0522: 1 and H0422: 1.Ser-172 to Trp-182,Ser-186 to Glu-194,Pro-286 to Pro-294.100H7TBC95865922110 3-704708Gln-154 to Ser-163.AR089: 1, AR061: 1S0198: 57, S0274: 12,S0252: 4, S0270: 3,S0264: 1, S0268: 1 andS0228: 1.908115421 3-7041019Gln-154 to Ser-163.101HAPPX52637493111 827-1222709Ser-59 to Ile-66,AR050: 16, AR054:Arg-73 to Gly-85.10, AR051: 5, AR061:2, AR089: 1H0305: 4, S0282: 1,H0575: 1, H0150: 1 andH0617: 1.872075422 3-3591020872076423400-2 1021His-73 to Phe-81,Thr-92 to Trp-102.102HBGSJ131152326112 1-756710Lys-1 to Lys-6,AR089: 1, AR061: 0Gln-25 to Asp-36,H0617: 2, H0013: 1,Ser-85 to Ile-96,H0271: 1, L0455: 1 andVal-115 to Ser-136,H0539: 1.Lys-172 to Trp-177,Pro-188 to Phe-201,Gly-217 to Ala-224,Asn-230 to Gly-239.878322424 1-6841022103HFKLX38880220113 3-308711AR089: 1, AR061: 1H0620: 2104HTLGP151165362114196-786712Gly-2 to Thr-10,AR089: 4, AR061: 3Glu-160 to Gly-175,T0010: 3, S0049: 2,Thr-189 to Glu-197.H0052: 2, L0415: 1,H0618: 1 and S0010: 1.880297425189-6621023Gly-2 to Thr-10,Glu-99 to Gly-104.105HMEGH461092158115 2-547713Asp-13 to Asp-19,AR054: 29, AR051:Lys-76 to Asp-84.12, AR061: 6, AR089:3, AR050: 2H0196: 1 and H0266:1.887791426 1-3151024Asp-13 to Asp-19,Lys-76 to Asn-83.106HE8PY291129488116 2-277714Asp-28 to Ser-36,AR061: 2, AR089: 1Glu-47 to Gln-60,H0013: 1 and S0126: 1.Phe-68 to Gly-77,Pro-81 to Val-86.887862427 2-2771025Asp-28 to Ser-36,Glu-47 to Gln-60,Phe-68 to Gly-77,Pro-81 to Val-86.107HTDAB17890384117 3-605715Asp-1 to Gly-14,AR089: 1, AR061: 06p24-p23125264,Ala-60 to Lys-71,L0747: 28, L0588: 22,134570,Gln-101 to Glu-118.L0757: 19, H0251: 15,600511,S0358: 14, S0045: 13,601556L0731: 12, H0551: 10,H0412: 10, L0771: 10,L0748: 9, L0758: 9,H0506: 9, H0556: 8,S0046: 8, H0622: 8,H0013: 7, H0623: 7,L0662: 7, S0192: 7,S0003: 6, L0659: 6,L0666: 6, S0328: 6,L0439: 6, L0750: 6,L0759: 6, L0599: 6,L0608: 6, S0040: 5,S0360: 5, H0581: 5,H0529: 5, L0763: 5,L0764: 5, L0664: 5,H0144: 5, S0026: 5,S0212: 4, H0486: 4,H0674: 4, L0776: 4,S0126: 4, H0672: 4,S0136: 4, L0752: 4,H0624: 3, S0420: 3,H0599: 3, H0004: 3,H0266: 3, H0615: 3,H0031: 3, H0553: 3,H0591: 3, H0264: 3,H0413: 3, H0494: 3,S0210: 3, L0770: 3,L0806: 3, H0519: 3,H0435: 3, L0740: 3,L0751: 3, L0749: 3,H0170: 2, H0657: 2,H0656: 2, S0356: 2,S0408: 2, H0619: 2,H0393: 2, H0333: 2,T0040: 2, H0427: 2,S0280: 2, H0156: 2,H0318: 2, H0596: 2,T0110: 2, H0545: 2,H0046: 2, H0009: 2,H0050: 2, L0471: 2,H0188: 2, H0328: 2,H0428: 2, L0483: 2,H0644: 2, H0038: 2,S0426: 2, L0772: 2,L0646: 2, L0766: 2,L0649: 2, L0651: 2,L0655: 2, L0789: 2,L0663: 2, L0665: 2,L0352: 2, H0658: 2,S0152: 2, H0521: 2,H0696: 2, S0404: 2,H0555: 2, S0028: 2,H0445: 2, L0591: 2,L0594: 2, H0543: 2,H0422: 2, H0171: 1,H0265: 1, S6024: 1,H0295: 1, T0049: 1,S0134: 1, H0661: 1,H0663: 1, H0664: 1,S0418: 1, L0005: 1,S0354: 1, S0376: 1,S0468: 1, H0351: 1,S0220: 1, H0431: 1,H0392: 1, H0403: 1,H0592: 1, H0587: 1,H0642: 1, H0574: 1,H0485: 1, L0021: 1,H0575: 1, H0274: 1,S0346: 1, T0048: 1,S0049: 1, H0434: 1,H0230: 1, H0196: 1,H0052: 1, H0263: 1,H0597: 1, H0572: 1,H0012: 1, H0620: 1,T0003: 1, H0024: 1,H0057: 1, H0051: 1,H0083: 1, H0510: 1,H0687: 1, H0288: 1,S0022: 1, H0039: 1,H0030: 1, H0628: 1,H0166: 1, H0212: 1,H0135: 1, H0163: 1,H0090: 1, H0040: 1,H0634: 1, H0087: 1,H0477: 1, H0488: 1,H0433: 1, H0268: 1,H0269: 1, H0056: 1,S0038: 1, H0100: 1,H0429: 1, S0450: 1,H0132: 1, H0633: 1,S0472: 1, H0647: 1,H0646: 1, H0652: 1,S0344: 1, L0640: 1,L0371: 1, L0372: 1,L0374: 1, L0767: 1,L0768: 1, L0364: 1,L0794: 1, L0650: 1,L0375: 1, L0378: 1,L0606: 1, L0656: 1,L0783: 1, L0647: 1,S0374: 1, T0068: 1,L0438: 1, H0547: 1,H0689: 1, H0711: 1,H0684: 1, H0659: 1,H0670: 1, H0648: 1,S0330: 1, S0378: 1,S0380: 1, H0709: 1,S0146: 1, S3012: 1,S0037: 1, S0206: 1,L0742: 1, L0744: 1,L0755: 1, H0707: 1,S0434: 1, S0436: 1,L0584: 1, L0593: 1,L0362: 1, S0011: 1,S0424: 1 and H0293: 1.108HCFCF471199931118 1-780716Arg-8 to Glu-15,AR089: 14, AR061: 7Gln-49 to Ala-58,H0341: 1 and H0422:Gly-175 to Gly-182,1.Arg-184 to Leu-191,Pro-198 to Phe-205.894415428 2-2981026Arg-1 to Glu-8.109HDQHB191194798119 2-709717Arg-1 to Thr-7,AR061: 3, AR089: 3Pro-19 to Ala-25,L0759: 12, L0439: 11,Pro-56 to Leu-64,L0766: 7, L0775: 5,His-72 to Asn-81,H0521: 5, L0755: 5,Phe-184 to Pro-192,L0748: 4, L0756: 4,Pro-218 to Val-226,L0777: 4, L0731: 4,Ser-229 to Arg-236.L0581: 4, L0619: 3,L0666: 3, L0779: 3,L0757: 3, L0588: 3,S0418: 2, L0618: 2,H0580: 2, L0055: 2,L0769: 2, L0773: 2,L0774: 2, L0791: 2,L0747: 2, L0750: 2,H0265: 1, H0663: 1,S0356: 1, H0208: 1,H0370: 1, H0108: 1,H0575: 1, H0618: 1,H0544: 1, H0545: 1,S0050: 1, H0510: 1,H0286: 1, H0031: 1,H0644: 1, H0068: 1,H0135: 1, L0564: 1,H0494: 1, L0475: 1,H0396: 1, S0144: 1,S0002: 1, S0426: 1,L0763: 1, L0761: 1,L0642: 1, L0764: 1,L0662: 1, L0768: 1,L0806: 1, L0661: 1,L0659: 1, L0367: 1,L0663: 1, H0519: 1,H0435: 1, H0658: 1,S3014: 1, L0751: 1,L0749: 1, L0603: 1,H0665: 1 and H0542: 1.895106429 2-5381027Pro-14 to Ala-20,Pro-51 to Leu-59,His-67 to Thr-77.110HAGDN531129154120 3-329718Gln-22 to Lys-30,AR050: 17, AR051:Phe-40 to Tyr-49,11, AR054: 2, AR089:Gln-70 to Trp-75,1, AR061: 0Arg-80 to Gln-87,S0010: 1 and S0027: 1.Gly-95 to Arg-101.895963430129-4281028Pro-9 to Gln-16,Phe-31 to Tyr-40,Gln-61 to Trp-66,Arg-71 to Gln-78,Gly-86 to Arg-92.111HUFDB741162672121 2-562719Gln-43 to Thr-58,AR061: 1, AR089: 1Asn-74 to His-79,H0575: 2, L0754: 2,Gly-109 to Trp-114,H0599: 1, T0048: 1,Asp-136 to Phe-145.L0163: 1, H0051: 1,H0188: 1, H0379: 1,L0438: 1, H0670: 1,H0672: 1, L0439: 1,L0747: 1, S0260: 1,L0591: 1 and H0506: 1.901451431 2-4121029Gln-43 to Thr-58,Asn-74 to His-79,Gly-109 to Trp-114.112HNHFH241092567122 28-516720His-8 to Gly-18,AR054: 20, AR050:Ala-39 to Gly-45,15, AR061: 7, AR089:Pro-94 to Glu-101,4, AR051: 1Pro-134 to Gly-142.S0053: 1903741432 28-4801030His-8 to Gly-18,Ala-39 to Gly-45,Pro-94 to Glu-101.113HBGQT031188175123 11-790721Glu-1 to Ala-12,AR061: 6, AR089: 3Glu-19 to Val-28,H0617: 10, L0665: 4,Glu-34 to Thr-45,H0333: 3, S0366: 3,Leu-140 to Asp-157,L0759: 3, H0599: 2,Thr-167 to Ala-198,L0648: 2, L0653: 2,Ala-211 to Asp-216.L0664: 2, H0519: 2,H0686: 1, H0484: 1,H0664: 1, H0392: 1,L0622: 1, S0280: 1,H0545: 1, T0010: 1,H0424: 1, H0031: 1,H0181: 1, H0708: 1,H0494: 1, H0633: 1,L0371: 1, L0764: 1,L0773: 1, L0768: 1,L0375: 1, L0651: 1,L0659: 1, L0783: 1,L0789: 1, L0438: 1,H0684: 1, H0670: 1,L0744: 1, L0780: 1,L0755: 1 and L0595: 1.908173433 3-7911031Lys-1 to Ala-15,Glu-22 to Val-31,Glu-37 to Thr-48,Leu-143 to Asp-160,Thr-170 to Ala-201,Ala-214 to Asp-219.114HETLF291103959124 3-482722Asp-119 to Tyr-124.AR061: 4, AR089: 2H0046: 1 and L0758:1.909762434 3-4161032115HOUGD291204931125 199-1821723Arg-9 to Gln-17,AR061: 6, AR089: 5Ile-33 to Asn-39,L0770: 4, L0789: 3,Gln-93 to Ser-104,L0439: 3, L0750: 3,Asp-141 to Leu-155,L0641: 2, L0747: 2,Ser-224 to Asn-234,L0758: 2, S0040: 1,Asn-243 to Lys-248,H0575: 1, T0010: 1,Ser-308 to Gln-320,H0087: 1, S0422: 1,Thr-350 to Glu-357,L0803: 1, L0375: 1,Ser-384 to Thr-390,L0776: 1, L0659: 1,Asp-435 to Ser-447,L0783: 1, H0144: 1,Ala-480 to Lys-487,L0352: 1, H0684: 1,Lys-496 to Leu-508,H0660: 1, S0027: 1,Ser-519 to Val-528,L0777: 1 and H0445: 1.Ser-533 to Gln-541.909797435199-9091033Arg-9 to Leu-15.116HTEMV091128254126 1-711724Asp-22 to Asp-28,AR089: 13, AR061: 13Leu-98 to Trp-103,L0666: 3, L0758: 3,Glu-123 to Trp-154,H0616: 2, L0779: 2,Pro-158 to Gln-178,S0036: 1, L0598: 1,Pro-180 to Met-189,L0766: 1, L0651: 1,Glu-207 to Lys-226,L0806: 1, L0776: 1,Ser-231 to Leu-237.H0144: 1, H0547: 1,H0672: 1 and H0555: 1.909843436 1-7111034Asp-22 to Asp-28,Leu-98 to Trp-103,Glu-123 to Trp-154.117HNTNB141128964127 3-614725Phe-2 to Gln-9,AR089: 1, AR061: 1Arg-22 to Val-29,S0007: 1, S0222: 1,Glu-51 to Leu-64,S0049: 1, L0438: 1,Tyr-73 to Ile-83,H0520: 1 and L0439: 1.Glu-98 to Thr-104,Ala-119 to Asp-126,Arg-155 to Ser-161.909942437 2-6581035Ala-2 to Gln-9,Arg-22 to Val-29,Glu-51 to Leu-64.118HE2KZ071149808128708-166726AR061: 9, AR089: 4H0624: 1909948438 2-7961036Leu-10 to Gly-16,Pro-37 to Glu-45,Glu-78 to Cys-87.119HSIGN571105444129 3-974727Val-10 to Gly-16,AR061: 2, AR089: 1Met-19 to Val-34,H0229: 1, H0590: 1,Ala-84 to Asp-90,S0049: 1, H0014: 1,Met-107 to Trp-120,H0560: 1, L0439: 1 andGln-191 to Ala-201,H0543: 1.Glu-223 to Val-229,Asn-309 to Gly-314.910078439 2-7601037Val-10 to Gly-16,Met-19 to Val-34.120HLHBC301106654130 39-512728Gln-7 to Glu-17,AR061: 8, AR089: 5Thr-36 to Asn-42,L0456: 3, H0024: 2Val-44 to Phe-49,and L0747: 1.Tyr-76 to Ile-85,Cys-94 to Glu-99,Pro-105 to Ser-110.910079440 39-5121038Gln-7 to Glu-17.121HFBDJ131195217131 172-1368729Ser-45 to Glu-53,S0007: 2, L0794: 2,Ile-78 to Asn-94,S0434: 2, S0354: 1,Leu-99 to Ser-104,N0006: 1, H0622: 1 andSer-110 to Trp-128,H0478: 1.Tyr-145 to Gly-153,Gln-168 to Trp-173,Leu-196 to Ala-205.911264441 3-4101039Ser-6 to Trp-24.122HTPGG251217208132 786-1628730Glu-29 to Arg-35,AR061: 2, AR089: 2Arg-50 to Leu-55,L0439: 6, L0777: 6,Leu-60 to Ser-69,H0052: 4, L0748: 4,Lys-102 to Asp-108,H0634: 3, L0662: 3,Pro-227 to Glu-233,L0805: 3, L0659: 3,Leu-249 to Glu-261.L0438: 3, H0547: 3,L0750: 3, L0758: 3,H0208: 2, H0123: 2,H0014: 2, H0617: 2,H0135: 2, L0769: 2,L0766: 2, L0803: 2,L0776: 2, L0666: 2,L0751: 2, L0745: 2,L0731: 2, H0265: 1,S0408: 1, H0549: 1,H0497: 1, L0622: 1,H0581: 1, H0194: 1,L0738: 1, H0546: 1,H0024: 1, S0362: 1,L0163: 1, T0010: 1,H0083: 1, H0510: 1,H0266: 1, H0428: 1,H0622: 1, H0673: 1,H0598: 1, S0036: 1,H0163: 1, H0413: 1,L0370: 1, T0041: 1,H0647: 1, L0637: 1,L0667: 1, L0772: 1,L0646: 1, L0800: 1,L0764: 1, L0649: 1,L0657: 1, L0809: 1,L0788: 1, L0663: 1,S0374: 1, H0520: 1,H0670: 1, H0666: 1,S0330: 1, H0539: 1,H0521: 1, H0696: 1,H0478: 1, S0028: 1,L0741: 1, L0747: 1,L0749: 1, L0780: 1,L0752: 1 and H0543: 1.911282442 3-3921040Pro-3 to Arg-8.123HSSMT34911294133 56-553731Glu-29 to Arg-35,AR061: 4, AR089: 3Arg-50 to Leu-55,L0439: 6, L0777: 6,Leu-60 to Ser-69,H0052: 4, L0748: 4,Lys-102 to Asp-108,H0634: 3, L0662: 3,Pro-133 to Gln-141.L0805: 3, L0659: 3,L0438: 3, H0547: 3,L0750: 3, L0758: 3,H0208: 2, H0123: 2,H0014: 2, H0617: 2,H0135: 2, L0769: 2,L0766: 2, L0803: 2,L0776: 2, L0666: 2,L0751: 2, L0745: 2,L0731: 2, H0265: 1,S0408: 1, H0549: 1,H0497: 1, L0622: 1,H0581: 1, H0194: 1,L0738: 1, H0546: 1,H0024: 1, S0362: 1,L0163: 1, T0010: 1,H0083: 1, H0510: 1,H0266: 1, H0428: 1,H0622: 1, H0673: 1,H0598: 1, S0036: 1,H0163: 1, H0413: 1,L0370: 1, T0041: 1,H0647: 1, L0637: 1,L0667: 1, L0772: 1,L0646: 1, L0800: 1,L0764: 1, L0649: 1,L0657: 1, L0809: 1,L0788: 1, L0663: 1,S0374: 1, H0520: 1,H0670: 1, H0666: 1,S0330: 1, H0539: 1,H0521: 1, H0696: 1,H0478: 1, S0028: 1,L0741: 1, L0747: 1,L0749: 1, L0780: 1,L0752: 1 and H0543: 1.124HWWDN341152430134 2-1012732Ser-15 to Leu-21,AR089: 1, AR061: 1Pro-24 to Val-30,S0354: 16, H0457: 7,Ser-91 to Lys-99,L0758: 3, H0555: 2,Thr-113 to Lys-120,H0170: 1, H0657: 1,Pro-168 to Gln-174,H0255: 1, H0662: 1,Glu-226 to Ser-231,S0360: 1, H0036: 1,Ser-296 to Gln-307,H0150: 1, H0051: 1,Asp-319 to Gly-328,H0553: 1, L0800: 1,Gly-330 to Ala-337.L0644: 1, L0771: 1,L0803: 1, L0787: 1,L0663: 1, H0144: 1,S0374: 1, H0670: 1,H0522: 1, L0749: 1,S0452: 1 and H0506: 1.911357443 2-10001041Ser-11 to Leu-17,Pro-20 to Val-26,Ser-87 to Lys-95,Thr-109 to Lys-116,Pro-164 to Gln-170,Glu-222 to Ser-227,Ser-292 to Gln-303,Asp-315 to Gly-324,Gly-326 to Ala-333.125HCEPW85911374135 3-314733Thr-2 to Gln-7.H0052: 1 and L0471:1.126HMTAW831071602136 1-363734Ile-26 to Trp-33,AR089: 0, AR061: 0Glu-52 to Leu-71.H0583: 1, H0644: 1,L0766: 1 and H0518: 1.911385444 1-3631042Ile-26 to Trp-33,Glu-52 to Leu-71.127HDMAV011194697137 1-657735Pro-1 to Glu-15,AR089: 2, AR061: 2Ala-26 to Lys-32,L0766: 5, L0776: 5,Glu-46 to Leu-65,L0754: 4, H0013: 3,Arg-82 to Cys-94,S0126: 3, L0742: 3,Leu-101 to Glu-107,L0750: 3, H0624: 2,Leu-146 to Asp-151,S0360: 2, H0560: 2,Gln-157 to Ser-162,L0769: 2, L0641: 2,Ser-165 to Ala-187,L0665: 2, S0330: 2,Phe-210 to Leu-217.L0756: 2, L0731: 2,L0759: 2, L0588: 2,H0171: 1, H0650: 1,H0402: 1, H0638: 1,H0340: 1, H0637: 1,H0351: 1, S0222: 1,H0581: 1, H0263: 1,H0545: 1, H0050: 1,S0051: 1, S0214: 1,H0039: 1, L0055: 1,H0090: 1, H0412: 1,H0022: 1, H0359: 1,H0561: 1, H0641: 1,L0770: 1, L0637: 1,L0646: 1, L0764: 1,L0773: 1, L0662: 1,L0768: 1, L0651: 1,L0653: 1, L0659: 1,L0792: 1, H0519: 1,H0522: 1, H0576: 1,S0028: 1, L0439: 1,L0740: 1, L0749: 1,L0777: 1, H0444: 1,L0596: 1, L0601: 1,H0542: 1 and H0543: 1.911386445 3-4281043Asp-1 to Glu-11,Ala-22 to Lys-28,Glu-42 to Leu-61,Arg-78 to Cys-90,Leu-97 to Glu-103.128HDPSR74911396138212-583736Leu-31 to Ser-39,AR050: 48, AR054:Val-57 to Trp-63,42, AR051: 35, AR089:Pro-103 to Gln-111,3, AR061: 1Leu-118 to Leu-124.H0575: 2, H0580: 1,S0002: 1, S0426: 1,H0521: 1, H0436: 1 andL0748: 1.129HHEZT5811606571391212-937 737Pro-4 to His-21,AR089: 8, AR061: 3Glu-35 to Gln-43.L0794: 3, L0758: 2,L0759: 2, H0624: 1,L0717: 1, T0082: 1,H0581: 1, H0553: 1,H0038: 1, T0067: 1,L0665: 1, H0436: 1,L0439: 1, L0745: 1 andH0543: 1.911416446 1-5581044Glu-9 to Lys-14,Gln-51 to Gln-57.130HTLDU05911649140 2-469738Pro-89 to Ala-97.AR061: 8, AR089: 79q31109400,H0253: 3, H0618: 1132800,and L0758: 1.132800,186855,223900,253800,253800,278700,602088131HTLET561189721141 144-1211739Ser-54 to Lys-61,AR061: 6, AR089: 5Pro-118 to Lys-128,H0253: 18, H0618: 7,Pro-233 to Val-244,L0794: 3, H0038: 1,Leu-262 to Ser-270,H0616: 1, L0788: 1 andSer-322 to Gly-344,L0758: 1.Pro-347 to Ser-353.911654447 95-8261045Ser-54 to Lys-61,Pro-118 to Lys-128,Thr-208 to Ser-213,Ser-218 to Ala-227,Pro-230 to Ser-236,Pro-238 to Ser-244.132HTLCA95911655142 38-1096740AR061: 373, AR089:188H0253: 3, H0618: 2,H0038: 2, H0616: 1 andL0758: 1.133HTEJT861090517143 1-1119741Ala-39 to Ala-45,AR089: 1, AR061: 1Gln-57 to Ser-63,L0794: 3, H0038: 2,Tyr-90 to Lys-95,H0265: 1, S0358: 1,Leu-113 to Ile-119.T0039: 1, H0616: 1,L0768: 1, L0804: 1,L0664: 1, L0777: 1,L0731: 1, L0758: 1 andL0465: 1.911656448 1-4111046Ala-39 to Ala-45,Gln-57 to Ser-63,Tyr-90 to Lys-95.134HTEMA541134919144 22-1359742Gly-23 to Asn-30,AR089: 19, AR061: 13Thr-58 to Val-79,H0618: 14, H0253: 12,Arg-101 to Ile-106,H0038: 11, H0616: 2,Thr-117 to Glu-126,L0794: 1, L0779: 1 andPro-184 to Lys-193,L0758: 1.Ile-298 to Val-303,Phe-381 to Leu-389.911666449 22-11671047Gly-23 to Asn-30,Arg-45 to Lys-50.135HTLGJ171135518145216-587743Gly-35 to Ser-44.AR089: 18, AR061: 5H0618: 5, H0549: 1and H0543: 1.915136450 93-4641048Val-6 to Arg-12.136HOUES64918119146 3-317744Pro-89 to Leu-102.AR061: 1, AR089: 0S0040: 1 and S0278: 1.137HMSCD15982250147507-1 745Asp-16 to Gln-22,AR089: 1, AR061: 1Val-44 to Ser-57.S0002: 2 and L0766: 1.918133451237-6351049138HDQDX201223474148 228-1715746Met-7 to Ser-12,AR089: 30, AR061: 4Ser-20 to Arg-30,H0521: 3, H0051: 2,Asp-85 to Ala-92,L0756: 2, H0590: 1,Met-119 to Asn-146,S0250: 1, L0772: 1,Pro-151 to Asp-161,H0522: 1, S0406: 1 andGln-253 to Glu-260,L0748: 1.Ile-333 to Val-342,Leu-396 to Ala-406.919027452 210-10371050Met-7 to Ser-12,Ser-20 to Arg-30,Asp-85 to Ala-92,Met-119 to Asn-146,Pro-151 to Asp-161.139HLTHP861110457149 3-1352747His-35 to Glu-44,AR089: 1, AR061: 1Lys-88 to Tyr-94,L0439: 3, L0438: 2,Asp-140 to Ser-152,S0028: 2, H0656: 1,Leu-166 to Lys-171,H0645: 1, H0369: 1,Glu-183 to Glu-197,S0222: 1, S0346: 1,Glu-210 to Leu-217,H0328: 1, H0029: 1,Pro-231 to Gln-236.H0644: 1, H0169: 1,H0591: 1, H0646: 1,H0520: 1, H0539: 1,L0746: 1 and L0366: 1.919354453 3-13101051140HMSOL5211827151501146-832 748Ser-1 to Gly-12,AR061: 5, AR089: 2Arg-30 to Pro-36,L0770: 4, L0803: 4,Thr-65 to Met-76,H0638: 1, H0123: 1,Pro-86 to Asp-95.S0426: 1, L0662: 1,H0648: 1, L0747: 1,L0756: 1, L0779: 1,L0752: 1 and L0759: 1.921126454 90-4731052Glu-29 to Gly-35,Arg-53 to Pro-59,Thr-88 to Met-99,Pro-109 to Asp-118.141HAHGD331219819151 3-1745749Gly-59 to Ser-68,AR061: 7, AR089: 5Ala-87 to Glu-98,H0039: 5, H0622: 5,Pro-106 to Asn-121,L0748: 4, H0667: 4,Ser-148 to Lys-159,H0255: 3, S0126: 3,Phe-207 to Ala-222,H0393: 2, S0278: 2,Cys-279 to Asn-285,H0599: 2, H0618: 2,Gly-322 to Pro-330,H0318: 2, H0123: 2,Glu-357 to Ala-364,H0050: 2, H0179: 2,Ile-402 to Asp-407,H0271: 2, S0036: 2,Pro-456 to Val-466,H0135: 2, H0634: 2,Ser-474 to Thr-487.H0087: 2, H0100: 2,H0633: 2, S0210: 2,S0002: 2, H0144: 2,L0438: 2, L0602: 2,L0744: 2, L0731: 2,L0595: 2, L0601: 2,H0665: 2, H0542: 2,H0556: 1, H0222: 1,H0294: 1, H0583: 1,H0650: 1, H0657: 1,H0484: 1, H0306: 1,S0418: 1, S0420: 1,S0354: 1, H0580: 1,S0007: 1, S0046: 1,H0619: 1, H0550: 1,H0392: 1, H0586: 1,H0333: 1, H0486: 1,H0122: 1, H0196: 1,H0597: 1, H0544: 1,H0009: 1, H0172: 1,L0471: 1, H0023: 1,H0071: 1, H0266: 1,H0290: 1, H0553: 1,H0628: 1, H0551: 1,H0056: 1, H0623: 1,S0038: 1, H0494: 1,H0625: 1, H0561: 1,H0386: 1, H0509: 1,H0131: 1, H0130: 1,H0646: 1, S0144: 1,S0426: 1, H0529: 1,L0565: 1, H0547: 1,H0689: 1, H0435: 1,H0670: 1, S0330: 1,H0521: 1, S0027: 1,S0028: 1, S0032: 1,L0439: 1, L0747: 1,L0759: 1, S0260: 1,H0445: 1, L0597: 1,L0604: 1, L0593: 1,L0366: 1, H0668: 1,S0242: 1 and H0422: 1.921782455 1-10201053Phe-22 to Ala-37,Cys-94 to Asn-100,Gly-137 to Pro-145,Glu-172 to Ala-179,Ile-217 to Asp-222.142HHEHC53921783152 3-908750Gly-59 to Ser-68,AR089: 3, AR061: 219pAla-87 to Glu-98,L0748: 8, H0039: 5,Pro-106 to Asn-121,H0622: 5, L0664: 5,Ser-148 to Lys-159,L0439: 5, L0779: 5,Phe-207 to Ala-222,L0731: 5, L0758: 5,Ile-284 to Lys-289.L0665: 4, L0744: 4,L0601: 4, H0667: 4,H0255: 3, H0618: 3,L0666: 3, L0438: 3,S0126: 3, L0602: 3,L0742: 3, L0604: 3,L0595: 3, H0542: 3,H0265: 2, S0358: 2,H0393: 2, S0278: 2,H0550: 2, H0333: 2,H0599: 2, H0318: 2,H0545: 2, H0123: 2,H0050: 2, H0620: 2,H0179: 2, H0271: 2,S0036: 2, H0135: 2,H0634: 2, H0087: 2,H0100: 2, H0633: 2,S0210: 2, S0002: 2,L0769: 2, L0646: 2,L0768: 2, L0774: 2,H0144: 2, L0565: 2,H0689: 2, S0027: 2,L0747: 2, L0755: 2,L0593: 2, H0665: 2,H0556: 1, T0002: 1,H0222: 1, H0685: 1,H0294: 1, S0430: 1,H0583: 1, H0650: 1,H0657: 1, S0212: 1,S0282: 1, H0484: 1,H0306: 1, S0418: 1,S0420: 1, S0354: 1,S0360: 1, H0580: 1,S0007: 1, S0046: 1,H0619: 1, H0351: 1,H0549: 1, H0392: 1,H0586: 1, H0486: 1,T0060: 1, L0022: 1,H0122: 1, H0196: 1,H0597: 1, H0544: 1,H0009: 1, H0172: 1,L0471: 1, H0023: 1,H0071: 1, H0266: 1,H0290: 1, S0022: 1,H0030: 1, H0553: 1,H0628: 1, H0182: 1,H0617: 1, H0606: 1,H0551: 1, H0413: 1,H0056: 1, H0623: 1,S0038: 1, H0494: 1,H0625: 1, H0561: 1,H0386: 1, H0509: 1,H0131: 1, H0130: 1,H0646: 1, S0144: 1,S0344: 1, S0426: 1,H0529: 1, L0763: 1,L0770: 1, L0637: 1,L0372: 1, L0662: 1,L0775: 1, L0776: 1,L0659: 1, L0383: 1,L0790: 1, H0547: 1,H0435: 1, H0658: 1,H0670: 1, S0330: 1,H0521: 1, H0436: 1,S0390: 1, S0028: 1,S0032: 1, L0750: 1,L0753: 1, L0757: 1,L0759: 1, S0260: 1,H0445: 1, H0595: 1,L0597: 1, L0366: 1,H0668: 1, S0242: 1,H0423: 1, H0422: 1 andH0352: 1.143HE2PB011206665153 695-1228751Arg-1 to Asn-12,AR061: 9, AR089: 6Leu-87 to Gln-92,L0754: 6, H0318: 3,Phe-111 to Ser-117,H0486: 2, H0013: 2,Arg-161 to Gly-168.H0014: 2, L0439: 2,L0777: 2, H0543: 2,H0171: 1, H0556: 1,S6024: 1, H0583: 1,H0650: 1, H0638: 1,S0354: 1, H0580: 1,H0619: 1, H0455: 1,H0009: 1, S0003: 1,L0483: 1, S0036: 1,H0591: 1, H0494: 1,S0014: 1, S0450: 1,L0520: 1, L0763: 1,L0769: 1, L0641: 1,L0521: 1, L0662: 1,L0803: 1, L0774: 1,L0776: 1, L0783: 1,L0663: 1, H0144: 1,H0520: 1, H0547: 1,H0519: 1, S0136: 1,H0521: 1, H0522: 1,H0478: 1, L0742: 1,L0780: 1, L0592: 1,S0011: 1, S0192: 1,S0424: 1 and H0506: 1.921850456 2-6911054Glu-1 to Gly-7,Gln-43 to Arg-50,Asp-60 to Gly-67,Phe-150 to Glu-156,Arg-176 to Lys-181.144HOUDP521219522154 3-1361752Lys-54 to Arg-60.AR061: 2, AR089: 1L0794: 7, L0743: 2,H0543: 2, S0040: 1,S0134: 1, S0356: 1,T0082: 1, H0251: 1,H0494: 1, H0625: 1,H0649: 1, L0806: 1,L0657: 1, L0565: 1,L0758: 1, L0608: 1 andS0026: 1.922102457 196-11041055145HHGAE471127881155 2-517753Gly-1 to Arg-19,AR061: 3, AR089: 2Asp-27 to Glu-34,L0769: 5, L0774: 5,Asp-40 to Lys-46,L0756: 4, H0624: 2,Pro-63 to Arg-70,S0358: 2, S0444: 2,Lys-97 to Lys-103,S0408: 2, H0587: 2,Asp-113 to Gly-118,L0764: 2, L0766: 2,Ala-148 to Tyr-158.L0775: 2, L0601: 2,H0170: 1, S0442: 1,S0410: 1, H0497: 1,H0333: 1, H0632: 1,H0156: 1, L0022: 1,L0738: 1, H0271: 1,H0039: 1, S0344: 1,L0637: 1, L0772: 1,L0646: 1, L0773: 1,L0662: 1, L0518: 1,L0783: 1, L0791: 1,L0663: 1, S0374: 1,H0593: 1, H0660: 1,H0648: 1, H0672: 1,H0696: 1, L0749: 1,L0750: 1, L0779: 1,L0752: 1, L0755: 1,L0599: 1 and H0667: 1.922194458 3-5031056Gly-25 to Arg-45,Asp-53 to Glu-60,Asp-66 to Lys-72,Arg-89 to Trp-106,Asn-121 to Gly-147,Val-152 to Gly-159,Ala-161 to Ser-166.146HMCGL451165349156 89-922754Glu-8 to Pro-16,AR061: 8, AR089: 5Gln-21 to Glu-26,L0769: 5, L0774: 5,Gly-105 to Arg-125,L0756: 4, H0624: 2,Asp-133 to Glu-140,S0358: 2, S0444: 2,Asp-146 to Lys-152,S0408: 2, H0587: 2,Pro-169 to Arg-176,L0764: 2, L0766: 2,Lys-203 to Lys-209,L0775: 2, L0601: 2,Asp-219 to Gly-224,H0170: 1, S0442: 1,Ala-254 to Tyr-264.S0410: 1, H0497: 1,H0333: 1, H0632: 1,H0156: 1, L0022: 1,L0738: 1, H0271: 1,H0039: 1, S0344: 1,L0637: 1, L0772: 1,L0646: 1, L0773: 1,L0662: 1, L0518: 1,L0783: 1, L0791: 1,L0663: 1, S0374: 1,H0593: 1, H0660: 1,H0648: 1, H0672: 1,H0696: 1, L0749: 1,L0750: 1, L0779: 1,L0752: 1, L0755: 1,L0599: 1 and H0667: 1.922195459442-8851057Gln-1 to Glu-10,Asp-16 to Lys-22,Pro-39 to Arg-46,Lys-73 to Lys-79,Asp-89 to Gly-94,Ala-124 to Tyr-134.147HELEF1111538841571310-501 755Ser-29 to Val-36,AR061: 1, AR089: 1Leu-217 to Ser-222,S0045: 1 and H0457: 1.Lys-255 to Ile-262.926930460 53-6251058Phe-21 to Lys-27.148HETJX041212235158 3-704756Asp-11 to Val-21,AR089: 4, AR061: 2Pro-27 to Thr-43,H0046: 1, H0032: 1,Trp-92 to Lys-97,H0040: 1 and L0565: 1.Pro-136 to Gly-149,Met-182 to Val-193,Thr-197 to Asn-203.927120461 3-7041059Asp-11 to Val-21,Pro-27 to Thr-43,Trp-92 to Lys-97,Pro-136 to Gly-149,Met-182 to Val-193,Thr-197 to Asn-203.149HSOBC041165357159 3-395757Asn-61 to Glu-70,AR061: 5, AR089: 2Ser-80 to Arg-85,L0747: 14, H0551: 9,Pro-88 to Ile-96,H0617: 7, S0022: 6,Gln-101 to Gly-109,H0135: 6, S3014: 6,Lys-117 to His-128.L0750: 6, L0757: 6,L0759: 6, H0545: 5,S0126: 5, H0124: 4,H0529: 4, L0769: 4,L0764: 4, L0665: 4,H0547: 4, S3012: 4,L0740: 4, L0731: 4,H0624: 3, H0294: 3,L0717: 3, H0251: 3,H0024: 3, S0250: 3,H0100: 3, L0375: 3,L0651: 3, L0758: 3,H0170: 2, S0040: 2,H0583: 2, H0550: 2,H0333: 2, H0041: 2,H0012: 2, H0620: 2,H0284: 2, H0553: 2,H0606: 2, H0130: 2,L0641: 2, L0662: 2,L0650: 2, L0783: 2,L0666: 2, L0664: 2,L0744: 2, L0439: 2,L0751: 2, L0755: 2,H0667: 2, S0418: 1,S0354: 1, S0222: 1,H0441: 1, H0370: 1,T0109: 1, H0156: 1,H0318: 1, H0544: 1,H0546: 1, H0123: 1,L0471: 1, T0003: 1,H0014: 1, T0010: 1,H0266: 1, S0312: 1,H0428: 1, H0048: 1,H0413: 1, H0059: 1,H0560: 1, S0150: 1,H0633: 1, S0210: 1,L0770: 1, L0639: 1,L0772: 1, L0773: 1,L0768: 1, L0649: 1,L0775: 1, L0653: 1,L0776: 1, L0657: 1,L0656: 1, L0659: 1,L0526: 1, L0384: 1,L0809: 1, T0068: 1,H0593: 1, H0689: 1,H0435: 1, H0660: 1,S0330: 1, H0539: 1,S0152: 1, L0743: 1,L0777: 1, H0595: 1,L0591: 1, L0601: 1,S0192: 1, S0242: 1,S0194: 1, S0196: 1 andH0352: 1.927280462 2-3881060Asn-59 to Glu-67.150HE8PW831069980160 1-546758Glu-69 to Gln-76.AR089: 0, AR061: 0L0748: 6, L0749: 6,L0803: 3, L0774: 3,L0775: 3, H0574: 1,H0632: 1, H0013: 1,L0789: 1, L0790: 1,H0144: 1 and L0581: 1.927532463 1-5461061Glu-69 to Gln-76.151HWLEA48927676161100-408759Pro-1 toThr-8.AR089: 1, AR061: 0S0354: 1 and L0596: 1.152HNHNP811129143162143-514760Ile-1 to Ser-16.AR051: 23, AR054:11, AR050: 9, AR061: 8, AR089: 5S0216: 1928378464143-5141062Ile-1 to Ser-16.153HFIDL6811236411631161-535 761Pro-18 to Pro-27,AR089: 7, AR061: 4,Glu-67 to Lys-73,AR050: 2, AR054: 2,Phe-147 to Tyr-165,AR051: 1Thr-203 to Ser-209.S0192: 1928475465 2-5291063Glu-40 to Lys-46,Phe-120 to Ser-132.154HUJCT051165261164 2-1618762Gly-1 to Glu-7,AR089: 8, AR061: 4Lys-16 to Leu-21,H0653: 2, H0650: 1,Ser-26 to Val-31,H0050: 1, L0370: 1,Asp-64 to Thr-70,L0800: 1, L0662: 1,Asp-131 to Asn-136,L0653: 1, H0436: 1 andLys-191 to Asp-197,L0749: 1.Ala-259 to Glu-264,Glu-273 to Gly-279,Gln-296 to Ala-305,Asn-317 to Ser-322,Asn-345 to Ser-352,Gln-384 to Asn-392,Asn-407 to Gly-412,Gly-434 to Pro-441,Lys-476 to Asp-481,Gln-497 to Asn-507,His-523 to Asn-528.929264466 2-5201064Lys-10 to Leu-15,Ser-20 to Val-25,Asp-58 to Thr-64,Asp-125 to Asn-130.155HTEGO05932583165 3-884763Pro-12 to Tyr-21.AR089: 1, AR061: 0H0038: 2, L0745: 2and H0616: 1.156HRDBH581226719166 183-2591764Asp-35 to Leu-41,AR089: 1, AR061: 0Val-45 to Ser-57,H0620: 3, L0794: 3,Glu-134 to Asp-139,S0212: 2, H0254: 2,Pro-253 to Leu-259,H0545: 2, H0266: 2,Ser-301 to Gly-306,L0639: 2, L0759: 2,Leu-324 to Arg-330,H0556: 1, H0657: 1,Val-374 to Tyr-381,S0418: 1, H0580: 1,Gly-422 to Gly-427,S0045: 1, H0619: 1,Gly-466 to Gly-481,H0550: 1, H0600: 1,Lys-500 to Asp-505,H0590: 1, H0253: 1,Pro-540 to Asn-554,H0581: 1, H0052: 1,Arg-610 to Ala-616,H0309: 1, H0085: 1,Pro-773 to Ala-780.H0083: 1, H0628: 1,H0617: 1, H0124: 1,H0059: 1, H0494: 1,S0144: 1, S0142: 1,S0426: 1, H0529: 1,L0796: 1, L0659: 1,L0790: 1, H0519: 1,H0711: 1, S0328: 1,H0521: 1, H0522: 1,S3014: 1, L0758: 1,S0260: 1, H0343: 1,S0434: 1, L0601: 1,H0668: 1 and H0542: 1.933364467 455-22391065157HSDGW22934467167124-324765Leu-49 toTyr-54.AR061: 0, AR089: 0L0794: 4, L0438: 4,L0761: 3, L0766: 3,L0748: 3, L0439: 3,H0556: 2, L0602: 2,L0754: 2, L0779: 2,H0580: 1, H0208: 1,H0013: 1, T0082: 1,S0010: 1, H0428: 1,H0553: 1, H0038: 1,H0616: 1, H0494: 1,L0796: 1, L0800: 1,L0773: 1, L0533: 1,L0803: 1, L0776: 1,L0657: 1, L0791: 1,H0520: 1, H0519: 1,H0521: 1, H0187: 1,L0731: 1, S0031: 1 andL0366: 1.158HNTMD791126594168 1-663766Thr-1 to Gly-11,AR089: 2, AR061: 2Thr-26 to Gly-34.H0519: 2, S0420: 1,T0114: 1, H0013: 1,S0346: 1, H0038: 1,S0142: 1, H0520: 1,H0521: 1 and H0136: 1.934522468182-5861066159HCE5J511197900169 2-703767Arg-20 to Ala-25,AR061: 4, AR089: 2Asp-56 to Val-62,H0052: 7, L0809: 4,Gln-88 to Ala-93,H0663: 3, L0439: 3,Thr-126 to Ala-132,L0752: 3, H0587: 2,Gln-142 to Asn-160,L0565: 2, H0550: 1,Ser-189 to Asn-196.H0194: 1, H0562: 1,H0571: 1, L0435: 1,L0769: 1, L0787: 1 andL0755: 1.934524469227-7811067Pro-15 to Cys-22.160HHEFQ421151482170 3-947768Val-57 to Tyr-65,AR089: 8, AR061: 3Asp-73 to Lys-81,L0803: 6, L0759: 6,Arg-118 to Arg-123,L0740: 4, S0410: 3,Asp-140 to Leu-147,L0764: 3, L0766: 3,Pro-151 to Thr-156,L0804: 3, H0144: 3,Ala-163 to Glu-168,S0406: 3, L0731: 3,Pro-177 to Thr-187,L0362: 3, S0358: 2,Asp-220 to Thr-229,S0444: 2, H0596: 2,Thr-283 to Thr-289.H0644: 2, H0124: 2,L0770: 2, L0663: 2,H0539: 2, L0747: 2,L0750: 2, L0779: 2,L0757: 2, L0758: 2,H0624: 1, H0171: 1,H0639: 1, L0717: 1,H0411: 1, S0222: 1,H0441: 1, H0431: 1,H0574: 1, H0013: 1,H0156: 1, H0085: 1,L0471: 1, T0023: 1,H0163: 1, H0130: 1,L0762: 1, L0763: 1,L0662: 1, L0794: 1,L0775: 1, L0375: 1,L0805: 1, L0659: 1,L0783: 1, L0666: 1,S0374: 1, H0520: 1,H0658: 1, H0672: 1,S0330: 1, L0743: 1,L0751: 1, L0777: 1 andH0542: 1.934527470 29-10721068161HLQDC551082368171 2-499769Gly-19 to Ile-27,AR061: 9, AR089: 3Thr-31 to Asp-41,H0574: 1 and S0344: 1.Asp-58 to Phe-67,Ser-79 to Lys-85,Leu-119 to Glu-127.934528471 1-4441069Gly-19 to Ile-27,Thr-31 to Asp-41,Asp-58 to Phe-67,Ser-79 to Lys-85,Leu-119 to Glu-127.162HFPHI621195825172 1-1119770Ala-5 to Gly-13,AR061: 226, AR089:Pro-31 to Gln-37,79Ala-46 to Ala-69,L0439: 8, H0052: 7,Tyr-81 to Ser-87,L0741: 7, L0756: 4,Ser-120 to Ile-137,S0010: 3, H0261: 2,Thr-148 to Thr-156,H0156: 2, S0049: 2,Lys-181 to Phe-191,L0770: 2, L0776: 2,Pro-275 to Asn-297,L0742: 2, L0745: 2,Asp-322 to Gly-331,L0366: 2, S0222: 1,Lys-352 to Glu-359,H0438: 1, H0390: 1,Gln-365 to Ser-372.S0346: 1, H0009: 1,L0455: 1, S0038: 1,L0789: 1 and L0758: 1.934529472 3-4101070Met-1 to Gln-6,Pro-38 to Asn-60.163HE8QH091152238173 1-597771Ala-46 to Ser-53,AR089: 3, AR061: 3Pro-63 to Leu-78,H0261: 1, H0013: 1,Asp-106 to Asp-114,H0052: 1, H0009: 1,Glu-129 to Leu-136,H0144: 1 and L0438: 1.Gly-144 to Asp-149.934532473 2-6251071Gly-4 to Thr-9.164HFAAX291128791174 1-585772Ala-108 to His-113,AR089: 12, AR061: 4Asp-149 to Asn-154,H0242: 2, S0040: 1,Cys-179 to Val-186.S6024: 1, S6014: 1,H0586: 1, H0013: 1,H0124: 1, L0756: 1,L0592: 1, L0366: 1 andH0542: 1.934540474 2-5651072165HHFOC791182276175157-681773Ala-1 to Trp-9,AR051: 14, AR089: 6,Pro-12 to Gln-17,AR061: 4, AR050: 2,Arg-37 to Pro-42,AR054: 2Thr-44 to Lys-51,L0744: 9, L0747: 8,Pro-66 to Pro-80,S3014: 7, L0740: 7,Thr-97 to Ala-106,S0192: 6, S0027: 5,Pro-120 to Trp-128,S0212: 4, H0124: 4,Leu-131 to Gly-137.L0731: 4, L0662: 3,L0743: 3, L0752: 3,L0759: 3, H0662: 2,S0418: 2, S0046: 2,H0575: 2, H0545: 2,H0041: 2, H0413: 2,L0775: 2, H0696: 2,S0037: 2, L0748: 2,L0751: 2, L0754: 2,L0749: 2, L0758: 2,H0445: 2, S0276: 2,H0624: 1, L0778: 1,L0005: 1, H0645: 1,H0441: 1, H0391: 1,S0005: 1, T0040: 1,H0069: 1, H0427: 1,S0280: 1, H0042: 1,T0048: 1, H0505: 1,H0309: 1, H0544: 1,H0009: 1, H0266: 1,H0617: 1, H0412: 1,H0623: 1, T0004: 1,L0564: 1, T0041: 1,H0494: 1, H0633: 1,H0646: 1, H0652: 1,L0769: 1, L0646: 1,L0655: 1, L0659: 1,L0546: 1, L0783: 1,L0809: 1, H0144: 1,L0565: 1, S0126: 1,H0689: 1, H0435: 1,H0659: 1, H0672: 1,S0378: 1, H0555: 1,S0206: 1, L0777: 1,L0780: 1, S0434: 1,S0011: 1, S0194: 1 andH0506: 1.935406475 3-4431073Glu-6 to Glu-15,Thr-21 to Asp-28,Ser-42 to Lys-47.166HOGEQ431223485176 1-2970774Gly-1 to Trp-10,AR089: 1, AR061: 0Glu-12 to Thr-48,H0457: 8, L0766: 7,Phe-94 to Gly-99,L0599: 6, H0677: 6,Tyr-105 to Ser-119,L0438: 5, L0779: 5,Thr-136 to Ala-147,H0012: 3, L0809: 3,Ser-165 to Ala-171,H0656: 2, H0620: 2,Glu-222 to Trp-227,L0771: 2, H0435: 2,His-233 to Trp-238,H0436: 2, L0748: 2,Glu-251 to Glu-258,L0439: 2, L0751: 2,Asn-265 to Ser-270,L0749: 2, S0134: 1,Gln-305 to Trp-310,H0645: 1, H0587: 1,Asn-325 to Phe-335,H0635: 1, H0581: 1,Ser-341 to Glu-349,H0546: 1, H0477: 1,Lys-378 to Trp-383,H0560: 1, H0641: 1,Ala-390 to Gly-398,S0422: 1, H0529: 1,Asn-451 to Trp-458,L0521: 1, L0662: 1,Met-476 to Trp-486,L0794: 1, L0774: 1,Gln-497 to Gly-505,L0775: 1, L0606: 1,Arg-567 to Glu-573,L0659: 1, L0647: 1,Gln-610 to Phe-618,L0789: 1, L0791: 1,Glu-687 to Arg-697,L0792: 1, L0666: 1,Gly-732 to Glu-742,L0663: 1, L0665: 1,Thr-797 to Asp-802,H0702: 1, H0547: 1,Arg-818 to Thr-827,H0576: 1, S0028: 1,Ile-840 to Lys-849,L0756: 1, L0777: 1,Ala-853 to Gly-860,L0755: 1, L0758: 1,Pro-879 to Tyr-886,H0543: 1 and H0506: 1.Ser-893 to Ile-901,Thr-904 to Phe-911,Asp-931 to Pro-937,Arg-952 to Thr-962.935465476 1-1501074Glu-1 to Thr-13.167HCECQ23938398177810-289775Pro-26 to Tyr-31.AR089: 1, AR061: 0L0769: 3, H0052: 2,L0439: 2, H0572: 1,H0015: 1, L0438: 1 andL0741: 1.168HTGAU791178621178 52-966776His-12 to Arg-20,AR061: 7, AR089: 4Pro-26 to Asp-43,H0551: 3, H0529: 3,Ala-62 to Glu-70,L0769: 3, L0758: 3,Arg-78 to Arg-83,S0418: 2, L0770: 2,Phe-100 to Gln-105,L0773: 2, L0521: 2,Gly-129 to Glu-136,H0701: 2, S0126: 2,Met-182 to Ile-187,L0747: 2, L0731: 2,Tyr-277 to Ala-284.L0759: 2, L0589: 2,L0601: 2, H0624: 1,H0149: 1, H0556: 1,H0295: 1, S0134: 1,H0583: 1, H0661: 1,H0592: 1, H0013: 1,H0635: 1, H0581: 1,S0250: 1, H0212: 1,H0412: 1, S0144: 1,L0763: 1, L0645: 1,L0764: 1, L0794: 1,L0766: 1, L0775: 1,L0783: 1, L0665: 1,H0519: 1, H0435: 1,H0672: 1, H0436: 1,S3014: 1, S0028: 1,L0750: 1, L0777: 1,L0366: 1, H0667: 1 andH0423: 1.940369477 63-9771075His-12 to Arg-20,Pro-26 to Asp-43,Ala-62 to Glu-70,Arg-78 to Arg-83,Phe-100 to Gln-105,Gly-129 to Glu-136.169HE9FI331156432179 2-286777Glu-65 to Pro-70.AR061: 6, AR089: 2L0749: 2, H0144: 1and L0748: 1.941348478 3-4341076170HNHCP79565781180 23-301778Gly-16 to Asn-21.AR051: 9, AR054: 9,AR050: 7, AR061: 3,AR089: 2H0271: 26, H0521: 26,H0046: 20, L0747: 20,S0278: 14, S0052: 14,L0754: 12, L0599: 12,S0142: 11, S0428: 11,H0179: 10, S0344: 10,L0776: 9, H0638: 8,L0771: 8, L0666: 8,S0360: 7, S0144: 7,L0775: 7, L0659: 7,H0422: 7, S0354: 6,H0580: 6, H0622: 6,H0641: 6, H0522: 6,L0740: 6, L0595: 6,H0581: 5, H0416: 5,H0673: 5, L0598: 5,L0774: 5, S3014: 5,L0777: 5, L0759: 5,L0362: 5, H0423: 5,H0069: 4, H0674: 4,L0770: 4, L0769: 4,L0750: 4, L0752: 4,L0731: 4, L0757: 4,L0603: 4, S0114: 3,S0134: 3, S0116: 3,H0341: 3, S0418: 3,S0358: 3, H0545: 3,H0050: 3, H0646: 3,L0768: 3, L0664: 3,S0053: 3, S0216: 3,S0374: 3, S0404: 3,S0206: 3, L0745: 3,L0756: 3, L0581: 3,H0170: 2, H0222: 2,L0785: 2, H0663: 2,S0376: 2, S0132: 2,S0222: 2, H0370: 2,H0486: 2, H0013: 2,H0635: 2, S0280: 2,H0575: 2, H0036: 2,H0618: 2, H0597: 2,H0014: 2, H0039: 2,L0142: 2, H0551: 2,H0056: 2, H0561: 2,S0426: 2, L0763: 2,L0761: 2, L0648: 2,L0662: 2, L0767: 2,L0655: 2, L0519: 2,L0665: 2, H0519: 2,H0435: 2, H0696: 2,S0027: 2, L0743: 2,L0751: 2, S0031: 2,S0260: 2, H0445: 2,S0434: 2, L0590: 2,S0276: 2, H0395: 1,H0556: 1, T0002: 1,H0685: 1, S0040: 1,H0294: 1, S0218: 1,S0001: 1, H0484: 1,H0483: 1, H0662: 1,H0176: 1, H0589: 1,H0459: 1, S0356: 1,S0408: 1, S0410: 1,L0717: 1, H0411: 1,H0549: 1, H0550: 1,H0431: 1, H0608: 1,H0409: 1, H0404: 1,H0587: 1, H0485: 1,H0250: 1, L0021: 1,H0590: 1, H0318: 1,T0071: 1, H0421: 1,H0263: 1, H0596: 1,H0150: 1, H0009: 1,L0471: 1, H0011: 1,S0051: 1, H0083: 1,H0510: 1, H0594: 1,S0318: 1, H0687: 1,H0286: 1, S0250: 1,H0328: 1, H0553: 1,L0055: 1, H0032: 1,H0169: 1, H0316: 1,H0135: 1, H0090: 1,H0591: 1, H0634: 1,H0413: 1, H0623: 1,H0059: 1, T0069: 1,S0038: 1, H0100: 1,T0041: 1, H0509: 1,S0150: 1, H0633: 1,S0002: 1, H0529: 1,L0762: 1, L0667: 1,L0772: 1, L0646: 1,L0643: 1, L0521: 1,L0766: 1, L0389: 1,L0653: 1, L0629: 1,L0527: 1, L0657: 1,L0517: 1, L0384: 1,L0809: 1, L0663: 1,H0144: 1, H0697: 1,S0126: 1, H0690: 1,H0670: 1, H0648: 1,S0378: 1, S0380: 1,H0518: 1, S0152: 1,S0013: 1, S0044: 1,H0214: 1, H0555: 1,H0436: 1, H0478: 1,S0432: 1, S3012: 1,S0032: 1, L0744: 1,L0439: 1, L0779: 1,L0758: 1, S0308: 1,S0436: 1, L0591: 1,L0593: 1, S0011: 1,H0543: 1 and S0458: 1.775293479138-2751077941862480 2-7481078171HTLIY521194806181 164-1351779Pro-3 to Gly-8,AR061: 0, AR089: 0Val-21 to Gly-30,H0618: 64, H0253: 52,Gly-68 to Ala-85,L0758: 6, L0779: 2,His-94 to Gly-99,H0392: 1, H0038: 1,Ala-105 to Arg-110,L0761: 1, L0803: 1,Ala-114 to Gln-138,L0806: 1 and L0697: 1.Arg-143 to Glu-155,Leu-202 to Arg-222,Arg-287 to Ser-292,Pro-325 to Arg-332,Arg-337 to Gly-351,Val-388 to Lys-396.942161481 1-13681079172HRAED74942527182289-651780His-9 to Ile-15.AR061: 1, AR089: 1S0222: 3, H0052: 3,L0361: 3, H0179: 2,L0769: 2, H0521: 2,H0555: 2, L0779: 2,L0758: 2, H0663: 1,H0549: 1, S0220: 1,H0586: 1, H0156: 1,S0010: 1, H0596: 1,S0051: 1, T0010: 1,H0271: 1, L0143: 1,H0617: 1, H0652: 1,L0764: 1, L0794: 1,L0806: 1, L0809: 1,H0518: 1, H0478: 1,L0751: 1, L0747: 1,L0750: 1, L0780: 1,L0731: 1 and L0366: 1.173HFKKN77943757183145-684781Thr-9 to Val-16.AR061: 6, AR089: 2H0620: 2, H0024: 2,H0208: 1, S0222: 1,H0194: 1, H0123: 1,H0051: 1 and S0052: 1.174HTEMU661205381184462-962782His-3 to Ser-14,AR061: 7, AR089: 5Thr-20 to Ser-27,H0616: 1Pro-41 to Asn-50,Glu-101 to Asp-109,Leu-149 to Ser-154.944419482454-9631080Ala-1 to Gln-7,Lys-24 to Ser-30,Pro-44 to Asn-53,Glu-104 to Asp-112,Leu-152 to Ser-157.175HWAGU621206797185 210-1511783Thr-1 to Leu-9,AR061: 5, AR089: 5Pro-34 to Lys-40,S0356: 9, L0803: 3,Glu-82 to Gln-87,L0766: 2, L0743: 2,Ala-216 to His-233,L0731: 2, L0785: 1,Met-235 to His-243,S0116: 1, S0354: 1,Pro-322 to Lys-327,S0358: 1, S0278: 1,Arg-346 to Trp-351.H0642: 1, H0486: 1,H0581: 1, H0596: 1,H0355: 1, S0003: 1,L0455: 1, H0090: 1,H0591: 1, S0142: 1,S0344: 1, S0422: 1,S0426: 1, L0598: 1,L0794: 1, L0804: 1,L0659: 1, L0789: 1,L0664: 1, H0547: 1,H0660: 1, S0330: 1,L0754: 1, L0779: 1,L0758: 1, L0608: 1,S0026: 1 and H0543: 1.945368483 1-15001081Pro-100 to Lys-106,Glu-148 to Gln-153.176HFPFB391198036186 93-1643784Glu-62 to Tyr-67,AR061: 6, AR089: 3Pro-169 to Lys-179,S0010: 4, S0222: 3,Pro-189 to Ala-201,H0455: 2, L0803: 2,Ala-218 to Arg-223,L0439: 2, L0745: 2,Tyr-324 to Asn-331,S0282: 1, S0400: 1,Gly-352 to Val-357,H0456: 1, H0441: 1,Leu-365 to Lys-371,S0346: 1, H0509: 1,His-393 to Ala-399,L0769: 1, L0438: 1,Asp-420 to Asn-425,L0756: 1 and S0106: 1.Thr-460 to Lys-473,Ser-488 to Gly-502.9461704841613-462 1082Pro-36 to Lys-46,Pro-56 to Ala-68,Ala-85 to Arg-90,Tyr-191 to Asn-198,Gly-219 to Val-224,Leu-232 to Lys-238,His-260 to Ala-266.177HPMFI381165993187 3-410785Pro-128 to Ser-134.AR061: 3, AR089: 2L0754: 3, H0644: 2,L0803: 2, L0748: 2,H0620: 1, H0031: 1,L0774: 1 and L0789: 1.946252485992-4951083Leu-143 to Thr-149,Gln-152 to Glu-157.178HBXDJ07946830188125-652786Glu-62 to Lys-68,AR061: 2, AR089: 2Asn-105 to Gly-113.L0439: 11, L0794: 5,L0666: 5, S0222: 4,H0052: 3, L0756: 3,H0624: 2, S6028: 2,S0038: 2, L0638: 2,L0805: 2, L0664: 2,L0438: 2, L0740: 2,H0171: 1, S6024: 1,H0013: 1, H0374: 1,H0050: 1, S0050: 1,H0051: 1, S0386: 1,L0769: 1, L0768: 1,L0776: 1, L0659: 1,L0789: 1, H0144: 1,L0745: 1 and L0746: 1.179HOFMS431152417189 1-1029787Pro-1 to Asp-16,AR051: 15, AR050: 9,Pro-60 to Asn-65,AR089: 7, AR061: 5,Tyr-83 to Tyr-89,AR054: 1Ser-102 to Pro-115,H0415: 1Pro-130 to Glu-141,Ser-151 to Glu-160,Trp-177 to Glu-183,Phe-191 to Arg-198,Phe-203 to Tyr-209,Asn-234 to Ala-240,Pro-266 to Pro-271,Ser-276 to Thr-311,Arg-338 to Gly-343.947973486 3-3591084Asp-1 to Asp-17,Pro-61 to Asn-66,Tyr-84 to Tyr-90,Ser-103 to Trp-110.180HOVCO141091087190 3-563788Arg-78 to His-85,AR089: 3, AR061: 2Leu-99 to Lys-104,S6016: 1 and H0428: 1.Lys-123 to His-132,Thr-164 to Pro-171.947999487 3-5391085Arg-78 to His-85,Leu-99 to Lys-104,Lys-123 to His-132,Ser-157 to Pro-174.181HTEPE351105272191 1-762789Tyr-1 to Lys-8,AR061: 4, AR089: 1Phe-19 to Ser-24,L0758: 7, L0768: 2,Thr-28 to Ser-34,H0616: 1 and L0151: 1.Pro-54 to Trp-70,Leu-110 to Asn-118,Ser-145 to Asp-151,Pro-162 to Val-172,Pro-180 to Thr-185.948475488839-78 1086Tyr-1 to Lys-8,Phe-19 to Ser-24,Thr-28 to Ser-34,Pro-54 to Trp-70.182HE8UA521229490192 1-1728790Pro-22 to Gly-32,AR061: 6, AR089: 1Arg-52 to Gly-60,L0752: 5, H0013: 2,Ser-78 to Met-89,L0780: 2, H0624: 1,Ile-100 to Ser-106,H0170: 1, H0645: 1,Asp-130 to Leu-137,H0318: 1, L0750: 1,Tyr-146 to Ala-151.L0779: 1 and L0777: 1.948509489 1-6151087Pro-22 to Gly-32,Arg-52 to Gly-60,Ser-78 to Met-89,Ile-100 to Ser-106,Asp-130 to Leu-137,Tyr-146 to Ser-152.183HOUBE501090776193 1-1188791Ser-50 to Glu-62.AR061: 1, AR089: 0S0040: 1, S0222: 1,L0471: 1 and L0517: 1.948519490 1-2431088Ser-50 to Ser-66.184HAJAV281165229194 3-1073792Pro-1 to Ala-12,AR089: 3, AR061: 1Ile-264 to Val-277,L0747: 12, L0755: 12,Gln-304 to Gln-309,L0766: 9, L0438: 9,Ile-324 to Leu-330.L0754: 7, H0046: 6,L0751: 6, L0752: 6,H0068: 5, L0775: 5,L0439: 5, S0010: 4,H0547: 4, S0152: 4,L0740: 4, L0779: 4,L0759: 4, H0591: 3,L0771: 3, L0662: 3,L0774: 3, L0666: 3,S0028: 3, L0748: 3,L0756: 3, L0731: 3,L0757: 3, H0624: 2,S0045: 2, H0619: 2,S0222: 2, S0049: 2,H0052: 2, H0615: 2,S0036: 2, T0041: 2,H0509: 2, S0002: 2,S0426: 2, L0769: 2,L0776: 2, L0659: 2,H0521: 2, H0707: 2,L0594: 2, L0362: 2,S0011: 2, H0170: 1,H0171: 1, H0685: 1,S0040: 1, T0049: 1,H0657: 1, S0001: 1,H0638: 1, S0358: 1,S0360: 1, S0408: 1,H0637: 1, S0007: 1,S0132: 1, S6022: 1,H0550: 1, H0431: 1,H0455: 1, H0574: 1,H0486: 1, T0114: 1,H0250: 1, H0069: 1,H0156: 1, L0105: 1,H0597: 1, H0546: 1,H0545: 1, H0050: 1,L0163: 1, H0594: 1,H0266: 1, H0290: 1,S0214: 1, H0328: 1,H0688: 1, H0622: 1,H0032: 1, H0673: 1,H0674: 1, S0364: 1,H0090: 1, H0040: 1,H0551: 1, T0067: 1,H0268: 1, H0100: 1,H0494: 1, H0560: 1,H0561: 1, H0633: 1,L0762: 1, L0763: 1,L0638: 1, L0772: 1,L0773: 1, L0521: 1,L0768: 1, L0794: 1,L0803: 1, L0809: 1,L0545: 1, L0664: 1,L0665: 1, H0144: 1,S0126: 1, H0660: 1,H0672: 1, S0378: 1,S0380: 1, S0350: 1,H0555: 1, H0436: 1,H0540: 1, S0390: 1,S0206: 1, S0032: 1,L0741: 1, L0749: 1,L0786: 1, L0777: 1,L0758: 1, S0026: 1 andH0506: 1.948630491 3-4641089Pro-1 to Ala-12.185HAQBZ891083554195 54-1034793Pro-43 to Asn-61,AR061: 9, AR089: 6Ala-77 to Arg-82,L0659: 4, L0758: 4,Glu-207 to His-212,L0777: 3, S0360: 2,Glu-252 to Glu-261,L0775: 2, L0750: 2,Asp-279 to Asn-284.L0731: 2, H0295: 1,S0218: 1, H0255: 1,H0402: 1, L0717: 1,H0411: 1, H0015: 1,H0030: 1, H0644: 1,H0673: 1, H0087: 1,L0770: 1, L0769: 1,L0637: 1, L0761: 1,L0646: 1, L0387: 1,L0766: 1, L0776: 1,L0655: 1, L0789: 1,H0683: 1, S0027: 1,L0748: 1, L0779: 1 andL0757: 1.949061492 2-3251090Ala-20 to Arg-25.186HELHF07949067196 65-340794Thr-16 to Pro-21.AR061: 5, AR089: 4S0045: 2187HE9QQ221127726197 41-436795AR061: 11, AR089: 3L0748: 3, H0144: 2,H0632: 1 and L0581: 1.949080493 3-5631091188HSDSB061128280198 3-863796Ile-25 to Asn-36,AR061: 4, AR089: 3Glu-54 to Val-63,H0590: 7, L0754: 5,Gly-81 to Glu-86,H0156: 3, L0731: 3,Gly-108 to Thr-114,L0600: 3, S0360: 2,Val-125 to Ser-131.H0339: 2, S0472: 2,L0803: 2, L0751: 2,L0779: 2, L0759: 2,S0031: 2, L0596: 2,S0212: 1, H0411: 1,S0222: 1, H0409: 1,H0601: 1, H0333: 1,H0632: 1, H0427: 1,L0021: 1, H0037: 1,H0596: 1, H0024: 1,H0239: 1, S6028: 1,H0266: 1, H0687: 1,H0328: 1, H0644: 1,H0674: 1, H0598: 1,T0067: 1, H0509: 1,L0763: 1, L0772: 1,L0764: 1, L0771: 1,L0773: 1, L0650: 1,L0806: 1, L0659: 1,L0547: 1, L0809: 1,L0666: 1, L0663: 1,L0665: 1, S0328: 1,S0380: 1, S0390: 1,S0032: 1, L0744: 1,L0745: 1, L0746: 1,L0747: 1, L0756: 1,L0777: 1, L0758: 1,L0588: 1, S0276: 1,S0196: 1, S0412: 1 andH0506: 1.949151494 3-8631092Ile-25 to Asn-36,Glu-54 to Val-63,Gly-81 to Glu-86,Gly-108 to Thr-114,Val-125 to Ser-131.189HACAD359491991991473-916 797Leu-9 to Leu-18,AR061: 2, AR089: 1Ala-49 to Gly-55,L0637: 2, L0783: 2,Gly-66 to Glu-74,L0777: 2, S6022: 1,Ala-95 to Gln-100.H0392: 1, H0586: 1,H0050: 1, L0809: 1,L0759: 1 and S0192: 1.190HEQAP17949358200819-295798AR051: 744, AR054:681, AR050: 564,AR061: 2, AR089: 1S0192: 3, H0544: 1,L0766: 1, L0804: 1,H0521: 1 and L0747: 1.191HMTBB171128589201 1-366799Arg-18 to Pro-23,AR089: 3, AR061: 3Pro-25 to Gly-37,L0438: 3, L0439: 3,Ile-48 to Ile-61,L0749: 3, L0758: 3,Asp-69 to Gly-74,L0766: 2, L0375: 2,Ser-105 to Asn-112.L0731: 2, L0759: 2,L0803: 1, L0655: 1,L0517: 1, L0666: 1,L0664: 1, H0518: 1,L0748: 1, L0779: 1,L0599: 1 and H0008: 1.950884495513-1001093Arg-34 to Pro-39,Pro-41 to Gly-53,Ile-64 to Ile-77,Asp-85 to Gly-90,Ser-121 to Asn-128.192HKGDE581129137202 2-541800Asp-1 to Cys-10,AR089: 2, AR061: 2Glu-31 to Pro-38,H0538: 1, L0803: 1Met-43 to Val-48,and L0731: 1.Asp-97 to Phe-110,Asp-119 to Gly-137.945039496 11-9371094Asp-17 to Cys-26,Glu-47 to Pro-54,Met-59 to Val-64,Asp-113 to Phe-126,Asp-135 to Gly-153.950885497469-1191095Ser-20 to Gly-32,Ile-43 to Ile-56,Asp-64 to Gly-69,Ser-100 to Asn-107.193HCHMW401144323203539-3 801Phe-1 to Trp-6,AR089: 9, AR061: 4Ser-41 to Arg-56,H0586: 14, H0587: 8,Pro-162 to Leu-174.L0763: 6, H0592: 4,H0484: 3, H0081: 3,H0063: 3, H0483: 2,H0664: 2, H0601: 1,H0600: 1, H0494: 1,L0648: 1, H0658: 1,S0328: 1 and L0747: 1.951518498 84-5721096Ser-7 to Gly-14,Leu-22 to Ala-28,Thr-57 to Ser-62.194HE8QZ341143411204 2-1306802Ser-85 to Arg-90,AR089: 4, AR061: 1His-99 to Met-105,H0046: 4, H0591: 2,Met-119 to Val-125,T0067: 2, L0766: 2,Lys-127 to Ile-133,H0144: 2, H0521: 2,Lys-215 to Tyr-221,L0744: 2, L0439: 2,Phe-239 to Lys-247,H0170: 1, H0013: 1,Asn-293 to Asp-298,H0599: 1, S0182: 1,Gln-404 to Tyr-411.H0051: 1, H0510: 1,S6028: 1, L0455: 1,H0616: 1, S0422: 1,S0374: 1, L0438: 1,S0390: 1, L0748: 1 andL0604: 1.952283499 3-10251097Ser-85 to Arg-90,His-99 to Met-105,Met-119 to Val-125,Lys-127 to Ile-133,Lys-215 to Tyr-221,Phe-239 to Lys-247,Asn-293 to Gly-298.195HWAFG041227627205 2-850803Gln-103 to Asp-113,AR089: 17, AR061: 8Ser-182 to Phe-200,L0789: 4, L0758: 4,Cys-211 to Ser-221,H0657: 3, H0052: 3,Gln-233 to Ala-238,L0438: 3, L0744: 3,Glu-256 to Ser-264.L0779: 3, L0005: 2,H0581: 2, H0194: 2,H0046: 2, H0038: 2,L0800: 2, L0659: 2,H0521: 2, L0743: 2,L0439: 2, H0556: 1,S0282: 1, S0358: 1,H0619: 1, H0586: 1,H0618: 1, H0231: 1,S0362: 1, H0622: 1,T0006: 1, H0616: 1,H0413: 1, H0623: 1,L0351: 1, S0150: 1,L0769: 1, L0372: 1,L0662: 1, L0794: 1,L0775: 1, L0651: 1,L0527: 1, L0657: 1,L0666: 1, H0547: 1,H0690: 1, H0658: 1,H0672: 1, H0539: 1,S0378: 1, H0555: 1,L0754: 1, L0747: 1,L0780: 1, L0596: 1,S0192: 1, H0542: 1 andH0423: 1.9528785001658-789 1098Gln-110 to Asp-120,Ser-189 to Phe-207,Cys-218 to Ser-228,Gln-240 to Ala-245,Glu-263 to Ser-271.196HTEKT331075836206 189-1268804Pro-19 to Gly-24,AR089: 15, AR061: 9Val-41 to Phe-47,L0766: 4, L0745: 3,Lys-75 to Asp-83,L0752: 3, S0360: 2,Ser-138 to Gln-154,L0748: 2, L0746: 2,Asp-230 to Ser-235,L0755: 2, H0624: 1,Asp-278 to Thr-283,S0114: 1, H0098: 1,Pro-315 to Ser-324,L0471: 1, H0083: 1,Trp-338 to Thr-344.H0428: 1, L0483: 1,H0090: 1, H0616: 1,H0494: 1, H0560: 1,H0509: 1, L0761: 1,L0772: 1, L0803: 1,L0776: 1, L0655: 1,L0792: 1, L0664: 1,S0374: 1, L0438: 1,H0520: 1, H0519: 1,H0435: 1, H0648: 1,S0152: 1, H0521: 1,H0478: 1, L0747: 1,L0756: 1, L0779: 1,L0758: 1, L0759: 1,H0667: 1, H0543: 1 andL0465: 1.953308501 200-14261099197HBXDM071206657207 1-1806805Leu-89 to Tyr-96,AR089: 9, AR061: 1Leu-195 to Glu-209,L0777: 7, L0747: 5,Val-253 to Ser-259,H0556: 4, L0794: 4,Ile-274 to Phe-279,L0592: 4, H0497: 3,Lys-317 to His-323,H0052: 3, L0803: 3,Lys-338 to Phe-345,H0087: 2, L0439: 2,Ile-361 to Ser-366,L0749: 2, L0593: 2,Glu-370 to Gln-375,H0459: 1, S0046: 1,Lys-398 to Lys-404,L0717: 1, H0575: 1,Val-482 to Arg-492,S0010: 1, H0390: 1,Met-539 to Glu-548,H0194: 1, H0050: 1,Gly-573 toTyr-578.H0375: 1, L0055: 1,S0036: 1, H0059: 1,S0038: 1, H0494: 1,S0002: 1, L0809: 1,L0789: 1, L0663: 1,L0665: 1, S0330: 1,H0521: 1, H0522: 1,H0696: 1, H0555: 1,S0028: 1, L0731: 1,L0759: 1, L0581: 1,H0542: 1 and H0422: 1.953622502 81-7161100198HFPFA83955614208187-735806Thr-9 to Val-16.AR054: 375, AR051:284, AR050: 235,AR061: 96, AR089: 33H0620: 2, H0024: 2,H0208: 1, S0222: 1,H0194: 1, H0123: 1,H0051: 1 and S0052: 1.199HKADO361189002209 3-815807Ala-6 to Pro-12,AR089: 2, AR061: 1Glu-22 to Ala-41,H0069: 2, H0497: 1,Ser-230 to Ala-238,H0494: 1, L0769: 1,Asp-257 to Ser-268.S0028: 1 and H0542: 1.956115503 2-2801101200HFXKG51956596210578-3 808His-13 to Asn-24,AR089: 2, AR061: 1Pro-147 to Asn-157,S0028: 2, S0282: 1 andGln-164 to Glu-169.S0051: 1.201HFPHR821187749211 3-1328809Ile-258 to Val-271,AR089: 3, AR061: 1Gln-298 to Gln-303,L0747: 12, L0755: 12,Ile-318 to Leu-324,L0766: 9, L0438: 9,Glu-353 to Leu-361,L0754: 7, H0046: 6,Ser-397 to Arg-408,L0751: 6, L0752: 6,Gly-427 to Leu-433.H0068: 5, L0775: 5,L0439: 5, S0010: 4,H0547: 4, S0152: 4,L0740: 4, L0779: 4,L0759: 4, H0591: 3,L0771: 3, L0662: 3,L0774: 3, L0666: 3,S0028: 3, L0748: 3,L0756: 3, L0731: 3,L0757: 3, H0624: 2,S0045: 2, H0619: 2,S0222: 2, S0049: 2,H0052: 2, H0615: 2,S0036: 2, T0041: 2,H0509: 2, S0002: 2,S0426: 2, L0769: 2,L0776: 2, L0659: 2,H0521: 2, H0707: 2,L0594: 2, L0362: 2,S0011: 2, H0170: 1,H0171: 1, H0685: 1,S0040: 1, T0049: 1,H0657: 1, S0001: 1,H0638: 1, S0358: 1,S0360: 1, S0408: 1,H0637: 1, S0007: 1,S0132: 1, S6022: 1,H0550: 1, H0431: 1,H0455: 1, H0574: 1,H0486: 1, T0114: 1,H0250: 1, H0069: 1,H0156: 1, L0105: 1,H0597: 1, H0546: 1,H0545: 1, H0050: 1,L0163: 1, H0594: 1,H0266: 1, H0290: 1,S0214: 1, H0328: 1,H0688: 1, H0622: 1,H0032: 1, H0673: 1,H0674: 1, S0364: 1,H0090: 1, H0040: 1,H0551: 1, T0067: 1,H0268: 1, H0100: 1,H0494: 1, H0560: 1,H0561: 1, H0633: 1,L0762: 1, L0763: 1,L0638: 1, L0772: 1,L0773: 1, L0521: 1,L0768: 1, L0794: 1,L0803: 1, L0809: 1,L0545: 1, L0664: 1,L0665: 1, H0144: 1,S0126: 1, H0660: 1,H0672: 1, S0378: 1,S0380: 1, S0350: 1,H0555: 1, H0436: 1,H0540: 1, S0390: 1,S0206: 1, S0032: 1,L0741: 1, L0749: 1,L0786: 1, L0777: 1,L0758: 1, S0026: 1 andH0506: 1.9575285041592-273 1102Ile-256 to Val-269,Gln-296 to Gln-301,Ile-316 to Leu-322,Glu-351 to Leu-359,Ser-395 to Arg-406,Gly-425 to Leu-431.202HISAF59959140212130-843810Gly-33 to Ser-48.AR089: 2, AR061: 2L0789: 4, L0758: 4,H0657: 3, H0052: 3,H0046: 3, L0438: 3,L0744: 3, L0779: 3,L0005: 2, H0586: 2,H0581: 2, H0194: 2,H0038: 2, L0800: 2,L0659: 2, H0521: 2,L0743: 2, L0439: 2,H0556: 1, S0282: 1,S0358: 1, H0619: 1,H0618: 1, H0231: 1,H0569: 1, S0362: 1,H0622: 1, T0006: 1,H0135: 1, H0616: 1,H0413: 1, H0623: 1,L0351: 1, S0150: 1,L0769: 1, L0372: 1,L0662: 1, L0794: 1,L0775: 1, L0651: 1,L0527: 1, L0657: 1,L0666: 1, H0144: 1,H0547: 1, H0690: 1,H0658: 1, H0672: 1,H0539: 1, S0378: 1,H0555: 1, L0754: 1,L0747: 1, L0780: 1,L0596: 1, S0192: 1,H0542: 1 and H0423: 1.203HCEHD661136122213 2-562811Ala-1 to Lys-6,AR061: 7, AR089: 3Leu-13 to Gln-25,L0769: 9, S0051: 4,Asp-70 to Gly-75,H0441: 3, S0036: 3,Gly-92 to Lys-97,L0809: 3, L0789: 3,Asp-106 to Tyr-112,L0438: 3, L0439: 3,Leu-135 to Val-146,L0731: 3, H0052: 2,Glu-165 to Asp-173.H0687: 2, H0181: 2,L0800: 2, L0794: 2,L0665: 2, L0741: 2,L0742: 2, L0756: 2,S0031: 2, H0171: 1,H0556: 1, S6024: 1,S0029: 1, H0411: 1,S0278: 1, H0455: 1,H0486: 1, L0109: 1,H0251: 1, L0163: 1,H0617: 1, H0413: 1,L0762: 1, L0638: 1,L0639: 1, L0761: 1,L0764: 1, L0662: 1,L0774: 1, L0807: 1,L0657: 1, S0053: 1,S0126: 1, H0626: 1,L0747: 1, L0757: 1,L0759: 1, L0597: 1 andL0608: 1.959160505 2-5831103Arg-1 to Lys-13,Leu-20 to Gln-32,Asp-77 to Gly-82,Gly-99 to Lys-104,Asp-113 to Tyr-119,Leu-142 to Val-153,Glu-172 to Asp-180.204HE8UY741163590214 1-492812AR061: 2, AR089: 1H0013: 1 and S0027: 1.960914506111-4551104205HAHIY08962113215 3-278813AR061: 10, AR089: 6206H2CBH451128919216 2-841814Leu-2 to Lys-10,AR061: 3, AR089: 3Ala-29 to Pro-43,H0437: 1, S0280: 1,Ser-63 to Thr-79,T0110: 1, H0622: 1,Pro-87 to Pro-104.L0745: 1, L0746: 1,L0731: 1 and L0596: 1.963811507 2-4211105Ala-1 to Met-18,Leu-20 to Asn-26,Val-38 to Leu-46,Pro-48 to Gly-53,Leu-81 to Gly-86,Gln-94 to Tyr-99,Glu-101 to Gly-109.207HMVAM091194828217 3-659815Asp-1 to Thr-7,AR089: 4, AR061: 1Asp-36 to Phe-51,L0731: 7, L0517: 5,Lys-61 to Gly-66,S0212: 3, L0775: 3,Gly-112 to Leu-120,L0740: 3, H0266: 2,Glu-134 to Tyr-150,L0809: 2, H0696: 2,Arg-193 to Lys-205.L0748: 2, S0132: 1,H0574: 1, H0013: 1,H0544: 1, H0023: 1,H0071: 1, H0286: 1,H0100: 1, H0494: 1,S0370: 1, L0770: 1,L0646: 1, L0764: 1,L0771: 1, L0363: 1,L0774: 1, L0659: 1,L0789: 1, L0666: 1,S0126: 1, H0522: 1,L0754: 1, L0747: 1 andL0755: 1.963814508 2-8021106208HFPEN041199663218 96-1646816Glu-62 to Tyr-67,AR061: 5, AR089: 2Pro-169 to Lys-179,S0010: 4, S0222: 3,Pro-189 to Ala-201,H0455: 2, L0803: 2,Ala-218 to Arg-223,L0439: 2, L0745: 2,Tyr-324 to Asn-331,S0282: 1, S0400: 1,Gly-352 to Val-357,H0456: 1, H0441: 1,Leu-365 to Lys-371,S0346: 1, H0509: 1,His-393 to Ala-399,L0769: 1, L0438: 1,Asp-420 to Asn-425,L0756: 1 and S0106: 1.Thr-460 to Lys-473,Ser-488 to Gly-502.964824509 88-5401107Glu-62 to Tyr-67,Ser-129 to Asp-135.209HSLJD021104452219 67-927817Leu-53 to Gln-58,AR054: 7, AR051: 1,Phe-162 to Gly-167,AR089: 1, AR061: 1,Gln-282 to Ala-287.AR050: 0S0390: 1965826510 47-9071108Leu-53 to Gln-58,Phe-162 to Gly-167,Gln-282 to Ala-287.210HDPFZ3012201642201637-819 818Leu-31 to Gly-41,AR089: 6, AR061: 2Arg-137 to Ser-143,H0250: 5, L0770: 2,Glu-241 to Glu-260.L0438: 2, L0439: 2,L0754: 2, S0114: 1,H0459: 1, H0489: 1,S0278: 1, H0069: 1,H0575: 1, H0318: 1,H0123: 1, L0471: 1,H0071: 1, H0328: 1,H0634: 1, T0067: 1,L0351: 1, H0560: 1,S0142: 1, S0344: 1,S0426: 1, L0763: 1,L0769: 1, L0761: 1,L0662: 1, L0363: 1,L0364: 1, L0805: 1,L0666: 1, L0664: 1,S0126: 1, H0658: 1,H0670: 1, H0521: 1,H0522: 1, S0044: 1,H0555: 1, H0576: 1,L0748: 1 and L0755: 1.9667525111637-819 1109Leu-31 to Gly-41,Arg-137 to Ser-143,Glu-241 to Glu-260.211HPJCR331217931221 140-1681819Ser-1 to Leu-13,AR089: 2, AR061: 1Pro-17 to Gly-31,L0754: 8, L0777: 8,Thr-44 to Leu-54,L0439: 5, H0266: 2,His-84 to Arg-95,L0438: 2, H0672: 2,Asn-105 to Gln-116,S0152: 2, L0745: 2,Pro-132 to Leu-138,L0758: 2, H0650: 1,Glu-148 to Gly-157,S0212: 1, S0045: 1,Arg-180 to Trp-185,S0046: 1, H0486: 1,Asn-340 to Glu-346,L0563: 1, H0051: 1,Asn-401 to Cys-412,H0644: 1, H0673: 1,Asp-430 to Ala-435,H0551: 1, H0269: 1,Thr-473 to Lys-478,S0344: 1, L0794: 1,Pro-490 to Tyr-498.L0766: 1, L0803: 1,L0805: 1, L0659: 1,H0547: 1, H0519: 1,S0126: 1, H0711: 1,H0528: 1, S0028: 1,L0750: 1, L0780: 1,L0757: 1, L0588: 1,L0591: 1, S0192: 1,H0423: 1 and H0293: 1.966758512 1-3751110212HTOAK341081321222784-599820Ser-10 to His-15.AR089: 1, AR061: 1L0766: 2, H0264: 1and H0521: 1.966800513 918-11961111Ser-67 to Trp-77.213HE8NI24971296223318-749821AR050: 3, AR051: 1,AR089: 0, AR061: 0H0013: 3, L0794: 2,L0439: 2, L0756: 2,L0779: 2, L0758: 2,S0001: 1, H0619: 1,L0638: 1, L0641: 1,L0776: 1 and H0435: 1.214HAMFM391055532224 782-2344822Gly-1 to Pro-7,AR050: 193, AR054:Gly-23 to Gly-50,122, AR051: 84,Ser-53 to Pro-84,AR089: 0, AR061: 0Ser-89 to Thr-129,H0255: 59, H0254: 10,Gly-140 to Gly-145,H0617: 9, L0747: 8,Pro-148 to Lys-158,S0358: 7, H0486: 6,Thr-161 to Ser-167,L0655: 6, H0208: 4,Leu-179 to Arg-189,H0545: 4, H0024: 4,Pro-203 to Lys-211,S0354: 3, H0250: 3,Glu-233 to Asp-240,H0123: 3, H0031: 3,Lys-261 to Gly-288,L0659: 3, S0328: 3,Arg-296 to Glu-305,L0731: 3, H0583: 2,Ala-315 to Arg-353,L0808: 2, L0785: 2,Glu-372 to Pro-382,H0662: 2, H0586: 2,Gln-395 to Glu-408,H0618: 2, H0253: 2,Asn-419 to Gly-427,H0424: 2, H0264: 2,Ala-458 to Gly-463,H0488: 2, H0100: 2,Pro-477 to Ala-483.L0771: 2, L0806: 2,L0809: 2, H0144: 2,H0689: 2, L0749: 2,L0750: 2, L0779: 2,L0777: 2, H0707: 2,L0595: 2, H0624: 1,H0341: 1, S0356: 1,S0360: 1, H0619: 1,H0411: 1, H0370: 1,H0485: 1, H0635: 1,H0025: 1, H0108: 1,H0318: 1, H0581: 1,T0110: 1, H0231: 1,L0738: 1, H0086: 1,H0271: 1, T0006: 1,H0644: 1, H0181: 1,H0124: 1, H0087: 1,T0067: 1, H0560: 1,H0646: 1, L0371: 1,L0800: 1, L0764: 1,L0768: 1, L0803: 1,L0774: 1, L0657: 1,L0368: 1, L0787: 1,L0666: 1, L0663: 1,L0665: 1, H0519: 1,H0414: 1, S0378: 1,S0380: 1, H0696: 1,S0044: 1, S0432: 1,L0439: 1, L0780: 1,L0755: 1, H0445: 1 andL0596: 1.9713475141121-29291112Gln-1 to Ala-7,Thr-36 to Trp-42,Gly-45 to Gly-52,Glu-77 to Pro-89,Gly-105 to Gly-132,Ser-135 to Glu-162.215HBGMG391126283225286-14 823Arg-27 to Gly-40,AR089: 1, AR061: 0Arg-67 to Asp-91.L0659: 12, L0769: 10,L0666: 8, L0747: 8,L0759: 7, L0439: 6,L0757: 6, L0756: 5,L0770: 4, L0761: 4,L0663: 4, L0665: 4,H0521: 4, L0749: 4,L0750: 4, L0777: 4,L0758: 4, H0550: 3,H0486: 3, H0544: 3,H0623: 3, L0662: 3,L0794: 3, L0766: 3,L0774: 3, L0664: 3,L0740: 3, L0779: 3,H0423: 3, S0418: 2,S0360: 2, L0717: 2,H0549: 2, H0618: 2,H0581: 2, H0545: 2,H0510: 2, H0617: 2,L0763: 2, L0772: 2,L0642: 2, L0764: 2,L0775: 2, L0655: 2,L0789: 2, S0374: 2,H0658: 2, H0522: 2,H0631: 2, L0745: 2,L0731: 2, H0556: 1,T0049: 1, H0656: 1,L0785: 1, H0483: 1,H0661: 1, H0664: 1,H0662: 1, S0420: 1,S0354: 1, S0358: 1,H0580: 1, S0468: 1,S0132: 1, S0222: 1,H0441: 1, H0586: 1,H0587: 1, H0497: 1,H0069: 1, H0427: 1,S0280: 1, H0046: 1,H0457: 1, H0081: 1,H0024: 1, T0010: 1,H0594: 1, H0188: 1,H0687: 1, H0553: 1,H0124: 1, H0494: 1,H0641: 1, S0422: 1,S0002: 1, S0426: 1,L0372: 1, L0646: 1,L0374: 1, L0648: 1,L0649: 1, L0803: 1,L0651: 1, L0653: 1,L0656: 1, L0635: 1,L0542: 1, L0526: 1,L0783: 1, L0809: 1,L0647: 1, L0791: 1,L0792: 1, H0698: 1,H0699: 1, H0693: 1,H0547: 1, H0689: 1,H0690: 1, H0683: 1,H0670: 1, S0378: 1,S0152: 1, H0555: 1,H0436: 1, S0392: 1,L0742: 1, L0751: 1,L0780: 1, H0668: 1,H0653: 1, S0242: 1,H0542: 1, H0543: 1 andS0460: 1.971414515 1-1951113Pro-1 to Asn-14,Lys-17 to Phe-23,Met-44 to Tyr-50.216HSXBV891128699226 1-2037824Gln-25 to Gly-33,AR054: 23, AR061: 4,Pro-49 to Gly-55,AR089: 3, AR050: 0Gly-89 to Glu-97,L0439: 31, L0741: 9,Ser-176 to Glu-183,L0438: 7, L0777: 6,Thr-231 to Gly-240,H0052: 5, H0617: 5,Pro-267 to Thr-275,L0748: 4, L0753: 4,Pro-297 to Asp-308,L0769: 3, L0775: 3,Asp-340 to Ser-345,L0776: 3, S0378: 3,Arg-353 to Leu-361,L0779: 3, S0040: 2,Pro-375 to Gly-382,L0103: 2, H0046: 2,Glu-393 to Trp-410,H0284: 2, T0006: 2,Gly-470 to Ser-475,S0036: 2, S0038: 2,Tyr-504 to Arg-516,L0351: 2, S0370: 2,Gly-531 to Thr-539,L0764: 2, H0670: 2,Pro-571 to Gln-580,L0602: 2, L0747: 2,Leu-591 to Glu-598,L0592: 2, S0342: 1,Gln-601 to Gly-611,S0282: 1, S0030: 1,Gly-649 to Ser-654,H0484: 1, S0007: 1,Asp-661 to Leu-666,S0278: 1, H0261: 1,Ala-669 to Glu-674.S0222: 1, H0441: 1,H0156: 1, T0082: 1,H0194: 1, T0010: 1,S6028: 1, H0271: 1,L0483: 1, H0424: 1,H0213: 1, H0181: 1,S0112: 1, S0144: 1,S0002: 1, L0520: 1,L0762: 1, L0763: 1,L0638: 1, L0772: 1,L0768: 1, L0653: 1,L0659: 1, L0636: 1,L0367: 1, L0791: 1,L0665: 1, L0352: 1,H0672: 1, H0539: 1,S0032: 1, L0742: 1,L0740: 1, L0758: 1 andH0667: 1.971821516 3-5091114Gln-20 to Gly-28,Pro-44 to Gly-50.217HBIOZ101143756227 3-572825Leu-50 to Asp-61,AR054: 189, AR051:Ser-100 to Leu-107,68, AR050: 35, AR089:Pro-119 to Leu-125.4, AR061: 3H0593: 1973131517 3-5031115Leu-50 to Asp-61,Ser-100 to Leu-107,Ala-120 to Thr-130.218HTLEJ111085651228 2-802826Tyr-52 to Gln-60.AR061: 3, AR089: 1H0618: 3 and H0253:1.973302518 2-8021116Tyr-52 to Gln-60,Phe-86 to Ala-94,Lys-111 to Arg-118,His-193 to Tyr-198.219HAWAM691207835229 1-1302827Val-2 to Cys-17,AR054: 334, AR050:Cys-41 to Gln-52,251, AR051: 249,Glu-70 to Phe-82,AR061: 6, AR089: 6Glu-89 to Ser-97,L0758: 12, L0662: 11,Lys-126 to Gly-132,H0251: 9, L0731: 9,Val-134 to Gly-145,S0360: 5, H0013: 5,Glu-167 to Arg-180,L0659: 5, L0747: 5,Glu-187 to Ser-200,H0252: 4, H0328: 4,Cys-204 to Ser-210,L0666: 4, L0439: 4,Glu-213 to Asp-221,H0135: 3, L0764: 3,Thr-260 to Ala-273,L0783: 3, L0749: 3,Ala-278 to Gln-290,S0358: 2, L0776: 2,Ser-317 to His-333,L0663: 2, H0651: 2,Leu-347 to Gly-356,L0744: 2, L0754: 2,Lys-358 to Phe-363,H0675: 1, H0329: 1,Leu-367 to Cys-376,H0619: 1, L0717: 1,Asp-385 to Ser-391,H0369: 1, H0550: 1,Glu-406 to Gly-434.H0333: 1, H0632: 1,H0486: 1, T0060: 1,H0042: 1, H0575: 1,H0618: 1, H0150: 1,H0123: 1, H0050: 1,H0105: 1, T0003: 1,H0024: 1, H0510: 1,H0594: 1, H0028: 1,H0644: 1, S0364: 1,S0366: 1, H0591: 1,H0100: 1, L0763: 1,L0631: 1, L0637: 1,L0646: 1, L0641: 1,L0644: 1, L0649: 1,L0803: 1, L0775: 1,L0782: 1, L0809: 1,L0519: 1, L0793: 1,L0665: 1, H0144: 1,L0438: 1, H0684: 1,H0672: 1, S0380: 1,L0748: 1, L0759: 1,L0596: 1, L0366: 1,L0600: 1 and H0352: 1.9431045191010-14411117Cys-38 to Gly-43,Gly-70 to Pro-82,Arg-129 to Glu-134,Gly-139 to Gly-144.973465520154-2 1118Leu-23 to Gly-32,Lys-34 to Lys-40.220HSCKD1110562882301238-2074828Ala-20 to Val-28,AR089: 1, AR061: 1Pro-60 to Cys-66,H0667: 1Ser-118 to Asp-123,Leu-225 to Asp-236,Thr-267 to His-274.973894521117-3261119Ala-6 to Ala-11,Phe-19 to Asn-24,Val-29 to Lys-34.221HDPLT621027241231195-800829Arg-9 to Leu-17,AR089: 1, AR061: 0Pro-90 to Asn-95,H0521: 1Lys-115 to Glu-125.973945522405-8091120222HTPFX16974296232 3-422830Asp-40 to Asn-49,AR061: 3, AR089: 2Cys-65 to Gly-71.L0750: 2, H0024: 1,H0039: 1, H0622: 1,H0040: 1 and S0434: 1.223HE9NO661079624233362-871831Phe-8 to Lys-27,AR061: 1, AR089: 1Ser-79 to Ser-87,L0774: 2 and H0144:Cys-102 to Val-116.2.974353523362-8711121Phe-8 to Lys-27,Ser-79 to Ser-87,Cys-102 to Val-116.224HSDJI441154068234 218-1921832Ala-324 to Phe-332,AR061: 0, AR089: 0Arg-336 to Thr-343,S0045: 6, H0255: 5,Pro-373 to Arg-384,S0028: 4, S0031: 2,Lys-424 to Asp-431.S0260: 2, H0341: 1,S0278: 1, H0333: 1,H0250: 1, S0050: 1,H0271: 1, H0100: 1,S0216: 1, S0044: 1 andS0390: 1.974784524 210-18471122Ala-324 to Phe-332,Arg-336 to Thr-343,Pro-373 to Arg-384,Lys-424 to Asp-431.225HFXDP531126294235 1-435833Gln-16 to Gly-25,AR061: 3, AR089: 1Thr-32 to Gly-42,S0001: 1Asn-46 to Asp-52.578868525 3-2721123Lys-1 to Arg-7,Phe-10 to Arg-19.226HWADY66734565236 1-186834AR061: 1, AR089: 1H0581: 1, H0494: 1,H0521: 1, H0444: 1,H0543: 1 and L0465: 1.227HLDBC631144557237 3-662835Phe-61 to Thr-68,AR061: 10, AR089: 4Arg-70 to Ser-76,L0754: 4, H0616: 1Gln-88 to Arg-93,and H0509: 1.Pro-145 to Gln-157,Glu-164 to Ser-171,Gly-215 to Thr-220.745061526 3-4161124Phe-61 to Thr-68,Arg-70 to Ser-76,Gln-88 to Arg-93.228HFIVB68978211238 3-665836Arg-20 to Thr-27,AR061: 1, AR089: 1Leu-40 to Gly-45,H0013: 2, S0468: 1,His-57 to Lys-74,S0046: 1, H0592: 1,Thr-97 to Ser-112,H0266: 1, S3014: 1,Asp-150 to Ser-155,S0028: 1 and S0196: 1.Leu-177 to Asp-182.752981527 82-6631125Thr-33 to Lys-47,Thr-70 to Ser-85,Asp-123 to Ser-128,Leu-150 to Asp-155.229HTLAC561181355239 3-698837AR061: 25, AR089: 15L0783: 3, S0007: 2,L0782: 2, H0539: 2,L0747: 2, H0333: 1,H0253: 1, H0052: 1,H0546: 1, T0006: 1,H0135: 1, L0770: 1,L0769: 1, L0776: 1,L0745: 1, L0777: 1 andL0753: 1.753093528 3-4221126230HSSAD41753094240 3-428838L0783: 3, S0007: 2,L0782: 2, H0539: 2,L0747: 2, H0333: 1,H0253: 1, H0052: 1,H0546: 1, T0006: 1,H0135: 1, L0770: 1,L0769: 1, L0776: 1,L0745: 1, L0777: 1 andL0753: 1.231HCFMT571218436241 3-1115839Ser-4 to Arg-15,AR061: 0, AR089: 0Glu-20 to Arg-62,L0157: 2, H0620: 2,Pro-107 to Gly-112,L0666: 2, S0001: 1,Gly-128 to Gly-134,L0717: 1, H0549: 1,Gln-137 to Arg-143,S0222: 1, H0581: 1,Ala-191 to Ala-212,H0194: 1, H0015: 1,Tyr-239 to Asn-247,H0399: 1, H0271: 1,Leu-300 to Pro-328,H0688: 1, H0428: 1,Pro-337 to Lys-350,H0124: 1, L0637: 1,Ser-356 to Leu-361,H0672: 1, L0439: 1,Gly-366 to Glu-371.L0750: 1 and H0423: 1.765375529380-3 1127Glu-5 to Arg-15,Glu-20 to Arg-62.232HDAAV611188787242 1-1332840Asp-34 to Asp-49,AR089: 25, AR061: 11Gly-276 to Ala-286,L0601: 5, H0266: 4,Ile-298 to Tyr-303,S0222: 3, H0265: 2,Leu-390 to Arg-395.H0556: 2, H0575: 2,H0052: 2, H0271: 2,S0114: 1, S0134: 1,S0420: 1, H0393: 1,H0550: 1, H0497: 1,H0318: 1, H0581: 1,H0251: 1, T0115: 1,H0014: 1, H0286: 1,H0494: 1, H0561: 1,L0766: 1, L0657: 1,H0698: 1, H0684: 1,S0330: 1, H0521: 1,S3014: 1, L0777: 1,S0260: 1, L0591: 1,L0594: 1 and H0543: 1.810305530 2-3431128Asp-90 to Lys-105.233HDPKD751096253243 2-637841Gly-2 to Asp-11,AR089: 4, AR061: 0Ala-23 to Asn-30,H0581: 1, H0494: 1,Phe-48 to Gly-56,H0521: 1, H0543: 1 andGlu-99 to His-105,L0465: 1.Glu-187 to Glu-192.810824531 2-4451129Ala-13 to Asn-20,Phe-38 to Gly-46,Glu-89 to His-95.234HTEON291126312244 1-516842Pro-27 to Ala-35,AR061: 6, AR089: 3Ser-138 to Asn-144.H0038: 4, L0758: 3,H0616: 2, L0794: 2,L0747: 2, L0803: 1,L0789: 1 and L0590: 1.815852532 2-5201130Pro-27 to Ala-35.235HSKAC241121800245 1-480843Ala-3 to Arg-20,AR061: 2, AR089: 1Ser-33 to Asp-39,H0370: 2, S0002: 1,Leu-70 to Ser-76,S0428: 1 and S0027: 1.Pro-117 to Tyr-122.823869533 98-4811131Ser-1 to Asp-7,Leu-38 to Ser-44,Pro-85 to Tyr-90.236HTJAA711216498246 25-1029844Arg-32 to Asn-39,AR089: 5, AR061: 4Leu-76 to Gly-82,S0328: 5, H0264: 4,Cys-112 to Ser-119,L0549: 3, S0306: 2,Gly-129 to Gly-135,H0379: 1, H0487: 1,Ala-141 to Val-167,S0448: 1, S0450: 1,Ser-181 to Ile-194,L0648: 1, L0551: 1 andSer-201 to Gly-239,S0330: 1.Ser-245 to Gln-250,Thr-256 to Thr-293,Ala-306 to Asp-335.846682534 23-3341132Arg-32 to Asn-39.237HTEKS201124378247 2-562845Pro-7 to Arg-12,AR061: 2, AR089: 1Phe-32 to Ile-37,L0758: 14, H0038: 5,Arg-39 to Lys-45,L0779: 4, L0794: 2 andLeu-47 to Gly-53,H0616: 1.Lys-102 to Lys-108,Asp-117 to Gly-122.846714535 3-5691133Pro-9 to Arg-14,Phe-34 to Ile-39,Arg-41 to Lys-47,Leu-49 to Gly-55,Lys-104 to Lys-110,Asp-119 to Gly-124.238HE9TK491125192248 3-863846Glu-134 to Glu-144,AR061: 3, AR089: 1Gln-151 to Arg-161,H0144: 2 and S0053: 1.Arg-167 to Gly-172,Tyr-183 to Asn-188,Asn-193 to Phe-209,Asp-261 to Trp-272.856343536 2-3281134239HCHAT011202275249 2-2269847Glu-75 to Glu-86,AR089: 1, AR061: 0Leu-176 to Gln-181,L0439: 12, L0748: 11,Ser-276 to Ala-282,L0751: 11, L0769: 7,Leu-320 to Lys-325,H0046: 6, L0756: 6,Met-366 to Ser-373,L0775: 5, L0666: 5,Leu-414 to Asp-419,L0747: 5, L0770: 4,Thr-471 to His-506,L0438: 4, L0740: 4,Lys-513 to Ile-522,L0777: 4, H0617: 3,Pro-526 to Gln-532,L0662: 3, L0774: 3,Asp-547 to Asp-552,L0776: 3, H0521: 3,Ala-576 to Cys-585,S0037: 3, L0749: 3,Glu-588 to His-598,L0731: 3, L0757: 3,Gly-637 to Pro-645,L0758: 3, S0212: 2,Leu-649 to Asp-657,S0222: 2, H0586: 2,Ile-733 to Phe-743,H0587: 2, H0333: 2,Ala-746 to Gly-753.H0156: 2, H0052: 2,S0388: 2, H0290: 2,L0640: 2, L0521: 2,L0766: 2, L0375: 2,L0659: 2, L0783: 2,H0144: 2, H0539: 2,L0755: 2, H0445: 2,L0596: 2, L0599: 2,H0149: 1, S0342: 1,H0294: 1, S0114: 1,H0484: 1, H0483: 1,H0664: 1, H0638: 1,S0418: 1, S0420: 1,L0005: 1, S0046: 1,S0300: 1, H0549: 1,H0550: 1, H0370: 1,H0497: 1, H0331: 1,H0486: 1, H0575: 1,S0010: 1, H0434: 1,H0327: 1, H0457: 1,H0041: 1, H0081: 1,H0620: 1, H0024: 1,H0057: 1, H0051: 1,H0083: 1, H0266: 1,H0188: 1, S0250: 1,H0688: 1, H0644: 1,H0674: 1, S0366: 1,H0087: 1, H0116: 1,H0488: 1, H0494: 1,H0131: 1, S0150: 1,H0633: 1, H0649: 1,H0652: 1, L0369: 1,L0638: 1, L0646: 1,L0641: 1, L0771: 1,L0773: 1, L0653: 1,L0658: 1, L0809: 1,L0789: 1, L0663: 1,L0664: 1, H0693: 1,H0520: 1, S0126: 1,H0682: 1, H0659: 1,S0330: 1, H0696: 1,S0174: 1, H0555: 1,S3012: 1, S0028: 1,L0742: 1, L0744: 1,L0745: 1, L0750: 1,L0786: 1, L0779: 1,L0752: 1, S0434: 1,L0366: 1, H0542: 1,H0423: 1 and H0352: 1.867209537 771-15561135Ser-20 to Ala-26,Leu-64 to Lys-69,Met-110 to Ser-117,Leu-158 to Asp-163.240HCEEN061150867250128-664848Pro-7 to Pro-14,AR089: 6, AR061: 3Asp-70 to Arg-80,L0666: 6, L0761: 4,Asp-145 to Gln-152.H0486: 3, L0794: 3,L0659: 3, H0255: 2,S0358: 2, H0052: 2,L0809: 2, L0743: 2,L0759: 2, H0692: 1,S0116: 1, H0581: 1,H0597: 1, L0055: 1,H0087: 1, L0763: 1,L0800: 1, L0644: 1,L0764: 1, L0766: 1,L0375: 1, L0655: 1,L0788: 1, L0663: 1,S0310: 1, H0672: 1,S0328: 1, H0539: 1 andH0436: 1.878658538 2-6191136Ser-1 to Ala-10,Cys-23 to Ala-29.241HDPKI83883382251101-673849Pro-19 to Lys-29,AR061: 3, AR089: 3His-38 to Phe-45,H0521: 7, H0580: 5,Ile-52 to Gln-66,L0665: 4, H0457: 3,Glu-123 to Glu-138.L0766: 3, L0745: 3,L0761: 2, L0806: 2,L0789: 2, L0750: 2,H0542: 2, H0650: 1,H0656: 1, H0581: 1,H0271: 1, H0553: 1,H0413: 1, H0641: 1,S0002: 1, L0774: 1,H0660: 1, H0555: 1,L0753: 1 and H0423: 1.242HSPBQ121152258252 1-588850His-4 to Gly-21,AR089: 2, AR061: 1Thr-55 to Ser-66,L0766: 19, L0439: 9,Pro-118 to Gly-130,L0803: 7, L0740: 7,Thr-168 to Ser-174.L0752: 7, L0770: 5,L0659: 5, L0731: 5,L0805: 4, L0777: 4,H0657: 3, H0373: 3,L0804: 3, S0152: 3,L0748: 3, L0749: 3,L0779: 3, H0650: 2,L0471: 2, S6028: 2,H0032: 2, L0783: 2,L0438: 2, H0521: 2,H0478: 2, L0744: 2,L0747: 2, L0750: 2,L0485: 2, S0424: 2,S0134: 1, S0354: 1,S0358: 1, H0580: 1,S0222: 1, H0013: 1,L0021: 1, H0575: 1,H0050: 1, H0014: 1,H0051: 1, H0031: 1,H0553: 1, H0165: 1,H0551: 1, H0509: 1,H0132: 1, H0652: 1,S0002: 1, H0529: 1,L0763: 1, L0772: 1,L0372: 1, L0771: 1,L0521: 1, L0662: 1,L0768: 1, L0386: 1,L0388: 1, L0774: 1,L0775: 1, L0655: 1,L0782: 1, L0809: 1,L0787: 1, L0789: 1,L0663: 1, L0664: 1,H0144: 1, L0352: 1,H0520: 1, H0435: 1,H0659: 1, H0658: 1,H0648: 1, H0672: 1,S0378: 1, S0380: 1,S0044: 1, L0780: 1,L0757: 1, L0758: 1 andH0543: 1.884004539 91-12871137243HPCID78886915253 1-429851Gly-1 to Cys-7.AR054: 2, AR051: 1,AR089: 1, AR061: 0H0642: 1244HDTKQ14886936254 1-555852Ser-60 to Thr-71,AR054: 60, AR051:Thr-82 to Leu-94,40, AR050: 36, AR089:Gln-113 to Asp-123,5, AR061: 2Val-125 to Tyr-133,H0521: 4, H0486: 2,Leu-144 to Gly-149.S0002: 2, L0770: 2,L0769: 2, L0766: 2,L0518: 2, L0783: 2,L0777: 2, L0731: 2,H0422: 2, H0556: 1,H0583: 1, H0650: 1,H0657: 1, H0179: 1,L0055: 1, H0488: 1,S0426: 1, L0662: 1,L0775: 1, L0655: 1,L0665: 1, S0053: 1,H0659: 1, L0754: 1,L0779: 1, L0759: 1 andH0543: 1.245HRACK83888037255 1-471853Gln-15 to Gln-21.AR089: 3, AR061: 2L0803: 4, L0758: 3,S0212: 2, S0358: 2,H0038: 2, L0770: 2,L0767: 2, L0766: 2,L0748: 2, L0751: 2,L0747: 2, L0759: 2,L0588: 2, L0599: 2,H0411: 1, H0392: 1,H0333: 1, L0021: 1,H0118: 1, T0115: 1,L0471: 1, L0163: 1,H0633: 1, L0769: 1,L0764: 1, L0775: 1,L0376: 1, L0806: 1,L0805: 1, L0807: 1,L0787: 1, H0547: 1,S0122: 1, H0555: 1,H0478: 1, L0744: 1,L0740: 1, L0749: 1,L0750: 1, L0755: 1 andL0595: 1.246HSIAO781156438256 2-631854Met-15 to Ser-20,AR089: 1, AR061: 0Asp-27 to Phe-37,S0354: 3, S0358: 3,Asp-53 to Tyr-59,H0587: 3, L0764: 3,Pro-86 to Asp-93,L0803: 3, L0758: 3,Pro-106 to Lys-129,H0036: 2, L0794: 2,Leu-139 to Ser-146,L0809: 2, S0374: 2,Thr-174 to Asp-183.S0376: 1, S0444: 1,S0408: 1, H0231: 1,L0783: 1, L0777: 1 andL0759: 1.889498540 2-6221138Asp-24 to Phe-34,Asp-50 to Tyr-56,Pro-83 to Asp-90,Pro-103 to Lys-126,Leu-136 to Ser-143,Thr-171 to Asp-180.247HWAGS73894404257 1-339855Val-14 to Lys-21,AR089: 2, AR061: 2Gln-41 to Trp-46,H0581: 3, H0622: 3,Ala-98 to Pro-103.H0575: 2, H0090: 2,L0777: 2, L0757: 2,S0114: 1, H0650: 1,H0255: 1, S0360: 1,S0278: 1, H0486: 1,H0318: 1, H0046: 1,H0457: 1, H0039: 1,H0553: 1, L0763: 1,L0761: 1, L0764: 1,L0789: 1, H0144: 1,S0374: 1, S0310: 1,H0555: 1, L0758: 1,H0445: 1 and S0276: 1.248HCMSL08898203258 1-1221856Gly-1 to Thr-10,AR051: 86, AR054:104770,Ala-14 to Gly-19,73, AR050: 67, AR089:107670,Pro-52 to Val-57,10, AR061: 5110700,Pro-85 to Gln-95,H0706: 8, S0366: 5,135940,Lys-198 to His-204,S0364: 4, L0485: 4,145001,Pro-254 to Glu-260,L0604; 4, L0777: 3,146790,Glu-269 to Ser-282,L0623: 2, S0362: 2,152445,Glu-302 to Gly-307,H0373: 2, L0520: 2,152445,Asp-320 to Asp-326,L0747: 2, H0624: 1,159001,Asp-373 to Ser-380,H0619: 1, H0550: 1,174000,Ile-396 to Asp-407.H0196: 1, L0646: 1,179755,L0809: 1, H0693: 1,182860,S0328: 1 and H0214: 1.182860,182860,191315,230800,230800,266200,600897,601105,601412,601652,6024919591765411372-569 1139Lys-59 to His-65,Pro-115 to Glu-121,Glu-130 to Ser-143,Glu-163 to Gly-168,Asp-181 to Asp-187,Asp-234 to Ser-241,Ile-257 to Asp-268.249HLWFN631101533259 404-2566857Thr-7 to Phe-29,AR051: 11, AR050: 9,Thr-37 to Lys-52,AR054: 5, AR089: 0,Glu-89 to Val-112.AR061: 0H0031: 5, S0222: 4,S0028: 4, H0662: 3,L0748: 3, S0260: 3,S0276: 3, S0282: 2,S0360: 2, S0046: 2,H0575: 2, H0196: 2,S0036: 2, H0268: 2,L0662: 2, S0027: 2,L0754: 2, L0747: 2,L0749: 2, L0756: 2,L0777: 2, L0604: 2,L0595: 2, H0171: 1,S0030: 1, S0029: 1,S0358: 1, H0619: 1,S0300: 1, L0717: 1,H0550: 1, H0441: 1,H0431: 1, H0392: 1,T0060: 1, S0010: 1,H0052: 1, H0309: 1,S6028: 1, S0250: 1,H0252: 1, H0553: 1,S0364: 1, S0366: 1,H0433: 1, H0269: 1,H0412: 1, L0372: 1,L0804: 1, L0789: 1,L0666: 1, L0663: 1,S0126: 1, S0044: 1,H0345: 1, S0390: 1,S0037: 1, S3014: 1,L0743: 1, L0439: 1,L0750: 1, L0779: 1,L0599: 1, L0593: 1,L0366: 1 and H0653: 1.908437542 404-25661140Thr-7 to Phe-29,Thr-37 to Lys-52,Glu-89 to Val-112.250HPWAY101128033260 397-1167858Gly-1 to Trp-7,AR089: 1, AR061: 0Leu-11 to Phe-21,H0341: 1, H0013: 1Glu-46 to His-52,and S0044: 1.Val-59 to Leu-73,Tyr-79 to Cys-91,His-111 toTyr-117,Ser-133 to Lys-149,His-167 to Tyr-173,His-195 to Tyr-201,His-251 to Lys-257.908549543149-4661141Cys-36 to Asn-43,Gln-74 to Trp-79.251HOUDH191153909261 3-404859Ser-2 to Gln-12,AR089: 1, AR061: 0Cys-14 to Met-19,S0040: 1, H0250: 1,Ser-34 to Leu-41,T0048: 1, L0761: 1,Pro-43 to Leu-48,L0764: 1, L0783: 1,Glu-89 to Asp-111,L0809: 1, L0789: 1 andIle-125 to Lys-134.L0757: 1.908588544 52-5731142Thr-8 to Gln-19,Lys-26 to Glu-33,Lys-41 to Ile-50.252HDPFF241194719262 124-1317860Ser-38 to Tyr-44,AR089: 4, AR061: 1Glu-72 to Pro-77,H0171: 5, S0026: 3,Asp-91 to Pro-113,S0400: 2, L0471: 2,Gln-124 to Asn-134,H0031: 2, H0553: 2,Ser-182 to Ile-190,H0547: 2, H0521: 2,Glu-215 to Gly-220,L0759: 2, H0423: 2,Leu-223 to Ser-229,H0170: 1, H0583: 1,Gln-243 to Ile-250,H0656: 1, S0001: 1,His-273 to His-290,S0358: 1, S0360: 1,His-301 to Lys-308,H0244: 1, H0349: 1,Thr-323 to Glu-336,H0590: 1, H0310: 1,Met-338 to Ser-343,H0014: 1, H0039: 1,Thr-358 to Tyr-363,S0366: 1, H0551: 1,His-385 to Asn-398.L0351: 1, H0509: 1,S0150: 1, L0369: 1,L0796: 1, L0773: 1,L0662: 1, L0766: 1,L0803: 1, L0635: 1,L0540: 1, H0519: 1,H0684: 1, H0660: 1,H0666: 1, S0044: 1,H0478: 1, H0479: 1,H0626: 1, L0748: 1,L0740: 1, L0777: 1,L0752: 1, L0755: 1 andH0543: 1.909232545104-4601143253HWLFH941152278263589-951861Glu-20 to Gly-25,AR089: 5, AR061: 2Gln-36 to Ser-48,S0358: 5, L0596: 3,Ser-56 to Val-62.L0771: 2, L0758: 2,S0354: 1, S0376: 1,T0109: 1, H0036: 1,H0590: 1, L0040: 1,H0038: 1, H0616: 1,L0646: 1, L0764: 1,L0768: 1, L0775: 1,L0659: 1 and S0404: 1.909682546134-5351144Ser-25 to Ala-52,Phe-64 to Glu-71.254HWMBM131152283264 3-557862Pro-17 to Ala-41,AR089: 2, AR061: 2Phe-53 to Glu-60,S0358: 6, L0794: 4,Glu-84 to Gly-89,L0758: 4, S0354: 3,Gln-100 to Ser-112,L0779: 3, L0596: 3,Ser-120 to Val-126.S0376: 2, H0036: 2,H0620: 2, H0063: 2,L0771: 2, L0803: 2,L0654: 2, L0659: 2,T0109: 1, H0013: 1,H0590: 1, H0052: 1,H0596: 1, T0110: 1,L0040: 1, H0090: 1,H0038: 1, H0040: 1,H0616: 1, H0429: 1,H0561: 1, L0646: 1,L0764: 1, L0768: 1,L0766: 1, L0775: 1,L0790: 1, L0792: 1,S0404: 1, S0390: 1,L0777: 1, L0755: 1,L0592: 1 and S0458: 1.909683547 3-5391145Pro-11 to Ala-35,Phe-47 to Glu-54,Glu-78 to Gly-83,Gln-94 to Ser-106,Ser-114 to Val-120.255HFIUE751172525265 79-1290863His-22 to Ile-35,AR089: 1, AR061: 1Phe-39 to Glu-47,L0748: 5, S0242: 3,Asp-65 to Ser-76,H0615: 2, S0376: 1,Ser-82 to Lys-89,S0360: 1, L0717: 1,Met-93 to Ser-98.L0641: 1, L0766: 1,L0664: 1, H0478: 1,L0593: 1 and S0196: 1.909758548 2-7751146Cys-1 to Val-10,Ala-14 to Met-22.256HNTCP131182313266 1-957864Leu-6 to Tyr-15,AR061: 3, AR089: 2Ser-48 to Phe-53,L0750: 4, H0519: 3,Asn-66 to Ser-71,L0666: 2, L0565: 2,Asp-101 to Pro-111,H0539: 2, L0742: 2,Gln-116 to Glu-124,L0744: 2, L0754: 2,Arg-203 to Asp-212.L0777: 2, L0759: 2,H0662: 1, S0045: 1,S0346: 1, H0251: 1,H0030: 1, H0628: 1,H0674: 1, H0529: 1,L0770: 1, L0764: 1,L0526: 1, L0783: 1,L0787: 1, H0547: 1,H0521: 1, H0696: 1,H0555: 1, L0747: 1,L0749: 1, L0786: 1,L0779: 1, L0780: 1,L0752: 1 and L0592: 1.909770549 1-9601147257HBIBQ891175111267 3-824865Gly-35 to Asp-41,AR089: 1, AR061: 0Phe-113 to Met-119,L0438: 6, L0751: 6,Pro-164 to Glu-170,L0439: 5, L0770: 4,Val-173 to Gly-178,H0052: 2, H0620: 2,Met-180 to Glu-190,H0521: 2, L0756: 2,Thr-192 to Gln-199,L0731: 2, L0758: 2,His-206 to Glu-211,L0588: 2, H0556: 1,Arg-244 to Ile-257.S0282: 1, H0662: 1,H0402: 1, S0418: 1,T0008: 1, S0222: 1,H0392: 1, H0333: 1,L0021: 1, H0581: 1,S0049: 1, L0471: 1,H0266: 1, L0351: 1,L0772: 1, L0766: 1,L0776: 1, L0659: 1,L0792: 1, H0522: 1,S0027: 1, L0779: 1 andS0011: 1.909782550 2-5771148258HWBEG181169125268439-161866Leu-42 to Ile-47.AR089: 82, AR061: 18H0580: 1909798551 55-6961149Trp-46 to Lys-51,Pro-109 to Asn-123,Phe-156 to Gly-165.259HTAHB431221956269 3-2174867AR089: 1, AR061: 0H0046: 34, L0731: 5,L0534: 4, L0769: 4,H0521: 4, S0356: 3,L0800: 3, L0794: 3,L0439: 3, L0749: 3,L0752: 3, L0759: 3,L0562: 2, H0486: 2,L0803: 2, L0805: 2,L0809: 2, L0789: 2,L0744: 2, L0485: 2,H0556: 1, H0657: 1,H0637: 1, H0580: 1,H0208: 1, H0261: 1,H0609: 1, H0455: 1,H0600: 1, H0586: 1,H0331: 1, H0635: 1,H0618: 1, H0544: 1,H0009: 1, H0050: 1,H0620: 1, H0288: 1,S0312: 1, S0314: 1,H0252: 1, H0688: 1,H0644: 1, S0366: 1,H0135: 1, H0063: 1,H0087: 1, H0551: 1,H0264: 1, S0002: 1,L0639: 1, L0771: 1,L0648: 1, L0766: 1,L0650: 1, L0378: 1,L0655: 1, H0699: 1,H0660: 1, L0743: 1,L0750: 1, L0777: 1,L0758: 1, L0097: 1,S0194: 1 and H0543: 1.909845552 18-9141150Gln-8 to Glu-13.260HSYBX32909846270 46-228868AR061: 0, AR089: 07q22-126650,H0046: 34, L0731: 5,q31.1126650,L0534: 4, L0769: 4,150240,H0521: 4, S0356: 3,154276,L0800: 3, L0794: 3,173360,L0439: 3, L0749: 3,173360,L0752: 3, L0759: 3,180105,L0562: 2, H0486: 2,222800,L0803: 2, L0805: 2,246900,L0809: 2, L0789: 2,602136,L0744: 2, L0485: 2,602136,H0556: 1, H0657: 1,602136,H0637: 1, H0580: 1,602447H0208: 1, H0261: 1,H0609: 1, H0455: 1,H0600: 1, H0586: 1,H0331: 1, H0635: 1,H0618: 1, H0544: 1,H0009: 1, H0050: 1,H0620: 1, H0288: 1,S0312: 1, S0314: 1,H0252: 1, H0688: 1,H0644: 1, S0366: 1,H0135: 1, H0063: 1,H0087: 1, H0551: 1,H0264: 1, S0002: 1,L0639: 1, L0771: 1,L0648: 1, L0766: 1,L0650: 1, L0378: 1,L0655: 1, H0699: 1,H0660: 1, L0743: 1,L0750: 1, L0777: 1,L0758: 1, L0097: 1,S0194: 1 and H0543: 1.261HCEHE351124531271 2-550869Arg-1 to Pro-15,AR061: 8, AR089: 3Asn-17 to Leu-25,S0222: 1, H0052: 1,Glu-27 to Pro-36,H0194: 1, H0290: 1 andPro-41 to Pro-55,H0264: 1.Glu-58 to Gln-79.909937553 3-3921151Asn-6 to Pro-13.262HFCBB561204971272112-753870Thr-37 to Ser-42,AR061: 1, AR089: 1Gln-48 to Pro-55,H0009: 1Ser-75 to Ala-80,Ser-95 to Val-111,Gln-113 to Gly-124,Glu-153 to Gly-161,Tyr-188 to Asp-193.910073554209-5651152263HAMFL82910074273 3-317871Glu-20 to Val-26.AR089: 9, AR061: 4H0229: 1, H0590: 1,S0049: 1, H0014: 1,H0560: 1, L0439: 1 andH0543: 1.264HBXCM381174533274 402-1349872AR061: 2, AR089: 1L0439: 6, S0038: 3,L0803: 3, H0455: 2,L0769: 2, L0809: 2,L0741: 2, L0756: 2,S6024: 1, S0001: 1,H0663: 1, S0222: 1,H0441: 1, H0438: 1,H0036: 1, S0049: 1,H0309: 1, H0566: 1,H0024: 1, S0388: 1,S0051: 1, T0010: 1,H0059: 1, L0645: 1,L0774: 1, L0790: 1,L0663: 1, L0665: 1,H0345: 1, L0742: 1,L0748: 1, L0749: 1,H0707: 1, L0595: 1 andL0366: 1.910086555 402-15351153Val-36 to Glu-43,Lys-66 to Glu-71.265HLHCR16910123275 2-3418873Pro-9 to Pro-15,AR050: 9, AR061: 2,Gly-49 to Trp-54,AR054: 2, AR089: 2,Ser-91 to Phe-96,AR051: 2Thr-109 to Asp-115,L0754: 14, L0777: 13,Cys-124 to Ile-130,H0553: 10, L0600: 7,Cys-164 to Trp-169,L0748: 6, L0803: 4,Thr-193 to Asp-207,L0749: 4, UNKWN: 4,Thr-215 to Tyr-220,H0624: 3, S0280: 3,Thr-228 to Ser-240,S0126: 3, L0747: 3,Glu-269 to Ser-276,S0282: 2, H0024: 2,Glu-327 to Ala-334,H0030: 2, H0031: 2,Asn-376 to Asp-392,H0040: 2, L0438: 2,Gln-420 to Asn-428,S0028: 2, L0743: 2,Tyr-547 to Ser-566,L0596: 2, L0603: 2,Ala-616 to Gly-623,S0212: 1, H0270: 1,Pro-625 to Ser-631,H0244: 1, H0427: 1,Ser-647 to Val-653,H0251: 1, H0309: 1,Gly-676 to Pro-681,S0338: 1, S0340: 1,Tyr-720 to Glu-740,S0250: 1, H0252: 1,Ile-742 to Lys-748,H0039: 1, L0143: 1,Asp-792 to Cys-804,H0038: 1, L0659: 1,Leu-841 to Val-848,L0565: 1, H0593: 1,Gln-850 to Gly-857,H0684: 1, H0518: 1,Asp-879 to Gly-886,S0390: 1, S0260: 1 andHis-906 to Trp-913,H0506: 1.Pro-968 to Thr-975,Gln-1051 to Ser-1057,Pro-1092 to Cys-1099,Lys-1113 to Cys-1120,Trp-1126 to Phe-1139.965511556 2-14921154Pro-9 to Pro-15,Gly-49 to Trp-54,Ser-91 to Phe-96,Thr-109 to Asp-115,Cys-124 to Ile-130,Cys-164 to Trp-169,Thr-193 to Asp-207,Thr-215 to Tyr-220,Thr-228 to Ser-240,Glu-269 to Ser-276,Glu-327 to Ala-334,Asn-376 to Asp-392,Gln-420 to Asn-428.266HE6GF021153883276 1-612874Gln-13 to Ser-18,AR061: 7, AR089: 4Glu-32 to Gly-37,H0100: 1 and H0521: 1Ala-44 to Trp-49,Glu-56 to Val-61,Gln-68 to Lys-74,Ala-83 to Glu-88,Arg-111 to Gly-117,Tyr-123 to His-143,Ser-167 to Met-201.911263557 1-2641155Gln-13 to Ser-18,Glu-32 to Gly-37,Ala-44 to Trp-49.267HOUFT361162680277 853-1563875Thr-70 to Ala-83,AR089: 2, AR061: 1Gly-105 to Asn-110,L0794: 6, L0598: 2,Ser-181 to Val-187.L0803: 2, L0748: 2,S0040: 1, S0046: 1,H0431: 1, H0318: 1,L0766: 1, L0606: 1,L0749: 1, L0758: 1 andS0192: 1.911293558160-8461156Lys-27 to Ile-43.268HAGGF841197460278 415-1530876Arg-24 to Arg-30,AR061: 3, AR089: 2Arg-39 to Tyr-44,L0766: 18, L0748: 11,Lys-78 to Glu-91,L0439: 9, L0749: 8,Val-215 to Lys-223.L0438: 5, L0750: 5,L0777: 4, L0759: 4,H0441: 3, H0052: 3,L0637: 3, L0761: 3,L0740: 3, L0747: 3,L0103: 2, H0574: 2,H0156: 2, H0597: 2,S0250: 2, L0649: 2,L0803: 2, L0806: 2,L0792: 2, S3014: 2,L0757: 2, L0485: 2,L0599: 2, H0171: 1,S6024: 1, L0002: 1,H0657: 1, H0341: 1,S0358: 1, S0360: 1,S0132: 1, L0717: 1,H0632: 1, H0013: 1,H0599: 1, S0010: 1,S0346: 1, H0318: 1,H0251: 1, T0115: 1,H0544: 1, L0471: 1,H0014: 1, S0362: 1,H0083: 1, H0188: 1,H0428: 1, H0646: 1,H0538: 1, L0598: 1,L0762: 1, L0763: 1,L0769: 1, L0662: 1,L0768: 1, L0776: 1,L0655: 1, L0659: 1,L0526: 1, L0783: 1,L0789: 1, L0665: 1,S0148: 1, H0520: 1,H0519: 1, S0330: 1,L0602: 1, S0152: 1,S0136: 1, S0350: 1,L0752: 1, H0343: 1,L0366: 1, S0011: 1,H0665: 1, S0196: 1,H0423: 1, L0697: 1 andS0462: 1.911312559 1-3331157Lys-14 to Glu-27.269HTTKP071119031279 2-571877Thr-15 to Asp-25,AR089: 1, AR061: 1Glu-69 to Leu-89,H0634: 2Glu-130 to Arg-137.911390560 2-3371158Thr-15 to Asp-25,Glu-69 to Leu-89.270HE9SE621171014280718-212878AR061: 16, AR089: 6L0804: 1, S0052: 1,H0144: 1 and H0659: 1.911476561 1-5641159271HUJAD241162674281 788-1255879Gln-49 to Thr-69,AR089: 1, AR061: 0His-129 to Cys-143.L0750: 3, H0650: 2,H0637: 2, H0265: 1,H0556: 1, S0222: 1,H0040: 1, H0280: 1,L0655: 1, L0789: 1 andL0666: 1.911498562 3-2931160272HWLFG751228123282 1-819880Val-15 to Gly-26,AR089: 1, AR061: 0Pro-43 to Ala-64,L0439: 7, L0770: 4,Arg-75 to Gly-82,L0771: 4, L0779: 4,Thr-115 to Thr-120,H0688: 3, H0617: 2,Leu-215 to Gly-222,L0533: 2, L0803: 2,Ser-230 to Trp-235,L0807: 2, L0791: 2,Pro-237 to Ala-248.L0666: 2, H0539: 2,H0624: 1, S0400: 1,H0125: 1, H0192: 1,S0356: 1, S0354: 1,S0376: 1, S0360: 1,S0278: 1, H0550: 1,H0333: 1, S0049: 1,H0205: 1, S0051: 1,L0142: 1, L0455: 1,L0769: 1, L0794: 1,L0658: 1, L0540: 1,L0518: 1, L0809: 1,L0663: 1, H0689: 1,S0332: 1, H0214: 1,S3014: 1, L0747: 1,L0749: 1, L0758: 1,S0260: 1 and S0458: 1.916563563 1-7501161Val-10 to Gly-21,Pro-38 to Ala-59,Arg-70 to Gly-77,Thr-110 to Thr-115,Leu-210 to Gly-217,Ser-225 to Trp-230,Pro-232 to Arg-239.273HT3BG121092417283 9-785881AR061: 8, AR089: 3L0758: 3, H0159: 2,S0001: 1, H0618: 1,H0660: 1 and L0779: 1.921593564 1-3811162Glu-1 to Ala-15,Lys-25 to Ser-32,Asp-45 to Thr-51,Pro-59 to Pro-65,Pro-78 to Ser-85.274HTLJC71922923284 3-1355882His-1 to Phe-9,AR061: 7, AR089: 5Cys-13 to Thr-18,H0618: 12, H0253: 8,Pro-35 to Gly-48,H0038: 6, L0758: 6,Glu-61 to Pro-68,L0779: 5, H0616: 3,Lys-105 to Ala-136,T0041: 1, L0776: 1,Thr-144 to Gln-154,S0274: 1 and H0543: 1.Leu-163 to Gly-171,Thr-205 to Gln-222,Pro-251 to Gln-257.275HCOMM051194701285 2-850883Gln-22 to Asp-41,AR089: 1, AR061: 1Pro-49 to Thr-58,H0670: 1Leu-99 to Gly-107,Ala-117 to Ala-122,Gln-128 to Trp-134,Pro-136 to Pro-144,Phe-147 to Glu-153,Glu-183 to Val-188,Glu-195 to Glu-200,Glu-257 to Leu-265,Met-275 to Ser-283.925952565 1-8401163Gln-19 to Asp-38,Pro-46 to Thr-55,Leu-96 to Gly-104,Ala-114 to Ala-119,Gln-125 to Trp-131,Pro-133 to Pro-141,Phe-144 to Glu-150,Glu-180 to Val-185,Glu-192 to Glu-197,Glu-254 to Leu-262,Met-272 to Ser-280.276HSLJE541229928286 650-1654884Ala-17 to Val-30,AR061: 0, AR089: 0Thr-32 to Arg-39,S0036: 1, H0521: 1,Arg-55 to Ser-60,H0436: 1 and S0390: 1.Ala-213 to Leu-219,Glu-236 to Glu-241,Ser-262 to Tyr-272,Pro-299 to Asn-305.926924566 3-7311164Arg-1 to Gly-7,Pro-25 to His-34,Leu-36 to Lys-49.277HTGED07927411287 16-483885Lys-35 to Asn-46.AR089: 33, AR061: 7278HOFNH30928365288 3-320886AR089: 4, AR061: 2H0415: 13, H0414: 2,H0355: 1, H0517: 1 andH0539: 1.279HWNCY051179767289 1-981887Lys-15 to Gly-23,AR061: 2, AR089: 1Glu-36 to His-47.H0656: 2, S0360: 2,H0657: 1, H0662: 1,S0420: 1, S0356: 1,S0358: 1, S0132: 1,H0392: 1, S0022: 1,H0144: 1, H0520: 1,H0659: 1, H0658: 1,H0660: 1, H0672: 1,S0380: 1, L0602: 1,H0653: 1 and H0677: 1.928789567 3-13191165Lys-11 to Gly-19,Glu-32 to His-43,Lys-60 to Glu-66,Pro-86 to Lys-98,Lys-118 to Leu-128,Thr-142 to Trp-148.280HDPDA47929193290103-906888Arg-17 to Leu-34,AR089: 11, AR061: 3Asp-44 to Ser-51,H0521: 7, H0581: 3,Asp-63 to Gly-72,H0422: 3, H0650: 2,Pro-74 to Gly-83,H0486: 2, S0002: 2,Thr-97 to Met-102.L0770: 2, L0769: 2,L0766: 2, L0518: 2,L0783: 2, L0777: 2,L0731: 2, H0445: 2,H0556: 1, H0583: 1,H0657: 1, H0656: 1,H0341: 1, H0575: 1,H0457: 1, H0179: 1,H0271: 1, L0055: 1,H0264: 1, H0488: 1,S0426: 1, L0662: 1,L0775: 1, L0655: 1,L0665: 1, S0053: 1,H0702: 1, H0701: 1,H0659: 1, L0754: 1,L0779: 1, L0759: 1 andH0543: 1.281HWMEV63931154291 2-454889His-9 to Asn-26,AR089: 1, AR061: 13q21-q25106165,Pro-47 to Ser-61,S0358: 1 and H0580: 1.117700,Arg-116 to Thr-122.117700,150210,169600,180380,180380,180380,190000,203500,222900,232050,276902,600882,601199,601199,601199,601471,601682282HCFAT251052857292 2-640890Pro-19 to Lys-34,AR061: 2, AR089: 2Arg-63 to Arg-72,S0358: 1, H0413: 1,Lys-76 to Pro-113,L0502: 1, L0657: 1,Gln-133 to Gln-150,H0522: 1 and H0422: 1.Gln-152 to Gln-163,Glu-167 to Arg-187.932068568 82-5881166Lys-15 to Ser-20,Arg-51 to Arg-60,Lys-64 to Pro-101.283HHEQV391165420293 1-711891Leu-7 to Phe-27,AR089: 3, AR061: 1Gln-50 to Gln-57.T0042: 1, H0543: 1and H0422: 1.932851569 1-7111167Leu-7 to Phe-27,Gln-50 to Gln-57.284HHFJH791228195294 608-2260892Ser-39 to Pro-44,AR061: 2, AR089: 2Pro-51 to Thr-56,H0424: 9, L0747: 7,Gly-78 to Cys-92,H0618: 5, H0620: 5,Pro-133 to Ser-146,L0809: 5, H0549: 4,Glu-211 to Ala-219,H0087: 4, L0655: 4,His-239 to Gln-250,L0750: 4, S0222: 3,Thr-280 to Glu-285,H0253: 3, S0346: 3,Asn-347 to Gly-371,H0150: 3, H0081: 3,Asp-378 to Asn-385,H0083: 3, H0188: 3,Arg-390 to Gly-398,H0428: 3, H0213: 3,Thr-406 to Gly-420,L0774: 3, L0805: 3,Pro-422 to Gly-427,L0749: 3, L0759: 3,Arg-440 to Leu-450,L0005: 2, H0619: 2,Pro-458 to Gly-463,S0278: 2, H0635: 2,Pro-485 to Leu-490,S0049: 2, H0251: 2,Gly-526 to Pro-533.H0546: 2, H0009: 2,H0012: 2, H0024: 2,H0617: 2, H0494: 2,L0769: 2, L0637: 2,L0772: 2, L0803: 2,L0518: 2, L0384: 2,L0665: 2, L0438: 2,H0547: 2, H0658: 2,S0152: 2, H0521: 2,L0439: 2, L0779: 2,L0758: 2, L0592: 2,L0599: 2, L0595: 2,H0543: 2, S0424: 2,H0352: 2, H0624: 1,H0686: 1, H0295: 1,S0114: 1, S0134: 1,H0341: 1, S0212: 1,H0254: 1, S0420: 1,S0354: 1, H0351: 1,H0550: 1, H0586: 1,H0333: 1, H0643: 1,L0623: 1, H0486: 1,T0039: 1, H0013: 1,T0082: 1, H0052: 1,H0597: 1, H0545: 1,H0178: 1, L0157: 1,H0594: 1, H0028: 1,H0604: 1, H0032: 1,H0674: 1, S0364: 1,H0361: 1, H0135: 1,H0551: 1, H0264: 1,H0100: 1, T0042: 1,H0561: 1, L0369: 1,L0770: 1, L0639: 1,L0800: 1, L0662: 1,L0768: 1, L0364: 1,L0794: 1, L0804: 1,L0375: 1, L0378: 1,L0653: 1, L0776: 1,L0659: 1, L0365: 1,L0542: 1, L0783: 1,L0789: 1, L0790: 1,L0791: 1, L0666: 1,S0052: 1, T0068: 1,H0689: 1, H0690: 1,H0684: 1, H0670: 1,H0660: 1, H0648: 1,H0672: 1, S0044: 1,L0741: 1, L0743: 1,L0748: 1, L0751: 1,L0756: 1, L0752: 1,L0731: 1, L0757: 1,H0665: 1, L0096: 1 andS0194: 1.933308570 2-8321168285HUCOW171155190295 1-993893Ser-30 to Ser-35.AR089: 4, AR061: 2L0439: 5, S0002: 3,L0604: 3, H0619: 2,H0024: 2, H0625: 2,L0768: 2, L0757: 2,H0638: 1, S0420: 1,S0360: 1, H0586: 1,L0163: 1, S0214: 1,L0143: 1, H0264: 1,L0769: 1, L0764: 1,L0774: 1, L0651: 1,L0659: 1, L0542: 1,L0789: 1, H0539: 1,H0521: 1, S0044: 1,L0777: 1, L0758: 1,L0599: 1 and H0422: 1.933357571155-8561169Gln-27 to Trp-45.286HFKIT061078092296 1-1119894Lys-6 to Trp-11,AR089: 0, AR061: 0Pro-26 to Pro-40,H0620: 2, L0761: 2,Pro-48 to Gln-53.L0766: 2, L0744: 2,L0754: 2, L0596: 2,H0686: 1, H0295: 1,H0657: 1, H0597: 1,H0009: 1, H0264: 1,S0002: 1, L0769: 1,L0774: 1, L0805: 1,L0657: 1, L0790: 1,H0690: 1 and H0521: 1.934019572 1-3001170Asp-2 to Pro-7,Pro-15 to Gln-20.287HDTBY881104159297 3-536895His-130 to Lys-140.AR089: 8, AR061: 2S0218: 1 and H0486: 1.934472573 3-4641171His-130 to Lys-140.288HWLHS821082268298 3-1349896Pro-22 to Thr-29,AR089: 2, AR061: 1Gly-65 to Lys-75.L0769: 3, S0354: 1,H0393: 1, H0355: 1 andH0124: 1.934505574 2-4271172Gly-34 to Lys-44,Glu-113 to Glu-118.289HDPNC961081629299 3-734897Val-2 to Gly-8,AR089: 1, AR061: 1Asp-20 to Gln-26.H0522: 2 and L0766:1.934520575 3-7341173Val-2 to Gly-8,Asp-20 to Gln-26.290HCE5I781197899300 2-1126898Ala-7 to Gly-15,AR061: 5, AR089: 2Pro-33 to Gln-39,L0439: 8, H0052: 7,Ala-48 to Ala-71,L0741: 7, L0756: 4,Tyr-83 to Ser-89,S0010: 3, H0261: 2,Ser-122 to Ile-139,H0156: 2, S0049: 2,Thr-150 to Thr-158,L0770: 2, L0776: 2,Lys-183 to Phe-193,L0742: 2, L0745: 2,Pro-277 to Asn-299,L0366: 2, S0222: 1,Asp-324 to Gly-333,H0438: 1, H0390: 1,Lys-354 to Glu-361,S0346: 1, H0009: 1,Gln-367 to Ser-374.L0455: 1, S0038: 1,L0789: 1 and L0758: 1.934531576 3-4221174Pro-14 to Gln-20,Ala-29 to Ala-52,Tyr-64 to Ser-70,Ser-103 to His-120.291HISDS621159625301182-862899Ser-39 to Trp-44,AR089: 2, AR061: 1Ile-48 to Trp-54,T0049: 1, S0278: 1,Asn-65 to Asp-87,H0031: 1 and H0539: 1.Pro-94 to Gln-100,Lys-129 to Asp-136,Asp-163 to His-174,Ser-193 to His-199.935932577 1-5191175Ser-11 to Trp-16,Ile-20 to Trp-26,Asn-37 to Ser-58,Leu-67 to Gln-72,Lys-101 to Asp-108,Asp-135 to Tyr-140.292HDQDV691212566302 210-1697900Met-7 to Ser-12,AR089: 46, AR061: 33Ser-20 to Arg-30,H0521: 4, H0051: 2,Asp-85 to Ala-92,L0803: 2, L0748: 2,Met-119 to Asn-146,L0740: 2, L0756: 2,Pro-151 to Asp-161,L0752: 2, L0755: 2,Gln-253 to Glu-260,H0590: 1, H0014: 1,Ile-333 to Val-342,S0250: 1, L0772: 1,Leu-396 to Ala-406.L0764: 1, L0804: 1,H0522: 1, S0406: 1,L0754: 1, L0779: 1,L0731: 1 and L0758: 1.937850578 2-8291176949702579551-3391177Lys-1 to Thr-7,Arg-34 to Pro-41.293HEMBT61939957303 1-351901AR061: 8, AR089: 4L0547: 2, S0046: 1,L0471: 1, L0772: 1,L0529: 1 and L0780: 1.294HRODZ701088554304 2-661902Lys-123 to Lys-128,AR089: 12, AR061: 4Trp-180 to Lys-186,H0598: 1 and H0135:Leu-204 to Thr-220.1.942673580 3-4401178Lys-49 to Lys-54,Trp-106 to Lys-112,Leu-130 to Gly-141.295HHERQ791184003305284-703903Ser-3 to Thr-11,AR089: 3, AR061: 2Lys-32 to Gly-39,H0597: 1, H0435: 1Thr-50 to Glu-57,and H0543: 1.Thr-83 to Gln-88.944057581 88-4741179Ser-3 to Thr-11,Lys-32 to Gly-39,Thr-50 to Glu-57,Thr-83 to Gln-88.296HCECM901031741306 2-577904Gly-12 to Gly-31,AR061: 2, AR089: 1Asn-38 to Gly-62,H0013: 3, L0439: 2,Asp-70 to Phe-84,H0624: 1, H0171: 1,Val-94 to Ser-101,S0040: 1, S0420: 1,Ala-112 to Ser-125,H0619: 1, H0156: 1,Lys-140 to Asn-145,H0575: 1, H0590: 1,Asn-175 to Tyr-180,H0052; 1, H0011: 1,Arg-187 to Thr-192.H0266: 1, H0494: 1,L0519: 1, H0519: 1,H0555: 1, L0777: 1,L0758: 1, S0436: 1 andH0506: 1.945088582 2-5771180Gly-12 to Gly-31,Asn-38 to Gly-62,Asp-70 to Phe-84,Val-94 to Ser-101,Ala-112 to Ser-125,Lys-140 to Asn-145,Asn-175 to Tyr-180,Arg-187 to Thr-192.297HWHGW721199614307 3-1208905Lys-63 to Pro-72,AR054: 23, AR050:Val-97 to Gly-102,16, AR051: 3, AR089:His-116 to Cys-123,1, AR061: 1Tyr-161 to Thr-167,H0586: 1 and L0375:Pro-204 to Lys-210,1.Ala-214 to Lys-222,Glu-276 to Lys-289,Tyr-305 to Thr-312,Pro-383 to Gly-398.945692583100-9391181947361584327-1 1182Gly-1 to Gly-7,Ala-13 to Gln-21,Ala-43 to Ser-48,Asn-67 to Gly-75,Pro-82 to Pro-90.298HPCRV841219890308123-875906Thr-1 to Leu-12,AR089: 0, AR061: 0Asp-107 to Thr-114,Pro-162 to Leu-170.945856585112-4171183Thr-1 to Leu-12.299HNSAA28946988309 85-1557907Glu-9 to Ser-20,AR050: 8, AR054: 6,Ile-23 to Gly-29,AR051: 3, AR089: 1,Pro-50 to Cys-66,AR061: 1Pro-74 to Glu-79,H0036: 2, L0766: 2,Glu-93 to Trp-98,H0686: 1, H0622: 1,Thr-121 to Ser-133,H0625: 1, L0791: 1,Leu-180 to Lys-196,L0779: 1 and S0434: 1.Thr-213 to Glu-225,Glu-234 to Glu-240,Arg-263 to Glu-270,Glu-283 to Ala-298,Lys-318 to Ala-336,Val-340 to Ala-351,Val-361 to Pro-372,Asn-445 to Pro-468,Pro-475 to Lys-491.972348586 3-4521184Thr-1 to Ala-10,Val-20 to Pro-31,Asn-104 to Thr-124.300HLWAR779474843101287-292 908Gln-97 to Pro-114,AR050: 21, AR054: 9,Trp-117 to Lys-129,AR051: 3, AR089: 1,Thr-166 to Gln-173,AR061: 1Ser-178 to Lys-183,H0553: 4 and L0759:Glu-250 to Phe-256,2.Ser-295 to His-301,Tyr-307 to Gln-316,Glu-322 to Ser-330.301HTTJW491127477311 423-1319909Phe-8 to Gln-13,AR061: 3, AR089: 2Arg-63 to Gly-69,L0769: 3, L0803: 3,Gly-135 to Lys-144,L0748: 3, L0749: 3,Ala-201 to Ala-211,H0574: 2, H0046: 2,Arg-248 to Thr-255,L0794: 2, L0776: 2,Leu-294 to Pro-299.L0439: 2, L0754: 2,L0747: 2, L0755: 2,L0605: 2, L0593: 2,H0686: 1, S0360: 1,L0717: 1, H0069: 1,H0575: 1, H0620: 1,H0024: 1, S0388: 1,H0510: 1, H0266: 1,H0644: 1, H0163: 1,H0090: 1, H0634: 1,H0561: 1, H0695: 1,L0763: 1, L0804: 1,L0774: 1, L0775: 1,L0659: 1, L0783: 1,L0809: 1, L0666: 1,L0665: 1, L0438: 1,H0519: 1, H0658: 1,H0539: 1, S0152: 1,H0522: 1, L0740: 1,L0777: 1, L0603: 1,S0276: 1 and H0542: 1.948107587 25-6601185Val-11 to Gly-21,Gly-72 to Thr-80.302HWAFS181155193312 54-1718910Pro-1 to Pro-7,AR089: 4, AR061: 3Leu-10 to Lys-18,H0581: 3, H0622: 3,Val-119 to Lys-126,H0575: 2, H0090: 2,Gln-146 to Trp-151.L0777: 2, L0757: 2,S0114: 1, H0650: 1,H0255: 1, S0360: 1,S0278: 1, H0486: 1,H0318: 1, H0457: 1,H0039: 1, H0553: 1,L0763: 1, L0761: 1,L0764: 1, L0789: 1,H0144: 1, S0374: 1,S0310: 1, H0555: 1,L0758: 1, H0445: 1 andS0276: 1.948434588 54-7911186Pro-1 to Pro-7,Leu-10 to Lys-18,Val-119 to Lys-126,Gln-146 to Trp-151,Asp-210 to Arg-216.303HFCBA441082762313 1-669911AR089: 14, AR061: 9H0457: 1, H0009: 1,L0666: 1, S0053: 1 andL0741: 1.948533589192-4941187304HVADT771180374314 1-249912Thr-28 to Lys-34,AR089: 4, AR061: 2Pro-36 to Asn-44,L0744: 9, L0747: 8,Lys-72 to Lys-83.S3014: 7, L0740: 7,S0192: 6, S0027: 5,S0212: 4, H0124: 4,L0731: 4, L0662: 3,L0743: 3, L0752: 3,L0759: 3, H0662: 2,S0418: 2, S0046: 2,H0575: 2, H0545: 2,H0041: 2, H0413: 2,L0775: 2, H0696: 2,L0748: 2, L0751: 2,L0754: 2, L0749: 2,L0758: 2, H0445: 2,S0276: 2, H0624: 1,L0778: 1, L0005: 1,H0645: 1, H0441: 1,H0391: 1, S0005: 1,T0040: 1, H0069: 1,H0427: 1, S0280: 1,H0042: 1, T0048: 1,H0505: 1, H0309: 1,H0544: 1, H0009: 1,H0266: 1, H0617: 1,H0412: 1, H0623: 1,T0004: 1, L0564: 1,T0041: 1, H0494: 1,H0633: 1, H0646: 1,H0652: 1, L0769: 1,L0646: 1, L0655: 1,L0659: 1, L0546: 1,L0783: 1, L0809: 1,H0144: 1, L0565: 1,S0126: 1, H0689: 1,H0435: 1, H0659: 1,H0672: 1, S0378: 1,H0555: 1, S0037: 1,S0206: 1, L0777: 1,L0780: 1, S0434: 1,S0011: 1, S0194: 1 andH0506: 1.948886590 1-3301188Thr-11 to Trp-25,Ser-35 to Arg-42,Asp-50 to Arg-56,Tyr-75 to Ser-81,Gly-89 to Gln-104.305HUFCN911189013315 25-858913Cys-7 to Ala-24,AR089: 2, AR061: 2Asn-30 to Asn-42,H0441: 5, H0134: 3,Ser-80 to Ser-89,H0050: 2, S0038: 2,Leu-130 to Arg-135,L0777: 2, H0583: 1,Leu-142 to Ser-150,H0650: 1, H0656: 1,Ser-206 to Leu-217,H0255: 1, H0125: 1,Arg-234 to Trp-240.H0192: 1, H0676: 1,H0438: 1, S0049: 1,H0038: 1, H0529: 1,H0690: 1, L0439: 1,H0677: 1 and H0506: 1.949137591 25-9241189Cys-7 to Ala-24,Asn-30 to Asn-42,Ser-80 to Ser-89,Leu-130 to Arg-135,Leu-142 to Ser-150,Tyr-161 to Arg-166.306HAGBX32951351316 3-509914Gly-14 to Gly-32,AR061: 4, AR089: 4108730,Pro-60 to Ala-70,L0439: 4, L0418: 1,147781,Thr-145 to Gly-153,S0010: 1, L0455: 1,172471,Ser-164 to Leu-169.S0028: 1 and L0741: 1.186580,264800,266600,278760,600760,600760,600761,600761,602066956281592473-1381190Phe-4 to Gly-12.307HWMIB811092933317 3-572915Ile-94 to Asp-99,AR061: 1, AR089: 1Asp-118 to Pro-123,L0748: 2, H0171: 1,Glu-131 to Ile-140,S0134: 1, S0354: 1,Tyr-143 to Asp-152,S0358: 1, H0014: 1,Glu-169 to Lys-179.H0083: 1, H0510: 1,L0764: 1, L0803: 1,L0789: 1, H0593: 1,H0659: 1, H0539: 1,H0555: 1, L0751: 1,L0758: 1, L0759: 1 andL0595: 1.9553365931491-922 1191Ile-94 to Asp-99,Asp-118 to Pro-123,Glu-131 to Ile-140,Tyr-143 to Asp-152,Glu-169 to Lys-179.308HCEMU861156430318 1-795916Ser-3 to Lys-8,AR061: 3, AR089: 2His-29 to Lys-38,L0439: 21, L0438: 12,Pro-201 to Thr-206.L0769: 9, T0010: 6,H0052: 5, L0776: 4,L0805: 3, S0126: 3,L0741: 3, L0589: 3,H0261: 2, T0006: 2,L0455: 2, L0659: 2,H0519: 2, L0742: 2,L0748: 2, L0751: 2,L0592: 2, S0276: 2,L0583: 1, L0418: 1,S0298: 1, S0001: 1,S0400: 1, L0394: 1,S0222: 1, H0455: 1,H0331: 1, H0156: 1,T0082: 1, N0006: 1,S6028: 1, H0644: 1,S0036: 1, H0100: 1,L0351: 1, S0352: 1,L0598: 1, L0369: 1,L0638: 1, L0666: 1,H0520: 1, L0777: 1,L0752: 1, L0753: 1,L0758: 1, L0593: 1 andL0366: 1.9568645942520-17351192His-26 to Lys-35,Pro-198 to Thr-203.309HRDAF831153911319368-3 917AR051: 23, AR050:20, AR054: 11, AR061:9, AR089: 5S0250: 8, S0126: 8,H0251: 3, H0545: 2,H0252: 2, L0794: 2,L0565: 2, L0744: 2,L0757: 2, S0040: 1,S0212: 1, S0418: 1,S0360: 1, H0549: 1,H0024: 1, L0053: 1,H0124: 1, S0208: 1,S0011: 1 and S0276: 1.957143595 36-3441193Asn-66 to Val-71,Glu-82 to Thr-91.963999596230-3 1194Ser-42 to Arg-50,Gln-66 to Val-73.310HUVGZ881204719320 503-1288918Pro-14 to His-20,AR089: 2, AR061: 2Gln-82 to Asp-92,L0789: 4, L0758: 4,Ser-161 to Phe-179,H0657: 3, H0052: 3,Cys-190 to Ser-200,L0438: 3, L0744: 3,Gln-212 to Ala-217,L0779: 3, L0005: 2,Glu-235 to Ser-243.H0581: 2, H0194: 2,H0046: 2, H0038: 2,L0800: 2, L0659: 2,H0521: 2, L0743: 2,L0439: 2, H0556: 1,S0282: 1, S0358: 1,H0619: 1, H0586: 1,H0618: 1, H0231: 1,S0362: 1, H0622: 1,T0006: 1, H0616: 1,H0413: 1, H0623: 1,L0351: 1, S0150: 1,L0769: 1, L0372: 1,L0662: 1, L0794: 1,L0775: 1, L0651: 1,L0527: 1, L0657: 1,L0666: 1, H0547: 1,H0690: 1, H0658: 1,H0672: 1, H0539: 1,S0378: 1, H0555: 1,L0754: 1, L0747: 1,L0780: 1, L0596: 1,S0192: 1, H0542: 1 andH0423: 1.959020597 83-4391195Asn-89 to Asn-95.311HSCKS551197921321 3-1646919Asp-1 to Gly-12,AR089: 13, AR061: 5Ala-24 to Gln-29,H0617: 6, H0556: 4,Ala-43 to Asn-61,H0305: 3, S0007: 3,Ala-68 to Gly-81,H0618: 3, H0521: 3,Pro-84 to Gln-99,L0439: 3, H0672: 2,Glu-105 to Gln-110,L0754: 2, L0600: 2,Ala-118 to Asp-123,S0442: 1, S0354: 1,Arg-170 to Leu-175,S0358: 1, S0045: 1,Pro-296 to Thr-306,S0046: 1, S0222: 1,Asn-311 to Gln-320,H0438: 1, H0587: 1,Arg-327 to Ala-335,H0599: 1, H0036: 1,Asp-382 to Gly-389,H0597: 1, H0530: 1,Ala-441 to Pro-451,L0118: 1, H0570: 1,Val-464 to Cys-491,H0023: 1, S0250: 1,Ser-495 to Gly-504,H0039: 1, H0181: 1,Asp-509 to Trp-516,H0674: 1, S0036: 1,Gly-518 to Pro-527.L0351: 1, T0041: 1,H0494: 1, H0509: 1,L0769: 1, L0761: 1,L0764: 1, L0768: 1,L0806: 1, H0519: 1,H0593: 1, H0670: 1,H0660: 1, S3014: 1,L0741: 1, L0779: 1 andH0667: 1.961074598 703-17041196Pro-72 to Thr-82,Asn-87 to Gln-96,Arg-103 to Ala-111,Asp-158 to Gly-165.312HOEET48963290322 2-1030920Ala-8 to Gly-14,AR061: 9, AR089: 5Gly-32 to Arg-48,S0356: 17, S0212: 6,Ala-58 to Asn-66,L0747: 6, S0360: 5,Glu-82 to Gln-92,H0486: 5, S0418: 3,Arg-101 to Gly-110,H0551: 3, S0040: 2,Thr-124 to Asp-131,S0354: 2, H0599: 2,Trp-137 to Gly-146,H0544: 2, H0617: 2,Leu-153 to His-160,H0413: 2, S0210: 2,Glu-171 to Lys-177,L0794: 2, S0126: 2,Asp-191 to Ser-196,S0037: 2, S0027: 2,Glu-225 to Gly-233,L0743: 2, H0665: 2,Glu-248 to Glu-253,S0192: 2, S0196: 2,Thr-259 to Trp-265,S0116: 1, H0662: 1,Arg-268 to Asp-277,S0420: 1, H0619: 1,Glu-303 to Arg-311,H0550: 1, H0013: 1,Ala-329 to Leu-343.H0618: 1, H0253: 1,H0251: 1, H0546: 1,H0545: 1, H0086: 1,H0123: 1, H0024: 1,H0286: 1, H0252: 1,H0628: 1, S0294: 1,L0372: 1, L0646: 1,L0773: 1, L0806: 1,L0654: 1, L0790: 1,L0565: 1, H0689: 1,H0670: 1, H0660: 1,S0028: 1, S0032: 1,L0751: 1, L0754: 1,L0749: 1, L0777: 1,L0780: 1, L0595: 1,H0668: 1, H0667: 1,S0276: 1, S0424: 1 andH0352: 1.313HBODE5111931493232704-1739921Phe-2 to Asp-13,AR089: 4, AR061: 1Ser-47 to Gly-52,L0751: 4, H0052: 3,Arg-161 to Asp-167,S0024: 3, S0364: 3,Leu-256 to Leu-261,L0438: 3, L0439: 3,Asp-288 to Asn-296.H0657: 2, L0415: 2,H0438: 2, H0156: 2,H0373: 2, L0455: 2,H0529: 2, L0664: 2,H0144: 2, L0749: 2,L0592: 2, H0422: 2,L0002: 1, H0583: 1,H0656: 1, S0045: 1,S0046: 1, L0717: 1,H0261: 1, H0455: 1,H0013: 1, H0575: 1,T0082: 1, S0665: 1,S0346: 1, H0581: 1,H0251: 1, H0046: 1,H0009: 1, H0050: 1,H0014: 1, T0010: 1,S0003: 1, S0214: 1,S0366: 1, H0316: 1,H0598: 1, L0351: 1,S0150: 1, L0643: 1,L0764: 1, L0662: 1,L0794: 1, L0805: 1,L0653: 1, L0659: 1,L0666: 1, L0665: 1,H0539: 1, H0521: 1,S0146: 1, H0436: 1,H0478: 1, H0345: 1,L0745: 1, L0758: 1,L0588: 1, L0366: 1,S0026: 1, H0667: 1,S0194: 1, H0542: 1,H0677: 1 and S0446: 1.964235599 2-20531197Gln-12 to His-20,Val-34 to Tyr-39,Asn-54 to Asn-59,Asp-105 to Gly-110,Gly-247 to Lys-256,Gln-314 to Gly-320,Arg-359 to Ser-366,Arg-420 to Gly-428,Ala-558 to Tyr-563,Leu-574 to Pro-579,Arg-592 to Phe-597,Ala-621 to Arg-630,Pro-636 to His-641.314HHFCK099653043242692-389 922Tyr-47 to Glu-58,AR089: 3, AR061: 2Lys-70 to Gly-77,L0666: 8, L0439: 6,Pro-121 to Leu-126,H0253: 5, H0046: 4,Leu-150 to Leu-158,L0769: 4, H0295: 3,Asn-166 to Glu-171,H0255: 3, L0747: 3,Arg-417 to Ser-425,L0756: 3, L0779: 3,Phe-465 to Cys-473,H0657: 2, H0618: 2,Ser-485 to Asn-492,H0318: 2, H0622: 2,Ser-497 to Ala-504,H0068: 2, L0667: 2,Gln-531 to Trp-537,L0772: 2, L0776: 2,Asp-557 to Glu-562.L0663: 2, H0520: 2,H0593: 2, H0670: 2,H0521: 2, L0750: 2,L0759: 2, L0593: 2,L0601: 2, S0116: 1,H0341: 1, S0212: 1,H0306: 1, H0402: 1,L0617: 1, S0358: 1,H0609: 1, H0592: 1,H0333: 1, T0040: 1,H0013: 1, H0635: 1,H0575: 1, H0036: 1,H0581: 1, H0123: 1,H0050: 1, H0012: 1,H0071: 1, T0010: 1,H0687: 1, H0290: 1,H0617: 1, H0606: 1,H0038: 1, H0487: 1,H0494: 1, H0334: 1,S0150: 1, H0647: 1,S0142: 1, L0640: 1,L0639: 1, L0637: 1,L0641: 1, L0768: 1,L0649: 1, L0514: 1,L0659: 1, L0783: 1,L0788: 1, L0664: 1,L0665: 1, L0438: 1,H0547: 1, H0435: 1,H0522: 1, H0696: 1,S0404: 1, H0478: 1,L0742: 1, L0740: 1,L0749: 1, L0758: 1,S0434: 1, S0194: 1,H0422: 1 and H0506: 1.315HCOOZ111110364325 89-943923Asp-43 to Glu-48.AR089: 15, AR061: 5H0662: 2, H0670: 1,L0756: 1 and L0759: 1.965306600 89-5921198Asp-43 to Glu-48.316HDPPO351119032326 72-1202924Lys-7 to Gly-69,AR089: 1, AR061: 0Lys-82 to Lys-88,H0521: 15, H0638: 5,Ser-94 to Asp-112,H0580: 5, H0271: 5,Ala-126 to Asp-131,H0641: 5, H0560: 4,Tyr-134 to Ser-140,H0090: 3, H0591: 3,Ser-147 to Phe-156,L0766: 3, H0542: 3,Asp-159 to Ser-165,H0543: 3, H0586: 2,Thr-176 to Asp-186,H0497: 2, H0581: 2,Glu-230 to Leu-250,L0655: 2, H0518: 2,Glu-291 to Arg-298,H0522: 2, L0754: 2,Gln-313 to Glu-320,L0747: 2, H0657: 1,Asn-331 to Gly-343,H0393: 1, H0431: 1,Ser-348 to Leu-363.H0250: 1, H0635: 1,L0021: 1, H0014: 1,H0179: 1, H0416: 1,H0488: 1, L0475: 1,H0359: 1, H0625: 1,S0426: 1, L0598: 1,L0667: 1, L0803: 1,L0804: 1, L0775: 1,L0651: 1, L0659: 1,L0792: 1, L0663: 1,S0428: 1, H0672: 1,H0555: 1, H0436: 1,L0779: 1, H0445: 1 andS0424: 1.966248601 72-12021199Lys-7 to Gly-69,Lys-82 to Lys-88,Ser-94 to Asp-112,Ala-126 to Asp-131,Tyr-134 to Ser-140,Ser-147 to Phe-156,Asp-159 to Ser-165,Thr-176 to Asp-186,Glu-230 to Leu-250,Glu-291 to Arg-298,Gln-313 to Glu-320,Asn-331 to Gly-343,Ser-348 to Leu-363.317HLWDZ531157542327 2-1126925AR089: 5, AR061: 2L0439: 13, L0752: 4,L0015: 3, H0144: 2,L0438: 2, L0742: 2,L0747: 2, L0758: 2,H0556: 1, L0785: 1,S0001: 1, H0664: 1,H0580: 1, H0486: 1,T0060: 1, H0253: 1,S0010: 1, H0009: 1,H0178: 1, H0564: 1,L0471: 1, S0051: 1,T0010: 1, H0553: 1,H0412: 1, L0370: 1,L0763: 1, L0768: 1,L0794: 1, L0806: 1,L0776: 1, L0657: 1,L0666: 1, H0520: 1,S0126: 1, H0682: 1,H0659: 1, H0187: 1,L0749: 1, L0755: 1 andH0445: 1.968602602 2-11261200318HEOPL361197910328100-501926Gly-11 to Thr-16,AR089: 18, AR061: 5Ser-35 to Ser-56,L0740: 11, L0439: 9,Thr-58 to Ser-73,L0748: 8, H0616: 5,Tyr-85 to Asp-91,L0666: 5, L0601: 5,Glu-100 to Glu-109.S0444: 4, L0776: 4,L0659: 4, L0744: 4,L0747: 4, L0749: 4,L0755: 4, H0457: 3,L0774: 3, L0750: 3,H0624: 2, T0002: 2,S0116: 2, S0358: 2,H0550: 2, T0040: 2,H0013: 2, H0599: 2,H0050: 2, H0673: 2,H0038: 2, H0040: 2,H0494: 2, L0770: 2,L0662: 2, L0364: 2,L0375: 2, L0809: 2,L0438: 2, H0547: 2,L0754: 2, L0756: 2,L0752: 2, L0731: 2,L0758: 2, L0485: 2,S0040: 1, H0583: 1,H0650: 1, H0657: 1,H0341: 1, H0663: 1,H0580: 1, H0619: 1,L0717: 1, H0574: 1,H0052: 1, H0263: 1,H0009: 1, H0172: 1,H0024: 1, T0010: 1,H0510: 1, H0644: 1,S0036: 1, H0551: 1,H0264: 1, H0488: 1,H0056: 1, H0100: 1,L0564: 1, T0041: 1,H0652: 1, S0344: 1,S0002: 1, L0763: 1,L0638: 1, L0761: 1,L0372: 1, L0643: 1,L0764: 1, L0768: 1,L0381: 1, L0775: 1,L0526: 1, L0782: 1,L0663: 1, L0665: 1,H0703: 1, H0520: 1,H0435: 1, H0521: 1,S0044: 1, L0751: 1,L0757: 1, L0759: 1,H0445: 1, L0584: 1,L0608: 1 and H0506: 1.968826603 85-4861201Gly-11 to Thr-16,Ser-35 to Ser-56,Thr-58 to Ser-73,Tyr-85 to Asp-91,Glu-100 to Glu-109.319HMCFS021152252329 3-734927Leu-42 to Gln-49,AR061: 2, AR089: 0Gln-59 to Thr-65,L0766: 10, L0794: 7,Pro-119 to Lys-128,L0758: 7, L0805: 6,Asn-134 to Phe-140,L0751: 4, L0754: 4,Arg-150 to Phe-155,L0803: 3, L0483: 2,Asp-205 to Gly-212.L0764: 2, L0659: 2,L0809: 2, L0790: 2,L0666: 2, L0755: 2,L0599: 2, H0170: 1,H0294: 1, H0583: 1,H0656: 1, S0282: 1,H0255: 1, S0420: 1,H0618: 1, H0688: 1,L0055: 1, S0344: 1,H0529: 1, L0761: 1,L0643: 1, L0645: 1,L0804: 1, L0806: 1,L0653: 1, L0776: 1,L0629: 1, L0636: 1,L0788: 1, L0789: 1,L0791: 1, L0665: 1,S0428: 1, H0702: 1,L0438: 1, S0330: 1,H0539: 1, H0478: 1,L0749: 1, L0750: 1,L0779: 1, L0731: 1,L0757: 1, H0665: 1 andH0423: 1.969326604 2-4961202Arg-3 to Lys-20,Phe-22 to Ser-28,Leu-50 to Gln-57.320HDPSR151194752330 178-1167928Pro-26 to Leu-34,AR061: 2, AR089: 2His-42 to Asn-51,L0759: 12, L0439: 11,Phe-154 to Pro-162,L0766: 7, L0775: 5,His-237 to Asp-246,H0521: 5, L0755: 5,Pro-263 to Lys-268,L0748: 4, L0756: 4,Lys-277 to Asp-282,L0777: 4, L0731: 4,Pro-285 to Leu-295,L0581: 4, L0619: 3,Pro-305 to Asp-312.L0666: 3, L0779: 3,L0757: 3, L0588: 3,S0418: 2, L0618: 2,H0580: 2, L0055: 2,L0769: 2, L0773: 2,L0774: 2, L0791: 2,L0747: 2, L0750: 2,H0265: 1, H0663: 1,S0356: 1, H0208: 1,H0370: 1, H0108: 1,H0575: 1, H0618: 1,H0544: 1, H0545: 1,S0050: 1, H0510: 1,H0286: 1, H0031: 1,H0644: 1, H0068: 1,H0135: 1, L0564: 1,H0494: 1, L0475: 1,H0396: 1, S0144: 1,S0002: 1, S0426: 1,L0763: 1, L0761: 1,L0642: 1, L0764: 1,L0662: 1, L0768: 1,L0806: 1, L0661: 1,L0659: 1, L0367: 1,L0663: 1, H0519: 1,H0435: 1, H0658: 1,S3014: 1, L0751: 1,L0749: 1, L0603: 1,H0665: 1 and H0542: 1.969666605168-7851203Pro-26 to Leu-34,His-42 to Asn-51.321HNTAV7812172313311104-697 929Pro-1 to Lys-6.AR054: 10, AR089: 2,AR061: 1, AR051: 1,AR050: 1H0305: 1, H0580: 1,H0428: 1, L0803: 1,L0809: 1 and H0519: 1.971315606 3-2661204Glu-52 to Leu-58,Arg-63 to Lys-71,Arg-83 to Val-88.322HFKDR141145842332 3-1319930Ala-2 to Pro-9,AR061: 3, AR089: 2Val-22 to Gly-28.L0792: 2, H0012: 1,H0100: 1, L0663: 1,L0756: 1 and L0780: 1.974255607 15-17331205Ala-5 to Gly-18.323HDPBI30974711333 182-1312931Asp-1 to Asn-10.AR051: 3, AR050: 1,AR089: 1, AR061: 0H0521: 3, H0656: 2,H0635: 2, H0549: 1,H0050: 1, H0413: 1,H0641: 1, L0387: 1,H0436: 1 and H0423: 1.324HODFF881094875334 14-544932His-8 to Gly-18,AR054: 34, AR051:Glu-150 to Leu-167.29, AR050: 23, AR089:4, AR061: 4H0615: 1974911608 14-5441206His-8 to Gly-18,Glu-150 to Leu-167.

[0040] The first column in Table 1A provides the gene number in the application corresponding to the clone identifier. The second column in Table 1A provides a unique “Clone ID NO:Z” for a cDNA clone related to each contig sequence disclosed in Table 1A. This clone ID references the cDNA clone which contains at least the 5′ most sequence of the assembled contig and at least a portion of SEQ ID NO:X was determined by directly sequencing the referenced clone. The reference clone may have more sequence than described in the sequence listing or the clone may have less. In the vast majority of cases, however, the clone is believed to encode a full-length polypeptide. In the case where a clone is not full-length, a full-length cDNA can be obtained by methods described elsewhere herein.

[0041] The third column in Table 1A provides a unique “Contig ID” identification for each contig sequence. The fourth column provides the “SEQ ID NO:” identifier for each of the contig polynucleotide sequences disclosed in Table 1A. The fifth column, “ORF (From-To)”, provides the location (i.e., nucleotide position numbers) within the polynucleotide sequence “SEQ ID NO:X” that delineate the preferred open reading frame (ORF) shown in the sequence listing and referenced in Table 1A, column 6, as SEQ ID NO:Y. Where the nucleotide position number “To” is lower than the nucleotide position number “From”, the preferred ORF is the reverse complement of the referenced polynucleotide sequence.

[0042] The sixth column in Table 1A provides the corresponding SEQ ID NO:Y for the polypeptide sequence encoded by the preferred ORF delineated in column 5. In one embodiment, the invention provides an amino acid sequence comprising, or alternatively consisting of, a polypeptide encoded by the portion of SEQ ID NO:X delineated by “ORF (From-To)”. Also provided are polynucleotides encoding such amino acid sequences and the complementary strand thereto.

[0043] Column 7 in Table 1A lists residues comprising epitopes contained in the polypeptides encoded by the preferred ORF (SEQ ID NO:Y), as predicted using the algorithm of Jameson and Wolf, (1988) Comp. Appl. Biosci. 4:181-186. The Jameson-Wolf antigenic analysis was performed using the computer program PROTEAN (Version 3.11 for the Power MacIntosh, DNASTAR, Inc., 1228 South Park Street Madison, Wis.). In specific embodiments, polypeptides of the invention comprise, or alternatively consist of, at least one, two, three, four, five or more of the predicted epitopes as described in Table 1A. It will be appreciated that depending on the analytical criteria used to predict antigenic determinants, the exact address of the determinant may vary slightly.

[0044] Column 8 in Table 1A provides an expression profile and library code: count for each of the contig sequences (SEQ ID NO:X) disclosed in Table 1A, which can routinely be combined with the information provided in Table 4 and used to determine the tissues, cells, and/or cell line libraries which predominantly express the polynucleotides of the invention. The first number in column 8 (preceding the colon), represents the tissue/cell source identifier code corresponding to the code and description provided in Table 4. For those identifier codes in which the first two letters are not “AR”, the second number in column 8 (following the colon) represents the number of times a sequence corresponding to the reference polynucleotide sequence was identified in the tissue/cell source. Those tissue/cell source identifier codes in which the first two letters are “AR” designate information generated using DNA array technology. Utilizing this technology, cDNAs were amplified by PCR and then transferred, in duplicate, onto the array. Gene expression was assayed through hybridization of first strand cDNA probes to the DNA array. cDNA probes were generated from total RNA extracted from a variety of different tissues and cell lines. Probe synthesis was performed in the presence of 33P dCTP, using oligo(dT) to prime reverse transcription. After hybridization, high stringency washing conditions were employed to remove non-specific hybrids from the array. The remaining signal, emanating from each gene target, was measured using a Phosphorimager. Gene expression was reported as Phosphor Stimulating Luminescence (PSL) which reflects the level of phosphor signal generated from the probe hybridized to each of the gene targets represented on the array. A local background signal subtraction was performed before the total signal generated from each array was used to normalize gene expression between the different hybridizations. The value presented after “[array code]:” represents the mean of the duplicate values, following background subtraction and probe normalization. One of skill in the art could routinely use this information to identify normal and/or diseased tissue(s) which show a predominant expression pattern of the corresponding polynucleotide of the invention or to identify polynucleotides which show predominant and/or specific tissue and/or cell expression.

[0045] Column 9 in Table 1A provides a chromosomal map location for certain polynucleotides of the invention. Chromosomal location was determined by finding exact matches to EST and cDNA sequences contained in the NCBI (National Center for Biotechnology Information) UniGene database. Each sequence in the UniGene database is assigned to a “cluster”; all of the ESTs, cDNAs, and STSs in a cluster are believed to be derived from a single gene. Chromosomal mapping data is often available for one or more sequence(s) in a UniGene cluster; this data (if consistent) is then applied to the cluster as a whole. Thus, it is possible to infer the chromosomal location of a new polynucleotide sequence by determining its identity with a mapped UniGene cluster.

[0046] A modified version of the computer program BLASTN (Altshul et al., J. Mol. Biol. 215:403-410 (1990); and Gish and States, Nat. Genet. 3:266-272 (1993)) was used to search the UniGene database for EST or cDNA sequences that contain exact or near-exact matches to a polynucleotide sequence of the invention (the ‘Query’). A sequence from the UniGene database (the ‘Subject’) was said to be an exact match if it contained a segment of 50 nucleotides in length such that 48 of those nucleotides were in the same order as found in the Query sequence. If all of the matches that met this criteria were in the same UniGene cluster, and mapping data was available for this cluster, it is indicated in Table 1A under the heading “Cytologic Band”. Where a cluster had been further localized to a distinct cytologic band, that band is disclosed; where no banding information was available, but the gene had been localized to a single chromosome, the chromosome is disclosed.

[0047] Once a presumptive chromosomal location was determined for a polynucleotide of the invention, an associated disease locus was identified by comparison with a database of diseases which have been experimentally associated with genetic loci. The database used was the Morbid Map, derived from OMIM™ (supra). If the putative chromosomal location of a polynucleotide of the invention (Query sequence) was associated with a disease in the Morbid Map database, an OMIM reference identification number was noted in column 10, Table 1A, labelled “OMIM Disease Reference(s)”. Table 5 is a key to the OMIM reference identification numbers (column 1), and provides a description of the associated disease in Column 2.

3TABLE 1BClone IDSEQ IDCONTIGBAC ID:SEQ IDEXONNO:ZNO:XID:ANO:BFrom-ToHCEPH7114522739AL3653191207 1-494HCEPH7114522739AL3907151208 1-494HLMDO9543928344AC0206411209 1-591 627-2046HTEAG4954954614AL3907961210 1-1310HTEAG4954954614AL3570451211 1-1310HTEAG4954954614AL3907961212 1-627HTEAG4954954614AL3570451213 1-627HACCH94103847143AL1614581214 1-1140HACCH94103847143AL1614581215 1-905811-6312HFKLX38113880220AL1363831216 1-321288-14541561-16463840-47005482-6798HTDAB17117890384AC0110781217 1-297359-4163247-36536083-6236 9753-1003611128-1123312148-1251412635-1314115604-1646319071-1919019476-2023220321-2063821200-2159421959-2221923120-2336223467-2414324766-2485325725-2614326310-2645527545-3061930708-31169HTLCA95142911655AC0126161218 1-1151HTLCA95142911655AC0126161219 1-284HHEHC53152921783AC0094271220 1-1001854-19423236-34634629-48685054-51815371-54765851-59536104-61496509-66127131-84158429-84928638-87488975-9440 9835-1049010606-1089911149-1128211382-1188112023-1207512172-1231512496-1255112638-1270612827-1299413077-13630HHEHC53152921783AC0094271221 1-428HHEHC53152921783AC0094271222 1-388466-526698-9061023-1922HELHF07196949067AC0736691223 1-597HELHF07196949067AC0236051224 1-583HELHF07196949067AC0742201225 1-362HELHF07196949067AC0742201226 1-105HACAD35199949199AC0073631227 1-983271-34134132-43577030-768211881-1200112874-13485HDTKQ14254886936AL3595421228 1-1401249-4264HDTKQ14254886936AL3595421229 1-499HDTKQ14254886936AL3595421230 1-145HWAGS73257894404AL0968701231 1-185 393-17431951-21182229-22952410-29063043-31073238-35193594-3970HWAGS73257894404AL0968701232 1-10802072-2811HSYBX32270909846AC0040841233 1-105 839-10212069-23022470-28553818-42654371-46104761-48105364-58025930-65177073-78078063-86188636-88759438-953710568-1077410897-1102511718-1232313749-1384913978-1418814474-1455416489-1662416924-1701917239-1745817908-1818519014-1926619356-1945119620-1987319893-2092021092-2124721512-2157921621-2175422001-2283122992-2351823710-2437024426-2459625213-2549325661-2619226588-2743327598-2774228073-2819928359-2865128777-2924929379-2950229646-2979429833-3003330085-3063030702-3266133104-3337433383-3366133808-3387133978-3744937587-3775438296-3843338597-3934340047-4039540462-4074340938-4103941187-4163442504-4268742703-4285042929-43475HSYBX32270909846AC0049511234 1-735 991-1547HSYBX32270909846AC0040841235 1-239HSYBX32270909846AC0040841236 1-283HSYBX32270909846AC0049511237 1-255HSYBX32270909846AC0049511238 1-318HTLJC71284922923AC0095161239 1-2009HTLJC71284922923AC0079571240 1-1747HTLJC71284922923AC0187511241 1-2009HTLJC71284922923AC0234901242 1-2009HTLJC71284922923AC0095161243 1-375HTLJC71284922923AC0095161244 1-494HTLJC71284922923AC0079571245 1-205HTLJC71284922923AC0187511246 1-494HTLJC71284922923AC0234901247 1-375HTLJC71284922923AC0187511248 1-375HWMEV63291931154AC0788161249 1-1574

[0048] Table 1B summarizes additional polynucleotides encompassed by the invention (including cDNA clones related to the sequences (Clone ID NO:Z), contig sequences (contig identifier (Contig ID:) contig nucleotide sequence identifiers (SEQ ID NO:X)), and genomic sequences (SEQ ID NO:B). The first column provides a unique clone identifier, “Clone ID NO:Z”, for a cDNA clone related to each contig sequence. The second column provides the sequence identifier, “SEQ ID NO:X”, for each contig sequence. The third column provides a unique contig identifier, “Contig ID:” for each contig sequence. The fourth column, provides a BAC identifier “BAC ID NO:A” for the BAC clone referenced in the corresponding row of the table. The fifth column provides the nucleotide sequence identifier, “SEQ ID NO:B” for a fragment of the BAC clone identified in column four of the corresponding row of the table. The sixth column, “Exon From-To”, provides the location (i.e., nucleotide position numbers) within the polynucleotide sequence of SEQ ID NO:B which delineate certain polynucleotides of the invention that are also exemplary members of polynucleotide sequences that encode polypeptides of the invention (e.g., polypeptides containing amino acid sequences encoded by the polynucleotide sequences delineated in column six, and fragments and variants thereof).

4TABLE 2SEQScore/Clone IDContigIDAnalysisPFam/NR AccessionPercentNTNTNO: ZID:NO: XMethodPFam/NR DescriptionNumberIdentityFromToHTPAD46503313335HMMERPFAM: Src homologyPF000184.141601861.8domain 3HCWFF88506577336HMMERPFAM: Src homologyPF000184.921401811.8domain 3HSSAX53507509337HMMERPFAM: Src homologyPF000184.362663311.8domain 3HCEPH7152273914HMMERPFAM: Src homologyPF000184.2233621.8domain 3HTEDF74522982338HMMERPFAM: C2 domainPF001686.981892331.8HTTEK47573649339HMMERPFAM: EF handPF0003610.822242891.8blastx.2DJ534K7.2 (novelsp|CAB92087|CAB92087100%11301protein).65%28337852%342410HTOBE75591896340HMMERPFAM: Sushi domainPF0008461.21002732.1.1(SCR repeat)HCFAT05592118341HMMERPFAM: Ion transportPF00520106.11373612.1.1proteinblastx.2potassium channel proteingb|AAA59457.1|67%134427[Homo sapiens]100%1813752%360491HFIAH37615597342HMMERPFAM: C2 domainPF001684.222412911.8HFTDF15657020343HMMERPFAM: Src homologyPF000184.851682031.8domain 3HPFCU80685294344HMMERPFAM: C2 domainPF0016842612961.8HSVAW49689674345HMMERPFAM: Src homologyPF0001836.33771691.8domain 3blastx.2(AF146277) adaptergb|AAD34595.1|AF146277_197%65166protein CMS [Homosapiens]HWHQC94111646323blastx.14(AC004472) P1.11659_3gi|2984587|gb|AAC07985.1|76%581874[Homo sapiens]41%476547HWHQC94715096346HMMERPFAM: C2 domainPF001684.172143001.8blastx.2Pig-o.sp|BAA96254|BAA9625464%1627HRSMD49723025347HMMERPFAM: Src homologyPF000184.761992701.8domain 3HFTDY67115122025blastx.14(AF182316) myoferlingi|6731235|gb|AAF27176.1|94%136852[Homo sapiens]AF182316_192%42133%2019442%63126%201145HFTDY67745221348HMMERPFAM: C2 domainPF0016820.0741441.8blastx.2Myoferlin.sp|AAF27177|AAF2717798%4225100%22429835%213263HYABL89786157349HMMERPFAM: C2 domainPF001686.052703171.8HCUEV29113779127blastx.14(AL110490) predictedgi|5824799|emb|CAB54442.1|53%96344using Genefinder53%387470[Caenorhabditis elegans]64%647HCUEV29816065350HMMERPFAM: EF handPF0003631.871432291.8blastx.2CG10641 PROTEIN.sp|Q9VJ26|Q9VJ2659%8928649%312500HCESP56827671351HMMERPFAM: EF handPF0003611.862403171.8blastx.2HYPOTHETICAL 27.4 KDAsp|Q9UJF6|Q9UJF6100%186452PROTEIN(FRAGMENT).HLQDT35839777352HMMERPFAM: Src homologyPF000183.853424191.8domain 3blastx.2(AK000579) unnameddbj|BAA91269.1|98%252458protein product [Homosapiens]HDPBS6484662430HMMERPFAM: ThioredoxinsPF00085116.871734931.8blastx.2ZK973.11 protein.sp|AAF40013|AAF4001332%182652HTBAB41867287353HMMERPFAM: C2 domainPF001685.32891571.8HTLGE3187024732HMMERPFAM: Sugar (and other)PF0008321.451151.8transportersHWLHK29115227933blastx.14(AF181098)gi|5823558|gb|AAD53186.1|27%474602synaptotagmin IVAF181098_129%198371[Drosophila melanogaster]30%77166HWLHK29876064354HMMERPFAM: C2 domainPF0016816.972224881.8HHEGG20110681634blastx.14(AF084205)gi|3452473|gb|AAC71014.1|84%3971serine/threonine proteinkinase TAO1 [Rattusnorvegicus]HHEGG20894409355HMMERPFAM: Eukaryotic proteinPF00069200.01265981.8kinase domainHDPRU43121703535blastx.14PUTATIVE RASGAP-sp|O43374|O4337497%1632460ACTIVATING-LIKE100%52174PROTEIN.HDPRU43909841356HMMERPFAM: C2 domainPF0016882.11583031.8HE8PK12909884357HMMERPFAM: Src homologyPF0001858.121973611.8domain 3blastx.2(AF136380) SH3P12gb|AAD27647.1|AF136380_182%59367protein [Homo sapiens]HE9HV92122751937blastx.14DIFFERENTIATIONsp|O97902|O9790253%1336ENHANCING FACTOR 1.64%820105037%47756347%29034036%766873HE9HV92911510358HMMERPFAM: Src homologyPF0001855.6786910421.8domain 3blastx.2DIFFERENTIATIONsp|O97902|O9790252%7071048ENHANCING FACTOR 1.44%841837%47556135%713871HOHCE47911566359HMMERPFAM: Eukaryotic proteinPF0006979.422114231.8kinase domainHSDII69917180360HMMERPFAM: Src homologyPF000184.093824291.8domain 3HKAKM10122763940blastx.14NUCLEAR BODYsp|Q9QUQ8|Q9QUQ888%92840ASSOCIATED KINASE2B.HKAKM10918685361HMMERPFAM: Eukaryotic proteinPF0006931.481272.1.1kinase domainHCEPU56920347362HMMERPFAM: C2 domainPF001683.932432931.8blastx.2HYPOTHETICAL 27.3 KDAsp|O60362|O6036282%219659PROTEIN.HUSHB5492805442HMMERPFAM: C2 domainPF0016820.522753551.8HLMDO9592834443HMMERPFAM: 7 transmembranePF0000143.252203691.8receptor (rhodopsinfamily)blastx.2Inflammation-related Gsp|AAF91467|AAF9146751%112375protein-coupled receptor95%375446EX33.HHASQ32928730363HMMERPFAM: CarnitatePF007553172508552.1.1acyltransferaseHARAB87116434045blastx.14neurotransmittergi|914028|gb|AAB32806.1|89%51350transporter rB21a [rats,83%349549brain, Peptide, 616 aa]90%1645[Rattus sp.]HARAB87933441364HMMERPFAM:PF0020979.62685702.1.1Sodium: neurotransmittersymporter familyHTNGF69933614365HMMERPFAM: C2 domainPF001683.945886591.8HMSJL96115478847blastx.14(AF104413) large tumorgi|4324434|gb|AAD16882.1|80%11245suppressor 1 [Homo86%12821371sapiens]41%10851171HMSJL96934483366HMMERPFAM: Eukaryotic proteinPF0006926.491993631.8kinase domainHDTBT06935404367HMMERPFAM: ActinPF00022110.85149032.1.1blastx.2actin-related protein - fruitpir|S44028|S4402847%52876fly (Drosophilamelanogaster)HTTIE47116536349blastx.14ZK550.2 [Caenorhabditisgi|3881745|emb|CAB05312.1|43%188382elegans]30%39854434%559627HTTIE47941834368HMMERPFAM: Sugar (and other)PF0008332.122955221.8transportersblastx.2predicted usingemb|CAB01157.1|38%151537Genefinder; Similarity toworm multidrug resistanceproteins [Caenorhabditiselegans]HHFBP4794666850HMMERPFAM: Sugar (and other)PF0008325.746963701.8transportersblastx.2(AL050350) dJ261K5.1emb|CAB56524.1|98%482955(novel organic cation95%135461transporter (BAC ORF100%38139RG331P03)) [Homo77%451516sapiens]HCCCC81108355351blastx.14(AL022605) putativegi|3080435|emb|CAA18752.1|50%125304protein [Arabidopsis68%428514thaliana]56%30239729%58868938%51459150%334369HCCCC81949062369HMMERPFAM:PF00202178.371878161.8Aminotransferases class-III pyridoxal-phosphateblastx.2hypothetical proteinpir|T25848|T2584846%190879T01B11.2 -Caenorhabditis elegansHPJEV71949153370HMMERPFAM: von WillebrandPF0009247.9886613751.8factor type A domainblastx.2CDNA FLJ10601 FIS,sp|BAA91707|BAA9170753%9741534CLONE NT2RP2005000.HTEIL07113612153blastx.14caltractin [Giardiagi|1399341|gb|AAB05594.1|28%9611128intestinalis]28%646792HTEIL07953803371HMMERPFAM: EF handPF0003611.271922631.8blastx.2Hypothetical 41.3 kDasp|CAB91065|CAB9106579%57392protein.HTEAG4995461454HMMERPFAM: Src homologyPF000184.513122381.8domain 3HSLCF9663767055HMMERPFAM: Sugar (and other)PF0008310.784155761.8transportersblastx.2(AE000352) putativegb|AAC75728.1|94%4151179transport protein100%297413[Escherichia coli]70%1101121120%40960026%10211179HSLCF96954777372HMMERPFAM: Sugar (and other)PF0008330.03129610151.8transportersblastx.2(AE000352) putativegb|AAC75728.1|96%3031127transport protein91%11471215[Escherichia coli]HNHCI3286167356HMMERPFAM: 7 transmembranePF00001133.171955451.8receptor (rhodopsinfamily)blastx.2G protein-coupledsp|AAF27279|AAF27279100%189551receptor 57.100%112186100%56112HNHCI32956105373HMMERPFAM: 7 transmembranePF00001133.179516011.8receptor (rhodopsinfamily)blastx.2(AF112461) G protein-gb|AAF27279.1|AF112461_1100%555917coupled receptor 57100%478552[Homo sapiens]100%422478HPMFL08959569374HMMERPFAM: Src homologyPF000184.972092381.8domain 3HTXRA1395962258HMMERPFAM: C2 domainPF0016851.35408092.1.1blastx.2GRANUPHILIN-A.sp|Q9R0Q1|Q9R0Q142%4291091HCE3H71119789859blastx.14SEIZURE-RELATEDsp|Q62223|Q6222389%413961PROTEIN 6 TYPE 295%12413PRECURSOR.27%12626634%15026356%36341039%36544841%21626643%27332036%38344846%41044857%410451HCE3H71961681375HMMERPFAM: Sushi domainPE0008479.23174962.1.1(SCR repeat)blastx.2seizure-related proteinpir|I52657|I5265783%5685SEZ-6 precursor - mouse64%56595730%9849648%9291000HUTSF11966029376HMMERPFAM: Eukaryotic proteinPF0006927.7431041.8kinase domainHTEGI48102123561blastx.14(AF074606) histonegi|4091980|gb|AAC99368.1|87%1525acetyltransferase [Homosapiens]HTEGI48530595377HMMERPFAM: Zinc finger,PF0153043.83444362.1.1C2HC typeHSFAM09573345378HMMERPFAM: Src homologyPF000185.331952181.8domain 3HNFHK77118228663blastx.14succinic semialdehydegi|147901|gb|AAC36831.1|100%180647dehydrogenase97%653898[Escherichia coli]HNFHK77576186379HMMERPFAM: AldehydePF0017130.22313262.1.1dehydrogenase familyHFXDO83101260264blastx.14No definition line foundgi|912476|gb|AAB18565.1|85%93596[Escherichia coli]HFXDO83578847380HMMERPFAM: AldehydePF0017146.2582072.1.1dehydrogenase familyHSDIW73110440665blastx.14(AE000506) putativegi|2367379|gb|AAC77312.1|100%11068transport protein, cryptic,orf, joins former yjiZ andyjjL [Escherichia coli]HSDIW73587311381HMMERPFAM: Sugar (and other)PF0008319.7155381.8transportersblastx.2(AE000506) putativegb|AAC77312.1|100%81075transport protein, cryptic,orf, joins former yjiZ andyjjL [Escherichia coli]HSDIW73954821382HMMERPFAM: Sugar (and other)PF0008311.23159510861.8transportersblastx.2(AE000506) putativegb|AAC77312.1|100%31049transport protein, cryptic,orf, joins former yjiZ andyjjL [Escherichia coli]HFVGD23676214383HMMERPFAM: CarnitatePF00755168.424452.1.1acyltransferaseHMSBZ24108236767blastx.14contains similarity to C2gi|1825586|gb|AAB42222.1|39%8241domains [Caenorhabditis30%548646elegans]43%55159826%617685HMSBZ24678707384HMMERPFAM: C2 domainPF0016835.4681811.8blastx.2CG15078 PROTEIN.sp|Q9V8M4|Q9V8M450%8289HWHHB69690442385HMMERPFAM: Src homologyPF0001831.65912551.8domain 3blastx.2(AF178432) SH3 proteingb|AAF35985.1|AF178432_170%91315[Homo sapiens]100%303329HFXLC69692773386HMMERPFAM: C2 domainPF0016817.77842001.8HBXBW40115676570blastx.14Protein Kinase [Rattusgi|2077934|dbj|BAA19880.1|93%214546norvegicus]78%546587HBXBW40706115387HMMERPFAM: Eukaryotic proteinPF0006934.012804231.8kinase domainHCE1L51114049871blastx.14carnitinegi|294521|gb|AAA40876.1|51%3233palmitoyltransferase I59%288434[Rattus norvegicus]HCE1L51715899388HMMERPFAM: CarnitatePF00755102.9334342.1.1acyltransferaseHRADM45717358389HMMERPFAM: Eukaryotic proteinPF0006923.7141241.8kinase domainblastx.2(AJ271722) putativeemb|CAB71146.1|98%2469serine/threonine proteinkinase MAK-V [Homosapiens]HTEFO45115391873blastx.14Munc13-3 [Rattusgi|1763306|gb|AAB39720.1|94%64597norvegicus]HTEFO45723446390HMMERPFAM: C2 domainPF0016830.641113831.8HOHBN82115227174blastx.14(AF182316) myoferlingi|6731235|gb|AAF27176.1|94%921408[Homo sapiens]AF182316_187%1418146533%1259136647%1397145926%12591315HOHBN82724322391HMMERPFAM: C2 domainPF0016849.95963471.8blastx.2Myoferlin.sp|AAF27176|AAF2717680%9362358%571606HWHGF52121702675blastx.14GUANINEsp|Q64096|DBS_MOUSE73%14271NUCLEOTIDE79%338439EXCHANGE FACTOR27%873DBS (DBLS BIGSISTER) (MCF2TRANSFORMINGSEQUENCE-LIKEPROTEIN).HWHGF52726102392HMMERPFAM: Src homologyPF000185.013253871.8domain 3blastx.2Dbs = Dbl guaninegb|AAB33461.1|74%3203nucleotide exchange72%319417factor homolog [mice,73%20325932D 1HBKDI30122386176blastx.14BA243J16.3 (similar tosp|CAC10006|CAC10006100%343672MYLK (myosin, light91%23307polypeptide 1HBKDI30729048393HMMERPFAM: Eukaryotic proteinPF0006942.2312131.8kinase domainHSQFR54118514377blastx.14vacuolar protein sortinggi|1477468|gb|AAC52985.1|86%52882homolog r-vps33a [Rattus94%11341640norvegicus]96%862113792%1640183731%13321466HSQFR54730964394HMMERPFAM: Sec1 familyPF0099566.122592.1.1blastx.2VACUOLAR PROTEINsp|Q63615|Q6361586%2310SORTING HOMOLOGR-VPS33A.HAGBA56110259378blastx.14(AF033655) Pftaire-1gi|2645810|gb|AAB87504.1|95%187735[Mus musculus]58%115379%132203HAGBA56732597395HMMERPFAM: Eukaryotic proteinPF0006964.91395162.1.1kinase domainHHSAE29122085179blastx.14probable sugar transportpir|S47743|S47743100%572946protein - Escherichia coli95%10211083HHSAE29743166396HMMERPFAM: Sugar (and other)PF0008344.67292501.8transportersHMSHO6474658280HMMERPFAM: Src homologyPF0001811.083164051.8domain 3blastx.2(AF030131) Plenty ofgb|AAC40070.1|47%1411SH3s; POSH [Musmusculus]HFPBW22750631397HMMERPFAM: C2 domainPF00168193234301.8HTLBH67751985398HMMERPFAM: Src homologyPF0001837.78161621.8domain 3HNTMH70114352383blastx.14similar to protein kinasesgi|1072163|gb|AAA81690.1|50%425667[Caenorhabditis elegans]40%10122029%34444547%62124HNTMH70757184399HMMERPFAM: Eukaryotic proteinPF0006994.551765771.8kinase domainHCETC59761881400HMMERPFAM: Sec1 familyPF0099560.41173052.1.1blastx.2Vacuolar protein sortingsp|AAF91174|AAF9117478%12336833B.100%6712364%311412HE8UX76116122385blastx.14(AB026803)gi|6136784|dbj|BAA85775.1|98%4371087synaptotagmin VI [Mus91%98454musculus]97%1129138336%69284134%11291242HE8UX76767871401HMMERPFAM: C2 domainPF001687.235926361.8HTLEN77113612486blastx.14(AF081671) VU91Dgi|3800851|gb|AAC68892.1|43%240392calmodulin [syntheticconstruct]HTLEN77772363402HMMERPFAM: EF handPF0003626.932943801.8blastx.2CALTRACTINsp|P53441|CATR_NAEGR30%111374(CENTRIN).HBGDI80112469587blastx.14(AL078627) actin-likegi|5051483|emb|CAB44762.1|46%5328protein; (2 actin domains)[Schizosaccharomycespombe]HBGDI80781600403HMMERPFAM: ActinPF0002233.6523182.1.1blastx.2CG7940 PROTEIN.sp|Q9VEC3|Q9VEC337%13336HELHB88811935404HMMERPFAM: EF handPF0003612.82473301.8blastx.2INTERSECTIN LONGsp|Q9UNK2|Q9UNK284%139567ISOFORM.46%145375100%7814630%361495HTEMV66115226189blastx.14contains EGF-like repeats;gi|495684|gb|AAA50735.1|56%265621highly similar to ZC84.1; 144%61195HTEMV66813038405HMMERPFAM: Eukaryotic proteinPF0006927.81543152.1.1kinase domainHMTAJ73813296406HMMERPFAM: Eukaryotic proteinPF0006921.3441141.8kinase domainHE9TD3181584591HMMERPFAM: EF handPF0003617.535196051.8blastx.2Intersectin 2 shortsp|AAF59904|AAF5990481%3626isoform.40%378626HGBDG55114136392blastx.14(AF004161) peroxisomalgi|2352427|gb|62%209529Ca-dependent soluteAAB69156.1|carrier [Oryctolaguscuniculus]HGBDG55815858407HMMERPFAM: EF handPF0003617.243023851.8blastx.2Calcium-bindingsp|AAF28888|AAF2888861%209388transporter (Fragment).64%38153037%284388HOUHL51112591493blastx.14(AJ249457) centrin,gi|5869981|emb|CAB55607.1|29%230487putative [Trichomonas25%173244vaginalis]HOUHL51815891408HMMERPFAM: EF handPF0003629.34295062.1.1blastx.2CENTRIN, PUTATIVE.sp|Q9U5I9|Q9U5I926%48506HEOPP67102011994blastx.14guanylate cyclasegi|623417|gb|AAA60716.1|31%169474activating protein [Musmusculus]HEOPP67827630409HMMERPFAM: EF handPF00036352333162.1.1blastx.2NADPH thyroid oxidasesp|AAF73922|AAF7392298%564332.HKAOV71116542395blastx.14(AF123303) calcium-gi|6841066|gb|AAF28888.1|88%61753binding transporter [HomoAF123303_193%711755sapiens]HKAOV71827679410HMMERPFAM: EF handPF0003650.72203002.1.1blastx.2Calcium-bindingsp|AAF28888|AAF2888888%61753transporter (Fragment).93%711755HDQID90113775296blastx.14(AF132480) Ese2 proteingi|4378889|gb|AAD19748.1|75%11268[Mus musculus]47%472522HDQID90831976411HMMERPFAM: EF handPF0003610.084134961.8blastx.2Intersectin 2.sp|AAF63600|AAF6360087%20669436%272565HFRBN81118255297blastx.14(AE000192) putativegi|1787127|gb|AAC73985.1|99%2163598transport [Escherichiacoli]HFRBN81833061412HMMERPFAM: Sugar (and other)PF0008335.69524201.8transportersblastx.2Hypothetical protein Ydbj|BAA35630.1|100%11116[Escherichia coli]HFRBN81973206413HMMERPFAM: Sugar (and other)PF0008330.84523931.8transportersblastx.2Hypothetical protein Ydbj|BAA35630.1|80%4546[Escherichia coli]HFRBN81973208414blastx.2Hypothetical protein Ydbj|BAA35630.1|97%2352[Escherichia coli]75%35158718%56229HFKJW01118713498blastx.14lactaldehydegi|145222|gb|AAA23427.1|100%3948dehydrogenase[Escherichia coli]HFKJW01836491415HMMERPFAM: AldehydePF00171174964402.1.1dehydrogenase familyblastx.2lactaldehydepir|A38165|A38165100%96440dehydrogenase (EC1.2.1.22) aldA -Escherichia coliHSDFL63836498416HMMERPFAM: AldehydePF00171127.412342.1.1dehydrogenase familyblastx.2RETINALDEHYDE-sp|O94788|DHAS_HUMAN100%1249SPECIFICDEHYDROGENASETYPE 2 (EC 1.2.1.—)(RALDH(II)) (RALDH-2).HDJET90836503100HMMERPFAM: AldehydePF00171150.4663712.1.1dehydrogenase familyHEMFC61836514101HMMERPFAM: AldehydePF00171397.1106422.1.1dehydrogenase familyblastx.2RETINALDEHYDE-sp|O94788|DHAS_HUMAN98%4642SPECIFICDEHYDROGENASETYPE 2 (EC 1.2.1.—)(RALDH(II)) (RALDH-2).HDTBR501174351102blastx.14intermediate chain 1gi|14817526|dbj|BAA09934.1|57%130306[Anthocidaris crassispina]44%327455HDTBR50846630417HMMERPFAM: ThioredoxinsPF0008529.851632971.8blastx.2NM23-H8.sp|AAF20909|AAF20909100%13032797%327467HACCH94847143103HMMERPFAM: 7 transmembranePF00001167.94107351.8receptor (rhodopsinfamily)blastx.2ORPHAN G PROTEIN-sp|O95853|O9585399%7879COUPLED RECEPTOR.HE8TI39849161418HMMERPFAM: EF handPF0003612.669861.8blastx.2CDNA FLJ11040 FIS,sp|BAA91969|BAA9196998%3371CLONE PLACE1004388.64%29968563%627719HEGAP32851207419HMMERPFAM: C2 domainPF0016833.03111721.8blastx.2SYNAPTOTAGMINsp|Q9R0N6|Q9R0N685%50238VIII.90%27029970%241270HCWFU66853005106HMMERPFAM: AldehydePF0017171.41052692.1.1dehydrogenase familyHUSYI29853149107HMMERPFAM: Sec1 familyPF00995108.833322.1.1blastx.2VACUOLAR PROTEINsp|Q63615|Q6361595%3314SORTING HOMOLOG93%403591R-VPS33A.90%33539431%87221HMEFT661134131108blastx.14(AF121859) sorting nexingi|4689258|gb|AAD27832.1|46%7549099 [Homo sapiens]AF121859_140%19332457%14218341%688738HMEFT66856149420HMMERPFAM: Src homologyPF0001828.5151361.8domain 3HKAAR71863023109HMMERPFAM: C2 domainPF0016816.263095511.8blastx.2TOLLIP PROTEIN.sp|Q9UJ69|Q9UJ6978%147959H7TBC95865922110HMMERPFAM: 7 transmembranePF00001189.536952.1.1receptor (rhodopsinfamily)blastx.2G-protein coupledsp|BAA93001|56%516701receptor SALPR.BAA9300161%5120641%303440H7TBC95908115421HMMERPFAM: 7 transmembranePF00001189.536952.1.1receptor (rhodopsinfamily)blastx.2angiotensin II receptorgb|AAC59635.1|34%6695[Xenopus laevis]HAPPX52872075422HMMERPFAM: Sugar (and other)PF0008337.84723591.8transportersHBGSJ131152326112blastx.14ferrienterobactin receptorgi|1778500|gb|AAB40783.1|94%1753precursor [Escherichiacoli]HBGSJ13878322424HMMERPFAM: Src homologyPF000184.074455101.8domain 3blastx.2ferrienterobactin receptorgb|AAB40783.1|92%64684precursor [Escherichiacoli]HFKLX38880220113HMMERPFAM: PMP-PF00822103.992992.1.122/EMP/MP20/Claudinfamilyblastx.2(AF087825) claudin-7gb|AAD09760.1|44%3299[Mus musculus]HTLGP151165362114blastx.14(AF060173) SV2 relatedgi|3901268|gb|AAC78627.1|88%301786protein [Rattusnorvegicus]HTLGP15880297425HMMERPFAM: Sugar (and other)PF0008334.172915901.8transportersHMEGH46887791426HMMERPFAM: C2 domainPF0016812.8110781.8blastx.2GLUT4 VESICLEsp|Q9Z1X5|Q9Z1X560%7216PROTEIN50%40216(FRAGMENT).HE8PY291129488116blastx.14(AF100751) FK506-gi|5410288|gb|AAD43015.1|100%2277binding protein FKB23isoform [Homo sapiens]HE8PY29887862427HMMERPFAM: EF handPF0003613.651912501.8blastx.2FK506-BINDINGsp|Q9Y6B0|Q9Y6B0100%2277PROTEIN.HTDAB17890384117HMMERPFAM: ThioredoxinPF00085107.92765332.1.1blastx.2CG1837 PROTEIN.sp|Q9VYV3|Q9VYV342%22551843%23153941%348533HCFCF47894415428HMMERPFAM: Eukaryotic proteinPF0006989.54202951.8kinase domainHDQHB19895106429HMMERPFAM: Eukaryotic proteinPF0006992.52605202.1.1kinase domainHAGDN531129154120blastx.2PALS1.sp|AAF63789|AAF6378997%3293HAGDN53895963430HMMERPFAM: Src homologyPF0001822.952703351.8domain 3blastx.2coded for by C. elegansgb|AAA96115.1|43%165455cDNA yk34a9.5; coded38%103156for by C. elegans 1elegans]HUFDB741162672121blastx.14phosphoinositide-specificgi|1195552|gb|AAA87954.1|58%170547phospholipase C [catfish,69%2160olfactory rosettes, PeptidePartial, 502 aa]HUFDB74901451431HMMERPFAM:PF0038789.421272.1.1Phosphatidylinositol-specific phospholipase C,Y domainHNHFH241092567122blastx.2PRO1722.sp|AAF69605|AAF6960558%90875668%763632HNHFH24903741432HMMERPFAM:PF0020937.22083062.1.1Sodium: neurotransmittersymporter familyblastx.14(AF075266) orphangi|3347930|gb|AAC27761.1|76%187327transporter isoform B927%414467[Mus musculus]HBGQT03908173433HMMERPFAM: SH3 domainPF0001868.56157852.1.1blastx.2(AF130979) SH3 domain-gb|AAF04472.1|AF130979_193%3791containing protein 6511[Homo sapiens]HETLF291103959124blastx.14(AJ250839)gi|7160989|emb|CAB76471.1|97%3482serine/threonine proteinkinase [Homo 1HETLF29909762434HMMERPFAM: Eukaryotic proteinPF00069143.1864161.8kinase domainblastx.14similar to cAMP-gi|3878636|emb|CAA88953.1|56%6416dependant protein kinase;cDNA EST 111HOUGD29909797435HMMERPFAM:PF00387118.22024532.1.1Phosphatidylinositol-specific phospholipase C,Y domainblastx.14(AF044576)gi|2957270|gb|AAC38963.1|42%202753phospholipase C PLC21035%757873[Caenorhabditis elegans]58%168203HTEMV091128254126blastx.14protein kinase I [Rattusgi|406113|gb|AAA19670.1|44%1321norvegicus]HTEMV09909843436HMMERPFAM: Eukaryotic proteinPF0006999.16193121.8kinase domainblastx.14protein kinase I [Rattusgi|406113|gb|AAA19670.1|44%1321norvegicus]HNTNB141128964127blastx.14calmodulin-bindinggi|349075|gb|AAA16633.1|98%42476protein [Rattus47%626676norvegicus]33%587676HNTNB14909942437HMMERPFAM: Eukaryotic proteinPF0006996.28383431.8kinase domainblastx.14calmodulin-bindinggi|349075|gb|AAA16633.197%41475protein [Rattus85%553657norvegicus]74%55365777%55365769%55965765%55365760%55365752%55365437%55365739%55363635%55364533%55965777%51253829%556657HE2KZ071149808128blastx.14(AB004267)gi|3135197|dbj|BAA28263.1|92%17508Ca2+/calmodulin-dependent protein kinase Ibeta 2 [Rattus norvegicus]HE2KZ07909948438HMMERPFAM: Eukaryotic proteinPF00069115.1952891.8kinase domainblastx.14(AB004267)gi|3135197|dbj|BAA28263.1|96%17433Ca2+/calmodulin-56%418507dependent protein kinase Ibeta 2 [Rattus norvegicus]HSIGN571105444129blastx.14(AB033615)gi|6705987|dbj|BAA89457.1|96%3962phospholipase C-L2 [Mus70%9821041musculus]HSIGN57910078439HMMERPFAM: PF00387159.31314842.1.1Phosphatidylinositol-specific phospholipase C,Y domainblastx.2PHOSPHOLIPASE C-L2.sp|Q9QYG1|Q9QYG183%2754HLHBC301106654130blastx.141-phosphatidylinositol-pir|S14113|S1411385%93324,5-bisphosphate77%408512phosphodiesterase 185%155175HLHBC30910079440HMMERPFAM: C2 domainPF00168104.78453051.8blastx.21-phosphatidylinositol-pir|S14113|S1411383%93324,5-bisphosphate74%408512phosphodiesterase 1HFBDJ131195217131blastx.14SH3 domains-containingpir|T09071|T0907190%1781182protein POSH - mouse56%1171136526%59869939%27237057%89893941%51460028%82389730%315383HFBDJ13911264441HMMERPFAM: SH3 domainPF0001878.61052692.1.1blastx.2(AF030131) Plenty ofgb|AAC40070.1|78%3473SH3s; POSH [Musmusculus]HTPGG25911282442HMMERPFAM: Eukaryotic proteinPF00069114.02723531.8kinase domainblastx.2(AL117482) hypotheticalemb|CAB55955.1|94%9353protein [Homo sapiens]92%35062263%258HSSMT34911294133HMMERPFAM: Eukaryotic proteinPF0006953.16952921.8kinase domainHWWDN341152430134blastx.14(AF144094)gi|6224683|gb|AAF05903.1|45%350814unconventional myosin-1551%128349[Homo sapiens]66%80088037%7215256%8713480%9612528%2820737%6914057%17821929%21628746%35439833%33940130%9919750%452487HWWDN34911357443HMMERPFAM: Src homologyPF0001814.096868531.8domain 3blastx.2(AF053130)gb|AAC40124.1|42%56874unconventional myosin66%788868MYO15 [Mus musculus]HCEPW85911374135HMMERPFAM: Eukaryotic proteinPF0006983.5232601.8kinase domainblastx.14predicted usinggi|3875903|emb|CAA94127.1|87%3260Genefinder; Similarity to111 cDNAHMTAW831071602136blastx.14(AF230904) c-Cbl-gi|7188749|gb|AAF37854.1|94%1354interacting protein [Homo 1AF230904_148%716851%8821059%78761%29835175%42546062%425448HMTAW83911385444HMMERPFAM: Src homologyPF0001876.1811591.8domain 3blastx.2(AF230904) c-Cbl-gb|AAF37854.1|AF230904_194%1354interacting protein [Homo52%7210sapiens]48%716861%29835175%425460HDMAV01911386445HMMERPFAM: Src homologyPF0001852.132644131.8domain 3blastx.2unnamed protein productemb|CAB42388.1|73%111410[unidentified]100%3116HDPSR74911396138HMMERPFAM: Src homologyPF0001847.192934601.8domain 3blastx.2(AF104246) enhancer ofgb|AAD11795.1|48%281553filamentation 1 homolog[Gallus gallus]HHEZT58911416446HMMERPFAM: Src homologyPF0001880.72774441.8domain 3blastx.14myosin I heavy chaingi|167839|gb|AAA33229.1|47%277435[Dictyosteliumdiscoideum]HTLDU05911649140HMMERPFAM: ActinsPF00022141.451254691.8 blastx.2ACTIN-LIKE-7-BETA.sp|Q9Y614|Q9Y61472%2580HTLET561189721141blastx.14actin [Filobasidiellagi|508701|gb|AAC49074.1|52%192854neoformans]48%932129130%837986HTLET56911654447HMMERPFAM: ActinsPF00022262.031347031.8blastx.2actin 1 - Trypanosomapir|A27724|A2772453%134715brucei39%71896340%9421022HTLCA95911655142HMMERPFAM: ActinPF00022345.217010962.1.1blastx.2actin - Phaffia rhodozymapir|S70377|S7037740%21096HTEJT861090517143blastx.14actin [Diphyllobothriumgi|1098579|gb|AAA82604.1|50%142684dendriticum]52%670111955%22129HTEJT86911656448HMMERPFAM: ActinPF00022106.843662.1.1blastx.2ACTIN.sp|Q9UVF3|Q9UVF344%2536966%366410HTEMA541134919144blastx.14(AF113526) actin-like-7-gi|5524058|gb|AAD44109.1|94%551359alpha [Homo sapiens]AF113526_1HTEMA54911666449HMMERPFAM: ActinPF00022320.724711612.1.1blastx.2ACTIN-LIKE-7-ALPHA.sp|Q9Y615|Q9Y61592%55116782%11211360HTLGJ171135518145blastx.14(AF191277) cytoplasmicgi|6478616|gb|AAF13923.1|59%360440actin [Cavia porcellus]AF191277_144%521574HTLGJ17915136450HMMERPFAM: ActinsPF0002225.122373171.8blastx.2DJ63M2.2 (similar tosp|CAC08484|CAC0848481%93413ACTIN) (Fragment).76%403465HOUES64918119146HMMERPFAM: AldehydePF00171138.532782.1.1dehydrogenase familyblastx.2lactaldehydepir|A38165|A3816598%3275dehydrogenase (EC1.2.1.22) aldA -Escherichia coliHMSCD15982250147blastx.14FBP 17 [Mus musculus]gi|1255033|gb|AAC52479.1|93%453635HMSCD15918133451HMMERPFAM: Src homologyPF0001841.064535991.8domain 3blastx.2(AK000975) unnameddbj|BAA91451.1|98%453635protein product [Homo29%387479sapiens]28%80175HDQDX20919027452HMMERPFAM: PX domainPF0078773.42465692.1.1blastx.14serine/threonine proteingi|294637|gb|AAA42137.1|78%633974kinase [Rattus norvegicus]44%465578HLTHP861110457149blastx.14(AF161420) HSPC302gi|6841254|gb|AAF28980.1|92%4981349[Homo sapiens]AF161420_176%13511833100%1811184952%12951351HLTHP86919354453HMMERPFAM: TBC domainPF0056669.485512742.1.1blastx.2(AF161420) HSPC302gb|AAF28980.1|AF161420_189%4561352[Homo sapiens]99%1309197452%12531309HMSOL52921126454HMMERPFAM: EF handPF0003612.432763591.8blastx.2CG11041 PROTEIN.sp|Q9V8Z6|Q9V8Z645%102464HAHGD33921782455HMMERPFAM: Eukaryotic proteinPF0006983.6845641.8kinase domainblastx.14(AF145690)gi|5052670|gb|AAD38665.1|68%1297BcDNA.LD28657AF145690_156%412609[Drosophila melanogaster]60%30442639%676804HHEHC53921783152HMMERPFAM: Eukaryotic proteinPF0006958.815077971.8kinase domainblastx.14(AF145690)gi|5052670|gb|AAD38665.1|79%567803BcDNA.LD28657AF145690_170%321563[Drosophila melanogaster]HE2PB01921850456HMMERPFAM: ActinPF0002235.9296162.1.1blastx.2HSPC281 (Fragment).sp|AAF28959|AAF28959100%2541HOUDP521219522154blastx.14CG7846 PROTEIN.sp|Q9VX09|Q9VX0943%2749152%1218136135%50175235%10021202HOUDP52922102457HMMERPFAM: ActinPF000223276010952.1.1blastx.2CG7846 PROTEIN.sp|Q9VX09|Q9VX0936%208110446%23190HHGAE471127881155blastx.14(AF187305) calmodulingi|5932428|gb|AAD56955.1|45%59499[Myxine glutinosa]AF187305_1HHGAE47922194458HMMERPFAM: EF handPF0003616.771712571.8blastx.2calmodulin [validated] -pir|S48728|MCHU48%310576human48%138260HMCGL451165349156blastx.14(AF187305) calmodulingi|5932428|gb|AAD56955.1|45%464904[Myxine glutinosa]AF187305_1HMCGL45922195459HMMERPFAM: EF handPF0003626.64605462.1.1blastx.2CALMODULIN.sp|Q9U6D3|Q9U6D345%460867HELEF111153884157blastx.14gamma-glutamylgi|1552811|gb|AAB08663.1|100%1283531phosphate reductase[Escherichia coli]HELEF11926930460HMMERPFAM: Pyridoxal-PF00282202.91465652.1.1dependent decarboxylaseconserved domainblastx.2glutamate decarboxylasepir|B43332|B4333281%131721(EC 4.1.1.15) beta -100%45152Escherichia coli56%59578047%564620HETJX041212235158blastx.14GRANUPHILIN-A.sp|Q9R0Q1|Q9R0Q194%668364%68581087%804827HETJX04927120461HMMERPFAM: C2 domainPF00168150.492602.1.1blastx.2GRANUPHILIN-A.sp|Q9R0Q1|Q9R0Q194%668357%68583148%719859HSOBC04927280462HMMERPFAM: EF handPF0003623.52783462.1.1blastx.2hypothetical proteinpir|T42709|T4270988%41388DKFZp586I0821.1 -human (fragment)HE8PW831069980160blastx.14(AB002584) beta-alanine-gi|1944136|dbj|BAA19549.1|86%4546pyruvate aminotransferase 1HE8PW83927532463HMMERPFAM:PF00202139.2744651.8Aminotransferases class-III pyridoxal-phosphateblastx.2ALANINE -sp|Q64565|AGT2_R83%4546GLYOXYLATEATAMINOTRANSFERASE2 PRECURSOR (EC 11HWLEA48927676161HMMERPFAM: Eukaryotic proteinPF0006932.821903811.8kinase domainblastx.2(AF169034) proteingb|AAF12757.2|AF169034_159%154429kinase [Homo sapiens]100%8916651%287415HNHNP811129143162blastx.14(AF091575) olfactorygi|3769641|gb|AAC64595.1|62%618896receptor [Rattus61%236505norvegicus]72%502621HNHNP81928378464HMMERPFAM: 7 transmembranePF0000158.092335111.8receptor (rhodopsinfamily)blastx.2OLFACTORYsp|Q9Z231|Q9Z23161%236505RECEPTOR52%502618(FRAGMENT).HFIDL681123641163blastx.14G protein-coupledgi|438129|emb|CAA80651.1|44%945742receptor [Lymnaea46%1086997stagnalis]HFIDL68928475465HMMERPFAM: 7 transmembranePF0000150.4283191.8receptor (rhodopsinfamily)blastx.2CG5042 PROTEIN.sp|Q9VBP0|Q9VBP038%8397HUJCT051165261164blastx.14(AF155116) NY-REN-60gi|5360127|gb|AAD42882.1|100%12771540antigen [Homo sapiens]AF155116_1HUJCT05929264466HMMERPFAM: EF handPF0003611.523594331.8blastx.2CG8334 PROTEINsp|Q9VW49|Q9VW4959%56499(FRAGMENT).HTEGO05932583165HMMERPFAM: Eukaryotic proteinPF0006950.832332.1.1kinase domainblastx.14male germ cell-associatedgi|205278|gb|AAA41562.1|85%3395kinase (mak) [Rattus64%489761norvegicus]85%76884838%10231100HRDBH581226719166blastx.14probable aldehydepir|II83136|H8313638%13201637dehydrogenase PA407340%21242309[imported] - Pseudomonas56%675770aeruginosa (strain PAO1)31%46263527%2328247123%894109440%1101119630%35446150%83187225%11251184HRDBH58933364467HMMERPFAM: AldehydePF00171175.4953615581.8dehydrogenasesblastx.2probable aldehydepir|H83136|H8313629%5691528dehydrogenase PA407334%244450[imported] - Pseudomonasaeruginosa (strain PAO1)HSDGW22934467167HMMERPFAM: Eukaryotic proteinPF00069441453182.1.1kinase domainblastx.14(AF071071) proteingi|5305333|gb|AAD41593.1|96%145318kinase Myak-S [MusAF071071_1100%9143musculus]100%303326HNTMD791126594168blastx.14(AB037134) IRE homologgi|6729348|dbj|BAA89784.1|58%2326031 [Arabidopsis thaliana]66%157228HNTMD79934522468HMMERPFAM: Eukaryotic proteinPF00069130.822035861.8kinase domainblastx.14LATS [Drosophilagi|903942|gb|AAA70336.1|52%239586melanogaster]33%7615657%16921022%64240HCE5J511197900169blastx.14SRG1.sp|P97610|P9761098%11703HCE5J51934524469HMMERPFAM: C2 domainPF0016882.593386041.8blastx.14Srg1 [Rattus norvegicus]gi|1772658|gb|AAC53019.1|94%1230286%29362288%625702HHEFQ421151482170blastx.14(AB025258) granuphilin-agi|5926736|dbj|BAA84656.1|40%57428[Mus musculus]56%81098048%61881542%63074331%18629946%567605HHEFQ42934527470HMMERPFAM: C2 domainPF00168128.82034722.1.1blastx.14(AB025258) granuphilin-agi|5926736|dbj|BAA84656.1|40%128505[Mus musculus]57%887105746%69589239%70782031%26337646%64468238%57261075%107130HLQDC551082368171blastx.14(AK002037) unnamedgi|7023676|dbj|BAA92048.1|49%332496protein product [Homo42%149253sapiens]HLQDC55934528471HMMERPFAM: C2 domainPF0016881.112162.1.1blastx.2CG15078 PROTEIN.sp|Q9V8M4|Q9V8M434%744143%123464%444494HFPHI621195825172blastx.14rabphilin-3A - bovinepir|A48097|A4809795%184111968%490HFPHI62934529472HMMERPFAM: C2 domainPF00168106.44272931.8blastx.2rabphilin-3A - mousepir|JX0338|JX033890%3410HE8QH091152238173blastx.14(AB000893)gi|1840399|dbj|BAA19204.1|95%28591synaptotagmin 3 [Mus96%585923musculus]39%4622833%19335139%453521HE8QH09934532473HMMERPFAM: C2 domainPF00168168.2773372.1.1blastx.14(AB000893)gi|1840399|dbj|BAA19204.1|92%56568synaptotagmin 3 [Mus98%564830musculus]42%40755633%57373132%17032834%56205100%215339%430498HFAAX291128791174blastx.14(AF000423)gi|2130632|gb|AAB58344.1|99%25570synaptotagmin XI [Rattus35%385510norvegicus]25%16933347%331387HFAAX29934540474HMMERPFAM: C2 domainPF00168115.21944632.1.1blastx.2SYNAPTOTAGMIN XI.sp|O08835|O0883598%855032%149490HHFOC791182276175blastx.14(AF081251) putative epsgi|3415099|gb|AAC31599.1|97%84296protein [Rattusnorvegicus]HIIFOC79935406475HMMERPFAM: EF handPF0003613.961862631.8blastx.2EH domain containing 2.sp|AAF40470|AAF4047098%54248HOGEQ43935465476HMMERPFAM: Src homologyPF0001828.13581321.8domain 3blastx.2(AF132480) Ese2 proteingb|AAD19748.1|93%37132[Mus musculus]IICECQ23938398177HMMERPFAM: CUB domainPF0043153.75674332.1.1blastx.2seizure-related proteinpir|I52657|I5265793%810370SEZ-6 precursor - mouse90%33523729%76537035%74457130%74757427%68748427%537409HTGAU791178621178blastx.14(AL157917) similarity togi|7106102|emb|CAB76028.1|50%745966endopeptidases 138%36156152%60772052%313363HTGAU79940369477HMMERPFAM: Eukaryotic proteinPF0006931.253157791.8kinase domainblastx.2(AL157917) similarity toemb|CAB76028.1|45%324977endopeptidases[Schizosaccharomyces 1HE9FI331156432179blastx.14putative integralgi|3004482|emb|CAA71076.1|59%14286membrane transport61%11531206protein [Rattusnorvegicus]HE9FI33941348478HMMERPFAM: Sugar (and other)PF0008317.622104221.8transportersblastx.2putative integralemb|CAA71076.1|53%3428membrane transportprotein [Rattusnorvegicus]HNHCP79941862480HMMERPFAM: 7 transmembranePF00001118.4726701.8receptor (rhodopsinfamily)blastx.14(AF102533) olfactorygi|3983394|gb|AAD13325.1|55%2658receptor F7 [Musmusculus]HTLIY521194806181blastx.14TESTIS-SPECIFICsp|Q61241|Q6124146%624956SERINE/THREONINE48%126398KINASE.45%41156342%549605HTLIY52942161481HMMERPFAM: Eukaryotic proteinPF00069251.191669331.8kinase domainblastx.2serine/threonine kinasegb|AAA99535.1|44%133936[Mus musculus]HRAED74942527182HMMERPFAM: Eukaryotic proteinPF0006959.64066121.8kinase domainblastx.2(AB023658)dbj|BAA75246.1|97%71346Ca/calmodulin-dependent81%388648protein kinase kinase71%342425alpha, CaM-kinase kinase88%662688alpha [Rattus norvegicus]HFKKN77943757183HMMERPEAM: 7 transmembranePF0000180.792745731.8receptor (rhodopsinfamily)blastx.2G-protein coupledpir|JC7289|JC728982%160714receptor, SREB3 - humanHTEMU661205381184blastx.14MEK KINASE ALPHA.sp|O96611|O9661151%71496266%633668HTEMU66944419482HMMERPFAM: Eukaryotic proteinPF00069114.856139631.8kinase domainblastx.2MEK Kinase 3 [Musgb|AAB03535.1|49%604948musculus]29%209340HWAGU621206797185blastx.14Zinc transporter like 1.sp|AAF43422|AAF4342295%26482796%969151193%2926233%22111HWAGU62945368483HMMERPFAM: Cation effluxPF0154515261913952.1.1familyblastx.2(AC007231) putativegb|AAD32753.1|AC007231_145%388858cation transport protein42%9101386[Arabidopsis thaliana]38%883951HFPFB391198036186blastx.14CG8745 PROTEIN.sp|Q9VU95|Q9VU95|65%15361460%69393565%945111866%1332141235%1161132850%609686HFPFB39946170484HMMERPFAM:PF00202235.0516137141.8Aminotransferases class-III pyridoxal-phosphateblastx.2hypothetical proteinpir|T25848|T2584849%1613651T01B11.2 -Caenorhabditis elegansHPMFI381165993187blastx.14(AB005451) RST [Musgi|2696709|dbj|BAA23875.1|51%30296musculus]73%382438HPMFI38946252485HMMERPFAM: Sugar (and other)PF0008332.168605641 .8transportersblastx.2renal organic anionemb|CAB09724.1|43%15488transporter[Pseudopleuronectesamericanus]HBXDJ07946830188HMMERPFAM: Synaptophysin/PF01284406.71256042.1.1synaptoporinblastx.2synaptoporin - ratpir|JH0300|JH030090%12564391%610921HOFMS431152417189blastx.14porcine membranegi|1018989|dbj|BAA20476.1|62%325738cofactor protein [Sus47%7312scrofa]52%32237829%67376536%68874433%512574HOFMS43947973486HMMERPFAM: Sushi domainPF00084641743022.1.1(SCR repeat)blastx.2PORCINE MEMBRANEsp|O02839|O0283947%12317COFACTOR PROTEIN.HOVCO141091087190blastx.2C4BP beta chain, leaderemb|CAA90392.1|34%3434[Rattus norvegicus]HOVCO14947999487HMMERPFAM: Sushi domainPF0008484211702.1.1(SCR repeat)blastx.2UNKNOWN PROTEINsp|Q28797|Q2879733%21347(FRAGMENT).34%2138038%42451647%424474HTEPE351105272191blastx.141-phosphatidylinositol-pir|S14113|S1411353%13574,5-bisphosphate51%451750phosphodiesterase 1HTEPE35948475488HMMERPFAM:PF00387163.88395072.1.1Phosphatidylinositol-specific phospholipase C,Y domainblastx.21-phosphatidylinositol-pir|S14113|S1411348%839904,5-bisphosphatephosphodiesterase 1HE8UA521229490192blastx.14collagen alpha 3(VI) chainpir|A37797|A3779734%121729precursor - chicken32%805135335%70081919%20833044%81486724%19630650%80886126%20833030%395484HE8UA52948509489HMMERPFAM: von WillebrandPF0009257.112085611.8factor type A domainblastx.2collagen alpha 3(VI) chain -pir|S32605|S3260541%121576mouse (fragment)HOUBE501090776193blastx.14neuroligin 3 [Rattusgi|1145791|gb|AAA97871.1|74%3761140norvegicus]67%3125277%27738168%11341181HOUBE50948519490HMMERPFAM: CarboxylesterasesPF0013555.97162431.8blastx.2Neuroligin 3 isoformsp|AAF71231|AAF7123170%31243HNL3s (Fragment).HAJAV281165229194blastx.14(AK000544) unnamedgi|7020711|dbj|BAA91243.1|95%961118protein product [Homo 100%10641345sapiens]HAJAV28948630491HMMERPFAM: ActinPF0002235.91202302.1.1blastx.2Uncharacterizedsp|AAF67655|AAF6765597%96458hypothalamus proteinHARP11.HAQBZ891083554195blastx.14strong similarity to class-gi|1707274|gb|AAB37999.1|49%249578III of 1 elegans]38%59488152%87399254%17258%186221HAQBZ89949061492HMMERPFAM:PF0020262.49683251.8Aminotransferases class-III pyridoxal-phosphateblastx.2CG8745 PROTEIN.sp|Q9VU95|Q9VU9552%8316HELHF07949067196HMMERPFAM:PF0020238.85952951.8Aminotransferases class-III pyridoxal-phosphateblastx.144-aminobutyrategi|1742132|dbj|BAA14871.1|85%83295aminotransferase (EC92%21982.6.1.19) 1145%246311aminotransferase).100%118[Escherichia coli]HE9QQ221127726197blastx.14(AB002584) beta-alanine-gi|1944136|dbj|BAA19549.1|77%92418pyruvate aminotransferase 180%41858585%587628100%677697HE9QQ22949080493HMMERPFAM:PF00202105.12855452.1.1Aminotransferases class-III pyridoxal-phosphateblastx.2ALANINE -sp|Q64565|AGT2—51%31070GLYOXYLATERAT43%5451003AMINOTRANSFERASE49%6829992 PRECURSOR (EC 11HSDSB06949151494HMMERPFAM: SH3 domainPF00018249.34836472.1.1blastx.2(AL133047) hypotheticalemb|CAB61374.1|98%3863protein [Homo sapiens]30%684833%222848HACAD35949199199HMMERPFAM: von WillebrandPF0009270.8146111742.1.1factor type A domainblastx.2SIMILAR TO COCH-sp|Q9UDN0|Q9UDN099%14649525B2.100%94764895%65745722%94466350%1005952HEQAP17949358200HMMERPFAM: 7 transmembranePF0000194.577414361.8receptor (rhodopsinfamily)blastx.2Orphan seven-sp|AAF59827|AAF5982784%786295transmembrane receptor.HMTBB17950884495HMMERPFAM: EF handPF0003615.742852021.8blastx.2CDNA FLJ10466 FIS,sp|BAA91628|BAA91628100%513100CLONE NT2RP1001665.HKGDE58945039496blastx.2CDNA FLJ10466 FIS,sp|BAA91628|BAA9162886%17835CLONE NT2RP1001665.30%28169155%69782536%69091435%32208HKGDE58950885497HMMERPFAM: EF handPF0003615.983042211.8blastx.14(AJ133836) calmodulin 2gi|4581211|emb|CAB40132.1|28%337179[Branchiostoma floridae]37%166119HCHMW401144323203blastx.14calmodulin [Plasmodiumgi|385234|gb|AAA29508.160%376564falciparum]49%13634838%34957336%15734532%481564HCHMW40951518498HMMERPFAM: EF handPF00036129.94865722.1.1blastx.2Calmodulin-like skinsp|AAF66821|AAF6682198%135572protein.HE8QZ341143411204blastx.14predicted usinggi|3875264|emb|CAB01132.1|39%9411201Genefinder; similar to EF50%437595hand (2 domains)40%695877[Caenorhabditis elegans]32%22741226%10821183HE8QZ34952283499HMMERPFAM: EF handPF0003612.975436171.8blastx.2CG4662 PROTEIN.sp|Q9VDT8|Q9VDT838%24998042%24987833%9771084HWAFG041227627205blastx.14PRO1038.sp|AAF71042|AAF7104256%555044%536745HWAFG04952878500HMMERPFAM: Eukaryotic proteinPF0006993.7416559451.8kinase domainblastx.14(AC002343) Ser/Thrgi|2262107|gb|AAB63615.141%16551383protein kinase isolog48%13191185[Arabidopsis thaliana]42%104693375%13551332HTEKT33953308501HMMERPFAM: Eukaryotic proteinPF00069200.5842813931.8kinase domainblastx.2(AC007661) putativegb|AAD32787.1|AC007661_2441%7221009protein kinase36%10701243[Arabidopsis thaliana]29%428628HBXDM07953622502HMMERPFAM: Sec1 familyPF0099596.72675752.1.1blastx.2Vacuolar protein sortingsp|AAF91174|AAF9117483%14172833B.97%114135%587706HFPFA83955614208HMMERPFAM: 7 transmembranePF00001107.63166811.8receptor (rhodopsinfamily)blastx.2G-protein coupledpir|JC7289|JC728998%202735receptor, SREB3 - humanHKADO36956115503HMMERPFAM: Sugar (and other)PF0008344.0322771.8transportersHFXKG51956596210HMMERPFAM: Sugar (and other)PF00083121.545061471.8transportersblastx.2No definition line foundgb|AAB18499.1|100%560982[Escherichia coli]HFPHR82957528504HMMERPFAM: ActinPF0002291.713223572.1.1blastx.2Uncharacterizedsp|AAF67655|AAF67655100%1523273hypothalamus proteinHARP11.HISAF59959140212HMMERPFAM: Eukaryotic proteinPF0006989.463407711.8kinase domainblastx.14(AC002343) Ser/Thrgi|2262107|gb|AAB63615.1|39%460768protein kinase isolog33%397468[Arabidopsis thaliana]HCEHD661136122213blastx.14neuronal calcium sensorgi|498032|gb|AAA88510.1|98%2562[Rattus norvegicus]HCEHD66959160505HMMERPFAM: EF handPF0003664.23113972.1.1blastx.2Neuronal calcium sensor-1.sp|AAD01642|AAD01642100%14583HE8UY741163590214blastx.14(AF080119) containsgi|3600036|gb|AAC35524.1|38%13291similarity to protein68%367441kinase 161%464502HE8UY74960914506HMMERPFAM: Eukaryotic proteinPF0006936.371144071.8kinase domainblastx.14(AF080119) containsgi|3600036|gb|AAC35524.1|36%117290similarity to protein45%13111kinase 173%36641037%467553HAHIY08962113215HMMERPFAM: Eukaryotic proteinPF0006974.92392781.8kinase domainblastx.14similar to tyrosine kinasegi|470364|gb|AAC47047.1|44%192278[Caenorhabditis elegans]64%189258%108179H2CBH45963811507HMMERPFAM: Src homologyPF00018131943101.8domain 3blastx.2Kryn [Mus musculus]dbj|BAA19686.1|85%237379%38146787%46048370%131160HMVAM09963814508HMMERPFAM: Src homologyPF000184.797288021.8domain 3blastx.2(AK001580) unnameddbj|BAA91769.1|96%20802protein product [Homosapiens]HFPEN041199663218blastx.14CG8745 PROTEIN.sp|Q9VU95|Q9VU9565%15661760%69693865%948112166%1335141535%1164133150%612689HFPEN04964824509HMMERPFAM:PF0020233.542594891.8Aminotransferases class-III pyridoxal-phosphateblastx.2CG8745 PROTEIN.sp|Q9VU95|Q9VU9562%148492HSLJD021104452219blastx.14UhpC protein [Escherichiagi|148114|gb|AAA24722.1|95%145927coli]HSLJD02965826510HMMERPFAM: Sugar (and other)PF0008319.534648741.8transportersblastx.2UhpC protein [Escherichiagb|AAA24722.1|100%56907coli]HDPFZ301220164220blastx.14Sulfate transporter.sp|CAC05432|CAC0543255%1684151748%1433133532%115499650%18013935%11991140HDPFZ30966752511HMMERPFAM: Sulfate transporterPF0091660.2149612332.1.1familyblastx.2(AF180728) sulfategb|AAD53951.1|28%175807transporter [Drosophila52%14196melanogaster]HPJCR33966758512HMMERPFAM: C2 domainPF0016831.15132671.8blastx.2E3 UBIQUITIN LIGASEsp|Q9UJT8|Q9UJT889%7375SMURF1 (FRAGMENT).HTOAK34966800513HMMERPFAM: Eukaryotic proteinPF0006932.41102011901.8kinase domainblastx.14(AF084205)gi|3452473|gb|AAC71014.1|75%9541190serine/threonine proteinkinase TAO1 [Rattusnorvegicus]HE8NI24971296223HMMERPFAM: 7 transmembranePF0000161.744537071.8receptor (rhodopsinfamily)blastx.2G-protein coupledpir|T47131|T4713193%345707receptor, SREB2 - human88%722748HAMFM391055532224blastx.14(AK001509) unnamedgi|7022807|dbj|BAA91729.1|53%38603432protein product [Homo100%41714106sapiens]HAMFM39971347514HMMERPFAM: Src homologyPF0001867.14113613061.8domain 3blastx.2(AK001509) unnameddbj|BAA91729.1|59%45114017protein product [Homosapiens]HBGMG39971414515HMMERPFAM: EF handPF0003610.69611411.8blastx.245 KDA CALCIUM-sp|Q61112|CB45_MOUSE94%7165BINDING PROTEINPRECURSOR(STROMAL 1HSXBV891128699226blastx.2TYPE Isp|Q9UJ47|Q9UJ4793%72037TRANSMEMBRANE29%3131044RECEPTOR29%7421464PRECURSOR.HSXBV89971821516HMMERPFAM: Sushi domainPF0008443.61232902.1.1(SCR repeat)blastx.2TYPE Isp|Q9UJ47|Q9UJ4782%3635TRANSMEMBRANE32%96536RECEPTORPRECURSOR.HBIOZ101143756227blastx.14(AF003134) stronggi|2088685|gb|AAB54139.1|43%3497similarity to theCDC2/CDX 1HBIOZ10973131517HMMERPFAM: Eukaryotic proteinPF00069121.133651.8kinase domainblastx.2(AF003134) stronggb|AAB54139.1|60%3305similarity to theCDC2/CDX subfamily ofser/thr protein kinases[Caenorhabditis elegans]HTLEJ111085651228blastx.14(AF144573) Mx-gi|4868443|gb|AAD31319.1|69%35268interacting protein kinaseAF144573_140%437592PKM [Mesocricetus42%293397auratus]38%877939HTLEJ11973302518HMMERPFAM: Eukaryotic proteinPF0006955.9442232.1.1kinase domainblastx.14(AF144573) Mx-gi|4868443|gb|AAD31319.1|69%35268interacting protein kinaseAF144573_140%437592PKM [Mesocricetus42%293397auratus]38%877939HAWAM691207835229blastx.14SPARC-RELATEDsp|Q9WVN9|Q9WVN963%580972PROTEIN.53%19343853%961122452%4919848%22333346%64075623%38262434%34341150%60465733%76083128%1025112038%22382300HAWAM69943104519blastx.2SPARC-RELATEDsp|Q9WVN9|Q9WVN951%31261PROTEIN.HAWAM69973465520HMMERPFAM: EF handPF0003610.1397261.8blastx.14(AF070470) SPARC-gi|5305327|gb|AAD41590.1|62%1335related protein [MusAF070470_1musculus]HSCKD111056288230blastx.14(AJ243342) nicotinicgi|6688136|emb|CAB65091.1|98%12921831acetylcholine receptor88%23472886alpha 9 subunit [Homo100%54203sapiens]33%3000310735%29382988HSCKD11973894521HMMERPFAM: Neurotransmitter-PF0006531.561472571.8gated ion-channelblastx.2(AJ243342) nicotinicemb|CAB65091.1|90%120296acetylcholine receptoralpha 9 subunit [Homosapiens]HDPLT62973945522HMMERPFAM: Neurotransmitter-PF00065102.124177461.8gated ion-channelblastxGABA receptor rho-3dbj|BAA09322.1|78%414797subunit precursor [Rattus92%262411norvegicus]HTPFX16974296232HMMERPFAM: PMP-PF0082250.2482992.1.122/EMP/MP20/Claudinfamilyblastx.2CLAUDIN-18.sp|P56857|CLDI_MOUSE67%3935944%316483HE9NO661079624233blastx.14(AB035267) Nck-gi|6472874|dbj|BAA87066.1|82%449775interacting kinase-like94%2283embryo specific kinase80%748990[Mus musculus]HE9NO66974353523HMMERPFAM: Eukaryotic proteinPF00069121.64737571.8kinase domainblastx.14(AB020741) NIK-relatedgi|6009519|dbj|BAA84943.173%449817kinase [Mus musculus]94%228379%748990HSDJI441154068234blastx.14(AE000180) 7,8-gi|1786991|gb|AAC73861.1|94%8031849diaminopelargonic acid98%18282088synthetase [Escherichiacoli]HSDJI44974784524HMMERPFAM:PF00202511.489418352.1.1Aminotransferases class-III pyridoxal-phosphateblastx.14(AE000180) 7,8-gi|1786991|gb|AAC73861.1|99%7951841diaminopelargonic acid100%18371860synthetase [Escherichiacoli]HFXDP53578868525HMMERPFAM: CUB domainPF0043111.7721771.8HWADY66734565236HMMERPFAM: Eukaryotic proteinPF0006928.8211741.8kinase domainHLDBC631144557237blastx.14carnitinegi|755646|gb|AAC41748.1|99%3629palmitoyltransferase I[Homo sapiens]HLDBC63745061526HMMERPFAM: CarnitatePF00755258.434102.1.1acyltransferaseHFIVB68752981527HMMERPFAM: C2 domainPF0016835.534065701.8blastx.2hypothetical proteinpir|T12449|T1244993%187615DKFZp564E1616.1 -97%692835human (fragments)97%58869296%10418468%310736%57366827%40654336%39746554%125157HTLAC561181355239blastx.14carnitinegi|755646|gb|AAC41748.1|54%6599palmitoyltransferase I[Homo sapiens]HTLAC56753093528HMMERPFAM: CarnitatePF00755143.364222.1.1acyltransferaseHSSAD41753094240HMMERPFAM: CarnitatePF0075590.8512992.1.1acyltransferaseHCFMT57765375529HMMERPFAM: Src homologyPF0001814.5510731.8domain 3blastx.2(AF039571) peripheralgb|AAD11957.1|96%3773benzodiazepine receptorinteracting protein; PBR-IP/PRAX1 [Homosapiens]HDAAV61810305530HMMERPFAM: Eukaryotic proteinPF0006941.11111451.8kinase domainHDPKD751096253243blastx.14(AF191838) TANKgi|6224868|gb|AAF05989.1|100%74637binding kinase TBK1AF191838_1[Homo sapiens]HDPKD75810824531HMMERPFAM: Eukaryotic proteinPF0006998.74684331.8kinase domainHTEON291126312244blastx.14(AB021866) CIB [Homogi|4092850|dbj|BAA36281.1|50%190339sapiens]47%39450157%10318648%1111HTEON29815852532HMMERPFAM: EF handPF0003622.292663491.8blastx.2CALCIUM-ANDsp|Q9R010|Q9R01041%2496INTEGRIN-BINDINGPROTEIN CIB.HSKAC241121800245blastx.14(AF144573) Mx-gi|4868443|gb|AAD31319.1|94%37453interacting protein kinaseAF144573_177%464490PKM [Mesocricetus100%1743auratus]88%484510HSKAC24823869533HMMERPFAM: Eukaryotic proteinPF0006979.361224541.8kinase domainHTJAA71846682534HMMERPFAM: S-100/ICaBP typePF0102336.3772082.1.1calcium binding domainblastx.2TUMOR RELATEDsp|Q9UBG3|Q9UBG370%68325PROTEIN.HTEKS201124378247blastx.14calcineurin [Bos taurus]gi|312969|emb|CAA50659.1|77%53562HTEKS20846714535HMMERPFAM: EF handPF0003684.74535392.1.1blastx.2calcineurin regulatorypir|A33391|A3339177%60569chain - humanHE9TK49856343536HMMERPFAM: Ion transportPF0052077.02112561.8proteinsblastx.2(AB012043) NBR13dbj|BAA36409.1|95%2256[Homo sapiens]50%25632737%259282HCHAT01867209537HMMERPFAM: EF handPF0003624.01122713041.8blastx.2AD 3 (FRAGMENT).sp|Q9UQ32|Q9UQ3247%795140972%1436757%47278379%375476HCEEN061150867250blastx.14(AB025258) granuphilin-agi|5926736|dbj|BAA84656.1|32%296490[Mus musculus]48%54865247%24229842%152229HCEEN06878658538HMMERPFAM: C2 domainPF0016851.792034661.8HDPKI83883382251HMMERPFAM: C2 domainPF0016813.475306011.8blastx.2MUNC13-4 PROTEIN.sp|Q9R189|Q9R18976%19463195%73579480%661738HSPBQ121152258252blastx.14copine I [Homo sapiens]gi|1791257|gb|AAC15920.1|73%5557063%570851HSPBQ12884004539HMMERPFAM: C2 domainPF0016842.063526241.8HPCID78886915253HMMERPFAM: Sulfate transporterPF0091626.42653812.1.1familyblastx.2(AF030880) pendringb|AAC51873.1|40%25375[Homo sapiens]HDTKQ14886936254HMMERPFAM: Src homologyPF0001812.874305461.8domain 3blastx.2(AL049683) hypotheticalemb|CAB41255.1|100%439555protein [Homo sapiens]56%76291HRACK83888037255HMMERPFAM: Eukaryotic proteinPF0006948.42114231.8kinase domainHSIAO78889498540HMMERPFAM: EF handPF0003619.913894631.8blastx.2HYPOTHETICAL 22.5 KDAsp|O43745|O4374597%38622PROTEIN.HWAGS73894404257HMMERPFAM: Eukaryotic proteinPF0006964.1742731.8kinase domainHCMSL08898203258HMMERPFAM: CalsequestrinPF012161001.15212212.1.1blastx.2calsequestrin precursor,pir|A60424|A6042495%1121197fast skeletal muscle -humanHCMSL08959176541HMMERPFAM: CalsequestrinPF01216697.413725692.1.1blastx.14calmitine; calsequestrinegi|688292|gb|AAB32063.1|93%1372593[Homo sapiens]HLWFN631101533259blastx.14(AL049683) hypotheticalgi|4678753|emb|CAB41255.1|45%470937protein [Homo sapiens]75%9891024HLWFN63908437542HMMERPFAM: Src homologyPF0001812.815156641.8domain 3blastx.2(AL049683) hypotheticalemb|CAB41255.1|44%4641024protein [Homo sapiens]HPWAY10908549543HMMERPFAM: KRAB boxPF01352156.32063942.1.1blastx.14zinc finger protein 30gi|456269|emb|CAA82913.1|70%152325[Mus musculus67%326454domesticus]HOUDH191153909261blastx.14(AC007842) BC331191_1gi|5080758|gb|AAD39268.1|100%57335[Homo sapiens]AC007842_3HOUDH19908588544HMMERPFAM: KRAB boxPF01352169.72414292.1.1blastx.2(AC007842) BC331191_1gb|AAD39268.1|AC007842_391%226549[Homo sapiens]HDPFF24909232545HMMERPFAM: KRAB boxPF01352121.31583492.1.1blastx.2(AC007228) R31665_2gb|AAD23606.1|AC007228_150%158457[AA 1-673] [Homosapiens]HWLFH941152278263blastx.14(AK000265) unnamedgi|7020230|dbj|BAA91041.1|41%739939protein product [Homo53%595690sapiens]57%335397HWLFH94909682546HMMERPFAM: Src homologyPF0001858.423084631.8domain 3blastx.2(AK000265) unnameddbj|BAA91041.1|40%215535protein product [Homosapiens]HWMBM131152283264blastx.14(AK000265) unnamedgi|7020230|dbj|BAA91041.1|56%153296protein product [Homo41%345545sapiens]HWMBM13909683547HMMERPFAM: Src homologyPF0001859.641262811.8domain 3blastx.2Eps8 [Mus musculus]gb|AAA16358.1|35%3331737%324527HFIUE751172525265blastx.14(AB037134) IRE homologgi|6729348|dbj|BAA89784.1|53%86811341 [Arabidopsis thaliana]40%1126125128%526684HFIUE75909758548HMMERPFAM: Eukaryotic proteinPF0006985.683776641.8kinase domainblastx.14(AD000092) hypotheticalgi|1905906|gb|AAB51171.1|43%362634human serine-threonine46%632715protein kinase R31240_147%724774[Homo sapiens]HNTCP13909770549HMMERPFAM: Eukaryotic proteinPF00069102.964459301.8kinase domainblastx.14(AC006530) unknowngi|4809337|gb|AAD30182.1|55%463957[Homo sapiens]AC006530_4HBIBQ89909782550HMMERPFAM: SH3 domainPF0001849.72123762.1.1blastx.2p115 [Homo sapiens]emb|CAA55394.1|41%14397HWBEG18909798551HMMERPFAM: EF handPF0003633.35055912.1.1blastx.2RAS ACTIVATORsp|Q9UNN9|Q9UNN955%103684RASGRP.71%69888948%2142HTAHB431221956269blastx.14PUTATIVE RASGAP-sp|O43374|O4337499%1532174ACTIVATING-LIKE100%21742356PROTEIN.97%4216434%4514031%42952457%14361477HTAHB43909845552HMMERPFAM: GTPase-activatorPF0061661.35197312.1.1protein for Ras-likeGTPaseblastx.2PUTATIVE RASGAP-sp|O43374|O4337497%39860ACTIVATING-LIKE100%133PROTEIN.HSYBX32909846270HMMERPFAM: C2 domainPF0016841.14461891.8blastx.2PUTATIVE RASGAP-sp|O43374|O4337498%49228ACTIVATING-LIKEPROTEIN.HCEHE35909937553HMMERPFAM: Eukaryotic proteinPF0006930.782103471.8kinase domainblastx.14protein kinase PRK2gi|914100|gb|AAB33346.1|66%204365[human, DX3 B-cellmyeloma cell line,Peptide, 984 aa] [Homosapiens]HFCBB561204971272blastx.14inositol 1,4,5-pir|S62358|S6235844%280387trisphosphate-binding31%439612protein, 130K - rat26%598744HFCBB56910073554HMMERPFAM: EF handPF0003623.954315141.8blastx.21-phosphatidylinositol-pir|S14113|S1411336%2755654,5-bisphosphatephosphodiesterase 1HAMFL82910074273HMMERPFAM: C2 domainPF0016873.492121.8blastx.2PHOSPHOLIPASE C-L2.sp|Q9QYG1|Q9QYG197%3317HBXCM381174533274blastx.14unnamed protein productgi|6740727|emb|CAB69447.1|97%4051349[unidentified]87%13396HBXCM38910086555HMMERPFAM: Src homologyPF0001855.89106212321.8domain 3blastx.2unnamed protein productemb|CAB69447.1|92%4021316[unidentified]87%1339677%12951348HLHCR16910123275HMMERPFAM: Sushi domainPF00084744.91973582.1.1(SCR repeat)blastx.2complement receptor 1 -pir|I36936|I3693629%7101600chimpanzee (fragment)30%1166192131%818163630%1958276428%710151329%1163192129%1757263228%1766263231%911163626%1970303127%1754260528%1166192130%2378303136%2056229%1244192126%2210316326%2210316332%1157132%1157132%710120432%1156232%710120430%2359530%992163032%710120429%2359536%2346033%2046027%3284429%728138730%983154928%728138731%713124632%2657312431%860138730%518122831%713124633%713111732%701111432%11357432%11357431%2417298330%1811224232%2642310934%8046032%1244163933%701111131%9554729%2219298331%2681313029%2618310930%734117131%1109153732%8045131%2219276132%725111425%1721245229%11357431%725111428%1550211326%2474331930%1472202627%1424211325%2474310025%1721245227%1100163929%893136335%1136426%1721232627%2555332226%1721244933%18257426%1100163029%2549293830%1997240130%3842432%1241165730%626101530%734109025%1250185224%1250185230%3837031%9538231%9538231%734109026%1769246431%725101531%725101529%1799205625%1472188230%59384722%371598HLHCR16965511556HMMERPFAM: Sushi domainPF00084357.81973582.1.1(SCR repeat)blastx.2furrowed [Drosophilagb|AAB36703.1|31%6381480melanogaster]28%254122834%737138728%254112035%8960432%908146834%692118930%740127030%419101832%1152927%1791627%1755334%1100148028%1144530%977148037%36557129%12201489HE6GF021153883276blastx.14(AJ007012) Fish proteingi|3702174|emb|CAA07416.1|75%10192[Mus musculus]66%20237839%20237539%119238%1316839%1016841%20537838%22337237%25332450%55060950%27332054%346378HE6GF02911263557HMMERPFAM: Src homologyPF0001851.15101741.8domain 3blastx.2(AJ007012) Fish proteinemb|CAA07416.1|77%10186[Mus musculus]44%201275HOUFT361162680277blastx.14(AF162130) MAGUKgi|6997272|gb|AAD45919.2|100%8681563protein TEM-61 [HomoAF162130_198%422868sapiens]87%31426HOUFT36911293558HMMERPFAM: PDZ domainPF0059535.33225582.1.1(Also known as DHR orGLGF).blastx.2(AF162130) MAGUKgb|AAD45919.2|AF162130_191%196846protein TEM-61 [Homo98%23193sapiens]HAGGF84911312559HMMERPFAM: Eukaryotic proteinPF00069105.85103181.8kinase domainblastx.14calmodulin-dependentgi|3241849|dbj|BAA28870.1|88%10363protein kinase II-delta87%366413dash [Oryctolagus100%320364cuniculus]HTTKP071119031279blastx.14(AL049683) hypotheticalgi|4678753|emb|CAB41255.1|63%8205protein [Homo sapiens]58%263451HTTKP07911390560HMMERPFAM: Src homologyPF0001815.82471961.8domain 3blastx.2(AL049683) hypotheticalemb|CAB41255.1|51%8289protein [Homo sapiens]56%292450HE9SE62911476561HMMERPFAM: Src homologyPF0001847.652684351.8domain 3blastx.2(AK000007) FLJ00007dbj|BAA92232.1|43%4435protein [Homo sapiens]64%877927HUJAD241162674281blastx.14serine/threonine kinasegi|2052191|emb|CAB06295.1|34%457777[Rattus norvegicus]48%36351234%79792857%28536251%14123372%1679173242%97124%7522472%22926142%180236HUJAD24911498562HMMERPFAM: Eukaryotic proteinPF0006934.7392151.8kinase domainblastx.14AMP-activated proteingi|758783|gb|AAA64850.1|45%336467kinase homolog [Homo45%123215sapiens]37%26733854%21124341%4595HWLFG751228123282blastx.14DJ63M2.4 (novel protein).sp|CAC08483|CAC0848381%47286191%8621074100%11401163HWLFG75916563563HMMERPFAM: EF handPF0003624.11872732.1.1blastx.2DJ63M2.4 (novel protein).sp|CAC08483|CAC0848389%720105875%457717100%11231146HT3BG12921593564HMMERPFAM: Eukaryotic proteinPF0006927.091091831.8kinase domainblastx.14CYCLIN-DEPENDENTgi|3715669|emb|CAA03585.1|85%1246KINASE (CDK)8[unidentified]HTLJC71922923284HMMERPFAM: Src homologyPF000189.14115213401.8domain 3blastx.2(AL133030) hypotheticalemb|CAB61362.1|94%31355protein [Homo sapiens]HCOMM051194701285blastx.14epidermal growth factorpir|I38728|I3872844%455721receptor kinase substrate -59%188370human46%72285023%5323236%12519063%10801112HCOMM05925952565HMMERPFAM: Src homologyPF0001859.481783421.8domain 3blastx.2epidermal growth factorgb|AAA62280.1|46%445840receptor kinase substrate43%115435[Homo sapiens]23%43222HSLJE54926924566HMMERPFAM: Pyridoxal-PF0028235.83425362.1.1dependent decarboxylaseconserved domainblastx.2CYSTEINE SULFINICsp|Q9UNJ5|Q9UNJ598%198548ACID92%542739DECARBOXYLASE-85%721885RELATED PROTEIN 4.100%885908HTGED07927411287HMMERPFAM: Sec1 familyPF00995128.9342972.1.1blastx.2VESICLE TRANSPORT-sp|Q9Y6A8|Q9Y6A889%25309RELATED PROTEIN.HOFNH30928365288HMMERPFAM: 7 transmembranePF0000124.5892481.8receptor (rhodopsinfamily)blastx.2CALCIUM-sp|Q9UBY5|Q9UBY575%18263MOBILIZING54%265375LYSOPHOSPHATIDICACID RECEPTOR 1HWNCY051179767289blastx.14GOK [Homo sapiens]gi|2264346|gb|AAC51627.1|65%10951HWNCY05928789567HMMERPFAM: EF handPF0003612.55181011.8blastx.2GOK.sp|Q13586|Q1358660%61292HDPDA47929193290HMMERPFAM: Src homologyPF0001812.526918101.8domain 3blastx.2(AL049683) hypotheticalemb|CAB41255.1|69%1451026protein [Homo sapiens]53%9451022HWMEV63931154291HMMERPFAM: 7 transmembranePF0000153.422622.1.1receptor (rhodopsinfamily)blastx.27 transmembrane G-sp|AAG09275|AAG0927575%2391protein coupled receptor.HCFAT251052857292blastx.14(AF096300) HPK/GCK-gi|4322936|gb|AAD16137.1|72%2412like kinase HGK [Homosapiens]HCFAT25932068568HMMERPFAM: Eukaryotic proteinPF0006926.61362312.1.1kinase domainblastx.14(AF096300) HPK/GCK-gi|4322936|gb|AAD16137.1|63%91456like kinase HGK [Homo72%60158sapiens]25%232312HHEQV391165420293blastx.14(AF037261) SH3-gi|3004948|gb|AAC09244.1|50%517606containing adaptor41%625696molecule-1 [Homo40%396470sapiens]36%141215HHEQV39932851569HMMERPFAM: Src homologyPF0001830.415267081.8domain 3HHFJH79933308570HMMERPFAM: von WillebrandPF0009212.76622321.8factor type A domainblastx.2R31181_2, PARTIALsp|O95783|O9578399%14316PROTEIN(FRAGMENT).HUCOW171155190295blastx.2RHOGAP PROTEIN.sp|Q98935|Q9893561%444162%787987HUCOW17933357571HMMERPFAM: Src homologyPF0001820.286477391.8domain 3blastx.2Graf protein [Homoemb|CAA71414.2|67%1261sapiens]50%60875183%75680940%187246HFKIT06934019572HMMERPFAM: Eukaryotic proteinPF0006934.651602701.8kinase domainblastx.14p58 galactosyltransferase-pir|A38282|A3828251%178270associated protein kinase -40%74118humanHDTBY881104159297blastx.14(AF130372) serine-gi|7108631|gb|AAF36509.1|87%186491threonine protein kinase 1AF130372_198%3170100%497535HDTBY88934472573HMMERPFAM: Eukaryotic proteinPF0006993.633022.1.1kinase domainblastx.14p56 KKIAMRE proteingi|1517820|gb|AAC50918.1|82%3170kinase [Homo sapiens]35%192458100%492509HWLHS821082268298blastx.2(AC005581) R31237_1,gb|AAC33487.1|93%162902partial CDS [Homo88%10491237sapiens]100%96170HWLHS82934505574HMMERPFAM: Eukaryotic proteinPF00069147.223192.1.1kinase domainblastx.2(AC005581) R31237_1,gb|AAC33487.1|90%68364partial CDS [Homo100%276sapiens]40%306422HDPNC961081629299blastx.14HUMAN NDRgi|2304746|emb|CAA03387.1|92%3734[unidentified]HDPNC96934520575HMMERPFAM: Eukaryotic proteinPF00069206.6337341.8kinase domainblastx.14HUMAN NDRgi|2304746|emb|CAA03387.1|92%3734[unidentified]HCE5I781197899300blastx.14rabphilin-3A - bovinepir|A48097|A4809795%191112668%1197HCE5I78934531576HMMERPFAM: C2 domainPF0016849.142134131.8blastx.2rabphilin-3A - bovinepir|A48097|A4809783%13540461%341HISDS621159625301blastx.14(AJ250425) Collybistin Igi|6706318|emb|CAB65966.1|90%185892[Rattus norvegicus]HISDS62935932577HMMERPFAM: RhoGEF domainPF0062151.32294862.1.1blastx.2(AJ250425) Collybistin Iemb|CAB65966.1|96%1483[Rattus norvegicus]HDQDV69937850578HMMERPFAM: Eukaryotic proteinPF00069212.5685982.1.1kinase domainblastx.2(AF169035) proteingb|AAF12758.1|AF169035_198%68829kinase [Homo sapiens]HEMBT61939957303HMMERPFAM: Eukaryotic proteinPF0006976.6162852.1.1kinase domainblastx.2(AD000092) hypotheticalgb|AAB51171.1|71%13441human serine-threonineprotein kinase R31240_1[Homo sapiens]HRODZ701088554304blastx.2kinase like proteinemb|CAB10257.1|39%254544[Arabidopsis thaliana]50%524601HRODZ70942673580HMMERPFAM: Eukaryotic proteinPF0006978.2332482.1.1kinase domainblastx.2kinase like proteinemb|CAB10257.1|39%33323[Arabidopsis thaliana]50%303380HHERQ79944057581HMMERPFAM: Eukaryotic proteinPF0006983.41334741.8kinase domainblastx.2(AB016589) inducibledbj|BAA85154.1|90%109471IKappaB kinase [Musmusculus]HCECM90945088582HMMERPFAM: Src homologyPF0001853.063925681.8domain 3HWHGW72945692583HMMERPFAM: ATP P2X receptorPF00864438.52478552.1.1blastx.2(AF190822) P2X2Agb|AAF19170.1|AF190822_191%190939receptor [Homo sapiens]HPCRV841219890308blastx.14MATERNALsp|Q61846|Q6184694%138839EMBRYONIC LEUCINEZIPPER KINASE(SERINE/THREONINE 1HPCRV84945856585HMMERPFAM: Eukaryotic proteinPF0006975.571573841.8kinase domainblastx.2similar to protein kinasedbj|BAA11492.1|78%127483of X. laevis, has putative 1HNSAA28946988309HMMERPFAM: SH3 domainPF000181497579152.1.1blastx.2(AF146277) adaptergb|AAD34595.1|AF146277_182%41554protein CMS [Homosapiens]HNSAA28972348586blastx.14(AF146277) adaptergi|4960047|gb|AAD34595.1|88%21449protein CMS [HomoAF146277_1sapiens]HLWAR77947484310HMMERPFAM: 7 transmembranePF00001214.212875531.8receptor (rhodopsinfamily)blastx.2G-protein coupledsp|AAF87078|AAF87078100%1287292receptor HLWAR77.HTTJW491127477311blastx.14(AF118838) citrin; adult-gi|5052319|gb|AAD38501.1|94%5161244onset type II citrullinemiaAF118838_197%101532protein [Homo sapiens]90%12321381100%1408150037%1244137840%1411147640%1200124460%1143117233%38348133%597695HTTJW49948107587HMMERPFAM: EF handPF0003611.982833481.8blastx.2CITRIN.sp|Q9UNI7|Q9UNI784%94627HWAFS181155193312blastx.14(AF156884) RIP-likegi|5059425|gb|AAD39005.1|89%1651718kinase [Homo sapiens]AF156884_1HWAFS18948434588HMMERPFAM: Eukaryotic proteinPF00069115.982256321.8kinase domainblastx.14(AF156884) RIP-likegi|5059425|gb|AAD39005.1|91%165632kinase [Homo sapiens]AF156884_166%702773100%632661HFCBA441082762313blastx.14(AB010633)gi|2810987|dbj|BAA24523.1|61%184639carboxylesterase precursor[Macaca fascicularis]HFCBA44948533589HMMERPFAM: CarboxylesterasesPF0013534.243154851.8blastx.2thiolesterase B (EC 3.—.—.—)pir|A47162|A4716256%2208precursor - mallard60%42348248%184264HVADT771180374314blastx.14alpha-3 collagen type VIgi|211622|gb|AAA03201.1|55%12119[Gallus gallus]HVADT77948886590HMMERPFAM: Kunitz/BovinePF0001474.411693211.8pancreatic trypsininhibitor domainblastx.2alpha-3 collagen type VIgb|AAA03201.1|43%130330[Gallus gallus]HUFCN911189013315blastx.14copine I [Homo sapiens]gi|1791257|gb|AAC15920.1|54%10354048%56178271%87493675%54058764%783833HUFCN91949137591HMMERPFAM: von WillebrandPF000929.835297291.8factor type A domainblastx.14copine I [Homo sapiens]gi|1791257|gb|AAC15920.1|56%10358848%971133348%56278357%1332143045%899100964%784834HAGBX32951351316HMMERPFAM: PMP-PF00822182.334762.1.122/EMP/MP20/Claudinfamilyblastx.2VOLTAGE-sp|O60359|CCG3_HUMAN89%12551DEPENDENTCALCIUM CHANNELGAMMA-3 SUBUNIT 1HWMIB81955336593HMMERPFAM: Eukaryotic proteinPF00069122.8514589341.8kinase domainblastx.2(AK000528) unnameddbj|BAA91232.1|100%3572protein product [Homosapiens]HCEMU861156430318blastx.14p87 = transporter-likegi|259174|gb|AAB24028.1|91%10795protein [cattle, Peptide,742 aa] [Bos taurus]HCEMU86956864594HMMERPFAM: Sugar (and other)PF0008324.49212417381.8transportersblastx.2p87 = transporter-likegb|AAB24028.1|99%3788protein [cattle, Peptide,742 aa] [Bos taurus]HRDAF831153911319blastx.14(AF137378) integrin alphagi|5915662|gb|AAD51919.2|93%4233811 subunit precursorAF137378_1[Homo sapiens]HRDAF83957143595HMMERPFAM: von WillebrandPF0009289.99723321.8factor type A domainblastx.2(AF137378) integrin alphagb|AAD51919.2|AF137378_192%3633211 subunit precursor[Homo sapiens]HUVGZ881204719320blastx.14PRO1038.sp|AAF71042|AAF7104250%52798844%974118350%44459330%73285175%713736HUVGZ88959020597HMMERPFAM: Eukaryotic proteinPF0006931.121824391.8kinase domainHSCKS55961074598HMMERPFAM: C2 domainPF001684.419319931.8HOEET48963290322HMMERPFAM: EF handPF00036266567272.1.1blastx.2Reticulocabin precursor.sp|AAG09692|AAG0969296%471030HBODE51964235599HMMERPFAM: MitochondrialPF00153235.2699518341.8carrier proteinsblastx.14aralar1 [Homo sapiens]gi|3559910|emb|CAA74834.1|93%202053HHFCK09965304324HMMERPFAM: TBC domainPF00566179.1230516552.1.1blastx.2(AL022238) dJ1042K10.2emb|CAA18266.1|97%26351268(supported by98%1276389GENSCAN, FGENES andGENEWISE) [Homosapiens]HCOOZ11965306600HMMERPFAM: Src homologyPF000185.221792141.8domain 3blastx.2(AL022238) dJ1042K10.2emb|CAA18266.1|100%182589(supported byGENSCAN, FGENES andGENEWISE) [Homosapiens]HDPPO351119032326blastx.14(AL049683) hypotheticalgi|4678753|emb|CAB41255.1|63%561758protein [Homo sapiens]65%81698071%8414665%30035945%1080113925%117245HDPPO35966248601HMMERPFAM: Src homologyPF0001814.076007491.8domain 3blastx.2(AL049683) hypotheticalemb|CAB41255.1|39%841148protein [Homo sapiens]HLWDZ53968602602HMMERPFAM: ActinPF00022291.17711112.1.1blastx.2Actin-related protein 3-sp|AAC98904|AAC9890499%951126beta.100%5498HEOPL36968826603HMMERPFAM: Src homologyPF0001879.813164831.8domain 3blastx.2(AL049758) dJ437M21.3emb|CAB51395.1|99%178486(protein kinase C andcasein kinase substrate inneurons 2) [Homosapiens]HMCFS021152252329blastx.14(AK000482) unnamedgi|7020600|dbj|BAA91194.1|59%72725protein product [Homo40%716781sapiens]HMCFS02969326604HMMERPFAM: C2 domainPF001688.053474571.8blastx.2CDNA FLJ20475 FIS,sp|BAA91194|BAA9119459%116496CLONE KAT07206.HDPSR15969666605HMMERPFAM: Eukaryotic proteinPF0006987.193516261.8kinase domainblastx.2(AB026289) proteindbj|BAA85045.1|95%6311158kinase SID6-1512 [Homo89%240692sapiens]HNTAV78971315606HMMERPFAM: 7 transmembranePF0000123.9231431.8receptor (rhodopsinfamily)blastx.2Cysteinyl leukotrienesp|BAB03601|BAB03601100%3266CysLT2 receptor.HFKDR141145842332blastx.14(AF128625) CDC42-gi|5006445|gb|AAD37506.1|98%901277binding protein kinaseAF128625_1100%12791320beta [Homo sapiens]HFKDR14974255607HMMERPFAM: Eukaryotic proteinPF00069244.2129710971.8kinase domainblastx.2(AF128625) CDC42-gb|AAD37506.1|AF128625_198%721733binding protein kinase22%15721706beta [Homo sapiens]HDPBI30974711333HMMERPFAM: 7 transmembranePF00001171.3138610961.8receptor (rhodopsinfamily)blastx.2G PROTEIN-COUPLEDsp|Q9UNW8|Q9UNW893%2061312RECEPTOR.HODFF881094875334blastx.2mixed-lineage proteinpir|S32467|JU022973%89493kinase 1 - human81%763969HODFF88974911608HMMERPFAM: Eukaryotic proteinPF00069101.43983701.8kinase domainblastx.14mixed-lineage proteinpir|S32467|JU022974%131493kinase 1 - human81%76392130%751915

[0049] Table 2 further characterizes certain encoded polypeptides of the invention, by providing the results of comparisons to protein and protein family databases. The first column provides a unique clone identifier, “Clone ID NO:”, corresponding to a cDNA clone disclosed in Table 1A. The second column provides the unique contig identifier, “Contig ID:” which allows correlation with the information in Table 1A. The third column provides the sequence identifier, “SEQ ID NO:”, for the contig polynucleotide sequences. The fourth column provides the analysis method by which the homology/identity disclosed in the Table was determined. The fifth column provides a description of the PFAM/NR hit identified by each analysis. Column six provides the accession number of the PFAM/NR hit disclosed in the fifth column. Column seven, score/percent identity, provides a quality score or the percent identity, of the hit disclosed in column five. Comparisons were made between polypeptides encoded by polynucleotides of the invention and a non-redundant protein database (herein referred to as “NR”), or a database of protein families (herein referred to as “PFAM”), as described below.

[0050] The NR database, which comprises the NBRF PIR database, the NCBI GenPept database, and the SIB SwissProt and TrEMBL databases, was made non-redundant using the computer program nrdb2 (Warren Gish, Washington University in Saint Louis). Each of the polynucleotides shown in Table 1A, column 3 (e.g., SEQ ID NO:X or the ‘Query’ sequence) was used to search against the NR database. The computer program BLASTX was used to compare a 6-frame translation of the Query sequence to the NR database (for information about the BLASTX algorithm please see Altshul et al., J. Mol. Biol. 215:403-410 (1990); and Gish and States, Nat. Genet. 3:266-272 (1993). A description of the sequence that is most similar to the Query sequence (the highest scoring ‘Subject’) is shown in column five of Table 2 and the database accession number for that sequence is provided in column six. The highest scoring ‘Subject’ is reported in Table 2 if (a) the estimated probability that the match occurred by chance alone is less than 1.0e-07, and (b) the match was not to a known repetitive element. BLASTX returns alignments of short polypeptide segments of the Query and Subject sequences which share a high degree of similarity; these segments are known as High-Scoring Segment Pairs or HSPs. Table 2 reports the degree of similarity between the Query and the Subject for each HSP as a percent identity in Column 7. The percent identity is determined by dividing the number of exact matches between the two aligned sequences in the HSP, dividing by the number of Query amino acids in the HSP and multiplying by 100. The polynucleotides of SEQ ID NO:X which encode the polypeptide sequence that generates an HSP are delineated by columns 8 and 9 of Table 2.

[0051] The PFAM database, PFAM version 2.1, (Sonnhammer et al., Nucl. Acids Res., 26:320-322, 1998)) consists of a series of multiple sequence alignments; one alignment for each protein family. Each multiple sequence alignment is converted into a probability model called a Hidden Markov Model, or HMM, that represents the position-specific variation among the sequences that make up the multiple sequence alignment (see, e.g., Durbin et al., Biological sequence analysis. probabilistic models of proteins and nucleic acids, Cambridge University Press, 1998 for the theory of HMMs). The program HMMER version 1.8 (Sean Eddy, Washington University in Saint Louis) was used to compare the predicted protein sequence for each Query sequence (SEQ ID NO:Y in Table 1A) to each of the HMMs derived from PFAM version 2.1. A HMM derived from PFAM version 2.1 was said to be a significant match to a polypeptide of the invention if the score returned by HMMER 1.8 was greater than 0.8 times the HMMER 1.8 score obtained with the most distantly related known member of that protein family. The description of the PFAM family which shares a significant match with a polypeptide of the invention is listed in column 5 of Table 2, and the database accession number of the PFAM hit is provided in column 6. Column 7 provides the score returned by HMMER version 1.8 for the alignment. Columns 8 and 9 delineate the polynucleotides of SEQ ID NO:X which encode the polypeptide sequence which show a significant match to a PFAM protein family.

[0052] As mentioned, columns 8 and 9 in Table 2, “NT From” and “NT To”, delineate the polynucleotides of “SEQ ID NO:X” that encode a polypeptide having a significant match to the PFAM/NR database as disclosed in the fifth column. In one embodiment, the invention provides a protein comprising, or alternatively consisting of, a polypeptide encoded by the polynucleotides of SEQ ID NO:X delineated in columns 8 and 9 of Table 2. Also provided are polynucleotides encoding such proteins, and the complementary strand thereto.

[0053] The nucleotide sequence SEQ ID NO:X and the translated SEQ ID NO:Y are sufficiently accurate and otherwise suitable for a variety of uses well known in the art and described further below. For instance, the nucleotide sequences of SEQ ID NO:X are useful for designing nucleic acid hybridization probes that will detect nucleic acid sequences contained in SEQ ID NO:X or the cDNA contained in Clone ID NO:Z. These probes will also hybridize to nucleic acid molecules in biological samples, thereby enabling immediate applications in chromosome mapping, linkage analysis, tissue identification and/or typing, and a variety of forensic and diagnostic methods of the invention. Similarly, polypeptides identified from SEQ ID NO:Y may be used to generate antibodies which bind specifically to these polypeptides, or fragments thereof, and/or to the polypeptides encoded by the cDNA clones identified in, for example, Table 1A.

[0054] Nevertheless, DNA sequences generated by sequencing reactions can contain sequencing errors. The errors exist as misidentified nucleotides, or as insertions or deletions of nucleotides in the generated DNA sequence. The erroneously inserted or deleted nucleotides cause frame shifts in the reading frames of the predicted amino acid sequence. In these cases, the predicted amino acid sequence diverges from the actual amino acid sequence, even though the generated DNA sequence may be greater than 99.9% identical to the actual DNA sequence (for example, one base insertion or deletion in an open reading frame of over 1000 bases).

[0055] Accordingly, for those applications requiring precision in the nucleotide sequence or the amino acid sequence, the present invention provides not only the generated nucleotide sequence identified as SEQ ID NO:X, and a predicted translated amino acid sequence identified as SEQ ID NO:Y, but also a sample of plasmid DNA containing cDNA Clone ID NO:Z (deposited with the ATCC on Oct. 5, 2000, and receiving ATCC designation numbers PTA 2574 and PTA 2575; deposited with the ATCC on Jan. 5, 2001, and having depositor reference numbers TS-1, TS-2, AC-1, and AC-2; and/or as set forth, for example, in Table 1A, 6 and 7). The nucleotide sequence of each deposited clone can readily be determined by sequencing the deposited clone in accordance with known methods. Further, techniques known in the art can be used to verify the nucleotide sequences of SEQ ID NO:X.

[0056] The predicted amino acid sequence can then be verified from such deposits. Moreover, the amino acid sequence of the protein encoded by a particular clone can also be directly determined by peptide sequencing or by expressing the protein in a suitable host cell containing the deposited human cDNA, collecting the protein, and determining its sequence.

[0057] RACE Protocol For Recovery of Full-Length Genes

[0058] Partial cDNA clones can be made full-length by utilizing the rapid amplification of cDNA ends (RACE) procedure described in Frohman, M. A., et al., Proc. Nat'l. Acad. Sci. USA, 85:8998-9002 (1988). A cDNA clone missing either the 5′ or 3′ end can be reconstructed to include the absent base pairs extending to the translational start or stop codon, respectively. In some cases, cDNAs are missing the start codon of translation, therefor. The following briefly describes a modification of this original 5′ RACE procedure. Poly A+ or total RNA is reverse transcribed with Superscript II (Gibco/BRL) and an antisense or complementary primer specific to the cDNA sequence. The primer is removed from the reaction with a Microcon Concentrator (Amicon). The first-strand cDNA is then tailed with dATP and terminal deoxynucleotide transferase (Gibco/BRL). Thus, an anchor sequence is produced which is needed for PCR amplification. The second strand is synthesized from the dA-tail in PCR buffer, Taq DNA polymerase (Perkin-Elmer Cetus), an oligo-dT primer containing three adjacent restriction sites (XhoI, SalI and ClaI) at the 5′ end and a primer containing just these restriction sites. This double-stranded cDNA is PCR amplified for 40 cycles with the same primers as well as a nested cDNA-specific antisense primer. The PCR products are size-separated on an ethidium bromide-agarose gel and the region of gel containing cDNA products the predicted size of missing protein-coding DNA is removed. cDNA is purified from the agarose with the Magic PCR Prep kit (Promega), restriction digested with XhoI or SalI, and ligated to a plasmid such as pBluescript SKII (Stratagene) at XhoI and EcoRV sites. This DNA is transformed into bacteria and the plasmid clones sequenced to identify the correct protein-coding inserts. Correct 5′ ends are confirmed by comparing this sequence with the putatively identified homologue and overlap with the partial cDNA clone. Similar methods known in the art and/or commercial kits are used to amplify and recover 3′ ends.

[0059] Several quality-controlled kits are commercially available for purchase. Similar reagents and methods to those above are supplied in kit form from Gibco/BRL for both 5′ and 3′ RACE for recovery of full length genes. A second kit is available from Clontech which is a modification of a related technique, SLIC (single-stranded ligation to single-stranded cDNA), developed by Dumas et al., Nucleic Acids Res., 19:5227-32 (1991). The major differences in procedure are that the RNA is alkaline hydrolyzed after reverse transcription and RNA ligase is used to join a restriction site-containing anchor primer to the first-strand cDNA. This obviates the necessity for the dA-tailing reaction which results in a polyT stretch that is difficult to sequence past.

[0060] An alternative to generating 5′ or 3′ cDNA from RNA is to use cDNA library double-stranded DNA. An asymmetric PCR-amplified antisense cDNA strand is synthesized with an antisense cDNA-specific primer and a plasmid-anchored primer. These primers are removed and a symmetric PCR reaction is performed with a nested cDNA-specific antisense primer and the plasmid-anchored primer.

[0061] RNA Ligase Protocol for Generating The 5′ or 3′ End Sequences to Obtain Full Length Genes

[0062] Once a gene of interest is identified, several methods are available for the identification of the 5′ or 3′ portions of the gene which may not be present in the original cDNA plasmid. These methods include, but are not limited to, filter probing, clone enrichment using specific probes and protocols similar and identical to 5′ and 3′ RACE. While the full length gene may be present in the library and can be identified by probing, a useful method for generating the 5′ or 3′ end is to use the existing sequence information from the original cDNA to generate the missing information. A method similar to 5′ RACE is available for generating the missing 5′ end of a desired full-length gene. (This method was published by Fromont-Racine et al., Nucleic Acids Res., 21(7):1683-1684 (1993)). Briefly, a specific RNA oligonucleotide is ligated to the 5′ ends of a population of RNA presumably containing full-length gene RNA transcript and a primer set containing a primer specific to the ligated RNA oligonucleotide and a primer specific to a known sequence of the gene of interest, is used to PCR amplify the 5′ portion of the desired full length gene which may then be sequenced and used to generate the full length gene. This method starts with total RNA isolated from the desired source, poly A RNA may be used but is not a prerequisite for this procedure. The RNA preparation may then be treated with phosphatase if necessary to eliminate 5′ phosphate groups on degraded or damaged RNA which may interfere with the later RNA ligase step. The phosphatase if used is then inactivated and the RNA is treated with tobacco acid pyrophosphatase in order to remove the cap structure present at the 5′ ends of messenger RNAs. This reaction leaves a 5′ phosphate group at the 5′ end of the cap cleaved RNA which can then be ligated to an RNA oligonucleotide using T4 RNA ligase. This modified RNA preparation can then be used as a template for first strand cDNA synthesis using a gene specific oligonucleotide. The first strand synthesis—reaction can then be used as a template for PCR amplification of the desired 5′ end using a primer specific to the ligated RNA oligonucleotide and a primer specific to the known sequence of the gene of interest. The resultant product is then sequenced and analyzed to confirm that the 5′ end sequence belongs to the relevant gene.

[0063] The present invention also relates to vectors or plasmids which include such DNA sequences, as well as the use of the DNA sequences. The material deposited with the ATCC (deposited with the ATCC on Oct. 5, 2000, and receiving ATCC designation numbers PTA 2574 and PTA 2575; deposited with the ATCC on Jan. 5, 2001, and receiving ATCC designation numbers TS-1, TS-2, AC-1, and AC-2; and/or as set forth, for example, in Table 1A, Table 6, or Table 7) is a mixture of cDNA clones derived from a variety of human tissue and cloned in either a plasmid vector or a phage vector, as described, for example, in Table 7. These deposits are referred to as “the deposits” herein. The tissues from which some of the clones were derived are listed in Table 7, and the vector in which the corresponding cDNA is contained is also indicated in Table 7. The deposited material includes cDNA clones corresponding to SEQ ID NO:X described, for example, in Table 1A (Clone ID NO:Z). A clone which is isolatable from the ATCC Deposits by use of a sequence listed as SEQ ID NO:X, may include the entire coding region of a human gene or in other cases such clone may include a substantial portion of the coding region of a human gene. Furthermore, although the sequence listing may in some instances list only a portion of the DNA sequence in a clone included in the ATCC Deposits, it is well within the ability of one skilled in the art to sequence the DNA included in a clone contained in the ATCC Deposits by use of a sequence (or portion thereof) described in, for example Tables 1A or 2 by procedures hereinafter further described, and others apparent to those skilled in the art.

[0064] Also provided in Table 7 is the name of the vector which contains the cDNA clone. Each vector is routinely used in the art. The following additional information is provided for convenience.

[0065] Vectors Lambda Zap (U.S. Pat. Nos. 5,128,256 and 5,286,636), Uni-Zap XR (U.S. Pat. Nos. 5,128,256 and 5,286,636), Zap Express (U.S. Pat. Nos. 5,128,256 and 5,286,636), pBluescript (pBS) (Short, J. M. et al., Nucleic Acids Res. 16:7583-7600 (1988); Alting-Mees, M. A. and Short, J. M., Nucleic Acids Res. 17:9494 (1989)) and pBK (Alting-Mees, M. A. et al., Strategies 5:58-61 (1992)) are commercially available from Stratagene Cloning Systems, Inc., 11011 N. Torrey Pines Road, La Jolla, Calif., 92037. pBS contains an ampicillin resistance gene and pBK contains a neomycin resistance gene. Phagemid pBS may be excised from the Lambda Zap and Uni-Zap XR vectors, and phagemid pBK may be excised from the Zap Express vector. Both phagemids may be transformed into E. coli strain XL-1 Blue, also available from Stratagene.

[0066] Vectors pSport1, pCMVSport 1.0, pCMVSport 2.0 and pCMVSport 3.0, were obtained from Life Technologies, Inc., P. O. Box 6009, Gaithersburg, Md. 20897. All Sport vectors contain an ampicillin resistance gene and may be transformed into E. coli strain DH10B, also available from Life Technologies. See, for instance, Gruber, C. E., et al., Focus 15:59-(1993). Vector lafmid BA (Bento Soares, Columbia University, New York, N.Y.) contains an ampicillin resistance gene and can be transformed into E. coli strain XL-1 Blue. Vector pCR®2.1, which is available from Invitrogen, 1600 Faraday Avenue, Carlsbad, Calif. 92008, contains an ampicillin resistance gene and may be transformed into E. coli strain DH10B, available from Life Technologies. See, for instance, Clark, J. M., Nuc. Acids Res. 16:9677-9686 (1988) and Mead, D. et al., Bio/Technology 9: (1991).

[0067] The present invention also relates to the genes corresponding to SEQ ID NO:X, SEQ ID NO:Y, and/or the deposited clone (Clone ID NO:Z). The corresponding gene can be isolated in accordance with known methods using the sequence information disclosed herein. Such methods include preparing probes or primers from the disclosed sequence and identifying or amplifying the corresponding gene from appropriate sources of genomic material.

[0068] Also provided in the present invention are allelic variants, orthologs, and/or species homologs. Procedures known in the art can be used to obtain full-length genes, allelic variants, splice variants, full-length coding portions, orthologs, and/or species homologs of genes corresponding to SEQ ID NO:X or the complement thereof, polypeptides encoded by genes corresponding to SEQ ID NO:X or the complement thereof, and/or the cDNA contained in Clone ID NO:Z, using information from the sequences disclosed herein or the clones deposited with the ATCC. For example, allelic variants and/or species homologs may be isolated and identified by making suitable probes or primers from the sequences provided herein and screening a suitable nucleic acid source for allelic variants and/or the desired homologue.

[0069] The polypeptides of the invention can be prepared in any suitable manner. Such polypeptides include isolated naturally occurring polypeptides, recombinantly produced polypeptides, synthetically produced polypeptides, or polypeptides produced by a combination of these methods. Means for preparing such polypeptides are well understood in the art.

[0070] The polypeptides may be in the form of the secreted protein, including the mature form, or may be a part of a larger protein, such as a fusion protein (see below). It is often advantageous to include an additional amino acid sequence which contains secretory or leader sequences, pro-sequences, sequences which aid in purification, such as multiple histidine residues, or an additional sequence for stability during recombinant production.

[0071] The polypeptides of the present invention are preferably provided in an isolated form, and preferably are substantially purified. A recombinantly produced version of a polypeptide, including the secreted polypeptide, can be substantially purified using techniques described herein or otherwise known in the art, such as, for example, by the one-step method described in Smith and Johnson, Gene 67:31-40 (1988). Polypeptides of the invention also can be purified from natural, synthetic or recombinant sources using techniques described herein or otherwise known in the art, such as, for example, antibodies of the invention raised against the polypeptides of the present invention in methods which are well known in the art.

[0072] The present invention provides a polynucleotide comprising, or alternatively consisting of, the nucleic acid sequence of SEQ ID NO:X, and/or the cDNA sequence contained in Clone ID NO:Z. The present invention also provides a polypeptide comprising, or alternatively, consisting of, the polypeptide sequence of SEQ ID NO:Y, a polypeptide encoded by SEQ ID NO:X or a complement thereof, a polypeptide encoded by the cDNA contained in Clone ID NO:Z, and/or the polypeptide sequence encoded by a nucleotide sequence in SEQ ID NO:B as defined in column 6 of Table 1B. Polynucleotides encoding a polypeptide comprising, or alternatively consisting of the polypeptide sequence of SEQ ID NO:Y, a polypeptide encoded by SEQ ID NO:X, a polypeptide encoded by the cDNA contained in Clone ID NO:Z, and/or a polypeptide sequence encoded by a nucleotide sequence in SEQ ID NO:B as defined in column 6 of Table 1B are also encompassed by the invention. The present invention further encompasses a polynucleotide comprising, or alternatively consisting of, the complement of the nucleic acid sequence of SEQ ID NO:X, a nucleic acid sequence encoding a polypeptide encoded by the complement of the nucleic acid sequence of SEQ ID NO:X, and/or the cDNA contained in Clone ID NO:Z.

[0073] Moreover, representative examples of polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more of the sequences delineated in Table 1B column 6, or any combination thereof. Additional, representative examples of polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more of the complementary strand(s) of the sequences delineated in Table 1B column 6, or any combination thereof. In further embodiments, the above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated in Table 1B, column 6, and have a nucleic acid sequence which is different from that of the BAC fragment having the sequence disclosed in SEQ ID NO:B (see Table 1B, column 5). In additional embodiments, the above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated in Table 1B, column 6, and have a nucleic acid sequence which is different from that published for the BAC clone identified as BAC ID NO:A (see Table 1B, column 4). In additional embodiments, the above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated in Table 1B, column 6, and have a nucleic acid sequence which is different from that contained in the BAC clone identified as BAC ID NO:A (see Table 1B, column 4). Polypeptides encoded by these polynucleotides, other polynucleotides that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides and polypeptides are also encompassed by the invention.

[0074] Further, representative examples of polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more of the sequences delineated in column 6 of Table 1B which correspond to the same Clone ID NO:Z (see Table 1B, column 1), or any combination thereof. Additional, representative examples of polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more of the complementary strand(s) of the sequences delineated in column 6 of Table 1B which correspond to the same Clone ID NO:Z (see Table 1B, column 1), or any combination thereof. In further embodiments, the above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated in column 6 of Table 1B which correspond to the same Clone ID NO:Z (see Table 1B, column 1) and have a nucleic acid sequence which is different from that of the BAC fragment having the sequence disclosed in SEQ ID NO:B (see Table 1B, column 5). In additional embodiments, the above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated in column 6 of Table 1B which correspond to the same Clone ID NO:Z (see Table 1B, column 1) and have a nucleic acid sequence which is different from that published for the BAC clone identified as BAC ID NO:A (see Table 1B, column 4). In additional embodiments, the above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated in column 6 of Table 1B which correspond to the same Clone ID NO:Z (see Table 1B, column 1) and have a nucleic acid sequence which is different from that contained in the BAC clone identified as BAC ID NO:A (see Table 1B, column 4). Polypeptides encoded by these polynucleotides, other polynucleotides that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides and polypeptides are also encompassed by the invention.

[0075] Further, representative examples of polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more of the sequences delineated in column 6 of Table 1B which correspond to the same contig sequence identifer SEQ ID NO:X (see Table 1B, column 2), or any combination thereof. Additional, representative examples of polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more of the complementary strand(s) of the sequences delineated in column 6 of Table 1B which correspond to the same contig sequence identifer SEQ ID NO:X (see Table 1B, column 2), or any combination thereof. In further embodiments, the above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated in column 6 of Table 1B which correspond to the same contig sequence identifer SEQ ID NO:X (see Table 1B, column 2) and have a nucleic acid sequence which is different from that of the BAC fragment having the sequence disclosed in SEQ ID NO:B (see Table 1B, column 5). In additional embodiments, the above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated in column 6 of Table 1B which correspond to the same contig sequence identifer SEQ ID NO:X (see Table 1B, column 2) and have a nucleic acid sequence which is different from that published for the BAC clone identified as BAC ID NO:A (see Table 1B, column 4). In additional embodiments, the above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated in column 6 of Table 1B which correspond to the same contig sequence identifer SEQ ID NO:X (see Table 1B, column 2) and have a nucleic acid sequence which is different from that contained in the BAC clone identified as BAC ID NO:A (See Table 1B, column 4). Polypeptides encoded by these polynucleotides, other polynucleotides that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides and polypeptides are also encompassed by the invention.

[0076] Moreover, representative examples of polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more of the sequences delineated in the same row of Table 1B column 6, or any combination thereof. Additional, representative examples of polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more of the complementary strand(s) of the sequences delineated in the same row of Table 1B column 6, or any combination thereof. In preferred embodiments, the polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more of the complementary strand(s) of the sequences delineated in the same row of Table 1B column 6, wherein sequentially delineated sequences in the table (i.e. corresponding to those exons located closest to each other) are directly contiguous in a 5′ to 3′ orientation. In further embodiments, above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated in the same row of Table 1B, column 6, and have a nucleic acid sequence which is different from that of the BAC fragment having the sequence disclosed in SEQ ID NO:B (see Table 1B, column 5). In additional embodiments, the above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated in the same row of Table 1B, column 6, and have a nucleic acid sequence which is different from that published for the BAC clone identified as BAC ID NO:A (see Table 1B, column 4). In additional embodiments, the above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated in the same row of Table 1B, column 6, and have a nucleic acid sequence which is different from that contained in the BAC clone identified as BAC ID NO:A (see Table 1B, column 4). Polypeptides encoded by these polynucleotides, other polynucleotides that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention.

[0077] In additional specific embodiments, polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more of the sequences delineated in column 6 of Table 1B, and the polynucleotide sequence of SEQ ID NO:X (e.g., as defined in Table 1B, column 2) or fragments or variants thereof. Polypeptides encoded by these polynucleotides, other polynucleotides that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention.

[0078] In additional specific embodiments, polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more of the sequences delineated in column 6 of Table 1B which correspond to the same Clone ID NO:Z (see Table 1B, column 1), and the polynucleotide sequence of SEQ ID NO:X (e.g., as defined in Table 1A or 1B) or fragments or variants thereof. In preferred embodiments, the delineated sequence(s) and polynucleotide sequence of SEQ ID NO:X correspond to the same Clone ID NO:Z. Polypeptides encoded by these polynucleotides, other polynucleotides that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention.

[0079] In further specific embodiments, polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more of the sequences delineated in the same row of column 6 of Table 1B, and the polynucleotide sequence of SEQ ID NO:X (e.g., as defined in Table 1A or 1B) or fragments or variants thereof. In preferred embodiments, the delineated sequence(s) and polynucleotide sequence of SEQ ID NO:X correspond to the same row of column 6 of Table 1B. Polypeptides encoded by these polynucleotides, other polynucleotides that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention.

[0080] In additional specific embodiments, polynucleotides of the invention comprise, or alternatively consist of a polynucleotide sequence in which the 3′ 10 polynucleotides of one of the sequences delineated in column 6 of Table 1B and the 5′ 10 polynucleotides of the sequence of SEQ ID NO:X are directly contiguous. Nucleic acids which hybridize to the complement of these 20 contiguous polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention. Polypeptides encoded by these polynucleotides and/or nucleic acids, other polynucleotides and/or nucleic acids that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides, nucleic acids, and polypeptides are also encompassed by the invention.

[0081] In additional specific embodiments, polynucleotides of the invention comprise, or alternatively consist of, a polynucleotide sequence in which the 3′ 10 polynucleotides of one of the sequences delineated in column 6 of Table 1B and the 5′ 10 polynucleotides of a fragment or variant of the sequence of SEQ ID NO:X are directly contiguous Nucleic acids which hybridize to the complement of these 20 contiguous polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention. Polypeptides encoded by these polynucleotides and/or nucleic acids, other polynucleotides and/or nucleic acids encoding these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides, nucleic acids, and polypeptides are also encompassed by the invention.

[0082] In specific embodiments, polynucleotides of the invention comprise, or alternatively consist of, a polynucleotide sequence in which the 3′ 10 polynucleotides of the sequence of SEQ ID NO:X and the 5′ 10 polynucleotides of the sequence of one of the sequences delineated in column 6 of Table 1B are directly contiguous. Nucleic acids which hybridize to the complement of these 20 contiguous polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention. Polypeptides encoded by these polynucleotides and/or nucleic acids, other polynucleotides and/or nucleic acids encoding these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides, nucleic acids, and polypeptides are also encompassed by the invention.

[0083] In specific embodiments, polynucleotides of the invention comprise, or alternatively consist of, a polynucleotide sequence in which the 3′ 10 polynucleotides of a fragment or variant of the sequence of SEQ ID NO:X and the 5′ 10 polynucleotides of the sequence of one of the sequences delineated in column 6 of Table 1B are directly contiguous. Nucleic acids which hybridize to the complement of these 20 contiguous polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention. Polypeptides encoded by these polynucleotides and/or nucleic acids, other polynucleotides and/or nucleic acids encoding these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides, nucleic acids, and polypeptides, are also encompassed by the invention.

[0084] In further specific embodiments, polynucleotides of the invention comprise, or alternatively consist of, a polynucleotide sequence in which the 3′ 10 polynucleotides of one of the sequences delineated in column 6 of Table 1B and the 5′ 10 polynucleotides of another sequence in column 6 are directly contiguous. Nucleic acids which hybridize to the complement of these 20 contiguous polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention. Polypeptides encoded by these polynucleotides and/or nucleic acids, other polynucleotides and/or nucleic acids encoding these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides, nucleic acids, and polypeptides are also encompassed by the invention.

[0085] In specific embodiments, polynucleotides of the invention comprise, or alternatively consist of, a polynucleotide sequence in which the 3′ 10 polynucleotides of one of the sequences delineated in column 6 of Table 1B and the 5′ 10 polynucleotides of another sequence in column 6 corresponding to the same Clone ID NO:Z (see Table 1B, column 1) are directly contiguous. Nucleic acids which hybridize to the complement of these 20 lower stringency conditions, are also encompassed by the invention. Polypeptides encoded by these polynucleotides and/or nucleic acids, other polynucleotides and/or nucleic acids encoding these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides, nucleic acids, and polypeptides are also encompassed by the invention.

[0086] In specific embodiments, polynucleotides of the invention comprise, or alternatively consist of, a polynucleotide sequence in which the 3′ 10 polynucleotides of one sequence in column 6 corresponding to the same contig sequence identifer SEQ ID NO:X (see Table 1B, column 2) are directly contiguous. Nucleic acids which hybridize to the complement of these 20 contiguous polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention. Polypeptides encoded by these polynucleotides and/or nucleic acids, other polynucleotides and/or nucleic acids encoding these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides, nucleic acids, and polypeptides are also encompassed by the invention.

[0087] In specific embodiments, polynucleotides of the invention comprise, or alternatively consist of a polynucleotide sequence in which the 3′ 10 polynucleotides of one of the sequences delineated in column 6 of Table 1B and the 5′ 10 polynucleotides of another sequence in column 6 corresponding to the same row are directly contiguous. In preferred embodiments, the 3′ 10 polynucleotides of one of the sequences delineated in column 6 of Table 1B is directly contiguous with the 5′ 10 polynucleotides of the next sequential exon delineated in Table 1B, column 6. Nucleic acids which hybridize to the complement of these 20 contiguous polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention. Polypeptides encoded by these polynucleotides and/or nucleic acids, other polynucleotides and/or nucleic acids encoding these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides, nucleic acids, and polypeptides are also encompassed by the invention.

[0088] Many polynucleotide sequences, such as EST sequences, are publicly available and accessible through sequence databases and may have been publicly available prior to conception of the present invention. Preferably, such related polynucleotides are specifically excluded from the scope of the present invention. Accordingly, for each contig sequence (SEQ ID NO:X) listed in the fourth column of Table 1A, preferably excluded are one or more polynucleotides comprising a nucleotide sequence described by the general formula of a-b, where a is any integer between 1 and the final nucleotide minus 15 of SEQ ID NO:X, b is an integer of 15 to the final nucleotide of SEQ ID NO:X, where both a and b correspond to the positions of nucleotide residues shown in SEQ ID NO:X, and where b is greater than or equal to a +14. More specifically, preferably excluded are one or more polynucleotides comprising a nucleotide sequence described by the general formula of a−b, where a and b are integers as defined in columns 4 and 5, respectively, of Table 3. In specific embodiments, the polynucleotides of the invention do not consist of at least one, two, three, four, five, ten, or more of the specific polynucleotide sequences referenced by the Genbank Accession No. as disclosed in column 6 of Table 3 (including for example, published sequence in connection with a particular BAC clone). In further embodiments, preferably excluded from the invention are the specific polynucleotide sequence(s) contained in the clones corresponding to at least one, two, three, four, five, ten, or more of the available material having the accession numbers identified in the sixth column of this Table (including for example, the actual sequence contained in an identified BAC clone). In no way is this listing meant to encompass all of the sequences which may be excluded by the general formula, it is just a representative example. All references available through these accessions are hereby incorporated by reference in their entirety.

5TABLE 3SEQClone IDIDContigEST DisclaimerNO: ZNO: XID:Range of aRange of bAccession #'sHTPAD461110489011-156115-1575AW026547, AL119230, AI992054,AI968455, AW193784, AW193801,AA386091, AA386130, AI825443,AA004475, and Z79235.HCWFF881210925661-63915-653HSSAX531311988891-34815-362HCEPH71145227391-43215-446AA326209, AA383931, AL365319, andAL390715.HTEDF741510834051-126515-1279AI948732, AW023798, AA058811,AA148838, C01936, and AA663155.HTTEK471611345341-122115-1235AI651805, AI989837, AI718022,AA400005, AI458374, AA401437,AW205244, AA883445, AI478808,AW134723, AI689951, AI269389,AA865056, AI640798, H11911, R44433,AI688852, AI183700, AA424180,AI277599, R46805, and AA771819.HTOBE751711638831-179715-1811AW192827, AA595431, AI251121,AI923335, AI284016, H30141, T70540,T90549, AI432106, AA953436, H27466,R50714, R50249, AI540363, T70809,AI659868, AW370667, Z99572, I36305,AL022146, M87861, M72332, M60235,L39075, M60230, M60228, M60231,M60229, M60232, M60227, and M60234.HCFAT051811563101-214515-2159AL133816, AA811374, AI095381,M38217, M55515, L23499, M85217,X16001, M30312, M31744, AR050270,U38240, U38182, U96110, and U45979.HFIAH371911890011-171215-1726AL134903, AI912243, AA976922,AI742663, AW028771, AW043595,AA478697, AA837145, W49831,AI354405, AA412384, AA602982, N62994,AA013476, AI674206, AA115419,AI377356, AW270325, AI479159,AA169423, AA133086, AA326624,H83962, R76363, AA587840, AI224540,H67227, H81547, AA665443, AA018206,AA662495, AW298791, AA251488,AA301274, AA132987, AI192416,AI300307, AA722928, AA478563, R76688,AA579347, AA501519, AA629042,AI299974, H70300, AI865166, AA252018,N79902, AI867958, AI300195, AA251195,AA251787, H65133, AW246148,AA644296, AA115418, AC004381, andAF227510.HFTDF152010848871-71515-729AA837715, AI623899, AA775049,AA317045, AA581914, AA679872,T03203, AA780929, AI634323, AA604843,AA904275, AW440633, AA362395,AA468486, AI582890, AA640979,AL046746, AA721615, H67866,AA632960, AI561116, AL120976,AI978792, AA663306, AF034193,AW302293, AF108083, AC005486,AL109985, AC007229, U47924,AC008249, AL031289, AC004031,AC003029, AC005899, AL022721,AC003982, Z97053, AC004134,AC004883, AC005839, AC003670,AC005746, AP000018, AL135744,AC005037, AL021808, Z95889,AC006071, AC005480, AC004815,AP000159, AC007551, AL031670,AC000117, I59642, AC005346, L39891,AF039906, AC008372, AL034420,AL078611, AC007055, AF134726,AL049643, AL121769, AL031447,AC003038, AL096701, AL031257,AL035690, AL050318, AL035681,AL049745, AL031589, AC005412,AC007792, AL049636, AL008583,AL021878, AL049569, AC007536,AF165926, AC005234, AL022237,AL031848, AL022149, AC004990,AC006501, AL023553, AC007363,AC005696, AL035455, AC006120,AL031282, AL050309, AC016830,AC005881, AC005529, AC004929,AC006459, AC007308, AC002319,AC002477, AL031277, AC003669,AC002470, AF196969, Z82215, AC006039,AL049780, AC006578, AF111168,AF207550, AF064864, AF165176,AL033521, AC004821, AC003663,AC004655, AF205588, AL096757,AC002429, AC005527, AL034402,AL031775, AC002077, AC009498,AC006006, AJ011930, AC005082,AL031650, AC005695, AL109627,AC000003, AC005264, AC007934,AC005086, Z83838, AC005777,AL133304, Z97630, AC003101,AC004034, AC004672, AL022320,AP000475, AC004905, Z98052, AL078581,AC004517, AL021997, AC006139,AC005996, AC005380, AC002395,AC006023, Z82244, AL034423, AF001550,AL031685, AL008723, AC005102,AC007199, AC007151, AC003043,AF037222, AL008710, AC007193,AC005081, AC004921, AL022329,AC005520, AC005039, AL035587,Z98051, AC020663, AC016027,AC006211, AC003104, Z69921,AL133246, AC005049, AL049869,AC002091, AC005225, and AL033504.HPFCU802110175931-32815-342AC005344.HSVAW492211539161-99115-1005AF146277, U95740, AL008731,AC002551, and AL079342.HWHQC942311164631-90415-918AI366191, AC004472, AC005259, andAW469987.HRSMD492410654581-44215-456AA136820.HFTDY672511512201-152315-1537AI335266, AI751901, AI751815,AI750604, AI040116, AW067945,AW239149, AI572373, N42174, N22119,N71503, AF182316, AF182317, AB033033,AR018882, and AB026436.HYABL892610907331-70015-714HCUEV292711377911-70315-717AI570209, AA583494, AW337550,AW087991, AI027766, H17041,AW410192, AI669151, T30350,AA335428, C16961, AA878169,AW411072, AW410900, T24722,AA365566, AA365567, AI418046,AA350018, AW246233, and AL031283.HCESP562811217511-50615-520AW247740, AW247029, AW204207,W39269, AA325536, R14422, W52568,Y16752, AL022170, and Z65186.HLQDT352911540641-130815-1322AI659435, AW006450, AI380742,AI953510, AI078578, AA707183,AI453381, AI445431, AW136858,AA807157, AW196880, AA131680,AI569636, AI140912, AA530976,AA410746, AA134742, AA152440,AA807317, AA283695, N66180, AI082380,AI269183, AA480063, AA635830,AA433870, AI631995, AI167742,AA292134, AA131985, AA923686,AI580936, N30879, AI358691, AA631878,AA485099, AA485100, AA165214,AA253107, H92198, AI160395, AA830846,AI753274, AA253052, AA152441,AA879095, R08557, R11497, AA358765,AA706241, AA134743, AI424722,AI800536, R08654, R10421, N99172,T89988, AA804424, W16996, AI934059,AA481922, AA292133, AA746933,N56752, AA290907, AI091625, andAL137699.HDPBS64308466241-74315-757AA888874, AA992389, and AI767840.HTBAB413110523881-78515-799AA382198, H48825, H58945, andAI359780.HTLGE31328702471-51915-533AA714179, AW051497, AI971919,AI094911, AW055123, AA293722,AI094408, and AA631985.HWLHK293311522791-94115-955AA243837, AA588755, AW137873,AI351894, AA360896, AI903764, R33743,AL045500, AL039086, AL036802,AL046849, AI906328, AL042628,AW129106, AL036638, AL119791,AI312428, AL036396, AW163554,AL041562, AI690887, AL079963,AL036631, AL037463, AL135661,AI581033, AW020397, AW022682,AL134598, AW238730, AW105601,AI349772, AW023338, AI349937,AI623941, AI537677, AA225339,AA470491, AW059828, AW020419,AL110306, AI279925, AI929108,AI580290, AI624859, AI538885,AI207510, AW268253, AL042166,AI312152, AL046944, AI613038,AI446373, AI590423, AA572758,AL041016, AI678446, AL042544, N29277,AL042382, AI340519, AI340603,AI580190, AI345111, AI907070,AL119748, AW044029, AI866691,AA640779, AL041150, AI433157,AI281867, AA641818, AI886594,AI345347, AI366974, AW162194,AI345415, AI473451, AI348897,AW020710, AI539800, AW149876,AI624943, AI251221, AI349933,AA420722, AI923989, AI284517,AA580663, AI500061, AI628325,AL135025, AW021717, AI868204,AI440263, AI699143, AI247193,AW051059, AI345735, AL042745,AL037454, AI344910, AI863014,AL047344, AW020693, AI538637,AI433976, AL121328, AW074869,AW189301, AW162189, AI801793,AI244249, AI064830, AL047042,AI445430, AW020460, AI357599,AI349256, AW021140, AI277008,AW022861, AW068845, AI698391,AI687568, AI815232, AW050578,AW023859, AI149592, AI307708,AL046931, AI680498, AA579232,AA635382, AL119399, AI340627,AW167918, AI349645, AW089572,AL120307, AL040241, AI690946,AW150578, AW151136, AI953765,AW403717, AI559752, AL121270,AI436429, AI590043, AI452857,AI798456, N57346, AI802542, AL042191,AA806719, AI567582, AI540674,AL038575, AL043070, AL040694,AA494167, AI916419, AW058233,AI573026, AI471227, AL118781,AI920968, AI589668, AI345612,AI345688, AW082532, F37439, AI610362,AL120736, AL036664, AL043168,AL119836, AI866465, AW074993,AW268072, AL043345, AI554343,AW191003, AL038605, AI866608,AI345608, AW268261, AI471898,AL036240, AW021373, AW020561,AL041772, AI114703, AI475371,AI582912, AL036736, AA693314,AI554821, AI287446, AI307569,AI345253, AA848053, AL048323,AA613907, AI345416, AW268067,AI313320, AW081383, AI624548,AL048340, AI343030, N22276,AW020095, AI310606, F35428, AL135022,AL036274, AW151138, AI915295,AL047422, AI345370, AW166645,AI312146, AI560012, AI312339,AI890907, AL036403, AW059638,AI921254, AW161202, AW022102,AI348854, AI345258, AI249946, W48671,AL119511, AL040243, AA528822,AI434223, AL037582, AL037602,AI612885, AW080402, AI801325,AA070889, AI273791, AW274192,AW163834, I48978, S78214, I48979,AR013797, AF090934, Y11587,AL133640, AF078844, AC006313,AF113691, AL137479, AL133016,AL050393, AL117457, AF215669,AL137459, AL137478, AF090900,AF118064, AL133075, AF081197,AF081195, A58524, A58523, AJ012755,AF126247, AF090943, AL117460, X84990,D16301, I89947, AF090901, U75932,Y10655, AL049938, AF118070, AL137529,AF090896, AL133606, AL122123, X99257,A08916, AF125949, AL137429, AL122050,AL122100, AL050146, A18777, I03321,AF111851, AL117629, I09499, AF158248,AF113013, X06146, X82434, Y16645,A08910, AF113019, A08909, AL049382,A08913, AL137705, AL122093,AL110196, AL049314, AL137527, I89931,AL137538, S61953, AF097996, E05822,E06743, L31396, AL050138, E02221,L31397, AJ242859, X70514, AF114170,AF090903, AL137294, AL080060, A08907,A08908, U77594, X80340, AL122111,AB007812, AL096744, AF039138,AF039137, AL133113, AF061573, X72889,I49625, AJ000937, AF069506, AF176651,AL137558, A93016, AL122110, AF113690,AF100931, AF017437, S78453, A65340,AL133093, AL080154, AL110197,AL117416, A07647, AL122118, AF030513,AR034821, AL137555, A03736,AR011880, AR038854, AL110159, S75997,Y11254, AF118090, AJ238278, AF017152,X65873, U42766, AF061943, AL050277,AF100781, AL050024, AF132676,AL122106, AF061836, AL122098,AF026124, AF057300, AL110225,AF057299, AL050092, AL096751,AF036941, AL133067, E02349, A08912,AL137548, AL110280, AF113694, S36676,A77033, A77035, AR000496, AF113699,U39656, X89102, U55017, X67688,AL133557, AL117649, I66342, AL137550,AL137656, S77771, AF106657, E01614,E13364, AF079765, AL137292, AB019565,U51587, AF113677, X59414, AL080158,U87620, AL050116, Z97214, AL137533,AF177401, AL117435, U72620, AL133560,AF113689, AL137526, AL080074, I33392,AL137271, AF192557, U96683, AF146568,AL137488, AL137521, X63574, AF102578,S76508, I89934, AF026816, A65341,AL049283, AL049466, AF200464,AF079763, AF111849, AL117585,AL049452, E07108, AL050108, A21101,AF180525, A52563, AL133619, L04504,X53587, A18788, AL080124, AL117648,AL137557, AF065135, AR038969,AL049430, X55446, AF090886, Y09972,AL133112, U00763, E03671, AL133080,X70685, U58996, AL137648, AL133565,AF104032, AL133081, AF031147,M96857, AL080137, X57961, AL133665,I68732, U66059, D83032, AF162270,AC004200, Y10080, AF111112, S83456,AL136884, AF003737, AL049300,AL117432, and AL050366.HHEGG203411068161-95815-972AF084205, and AW473942.HDPRU433512170351-293915-2953AI912020, AI738591, AI673200,AW195629, AI914327, AW207103,AI858984, AI016102, AI948562,AI021976, AA515654, AA824295,AI288261, AW003109, AA029227,AI332790, AI863407, AW271426,AA447206, AW197033, AI673222,AA846300, AI093417, AW274813,AA917651, AI989749, AI198249,AI933079, AW337461, AA149282,AA476264, AA029228, N89854, AI650694,Z28929, AA149376, R53155, AA805734,AI862408, AW374891, AA010232,AW137892, AW250017, AA077657,H43959, AA077229, AI400383, AA077669,AI971136, AA076731, AW405369,AI862409, AW386712, AW386708,R49809, R43152, H22270, AA077893,AA889934, F00253, AF139794, M91506,AA077980, AI282751, AA077825,AA010157, F01048, AW393714,AA078429, AA077544, AI768799,AA076983, AA076893, AI125178,AW205906, AW373785, AI963990,AB011110, AC004084, and AC004985.HE8PK123612276471-367815-3692AI675352, AA404223, N21458,AW377007, AW055240, AW376965,W72194, AI744480, AW377009,AA460033, AA689223, AI151138,AW367398, AI204462, AW367559,AI814765, AA514481, AA514486,AL041934, AI378530, AI127602,AI805975, AI816548, AA459944,AI079813, H99690, N69075, AI220492,AI924878, AA894562, AA227526,AW440516, AA652186, W78757,AI769346, AI223789, AA663829, R74386,AI129571, AI679615, T08944, W80492,N35223, AI366554, AA884120, AA227665,W04635, AI025087, AI141989, AI276061,AI131356, AI057082, N76793, AW089172,AA758991, AI149095, AW089170,N69065, H82484, AI366370, W31730,N44887, R78350, AI868614, M85694,T07736, AA404607, AI167295, H13964,AI351984, AA704854, AI832531, Z43566,AW008608, T30004, T35144, AW023452,R62595, R83633, AA978218, R77974,R62643, R84838, R73944, D62538,C04805, W77830, Z39632, AA663708,AA258404, AA904920, AI915380, T30003,T30824, AA016152, AA296029, AI139527,H22033, AA555134, AA776071, D50973,AA216002, AW148599, AA249464,H22108, AA369404, AA194728,AW367406, D79628, AI800441,AL117472, AL122082, U58883, AF136380,AF136381, AF078667, and AF078666.HE9HV923712275191-342215-3436AW451462, AW389371, AA469982,AW389370, AW389367, AI625782,N62808, AW389369, AW367986,AW389368, AW389596, AW389606,AW044368, AA679038, AI493173,AW083020, AI280052, AI634922,AA402837, AW377678, AW389366,N33240, AI805314, AW004651, AI141651,N93923, AI000968, AF114067, AI922980,AA910867, AA918883, AW389616,H98243, H29386, H29296, H98939,AW367988, AW028291, F11743,AW002598, AI143427, AI679840,AA709068, AA402687, AA927333,AI183648, W01827, AW367982, Z45487,F06551, T65373, AW367985, AW389594,AW367974, AI679265, AW367968,AI569013, N79690, AA412251, N70773,AA994823, AI588847, AW377594,AA886715, AW139939, Z39513, F04059,F09403, F04352, AA348028, AI700831,R12190, AW139934, AI275660, AI253408,AI423822, R35809, AW389634,AW389639, Z45167, AW295763,AW389633, AW389637, R39914,AW389632, AA336725, T65450,AW389643, N41955, M78477, AW389640,AW389626, N56035, AW083017,AA343776, AW389615, AW389623,AW389628, AI802303, AW389641,AW389630, AW389359, AW389354,AA961766, AB007860, I74314, I74317,AF069506, AW627522, and AW630555.HOHCE473812170591-213115-2145AI902550, AA852921, AW449324, andAB018324.HSDII693911540671-141815-1432AA203346, AA203330, AI024792,AI383978, AA489694, AA658936,AI912487, AA640288, AW022618,AA115749, AA669824, AW296909,AW024848, AA133454, AA552781,AI332862, T86475, AI332863, AA665267,AA640358, AI625278, AA878769, T31809,AA318980, T86474, Z24863, andAL049423.HKAKM104012276391-392115-3935AW166113, AI762270, AI761800,AI968494, AI935006, AI393355,AA885443, AA847857, AW044423,AI188200, AW085175, T66118, AI935038,AW003121, AI356390, AA588552,AI475498, AI139170, N63644, AW450561,AW088729, R70631, AI203354, AI239443,W45495, AA654175, H95337, AI002826,AA578373, F10209, AI468683, F09647,T83850, AA317152, T65033, AA641861,AW196381, AI350190, F12591, AI383380,W40418, F12002, T74348, W23298,R70529, T89046, N80491, AI474886,R88730, AB014530, AF071070, AF170304,AF077658, AF170303, and AF071071.HCEPU564112261201-275415-2768AL134903, AW043595, AI912243,AA602982, AI674206, AA976922,AI224540, AA662495, AI742663,AW028771, AA478697, AA837145,AI354405, W49831, AA132987,AA412384, N62994, AI300307, AA013476,AA115419, AA722928, AI377356,AA478563, AW270325, AI479159,R76688, AA579347, Z43536, AA629042,AA169423, AA326624, H83962,AA133086, R76363, AA665443,AA587840, H67227, H81547, AA018206,AW298791, AA251488, AA301274,AI300195, AI192416, AI867958,AA501519, AI299974, H70300, AA644296,AI865166, AA252018, N79902, AA251195,AA251787, H65133, T26917, N83883,T26928, AL045327, AL045328, AL134524,AL134110, AL038878, AL047163,AW246148, AL042898, U46344,AI318479, AL037295, AL038838,AL048677, AL037343, AL038651,AI547295, AL038983, AI142134, D29033,AL037436, AL037335, AL037323,AL135012, AL037727, AL037443,AL038532, AL038822, AL038761,AL037435, AL040472, AL043941,AL039432, AL045494, AL044125,AL043923, AL043814, AL047012,AL041238, AL044186, AL040617,AL043845, AL041347, AL038024,AL040576, AL040193, AL045753,AL042523, AL041955, AL040463,AL047170, AL048657, AL044037,AL041635, AL040294, AL044064,AL041459, AL041577, AL044162,AL040464, AL047219, AL040625,AL045684, AL041752, AL046850,AL040768, AL045671, AL046994,AL048714, AL046914, AL045891,AL039360, AL038745, AL040052,AL043496, AL040444, AL043538,AL040621, AL040510, AL043467,AL043677, AL040839, AL043492,AL041602, AL044074, AL041730,AL041523, AL043627, AL041374,AL043848, AL043570, AL047183,AL042135, AL042655, AL046442,AL041324, AA115418, AL041133,AL039643, AL039316, AL041098,AL040322, AL046392, AL043089,AL040119, AL044272, AL044258,AL042420, AL037341, AL042468,AL038040, AL041096, AL042096,AL045817, AL041168, AL049018,AL079852, AL043321, AL041163,AL041159, AW363350, AL045920,AL040148, AL042741, AL047057,AL038041, AL040458, AL044187,AL046356, AF227510, AC004381,AR066494, AJ238010, A93923, A93916,D17247, A93931, AR064707, AL133053,AL122101, A85203, AR023813,AW611706, and AW769812.HUSHB54429280541-34315-357AA348022.HLMDO95439283441-46915-483AC020641.HHASQ324411989021-88015-894H00195, AI251764, AL042753,AW117882, AI349772, AL047763,AL121270, AI868831, AI433976,AW162071, AL047042, AI064830,AI436456, AI815383, AI433157,AL135661, AL036146, AI500077,AL119791, AL036396, AW268253,AL045500, AL046849, AA640779,AI349645, AI500553, AL036802,AW071349, AI687376, AI679724,AL119049, AI568870, AI863014,AI608667, AI690751, AI702406,AA613907, AW303152, AL121365,AI567351, AI349933, AI920968,AW238730, AI207510, AI873731,AL040169, AI687728, AI538716,AI469532, AI580190, AL036759,AI567632, AL119748, AW103371,AI673256, AL120854, AI934036,AI969601, AW166645, AI340582,AW080838, AW089572, AI907070,AI687415, AI521012, AI440426,AI906328, AW074993, AL036980,AI499393, AI349256, AI312152,AI753683, AW274192, AI969567,AI349937, AW302965, AI349614,AI349004, AI349598, AL038605,AI889203, AA585422, AI251485,AI343112, AL036274, AI343059,AL120736, AL036240, AI818683,AW132121, AW301409, AI282655,AA572758, AI609592, AI678302,AW235035, AI285735, AI250293,AI799305, AI309401, AI631107,AW195957, AI366549, AI281779,AW071417, AI149592, AI583316,AI690835, AI340519, AI345860,AI909666, AI686926, AI345744,AI475371, AI345735, AW087445,AI696846, AI498579, AI439087,AI497733, AI636456, AI813914,AI307466, AI366991, AI682743,AI907061, AI866608, AI275175,AI597918, AI635461, AI345111,AI909662, AW169653, AI635942,AI624859, AI671679, AA528491,AW068845, AI564719, AI699857,AI560012, AI568854, AI620284,AI220734, AA938383, AI446606,AI312428, AI590128, AL040243,AI445432, AI625079, AA528822,AI697137, AI857296, AI919058,AI540832, AI613017, AI249257,AI702433, AW148320, AI800453,AI800433, AI609580, AI866780,AI499463, AL038778, AI610307,AI758437, AI597750, AI633419,AI866002, AI684265, AA603930,AL043326, AW074869, AI224992,AL048871, AI687362, AI434281,AI281773, AI680113, AI344182,AI952114, AI874109, AI862142,AI440239, AI271786, AI499131,AL038779, AI682841, AI269696,AI307558, AL042538, AL036247,AI348897, AI475134, AI610645,AI682106, AI889839, AI866887,AW301300, AI628205, AI800411,AI539771, AI500659, AI469811,AI754897, AI064787, AI569616,AI281762, AI624668, AI580984,AA508692, AA528529, AI570384,AW075351, AW167776, AI802542,AW026882, AL036260, AI907056,AL047041, AI921379, AI610756,AI886532, AI818206, AI811863,AI334902, AI492540, AI445025,AL044207, AI636445, AW104724,AI439745, AW002342, AI888953,AI687375, AI282281, AI620868,AL036361, AI493248, AI340603,AW085799, AI349226, AW118557,AI804585, AI609331, AW268768,AI283941, AC005045, AF073770,AF168793, U65745, I48979, AF118064,AL049430, Y11587, S78214, AL133640,AF090934, AL050393, AL133016,AF113013, AF078844, AF118070, A93016,AF113691, AF090900, AF125949,AL050146, AF090901, AF090943, L31396,L31397, AJ242859, AL080060, AL110196,AF113676, AL133606, AL049938,AF104032, AF090903, AL117460,AL117457, AL049452, AF113689,AL110221, AR059958, AF113694,AL137527, I89947, AC002467, A08916,S68736, AF091512, AF113690, I89931,U95739, AC006371, AL122050,AL050149, AF090896, AF106862,AC007298, X84990, AL050108, U42766,AL096744, AB019565, AC004987,AL133075, AL050116, A08913,AL049314, AL096776, AL050277,AL122093, AF113677, AC005015,AF113019, Y16645, AC004690,AL049466, AC006336, I48978, AC005242,AF113699, AF017152, AL080137, S61953,AC006115, AC006944, AF042090,AC008394, AL133557, AC005886,AC007151, AR011880, E03348,AL137283, AL133093, AC005291,X63574, AL022147, AC007390,AL035587, AL049776, AC004520,AL080124, AL133080, AL022315,AL137557, AC006480, AC005520,AC010072, AF158248, AC005488,AL117394, AL034451, AL121934,Y11254, AL137459, AL133565,AC004686, AC004491, AL031283,AF162270, AF097996, AP000013,AL030999, U91329, AL122121, AF146568,AC004883, AC005175, Z98036, AJ000937,Z82206, AC006840, AC004383,AL122123, E07361, AF111851, AC007676,AP000201, AC009233, AF003737, U66059,AC002504, AF196779, AL109827, Z68164,AL050138, AF091084, AP000097,AL035067, AL033521, AL078593,AL008735, AC005048, AL110225,AC006157, AF079765, AP000240, andX82434.HARAB874511643401-53615-550AI733761, AA121199, AA296916,AI770025, AI733898, AA366952,AA149362, AI288862, AC005669, S76742,I26666, and AF075261.HTNGF694611522681-82415-838AI139022, AA552953, AI204346,AI741884, AI042245, AW205544,AI017548, AA258321, AI571085,AA478401, AA258530, AA701456,AA454012, AA402279, AA873796,Z41600, AA688162, AI799203, andC00673.HMSJL964711547881-189815-1912U69566, AI478738, AI700975, AW270625,AA558431, AI955662, AI796542,AA620937, AW073767, AA928103,AI087266, AI949116, AI273460,AI766325, AW297573, AA873139,AI350929, AA770419, AA954532, N64139,R75698, N87915, R01072, H26525,R01798, AA773665, AI003807, AA318420,and AA318524.HDTBT064812052611-143415-1448AA443164, AW020571, AA532437,AW118680, AI890631, AI167234,AA744921, AI401545, AW152040,AW074628, H06431, R74385, AA862392,AA649701, AA160546, AI880570, D62917,H23246, H46816, AA973615, AI819867,H00507, R21742, AA463453, C05953,R25958, R26764, AA639080, AW195349,AA315526, AW362722, AI494615,AW070869, H71752, T84284, AA767232,AI824357, and AF161399.HTTIE474911653631-87815-892R29204, W60551, and AC004386.HHFBP47509466681-119315-1207AW341717, AA001259, AA778598,AA203115, AI632467, AA707336, H69425,H68732, AI992354, AA346824, AI541205,AI557731, AI525500, AI557533,AI525556, AI557262, AI541321,AI557082, H65400, AI557238, AI557602,T18597, AI540890, AW021281, AI541056,AI557258, AI541048, AI525856,AI557084, AI535660, AI525656, D50992,Z32887, AW021585, AI526078, Z33559,AI557234, D59751, AI535639, AW023235,AI525757, AW022727, AL050350,AC002464, AR050070, S68736, A62298,A82595, A82593, Z30183, U94592,Y08991, A62300, AF006072, U45328, andAR025466.HCCCC815110835531-71015-724W16450, AL042537, AA367722,AI436552, and AA236375.HPJEV715211978411-183015-1844AW239092, AA174017, AA303007,AL038072, AW103758, AI923333,AA576336, AA704393, AI654285,AI859946, AI888468, AA837778,AI254913, AW089101, AA584145,AA468131, F08230, AA302973,AA847952, AI620992, AI923126,AI800168, AI278972, AI207465,AW023149, H06140, AI751216, AI800180,AW131208, AI824706, AW440545,AP001037, AC006211, AL080243,AL121892, AP000525, AC006480,AC006288, AL031321, AL008731,AC004466, AC005870, AC008134,AC000353, AL117352, AC003049,AC007227, AF008191, AC004836,AC005041, AC005209, Z98941,AL049690, AC005952, AC007216,AL034417, AC004851, X05151,AC005480, AC006080, AL049830,AC004837, U95742, AC003684,AC005324, AC005529, AF059650,AF050154, Z85987, AL008638, AC005886,AL049636, AC007114, AC005368,AC007056, AC012099, Z97989,AL133249, AP000555, AC007386,AC004924, AC005728, AC006946,AL133246, AL022323, AC006961,AL022302, Z99755, AL031427, Z81014,AC002350, AL021918, Z97876, AL035417,AC005484, AP000228, Z98750, AC003670,AP001054, AL035555, Z83826, AL078463,U95743, AC004882, AC004622,AL024494, Z69707, U95090, AC006133,AL032821, AC007738, AL022238, Z71182,AC005920, AL034423, X12919,AC004475, AC000066, AC002126,AC007011, AC002996, AP000140,AF157623, AC005318, AL035659,AC002980, AC007537, AC016027,AC016830, AL133244, Z82206,AC005514, AL096775, AF070718,AC004905, AP000035, AC004696,AP000350, Z97054, AC005089, AC005587,AL031575, AL031281, Z82214, AC006230,AC006064, AC001644, AP000466,AP000088, AC002325, L29252, AC000082,Z73965, AL031767, AC002553, andAC007312.HTEIL075311361211-121615-1230AA725842, AW139921, AI651885,AI971598, AA453466, AL110422,AI027229, AI026797, AA778573,AI279962, AI073425, AI208767, H55405,AA453966, H55431, AL118498, andAL031843.HTEAG49549546141-128915-1303AW452652, AI039005, AA780077,AW316890, AI337290, AA463229,AA463230, AI423317, AI468158,AA382497, N66986, AF041822,AL390796, AL390796, AL357045, andAL357045.HSLCF96556376701-157515-1589HNHCI32568616731-58615-600AF112462, and AR035954.HPMFL085710506841-46315-477AA555286, AA640814, AI281916,AW073979, AI378363, R70468,AW242350, AW013856, AA644290,AW449140, Z93016, AC012384,AL035541, AC005228, AC003662, andAC009300.HTXRA13589596221-115015-1164AI701008, N92159, AA363885, AI990716,AW197467, and AW291923.HCE3H715911978981-205915-2073AI638508, AW139057, AW149807,AI654790, AI419465, AI569283, H51217,R90888, R85125, H29245, H29244,AA488106, AI480182, H05353, AI500178,AI873131, AI499775, F11386, H05303,AI458144, H51247, F05862, R44423,F09051, R14779, R51725, R51648,R42549, AA338672, Z40525, F07470,R19635, W22746, R49451, R35934,R85080, F02105, AW169213, AL046466,AW162194, AI352326, AI700159,AI955945, AW192701, AI890507,AI978703, AI263331, AI800159,AI573167, AI571980, AI961589,AW169671, AW188693, AI634224,AI274507, AI690748, AI758309,AI648436, AL039716, AI590043,AI911648, AI690687, AW075484,AI917963, AI648684, AI698391,AI671642, AI619607, AW168031,AW168296, AI572787, AI918435,AL119863, AI564426, AI814087,AI470648, AI452556, AW029611,AI921254, AW054960, AI365256,AI564234, AI597758, AI919500,AI624120, AI686716, AI621341,AI309306, AL079799, AW071349,AW008781, AI926878, AI564448,AW024594, AI933992, AI540676,AW051088, AW198075, AI921155,AI536685, AI561356, AI922812,AI538850, AI468872, AW022699,AW089275, AI919593, AW188382,AI866770, AI434134, AW264019,AA835966, AI690620, AI950729,AI564765, AI813868, AI925680,AI829327, AI417790, AI540759,AI630876, AI570872, AI744279,AL046595, AI439452, AI566386,AI354630, AI609409, AI681985,AL040011, AI784219, AI247193,AI590781, AW080746, AI874261,AI249877, AA279795, AI470284,AI589428, AI445611, AI590415,AI620284, AI887389, AI554485,AI963625, AI522052, AI654672,AI520809, AI889306, AI540674,AI783821, AI281757, AI474646,AI889376, AA983883, AI567769,AA916133, AI670009, AI783504,AL047675, AI805671, AI366900,AW130307, AI289310, AI553645,AW022093, AI797538, AW029197,AI696570, AI624293, AI345688,AI872423, AI362580, AI580674,AW087199, AL135517, AW170725,AA687501, AI479577, AI868204,AI863241, AW194185, AI572021,AI492540, AA502794, AW080346,AI419650, AI679550, AI632851,AI920835, AW188539, AI241923,AI587121, AW163823, AI433611,AI697157, AI621209, AW008166,AI862139, AI612750, AI147686,AA806719, AW102902, AI273901,AI306613, AI679672, AW152182,AI270295, AI422688, AW024793,AI288305, AI695129, AI569945,AL036673, AI918449, AW088560,AI889189, AW148423, AI620075,AI357599, AI269205, AW167228,AI277325, AL138386, AI439745,AW193467, AI590830, AI623941,AW117997, AI797908, AI963224,AL038665, AI524179, AI818353,AW075413, AI954507, AI683099,AI524671, AI635478, AI653578,AI623379, AI860783, AW007309,AI635639, AI564749, AI830259,AW029533, AI627360, AI874166,AI582932, D29763, D64009, D64010,I89947, AL137271, AF067728, AL137665,AR038854, AL050155, AL117435,AJ003118, AF177401, AJ005690, I48978,AF113690, AL137463, AL050277, A65340,AL137479, AL133560, S75997, AF028823,S36676, AF111849, E01314, AL080126,AL110296, Z97214, U75932, AF151109,X52128, AL133606, AF106862, A18777,AL137459, AL122100, AF061795,AF151685, A45787, AL110225, I32738,AR034821, A15345, AL080124,AL137550, AL117416, AF183393, E02349,A08913, AL137574, AL133568,AL137480, AL133112, AL137478,A08916, U86379, AL133557, M27260,A08912, AL110280, A52563, AL133637,I89931, X93495, AL110222, A08910,X72889, A21103, I49625, AF091084,A08909, AJ238278, AL122098, AL117394,AL080148, X63574, AF061573, AF113677,A08908, AF031147, AL137533, AL137294,AL050138, AJ006417, AF026816, I89934,AF017437, AL050149, AL133010, U35846,E01614, E13364, AL049339, A07588,AL133558, Y11254, Y10655, AL122110,AR020905, A65341, A77033, A77035,AL137641, AL023657, I48979, S78453,AL080159, AF087943, Y14314, AL137558,AF158248, AF061981, AL133619,AL117463, AF079763, U87620, U80742,S76508, AL049314, AL080154, AF111851,AL110218, A90832, AL117460, AF094480,AF090900, U66274, AL133016, A23630,I00734, AF113694, L19437, AF090943,AF069506, Z82022, AF017152, AF205861,E00617, E00717, E00778, AF016271,AF100781, AL137292, AL137537, I89944,L04849, X82434, X79812, Y11587,AF118070, AL122050, AL117583,AF159615, AF017790, I17544, AL137529,AL110221, AF090903, Y09972, AF026124,U42766, AF051325, D83032, AL049464,AL137560, AL137530, AL049452,U55017, X67688, A18788, E12747,X63410, AF113019, AL137523, AF057300,AF057299, AL110171, AF118094,AL049283, E04233, X80340, AR059958,AL080127, AF125948, AF185576,AF113691, A58524, A58523, AL137276,AL137429, AL137283, X76228, S63521,AL133080, X70685, M30514, E06743,AJ242859, AL137256, L31396, U68387,AL137656, AF146568, AL122093, L31397,U91329, AJ012755, M92439, AF003737,AL080163, I96214, AR034830, M86826,S82852, AF137367, AF115392, AL122106,U00763, AF106697, U57352, S68736,AL137488, AL133113, A03736, AF032666,AF111112, A08907, Y10823, AL049430,AL133640, AF118090, M96857, E07108,AL117457, D16301, I66342, AL133665,AL050092, AL049465, AR011880,A08911, and AL133067.HUTSF116011349141-43715-451AI384010, AI288640, Z20435, andA74523.HTEGI486110212351-53615-550AA102044, H93506, AF140360, andAF074606.HSFAM096211539131-28115-295HNFHK776311822861-127215-1286HFXDO836410126021-60515-619T03269, D58283, D59859, D80022,D80166, D80195, D80193, D59927,D59467, D51423, D59619, D80210,D51799, D80391, D80164, D59275,D80240, D80253, D80043, D59787,D80227, D59502, C14331, D80038,AW177440, AW378532, AW179328,AA305409, D80134, C14389, D51097,C75259, AW360811, AW366296,AW375405, D58253, AA305578,AW375406, AW378534, AW179332,AW377672, AW179023, AW178905,D80132, AW352170, AW377676,AW178906, AW177733, AA033512,D89785, D34614, X67155, X82626,A84916, A67220, A62300, A62298,Y17188, A78862, D26022, D88547,AJ132110, A25909, AR018138, AB012117,AR025207, A85396, A44171, A85477,AR066482, A86792, I19525, X93549,AF058696, Y12724, AR008278,AB028859, D88507, A94995, AF135125,U87250, I18367, U87247, AB033111, andAR064240.HSDIW736511044061-160215-1616HFVGD236611996451-184215-1856W78862, W19697, H14921, AA576940,H14628, H54580, AW119184, T93838,AA056294, R53170, R99389, R11968,F11919, F11380, T79980, AA043550,T65194, R09850, AI948441, AI206824,AW190832, AI199930, R55990,AA013266, AI097591, AI754286,AF073770, AC005045, AF052187,AF168793, U65745, U26033, J02844, andZ98742.HMSBZ246710823671-70615-720H14236.HWHHB696812170421-289215-2906AL045925, AL042685, AW245943,AW245500, AI827711, AW247149,AI589761, AI422317, AI076031, N94372,AA708691, AA702609, AA702730,AI912816, AI984055, AL042686,AI420887, AL045926, AW272712,AW237553, AI469477, AA494501,AW374634, AI863106, AA181277,T15903, AI476561, R06007, AA380280,AA322751, C04187, F36452, AW248180,AA883148, AI767541, T82100, AA506918,AA227226, C01255, N57562, andAJ242655.HFXLC696911625431-163215-1646AA296887, AA297110, AF065216,AF121908, and AF065215.HBXBW407011567651-57415-588AL023754, AL049688, and D86557.HCE1L517111404981-42015-434AA349751.HRADM457211480461-46815-482AA418916, AA426580, AJ271722,AP000260, AP000036, AF055919,AP000099, and AP000098.HTEFO457311539181-89915-913AA620907, AA758187, AA015938,R84831, AA412093, R58215, and U75361.HOHBN827411522711-145315-1467AI335266, AI751901, AI751815,AI750604, AI040116, AW067945,AW239149, AI572373, N42174, N22119,N71503, AF182316, AF182317, AB033033,and AR018882.HWHGF527512170261-222815-2242AW044636, AI216076, AA846742,W22035, AW172841, AI291112,AI681233, AW080046, AI399893, H28922,AA614450, AA536156, AI278855, N39741,AI183847, H21177, AA313439, N20038,AA470881, AI088477, AA032036,AW007815, AA515917, AA348553,R86198, AW006990, AI094006, D61068,AI282605, R48136, AA971399, AA333772,AA365066, N26782, AA034263,AA377509, AA804676, AI242834, R48135,AA576728, D60948, D60442, H46369,R49863, D81590, AI183862, AA894892,AA886813, D53429, AA484583, R49862,AA907007, AA339653, AI183749,AA348552, AI167561, AI869376,AA223889, M78928, AI241882,AB002360, and Z35654.HBKDI307612238611-162615-1640F37398, AA197072, F22840, AW444680,AA196995, F31270, R02824, F37397,AA112849, F17211, F37676, F30884,F35768, F35763, J05194, and J03886.HSQFR547711851431-201215-2026AA811369, AA873372, AA209183,AA304836, W26880, AW189673,AI459087, AA326319, T11336, H00191,AL120282, and U35244.HAGBA567811025931-127315-1287AA812064, AI803142, AA430303,AA430200, AI425013, AA954361,AA012835, T07365, AA984341, AI207984,AB020641, AF033655, U62391,AC000057, AC006036, and AC002458.HHSAE297912208511-119315-1207HMSHO64807465821-39815-412HFPBW228111547861-105115-1065AA583561, AI685755, and AI971263.HTLBH678212243711-287215-2886AI700778, AI651596, AA203283,AI079116, AI269328, AA766256,AA846207, W94671, W95139, AI765522,AI123765, AW337193, AI693824, T15539,AI269741, W19592, AW449541,AW407143, AA045079, AI204381,AA045395, AW305104, AI742830,AW052085, AI858504, AI587476,AW135172, AI887064, AW380082,AW295490, AI453541, AI955603,AA971802, AA485799, AA287153,AI761438, AA485669, AA781274,AA214554, AI865011, AA284733,AW341388, AA814271, AA447822,AA256234, AA368228, AC005368,AF086518, and AF059650.HNTMH708311435231-107315-1087AL045099, AL110384, AA477597,AA485587, R25821, AL134410, R55476,AA584162, H19102, AI383263, AA349309,H42360, AA953198, AI688425, W22471,AI537054, AA736967, R56322, AA526482,H29361, AI699883, F06802, Z78292,AA081493, AW168512, AA035215,AA078393, AA077854, H21589, AI004027,AA836778, D44975, AA581900, R17223,H18689, AA507183, AI570959, AL134844,R87452, AA569527, AA491053,AA077436, R28228, R49752, F00981,W38765, AA584770, AA601420,AA374571, R87607, AA601480,AA310271, R26668, R11721, AW298136,AA780824, F12062, AA164247,AA062928, AI539135, AI752143, T66331,AA868891, AI298079, AA582543,AC005726, AC005785, AF196970,AL034374, AL049822, Y18000,AC005529, AC005288, AC002553,AC002395, AF052041, AP000087,AL049594, AC015853, AC007999,AL049839, AC005531, AC008038,AF095725, AC002351, AL034394,AC005694, AC004860, AC005343,Z98304, AL031650, AL109847, AL021938,AP000225, AC002524, AC007372,AL050321, AP000555, AC005829,AC003663, AL035681, AP000350,AL049837, AL118497, AC005013, Z98742,AL031685, AC003991, AC006449,AB000882, AL031057, U91326,AL024507, AC002544, AL109662,AC005206, AC006992, AL049632,AL133245, AL109837, AC005667,AP000506, AL080233, AL117356,AC007376, AB014080, AL021154,AC004814, AL121653, AC004206,AC006236, AL034430, AL121756,AL049757, AC004757, AL049552,AC004073, AC006557, AL117258,AP000514, AL031671, AC004167,AJ011930, AC004849, AB000879,AL021397, AL049539, AC005535,AC004079, AL031668, AL031430,AC005971, AC004558, Z69654,AL080112, AL096773, AC005681,AF165147, AC005006, AP000502,AC002525, AC005962, Z83840,AC004448, AC007172, AC008372,AL031602, Z82976, AL035045, AF129077,Z97054, AC007560, AC004021,AL121577, AC003005, Z94054, AL023575,AC002531, AC005156, AC004196,Z94044, AL033527, AF165926, AP000520,L09706, AC008249, AC004474, AF029308,Z75746, AC006132, Z73900, AC005046,AB023056, AF019413, AC007327,AF064859, AC005069, AC007126, Z11739,AP000510, AL133246, AL021394,AC004520, AC002556, AC007066,AC003071, AP000553, AC005684, Z11740,Z98172, AB023048, AC009516, Z95437,AC004065, AC007546, AC004973,AL031224, AC006427, AL049842,AL022101, AL035078, AC005332, Z83844,AC003102, AC004169, AP000694,AC004601, AC008417, AC006313,AC007967, AL049832, Z83841,AC005510, AC004462, AP000501,AL133304, AC007387, Z84487, AL096703,AC002463, AC004213, AL133243,AC004762, AL137191, AC006539,AL022476, D10855, AP000354, AL022723,AL031391, AC002991, L77570, Z82203,AC005067, AC004067, AL022170,AC005320, AC005527, AC002400,Z75889, AL023875, AC007666, AL135745,AC000052, AD000813, AC004144,AF187850, AC004019, AC005940,AL031229, AF109076, AL049646,AC007435, Z82171, AC005570,AC004531, AC000025, AL050343,AF107885, AC005828, Z82196, AL022332,Z93403, AF117829, AL031275, AL021877,AC006077, Z98941, AC004858,AC000387, AC006222, AC006952,AC006001, AL022721, AC004885,AL035458, AP000352, AC002402,AF109907, AC000003, AC007687, Z97056,AC005192, AC005994, AL109802,AL008583, AL022400, AL049843,AL079342, AC002996, AP001053,AC006039, AC005317, AC006008, andAL135879.HCETC598411833341-40115-415AA346622, AA085022, and U35245.HE8UX768511612231-176315-1777AI863338, AA703679, AI223268, F12220,F09849, R22646, H04918, H04815,T66347, AB026810, AB026803,AB026809, U20105, and Y19241.HTLEN778611361241-118015-1194T89857, T89583, AI349772, AL121365,AL047042, AL036396, AL121270,AI687376, AL119791, AW238730,AW268253, AI636456, AI349004,AI868831, AL045500, AI521012,AI285735, AW301409, AL135661,AL038778, AW103371, AL036802,AL047763, AI433976, AI475371,AW071349, AI436456, AL036146,AI687728, AI064830, AI349645,AI445432, AI635461, AI625079,AI250293, AI568870, AI702406,AI538716, AW195957, AI439087,AW162071, AI678302, AI564719,AL119049, AI815383, AI620284,AI498579, AI433157, AW169653,AI340582, AI349933, AI613017,AL045903, AL046849, AI866608,AL036274, AL040243, AW274192,AI590128, AI699857, AW071417,AI540832, AI440426, AI697137,AI249257, AI275175, AI281779,AI857296, AI702433, AW148320,AI800453, AI800433, AL038605,AI500553, AI920968, AI500077,AI580190, AI567351, AW074993,AI439745, AI312152, AI345735,AI349937, AW089572, AI207510,AI906328, AI690751, AW068845,AI758437, AI597750, AI499463,AI682743, AL120854, AW166645,AI497733, AI673256, AI679724,AI633419, AI866002, AI863014,AI349614, AI499131, AW080838,AI686926, AW117882, AL043326,AI440239, AI873731, AI567632,AW303152, AI282655, AI281773,AI610307, AI907070, AI631107,AI568855, AI818683, AI499393,AI224992, AL036759, AL038779,AI343112, AI866887, AA613907,AW074869, AI349598, AL120736,AI349256, AI608667, AI866780,AL048871, AW087445, AI348897,AL040169, AI682841, AI271786AA572758, AI560012, AI800411,AA640779, AI628205, AI597918,AI921379, AI690835, AI469532,AI568854, AI434281, AW132121,AI969601, AI580984, AI366549,AI251485, AL036980, AI475134,AW268768, AI813914, AI281762,AL119748, AW085799, AI811863,AI469811, AI862142, AI680113,AI889203, AI874109, AI493248,AI802542, AW301300, AI687465,AW026882, AI934036, AI269696,AI889839, AI609592, AI583316,AI492540, AI671679, AI570384,AA528822, AW167776, AI753683,AI610645, AI696398, AW235035,AL042753, AI539771, AI969567,AI919058, AI500659, AA508692,AI445025, AL044207, AI349226,AI687375, AW104724, AI591311,AI624668, AI754897, AI569616,AL121014, AI684265, AI687415,AI818206, AI952114, AI149592,AI536685, AI872711, AL047041,AI318569, AW075351, AI620868,AI909666, AI612913, AW118557,AI859733, AI609580, AI345860,AA585422, AI609331, AI446628,AW183130, AI696846, AI343059,AI635942, AI432969, AI432229,AI690426, AI679764, AI345744,AI624859, AI636445, AI334902,AI340519, AW302992, AW002342,AI619502, AI446606, AA938383,AI282281, AI907061, AL119828,AI269205, AL036240, AI282903,AI800384, AI583445, AI366991,AI804585, AI309401, AL041573, I48979,AL050393, AF090900, L31396, L31397,AF125949, S78214, AF090934, I89947,AF113013, AF118070, AF078844,AL080060, Y11587, AF090901, AF118064,AF113694, AL133016, AF113691,AL133640, A93016, AF090943, AL117457,AL133606, AF104032, AR059958,AL117460, AF090903, AL050146,AL137527, AF113689, AL110196,AF113690, AL049938, AL049452,AL050149, AL110221, S68736, AJ242859,AL122050, AB019565, AF090896, I89931,AF113676, X84990, AL050108, U42766,A08916, AF106862, AL133075, AL050116,AF113677, AL049314, AL122093,AF113019, AL049466, AL133557,AL096744, A08913, AF017152, AL137283,AL122121, AL080124, AL080137,AL050277, AL133565, E03348, AL137557,AL133093, I48978, AF158248, AF113699,AL137459, Y16645, Y11254, AJ000937,AR011880, AL133080, X63574,AF177401, AF111851, E07361, AL049430,AL122123, U91329, X82434, AF091084,AL050138, AF125948, AF146568,AF097996, AL117394, AL110225, E07108,AF079765, AL049300, AL137550, I49625,AL117583, U00763, AL133560, A65341,S61953, AF017437, AL117585, X70685,E02349, AJ238278, A08910, AL050024,AL117435, AL049464, A08912,AL049382, AF067728, AF118094, I33392,A58524, A58523, AL133113, AL137538,AC007390, AF183393, U72620, A77033,A77035, Z82022, E05822, A08909,AL122110, AL137271, AL122098,A03736, AF091512, AC002467, A12297,AL049283, X72889, AL137648,AL137463, AF061943, U80742, I03321,AF087943, Y09972, I09360, X96540,I42402, AL080127, X93495, AL110197,AL096776, X57961, U35846, X65873,AL080159, U67958, I00734, AL137521,AC006336, E00617, E00717, E00778,AC006371, AF111112, AL110280,X98834, U49908, AC004093, AJ012755,AL133014, AL133104, I26207, AR013797,AC007172, AL022147, AL133568,A93350, E08263, E08264, I17767,AC004686, AL133072, AC006840,AC004690, L13297, I66342, AL137523,AL137560, E15569, AR000496, U39656,U58996, AF026816, Y14314, AC007298,AL137533, X83508, AC004987,AF153205, AF061981, Z37987, AL133077,AL137476, AF119337, AL137556,AF100931, A08911, AL122049, AL137526,Z72491, M30514, U95739, AL133445,AF000145, AF057300, AF057299, A45787,AL050172, AF026124, and AC004200.HBGDI808711246951-106915-1083AI950010, AW014786, AI694670,AA171820, AI862334, AA604593,AA171724, AI697768, T11223, R41784,AI023823, T10688, and AI802300.HELHB888812256321-330515-3319AI671143, AI580905, AI632369,AA773823, AW370310, AA773263,AA313515, AA985166, AL121153,AI382884, AA181724, AI469456,AA024936, AI750406, AAI55593,AA024853, AL120063, T78993, T78788,T82245, AA155607, AI750407, AL134372,AA670080, L25219, AA247838,AF114487, AF114488, AF064243,AF064244, U61166, AF127798, AF132672,AF132478, AF132481, AP000311,AP000117, AP000193, AP000049,AP000116, AP000050, AP000048,AP000308, and AP000115.HTEMV668911522611-84715-861AI149647, AA430041, AA430250, andAW372558.HMTAJ739010639891-95415-968AI831613, AI870169, AI924408,AI368905, AI284115, AW168626,AA678670, AW068406, AA568895,AW198110, AA627558, AA857431,AW068493, AI284116, H19069, AJ010119,AF074714, and AF074715.HE9TD31918158451-94115-955AI475682, AI439613, AA815076,AB033082, AF132480, and AF132479.HGBDG559211413631-52115-535AA368408, and AW614226.HOUHL519311259141-63315-647AA431822, AI341790, AW295199,AI656610, AW292290, AA037543,AA431419, AA974280, AA815270,AA037457, AI651702, AA583011,AI208605, AI419858, AA620408,AA417333, AA417321, W28051,AA251183, AI917695, W28536, andAI024754.HEOPP679410201191-95915-973AA641653, AF181972, AF181973, andAL137592.HKAOV719511654231-74315-757AF123303, and AF004161.HDQID909611377521-100715-1021AW300598, AA669095, AI948608,AI797687, AI718165, AI129358,AI765613, AA114888, AA504203,AI701050, AI890342, AA974370,AI023212, AA504439, AI935316,AA252310, AA464174, D57415,AA280044, W46279, D54675, AA165321,AI420451, T71333, N69756, AA832206,AA521314, AA114887, AA877638,AW196653, AI027401, AI807828,AA863081, AA995204, AI831132, Z28882,Z40146, D57019, AA464762, Z25261,N87679, Z19443, AI918466, T71487,F00129, D56990, AL047889, AA278335,AW369458, W46278, AA743770,AL047888, AI809238, AA767219,AW025464, AL110306, AW302960,AW162194, AL036802, AW087445,AW071417, AI699865, AW163823,AL036274, AL041562, AI254727,AL119863, AW238730, AL045500,AI783861, AI909697, AI340603,AI537677, AI433157, AW163554,AI698391, AI929108, AI619502,AI802542, AW026882, AI783504,AI620284, AW198075, AI860783,AL079963, AA640779, AL047344,AW051088, AA572758, AL041772,AI699011, AI345180, AW082113,AW269098, AW268251, AI064830,AW161156, AL039086, AW020693,AI349645, AW268768, AW300782,AI349933, AL036403, AL119828,AI923989, AW300889, AL119791,AI497733, R36271, AL036396,AW172745, AI521012, AL041150,AW150578, AL079960, AI340519,AL048656, AL135022, AL080046,AI610293, AI567351, AW162071,AW079572, AW169653, AW074993,AI636456, AI284517, AI433976,AI697324, AW160916, AW268253,AI569583, AL038505, AL040169,AI312152, AW301409, AI345735,AI468872, AI524671, AI349937,AL036638, AA613907, AW089572,AI863382, AL047042, AW080402,AI270183, AI884318, AL037030,AI926878, AW023859, AI890507,AI564719, AI568114, AW068845,AI815232, AI868931, AW020419,AI624293, AI683395, AL039716,AI499285, AI886753, AL037582,AL037602, AL037463, AI815855,AI933589, AW004886, AW161579,AI312428, AI538850, AI307604,AI702073, AA427700, AI873644,AL046931, AW059828, AI499963,AI866469, AI610645, AL037454,AI349614, AL038605, AI343112,AL080045, Z99428, AW129106,AL134830, AI348897, AI862144,AL040827, AL121014, AI349598,AW118518, AI560012, AI473536,AI633125, AI307285, AI539153,AI627988, AI309401, AI567582,AI539771, AI500523, AI335208,AI866608, AI537244, AL036146,AL119049, AI521560, AI889189,AA449768, AI340582, AI500659,AI269862, AI287449, AI625464,AL121365, AI801325, AA804740,AW103371, AW022808, AI358701,AI613038, AI859991, AI538980,AI754897, AA580663, AI813914,AW078839, AA528822, AI866770,AI500662, AI917963, AL040241,AI349004, AI570807, AI352274, F27788,AL046944, AL043975, AI570966,AI635492, AL042745, AI682841,AI475371, AW149236, AI538716,AC002350, AB033082, AF132480,AF132479, I48979, U35846, I09499,I89947, AL122050, I48978, Y16645,AL096744, AF090903, AL110196,AL117457, Y11587, AF177401, AF090943,S78214, AL122093, A08910, AF100931,AL133075, AF146568, AL050116,AF090901, AL137550, AF090900, A08916,I89931, AL133606, AF078844, A77033,A77035, AL137271, AL133560, A08909,AL049382, AL137538, E07108, AF090934,AF183393, AL133565, AL137557,AL133640, AL110221, AL133016,A08913, E12747, AF113019, AL133557,AF113013, AF090896, AR013797,AL049452, AF113691, AF079763,AL137533, AF104032, AF113694, U87620,AF125949, AF097996, AL049938,AL137478, AJ000937, AL049283,AF125948, AL050146, I00734, S68736,AL117435, AB019565, AL049300, E05822,AL137459, AL122100, E00617, E00717,E00778, AL080124, AF067728, AL050149,AL050277, E06743, AL133080, I33392,U58996, AL117460, AL137488,AL080148, E02221, AF091084, AF113690,X70685, E02349, Y11254, AL137480,AL049430, AF111851, L31396, AL137527,AL050393, AL137548, L31397, AL122121,AF118090, M27260, AL050108, U42766,A58524, A58523, AR038854, AF017437,I49625, AF118064, A65341, AL050024,AF118070, AF111849, X63574, X72889,AL080060, AF106862, AR059958,AL133113, AF079765, AF057300,AF057299, AF017152, AF113699,AL050172, Z82022, AJ242859, X84990,A12297, AL110225, AL137294, AF032666,AL137479, A93016, AL122110,AL137283, AF118094, A65340, AF158248,AR011880, AL133067, AF031147,AL122098, AL137529, A03736,AL137521, AF067790, AF087943,AL049314, AL117649, Z37987, U80742,U91329, X83508, U72620, AL049466,AF113677, X79812, A08908, AL096720,A08912, X82434, AF113689, AJ238278,AF113676, AJ005690, AR020905,AL117583, U78525, AF126247, AF153205,AF026124, U68387, AF081197, AF081195,S61953, AF159615, Y09972, AL050366,AL117416, A07647, Y07905, AL122123,AF210052, AL080158, AL023657,AL080137, AL050138, I68732, AF100781,AL137526, X87582, AF054599, U96683,AL133568, AF185576, AL117394, A21103,AL080074, AF106657, AF030513,AL137292, AB016226, A18777,AL133093, AL080159, Y14314, AF061981,AL050155, X65873, AF119337, E03348,U67958, AL137560, AF106697, AF139986,AL050092, AL049464, X53587, U88966,AL080234, AB007812, AL110222,X98834, AL137463, X81464, AJ012755,AR038969, X80340, U00763, E15569,AJ006417, AF026816, E07361, S36676,and AR000496.HFRBN819711825521-409915-4113HFKJW019811871341-47515-489HSDFL639912193001-257815-2592AI811010, AW182310, AI669944,AA553658, AI149982, AW002490,AI138253, AI015322, AI417803,AW262574, AI338004, AW072451,AW073593, AI093838, AI027630,AI655339, AA922787, AI638037,AI095502, AW439551, AA913807,AW305307, AL042064, AA977416,AI205666, AA036837, AA036780,AA447978, AA356059, AA382934,AI611255, AI401469, AI280408,AI522175, AW263492, AA320515,AA382259, AA382933, AW450423,AA927003, AI933504, AI990172,AA552283, AA330729, AA382258,AB015228, AL110274, AL110299,AL137418, AB015226, AB015227,U60063, and X99273.HPJET901008365031-54215-556T03269, D58283, D80022, D80195,D80193, D59927, D51423, D59619,D80210, D51799, D80391, D80240,D81030, D80253, D80227, D80196,D80188, D59275, D80219, D80366,D80043, D80038, D59859, D80166,D80212, AI905856, D80269, D50995,D59889, C14429, D50979, D80134,D80378, D80024, D80045, C75259,AW178893, C14014, D59502, D57483,D59787, D59610, D51097, D80164,D80241, F13647, D80268, C14331,D51060, AW177440, D58253, C15076,D59467, D80949, D80168, C14227,AA305409, AW378532, AW178775,AW179328, C14389, AW352117,AW352158, AW177501, AW177511,D51079, AW178762, D51022, AW366296,AW360811, AW176467, AW375405,AA305578, AW377671, D59695,AW179220, AW360844, AW360817,D81026, AW375406, D80248, AW378534,AW179332, AW377672, AW179023,AW178905, AA514188, D52291, D80251,D80132, AW352170, AW352171, D80522,AW377676, AW177731, AW178907,AW179019, AW179024, AA514186,D80133, AW178906, AW177505,AW179020, AW178909, AW177456,AW179329, AW178980, AW177733,AW378528, AW178908, AW178754,AW179018, AW179004, AW178914,AW178911, AW367967, AA033512,AW352174, D80302, AW178774,AW177723, D80439, D80247, T48593,AW178983, D51103, AI535850,AW367950, C14975, AW178986, D45260,A67220, A62300, A62298, D34614,D26022, X67155, A25909, A84916,D89785, A78862, AR025207, D88547,AJ132110, AB012117, Y17188, A85396,AR066482, X82626, A85477, A86792,X93549, I19525, A44171, AR018138,X68127, AF135125, U87250, AF058696,AR008278, Y12724, AB028859,AR064240, A94995, D88507, AR008443,AB033111, I18367, I50126, I50132,I50128, I50133, AR066488, A82595,AR066490, AR016514, D50010, D13509,AR060138, A45456, A26615, AR052274,Y09669, AR060385, AB002449,AR066487, A43192, Z32749, A43190,AR038669, A30438, AR008408, U87247,U79457, AB023656, AR060133, X93535,and AR008382.HEMFC611018365141-70115-715AB015226, AB015227, AB015228,AL110299, X99273, and U60063.HDTBR5010211743511-45315-467HACCH941038471431-139915-1413AI093369, AW292321, AA972431,N40174, AA746376, AA130392,AA286750, AA287684, R71586, R71568,R71587, H03136, H03946, R71567,AI471079, H97311, AA365025, AF039686,AF118670, AR034800, AF081916,AL161458, and AL161458.HE8TI3910412234811-282015-2834AI681280, AA251688, W60548, Z45463,H86769, F07768, H86760, Z42543,R15292, AW339546, H86765, Z44339,H86774, H86962, AA091738, R58217,F06562, AW377760, and R21166.HEGAP3210511403931-51215-526AI814253, AW338148, AW192239,H72014, and AL137708.HCWFU661068530051-39215-406HUSYI291078531491-76315-777AA209183, AA811369, AA873372,AA304836, AW189673, AA326319,T11336, AI459087, H00191, andAL120282.HMEFT6610811341311-91915-933AI269592.HKAAR711098630231-96315-977AW068487, AW239526, T19084, R73854,AA984561, AA190644, R22358, R14385,AI968382, AA773650, AA425030,AW237628, AJ242972, and AJ242971.H7TBC951108659221-69215-706HAPPX521116374931-236815-2382T47026.HBGSJ1311211523261-74415-758W22401, W22336, and W26219.HFKLX381138802201-29515-309AL136383.HTLGP1511411653621-94915-963AW173250, T08271, AI372624,AA350545, T10079, AA349847,AA351693, and AF060173.HMEGH4611510921581-71315-727AB033054.HE8PY2911611294881-71015-724AI271550, AI753504, AA809220,AW081079, AI264068, AW386283,AI219556, AW082138, AA455733,AI382746, AA548778, W78099,AA431230, AF092137, AF100751, andAF040252.HTDAB171178903841-59115-605AW410520, Z21669, AA334183,AW376427, and AC011078.HCFCF4711811999311-184215-1856AL038713, AI264673, AI685116,AI887321, AI924175, AA189081,AI610776, AI818151, AI367384,AW157413, AI925647, AA767353,AA778304, AI559442, AI147839,AI110720, AL133942, AW089655,AI811494, W49501, AW084901,AI376984, AA085707, AI110640,AI132962, AW177120, AA493998,AI862874, N64574, AI675848, AI858607,N24958, AA807609, AA902828, AI749571,AI568919, AW074001, AI827133,AI817158, AI561208, AI250812,AI983921, AW177226, AI082077,AA130536, AI819528, AW139132,AA055654, AI027421, AI801377,AA724159, AI610326, AA932087,AA846188, AA864823, AW085676,W58442, AI453790, AI499286, AA659014,AW151307, T16214, AW090210,AA581433, AW262471, AI973178,AI189033, N26540, N76274, AI984510,AA709024, AI733728, AA581340,AW177317, AL044349, AW167452,AA130476, AA160519, AI811854,AI921101, AI369914, AW168798,AI479035, AA911409, AI346802,AI439860, AI034217, AW193992,AA493735, AI927861, AA287329,AW440317, AA129986, AI683685,AW102963, AA854527, AI246569,AW177237, AA346162, AA745961,T62931, AA889273, AI872415, W85828,AA847621, AA946637, AI197934,AW167319, AA907819, AA953572,AW272376, AL041411, AI862212,AI025602, AA092309, AI334099, C17235,AI439415, C06012, AI632138, AW130042,H20876, AI937684, AI538654, AA470572,AW023397, AI061131, AA811111,T69889, AA342331, AI874256, AI564253,AI440018, AI095849, AA157033,AI051341, AA771730, T57073, AA771711,AI926394, AI358417, AA174085,AI952804, AI860964, AI499811,AA368610, AI591332, AA136576,AA633390, W58428, AI619818,AA782144, AI291353, AA604138,AI393148, AA862481, AI863415,AA694579, AA176355, AI917243,AA169142, W33199, AA643785, W87732,AI285660, AA099788, AI955464,AI088796, AA347740, AA350922,AA961590, AA166854, AI871616,AA779937, AI290861, N77920, AA070814,AA508451, AI869945, AI051363, T06932,AI271865, C17730, AI813517, AI627862,AA132716, AI457328, AW235478,N20521, AA460715, H06476, AI336355,AA121916, AL119355, AA505616,AA598990, AI567544, AA559950,AW090739, AI949496, AI362951,AW168195, AA041332, N94967,AI004961, W31567, AA501873,AA629837, W19865, AA328438, W04638,AI310239, AI540098, AI375417,AW272291, AI086865, AI803229,AA876415, AA132536, T06365,AL031656, AL033379, AC002564,AC007381, AC005484, Z84719,AL023513, AL049734, AC007514,AC003084, AC004069, AL049867,AL049635, AC005181, AB017651,AC004050, AC004384, AL031407,AB019441, AC002349, AC007630,AC004650, AC004748, AC006463,AB020872, AC009405, AC003029,AL078477, AC006327, AL078638,AL135960, AJ131016, AC002385,AC003001, AL031054, Z98052, AL050348,AC006387, AC006198, AL132800,AC002079, AF207955, AP000355,AC004677, Z95325, AC007677,AC007358, AP000365, AL078604,AC005562, AC004875, AP000014,AB014082, AL121577, Z95328,AC003091, AC004022, AP000532,AC006396, Z97181, AC006374,AC005901, AC003085, AC000119,AL110505, AC006370, AL035542,AL009029, AL021940, AC007262,AC004891, AC003106, AC002122,AC004813, U96409, AL035088,AC003109, AL035397, AL035555,AC002458, AC005682, AC004053,AP000946, AJ007973, AC007671, Z70758,AL109759, AL121652, AC007402,AC008173, Z94721, AC004869, AP000962,AC006026, AC004536, AC002416,AL034377, AC007649, AC004664,AL080239, AC011198, AL033522,AC006296, AL109941, U70984,AC004822, AL136297, AC004757,AC005670, AL031671, AF130343,AC002471, AC002528, AC003013,AC005374, AC005993, AF117829,AL035633, AL049733, AL136018,AP000023, AL122007, AF095725,AL022100, AC004388, AL121578,AC004911, AL008722, AC004381,AC004701, AC007392, AC003100,AL023806, AF146191, AL033525, Z98304,AL022099, AL031115, AC007971,AL096867, Z94056, AC006239,AC005191, AC005053, AC004111,AD000091, AF121898, AC005799,AC007364, AC002485, AC002041,AC005521, AF130342, AL132994,AC004460, AC004999, AL109611,Z92542, AC005791, AC005082,AC004047, AL022401, AF152364,AL022097, AL121767, AL031313,AC006525, AJ251973, AC004987,AL031285, AC005820, AC006924,AC004749, AC006502, AF129078,AC005962, AC006971, AL078644,AP000473, AL034406, AP000455,AL022394, Z93019, AL021877, AC005297,AJ239321, AC006986, AF123462,AB023056, AC005922, AC007870,AC004223, AC003687, AL034399,AC007486, AB020868, AC006382,AL109659, AL049544, AL008627,AF207550, AF044083, AC005367,AC011594, AC004909, AL031684,AL031430, AL096772, AL050339,AC005681, AF055066, AC004894,AL049835, AL031737, AC003992,AC010722, AC007012, AC002379,AC004910, AC002492, AC006581,AL033380, AC007436, AC006369,AF070718, AC003692, AC004103,AC006504, AC006315, Z82212,AL009047, AC005284, AC004073,AC000114, AC004506, AL136295,AC008014, AL035700, AP000949,AC000109, U80017, AC006961,AP000033, AC003667, AC005913,AC005164, Z93018, AC006344,AC007377, and AC004178.HDQHB1911911947981-158615-1600AW195239, AW149418, AA461376,AW005579, AI392913, AI378013,AA461199, AA779830, AI860240,AI436586, AA576717, AA147800,AI276889, AW337924, AA459985,AI264931, AI203549, AW104319,AA460078, AI377235, AI291778,AI925811, AI094031, AI612894,AA147758, AA639492, AA037273,R61563, AA463275, AA767986, AI289261,AA768536, AW166255, AA832298,H59980, N47107, AA157070, AA508841,AI360737, AA417605, AA628790,AA731955, R35919, AI273576, AI301339,AI682196, AA463188, AI208175, H59937,D20738, AI391726, W42645, R14353,AI271983, N30324, T53535, AI073411,H57854, N56657, AW001677, R35723,N50317, AA514325, AI242094, AA969269,AI808646, AI280324, AA360254,AI699261, AA263061, T77863, AA551384,AA741518, AA906804, AI091790, T11446,AI208678, AA031793, AA505982,AA214523, AA428834, T12550, T11445,AB026289, and AR044150.HAGDN5312011291541-155315-1567AI671311.HUFDB7412111626721-153515-1549AW411339, AI978930, AW090612,AI521584, C05134, AA025277, AI281896,AA190389, R80748, F13061, AA190814,T75371, R80747, AA074211, AI570569,AA296815, AA024667, F10653, AI370565,AW197990, AI254860, AC002117,AC002425, AC004671, AC005736,AC005899, AC005757, AL034549,AC003048, AF001552, AL031591,AC007637, AC009363, AC005837,AC004813, Z83822, AC005568,AC003101, AC005669, AC003010,AC004887, AL035684, AC008079,AC006312, AL022319, AC002302,M19364, AL109758, AL008583,AF111168, AC005071, AC004916,AC005088, AC002565, AP000252,AC005363, AC007685, AL009051,AL034417, AC007565, Z82245, Y14768,AC005606, L34079, AL022396, AL034423,AP000505, AL049871, AL117337,AC004104, AL024509, AC006026,AC002314, AC002352, AC005295,AL080243, AC009288, AC004963,AC004876, U63721, AL031737, Z82206,AC005666, AL021977, AC002472,AC004216, U91321, AP000501,AC006040, AC005183, Z98200,AL049709, AC007981, AC009464,Z97054, and AC005585.HNHFH2412210925671-101515-1029AW023111, AI792578, AA570740,AA483606, AW069227, AI634187,AA515728, AA084609, AA565911,AI457313, AA568204, AI187148,AA856961, AA644090, AA557136,W96522, AI192440, AI440117, AI369580,AI610468, AW161016, AI054419,AI245693, N34689, AW189113, AI889579,AW245860, AA602906, AI583142,AI275982, AI494417, AI679294,AI755057, AI679871, AA126005,AA502207, AI362442, AA653823,AI342183, AL037714, AI636734,AA622801, AI866970, AI185394,AA469282, F28845, AA526099,AA714011, AA501461, AA133332,AA449997, AA635433, AW274072,AA191418, AI434513, R98218, AI925869,AA808861, AA279649, AA598892,AC004656, AC005736, AL049831,AC007371, AC005777, AC007993,AC002470, AF124523, AL117694,AC004975, AF104455, AL078477,AC003043, AL049539, AC005037,AF196972, AC005274, AC007542,AL109984, D87675, AC007917,AC005209, U78027, AC005696, U47924,AC005920, AC007298, AC005701,AC005500, AL035420, AL035422,AC005399, AC008064, AC006006,AL034423, AC004491, Z94056,AC005523, AC003982, AP000103,AP000032, AC007546, AC005972,AC005295, AP000555, AL035209,AF111169, AC004919, AC002091,AC004905, AC006486, AC005324,AL133448, AC003108, AC005004,AC004659, AC000025, AC004816,AC005527, AC005591, AC007690,AC004815, AC007327, Z82244, U91322,AL021154, AC006961, AC005529,AC004551, AC003110, AC005536,AC005730, AL031767, AL022313,AC006354, AC003681, AC005189,AC007386, AL049570, AP000263,AC006285, AC005280, AC005599,AP000100, AC005277, AL031228,AP000210, AP000132, AL022334,AC007934, AL121769, AL109801,AF196779, AL050343, AP000503,AL049776, AC005200, AC005075,AL031597, AP000193, AC006312,U62293, AC009247, Z93241, AC004983,AC005726, AL136295, AL035697,AC007731, AP000117, AC005952,AL031431, AL109952, Z84466, AC002301,Z93023, AC005513, AP000036, AC005409,AP000313, AP000050, AC008012,AC007437, Z82206, AC007242,AL034402, AC004020, AC006487,AC005901, AC006480, AL035683,AP000141, AC005330, AC004647,AF134726, AF031078, AC002316,AC007842, AF030876, AC005225,AC002476, AC006057, AC005067,AC006271, AC004796, AC004813,AL022394, AC003029, AL078611,AC005251, AC007041, AC007421,Z83844, AL034549, AC004383,AC003035, AC005180, AC005625,AP000228, AP000140, AP000088, U89337,AC005231, AC004805, AC004967,AC002477, U76667, AC007664,AL022476, Z49236, AL035086, AC004526,AP000212, AP000134, Z84467, AL033397,AL034421, AP000248, AC007773,AL109963, AF064857, AF042484,AP000497, AC007021, AL035684,AL021546, AC006597, AC004025,AP000302, AC004859, AC000134,AC006449, Z97630, AC003665, Z99714,AC004033, AC005095, AC005915,AP000558, AC005837, AL117352,AC000111, AC007938, AC005300,AL022326, AL033527, AL078584,AL033521, AL023553, AC005318,AC004673, AC004821, U73630,AC006255, AP000114, AP000046,AC002559, AC007052, AP000694,AC005682, AC003046, AC005531,AC006211, AL031589, AC005484,Z97876, AL050341, Z98036, AL049757,AC005245, AC004106, AC006077,AC003070, AC005013, AL049643,AC005332, AC005058, and AC006035.HBGQT0312311881751-120415-1218AW193981, AA576536, AW439879,AA218860, AA587394, AW206358,AI735027, AI075695, AI749755,AW328242, AI073515, AI828816,AI283940, AA452508, AI741698, F25077,AA454093, AI280249, AI826261,AI567379, AA350150, AI251129, F26225,AI354257, AW129660, AA171893,AI357160, F26293, F36700, H24638,AI270014, AI952189, AA834233,AI689497, AI688448, F17480, Z38509,T11668, N93072, AW362737, T11669,AW273866, N93071, AW328241, andAF130979.HETLF2912411039591-46915-483AA960957, AI001155, and AC004664.HOUGD2912512049311-227415-2288AW243053, AI674036, AI160750,AA426073, AI299702, AI079952,AA405859, AI805286, AI828341,AW022161, W24408, AI421345,AW206919, AA351454, AW118669,AI263265, AA640181, AW134737,R13311, AW135904, AW008702,AA041245, R40791, T16675, AI418741,AI572229, AA040806, AI057025, N93105,AA954204, AF117948, and AL050031.HTEMV0912611282541-134715-1361AI818734, AA454060, AW268879,AA453640, AI377304, AI818733,AI681535, AI818743, AI741915,AA948041, AI198872, AW341578,AI267885, AA767746, AI677678,AI829855, AI677729, AW129267,AA947425, AA297313, AL041049,N67346, and AA889773.HNTNB1412711289641-71315-727F12661, T74243, R60839, AA349498,AA082976, and L22557.HE2KZ0712811498081-69515-709AA114993, AI696123, AI141657,AI986390, AI307294, AB004267,AF181984, AB023027, and D86556.HSIGN5712911054441-108615-1100AW408450, T78839, and AB029015.HLHBC3013011066541-77015-784AI366170, W21960, AA047627, W22094,AA047570, W22769, AA359453,AA359449, AA128575, AI220321, andW22833.HFBDJ1313111952171-135515-1369AI637673, AI982948, M86084, AW161837,AA568170, AA044809, AW157364,AI686957, AA191323, AI161414,AI140787, AI752687, R17699, AA742642,and AF030131.HTPGG2513212172081-277115-2785AI681123, AI741848, AI636347,AI973055, AI554720, AI768326,AI871117, AI333117, AI745311,AW340966, AW192924, AA706712,AI091179, AI677802, AI889659,AI688189, AI804323, AI298377,AA535027, AI830304, AI139157,AW089901, AA018361, AA410579,AA416567, AW073842, AW316637,AI827376, AA417232, AI372513,AA411560, AW001905, AI796719,AI400032, AI334363, AI452964,AI085075, AI888902, AI400560,AA308319, T33187, AI372512, AI332395,AA877699, AA485507, AA017127,R85136, H94860, T33188, AW016699,AA324901, AA988884, R18537, AI925753,AA993373, AI263531, W05059,AA282629, F29641, AA378627,AA625328, H58095, R17859, AA354334,AW337874, AI559961, AA282410,AA126985, AI014243, AI671403, F07835,R01402, AA485352, R41526, Z39066,F04091, R43109, W04796, R01401,Z42943, AA128150, AW375092, C04525,T25085, F17839, N74669, AW371533,AW058382, AW371557, AL117482,Z61430, AW612722, and AW662030.HSSMT341339112941-54015-554AA378627, F07835, and AL117482.HWWDN313411524301-125415-1268AI671062, AI023330, AW243448,AI990947, AW081367, AW391909,AA448391, AI984688, AA448394,AI283270, AW014216, AI344135,AA127530, AA335984, AA377148,Z42084, R12430, and AA400585.HCEPW851359113741-30215-316N83965, AA326737, and H14153.HMTAW8313610716021-48715-501AI908321, AA831896, AR058970,AR058968, A68194, and AR058969.HDMAV0113711946971-179615-1810AA639636, AI631313, AW295120,AA182022, AW085587, AI805706,AA181842, AA732237, AW275826,AI215758, AA482475, AI802881, N50527,AI582673, AI581648, AA687232,AA742653, AI678872, AI937512,AA043083, N95362, AA130860,AW303909, N50583, AA186576,AW149048, AA743626, AA972227,R82179, AA988476, H19646, AA872660,AA476734, AA040856, AA074515,W93062, AA649136, AA186539,AA443881, AW021342, AI039487,AA252656, H22712, AW006472,AI342673, N72971, AW316519, H01103,R82221, AW079113, W02736, H19647,H43529, F23483, AI681977, AA989257,AA357239, H01104, W21036, AA872966,AI908321, N93561, R57677, AA984133,AA306556, AI423661, AL120709, D19838,AW295339, AW403073, AA100099,H38954, AA297943, AA130916,AW007473, A68194, AR058970,AR058968, and AR058969.HDPSR741389113961-70915-723HHEZT5813911606571-219715-2211AI653410, AI808115, AW103846,AI954664, AI809351, AL038027,AW365646, AA598677, AW118178,AA461436, AA461119, AI915224,AI452666, AI962619, AA384806,AA021033, AA317505, AW179253,AA054648, AI949041, R59010, H21414,AW450327, AI084523, AA813068,AW021195, AI242505, AA768247,AL046262, AL138455, AL035847,AA714441, AL042853, AI570389,AI821062, AA804728, AA760655,AA665057, AL042753, AL045943,AI493858, AW079656, AI027175,AA504562, AL049552, AF162270,AC004834, AP000208, AP000130,AP000247, AL049557, D83989, AL022315,AC009501, AL031281, AL109758,AC006313, AC002464, AC005291,AC002565, AC004837, AL022313,AL031393, AC007172, AC018767,AL021393, AL022165, AF222686,AC004987, AC005091, AC006112,AL110269, AC006115, AC011331,AC005156, AC002430, AC005088,AC004686, AC007384, AC005902,AP000697, AC004883, AC006501,AC006039, L30117, AL035458, AL035461,AC010206, AL035067, AL034417,AC005057, AC002472, AP000009,U96629, Z83840, AF053356, AC004383,AF091512, AP000344, U67211, U66059,AC000004, AC003070, AL096791,Z82206, AC005081, AC008067,AL034400, and AC007284.HTLDU051409116491-58915-603AA437044, AF113527, AB023062, andAF113520.HTLET5614111897211-139015-1404AI968198, AI655275, AL044119,AW003563, AA397903, AL044168,AL044118, AW016204, AA625705, andZ74696.HTLCA951429116551-114815-1162AI028227, AI798166, AI968058,AI962770, AC012616, and AC012616.HTEJT8614310905171-164415-1658AI290635, AI097065, AI811210,AI655508, AI655489, AA402182,AA373381, AA948283, Z21336, AI984919,AI825441, Z21335, and AW467143.HTEMA5414411349191-148315-1497AI954673, AI220421, AA813119,AA382989, AI024406, AF113526,AB023063, and AF113519.HTLGJ1714511355181-57315-587HOUES641469181191-30415-318HMSCD151479822501-64215-656AA864599, AA828277, AW270419,AA761244, AA262754, AA779760,W37119, AA206843, Z42584, AA206842,and AB011126.HDQDX2014812234741-268815-2702AI905612, N63562, N75655, N94726,AA297704, N64807, AW265468,AA601376, AW162314, AW162332,AI318548, AW410844, AL042753,AL121039, AI702049, AI547110,AI919048, AA845690, AW021674,AA280886, AW328185, AI076729,AW327673, AL138455, AA526542,AL118628, AW148821, AI754926,AI344906, AI064968, AW419201,AI065031, AI129421, AL041924,AI090377, AI252005, AI254463,AW409621, AA313025, AA610644,AA760655, AI797998, AI821901,AA557945, AA595661, AI860423,AA831426, AW021399, AA765899,AI174827, AI251024, AL042667,AL042670, AI003068, AW439224,AL042377, AI567676, AL134524,AI905408, AI570067, AA527633,AW022796, AI025355, AA525753,AI133355, AI224583, AI609992, N95424,AI281622, AI446618, AI745666,AA632355, AI791659, AW085626,AI815425, AI819419, AI884404,AA828840, AA631915, AI049845,AA133568, AI921744, AI114755,AL044966, W27084, AA524604,AL039436, AA084439, AI279417,AW023975, AW239465, AI753131,AW275432, AA847341, AW177869,AW192930, AW409626, AI049643,AW338525, AI828721, AA935827,AI857834, AI348780, AI744963, F35374,AI434103, AA601712, AL138262,AL138431, AW272815, AI445699,AI270280, AI888050, AW029626,AI150934, AL042731, AI003391,AL044701, AI924950, AA535558, H86399,AI733523, AW020612, AW069769,AA568433, H47461, AA167656,AL079698, H62123, AA196994, T03928,AI078409, AA015948, AI049892,AI815770, AW303052, AA558488,AA315052, AF169035, AF085233,AL049694, AC006084, AC006581,U93163, AC007298, AL022315,AC007050, L78810, AF196779, AC005081,AL031680, AC004891, AF113676,AC004895, AC005921, AC005015,AC005701, AC005291, AF207550, Z98941,AC006079, Z83844, U95739, AC005553,AL031846, AL021391, AC004263,AC004808, Z99716, AL008582,AC006312, Z93241, AC005519,AL121603, AC006115, AL080317,AL049538, AC002558, AL121595,AC004703, AC007172, AC006441,AL022238, AC007227, Z82206,AC004702, AP000116, AC007392, Z98752,AC005730, AL031311, AL022165,AC005039, AL021155, AL035551,AC005829, AC007055, AP000555,AC007237, AC005484, AF053356,AC005365, AC003982, AF038458,AL137100, AC004966, Z95152, AL049872,AL122020, AL049869, AL049557,AC005224, AC004797, AC005971,U85195, AC006538, AL078463,AC005585, AL031283, AL034549,AC007193, AF001549, AE000658,AC008372, AC006978, AC005251,AC007536, AC005695, Z82184,AL022723, AC007384, Z98949, AP000692,AL035587, AC006953, AL096701,AC005409, AC006013, AC005944,AL031433, AC005911, AL096703,AC005839, AP000210, AP000132,AC003108, AC006039, Z93017, AP000516,AC004659, AC006111, AL022721,Z77249, AF111168, Z94721, AP000212,AP000134, AL050338, AC005527,AC007390, AC005216, AF187320,AC002036, AC009510, AC005666,AC004673, AL021808, AC005225,AC004213, AL021453, AL008725,AC007242, AL008726, AP001046,AC005618, AC007878, AL031575,AC002350, AL049699, AL121653,AC003013, AC006966, AL031228,AL109984, AC002351, AC006509,AC005175, AL049569, AC006211,AC003037, Z98200, AC004805, AP000201,AL031229, AC006196, AC004913,AP000248, Z84466, AL022476, AL034379,AC005327, AL008712, AC004663,AC005017, AL021579, Z95114,AC002477, AC007537, U91322,AC007676, AC005900, AC002565,AP000097, AP000348, AC006277,AC011311, AC007546, U07563, L44140,AC007684, Z68284, AC003111,AC005529, AC005368, U07561,AC004000, AC004883, AL022316,AC005500, AC005369, Z82976,AC006571, AP000505, AC005821,AL034369, AC007057, AP000049,AL034421, AC007510, AL031594,AC005041, AC004821, AP000338,AC005071, AC008134, AP000030,AP000104, AL035086, AL132712,AL096803, AF064862, AC004383,D84401, AC003689, AF024533,AC002980, Z82211, AC005156,AC006539, AC007731, AC000090,AP000511, AP000211, AP000133,AC002306, AC003029, AD000092,AC003043, AC006275, AP000216,AC006157, Z84474, Z82208, AC005048,AC002133, AC006458, and AC005697.HLTHP8614911104571-183715-1851AA663589, H16878, R56761, H84971,H16267, Z44040, Z44727, AA363673,T95749, H09671, AA809486, R91239,N50504, AA834548, N58608, AA872305,and AF161420.HMSOL5215011827151-128915-1303AI911515, AI360955, AW028045,AI796049, AI609712, AW195544,AI184337, AI470056, AI361065, N34939,AI017177, AI038779, AI440241,AI651451, AA789292, AA854683,AI765258, AI702748, and AA384884.HAHGD3315112198191-221115-2225AA058874, AA778668, AW408302,AA205441, AI127967, AI887424,AI479839, AI697118, AI313146,AI992196, W01185, AI828049, AA196613,AA988948, AW410815, AI569584,W44348, AI754108, AI858157,AW161181, AI809391, AI418172,AW378448, AW378426, AI554365,AA064738, N41640, AI129499,AW080159, AI079282, AI624100, Z43369,N29634, AA160313, AI476090, AI748939,H40577, W52734, AI828528, AA984486,AA723784, W73106, AI811703, AI982708,AA610087, H45144, AA975653, D31100,AA064739, AA226860, W79308,AA976982, AA961166, T35036,AA044358, AA969553, AW380268,AA961174, AW380277, AA192146,AA348984, C21229, R16767, AA348983,AI536815, W59987, AI632654, T35774,AW292300, T08259, AI368455, AI701960,AI969407, AI559435, AI285876,AI243806, AA232389, AI476704,AW410816, AA588076, AA918087,AA989405, AA639598, AA862740,AA160321, AI904952, Z39439, H46434,R10018, AA227210, N78192, AA977128,F35561, R40005, AW050898, H14918,AA852243, AA043840, N74228,AA719438, W45701, AI879824,AA249020, H79060, AA852244, T80752,H45445, AA353126, AA320390, R45977,AI187729, AW385198, AI364441,AI284294, AA862396, T81225, H46980,AA642250, R57404, H14625, AA295773,W79389, T12193, W52140, T81300,R13945, AI918467, T47464, AA043841,Z43504, AA404490, AA627362, D45662,AA205277, AA196528, C21486, AI739190,AI123849, AI205909, AI025702, andAF113249.HHEHC531529217831-89615-910AW408302, AW410815, AW161181,AA160313, AA226860, AA044358,AI632654, AA232389, Z43369, AA249020,T35774, AA852244, AA064738,AA295773, D31100, R13945, AA205277,T47464, T08259, AI904952, AF113249,AC009427, AC009427, and AC009427.HE2PB0115312066651-175315-1767AA443164, AW020571, AA532437,AA315526, AW118680, AI167234,AI890631, AA744921, AI401545,AW195349, AW152040, AW074628,H06431, R74385, AA862392, H23357,AA649701, AW362722, AA160546,AI880570, D62917, AA973615, H23246,H46816, AI819867, H00507, AA329368,R21787, R21742, AA463453, C05953,R25958, R26764, AA639080, AI494615,AW070869, H71752, T84284, AA767232,AA160663, AI824357, and AF161399.HOUDP5215412195221-173515-1749AA126458, AI091270, AA535353,AW129933, AI653554, AA809485,AI638693, AI208921, R73542, AW088345,R72907, AW105725, AA483641,AI828781, AI350843, AI970412,AI971578, AA121009, AI989884,AI671096, AI962165, AI632336,AI241787, AI580332, AA991236,AI587241, AA317304, AI655218,AA853441, and AI971684.HHGAE4715511278811-73315-747AW025529, AI475932, AW026010,AA886335, AA662803, AI056120,AW050607, AI885090, AI244837,AW449834, AI375435, AA878578,AA922036, AW197722, AA161103,AW058170, AI393408, AI056614,AA643750, AI560410, AI749095,AI720931, AI446208, AI913781, N52768,AI268967, AI277003, AA657904,AI914599, AA910277, AA485405,AI192693, AW050712, N52783, AI673692,AA631339, AA485566, AA158820,AI619710, AI560351, AI919380, N57590,AI832600, and N57604.HMCGL4515611653491-113615-1150AW025529, AW026010, AI475932,AA886335, AA662803, AI056120,AI885090, AW050607, AI375435,AW449834, AI244837, AA878578,AI056614, AA922036, AW058170,AW197722, AI393408, AA161103,AI560410, AA643750, AI749095,AI720931, AI446208, AI913781, N52768,AA910277, AI277003, AI268967,AA657904, AI914599, AI192693,AA485405, AI673692, N52783,AW050712, AA158820, AA631339,AA485566, AI619710, AI560351,AI919380, N57590, AI832600, N57604,T25136, and AA299927.HELEF1115711538841-132415-1338HETJX0415812122351-84415-858AA853282, AA370481, AA625156,AB025258, and AB025259.HSOBC0415911653571-119415-1208AA115298, AI741325, AI688227,AI819333, AA452504, AI742595,AI925664, AI174530, AA115338,AA563582, AA461615, AA807844,N94422, W58424, AA569395, AI095261,AI142563, AA687480, AI567500,AA479551, AA582573, AA779677,N24393, AI280806, AI081428, AI863187,AA988617, AA834079, AW302361,AI362861, AW273442, AA553678,AA150123, AI752480, AI312661, W52661,AI298150, AA463418, W72509,AA024450, W72139, W79868, AI037968,H39596, AI028169, AA477651, W02690,AI198327, AI952450, AA926794,AI087245, W74236, H98040, AW004736,AI870989, AI689546, W76066, AI332748,AA150031, AI349417, W80872, H99144,AI334346, W52767, AA496878,AW166280, R40403, H25985, AI357863,R55375, AA363023, AA378409,AA595996, AW104147, AI687489,AI376184, T32290, AI979074, N26307,H97338, N95244, W77880, AI917258,W25604, AA024802, AI536791,AA577352, AA328156, AA359865,AA367475, AA358275, AA461442,W80763, T09474, AA987427, AI611160,AA888165, AA595303, AI918172,W30769, AI201782, W21074, AAI87662,AA411955, AA090719, AA411956,AA935961, AA451977, AI371307,AW074526, N79974, N39751, AI635472,AI612934, AA478489, AA102215,AI802295, AI750502, AA496836,AL133116, and L07063.HE8PW8316010699801-121915-1233AA203427, AW024967, AA707382,N74230, AA875830, AI051580, AA975082,AI650913, AI651056, AA707184, N69106,AA938679, N69120, AA704705, AI283739,T72088, AW135507, R10286, R10287,AA010205, AW296131, AI085405,AA333931, AA873194, W01187,AF121182, AB002584, D38100, andAW611663.HWLEA481619276761-41515-429AA130828, AF169034, Z98752, andAF169033.HNHNP8116211291431-103615-1050AW366296, D80045, AW375405, D51060,C14014, D58283, AA305409, D80253,D80024, D80166, D59619, D80210,D80240, D80366, AA514186, C14389,D80043, D81030, D80133, D80247,D59859, D80212, D51799, D80164,D80219, D51423, D80022, D80391,D59787, D80195, D80188, D80248,C14331, D59502, D59467, D57483,D59275, D59610, D80227, D81026,D50995, D80196, T48593, D80439,D80251, D80269, D59889, D51022,AI905856, D50979, D80268, D80522,C15076, D59927, AA305578, D80038,D80193, D80241, AA514188, AW375406,AW360811, D80378, AW177440, D80302,C05695, C14429, AW178893, AW377671,D80157, T03269, C75259, AW178906,AW179328, AW360844, AW360817,D59373, D51103, AW378534, AW179332,AW377672, AW179023, AW178905,D51759, AW360841, AW378532,AW177731, AW177501, AW177511,D58253, AW352171, AW377676,AW352170, AW178907, AW378528,AW178762, AW179019, D80132,AW179024, D80134, D51250, AW176467,AW178983, AW177505, D59653,AW179020, AW178775, AW178909,AW177456, AW179329, AW178980,AW178914, AW177733, AW178908,AW178754, AW179018, C06015,AW367967, AW352158, AW369651,AW352117, AW178774, AW352120,D45260, AW179004, D80949, AW179012,D81111, AW378525, D59695, AW352163,D52291, AW360834, D59627, D51079,AW378543, D80064, AW177728, H67854,D80168, AW179009, AI525923, C14298,AW352174, D80258, AW367950,AW178911, AW177722, AI910186,AW378540, C03092, H67866, C14227,AA809122, D51221, D59503, AW178781,D58246, T11417, D58101, AW177508,F13647, AI525917, C14407, AW178986,T03116, D80228, AI535686, Z21582,C14077, AW177497, D59317, D80014,D59474, AW177734, AI525920,AW177723, C14973, AI535850, C14344,AW378533, AA514184, D51213, D59551,D45273, C14957, AI525215, D60010,AI525235, D60214, AI525227, C14046,AI557774, AI525912, AI525925, D51097,AI525242, T03048, AA285331,AW378542, AI557751, AI525222,AW378539, Z30160, C16955, C05763,Z33452, T02974, D51053, AW360855,H67858, AI525237, C04682, T02868,C13958, A62298, A62300, AB028859,A82595, AR018138, A94995, AJ132110,A84916, AR008278, Y17188, AF058696,AR016514, AR060385, Y17187,AB002449, A43190, X67155, D26022,Y12724, A25909, D89785, AR038669,A67220, A78862, D34614, AR008443,A30438, I50126, I50132, I50128, I50133,D88547, AR066488, AR060138, A45456,A26615, AR052274, X82626, Y09669,A43192, AR066487, I14842, AR025207,AR054175, D50010, AR066490,AR008277, AR008281, A63261, I18367,AR008408, AR062872, A70867,AR016691, AR016690, U46128, D13509,AR060133, AB012117, A64136, A68321,I79511, X68127, U79457, AF123263,AR032065, Z82022, A63887, andAR008382.HFIDL6816311236411-114815-1162AI805323, AI375172, AL119457,AL119399, AL119511, AL119324,AL134524, AL042544, AL043152, Z99396,AL042382, AL043168, AI431323,AI582910, AL038837, AL079794,AL037051, AL036725, AL037081,AL046052, AA631969, AL039074,AW151132, AL036418, AL042866,AW058275, AL045413, AI282249,AL119443, AL039085, AL039564,AL039456, AA580663, AL036858,AL039156, AW151974, AI927233,AL039108, AL038509, AI539771,AI432644, AI537677, AI494201,AL039109, AI804505, AI500659,AI815239, AL039128, AI446536,AI866465, AI815232, AI866691,AI801325, AI500523, AI538850,AW392670, AI358271, AI887775,AI582932, AI923989, AI590043,AI872423, AI284517, AI500706,AI635082, AI445237, AI491776,AW151138, AI289791, AI926593,AI889189, AI521560, AI500662,AI285417, AI582912, AI284509,AI539800, AW172723, AI440263,AI538885, AI889168, AI866573,AI633493, AI434256, AI866469,AI805769, AI434242, AI888661,AI500714, AI284513, AI888118,AI285439, AI859991, AI436429,AI889147, AI355779, AI623736,AI581033, AI371228, AI932794,AI491710, AI440252, AI431307,AI440238, AW089844, AI567971,AI866786, AI860003, AI610557,AI431316, AI242736, AI539260,AI828574, AI698391, AI887499,AW151979, AI539781, AI702065,AI539707, AI801286, AI885949,AI638644, AW089557, AI285419,AI559957, AI521571, AI469775,AI866581, AW074057, AI567953,AI815150, AI446495, AI867068,AI952433, AI889191, AI225248,AI698352, AI371229, AL036924,AW151136, AI440260, AI584130,AI355126, AI440236, AA761608,AL036241, AW189301, AI623302,AI866458, AI431238, AI469784,AW194509, AI370623, AL079741,AL042365, AI345688, AI436438,AI890907, AI357672, AI623941,AI554821, AI561170, AI586931,AI859644, AI924051, AI690946,AI371251, AI869377, AI866510,AI648567, AL119791, AL047849,AI866461, AI923046, AI433157,AI457113, AI434240, AI632036,AL047422, AI954200, AA641818,AI345010, AL048403, AW023072,AI610714, F36033, AL037094, AI250282,AL047398, AI648699, AI610616,AI887163, AW384394, AI049850,AI469764, AI284516, AL045166,AW019988, AI354981, AL039659,AI798359, AL038531, AI653829,AW074651, AI383510, AW238753,AI469754, AW075382, AW083572,AI371243, AI419455, AI537925,AW151970, AI049859, N25033,AL036196, AL119863, AW162194,AI560954, AL135012, AI439995,AI887785, AL039390, AI493559,AI963849, AI955221, AI566386,AI559976, AI250353, AI274759,AI432666, AL049048, T66952, N29277,AW245729, AI866419, AI433976,AW363220, AL038986, AL047611,AI349256, AI798456, AR060234,AR066494, AR023813, U77594, A81671,AR064707, AR069079, S75997, L10353,Y11587, AL133084, AF115410, AL133559,S82852, M85164, U72621, U30290,AL137479, AC002464, AF090943,AL133070, AJ005690, AF176651,AJ238278, AL137529, AF132676,AF061836, I09499, U83172, AR029490,AL133016, X70685, E12580, AL137533,AL122101, AL133049, AL049423,AB007812, AF180525, Z35309, AF013214,AF079763, AL023657, AR038854,AL133640, AL133655, AF090900,AF043642, AL110199, AF113694,AL122104, AL133053, AF060866, I25049,A03736, AL049276, AR020905,AF161699, AF067728, E04233, AL137554,AF146568, E13998, AL137521, M79462,AL133560, AF119336, I28326, D44497,Y09972, A07647, AL049466, A08910,AL110218, S54890, X61970, AF002985,A08909, U49434, AR034821, AL080129,U49908, A60092, A60094, AF031572,AF077051, A08908, X66862, AL110196,A77033, A77035, AF113699, X63162,AF068615, U51123, AF177401, S69407,AL117435, AL137268, AF098162,AL137550, AL049447, AL136884,AF113677, AB028451, AL133557,AL133015, AL133608, S53987, I32738,A18777, AF076633, AF067790, M19658,Y16645, AL080159, AL050172, AF106697,S77771, AR068466, AL133619, E02221,Z97214, AL080118, E05822, AL117587,E12579, A08913, AF080622, I89947,AL117443, AL110269, AF182215,AR068751, AL122049, Y10823,AF131821, AF112208, X67813, AC004213,AF026816, A08907, U76419, Z13966,A08912, AF090903, I25048, AL110280,M80340, D83032, A08911, AC004200,E12747, AB016226, AL133067, I48978,Y11254, AL122050, AL117460, AF061795,A58545, AF151685, I03321, AL137292,AF032666, AF090896, AF047716, I89931,AF113690, AL117438, I42402, Y10080,Z82022, AL117582, AF100931, AL137281,AL133062, AJ010277, AF016271,AF004162, U78525, AL050155, AL096751,S76508, AF061573, AB026128, X83508,U72620, I89934, I89944, I49625, A32826,A32827, X60769, AF148129, AL137560,AF026030, AF111849, AL117394,AF030513, AF002672, AL137555, X65873,AF104032, A18788, AF106945, E02914,AF175903, AC018767, AL137478,A49139, AL137459, AF094480, AF126488,AF124728, AL080140, AL136842, U73682,Y08616, AL096728, A27171, AL137259,AL133565, E01614, E13364, X06146,AF102578, M92439, L24896, Y13350,AF118090, AL137271, AF111851, L13297,U86379, and X59812.HUJCT0516411652611-163215-1646AW268357, AI889091, C15588, AI949350,AI056961, AI124874, R39133, AA887911,AW023386, AC003962, and AF155116.HTEGO051659325831-108615-1100AA059465, AA059211, AA731209,AA236961, T86500, T87461, AL024498,and M35862.HRDBH5816612267191-273115-2745AI863355, AI863364, AI674922,AI754389, AI056058, AW295190,AI623178, AW131720, AI949042,AI056059, AA356949, AI668970,AA620354, C14389, C15076, D59467,D80164, D59787, D81026, C14331,D51423, D81030, D80195, D80391,D80166, C14429, D59275, D58283,D59859, D59927, D80196, D80022,D80253, D59889, D59619, D80210,D51799, D80240, D80043, D80227,D59502, D59610, D50995, D80212,D80038, D80188, D80219, D57483,D80366, D80269, D80193, D51022,D50979, AW177440, D80251, D80241,AW378532, AI557751, AA305409,D80024, D80378, D80045, T03269,AW178893, AA305578, D80522, D51060,C75259, AW179328, D80248, C14014,AW352158, T11417, D51250, AA514188,D80134, C14407, AW369651, AW178762,AW178775, AW177501, AW177511,AW176467, AW360811, AA514186,D80133, D58253, D80268, F13647,AW352117, AW377671, AW375405,C05695, AW378540, AI910186,AW366296, AW360844, AW360817,AW377676, AW375406, AW378534,AW179332, AW377672, AW179023,AW178905, D80439, AW352170,AI905856, D80302, AW177505, D80132,AW352171, AW178906, AW177731,AW178907, AW179019, AW179024,D80247, AW360841, AW179020, C06015,D51213, AW178909, AW177456,AW179329, AW178980, AW177733,AW378528, AW178908, AW178754,AW179018, AW360834, C14227, Z21582,AA809122, AW179004, AW179012,D80014, AW352174, AW178914,AW378525, D81111, D51759, D51097,T02974, D51103, AW177722, AW177728,D80157, AW179009, D80258, AW178774,AW178911, AW378543, AA285331,AW352163, AW367967, AW178983,AW352120, AI557774, AW178781,T48593, D59627, D58246, AW177723,D59503, D59653, D45260, D58101,C03092, D80168, AW367950, AW177508,C14344, AI535850, H67854, AW378533,D80949, H67866, C14975, AW178986,D80064, C14973, AW177497, AI525923,AI535686, AW177734, AI525917, D51079,T03116, D59317, D51221, AA514184,AI525227, D59474, AI535959, AI525920,D60010, AI535961, AW378539, D45273,D59551, C14046, D60214, AI525235,AW378542, AI525242, AI525912,AI525925, AI525215, C16955, A84916,A62298, AR018138, Y17188, A62300,AJ132110, A25909, X67155, AF058696,D26022, AR008278, A67220, D89785,A78862, D34614, AB028859, X82626,D88547, Y12724, A82595, I82448,AR025207, X68127, AR008277,AR008281, A30438, A94995, AR060385,AR008443, AB002449, AB012117, I50126,I50132, I50128, I50133, A85396,AR066482, A44171, AR066488,AR016514, A85477, AR060138, A45456,I19525, A86792, Y17187, X93549,A26615, AR052274, Y09669, U79457,A43192, A43190, AR038669, U87250,AR066487, X64588, AR066490, D88507,AR054175, D50010, I14842, I18367,AF135125, A63261, U46128, AR016691,AR016690, AR008408, AR062872,A70867, D13509, A64136, A68321,AR060133, I79511, AB023656, AF123263,AB033111, AR032065, X93535,AR008382, AA159456, and AI990060.HSDGW221679344671-31215-326AF071071, AF170303, AF170304,AF077658, and AF071070.HNTMD7916811265941-64915-663AA305176.HCE5J5116911979001-279615-2810AA622725, AI022884, AI003586,AI420804, AI682458, H19352, AI808077,AA324320, AA855153, W19561,AA325225, AA324316, AA853261,H18913, H11459, AW014428, AI148776,AI659608, AI675556, AW006008, T26639,AA853262, and U71294.HHEFQ4217011514821-126115-1275N21687, AA447158, W42824, AA046565,AI693814, AI092205, AW377670,AA443786, AI160460, AI808004,AA160291, N34495, R70372, AW008266,AA393368, AA769319, AA977890,AI332771, AA089858, AA367462,AI806021, AA160290, AA164499,AA357550, AA909368, AI337806,W42734, N31093, AA521439, AA600712,AI587601, AI491781, AA047086,AA933922, AA078980, N49165, AI623941,AW163834, AW020328, AL036705,AW151974, AI249946, AI588892,AL138406, AW021717, AI648494,AW051088, AI345131, AI619820,AI500061, AI434731, AA641818,AI524654, AI860027, AW020046,AL120307, AL038505, AW087987,AI919500, AI334893, AW020397,AI401697, AA834534, AI538850,AI698391, AI659043, AL036901,AI440238, AI890907, AW075382,AI799313, AI866465, AI696340,AW151132, AW083111, AI590043,AI627714, AL134840, AW195253,AI473536, AI499325, AL138399,AL047100, AI884318, AI619525,AI249389, AI244249, AL048323,AI590755, AW161098, AW129117,AL048340, R36271, AI635634, AA765029,AA229532, AI491852, AI859991,AI690813, AL039730, AA503384,AW196720, AI373276, AI741158,AI289791, AI699823, AI345745,AI431323, AI538764, AI538885,AI301710, AA857847, AI244343,AI287233, AI241678, AW132034,AI277325, AA977968, AA808175,AA575874, AI559752, AA001397,AL037602, AL045413, AI499986,AI923989, AL118781, AL037582,AI310575, AI863665, AW188525,AI310582, AW020419, AW161579,AI879377, AI285439, AI472487,AW022093, AW020095, AA648480,AI340533, AW170787, AA669025,AI685005, AI343325, AL041587,AI872104, AI419826, AI345010,AI955441, AI581033, AA928539,AA127565, AI567971, AI953562, T69241,AI784214, AI553645, N33175, AI470717,AI499570, AI872423, AI370623,AL037454, AI538692, N99092, AL121014,AW191844, R32821, AI621341, AI633125,AA848053, AW022602, AI287476,AI718513, AI538564, AI350489,AA502794, AW021464, AI796113,AI915291, AA580663, AW152182,AL118752, W45039, AL047344,AW162189, AI355779, AI312210,AI582932, AI688848, AI620864,AI634736, AI524179, AI583578,AI889189, AI521560, W46378,AW089844, N75779, AI539800,AW082607, AI800473, AW104141,AI309306, AW029457, AI927233,AA761557, AA939199, AI805603,AI858827, AA470523, AI479292,AL043981, AI866469, AA830406,AW263569, AI801325, AI500523,AI886355, AR000496, U39656, X65873,AF126372, AF069506, A65965, A65943,AF201468, AF141289, A49139, AJ005690,AL136884, X06146, A17115, A18079,AR013797, Y16645, AF126247, A77033,A77035, AF177401, AL137479,AR038854, AL096744, AL110269,X66871, AL122106, AL133113,AL137627, AF087943, I32738, AR068466,A58524, A58523, AF113690, S77771,AL137533, AL050277, U02475,AL133557, Y07905, I48978, AL080159,E12580, A08913, M85165, AF039138,AF039137, A08907, E05822, X70685,AJ238278, A08912, AL117457, AF125948,A08910, A08911, A08909, AL110158,AL049276, A08908, AF145233, U70981,AL110196, E12579, AL137459, X01775,AF151109, S76508, D16301, A48221,X76228, AB028451, AF031147, AF061981,AL050155, AL080148, E06743, Z97214,A48220, I89947, AL122110, X56039,AR068753, X59812, AF090903, AF140224,D83032, A15345, A23327, AF090934,AF067728, AL122100, AL137275,AL137488, AL080129, U91329,AL133665, A18777, I89931, A08456,AF031903, AF017437, AL137478, A76335,AL137271, U58996, I52013, AF079763,AL117460, AL137529, L04504, AL080074,U88966, I08319, I89934, I89944, L24896,I49625, AL049465, AL122104, I33392,AL137258, U55017, X67688, M27260,AL050149, U62807, I18358, I34395,AF047443, U42766, E08516, AL049447,X97332, AF091084, AF097996, AL117438,E02349, AL133081, AF200464, AF183393,A91160, I66342, S82852, AF004162,AL117435, A91162, M79462, A65341,AL137530, AL050172, L13297, AF017152,M96857, AL133075, AL023657,AL080126, AF061573, AF106862,AB026128, AL110280, I68732, AF035161,AL133560, AF026816, I79595, AF002985,Y13653, S63521, A65340, AL050024,AL122050, AL133640, X72624, I36502,AF061795, AF151685, AJ010277,AL137294, AF139373, AL137267, A18788,X57961, AF047716, X72387, AL080118,E02152, S61953, AL117648, AF113694,S78453, AL049300, A86558, AL137298,X83544, AF107847, Z82022, M85164,X84990, AL137554, AB007812,AF090886, AL050116, AL133016, U37359,AL137256, AL117440, AL137722,AF112208, AF081571, AL122093,AL110222, AF182215, AF061943,AF000167, AL080060, AL049339, S36676,AL049430, AF131821, I46765, AF111849,AL117649, U83172, X99717, AF158248,AL122118, AF180525, AF030513, A12297,AL137254, AL137521, A23630, AF113013,AL137537, AF111112, AR020905, andA57389.HLQDC5517110823681-69115-705HFPHI6217211958251-314615-3160AA988953, N63548, AA663569, H38453,H15532, H41422, AA976078, N94873,H46380, N51089, R88023, H41541,H48172, H38492, R18083, AI423964,AI003013, H11023, AI421373, R90757,R88479, H85702, R40487, AI208504,R42977, R87980, AA338374, AW207073,H15589, AA319224, AA325807, H84794,H10745, AA338373, W28283, R13057,AA325143, N54215, AB023202, D13613,AC004551, D29965, U12571, andAC004465.HE8QH0917311522381-143915-1453AW451023, F08666, Z41650, AA594745,D45858, D28512, and AB000893.HFAAX2917411287911-84215-856AL119825, AW367632, AA333024,AA164770, N53725, AA984472, N44616,AF000423, AB026808, and D38522.HHFOC7917511822761-209115-2105W37105, W72587, AI417917, N40695,N40709, AI750977, AA399093, AA528204,AI160861, AI167229, W56631, AA845109,AA450162, AA740816, AA708621,AA034924, AI610231, AI926222,AA600107, AA034932, AI569931,W79446, AA450099, AA405199, H99848,AI148368, AI755034, H13127, H21360,H45102, AI144154, AI022674, R48598,N24731, AI750978, H40009, N27945,R48134, H44414, M79188, R73380,H45412, AA375133, W56545, AI868381,H21461, AW015358, AA995764, H14443,H28019, R48457, H26214, W15232,AA405198, AA852302, AI865401, R11082,N27934, AA045985, W76150, R48700,H27127, AI082451, N28528, R48133,AA046015, AA358230, C17362, H27122,AA404366, AA699711, W37806, T30018,AA398203, R32099, AI870723, AA483606,AA570740, AA568204, AA502532,AW057716, AA491767, AA584756,AW089016, F11718, AA572813,AI755214, AI754567, AI815665,AA089756, AW163795, AI915081,AI754105, AW022934, AA447247,AI249688, AA832145, AI280266,AA604904, AI270476, AI080732,AA630854, AI432298, H27788, AA772906,AI380617, AA846046, AI038304,AI679221, AI141130, AW131356,AL079734, F16848, AL079894, AI431513,F35097, AA491955, AW188742,AW192599, AI569100, AW407632,AW194077, AI859438, AW157005,AI538491, AI623563, AL121657,AC007011, AC004655, AL049780,AC005015, AC004912, Z97055,AL009181, AL034429, Z98949, AC006538,AL023575, AL034420, AC007690,AC005081, AC002425, AC006449,AC004876, AC004797, AC005409,AC004491, AC004000, AC005231,AL049743, Z99755, U52112, AB023054,AC004929, AL035088, AL022727,AC006040, AC005578, Z93930,AL121754, AC004812, AP000014,AC005399, AL049636, AC007193,AC006530, AC008072, AC004084,AC005197, AL034549, AC005747,AC006441, AC004408, AL020997,AC003684, AF165926, AC005007,AC006211, AC005701, AL031273,AC004531, AC004216, L78810,AC005695, Z73417, Z82195, AC007637,AL049759, AL034417, AC004263,AL133445, AC005971, AC000353,AF200465, U91321, AL035420,AC005102, AC002300, U91318,AL096775, AL033524, AC005514, Z83822,AC002477, AL133216, AL032821,AF207550, Z82203, AL022333, AC005722,AL009172, AF109907, AC005940,AC005071, AC008044, AC003663,AC005920, AC007151, AC005752,AC006115, AL133448, AF088219,AC002070, AC005666, AP000511,AC004895, AC004675, AL050307,AL122020, AC003042, AC005815,AC006487, AJ003147, AC006965, Z77249,AL008718, AL022330, AC004770,AC007388, AC004967, AL035457,AC005280, AF111168, AC004914,AC006241, AP000501, AC003049,AC007792, AC004638, AC007263,AC005523, AC006946, AC007461,AC003009, AB003151, AC000115,AL034548, Z99716, AL078477, AC002351,AC002301, AL109627, AP000098,AC012085, U95739, AC005803,AL031291, AC003070, AC002492,AC004962, AC002378, AC005366,AC008372, AC005808, AC004051,U91323, AP000517, AP000697, AC004032,AC007225, AC007686, AC005486,AL031778, AC005620, AC004938,AC005921, AL132712, AC005829,AC005740, AC004659, AC005837,AC003010, AC005291, AL049539,AC004231, AC007298, AC006536,AC007425, AL022318, AC002470,AC003104, AC004910, AL109865,AL049761, AC005207, AL031681, Z98200,AL121748, AC005878, AC006120,U82668, AC004835, AC007055,AC004913, AL008725, AC005476,AC007344, AC004020, AL132777,AL021977, AC005017, AC002504,AC002310, AC005088, AC004844,Z85996, AL031680, AC005943, AL133371,AC005288, Z98051, AC002347,AC006974, AC004745, AC006065,Z98941, AL023877, AP000103, AF064858,AF015416, Z68869, AL022315, AC007688,AC004383, AC007376, AL109628,AL031602, AL049814, AF111167,AF001549, AP000474, AC002375,AL022323, AP000961, U95090,AC012384, L44140, AF196972, andAL109984.HOGEQ4317612234851-373215-3746AW001408, AW025576, AI167306,AW183595, AW206547, AA421304,AI884557, N53420, AI961482, AI366803,AI471722, AI277353, AI208800,AI285232, AI923048, AI002657,AA905774, AW444453, AW072850,AI002663, AA917870, AI420232, Z40060,AA995040, Z44009, AA743874, T91698,T91710, AA421383, T77085, AI536628,AA768502, T92932, F07259, AA465719,F13268, AW197122, AI222859, T92460,T93049, R13318, AW385033, T92477,AI078087, T89796, AA780031, AI680633,T89430, AI572783, F03531, AA747975,AA465126, AA361777, R57124, AI417757,AA808475, AI805839, AA324494, N94713,AW384784, AA838146, AA971578,AI422164, AA835936, AB033082,AF001630, AF038189, AF132479,AF132480, and U61167.HCECQ231779383981-79615-810AI480182, AI500178, AI873131,AA322958, AA322718, AI936088, R85125,R90888, H05353, AA338672, H51247,R35934, AW139057, H05303, R49451,R42549, H29245, AI654790, AI638508,R51648, AW149807, R44423, AI458144,AI419465, F02105, AI499775, Z40525,F09051, R85080, H51217, AI569283,D29763, D64009, and D64010.HTGAU7917811786211-108115-1095AA579641, AI149891, AI028588,AA659710, AI889233, AA461487,AA460132, AA682750, AA113277,AI753378, AA985248, AI421205,AI914588, AI493749, AA633392,AA936668, AA722409, AI424537,AA233726, AW268600, AA516077,AA483490, AW082930, AA074311,AI752615, AI989665, AA046575, T87841,AI087102, and AL031055.HE9FI3317911564321-155015-1564AA010320, AA010384, and R01100.HNHCP791805657811-28815-302HTLIY5218111948061-133815-1352AI827749, AI580407, AI819667,AI025487, AI223109, AI150036,AI024234, AW087713, T18864, AI479322,AA883975, AW341589, AA860213,AI831802, AA913074, AA608857,AI050685, AA860223, AA948538, andAI075930.HRAED741829425271-69115-705AC005940, L42810, S83194, AF117384,and AB023658.HFKKN771839437571-71915-733HTEMU6618412053811-107715-1091AL039924, AL045794, AW013814,AL036630, T02921, AL044412, AL044364,T24119, T24112, AW450335, AL039476,D51250, AL039521, AL045341,AL039386, D80043, AL040992,AL039109, AL038531, AL037726,AL039629, AL039659, AL039625,AL039648, AL038837, AL039074,AL039566, AL039678, AL039108,AL039538, AL039564, AL039509,D80253, AL039156, H00069, AL039128,AL044407, AL036973, AL042909,AL045337, AL037051, AL045353,AL039423, D59787, AL039410,AL039150, AL038025, AL044530,AL039459, AL038821, D80219,AL036725, AI535983, AL043422,AL043445, D59275, AL043586,AL043423, D80240, AL043441,AL036650, T23947, D80210, D51423,AL036196, D80134, AL037639,AW451070, D59619, D80391, D80227,AL037615, D80193, AL036767, AI535783,D80196, AL039085, AL036117,AL042334, AW452756, R47228,AL037526, D80366, D59927, AL036238,AL036679, AL039504, AL037601,AL039842, AL036964, AL036733,AL037027, AL036158, AL036924, C14014,AL037054, D50995, AL036765, D80045,C75259, T23658, AL044413, T23659,AL037177, AL036998, AL038851,AW293068, AL036227, AL036418,AL037047, D59889, AL037643, C15076,D81026, AL036133, AL036163, C14227,C14389, D50979, AL037082, AL036167,AL037124, AL037679, AL036207,AL036132, AL037049, AL036191,AL037021, D80022, AL037600,AL036190, T11417, D59467, AL036139,D81030, D80038, T23656, AL036152,D58283, D80195, C14429, F13647,D80188, AL036900, D52291, T48598,T03269, D51799, D80168, D80522,AI557751, AL048425, D80378,AW451416, AL037178, AL043613,C14298, AL039555, D59502, AA514190,AL043585, D80164, D80166, D59859,D80212, AL037085, Z25782, Z21582,D80269, C14331, D59695, D80024,D57483, AA285331, AL036953, D59610,AL036808, AW450376, D80064, D58253,D80241, AL036174, AL037569,AL044178, D80949, AL036150, Z99396,D80268, D51060, AA305409, AW178893,D81111, D51079, H00072, D59627,AL036268, AL038043, N47620, D80251,D51022, AW177440, C14407, AW179328,AI910186, AW178775, AL044447,AW378532, AA305578, AW352158,D80248, AW377671, AI905856,AW369651, D51097, AW178762,AA514188, AL043840, AW177501,AW177511, D50981, AL037002,AW360834, AL037077, AA514186,D80133, AL039417, AW360811, C05695,AW352117, Z25783, AW135155,AW176467, AW375405, AW378540,AA809122, AW366296, AW360844,AW360817, AW375406, D80132, A85396,A25909, A85477, A44171, A86792,X68127, AR062871, A84772, A84776,A84773, A84775, A84774, AR037157,AR017907, AR062872, AR062873,AR067731, AR067732, A58522, A91750,A20702, A43189, A43188, A20700,AR036905, A10361, AR038762, A98767,A93963, A93964, A95052, A38214,I03343, AR043602, AR043603, AR043601,A95117, A95051, A18053, AR022240,A18050, AR054109, A23334, A75888,I70384, I56772, I95540, AR018924,A63067, A51047, A63064, AR031374,AR018923, A60111, A48774, A23633,A63072, AR007512, A49700, A48775,A23998, AR068507, AR068506, A81878,AR015960, I60241, I60242, AR000007,AR015961, A58521, I63120, A24783,A24782, AR020969, E12615, AR035193,A92133, E14304, A27396, AR027100,I28266, A49045, E16678, A82653, E16636,AR031375, A93016, A58524, A58523,I25027, I26929, E13740, I44515, I26928,I26930, I26927, A58525, I06859,AR000006, I49890, I44516, A58526,A91753, AJ244003, AR025207, I18371,E06034, AF156296, A13038, A29289,AR029417, AF156294, AR067733,AR029418, AR067734, AR028669,AR028668, AR028667, AR028670,AR017908, A68112, A68104, A98467,I62368, AR031488, I13521, I52048,I44531, A84746, A17115, AR028672,A18079, AR038066, I50882, I15353,A64081, I66495, I66494, I66498, I66497,I66496, I66486, I66487, A71440, A71435,I13349, A07699, E08322, I74623, A60109,A67220, AJ244004, U87250, I00081,A70040, AR028564, A83643, A98420,A98423, A98432, A98436, A98417,A98427, A83151, I01968, A13388, E00974,A02228, E00954, E00952, E00953, E00955,I08049, I43960, AR021440, E03165,E02221, E13364, A10360, E02679, E02104,E02098, A92666, E02001, E01718, E02003,E02102, E03550, A28163, E02100, E01997,A58998, E02291, E02292, E02293, E01999,E02396, E02327, E01563, E02431, E01693,E01696, A92668, AR005163, AR005154,AR005157, AR005160, I09250, AR005153,A92667, E01024, AR050583, AR050584,E02430, E01148, E12527, E01503,AR005165, E12523, E02432, A49701,A62009, E01216, I77211, A95096, I70974,I31847, I31848, AR060673, AR060676,A95106, A95105, A80476, A91965,E01324, I08638, A80474, A94048, A94061,A80477, AR035224, A80475, A94046,A94054, I63560, AR031529, A49428,I63561, I63563, and E16036.HWAGU6218512067971-219215-2206AI913535, AI762854, AI758705,AI677912, AI740876, AI825702,AA412665, AI800271, AI525869,AA883055, AI823434, R77865,AW204604, AA491093, AA134753,AA598723, AA845774, R73497,AA134752, AA993841, R73498, D62016,AI332677, AI535824, AI598069,AI535821, N68128, R77771, AA761684,W28537, AW370473, AW370408,AI758562, AI754802, AA293681,AA075272, AI687944, AI916337,AI633125, AW198090, AI620056,AI608882, AI479292, AI890223,AI859932, AW151893, AW152182,AW190194, AI802542, AI625079,AI677796, AI554821, AI701097,AW409867, AA830709, AI687362,AI584130, AW073677, AW083572,AI819326, AA811202, AI538564,AI571439, AW169604, AW020619,AI653979, AI702073, AA743430,AI673140, AI433157, AW157096,AA937566, AI049733, AI636588,AW195169, AI241744, AI635032,AW166870, AI445025, AI567128,AI973152, AW002362, AI637584,AI469532, AI635016, AI247293,AI619662, AW083374, AI564719,AW129722, AI952471, AI811644,AI620003, AI357940, AI376425,AW104724, AC007785, AF100752,X78627, AF124435, AL133049, AF222801,X59812, A15345, X60769, AR050959,S36676, D44497, AL137550, Z82022,AL137476, A60092, A60094, AF031572,AC006288, X63162, AF084644, AF084645,A27171, A52184, L35261, AR038854,AF115410, AL137560, AF124396,AL136884, AF111849, U49434, U95114,X68497, X52220, X98066, U97675,AL023657, A83556, AL133607, AF082324,and X68249.HFPFB3918611980361-209015-2104AI631883, AI767614, AA018867,AA019175, R56792, AI497937, N95592,R44001, AW271411, R56793, Z38315, andZ42023.HPMFI3818711659931-97515-989AI765950, N54154, H12876, R93383,R56658, R16970, AW205252, R25871,AA886432, R93384, R16914, and N58678.HBXDJ071889468301-147015-1484H11405, R55569, N27906, H20863,N25140, and U27708.HOFMS4318911524171-106515-1079HOVCO1419010910871-59215-606AI701529, AA994711, AI192036, andAC007198.HTEPE3519111052721-82515-839AI935040, AA861064, AA933697,AA707583, AA872105, AA398866,AA609626, AA620685, AA435866,AA812556, AA781805, AA993718,AF012362, AI217888, and AA824315.HE8UA5219212294901-340815-3422AI742521, AW274710, AA723595,AW365665, AI141346, AW043776,AI469940, AI378321, AI079805,AI168469, AI142001, AA772045,AI337483, AA478634, AI051463,AA912545, AW149092, AI249877,AI583065, AI873923, AL040011,AI700159, AI805638, AA427373,AI560683, AI287793, N42321, AI633125,AI499325, AW170635, AW151714,AI587114, AI872804, AW087455,AI631216, AI669459, N29277, AI921281,AI916419, AI538850, AI476478,AI242248, AW193231, AW162194,AI677646, AA580663, AW192228,AI677796, AI434038, AI494201,AL039753, AI583308, AW079859,AI357599, AI473536, AW152182,AW021373, AI884318, AI655841,AI241744, N71180, AI468959, AI538564,AI564723, AI685798, AI340519,AW081311, AW169653, AI863382,AI619691, AI432030, AL134830,AI620643, AI915291, N36182, AW263804,AI590423, AW191844, AA464646,AI636457, AI871923, AI631264,AI345688, AI933992, AL120526,AI690813, AL047422, AI868204,AI282307, AW243886, AW149311,AI718707, AI554343, AI446373,AW265004, AI344817, AI635216,AA019328, AW084117, AI687295,AI345224, AI679098, AW081322,AW104141, AI344935, T99953,AA502794, N22406, AI538716, AI539771,AI673710, AI624963, AI311892,AI579901, AI307736, AI472536,AW085673, AI611743, AI491710,AI249962, AW189270, AI281867,AW059828, N75771, AI635478,AA494167, AI684305, AI798456,AI554186, AI499131, AW105383,AI468930, AW089572, AW084097,AI648567, AI610690, AI890954,AI627893, AI932794, AL039858,AI653979, AW166182, AI890907,AW130134, AI951868, AW129916,AI670009, AI520862, AI310575,AW163554, AI570989, AI445663,AA812110, AL042628, AI499947,AI702073, AI569579, AI565031,AW102864, AA468418, AA835801,AI591387, AI340533, AI439527,AI890183, AI583085, AI924911,AI280732, AA641818, AI249946,AW020397, AI537837, AI566003,AI690748, AI698391, AI554821,AW058233, AI633000, AI623682,AI499393, AI801112, AW020693,AL037602, H41759, AI433157, AI620859,AW079119, AW130681, AI349957,AI922707, N29481, AI889189, AL040243,AW172723, AI889168, AI440263,AI700305, AI340982, AW169527,AI345005, AI357316, AI348870, H89138,AI340627, AI469505, AI274785,AW163834, AL047344, AW023338,AL119748, AL046618, AL079963,AW129929, AW104724, AI470651,AI582912, AI453328, AA833760,AL037582, AI679211, AW237857,AI445829, AI539707, AI637584,AI828705, AI340603, AI889147,AI659334, AW105431, AI288285,AI564311, AI678496, AI862142,AI567612, AW192701, AW083804,AI439478, AI096534, AW192042,AW152459, AI621362, AA635382,AI866465, AI689420, AI918554,AI273179, AI815232, AW302924,AI251221, AW198090, A21103,AL122045, AL122111, AL137529,AF183393, AL110225, AF061981,AL050149, A18777, I48978, I89947,A08913, A08910, AR038854, A08909,AL117460, AL049452, A08912, U49434,A08908, I26207, S76508, I89931,AL080126, I89934, I89944, A15345,I49625, A08916, A58524, A58523,AF090900, X63410, AF113677, L13297,S77771, I68732, A18788, AF026816,AF118094, X70685, AF090903, AJ003118,D83032, S61953, AL133558, X80340,AL137533, AL137488, E01573, E02319,A08907, AL049464, L19437, AL049283,Y10936, AL137523, A90832, AL137463,AL110171, AL137526, AL050277,A83556, I46765, AF079763, L31396,AL080148, L31397, X57961, E12747,AR068751, E05822, AL133640, AL049314,AJ242859, AF159615, M96857, AF000301,I09499, U80742, AF090901, AB029065,AF113694, Y10080, AL122049, AF118064,AF118070, AF107847, AL133098, X52128,AL133557, AL133016, AL133014,Y11587, X83544, AL122106, Z82022,AL137648, AF153205, Z37987, AL050116,AF146568, A52563, A12297, AL117435,AF085809, AJ006417, X98834, AL133568,AL122110, AF090943, Y11254, AL080159,AL137560, M86826, AF017790,AL080127, AL133113, AF125948, I03321,U68387, AF090896, AL133606, AL080163,AF032666, AF057300, AF057299, A23630,AL137550, AF091084, AF100781,AB016226, AF100931, Y16645, U00763,AF215669, AJ000937, X62580, I33392,AF051325, AL137538, X84990, AL122098,AB007812, AL117457, AL049466,AL137558, AL133565, AL137479,AL049347, I00734, AL110159, AL049465,AL137276, Y10655, S75997, AL049996,AF113019, AL049938, AF067728, X79812,A65341, AL122100, AL050108,AL137656, AF139986, AF137367,AL110222, AL133112, U72620, S68736,AL137292, I41145, AL080086, AL133104,AF003737, AF017437, X87582, AL122050,AL117583, AL117416, U35146,AL117585, AL080158, X92070, S69510,E08631, U00686, AF113676, AF040751,AL122118, U35846, AL137521, AF106862,AR054987, AL133665, Z97214,AR013797, U68233, AF113690, I92592,AL137283, AL049300, AL050024,AL110196, AL049430, A76335, AF118090,AF106827, A49139, I17767, A93350,Y09972, A07647, A45787, AL136842,AL050146, AF177401, AL117394,AL122093, AF106657, AL050092,AL133031, AF079765, X63574, AL137480,AF061573, AJ012755, X83508, AL110218,A08911, I48979, AL080124, AR020905,AF126247, AF162270, U67958, A77033,A77035, AF113699, AF158248, AF111849,AF026124, I80064, E01314, AL110221,AL137712, AL137273, AL050138, andAL137641.HOUBE5019310907761-117515-1189AL120487, AA578623, N85038,AB033086, and AB023168.HAJAV2819411652291-158715-1601AA913364, AW027373, AW305275,AI799031, AA588138, AA775450,AW117480, AW190848, AA411334,AA866178, W61038, AA411335,AA775769, AA769134, N30274, AI493881,AA075643, AA614747, AA627544,AI147666, AI025647, N30309, AA075644,AA812101, N94919, AW339918,AI140386, AI138196, AA868924,AA284960, AI554835, AI343592,AI128292, AI143201, AI017993,AI492556, AI347094, N42630, W72001,N20511, AI679099, AI933338, AA040263,AI268560, W69402, AI354734, T32745,N29505, AA872581, AI017992, N41996,AA863176, AA719956, F28362,AA448442, AI679673, AI142803,AA732322, AA993672, AA444022,AI624135, AA846235, AI004213, W44778,W44314, AA305351, AI285067, AI304774,AA448307, W65308, AW274816, W37097,AI475118, N25416, D79265, AI872626,AI186604, AA682603, AA285117,AI149185, AI339799, AI344717,AA996037, AA444000, AI128110,AA557809, H13966, AA922743,AA515842, F37123, AA732278, W65338,N56641, AI239957, AI580506, AA989130,R21057, W45686, R96887, AA721150,AA034909, F09166, H12989, N26843,N57447, W69363, W19542, AW183468,AI685763, R96888, C04314, H87003,AI719584, R24370, T35889, AA081676,T34115, N40568, AW023063, N63592,AA010732, T35834, AW089136, Z42311,F11505, W76570, T35841, AA913531,T30076, AI366940, AA505347, T30517,H05630, H27007, Z42248, F09167,H87002, W56871, T36163, AA040400,W19619, AA721261, AA649338, F34780,T35871, H13965, R46237, T35833,AA034976, AA863208, AA913094,AI274646, N63619, AA890331, AA082031,F05477, AI564372, AA603299, AA322616,AA603386, R27044, Z38484, R27043,R27029, F01840, AI859366, R27028,AA748381, F01739, AA355325, H53598,AI859362, N46608, AA339209, andAF190797.HAQBZ8919510835541-140315-1417AI436552, AW071796, AI299053,AI334145, AI613263, AI921361,AI161282, AI018067, AI984679,AI934889, W52097, AI281829, AA236375,AA121294, AI342850, W16450,AA729045, AA608803, AW135827,N78654, AW193167, AI689644, N46334,AI159772, AA456075, AA130122, N63941,N58535, AL042537, AA367722,AA969946, AI432541, AA829498,AA781924, AW304842, AI299054,T69736, AW194058, AW170746,AI382899, AA862441, AA257021,AA345125, AI049756, AI983846,AI129698, T70395, AA130159, AA455578,AA833560, AW351523, AA451639,AI205015, and AW512516.HELHF071969490671-32715-341AC073669, AC074220, AC074220, andAC023605.HE9QQ2219711277261-74815-762N90644, T72234, R83190, AA010242,N90655, N90629, N94219, N65959,R83191, D50995, D80164, C15076,C14014, D59502, D80247, D59275,D80195, C14389, D81026, AA305409,D59467, D80227, D50979, D80269,D51799, D80022, D80166, D58283,D80193, D59619, D59859, D80210,D80391, D80240, D80045, C14331,D59787, AA514188, D51423, D80253,D80043, AA305578, D81030, D80188,D80439, D51060, D80038, D80212,D80196, D80248, D80219, D80366,D51022, AA514186, D80522, D59927,D80302, D80024, D51103, D57483,D59610, D80378, D80133, D59889,AW360811, D80268, AW177440, D80251,D80241, C14429, AW178893, AW377671,AW375405, D80157, T03269, AW178906,D51759, AW366296, AW179328,AW360844, AW360817, C75259,AW375406, AW378534, AW179332,AW377672, AW179023, AW178905,D59373, C05695, AW378528, AW178762,C06015, AW378532, AW176467,AW177501, AW177511, T48593, D59695,AW352171, AW377676, D59627,AW352170, AW177731, D80132,AW178907, AW179019, AW179024,D51250, AW178983, D80134, AW360841,D59653, AW177505, H67854, AW352120,AW179020, AW178775, D58253,AW178909, AW367967, AW177456,AW369651, AW179329, AW178980,AW178914, AW178774, AW177733,AW178908, AW178754, AW179018,AW352158, AW179009, D45260,AW352117, AI525923, D81111,AA809122, AW360834, C03092, D52291,AW179004, AW367950, AW179012,D51079, AW378543, AW378525,AW378540, AW352163, D59503,AW177728, D59551, H67866, T03116,D80949, D80258, D80168, T11417,AW352174, AW178781, AW178911,AI535686, AW177722, F13647, D59317,AI910186, AW178986, T74930, AI905856,D58246, C14298, AI525917, C14077,C14227, D80064, D80014, C14407,D51221, C14973, C14344, AW378533,AI535850, AW177734, AI525925,AI525920, D59474, D60010, AW177508,AA514184, D51213, D60214, AI525227,AI557774, D58101, AW177723,AW177497, C14957, C14046, Z21582,AI525242, AI525235, T03048, D45273,AI525237, AI525215, AI525912,AA285331, AW378542, AI557751,AW378539, C16955, D51053, C05763,Z33452, T02974, D31458, AI525222,C13958, AW360855, D51097, Z30160,T02868, H67858, D38100, AB002584,AR018138, AB028859, AJ132110, A84916,A82595, A62300, A62298, AR008278,AF058696, AB002449, AR060385,X67155, Y17188, A94995, D26022,Y12724, A25909, I50126, I50132, I50128,I50133, A67220, D89785, A78862,D34614, AR008443, AR066488,AR016514, AR060138, A45456, A26615,AR052274, D88547, Y09669, A43192,A43190, AR038669, I14842, AR066487,X82626, A30438, I82448, AR016808,AR054175, D50010, Y17187, AR025207,A63261, AR008277, AR008281,AR066490, AR008408, AR062872,A70867, I18367, AR016691, AR016690,U46128, I79511, D13509, A64136,A68321, AR060133, X68127, AB012117,AF123263, AR032065, U79457,AR060382, Z82022, A63887, andAR008382.HSDSB0619811282801-267415-2688AI766068, AW009631, AI817090,AI765056, AA127727, AW069132,AI372853, AA877550, AI432584,AI129942, AA610281, AA102362,AA625117, AA948383, AA430522,AA447454, AW194550, AI635683,AA532394, AA446651, AA724535,AI220147, AW263019, AA430607,AA019158, AI752984, AI198643,AW389353, AA045561, AA516463,AW197881, AA186967, R67323, H86071,H67029, AA588072, C01179, AI457685,H22246, H12434, AW378928, H12433,AW050669, H68111, AA768085,AA054561, R33871, R66487, AA054621,AA478635, N55248, AA359925,AA385529, R33870, W68645, AA961423,AW002948, AI802284, AA377365,D31590, AI742590, AW275740, AI819328,AL133047, D89677, AF003234, andAW517631.HACAD351999491991-146015-1474AI817458, AI810494, AA442536,AI271432, AA228126, AA227978,AA687616, AW316554, AW351633,D62849, AA346825, AW262553,AC007363, and AC007363.HEQAP172009493581-80715-821AI131555, AI769466, AA215577,AW190975, AA258335, AA258499,AL044652, S63848, Y17793, and A49045.HMTBB1720111285891-46215-476AA582539, AI963340, AI097093,AA286856, AI761614, AI149781,AI460219, AI032670, AI636161,AI819154, AI089302, H12042, AI811219,H05308, T95010, AA836993, AW271462,R37000, AI001803, AA904906, AA743196,AI015200, AA453607, F05000, AA578803,AI241466, AI033193, AA330970, F03322,F01968, AA037601, T75492, N47542,AW183219, AI288171, AA054759,F01965, AA651907, AL122084, andAL049611.HKGDE5820211291371-132515-1339AW271462, AA582539, AI963340,AI097093, AI460219, AA286856,AI761614, AI149781, AA448686,AI032670, AI819154, AI089302,AI636161, H12042, AI811219, AA287162,AA836993, T95010, AI001803, AA904906,AA743196, AI015200, R37000, H05308,AA453607, F05000, AA578803, AI241466,AI033193, F03322, AA037601, AA330970,F01968, R13858, T75535, H12041, T75492,AA053290, Z46111, F07044, T94956,H05358, N47542, AW183219, AA454139,F05720, AI288171, AA651907, AA054759,F01965, F05717, AL122084, andAL049611.HCHMW4020311443231-94115-955AI732539, AI791495, AI791325,AA709067, AW082062, AI791964,AI732667, AA505923, AW057561,AI909857, AI909862, AA601601,AI909853, AW082594, AI812032,AI310332, AI610362, AI627880,AW403717, AA427700, AI687065,AW151136, AI783504, AW071417,AI446373, AI280670, AI886022,AI345253, AI345677, AW189415,AI800453, AI284131, AI285826,AW051088, AW022682, AL134999,AI280732, AI680162, AW168485,AI610645, AL079963, AI680498,AI805688, AI886181, AL040241,AI312428, AW020095, AI537677,AI498579, AI400725, AI866741,AI251221, AI932915, AI589947,AI343059, AI349933, AL119863,AL036548, AI345608, AI690410,AI571909, AI478123, AI335426,AI348777, AL041772, AW274192,AI590686, AI869377, AI364788,AL036638, AI310606, AW301344,AI568765, AI922901, AI345471,AI366992, AI863321, AI336575,AI950877, AI334450, AL120853,AI932794, AL110306, AI929108,AA908294, AA225339, AI608936,AI345746, AI251205, AW081255,AL045266, AW088134, AI824576,AI280661, AI659795, AI537617,AI520809, AI812015, AI569309,AI828568, AW087915, AI434741,AI648502, AI919107, AI352497,AI678357, AI888944, AI308035,AI886123, AW198075, AI174394,AI933589, AI686552, AW168709,AI587606, AI783861, AI468872,AW083778, AW163823, AW084219,AW103886, AI952920, AI955866,AI628217, AI933785, AI697324, N80094,AI633419, AI554218, AW151785,AI963216, AI445165, AI590120,AI431909, AI866002, AI829327,AW268302, AI433976, AW168373,AW149227, AL119791, AI282326,AI828731, AW302973, AI499463,AW079159, AI251830, AI619716,AW268060, AI288285, AW072719,AI539028, AW023590, AA493923,AI702019, AI744256, AI432736,AW148716, AW168031, AI648684,AI612759, AW152469, AI249962,AW089179, AI628316, AW151729,AI867042, AI696819, AI566630,AI619754, AI623682, AW163834,AI699011, AW088903, AW151714,AI919345, AW129230, AI698401,AI922577, AI249877, AI471361,AW088899, AW148356, AI366549,AI636719, AI539153, AI073952,AL036214, AI539771, AI916419,AI439762, AI431424, AI866608,AI349645, AI611743, AI800433,AW268220, AI689175, AW083804,AI590423, AI886192, AI888621,AI696626, AI589993, AW059713,AI365256, AI271796, AI431408,AW167882, AI348897, AI537515,AW029611, AI801213, AI863382,AI802654, AI783997, AI620639,AI310575, AW080135, AI805769,AL036396, AI340627, AI791396, F34698,AI493559, AI285431, AI340533,AI089970, AI564749, AI308032,AI800138, AW087445, AI610557,AI348854, AW087938, AI343091,AI344785, AI889953, AI680457,AI921386, AI362580, AW190042,AL036718, AI383919, AI886124,AI250663, AI824375, AA640779,AI285732, Z82022, AF111112, E03348,E04233, AL137521, AJ242859, AL133093,I89947, I48978, A08913, X87582,AL080127, A08916, AL117435, U91329,AL080060, I89931, AL133016, A08910,I49625, A08909, I26207, AL050024,X84990, A45787, X53587, AL050138,A03736, AF113690, Y11587, AF090900,AF008439, A93016, AL137556, E05822,AL080124, AF078844, AL137538, U96683,A08912, I00734, U88966, AF162270,I42402, E00617, E00717, E00778,AL133077, I48979, AR038969, AL137526,AF090943, AR059958, AL133557,AF113691, AL137463, AL122121,AF067790, AR038854, AL110196,AL080159, AL049382, AL049314,AF183393, AJ238278, AL117585,AL133075, AF090903, Y14314, AL050149,U80742, X65873, AJ012755, AL110280,AL137476, AF090934, AF118094, I09360,L19437, AL080074, AL133640, E15569,AF153205, AF017152, AL080137,AL137527, X96540, AF081197, AL049466,AF104032, S68736, A93350, AL050277,AL137550, AR000496, AL049430,U39656, AL137523, AF026124, AL050393,X82434, AF113689, AF113699, AF132676,X70685, AL137271, AF061836, AF111851,L30117, S78214, AL110221, Y09972,AL117440, AL133014, AF146568,AL122093, A12297, X93495, AJ006417,S61953, AR011880, AF113694, AF113019,U67958, AF113676, AL137533, AF158248,AL133113, AL133565, U00763, U72620,AL122049, AF118064, A90832, A07647,AF118070, AL122050, AL133098,AL122098, AL096744, AL050146,AL133568, AF090896, U42766, AF106862,I03321, I33392, E02349, AL050092,AL133560, AF097996, AF087943,AF061573, AL117460, E07108, AF177401,AL137273, AF026816, AL133606,AB019565, AL122110, AF091084,AL133067, AF067728, A65341, A77033,A77035, AL117583, AL049464,AL117457, AF125948, AF185576, U78525,AF090901, U35846, X72889, AF119337,AL133104, AF017437, AF113677, Y16645,AL049452, AF113013, AF125949,AF079765, AL117432, AF003737,AL049300, AL137557, Y11254, AJ000937,AL133080, AL137560, Z72491, M30514,U58996, AL137648, AL137459,AL050116, L31396, U68387, AL050108,AL110225, AL133072, L31397, AL117394,X63574, AF057300, AF057299, X98834,AF061943, AF081195, AL122123, E08263,E08264, E07361, A58524, A58523,AL137478, AL133645, AF061795,AF151685, A18777, AL137480, Z37987,E08631, E02221, AL049283, AF079763,AF111849, AL023657, AF030513, Y07905,E06743, AL050172, AL117649, andAL137294.HE8QZ3420411434111-187315-1887AI346431, AA600828, AI654068,AI627476, AI692275, AI417541,AI033899, AA127745, AI634873,AI653646, AI564494, AI635599,AI434505, AA127671, AI818370,AI912908, AI311461, AI914324,AI371325, AA622400, AA225468,AA811711, AA593299, AA633708,AA768315, AW236744, W28575,AA210809, AW071534, AA565871,AI371817, AA287455, AA210690,AA225526, W00592, AA085933, N68915,AA908320, AA281938, AA112126,R35313, AA225852, AI277214, AA465202,AA580581, R86156, W19885, AA455978,AA286753, AW020244, AA636108,AW405566, AA452896, AI863445,AI055868, AA594111, AA772109, T06553,AW195517, AA385668, AI673762,AA384297, AA730967, Z41832,AW338426, R61806, H25863, AA370592,R43790, AA361533, AA449099, H50786,H89555, AI312685, R25032, AW118452,H89367, T35293, Z46079, R20776,AA334931, AA490204, R51228, AI874163,AI446400, AI475237, AA296796,AA249741, C01856, AI630454, AA449535,AW073930, AA770110, AW367213,AI633501, and AI567975.HWAFG0420512276271-138515-1399AW239548, R23870, F07734, D54154,AA297059, C14980, AI418298, AA088799,C15480, R46685, AA325059, AA974700,H42947, AI302185, AI652375, AI394630,AA364780, AI936871, AA382960,AI559159, AI025217, AI983297,AI025310, AW002416, AW206793,AA593826, D57560, AW140018,AW192088, AI017077, AW401787, andAW373232.HTEKT3320610758361-210515-2119AL044351, AI817283, AI024728,AI744491, AW072402, AW378258,AW072368, AL044352, AW192336,AI337979, AI627480, AI696944,AW369691, AI860188, AW378250,AW378255, AI627747, AL042027,AI926480, AW378166, AW068812,AL047150, AW029453, AA464338,AI801683, AI890113, AI745169,AI623625, AW069248, AA573520,AW378242, AW369645, AA722200,AW152579, AI679464, AI754467,AW069250, AI863890, AI420021,AI754556, AI017236, AA490151,AW337220, AI423346, AW385997,AW074087, AI015943, AI689287,AI587461, AW369622, AA428544,AI222770, AI143319, AI720991,AI754784, AW152549, AI143344,AI589285, AA582432, AA826662,AI587096, AI753623, W68196, AI754168,AW383083, AW383201, AA045260,AA844711, AW378254, AI753721,AA595042, AA432225, AI079642,AI754630, T63365, AA598475, AI962914,AW378175, T51018, AA634355,AI752372, AI922832, AI696875,AI675948, AI371134, AI598124,AI371125, AI143853, AI143041,AA779030, AI281593, AI216596,AI753783, AI313439, AI571587,AA931107, AA598793, W68075,AI339836, AW378191, AW351438,AA045150, AW169487, AI751039,AI051715, AI148583, AI970742,AI338164, AW007286, AA541755,H12770, AI694286, AI309918, AI358476,AI582893, AI304450, AA026628, N78364,D53342, AI041911, AW024056,AA935545, AW130479, W39177,AI354525, AI376810, N34461, AI808534,AA778065, AW192368, AW244107,AI589361, AI282168, AW028623,AA778055, AW150321, C17178,AI478760, AI244443, W86793, AI087238,W00916, AI446098, AA719311, H24846,AI209065, D57494, AA599264, W94304,AI083988, AI278860, AI935852,AI091878, AI829367, AI751862,AI278832, AW382654, AA620910,AW262984, W80605, AA834064, N94128,N62598, W72648, AW383076, AA026668,AW193117, AA084560, AI752585,AI624605, H49416, N55278, AI360559,AI520753, AW068113, AW370175,AL046974, AI815020, AI753242,AI963416, AI147036, H42782, AI571697,AI751040, AW242018, AI570991, H94334,AI953531, N57952, AI141927, AI752371,AW242264, D57929, W79127, W01207,AL048585, AW068071, AI624798,AA564187, W39458, W44950, AA358927,W02137, AW239018, AI751675,AA485695, AW262921, N67498, H24847,AI889395, W01047, AA329561,AA777026, N65969, AL048586, H64363,AA916595, W32452, AA455154,AI539001, AI702017, AA334020,AW382258, R77374, AA541671,AA505619, R83643, AW024024,AA953041, AI636202, R19771, R83636,F27591, F07112, F27593, AI751863,AW068339, AI920900, F06387, AI014855,H63926, H78881, AA953591, D51304,AI879452, AA718967, W87292, D56506,AW365001, N70195, M14219, AF138300,AR012319, AC007115, AF038127,U83141, AF125537, S76584, Y00712,AF125041, AF140270, U03394, X53929,L01131, AF138302, M98262, L75825,L01125, AF138304, L01127, L01130,L01128, L01126, L01129, A62298,A82595, AR050070, and A82593.HBXDM0720712066571-211015-2124AI672363, AI589203, AI631066,AI810806, AI654696, AA700425,AW249815, AI271343, AI127886,AI581871, AW044228, AA188637,AI769068, AI142895, AA084919,AA451876, AA482390, D20750,AA463624, AI351007, AI240526,AA074549, AW090810, AI143138,AI762690, AI752042, AI912224, R56540,AI127226, AA626231, R56145, AA190610,AA085022, AI336960, AI962785,AI690749, AA035626, AI762681,AA346622, AA847532, AW139485,AA426415, AI625813, AA443041,AI940415, AA062842, AA599682,AW374890, AI659071, AA953644, andU35245.HFPFA832089556141-72315-737C14389, C15076, D59467, D58283,D50979, D80522, D80164, D80166,D80195, D80043, D80227, D81030,D59275, D59502, D80188, D59859,D80022, C14331, D51423, D59619,D80210, D51799, D80391, D80240,D80253, D80038, D80269, D59787,D80193, D59610, D80212, D80196,D80219, D81026, D59927, D57483,D80378, AW177440, D80366, D80251,AA305409, AA305578, D59889, D50995,D80024, D80241, D51022, D80045,C14429, D51060, C75259, T03269,AW178893, AW179328, AA514188,AW378532, D80248, C14014, AW377671,D51250, AW369651, AW178762,AW178775, AW177501, D80134,AW177511, AA514186, D80133,AW176467, D58253, AW360811,AW352117, C05695, AW375405,AW352158, D80268, AI910186, D80132,AW366296, AW178906, AW360844,AW360817, AW375406, AW378534,AW179332, AW377672, AW179023,AW178905, D80302, D59627, AI905856,AW378540, AW352171, D80258, D80439,AW377676, AW352170, AW177731,AW178907, AW179019, AW179024,D59373, D80247, AW177505, AW179020,AW360841, AW178909, AW177456,AW179329, AW178980, AW177733,AW378528, AW178908, AW178754,AW179018, AW352174, Z21582,AW360834, D51103, AW179004,AW179012, C06015, AW178914,AW378525, AW367967, D80157,AW177722, D51759, AW177728,AW179009, AA285331, AW178774,AW178911, D51097, AW378543,AW352163, D58101, D80064, D58246,D80014, D59503, AW178983, AW352120,AW178781, T48593, AI535850,AW177723, T11417, D59653, AA809122,AW177508, D45260, D59317, C14975,AW378533, AW367950, F13647, D81111,H67854, C03092, C14227, H67866,AI557774, AI525923, AW177497, T02974,AI557751, AW178986, T03116, C14298,D45273, D52291, AW177734, D59474,AI525917, AI525227, D59695, D60010,C14973, AI535961, C14344, C14407,AI535686, C14957, D51221, D59551,AI525920, AA514184, AI525242, D60214,T03048, C14046, AI525912, AI525235,C16955, AI525925, AI525222, D80168,AW378542, AW378539, AI525215,AI525237, C05763, Z33452, AI525928,AW360855, T02868, D51213, D31458,H67858, AR018138, AJ132110, A84916,A62300, A62298, AR008278, AF058696,AB028859, X67155, Y17188, D26022,A25909, A67220, D89785, A78862,D34614, D88547, I82448, Y12724,X82626, AR025207, AR016808, A82595,AR060385, A94995, AB002449,AR008443, AB012117, I50126, I50132,I50128, I50133, AR066488, AR016514,AR060138, A45456, A26615, AR052274,A85396, AR066482, A44171, A85477,I19525, A86792, Y09669, A43192,A43190, AR038669, AR066490, U87250,AR066487, X93549, I14842, A30438,I18367, D88507, AR054175, D50010,Y17187, A63261, AR008277, AR008281,AR008408, AR062872, A70867,AR016691, AR016690, U46128, D13509,I79511, A64136, A68321, AR060133,X68127, AF135125, U79457, AF123263,AB023656, AR032065, AB033111,X93535, and AR008382.HKADO3620911890021-136415-1378AI886975, AJ011372, AC001644, andAC002325.HFXKG512109565961-111415-1128R29445, AA585101, AA585476, Z28355,Z30131, T11028, D57491, AI546999,D57186, AI541365, T18597, AI541374,C16300, AI525431, AI546945, R28735,R45895, AI525306, AA585098, AI525556,AI541523, AI547006, R29218, D55233,AI557262, R28892, AA283326, AI526184,D61254, AI546875, AA170832, AI540967,Z32822, R28895, R28967, R28965,AI557734, AI541535, C15406, AI547039,AI557763, D60765, C16305, R29177,AA585439, AI557787, AI526194,AI541205, D61185, AI526140, AI557740,AI546891, C16293, C15069, AA585329,AI541307, AI526073, AI557731,AA585325, AI541514, AI541013,AI557727, AI557807, AI541346,AI541517, D60844, D59436, AI546996,AI547250, AI557864, AI557084,AI525316, R29172, AI557408, AA174170,AL043444, AL044125, AI541356, T41289,AA585453, AI541510, D53472, C15737,AI546971, AI556967, AI526176,AI525500, AI541345, AI546921,AI546828, AI525320, AL041347,AL044529, AI540974, R29179, AI526180,AI557602, C14723, R29262, AJ239433,AL041233, AI557808, AL041296,AI526195, AL041086, AL043496,AI541390, AL044162, D54897, C15120,AI546829, D53161, AI526113, D59751,AL040193, C16294, AL041324, AI546855,AL043538, AL040621, C16292, AL041098,AL041163, AI526187, AL041277,AI541508, AL041358, AL041096,AL047012, R29657, AL040464, AL041346,AI535813, AL040155, T41329, AL041197,AI526158, AL043612, AL039915,AL040463, AI557155, AL047219,AL041227, AL047057, AL047170,AL040119, AL047036, AL041292,AL041051, AL047183, AL040322,AL041131, AL046330, AL041133,AL041159, AL041238, AL041142,AL045817, AI541534, AA585356,AL040625, AL040510, AL043467,AL044186, AL040529, AI541383,AL044037, AL040091, AL040128,AL040168, AL040255, AL040285,AL040342, AL040332, AL040617,AL045684, AL040745, AL040370,AL043677, AL046442, AL040839,AL041752, AL041168, AL043775,AL044165, AL041140, AL043492,AL041602, T23985, AL045920, AL037436,AL038838, AL044074, AL041635,AL045990, AL040458, AL044199,AL044187, AL040090, AL040263,AL040294, AL040329, AL040082,AL044272, AL040444, AL041186,AL040148, AL045725, AL041730,AL041523, AL043627, AL079878,AL046392, AL041374, D53447,AL040052, AL043845, AI525339,AL039338, AL042135, AL044064,AL038983, AL039316, AL043923,AL043814, AA585155, AL043537,AL043848, AL037435, AL041459,AL043570, AL041577, C15762, AL044201,AL044258, AL046850, AL038532,AL040768, AL037727, AI557238,AL046994, AL040253, AL040414,AI541017, AL040571, AL046914,AI142134, AL040576, T23957, AL045753,AR062871, A25909, AR038855, AJ244005,Y16359, AJ244004, D50010, Y09813,AF082186, AR062872, AJ244003,AR062873, A20702, D78345, AR017907,A20700, A43189, A43188, X81969,A85395, A85476, D13509, Z32836,AJ244007, AR037157, A98420, A98423,A98432, A98436, A98417, A98427,A98767, AR038762, A93963, A93964,I63120, I05558, I44681, X83865, A84772,A70872, I08396, I08389, A84776, A84773,A84775, A84774, AR054109, AR067731,I15718, AR067732, A86792, A58522,A91750, AR008429, AB025273, I13349,A18053, A60212, A60209, A60210,A60211, E03627, I84553, I19525, I84554,I48927, I15717, AR031566, A90655,D13316, AC005913, A35536, A35537,A02135, A02136, A04663, A04664,A02712, A77094, A77095, M28262,A81878, I00682, A95051, I06859, A18050,A23334, A75888, I70384, A64973,A60111, A23633, AR007512, A11623,A11624, E00609, E13740, A11178,E01007, A10361, AF149828, I08395,AR043601, A11245, AR054723, I03331,A02710, E12615, AR035193, A92133,E14304, A07700, A13392, A13393, I62368,AR031488, I13521, I52048, A27396,AR027100, I49890, I44531, I28266,I21869, A91965, I44516, A70040, E16678,I01995, A82653, E16636, A95117,AJ230935, A93016, A24783, A24782,AR051957, I25027, A58524, A58523,I26929, I44515, I26928, I26930, I26927,AR038066, I08051, X55486, A22738,I66498, I66497, I66496, I66486, I60241,I60242, A20699, E00696, AJ231028,E00697, E03813, I66482, AR009151,I66485, E17098, I66483, I66484,AR027099, I66487, AJ230902, AJ230972,AJ230951, AJ230867, AR051652,AR063812, S60422, AR051651, A06631,X82786, I66495, I66494, AJ231009,Y14219, A22734, X76012, AJ230845,I05845, AR035975, AR035977, AJ244006,AR022273, AJ243486, AR051865, I03669,I03668, A16035, I18895, AJ231011,I33632, AR003381, AJ230996, Y09940,AR035974, AR035976, AR035978,E03654, AR031358, AR031365, I36244,AR051864, A22739, AR017826, A05993,and A05991.HFPHR8221111877491-159715-1611AA913364, AW027373, AW305275,AI799031, AW117480, AA588138,AA775450, AW190848, AA411334,AA866178, W61038, AA411335,AA775769, AA769134, AA075643,N30274, AA627544, AI493881, AI147666,AA075644, AA614747, N30309,AA812101, AI025647, AA868924,AW339918, AI140386, N94919, AI143201,AI347094, AI138196, AA284960,AI933338, AI343592, AI128292,AI017993, AI492556, N20511, N41996,N42630, W72001, W69402, AI554835,AI679099, AA040263, T32745, AA872581,N29505, AI354734, AI268560, AI017992,F28362, AA448442, AI679673, AA863176,AA719956, AA444022, AI142803,AA732322, AI304774, AI475118,AA993672, AI624135, AA846235,W44314, W44778, W37097, AA682603,W65308, AA448307, AW274816, N25416,AI339799, AI128110, D79265, AI004213,AA305351, AA285117, AI149185,AI285067, AI872626, AI186604,AA557809, AA444000, AI344717,AA922743, N56641, AA996037, H13966,AA515842, F37123, AA732278,AA989130, W65338, W76570, R21057,N26843, AA721150, AI239957, AI580506,R96887, AA034909, W45686, W69363,F09166, H12989, W19542, N57447,R96888, C04314, AW183468, H87003,AI719584, R24370, T35889, AI685763,AA081676, T34115, N40568, AA010732,AW023063, N63592, T35834, AW089136,AA913531, Z42311, F11505, T35841,T30076, H05630, AA863208, AA505347,T30517, H27007, AI366940, Z42248,F09167, H87002, W56871, T36163,AA040400, W19619, AA721261,AA649338, F34780, T35871, R46237,H13965, AA913094, T35833, AA034976,AA890331, AI274646, N63619, AA082031,F05477, AA603299, AA322616,AA603386, R27044, Z38484, AI564372,AI859366, F01840, R27043, R27029,R27028, AA748381, F01739, AA355325,AI859362, H53598, N46608, AA339209,and AF190797.HISAF592129591401-89915-913AW401787, AI394630, AI418298,AW375742, T30407, Z44281, F07299,R25015, T32685, AA974700, F07734,AA297059, AW239548, AA897415,R45025, AI807678, AI343378, AW206793,AW138409, AW163027, AI815476,AA503315, AA047793, AW137324,AW140018, AI936871, AI015047,AI017077, AI168175, AI302185,AI025217, F03423, R46686, AI073417,Z40806, AA026054, AW002416,AI652375, F03562, T03397, AI983297,H42881, T82311, AI025310, AI831833,R08769, AI911100, AA471062,AW157059, AA382959, H22172,AI356604, AI537006, AI825970,AW338394, AW192088, AI559159,AA593826, AW078709, and Z61277.HCEHD6621311361221-92615-940AI968437, AI824971, AW104052,AI762197, AI598138, AI088543,AW135225, AI827280, AW007187,AI391466, AA534403, AW028554,AI492390, AI369729, AA135928,AA507443, AW137272, AW381735,AW006062, AW138526, W81153,AA918755, AA085774, AA058923,C03738, R85039, AI937792, AI867512,N62215, AI864402, N62216, D44882,H14329, H40979, AI808139, AA463408,AI744140, H40980, AI479413, N62166,AA319197, W30972, AA460467,AA135929, AI400940, N62223, AF186409,L27421, and AC006241.HE8UY7421411635901-59115-605N23547, H06088, Z24919, AA010516,R94366, AA004981, AA304780,AA343256, and N31310.HAHIY082159621131-26515-279AA100160, AA307684, AA244505,R57782, AA864846, AR044133, andAR044123.H2CBH4521611289191-98315-997AA307462, H86407, AA019170,AW351511, AA036880, AA045560,D89677, and AL133047.HMVAM0921711948281-183515-1849AI685410, AA873182, AI969804, N21495,AA708519, AA581446, AA035001,AI088511, N20223, AA101798, AA534317,AA397365, AA621392, AA397470,AI343916, AA035462, AI682643,AA397454, AA565111, AI096942, N68558,AA101799, AW029215, AI802673,AI130817, N73321, AA470951, AA470484,AI302901, N29290, AW024919,AA358533, AW135812, AI376856,AA740383, AA349063, AW051031, andAI276887.HFPEN0421811996631-209015-2104AI631883, AI767614, AA018867,AA019175, R56792, AI497937, N95592,R44001, AW271411, R56793, Z38315, andZ42023.HSLJD0221911044521-96515-979AI905856, AW178893, T03269, D80164,D58283, D59859, D80022, D80166,D80195, D80193, D59927, D51423,D59619, D80210, D51799, D80391,D59275, D80240, D80253, D80043,D59787, D80227, D59502, AW177440,D59467, D81030, AW179328, D80212,D51060, D80196, D80188, D80219,C14014, C14331, D80038, D80269,D80366, D57483, C15076, AW178775,D50979, D50995, D59889, AW360811,D80134, C14389, AW366296, AW375405,D80045, AW378532, D51097, AA305409,AW177501, AW177511, AW178762,AW377671, D51022, AW375406,AW378534, AW179332, AW377672,AW179023, AW178905, AW352170,AW367967, D58253, D81026, AW178980,AA305578, AW377676, AW352171,AW177731, AW178907, AW178906,D80248, D80132, AW179019, AW179024,D80522, D80133, AA514186, AA514188,AW179329, AW378528, D80247, D80439,T48593, D89785, D34614, X67155,AR018138, D88547, X82626, A84916,A67220, A62300, A62298, Y17188,A78862, D26022, AJ132110, A25909,AB012117, AR025207, A85396, A44171,I19525, A85477, AR066482, A86792,Y12724, AF058696, AB028859,AR008278, X93549, A94995, I18367,AR066490, D88507, AR008443,AR066488, D13509, A82595, A26615,AR052274, AR016514, D50010,AR060138, A45456, Y09669, AR060385,AB002449, and AR008408.HDPFZ3022012201641-168315-1697AA582196, AI690196, AW207633,AI417381, AI494563, H15131, AI220943,AI963799, AW055140, AI680222, N54913,AW008673, AI000592, H04038,AA385773, H03355, AW374840,AI014381, H15190, Z41172, AW374989,F08100, AA348883, N88565, AI924734,T03532, and AA348882.HPJCR3322112179311-240415-2418AA630312, AA399309, AI566447,W45583, AA148064, AI379707, AI041054,AA478756, AI581333, AI745305,AA253311, AA947298, AI744665,AA545782, AA194036, AA479976,R61402, AW297108, AI083947,AA398243, AA235081, AI700620,AI458557, W45546, AI433426, AI092537,AA195635, Z46141, H15959, AA235210,AA743383, AI656322, AA015701,AA766679, R23558, AI702152, AI370616,R61360, AI038624, AA018978, Z41774,R60379, AI535639, R14628, R68398,R31614, H01672, AA585439, Z28355,M85798, R31659, AA585101, R45613,Z30131, N87140, AI525556, AI546999,AI541365, AI525306, R68600, AI557731,AA585453, AI525431, AI535660,AI536138, AI547039, AI526194,AI540967, AI546855, AI541535,AI557807, T11028, AI557262, AI541374,AI541514, AI556967, AI526140,AI541510, R29445, D61254, AI525316,AI541523, H01673, AI546828, R28735,AI541013, AI557787, AA585440, D57491,AA585434, AL041159, AI541509, R32345,AA585476, AL041142, AL041238,AL041133, AL047183, AL040322,AL041131, AL046330, AL041051,AL041292, AL040119, AL047036,AL047170, AL047057, AL047219,AL041227, AL040463, AL039915,AL043612, AL045817, AL041197,AL040155, AL041346, AL040529,AL041096, AL047012, AL041358,AL041277, AL041163, AL041098,AL040621, AL043538, AL041324,AL040464, AL044162, AL041086,AL043496, AL041296, AL041233,AL041278, AL045725, AL041140, T23985,AL040193, AL040625, AI541534,AL040510, C16300, AL043467, AL044186,AL040553, AL038761, AL044037,AL040091, AL040128, AL040168,AL040255, AL040285, AL040342,AL040332, AL040617, AL045684,AL040149, AL040745, AL040370,AL043677, AL046442, AL040839,AL041752, AL043775, AL044165,AL043492, AL041602, AL045920,AL038838, AL044074, AL041635,AL045990, AL040458, AL044199,AL044187, AL040090, AL040263,AL040294, AL040329, AL040082,AL044272, AL041186, AL040148,AL041730, AL041523, AL043627,AL040253, AL046392, AL041374,AI535813, AL040052, AL043845,AL043537, AL039338, AL042135,AL044064, AL038983, AL039316,AL043923, AL043814, AL043848,AL041459, AL043570, AL041577,AL044201, AL044258, AL046850,AL038532, AL040768, AL037727,AI546891, AL046994, AL040414,AL040571, AL046914, AI142134,AL040576, AL045753, AL044274,AL079878, AL049018, AL040444,AL045857, AL041168, AI541205,AL039744, AL038822, AL045671,AI541508, T41289, AI525320, AL046327,AI557796, AL040238, C16305, AL049069,AL043444, AL041246, AL040472,AL040075, AL041347, AL041955,AI557799, T23957, AL080031, AR038855,AR062871, A25909, A85395, A85476,AR008429, AR062872, AF082186,AR062873, AR017907, AJ244004,AJ244005, A20702, A20700, A43189,A43188, AJ244003, AR037157, A98420,A98423, A98432, A98436, A98417,A98427, I63120, A98767, I18895, A93963,A93964, A58522, Y16359, I44681,AR038762, A84772, A84774, A84776,A84773, A84775, AR054109, AR067731,AR067732, A91750, X83865, D78345,A86792, AI8053, M28262, I06859, I13349,I15717, I15718, E03627, U94592,AJ244007, I48927, I19525, A02712,A77094, A77095, I84553, A81878,A95051, I84554, A18050, A23334,A75888, I70384, A64973, A60111,A23633, AR007512, I08396, A60212,A60209, A60210, A60211, I00682,A11624, A11623, E00609, E13740,A11178, E01007, A10361, A91965, I49890,A93016, A35537, A35536, A02136,A04664, A02135, A04663, I08395,AR043601, A11245, I03331, E12615,A02710, AR035193, E14304, A07700,AI3393, AI3392, I62368, AR031488,I13521, I52048, A27396, AR027100,I44531, I28266, I21869, A70040, E16678,A82653, E16636, A24783, A24782,A92133, A95117, A58524, A58523,A90655, I44516, AF149828, I01995,I08051, AR031566, I25027, I26929,I44515, I26928, I26930, I26927, I60241,I60242, AR038066, A20699, E00696,E00697, E03813, I66482, AR009151,I66485, I66483, I66484, I66497, I66496,AR027099, I66486, I66487, Y09813,I66498, AR051652, AR051651, Z32836,A62298, X81969, AJ230935, D50010,AJ230902, I05558, AJ230972, A62300,A70872, A82595, AJ230951, U87250,A84916, I08389, A22738, D13316,X07299, A70869, D13509, AR035975,AR035977, AB025273, AR051957,E12584, I18302, AR018138, AC005332,Y17188, AR035974, AR035976,AR035978, AR016808, I66495, I66494,AJ132110, E17098, I66481, A83642,A83643, I66488, I66489, I66490, I66491,I66492, I66493, A83151, AR008278,X55486, A22734, AR022273, AJ231028,AB028859, AF058696, AC005913,AR054723, AR064707, Y17187, AJ230867,AJ230845, AI231009, A94995, X67155,D26022, A67220, D89785, A78862,D34614, I36244, AF006072, AR051864,D88547, Y12724, A06631, X82786,S60422, AR051865, R32371, andAI191037.HTOAK3422210813211-82715-841AA491322, AA505126, and AW408167.HE8N1242239712961-73715-751AA883367, AA332611, AA732890,AI283442, AI673342, AI631153,AI200800, AI910962, T11417, D80258,D59503, D80014, D81111, C14227,D80064, AI557751, D58246, C06015,AA514184, AI535959, AW178893,AW178907, AW375405, AW177440,AI535686, AW360834, AW178908,AW360811, D80314, AA809122, D80251,D80253, C03092, D80247, D80043,AA285331, AW176467, C14389,AW179328, T48593, AW375406, D80439,AW378534, AW179332, D58283,AW377672, AW179023, AW178905,D59859, D80022, C14331, D80166,AW177731, D80195, AA305578, D80193,D59927, T03269, D59467, D51423,D59619, AW378528, D80210, AW178906,D51799, D80391, D80164, D59275,AW178762, D80240, D80038, AW179019,D59787, D80227, AW378533, D59502,AA305409, AW378532, F13647, D45260,AW178914, AW378542, AW360855,AW377676, I50126, I50132, I50128,I50133, AF123263, A70867, D88547,AR062872, AR066488, AR016514,A62300, D50010, X82626, AR066487,Y17187, AR060138, A84916, A45456,A67220, D89785, A62298, Y09669,Y17188, AB028859, A82595, A78862,D34614, A94995, D26022, AR060385,A30438, AJ132110, AR018138, A26615,AR052274, A43192, AR008278, X67155,Y12724, A63261, A43190, AR038669,AF058696, A25909, X68127, AR008443,AB002449, AR025207, AR016691,AR016690, and U46128.HAMFM3922410555321-462015-4634AI660231, AI304355, AI745391,AI951619, AI814592, AI467782,AI274105, AA426190, AW301083,AI375593, AI922346, AI829867,AW136366, AA227834, AW105735,AI304726, AW194167, AW297557,AI971865, AA151872, AI985643,AA757072, AW028756, AI202419,AI346603, AW295401, AW176248,AI659079, AA149658, AA425159,AI870033, AW194075, AA233413,AW102818, AI765117, R61588,AA365664, AA365663, AA601170,R61532, AA357346, and AA551861HBGMG3922511262831-34315-357AI497641, AW069135, AI623708,AW084071, AW339039, AI569972,AL036585, AW192429, AI458409,AI379125, AI066465, AW003414,AI742565, AI740930, AI743686,AI168481, AI991329, AI061450,AW007899, AW102701, AI342244,AI609020, AI453165, AI248142,AW044270, AI761292, AI693756,AI870883, AI129686, AI992036,AW057507, AW151858, AI362058,AI475537, AI864435, AI479128,AI378430, AW008808, AI080156,AW191953, AW080685, AI627434,AI369151, AI926062, AI963067,AI923881, AI740972, AL045227,AI351617, AI334039, AI452903,AI700412, AI469546, AI187911,AI066744, AA551905, AW132118,AI190677, AI812069, AI081231,AI050026, AA577674, AI890929,AI096904, AI554840, AW073739,AW190499, AW250073, AI524714,AI884727, AW168902, AI858574,AI422058, AA304774, AI890721,AI052823, AI683215, AA682804,AI087154, AW004991, AI640737,AA461453, AI668789, AA532435,AA912478, AW079985, AA054275,AA970861, AI819302, AW085591,AA010790, AI017023, AI217784,AI584163, AI581899, AI569968,AA984939, AI469425, AA709077,AA744639, AA779825, AI000961,AI095374, AA864689, AI564659,AA969103, AA827905, AA918799,AA862362, AA446048, AA024454,AW087352, N53410, AI672825, W48745,N20167, AA931463, AI587533, AI193638,AA612750, AW150032, AI961508,AA024670, AI619904, AA429899,W02113, AA969104, AW190663,AA765028, AA780072, N50640,AA533847, AW182184, W32574,AW073566, AI933617, AL041483,AI952283, AA937508, AI095341,AI917215, N48059, T23065, AI869852,N78603, AW444850, AA877348,AA010101, AA252354, AW297832,AA011219, AW204879, AA506948,AI167689, N53257, AI917614, AI937775,AW168023, AI858564, AW043696,AA450098, AI263865, AI885655,AW139189, AA883859, AI801558,AI859652, AI554186, AA151875,AA187616, AI828806, AI095024,AI628324, AI678623, AW161202,AI560004, AI393038, AI584130,AI687568, H89138, AA835947, AI673140,AI521005, AI287476, AI678324, E12258,AF153686, E12259, E12260, D44497,AC007559, A15345, X99971, AL080159,AL133062, AF061981, AF111849, A21103,and S36676.HSXBV8922611286991-223315-2247Z99410, AW373793, AA553665,AI190674, AI366341, AI184600,AW004692, H10539, AA482462, R45646,AI521134, AA973962, R42556, AA720003,H11452, AA211375, R22747, AW009222,AA417872, AA632195, W39346, R52677,R61178, AA720022, R61892, T80548,R20364, H46470, F07364, R86859,R55930, Z41453, AI360146, R87776,H08592, AA483558, W38313, R55720,R43749, T30657, AA426408, H95699,AA363950, R18352, R15260, R18538,R87830, F05821, R87766, R10214,R90750, T15918, F03602, T34747, H95698,W84692, R87845, T78493, F08462,T16643, AA210976, R13935, R25677,H50912, AA349344, AI302317, Z99411,F07877, AA643729, R52724, R43750,AA522619, F02068, R43495, R41527,H52035, AI092517, AI291295, AI570476,AI951902, AI906668, AI906666, H38986,AI553790, AW268395, AI312547,AW450243, AI681390, AI344432,AI343814, N44903, W84640, AA614791,AJ245820, AJ245821, AJ245822, andAF131749.HBIOZ1022711437561-55915-573HTLEJ1122810856511-95615-970M62294.HAWAM6922912078351-294415-2958AW385785, AA430300, AA541688,AA776700, AA679037, AA573270,AA126614, AL045796, AI268236,AA682186, AI963606, AI926591,AW192904, AI924827, AI922590,AI032288, AI375804, AA705172,AW081541, AA694514, AI130883,AI800450, AA931725, N25288, AI270687,AI366906, AW058362, AI683319,AA436891, R59176, AI597744, AI446542,W69578, AW453004, AI911821,AI095665, AA687634, AI130013, W69579,R59232, AA722782, AI587015, AI191864,AA398533, AA676733, AI476374,AA115447, AA554327, AA759328,AW242281, AI139766, AA042956,AA886732, AA664356, AA358590,AA135916, AI565897, AW304844,AA618576, AA916086, R66162, H71919,AA363371, AA430199, AI370031, Z44808,AA320329, AI934183, AA393105,AI004140, AA135927, AA042816,AW452852, AA135926, H44791,AA813424, AI865731, T35731, R42647,R27785, T32691, AI857286, AW008428,AI631988, AA115446, AA678468,AW075384, AI569918, H44790, AI918635,AA601518, AA603858, H42641, AI745618,AI445766, R27874, AI939990, AA677131,AW364938, AI569374, AW029062,AL042382, AL119511, AL119457,AL042544, AL119399, AL079794, C01947,AL043152, AI863382, AL043168,AL119324, AI670009, AI698391,AL036403, AI358701, AI909661,AL046944, AL037454, N42321,AW082113, AI886124, AI590120,AI978703, AI525669, AW167228,AI525653, AI254727, AI637584,AI783504, AL079963, AI557808,AL119863, AI609409, AA127565,AW020397, AI247193, AI889189,AL039086, AI866608, AI590686,AL045500, AL121306, AI612885,AI364788, AI433157, AI702073,AI888661, AW238730, AL043975,AI541027, AI633125, AI610357,AI673278, AL079741, AL121328,AW022682, AL045997, AI859991,AI686906, AI923989, AI621341,AI269862, AL042745, AI620284,AI915291, AW020419, AI340519,AW023338, AI522052, AI874166,AI866770, AI679550, AI284509,AI541056, AI801793, AW163464,AI280637, AI744243, AW104641,AA640779, AI537677, AL045163,AL041150, AI571439, AI538885,AI557238, AL121365, AW411235,AA493647, AI491842, AL080011,AI627988, AI340603, AL121270,AI623941, AI583578, AA420758,AI538850, AA420722, AI624693,AI499285, AL047100, AI590601,AI669864, AL110306, AW020592,AW051088, AW021717, AI554821,AI866573, AW161579, AI801325,AI539771, AI929108, AW162189,AW198090, AI521560, AI500662,AI619502, AI500659, AI285448,AI541048, AI538008, AW160916, D50977,AI500523, AW079654, AW162194,AI284517, AI500706, AI445237,AI491776, AW151138, AL036548,AW409775, AI922707, AI633493,AI620003, AI434256, AI591228,AL138386, AI284513, AI888118,AI859464, AW104056, AI582932,AW303089, AI582558, U77594, Y11587,S68736, AL050277, AF118094, I48978,AL133072, AR011880, AL122110,AL137550, Z97214, AF182215, I89947,AL122093, AF126247, AL133640, A91160,AF146568, AL133080, A93016, AL049382,AR029490, A08916, A08913, I48979,AF100781, Y10655, AF113694, AF177401,AL137479, AF207750, X56039, AL050393,AR034821, S61953, X82434, Y11254,AF079763, AF159615, AL050149,AL096744, A08910, A08909, A77033,A77035, Z82022, AL137294, AL117460,U35846, AF113019, AF114170, AL050024,AL137459, AL080126, A65341, I03321,AJ005690, AF090934, I33392, AL133075,I68732, X63410, I89931, X79812, M27260,I00734, I49625, E00617, E00717, E00778,E01614, E13364, A76335, AB007812,AL117435, A03736, AF057300, AF057299,X72889, AF139986, A57389, AL122049,A07588, AF067728, AF111851, AF183393,AF091084, AL110221, AF090900,AL137548, A18777, AL049283,AL137560, X70685, AL133557, Z37987,AF090903, S78214, D83032, AF017437,Y16645, AF090886, I66342, AF104032,AR068751, U88966, AR038854,AF065135, AF087943, AL110197,AL133016, I09499, AL133113, A58524,A58523, AL122050, AL049430, AF113699,AL137271, E02349, I92592, AF026124,A08912, X65873, AL122121, AJ012755,AL137463, X81464, AF028823, A08908,AF069506, AL122100, AL137533,AF079765, AL137521, AF008439,AL117457, AL133558, AF017152,M96857, U80742, AF090901, U75932,E03348, X66862, AL137478, I17544,X63574, AF061943, AL080074, AL049300,AL049466, X84990, A93350, I30339,I30334, AL117394, AF106657, AL050138,AL122123, AL133560, AL133637,AF113689, X72387, U00763, AL137523,AJ003118, AL137557, AL137300,AL110225, AF081571, AL137705, L04504,X96540, I89934, AR013797, AF113690,AL137283, AF118070, AL110196,AL049314, AL049452, AR059958,AL137538, AJ238278, AL122098,AL050146, U78525, AR068753,AL137526, AJ000937, AL080158, E07108,AL133606, AF106862, AB019565,AF113677, I09360, AF097996, E06743,AL137648, AF031147, AL117585,AL137558, AL050108, AF090896,AL133010, U42766, X83508, AL137476,AF111112, AF111849, AL133665,AL133568, E15569, S76508, AF113691,AL137539, AL080124, L04849, A65340,AL137530, I42402, AL080159, AL117583,M86826, X92070, Y14314, AL050116,AF125948, AL137660, A23630, AL137488,AL050155, AF113013, AL049464,AF078844, X06146, AF100931, A86558,AF090943, X87582, U67958, AL122106,U87620, AF125949, L31396, AL080148,and AW664335.HSCKD1123010562881-371415-3728AL037558, AA417129, AW021717,AL042365, AI440238, AI885989,AA641818, AI378123, AW088560,AI648699, AI673278, AI572717,AI559752, AW023338, AI280732,AW238730, AI538885, AI866608,AA805434, AI923989, AI249877,AW022636, AI697324, AL118781,AL120995, AA514684, AI349226,AL048323, AL079963, AI929108,AL048340, AI697420, AI784252,AI318280, AW151714, AI811192,AW022102, AW161098, AI565031,AW029457, AI471429, AI862880,AW163834, AI536836, AI950729,AI582932, AI521594, AW088903,AI862144, AI954183, AI288285,AL037592, AW118448, AI348777,AI335426, AW082532, AI537677,AI611743, AW166742, AW161579,AI567971, AI954422, AI633125,AI925404, AI809062, AI860027,AI815232, AW152182, N22276,AW162194, AI538716, AI345688,AW366372, AI334893, AI440260,AI241678, AI335214, AI493836,AI499285, AL041220, AI432644,AI798456, AI702091, AW021189,AW073697, AW083573, AI537261,AW089840, AI252023, AI586931,AW151132, F26535, AW020480,AL047344, AI340603, AI963668,AW161402, AI589004, AI473208,AI524654, AI621341, AW104141,AI338427, AI287233, AI349246,AA908294, AA420722, AI886123,AI623941, AI340519, AI690813,AW170787, AI254042, AI627714,AW051088, AW020381, AI680165,AI473536, AW020397, AI240978,AW079432, AI365256, AL036923,AI269862, AI635016, AL038605,AI368579, AI623736, AI500061,AI699020, AW129456, AW087336,AI289791, AI138480, AI567351,AW029216, AI927233, AW059828,AI866465, R39624, AI969641, AI589428,AW022699, AI915291, AW020419,AI470651, AL048496, AW169234,AI421328, AI587606, AI439452,AI539771, AW169604, AI540674,AI745329, AI282652, AW054931,AW020561, AI538829, AI887163,AI525653, AW129202, AI969655,AI610086, AI348854, AL121270,AI523806, AI432532, AI282930,AA602414, AW090768, H89138,AI873644, AW189473, AW162189,AI536601, AW079706, AW148356,AI345415, AI355779, AW074605,AI254420, AL110306, AI625467,AL045349, AA806719, AA833760,AI620056, AI591101, AW118237,AA736984, AI680280, AI538764, F27788,AA503384, AI623396, AW078895,AI868831, AL135024, AI758924,AA829061, W45039, AI472487, AI434223,AI289436, AI962858, AI673363,AW129230, AW194014, AW022300,AL039086, AL043981, AW090103,AI890907, AW025464, AW103628,AI554821, H43387, AW029186, AI499325,AW080766, AA808175, AL046942,AW090492, AI698391, AI863479,AI612885, AW087200, AI921244,AI804531, AA001397, AI340627,AL036705, AI677796, AW079572,AI696358, AI247293, AI452560,AW019985, AI620287, AI246319,AA764968, AJ243342, I72363, U12336,S53987, M30514, AL050116, AL137256,AL133080, I89934, AC005291, A65340,AF017437, AF200464, AF058921,AL133560, AF047716, AL122110,AF026008, AL050280, AF113676, A08910,A08909, M79462, U78525, I48978,Y16645, AL137258, AF124728, AL080148,AL137276, S36676, AJ005690, AL133113,I00734, AC004987, AL050393, U72620,E01356, AL080118, AF008439, AL133557,AF113690, AF090896, AF002672,AL133010, AF102578, AR038854, I89947,AF000167, AF082526, AF118094, L40363,A08913, AL137459, AL117460,AR034821, AL137550, AL137463,AF100931, A86558, AL133070, AL080159,U73682, X80340, U58996, X89102,X59812, AF043493, AF090934, Y13350,AF145233, AL110158, AF090901, E00617,E00717, E00778, AL122104, X62580,AF069506, AF183393, AF185614, I66342,AL050172, L13297, Y10936, A08912,Y07905, M92439, A08911, A08907,AF038847, I18358, I34395, AL080074,AL110228, AR068466, U88966,AL080057, S54890, I89931, AL133067,AL133084, X76228, AF097996, AF051325,S76508, X72387, U51587, AL137558,AL133623, I49625, AF106862, AF090923,I33391, AR029490, M85164, AF080622,AL122093, AL137479, AL122123, I79595,AF002985, AL137657, AF175903, E12580,AL137648, AL117629, AF158248,AF180525, I32738, AB008792, AL122118,U75604, AB008791, U42766, AF182215,Y13653, A91160, I28326, A77033,A77035, U68233, I92592, Z37987, A03736,A18777, A08916, AL117463, AL137478,AL122050, AL133081, AL117416, S77771,AF118558, AL133568, AR050959,AF111112, AF067790, M19658,AR020905, AL133104, AL137429,A08908, Y10823, AF065135, E12579,AF090900, U37359, X99717, AL080150,AR022283, S78214, AL049347, AF026816,AL136884, and X82397.HDPLT6223110272411-269315-2707AI279417, AL079734, AA502991,AA410788, AI755214, AA643770,AW023111, AI754105, AI754567,AI805363, AW328331, AA632765,AL042373, AI254779, AA704393,AI962030, AA659832, AA507042,AI253376, AI380617, AI923052,AA524229, AA904275, AI460050,AA714110, AI345695, AL119247, N23913,AA572813, AW008169, AA470933,H45698, AA169245, AA477503,AA829036, AA535216, AA452887,AA809546, T71936, AI311647, AI278972,AI783911, AI674840, AW409710,AA581247, AI620585, AI754767,AA524616, AI114733, AI677655,AW265688, AI133552, AA618316,AI627614, AI491765, AW419389,AI053696, AA632993, AA601394,AI056046, T07225, AI270602, AA837686,AL120343, AW083934, AL135108,AW193493, AA714288, T74524,AI912401, AA228778, AI921765, R99034,AL041894, AI242011, Y18994, D50671,AP000031, AF053356, AC006333,AP000252, AL008718, AC004858,AC004999, AL049757, AL121653,AC006538, AC006449, Z82172,AC005751, AC005480, AB023049,U80017, U62293, AL049759, AL049830,Z81369, AC020663, AL034548,AC005057, AL096701, AC000004,Z95114, AL079295, U91326, AL035086,AL021977, AC002350, AC010582,U63721, AL023655, AL023553,AC005821, AP000134, AP000212,AP000116, AC004991, AC000353,AB003151, AC005031, AL031295,AC010205, AC005736, AL139054,AL133245, AC006480, AC005288,AF130343, AL022238, AC006057,AC004895, AC005363, AC007637,AP000304, AC005274, L11910, Z95118,AC005725, AF111168, AC007707,AC007041, AL031662, AC007216,AC004148, AF064858, AL031311,AC003042, AC005695, AC007688,Z97205, AC007283, AL031005,AC005104, AC005678, AF109907,AC005777, AC003007, AC007371,AP000688, AC006088, AC007450,AC003957, AL034420, AC006537,AL096791, AC009516, AL031276, Z99716,AC004905, AL049713, AL022723,U91318, AF001550, AC004859,AC004598, AC007774, AL021154,AC005037, AC007899, AC005067,AC005046, AC000118, AL132712,AL031770, AC005332, Z86061, Z98946,AL049776, AC005779, AP000044,AP000112, AL031779, AP000513,AC005365, AC004655, AC005291,AL135744, AL008582, AC004755,AC006344, AL022316, AP000344,AC006084, AC004408, AL080317,AC007686, AC006285, AL078638,AC004025, Z83840, AF196969, AC007731,AC005619, AC006199, AC004771,AL031281, AC004797, AC016025,AC006046, AF001549, AC007014,AC000026, AL121652, AC002352,AC000062, AC002431, AL020997,AC005041, AL132994, AJ246003,AC004526, Z84483, AC004890,AC005231, AC004883, Z84487, AP000065,AC004821, AC002558, AL009179,AC002059, AC007193, AC002544,AF196779, AC005081, AC004878,AL133163, AC008115, Z99755,AC005565, AC005899, Z95115,AC004834, AF190465, AL031768,AL035697, AL022336, AC005632,AL078581, AL022311, AC005409,AJ003147, AP000545, AC005005,AL035495, D84394, AC005911,AC000003, Z86090, AL121658, AL049869,AC007766, AC002364, AL035691,AL031230, AC002565, AC004129,AC008126, AL031123, AC005839,AC005207, L78810, AC005089,AC006160, AC005902, AC002477,AF124730, AC004650, AC005944,AC005519, AL031848, AC002351,AC002429, AC004832, AL023281,AC006012, AC005783, AL009173,AC004765, AC004601, AC005378,AL021453, Z98941, U95742, AL033376,AL121825, AL024498, AC006241, Z85996,AC006120, and AL021391.HTPFX162329742961-47015-484HE9NO6623310796241-97615-990AI732997, AA865818, AA977633, Z69734,AB035267, AB020741, and Z68339.HSDJI4423411540681-209315-2107A38246, A11524, AR029497, E05333, andE00893.HFXDP5323511262941-156715-1581AR001481.HWADY662367345651-49015-504AA923698, AI673803, AI302688,AF191838, AR016417, AF145705,AF191839, and AC000025.HLDBC6323711445571-85915-873AW374344, AI424750, H12370, R32561,H12318, R28631, AI638545, AI760745,L39211, L07736, U88294, and Z56143.HFIVB682389782111-82315-837AA332003, AB033033, AF182316,AF182317, and AL096713.HTLAC5623911813551-83615-850AA614273, AA149526, AA722818,AA151569, AI299133, AI150935,AA542982, AI360422, AI079938,AA603610, AA716009, AI751611,W79786, AI083848, M78943, H40867,T31238, AA662256, AA298393,AA297499, AA541718, W79587, andAI074815.HSSAD412407530941-56615-580AA149526, AA151569, AA722818,AA614273, AI299133, AI150935,AA542982, AI360422, AI079938,AA716009, AI751611, W79786, AI083848,H40867, M78943, AA603610, T31238,AA662256, W79587, AA541718, andAA298393.HCFMT5724112184361-215515-2169AI452994, AW370315, AI333235,AI983346, AI811889, AA598963,AW370305, D81728, D81129, AA344996,R14748, AI699873, AC004687, AB014512,and AF039571.HDAAV6124211887871-131915-1333AA314786, AA160847, AA158845,AA157440, AA083972, AA159380,AI762433, AW404501, AI191825,AW105372, AW386309, AA157878,AI140935, AI922109, AW386330,AA442139, AA159268, AA083866,AW390887, AA100183, AW390888,AA262414, AA041219, AA488548,AA158846, AA158270, AI187149,AW076076, AI167533, AA743207,AA837990, AA442140, AA488413,AF125532, Y11092, AL137565,AC007136, and Z25424.HDPKD7524310962531-62315-637AA923698, AL040000, Z21326,AR016417, AF191838, AF191839, andAF145705.HTEON2924411263121-53815-552AW004028, AA432290, AI968030,AW237673, AW138422, AA428635, andAA861634.HSKAC2424511218001-49715-511AF170301, AF170302, AF077659, andAF144573.HTJAA7124612164981-172115-1735AW238721, AW265324, AW238371,AW238695, AW238323, AW084388,AF077831, and AF185276.HTEKS2024711243781-106115-1075AI936596, AA868353, AI797296,AA725553, AI221970, AI073397,AA428462, AA429551, AA431190,AA629305, AA629047, AW235895,AI123443, AI808267, AA609412,AI914363, AA953895, AI214385,AA431516, AA911681, AA781953,AI825106, AA298758, AI215028,AA909534, AA723768, D10393, andS63991.HE9TK4924811251921-135315-1367AA086273, AF126965, AF126966,AC004590, AF124351, AB012043,AF134985, AF134986, AF125161,AF027984, and AJ012569.HCHAT0124912022751-297715-2991AL079756, W80383, AA570709,AA746031, AA316540, N24219, W78982,AA740804, M78765, AA985314,AW337818, AA348323, H23489, W01009,AW273022, F10250, AW050885, H30484,AI494441, AA732761, N41828, Z41805,D51145, AL041943, AW079303, Z46179,H20834, H12176, H90615, AA641974,N28433, N45481, T31584, F12634,AA342119, R43266, AW365055,AA459071, AA464017, AA464078,R43775, T78049, T31949, AA046221,AA477480, AW139948, AW372243,AA143521, AW372245, R17167,AI655751, AA496370, R43146, T31978,AA496647, R25017, AA348324, H46223,T35092, AW071171, AA150812,AI085437, N99421, AA342120, AA131786,R08999, R09622, AW020710, andAB014576.HCEEN0625011508671-75215-766AW249673, AI568057, AI762163,AW300914, AI760528, AI341537,AI685427, AI572091, AI560209,AI453713, AI025000, AI761048,AI972204, AW170258, AW009963,AA864405, AI343959, AI912498,AI079913, AI559574, AI418784,AI438969, AI559101, AI973222,AI660152, AW016547, AI986363,AA806938, AI656111, AW003957,AI018296, AW028203, AI652597, R52151,H27175, D20315, H27174, and H77974.HDPKI832518833821-78115-795AA287508, W15561, N80792, AA922507,and AF159356.HSPBQ1225211522581-113015-1144W02910, AA282287, N72351, AA829957,AW205583, AW408526, AA282483,W86878, AA249705, R97467, AI090365,AB014536, and AF077226.HPCID782538869151-79315-807HDTKQ142548869361-54115-555AL023653, AL049683, AL359542,AL359542, and AL359542.HRACK832558880371-56615-580AC005832.HSIAO7825611564381-199015-2004AA527435, AW195324, AI653000,AW051613, AA514619, AI652532,AI675204, AA435717, AI796596,AI273289, AI659333, AI880669,AI826786, AA889355, AI174916,AW004627, AA377072, AA255838,AA397980, AA430523, AI565825,AI435476, AW001866, N52904,AA430608, AI760594, AA298640,W69756, AA594479, AI149418, AI911011,AI871818, N71537, AI089421, AA400874,AI038591, AA854839, AW044396,AI565867, AI131012, AI144119, H65663,N47230, AI732273, AW079534,AA847967, AW027678, AL044698,AA224892, T36269, AA009702, AI668849,AW182206, AA011130, N78511,AI676028, AA968449, AI984040,AA207018, AA658246, N73670, AI937659,R53598, AA453038, AA904224,AW293549, R48261, AA775033, H52314,R38289, H48428, AW083969, AA588654,F10880, AA578060, AW298073, W25831,AA889378, AA483944, AC002302,X62260, AC002288, AL035588,AC007425, AC004216, AF181896,AP000280, AP000038, AP000107,AF003528, AL033525, AC009498,AC007676, AC006080, AC005704,AL022328, AL118497, AC005332,AC007221, AL132987, AC005011,AC004874, U69570, Z48484, AB020858,AC004382, AC005213, AL049753,AC005771, AC004894, AC006023,AC003976, AL035668, D83402,AL139165, K03021, AC005046,AL035671, AC005184, AC005235,AC005183, AC005004, AC002541,AC005161, AL117338, AC004100,AF042484, AL117436, AC006354,AC004671, AC002546, AC007637,AC004129, AC004875, AL021877,AC006599, Z83822, AP000030, U61375,AC003693, AL009183, AC005922,AC004098, AC004982, Z75741, AJ006345,AL049821, AL035663, Z95114, AL080286,AL021395, AC008012, AC007529,Z84484, AF095703, AC006973, AL035696,AL078638, AP000252, AC007392,AC004061, AL117375, AL133162,AL031123, AP000466, AC007680,AC006952, Z82246, Z86062, AL035427,AF001550, AP000134, AP000212,AC011605, AL035106, AC006360,AC012088, AC000378, AC004984,AC005060, AL022159, AF001548,AC011594, AL135746, AB014079,AC004616, Z96074, AC007543,AC004015, AC004530, AL024508,AL031390, AC008069, AC005533,AC009263, AC000119, Z97876,AC004838, Z68324, AL031737, AP000088,AC006398, AC006139, AP000140,AP000146, AC005086, AP000228,AL132718, AC007198, AC007450,AP000299, AC004889, AC004055,AL035633, AC009275, AC004882,AL021707, AC007314, AC007156,AC006511, AC002563, AF152365,AP000514, AC005553, AC005690,AF047825, AC006367, AL034425,AL117347, AB020865, AL020989, Z84480,AC004888, AC005562, AP000113,AP000045, AC005529, AC005393,AC005538, AL021407, AL049839,U95740, AL031120, AC005701, Z75957,AC007617, AC000072, AC012083,AL008635, AC004381, L77569, AL031056,AC009464, Z84488, AC011700,AL110120, AL109914, AL035072,AC007262, AC007455, AL022162,U91319, AL118512, AL022329,AL121866, AL034555, AC005587,AL023806, AJ006995, AC006196,AC006213, AP000536, AC004968,AC006998, AC006064, AC007628, andAC002132.HWAGS732578944041-58715-601AF156884, AL096870, and AL096870.HCMSL082588982031-174015-1754AI141895, AI201842, AI351867,AA251985, AA197256, F20444,AA194198, AI800873, F36894, F26457,AA482914, AA197255, AA482932,F30374, F32249, F27719, F32998, F01019,F01208, F31596, F34858, F00856, F29622,AA194227, F29621, F36047, AA016148,F00201, AA178901, F34860, AA176342,F35465, AA252091, F21999, AA192776,AI023647, F34620, S73775, U93291,M15747, M22717, M22712, M20142,M22714, and M22713.HLWFN6325911015331-308915-3103AA707313, AI880426, AI684827,AI744551, AI307796, AA101249,AI284152, AA007399, N98643, AI375268,N66095, R71685, R02817, AA085724,AI221876, AI061056, AW207571,AA111956, AI460369, AI333887,AA594062, R18624, R62793, W22434,AW007868, AA776586, T70023, R71720,H70803, AA101290, AA323135,AA029721, AA320669, AI193496, R07828,AA007478, AI915644, AI932703, T69946,R62792, AA029660, AI859215, AA205667,AI625446, AI273982, AB018333, andAC006599.HPWAY1026011280331-115415-1168AI027613, AA332875, AW367440,AW376896, AA330257, AL042522,AW367550, AW304212, H80390,AA447076, AI923485, AW316546,R08512, AB023178, Z30174, AC007676,AC007842, AC004696, AC005498,AB021641, and X78933.HOUDH1926111539091-39215-406AW207457, AI084622, AI392839,AA628082, AA534387, AW243188,AW129204, AI695812, AI572814,AI948743, AA651748, AA301865,AW371758, AC007842, and AC005614.HDPFF2426211947191-177415-1788AI279875, AW369718, AI800428,AI566117, AA446945, AI246362,AA565728, AI436697, AA443369,AI761945, AW237445, AI889318, N66534,AW103788, N52334, AW302153,AW131719, AA588564, AA261806,H56725, AA199832, AA287107,AW419044, AI015242, AI807571, T80574,AI929099, N75104, AI269178, AA207149,AA912487, AA086082, AA780112,AI928828, R38930, AW450464,AA766626, AI904377, AA086081,AA365879, N88094, R68759, AA662606,AA281290, AA577126, R58834, H67457,D58175, N87549, H56648, T03486,AA261992, N28632, and AI913676.HWLFH9426311522781-126815-1282AI339104, AA861042, AA134985,AA868144, AA134946, AI626100,AA922724, AA535447, AA308766,AA056635, D25742, AA916634,AA551763, AA873574, AW192836, andAR044148.HWMBM1326411522831-87915-893AI339104, AA861042, AA134985,AA868144, AA134946, AI626100,AA922724, AA535447, AA056635,AA308766, D25742, AA916634,AA551763, AA873574, AW192836, andAR044148.HFIUE7526511725251-159615-1610AA745592, AW408392, AA780791,AI680317, AA205127, R06019,AW074511, T76970, T86065, AI709216,T77135, R05922, T85884, R77022,AA730855, C14389, C15076, R65976,D81026, D59467, D80164, D81030,D59787, D80166, D59619, D59502,D80210, D80240, D80212, D80219,D80188, D80022, D59859, D51423,C14331, D51799, D80253, D80043,D59610, D58283, D80195, D80391,D59275, D80227, D57483, D80366,D80193, D80196, D59889, D59927,AA305409, D80241, D50995, D80251,D80269, D80038, D50979, D80522,C14429, D80024, AW177440, D51022,D80045, AA305578, D51060, D80378,C14014, AW378532, T03269, C75259,AW178893, D80248, T11417, D80133,AW179328, AA514188, AA514186,AW377671, D80134, D80268, AW177501,D51250, AW177511, C05695, AW360811,AW178762, D58253, AW369651, C14077,AW352158, AW178775, D59373,AW176467, AW375405, AW352117,AI910186, F13647, D80439, D80132,D80247, AW366296, AW360844,AW360817, AW375406, AW378534,AW179332, AW377672, AW179023,AW178905, D81111, AW378540, C06015,D51213, D80302, AW352171, AW377676,AW178906, AW352170, AW177731,AW178907, AW179019, AW179024,AW360834, AI905856, AW177505,AW360841, Z21582, AW179020,AW178909, D80157, AW177456,AW179329, AW178980, AW177733,AW378528, AW178908, AW178754,AW179018, D80014, AW352174, C14227,AW179004, AW179012, AW178914,AW378525, D59627, D51103, D51759,AW177722, AW177728, T02974,AW179009, AW367967, D59503,AW178774, AW178911, AW378543,D58246, AW352163, D51097, D80258,D59653, AW352120, AA285331,AW178983, D58101, AW178781, T48593,D80064, AW177723, D45260, D80168,H67854, C03092, AI557774, AW177508,C14975, AI535850, D80949, AW367950,D51079, H67866, AW378533, C14407,AA809122, AI525917, C14344, AI525923,AW178986, AW177497, D51221,AI535961, T03116, D80228, AW177734,D59317, D59474, C14973, AI525920,AA514184, D59551, C14298, C14957,D45273, AI525215, D60010, AI535686,D50981, AI525235, D60214, AW378539,AI525227, C14046, AI525912, D51053,AI557751, T03048, AI525242, AW378542,AR018138, A62298, A84916, A62300,AJ132110, AF058696, AR008278, A25909,X67155, D26022, Y17188, AB028859,A67220, D89785, A78862, D34614,D88547, Y12724, X82626, A82595,AR025207, AB002449, A94995,AR060385, AR008443, AB012117, I50126,I50132, I50128, I50133, AR066488,AR016514, X68127, AR060138, A45456,A26615, AR052274, A85396, AR066482,A44171, A85477, A30438, Y09669,I19525, A43192, A43190, AR038669,A86792, I18367, AR066487, I14842,X93549, AR066490, AR008277,AR008281, U87250, D88507, AR054175,D50010, Y17187, A63261, AR008408,AR062872, A70867, AR016691,AR016690, U46128, U79457, D13509,AR060133, A64136, A68321, I79511,AF135125, AF123263, AR032065, andAR008382.HNTCP1326611823131-224315-2257AA463847, AI479379, AW273740,AI740675, AI922082, AW009462,AI014722, AA463334, AI073540, N95224,AA007373, AI190238, AI798079,AA476563, AA670286, H02882, N92851,AA652716, H45475, W25554, AA774170,AW016339, H45576, AI370125, H03781,AW119159, AI811794, H20952,AA853882, AA853883, AI471060,AW382128, AW371996, W21053, H20991,AA368628, AW138258, AA476448,AA876335, AA788825, AF037447, andAC004486.HB1BQ8926711751111-284915-2863AA496424, AW296705, AA292435,AA768388, AI359369, AA136659,AA435941, AW192609, AI361569,N62073, AA136739, AA411140, AI940074,AA101178, H09588, AA419044, AI358988,AA399613, AA323662, AL118665,AI268044, AA436039, H09587, AA627787,N55651, AA325534, T15506, AA082253,AA423798, AA904377, Z41610, R19206,F01655, F11248, R44468, C00466, Z42117,N77119, F08911, AA767800, AI418714,F05395, T35421, and AB007925.HWBEG1826811691251-44815-462AI114866, AI500518, AW327796,AW328350, AI755116, AW328609,AW410322, AW409642, AI754439,AW409590, AI754460, AI287514,AA551550, AA483482, AW205951,AA535393, AI201181, AI699829,AW409683, AW328376, AW328614,AW087373, AI827247, AW328444,AW327862, AW328440, AW328380,AW007896, AI971783, AW328007,AI628924, AI610070, AW080272,AW134985, AI440000, AA603360,AA501684, AI560651, AI559540,AA574413, AI818460, AA854768,AI754142, AA283266, AI497632,AA772414, AA496883, AA679713,AI050044, AI619744, AI339813,AW250421, AI274211, AW020501,F32918, AA579416, AA652660, AI567937,AA632536, AI831479, AI186976,AI151481, AW328320, AA877933,AI185119, AA513486, AI620681,AI690593, AA558105, AA610650,AA632560, AI922235, AA843775,AI924171, AI961721, AW250755,AW090148, AA491636, AI338728,AI123375, AW081336, AW090155,AI539209, AI890302, AW137738,AA847963, AA632723, AW007719,AA757769, AI573062, AA886011,AW245791, AI028444, AI863898,AI439763, AA886913, AA508174,AI749978, AA863478, AW261931,AA714364, AI634383, AI191638,AI198771, AI719450, AI344453,AI718439, AI268677, AI253560,AW250772, AI796657, AI347293,AI619835, AA569292, AW169077,AI570813, AI697471, AI149358,AI355377, AA610275, AI565047,AI193415, AI571454, AW262848,AA954603, AA600356, AI674831,AW316876, AA536172, AI185211,AI660181, AW273029, AI818029,AW192285, AA580796, AI719806,AI813549, AW242012, AI620986,AI830017, AI478688, AI925379,AI683998, R02544, AI745129, AI491901,AW338471, AI582160, N91538,AW090784, AI610180, AI697356,AI660159, AI925537, AI224078,AI859783, AW028278, AA598891,AI281231, AI289421, AW105711,AW305195, AA908802, AW073669,AI800405, AI342580, AI432916,AW170472, AA513180, AA653476,AI272858, AI963461, W73177, AI620289,AI523503, AI891159, AI475307, F24388,N32326, AW008147, AI270199, AI924530,AA448266, N64026, AW118551,AI560707, F20364, AA776791, AI206373,AI189997, AI510744, AA723534,AI631303, AA991428, AI979037,AI689023, AA505910, AA304640,AI962912, AA879052, AW073952,AI673755, AI951247, AA737215, F21939,AW005146, AI624705, AA879000,AA662258, AW023162, AA609197,AA984855, AI983037, AI734902,AI921779, N32870, AI718498, AW170473,AI924173, N34014, AI860956, AI523495,AW166489, AI818256, N31753, AI583997,AI000938, AI749136, AI333494,AA580751, W85708, AW328378,AA580635, AW328608, AI439940,AI830217, AI924195, AA491865,AI986358, AA312014, AW440520,AI860497, AI557538, AI185035, W37825,AI160628, AW148607, H73192,AA888879, AW273989, M36072,AC000089, AJ224080, AC004217, X06705,X61923, X52138, AC002107, AB023058,AP000521, AF055066, AL022723,AC004192, AL034417, AJ224082,AC004172, AJ224081, X15013, AC000399,Z84469, D63790, AC005042, AL031736,AC004129, AL078595, AC007110,AB020653, AC002452, and Y17212.HTAHB4326912219561-234415-2358R53155, AW374891, AA029227,AA010232, AA149376, AA077657,AA077229, AA077669, AA076731,AW405369, AW386712, AW386708,AA077893, AF139794, AA077980,AA077825, AA078429, AI768799,AI912020, AA077544, AI738591,AA076983, AA076893, AI125178, F01048,AW373785, AI963990, AB011110,AC004084, and AC004985.HSYBX322709098461-21615-230AA078429, AC004084, AB011110,AC004951, AC004951, AC004951,AC004084, AC004084, and AC004084.HCEHE3527111245311-53715-551AB019692.HFCBB5627212049711-214415-2158AA547979, AI696793, AA847499,AI889995, AI279417, AA669155,AA833875, AA833896, AL036909,AW162288, AW237905, AA862183,AA019542, AW408767, AA491955,AA904211, AW265688, AI366555,AI620992, AI251034, AI250552,AI284543, AW303098, AI251203, F13749,AW407889, AW026305, AI251284,AI421950, AI628922, AA831426,AA916430, AA828834, AA339423,AI493583, AL038606, AA614214,AI223626, AI224619, AW005974,AI492579, AA572983, AL047247,AA864271, AA053463, AL042667,AL042670, T57767, AI254770, AW316599,AA600202, AA832016, AA565232,AI278372, AI791185, AI753969,AL022316, Z84469, AC007934,AC000025, AL122020, AF015262,AL049869, AJ229043, AL021977,AC009516, AC005800, AC004821,Z83844, Z83822, AL031681, AL033392,AC005726, AC007298, AL133163,AC002549, AC006001, AL034429,AC006530, AL031311, AP000558,AL034400, AP000310, AC002395,AL022067, AC006441, AC002565,Y07848, AC005531, AC005527,AC004991, AP000155, AC005871,AL049758, AL049761, AC005736, Z95114,AC005529, AL008631, AL031680,AL133353, AC002472, AC009509,AP000116, AC012384, AC007325,AL022322, AL139054, AP000501,AC000035, AP000704, AC005972,AP000210, AC004815, AC002477,AC005484, AC012085, AC011311,AL031659, AC002430, AC005300,AC005229, AL050318, AC004895,AL008731, AC006241, AC005089,Z95115, AC005243, AC007384,AC007242, AC005520, AC002492,AC016831, AC002312, AC003664,AC020663, AC002091, AC005088,AL049712, Z73359, AC002381,AC005876, AC005480, AC005837,AF165926, AL135744, AC005952,AC004263, AC005829, AC004494,AC005412, AL031657, AL022238,AP000030, AC005697, AL049872,AC004699, AC002300, Z94056,AL022315, AC005755, AP000132,AC008040, AC004883, AL020997,AC005670, AL022163, AC003684,Z98047, AC007546, AC007065,AC005231, AC005220, AL049874,AC004859, Z85986, AC016025, AF196969,Z86090, AL096768, AP000014, AL031587,AC004552, AC000026, AC005874,AF134471, AC005901, AF111168,AL031282, AC004921, AC005399,AL121658, AL049709, AL031848,AC004167, AL030996, AF064861,AL034420, AL035086, AL023877,AF111167, AC004659, AL132777,AC006160, AC003962, Z98884,AL022165, AC002350, AL031431,AC004884, AC006974, AL133243,AC005037, Z95331, AC005859,AL031295, AC007055, U91322,AC000003, AL096791, Z82189,AC004149, AC005667, AF001550,AF207550, AL035423, AL031591,AC008928, AC009946, AL121603,AC004383, AL009172, U91321,AC006065, AC008018, AC006285,AL096701, AC004812, AC007130,AC004231, AL133448, AF117829,AC007435, AC007227, AL121655,AC003043, AL022476, AC005011,AC005031, AC005722, AC005808,AL035555, AC006275, AL049829,AL078477, AL049538, AC005703,AC006958, AC007406, AC007631,Z95113, AC008179, AC005592,AC005295, U52112, AC002310,AC004675, AC005996, AC006013,AF205588, AF003626, AC002369,AC007308, AL031585, AC006377,AC007880, AP000694, AC007948,AC007563, AC007216, U52111,AC004865, AC004150, AC004084,AC005216, AL135783, AF088219,AC012330, Z83826, AL031588,AC004020, AC005377, AC005206,AP000088, AC002059, AC005358,AL031280, AL050341, Z97056, AC005754,AC004773, AL034406, AP000065, andAC000379.HAMFL822739100741-48915-503T78839, and AB029015.HBXCM3827411745331-216115-2175AI752485, AI804792, AI439106,AI971133, AI991958, AI752484,AI432296, AI478420, AW082819,AI912373, R89026, AI554161, AA894797,AI752414, H13307, AI249165, R61527,N62403, R89727, N47856, AI689339,R61583, AI368569, AI984780, H44175,AA219502, T32963, AI802627, AI752415,AW295386, AA985168, H06745, R40750,M79099, AA203312, R00511, A91842,A91846, A91844, and A91848.HLHCR162759101231-379015-3804AA402528, AI379350, AA716107,AI123557, AI127175, AA234106,AA234698, AI039768, N77999, AI580137,AA424560, AA419490, AI334141, R71349,AI224976, AI417798, AI080508, N58410,AI818475, AA424657, N91089, AA399612,AI144265, AA399137, AI498363,AA410986, AA235306, AA399148,AW292497, AI249102, W86869,AA115407, H81257, AI077499, H00194,H45499, R01206, AW104245, AA234880,AA367417, AA852175, F05822,AA853077, AA852176, R82875, H81245,AA298421, R71350, R22096, H81585,AI985171, T49265, H81591, AA115408,R64037, T54283, AI492930, AI492932,R82876, R81695, R33935, R34138,R25630, H81595, R32025, R81696,T49264, R95688, R01319, R22040,Z21579, R33043, Z38741, AA514393,T97662, T34993, AA705441, D79105,R32078, H45500, AA707178, R30943,H81576, H81250, AI032721, AL079279,and I76197.HE6GF0227611538831-59915-613HOUFT3627711626801-184215-1856AI190165, AI979249, AI917302,AI806483, AI633819, AI624750,AI147946, AI471728, AW196791,AI471611, AI985423, AA609421,AA256164, AA705338, N22327,AA811162, N75202, AW236751,AI922484, H79904, AA057615, H79810,AW362445, AW407702, AA542823,AF162130, AF161181, and AC005084.HAGGF8427811974601-406415-4078Z99408, AI829910, AI004608, AI263188,AI262955, AA777512, AI625727, Z99409,AI436555, AI494561, AI085959,AI540110, AA481595, AA521313,AA283109, AL135568, AA988090,AA810697, AA233122, AI933149,AI754128, AA010623, AA806759,AI818869, AA809487, AA766911,AA316038, AA010624, AI750702,AA235130, W88865, AW296786,AI537382, AI432148, AA837093,AW385431, AW190812, AI033124,AI002712, W88760, AA283596,AA234956, AW295662, AA788729,N44843, AA777583, AA554176,AA889979, AA446963, AA448956,N55182, AI374973, AA447669, AI690467,AA283023, AA233206, AI073663, N33311,W27009, AA490727, R44959, AW135448,H12918, AA683306, AW021785, N93004,AA491218, W05278, AI393402, R37684,W30935, H54295, W45465, AA865595,AA765107, AA614573, AA767313,AA009487, H17018, AA767430,AW022593, R13824, AA736921,AA282705, H54372, AL120285, H67974,F00924, W56321, AA447819, W02795,AI124777, AA665370, AA687964, F00219,R20172, AW239099, N77241, H17017,AW262494, AA449684, AA481529,T39306, AI810957, W05327, AI750703,H87993, AW263323, F02227, F05900,Z33528, N75469, AA782796, AW262536,N75534, H05503, N37017, W56241,H54503, F02057, F05985, T40514,AA009901, F05811, AA322373,AA476641, R00735, R58669, H67654,AA732599, R00734, AA766461, AI381341,AW130224, R08857, AA330053,AA400645, AA730271, R08964, N88052,T19108, AI432714, AA424457, AI051190,W21355, AW148465, AF071569, D14906,U73504, AJ252239, J05072, X63615,L07044, AC004168, AC004056, U66064,X77192, X75774, X77193, X77195,X77194, L13406, L13407, L13408,AF059029, U50361, U73738, AF067728,AC006115, AC006112, AF107018,Y00093, AF179633, AC005156,AL133088, AW627616, AND AW630893.HTTKP0727911190311-59815-612AI640500, AA035703, AF130247, andAF165138.HE9SE6228011710141-70615-720AI765247, AW021430, AI822051,AI822104, AA010459, N70537, AL133567,and AB018312.HUJAD2428111626741-173215-1746AI923935, AI336906, AI333385,AI312076, AW204515, and AI203953.HWLFG7528212281231-202415-2038AI356559, AW163067, AI937030,AI652337, AW028706, AW157098,AW028808, AA004795, AA443325,AW005140, AW173645, R60229,AA442531, AI274924, AI810652,AI924004, AI572794, AI336556,AI672253, AI147260, AI872258,AI347103, AA467751, AA724594,AI280850, R52646, AA536110, H16834,AW450707, AW444512, AI376913,AA468349, AI807962, AA927875, R42625,AA609873, W28566, AI918962,AA578362, AA578062, R17389, C18386,R15375, AI016851, R60462, H16941,AI423739, AA467933, AA740299,AA025666, R42116, AA978110, AI423740,AW117517, AI886594, AA443338,AI857296, AW071349, AL048871,AL047763, AI702406, AI250293,AW117882, AI702433, AW195957,AI439087, AL119791, AI568870,AI433976, AI499463, AI538716,AI633419, AI349933, AI678302,AW274192, AI868831, AI498579,AI613017, AW162071, AI699857,AW071417, AW235035, AI540832,AI863014, AI628205, AW169653,AI249257, AW238730, AI349004,AI690835, AI224992, AI349772,AW301409, AW103371, AI866608,AL120736, AI564719, AL135661,AI866002, AI440426, AI590128,AI620284, AI275175, AI281779,AI433157, AL036146, AL036396,AI340582, AI521012, AI500077,AL040243, AI567351, AW074993,AI349645, AI500553, AW268253,AL045500, AI312152, AI345735,AI349937, AW089572, AI366549,AW068845, AI568854, AI597918,AI673256, AL038605, AI497733,AL047042, AI969601, AW148320,AI800453, AI800433, AI636456,AI635461, AL121270, AL036802,AI800411, AI682841, AL119049,AI610645, AI282655, AI597750,AI921379, AI758437, AI866887,AI432229, AI625079, AI690751,AI343112, AL036274, AI349256,AI567632, AI349614, AI570384,AI281773, AI434281, AI686926,AI687728, AI282903, AW303152,AL043326, AI207510, AI697137,AA640779, AI445432, AL121365,AA613907, AA572758, AL036759,AI679724, AI499393, AL038779,AI539771, AI811863, AL036980,AW074869, AW085799, AI475371,AI620868, AI815383, AL119748,AI569616, AI610307, AI436456,AI064830, AI631107, AI969567,AI608667, AI818683, AI687376,AI285735, AI439745, AI348897,AL046849, AL040169, AW002342,AL038778, AI920968, AI475134,AI469532, AI872711, AW301300,AI889203, AW168384, AI580984,AL047041, AI580190, AI873731,AI269696, AI680113, AI889839,AI671679, AI874109, AI753683,AI610756, AL120854, AI934036,AI609592, AI583316, AW166645,AW167776, AW080838, AI343059,AI446606, AI799199, AI499131,AW196141, AI255071, AI866780,AI919058, AI539153, AI687415,AI818206, AI345860, AI952114,AI687375, AL042753, AW087445,AI149592, AI906328, AI907070,AI804585, AI492540, AI469811,AI636445, AI345111, AI500659,AA508692, AI349226, AL080060, I48979,AL133016, Y11587, I89947, S68736,AR059958, AF090901, AF113691,AL110221, AL137527, AF113689, A93016,AF113676, S78214, AL049452, AL050393,AF090934, AF104032, AL133640,AF113013, AF118064, AF118070,AF078844, AF090900, AF090903,AF125949, AF113694, AF090943,AL049938, L31396, AJ242859, AL050146,AL133606, L31397, AL117460, AF113690,AL050149, AF106862, AL110196, X84990,AL117457, I89931, AF090896, A08916,AL050116, AL050108, AL050138,U42766, AL049466, AL133075,AB019565, AL049314, E03348, AL122050,AF113677, AF113019, AL080137, A08913,AF017152, AL080124, AL137283,AL133557, AL122093, AL050277,AL096744, I48978, U91329, AR011880,AL133093, AL122121, AF158248, X63574,AL049430, AF113699, AL137557,AL133565, Y11254, AL133080, X82434,Y16645, AJ000937, E07361, AL122123,AF091084, AF111851, AL137459,AF177401, AL117394, AC002467,AF146568, AL117583, AF125948,AF097996, AL110225, AL021393,AF079765, I49625, S61953, I33392,AF017437, A08912, U80742, U00763,AL050024, AL117585, AL049382, E07108,AL133560, AL137550, A65341, AF000145,A08910, AF095901, AL049464, AJ238278,E02349, AL049300, AL117435, AF067728,X70685, X72889, Z82022, AL122098,AF183393, AC006222, AC006371,AC007390, AF118094, U62317, A58524,A58523, AL137271, U72620, A08909,I03321, AC002464, AL137463, AL096776,I09360, AL137538, AL022147, AL133113,AC004200, AL122110, AL049283,A03736, AL080127, AC007172, A77033,A77035, E05822, A12297, AP000344,U35846, AF087943, AL137648, AL031281,X96540, AC010077, AF061943, A08911,X65873, U95738, AC004690, AC009233,AL110197, AC005291, X93495,AC007458, AC005057, AC006336,AF177767, L13297, AC004093, U67958,AL080159, AJ012755, U95739, AC005048,I42402, X98834, AL022723, AL049557,AF111112, A21622, AL031732, AL078630,AC006112, AC004686, U66059, A21625,Y09972, AL133072, AL137521, AF132341,AC007298, AL080154, M30514,AC003977, AF093117, and I17767.HT3BG1228310924171-105015-1064AI028599, AA774434, AA905685,AI393349, AA991810, D80022, D80268,D58283, D80043, D80166, C14389,D50995, AW360811, AW178893, C14331,T03269, AW177440, D59927, D50979,D57483, D59859, D80195, D51423,D59619, D80210, D51799, D80391,D59275, D80240, D80253, D59787,D80227, D80522, D59502, AA305409,D80164, D59467, AW178906, D81030,D81026, D51060, AW377671, AW375405,C14429, D80188, D80212, D80193,D80196, D80219, AW179328, D80247,D80038, D80269, D80366, D59889,C15076, D80024, D80439, D80248,AA514186, D80045, AW366296, D59610,AW360844, AW360817, D80378,AW375406, AW378534, AW352158,AW179332, AW377672, AW179023,AW178905, AW378532, D51022,AW378528, AW178762, C14014,AA305578, T48593, D80133, AW176467,D80302, AW369651, AW178775,AW177501, D80241, AW177511,AW177731, AW352120, AA514188,AW352171, AW377676, D51759,AW352170, AW178907, AW179019,AW179024, D51250, AW178983,AW179020, D80251, AW360834,AW360841, D51103, AW178908,AW177505, C75259, AW352117,AW178909, AW177456, AW179329,D80157, AW178980, AW178914,AW178774, AW367967, AW177733,AW178754, AW179018, AW367950,AW179004, AW178986, AI905856,AW378543, AI910186, AW378540,AW177728, AW378525, AW352163,D80134, AW178911, D80132, AW178781,C06015, C05695, AW177734, AW378533,D51097, D45260, D58253, T11417,C03092, AW177723, H67866, H67854,D80064, AA809122, F13647, AA285331,C14227, AW378542, D59317, AW378539,AI525923, D80014, AA514184, AI525917,D59474, T03116, AI535686, AW360855,D58246, D81111, D80258, D59503,C14973, D80314, AI525920, AI525913,AI525227, D58101, AI525242, D59551,AI557751, AI557774, AI525912,AI525235, AI535959, AI525925,AB028951, AL122055, A78862, A62300,AJ132110, AR018138, X67155, A67220,D89785, D34614, A84916, A62298,AF058696, Y17188, D26022, A25909,AB028859, AR008278, I50126, I50132,I50128, I50133, A94995, Y12724,AR066488, A82595, D88547, AR008443,AR016514, AR060138, A45456, Y09669,A26615, AR052274, AR060385,AR066487, X82626, AB002449, A30438,A43192, A43190, AR038669, AR025207,D50010, Y17187, A70867, A63261,AR016691, AR016690, U46128,AR008408, AR062872, AR066490,D13509, AR060133, AR054175, I18367,A64136, A68321, I14842, AB012117,X68127, AR008277, AR008281,AR050680, AF123263, AR032065,U79457, and AR008382.HTLJC712849229231-173815-1752AL039539, AL045443, AI336919,AA406128, AA405229, AL042307,AA431504, AA311249, AW086440,AA813520, AI240644, AA897733,AW268487, AA782009, AW172455,AI301209, AI014598, AA969918,AL041043, AA431178, AL039540,AA973051, AI221826, AL133030,AC009516, AP000552, AP000556,AP000557, AL117509, AC023490,AC023490, AC009516, AC009516,AC009516, AC018751, AC018751,AC018751, AC007957, and AC007957.HCOMM0528511947011-162415-1638AI681802, AA534542, AL044632,AI912061, AI912050, AI298280,AI271683, AA587766, AA143726,AA565889, AI052230, AI983603,AI148467, AI028441, AI369272,AA079779, N93876, AI950520,AW024410, AI933105, AW138648,AI244430, AA583571, AA436795,AA158795, AA079751, AA595313,AA947417, R27340, AI373637, AI869920,AI244362, AW182724, AW381284,AI991003, AI250467, AI252203,AI252717, AW086414, AI252210,AI611516, AI345868, AI348932,AI308005, AI345138, AI310659,AI250541, AI311504, AW303046,AW301779, AI311180, AI250964,AI335365, AI250009, AI348928,AW086115, AI862506, AI305732,AI335130, AI311179, AI269522,AI305978, AI371584, AW301495,AI251162, AI254155, AI310038,AI311043, AI251045, AI335320,AI590436, AA158794, AW301422,AI224292, AI310178, AI284537,AI252538, D31472, AI053635, AI540612,AI685886, AI862262, AW268255,AW371409, W40369, AW071370,AI886550, AW371414, AI955911,AI024888, AA143739, AI254135,AI246935, AW177721, T35070, C00345,AW371416, AA902858, AW369434,AA581215, R34522, AA835966,AA083230, AW078797, AI475833, andR35706.HSLJE5428612299281-201515-2029AA224020, AI906305, AA325180,AI282479, AW237905, AL038842,AW410354, AW069227, AW327624,AW340905, AA904211, AI634187,AW162288, AW023111, AI696793,AA584489, AI278972, AA558404,AI340641, AW275432, T05834,AA640430, AI635028, AI457313,AW272294, AA225406, AI076236,AA916430, AI340832, AI284640,AI358712, AL046409, AA640410,AI891080, AI280266, AL039041,AL039042, AW276827, AI753365,AI609972, H73550, AW021917, F13749,AA742815, AI192440, H71738,AW264901, AA829065, AA557982,AW271917, AW243793, AI362442,AA704393, AW303196, AA133472,AW274349, AI560085, AI755057,AA503018, AL079869, AI587583,AA747757, AW243831, AI587565,AI890324, AA832175, T47138, AI694784,AI246796, AI628859, AI570261,AW105346, AI250552, AA557945,AW301350, AI284543, AW408767,AW157731, AW193265, AA828619,AI620992, AW341903, AI859438,W63553, AA565911, AA182731,AW084445, AW068596, AA639155,AW029038, AI254770, AI245693,AI909199, AI570943, AA169245,AW328446, AI457597, AI251284,AI251034, AI251203, AI223626,AA502532, AI310464, AI431303,AA659832, AA772906, AA604831,AI251104, AI635196, H71678, AA653823,AA469327, AW078909, AF116548,AF116546, AF116547, X94152, AJ132661,E13557, M64755, AF116545, AF115343,AF115344, AL049779, AC005722,AP000692, AC007363, AL035462,AC007899, AC007707, AL049760,AL023553, AP000228, AP000140,AC007546, AC007201, AF129756,AP000555, AP000088, AC004552,AP001052, AC005529, AC006441, Z84487,AC005740, AC006285, AF107045,AC000118, AF031076, AL022718,AC009044, AC005233, AL031984,AL049548, AC005015, AL133485,AC005081, AC006130, X96421,AC007285, AC005837, AL008726,AC005881, AL031311, AC010205,AL049869, AC005409, AC005921,AC004778, AC005284, AF031078,AC003104, AF134726, Z85996, AC006965,AC005261, AC005363, AC009247,AF003626, AL109799, Z93023, Z95115,Z94056, AC005274, AC005527,AC005520, AC004534, AC002037,AC005049, AC005300, AF047825, Z93244,AC005899, AL049776, AC007253,AC005200, AL049576, AL035361,AL031432, AC003957, AC005598,AL031427, AC005620, AC004841,AC006312, AP000553, AF190465,AL031767, AC005004, Y18000,AC000068, AL096701, AL031286, Z93930,AC005775, AC005666, AC004922,AL122020, AC005288, AC005345,AL022320, M87889, AC004859,AL031681, AC009516, AF001549,AC006125, AC007327, Z93096,AC005011, AC005839, AC004525,AC007868, AL080243, AC004776,AL031774, Z98051, AL121658, L35532,AL031729, AF077058, AC005763,AC006040, AF030876, AC005694,AC004819, L78810, AL022336, Z82201,AL035461, X53550, AC005874,AF134471, AC006241, AC002070,U96629, AL109798, AF196969,AC007541, AL049743, AC003683,AC004598, AP001053, AC007649,AL031587, AC004743, AJ246003,AL031230, Z81314, AC005988,AC005189, AC007510, AC003665,AC003070, AF015160, AC008064,AC002394, AC007386, AP000065, Z97053,AC006101, AL009181, AC000353,AL022165, AC007406, AL080317,AC005962, AL021397, AC006064,AL049748, AL035530, AL109753,AC004659, AC005544, AC006468,AC006006, D83989, AC007435,AL049766, AC005229, AC005378,AL021918, AL023582, AC003035,AP000213, AL049823, AC005476,AC004211, AP000313, AP000135, X75335,AC004757, Z83846, I34294, AC002306,AC002477, AC002347, U57008,AP000303, AL035413, AL024508,AC002350, AC002365, AL008725,AC004760, AP000031, AC003080,AC004752, AB001523, AP000117,AC006372, AC004884, AC004668,Z95152, AL121603, and AC003010.HTGED072879274111-48915-503AA565765, AA565751, AI765474,AA085796, AA988365, AI380028,AI243840, AA976164, F05393, H13329,Z78407, T78216, D82559, AI868591,AL121852, AL117355, AB020724,AF092139, and AF110646.HOFNH302889283651-36215-376AF186380, and AF127138.HWNCY0528911797671-96815-982W40569, AW025860, D63226, AA334307,and AC006928.HDPDA472909291931-103615-1050AW402583, AL049683, and AL023653.HWMEV632919311541-44015-454D13626, and AC078816.HCFAT2529210528571-62615-640AI287912, AI349658, AI792640,AF096300, AB014587, AC005035,AL137755, and U88984.HHEQV3929311654201-87315-887AA355773, and AA355926.HHFJH7929412281951-230315-2317AW370568, AI684034, AI567533,AW361753, R17150, AA554360,AW245518, AW245843, AL046054,AI185853, AI814502, AI830917,AW027953, AI423947, AI761370, R77851,AW028779, AA338439, R59685, R25627,AI244276, AA587224, AI342419,AI377795, AW188290, AI361824,AI249713, AW008444, AW173497,Z45900, AA339575, AA232181, AI206821,AA142987, H11878, AA470821, AI933390,AI636179, AA317977, AA363355,AA373480, U79287, AC006942, andAL050131.HUCOW1729511551901-169315-1707AI245305, AA307147, AW139688,W52616, AA416742, AA102287, R60274,AI085546, AI167698, R60782, H17001,AI400152, H15571, D78731, AI264789,H17000, H15631, AA192581, C03464,AI571949, AA872017, AI694004,AA486536, AA112946, AA533831,AA872016, and AI357709.HFKIT0629610780921-155415-1568AA747315, AI392901, AI697980,AW295170, AA148983, AI283647,AA148982, AI393927, AA687243, R70140,H44117, H44044, AI783873, AI918219,R70141, AI741034, D80164, D80227,D51799, C14331, D80022, D59467,D80195, D59275, D59787, D59502,D80269, D59610, D58283, C15076,D59859, D81030, D80166, D51423,D59619, D80210, D80391, D80240,D80253, D80043, D80378, C14429,D80038, D80212, D50979, D80193,D80196, D80188, D80024, D80219,D59927, D80241, D57483, D50995,D80366, D59889, D51060, AA305409,AA305578, C14389, D80045, C14014,T03269, AW178893, C75259, AW177440,D51022, AW378532, D81026, D52291,AW352158, AA514188, D80134,AW179328, AW178775, D51250, D80251,D59695, AA514186, AW369651, C14407,D80522, F13647, D80248, D80133,D58253, D80168, AW178762, AW177501,AW177511, C14227, AW360811,AW378540, D81111, C14298, AI557751,AI910186, D80268, D80064, AW176467,C05695, AI905856, AW352117,AW375405, D80439, AW377671, D80247,D80302, Z21582, AW360834, AW366296,AW360844, AW360817, AW375406,AW378534, AW179332, AW377672,AW179023, AW178905, AW178906,D80132, D51103, AW352171, D51097,AW377676, AW352170, AW177731,AW178907, AW179019, AW179024,D59373, AA285331, D80157, AW177505,AW360841, AW179020, AW178909,AW177456, AW352120, AW179329,D51759, AW178980, AW177733,AW378528, AW178908, AW178754,AW179018, T11417, AA809122, C06015,AW177728, AW179004, AW179012,AW367967, AW178914, AW178774,AW378543, AW378525, T48593,AW178983, AW177722, AW352163,AW352174, AW179009, AW178781,AW178911, D59653, AI535686, D45260,D59503, AW177508, D58246, AI525923,AW367950, D59627, H67854, C03092,H67866, D58101, C14344, AW177723,AI557774, T03116, AW177497, D59317,AW178986, D80258, AI525920, D45273,AI525917, AI535850, D80014, C14973,AW378533, D51213, AI525227, D80228,D60214, D51221, D59551, D59474,AW177734, D60010, AA514184, N66429,C14957, C14046, AI525242, AI525235,Z33452, T03048, AI525925, AI525912,AI525237, AW378542, AW378539,AI525215, C16955, AF075060, A62300,A62298, A84916, AJ132110, AR018138,Y17188, A78862, X67155, A25909,D26022, A67220, D89785, D34614,AF058696, AB028859, D88547, X82626,AR008278, AR025207, I82448, A82595,Y12724, AR060385, AR008443,AB002449, AB012117, A94995, X68127,AR016808, I50126, I50132, I50128,I50133, A85396, AR066482, A45456,A44171, A85477, AR066488, I19525,AR016514, A86792, AR060138, A26615,AR052274, X93549, I14842, A30438,Y09669, A43192, A43190, AR038669,AR054175, AR066487, AR062872,Y17187, U46128, D50010, AR066490,AR008277, AR008281, A63261, I18367,Z82022, AR016691, AR016690,AR008408, D88507, A70867, U79457,I79511, D13509, A64136, A68321,AR060133, AF135125, AF123263,AR032065, AR060382, and AR008382.HDTBY8829711041591-52315-537AA868305, AI700890, AA789239,AI803004, AI694352, AA043382, F08474,R21498, AF112183, AF112184, andAC005354.HWLHS8229810822681-133515-1349AI939391, AW401390, AI202873,AI869600, AW411117, AI142585, Z99396,H50446, AL119457, AL119324,AL036418, AL038837, AL037051,AL036725, AW392670, AA631969,AL119443, AL039074, AW384394,AL036924, AW372827, AW363220,AL036858, AL119497, AL119355,AL037094, AL038509, AL039564,AL119483, AL039085, AL119335,U46351, AL119319, AL036196,AL039156, AL039108, AL039109,AL039128, AL036190, U46349,AL119341, AL119484, AL119363,AL119391, AL042544, AL119399,AL036767, AL039659, U46350,AL119418, AL037082, AL119522,AL037526, U46341, AL134902,AL037639, AL119396, AL038531,AL042984, AL119496, U46347,AL037085, AL119444, AL036268,AL134533, AL039625, AL039648,AL037077, AL045337, AL119401,AL036238, AL037205, AL119439,U46346, AL134536, AL038447,AL042450, AL042909, AL043033,AL039386, AL039678, AL039629,AL119464, AL134538, AL042614,AL036733, AL039423, AL038520,AL042965, AL042975, AL039150,AL134525, U46345, AL040992,AL043029, AL042551, AL037027,AL036998, AL037615, AL038851,AL043019, AL037178, AL134542,AL043011, AL042542, AL036765,AL037726, AL036719, AL036191,AL039410, AL043003, AL036679,AL036774, AC005581, AC005781,AR060234, AR066494, A81671,AR023813, AR069079, AR064707,AB026436, and AR054110.HDPNC9629910816291-72015-734AA256100, and AB023182.HCE5I7830011978991-314915-3163AA988953, N63548, AA663569, H15532,H38453, AA976078, N94873, H41422,H46380, R88023, N51089, H41541,H48172, AI423964, H38492, AI421373,R18083, AI208504, AI003013, R90757,R88479, H11023, R40487, R42977,H85702, AW207073, R87980, AA338374,H84794, H15589, AA325807, H10745,AA319224, AA338373, W28283, R13057,AA325143, N54215, AB023202, D13613,AC004551, D29965, U12571, andAC004465.HISDS6230111596251-88815-902AA126105, AA306119, W27339, D80253,D80366, D80043, D58283, D80188,D59502, D80166, D80195, D51423,D59619, D57483, D80210, D51799,D80240, D59859, D80391, D80212,D80219, D80227, D59889, D80196,D51060, D81030, D59927, D59610,D80269, D80038, D80022, D80024,D80193, D59275, C14389, D50979,D59787, D50995, D80241, C14014,C75259, D80045, C14429, D80164,D80378, D59467, T03269, C14331,C15076, AA305409, D80251, AW178893,D80134, AW177440, D51022, D80949,D51079, D58253, AW178775, AW179328,AA305578, AI905856, AW378532,D80248, D51097, D80522, AW352158,D81026, D59695, AA514186, AA514188,AW177501, AW177511, AW178762,D80268, AW360811, D80133, R58551,AW352117, AW375405, AW366296,AW377671, AW360844, AW360817,AW375406, AW378534, AW179332,AW377672, AW179023, AW178905,D80302, D80439, D80247, D80132,AW352171, AW377676, AW352172,AW178906, AW352170, AW177731,AW178907, W27700, AW179019,AW179024, AW177505, AW179020,AW178909, AW177456, AW179220,AW179329, AW178980, AW177733,AW378528, AW178908, AW178754,AW179018, AW179004, AW178914,AW367967, C14975, AW352174,AA033512, AI535850, AB007884,AJ250425, AJ132110, A62298, A84916,A62300, AR018138, Y17188, X67155,A67220, D89785, A78862, D26022,A25909, D34614, D88547, AR008278,AR025207, AF058696, X82626,AB028859, AB012117, A85396,AR066482, A44171, A85477, I19525,A86792, X93549, Y12724, AF135125,A94995, D88507, AR008443, U87250,AB033111, AR064240, I18367, AR066490,U87247, U79457, and X93535.HDQDV6930212125661-255815-2572N63562, AI905612, N75655, N94726,AA297704, N64807, AI919048, AI567676,AI884404, AI797998, AI570067,AA765899, W27084, AW162314,AA632355, AI174827, AI281622,AW021674, AW085626, H86399,AA196994, AI745666, AI888050,AW029626, AA527633, AI150934,AW303052, AI065031, AI003391,AI049845, AW410844, AL121039,AW192930, AL138262, AI702049,AI307563, AI445699, AW162332, H47461,AI434103, T03928, AA015948,AW265468, AW328185, AA828840,AI538404, AA610644, W02419, AI547110,AI828721, AI370470, AI890857,AI744259, AI302350, AA847341,AI268465, AI003086, AI733523,AI064968, AW020612, AL118628,AW148821, H81270, AA568433,AI889177, AI857834, AI312614,AI280566, AA557945, AW149241,AI624191, AW327673, N49298, M77888,AA846046, AI744963, AW239465,AI311796, AI003068, AL044966,AA748071, AI791659, AI590404, N55076,H53109, AA947352, AA814719, AI801563,AA525953, AA199582, AA598608,AA280886, AI926656, AW439224,AI167715, AI207534, AA152398,AA167656, AW070901, AA133568,AI446574, AI905408, AI815770,AI934664, AA969564, AA846014,AI344906, AI318548, AA601712,AI819419, H48017, AW028376,AW419201, AI683079, AI376687,AA676462, AA313025, AA935827,AI969090, AI342863, AA715848,AA604149, AI754926, AA112864,AA018258, AA315052, AI350189,AI049999, AW152451, AI076729,AA774223, AA487053, AI800706, H62123,AI221027, AA669165, AA668727,AI039257, AI860423, AA601718,AA421536, AA602458, AA084439,AI620666, AI038990, AW131394,AA493245, AA526542, AL044701,T10218, AI872229, W45215, AW023975,AA525753, AL133768, AI921744,AI282488, AW275432, N53783, AI274480,AA729004, AI679759, AF169035,AF085233, Z98752, AL050338, AC005701,AC002375, AL133448, AC007537,AF196779, AC005519, AC004808,AC005822, AC004685, AC006079,AC004765, AC008044, U85195,AF176815, AE000658, AC006581,AC003665, AL049694, AP000076,AF207550, AC005089, AF001549,AC004033, AC002036, AC007731,AC005500, AC004263, AC004531,AC002558, AF109907, AF165926,AC006006, Z75744, U91327, AC003982,AP000692, AC005175, AC006511,AP001068, AL050321, AC005412,AC002310, AC005696, Z83844, AF024533,AJ003147, AC004019, AC004890, Z98941,AC002996, AC005632, Z84474,AL033527, AC005484, AL049538,AL049776, AC002472, AL022238,AF107885, AL021707, AC005399,AC006539, AL031680, AL109852,AC004997, AF104670, AP000030,AC012627, AC009247, AC009784,AC004233, AC004643, AC005920,AC002094, U93163, AL021579,AC005585, AC006597, AC006455,AC000353, AR036572, U91328,AC007036, AC005261, AC004771,AC005821, AC002477, AC005003,Z92542, AC004491, AP000555, AC007664,AL008712, AC007050, AL035445,AC006509, AL022323, AF064861,AL031864, Z80232, AL035420, AC005529,AC002351, AL022313, AC005914,AC004673, AP000251, AL121603, Z73979,AP000503, AC006071, U73636,AC007243, AC003108, AC004950,AC004703, AC004891, AC005393,AC000070, AL022241, AL121825,AL035079, AC007934, AC000082,AC004552, AC005859, AL035551,AL030996, Z83838, AC005598, AL031311,AC005365, AL080317, AC006277,AC002991, AP000365, AP000547,AF134726, AD000092, AL022334,AC005212, AL049839, AL109952, L78810,AC007227, AC005071, AC003037,AP000141, AC007993, AC005057,AC005355, AC003957, AC001052,AL136130, AP001046, AL049562,AC004099, Z84469, AL050318, M89651,AC005736, Z84719, AC010205, AJ011930,AL096701, AC000040, AP000116,AC005535, AC006013, AC004973,AC002306, AC005409, AP000049,AL024507, AC005081, AC000025,Z82201, AL022315, AC004751, D16583,AC007911, AC004500, AF043945,AL109627, AC005527, Z84480, AL021453,AP000309, AC005755, AC004783,AC004006, AP000694, AC002476,AL034417, AP000689, AF038458,AP000311, AF045555, Z94721, AL022333,AC005064, AP000038, AP000106,AC006211, AC007066, Z79997,AC004659, AB003151, Z84492, Z84486,U95739, AL135744, AC005220,AC004821, AL008582, AC005303,AP000211, AP000133, Z81364, AC006116,AC005512, AL050348, AC006274,U22376, AC003682, AL109802,AL133355, AC002544, AC006953,AC006968, U78027, AL031281, M27835,Z93241, AP000048, and AA887783.HEMBT613039399571-44915-463N86549, AW369713, and AB002301.HRODZ7030410885541-92715-941AA167766, AA167655, AA292911,H97685, AA635138, Z41812, AA507096,R40592, T17069, and AB007941.HHERQ7930511840031-68915-703AW340333, AW407893, AI806295,AW268810, AA827664, AA829237,AA909185, AA919008, AA604425,AI539428, AA011359, D63485, AB016590,and AB016589.HCECM9030610317411-137915-1393AA463356, AA453500, AA322899,AA340682, H24259, AA603868,AA330182, R19782, and AB023227.HWHGW7230711996141-162515-1639AA961509, AF190823, AF109388,AF190822, AF109387, AF190825,AF053328, AF053327, AF190824, U14414,AF053329, Y10473, Y10475, Y10474,AF020759, AF064549, AF190826,AF020756, and Y09910.HPCRV8430812198901-86315-877AA307070, D79997, L76158, and X95351.HNSAA283099469881-154415-1558AA713959, AI564093, AA768779,AA825697, AA808021, AA808149,AI401490, AW181992, AW444640,AI018159, AF146277, and AF077003.HLWAR773109474841-127515-1289AA449919, AA449920, and AF119815.HTTJW4931111274771-148615-1500AW268880, AA199865, AW444872,AI769428, AI061340, AA707168,AI970984, AW236544, AI884812,AI479954, AI356088, AI701720,AA722812, AI989992, AA410516,AI765045, AI267987, AI298592,AA005114, AI865503, AW389168,AA360112, AI633370, AI498423, T55265,AA878382, AF118838, Y17571,AF164632, AC002450, AF164529,AF164526, AF164530, AF164528,AF164527, AC004458, AC002540, andAF164525.HWAFS1831211551931-187315-1887AW009940, AI394293, AI082857,AW207417, AA227673, N99896,AA227560, AA491391, AA361942,AI000839, W25237, AA491389, andAF156884.HFCBA4431310827621-76915-783R85476, AW293022, AA339398, D80166,D80193, D81030, D59502, C14389,D80195, C15076, D80164, D59619,D80210, D80240, D80045, D80022,D59467, D59275, D80219, D80227,D80212, D80269, D58283, D59859,D80391, C14331, D59787, D51423,D51799, D80253, D80038, D80043,D80196, D81026, D50979, D80366,D80188, D59927, AA305409, D57483,D80378, D59889, D50995, D80024,D59610, D80133, D51060, C14014,AA305578, C14429, AW178893, D80248,D80522, D80241, AA514188, D51022,T03269, AW360811, AW177440, D80302,D80251, C75259, AA514186, D80439,AW377671, AW375405, AW179328,D80268, D80247, AW378532, AW352158,AW366296, AW360844, AW360817,AW375406, AW378534, AW179332,AW377672, AW179023, AW178905,AW178775, AW178762, D59373, D80134,AW369651, AW377676, D80157,AW177501, AW177511, C05695, D51250,D51103, AW352171, AW352170, D59695,AW178906, AW177731, AW178907,AW378528, AW179019, AW179024,D80132, D51759, D52291, AW176467,D80949, AW360841, AW177505, D80168,AW179020, C06015, D80064, AW178909,AW177456, AW179329, AW178980,AW177733, AW178908, AW178754,AW179018, C14298, AW352117,AW378540, D59653, D58253, C14227,D59503, AW179004, AI905856,AW179012, AI910186, AW178914,AW378525, AW352163, C14407, T48593,AI525923, D80258, AW360834,AW178983, C03092, AW179009, D45260,AW178774, AW178911, AW378543,AW177722, AW177728, AW352120,D58246, D59627, AA809122, H67854,H67866, AW178781, AI525917, D59317,T03116, D58101, C14077, T11417,AW367950, D51097, F13647, AW178986,AI525925, AW177723, AI535686, D81111,AI525920, D80014, D59551, C14973,AI525227, C14344, AW378533, AI557774,D51221, AA514184, D59474, D60010,D51213, D60214, AI525242, AI525235,AW177734, C14957, C14046, T03048,AI525912, AI525215, AI525222, D45273,AI525237, AA285331, AW378542,AI557751, AW378539, C13958, C16955,AI525928, C05763, Z33452, T02974,Z21582, AW360855, T02868, D51053,A84916, AJ132110, A62300, A62298,AR018138, AB028859, AR008278,AF058696, AR060385, X67155, Y17188,D26022, A25909, A82595, A67220,D89785, A78862, D34614, AB002449,Y12724, D88547, A45456, A94995,I82448, X82626, I50126, I50132, I50128,I50133, AR025207, AR008443, AR016808,AR016514, AR060138, I14842, AR066488,A26615, AR052274, Y09669, A43192,A43190, AR038669, AR066487, A30438,AB012117, AR054175, Y17187, D50010,A63261, AR008408, X64588, AR008277,AR008281, A85396, AR066482, A44171,I18367, AR062872, A70867, A85477,I19525, A86792, AR016691, AR016690,U46128, X93549, I79511, A64136,A68321, D88507, I50121, I50125, D13509,AR060133, X68127, AF123263,AR032065, and AW512144.HVADT7731411803741-23615-250AI217375, AI554767, AI948424,AI693185, AL134259, and AC006356.HUFCN9131511890131-253415-2548AW401749, AW451778, AW450685,AW268855, AW152133, AI983406,AI656119, AI961994, AW448985,AI831678, AW403024, AW296492,AA195250, H09181, AI433189, AA258743,AA258380, AA866028, AI982819,AI919443, H09127, C01544, AA262275,and Z85996.HAGBX323169513511-62415-638W29095, H04905, H11833, AF100346, andAC004125.HWMIB8131710929331-160415-1618AW380440, AW299858, AW391525,H78769, H78659, H53674, AA628987,AA447173, AW204470, AA343468,AA480342, AA114131, and AF155118.HCEMU8631811564301-214515-2159W28902, AI694781, AA780441, W26632,AA861408, AI598157, R87939, AI360413,AA206559, H06785, AA496386, H14668,AA443069, AA437394, AW007070,AA351204, AI499867, AI982847, H14395,AW007079, T08889, AI933154,AA082863, AA085216, F07213, T33930,AA324000, AI366734, AI683546, R35601,T08328, R85233, AA325921, H09089,F07449, AA931914, T09198, AA297096,T07392, H14346, R40510, F02224,R13080, AA975823, R24819, H09438,H06678, F05981, F09630, F11982,AA551277, T83837, T30652, H21015,T65199, F08411, F08469, T33716, F13035,T08309, D54295, R58912, T05132,R51804, AI567931, F10628, T33189,T03545, H06677, R41862, Z19847, F04626,AI468068, AI803000, AA987266, R16332,AA365773, AI275574, AI092574, T65130,AI254357, AW183676, AA496337,R38256, H06605, AB018279, S47919,L05435, L01788, and A75140.HRDAF8331911539111-121115-1225AI825217, AA316377, AW302711,AI144081, AA488067, AI587537,AI253519, AI953573, AW339743,AW302753, AA828046, AA292719,F35011, R70326, AA601230, T96582,AI053560, AW301906, AI133727,AI973207, AA205743, AW292981,H80463, AW085744, AA297496,AA502223, AA730601, H73438,AW438539, AA262086, AW301854,AA654849, AA491864, AA640495,AA363225, AI435248, AL041325, R21949,AA904231, AI696343, AA159046,AI745457, AI754567, AI754105,AI755214, AA669132, AA652267,AA984920, N25819, AA479930, AI734052,AI625693, AF109681, AF137378,AL135747, AL035251, AL049553, Z99128,AC002431, AC003046, AP000704,AC005993, AL020993, AC005844,Z95113, L44140, AC004068, AF002223,AP000500, AC004910, AL133163, Z84489,AC003012, Y10196, AC003664, L40817,X92763, Z73359, AC004757, AL035458,AF051976, AC003044, AF196971,AL109758, AC007298, AC006559,AC000026, AL031985, AC004150,AC005668, AL022326, AC007707,AL049653, AC002059, AP000212,AP000134, Z83855, AL023285, AC005193,AP000030, AL035106, Z49250, AP000251,AC003007, AC004675, AC003084,AC007934, AC005083, Z83845, AP000402,L42103, AL049778, AL117592, AC005189,AC002457, AP000013, L48038, AL121577,AL049709, AC005608, AC005304,AL109865, Z23091, AF190465, AC005212,Z97053, AL023653, AL049538, Z93244,AL133245, AC006312, AC002470,AC010205, AC005632, AC005773,AC005409, AL109627, AF054589,AL034548, AL121655, AC002430,AC006145, AL139054, AL109977,AC007878, AC005871, AP000459,AL031602, AL078477, AL080243,AC007384, Z68756, AC006965,AC004966, AL031657, AL035089,AL035249, AF020503, AC009294,AL034429, AC001231, AC018633,AC005696, AL022721, AL031289,AC005730, AC005011, AP000155,AC004094, AC005949, AC005969,AC004673, AP000565, U95739,AL049733, AC005484, AC005015,AC002094, AL050307, AL049557,AC000003, AF087970, AC010170,AC005821, Z94801, AF088219, AC004629,AB001523, AC005393, AC004386,AC002300, U73168, AL132777,AC002418, AL022313, AL022333,AC007226, AC002407, AC006237,AC007510, Z98750, AP000355, AC010209,AP000511, Z95116, AC004953, AC007639,AC007388, AL022165, AC020663,AL022163, AL049694, AL109946,AC005138, Z84481, AL117340,AC003956, AC005105, AC005952,U73166, AB023048, and AL121652.HUVGZ8832012047191-207215-2086AI302185, AI652375, AI418298,AI936871, AI394630, AW206793,AI025217, AI983297, AI025310,AW002416, AI559159, AA593826,AW140018, AW401787, AI017077,AW239548, D54154, T03397, H42947,D57560, AW192088, R23870, AA026054,AA503315, Z40806, AW375742, F07734,AI807678, C14980, AA297059,AW338394, AI073417, AW138409,AA088799, AI356604, C15480, AI825970,AI537006, AI168175, R46685, AW078709,AA364780, AI343378, R46686,AW137324, AI015047, AA974700,H42881, AA325059, AI831833, H22172,T30407, AA897415, Z44281, F03562,R45025, F03423, F07299, R25015,AA382960, T32685, AW163027,AI815476, AA471062, T82311, AI911100,AW157059, R08769, AA047793,AA382959, and AW373232.HSCKS5532111979211-227815-2292AW183030, AA535809, AI885834,AI831611, AA534906, T15603, AI364740,H29399, N27334, AI689402, H17545,AA353936, AA903747, R81708, D31451,R33400, AI707698, N75704, AI735097,R31870, AI608711, AI357830, R87560,AA954380, AI904965, AI289077, Z78378,AL046356, AL045891, AL041862,AI432666, AI434223, AL047675,AW172723, AL119748, AW151138,AL042787, AI366900, AI815232,AI805769, AI433157, AW151136,AI539771, AL042551, AI537677,AI494201, AI866786, AI500659,AI801325, AI500523, AI436429,AI582932, AI284517, AI923989,AI500706, AI445237, AI491776,AI521560, AI889189, AI500662,AI284509, AI860003, AI889168,AI866573, AI633493, AI434256,AI888661, AI284513, AI888118,AI889147, AI371228, AI440252,AI610557, AI887499, AI433976,AI554821, AL045163, AI440263,AL042488, AI432656, AL045500,AI866465, AA483831, AL047092,AI890907, AW172745, AL040207,AI866510, AI872423, AI923046,AL045620, AW191003, AI859991,AL047163, AI275175, AI934147,AL039276, AL047422, AL079977,AL042745, AL043089, AI371251,AI499463, AI610362, AL046990,AI440239, AI628850, AI620284,AW197139, AI537273, AI371265,AI436456, AI433968, AL046926,AI963846, AI567940, AI817244,AI612913, AI285826, AI863014,AI521594, AI499512, AI889133,AI866469, AI537515, AI610429,AI539632, AI889148, AW118237,AI539847, AI828583, AL042538,AI872300, AI354998, AI567935,AI805762, AL042365, AL135012, N80094,AI400725, AI582483, AI434242,AI608936, AI866608, AL042572,AI251205, AI800138, AI610402,AI242736, AI857724, AI866820,AI627880, AI274759, AI866741,AL042440, AL042557, AI538850,AI623682, AI818578, AI696378,AI866457, AI499381, AW082113,AI636268, AI049851, AI343091,AI567993, AL135025, F27438, AL042628,AW087938, AI654276, AI269862,AL048323, AW168485, AI431909,AL041772, AL048375, AI364788,AI922577, AI886022, AI538878,AI689175, AI445165, AW082594,AI919107, AW149227, AI873644,AI687065, AL047037, AI250663,AI824557, AI560023, AI812015,AW083804, AL043321, AL038445,AI697243, AL040243, AI916419,AI499986, AI590423, AA715307,AL036214, AI801561, AI355779,AI307210, AI823670, AL042377,AI343059, AL121270, AI538885,AI612885, AA420758, AI280670,AI349933, AI432644, AI097410,AL048312, AW078680, AI521596,AL048496, AI282326, AI439995,AW023590, AI888953, AW088899,AL042744, AA911767, AI919345,AI829327, AI349598, AL048377,AI619716, AI648567, AW269097,AI345347, AI686073, AI824375,AI334930, AI950664, AL118781,AI345608, AL038565, AI554218,AA427700, AI866002, AI471361,AW079159, AL122049, I48978, AL133565,AL133077, AL137550, A08916,AL133049, AL050277, AL117432, I89947,A08913, A08910, A08909, I89931, E15569,AL096744, I49625, AF125949, I48979,AL133093, AF162270, X70685, AL117585,AL122098, AL050108, AL133104,AL050116, L30117, AL137527, AF079765,E03348, AL110196, AF113013, AF090896,AL137459, AR059958, AF017152,A08912, S61953, AL122110, AR038854,AF113689, AL050024, Y11587, AL133080,AL122050, AR000496, U39656, M30514,AL117440, X93495, E07361, AF090943,AL122101, U80742, AL080137,AL117583, AF111112, AL137521,AF026816, Y16645, AL133072, I33392,Z72491, AL050172, AJ242859, AL049464,AL080127, U96683, AL137273,AL133606, AJ012755, AL080060,AL137523, AF153205, AL133557, X65873,E02221, AF003737, AL049466, AC002467,AL137526, AL049300, AL080159,AL133640, AL137271, Y14314,AL133016, AF146568, AL133113,AL122121, AL137476, AL122123, S68736,X72889, AF113694, AL049430, AL080074,AL049314, S78214, AF113676, AL050146,AL122118, U35846, U42766, AB019565,AF113699, I42402, AR011880, AF158248,AF104032, AF119337, A93016, AF091084,A65341, AF118070, A93350, AL110221,AL110225, AL122093, AL117435,U72620, AF078844, I26207, AR038969,AF113690, AF113019, AL137556,AF090934, AF118064, AF067728, E04233,AL049382, AF111851, Z82022, AF183393,AL137538, AL117460, Z37987, Y09972,U78525, AF090901, AL050393, X63574,AL137463, AF061943, U00763, AF113677,L19437, I09360, A90832, X87582, E05822,Y11254, U67958, E02349, AL133098,X84990, AL133075, AF090900, AL117457,A45787, L31396, AL133568, AL133014,AL050138, AL133560, L31397, AF106862,A58524, A58523, AL133067, AF118094,E08631, X98834, AF097996, AL137557,AJ000937, AL137648, E07108, AF090903,A12297, Y07905, U91329, AF057300,AF057299, X96540, I03321, AL049452,AJ238278, AF177401, AL080124, X82434,AF017437, A77033, A77035, AF087943,AL137560, U58996, AF111849, AF026124,AF113691, AL050149, AF125948,AL137533, I00734, AF185576, A03736,AF008439, AL110280, AF067790,AL049283, AL137478, AF079763, A07647,E00617, E00717, E00778, U68387,AL137480, E08263, E08264, AL117394,AL080086, X62580, AF081197, AL049938,AF132676, AL133081, AF061836,AJ006417, AL137292, AF081195,AF100931, AF210052, AL133053, X92070,AF061795, AF151685, X83508, AL137283,AF051325, AL137300, X53587, E06743,AL137294, AF061573, AF032666, I68732,and AL122111.HOEET483229632901-146615-1480AI797684, AW239200, AA456267,AI478733, AI751749, AI990902,AA427646, AI379565, AI970534, W95460,AA788855, AA405402, AW068453,AW294114, AI751750, AA594137,AA947297, AW177719, AI057073,AA427487, AI341112, AA232452,AA041304, AW068711, H73236,AA041328, W95567, AW167569,AA853047, AI652166, W02069, H74164,R34003, AI341381, AW176526,AA580289, D30965, D31176, AA367502,and AR035969.HBODE5132311931491-277415-2788AW411466, AW137475, AA922772,AA974304, AW204679, H05447,AI357778, AL044000, AL138265,AI038990, AW131249, D63198,AL120269, AI951863, AI732911,AI471543, AA601355, AL048969,AW102811, AA722372, AW248523,N66012, AW055226, F28204, AI457389,AA837677, AI952885, N49425,AW188427, AI863046, AW080062,AW070703, AA618412, AL079869,AL137984, AW167374, AL046746,AI791227, AL135698, AA610271,AA631507, AA604395, AA593752,AA706495, AL119123, N80210,AW204532, AW085751, AL046457,AA603421, AL042539, AA130901,AI313166, AA223932, AW008089,AA515250, AI859742, AL038705,AA634991, AI821044, AA577719,AI028148, AI569982, AI583978,AA744826, AA121815, AA679794,Y16610, AC004491, AF134726,AC005529, AC005015, AL022316,AC005081, AC009247, AC004383,AC002310, AC004967, AC005694,AC006312, AC007225, AF111169,U47924, AP000553, AC005484,AC004821, AC005531, AC005011,AL109984, AC004638, AL009181,AC006211, AC004851, AL096791,AP000688, AC004678, AC006285,AL031311, AC004263, AC007216,AC002316, AC006120, AC005837,AC005037, AC004019, AB023049,AC000025, AC007055, AL031681,AF001548, AL049795, AC002300,AC004814, AL031005, AL031848,AL024498, AL139054, AL034549,AC005225, AL109628, AC003101,AL135744, AC003070, AC006071,AP000557, AL133448, AC007686,AC007193, AC002301, AL035249,U91326, AC006538, AC007227,AF030876, AF030453, AC004812,AC007666, AF080517, AC006111,AC005527, AF080518, AP000512,AC004765, AC005972, AF196971,AL121603, AC007993, AL096701,AC016025, AC005088, AC005488,AC005480, AC005180, AL035683,AL049569, AL121653, AF196969,AC005620, AC007371, AC005089,AF053356, Z95114, AF001549, Z98941,AC007226, AC006064, AC004707,AC005049, AC003982, AL022476,AC007057, AL035659, AC000353,AC020663, AC002365, AL031602,AL033527, AL049776, AC002126,AC016830, AF196779, AC000026, Z93241,AF031078, AC005846, AF045555,AC005500, AP000350, AC006160,AC007731, AC004150, AC005755,AC009516, AL050318, AC005736,AL031670, AC007688, AL133355,AP000117, AC004887, AC004953,AC004832, AC007707, AC006088,AL021155, AC005899, AC006441,AC005071, AL049780, AC008040,AP000555, Z99128, AC000159, AL035684,AC002059, AC005921, AL022315,AC007536, AL020997, AL034420,U95742, AC004895, AL033392,AL034379, AC002400, AC006511,Z85996, AL109839, AL080242, AC005821,Z85987, AL022165, AL049830, AL008718,AC004217, AL035413, AP000503,AC005839, AC005516, Z97054, U62317,AC002425, AC005971, AC007917,AC006241, AC005746, AC006449,AP000356, AL021154, AC002470,AC002477, Z95115, AL049843, AL031667,U78027, AC004386, AC005520,AL031276, AC005519, U80017,AL117258, AL031295, AL031283,AC005004, AC004167, AC007308,U96629, AC004883, AC005914,AL022323, AC006013, AC005778,AC005696, AL022163, Z94044,AC007676, AF111168, AC000052,AF088219, AL031577, AC006057,AC004675, AC003029, Z84466,AC005328, U95740, AC002350,AC007842, AL031427, AL031733,AL021453, AL109759, AL035086,AC002544, AC006581, AC002288,AC005800, AL022328, Z82190,AC003043, AP000692, AL132987,AL049759, AF196972, AC006121,AC004882, AL133245, AC005231,AP000045, AL049761, AC004922,AL023807, Y14768, AL034429,AL022238, AL031255, AC000003, andW26963.HHFCK093249653041-278915-2803AI218626, AI076006, AW162820,AI797880, AI922744, AI872391,AI559566, AL045117, AW161046,AW162613, AI565503, AW183962,AI857802, AA460810, AI884907,AI371131, AW248493, AI081779,AA460372, AA679085, N27884,AA581796, AA074070, AA971563,AI292006, AI922373, W76538, N93245,AI609183, AW172513, AI904299,AI682939, AA075764, AI885613,AA747871, AA449042, AA928020,AW401847, AA449757, AW268637,AW073851, AW304978, AI683858,AA568598, W74167, AI367698,AW191998, N62781, AW016535,AI902503, AA347639, AA297591,AA379280, AA568887, AA649970,AW264577, AI221886, H20460,AW387087, AW000860, AI275195,AA341002, T32918, AW162711, W25103,AI699657, R42681, AW243790,AA768740, T78554, AI279653, AI560482,AI696251, AI951374, Z45830, AA147203,AI499410, R43259, AI350354, AA732831,AW079129, AA375228, F08622,AI475009, R56337, AA379846, R17163,AW380349, AA783050, AW247402,N47545, R35508, R51077, AI474934,N79729, D61534, Z41466, AI678630,AA339343, AW367003, AA160401,Z41592, AW079321, N47546, AI252528,R58857, T16943, H55297, AL022238,AL137499, and AJ236700.HCOOZ1132511103641-132115-1335AI218626, AI076006, AI797880,AI872391, AI922744, AA449042,AI559566, AW162613, AI565503,AI857802, AW248493, AI922373,AI292006, AA679085, AI904299,AA074070, W76538, AI682939,AA928020, AI609183, AW172513,N79729, AA747871, AI885613, AA449757,AW268637, AW183962, AW304978,AA568598, AI683858, AA379280,AI371131, AI884907, N62781, H20460,AI367698, N93245, AW073851,AW191998, AA581796, AA568887,AW016535, T78554, AI081779, AI221886,AW264577, AA649970, AI350354,AI275195, AI902503, AW000860,AW162711, AI699657, AA460372,AA379846, AA147203, AI951374,AI560482, AW243790, R42681,AA375228, AI499410, AI279653,AA732831, AI475009, N47545, AI696251,R43259, D61534, AA783050, AW079129,AI474934, R51077, AA160401, Z41466,W74167, AI678630, Z41592, AW079321,N47546, AL022238, and AL137499.HDPPO3532611190321-188915-1903AI640500, AW439548, AI823872,AW297416, AA831672, AI815031,AA994323, AA741162, AA471280,AW339548, AI223999, AW235171,AI635436, AA035703, AA747998,AI371399, N67227, AA361754, AI536057,AA063573, AI357169, R33401, C01451,R33402, AA825399, AF165138, andAF130247.HLWDZ5332711575421-200115-2015AA772242, AI309977, AI268079,AW152237, AI923556, AI968056,AA394106, AI985775, AI124863, T66217,AA700659, AA398135, AA927679,W68401, AW105606, AA989587, Y08708,AA559050, W68288, H11047, R23506,AI277174, F09831, H08254, R37378,AA339671, H11135, Y08709, H08154,H46511, R50952, H47050, F12201,R13468, AA320227, W31952, Y08714,R44639, R35282, H58759, H11255,T66150, W31329, AI910241, AA772192,AA662681, N84315, Y08707, T08656,AP000546, and AC002038.HEOPL3632811979101-212215-2136AI061632, AW316850, AW367730,AA814516, AW367737, AI754746,AA777525, AW367804, AI097103,AW367782, AW367735, AW367773,N30303, AA847688, AW404639, N31888,AA779663, AW367809, AW367777,AW367785, AI923024, AA953814,AW367237, AI305265, AI624406,AA582843, N20267, AI827125, AI093328,AA911743, AI292227, AI684334,AI554543, AA290888, AW367806,AA503789, AW367741, H29119,AW338358, AW342022, R55898,AI401634, W79584, N45284, H25738,AA626619, AI024861, N31823, N42550,N33984, H17043, H41983, H99343,AI002168, AI979180, AI311586, N78814,AA564287, AA351508, AA968656,W20290, N31566, R75948, AI372938,AI277345, Z44878, AW367805, H29018,H60714, H17044, H71114, AI567783,H53271, AA599764, R60811, AI673673,T35639, AA158814, AI129803, H60713,H25974, AI423976, R61308, T30051,AA582155, AI174537, AI186537,AA705042, M85360, W79783, N29173,AA161097, R00089, N26452, Z17839,F01057, T34112, N54731, AI471185,AI023664, AW130265, W31954, H41935,T33806, R59766, AA826327, AI244584,AI202016, AI077600, N90670, AW009848,AA987544, N24540, AI432312, R09698,AA777544, R76786, AW081089, T31022,AW089365, AI468541, Z40650, AI217229,AI371184, T77157, Z28806, AA069041,AA505841, N42027, N42580, AI220752,AA346958, AI497869, AA160430, T32679,R09699, R55813, AI400851, AA298583,AI919072, T33966, AA159592,AW196132, AA743892, AI689177,W31331, T34006, AA911196, AW371882,AA301187, AW367265, AI217099,AA856916, AA879055, N29277, AI285514,H89138, AI819663, AA830044, AI687568,AW411043, AI583578, N49165,AW020381, AW151132, AW080076,AI954721, AI610426, AW198112,AI306705, AI306613, AI656270,AI628875, AI811373, AA737649,AI612885, AW189196, AA838230,AI274527, T66952, AI421662, AI249946,AL039011, AI470717, AI624529,AI318603, AW059828, AW303152,AI380329, AI225000, AI345778,AL037626, AI702527, AI627714,AI796743, AI056328, AI473652,AI280561, AW090550, AL045983,AI491904, AI799273, AI886355,AI933992, AI114703, AI678446,AW409775, AI479292, AI648699,AI250282, AI868204, AW022907, F37409,AI590043, AI500061, AW188388,AI553926, AI524654, AA579232,AI401697, AW080157, AW083484,AI815232, F26535, AA420722, AI471909,AA514684, AI370623, AI538885,AA761557, AW150762, AI540382,AW021189, AI682968, U69181,AL036980, AI860027, AW166870,AI452857, AW149849, AI628325,AI537187, AA746607, AI349957,AI804505, AF128536, AL049758,AF128535, AF139495, AF139493,AF139494, AF139492, AR030257,AL133624, AL133047, AF185614, U70981,AL137271, AL137555, U72621,AL049276, AR060156, AL133014,AJ010277, U57352, Y14634, I48978,AL080234, A07647, AF036941, AF115392,AR029580, L13297, AL122050, AL110222,AF102578, AF124728, AL137298, A58524,A58523, AF090943, AF038847, AF210052,AL050116, AF115410, AF113013,AF030165, AF111851, E12580, X53587,A07588, AJ238278, S61953, AF091084,AF120268, U42031, AF090886, AL110225,E02221, I89947, AF167995, S36676,AF097996, AL117457, I66342, AL137548,A08913, AF159148, AF044221, AL110221,AF015958, AR068466, I89931, AF090934,Y16645, AF118094, AL110224, A08910,AF081195, I49625, AR016469, A08909,AR012379, U02475, S77771, AL137665,AR050959, A18777, AR020905,AL049464, U89906, Y13350, U95114,AF125948, S68736, AL133010, AJ012755,AL133665, A70386, AF161699, U77594,U87620, AL080140, AL117394, X59414,AF043642, AF032666, I68732, AL137550,AF035161, AL137539, AF013214, A65340,AJ000937, AF118090, A93914, S82852,AF106657, AF061573, AF019298, X83508,X72889, AF113691, I00734, AF113690,AB016226, AF119336, AF176651, E00617,E00717, E00778, AL133049, AR011880,S78214, AL137463, A26498, AF067790,U92992, X82434, AF100931, AF017437,E03671, E12579, AF200464, AF183393,X52128, A91160, AL137711, AF090903,AF118558, AL050149, L31396, AL136842,AL080147, U42766, L31397, AF124435,A91162, Z97214, AL110280, U72620,I48979, AF118070, A77033, A77035,X62580, AL117460, AL122093,AL049452, X98834, AF067728, Y11254,AL050277, X80340, E07108, AL137716,AL133560, AF078844, AF031903,AL137557, Y18680, U92068, AL137560,X70685, AL110296, AJ242859, M96857,AL137281, U80919, U91329, AF057300,U88966, L24896, AL122110, AL136884,AF205861, AL035458, AL080159, Z82022,U77351, AL133557, U00763, U80742,AJ012582, X93495, S69510, A08916,E12747, AL080124, AL096728, X66862,AF070632, AL137478, AL117587,AL049382, AL133640, AL137459,AF111849, AL137554, AB007812,A93350, I09499, U73682, AL137658,AF146568, AL117435, AL133031, A03736,AF026816, A32826, A30330, A32827,A30331, and U01145.HMCFS0232911522521-143215-1446AL119990, AI459232, AI990971,AW274849, AA524531, AI703399,AW295852, AI086621, AI582316,AI887899, AI198620, AW444954,AI201159, AI761662, AA769116,AI678189, AI769819, AA908833,AW001104, AI797112, AW339272,AW151222, AI025272, AI638810,AA291544, AW083242, AA781447,AW003604, AW015232, N30310,AA831760, AI624665, AI027612,AW276610, AW168171, AA442455,AW170708, AI807572, AW080508,T62956, AI052790, AA827853, AA737447,AI969056, AA465529, H03505, R78862,AW378235, AA465115, AW205495,AA515398, AI972735, AA805386,AA436647, H03615, N56642, AI825714,AA293351, AI473377, AA404571,AI349471, AW292091, T63206,AA252574, R79349, Z38283, AA399447,AI582475, AI870404, AA252529,AA708728, AI670981, AA429843, C06324,C06235, T88738, AA429867, AA748807,T10782, AW392315, AI650622, andD20600.HDPSR1533011947521-168215-1696AW195239, AW149418, AA461376,AW005579, AI392913, AI378013,AA461199, AI860240, AA779830,AI436586, AA147800, AA576717,AI276889, AW337924, AI264931,AI203549, AA459985, AW104319,AA460078, AI377235, AI925811,AI094031, AI291778, AI612894,AA147758, AA639492, AA767986,AA037273, R61563, AA463275, H59980,AA832298, AA417605, AA628790,AW166255, AA768536, R35919,AI289261, AA157070, AA731955,AI273576, AA508841, AI360737, N47107,AI301339, AI682196, AA463188, H59937,AI208175, D20738, AI391726, W42645,R14353, AI271983, AI073411, N30324,N56657, T53535, H57854, AW001677,R35723, AI242094, AA969269, N50317,AI808646, AA514325, AI280324,AA360254, AI699261, AA551384, T77863,AA263061, AA906804, AA741518,T11446, AI091790, AA505982, AI208678,AA031793, AA214523, AA428834,T12550, T11445, AB026289, andAR044150.HNTAV7833112172311-373315-3747AI417713, AW235714, AI537274,AL042544, AL119457, AL119511,AL119399, AL043152, AL042382,AL119324, AL134524, AL079794,AI431323, AL043168, AI889189,AI521560, AI433157, AI539771,AI537677, AI500659, AI801325,AI500523, AI284517, AI500706,AI445237, AI491776, AW151138,AI500662, AI284509, AI633493,AI538885, AI866573, AI434256,AI888661, AI284513, AI888118,AI582932, AL045500, AI537273,AI815232, AI433976, AI567935,AI554821, AI889376, AI371251,AI889168, AI620284, AI275175,AW151136, AL042551, AI866510,AL042365, AI923989, AI440252,AL110306, AI499463, AI929108,AI610362, AI440239, AI927233,AI436456, AI432656, AI567940,AI817244, AL042787, AL119863,AI612913, AL041862, AI285826,AI863014, AI499512, AI889133,AI610402, AI434223, AI610429,AI963846, AL042538, AI432666,AI805769, AI889148, AI628850,AI633125, AW161579, AI866608,AI859991, AI567993, AI860003,AI364788, AL036980, AI922901,AI491710, AL042866, AL134259,AW022682, AI889147, AL047422,AW083804, AL045163, AI866457,AA572758, AI446373, AI654276,AI866786, AI612885, AA420758,AI874166, AI494201, AW089272,AW021717, AI805762, AL048496,AI866469, AL037454, AI872423,AW301300, AI349598, AW172723,AI702073, AI440263, AW269097,AI434242, AW051258, AW191003,AI436429, AI371228, AI872300,AI610557, AI242736, AI887499,AI539632, AI539847, AL045620,AL042627, AL042557, AI433037,AI500714, AI567961, AW081255,AL079741, AI538850, AI955441,AI345010, AW129106, AL042745,AI312428, AL036705, AL039086,AW020693, AI289791, AI890907,AI521594, AW087445, AA613907,AI348897, AI635492, AI670009,AI673278, AW023072, AL119836,AI269862, AI637584, AL119828,AI432644, AI340603, AL036631,AI582926, AA635382, AA259207,AI567953, AI537515, AI318280,AI537191, AA420722, AW082113,AI499131, AL119791, AL040241,AI866465, AL119748, AI274759,AI916419, AL047387, AI811344,AL046926, AW268220, AL041573,AI431307, AW073865, AI049851,AI648567, AI690946, AL022726,AC006296, AL031295, AL030998,AC007392, AC005224, AL096709,AC005723, AC008394, AC010175,AC006115, AC004062, AL009029,AL031123, AC004808, and Z83840.HFKDR1433211458421-130815-1322AI761729, AW104395, AW298361,AI073443, N40162, AW162515, AI827518,AW297353, R52045, AI342317, R71958,AF128625, AF021936, and AW517595.HDPBI303339747111-291115-2925AA714520, N78665, W15172, AL134531,AA074818, AI251157, AI311635,AA079403, AW130754, AI935943,AF083955, AC005015, AL034423,AP000030, AC002992, AC004216,AC003013, U91321, AC003684,AC002528, AL117258, AL021155,AP000045, AF053356, AL033521,AC004598, U91326, AL035072,AD000091, U82668, AC012384, L44140,AF006752, AL034350, AC006039,AC005756, AC005072, AL034429,AC002352, AC005682, AC003663,AC005049, AC007298, AC005620,AC004887, AL117694, AC005911,AC007688, AC006014, AC004797,AL031186, AL031283, AC004963, L47234,Z84466, AC004125, AC005529,AL031293, AC006276, AL034400,AC004099, AC005089, AL049871,AC004893, AL080243, AC007021,AL049712, AC007993, AC006581,AC005837, AF139813, M13792,AC005086, AL096791, AJ251973,AC002301, AC006139, AC005488,L78810, AC006115, AC004966,AC006538, Z93244, AC004834,AL049570, AC004084, AP000113,AP000251, and AC005696.HODFF8833410948751-184315-1857D80164, D59502, D80193, D80195,D59275, C15076, D80227, D58283,D80022, D80166, D81030, D59859,D51799, D59619, D80210, D80391,D80240, D59787, D51423, D80253,D80043, D80269, D50979, D80212,D80038, D80196, D80024, D80219,D80188, C14331, D59467, D57483,D59927, D80378, D80366, C14389,D59889, D50995, D80045, D59610,AA305409, C14429, D80241, D51060,T03269, C14014, AW178893, C75259,AA305578, D81026, D59695, D51022,AW179328, D81111, AW178775, D80134,AW378532, AW177440, D51250,AW352158, D80268, F13647, AA514188,AW369651, D80251, D80522, D51079,D80248, D80949, D58253, AW178762,D80168, D52291, C14227, AA514186,AI905856, AW177501, AW177511,D80133, Z21582, AW360811, C05695,C14298, AW352117, D80064, AW176467,AW375405, AW378540, C14407,AW377671, D51097, AW366296, D80302,AW360844, AW360817, AW375406,AW378534, AW179332, AW377672,AW179023, AW178905, D80132,AW360834, AA285331, D80439,AW352171, AW377676, AW178906,AW352170, AW177731, D80247,AW178907, AW179019, AW179024,D51103, AW177505, AW360841,AW179020, AW178909, AW177456,AW179329, AW178980, AW177733,AW378528, AW178908, AW178754,AW179018, AW179220, AI557751,AW179004, AW178914, AW378525,AW352174, T11417, D80157, AW177728,D59627, D51759, AW367967, AW178774,AW178911, AW378543, AW352163,D59503, D80258, D80014, C06015,AI557774, AW178983, AW352120,T03116, AW178781, T48593, D58246,C14077, D59653, AW177723, D58101,D45260, AI525923, AW178986,AW367950, C03092, AA809122, H67854,D59551, H67866, C14975, T02974,AW378533, AW378539, D51213,AW177734, AI535686, D59317, D51221,AI525917, C14973, AA514184, C14344,D45273, AI525925, AI525920, D59474,AI525227, D31458, C14046, AI525242,AI525235, T03048, AI525912, AW378542,AI525215, AI525237, C16955, C05763,Z33452, AI535850, AI535961, A84916,AJ132110, A62300, A62298, AR018138,X67155, Y17188, D26022, A25909,A67220, D89785, A78862, D34614,D88547, AF058696, X82626, AR008278,AB028859, AR025207, I82448, Y12724,A82595, AB012117, AR060385,AB002449, A85396, AR066482, A44171,A85477, A94995, X68127, I19525,A86792, X93549, AR008443, AR016808,U87250, I50133, I50126, I50132, I50128,AR066488, AR016514, AR060138,A45456, A26615, AR052274, I14842,Y09669, A43192, A43190, AR038669,AR066487, A30438, AF135125, D88507,AR066490, D50010, AR054175, I18367,Y17187, A63261, AR008277, AR008281,AR008408, AR062872, A70867,AB033111, AR016691, AR016690,U46128, D13509, AR060133, I79511,AR064240, A64136, A68321, U87247,AB023656, U79457, AF123263, X93535,and AR008382.

[0089]

6

TABLE 4

Code
Description
Tissue
Organ
Cell Line
Disease
Vector

AR022
a_Heart
a_Heart

AR023
a_Liver
a_Liver

AR024
a_mammary gland
a_mammary gland

AR025
a_Prostate
a_Prostate

AR026
a_small intestine
a_small intestine

AR027
a_Stomach
a_Stomach

AR028
Blood B cells
Blood B cells

AR029
Blood B cells activated
Blood B cells

activated

AR030
Blood B cells resting
Blood B cells

resting

AR031
Blood T cells activated
Blood T cells

activated

AR032
Blood T cells resting
Blood T cells resting

AR033
brain
brain

AR034
breast
breast

AR035
breast cancer
breast cancer

AR036
Cell Line CAOV3
Cell Line CAOV3

AR037
cell line PA-1
cell line PA-1

AR038
cell line transformed
cell line transformed

AR039
colon
colon

AR040
colon (9808co65R)
colon (9808co65R)

AR041
colon (9809co15)
colon (9809co15)

AR042
colon cancer
colon cancer

AR043
colon cancer (9808co64R)
colon cancer

(9808co64R)

AR044
colon cancer 9809co14
colon cancer

9809co14

AR045
corn clone 5
corn clone 5

AR046
corn clone 6
corn clone 6

AR047
corn clone 2
corn clone 2

AR048
corn clone 3
corn clone 3

AR049
Corn Clone 4
Corn Clone 4

AR050
Donor II B Cells 24 hrs
Donor II B Cells

24 hrs

AR051
Donor II B Cells 72 hrs
Donor II B Cells

72 hrs

AR052
Donor II B-Cells 24 hrs.
Donor II B-Cells 24

hrs.

AR053
Donor II B-Cells 72 hrs
Donor II B-Cells

72 hrs

AR054
Donor II Resting B Cells
Donor II Resting B

Cells

AR055
Heart
Heart

AR056
Human Lung (clonetech)
Human Lung

(clonetech)

AR057
Human Mammary
Human Mammary

(clontech)
(clontech)

AR058
Human Thymus
Human Thymus

(clonetech)
(clonetech)

AR059
Jurkat (unstimulated)
Jurkat

(unstimulated)

AR060
Kidney
Kidney

AR061
Liver
Liver

AR062
Liver (Clontech)
Liver (Clontech)

AR063
Lymphocytes chronic
Lymphocytes

lymphocytic leukaemia
chronic lymphocytic

leukaemia

AR064
Lymphocytes diffuse large
Lymphocytes

B cell lymphoma
diffuse large B cell

lymphoma

AR065
Lymphocytes follicular
Lymphocytes

lymphoma
follicular lymphoma

AR066
normal breast
normal breast

AR067
Normal Ovarian
Normal Ovarian

(4004901)
(4004901)

AR068
Normal Ovary 9508G045
Normal Ovary

9508G045

AR069
Normal Ovary 9701G208
Normal Ovary

9701G208

AR070
Normal Ovary 9806G005
Normal Ovary

9806G005

AR071
Ovarian Cancer
Ovarian Cancer

AR072
Ovarian Cancer
Ovarian Cancer

(9702G001)
(9702G001)

AR073
Ovarian Cancer
Ovarian Cancer

(9707G029)
(9707G029)

AR074
Ovarian Cancer
Ovarian Cancer

(9804G011)
(9804G011)

AR075
Ovarian Cancer
Ovarian Cancer

(9806G019)
(9806G019)

AR076
Ovarian Cancer
Ovarian Cancer

(9807G017)
(9807G017)

AR077
Ovarian Cancer
Ovarian Cancer

(9809G001)
(9809G001)

AR078
ovarian cancer 15799
ovarian cancer

15799

AR079
Ovarian Cancer
Ovarian Cancer

17717AID
17717AID

AR080
Ovarian Cancer
Ovarian Cancer

4004664B1
4004664B1

AR081
Ovarian Cancer
Ovarian Cancer

4005315A1
4005315A1

AR082
ovarian cancer 94127303
ovarian cancer

94127303

AR083
Ovarian Cancer 96069304
Ovarian Cancer

96069304

AR084
Ovarian Cancer 9707G029
Ovarian Cancer

9707G029

AR085
Ovarian Cancer 9807G045
Ovarian Cancer

9807G045

AR086
ovarian cancer 9809G001
ovarian cancer

9809G001

AR087
Ovarian Cancer
Ovarian Cancer

9905C032RC
9905C032RC

AR088
Ovarian cancer 9907 C00
Ovarian cancer 9907

3rd
C00 3rd

AR089
Prostate
Prostate

AR090
Prostate (clonetech)
Prostate (clonetech)

AR091
prostate cancer
prostate cancer

AR092
prostate cancer #15176
prostate cancer

#15176

AR093
prostate cancer #15509
prostate cancer

#15509

AR094
prostate cancer #15673
prostate cancer

#15673

AR095
Small Intestine (Clontech)
Small Intestine

(Clontech)

AR096
Spleen
Spleen

AR097
Thymus T cells activated
Thymus T cells

activated

AR098
Thymus T cells resting
Thymus T cells

resting

AR099
Tonsil
Tonsil

AR100
Tonsil geminal center
Tonsil geminal

centroblast
center centroblast

AR101
Tonsil germinal center B
Tonsil germinal

cell
center B cell

AR102
Tonsil lymph node
Tonsil lymph node

AR103
Tonsil memory B cell
Tonsil memory B

cell

AR104
Whole Brain
Whole Brain

AR105
Xenograft ES-2
Xenograft ES-2

AR106
Xenograft SW626
Xenograft SW626

H0004
Human Adult Spleen
Human Adult
Spleen

Uni-ZAP XR

Spleen

H0008
Whole 6 Week Old

Uni-ZAP XR

Embryo

H0009
Human Fetal Brain

Uni-ZAP XR

H0011
Human Fetal Kidney
Human Fetal Kidney
Kidney

Uni-ZAP XR

H0012
Human Fetal Kidney
Human Fetal Kidney
Kidney

Uni-ZAP XR

H0013
Human 8 Week Whole
Human 8 Week Old
Embryo

Uni-ZAP XR

Embryo
Embryo

H0014
Human Gall Bladder
Human Gall Bladder
Gall Bladder

Uni-ZAP XR

H0015
Human Gall Bladder,
Human Gall Bladder
Gall Bladder

Uni-ZAP XR

fraction II

H0022
Jurkat Cells
Jurkat T-Cell Line

Lambda

ZAP II

H0023
Human Fetal Lung

Uni-ZAP XR

H0024
Human Fetal Lung III
Human Fetal Lung
Lung

Uni-ZAP XR

H0025
Human Adult Lymph
Human Adult
Lymph Node

Lambda

Node
Lymph Node

ZAP II

H0027
Human Ovarian Cancer

disease
Uni-ZAP XR

H0028
Human Old Ovary
Human Old Ovary
Ovary

pBluescript

H0029
Human Pancreas
Human Pancreas
Pancreas

Uni-ZAP XR

H0030
Human Placenta

Uni-ZAP XR

H0031
Human Placenta
Human Placenta
Placenta

Uni-ZAP XR

H0032
Human Prostate
Human Prostate
Prostate

Uni-ZAP XR

H0036
Human Adult Small
Human Adult Small
Small Int.

Uni-ZAP XR

Intestine
Intestine

H0037
Human Adult Small
Human Adult Small
Small Int.

pBluescript

Intestine
Intestine

H0038
Human Testes
Human Testes
Testis

Uni-ZAP XR

H0039
Human Pancreas Tumor
Human Pancreas
Pancreas

disease
Uni-ZAP XR

Tumor

H0040
Human Testes Tumor
Human Testes
Testis

disease
Uni-ZAP XR

Tumor

H0041
Human Fetal Bone
Human Fetal Bone
Bone

Uni-ZAP XR

H0042
Human Adult Pulmonary
Human Adult
Lung

Uni-ZAP XR

Pulmonary

H0046
Human Endometrial
Human Endometrial
Uterus

disease
Uni-ZAP XR

Tumor
Tumor

H0048
Human Pineal Gland
Human Pineal Gland

Uni-ZAP XR

H0050
Human Fetal Heart
Human Fetal Heart
Heart

Uni-ZAP XR

H0051
Human Hippocampus
Human
Brain

Uni-ZAP XR

Hippocampus

H0052
Human Cerebellum
Human Cerebellum
Brain

Uni-ZAP XR

H0056
Human Umbilical Vein,
Human Umbilical
Umbilical

Uni-ZAP XR

Endo. remake
Vein Endothelial
vein

Cells

H0057
Human Fetal Spleen

Uni-ZAP XR

H0059
Human Uterine Cancer
Human Uterine
Uterus

disease
Lambda

Cancer

ZAP II

H0063
Human Thymus
Human Thymus
Thymus

Uni-ZAP XR

H0068
Human Skin Tumor
Human Skin Tumor
Skin

disease
Uni-ZAP XR

H0069
Human Activated T-Cells
Activated T-Cells
Blood
Cell Line

Uni-ZAP XR

H0071
Human Infant Adrenal
Human Infant
Adrenal

Uni-ZAP XR

Gland
Adrenal Gland
gland

H0075
Human Activated T-Cells
Activated T-Cells
Blood
Cell Line

Uni-ZAP XR

(II)

H0081
Human Fetal Epithelium
Human Fetal Skin
Skin

Uni-ZAP XR

(Skin)

H0083
HUMAN JURKAT
Jurkat Cells

Uni-ZAP XR

MEMBRANE BOUND

POLYSOMES

H0085
Human Colon
Human Colon

Lambda

ZAP II

H0086
Human epithelioid
Epithelioid
Sk Muscle

disease
Uni-ZAP XR

sarcoma
Sarcoma, muscle

H0087
Human Thymus
Human Thymus

pBluescript

H0090
Human T-Cell Lymphoma
T-Cell Lymphoma
T-Cell

disease
Uni-ZAP XR

H0098
Human Adult Liver,
Human Adult Liver
Liver

Uni-ZAP XR

subtracted

H0100
Human Whole Six Week
Human Whole Six
Embryo

Uni-ZAP XR

Old Embryo
Week Old

Embryo

H0101
Human 7 Weeks Old
Human Whole 7
Embryo

Lambda

Embryo, subtracted
Week Old Embryo

ZAP II

H0102
Human Whole 6 Week
Human Whole Six
Embryo

pBluescript

Old Embryo (II), subt
Week Old Embryo

H0105
Human Fetal Heart,
Human Fetal Heart
Heart

pBluescript

subtracted

H0108
Human Adult Lymph
Human Adult
Lymph Node

Uni-ZAP XR

Node, subtracted
Lymph Node

H0116
Human Thymus Tumor,
Human Thymus
Thymus

pBluescript

subtracted
Tumor

H0118
Human Adult Kidney
Human Adult
Kidney

Uni-ZAP XR

Kidney

H0122
Human Adult Skeletal
Human Skeletal
Sk Muscle

Uni-ZAP XR

Muscle
Muscle

H0123
Human Fetal Dura Mater
Human Fetal Dura
Brain

Uni-ZAP XR

Mater

H0124
Human
Human
Sk Muscle

disease
Uni-ZAP XR

Rhabdomyosarcoma
Rhabdomyosarcoma

H0125
Cem cells cyclohexamide
Cyclohexamide
Blood
Cell Line

Uni-ZAP XR

treated
Treated Cem, Jurkat,

Raji, and Supt

H0130
LNCAP untreated
LNCAP Cell Line
Prostate
Cell Line

Uni-ZAP XR

H0131
LNCAP + o.3 nM R1881
LNCAP Cell Line
Prostate
Cell Line

Uni-ZAP XR

H0132
LNCAP + 30 nM R1881
LNCAP Cell Line
Prostate
Cell Line

Uni-ZAP XR

H0134
Raji Cells, cyclohexamide
Cyclohexamide
Blood
Cell Line

Uni-ZAP XR

treated
Treated Cem, Jurkat,

Raji, and Supt

H0135
Human Synovial Sarcoma
Human Synovial
Synovium

Uni-ZAP XR

Sarcoma

H0136
Supt Cells, cyclohexamide
Cyclohexamide
Blood
Cell Line

Uni-ZAP XR

treated
Treated Cem, Jurkat,

Raji, and Supt

H0141
Activated T-Cells, 12 hrs.
Activated T-Cells
Blood
Cell Line

Uni-ZAP XR

H0144
Nine Week Old Early
9 Wk Old Early
Embryo

Uni-ZAP XR

Stage Human
Stage Human

H0149
7 Week Old Early Stage
Human Whole 7
Embryo

Uni-ZAP XR

Human, subtracted
Week Old Embryo

H0150
Human Epididymus
Epididymis
Testis

Uni-ZAP XR

H0154
Human Fibrosarcoma
Human Skin
Skin

disease
Uni-ZAP XR

Fibrosarcoma

H0156
Human Adrenal Gland
Human Adrenal
Adrenal

disease
Uni-ZAP XR

Tumor
Gland Tumor
Gland

H0159
Activated T-Cells, 8 hrs.,
Activated T-Cells
Blood
Cell Line

Uni-ZAP XR

ligation 2

H0163
Human Synovium
Human Synovium
Synovium

Uni-ZAP XR

H0165
Human Prostate Cancer,
Human Prostate
Prostate

disease
Uni-ZAP XR

Stage B2
Cancer, stage B2

H0166
Human Prostate Cancer,
Human Prostate
Prostate

disease
Uni-ZAP XR

Stage B2 fraction
Cancer, stage B2

H0169
Human Prostate Cancer,
Human Prostate
Prostate

disease
Uni-ZAP XR

Stage C fraction
Cancer, stage C

H0170
12 Week Old Early Stage
Twelve Week Old
Embryo

Uni-ZAP XR

Human
Early Stage Human

H0171
12 Week Old Early Stage
Twelve Week Old
Embryo

Uni-ZAP XR

Human, II
Early Stage Human

H0172
Human Fetal Brain,
Human Fetal Brain
Brain

Lambda

random primed

ZAP II

H0175
H. Adult Spleen, ziplox

pSport1

H0176
CAMA1Ee Cell Line
CAMA1Ee Cell
Breast
Cell Line

Uni-ZAP XR

Line

H0178
Human Fetal Brain
Human Fetal Brain
Brain

Uni-ZAP XR

H0179
Human Neutrophil
Human Neutrophil
Blood
Cell Line

Uni-ZAP XR

H0181
Human Primary Breast
Human Primary
Breast

disease
Uni-ZAP XR

Cancer
Breast Cancer

H0182
Human Primary Breast
Human Primary
Breast

disease
Uni-ZAP XR

Cancer
Breast Cancer

H0187
Resting T-Cell
T-Cells
Blood
Cell Line

Lambda

ZAP II

H0188
Human Normal Breast
Human Normal
Breast

Uni-ZAP XR

Breast

H0192
Cem Cells, cyclohexamide
Cyclohexamide
Blood
Cell Line

Uni-ZAP XR

treated, subtra
Treated Cem, Jurkat,

Raji, and Supt

H0194
Human Cerebellum,
Human Cerebellum
Brain

pBluescript

subtracted

H0196
Human Cardiomyopathy,
Human
Heart

Uni-ZAP XR

subtracted
Cardiomyopathy

H0201
Human Hippocampus,
Human
Brain

pBluescript

subtracted
Hippocampus

H0205
Human Colon Cancer,
Human Colon
Colon

pBluescript

differential
Cancer

H0208
Early Stage Human Lung,
Human Fetal Lung
Lung

pBluescript

subtracted

H0212
Human Prostate,
Human Prostate
Prostate

pBluescript

subtracted

H0213
Human Pituitary,
Human Pituitary

Uni-ZAP XR

subtracted

H0214
Raji cells, cyclohexamide
Cyclohexamide
Blood
Cell Line

pBluescript

treated, subtracted
Treated Cem, Jurkat,

Raji, and Supt

H0222
Activated T-Cells, 8 hrs,
Activated T-Cells
Blood
Cell Line

Uni-ZAP XR

subtracted

H0229
Early Stage Human Brain,
Early Stage Human
Brain

Lambda

random primed
Brain

ZAP II

H0230
Human Cardiomyopathy,
Human
Heart

disease
Uni-ZAP XR

diff exp
Cardiomyopathy

H0231
Human Colon, subtraction
Human Colon

pBluescript

H0239
Human Kidney Tumor
Human Kidney
Kidney

disease
Uni-ZAP XR

Tumor

H0242
Human Fetal Heart,
Human Fetal Heart
Heart

pBluescript

Differential (Fetal-

Specific)

H0244
Human 8 Week Whole
Human 8 Week Old
Embryo

Uni-ZAP XR

Embryo, subtracted
Embryo

H0250
Human Activated
Human Monocytes

Uni-ZAP XR

Monocytes

H0251
Human Chondrosarcoma
Human
Cartilage

disease
Uni-ZAP XR

Chondrosarcoma

H0252
Human Osteosarcoma
Human
Bone

disease
Uni-ZAP XR

Osteosarcoma

H0253
Human adult testis, large
Human Adult Testis
Testis

Uni-ZAP XR

inserts

H0254
Breast Lymph node cDNA
Breast Lymph Node
Lymph Node

Uni-ZAP XR

library

H0255
breast lymph node CDNA
Breast Lymph Node
Lymph Node

Lambda

library

ZAP II

H0261
H. cerebellum, Enzyme
Human Cerebellum
Brain

Uni-ZAP XR

subtracted

H0263
human colon cancer
Human Colon
Colon

disease
Lambda

Cancer

ZAP II

H0264
human tonsils
Human Tonsil
Tonsil

Uni-ZAP XR

H0265
Activated T-Cell
T-Cells
Blood
Cell Line

Uni-ZAP XR

(12hs)/Thiouridine

labelledEco

H0266
Human Microvascular
HMEC
Vein
Cell Line

Lambda

Endothelial Cells, fract. A

ZAP II

H0268
Human Umbilical Vein
HUVE Cells
Umbilical
Cell Line

Lambda

Endothelial Cells, fract. A

vein

ZAP II

H0269
Human Umbilical Vein
HUVE Cells
Umbilical
Cell Line

Lambda

Endothelial Cells, fract. B

vein

ZAP II

H0270
HPAS (human pancreas,
Human Pancreas
Pancreas

Uni-ZAP XR

subtracted)

H0271
Human Neutrophil,
Human Neutrophil -
Blood
Cell Line

Uni-ZAP XR

Activated
Activated

H0272
HUMAN TONSILS,
Human Tonsil
Tonsil

Uni-ZAP XR

FRACTION 2

H0274
Human Adult Spleen,
Human Adult
Spleen

Uni-ZAP XR

fractionII
Spleen

H0280
K562 + PMA (36 hrs)
K562 Cell line
cell line
Cell Line

ZAP Express

H0284
Human OB MG63 control
Human
Bone
Cell Line

Uni-ZAP XR

fraction I
Osteoblastoma

MG63 cell line

H0286
Human OB MG63 treated
Human
Bone
Cell Line

Uni-ZAP XR

(10 nM E2) fraction I
Osteoblastoma

MG63 cell line

H0288
Human OB HOS control
Human
Bone
Cell Line

Uni-ZAP XR

fraction I
Osteoblastoma HOS

cell line

H0290
Human OB HOS treated
Human
Bone
Cell Line

Uni-ZAP XR

(1 nM E2) fraction I
Osteoblastoma HOS

cell line

H0292
Human OB HOS treated
Human
Bone
Cell Line

Uni-ZAP XR

(10 nM E2) fraction I
Osteoblastoma HOS

cell line

H0293
WI 38 cells

Uni-ZAP XR

H0294
Amniotic Cells - TNF
Amniotic Cells -
Placenta
Cell Line

Uni-ZAP XR

induced
TNF induced

H0295
Amniotic Cells - Primary
Amniotic Cells -
Placenta
Cell Line

Uni-ZAP XR

Culture
Primary Culture

H0305
CD34 positive cells (Cord
CD34 Positive Cells
Cord Blood

ZAP Express

Blood)

H0306
CD34 depleted Buffy Coat
CD34 Depleted
Cord Blood

ZAP Express

(Cord Blood)
Buffy Coat (Cord

Blood)

H0309
Human Chronic Synovitis
Synovium, Chronic
Synovium

disease
Uni-ZAP XR

Synovitis/

Osteoarthritis

H0310
human caudate nucleus
Brain
Brain

Uni-ZAP XR

H0316
HUMAN STOMACH
Human Stomach
Stomach

Uni-ZAP XR

H0318
HUMAN B CELL
Human B Cell
Lymph Node

disease
Uni-ZAP XR

LYMPHOMA
Lymphoma

H0320
Human frontal cortex
Human Frontal
Brain

Uni-ZAP XR

Cortex

H0327
human corpus colosum
Human Corpus
Brain

Uni-ZAP XR

Callosum

H0328
human ovarian cancer
Ovarian Cancer
Ovary

disease
Uni-ZAP XR

H0329
Dermatofibrosarcoma
Dermatofibrosarcoma
Skin

disease
Uni-ZAP XR

Protuberance
Protuberans

H0331
Hepatocellular Tumor
Hepatocellular
Liver

disease
Lambda

Tumor

ZAP II

H0333
Hemangiopericytoma
Hemangiopericytoma
Blood vessel

disease
Lambda

ZAP II

H0334
Kidney cancer
Kidney Cancer
Kidney

disease
Uni-ZAP XR

H0339
Duodenum
Duodenum

Uni-ZAP XR

H0340
Corpus Callosum
Corpus Collosum-

Uni-ZAP XR

93052

H0341
Bone Marrow Cell Line
Bone Marrow Cell
Bone Marrow
Cell Line

Uni-ZAP XR

(RS4; 11)
Line RS4; 11

H0343
stomach cancer (human)
Stomach Cancer -

disease
Uni-ZAP XR

5383A (human)

H0345
SKIN
Skin - 4000868H
Skin

Uni-ZAP XR

H0349
human adult liver cDNA
Human Adult Liver
Liver

pCMVSport

library

1

H0351
Glioblastoma
Glioblastoma
Brain

disease
Uni-ZAP XR

H0352
wilm″s tumor
Wilm″s Tumor

disease
Uni-ZAP XR

H0355
Human Liver
Human Liver,

pCMVSport

normal Adult

1

H0359
KMH2 cell line
KMH2

ZAP Express

H0361
Human rejected kidney
Human Rejected

disease
pBluescript

Kidney

H0364
Human Osteoclastoma,
Human

disease
pBluescript

excised
Osteoclastoma

H0369
H. Atrophic Endometrium
Atrophic

Uni-ZAP XR

Endometrium and

myometrium

H0370
H. Lymph node breast
Lymph node with

disease
Uni-ZAP XR

Cancer
Met. Breast Cancer

H0373
Human Heart
Human Adult Heart
Heart

pCMVSport

1

H0374
Human Brain
Human Brain

pCMVSport

1

H0375
Human Lung
Human Lung

pCMVSport

1

H0379
Human Tongue, frac 1
Human Tongue

pSport1

H0386
Leukocyte and Lung; 4
Human Leukocytes
Blood
Cell Line

pCMVSport

screens

1

H0390
Human Amygdala
Human Amygdala

disease
pBluescript

Depression, re-excision
Depression

H0391
H. Meniingima, M6
Human Meningima
brain

pSport1

H0392
H. Meningima, M1
Human Meningima
brain

pSport1

H0393
Fetal Liver, subtraction II
Human Fetal Liver
Liver

pBluescript

H0394
A-14 cell line
Redd-Sternberg cell

ZAP Express

H0395
A1-CELL LINE
Redd-Sternberg cell

ZAP Express

H0396
L1 Cell line
Redd-Sternberg cell

ZAP Express

H0399
Human Kidney Cortex, re-
Human Kidney

Lambda

rescue
Cortex

ZAP II

H0402
CD34 depleted Buffy Coat
CD34 Depleted
Cord Blood

ZAP Express

(Cord Blood), re-excision
Buffy Coat (Cord

Blood)

H0403
H. Umbilical Vein
HUVE Cells
Umbilical
Cell Line

Uni-ZAP XR

Endothelial Cells, IL4

vein

induced

H0404
H. Umbilical Vein
HUVE Cells
Umbilical
Cell Line

Uni-ZAP XR

endothelial cells,

vein

uninduced

H0409
H. Striatum Depression,
Human Brain,
Brain

pBluescript

subtracted
Striatum Depression

H0411
H Female Bladder, Adult
Human Female
Bladder

pSport1

Adult Bladder

H0412
Human umbilical vein
HUVE Cells
Umbilical
Cell Line

pSport1

endothelial cells, IL-4

vein

induced

H0413
Human Umbilical Vein
HUVE Cells
Umbilical
Cell Line

pSport1

Endothelial Cells,

vein

uninduced

H0414
Ovarian Tumor I, OV5232
Ovarian Tumor,
Ovary

disease
pSport1

OV5232

H0415
H. Ovarian Tumor, II,
Ovarian Tumor,
Ovary

disease
pCMVSport

OV5232
OV5232

2.0

H0416
Human Neutrophils,
Human Neutrophil -
Blood
Cell Line

pBluescript

Activated, re-excision
Activated

H0421
Human Bone Marrow, re-
Bone Marrow

pBluescript

excision

H0422
T-Cell PHA 16 hrs
T-Cells
Blood
Cell Line

pSport1

H0423
T-Cell PHA 24 hrs
T-Cells
Blood
Cell Line

pSport1

H0424
Human Pituitary, subt IX
Human Pituitary

pBluescript

H0427
Human Adipose
Human Adipose, left

pSport1

hiplipoma

H0428
Human Ovary
Human Ovary
Ovary

pSport1

Tumor

H0429
K562 + PMA (36 hrs), re-
K562 Cell line
cell line
Cell Line

ZAP Express

excision

H0431
H. Kidney Medulla, re-
Kidney medulla
Kidney

pBluescript

excision

H0433
Human Umbilical Vein
HUVE Cells
Umbilical
Cell Line

pBluescript

Endothelial cells, frac B,

vein

re-excision

H0434
Human Brain, striatum,
Human Brain,

pBluescript

re-excision
Striatum

H0435
Ovarian Tumor 10-3-95
Ovarian Tumor,
Ovary

pCMVSport

OV350721

2.0

H0436
Resting T-Cell Library, II
T-Cells
Blood
Cell Line

pSport1

H0437
H Umbilical Vein
HUVE Cells
Umbilical
Cell Line

Lambda

Endothelial Cells, frac A,

vein

ZAP II

re-excision

H0438
H. Whole Brain #2, re-
Human Whole Brain

ZAP Express

excision
#2

H0441
H. Kidney Cortex,
Kidney cortex
Kidney

pBluescript

subtracted

H0444
Spleen metastic melanoma
Spleen, Metastic
Spleen

disease
pSport1

malignant

melanoma

H0445
Spleen, Chronic
Human Spleen, CLL
Spleen

disease
pSport1

lymphocytic leukemia

H0455
H. Striatum Depression,
Human Brain,
Brain

pBluescript

subt
Striatum Depression

H0456
H Kidney Cortex,
Human Kidney

pBluescript

subtracted III
Cortex

H0457
Human Eosinophils
Human Eosinophils

pSport1

H0459
CD34 + cells, II,
CD34 positive cells

pCMVSport

FRACTION 2

2.0

H0477
Human Tonsil, Lib 3
Human Tonsil
Tonsil

pSport1

H0478
Salivary Gland, Lib 2
Human Salivary
Salivary

pSport1

Gland
gland

H0479
Salivary Gland, Lib 3
Human Salivary
Salivary

pSport1

Gland
gland

H0483
Breast Cancer cell line,
Breast Cancer Cell

pSport1

MDA 36
line, MDA 36

H0484
Breast Cancer Cell line,
Breast Cancer Cell

pSport1

angiogenic
line, Angiogenic,

36T3

H0485
Hodgkin″s Lymphoma I
Hodgkin″s

disease
pCMVSport

Lymphoma I

2.0

H0486
Hodgkin″s Lymphoma II
Hodgkin″s

disease
pCMVSport

Lymphoma II

2.0

H0487
Human Tonsils, lib I
Human Tonsils

pCMVSport

2.0

H0488
Human Tonsils, Lib 2
Human Tonsils

pCMVSport

2.0

H0489
Crohn″s Disease
Ileum
Intestine

disease
pSport1

H0494
Keratinocyte
Keratinocyte

pCMVSport

2.0

H0497
HEL cell line
HEL cell line

HEL

pSport1

92.1.7

H0505
Human Astrocyte
Human Astrocyte

pSport1

H0506
Ulcerative Colitis
Colon
Colon

pSport1

H0509
Liver, Hepatoma
Human Liver,
Liver

disease
pCMVSport

Hepatoma, patient 8

3.0

H0510
Human Liver, normal
Human Liver,
Liver

pCMVSport

normal, Patient #8

3.0

H0517
Nasal polyps
Nasal polyps

pCMVSport

2.0

H0518
pBMC stimulated w/poly
pBMC stimulated

pCMVSport

I/C
with poly I/C

3.0

H0519
NTERA2, control
NTERA2,

pCMVSport

Teratocarcinoma

3.0

cell line

H0520
NTERA2 + retinoic acid,
NTERA2,

pSport1

14 days
Teratocarcinoma

cell line

H0521
Primary Dendritic Cells,
Primary Dendritic

pCMVSport

lib 1
cells

3.0

H0522
Primary Dendritic
Primary Dendritic

pCMVSport

cells,frac 2
cells

3.0

H0528
Poly[I]/Poly[C] Normal
Poly[I]/Poly[C]

pCMVSport

Lung Fibroblasts
Normal Lung

3.0

Fibroblasts

H0529
Myoloid Progenitor Cell
TF-1 Cell Line;

pCMVSport

Line
Myoloid progenitor

3.0

cell line

H0530
Human Dermal
Human Dermal

pSport1

Endothelial
Endothelial Cells;

Cells, untreated
untreated

H0538
Merkel Cells
Merkel cells
Lymph node

pSport1

H0539
Pancreas Islet Cell Tumor
Pancreas Islet Cell
Pancreas

disease
pSport1

Tumour

H0540
Skin, burned
Skin, leg burned
Skin

pSport1

H0542
T Cell helper I
Helper T cell

pCMVSport

3.0

H0543
T cell helper II
Helper T cell

pCMVSport

3.0

H0544
Human endometrial
Human endometrial

pCMVSport

stromal cells
stromal cells

3.0

H0545
Human endometrial
Human endometrial

pCMVSport

stromal cells-treated with
stromal cells-treated

3.0

progesterone
with proge

H0546
Human endometrial
Human endometrial

pCMVSport

stromal cells-treated with
stromal cells-treated

3.0

estradiol
with estra

H0547
NTER A2 teratocarcinoma
NTERA2,

pSport1

cell line + retinoic acid (14
Teratocarcinoma

days)
cell line

H0549
H. Epididiymus, caput &
Human

Uni-ZAP XR

corpus
Epididiyrnus, caput

and corpus

H0550
H. Epididiymus, cauda
Human

Uni-ZAP XR

Epididiymus, cauda

H0551
Human Thymus Stromal
Human Thymus

pCMVSport

Cells
Stromal Cells

3.0

H0553
Human Placenta
Human Placenta

pCMVSport

3.0

H0555
Rejected Kidney, lib 4
Human Rejected
Kidney

disease
pCMVSport

Kidney

3.0

H0556
Activated T-
T-Cells
Blood
Cell Line

Uni-ZAP XR

cell(12 h)/Thiouridine-re-

excision

H0559
HL-60, PMA 4H, re-
HL-60 Cells, PMA
Blood
Cell Line

Uni-ZAP XR

excision
stimulated 4H

H0560
KMH2
KMH2

pCMVSport

3.0

H0561
L428
L428

pCMVSport

3.0

H0562
Human Fetal Brain,
Human Fetal Brain

pCMVSport

normalized c5-11-26

2.0

H0563
Human Fetal Brain,
Human Fetal Brain

pCMVSport

normalized 50021F

2.0

H0564
Human Fetal Brain,
Human Fetal Brain

pCMVSport

normalized C5001F

2.0

H0566
Human Fetal
Human Fetal Brain

pCMVSport

Brain,normalized c50F

2.0

H0569
Human Fetal Brain,
Human Fetal Brain

pCMVSport

normalized CO

2.0

H0570
Human Fetal Brain,
Human Fetal Brain

pCMVSport

normalized C500H

2.0

H0571
Human Fetal Brain,
Human Fetal Brain

pCMVSport

normalized C500HE

2.0

H0572
Human Fetal Brain,
Human Fetal Brain

pCMVSport

normalized AC5002

2.0

H0574
Hepatocellular Tumor; re-
Hepatocellular
Liver

disease
Lambda

excision
Tumor

ZAP II

H0575
Human Adult
Human Adult
Lung

Uni-ZAP XR

Pulmonary; re-excision
Pulmonary

H0576
Resting T-Cell; re-
T-Cells
Blood
Cell Line

Lambda

excision

ZAP II

H0579
Pericardium
Pericardium
Heart

pSport1

H0580
Dendritic cells, pooled
Pooled dendritic

pCMVSport

cells

3.0

HO581
Human Bone Marrow,
Human Bone
Bone Marrow

pCMVSport

treated
Marrow

3.0

H0583
B Cell lymphoma
B Cell Lymphoma
B Cell

disease
pCMVSport

3.0

H0586
Healing groin wound, 6.5
healing groin
groin

disease
pCMVSport

hours post incision
wound, 6.5 hours

3.0

post incision - 2/

H0587
Healing groin wound; 7.5
Groin-2/19/97
groin

disease
pCMVSport

hours post incision

3.0

H0589
CD34 positive cells (cord
CD34 Positive Cells
Cord Blood

ZAP Express

blood), re-ex

H0590
Human adult small
Human Adult Small
Small Int.

Uni-ZAP XR

intestine, re-excision
Intestine

H0591
Human T-cell
T-Cell Lymphoma
T-Cell

disease
Uni-ZAP XR

lymphoma; re-excision

H0592
Healing groin wound -
HGS wound healing

disease
pCMVSport

zero hr post-incision
project; abdomen

3.0

(control)

H0593
Olfactory
Olfactory epithelium

pCMVSport

epithelium; nasalcavity
from roof of left

3.0

nasal cacit

H0594
Human Lung Cancer; re-
Human Lung Cancer
Lung

disease
Lambda

excision

ZAP II

H0595
Stomach cancer
Stomach Cancer -

disease
Uni-ZAP XR

(human); re-excision
5383A (human)

H0596
Human Colon Cancer; re-
Human Colon
Colon

Lambda

excision
Cancer

ZAP II

H0597
Human Colon; re-excision
Human Colon

Lambda

ZAP II

H0598
Human Stomach; re-
Human Stomach
Stomach

Uni-ZAP XR

excision

H0599
Human Adult Heart; re-
Human Adult Heart
Heart

Uni-ZAP XR

excision

H0600
Healing Abdomen
Abdomen

disease
pCMVSport

wound; 70&90 min post

3.0

incision

H0601
Healing Abdomen
Abdomen

disease
pCMVSport

Wound; 15 days post

3.0

incision

H0604
Human Pituitary, re-
Human Pituitary

pBluescript

excision

H0606
Human Primary Breast
Human Primary
Breast

disease
Uni-ZAP XR

Cancer; re-excision
Breast Cancer

H0608
H. Leukocytes, control
H. Leukocytes

pCMVSport

1

H0609
H. Leukocytes,
H. Leukocytes

pCMVSport

normalized cot > 500 A

1

H0614
H. Leukocytes,
H. Leukocytes

pCMVSport

normalized cot 500 A

H0615
Human Ovarian Cancer
Ovarian Cancer
Ovary

disease
Uni-ZAP XR

Reexcision

H0616
Human Testes, Reexcision
Human Testes
Testis

Uni-ZAP XR

H0617
Human Primary Breast
Human Primary
Breast

disease
Uni-ZAP XR

Cancer Reexcision
Breast Cancer

H0618
Human Adult Testes,
Human Adult Testis
Testis

Uni-ZAP XR

Large Inserts, Reexcision

H0619
Fetal Heart
Human Fetal Heart
Heart

Uni-ZAP XR

H0620
Human Fetal Kidney;
Human Fetal Kidney
Kidney

Uni-ZAP XR

Reexcision

H0622
Human Pancreas Tumor;
Human Pancreas
Pancreas

disease
Uni-ZAP XR

Reexcision
Tumor

H0623
Human Umbilical Vein;
Human Umbilical
Umbilical

Uni-ZAP XR

Reexcision
Vein Endothelial
vein

Cells

H0624
12 Week Early Stage
Twelve Week Old
Embryo

Uni-ZAP XR

Human II; Reexcision
Early Stage Human

H0625
Ku 812F Basophils Line
Ku 812F Basophils

pSport1

H0626
Saos2 Cells; Untreated
Saos2 Cell Line;

pSport1

Untreated

H0628
Human Pre-Differentiated
Human Pre-

Uni-ZAP XR

Adipocytes
Differentiated

Adipocytes

H0631
Saos2, Dexamethosome
Saos2 Cell Line;

pSport1

Treated
Dexamethosome

Treated

H0632
Hepatocellular Tumor; re-
Hepatocellular
Liver

Lambda

excision
Tumor

ZAP II

H0633
Lung Carcinoma A549
TNF alpha activated

disease
pSport1

TNF alpha activated
A549--Lung

Carcinoma

H0634
Human Testes Tumor, re-
Human Testes
Testis

disease
Uni-ZAP XR

excision
Tumor

H0635
Human Activated T-Cells,
Activated T-Cells
Blood
Cell Line

Uni-ZAP XR

re-excision

H0637
Dendritic Cells From
Dentritic cells from

pSport1

CD34 Cells
CD34 cells

H0638
CD40 activated monocyte
CD40 activated

pSport1

dendridic cells
monocyte dendridic

cells

H0639
Ficolled Human Stromal
Ficolled Human

Other

Cells, 5Fu treated
Stromal Cells, 5Fu

treated

H0641
LPS activated derived
LPS activated

pSport1

dendritic cells
monocyte derived

dendritic cells

H0642
Hep G2 Cells, lambda
Hep G2 Cells

Other

library

H0643
Hep G2 Cells, PCR library
Hep G2 Cells

Other

H0644
Human Placenta (re-
Human Placenta
Placenta

Uni-ZAP XR

excision)

H0645
Fetal Heart, re-excision
Human Fetal Heart
Heart

Uni-ZAP XR

H0646
Lung, Cancer (4005313
Metastatic

pSport1

A3): Invasive Poorly
squamous cell lung

Differentiated Lung
carcinoma, poorly di

Adenocarcinoma,

H0647
Lung, Cancer (4005163
Invasive poorly

disease
pSport1

B7): Invasive, Poorly Diff,
differentiated lung

Adenocarcinoma,
adenocarcinoma

Metastatic

H0648
Ovary, Cancer: (4004562
Papillary Cstic

disease
pSport1

B6) Papillary Serous
neoplasm of low

Cystic Neoplasm, Low
malignant potentia

Malignant Pot

H0649
Lung, Normal: (4005313
Normal Lung

pSport1

B1)

H0650
B-Cells
B-Cells

pCMVSport

3.0

H0651
Ovary, Normal:
Normal Ovary

pSport1

(9805C040R)

H0652
Lung, Normal: (4005313B1)
Normal Lung

pSport1

H0653
Stromal Cells
Stromal Cells

pSport1

H0656
B-cells (unstimulated)
B-cells

pSport1

(unstimulated)

H0657
B-cells (stimulated)
B-cells (stimulated)

pSport1

H0658
Ovary, Cancer
9809C332-Poorly
Ovary &

disease
pSport1

(9809C332): Poorly
differentiate
Fallopian

differentiated

Tubes

adenocarcinoma

H0659
Ovary, Cancer
Grade II Papillary
Ovary

disease
pSport1

(15395A1F): Grade II
Carcinoma, Ovary

Papillary Carcinoma

H0660
Ovary, Cancer:
Poorly differentiated

disease
pSport1

(15799A1F) Poorly
carcinoma, ovary

differentiated carcinoma

H0661
Breast, Cancer: (4004943A5)
Breast cancer

disease
pSport1

H0662
Breast, Normal:
Normal Breast -
Breast

pSport1

(4005522B2)
#4005522(B2)

H0663
Breast, Cancer: (4005522A2)
Breast Cancer -
Breast

disease
pSport1

#4005522(A2)

H0664
Breast, Cancer:
Breast Cancer
Breast

disease
pSport1

(9806C012R)

H0665
Stromal cells 3.88
Stromal cells 3.88

pSport1

H0666
Ovary, Cancer: (4004332A2)
Ovarian Cancer,

disease
pSport1

Sample

#4004332A2

H0667
Stromal cells(HBM3.18)
Stromal cell(HBM

pSport1

3.18)

H0668
stromal cell clone 2.5
stromal cell clone

pSport1

2.5

H0670
Ovary, Cancer(4004650A3):
Ovarian Cancer -

pSport1

Well-Differentiated
4004650A3

Micropapillary Serous

Carcinoma

H0672
Ovary, Cancer: (4004576A8)
Ovarian
Ovary

pSport1

Cancer(4004576A8)

H0673
Human Prostate Cancer,
Human Prostate
Prostate

Uni-ZAP XR

Stage B2; re-excision
Cancer, stage B2

H0674
Human Prostate Cancer,
Human Prostate
Prostate

Uni-ZAP XR

Stage C; re-excission
Cancer, stage C

H0675
Colon, Cancer:
Colon Cancer

pCMVSport

(9808C064R)
9808C064R

3.0

H0676
Colon, Cancer:
Colon Cancer

pCMVSport

(9808C064R)-total RNA
9808C064R

3.0

H0677
TNFR degenerate oligo
B-Cells

PCRII

H0682
Serous Papillary
serous papillary

pCMVSport

Adenocarcinoma
adenocarcinoma

3.0

(9606G304SPA3B)

H0683
Ovarian Serous Papillary
Serous papillary

pCMVSport

Adenocarcinoma
adenocarcinoma,

3.0

stage 3C (9804G01

H0684
Serous Papillary
Ovarian Cancer-
Ovaries

pCMVSport

Adenocarcinoma
9810G606

3.0

H0685
Adenocarcinoma of
Adenocarcinoma of

pCMVSport

Ovary, Human Cell Line,
Ovary, Human Cell

3.0

# OVCAR-3
Line, # OVCAR-

H0686
Adenocarcinoma of
Adenocarcinoma of

pCMVSport

Ovary, Human Cell Line
Ovary, Human Cell

3.0

Line, # SW-626

H0687
Human normal
Human normal
Ovary

pCMVSport

ovary (#9610G215)
ovary (#9610G215)

3.0

H0688
Human Ovarian
Human Ovarian

pCMVSport

Cancer (#9807G017)
cancer (#9807G017),

3.0

mRNA from Maura

Ru

H0689
Ovarian Cancer
Ovarian Cancer,

pCMVSport

#9806G019

3.0

H0690
Ovarian Cancer, #
Ovarian Cancer,

pCMVSport

9702G001
#9702G001

3.0

H0692
BLyS Receptor from
B Cell Lymphoma
B Cell

pCMVSport

Expression Cloning

3.0

H0693
Normal Prostate
Normal Prostate

pCMVSport

#ODQ3958EN
Tissue

3.0

#ODQ3958EN

H0695
mononucleocytes from
mononucleocytes

pCMVSport

patient
from patient at

3.0

Shady Grove Hospit

N0006
Human Fetal Brain
Human Fetal Brain

S0001
Brain frontal cortex
Brain frontal cortex
Brain

Lambda

ZAP II

S0002
Monocyte activated
Monocyte-activated
blood
Cell Line

Uni-ZAP XR

S0003
Human Osteoclastoma
Osteoclastoma
bone

disease
Uni-ZAP XR

S0005
Heart
Heart-left ventricle
Heart

pCDNA

S0007
Early Stage Human Brain
Human Fetal Brain

Uni-ZAP XR

S0010
Human Amygdala
Amygdala

Uni-ZAP XR

S0011
STROMAL -
Osteoclastoma
bone

disease
Uni-ZAP XR

OSTEOCLASTOMA

S0013
Prostate
Prostate
prostate

Uni-ZAP XR

S0014
Kidney Cortex
Kidney cortex
Kidney

Uni-ZAP XR

S0022
Human Osteoclastoma
Osteoclastoma

Uni-ZAP XR

Stromal Cells -
Stromal Cells

unamplified

S0023
Human Kidney Cortex -
Human Kidney

unamplified
Cortex

S0024
Human Kidney Medulla -
Human Kidney

unamplified
Medulla

S0026
Stromal cell TF274
stromal cell
Bone marrow
Cell Line

Uni-ZAP XR

S0027
Smooth muscle, serum
Smooth muscle
Pulmanary
Cell Line

Uni-ZAP XR

treated

artery

S0028
Smooth muscle, control
Smooth muscle
Pulmanary
Cell Line

Uni-ZAP XR

artery

S0029
brain stem
Brain stem
brain

Uni-ZAP XR

S0030
Brain pons
Brain Pons
Brain

Uni-ZAP XR

S0031
Spinal cord
Spinal cord
spinal cord

Uni-ZAP XR

S0032
Smooth muscle-ILb
Smooth muscle
Pulmanary
Cell Line

Uni-ZAP XR

induced

artery

S0036
Human Substantia Nigra
Human Substantia

Uni-ZAP XR

Nigra

S0037
Smooth muscle, IL1b
Smooth muscle
Pulmanary
Cell Line

Uni-ZAP XR

induced

artery

S0038
Human Whole Brain #2 -
Human Whole Brain

ZAP Express

Oligo dT > 1.5 Kb
#2

S0040
Adipocytes
Human Adipocytes

Uni-ZAP XR

from Osteoclastoma

S0042
Testes
Human Testes

ZAP Express

S0044
Prostate BPH
prostate BPH
Prostate

disease
Uni-ZAP XR

S0045
Endothelial cells-control
Endothelial cell
endothelial
Cell Line

Uni-ZAP XR

cell-lung

S0046
Endothelial-induced
Endothelial cell
endothelial
Cell Line

Uni-ZAP XR

cell-lung

S0049
Human Brain, Striatum
Human Brain,

Uni-ZAP XR

Striatum

S0050
Human Frontal Cortex,
Human Frontal

disease
Uni-ZAP XR

Schizophrenia
Cortex,

Schizophrenia

S0051
Human
Human

disease
Uni-ZAP XR

Hypothalmus,
Hypothalamus,

Schizophrenia
Schizophrenia

S0052
neutrophils control
human neutrophils
blood
Cell Line

Uni-ZAP XR

S0053
Neutrophils IL-1 and LPS
human neutrophil
blood
Cell Line

Uni-ZAP XR

induced
induced

S0106
STRIATUM

BRAIN

disease
Uni-ZAP XR

DEPRESSION

S0112
Hypothalamus

Brain

Uni-ZAP XR

S0114
Anergic T-cell
Anergic T-cell

Cell Line

Uni-ZAP XR

S0116
Bone marrow
Bone marrow
Bone marrow

Uni-ZAP XR

S0122
Osteoclastoma-normalized
Osteoclastoma
bone

disease
pBluescript

A

S0126
Osteoblasts
Osteoblasts
Knee
Cell Line

Uni-ZAP XR

S0132
Epithelial-TNFa and INF
Airway Epithelial

Uni-ZAP XR

induced

S0134
Apoptotic T-cell
apoptotic cells

Cell Line

Uni-ZAP XR

S0136
PERM TF274
stromal cell
Bone marrow
Cell Line

Lambda

ZAP II

S0142
Macrophage-oxLDL
macrophage-
blood
Cell Line

Uni-ZAP XR

oxidized LDL

treated

S0144
Macrophage (GM-CSF
Macrophage (GM-

Uni-ZAP XR

treated)
CSF treated)

S0146
prostate-edited
prostate BPH
Prostate

Uni-ZAP XR

S0148
Normal Prostate
Prostate
prostate

Uni-ZAP XR

S0150
LNCAP prostate cell line
LNCAP Cell Line
Prostate
Cell Line

Uni-ZAP XR

S0152
PC3 Prostate cell line
PC3 prostate cell

Uri-ZAP XR

line

S0168
Prostate/LNCAP,
PC3 prostate cell

pBluescript

subtraction I
line

S0174
Prostate-BPH subtracted II
Human Prostate

pBluescript

BPH

S0182
Human B Cell 8866
Human B- Cell 8866

Uni-ZAP XR

S0192
Synovial Fibroblasts
Synovial Fibroblasts

pSport1

(control)

S0194
Synovial hypoxia
Synovial Fibroblasts

pSport1

S0196
Synovial IL-1/TNF
Synovial Fibroblasts

pSport1

stimulated

S0198
7TM-pbfd
PBLS, 7TM

PCRII

receptor enriched

S0206
Smooth Muscle- HASTE
Smooth muscle
Pulmanary
Cell Line

pBluescript

normalized

artery

S0208
Messangial cell, frac 1
Messangial cell

pSort1

S0210
Messangial cell, frac 2
Messangial cell

pSport1

S0212
Bone Marrow Stromal
Bone Marrow

pSport1

Cell, untreated
Stromal

Cell, untreated

S0214
Human Osteoclastoma, re-
Osteoclastoma
bone

disease
Uni-ZAP XR

excision

S0216
Neutrophils IL-1 and LPS
human neutrophil
blood
Cell Line

Uni-ZAP XR

induced
induced

S0218
Apoptotic T-cell, re-
apoptotic cells

Cell Line

Uni-ZAP XR

excision

S0220
H. hypothalamus, frac
Hypothalamus
Brain

ZAP Express

A; re-excision

S0222
H. Frontal
H. Brain, Frontal
Brain

disease
Uni-ZAP XR

cortex, epileptic; re-
Cortex, Epileptic

excision

S0228
PSMIX
PBLS, 7TM

PCRII

receptor enriched

S0242
Synovial Fibroblasts
Synovial Fibroblasts

pSport1

(II1/TNF), subt

S0250
Human Osteoblasts II
Human Osteoblasts
Femur

disease
pCMVSport

2.0

S0252
7TM-PIMIX
PBLS, 7TM

PCRII

receptor enriched

S0260
Spinal Cord, re-excision
Spinal cord
spinal cord

Uni-ZAP XR

S0264
PPMIX
PPMIX (Human
Pituitary

PCRII

Pituitary)

S0268
PRMIX
PRMIX (Human
prostate

PCRII

Prostate)

S0270
PTMIX
PTMIX (Human
Thymus

PCRII

Thymus)

S0274
PCMIX
PCMIX (Human
Brain

PCRII

Cerebellum)

S0276
Synovial hypoxia-RSF
Synovial fobroblasts
Synovial

pSport1

subtracted
(rheumatoid)
tissue

S0278
H Macrophage (GM-CSF
Macrophage (GM-

Uni-ZAP XR

treated), re-excision
CSF treated)

S0280
Human Adipose Tissue,
Human Adipose

Uni-ZAP XR

re-excision
Tissue

S0282
Brain Frontal Cortex, re-
Brain frontal cortex
Brain

Lambda

excision

ZAP II

S0294
Larynx tumor
Larynx tumor
Larynx, vocal

disease
pSport1

cord

S0298
Bone marrow
Bone marrow
Bone marrow

pSport1

stroma, treated
stroma, treatedSB

S0300
Frontal lobe, dementia; re-
Frontal Lobe
Brain

Uni-ZAP XR

excision
dementia/Alzheimer′′s

S0306
Larynx normal #10 261-
Larynx normal

pSport1

273

S0308
Spleen/normal
Spleen normal

pSport1

S0310
Normal trachea
Normal trachea

pSport1

S0312
Human
Human

disease
pSport1

osteoarthritic; fraction II
osteoarthritic

cartilage

S0314
Human
Human

disease
pSport1

osteoarthritis; fraction I
osteoarthritic

cartilage

S0316
Human Normal
Human Normal

pSport1

Cartilage, Fraction I
Cartilage

S0318
Human Normal Cartilage
Human Normal

pSport1

Fraction II
Cartilage

S0328
Palate carcinoma
Palate carcinoma
Uvula

disease
pSport1

S0330
Palate normal
Palate normal
Uvula

pSport1

S0332
Pharynx carcinoma
Pharynx carcinoma
Hypopharynx

pSport1

S0334
Human Normal Cartilage
Human Normal

pSport1

Fraction III
Cartilage

S0338
Human Osteoarthritic
Human

disease
pSport1

Cartilage Fraction III
osteoarthritic

cartilage

S0340
Human Osteoarthritic
Human

disease
pSport1

Cartilage Fraction IV
osteoarthritic

cartilage

S0342
Adipocytes; re-excision
Human Adipocytes

Uni-ZAP XR

from Osteoclastoma

S0344
Macrophage-oxLDL; re-
macrophage-
blood
Cell Line

Uni-ZAP XR

excision
oxidized LDL

treated

S0346
Human Amygdala; re-
Amygdala

Uni-ZAP XR

excision

S0350
Pharynx Carcinoma
Pharynx carcinoma
Hypopharynx

disease
pSport1

S0352
Larynx Carcinoma
Larynx carcinoma

disease
pSport1

S0354
Colon Normal II
Colon Normal
Colon

pSport1

S0356
Colon Carcinoma
Colon Carcinoma
Colon

disease
pSport1

S0358
Colon Normal III
Colon Normal
Colon

pSport1

S0360
Colon Tumor II
Colon Tumor
Colon

disease
pSport1

S0362
Human Gastrocnemius
Gastrocnemius

pSport1

muscle

S0364
Human Quadriceps
Quadriceps muscle

pSport1

S0366
Human Soleus
Soleus Muscle

pSport1

S0370
Larynx carcinoma II
Larynx carcinoma

disease
pSport1

S0374
Normal colon
Normal colon

pSport1

S0376
Colon Tumor
Colon Tumor

disease
pSport1

S0378
Pancreas normal PCA4
Pancreas Normal

pSport1

No
PCA4 No

S0380
Pancreas Tumor PCA4 Tu
Pancreas Tumor

disease
pSport1

PCA4 Tu

S0386
Human Whole Brain, re-
Whole brain
Brain

ZAP Express

excision

S0388
Human
Human

disease
Uni-ZAP XR

Hypothalamus,
Hypothalamus,

schizophrenia,
Schizophrenia

re-excision

S0390
Smooth muscle, control;
Smooth muscle
Pulmanary
Cell Line

Uni-ZAP XR

re-excision

artery

S0392
Salivary Gland
Salivary gland;

pSport1

normal

S0400
Brain; normal
Brain; normal

pSport1

S0404
Rectum normal
Rectum, normal

pSport1

S0406
Rectum tumour
Rectum tumour

pSport1

S0408
Colon, normal
Colon, normal

pSport1

S0410
Colon, tumour
Colon, tumour

pSport1

S0412
Temporal cortex-
Temporal cortex,

disease
Other

Alzheizmer; subtracted
alzheimer

S0414
Hippocampus, Alzheimer
Hippocampus,

Other

Subtracted
Alzheimer

Subtracted

S0418
CHME Cell Line; treated 5
CHME Cell Line;

pCMVSport

hrs
treated

3.0

S0420
CHME Cell
CHME Cell line,

pSport1

Line, untreated
untreatetd

S0422
Mo7e Cell Line GM-CSF
Mo7e Cell Line

pCMVSport

treated (1 nglml)
GM-CSF treated

3.0

(1 ng/ml)

S0424
TF-1 Cell Line GM-CSF
TF-1 Cell Line

pSport1

Treated
GM-CSF Treated

S0426
Monocyte activated; re-
Monocyte-activated
blood
Cell Line

Uni-ZAP XR

excision

S0428
Neutrophils control; re-
human neutrophils
blood
Cell Line

Uni-ZAP XR

excision

S0430
Aryepiglottis Normal
Aryepiglottis

pSport1

Normal

S0432
Sinus piniformis Tumour
Sinus piniformis

pSport1

Tumour

S0434
Stomach Normal
Stomach Normal

disease
pSport1

S0436
Stomach Tumour
Stomach Tumour

disease
pSport1

S0438
Liver Normal Met5No
Liver Normal

pSport1

Met5No

S0440
Liver Tumour Met 5 Tu
Liver Tumour

pSport1

S0442
Colon Normal
Colon Normal

pSport1

S0444
Colon Tumor
Colon Tumour

disease
pSport1

S0446
Tongue Tumour
Tongue Tumour

pSport1

S0448
Larynx Normal
Larynx Normal

pSport1

S0450
Larynx Tumour
Larynx Tumour

pSport1

S0452
Thymus
Thymus

pSport1

S0456
Tongue Normal
Tongue Normal

pSport1

S0458
Thyroid Normal (SDCA2
Thyroid normal

pSport1

No)

S0460
Thyroid Tumour
Thyroid Tumour

pSport1

S0462
Thyroid Thyroiditis
Thyroid Thyroiditis

pSport1

S0464
Larynx Normal
Larynx Normal

pSport1

S0468
Ea.hy.926 cell line
Ea.hy.926 cell line

pSport1

S0472
Lung Mesothelium
PYBT

pSport1

S0474
Human blood platelets
Platelets
Blood

Other

platelets

S0665
Human Amygdala; re-
Amygdala

Uni-ZAP XR

excission

S3012
Smooth Muscle Serum
Smooth muscle
Pulmanary
Cell Line

pBluescript

Treated, Norm

artery

S3014
Smooth muscle, serum
Smooth muscle
Pulmanary
Cell Line

pBluescript

induced, re-exc

artery

S6014
H. hypothalamus, frac A
Hypothalamus
Brain

ZAP Express

S6016
H Frontal Cortex,
H. Brain, Frontal
Brain

disease
Uni-ZAP XR

Epileptic
Cortex, Epileptic

S6022
H. Adipose Tissue
Human Adipose

Uni-ZAP XR

Tissue

S6024
Alzheimers, spongy
Alzheimer″s/Spongy
Brain

disease
Uni-ZAP XR

change
change

S6026
Frontal Lobe, Dementia
Frontal Lobe
Brain

Uni-ZAP XR

dementia/Alzheimer′′s

S6028
Human Manic Depression
Human Manic
Brain

disease
Uni-ZAP XR

Tissue
depression tissue

T0002
Activated T-cells
Activated T-Cell,
Blood
Cell Line

pBluescript

PBL fraction

SK−

T0003
Human Fetal Lung
Human Fetal Lung

pBluescript

SK−

T0004
Human White Fat
Human White Fat

pBluescript

SK−

T0006
Human Pineal Gland
Human Pinneal

pBluescript

Gland

SK−

T0008
Colorectal Tumor
Colorectal Tumor

disease
pBluescript

SK−

T0010
Human Infant Brain
Human Infant Brain

Other

T0023
Human Pancreatic
Human Pancreatic

disease
pBluescript

Carcinoma
Carcinoma

SK−

T0039
HSA 172 Cells
Human HSA172 cell

pBluescript

line

SK−

T0040
HSC172 cells
SA172 Cells

pBluescript

SK−

T004l
Jurkat T-cell G1 phase
Jurkat T-cell

pBluescript

SK−

T0042
Jurkat T-Cell, S phase
Jurkat T-Cell Line

pBluescript

SK−

T0048
Human Aortic
Human Aortic

pBluescript

Endothelium
Endothilium

SK−

T0049
Aorta endothelial cells +
Aorta endothelial

pBluescript

TNF-a
cells

SK−

T0060
Human White Adipose
Human White Fat

pBluescript

SK−

T0067
Human Thyroid
Human Thyroid

pBluescript

SK−

T0068
Normal Ovary,
Normal Ovary,

pBluescript

Premenopausal
Premenopausal

SK−

T0069
Human Uterus, normal
Human Uterus,

pBluescript

normal

SK−

T0071
Human Bone Marrow
Human Bone

pBluescript

Marrow

SK−

T0082
Human Adult Retina
Human Adult Retina

pBluescript

SK−

T0109
Human (HCC) cell line

pBluescript

liver (mouse) metastasis,

SK−

remake

T0110
Human colon carcinoma

pBluescript

(HCC) cell line, remake

SK−

T0114
Human (Caco-2) cell line,

pBluescript

adenocarcinoma, colon,

SK−

remake

T0115
Human Colon Carcinoma

pBluescript

(HCC) cell line

SK−

L0002
Atrium cDNA library

Human heart

L0004
ClonTech HL 1065a

L0005
Clontech human aorta

polyA+ mRNA (#6572)

L0015
Human

L0021
Human adult (K. Okubo)

L0022
Human adult lung 3″

directed MboI CDNA

L0040
Human colon mucosa

L0053
Human pancreatic tumor

L0055
Human promyelocyte

L0096
Subtracted human retina

L0097
Subtracted human retinal

pigment epithelium (RPE)

L0l03
DKFZphamy1
amygdala

L0105
Human aorta polyA+
aorta

(TFujiwara)

L0109
Human brain cDNA
brain

L0118
Human fetal brain S.
brain

Meier-Ewert

L0142
Human placenta eDNA
placenta

(TFujiwara)

L0143
Human placenta polyA+
placenta

(TFujiwara)

L0151
Human testis (C. De
testis

Smet)

L0157
Human fetal brain
brain

(TFujiwara)

L0163
Human heart cDNA

heart

(YNakamura)

L0351
Infant brain, Bento Soares

BA, M13-

derived

L0352
Normalized infant brain,

BA, M13-

Bento Soares

derived

L0356
S, Human foetal Adrenals

Bluescript

tissue

L0361
Stratagene ovary

ovary

Bluescript

(#937217)

SK

L0362
Stratagene ovarian cancer

Bluescript

(#937219)

SK−

L0363
NCI_CGAP_GC2
germ cell tumor

Bluescript

SK−

L0364
NCI_CGAP_GC5
germ cell tumor

Bluescript

SK−

L0365
NCI_CGAP_Phe1
pheochromocytoma

Bluescript

SK−

L0366
Stratagene schizo brain
schizophrenic brain

Bluescript

S11
S-11 frontal lobe

SK−

L0367
NCI_CGAP_Sch1
Schwannoma tumor

Bluescript

SK−

L0368
NCI_CGAP_SS1
synovial sarcoma

Bluescript

SK−

L0369
NCI_CGAP_AAI
adrenal adenoma
adrenal gland

Bluescript

SK−

L0370
Johnston frontal cortex
pooled frontal lobe
brain

Bluescript

SK−

L0371
NCI_CGAP_Br3
breast tumor
breast

Bluescript

SK−

L0372
NCI_CGAP_Col2
colon tumor
colon

Bluescript

SK−

L0373
NCI_CGAP_Co11
tumor
colon

Bluescript

SK−

L0374
NCI_CGAP_Co2
tumor
colon

Bluescript

SK−

L0375
NCI_CGAP_Kid6
kidney tumor
kidney

Bluescript

SK−

L0376
NCI_CGAP_Lar1
larynx
larynx

Bluescript

SK−

L0378
NCI_CGAP_Lu1
lung tumor
lung

Bluescript

SK−

L0381
NCI_CGAP_HN4
squamous cell
pharynx

Bluescnipt

carcinoma

SK−

L0383
NCI_CGAP_Pr24
invasive tumor (cell
prostate

Bluescript

line)

SK−

L0384
NCI_CGAP_Pr23
prostate tumor
prostate

Bluescript

SK−

L0386
NCI_CGAP_HN3
squamous cell
tongue

Bluescnipt

carcinoma from base

SK−

of tongue

L0387
NCI_CGAP_GCB0
germinal center B-
tonsil

Bluescnipt

cells

SK−

L0388
NCI_CGAP_HN6
normal gingiva (cell

Bluescript

line from

SK−

immortalized kerati

L0389
NCI_CGAP_HN5
normal gingiva (cell

Bluescnipt

line from primary

SK−

keratinocyt

L0394
H, Human adult Brain

gt11

Cortex tissue

L0415
b4HB3MA Cot8-HAP-Ft

Lafmid BA

L0418
b4HB3MA-Cot109 + l0−

Lafmid BA

Bio

L0435
Infant brain, LLNL array

lafmid BA

of Dr. M. Soares 1NIB

L0438
normalized infant brain
total brain
brain

lafmid BA

cDNA

L0439
Soares infant brain 1NIB

whole brain

Lafmid BA

L0455
Human retina cDNA
retina
eye

lambda gt10

randomly primed

sublibrary

L0456
Human retina cDNA
retina
eye

lambda gt10

Tsp5091-cleaved

sublibrary

L0462
WATM1

lambda gt11

L0465
TEST1, Human adult

lambda

Testis tissue

nm1149

L0471
Human fetal heart,

Lambda

Lambda ZAP Express

ZAP Express

L0475
KG1-a Lambda Zap

KG1-a

Lambda Zap

Express cDNA library

Express

(Stratagene)

L0483
Human pancreatic islet

Lambda

ZAPII

L0485
STRATAGENE Human
skeletal muscle
leg muscle

Lambda

skeletal muscle cDNA

ZAPII

library, cat. #936215.

L0502
NCI_CGAP_Brl5
adenocarcinoma
breast

pAMP1

L0514
NCI_CGAP_Ov31
papillary serous
ovary

pAMP1

carcinoma

L0517
NCI_CGAP_Pr1

pAMP10

L0518
NCI_CGAP_Pr2

pAMP10

L0519
NCI_CGAP_Pr3

pAMP10

L0520
NCI_CGAP_Alv1
alveolar

pAMP10

rhabdomyosarcoma

L0521
NCI_CGAP_Ew1
Ewing″s sarcoma

pAMP10

L0523
NCI_CGAP_Lip2
liposarcoma

pAMP10

L0526
NCI_CGAP_Pr12
metastatic prostate

pAMP10

bone lesion

L0527
NCI_CGAP_Ov2
ovary

pAMP10

L0528
NCI_CGAP_Pr5
prostate

pAMP10

L0529
NCI_CGAP_Pr6
prostate

pAMP10

L0530
NCI_CGAP_Pr8
prostate

pAMP10

L0533
NCI_CGAP_HSC1
stem cells
bone marrow

pAMP10

L0534
Chromosome 7 Fetal
brain
brain

pAMP10

Brain cDNA Library

L0540
NCI_CGAP_Pr10
invasive prostate
prostate

pAMP10

tumor

L0542
NCI_CGAP_Pr11
normal prostatic
prostate

pAMP10

epithelial cells

L0545
NCI_CGAP_Pr4.1
prostatic
prostate

pAMP10

intraepithelial

neoplasia - high

grade

L0546
NCI_CGAP_Pr18
stroma
prostate

pAMP10

L0547
NCI_CGAP_Pr16
tumor
prostate

pAMP10

L0549
NCI_CGAP_HN10
carcinoma in situ

pAMP10

from retromolar

trigone

L0551
NCI_CGAP_HN7
normal squamous

pAMP10

epithelium, floor of

mouth

L0558
NCI_CGAP_Ov40
endometrioid
ovary

pAMP10

ovarian metastasis

L0562
Chromosome 7 HeLa

HeLa cell

pAMP10

cDNA Library

line;

ATCC

L0563
Human Bone Marrow
bone marrow

pBluescript

Stromal Fibroblast

L0564
Jia bone marrow stroma
bone marrow stroma

pBluescript

L0565
Normal Human
Bone
Hip

pBluescript

Trabecular Bone Cells

L0581
Stratagene liver (#937224)

liver

pBluescript

SK

L0584
Stratagene cDNA library

pBluescript

Human heart, cat#936208

SK(+)

L0588
Stratagene endothelial cell

pBluescript

937223

SK−

L0589
Stratagene fetal retina

pBluescript

937202

SK−

L0590
Stratagene fibroblast

pBluescript

(#937212)

SK−

L0591
Stratagene HeLa cell s3

pBluescript

937216

SK−

L0592
Stratagene h, NT neuron

pBluescript

(#937233)

SK−

L0593
Stragene

pBluescript

neuroepithelium

SK−

(#937231)

L0594
Stratagene

pBluescript

neuroepithelium

SK−

NT2RAMI 937234

L0595
Stratagene NT2 neuronal
neuroepithelial cells
brain

pBluescript

precursor 937230

SK−

L0596
Stratagene colon

colon

pBluescript

(#937204)

SK−

L0597
Stratagene corneal stroma

cornea

pBluescript

(#937222)

SK−

L0598
Morton Fetal Cochlea
cochlea
ear

pBluescript

SK−

L0599
Stratagene lung (#937210)

lung

pBluescript

SK−

L0600
Weizmann Olfactory
olfactory epithelium
nose

pBluescript

Epithelium

SK−

L0601
Stratagene pancreas

pancreas

pBluescript

(#937208)

SK−

L0602
Pancreatic Islet
pancreatic islet
pancreas

pBluescript

SK−

L0603
Stratagene placenta

placenta

pBluescript

(#937225)

SK−

L0604
Stratagene muscle 937209
muscle
skeletal

pBluescript

muscle

SK−

L0605
Stratagene fetal spleen
fetal spleen
spleen

pBluescript

(#937205)

SK−

L0606
NCI_CGAP_Lym5
follicular lymphoma
lymph node

pBluescript

SK−

L0608
Stratagene lung carcinoma
lung carcinoma
lung
NCI-H69

pBluescript

937218

SK−

L0615
22 week old human fetal

pBluescriptII

liver cDNA library

SK(-)

L0617
Chromosome 22 exon

pBluescriptII

KS+

L0618
Chromosome 9 exon

pBluescriptII

KS+

L0619
Chromosome 9 exon II

pBluescriptII

KS+

L0622
HM1

pcDNAII

(Invitrogen)

L0623
HM3
pectoral muscle

pcDNAII

(after mastectomy)

(Invitrogen)

L0629
NCI_CGAP_Me13
metastatic
bowel (skin

pCMV-

melanoma to bowel
primary)

SPORT4

L0630
NCI_CGAP_CNS1
substantia nigra
brain

pCMV-

SPORT4

L0631
NCI_CGAP_Br7

breast

pCMV-

SPORT4

L0635
NCI_CGAP_PNS1
dorsal root ganglion
peripheral

pCMV-

nervous

SPORT4

system

L0636
NCI_CGAP_Pit1
four pooled pituitary
brain

pCMV-

adenomas

SPORT6

L0637
NCI_CGAP_Brn53
three pooled
brain

pCMV-

meningiomas

SPORT6

L0638
NCI_CGAP_Brn35
tumor, 5 pooled (see
brain

pCMV-

description)

SPORT6

L0639
NCI_CGAP_Brn52
tumor, 5 pooled (see
brain

pCMV-

description)

SPORT6

L0640
NCI_CGAP_Brl8
four pooled high-
breast

pCMV-

grade tumors,

SPORT6

including two prima

L0641
NCI_CGAP_Co17
juvenile granulosa
colon

pCMV-

tumor

SPORT6

L0642
NCI_CGAP_Co18
moderately
colon

pCMV-

differentiated

SPORT6

adenocarcinoma

L0643
NCI_CGAP_Co19
moderately
colon

pCMV-

differentiated

SPORT6

adenocarcinoma

L0644
NCI_CGAP_Co20
moderately
colon

pCMV-

differentiated

SPORT6

adenocarcinoma

L0645
NCI_CGAP_Co21
moderately
colon

pCMV-

differentiated

SPORT6

adenocarcinoma

L0646
NCI_CGAP_Co14
moderately-
colon

pCMV-

differentiated

SPORT6

adenocarcinoma

L0647
NCI_CGAP_Sar4
five pooled
connective

pCMV-

sarcomas, including
tissue

SPORT6

myxoid liposarcoma

L0648
NCI_CGAP_Eso2
squamous cell
esophagus

pCMV-

carcinoma

SPORT6

L0649
NCI_CGAP_GU1
2 pooled high-grade
genitourinary

pCMV-

transitional cell
tract

SPORT6

tumors

L0650
NCI_CGAP_Kid13
2 pooled Wilms″
kidney

pCMV-

tumors, one primary

SPORT6

and one metast

L0651
NCI_CGAP_Kid8
renal cell tumor
kidney

pCMV-

SPORT6

L0653
NCI_CGAP_Lu28
two pooled
lung

pCMV-

squamous cell

SPORT6

carcinomas

L0654
NCI_CGAP_Lu31

lung, cell line

pCMV-

SPORT6

L0655
NCI_CGAP_Lym12
lymphoma,
lymph node

pCMV-

follicular mixed

SPORT6

small and large cell

L0656
NCI_CGAP_Ov38
normal epithelium
ovary

pCMV-

SPORT6

L0657
NCI_CGAP_Ov23
tumor, 5 pooled (see
ovary

pCMV-

description)

SPORT6

L0658
NCI_CGAP_Ov35
tumor, 5 pooled (see
ovary

pCMV-

description)

SPORT6

L0659
NCI_CGAP_Pan1
adenocarcinoma
pancreas

pCMV-

SPORT6

L0661
NCI_CGAP_Me115
malignant
skin

pCMV-

melanoma,

SPORT6

metastatic to lymph

node

L0662
NCI_CGAP_Gas4
poorly differentiated
stomach

pCMV-

adenocarcinoma

SPORT6

with signet r

L0663
NCI_CGAP_Ut2
moderately-
uterus

pCMV-

differentiated

SPORT6

endometrial

adenocarcino

L0664
NCI_CGAP_Ut3
poorly-differentiated
uterus

pCMV-

endometrial

SPORT6

adenocarcinoma,

L0665
NCI_CGAP_Ut4
serous papillary
uterus

pCMV-

carcinoma, high

SPORT6

grade, 2 pooled t

L0666
NCI_CGAP_Ut1
well-differentiated
uterus

pCMV-

endometrial

SPORT6

adenocarcinoma, 7

L0667
NCI_CGAP_CML1
myeloid cells, 18
whole blood

pCMV-

pooled CML cases,

SPORT6

BCR/ABL rearra

L0697
Testis 1

PGEM

5zf(+)

L0717
Gessler Wilms tumor

pSPORT1

L0731
Soares_pregnant_uterus—

uterus

pT7T3-Pac

NbHPU

L0738
Human colorectal cancer

pT7T3D

L0740
Soares melanocyte
melanocyte

pT7T3D

2NbHM

(Pharmacia)

with a

modified

polylinker

L0741
Soares adult brain

brain

pT7T3D

N2b4HB55Y

(Pharmacia)

with a

modified

polylinker

L0742
Soares adult brain
brain

pT7T3D

N2b5HB55Y

(Pharmacia)

with a

modified

polylinker

L0743
Soares breast 2NbHBst

breast

pT7T3D

(Pharmacia)

with a

modified

polylinker

L0744
Soares breast 3NbHBst

breast

pT7T3D

(Pharmacia)

with a

modified

polylinker

L0745
Soares retina N2b4HR
retina
eye

pT7T3D

(Pharmacia)

with a

modified

polylinker

L0746
Soares retina N2b5HR
retina
eye

pT7T3D

(Pharmacia)

with a

modified

polylinker

L0747
Soares_fetal_heart—

heart

pT7T3D

NbHH19W

(Pharmacia)

with a

modified

polylinker

L0748
Soares fetal liver spleen

Liver and

pT7T3D

1NFLS

Spleen

(Pharmacia)

with a

modified

polylinker

L0749
Soares_fetal_liver_spleen—

Liver and

pT7T3D

1NFLS_S1

Spleen

(Pharmacia)

with a

modified

polylinker

L0750
Soares_fetal_lung—

lung

pT7T3D

NbHL19W

(Pharmacia)

with a

modified

polylinker

L0751
Soares ovary tumor
ovarian tumor
ovary

pT7T3D

NbHOT

(Pharmacia)

with a

modified

polylinker

L0752
Soares_parathyroid_tumor—
parathyroid tumor
parathyroid

pT7T3D

NbHPA

gland

(Pharmacia)

with a

modified

polylinker

L0753
Soares_pineal_gland—

pineal gland

pT7T3D

N3HPG

(Pharmacia)

with a

modified

polylinker

L0754
Soares placenta Nb2HP

placenta

pT7T3D

(Pharmacia)

with a

modified

polylinker

L0755
Soares_placenta_8to9

placenta

pT7T3D

weeks_2NbHP8to9W

(Pharmacia)

with a

modified

polylinker

L0756
Soares_multiple_sclerosis—
multiple sclerosis

pT7T3D

2NbHMSP
lesions

(Pharmacia)

with a

modified

polylinker

V_TYPE

L0757
Soares_senescent—
senescent fibroblast

pT7T3D

fibroblasts_NbHSF

(Pharmacia)

with a

modified

polylinker

V_TYPE

L0758
Soares_testis_NHT

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0759
Soares_total_fetus—

pT7T3D-Pac

Nb2HF8_9w

(Pharmacia)

with a

modified

polylinker

L0761
NCI_CGAP_CLL1
B-cell, chronic

pT7T3D-Pac

lymphotic leukemia

(Pharmacia)

with a

modified

polylinker

L0762
NCI_CGAP_Br1.1
breast

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0763
NCI_CGAP_Br2
breast

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0764
NCI_CGAP_Co3
colon

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0766
NCI_CGAP_GCB1
germinal center B

pT7T3D-Pac

cell

(Pharmacia)

with a

modified

polylinker

L0767
NCI_CGAP_GC3
pooled germ cell

pT7T3D-Pac

tumors

(Pharmacia)

with a

modified

polylinker

L0768
NCI_CGAP_GC4
pooled germ cell

pT7T3D-Pac

tumors

(Pharmacia)

with a

modified

polylinker

L0769
NCI_CGAP_Brn25
anaplastic
brain

pT7T3D-Pac

oligodendroglioma

(Pharmacia)

with a

modified

polylinker

L0770
NCI_CGAP_Brn23
glioblastoma
brain

pT7T3D-Pac

(pooled)

(Pharmacia)

with a

modified

polylinker

L0771
NCI_CGAP_Co8
adenocarcinoma
colon

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0772
NCI_CGAP_Co10
colon tumor RER+
colon

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0773
NCI_CGAP_Co9
colon tumor RER+
colon

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0774
NCI_CGAP_Kid3

kidney

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0775
NCI_CGAP_Kid5
2pooled tumors
kidney

pT7T3D-Pac

(clear cell type)

(Pharmacia)

with a

modified

polylinker

L0776
NCI_CGAP_Lu5
carcinoid
lung

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0777
Soares_NhHMPu_S1
Pooled human
mixed (see

pT7T3D-Pac

melanocyte, fetal
below)

(Pharmacia)

heart, and pregnant

with a

modified

polylinker

L0778
Barstead pancreas

pancreas

pT7T3D-Pac

HPLRB1

(Pharmacia)

with a

modified

polylinker

L0779
Soares_NFL_T_GBC_S1
pooled

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0780
Soares_NSF_FS_9W_OT—

pooled

pT7T3D-Pac

PA_P_S1

(Pharmacia)

with a

modified

polylinker

L0782
NCI_CGAP_Pr21
normal prostate
prostate

pT7T3D-PaC

(Pharmacia)

with a

modified

polylinker

L0783
NCI_CGAP_Pr22
normal prostate
prostate

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0785
Barstead spleen HPLRB2

spleen

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0786
Soares_NbHFB

whole brain

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0787
NCI_CGAP_Sub1

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0788
NCI_CGAP_Sub2

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0789
NC_CGAP_Sub3

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0790
NCI_CGAP_Sub4

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0791
NCI_CGAP_Sub5

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0792
NCI_CGAP_Sub6

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0793
NCI_CGAP_Sub7

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0794
NCI_CGAP_GC6
pooled germ cell

pT7T3D-Pac

tumors

(Pharmacia)

with a

modified

polylinker

L0796
NCI_CGAP_Brn50
medulloblastoma
brain

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0800
NCI_CGAP_Co16
colon tumor, RER+
colon

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0803
NCI_CGAP_Kid11

kidney

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0804
NCI_CGAP_Kid12
2 pooled tumors
kidney

pT7T3D-Pac

(clear cell type)

(Pharmacia)

with a

modified

polylinker

L0805
NCI_CGAP_Lu24
carcinoid
lung

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0806
NCI_CGAP_Lu19
squamous cell
lung

pT7T3D-Pac

carcinoma, poorly

(Pharmacia)

differentiated (4

with a

modified

polylinker

L0807
NCI_CGAP_Ov18
fibrotheoma
ovary

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L0808
Barstead prostate

prostate

pT7T3D-Pac

BPHHPLRB4 1

(Pharmacia)

with a

modified

polylinker

L0809
NCI_CGAP_Pr28

prostate

pT7T3D-Pac

(Pharmacia)

with a

modified

polylinker

L2251
Human fetal lung
Fetal lung

[0090]

7

TABLE 5

OMIM

Refer-

ence
Description

104770
Amyloidosis, secondary, susceptibility to

106165
Hypertension, essential, 145500

107670
Apolipoprotein A-II deficiency

108730
Brodymyopathy, 601003

109400
Basal cell nevus syndrome

110700
Vivax malaria, susceptibility to

117700
[Hypoceruloplasminemia, hereditary]

117700
Hemosiderosis, systemic, due to aceruloplasminemia

125264
Leukemia, acute nonlymphocytic

126650
Chloride diarrhea, congenital, Finnish type, 214700

126650
Colon cancer

132800
Basal cell carcinoma

132800
Epithelioma, self-healing, squamous 1, Ferguson-Smith type

134570
Factor XIIIA deficiency

135940
Ichthyosis vulgaris, 146700

145001
Hyperparathyroidism-jaw tumor syndrome

146790
Lupus nephritis, susceptibility to

147781
Atopy, susceptibility to

150210
Lactoferrin-deficient neutrophils, 245480

150240
Cutis laxa, marfanoid neonatal type

152445
Vohwinkel syndrome, 124500

152445
Erythrokeratoderma, progressive symmetric, 602036

154276
Malignant hyperthermia susceptibility 3

159001
Muscular dystrophy, limb-girdle, type lB

169600
Hailey-Hailey disease

172471
Glycogenosis, hepatic, autosomal

173360
Thrombophilia due to excessive plasminogen activator inhibitor

173360
Hemorrhagic diathesis due to PAI1 deficiency

174000
Medullary cystic kidney disease, AD

179755
Renal cell carcinoma, papillary, 1

180105
Retinitis pigmentosa-l0

180380
Night blindness, congenital stationery, rhodopsin-related

180380
Retinitis pigmentosa, autosomal recessive

180380
Retinitis pigmentosa-4, autosomal dominant

182860
Pyropoikilocytosis

182860
Spherocytosis, recessive

182860
Elliptocytosis-2

186580
Arthrocutaneouveal granulomatosis

186855
Leukemia-2, T-cell acute lymphoblastic

190000
Atransferrinemia

191315
Insensitivity to pain, congenital, with anhidrosis, 256800

203500
Alkaptonuria

222800
Hemolytic anemia due to bisphosphoglycerate mutase deficiency

222900
Sucrose intolerance

223900
Dysautonomia, familial

230800
Gaucher disease

230800
Gaucher disease with cardiovascular calcification

232050
Propionicacidemia, type II or pccB type

246900
Lipoamide dehydrogenase deficiency

253800
Walker-Warburg syndrome, 236670

253800
Fukuyama type congenital muscular dystrophy

264800
Pseudoxanthoma elasticum

266200
Anemia, hemolytic, due to PK deficiency

266600
Inflammatory bowel disease-1

276902
Usher syndrome, type 3

278700
Xeroderma pigmentosum, group A

278760
Xeroderma pigmentosum, group F

600511
Schizophrenia-3

600760
Pseudohypoaldosteronism, type I, 264350

600760
Liddle syndrome, 177200

600761
Pseudohypoaldosteronism, type I, 264350

600761
Liddle syndrome, 177200

600882
Charcot-Marie-Tooth neuropathy-2B

600897
Cataract, zonular pulverulent-1, 116200

601105
Pycnodysostosis, 265800

601199
Neonatal hyperparathyroidism, 239200

601199
Hypocalcemia, autosomal dominant, 601198

601199
Hypocalciuric hypercalcemia, type I, 145980

601412
Deafness, autosomal dominant 7

601471
Moebius syndrome-2

601556
Spinocerebellar ataxia-1, 164400

601652
Glaucoma 1A, primary open angle, juvenile-onset, 137750

601682
Glaucoma 1C, primary open angle

602066
Convulsions, infantile and paroxysmal choreoathetosis

602088
Nephronophthisis, infantile

602136
Refsum disease, infantile, 266510

602136
Zellweger syndrome-1, 214100

602136
Adrenoleukodystrophy, neonatal, 202370

602447
Coronary artery disease, susceptibility to

602491
Hyperlipidemia, familial combined, 1

[0091] Polynucleotide and Polypeptide Variants

[0092] The present invention is directed to variants of the polynucleotide sequence disclosed in SEQ ID NO:X or the complementary strand thereto, nucleotide sequences encoding the polypeptide of SEQ ID NO:Y, the nucleotide sequence of SEQ ID NO:X encoding the polypeptide sequence as defined in column 7 of Table 1A, nucleotide sequences encoding the polypeptide as defined in column 7 of Table 1A, the nucleotide sequence as defined in columns 8 and 9 of Table 2, nucleotide sequences encoding the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2, the nucleotide sequence as defined in column 6 of Table 1B, nucleotide sequences encoding the polypeptide encoded by the nucleotide sequence as defined in column 6 of Table 1B, the cDNA sequence contained in Clone ID NO:Z, and/or nucleotide sequences encoding the polypeptide encoded by the cDNA sequence contained in Clone ID NO:Z.

[0093] The present invention also encompasses variants of the polypeptide sequence disclosed in SEQ ID NO:Y, the polypeptide sequence as defined in column 7 of Table 1A, a polypeptide sequence encoded by the polynucleotide sequence in SEQ ID NO:X, a polypeptide sequence encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2, a polypeptide sequence encoded by the nucleotide sequence as defined in column 6 of Table 1B, a polypeptide sequence encoded by the complement of the polynucleotide sequence in SEQ ID NO:X, and/or a polypeptide sequence encoded by the cDNA sequence contained in Clone ID NO:Z.

[0094] “Variant” refers to a polynucleotide or polypeptide differing from the polynucleotide or polypeptide of the present invention, but retaining essential properties thereof. Generally, variants are overall closely similar, and, in many regions, identical to the polynucleotide or polypeptide of the present invention.

[0095] Thus, one aspect of the invention provides an isolated nucleic acid molecule comprising, or alternatively consisting of, a polynucleotide having a nucleotide sequence selected from the group consisting of: (a) a nucleotide sequence described in SEQ ID NO:X or contained in the cDNA sequence of Clone ID NO:Z; (b) a nucleotide sequence in SEQ ID NO:X or the cDNA in Clone ID NO:Z which encodes the complete amino acid sequence of SEQ ID NO:Y or the complete amino acid sequence encoded by the cDNA in Clone ID NO:Z; (c) a nucleotide sequence in SEQ ID NO:X or the cDNA in Clone ID NO:Z which encodes a mature polypeptide; (d) a nucleotide sequence in SEQ ID NO:X or the cDNA sequence of Clone ID NO:Z, which encodes a biologically active fragment of a polypeptide; (e) a nucleotide sequence in SEQ ID NO:X or the cDNA sequence of Clone ID NO:Z, which encodes an antigenic fragment of a polypeptide; (f) a nucleotide sequence encoding a polypeptide comprising the complete amino acid sequence of SEQ ID NO:Y or the complete amino acid sequence encoded by the cDNA in Clone ID NO:Z; (g) a nucleotide sequence encoding a mature polypeptide of the amino acid sequence of SEQ ID NO:Y or the amino acid sequence encoded by the cDNA in Clone ID NO:Z; (h) a nucleotide sequence encoding a biologically active fragment of a polypeptide having the complete amino acid sequence of SEQ ID NO:Y or the complete amino acid sequence encoded by the cDNA in Clone ID NO:Z; (i) a nucleotide sequence encoding an antigenic fragment of a polypeptide having the complete amino acid sequence of SEQ ID NO:Y or the complete amino acid sequence encoded by the cDNA in Clone ID NO:Z; and (j) a nucleotide sequence complementary to any of the nucleotide sequences in (a), (b), (c), (d), (e), (f), (g), (h), or (i) above.

[0096] The present invention is also directed to nucleic acid molecules which comprise, or alternatively consist of, a nucleotide sequence which is at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100%, identical to, for example, any of the nucleotide sequences in (a), (b), (c), (d), (e), (f), (g), (h), (i), or (j) above, the nucleotide coding sequence in SEQ ID NO:X or the complementary strand thereto, the nucleotide coding sequence of the cDNA contained in Clone ID NO:Z or the complementary strand thereto, a nucleotide sequence encoding the polypeptide of SEQ ID NO:Y, a nucleotide sequence encoding a polypeptide sequence encoded by the nucleotide sequence in SEQ ID NO:X, a polypeptide sequence encoded by the complement of the polynucleotide sequence in SEQ ID NO:X, a nucleotide sequence encoding the polypeptide encoded by the cDNA contained in Clone ID NO:Z, the nucleotide coding sequence in SEQ ID NO:X as defined in columns 8 and 9 of Table 2 or the complementary strand thereto, a nucleotide sequence encoding the polypeptide encoded by the nucleotide sequence in SEQ ID NO:X as defined in columns 8 and 9 of Table 2 or the complementary strand thereto, the nucleotide coding sequence in SEQ ID NO:B as defined in column 6 of Table 1B or the complementary strand thereto, a nucleotide sequence encoding the polypeptide encoded by the nucleotide sequence in SEQ ID NO:B as defined in column 6 of Table 1B or the complementary strand thereto, the nucleotide sequence in SEQ ID NO:X encoding the polypeptide sequence as defined in column 7 of Table 1 A or the complementary strand thereto, nucleotide sequences encoding the polypeptide as defined in column 7 of Table 1A or the complementary strand thereto, and/or polynucleotide fragments of any of these nucleic acid molecules (e.g., those fragments described herein). Polynucleotides which hybridize to the complement of these nucleic acid molecules under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention, as are polypeptides encoded by these polynucleotides and nucleic acids.

[0097] In a preferred embodiment, the invention encompasses nucleic acid molecules which comprise, or alternatively, consist of a polynucleotide which hybridizes under stringent hybridization conditions, or alternatively, under lower stringency conditions, to a polynucleotide in (a), (b), (c), (d), (e), (f), (g), (h), or (i), above, as are polypeptides encoded by these polynucleotides. In another preferred embodiment, polynucleotides which hybridize to the complement of these nucleic acid molecules under stringent hybridization conditions, or alternatively, under lower stringency conditions, are also encompassed by the invention, as are polypeptides encoded by these polynucleotides.

[0098] In another embodiment, the invention provides a purified protein comprising, or alternatively consisting of, a polypeptide having an amino acid sequence selected from the group consisting of: (a) the complete amino acid sequence of SEQ ID NO:Y or the complete amino acid sequence encoded by the cDNA in Clone ID NO:Z; (b) the amino acid sequence of a mature form of a polypeptide having the amino acid sequence of SEQ ID NO:Y or the amino acid sequence encoded by the cDNA in Clone ID NO:Z; (c) the amino acid sequence of a biologically active fragment of a polypeptide having the complete amino acid sequence of SEQ ID NO:Y or the complete amino acid sequence encoded by the cDNA in Clone ID NO:Z; and (d) the amino acid sequence of an antigenic fragment of a polypeptide having the complete amino acid sequence of SEQ ID NO:Y or the complete amino acid sequence encoded by the cDNA in Clone ID NO:Z.

[0099] The present invention is also directed to proteins which comprise, or alternatively consist of, an amino acid sequence which is at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100%, identical to, for example, any of the amino acid sequences in (a), (b), (c), or (d), above, the amino acid sequence shown in SEQ ID NO:Y, the amino acid sequence encoded by the cDNA contained in Clone ID NO:Z, the amino acid sequence of the polypeptide encoded by the nucleotide sequence in SEQ ID NO:X as defined in columns 8 and 9 of Table 2, the amino acid sequence of the polypeptide encoded by the nucleotide sequence in SEQ ID NO:B as defined in column 6 of Table 1B, the amino acid sequence as defined in column 7 of Table 1A, an amino acid sequence encoded by the nucleotide sequence in SEQ ID NO:X, and an amino acid sequence encoded by the complement of the polynucleotide sequence in SEQ ID NO:X. Fragments of these polypeptides are also provided (e.g., those fragments described herein). Further proteins encoded by polynucleotides which hybridize to the complement of the nucleic acid molecules encoding these amino acid sequences under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention, as are the polynucleotides encoding these proteins.

[0100] By a nucleic acid having a nucleotide sequence at least, for example, 95% “identical” to a reference nucleotide sequence of the present invention, it is intended that the nucleotide sequence of the nucleic acid is identical to the reference sequence except that the nucleotide sequence may include up to five point mutations per each 100 nucleotides of the reference nucleotide sequence encoding the polypeptide. In other words, to obtain a nucleic acid having a nucleotide sequence at least 95% identical to a reference nucleotide sequence, up to 5% of the nucleotides in the reference sequence may be deleted or substituted with another nucleotide, or a number of nucleotides up to 5% of the total nucleotides in the reference sequence may be inserted into the reference sequence. The query sequence may be an entire sequence referred to in Table 1A or 2 as the ORF (open reading frame), or any fragment specified as described herein.

[0101] As a practical matter, whether any particular nucleic acid molecule or polypeptide is at least 80%, 85%, 90%, 95%, 96%, 97%, 98% or 99% identical to a nucleotide sequence of the present invention can be determined conventionally using known computer programs. A preferred method for determining the best overall match between a query sequence (a sequence of the present invention) and a subject sequence, also referred to as a global sequence alignment, can be determined using the FASTDB computer program based on the algorithm of Brutlag et al. (Comp. App. Biosci. 6:237-245 (1990)). In a sequence alignment the query and subject sequences are both DNA sequences. An RNA sequence can be compared by converting U's to T's. The result of said global sequence alignment is expressed as percent identity. Preferred parameters used in a FASTDB alignment of DNA sequences to calculate percent identity are: Matrix=Unitary, k-tuple=4, Mismatch Penalty=1, Joining Penalty=30, Randomization Group Length=0, Cutoff Score=1, Gap Penalty=5, Gap Size Penalty 0.05, Window Size=500 or the length of the subject nucleotide sequence, whichever is shorter.

[0102] If the subject sequence is shorter than the query sequence because of 5′ or 3′ deletions, not because of internal deletions, a manual correction must be made to the results. This is because the FASTDB program does not account for 5′ and 3′ truncations of the subject sequence when calculating percent identity. For subject sequences truncated at the 5′ or 3′ ends, relative to the query sequence, the percent identity is corrected by calculating the number of bases of the query sequence that are 5′ and 3′ of the subject sequence, which are not matched/aligned, as a percent of the total bases of the query sequence. Whether a nucleotide is matched/aligned is determined by results of the FASTDB sequence alignment. This percentage is then subtracted from the percent identity, calculated by the above FASTDB program using the specified parameters, to arrive at a final percent identity score. This corrected score is what is used for the purposes of the present invention. Only bases outside the 5′ and 3′ bases of the subject sequence, as displayed by the FASTDB alignment, which are not matched/aligned with the query sequence, are calculated for the purposes of manually adjusting the percent identity score.

[0103] For example, a 90 base subject sequence is aligned to a 100 base query sequence to determine percent identity. The deletions occur at the 5′ end of the subject sequence and therefore, the FASTDB alignment does not show a matched/alignment of the first 10 bases at 5′ end. The 10 unpaired bases represent 10% of the sequence (number of bases at the 5′ and 3′ ends not matched/total number of bases in the query sequence) so 10% is subtracted from the percent identity score calculated by the FASTDB program. If the remaining 90 bases were perfectly matched the final percent identity would be 90%. In another example, a 90 base subject sequence is compared with a 100 base query sequence. This time the deletions are internal deletions so that there are no bases on the 5′ or 3′ of the subject sequence which are not matched/aligned with the query. In this case the percent identity calculated by FASTDB is not manually corrected. Once again, only bases 5′ and 3′ of the subject sequence which are not matched/aligned with the query sequence are manually corrected for. No other manual corrections are to be made for the purposes of the present invention.

[0104] By a polypeptide having an amino acid sequence at least, for example, 95% “identical” to a query amino acid sequence of the present invention, it is intended that the amino acid sequence of the subject polypeptide is identical to the query sequence except that the subject polypeptide sequence may include up to five amino acid alterations per each 100 amino acids of the query amino acid sequence. In other words, to obtain a polypeptide having an amino acid sequence at least 95% identical to a query amino acid sequence, up to 5% of the amino acid residues in the subject sequence may be inserted, deleted, (indels) or substituted with another amino acid. These alterations of the reference sequence may occur at the amino or carboxy terminal positions of the reference amino acid sequence or anywhere between those terminal positions, interspersed either individually among residues in the reference sequence or in one or more contiguous groups within the reference sequence.

[0105] As a practical matter, whether any particular polypeptide is at least 80%, 85%, 90%, 95%, 96%, 97%, 98% or 99% identical to, for instance, the amino acid sequence of a polypeptide referred to in Table 1A (e.g., the amino acid sequence identified in column 6) or Table 2 (e.g., the amino acid sequence of the polypeptide encoded by the polynucleotide sequence defined in columns 8 and 9 of Table 2) or a fragment thereof, the amino acid sequence of the polypeptide encoded by the polynucleotide sequence in SEQ ID NO:B as defined in column 6 of Table 1B or a fragment thereof, the amino acid sequence of the polypeptide encoded by the nucleotide sequence in SEQ ID NO:X or a fragment thereof, or the amino acid sequence of the polypeptide encoded by cDNA contained in Clone ID NO:Z, or a fragment thereof, can be determined conventionally using known computer programs. A preferred method for determining the best overall match between a query sequence (a sequence of the present invention) and a subject sequence, also referred to as a global sequence alignment, can be determined using the FASTDB computer program based on the algorithm of Brutlag et al. (Comp. App. Biosci.6:237-245 (1990)). In a sequence alignment the query and subject sequences are either both nucleotide sequences or both amino acid sequences. The result of said global sequence alignment is expressed as percent identity. Preferred parameters used in a FASTDB amino acid alignment are: Matrix=PAM 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 amino acid sequence, whichever is shorter.

[0106] If the subject sequence is shorter than the query sequence due to N- or C-terminal deletions, not because of internal deletions, a manual correction must be made to the results. This is because 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 is corrected by calculating the number of residues of the query sequence that are N- and C-terminal of the subject sequence, which are not matched/aligned with a corresponding subject residue, as a percent of the total bases of the query sequence. Whether a residue is matched/aligned is determined by results of the FASTDB sequence alignment. This percentage is then subtracted from the percent identity, calculated by the above FASTDB program using the specified parameters, to arrive at a final percent identity score. This final percent identity score is what is used for the purposes of the present invention. Only residues to the N- and C-termini of the subject sequence, which are not matched/aligned with the query sequence, are 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.

[0107] For example, a 90 amino acid residue subject sequence is 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% is subtracted from the percent identity score calculated by the FASTDB program. If the remaining 90 residues were perfectly matched the final percent identity would be 90%. In another example, a 90 residue subject sequence is compared with a 100 residue query sequence. This time the deletions are internal deletions so there are no residues at the N- or C-termini of the subject sequence which are not matched/aligned with the query. In this case the percent identity calculated by FASTDB is 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 are not matched/aligned with the query sequnce are manually corrected for. No other manual corrections are to made for the purposes of the present invention.

[0108] The polynucleotide variants of the invention may contain alterations in the coding regions, non-coding regions, or both. Especially preferred are polynucleotide variants containing alterations which produce silent substitutions, additions, or deletions, but do not alter the properties or activities of the encoded polypeptide. Nucleotide variants produced by silent substitutions due to the degeneracy of the genetic code are preferred. Moreover, polypeptide variants in which less than 50, less than 40, less than 30, less than 20, less than 10, or 5-50, 5-25, 5-10, 1-5, or 1-2 amino acids are substituted, deleted, or added in any combination are also preferred. Polynucleotide variants can be produced for a variety of reasons, e.g., to optimize codon expression for a particular host (change codons in the human mRNA to those preferred by a bacterial host such as E. coli).

[0109] Naturally occurring variants are called “allelic variants,” and refer to one of several alternate forms of a gene occupying a given locus on a chromosome of an organism. (Genes II, Lewin, B., ed., John Wiley & Sons, New York (1985)). These allelic variants can vary at either the polynucleotide and/or polypeptide level and are included in the present invention. Alternatively, non-naturally occurring variants may be produced by mutagenesis techniques or by direct synthesis.

[0110] Using known methods of protein engineering and recombinant DNA technology, variants may be generated to improve or alter the characteristics of the polypeptides of the present invention. For instance, one or more amino acids can be deleted from the N-terminus or C-terminus of the polypeptide of the present invention without substantial loss of biological function. As an example, Ron et al. (J. Biol. Chem. 268: 2984-2988 (1993)) reported variant KGF proteins having heparin binding activity even after deleting 3, 8, or 27 amino-terminal amino acid residues. Similarly, Interferon gamma exhibited up to ten times higher activity after deleting 8-10 amino acid residues from the carboxy terminus of this protein. (Dobeli et al., J. Biotechnology 7:199-216 (1988).)

[0111] Moreover, ample evidence demonstrates that variants often retain a biological activity similar to that of the naturally occurring protein. For example, Gayle and coworkers (J. Biol. Chem. 268:22105-22111 (1993)) conducted extensive mutational analysis of human cytokine IL-1a. They used random mutagenesis to generate over 3,500 individual IL-1a mutants that averaged 2.5 amino acid changes per variant over the entire length of the molecule. Multiple mutations were examined at every possible amino acid position. The investigators found that “[m]ost of the molecule could be altered with little effect on either [binding or biological activity].” In fact, only 23 unique amino acid sequences, out of more than 3,500 nucleotide sequences examined, produced a protein that significantly differed in activity from wild-type.

[0112] Furthermore, even if deleting one or more amino acids from the N-terminus or C-terminus of a polypeptide results in modification or loss of one or more biological functions, other biological activities may still be retained. For example, the ability of a deletion variant to induce and/or to bind antibodies which recognize the secreted form will likely be retained when less than the majority of the residues of the secreted form are removed from the N-terminus or C-terminus. Whether a particular polypeptide lacking N- or C-terminal residues of a protein retains such immunogenic activities can readily be determined by routine methods described herein and otherwise known in the art.

[0113] Thus, the invention further includes polypeptide variants which show a functional activity (e.g., biological activity) of the polypeptides of the invention. Such variants include deletions, insertions, inversions, repeats, and substitutions selected according to general rules known in the art so as have little effect on activity.

[0114] The present application is directed to nucleic acid molecules at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to the nucleic acid sequences disclosed herein, (e.g., encoding a polypeptide having the amino acid sequence of an N and/or C terminal deletion), irrespective of whether they encode a polypeptide having functional activity. This is because even where a particular nucleic acid molecule does not encode a polypeptide having functional activity, one of skill in the art would still know how to use the nucleic acid molecule, for instance, as a hybridization probe or a polymerase chain reaction (PCR) primer. Uses of the nucleic acid molecules of the present invention that do not encode a polypeptide having functional activity include, inter alia, (1) isolating a gene or allelic or splice variants thereof in a cDNA library; (2) in situ hybridization (e.g., “FISH”) to metaphase chromosomal spreads to provide precise chromosomal location of the gene, as described in Verma et al., Human Chromosomes: A Manual of Basic Techniques, Pergamon Press, New York (1988); (3) Northern Blot analysis for detecting mRNA expression in specific tissues (e.g., normal or diseased tissues); and (4) in situ hybridization (e.g., histochemistry) for detecting mRNA expression in specific tissues (e.g., normal or diseased tissues).

[0115] Preferred, however, are nucleic acid molecules having sequences at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99% or 100% identical to the nucleic acid sequences disclosed herein, which do, in fact, encode a polypeptide having functional activity. By a polypeptide having “functional activity” is meant, a polypeptide capable of displaying one or more known functional activities associated with a full-length (complete) protein of the invention. Such functional activities include, but are not limited to, biological activity, antigenicity [ability to bind (or compete with a polypeptide of the invention for binding) to an anti-polypeptide of the invention antibody], immunogenicity (ability to generate antibody which binds to a specific polypeptide of the invention), ability to form multimers with polypeptides of the invention, and ability to bind to a receptor or ligand for a polypeptide of the invention.

[0116] The functional activity of the polypeptides, and fragments, variants and derivatives of the invention, can be assayed by various methods.

[0117] For example, in one embodiment where one is assaying for the ability to bind or compete with a full-length polypeptide of the present invention for binding to an anti-polypetide antibody, various immunoassays known in the art can be used, including but not limited to, competitive and non-competitive assay systems using techniques such as radioimmunoassays, ELISA (enzyme linked immunosorbent assay), “sandwich” immunoassays, immunoradiometric assays, gel diffusion precipitation reactions, immunodiffusion assays, in situ immunoassays (using colloidal gold, enzyme or radioisotope labels, for example), western blots, precipitation reactions, agglutination assays (e.g., gel agglutination assays, hemagglutination assays), complement fixation assays, immunofluorescence assays, protein A assays, and immunoelectrophoresis assays, etc. In one embodiment, antibody binding is detected by detecting a label on the primary antibody. In another embodiment, the primary antibody is detected by detecting binding of a secondary antibody or reagent to the primary antibody. In a further embodiment, the secondary antibody is labeled. Many means are known in the art for detecting binding in an immunoassay and are within the scope of the present invention.

[0118] In another embodiment, where a ligand is identified, or the ability of a polypeptide fragment, variant or derivative of the invention to multimerize is being evaluated, binding can be assayed, e.g., by means well-known in the art, such as, for example, reducing and non-reducing gel chromatography, protein affinity chromatography, and affinity blotting. See generally, Phizicky et al., Microbiol. Rev. 59:94-123 (1995). In another embodiment, the ability of physiological correlates of a polypeptide of the present invention to bind to a substrate(s) of the polypeptide of the invention can be routinely assayed using techniques known in the art.

[0119] In addition, assays described herein (see Examples) and otherwise known in the art may routinely be applied to measure the ability of polypeptides of the present invention and fragments, variants and derivatives thereof to elicit polypeptide related biological activity (either in vitro or in vivo). Other methods will be known to the skilled artisan and are within the scope of the invention.

[0120] Of course, due to the degeneracy of the genetic code, one of ordinary skill in the art will immediately recognize that a large number of the nucleic acid molecules having a sequence at least 80%, 85%, 90%, 95%, 96%, 97%, 98%, 99%, or 100% identical to, for example, the nucleic acid sequence of the cDNA contained in Clone ID NO:Z, the nucleic acid sequence referred to in Table 1A (SEQ ID NO:X), the nucleic acid sequence disclosed in Table 2 (e.g,. the nucleic acid sequence delineated in columns 8 and 9) or fragments thereof, will encode polypeptides “having functional activity.” In fact, since degenerate variants of any of these nucleotide sequences all encode the same polypeptide, in many instances, this will be clear to the skilled artisan even without performing the above described comparison assay. It will be further recognized in the art that, for such nucleic acid molecules that are not degenerate variants, a reasonable number will also encode a polypeptide having functional activity. This is because the skilled artisan is fully aware of amino acid substitutions that are either less likely or not likely to significantly effect protein function (e.g., replacing one aliphatic amino acid with a second aliphatic amino acid), as further described below.

[0121] For example, guidance concerning how to make phenotypically silent amino acid substitutions is provided in Bowie et al., “Deciphering the Message in Protein Sequences: Tolerance to Amino Acid Substitutions,” Science 247:1306-1310 (1990), wherein the authors indicate that there are two main strategies for studying the tolerance of an amino acid sequence to change.

[0122] The first strategy exploits the tolerance of amino acid substitutions by natural selection during the process of evolution. By comparing amino acid sequences in different species, conserved amino acids can be identified. These conserved amino acids are likely important for protein function. In contrast, the amino acid positions where substitutions have been tolerated by natural selection indicates that these positions are not critical for protein function. Thus, positions tolerating amino acid substitution could be modified while still maintaining biological activity of the protein.

[0123] The second strategy uses genetic engineering to introduce amino acid changes at specific positions of a cloned gene to identify regions critical for protein function. For example, site directed mutagenesis or alanine-scanning mutagenesis (introduction of single alanine mutations at every residue in the molecule) can be used. See Cunningham and Wells, Science 244:1081-1085 (1989). The resulting mutant molecules can then be tested for biological activity.

[0124] As the authors state, these two strategies have revealed that proteins are surprisingly tolerant of amino acid substitutions. The authors further indicate which amino acid changes are likely to be permissive at certain amino acid positions in the protein. For example, most buried (within the tertiary structure of the protein) amino acid residues require nonpolar side chains, whereas few features of surface side chains are generally conserved. Moreover, tolerated conservative amino acid substitutions involve replacement of the aliphatic or hydrophobic amino acids Ala, Val, Leu and Ile; replacement of the hydroxyl residues Ser and Thr; replacement of the acidic residues Asp and Glu; replacement of the amide residues Asn and Gln, replacement of the basic residues Lys, Arg, and His; replacement of the aromatic residues Phe, Tyr, and Trp, and replacement of the small-sized amino acids Ala, Ser, Thr, Met, and Gly. Besides conservative amino acid substitution, variants of the present invention include (i) substitutions with one or more of the non-conserved amino acid residues, where the substituted amino acid residues may or may not be one encoded by the genetic code, or (ii) substitutions with one or more of the amino acid residues having a substituent group, or (iii) fusion of the mature polypeptide with another compound, such as a compound to increase the stability and/or solubility of the polypeptide (for example, polyethylene glycol), (iv) fusion of the polypeptide with additional amino acids, such as, for example, an IgG Fc fusion region peptide, serum albumin (preferably human serum albumin) or a fragment thereof, or leader or secretory sequence, or a sequence facilitating purification, or (v) fusion of the polypeptide with another compound, such as albumin (including but not limited to recombinant albumin (see, e.g., U.S. Pat. No. 5,876,969, issued Mar. 2, 1999, EP Patent 0 413 622, and U.S. Pat. No. 5,766,883, issued Jun. 16, 1998, herein incorporated by reference in their entirety)). Such variant polypeptides are deemed to be within the scope of those skilled in the art from the teachings herein.

[0125] For example, polypeptide variants containing amino acid substitutions of charged amino acids with other charged or neutral amino acids may produce proteins with improved characteristics, such as less aggregation. Aggregation of pharmaceutical formulations both reduces activity and increases clearance due to the aggregate's immunogenic activity. See Pinckard et al., Clin. Exp. Immunol. 2:331-340 (1967); Robbins et al., Diabetes 36: 838-845 (1987); Cleland et al., Crit. Rev. Therapeutic Drug Carrier Systems 10:307-377 (1993).

[0126] A further embodiment of the invention relates to polypeptides which comprise the amino acid sequence of a polypeptide having an amino acid sequence which contains at least one amino acid substitution, but not more than 50 amino acid substitutions, even more preferably, not more than 40 amino acid substitutions, still more preferably, not more than 30 amino acid substitutions, and still even more preferably, not more than 20 amino acid substitutions from a polypeptide sequence disclosed herein. Of course it is highly preferable for a polypeptide to have an amino acid sequence which comprises the amino acid sequence of a polypeptide of SEQ ID NO:Y, an amino acid sequence encoded by SEQ ID NO:X, an amino acid sequence encoded by the portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2, an amino acid sequence encoded by the complement of SEQ ID NO:X, and/or an amino acid sequence encoded by cDNA contained in Clone ID NO:Z which contains, in order of ever-increasing preference, at least one, but not more than 10, 9, 8, 7, 6, 5, 4, 3, 2 or 1 amino acid substitutions.

[0127] In specific embodiments, the polypeptides of the invention comprise, or alternatively, consist of, fragments or variants of a reference amino acid sequence selected from: (a) the amino acid sequence of SEQ ID NO:Y or fragments thereof (e.g., the mature form and/or other fragments described herein); (b) the amino acid sequence encoded by SEQ ID NO:X or fragments thereof; (c) the amino acid sequence encoded by the complement of SEQ ID NO:X or fragments thereof; (d) the amino acid sequence encoded by the portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2 or fragments thereof; and (e) the amino acid sequence encoded by cDNA contained in Clone ID NO:Z or fragments thereof; wherein the fragments or variants have 1-5,5-10, 5-25, 5-50, 10-50 or 50-150, amino acid residue additions, substitutions, and/or deletions when compared to the reference amino acid sequence. In preferred embodiments, the amino acid substitutions are conservative. Polynucleotides encoding these polypeptides are also encompassed by the invention.

[0128] Polynucleotide and Polypeptide Fragments

[0129] The present invention is also directed to polynucleotide fragments of the polynucleotides (nucleic acids) of the invention. In the present invention, a “polynucleotide fragment” refers to a polynucleotide having a nucleic acid sequence which, for example: is a portion of the cDNA contained in Clone ID NO:Z or the complementary strand thereto; is a portion of the polynucleotide sequence encoding the polypeptide encoded by the cDNA contained in Clone ID NO:Z or the complementary strand thereto; is a portion of a polynucleotide sequence encoding the amino acid sequence encoded by the region of SEQ ID NO:X as defined in columns 8 and 9 of Table 2 or the complementary strand thereto; is a portion of the polynucleotide sequence of SEQ ID NO:X as defined in columns 8 and 9 of Table 2 or the complementary strand thereto; is a portion of the polynucleotide sequence in SEQ ID NO:X or the complementary strand thereto; is a polynucleotide sequence encoding a portion of the polypeptide of SEQ ID NO:Y; is a polynucleotide sequence encoding a portion of a polypeptide encoded by SEQ ID NO:X; is a polynucleotide sequence encoding a portion of a polypeptide encoded by the complement of the polynucleotide sequence in SEQ ID NO:X; is a portion of a polynucleotide sequence encoding the amino acid sequence encoded by the region of SEQ ID NO:B as defined in column 6 of Table 1B or the complementary strand thereto; or is a portion of the polynucleotide sequence of SEQ ID NO:B as defined in column 6 of Table 1B or the complementary strand thereto.

[0130] The polynucleotide fragments of the invention are preferably at least about 15 nt, and more preferably at least about 20 nt, still more preferably at least about 30 nt, and even more preferably, at least about 40 nt, at least about 50 nt, at least about 75 nt, or at least about 150 nt in length. A fragment “at least 20 nt in length,” for example, is intended to include 20 or more contiguous bases from the cDNA sequence contained in Clone ID NO:Z, or the nucleotide sequence shown in SEQ ID NO:X or the complementary stand thereto. In this context “about” includes the particularly recited value or a value larger or smaller by several (5, 4, 3, 2, or 1) nucleotides, at either terminus or at both termini. These nucleotide fragments have uses that include, but are not limited to, as diagnostic probes and primers as discussed herein. Of course, larger fragments (e.g., at least 160, 170, 180, 190, 200, 250, 500, 600, 1000, or 2000 nucleotides in length) are also encompassed by the invention.

[0131] Moreover, representative examples of polynucleotide fragments of the invention comprise, or alternatively consist of, a sequence from about nucleotide number 1-50, 51-100, 101-150, 151-200, 201-250, 251-300, 301-350, 351-400, 401-450, 451-500, 501-550, 551-600, 601-650, 651-700, 701-750, 751-800, 801-850, 851-900, 901-950, 951-1000, 1001-1050, 1051-1100, 1101-1150, 1151-1200, 1201-1250, 1251-1300, 1301-1350, 1351-1400, 1401-1450, 1451-1500, 1501-1550, 1551-1600, 1601-1650, 1651-1700, 1701-1750, 1751-1800, 1801-1850, 1851-1900, 1901-1950, 1951-2000, 2001-2050, 2051-2100, 2101-2150, 2151-2200, 2201-2250, 2251-2300, 2301-2350, 2351-2400, 2401-2450, 2451-2500, 2501-2550, 2551-2600, 2601-2650, 2651-2700, 2701-2750, 2751-2800, 2801-2850, 2851-2900, 2901-2950, 2951-3000, 3001-3050, 3051-3100, 3101-3150, 3151-3200, 3201-3250, 3251-3300, 3301-3350, 3351-3400, 3401-3450, 3451-3500, 3501-3550, 3551-3600, 3601-3650, 3651-3700, 3701-3750, 3751-3800, 3801-3850, 3851-3900, 3901-3950, 3951-4000, 4001-4050, 4051-4100, 4101-4150, 4151-4200, 4201-4250, 4251-4300, 4301-4350, 4351-4400, 4401-4450, 4451-4500, 4501-4550, 4551-4600, 4601-4650, 4651-4700, 4701-4750, 4751-4800, 4801-4850, 4851-4900, 4901-4950, 4951-5000, 5001-5050, 5051-5100, 5101-5150, 5151-5200, 5201-5250, 5251-5300, 5301-5350, 5351-5400, 5401-5450, 5451-5500, 5501-5550, 5551-5600, 5601-5650, 5651-5700, 5701-5750, 5751-5800, 5801-5850, 5851-5900, 5901-5950, 5951-6000, 6001-6050, 6051-6100, 6101-6150, 6151-6200, 6201-6250, 6251-6300, 6301-6350, 6351-6400, 6401-6450, 6451-6500, 6501-6550, 6551-6600, 6601-6650, 6651-6700, 6701-6750, 6751-6800, 6801-6850, 6851-6900, 6901-6950, 6951-7000, 7001-7050, 7051-7100, 7101-7150, 7151-7200, 7201-7250, 7251-7300 or 7301 to the end of SEQ ID NO:X, or the complementary strand thereto. In this context “about” includes the particularly recited range or a range larger or smaller by several (5, 4, 3, 2, or 1) nucleotides, at either terminus or at both termini. Preferably, these fragments encode a polypeptide which has a functional activity (e.g., biological activity). More preferably, these polynucleotides can be used as probes or primers as discussed herein. Polynucleotides which hybridize to one or more of these polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions are also encompassed by the invention, as are polypeptides encoded by these polynucleotides.

[0132] Further representative examples of polynucleotide fragments of the invention comprise, or alternatively consist of, a sequence from about nucleotide number 1-50, 51-100, 101-150, 151-200, 201-250, 251-300, 301-350, 351-400, 401-450, 451-500, 501-550, 551-600, 601-650, 651-700, 701-750, 751-800, 801-850, 851-900, 901-950, 951-1000, 1001-1050, 1051-1100, 1101-1150, 1151-1200, 1201-1250, 1251-1300, 1301-1350, 1351-1400, 1401-1450, 1451-1500, 1501-1550, 1551-1600, 1601-1650, 1651-1700, 1701-1750, 1751-1800, 1801-1850, 1851-1900, 1901-1950, 1951-2000, 2001-2050, 2051-2100, 2101-2150, 2151-2200, 2201-2250, 2251-2300, 2301-2350, 2351-2400, 2401-2450, 2451-2500, 2501-2550, 2551-2600, 2601-2650, 2651-2700, 2701-2750, 2751-2800, 2801-2850, 2851-2900, 2901-2950, 2951-3000, 3001-3050, 3051-3100, 3101-3150, 3151-3200, 3201-3250, 3251-3300, 3301-3350, 3351-3400, 3401-3450, 3451-3500, 3501-3550, 3551-3600, 3601-3650, 3651-3700, 3701-3750, 3751-3800, 3801-3850, 3851-3900, 3901-3950, 3951-4000, 4001-4050, 4051-4100, 4101-4150, 4151-4200, 4201-4250, 4251-4300, 4301-4350, 4351-4400, 4401-4450, 4451-4500, 4501-4550, 4551-4600, 4601-4650, 4651-4700, 4701-4750, 4751-4800, 4801-4850, 4851-4900, 4901-4950, 4951-5000, 5001-5050, 5051-5100, 5101-5150, 5151-5200, 5201-5250, 5251-5300, 5301-5350, 5351-5400, 5401-5450, 5451-5500, 5501-5550, 5551-5600, 5601-5650, 5651-5700, 5701-5750, 5751-5800, 5801-5850, 5851-5900, 5901-5950, 5951-6000, 6001-6050, 6051-6100, 6101-6150, 6151-6200, 6201-6250, 6251-6300, 6301-6350, 6351-6400, 6401-6450, 6451-6500, 6501-6550, 6551-6600, 6601-6650, 6651-6700, 6701-6750, 6751-6800, 6801-6850, 6851-6900, 6901-6950, 6951-7000, 7001-7050, 7051-7100, 7101-7150, 7151-7200, 7201-7250, 7251-7300 or 7301 to the end of the cDNA sequence contained in Clone ID NO:Z, or the complementary strand thereto. In this context “about” includes the particularly recited range or a range larger or smaller by several (5, 4, 3, 2, or 1) nucleotides, at either terminus or at both termini. Preferably, these fragments encode a polypeptide which has a functional activity (e.g., biological activity). More preferably, these polynucleotides can be used as probes or primers as discussed herein. Polynucleotides which hybridize to one or more of these polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions are also encompassed by the invention, as are polypeptides encoded by these polynucleotides.

[0133] Moreover, representative examples of polynucleotide fragments of the invention comprise, or alternatively consist of, a nucleic acid sequence comprising one, two, three, four, five, six, seven, eight, nine, ten, or more of the above described polynucleotide fragments of the invention in combination with a polynucleotide sequence delineated in Table 1B column 6. Additional, representative examples of polynucleotide fragments of the invention comprise, or alternatively consist of, a nucleic acid sequence comprising one, two, three, four, five, six, seven, eight, nine, ten, or more of the above described polynucleotide fragments of the invention in combination with a polynucleotide sequence that is the complementary strand of a sequence delineated in column 6 of Table 1B. In further embodiments, the above-described polynucleotide fragments of the invention comprise, or alternatively consist of, sequences delineated in Table 1B, column 6, and have a nucleic acid sequence which is different from that of the BAC fragment having the sequence disclosed in SEQ ID NO:B (see Table 1B, column 5). In additional embodiments, the above-described polynucleotide fragments of the invention comprise, or alternatively consist of, sequences delineated in Table 1B, column 6, and have a nucleic acid sequence which is different from that published for the BAC clone identified as BAC ID NO:A (see Table 1B, column 4). In additional embodiments, the above-described polynucleotides of the invention comprise, or alternatively consist of, sequences delineated Table 1B, column 6, and have a nucleic acid sequence which is different from that contained in the BAC clone identified as BAC ID NO:A (see Table 1B, column 4). Polypeptides encoded by these polynucleotides, other polynucleotides that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides and polypeptides are also encompassed by the invention.

[0134] In additional specific embodiments, polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more fragments of the sequences delineated in column 6 of Table 1B, and the polynucleotide sequence of SEQ ID NO:X (e.g., as defined in Table 1B, column 2) or fragments or variants thereof. Polypeptides encoded by these polynucleotides, other polynucleotides that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention.

[0135] In additional specific embodiments, polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more fragments of the sequences delineated in column 6 of Table 1B which correspond to the same Clone ID NO:Z (see Table 1B, column 1), and the polynucleotide sequence of SEQ ID NO:X (e.g., as defined in Table 1A or 1B) or fragments or variants thereof. Polypeptides encoded by these polynucleotides, other polynucleotides that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention.

[0136] In further specific embodiments, polynucleotides of the invention comprise, or alternatively consist of, one, two, three, four, five, six, seven, eight, nine, ten, or more fragments of the sequences delineated in the same row of column 6 of Table 1B, and the polynucleotide sequence of SEQ ID NO:X (e.g., as defined in Table 1A or 1B) or fragments or variants thereof. Polypeptides encoded by these polynucleotides, other polynucleotides that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention.

[0137] In additional specific embodiments, polynucleotides of the invention comprise, or alternatively consist of a polynucleotide sequence in which the 3′ 10 polynucleotides of one of the sequences delineated in column 6 of Table 1B and the 5′ 10 polynucleotides of the sequence of SEQ ID NO:X are directly contiguous. Nucleic acids which hybridize to the complement of these 20 contiguous polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention. Polypeptides encoded by these polynucleotides and/or nucleic acids, other polynucleotides and/or nucleic acids that encode these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides, nucleic acids, and polypeptides are also encompassed by the invention.

[0138] In additional specific embodiments, polynucleotides of the invention comprise, or alternatively consist of a polynucleotide sequence in which the 3′ 10 polynucleotides of one of the sequences delineated in column 6 of Table 1B and the 5′ 10 polynucleotides of a fragment or variant of the sequence of SEQ ID NO:X (e.g., as described herein) are directly contiguous Nucleic acids which hybridize to the complement of these 20 contiguous polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention. Polypeptides encoded by these polynucleotides and/or nucleic acids, other polynucleotides and/or nucleic acids encoding these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides, nucleic acids, and polypeptides are also encompassed by the invention.

[0139] In further specific embodiments, polynucleotides of the invention comprise, or alternatively consist of a polynucleotide sequence in which the 3′ 10 polynucleotides of a fragment or variant of the sequence of SEQ ID NO:X and the 5′ 10 polynucleotides of the sequence of one of the sequences delineated in column 6 of Table 1B are directly contiguous. Nucleic acids which hybridize to the complement of these 20 contiguous polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention. Polypeptides encoded by these polynucleotides and/or nucleic acids, other polynucleotides and/or nucleic acids encoding these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides, nucleic acids, and polypeptides are also encompassed by the invention.

[0140] In specific embodiments, polynucleotides of the invention comprise, or alternatively consist of a polynucleotide sequence in which the 3′ 10 polynucleotides of one of the sequences delineated in column 6 of Table 1B and the 5′ 10 polynucleotides of another sequence in column 6 are directly contiguous. In preferred embodiments, the 3′ 10 polynucleotides of one of the sequences delineated in column 6 of Table 1B is directly contiguous with the 5′ 10 polynucleotides of the next sequential exon delineated in Table 1B, column 6. Nucleic acids which hybridize to the complement of these 20 contiguous polynucleotides under stringent hybridization conditions or alternatively, under lower stringency conditions, are also encompassed by the invention. Polypeptides encoded by these polynucleotides and/or nucleic acids, other polynucleotides and/or nucleic acids encoding these polypeptides, and antibodies that bind these polypeptides are also encompassed by the invention. Additionally, fragments and variants of the above-described polynucleotides, nucleic acids, and polypeptides are also encompassed by the invention.

[0141] In the present invention, a “polypeptide fragment” refers to an amino acid sequence which is a portion of that contained in SEQ ID NO:Y, a portion of an amino acid sequence encoded by the portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2, a portion of an amino acid sequence encoded by the polynucleotide sequence of SEQ ID NO:X, a portion of an amino acid sequence encoded by the complement of the polynucleotide sequence in SEQ ID NO:X, and/or a portion of an amino acid sequence encoded by the cDNA contained in Clone ID NO:Z. Protein (polypeptide) fragments may be “free-standing,” or comprised within a larger polypeptide of which the fragment forms a part or region, most preferably as a single continuous region. Representative examples of polypeptide fragments of the invention, include, for example, fragments comprising, or alternatively consisting of, from about amino acid number 1-20, 21-40, 41-60, 61-80, 81-100, 101-120, 121-140, 141-160, 161-180, 181-200, 201-220, 221-240, 241-260, 261-280, 281-300, 301-320, 321-340, 341-360, 361-380, 381-400, 401-420, 421-440, 441-460, 461-480, 481-500, 501-520, 521-540, 541-560, 561-580, 581-600, 601-620, 621-640, 641-660, 661-680, 681-700, 701-720, 721-740, 741-760, 761-780, 781-800, 801-820, 821-840, 841-860, 861-880, 881-900, 901-920, 921-940, 941-960, 961-980, 981-1000, 1001-1020, 1021-1040, 1041-1060, 1061-1080, 1081-1100, 1101-1120, 1121-1140, 1141-1160, 1161-1180, 1181-1200, 1201-1220, 1221-1240, 1241-1260, 1261-1280, 1281-1300, 1301-1320, 1321-1340, 1341-1360, 1361-1380, 1381-1400, 1401-1420, 1421-1440, or 1441 to the end of the coding region of cDNA and SEQ ID NO: Y. In a preferred embodiment, polypeptide fragments of the invention include, for example, fragments comprising, or alternatively consisting of, from about amino acid number 1-20, 21-40, 4L-60, 61-80, 81-100, 101-120, 121-140, 141-160, 161-180, 181-200, 201-220, 221-240, 241-260, 261-280, 281-300, 301-320, 321-340, 341-360, 361-380, 381-400, 401-420, 421-440, 441-460, 461-480, 481-500, 501-520, 521-540, 541-560, 561-580, 581-600, 601-620, 621-640, 641-660, 661-680, 681-700, 701-720, 721-740, 741-760, 761-780, 781-800, 801-820, 821-840, 841-860, 861-880, 881-900, 901-920, 921-940, 941-960, 961-980, 981-1000, 1001-1020, 1021-1040, 1041-1060, 1061-1080, 1081-1100, 1101-1120, 1121-1140, 1141-1160, 1161-1180, 1181-1200, 1201-1220, 1221-1240, 1241-1260, 1261-1280, 1281-1300, 1301-1320, 1321-1340, 1341-1360, 1361-1380, 1381-1400, 1401-1420, 1421-1440, or 1441 to the end of the coding region of SEQ ID NO:Y. Moreover, polypeptide fragments of the invention may be at least about 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 100, 110, 120, 130, 140, or 150 amino acids in length. In this context “about” includes the particularly recited ranges or values, or ranges or values larger or smaller by several (5, 4, 3, 2, or 1) amino acids, at either extreme or at both extremes. Polynucleotides encoding these polypeptide fragments are also encompassed by the invention.

[0142] Even if deletion of one or more amino acids from the N-terminus of a protein results in modification of loss of one or more biological functions of the protein, other functional activities (e.g., biological activities, ability to multimerize, ability to bind a ligand) may still be retained. For example, the ability of shortened muteins to induce and/or bind to antibodies which recognize the complete or mature forms of the polypeptides generally will be retained when less than the majority of the residues of the complete or mature polypeptide are removed from the N-terminus. Whether a particular polypeptide lacking N-terminal residues of a complete polypeptide retains such immunologic activities can readily be determined by routine methods described herein and otherwise known in the art. It is not unlikely that a mutein with a large number of deleted N-terminal amino acid residues may retain some biological or immunogenic activities. In fact, peptides composed of as few as six amino acid residues may often evoke an immune response.

[0143] Accordingly, polypeptide fragments include the secreted protein as well as the mature form. Further preferred polypeptide fragments include the secreted protein or the mature form having a continuous series of deleted residues from the amino or the carboxy terminus, or both. For example, any number of amino acids, ranging from 1-60, can be deleted from the amino terminus of either the secreted polypeptide or the mature form. Similarly, any number of amino acids, ranging from 1-30, can be deleted from the carboxy terminus of the secreted protein or mature form. Furthermore, any combination of the above amino and carboxy terminus deletions are preferred. Similarly, polynucleotides encoding these polypeptide fragments are also preferred.

[0144] The present invention further provides polypeptides having one or more residues deleted from the amino terminus of the amino acid sequence of a polypeptide disclosed herein (e.g., a polypeptide of SEQ ID NO:Y, a polypeptide encoded by the polynucleotide sequence contained in SEQ ID NO:X or the complement thereof, a polypeptide encoded by the portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2, a polypeptide encoded by the portion of SEQ ID NO:B as defined in column 6 of Table 1B, and/or a polypeptide encoded by the cDNA contained in Clone ID NO:Z). In particular, N-terminal deletions may be described by the general formula m−q, where q is a whole integer representing the total number of amino acid residues in a polypeptide of the invention (e.g., the polypeptide disclosed in SEQ ID NO:Y, or the polypeptide encoded by the portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2), and m is defined as any integer ranging from 2 to q−6. Polynucleotides encoding these polypeptides are also encompassed by the invention.

[0145] The present invention further provides polypeptides having one or more residues from the carboxy terminus of the amino acid sequence of a polypeptide disclosed herein (e.g., a polypeptide of SEQ ID NO:Y, a polypeptide encoded by the polynucleotide sequence contained in SEQ ID NO:X, a polypeptide encoded by the portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2, and/or a polypeptide encoded by the cDNA contained in Clone ID NO:Z). In particular, C-terminal deletions may be described by the general formula 1−n, where n is any whole integer ranging from 6 to q−1, and where n corresponds to the position of amino acid residue in a polypeptide of the invention. Polynucleotides encoding these polypeptides are also encompassed by the invention.

[0146] In addition, any of the above described N- or C-terminal deletions can be combined to produce a N- and C-terminal deleted polypeptide. The invention also provides polypeptides having one or more amino acids deleted from both the amino and the carboxyl termini, which may be described generally as having residues m-n of a polypeptide encoded by SEQ ID NO:X (e.g., including, but not limited to, the preferred polypeptide disclosed as SEQ ID NO:Y and the polypeptide encoded by the portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2), the cDNA contained in Clone ID NO:Z, and/or the complement thereof, where n and m are integers as described above. Polynucleotides encoding these polypeptides are also encompassed by the invention.

[0147] Also as mentioned above, even if deletion of one or more amino acids from the C-terminus of a protein results in modification of loss of one or more biological functions of the protein, other functional activities (e.g., biological activities, ability to multimerize, ability to bind a ligand) may still be retained. For example the ability of the shortened mutein to induce and/or bind to antibodies which recognize the complete or mature forms of the polypeptide generally will be retained when less than the majority of the residues of the complete or mature polypeptide are removed from the C-terminus. Whether a particular polypeptide lacking C-terminal residues of a complete polypeptide retains such immunologic activities can readily be determined by routine methods described herein and otherwise known in the art. It is not unlikely that a mutein with a large number of deleted C-terminal amino acid residues may retain some biological or immunogenic activities. In fact, peptides composed of as few as six amino acid residues may often evoke an immune response.

[0148] The present application is also directed to proteins containing polypeptides at least 80%, 85%, 90%, 95%, 96%, 97%, 98% or 99% identical to a polypeptide sequence set forth herein. In preferred embodiments, the application is directed to proteins containing polypeptides at least 80%, 85%, 90%, 95%, 96%, 97%, 98% or 99% identical to polypeptides having the amino acid sequence of the specific N- and C-terminal deletions. Polynucleotides encoding these polypeptides are also encompassed by the invention.

[0149] Any polypeptide sequence encoded by, for example, the polynucleotide sequences set forth as SEQ ID NO:X or the complement thereof, (presented, for example, in Tables 1A and 2), the cDNA contained in Clone ID NO:Z, or the polynucleotide sequence as defined in column 6 of Table 1B, may be analyzed to determine certain preferred regions of the polypeptide. For example, the amino acid sequence of a polypeptide encoded by a polynucleotide sequence of SEQ ID NO:X (e.g., the polypeptide of SEQ ID NO:Y and the polypeptide encoded by the portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2) or the cDNA contained in Clone ID NO:Z may be analyzed using the default parameters of the DNASTAR computer algorithm (DNASTAR, Inc., 1228 S. Park St., Madison, Wis. 53715 USA; http://www.dnastar.com/).

[0150] Polypeptide regions that may be routinely obtained using the DNASTAR computer algorithm include, but are not limited to, Garnier-Robson alpha-regions, beta-regions, turn-regions, and coil-regions; Chou-Fasman alpha-regions, beta-regions, and turn-regions; Kyte-Doolittle hydrophilic regions and hydrophobic regions; Eisenberg alpha- and beta-amphipathic regions; Karplus-Schulz flexible regions; Emini surface-forming regions; and Jameson-Wolf regions of high antigenic index. Among highly preferred polynucleotides of the invention in this regard are those that encode polypeptides comprising regions that combine several structural features, such as several (e.g., 1, 2, 3 or 4) of the features set out above.

[0151] Additionally, Kyte-Doolittle hydrophilic regions and hydrophobic regions, Emini surface-forming regions, and Jameson-Wolf regions of high antigenic index (i.e., containing four or more contiguous amino acids having an antigenic index of greater than or equal to 1.5, as identified using the default parameters of the Jameson-Wolf program) can routinely be used to determine polypeptide regions that exhibit a high degree of potential for antigenicity. Regions of high antigenicity are determined from data by DNASTAR analysis by choosing values which represent regions of the polypeptide which are likely to be exposed on the surface of the polypeptide in an environment in which antigen recognition may occur in the process of initiation of an immune response.

[0152] Preferred polypeptide fragments of the invention are fragments comprising, or alternatively, consisting of, an amino acid sequence that displays a functional activity (e.g. biological activity) of the polypeptide sequence of which the amino acid sequence is a fragment. By a polypeptide displaying a “functional activity” is meant a polypeptide capable of one or more known functional activities associated with a full-length protein, such as, for example, biological activity, antigenicity, immunogenicity, and/or multimerization, as described herein.

[0153] Other preferred polypeptide fragments are biologically active fragments. Biologically active fragments are those exhibiting activity similar, but not necessarily identical, to an activity of the polypeptide of the present invention. The biological activity of the fragments may include an improved desired activity, or a decreased undesirable activity.

[0154] In preferred embodiments, polypeptides of the invention comprise, or alternatively consist of, one, two, three, four, five or more of the antigenic fragments of the polypeptide of SEQ ID NO:Y, or portions thereof. Polynucleotides encoding these polypeptides are also encompassed by the invention.

[0155] The present invention encompasses polypeptides comprising, or alternatively consisting of, an epitope of: the polypeptide sequence shown in SEQ ID NO:Y; a polypeptide sequence encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide sequence encoded by the portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2; the polypeptide sequence encoded by the portion of SEQ ID NO:B as defined in column 6 of Table 1B or the complement thereto; the polypeptide sequence encoded by the cDNA contained in Clone ID NO:Z; or the polypeptide sequence encoded by a polynucleotide that hybridizes to the sequence of SEQ ID NO:X, the complement of the sequence of SEQ ID NO:X, the complement of a portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2, or the cDNA sequence contained in Clone ID NO:Z under stringent hybridization conditions or alternatively, under lower stringency hybridization as defined supra. The present invention further encompasses polynucleotide sequences encoding an epitope of a polypeptide sequence of the invention (such as, for example, the sequence disclosed in SEQ ID NO:X, or a fragment thereof), polynucleotide sequences of the complementary strand of a polynucleotide sequence encoding an epitope of the invention, and polynucleotide sequences which hybridize to the complementary strand under stringent hybridization conditions or alternatively, under lower stringency hybridization conditions defined supra.

[0156] The term “epitopes,” as used herein, refers to portions of a polypeptide having antigenic or immunogenic activity in an animal, preferably a mammal, and most preferably in a human. In a preferred embodiment, the present invention encompasses a polypeptide comprising an epitope, as well as the polynucleotide encoding this polypeptide. An “immunogenic epitope,” as used herein, is defined as a portion of a protein that elicits an antibody response in an animal, as determined by any method known in the art, for example, by the methods for generating antibodies described infra. (See, for example, Geysen et al., Proc. Natl. Acad. Sci. USA 81:3998-4002 (1983)). The term “antigenic epitope,” as used herein, is defined as a portion of a protein to which an antibody can immunospecifically bind its antigen as determined by any method well known in the art, for example, by the immunoassays described herein. Immunospecific binding excludes non-specific binding but does not necessarily exclude cross-reactivity with other antigens. Antigenic epitopes need not necessarily be immunogenic.

[0157] Fragments which function as epitopes may be produced by any conventional means. (See, e.g., Houghten, R. A., Proc. Natl. Acad. Sci. USA 82:5131-5135 (1985) further described in U.S. Pat. No. 4,631,211.)

[0158] In the present invention, antigenic epitopes preferably contain a sequence of at least 4, at least 5, at least 6, at least 7, more preferably at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 20, at least 25, at least 30, at least 40, at least 50, and, most preferably, between about 15 to about 30 amino acids. Preferred polypeptides comprising immunogenic or antigenic epitopes are at least 10, 15, 20, 25, 30, 35, 40, 45, 50, 55, 60, 65, 70, 75, 80, 85, 90, 95, or 100 amino acid residues in length. Additional non-exclusive preferred antigenic epitopes include the antigenic epitopes disclosed herein, as well as portions thereof. Antigenic epitopes are useful, for example, to raise antibodies, including monoclonal antibodies, that specifically bind the epitope. Preferred antigenic epitopes include the antigenic epitopes disclosed herein, as well as any combination of two, three, four, five or more of these antigenic epitopes. Antigenic epitopes can be used as the target molecules in immunoassays. (See, for instance, Wilson et al., Cell 37:767-778 (1984); Sutcliffe et al., Science 219:660-666 (1983)).

[0159] Non-limiting examples of epitopes of polypeptides that can be used to generate antibodies of the invention include a polypeptide comprising, or alternatively consisting of, at least one, two, three, four, five, six or more of the portion(s) of SEQ ID NO:Y specified in column 7 of Table 1A. These polypeptide fragments have been determined to bear antigenic epitopes of the proteins of the invention by the analysis of the Jameson-Wolf antigenic index which is included in the DNAStar suite of computer programs. By “comprise” it is intended that a polypeptide contains at least one, two, three, four, five, six or more of the portion(s) of SEQ ID NO:Y shown in column 7 of Table 1A, but it may contain additional flanking residues on either the amino or carboxyl termini of the recited portion. Such additional flanking sequences are preferably sequences naturally found adjacent to the portion; i.e., contiguous sequence shown in SEQ ID NO:Y. The flanking sequence may, however, be sequences from a heterolgous polypeptide, such as from another protein described herein or from a heterologous polypeptide not described herein. In particular embodiments, epitope portions of a polypeptide of the invention comprise one, two, three, or more of the portions of SEQ ID NO:Y shown in column 7 of Table 1A.

[0160] Similarly, immunogenic epitopes can be used, for example, to induce antibodies according to methods well known in the art. See, for instance, Sutcliffe et al., supra; Wilson et al., supra; Chow et al., Proc. Natl. Acad. Sci. USA 82:910-914; and Bittle et al., J. Gen. Virol. 66:2347-2354 (1985). Preferred immunogenic epitopes include the immunogenic epitopes disclosed herein, as well as any combination of two, three, four, five or more of these immunogenic epitopes. The polypeptides comprising one or more immunogenic epitopes may be presented for eliciting an antibody response together with a carrier protein, such as an albumin, to an animal system (such as rabbit or mouse), or, if the polypeptide is of sufficient length (at least about 25 amino acids), the polypeptide may be presented without a carrier. However, immunogenic epitopes comprising as few as 8 to 10 amino acids have been shown to be sufficient to raise antibodies capable of binding to, at the very least, linear epitopes in a denatured polypeptide (e.g., in Western blotting).

[0161] Epitope-bearing polypeptides of the present invention may be used to induce antibodies according to methods well known in the art including, but not limited to, in vivo immunization, in vitro immunization, and phage display methods. See, e.g., Sutcliffe et al., supra; Wilson et al., supra, and Bittle et al., J. Gen. Virol., 66:2347-2354 (1985). If in vivo immunization is used, animals may be immunized with free peptide; however, anti-peptide antibody titer may be boosted by coupling the peptide to a macromolecular carrier, such as keyhole limpet hemacyanin (KLH) or tetanus toxoid. For instance, peptides containing cysteine residues may be coupled to a carrier using a linker such as maleimidobenzoyl-N-hydroxysuccinimide ester (MBS), while other peptides may be coupled to carriers using a more general linking agent such as glutaraldehyde. Animals such as rabbits, rats and mice are immunized with either free or carrier-coupled peptides, for instance, by intraperitoneal and/or intradermal injection of emulsions containing about 100 μg of peptide or carrier protein and Freund's adjuvant or any other adjuvant known for stimulating an immune response. Several booster injections may be needed, for instance, at intervals of about two weeks, to provide a useful titer of anti-peptide antibody which can be detected, for example, by ELISA assay using free peptide adsorbed to a solid surface. The titer of anti-peptide antibodies in serum from an immunized animal may be increased by selection of anti-peptide antibodies, for instance, by adsorption to the peptide on a solid support and elution of the selected antibodies according to methods well known in the art.

[0162] As one of skill in the art will appreciate, and as discussed above, the polypeptides of the present invention (e.g., those comprising an immunogenic or antigenic epitope) can be fused to heterologous polypeptide sequences. For example, polypeptides of the present invention (including fragments or variants thereof), may be fused with the constant domain of immunoglobulins (IgA, IgE, IgG, IgM), or portions thereof (CH1, CH2, CH3, or any combination thereof and portions thereof, resulting in chimeric polypeptides. By way of another non-limiting example, polypeptides and/or antibodies of the present invention (including fragments or variants thereof) may be fused with albumin (including but not limited to recombinant human serum albumin or fragments or variants thereof (see, e.g., U.S. Pat. No. 5,876,969, issued Mar. 2, 1999, EP Patent 0 413 622, and U.S. Pat. No. 5,766,883, issued Jun. 16, 1998, herein incorporated by reference in their entirety)). In a preferred embodiment, polypeptides and/or antibodies of the present invention (including fragments or variants thereof) are fused with the mature form of human serum albumin (i.e., amino acids 1-585 of human serum albumin as shown in FIGS. 1 and 2 of EP Patent 0 322 094) which is herein incorporated by reference in its entirety. In another preferred embodiment, polypeptides and/or antibodies of the present invention (including fragments or variants thereof) are fused with polypeptide fragments comprising, or alternatively consisting of, amino acid residues 1−z of human serum albumin, where z is an integer from 369 to 419, as described in U.S. Pat. No. 5,766,883 herein incorporated by reference in its entirety. Polypeptides and/or antibodies of the present invention (including fragments or variants thereof) may be fused to either the N- or C-terminal end of the heterologous protein (e.g., immunoglobulin Fc polypeptide or human serum albumin polypeptide). Polynucleotides encoding fusion proteins of the invention are also encompassed by the invention.

[0163] Such fusion proteins as those described above may facilitate purification and may increase half-life in vivo. This has been shown for chimeric proteins consisting of the first two domains of the human CD4-polypeptide and various domains of the constant regions of the heavy or light chains of mammalian immunoglobulins. See, e.g., EP 394,827; Traunecker et al., Nature, 331:84-86 (1988). Enhanced delivery of an antigen across the epithelial barrier to the immune system has been demonstrated for antigens (e.g., insulin) conjugated to an FcRn binding partner such as IgG or Fc fragments (see, e.g., PCT Publications WO 96/22024 and WO 99/04813). IgG fusion proteins that have a disulfide-linked dimeric structure due to the IgG portion desulfide bonds have also been found to be more efficient in binding and neutralizing other molecules than monomeric polypeptides or fragments thereof alone. See, e.g., Fountoulakis et al., J. Biochem., 270:3958-3964 (1995). Nucleic acids encoding the above epitopes can also be recombined with a gene of interest as an epitope tag (e.g., the hemagglutinin (HA) tag or flag tag) to aid in detection and purification of the expressed polypeptide. For example, a system described by Janknecht et al. allows for the ready purification of non-denatured fusion proteins expressed in human cell lines (Janknecht et al., 1991, Proc. Natl. Acad. Sci. USA 88:8972-897). In this system, the gene of interest is subcloned into a vaccinia recombination plasmid such that the open reading frame of the gene is translationally fused to an amino-terminal tag consisting of six histidine residues. The tag serves as a matrix binding domain for the fusion protein. Extracts from cells infected with the recombinant vaccinia virus are loaded onto Ni2+ nitriloacetic acid-agarose column and histidine-tagged proteins can be selectively eluted with imidazole-containing buffers.

[0164] Fusion Proteins

[0165] Any polypeptide of the present invention can be used to generate fusion proteins. For example, the polypeptide of the present invention, when fused to a second protein, can be used as an antigenic tag. Antibodies raised against the polypeptide of the present invention can be used to indirectly detect the second protein by binding to the polypeptide. Moreover, because secreted proteins target cellular locations based on trafficking signals, polypeptides of the present invention which are shown to be secreted can be used as targeting molecules once fused to other proteins.

[0166] Examples of domains that can be fused to polypeptides of the present invention include not only heterologous signal sequences, but also other heterologous functional regions. The fusion does not necessarily need to be direct, but may occur through linker sequences.

[0167] In certain preferred embodiments, proteins of the invention are fusion proteins comprising an amino acid sequence that is an N and/or C-terminal deletion of a polypeptide of the invention. In preferred embodiments, the invention is directed to a fusion protein comprising an amino acid sequence that is at least 90%, 95%, 96%, 97%, 98% or 99% identical to a polypeptide sequence of the invention. Polynucleotides encoding these proteins are also encompassed by the invention.

[0168] Moreover, fusion proteins may also be engineered to improve characteristics of the polypeptide of the present invention. For instance, a region of additional amino acids, particularly charged amino acids, may be added to the N-terminus of the polypeptide to improve stability and persistence during purification from the host cell or subsequent handling and storage. Also, peptide moieties may be added to the polypeptide to facilitate purification. Such regions may be removed prior to final preparation of the polypeptide. The addition of peptide moieties to facilitate handling of polypeptides are familiar and routine techniques in the art.

[0169] As one of skill in the art will appreciate that, as discussed above, polypeptides of the present invention, and epitope-bearing fragments thereof, can be combined with heterologous polypeptide sequences. For example, the polypeptides of the present invention may be fused with heterologous polypeptide sequences, for example, the polypeptides of the present invention may be fused with the constant domain of immunoglobulins (IgA, IgE, IgG, IgM) or portions thereof (CH1, CH2, CH3, and any combination thereof, including both entire domains and portions thereof), or albumin (including, but not limited to, native or recombinant human albumin or fragments or variants thereof (see, e.g., U.S. Pat. No. 5,876,969, issued Mar. 2, 1999, EP Patent 0 413 622, and U.S. Pat. No. 5,766,883, issued Jun. 16, 1998, herein incorporated by reference in their entirety)), resulting in chimeric polypeptides. For example, EP-A-O 464 533 (Canadian counterpart 2045869) discloses fusion proteins comprising various portions of constant region of immunoglobulin molecules together with another human protein or part thereof. In many cases, the Fc part in a fusion protein is beneficial in therapy and diagnosis, and thus can result in, for example, improved pharmacokinetic properties (EP-A 0232 262). Alternatively, deleting the Fe part after the fusion protein has been expressed, detected, and purified, would be desired. For example, the Fe portion may hinder therapy and diagnosis if the fusion protein is used as an antigen for immunizations. In drug discovery, for example, human proteins, such as hIL-5, have been fused with Fe portions for the purpose of high-throughput screening assays to identify antagonists of hIL-5. See, D. Bennett et al., J. Molecular Recognition 8:52-58 (1995); K. Johanson et al., J. Biol. Chem. 270:9459-9471 (1995).

[0170] Moreover, the polypeptides of the present invention can be fused to marker sequences, such as a polypeptide which facilitates purification of the fused polypeptide. In preferred embodiments, the marker amino acid sequence is a hexa-histidine peptide, such as the tag provided in a pQE vector (QIAGEN, Inc., 9259 Eton Avenue, Chatsworth, Calif., 91311), among others, many of which are commercially available. As described in Gentz et al., Proc. Natl. Acad. Sci. USA 86:821-824 (1989), for instance, hexa-histidine provides for convenient purification of the fusion protein. Another peptide tag useful for purification, the “HA” tag, corresponds to an epitope derived from the influenza hemagglutinin protein (Wilson et al., Cell 37:767 (1984)).

[0171] Additional fusion proteins of the invention may be generated through the techniques of gene-shuffling, motif-shuffling, exon-shuffling, and/or codon-shuffling (collectively referred to as “DNA shuffling”). DNA shuffling may be employed to modulate the activities of polypeptides of the invention, such methods can be used to generate polypeptides with altered activity, as well as agonists and antagonists of the polypeptides. See, generally, U.S. Pat. Nos. 5,605,793; 5,811,238; 5,830,721; 5,834,252; and 5,837,458, and Patten et al., Curr. Opinion Biotechnol. 8:724-33 (1997); Harayama, Trends Biotechnol. 16(2):76-82 (1998); Hansson, et al., J. Mol. Biol. 287:265-76 (1999); and Lorenzo and Blasco, Biotechniques 24(2):308-13 (1998) (each of these patents and publications are hereby incorporated by reference in its entirety). In one embodiment, alteration of polynucleotides corresponding to SEQ ID NO:X and the polypeptides encoded by these polynucleotides may be achieved by DNA shuffling. DNA shuffling involves the assembly of two or more DNA segments by homologous or site-specific recombination to generate variation in the polynucleotide sequence. In another embodiment, polynucleotides of the invention, or the encoded polypeptides, may be altered by being subjected to random mutagenesis by error-prone PCR, random nucleotide insertion or other methods prior to recombination. In another embodiment, one or more components, motifs, sections, parts, domains, fragments, etc., of a polynucleotide encoding a polypeptide of the invention may be recombined with one or more components, motifs, sections, parts, domains, fragments, etc. of one or more heterologous molecules.

[0172] Thus, any of these above fusions can be engineered using the polynucleotides or the polypeptides of the present invention.

[0173] Recombinant and Synthetic Production of Polypeptides of the Invention

[0174] The present invention also relates to vectors containing the polynucleotide of the present invention, host cells, and the production of polypeptides by synthetic and recombinant techniques. The vector may be, for example, a phage, plasmid, viral, or retroviral vector. Retroviral vectors may be replication competent or replication defective. In the latter case, viral propagation generally will occur only in complementing host cells.

[0175] The polynucleotides of the invention may be joined to a vector containing a selectable marker for propagation in a host. Generally, a plasmid vector is introduced in a precipitate, such as a calcium phosphate precipitate, or in a complex with a charged lipid. If the vector is a virus, it may be packaged in vitro using an appropriate packaging cell line and then transduced into host cells.

[0176] The polynucleotide insert should be operatively linked to an appropriate promoter, such as the phage lambda PL promoter, the E. coli lac, trp, phoA and tac promoters, the SV40 early and late promoters and promoters of retroviral LTRs, to name a few. Other suitable promoters will be known to the skilled artisan. The expression constructs will further contain sites for transcription initiation, termination, and, in the transcribed region, a ribosome binding site for translation. The coding portion of the transcripts expressed by the constructs will preferably include a translation initiating codon at the beginning and a termination codon (UAA, UGA or UAG) appropriately positioned at the end of the polypeptide to be translated.

[0177] As indicated, the expression vectors will preferably include at least one selectable marker. Such markers include dihydrofolate reductase, G418, glutamine synthase, or neomycin resistance for eukaryotic cell culture, and tetracycline, kanamycin or ampicillin resistance genes for culturing in E. coli and other bacteria. Representative examples of appropriate hosts include, but are not limited to, bacterial cells, such as E. coli, Streptomyces and Salmonella typhimurium cells; fungal cells, such as yeast cells (e.g., Saccharomyces cerevisiae or Pichia pastoris (ATCC Accession No. 201178)); insect cells such as Drosophila S2 and Spodoptera Sf9 cells; animal cells such as CHO, COS, 293, and Bowes melanoma cells; and plant cells. Appropriate culture mediums and conditions for the above-described host cells are known in the art.

[0178] Among vectors preferred for use in bacteria include pQE70, pQE60 and pQE-9, available from QIAGEN, Inc.; pBluescript vectors, Phagescript vectors, pNH8A, pNH16a, pNH 18A, pNH46A, available from Stratagene Cloning Systems, Inc.; and ptrc99a, pKK223-3, pKK233-3, pDR540, pRIT5 available from Pharmacia Biotech, Inc. Among preferred eukaryotic vectors are pWLNEO, pSV2CAT, pOG44, pXT1 and pSG available from Stratagene; and pSVK3, pBPV, pMSG and pSVL available from Pharmacia. Preferred expression vectors for use in yeast systems include, but are not limited to pYES2, pYD1, pTEF1/Zeo, pYES2/GS, pPICZ, pGAPZ, pGAPZalph, pPIC9, pPIC3.5, pHIL-D2, pHIL-S1, pPIC3.5K, pPIC9K, and PA0815 (all available from Invitrogen, Carlbad, Calif.). Other suitable vectors will be readily apparent to the skilled artisan.

[0179] Vectors which use glutamine synthase (GS) or DHFR as the selectable markers can be amplified in the presence of the drugs methionine sulphoximine or methotrexate, respectively. An advantage of glutamine synthase based vectors are the availabilty of cell lines (e.g., the murine myeloma cell line, NS0) which are glutamine synthase negative. Glutamine synthase expression systems can also function in glutamine synthase expressing cells (e.g., Chinese Hamster Ovary (CHO) cells) by providing additional inhibitor to prevent the functioning of the endogenous gene. A glutamine synthase expression system and components thereof are detailed in PCT publications: WO87/04462; WO86/05807; WO89/01036; WO89/10404; and WO91/06657, which are hereby incorporated in their entireties by reference herein. Additionally, glutamine synthase expression vectors can be obtained from Lonza Biologics, Inc. (Portsmouth, N.H.). Expression and production of monoclonal antibodies using a GS expression system in murine myeloma cells is described in Bebbington et al., Bio/technology 10:169(1992) and in Biblia and Robinson Biotechnol. Prog. 11:1 (1995) which are herein incorporated by reference.

[0180] The present invention also relates to host cells containing the above-described vector constructs described herein, and additionally encompasses host cells containing nucleotide sequences of the invention that are operably associated with one or more heterologous control regions (e.g., promoter and/or enhancer) using techniques known of in the art. The host cell can be a higher eukaryotic cell, such as a mammalian cell (e.g., a human derived cell), or a lower eukaryotic cell, such as a yeast cell, or the host cell can be a prokaryotic cell, such as a bacterial cell. A host strain may be chosen which modulates the expression of the inserted gene sequences, or modifies and processes the gene product in the specific fashion desired. Expression from certain promoters can be elevated in the presence of certain inducers; thus expression of the genetically engineered polypeptide may be controlled. Furthermore, different host cells have characteristics and specific mechanisms for the translational and post-translational processing and modification (e.g., phosphorylation, cleavage) of proteins. Appropriate cell lines can be chosen to ensure the desired modifications and processing of the foreign protein expressed.

[0181] Introduction of the nucleic acids and nucleic acid constructs of the invention into the host cell can be effected by calcium phosphate transfection, DEAE-dextran mediated transfection, cationic lipid-mediated transfection, electroporation, transduction, infection, or other methods. Such methods are described in many standard laboratory manuals, such as Davis et al., Basic Methods In Molecular Biology (1986). It is specifically contemplated that the polypeptides of the present invention may in fact be expressed by a host cell lacking a recombinant vector.

[0182] In addition to encompassing host cells containing the vector constructs discussed herein, the invention also encompasses primary, secondary, and immortalized host cells of vertebrate origin, particularly mammalian origin, that have been engineered to delete or replace endogenous genetic material (e.g., the coding sequence), and/or to include genetic material (e.g., heterologous polynucleotide sequences) that is operably associated with polynucleotides of the invention, and which activates, alters, and/or amplifies endogenous polynucleotides. For example, techniques known in the art may be used to operably associate heterologous control regions (e.g., promoter and/or enhancer) and endogenous polynucleotide sequences via homologous recombination (see, e.g., U.S. Pat. No. 5,641,670, issued Jun. 24, 1997; International Publication Number WO 96/29411; International Publication Number WO 94/12650; Koller et al., Proc. Natl. Acad. Sci. USA 86:8932-8935 (1989); and Zijlstra et al., Nature 342:435-438 (1989), the disclosures of each of which are incorporated by reference in their entireties).

[0183] Polypeptides of the invention can be recovered and purified from recombinant cell cultures by well-known methods including ammonium sulfate or ethanol precipitation, acid extraction, anion or cation exchange chromatography, phosphocellulose chromatography, hydrophobic interaction chromatography, affinity chromatography, hydroxylapatite chromatography and lectin chromatography. Most preferably, high performance liquid chromatography (“HPLC”) is employed for purification.

[0184] Polypeptides of the present invention can also be recovered from: products purified from natural sources, including bodily fluids, tissues and cells, whether directly isolated or cultured; products of chemical synthetic procedures; and products produced by recombinant techniques from a prokaryotic or eukaryotic host, including, for example, bacterial, yeast, higher plant, insect, and mammalian cells. Depending upon the host employed in a recombinant production procedure, the polypeptides of the present invention may be glycosylated or may be non-glycosylated. In addition, polypeptides of the invention may also include an initial modified methionine residue, in some cases as a result of host-mediated processes. Thus, it is well known in the art that the N-terminal methionine encoded by the translation initiation codon generally is removed with high efficiency from any protein after translation in all eukaryotic cells. While the N-terminal methionine on most proteins also is efficiently removed in most prokaryotes, for some proteins, this prokaryotic removal process is inefficient, depending on the nature of the amino acid to which the N-terminal methionine is covalently linked.

[0185] In one embodiment, the yeast Pichia pastoris is used to express polypeptides of the invention in a eukaryotic system. Pichia pastoris is a methylotrophic yeast which can metabolize methanol as its sole carbon source. A main step in the methanol metabolization pathway is the oxidation of methanol to formaldehyde using O2. This reaction is catalyzed by the enzyme alcohol oxidase. In order to metabolize methanol as its sole carbon source, Pichia pastoris must generate high levels of alcohol oxidase due, in part, to the relatively low affinity of alcohol oxidase for O2. Consequently, in a growth medium depending on methanol as a main carbon source, the promoter region of one of the two alcohol oxidase genes (AOX1) is highly active. In the presence of methanol, alcohol oxidase produced from the AOX1 gene comprises up to approximately 30% of the total soluble protein in Pichia pastoris. See Ellis, S. B., et al., Mol. Cell. Biol. 5:1111-21 (1985); Koutz, P. J, et al., Yeast 5:167-77 (1989); Tschopp, J. F., et al., Nucl. Acids Res. 15:3859-76 (1987). Thus, a heterologous coding sequence, such as, for example, a polynucleotide of the present invention, under the transcriptional regulation of all or part of the AOX1 regulatory sequence is expressed at exceptionally high levels in Pichia yeast grown in the presence of methanol.

[0186] In one example, the plasmid vector pPIC9K is used to express DNA encoding a polypeptide of the invention, as set forth herein, in a Pichea yeast system essentially as described in “Pichia Protocols: Methods in Molecular Biology,” D. R. Higgins and J. Cregg, eds. The Humana Press, Totowa, N.J., 1998. This expression vector allows expression and secretion of a polypeptide of the invention by virtue of the strong AOX1 promoter linked to the Pichia pastoris alkaline phosphatase (PHO) secretory signal peptide (i.e., leader) located upstream of a multiple cloning site.

[0187] Many other yeast vectors could be used in place of pPIC9K, such as, pYES2, pYD1, pTEF1/Zeo, pYES2/GS, pPICZ, pGAPZ, pGAPZalpha, pPIC9, pPIC3.5, pHIL-D2, pHIL-S1, pPIC3.5K, and PAO815, as one skilled in the art would readily appreciate, as long as the proposed expression construct provides appropriately located signals for transcription, translation, secretion (if desired), and the like, including an in-frame AUG as required.

[0188] In another embodiment, high-level expression of a heterologous coding sequence, such as, for example, a polynucleotide of the present invention, may be achieved by cloning the heterologous polynucleotide of the invention into an expression vector such as, for example, pGAPZ or pGAPZalpha, and growing the yeast culture in the absence of methanol.

[0189] In addition to encompassing host cells containing the vector constructs discussed herein, the invention also encompasses primary, secondary, and immortalized host cells of vertebrate origin, particularly mammalian origin, that have been engineered to delete or replace endogenous genetic material (e.g., coding sequence), and/or to include genetic material (e.g., heterologous polynucleotide sequences) that is operably associated with polynucleotides of the invention, and which activates, alters, and/or amplifies endogenous polynucleotides. For example, techniques known in the art may be used to operably associate heterologous control regions (e.g., promoter and/or enhancer) and endogenous polynucleotide sequences via homologous recombination (see, e.g., U.S. Pat. No. 5,641,670, issued Jun. 24, 1997; International Publication No. WO 96/29411, published Sep. 26, 1996; International Publication No. WO 94/12650, published Aug. 4, 1994; Koller et al., Proc. Natl. Acad. Sci. USA 86:8932-8935 (1989); and Zijlstra et al., Nature 342:435-438 (1989), the disclosures of each of which are incorporated by reference in their entireties).

[0190] In addition, polypeptides of the invention can be chemically synthesized using techniques known in the art (e.g., see Creighton, 1983, Proteins: Structures and Molecular Principles, W. H. Freeman & Co., N.Y., and Hunkapiller et al., Nature, 310:105-111 (1984)). For example, a polypeptide corresponding to a fragment of a polypeptide can be synthesized by use of a peptide synthesizer. Furthermore, if desired, nonclassical amino acids or chemical amino acid analogs can be introduced as a substitution or addition into the polypeptide sequence. Non-classical amino acids include, but are not limited to, to the D-isomers of the common amino acids, 2,4-diaminobutyric acid, a-amino isobutyric acid, 4-aminobutyric acid, Abu, 2-amino butyric acid, g-Abu, e-Ahx, 6-amino hexanoic acid, Aib, 2-amino isobutyric acid, 3-amino propionic acid, omithine, norleucine, norvaline, hydroxyproline, sarcosine, citrulline, homocitrulline, cysteic acid, t-butylglycine, t-butylalanine, phenylglycine, cyclohexylalanine, b-alanine, fluoro-amino acids, designer amino acids such as b-methyl amino acids, Ca-methyl amino acids, Na-methyl amino acids, and amino acid analogs in general. Furthermore, the amino acid can be D (dextrorotary) or L (levorotary).

[0191] The invention encompasses polypeptides of the present invention which are differentially modified during or after translation, e.g., by glycosylation, acetylation, phosphorylation, amidation, derivatization by known protecting/blocking groups, proteolytic cleavage, linkage to an antibody molecule or other cellular ligand, etc. Any of numerous chemical modifications may be carried out by known techniques, including but not limited, to specific chemical cleavage by cyanogen bromide, trypsin, chymotrypsin, papain, V8 protease, NaBH4; acetylation, formylation, oxidation, reduction; metabolic synthesis in the presence of tunicamycin; etc.

[0192] Additional post-translational modifications encompassed by the invention include, for example, e.g., N-linked or O-linked carbohydrate chains, processing of N-terminal or C-terminal ends), attachment of chemical moieties to the amino acid backbone, chemical modifications of N-linked or O-linked carbohydrate chains, and addition or deletion of an N-terminal methionine residue as a result of procaryotic host cell expression. The polypeptides may also be modified with a detectable label, such as an enzymatic, fluorescent, isotopic or affinity label to allow for detection and isolation of the protein.

[0193] Examples of suitable enzymes include horseradish peroxidase, alkaline phosphatase, beta-galactosidase, or acetylcholinesterase; examples of suitable prosthetic group complexes include streptavidin/biotin and avidin/biotin; examples of suitable fluorescent materials include umbelliferone, fluorescein, fluorescein isothiocyanate, rhodamine, dichlorotriazinylamine fluorescein, dansyl chloride or phycoerythrin; an example of a luminescent material includes luminol; examples of bioluminescent materials include luciferase, luciferin, and aequorin; and examples of suitable radioactive material include iodine (121I, 123I, 125I, 131I), carbon (14C), sulfur (35S), tritium (3H), indium (111In, 112In, 113mIn, 115mIn), technetium (99Tc, 99mTc), thallium (201Ti), gallium (68Ga, 67Ga), palladium (103Pd), molybdenum (99Mo), xenon (133Xe), fluorine (18F), 153Sm, 177Lu, 159Gd, 149Pm, 140La, 175Yb, 166Ho, 90Y, 47Sc, 186Re, 188Re, 142Pr, 105Rh, and 97Ru.

[0194] In specific embodiments, a polypeptide of the present invention or fragment or variant thereof is attached to macrocyclic chelators that associate with radiometal ions, including but not limited to, 177Lu, 90Y, 166Ho, and 153Sm, to polypeptides. In a preferred embodiment, the radiometal ion associated with the macrocyclic chelators is 111In. In another preferred embodiment, the radiometal ion associated with the macrocyclic chelator is 90Y. In specific embodiments, the macrocyclic chelator is 1,4,7,10-tetraazacyclododecane-N,N′,N″,N′″-tetraacetic acid (DOTA). In other specific embodiments, DOTA is attached to an antibody of the invention or fragment thereof via a linker molecule. Examples of linker molecules useful for conjugating DOTA to a polypeptide are commonly known in the art—see, for example, DeNardo et al., Clin Cancer Res. 4(10):2483-90 (1998); Peterson et al., Bioconjug. Chem. 10(4):553-7 (1999); and Zimmerman et al, Nucl. Med. Biol. 26(8):943-50 (1999); which are hereby incorporated by reference in their entirety.

[0195] As mentioned, the proteins of the invention may be modified by either natural processes, such as posttranslational processing, or by chemical modification techniques which are well known in the art. It will be appreciated that the same type of modification may be present in the same or varying degrees at several sites in a given polypeptide. Polypeptides of the invention may be branched, for example, as a result of ubiquitination, and they may be cyclic, with or without branching. Cyclic, branched, and branched cyclic polypeptides may result from posttranslation natural processes or may be made by synthetic methods. Modifications include acetylation, acylation, ADP-ribosylation, amidation, covalent attachment of flavin, covalent attachment of a heme moiety, covalent attachment of a nucleotide or nucleotide derivative, covalent attachment of a lipid or lipid derivative, covalent attachment of phosphotidylinositol, cross-linking, cyclization, disulfide bond formation, demethylation, formation of covalent cross-links, formation of cysteine, formation of pyroglutamate, formylation, gamma-carboxylation, glycosylation, GPI anchor formation, hydroxylation, iodination, methylation, myristoylation, oxidation, pegylation, proteolytic processing, phosphorylation, prenylation, racemization, selenoylation, sulfation, transfer-RNA mediated addition of amino acids to proteins such as arginylation, and ubiquitination. (See, for instance, PROTEINS—STRUCTURE AND MOLECULAR PROPERTIES, 2nd Ed., T. E. Creighton, W. H. Freeman and Company, New York (1993); POSTTRANSLATIONAL COVALENT MODIFICATION OF PROTEINS, B. C. Johnson, Ed., Academic Press, New York, pgs. 1-12 (1983); Seifter et al., Meth. Enzymol. 182:626-646 (1990); Rattan et al., Ann. N.Y. Acad. Sci. 663:48-62 (1992)).

[0196] Also provided by the invention are chemically modified derivatives of the polypeptides of the invention which may provide additional advantages such as increased solubility, stability and circulating time of the polypeptide, or decreased immunogenicity (see U.S. Pat. No. 4,179,337). The chemical moieties for derivitization may be selected from water soluble polymers such as polyethylene glycol, ethylene glycol/propylene glycol copolymers, carboxymethylcellulose, dextran, polyvinyl alcohol and the like. The polypeptides may be modified at random positions within the molecule, or at predetermined positions within the molecule and may include one, two, three or more attached chemical moieties.

[0197] The polymer may be of any molecular weight, and may be branched or unbranched. For polyethylene glycol, the preferred molecular weight is between about 1 kDa and about 100 kDa (the term “about” indicating that in preparations of polyethylene glycol, some molecules will weigh more, some less, than the stated molecular weight) for ease in handling and manufacturing. Other sizes may be used, depending on the desired therapeutic profile (e.g., the duration of sustained release desired, the effects, if any on biological activity, the ease in handling, the degree or lack of antigenicity and other known effects of the polyethylene glycol to a therapeutic protein or analog). For example, the polyethylene glycol may have an average molecular weight of about 200, 500, 1000, 1500, 2000, 2500, 3000, 3500, 4000, 4500, 5000, 5500, 6000, 6500, 7000, 7500, 8000, 8500, 9000, 9500, 10,000, 10,500, 11,000, 11,500, 12,000, 12,500, 13,000, 13,500, 14,000, 14,500, 15,000, 15,500, 16,000, 16,500, 17,000, 17,500, 18,000, 18,500, 19,000, 19,500, 20,000, 25,000, 30,000, 35,000, 40,000, 45,000, 50,000, 55,000, 60,000, 65,000, 70,000, 75,000, 80,000, 85,000, 90,000, 95,000, or 100,000 kDa.

[0198] As noted above, the polyethylene glycol may have a branched structure. Branched polyethylene glycols are described, for example, in U.S. Pat. No. 5,643,575; Morpurgo et al., Appl. Biochem. Biotechnol. 56:59-72 (1996); Vorobjev et al., Nucleosides Nucleotides 18:2745-2750 (1999); and Caliceti et al., Bioconjug. Chem. 10:638-646 (1999), the disclosures of each of which are incorporated herein by reference.

[0199] The polyethylene glycol molecules (or other chemical moieties) should be attached to the protein with consideration of effects on functional or antigenic domains of the protein. There are a number of attachment methods available to those skilled in the art, such as, for example, the method disclosed in EP 0 401 384 (coupling PEG to G-CSF), herein incorporated by reference; see also Malik et al., Exp. Hematol. 20:1028-1035 (1992), reporting pegylation of GM-CSF using tresyl chloride. For example, polyethylene glycol may be covalently bound through amino acid residues via a reactive group, such as a free amino or carboxyl group. Reactive groups are those to which an activated polyethylene glycol molecule may be bound. The amino acid residues having a free amino group may include lysine residues and the N-terminal amino acid residues; those having a free carboxyl group may include aspartic acid residues glutamic acid residues and the C-terminal amino acid residue. Sulfhydryl groups may also be used as a reactive group for attaching the polyethylene glycol molecules. Preferred for therapeutic purposes is attachment at an amino group, such as attachment at the N-terminus or lysine group.

[0200] As suggested above, polyethylene glycol may be attached to proteins via linkage to any of a number of amino acid residues. For example, polyethylene glycol can be linked to proteins via covalent bonds to lysine, histidine, aspartic acid, glutamic acid, or cysteine residues. One or more reaction chemistries may be employed to attach polyethylene glycol to specific amino acid residues (e.g., lysine, histidine, aspartic acid, glutamic acid, or cysteine) of the protein or to more than one type of amino acid residue (e.g., lysine, histidine, aspartic acid, glutamic acid, cysteine and combinations thereof) of the protein.

[0201] One may specifically desire proteins chemically modified at the N-terminus. Using polyethylene glycol as an illustration of the present composition, one may select from a variety of polyethylene glycol molecules (by molecular weight, branching, etc.), the proportion of polyethylene glycol molecules to protein (polypeptide) molecules in the reaction mix, the type of pegylation reaction to be performed, and the method of obtaining the selected N-terminally pegylated protein. The method of obtaining the N-terminally pegylated preparation (i.e., separating this moiety from other monopegylated moieties if necessary) may be by purification of the N-terminally pegylated material from a population of pegylated protein molecules. Selective proteins chemically modified at the N-terminus modification may be accomplished by reductive alkylation which exploits differential reactivity of different types of primary amino groups (lysine versus the N-terminal) available for derivatization in a particular protein. Under the appropriate reaction conditions, substantially selective derivatization of the protein at the N-terminus with a carbonyl group containing polymer is achieved.

[0202] As indicated above, pegylation of the proteins of the invention may be accomplished by any number of means. For example, polyethylene glycol may be attached to the protein either directly or by an intervening linker. Linkerless systems for attaching polyethylene glycol to proteins are described in Delgado et al., Crit. Rev. Thera. Drug Carrier Sys. 9:249-304 (1992); Francis et al., Intern. J. of Hematol. 68:1-18 (1998); U.S. Pat. No. 4,002,531; U.S. Pat. No. 5,349,052; WO 95/06058; and WO 98/32466, the disclosures of each of which are incorporated herein by reference.

[0203] One system for attaching polyethylene glycol directly to amino acid residues of proteins without an intervening linker employs tresylated MPEG, which is produced by the modification of monmethoxy polyethylene glycol (MPEG) using tresylchloride (ClSO2CH2CF3). Upon reaction of protein with tresylated MPEG, polyethylene glycol is directly attached to amine groups of the protein. Thus, the invention includes protein-polyethylene glycol conjugates produced by reacting proteins of the invention with a polyethylene glycol molecule having a 2,2,2-trifluoreothane sulphonyl group.

[0204] Polyethylene glycol can also be attached to proteins using a number of different intervening linkers. For example, U.S. Pat. No. 5,612,460, the entire disclosure of which is incorporated herein by reference, discloses urethane linkers for connecting polyethylene glycol to proteins. Protein-polyethylene glycol conjugates wherein the polyethylene glycol is attached to the protein by a linker can also be produced by reaction of proteins with compounds such as MPEG-succinimidylsuccinate, MPEG activated with 1,1′-carbonyldiimidazole, MPEG-2,4,5-trichloropenylcarbonate, MPEG-p-nitrophenolcarbonate, and various MPEG-succinate derivatives. A number of additional polyethylene glycol derivatives and reaction chemistries for attaching polyethylene glycol to proteins are described in International Publication No. WO 98/32466, the entire disclosure of which is incorporated herein by reference. Pegylated protein products produced using the reaction chemistries set out herein are included within the scope of the invention.

[0205] The number of polyethylene glycol moieties attached to each protein of the invention (i.e., the degree of substitution) may also vary. For example, the pegylated proteins of the invention may be linked, on average, to 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 12, 15, 17, 20, or more polyethylene glycol molecules. Similarly, the average degree of substitution within ranges such as 1-3,2-4, 3-5,4-6, 5-7,6-8, 7-9,8-10, 9-11, 10-12, 11-13, 12-14, 13-15, 14-16, 15-17, 16-18, 17-19, or 18-20 polyethylene glycol moieties per protein molecule. Methods for determining the degree of substitution are discussed, for example, in Delgado et al., Crit. Rev. Thera. Drug Carrier Sys. 9:249-304 (1992).

[0206] The polypeptides of the invention can be recovered and purified from chemical synthesis and recombinant cell cultures by standard methods which include, but are not limited to, ammonium sulfate or ethanol precipitation, acid extraction, anion or cation exchange chromatography, phosphocellulose chromatography, hydrophobic interaction chromatography, affinity chromatography, hydroxylapatite chromatography and lectin chromatography. Most preferably, high performance liquid chromatography (“HPLC”) is employed for purification. Well known techniques for refolding protein may be employed to regenerate active conformation when the polypeptide is denatured during isolation and/or purification.

[0207] The polypeptides of the invention may be in monomers or multimers (i.e., dimers, trimers, tetramers and higher multimers). Accordingly, the present invention relates to monomers and multimers of the polypeptides of the invention, their preparation, and compositions (preferably, Therapeutics) containing them. In specific embodiments, the polypeptides of the invention are monomers, dimers, trimers or tetramers. In additional embodiments, the multimers of the invention are at least dimers, at least trimers, or at least tetramers.

[0208] Multimers encompassed by the invention may be homomers or heteromers. As used herein, the term homomer refers to a multimer containing only polypeptides corresponding to a protein of the invention (e.g., the amino acid sequence of SEQ ID NO:Y, an amino acid sequence encoded by SEQ ID NO:X or the complement of SEQ ID NO:X, the amino acid sequence encoded by the portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2, and/or an amino acid sequence encoded by cDNA contained in Clone ID NO:Z (including fragments, variants, splice variants, and fusion proteins, corresponding to these as described herein)). These homomers may contain polypeptides having identical or different amino acid sequences. In a specific embodiment, a homomer of the invention is a multimer containing only polypeptides having an identical amino acid sequence. In another specific embodiment, a homomer of the invention is a multimer containing polypeptides having different amino acid sequences. In specific embodiments, the multimer of the invention is a homodimer (e.g., containing two polypeptides having identical or different amino acid sequences) or a homotrimer (e.g., containing three polypeptides having identical and/or different amino acid sequences). In additional embodiments, the homomeric multimer of the invention is at least a homodimer, at least a homotrimer, or at least a homotetramer.

[0209] As used herein, the term heteromer refers to a multimer containing one or more heterologous polypeptides (i.e., polypeptides of different proteins) in addition to the polypeptides of the invention. In a specific embodiment, the multimer of the invention is a heterodimer, a heterotrimer, or a heterotetramer. In additional embodiments, the heteromeric multimer of the invention is at least a heterodimer, at least a heterotrimer, or at least a heterotetramer.

[0210] Multimers of the invention may be the result of hydrophobic, hydrophilic, ionic and/or covalent associations and/or may be indirectly linked by, for example, liposome formation. Thus, in one embodiment, multimers of the invention, such as, for example, homodimers or homotrimers, are formed when polypeptides of the invention contact one another in solution. In another embodiment, heteromultimers of the invention, such as, for example, heterotrimers or heterotetramers, are formed when polypeptides of the invention contact antibodies to the polypeptides of the invention (including antibodies to the heterologous polypeptide sequence in a fusion protein of the invention) in solution. In other embodiments, multimers of the invention are formed by covalent associations with and/or between the polypeptides of the invention. Such covalent associations may involve one or more amino acid residues contained in the polypeptide sequence (e.g., that recited in SEQ ID NO:Y, encoded by the portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2, and/or encoded by the cDNA contained in Clone ID NO:Z). In one instance, the covalent associations are cross-linking between cysteine residues located within the polypeptide sequences which interact in the native (i.e., naturally occurring) polypeptide. In another instance, the covalent associations are the consequence of chemical or recombinant manipulation. Alternatively, such covalent associations may involve one or more amino acid residues contained in the heterologous polypeptide sequence in a fusion protein. In one example, covalent associations are between the heterologous sequence contained in a fusion protein of the invention (see, e.g., U.S. Pat. No. 5,478,925). In a specific example, the covalent associations are between the heterologous sequence contained in a Fe fusion protein of the invention (as described herein). In another specific example, covalent associations of fusion proteins of the invention are between heterologous polypeptide sequence from another protein that is capable of forming covalently associated multimers, such as for example, osteoprotegerin (see, e.g., International Publication NO: WO 98/49305, the contents of which are herein incorporated by reference in its entirety). In another embodiment, two or more polypeptides of the invention are joined through peptide linkers. Examples include those peptide linkers described in U.S. Pat. No. 5,073,627 (hereby incorporated by reference). Proteins comprising multiple polypeptides of the invention separated by peptide linkers may be produced using conventional recombinant DNA technology.

[0211] Another method for preparing multimer polypeptides of the invention involves use of polypeptides of the invention fused to a leucine zipper or isoleucine zipper polypeptide sequence. Leucine zipper and isoleucine zipper domains are polypeptides that promote multimerization of the proteins in which they are found. Leucine zippers were originally identified in several DNA-binding proteins (Landschulz et al., Science 240:1759, (1988)), and have since been found in a variety of different proteins. Among the known leucine zippers are naturally occurring peptides and derivatives thereof that dimerize or trimerize. Examples of leucine zipper domains suitable for producing soluble multimeric proteins of the invention are those described in PCT application WO 94/10308, hereby incorporated by reference. Recombinant fusion proteins comprising a polypeptide of the invention fused to a polypeptide sequence that dimerizes or trimerizes in solution are expressed in suitable host cells, and the resulting soluble multimeric fusion protein is recovered from the culture supernatant using techniques known in the art.

[0212] Trimeric polypeptides of the invention may offer the advantage of enhanced biological activity. Preferred leucine zipper moieties and isoleucine moieties are those that preferentially form trimers. One example is a leucine zipper derived from lung surfactant protein D (SPD), as described in Hoppe et al. (FEBS Letters 344:191, (1994)) and in U.S. patent application Ser. No. 08/446,922, hereby incorporated by reference. Other peptides derived from naturally occurring trimeric proteins may be employed in preparing trimeric polypeptides of the invention.

[0213] In another example, proteins of the invention are associated by interactions between Flag® polypeptide sequence contained in fusion proteins of the invention containing Flag® polypeptide sequence. In a further embodiment, proteins of the invention are associated by interactions between heterologous polypeptide sequence contained in Flag® fusion proteins of the invention and anti-Flag® antibody.

[0214] The multimers of the invention may be generated using chemical techniques known in the art. For example, polypeptides desired to be contained in the multimers of the invention may be chemically cross-linked using linker molecules and linker molecule length optimization techniques known in the art (see, e.g., U.S. Pat. No. 5,478,925, which is herein incorporated by reference in its entirety). Additionally, multimers of the invention may be generated using techniques known in the art to form one or more inter-molecule cross-links between the cysteine residues located within the sequence of the polypeptides desired to be contained in the multimer (see, e.g., U.S. Pat. No. 5,478,925, which is herein incorporated by reference in its entirety). Further, polypeptides of the invention may be routinely modified by the addition of cysteine or biotin to the C-terminus or N-terminus of the polypeptide and techniques known in the art may be applied to generate multimers containing one or more of these modified polypeptides (see, e.g., U.S. Pat. No. 5,478,925, which is herein incorporated by reference in its entirety). Additionally, techniques known in the art may be applied to generate liposomes containing the polypeptide components desired to be contained in the multimer of the invention (see, e.g., U.S. Pat. No. 5,478,925, which is herein incorporated by reference in its entirety).

[0215] Alternatively, multimers of the invention may be generated using genetic engineering techniques known in the art. In one embodiment, polypeptides contained in multimers of the invention are produced recombinantly using fusion protein technology described herein or otherwise known in the art (see, e.g., U.S. Pat. No. 5,478,925, which is herein incorporated by reference in its entirety). In a specific embodiment, polynucleotides coding for a homodimer of the invention are generated by ligating a polynucleotide sequence encoding a polypeptide of the invention to a sequence encoding a linker polypeptide and then further to a synthetic polynucleotide encoding the translated product of the polypeptide in the reverse orientation from the original C-terminus to the N-terminus (lacking the leader sequence) (see, e.g., U.S. Pat. No. 5,478,925, which is herein incorporated by reference in its entirety). In another embodiment, recombinant techniques described herein or otherwise known in the art are applied to generate recombinant polypeptides of the invention which contain a transmembrane domain (or hydrophobic or signal peptide) and which can be incorporated by membrane reconstitution techniques into liposomes (see, e.g., U.S. Pat. No. 5,478,925, which is herein incorporated by reference in its entirety).

[0216] Antibodies

[0217] Further polypeptides of the invention relate to antibodies and T-cell antigen receptors (TCR) which immunospecifically bind a polypeptide, polypeptide fragment, or variant of the invention (e.g., a polypeptide or fragment or variant of the amino acid sequence of SEQ ID NO:Y or a polypeptide encoded by the cDNA contained in Clone ID No:Z, and/or an epitope, of the present invention) as determined by immunoassays well known in the art for assaying specific antibody-antigen binding. Antibodies of the invention include, but are not limited to, polyclonal, monoclonal, multispecific, human, humanized or chimeric antibodies, single chain antibodies, Fab fragments, F(ab′) fragments, fragments produced by a Fab expression library, anti-idiotypic (anti-Id) antibodies (including, e.g., anti-Id antibodies to antibodies of the invention), intracellularly-made antibodies (i.e., intrabodies), and epitope-binding fragments of any of the above. The term “antibody,” as used herein, refers to immunoglobulin molecules and immunologically active portions of immunoglobulin molecules, i.e., molecules that contain an antigen binding site that immunospecifically binds an antigen. The immunoglobulin molecules of the invention can be of any type (e.g., IgG, IgE, IgM, IgD, IgA and IgY), class (e.g., IgG1, IgG2, IgG3, IgG4, IgA1 and IgA2) or subclass of immunoglobulin molecule. In preferred embodiments, the immunoglobulin molecules of the invention are IgG1. In other preferred embodiments, the immunoglobulin molecules of the invention are IgG4.

[0218] Most preferably the antibodies are human antigen-binding antibody fragments of the present invention and include, but are not limited to, Fab, Fab′ and F(ab′)2, Fd, single-chain Fvs (scFv), single-chain antibodies, disulfide-linked Fvs (sdFv) and fragments comprising either a VL or VH domain. Antigen-binding antibody fragments, including single-chain antibodies, may comprise the variable region(s) alone or in combination with the entirety or a portion of the following: hinge region, CH1, CH2, and CH3 domains. Also included in the invention are antigen-binding fragments also comprising any combination of variable region(s) with a hinge region, CH1, CH2, and CH3 domains. The antibodies of the invention may be from any animal origin including birds and mammals. Preferably, the antibodies are human, murine (e.g., mouse and rat), donkey, ship rabbit, goat, guinea pig, camel, horse, or chicken. As used herein, “human” antibodies include antibodies having the amino acid sequence of a human immunoglobulin and include antibodies isolated from human immunoglobulin libraries or from animals transgenic for one or more human immunoglobulin and that do not express endogenous immunoglobulins, as described infra and, for example in, U.S. Pat. No. 5,939,598 by Kucherlapati et al.

[0219] The antibodies of the present invention may be monospecific, bispecific, trispecific or of greater multispecificity. Multispecific antibodies may be specific for different epitopes of a polypeptide of the present invention or may be specific for both a polypeptide of the present invention as well as for a heterologous epitope, such as a heterologous polypeptide or solid support material. See, e.g., PCT publications WO 93/17715; WO 92/08802; WO 91/00360; WO 92/05793; Tutt, et al., J. Immunol. 147:60-69 (1991); U.S. Pat. Nos. 4,474,893; 4,714,681; 4,925,648; 5,573,920; 5,601,819; Kostelny et al., J. Immunol. 148:1547-1553 (1992).

[0220] Antibodies of the present invention may be described or specified in terms of the epitope(s) or portion(s) of a polypeptide of the present invention which they recognize or specifically bind. The epitope(s) or polypeptide portion(s) may be specified as described herein, e.g., by N-terminal and C-terminal positions, or by size in contiguous amino acid residues, or listed in the Tables and Figures. Preferred epitopes of the invention include the predicted epitopes shown in column 7 of Table 1A, as well as polynucleotides that encode these epitopes. Antibodies which specifically bind any epitope or polypeptide of the present invention may also be excluded. Therefore, the present invention includes antibodies that specifically bind polypeptides of the present invention, and allows for the exclusion of the same.

[0221] Antibodies of the present invention may also be described or specified in terms of their cross-reactivity. Antibodies that do not bind any other analog, ortholog, or homolog of a polypeptide of the present invention are included. Antibodies that bind polypeptides with at least 95%, at least 90%, at least 85%, at least 80%, at least 75%, at least 70%, at least 65%, at least 60%, at least 55%, and at least 50% identity (as calculated using methods known in the art and described herein) to a polypeptide of the present invention are also included in the present invention. In specific embodiments, antibodies of the present invention cross-react with murine, rat and/or rabbit homologs of human proteins and the corresponding epitopes thereof. Antibodies that do not bind polypeptides with less than 95%, less than 90%, less than 85%, less than 80%, less than 75%, less than 70%, less than 65%, less than 60%, less than 55%, and less than 50% identity (as calculated using methods known in the art and described herein) to a polypeptide of the present invention are also included in the present invention. In a specific embodiment, the above-described cross-reactivity is with respect to any single specific antigenic or immunogenic polypeptide, or combination(s) of 2, 3, 4, 5, or more of the specific antigenic and/or immunogenic polypeptides disclosed herein. Further included in the present invention are antibodies which bind polypeptides encoded by polynucleotides which hybridize to a polynucleotide of the present invention under stringent hybridization conditions (as described herein). Antibodies of the present invention may also be described or specified in terms of their binding affinity to a polypeptide of the invention. Preferred binding affinities include those with a dissociation constant or Kd less than 5×10−2 M, 10−2 M, 5×10−3 M, 10−3 M, 5×10−−4 M, 10−4 M, 5×10−5 M, 10−5 M, 5×10−6 M, 10−6M, 5×10−7 M, 107 M, 5×10−8 M, 10−8 M, 5×10−9 M, 10−9 M, 5×10−10 M, 10−10 M, 5×10−11 M, 10−11 M, 5×10−12 M, 10−12 M, 5×10−13 M, 10−13 M, 5×10−14 M, 10−14 M, 5×10−15 M, or 10−15 M.

[0222] The invention also provides antibodies that competitively inhibit binding of an antibody to an epitope of the invention as determined by any method known in the art for determining competitive binding, for example, the immunoassays described herein. In preferred embodiments, the antibody competitively inhibits binding to the epitope by at least 95%, at least 90%, at least 85%, at least 80%, at least 75%, at least 70%, at least 60%, or at least 50%.

[0223] Antibodies of the present invention may act as agonists or antagonists of the polypeptides of the present invention. For example, the present invention includes antibodies which disrupt the receptor/ligand interactions with the polypeptides of the invention either partially or fully. Preferably, antibodies of the present invention bind an antigenic epitope disclosed herein, or a portion thereof. The invention features both receptor-specific antibodies and ligand-specific antibodies. The invention also features receptor-specific antibodies which do not prevent ligand binding but prevent receptor activation. Receptor activation (i.e., signaling) may be determined by techniques described herein or otherwise known in the art. For example, receptor activation can be determined by detecting the phosphorylation (e.g., tyrosine or serine/threonine) of the receptor or its substrate by immunoprecipitation followed by western blot analysis (for example, as described supra). In specific embodiments, antibodies are provided that inhibit ligand activity or receptor activity by at least 95%, at least 90%, at least 85%, at least 80%, at least 75%, at least 70%, at least 60%, or at least 50% of the activity in absence of the antibody.

[0224] The invention also features receptor-specific antibodies which both prevent ligand binding and receptor activation as well as antibodies that recognize the receptor-ligand complex, and, preferably, do not specifically recognize the unbound receptor or the unbound ligand. Likewise, included in the invention are neutralizing antibodies which bind the ligand and prevent binding of the ligand to the receptor, as well as antibodies which bind the ligand, thereby preventing receptor activation, but do not prevent the ligand from binding the receptor. Further included in the invention are antibodies which activate the receptor. These antibodies may act as receptor agonists, i.e., potentiate or activate either all or a subset of the biological activities of the ligand-mediated receptor activation, for example, by inducing dimerization of the receptor. The antibodies may be specified as agonists, antagonists or inverse agonists for biological activities comprising the specific biological activities of the peptides of the invention disclosed herein. The above antibody agonists can be made using methods known in the art. See, e.g., PCT publication WO 96/40281; U.S. Pat. No. 5,811,097; Deng et al., Blood 92(6):1981-1988 (1998); Chen et al., Cancer Res. 58(16):3668-3678 (1998); Harrop et al., J. Immunol. 161(4):1786-1794 (1998); Zhu et al., Cancer Res. 58(15):3209-3214 (1998); Yoon et al., J. Immunol. 160(7):3170-3179 (1998); Prat et al., J. Cell. Sci. 111(Pt2):237-247 (1998); Pitard et al., J. Immunol. Methods 205(2):177-190 (1997); Liautard et al., Cytokine 9(4):233-241 (1997); Carlson et al., J. Biol. Chem. 272(17):11295-11301 (1997); Taryman et al., Neuron 14(4):755-762 (1995); Muller et al., Structure 6(9):1153-1167 (1998); Bartunek et al., Cytokine 8(1):14-20 (1996) (which are all incorporated by reference herein in their entireties).

[0225] Antibodies of the present invention may be used, for example, to purify, detect, and target the polypeptides of the present invention, including both in vitro and in vivo diagnostic and therapeutic methods. For example, the antibodies have utility in immunoassays for qualitatively and quantitatively measuring levels of the polypeptides of the present invention in biological samples. See, e.g., Harlow et al., Antibodies: A Laboratory Manual, (Cold Spring Harbor Laboratory Press, 2nd ed. 1988); incorporated by reference herein in its entirety.

[0226] As discussed in more detail below, the antibodies of the present invention may be used either alone or in combination with other compositions. The antibodies may further be recombinantly fused to a heterologous polypeptide at the N- or C-terminus or chemically conjugated (including covalent and non-covalent conjugations) to polypeptides or other compositions. For example, antibodies of the present invention may be recombinantly fused or conjugated to molecules useful as labels in detection assays and effector molecules such as heterologous polypeptides, drugs, radionuclides, or toxins. See, e.g., PCT publications WO 92/08495; WO 91/14438; WO 89/12624; U.S. Pat. No. 5,314,995; and EP 396,387; the disclosures of which are incorporated herein by reference in their entireties.

[0227] The antibodies of the invention include derivatives that are modified, i.e, by the covalent attachment of any type of molecule to the antibody such that covalent attachment does not prevent the antibody from generating an anti-idiotypic response. For example, but not by way of limitation, the antibody derivatives include antibodies that have been modified, e.g., by glycosylation, acetylation, pegylation, phosphylation, amidation, derivatization by known protecting/blocking groups, proteolytic cleavage, linkage to a cellular ligand or other protein, etc. Any of numerous chemical modifications may be carried out by known techniques, including, but not limited to specific chemical cleavage, acetylation, formylation, metabolic synthesis of tunicamycin, etc. Additionally, the derivative may contain one or more non-classical amino acids.

[0228] The antibodies of the present invention may be generated by any suitable method known in the art. Polyclonal antibodies to an antigen-of-interest can be produced by various procedures well known in the art. For example, a polypeptide of the invention can be administered to various host animals including, but not limited to, rabbits, mice, rats, etc. to induce the production of sera containing polyclonal antibodies specific for the antigen. Various adjuvants may be used to increase the immunological response, depending on the host species, and include but are not limited to, Freund's (complete and incomplete), mineral gels such as aluminum hydroxide, surface active substances such as lysolecithin, pluronic polyols, polyanions, peptides, oil emulsions, keyhole limpet hemocyanins, dinitrophenol, and potentially useful human adjuvants such as BCG (bacille Calmette-Guerin) and corynebacterium parvum. Such adjuvants are also well known in the art.

[0229] Monoclonal antibodies can be prepared using a wide variety of techniques known in the art including the use of hybridoma, recombinant, and phage display technologies, or a combination thereof. For example, monoclonal antibodies can be produced using hybridoma techniques including those known in the art and taught, for example, in Harlow et al., Antibodies: A Laboratory Manual, (Cold Spring Harbor Laboratory Press, 2nd ed. 1988); Hammerling, et al., in: Monoclonal Antibodies and T-Cell Hybridomas 563-681 (Elsevier, N.Y., 1981) (said references incorporated by reference in their entireties). The term “monoclonal antibody” as used herein is not limited to antibodies produced through hybridoma technology. The term “monoclonal antibody” refers to an antibody that is derived from a single clone, including any eukaryotic, prokaryotic, or phage clone, and not the method by which it is produced.

[0230] Methods for producing and screening for specific antibodies using hybridoma technology are routine and well known in the art and are discussed in detail in the Examples. In a non-limiting example, mice can be immunized with a polypeptide of the invention or a cell expressing such peptide. Once an immune response is detected, e.g., antibodies specific for the antigen are detected in the mouse serum, the mouse spleen is harvested and splenocytes isolated. The splenocytes are then fused by well known techniques to any suitable myeloma cells, for example cells from cell line SP20 available from the ATCC. Hybridomas are selected and cloned by limited dilution. The hybridoma clones are then assayed by methods known in the art for cells that secrete antibodies capable of binding a polypeptide of the invention. Ascites fluid, which generally contains high levels of antibodies, can be generated by immunizing mice with positive hybridoma clones.

[0231] Accordingly, the present invention provides methods of generating monoclonal antibodies as well as antibodies produced by the method comprising culturing a hybridoma cell secreting an antibody of the invention wherein, preferably, the hybridoma is generated by fusing splenocytes isolated from a mouse immunized with an antigen of the invention with myeloma cells and then screening the hybridomas resulting from the fusion for hybridoma clones that secrete an antibody able to bind a polypeptide of the invention.

[0232] Another well known method for producing both polyclonal and monoclonal human B cell lines is transformation using Epstein Barr Virus (EBV). Protocols for generating EBV-transformed B cell lines are commonly known in the art, such as, for example, the protocol outlined in Chapter 7.22 of Current Protocols in Immunology, Coligan et al., Eds., 1994, John Wiley & Sons, NY, which is hereby incorporated in its entirety by reference. The source of B cells for transformation is commonly human peripheral blood, but B cells for transformation may also be derived from other sources including, but not limited to, lymph nodes, tonsil, spleen, tumor tissue, and infected tissues. Tissues are generally made into single cell suspensions prior to EBV transformation. Additionally, steps may be taken to either physically remove or inactivate T cells (e.g., by treatment with cyclosporin A) in B cell-containing samples, because T cells from individuals seropositive for anti-EBV antibodies can suppress B cell immortalization by EBV.

[0233] In general, the sample containing human B cells is innoculated with EBV, and cultured for 3-4 weeks. A typical source of EBV is the culture supernatant of the B95-8 cell line (ATCC #VR-1492). Physical signs of EBV transformation can generally be seen towards the end of the 3-4 week culture period. By phase-contrast microscopy, transformed cells may appear large, clear, hairy and tend to aggregate in tight clusters of cells. Initially, EBV lines are generally polyclonal. However, over prolonged periods of cell cultures, EBV lines may become monoclonal or polyclonal as a result of the selective outgrowth of particular B cell clones. Alternatively, polyclonal EBV transformed lines may be subcloned (e.g., by limiting dilution culture) or fused with a suitable fusion partner and plated at limiting dilution to obtain monoclonal B cell lines. Suitable fusion partners for EBV transformed cell lines include mouse myeloma cell lines (e.g., SP2/0, X63-Ag8.653), heteromyeloma cell lines (human x mouse; e.g, SPAM-8, SBC-H20, and CB-F7), and human cell lines (e.g., GM 1500, SKO-007, RPMI 8226, and KR-4). Thus, the present invention also provides a method of generating polyclonal or monoclonal human antibodies against polypeptides of the invention or fragments thereof, comprising EBV-transformation of human B cells.

[0234] Antibody fragments which recognize specific epitopes may be generated by known techniques. For example, Fab and F(ab′)2 fragments of the invention may be produced by proteolytic cleavage of immunoglobulin molecules, using enzymes such as papain (to produce Fab fragments) or pepsin (to produce F(ab′)2 fragments). F(ab′)2 fragments contain the variable region, the light chain constant region and the CH1 domain of the heavy chain.

[0235] For example, the antibodies of the present invention can also be generated using various phage display methods known in the art. In phage display methods, functional antibody domains are displayed on the surface of phage particles which carry the polynucleotide sequences encoding them. In a particular embodiment, such phage can be utilized to display antigen binding domains expressed from a repertoire or combinatorial antibody library (e.g., human or murine). Phage expressing an antigen binding domain that binds the antigen of interest can be selected or identified with antigen, e.g., using labeled antigen or antigen bound or captured to a solid surface or bead. Phage used in these methods are typically filamentous phage including fd and M13 binding domains expressed from phage with Fab, Fv or disulfide stabilized Fv antibody domains recombinantly fused to either the phage gene III or gene VIII protein. Examples of phage display methods that can be used to make the antibodies of the present invention include those disclosed in Brinkman et al., J. Immunol. Methods 182:41-50 (1995); Ames et al., J. Immunol. Methods 184:177-186 (1995); Kettleborough et al., Eur. J. Immunol. 24:952-958 (1994); Persic et al., Gene 187 9-18 (1997); Burton et al., Advances in Immunology 57:191-280 (1994); PCT application No. PCT/GB91/01134; PCT publications WO 90/02809; WO 91/10737; WO 92/01047; WO 92/18619; WO 93/11236; WO 95/15982; WO 95/20401; and U.S. Pat. Nos. 5,698,426; 5,223,409; 5,403,484; 5,580,717; 5,427,908; 5,750,753; 5,821,047; 5,571,698; 5,427,908; 5,516,637; 5,780,225; 5,658,727; 5,733,743 and 5,969,108; each of which is incorporated herein by reference in its entirety.

[0236] As described in the above references, after phage selection, the antibody coding regions from the phage can be isolated and used to generate whole antibodies, including human antibodies, or any other desired antigen binding fragment, and expressed in any desired host, including mammalian cells, insect cells, plant cells, yeast, and bacteria, e.g., as described in detail below. For example, techniques to recombinantly produce Fab, Fab′ and F(ab′)2 fragments can also be employed using methods known in the art such as those disclosed in PCT publication WO 92/22324; Mullinax et al., BioTechniques 12(6):864-869 (1992); and Sawai et al., AJRI 34:26-34 (1995); and Better et al., Science 240:1041-1043 (1988) (said references incorporated by reference in their entireties).

[0237] Examples of techniques which can be used to produce single-chain Fvs and antibodies include those described in U.S. Pat. Nos. 4,946,778 and 5,258,498; Huston et al., Methods in Enzymology 203:46-88 (1991); Shu et al., PNAS 90:7995-7999 (1993); and Skerra et al., Science 240:1038-1040 (1988). For some uses, including in vivo use of antibodies in humans and in vitro detection assays, it may be preferable to use chimeric, humanized, or human antibodies. A chimeric antibody is a molecule in which different portions of the antibody are derived from different animal species, such as antibodies having a variable region derived from a murine monoclonal antibody and a human immunoglobulin constant region. Methods for producing chimeric antibodies are known in the art. See e.g., Morrison, Science 229:1202 (1985); Oi et al., BioTechniques 4:214 (1986); Gillies et al., (1989) J. Immunol. Methods 125:191-202; U.S. Pat. Nos. 5,807,715; 4,816,567; and 4,816397, which are incorporated herein by reference in their entirety. Humanized antibodies are antibody molecules from non-human species antibody that binds the desired antigen having one or more complementarity determining regions (CDRs) from the non-human species and a framework regions from a human immunoglobulin molecule. Often, framework residues in the human framework regions will be substituted with the corresponding residue from the CDR donor antibody to alter, preferably improve, antigen binding. These framework substitutions are identified by methods well known in the art, e.g., by modeling of the interactions of the CDR and framework residues to identify framework residues important for antigen binding and sequence comparison to identify unusual framework residues at particular positions. (See, e.g., Queen et al., U.S. Pat. No. 5,585,089; Riechmann et al., Nature 332:323 (1988), which are incorporated herein by reference in their entireties.) Antibodies can be humanized using a variety of techniques known in the art including, for example, CDR-grafting (EP 239,400; PCT publication WO 91/09967; U.S. Pat. Nos. 5,225,539; 5,530,101; and 5,585,089), veneering or resurfacing (EP 592,106; EP 519,596; Padlan, Molecular Immunology 28(4/5):489-498 (1991); Studnicka et al., Protein Engineering 7(6):805-814 (1994); Roguska. et al., PNAS 91:969-973 (1994)), and chain shuffling (U.S. Pat. No. 5,565,332).

[0238] Completely human antibodies are particularly desirable for therapeutic treatment of human patients. Human antibodies can be made by a variety of methods known in the art including phage display methods described above using antibody libraries derived from human immunoglobulin sequences. See also, U.S. Pat. Nos. 4,444,887 and 4,716,111; and PCT publications WO 98/46645, WO 98/50433, WO 98/24893, WO 98/16654, WO 96/34096, WO 96/33735, and WO 91/10741; each of which is incorporated herein by reference in its entirety.

[0239] Human antibodies can also be produced using transgenic mice which are incapable of expressing functional endogenous immunoglobulins, but which can express human immunoglobulin genes. For example, the human heavy and light chain immunoglobulin gene complexes may be introduced randomly or by homologous recombination into mouse embryonic stem cells. Alternatively, the human variable region, constant region, and diversity region may be introduced into mouse embryonic stem cells in addition to the human heavy and light chain genes. The mouse heavy and light chain immunoglobulin genes may be rendered non-functional separately or simultaneously with the introduction of human immunoglobulin loci by homologous recombination. In particular, homozygous deletion of the JH region prevents endogenous antibody production. The modified embryonic stem cells are expanded and microinjected into blastocysts to produce chimeric mice. The chimeric mice are then bred to produce homozygous offspring which express human antibodies. The transgenic mice are immunized in the normal fashion with a selected antigen, e.g., all or a portion of a polypeptide of the invention. Monoclonal antibodies directed against the antigen can be obtained from the immunized, transgenic mice using conventional hybridoma technology. The human immunoglobulin transgenes harbored by the transgenic mice rearrange during B cell differentiation, and subsequently undergo class switching and somatic mutation. Thus, using such a technique, it is possible to produce therapeutically useful IgG, IgA, IgM and IgE antibodies. For an overview of this technology for producing human antibodies, see Lonberg and Huszar, Int. Rev. Immunol. 13:65-93 (1995). For a detailed discussion of this technology for producing human antibodies and human monoclonal antibodies and protocols for producing such antibodies, see, e.g., PCT publications WO 98/24893; WO 92/01047; WO 96/34096; WO 96/33735; European Patent No. 0 598 877; U.S. Pat. Nos. 5,413,923; 5,625,126; 5,633,425; 5,569,825; 5,661,016; 5,545,806; 5,814,318; 5,885,793; 5,916,771; 5,939,598; 6,075,181; and 6,114,598, which are incorporated by reference herein in their entirety. In addition, companies such as Abgenix, Inc. (Freemont, Calif.) and Genpharm (San Jose, Calif.) can be engaged to provide human antibodies directed against a selected antigen using technology similar to that described above.

[0240] Completely human antibodies which recognize a selected epitope can be generated using a technique referred to as “guided selection.” In this approach a selected non-human monoclonal antibody, e.g., a mouse antibody, is used to guide the selection of a completely human antibody recognizing the same epitope. (Jespers et al., Bio/technology 12:899-903 (1988)).

[0241] Further, antibodies to the polypeptides of the invention can, in turn, be utilized to generate anti-idiotype antibodies that “mimic” polypeptides of the invention using techniques well known to those skilled in the art. (See, e.g., Greenspan & Bona, FASEB J. 7(5):437-444; (1989) and Nissinoff, J. Immunol. 147(8):2429-2438 (1991)). For example, antibodies which bind to and competitively inhibit polypeptide multimerization and/or binding of a polypeptide of the invention to a ligand can be used to generate anti-idiotypes that “mimic” the polypeptide multimerization and/or binding domain and, as a consequence, bind to and neutralize polypeptide and/or its ligand. Such neutralizing anti-idiotypes or Fab fragments of such anti-idiotypes can be used in therapeutic regimens to neutralize polypeptide ligand(s)/receptor(s). For example, such anti-idiotypic antibodies can be used to bind a polypeptide of the invention and/or to bind its ligand(s)/receptor(s), and thereby block its biological activity. Alternatively, antibodies which bind to and enhance polypeptide multimerization and/or binding, and/or receptor/ligand multimerization, binding and/or signaling can be used to generate anti-idiotypes that function as agonists of a polypeptide of the invention and/or its ligand/receptor. Such agonistic anti-idiotypes or Fab fragments of such anti-idiotypes can be used in therapeutic regimens as agonists of the polypeptides of the invention or its ligand(s)/receptor(s). For example, such anti-idiotypic antibodies can be used to bind a polypeptide of the invention and/or to bind its ligand(s)/receptor(s), and thereby promote or enhance its biological activity.

[0242] Intrabodies of the invention can be produced using methods known in the art, such as those disclosed and reviewed in Chen et al., Hum. Gene Ther. 5:595-601 (1994); Marasco, W. A., Gene Ther. 4:11-15 (1997); Rondon and Marasco, Annu. Rev. Microbiol. 51:257-283 (1997); Proba et al., J. Mol. Biol. 275:245-253 (1998); Cohen et al., Oncogene 17:2445-2456 (1998); Ohage and Steipe, J. Mol. Biol. 291:1119-1128 (1999); Ohage et al., J. Mol. Biol. 291:1129-1134 (1999); Wirtz and Steipe, Protein Sci. 8:2245-2250 (1999); Zhu et al., J. Immunol. Methods 231:207-222 (1999); and references cited therein.

[0243] Polynucleotides Encoding Antibodies

[0244] The invention further provides polynucleotides comprising a nucleotide sequence encoding an antibody of the invention and fragments thereof. The invention also encompasses polynucleotides that hybridize under stringent or alternatively, under lower stringency hybridization conditions, e.g., as defined supra, to polynucleotides that encode an antibody, preferably, that specifically binds to a polypeptide of the invention, preferably, an antibody that binds to a polypeptide having the amino acid sequence of SEQ ID NO:Y, to a polypeptide encoded by a portion of SEQ ID NO:X as defined in columns 8 and 9 of Table 2, and/or to a polypeptide encoded by the cDNA contained in Clone ID NO:Z.

[0245] The polynucleotides may be obtained, and the nucleotide sequence of the polynucleotides determined, by any method known in the art. For example, if the nucleotide sequence of the antibody is known, a polynucleotide encoding the antibody may be assembled from chemically synthesized oligonucleotides (e.g., as described in Kutmeier et al., BioTechniques 17:242 (1994)), which, briefly, involves the synthesis of overlapping oligonucleotides containing portions of the sequence encoding the antibody, annealing and ligating of those oligonucleotides, and then amplification of the ligated oligonucleotides by PCR.

[0246] Alternatively, a polynucleotide encoding an antibody may be generated from nucleic acid from a suitable source. If a clone containing a nucleic acid encoding a particular antibody is not available, but the sequence of the antibody molecule is known, a nucleic acid encoding the immunoglobulin may be chemically synthesized or obtained from a suitable source (e.g., an antibody cDNA library, or a cDNA library generated from, or nucleic acid, preferably poly A+ RNA, isolated from, any tissue or cells expressing the antibody, such as hybridoma cells selected to express an antibody of the invention) by PCR amplification using synthetic primers hybridizable to the 3′ and 5′ ends of the sequence or by cloning using an oligonucleotide probe specific for the particular gene sequence to identify, e.g., a cDNA clone from a cDNA library that encodes the antibody. Amplified nucleic acids generated by PCR may then be cloned into replicable cloning vectors using any method well known in the art.

[0247] Once the nucleotide sequence and corresponding amino acid sequence of the antibody is determined, the nucleotide sequence of the antibody may be manipulated using methods well known in the art for the manipulation of nucleotide sequences, e.g., recombinant DNA techniques, site directed mutagenesis, PCR, etc. (see, for example, the techniques described in Sambrook et al., 1990, Molecular Cloning, A Laboratory Manual, 2d Ed., Cold Spring Harbor Laboratory, Cold Spring Harbor, N.Y. and Ausubel et al., eds., 1998, Current Protocols in Molecular Biology, John Wiley & Sons, NY, which are both incorporated by reference herein in their entireties), to generate antibodies having a different amino acid sequence, for example to create amino acid substitutions, deletions, and/or insertions.

[0248] In a specific embodiment, the amino acid sequence of the heavy and/or light chain variable domains may be inspected to identify the sequences of the complementarity determining regions (CDRs) by methods that are well know in the art, e.g., by comparison to known amino acid sequences of other heavy and light chain variable regions to determine the regions of sequence hypervariability. Using routine recombinant DNA techniques, one or more of the CDRs may be inserted within framework regions, e.g., into human framework regions to humanize a non-human antibody, as described supra. The framework regions may be naturally occurring or consensus framework regions, and preferably human framework regions (see, e.g., Chothia et al., J. Mol. Biol. 278: 457-479 (1998) for a listing of human framework regions). Preferably, the polynucleotide generated by the combination of the framework regions and CDRs encodes an antibody that specifically binds a polypeptide of the invention. Preferably, as discussed supra, one or more amino acid substitutions may be made within the framework regions, and, preferably, the amino acid substitutions improve binding of the antibody to its antigen. Additionally, such methods may be used to make amino acid substitutions or deletions of one or more variable region cysteine residues participating in an intrachain disulfide bond to generate antibody molecules lacking one or more intrachain disulfide bonds. Other alterations to the polynucleotide are encompassed by the present invention and within the skill of the art.

[0249] In addition, techniques developed for the production of “chimeric antibodies” (Morrison et al., Proc. Natl. Acad. Sci. 81:851-855 (1984); Neuberger et al., Nature 312:604-608 (1984); Takeda et al., Nature 314:452-454 (1985)) by splicing genes from a mouse antibody molecule of appropriate antigen specificity together with genes from a human antibody molecule of appropriate biological activity can be used. As described supra, a chimeric antibody is a molecule in which different portions are derived from different animal species, such as those having a variable region derived from a murine mAb and a human immunoglobulin constant region, e.g., humanized antibodies.

[0250] Alternatively, techniques described for the production of single chain antibodies (U.S. Pat. No. 4,946,778; Bird, Science 242:423-42 (1988); Huston et al., Proc. Natl. Acad. Sci. USA 85:5879-5883 (1988); and Ward et al., Nature 334:544-54 (1989)) can be adapted to produce single chain antibodies. Single chain antibodies are formed by linking the heavy and light chain fragments of the Fv region via an amino acid bridge, resulting in a single chain polypeptide. Techniques for the assembly of functional Fv fragments in E. coli may also be used (Skerra et al., Science 242:1038-1041 (1988)).

[0251] Methods of Producing Antibodies

[0252] The antibodies of the invention can be produced by any method known in the art for the synthesis of antibodies, in particular, by chemical synthesis or preferably, by recombinant expression techniques. Methods of producing antibodies include, but are not limited to, hybridoma technology, EBV transformation, and other methods discussed herein as well as through the use recombinant DNA technology, as discussed below.

[0253] Recombinant expression of an antibody of the invention, or fragment, derivative or analog thereof, (e.g., a heavy or light chain of an antibody of the invention or a single chain antibody of the invention), requires construction of an expression vector containing a polynucleotide that encodes the antibody. Once a polynucleotide encoding an antibody molecule or a heavy or light chain of an antibody, or portion thereof (preferably containing the heavy or light chain variable domain), of the invention has been obtained, the vector for the production of the antibody molecule may be produced by recombinant DNA technology using techniques well known in the art. Thus, methods for preparing a protein by expressing a polynucleotide containing an antibody encoding nucleotide sequence are described herein. Methods which are well known to those skilled in the art can be used to construct expression vectors containing antibody coding sequences and appropriate transcriptional and translational control signals. These methods include, for example, in vitro recombinant DNA techniques, synthetic techniques, and in vivo genetic recombination. The invention, thus, provides replicable vectors comprising a nucleotide sequence encoding an antibody molecule of the invention, or a heavy or light chain thereof, or a heavy or light chain variable domain, operably linked to a promoter. Such vectors may include the nucleotide sequence encoding the constant region of the antibody molecule (see, e.g., PCT Publication WO 86/05807; PCT Publication WO 89/01036; and U.S. Pat. No. 5,122,464) and the variable domain of the antibody may be cloned into such a vector for expression of the entire heavy or light chain.

[0254] The expression vector is transferred to a host cell by conventional techniques and the transfected cells are then cultured by conventional techniques to produce an antibody of the invention. Thus, the invention includes host cells containing a polynucleotide encoding an antibody of the invention, or a heavy or light chain thereof, or a single chain antibody of the invention, operably linked to a heterologous promoter. In preferred embodiments for the expression of double-chained antibodies, vectors encoding both the heavy and light chains may be co-expressed in the host cell for expression of the entire immunoglobulin molecule, as detailed below.

[0255] A variety of host-expression vector systems may be utilized to express the antibody molecules of the invention. Such host-expression systems represent vehicles by which the coding sequences of interest may be produced and subsequently purified, but also represent cells which may, when transformed or transfected with the appropriate nucleotide coding sequences, express an antibody molecule of the invention in situ. These include but are not limited to microorganisms such as bacteria (e.g., E. coli, B. subtilis) transformed with recombinant bacteriophage DNA, plasmid DNA or cosmid DNA expression vectors containing antibody coding sequences; yeast (e.g., Saccharomyces, Pichia) transformed with recombinant yeast expression vectors containing antibody coding sequences; insect cell systems infected with recombinant virus expression vectors (e.g., baculovirus) containing antibody coding sequences; plant cell systems infected with recombinant virus expression vectors (e.g., cauliflower mosaic virus, CaMV; tobacco mosaic virus, TMV) or transformed with recombinant plasmid expression vectors (e.g., Ti plasmid) containing antibody coding sequences; or mammalian cell systems (e.g., COS, CHO, BHK, 293, 3T3 cells) harboring recombinant expression constructs containing promoters derived from the genome of mammalian cells (e.g., metallothionein promoter) or from mammalian viruses (e.g., the adenovirus late promoter; the vaccinia virus 7.5K promoter). Preferably, bacterial cells such as Escherichia coli, and more preferably, eukaryotic cells, especially for the expression of whole recombinant antibody molecule, are used for the expression of a recombinant antibody molecule. For example, mammalian cells such as Chinese hamster ovary cells (CHO), in conjunction with a vector such as the major intermediate early gene promoter element from human cytomegalovirus is an effective expression system for antibodies (Foecking et al., Gene 45:101 (1986); Cockett et al., Bio/Technology 8:2 (1990)).

[0256] In bacterial systems, a number of expression vectors may be advantageously selected depending upon the use intended for the antibody molecule being expressed. For example, when a large quantity of such a protein is to be produced, for the generation of pharmaceutical compositions of an antibody molecule, vectors which direct the expression of high levels of fusion protein products that are readily purified may be desirable. Such vectors include, but are not limited, to the E. coli expression vector pUR278 (Ruther et al., EMBO J. 2:1791 (1983)), in which the antibody coding sequence may be ligated individually into the vector in frame with the lac Z coding region so that a fusion protein is produced; pIN vectors (Inouye & Inouye, Nucleic Acids Res. 13:3101-3109 (1985); Van Heeke & Schuster, J. Biol. Chem. 24:5503-5509 (1989)); and the like. pGEX vectors may also be used to express foreign polypeptides as fusion proteins with glutathione S-transferase (GST). In general, such fusion proteins are soluble and can easily be purified from lysed cells by adsorption and binding to matrix glutathione-agarose beads followed by elution in the presence of free glutathione. The pGEX vectors are designed to include thrombin or factor Xa protease cleavage sites so that the cloned target gene product can be released from the GST moiety.

[0257] In an insect system, Autographa californica nuclear polyhedrosis virus (AcNPV) is used as a vector to express foreign genes. The virus grows in Spodoptera frugiperda cells. The antibody coding sequence may be cloned individually into non-essential regions (for example the polyhedrin gene) of the virus and placed under control of an AcNPV promoter (for example the polyhedrin promoter).

[0258] In mammalian host cells, a number of viral-based expression systems may be utilized. In cases where an adenovirus is used as an expression vector, the antibody coding sequence of interest may be ligated to an adenovirus transcription/translation control complex, e.g., the late promoter and tripartite leader sequence. This chimeric gene may then be inserted in the adenovirus genome by in vitro or in vivo recombination. Insertion in a non-essential region of the viral genome (e.g., region E1 or E3) will result in a recombinant virus that is viable and capable of expressing the antibody molecule in infected hosts. (e.g., see Logan & Shenk, Proc. Natl. Acad. Sci. USA 81:355-359 (1984)). Specific initiation signals may also be required for efficient translation of inserted antibody coding sequences. These signals include the ATG initiation codon and adjacent sequences. Furthermore, the initiation codon must be in phase with the reading frame of the desired coding sequence to ensure translation of the entire insert. These exogenous translational control signals and initiation codons can be of a variety of origins, both natural and synthetic. The efficiency of expression may be enhanced by the inclusion of appropriate transcription enhancer elements, transcription terminators, etc. (see Bittner et al., Methods in Enzymol. 153:51-544 (1987)).

[0259] In addition, a host cell strain may be chosen which modulates the expression of the inserted sequences, or modifies and processes the gene product in the specific fashion desired. Such modifications (e.g., glycosylation) and processing (e.g., cleavage) of protein products may be important for the function of the protein. Different host cells have characteristic and specific mechanisms for the post-translational processing and modification of proteins and gene products. Appropriate cell lines or host systems can be chosen to ensure the correct modification and processing of the foreign protein expressed. To this end, eukaryotic host cells which possess the cellular machinery for proper processing of the primary transcript, glycosylation, and phosphorylation of the gene product may be used. Such mammalian host cells include but are not limited to CHO, VERY, BHK, Hela, COS, MDCK, 293, 3T3, WI38, and in particular, breast cancer cell lines such as, for example, BT483, Hs578T, HTB2, BT20 and T47D, and normal mammary gland cell line such as, for example, CRL7030 and Hs578Bst.

[0260] For long-term, high-yield production of recombinant proteins, stable expression is preferred. For example, cell lines which stably express the antibody molecule may be engineered. Rather than using expression vectors which contain viral origins of replication, host cells can be transformed with DNA controlled by appropriate expression control elements (e.g., promoter, enhancer, sequences, transcription terminators, polyadenylation sites, etc.), and a selectable marker. Following the introduction of the foreign DNA, engineered cells may be allowed to grow for 1-2 days in an enriched media, and then are switched to a selective media. The selectable marker in the recombinant plasmid confers resistance to the selection and allows cells to stably integrate the plasmid into their chromosomes and grow to form foci which in turn can be cloned and expanded into cell lines. This method may advantageously be used to engineer cell lines which express the antibody molecule. Such engineered cell lines may be particularly useful in screening and evaluation of compounds that interact directly or indirectly with the antibody molecule.

[0261] A number of selection systems may be used, including but not limited to the herpes simplex virus thymidine kinase (Wigler et al., Cell 11:223 (1977)), hypoxanthine-guanine phosphoribosyltransferase (Szybalska & Szybalski, Proc. Natl. Acad. Sci. USA 48:202 (1992)), and adenine phosphoribosyltransferase (Lowy et al., Cell 22:817 (1980)) genes can be employed in tk-, hgprt- or aprt-cells, respectively. Also, antimetabolite resistance can be used as the basis of selection for the following genes: dhfr, which confers resistance to methotrexate (Wigler et al., Natl. Acad. Sci. USA 77:357 (1980); O'Hare et al., Proc. Natl. Acad. Sci. USA 78:1527 (1981)); gpt, which confers resistance to mycophenolic acid (Mulligan & Berg, Proc. Natl. Acad. Sci. USA 78:2072 (1981)); neo, which confers resistance to the aminoglycoside G-418 Clinical Pharmacy 12:488-505; Wu and Wu, Biotherapy 3:87-95 (1991); Tolstoshev, Ann. Rev. Pharmacol. Toxicol. 32:573-596 (1993); Mulligan, Science 260:926-932 (1993); and Morgan and Anderson, Ann. Rev. Biochem. 62:191-217 (1993); May, 1993, TIB TECH 11(5):155-215 (1993)); and hygro, which confers resistance to hygromycin (Santerre et al., Gene 30:147 (1984)). Methods commonly known in the art of recombinant DNA technology may be routinely applied to select the desired recombinant clone, and such methods are described, for example, in Ausubel et al. (eds.), Current Protocols in Molecular Biology, John Wiley & Sons, NY (1993); Kriegler, Gene Transfer and Expression, A Laboratory Manual, Stockton Press, NY (1990); and in Chapters 12 and 13, Dracopoli et al. (eds), Current Protocols in Human Genetics, John Wiley & Sons, NY (1994); Colberre-Garapin et al., J. Mol. Biol. 150:1 (1981), which are incorporated by reference herein in their entireties.

[0262] The expression levels of an antibody molecule can be increased by vector amplification (for a review, see Bebbington and Hentschel, The use of vectors based on gene amplification for the expression of cloned genes in mammalian cells in DNA cloning, Vol.3. (Academic Press, New York, 1987)). When a marker in the vector system expressing antibody is amplifiable, increase in the level of inhibitor present in culture of host cell will increase the number of copies of the marker gene. Since the amplified region is associated with the antibody gene, production of the antibody will also increase (Crouse et al., Mol. Cell. Biol. 3:257 (1983)).

[0263] Vectors which use glutamine synthase (GS) or DHFR as the selectable markers can be amplified in the presence of the drugs methionine sulphoximine or methotrexate, respectively. An advantage of glutamine synthase based vectors are the availabilty of cell lines (e.g., the murine myeloma cell line, NS0) which are glutamine synthase negative. Glutamine synthase expression systems can also function in glutamine synthase expressing cells (e.g. Chinese Hamster Ovary (CHO) cells) by providing additional inhibitor to prevent the functioning of the endogenous gene. A glutamine synthase expression system and components thereof are detailed in PCT publications: WO87/04462; WO86/05807; WO89101036; WO89/10404; and WO91/06657 which are incorporated in their entireties by reference herein. Additionally, glutamine synthase expression vectors that may be used according to the present invention are commercially available from suplliers, including, for example Lonza Biologics, Inc. (Portsmouth, N.H.). Expression and production of monoclonal antibodies using a GS expression system in murine myeloma cells is described in Bebbington et al., Bio/technology 10:169(1992) and in Biblia and Robinson Biotechnol. Prog. 11:1 (1995) which are incorporated in their entirities by reference herein.

[0264] The host cell may be co-transfected with two expression vectors of the invention, the first vector encoding a heavy chain derived polypeptide and the second vector encoding a light chain derived polypeptide. The two vectors may contain identical selectable markers which enable equal expression of heavy and light chain polypeptides. Alternatively, a single vector may be used which encodes, and is capable of expressing, both heavy and light chain polypeptides. In such situations, the light chain should be placed before the heavy chain to avoid an excess of toxic free heavy chain (Proudfoot, Nature 322:52 (1986); Kohler, Proc. Natl. Acad. Sci. USA 77:2197 (1980)). The coding sequences for the heavy and light chains may comprise cDNA or genomic DNA.

[0265] Once an antibody molecule of the invention has been produced by an animal, chemically synthesized, or recombinantly expressed, it may be purified by any method known in the art for purification of an immunoglobulin molecule, for example, by chromatography (e.g., ion exchange, affinity, particularly by affinity for the specific antigen after Protein A, and sizing column chromatography), centrifugation, differential solubility, or by any other standard technique for the purification of proteins. In addition, the antibodies of the present invention or fragments thereof can be fused to heterologous polypeptide sequences described herein or otherwise known in the art, to facilitate purification.

[0266] The present invention encompasses antibodies recombinantly fused or chemically conjugated (including both covalently and non-covalently conjugations) to a polypeptide (or portion thereof, preferably at least 10, 20, 30, 40, 50, 60, 70, 80, 90 or 100 amino acids of the polypeptide) of the present invention to generate fusion proteins. The fusion does not necessarily need to be direct, but may occur through linker sequences. The antibodies may be specific for antigens other than polypeptides (or portion thereof, preferably at least 10, 20, 30, 40, 50, 60, 70, 80, 90 or 100 amino acids of the polypeptide) of the present invention. For example, antibodies may be used to target the polypeptides of the present invention to particular cell types, either in vitro or in vivo, by fusing or conjugating the polypeptides of the present invention to antibodies specific for particular cell surface receptors. Antibodies fused or conjugated to the polypeptides of the present invention may also be used in in vitro immunoassays and purification methods using methods known in the art. See e.g., Harbor et al., supra, and PCT publication WO 93/21232; EP 439,095; Naramura et al., Immunol. Lett. 39:91-99 (1994); U.S. Pat. No. 5,474,981; Gillies et al., PNAS 89:1428-1432 (1992); Fell et al., J. Immunol. 146:2446-2452 (1991), which are incorporated by reference in their entireties.

[0267] The present invention further includes compositions comprising the polypeptides of the present invention fused or conjugated to antibody domains other than the variable regions. For example, the polypeptides of the present invention may be fused or conjugated to an antibody Fc region, or portion thereof. The antibody portion fused to a polypeptide of the present invention may comprise the constant region, hinge region, CH1 domain, CH2 domain, and CH3 domain or any combination of whole domains or portions thereof. The polypeptides may also be fused or conjugated to the above antibody portions to form multimers. For example, Fc portions fused to the polypeptides of the present invention can form dimers through disulfide bonding between the Fc portions. Higher multimeric forms can be made by fusing the polypeptides to portions of IgA and IgM. Methods for fusing or conjugating the polypeptides of the present invention to antibody portions are known in the art. See, e.g., U.S. Pat. Nos. 5,336,603; 5,622,929; 5,359,046; 5,349,053; 5,447,851; 5,112,946; EP 307,434; EP 367,166; PCT publications WO 96/04388; WO 91/06570; Ashkenazi et al., Proc. Natl. Acad. Sci. USA 88:10535-10539 (1991); Zheng et al., J. Immunol. 154:5590-5600 (1995); and Vil et al., Proc. Natl. Acad. Sci. USA 89:11337-11341 (1992) (said references incorporated by reference in their entireties).

[0268] As discussed, supra, the polypeptides corresponding to a polypeptide, polypeptide fragment, or a variant of SEQ ID NO:Y may be fused or conjugated to the above antibody portions to increase the in vivo half life of the polypeptides or for use in immunoassays using methods known in the art. Further, the polypeptides corresponding to SEQ ID NO:Y may be fused or conjugated to the above antibody portions to facilitate purification. One reported example describes chimeric proteins consisting of the first two domains of the human CD4-polypeptide and various domains of the constant regions of the heavy or light chains of mammalian immunoglobulins. See EP 394,827; and Traunecker et al., Nature 331:84-86 (1988). The polypeptides of the present invention fused or conjugated to an antibody having disulfide-linked dimeric structures (due to the IgG) may also be more efficient in binding and neutralizing other molecules, than the monomeric secreted protein or protein fragment alone. See, for example, Fountoulakis et al., J. Biochem. 270:3958-3964 (1995). In many cases, the Fc part in a fusion protein is beneficial in therapy and diagnosis, and thus can result in, for example, improved pharmacokinetic properties. See, for example, EP A 232,262. Alternatively, deleting the Fc part after the fusion protein has been expressed, detected, and purified, would be desired. For example, the Fc portion may hinder therapy and diagnosis if the fusion protein is used as an antigen for immunizations. In drug discovery, for example, human proteins, such as hIL-5, have been fused with Fc portions for the purpose of high-throughput screening assays to identify antagonists of hIL-5. (See, Bennett et al., J. Molecular Recognition 8:52-58 (1995); Johanson et al., J. Biol. Chem. 270:9459-9471 (1995)).

[0269] Moreover, the antibodies or fragments thereof of the present invention can be fused to marker sequences, such as a peptide to facilitate purification. In preferred embodiments, the marker amino acid sequence is a hexa-histidine peptide, such as the tag provided in a pQE vector (QIAGEN, Inc., 9259 Eton Avenue, Chatsworth, Calif., 91311), among others, many of which are commercially available. As described in Gentz et al., Proc. Natl. Acad. Sci. USA 86:821-824 (1989), for instance, hexa-histidine provides for convenient purification of the fusion protein. Other peptide tags useful for purification include, but are not limited to, the “HA” tag, which corresponds to an epitope derived from the influenza hemagglutinin protein (Wilson et al., Cell 37:767 (1984)) and the “flag” tag.

[0270] The present invention further encompasses antibodies or fragments thereof conjugated to a diagnostic or therapeutic agent. The antibodies can be used diagnostically to, for example, monitor the development or progression of a tumor as part of a clinical testing procedure to, e.g., determine the efficacy of a given treatment regimen. Detection can be facilitated by coupling the antibody to a detectable substance. Examples of detectable substances include various enzymes, prosthetic groups, fluorescent materials, luminescent materials, bioluminescent materials, radioactive materials, positron emitting metals using various positron emission tomographies, and nonradioactive paramagnetic metal ions. The detectable substance may be coupled or conjugated either directly to the antibody (or fragment thereof) or indirectly, through an intermediate (such as, for example, a linker known in the art) using techniques known in the art. See, for example, U.S. Pat. No. 4,741,900 for metal ions which can be conjugated to antibodies for use as diagnostics according to the present invention. Examples of suitable enzymes include horseradish peroxidase, alkaline phosphatase, beta-galactosidase, or acetylcholinesterase; examples of suitable prosthetic group complexes include streptavidin/biotin and avidin/biotin; examples of suitable fluorescent materials include umbelliferone, fluorescein, fluorescein isothiocyanate, rhodamine, dichlorotriazinylamine fluorescein, dansyl chloride or phycoerythrin; an example of a luminescent material includes luminol; examples of bioluminescent materials include luciferase, luciferin, and aequorin; and examples of suitable radioactive material include 125I, 131I, 111In or 99Tc.

[0271] Further, an antibody or fragment thereof may be conjugated to a therapeutic moiety such as a cytotoxin, e.g., a cytostatic or cytocidal agent, a therapeutic agent or a radioactive metal ion, e.g., alpha-emitters such as, for example, 213Bi. A cytotoxin or cytotoxic agent includes any agent that is detrimental to cells. Examples include paclitaxol, cytochalasin B, gramicidin D, ethidium bromide, emetine, mitomycin, etoposide, tenoposide, vincristine, vinblastine, colchicin, doxorubicin, daunorubicin, dihydroxy anthracin dione, mitoxantrone, mithramycin, actinomycin D, 1-dehydrotestosterone, glucocorticoids, procaine, tetracaine, lidocaine, propranolol, and puromycin and analogs or homologs thereof. Therapeutic agents include, but are not limited to, antimetabolites (e.g., methotrexate, 6-mercaptopurine, 6-thioguanine, cytarabine, 5-fluorouracil decarbazine), alkylating agents (e.g., mechlorethamine, thioepa chlorambucil, melphalan, carmustine (BSNU) and lomustine (CCNU), cyclothosphamide, busulfan, dibromomannitol, streptozotocin, mitomycin C, and cis-dichlorodiamine platinum (II) (DDP) cisplatin), anthracyclines (e.g., daunorubicin (formerly daunomycin) and doxorubicin), antibiotics (e.g., dactinomycin (formerly actinomycin), bleomycin, mithramycin, and anthramycin (AMC)), and anti-mitotic agents (e.g., vincristine and vinblastine).

[0272] The conjugates of the invention can be used for modifying a given biological response, the therapeutic agent or drug moiety is not to be construed as limited to classical chemical therapeutic agents. For example, the drug moiety may be a protein or polypeptide possessing a desired biological activity. Such proteins may include, for example, a toxin such as abrin, ricin A, pseudomonas exotoxin, or diphtheria toxin; a protein such as tumor necrosis factor, a-interferon, β-interferon, nerve growth factor, platelet derived growth factor, tissue plasminogen activator, an apoptotic agent, e.g., TNF-alpha, TNF-beta, AIM I (See, International Publication No. WO 97/33899), AIM II (See, International Publication No. WO 97/34911), Fas Ligand (Takahashi et al., Int. Immunol., 6:1567-1574 (1994)), VEGI (See, International Publication No. WO 99/23105), a thrombotic agent or an anti-angiogenic agent, e.g., angiostatin or endostatin; or, biological response modifiers such as, for example, lymphokines, interleukin-1 (“IL-1”), interleukin-2 (“IL-2”), interleukin-6 (“IL-6”), granulocyte macrophage colony stimulating factor (“GM-CSF”), granulocyte colony stimulating factor (“G-CSF”), or other growth factors.

[0273] Antibodies may also be attached to solid supports, which are particularly useful for immunoassays or purification of the target antigen. Such solid supports include, but are not limited to, glass, cellulose, polyacrylamide, nylon, polystyrene, polyvinyl chloride or polypropylene.

[0274] Techniques for conjugating such therapeutic moiety to antibodies are well known. See, for example, Arnon et al., “Monoclonal Antibodies For Immunotargeting Of Drugs In Cancer Therapy”, in Monoclonal Antibodies And Cancer Therapy, Reisfeld et al. (eds.), pp. 243-56 (Alan R. Liss, Inc. 1985); Hellstrom et al., “Antibodies For Drug Delivery”, in Controlled Drug Delivery (2nd Ed.), Robinson et al. (eds.), pp. 623-53 (Marcel Dekker, Inc. 1987); Thorpe, “Antibody Carriers Of Cytotoxic Agents In Cancer Therapy: A Review”, in Monoclonal Antibodies '84: Biological And Clinical Applications, Pinchera et al. (eds.), pp. 475-506 (1985); “Analysis, Results, And Future Prospective Of The Therapeutic Use Of Radiolabeled Antibody In Cancer Therapy”, in Monoclonal Antibodies For Cancer Detection And Therapy, Baldwin et al. (eds.), pp. 303-16 (Academic Press 1985), and Thorpe et al., “The Preparation And Cytotoxic Properties Of Antibody-Toxin Conjugates”, Immunol. Rev. 62:119-58 (1982).

[0275] Alternatively, an antibody can be conjugated to a second antibody to form an antibody heteroconjugate as described by Segal in U.S. Pat. No. 4,676,980, which is incorporated herein by reference in its entirety.

[0276] An antibody, with or without a therapeutic moiety conjugated to it, administered alone or in combination with cytotoxic factor(s) and/or cytokine(s) can be used as a therapeutic.

[0277] Immunophenotyping

[0278] The antibodies of the invention may be utilized for immunophenotyping of cell lines and biological samples. Translation products of the gene of the present invention may be useful as cell-specific markers, or more specifically as cellular markers that are differentially expressed at various stages of differentiation and/or maturation of particular cell types. Monoclonal antibodies directed against a specific epitope, or combination of epitopes, will allow for the screening of cellular populations expressing the marker. Various techniques can be utilized using monoclonal antibodies to screen for cellular populations expressing the marker(s), and include magnetic separation using antibody-coated magnetic beads, “panning” with antibody attached to a solid matrix (i.e., plate), and flow cytometry (See, e.g., U.S. Pat. No. 5,985,660; and Morrison et al., Cell, 96:737-49 (1999)).

[0279] These techniques allow for the screening of particular populations of cells, such as might be found with hematological malignancies (i.e. minimal residual disease (MRD) in acute leukemic patients) and “non-self” cells in transplantations to prevent Graft-versus-Host Disease (GVHD). Alternatively, these techniques allow for the screening of hematopoietic stem and progenitor cells capable of undergoing proliferation and/or differentiation, as might be found in human umbilical cord blood.

[0280] Assays For Antibody Binding

[0281] The antibodies of the invention may be assayed for immunospecific binding by any method known in the art. The immunoassays which can be used include but are not limited to competitive and non-competitive assay systems using techniques such as western blots, radioimmunoassays, ELISA (enzyme linked immunosorbent assay), “sandwich” immunoassays, immunoprecipitation assays, precipitin reactions, gel diffusion precipitin reactions, immunodiffusion assays, agglutination assays, complement-fixation assays, immunoradiometric assays, fluorescent immunoassays, and protein A immunoassays, to name but a few. Such assays are routine and well known in the art (see, e.g., Ausubel et al, eds, 1994, Current Protocols in Molecular Biology, Vol. 1, John Wiley & Sons, Inc., New York, which is incorporated by reference herein in its entirety). Exemplary immunoassays are described briefly below (but are not intended by way of limitation).

[0282] Immunoprecipitation protocols generally comprise lysing a population of cells in a lysis buffer such as RIPA buffer (1% NP-40 or Triton X-100, 1% sodium deoxycholate, 0.1% SDS, 0.15 M NaCl, 0.01 M sodium phosphate at pH 7.2, 1% Trasylol) supplemented with protein phosphatase and/or protease inhibitors (e.g., EDTA, PMSF, aprotinin, sodium vanadate), adding the antibody of interest to the cell lysate, incubating for a period of time (e.g., 1-4 hours) at 4° C., adding protein A and/or protein G sepharose beads to the cell lysate, incubating for about an hour or more at 4° C., washing the beads in lysis buffer and resuspending the beads in SDS/sample buffer. The ability of the antibody of interest to immunoprecipitate a particular antigen can be assessed by, e.g., western blot analysis. One of skill in the art would be knowledgeable as to the parameters that can be modified to increase the binding of the antibody to an antigen and decrease the background (e.g., pre-clearing the cell lysate with sepharose beads). For further discussion regarding immunoprecipitation protocols see, e.g., Ausubel et al., eds., (1994), Current Protocols in Molecular Biology, Vol. 1, John Wiley & Sons, Inc., New York, section 10. 16.1.

[0283] Western blot analysis generally comprises preparing protein samples, electrophoresis of the protein samples in a polyacrylamide gel (e.g., 8%-20% SDS-PAGE depending on the molecular weight of the antigen), transferring the protein sample from the polyacrylamide gel to a membrane such as nitrocellulose, PVDF or nylon, blocking the membrane in blocking solution (e.g., PBS with 3% BSA or non-fat milk), washing the membrane in washing buffer (e.g., PBS-Tween 20), blocking the membrane with primary antibody (the antibody of interest) diluted in blocking buffer, washing the membrane in washing buffer, blocking the membrane with a secondary antibody (which recognizes the primary antibody, e.g., an anti-human antibody) conjugated to an enzymatic substrate (e.g., horseradish peroxidase or alkaline phosphatase) or radioactive molecule (e.g., 32P or 125I) diluted in blocking buffer, washing the membrane in wash buffer, and detecting the presence of the antigen. One of skill in the art would be knowledgeable as to the parameters that can be modified to increase the signal detected and to reduce the background noise. For further discussion regarding western blot protocols see, e.g., Ausubel et al, eds, (1994), Current Protocols in Molecular Biology, Vol. 1, John Wiley & Sons, Inc., New York, section 10.8.1.

[0284] ELISAs comprise preparing antigen, coating the well of a 96 well microtiter plate with the antigen, adding the antibody of interest conjugated to a detectable compound such as an enzymatic substrate (e.g., horseradish peroxidase or alkaline phosphatase) to the well and incubating for a period of time, and detecting the presence of the antigen. In ELISAs the antibody of interest does not have to be conjugated to a detectable compound; instead, a second antibody (which recognizes the antibody of interest) conjugated to a detectable compound may be added to the well. Further, instead of coating the well with the antigen, the antibody may be coated to the well. In this case, a second antibody conjugated to a detectable compound may be added following the addition of the antigen of interest to the coated well. One of skill in the art would be knowledgeable as to the parameters that can be modified to increase the signal detected as well as other variations of ELISAs known in the art. For further discussion regarding ELISAs see, e.g., Ausubel et al, eds, (1994), Current Protocols in Molecular Biology, Vol. 1, John Wiley & Sons, Inc., New York, section 11.2.1.

[0285] The binding affinity of an antibody to an antigen and the off-rate of an antibody-antigen interaction can be determined by competitive binding assays. One example of a competitive binding assay is a radioimmunoassay comprising the incubation of labeled antigen (e.g., 3H or 125I) with the antibody of interest in the presence of increasing amounts of unlabeled antigen, and the detection of the antibody bound to the labeled antigen. The affinity of the antibody of interest for a particular antigen and the binding off-rates can be determined from the data by scatchard plot analysis. Competition with a second antibody can also be determined using radioimmunoassays. In this case, the antigen is incubated with antibody of interest conjugated to a labeled compound (e.g., 3H or 125I) in the presence of increasing amounts of an unlabeled second antibody.

[0286] Antibodies of the invention may be characterized using immunocytochemisty methods on cells (e.g., mammalian cells, such as CHO cells) transfected with a vector enabling the expression of an antigen or with vector alone using techniques commonly known in the art. Antibodies that bind antigen transfected cells, but not vector-only transfected cells, are antigen specific.

[0287] Therapeutic Uses

[0288] The present invention is further directed to antibody-based therapies which involve administering antibodies of the invention to an animal, preferably a mammal, and most preferably a human, patient for treating one or more of the disclosed diseases, disorders, or conditions. Therapeutic compounds of the invention include, but are not limited to, antibodies of the invention (including fragments, analogs and derivatives thereof as described herein) and nucleic acids encoding antibodies of the invention (including fragments, analogs and derivatives thereof and anti-idiotypic antibodies as described herein). The antibodies of the invention can be used to treat, inhibit or prevent diseases, disorders or conditions associated with aberrant expression and/or activity of a polypeptide of the invention, including, but not limited to, any one or more of the diseases, disorders, or conditions described herein. The treatment and/or prevention of diseases, disorders, or conditions associated with aberrant expression and/or activity of a polypeptide of the invention includes, but is not limited to, alleviating symptoms associated with those diseases, disorders or conditions. Antibodies of the invention may be provided in pharmaceutically acceptable compositions as known in the art or as described herein.

[0289] In a specific and preferred embodiment, the present invention is directed to antibody-based therapies which involve administering antibodies of the invention to an animal, preferably a mammal, and most preferably a human, patient for treating one or more diseases, disorders, or conditions, including but not limited to: neural disorders, immune system disorders, muscular disorders, reproductive disorders, gastrointestinal disorders, pulmonary disorders, cardiovascular disorders, renal disorders, proliferative disorders, and/or cancerous diseases and conditions., and/or as described elsewhere herein. Therapeutic compounds of the invention include, but are not limited to, antibodies of the invention (e.g., antibodies directed to the full length protein expressed on the cell surface of a mammalian cell; antibodies directed to an epitope of a polypeptide of the invention (such as, for example, a predicted linear epitope shown in column 7 of Table 1A; or a conformational epitope, including fragments, analogs and derivatives thereof as described herein) and nucleic acids encoding antibodies of the invention (including fragments, analogs and derivatives thereof and anti-idiotypic antibodies as described herein). The antibodies of the invention can be used to treat, inhibit or prevent diseases, disorders or conditions associated with aberrant expression and/or activity of a polypeptide of the invention, including, but not limited to, any one or more of the diseases, disorders, or conditions described herein. The treatment and/or prevention of diseases, disorders, or conditions associated with aberrant expression and/or activity of a polypeptide of the invention includes, but is not limited to, alleviating symptoms associated with those diseases, disorders or conditions. Antibodies of the invention may be provided in pharmaceutically acceptable compositions as known in the art or as described herein.

[0290] A summary of the ways in which the antibodies of the present invention may be used therapeutically includes binding polynucleotides or polypeptides of the present invention locally or systemically in the body or by direct cytotoxicity of the antibody, e.g. as mediated by complement (CDC) or by effector cells (ADCC). Some of these approaches are described in more detail below. Armed with the teachings provided herein, one of ordinary skill in the art will know how to use the antibodies of the present invention for diagnostic, monitoring or therapeutic purposes without undue experimentation.

[0291] The antibodies of this invention may be advantageously utilized in combination with other monoclonal or chimeric antibodies, or with lymphokines or hematopoietic growth factors (such as, e.g., IL-2, IL-3 and IL-7), for example, which serve to increase the number or activity of effector cells which interact with the antibodies.

[0292] The antibodies of the invention may be administered alone or in combination with other types of treatments (e.g., radiation therapy, chemotherapy, hormonal therapy, immunotherapy and anti-tumor agents). Generally, administration of products of a species origin or species reactivity (in the case of antibodies) that is the same species as that of the patient is preferred. Thus, in a preferred embodiment, human antibodies, fragments derivatives, analogs, or nucleic acids, are administered to a human patient for therapy or prophylaxis.

[0293] It is preferred to use high affinity and/or potent in vivo inhibiting and/or neutralizing antibodies against polypeptides or polynucleotides of the present invention, fragments or regions thereof, for both immunoassays directed to and therapy of disorders related to polynucleotides or polypeptides, including fragments thereof, of the present invention. Such antibodies, fragments, or regions, will preferably have an affinity for polynucleotides or polypeptides of the invention, including fragments thereof. Preferred binding affinities include those with a dissociation constant or Kd less than 5×10−2 M, 10−2 M, 5×10−3 M, 10−3 M, 5×10−4 M, 10−4 M, 5×10−5 M, 10−5 M, 5×10−6 M, 10−6 M, 5×10−7 M, 10−7 M, 5×10−8 M, 10−8 M, 5×10−9 M, 10−9 M, 5×10−10 M, 10−10 M, 5×10−11 M, 10−11 M, 5×10−12 M, 10−12 M, 5×10−13 M, 10−13 M, 5×10−14 M, 10−14 M, 5×10−15M, and 10−15 M.

[0294] Gene Therapy

[0295] In a specific embodiment, nucleic acids comprising sequences encoding antibodies or functional derivatives thereof, are administered to treat, inhibit or prevent a disease or disorder associated with aberrant expression and/or activity of a polypeptide of the invention, by way of gene therapy. Gene therapy refers to therapy performed by the administration to a subject of an expressed or expressible nucleic acid. In this embodiment of the invention, the nucleic acids produce their encoded protein that mediates a therapeutic effect.

[0296] Any of the methods for gene therapy available in the art can be used according to the present invention. Exemplary methods are described below.

[0297] For general reviews of the methods of gene therapy, see Goldspiel et al., Clinical Pharmacy 12:488-505 (1993); Wu and Wu, Biotherapy 3:87-95 (1991); Tolstoshev, Ann. Rev. Pharmacol. Toxicol. 32:573-596 (1993); Mulligan, Science 260:926-932 (1993); and Morgan and Anderson, Ann. Rev. Biochem. 62:191-217 (1993); May, TIBTECH 11(5):155-215 (1993). Methods commonly known in the art of recombinant DNA technology which can be used are described in Ausubel et al. (eds.), Current Protocols in Molecular Biology, John Wiley & Sons, NY (1993); and Kriegler, Gene Transfer and Expression, A Laboratory Manual, Stockton Press, NY (1990).

[0298] In a preferred embodiment, the compound comprises nucleic acid sequences encoding an antibody, said nucleic acid sequences being part of expression vectors that express the antibody or fragments or chimeric proteins or heavy or light chains thereof in a suitable host. In particular, such nucleic acid sequences have promoters operably linked to the antibody coding region, said promoter being inducible or constitutive, and, optionally, tissue-specific. In another particular embodiment, nucleic acid molecules are used in which the antibody coding sequences and any other desired sequences are flanked by regions that promote homologous recombination at a desired site in the genome, thus providing for intrachromosomal expression of the antibody encoding nucleic acids (Koller and Smithies, Proc. Natl. Acad. Sci. USA 86:8932-8935 (1989); Zijlstra et al., Nature 342:435-438 (1989). In specific embodiments, the expressed antibody molecule is a single chain antibody; alternatively, the nucleic acid sequences include sequences encoding both the heavy and light chains, or fragments thereof, of the antibody.

[0299] Delivery of the nucleic acids into a patient may be either direct, in which case the patient is directly exposed to the nucleic acid or nucleic acid-carrying vectors, or indirect, in which case, cells are first transformed with the nucleic acids in vitro, then transplanted into the patient. These two approaches are known, respectively, as in vivo or ex vivo gene therapy.

[0300] In a specific embodiment, the nucleic acid sequences are directly administered in vivo, where it is expressed to produce the encoded product. This can be accomplished by any of numerous methods known in the art, e.g., by constructing them as part of an appropriate nucleic acid expression vector and administering it so that they become intracellular, e.g., by infection using defective or attenuated retrovirals or other viral vectors (see U.S. Pat. No. 4,980,286), or by direct injection of naked DNA, or by use of microparticle bombardment (e.g., a gene gun; Biolistic, Dupont), or coating with lipids or cell-surface receptors or transfecting agents, encapsulation in liposomes, microparticles, or microcapsules, or by administering them in linkage to a peptide which is known to enter the nucleus, by administering it in linkage to a ligand subject to receptor-mediated endocytosis (see, e.g., Wu and Wu, J. Biol. Chem. 262:4429-4432 (1987)) (which can be used to target cell types specifically expressing the receptors), etc. In another embodiment, nucleic acid-ligand complexes can be formed in which the ligand comprises a fusogenic viral peptide to disrupt endosomes, allowing the nucleic acid to avoid lysosomal degradation. In yet another embodiment, the nucleic acid can be targeted in vivo for cell specific uptake and expression, by targeting a specific receptor (see, e.g., PCT Publications WO 92/06180; WO 92/22635; WO92/20316; WO93/14188, WO 93/20221). Alternatively, the nucleic acid can be introduced intracellularly and incorporated within host cell DNA for expression, by homologous recombination (Koller and Smithies, Proc. Natl. Acad. Sci. USA 86:8932-8935 (1989); Zijlstra et al., Nature 342:435-438 (1989)).

[0301] In a specific embodiment, viral vectors that contains nucleic acid sequences encoding an antibody of the invention are used. For example, a retroviral vector can be used (see Miller et al., Meth. Enzymol. 217:581-599 (1993)). These retroviral vectors contain the components necessary for the correct packaging of the viral genome and integration into the host cell DNA. The nucleic acid sequences encoding the antibody to be used in gene therapy are cloned into one or more vectors, which facilitates delivery of the gene into a patient. More detail about retroviral vectors can be found in Boesen et al., Biotherapy 6:291-302 (1994), which describes the use of a retroviral vector to deliver the mdr1 gene to hematopoietic stem cells in order to make the stem cells more resistant to chemotherapy. Other references illustrating the use of retroviral vectors in gene therapy are: Clowes et al., J. Clin. Invest. 93:644-651 (1994); Kiem et al., Blood 83:1467-1473 (1994); Salmons and Gunzberg, Human Gene Therapy 4:129-141 (1993); and Grossman and Wilson, Curr. Opin. in Genetics and Devel. 3:110-114 (1993).

[0302] Adenoviruses are other viral vectors that can be used in gene therapy. Adenoviruses are especially attractive vehicles for delivering genes to respiratory epithelia. Adenoviruses naturally infect respiratory epithelia where they cause a mild disease. Other targets for adenovirus-based delivery systems are liver, the central nervous system, endothelial cells, and muscle. Adenoviruses have the advantage of being capable of infecting non-dividing cells. Kozarsky and Wilson, Current Opinion in Genetics and Development 3:499-503 (1993) present a review of adenovirus-based gene therapy. Bout et al., Human Gene Therapy 5:3-10 (1994) demonstrated the use of adenovirus vectors to transfer genes to the respiratory epithelia of rhesus monkeys. Other instances of the use of adenoviruses in gene therapy can be found in Rosenfeld et al., Science 252:431-434 (1991); Rosenfeld et al., Cell 68:143-155 (1992); Mastrangeli et al., J. Clin. Invest. 91:225-234 (1993); PCT Publication WO94/12649; and Wang, et al., Gene Therapy 2:775-783 (1995). In a preferred embodiment, adenovirus vectors are used.

[0303] Adeno-associated virus (AAV) has also been proposed for use in gene therapy (Walsh et al., Proc. Soc. Exp. Biol. Med. 204:289-300 (1993); U.S. Pat. No. 5,436,146).

[0304] Another approach to gene therapy involves transferring a gene to cells in tissue culture by such methods as electroporation, lipofection, calcium phosphate mediated transfection, or viral infection. Usually, the method of transfer includes the transfer of a selectable marker to the cells. The cells are then placed under selection to isolate those cells that have taken up and are expressing the transferred gene. Those cells are then delivered to a patient.

[0305] In this embodiment, the nucleic acid is introduced into a cell prior to administration in vivo of the resulting recombinant cell. Such introduction can be carried out by any method known in the art, including but not limited to transfection, electroporation, microinjection, infection with a viral or bacteriophage vector containing the nucleic acid sequences, cell fusion, chromosome-mediated gene transfer, microcell-mediated gene transfer, spheroplast fusion, etc. Numerous techniques are known in the art for the introduction of foreign genes into cells (see, e.g., Loeffler and Behr, Meth. Enzymol. 217:599-618 (1993); Cohen et al., Meth. Enzymol. 217:618-644 (1993); Cline, Pharmac. Ther. 29:69-92m (1985) and may be used in accordance with the present invention, provided that the necessary developmental and physiological functions of the recipient cells are not disrupted. The technique should provide for the stable transfer of the nucleic acid to the cell, so that the nucleic acid is expressible by the cell and preferably heritable and expressible by its cell progeny.

[0306] The resulting recombinant cells can be delivered to a patient by various methods known in the art. Recombinant blood cells (e.g., hematopoietic stem or progenitor cells) are preferably administered intravenously. The amount of cells envisioned for use depends on the desired effect, patient state, etc., and can be determined by one skilled in the art.

[0307] Cells into which a nucleic acid can be introduced for purposes of gene therapy encompass any desired, available cell type, and include but are not limited to epithelial cells, endothelial cells, keratinocytes, fibroblasts, muscle cells, hepatocytes; blood cells such as T lymphocytes, B lymphocytes, monocytes, macrophages, neutrophils, eosinophils, megakaryocytes, granulocytes; various stem or progenitor cells, in particular hematopoietic stem or progenitor cells, e.g., as obtained from bone marrow, umbilical cord blood, peripheral blood, fetal liver, etc. 1290 In a preferred embodiment, the cell used for gene therapy is autologous to the patient.

[0308] In an embodiment in which recombinant cells are used in gene therapy, nucleic acid sequences encoding an antibody are introduced into the cells such that they are expressible by the cells or their progeny, and the recombinant cells are then administered in vivo for therapeutic effect. In a specific embodiment, stem or progenitor cells are used. Any stem and/or progenitor cells which can be isolated and maintained in vitro can potentially be used in accordance with this embodiment of the present invention (see e.g. PCT Publication WO 94/08598; Stemple and Anderson, Cell 71:973-985 (1992); Rheinwald, Meth. Cell Bio. 21A:229 (1980); and Pittelkow and Scott, Mayo Clinic Proc. 61:771 (1986)).

[0309] In a specific embodiment, the nucleic acid to be introduced for purposes of gene therapy comprises an inducible promoter operably linked to the coding region, such that expression of the nucleic acid is controllable by the presence or absence of an appropriate inducer of transcription.

[0310] Demonstration of Therapeutic or Prophylactic Activity

[0311] The compounds or pharmaceutical compositions of the invention are preferably tested in vitro, and then in vivo for the desired therapeutic or prophylactic activity, prior to use in humans. For example, in vitro assays to demonstrate the therapeutic or prophylactic utility of a compound or pharmaceutical composition include, the effect of a compound on a cell line or a patient tissue sample. The effect of the compound or composition on the cell line and/or tissue sample can be determined utilizing techniques known to those of skill in the art including, but not limited to, rosette formation assays and cell lysis assays. In accordance with the invention, in vitro assays which can be used to determine whether administration of a specific compound is indicated, include in vitro cell culture assays in which a patient tissue sample is grown in culture, and exposed to or otherwise administered a compound, and the effect of such compound upon the tissue sample is observed.

[0312] Therapeutic/Prophylactic Administration and Composition

[0313] The invention provides methods of treatment, inhibition and prophylaxis by administration to a subject of an effective amount of a compound or pharmaceutical composition of the invention, preferably a polypeptide or antibody of the invention. In a preferred embodiment, the compound is substantially purified (e.g., substantially free from substances that limit its effect or produce undesired side-effects). The subject is preferably an animal, including but not limited to animals such as cows, pigs, horses, chickens, cats, dogs, etc., and is preferably a mammal, and most preferably human.

[0314] Formulations and methods of administration that can be employed when the compound comprises a nucleic acid or an immunoglobulin are described above; additional appropriate formulations and routes of administration can be selected from among those described herein below.

[0315] Various delivery systems are known and can be used to administer a compound of the invention, e.g., encapsulation in liposomes, microparticles, microcapsules, recombinant cells capable of expressing the compound, receptor-mediated endocytosis (see, e.g., Wu and Wu, J. Biol. Chem. 262:4429-4432 (1987)), construction of a nucleic acid as part of a retroviral or other vector, etc. Methods of introduction include but are not limited to intradermal, intramuscular, intraperitoneal, intravenous, subcutaneous, intranasal, epidural, and oral routes. The compounds or compositions may be administered by any convenient route, for example by infusion or bolus injection, by absorption through epithelial or mucocutaneous linings (e.g., oral mucosa, rectal and intestinal mucosa, etc.) and may be administered together with other biologically active agents. Administration can be systemic or local. In addition, it may be desirable to introduce the pharmaceutical compounds or compositions of the invention into the central nervous system by any suitable route, including intraventricular and intrathecal injection; intraventricular injection may be facilitated by an intraventricular catheter, for example, attached to a reservoir, such as an Ommaya reservoir. Pulmonary administration can also be employed, e.g., by use of an inhaler or nebulizer, and formulation with an aerosolizing agent.

[0316] In a specific embodiment, it may be desirable to administer the pharmaceutical compounds or compositions of the invention locally to the area in need of treatment; this may be achieved by, for example, and not by way of limitation, local infusion during surgery, topical application, e.g., in conjunction with a wound dressing after surgery, by injection, by means of a catheter, by means of a suppository, or by means of an implant, said implant being of a porous, non-porous, or gelatinous material, including membranes, such as sialastic membranes, or fibers. Preferably, when administering a protein, including an antibody, of the invention, care must be taken to use materials to which the protein does not absorb.

[0317] In another embodiment, the compound or composition can be delivered in a vesicle, in particular a liposome (see Langer, Science 249:1527-1533 (1990); Treat et al., in Liposomes in the Therapy of Infectious Disease and Cancer, Lopez-Berestein and Fidler (eds.), Liss, New York, pp. 353-365 (1989); Lopez-Berestein, ibid., pp. 317-327; see generally ibid.)

[0318] In yet another embodiment, the compound or composition can be delivered in a controlled release system. In one embodiment, a pump may be used (see Langer, supra; Sefton, CRC Crit. Ref. Biomed. Eng. 14:201 (1987); Buchwald et al., Surgery 88:507 (1980); Saudek et al., N. Engl. J. Med. 321:574 (1989)). In another embodiment, polymeric materials can be used (see Medical Applications of Controlled Release, Langer and Wise (eds.), CRC Pres., Boca Raton, Fla. (1974); Controlled Drug Bioavailability, Drug Product Design and Performance, Smolen and Ball (eds.), Wiley, New York (1984); Ranger and Peppas, J., Macromol. Sci. Rev. Macromol. Chem. 23:61 (1983); see also Levy et al., Science 228:190 (1985); During et al., Ann. Neurol. 25:351 (1989); Howard et al., J. Neurosurg. 71:105 (1989)). In yet another embodiment, a controlled release system can be placed in proximity of the therapeutic target, e.g., the brain, thus requiring only a fraction of the systemic dose (see, e.g., Goodson, in Medical Applications of Controlled Release, supra, vol. 2, pp. 115-138 (1984)).

[0319] Other controlled release systems are discussed in the review by Langer (Science 249:1527-1533 (1990)).

[0320] In a specific embodiment where the compound of the invention is a nucleic acid encoding a protein, the nucleic acid can be administered in vivo to promote expression of its encoded protein, by constructing it as part of an appropriate nucleic acid expression vector and administering it so that it becomes intracellular, e.g., by use of a retroviral vector (see U.S. Pat. No. 4,980,286), or by direct injection, or by use of microparticle bombardment (e.g., a gene gun; Biolistic, Dupont), or coating with lipids or cell-surface receptors or transfecting agents, or by administering it in linkage to a homeobox-like peptide which is known to enter the nucleus (see e.g., Joliot et al., Proc. Natl. Acad. Sci. USA 88:1864-1868 (1991)), etc. Alternatively, a nucleic acid can be introduced intracellularly and incorporated within host cell DNA for expression, by homologous recombination.

[0321] The present invention also provides pharmaceutical compositions. Such compositions comprise a therapeutically effective amount of a compound, and a pharmaceutically acceptable carrier. In a specific embodiment, the term “pharmaceutically acceptable” means approved by a regulatory agency of the Federal or a state government or listed in the U.S. Pharmacopeia or other generally recognized pharmacopeia for use in animals, and more particularly in humans. The term “carrier” refers to a diluent, adjuvant, excipient, or vehicle with which the therapeutic is administered. Such pharmaceutical carriers can be sterile liquids, such as water and oils, including those of petroleum, animal, vegetable or synthetic origin, such as peanut oil, soybean oil, mineral oil, sesame oil and the like. Water is a preferred carrier when the pharmaceutical composition is administered intravenously. Saline solutions and aqueous dextrose and glycerol solutions can also be employed as liquid carriers, particularly for injectable solutions. Suitable pharmaceutical excipients include starch, glucose, lactose, sucrose, gelatin, malt, rice, flour, chalk, silica gel, sodium stearate, glycerol monostearate, talc, sodium chloride, dried skim milk, glycerol, propylene, glycol, water, ethanol and the like. The composition, if desired, can also contain minor amounts of wetting or emulsifying agents, or pH buffering agents. These compositions can take the form of solutions, suspensions, emulsion, tablets, pills, capsules, powders, sustained-release formulations and the like. The composition can be formulated as a suppository, with traditional binders and carriers such as triglycerides. Oral formulation can include standard carriers such as pharmaceutical grades of mannitol, lactose, starch, magnesium stearate, sodium saccharine, cellulose, magnesium carbonate, etc. Examples of suitable pharmaceutical carriers are described in “Remington's Pharmaceutical Sciences” by E. W. Martin. Such compositions will contain a therapeutically effective amount of the compound, preferably in purified form, together with a suitable amount of carrier so as to provide the form for proper administration to the patient. The formulation should suit the mode of administration.

[0322] In a preferred embodiment, the composition is formulated in accordance with routine procedures as a pharmaceutical composition adapted for intravenous administration to human beings. Typically, compositions for intravenous administration are solutions in sterile isotonic aqueous buffer. Where necessary, the composition may also include a solubilizing agent and a local anesthetic such as lignocaine to ease pain at the site of the injection. Generally, the ingredients are supplied either separately or mixed together in unit dosage form, for example, as a dry lyophilized powder or water free concentrate in a hermetically sealed container such as an ampoule or sachette indicating the quantity of active agent. Where the composition is to be administered by infusion, it can be dispensed with an infusion bottle containing sterile pharmaceutical grade water or saline. Where the composition is administered by injection, an ampoule of sterile water for injection or saline can be provided so that the ingredients may be mixed prior to administration.

[0323] The compounds of the invention can be formulated as neutral or salt forms. Pharmaceutically acceptable salts include those formed with anions such as those derived from hydrochloric, phosphoric, acetic, oxalic, tartaric acids, etc., and those formed with cations such as those derived from sodium, potassium, ammonium, calcium, ferric hydroxides, isopropylamine, triethylamine, 2-ethylamino ethanol, histidine, procaine, etc.

[0324] The amount of the compound of the invention which will be effective in the treatment, inhibition and prevention of a disease or disorder associated with aberrant expression and/or activity of a polypeptide of the invention can be determined by standard clinical techniques. In addition, in vitro assays may optionally be employed to help identify optimal dosage ranges. The precise dose to be employed in the formulation will also depend on the route of administration, and the seriousness of the disease or disorder, and should be decided according to the judgment of the practitioner and each patient's circumstances. Effective doses may be extrapolated from dose-response curves derived from in vitro or animal model test systems.

[0325] For antibodies, the dosage administered to a patient is typically 0.1 mg/kg to 100 mg/kg of the patient's body weight. Preferably, the dosage administered to a patient is between 0.1 mg/kg and 20 mg/kg of the patient's body weight, more preferably 1 mg/kg to 10 mg/kg of the patient's body weight. Generally, human antibodies have a longer half-life within the human body than antibodies from other species due to the immune response to the foreign polypeptides. Thus, lower dosages of human antibodies and less frequent administration is often possible. Further, the dosage and frequency of administration of antibodies of the invention may be reduced by enhancing uptake and tissue penetration (e.g., into the brain) of the antibodies by modifications such as, for example, lipidation.

[0326] The invention also provides a pharmaceutical pack or kit comprising one or more containers filled with one or more of the ingredients of the pharmaceutical compositions of the invention. Optionally associated with such container(s) can be a notice in the form prescribed by a governmental agency regulating the manufacture, use or sale of pharmaceuticals or biological products, which notice reflects approval by the agency of manufacture, use or sale for human administration.

[0327] Diagnosis and Imaging

[0328] Labeled antibodies, and derivatives and analogs thereof, which specifically bind to a polypeptide of interest can be used for diagnostic purposes to detect, diagnose, or monitor diseases, disorders, and/or conditions associated with the aberrant expression and/or activity of a polypeptide of the invention. The invention provides for the detection of aberrant expression of a polypeptide of interest, comprising (a) assaying the expression of the polypeptide of interest in cells or body fluid of an individual using one or more antibodies specific to the polypeptide interest and (b) comparing the level of gene expression with a standard gene expression level, whereby an increase or decrease in the assayed polypeptide gene expression level compared to the standard expression level is indicative of aberrant expression.

[0329] The invention provides a diagnostic assay for diagnosing a disorder, comprising (a) assaying the expression of the polypeptide of interest in cells or body fluid of an individual using one or more antibodies specific to the polypeptide interest and (b) comparing the level of gene expression with a standard gene expression level, whereby an increase or decrease in the assayed polypeptide gene expression level compared to the standard expression level is indicative of a particular disorder. With respect to cancer, the presence of a relatively high amount of transcript in biopsied tissue from an individual may indicate a predisposition for the development of the disease, or may provide a means for detecting the disease prior to the appearance of actual clinical symptoms. A more definitive diagnosis of this type may allow health professionals to employ preventative measures or aggressive treatment earlier thereby preventing the development or further progression of the cancer.

[0330] Antibodies of the invention can be used to assay protein levels in a biological sample using classical immunohistological methods known to those of skill in the art (e.g., see Jalkanen et al., J. Cell. Biol. 101:976-985 (1985); Jalkanen et al., J. Cell . Biol. 105:3087-3096 (1987)). Other antibody-based methods useful for detecting protein gene expression include immunoassays, such as the enzyme linked immunosorbent assay (ELISA) and the radioimmunoassay (RIA). Suitable antibody assay labels are known in the art and include enzyme labels, such as, glucose oxidase; radioisotopes, such as iodine (125I, 121I), carbon (14C), sulfur (35S), tritium (3H), indium (112In), and technetium (99Tc); luminescent labels, such as luminol; and fluorescent labels, such as fluorescein and rhodamine, and biotin.

[0331] One facet of the invention is the detection and diagnosis of a disease or disorder associated with aberrant expression of a polypeptide of interest in an animal, preferably a mammal and most preferably a human. In one embodiment, diagnosis comprises: a) administering (for example, parenterally, subcutaneously, or intraperitoneally) to a subject an effective amount of a labeled molecule which specifically binds to the polypeptide of interest; b) waiting for a time interval following the administering for permitting the labeled molecule to preferentially concentrate at sites in the subject where the polypeptide is expressed (and for unbound labeled molecule to be cleared to background level); c) determining background level; and d) detecting the labeled molecule in the subject, such that detection of labeled molecule above the background level indicates that the subject has a particular disease or disorder associated with aberrant expression of the polypeptide of interest. Background level can be determined by various methods including, comparing the amount of labeled molecule detected to a standard value previously determined for a particular system.

[0332] It will be understood in the art that the size of the subject and the imaging system used will determine the quantity of imaging moiety needed to produce diagnostic images. In the case of a radioisotope moiety, for a human subject, the quantity of radioactivity injected will normally range from about 5 to 20 millicuries of 99 mTc. The labeled antibody or antibody fragment will then preferentially accumulate at the location of cells which contain the specific protein. In vivo tumor imaging is described in S. W. Burchiel et al., “Immunopharmacokinetics of Radiolabeled Antibodies and Their Fragments.” (Chapter 13 in Tumor Imaging: The Radiochemical Detection of Cancer, S. W. Burchiel and B. A. Rhodes, eds., Masson Publishing Inc. (1982)).

[0333] Depending on several variables, including the type of label used and the mode of administration, the time interval following the administration for permitting the labeled molecule to preferentially concentrate at sites in the subject and for unbound labeled molecule to be cleared to background level is 6 to 48 hours or 6 to 24 hours or 6 to 12 hours. In another embodiment the time interval following administration is 5 to 20 days or 5 to 10 days.

[0334] In an embodiment, monitoring of the disease or disorder is carried out by repeating the method for diagnosing the disease or disease, for example, one month after initial diagnosis, six months after initial diagnosis, one year after initial diagnosis, etc.

[0335] Presence of the labeled molecule can be detected in the patient using methods known in the art for in vivo scanning. These methods depend upon the type of label used. Skilled artisans will be able to determine the appropriate method for detecting a particular label. Methods and devices that may be used in the diagnostic methods of the invention include, but are not limited to, computed tomography (CT), whole body scan such as position emission tomography (PET), magnetic resonance imaging (MRI), and sonography.

[0336] In a specific embodiment, the molecule is labeled with a radioisotope and is detected in the patient using a radiation responsive surgical instrument (Thurston et al., U.S. Pat. No. 5,441,050). In another embodiment, the molecule is labeled with a fluorescent compound and is detected in the patient using a fluorescence responsive scanning instrument. In another embodiment, the molecule is labeled with a positron emitting metal and is detected in the patent using positron emission-tomography. In yet another embodiment, the molecule is labeled with a paramagnetic label and is detected in a patient using magnetic resonance imaging (MRI).

[0337] Kits

[0338] The present invention provides kits that can be used in the above methods. In one embodiment, a kit comprises an antibody of the invention, preferably a purified antibody, in one or more containers. In a specific embodiment, the kits of the present invention contain a substantially isolated polypeptide comprising an epitope which is specifically immunoreactive with an antibody included in the kit. Preferably, the kits of the present invention further comprise a control antibody which does not react with the polypeptide of interest. In another specific embodiment, the kits of the present invention contain a means for detecting the binding of an antibody to a polypeptide of interest (e.g., the antibody may be conjugated to a detectable substrate such as a fluorescent compound, an enzymatic substrate, a radioactive compound or a luminescent compound, or a second antibody which recognizes the first antibody may be conjugated to a detectable substrate).

[0339] In another specific embodiment of the present invention, the kit is a diagnostic kit for use in screening serum containing antibodies specific against proliferative and/or cancerous polynucleotides and polypeptides. Such a kit may include a control antibody that does not react with the polypeptide of interest. Such a kit may include a substantially isolated polypeptide antigen comprising an epitope which is specifically immunoreactive with at least one anti-polypeptide antigen antibody. Further, such a kit includes means for detecting the binding of said antibody to the antigen (e.g., the antibody may be conjugated to a fluorescent compound such as fluorescein or rhodamine which can be detected by flow cytometry). In specific embodiments, the kit may include a recombinantly produced or chemically synthesized polypeptide antigen. The polypeptide antigen of the kit may also be attached to a solid support.

[0340] In a more specific embodiment the detecting means of the above-described kit includes a solid support to which said polypeptide antigen is attached. Such a kit may also include a non-attached reporter-labeled anti-human antibody. In this embodiment, binding of the antibody to the polypeptide antigen can be detected by binding of the said reporter-labeled antibody.

[0341] In an additional embodiment, the invention includes a diagnostic kit for use in screening serum containing antigens of the polypeptide of the invention. The diagnostic kit includes a substantially isolated antibody specifically immunoreactive with polypeptide or polynucleotide antigens, and means for detecting the binding of the polynucleotide or polypeptide antigen to the antibody. In one embodiment, the antibody is attached to a solid support. In a specific embodiment, the antibody may be a monoclonal antibody. The detecting means of the kit may include a second, labeled monoclonal antibody. Alternatively, or in addition, the detecting means may include a labeled, competing antigen.

[0342] In one diagnostic configuration, test serum is reacted with a solid phase reagent having a surface-bound antigen obtained by the methods of the present invention. After binding with specific antigen antibody to the reagent and removing unbound serum components by washing, the reagent is reacted with reporter-labeled anti-human antibody to bind reporter to the reagent in proportion to the amount of bound anti-antigen antibody on the solid support. The reagent is again washed to remove unbound labeled antibody, and the amount of reporter associated with the reagent is determined. Typically, the reporter is an enzyme which is detected by incubating the solid phase in the presence of a suitable fluorometric, luminescent or colorimetric substrate (Sigma, St. Louis, Mo.).

[0343] The solid surface reagent in the above assay is prepared by known techniques for attaching protein material to solid support material, such as polymeric beads, dip sticks, 96-well plate or filter material. These attachment methods generally include non-specific adsorption of the protein to the support or covalent attachment of the protein, typically through a free amine group, to a chemically reactive group on the solid support, such as an activated carboxyl, hydroxyl, or aldehyde group. Alternatively, streptavidin coated plates can be used in conjunction with biotinylated antigen(s).

[0344] Thus, the invention provides an assay system or kit for carrying out this diagnostic method. The kit generally includes a support with surface-bound recombinant antigens, and a reporter-labeled anti-human antibody for detecting surface-bound anti-antigen antibody.

[0345] Uses of the Polynucleotides

[0346] Each of the polynucleotides identified herein can be used in numerous ways as reagents. The following description should be considered exemplary and utilizes known techniques.

[0347] The polynucleotides of the present invention are useful for chromosome identification. There exists an ongoing need to identify new chromosome markers, since few chromosome marking reagents, based on actual sequence data (repeat polymorphisms), are presently available. Each sequence is specifically targeted to and can hybridize with a particular location on an individual human chromosome, thus each polynucleotide of the present invention can routinely be used as a chromosome marker using techniques known in the art. Table 1A, column 9 provides the chromosome location of some of the polynucleotides of the invention.

[0348] Briefly, sequences can be mapped to chromosomes by preparing PCR primers (preferably at least 15 bp (e.g., 15-25 bp) from the sequences shown in SEQ ID NO:X. Primers can optionally be selected using computer analysis so that primers do not span more than one predicted exon in the genomic DNA. These primers are then used for PCR screening of somatic cell hybrids containing individual human chromosomes. Only those hybrids containing the human gene corresponding to SEQ ID NO:X will yield an amplified fragment.

[0349] Similarly, somatic hybrids provide a rapid method of PCR mapping the polynucleotides to particular chromosomes. Three or more clones can be assigned per day using a single thermal cycler. Moreover, sublocalization of the polynucleotides can be achieved with panels of specific chromosome fragments. Other gene mapping strategies that can be used include in situ hybridization, prescreening with labeled flow-sorted chromosomes, preselection by hybridization to construct chromosome specific-cDNA libraries, and computer mapping techniques (See, e.g., Shuler, Trends Biotechnol 16:456-459 (1998) which is hereby incorporated by reference in its entirety).

[0350] Precise chromosomal location of the polynucleotides can also be achieved using fluorescence in situ hybridization (FISH) of a metaphase chromosomal spread. This technique uses polynucleotides as short as 500 or 600 bases; however, polynucleotides 2,000-4,000 bp are preferred. For a review of this technique, see Verma et al., “Human Chromosomes: a Manual of Basic Techniques,” Pergamon Press, New York (1988).

[0351] For chromosome mapping, the polynucleotides can be used individually (to mark a single chromosome or a single site on that chromosome) or in panels (for marking multiple sites and/or multiple chromosomes).

[0352] Thus, the present invention also provides a method for chromosomal localization which involves (a) preparing PCR primers from the polynucleotide sequences in Table 1A and/or Table 2 and SEQ ID NO:X and (b) screening somatic cell hybrids containing individual chromosomes.

[0353] The polynucleotides of the present invention would likewise be useful for radiation hybrid mapping, HAPPY mapping, and long range restriction mapping. For a review of these techniques and others known in the art, see, e.g. Dear, “Genome Mapping: A Practical Approach,” IRL Press at Oxford University Press, London (1997); Aydin, J. Mol. Med. 77:691-694 (1999); Hacia et al., Mol. Psychiatry 3:483-492 (1998); Herrick et al., Chromosome Res. 7:409-423 (1999); Hamilton et al., Methods Cell Biol. 62:265-280 (2000); and/or Ott, J. Hered. 90:68-70 (1999) each of which is hereby incorporated by reference in its entirety.

[0354] Once a polynucleotide has been mapped to a precise chromosomal location, the physical position of the polynucleotide can be used in linkage analysis. Linkage analysis establishes coinheritance between a chromosomal location and presentation of a particular disease. (Disease mapping data are found, for example, in V. McKusick, Mendelian Inheritance in Man (available on line through Johns Hopkins University Welch Medical Library)). Column 10 of Table 1A provides an OMIM reference identification number of diseases associated with the cytologic band disclosed in column 9 of Table 1A, as determined using techniques described herein and by reference to Table 5. Assuming 1 megabase mapping resolution and one gene per 20 kb, a cDNA precisely localized to a chromosomal region associated with the disease could be one of 50-500 potential causative genes.

[0355] Thus, once coinheritance is established, differences in a polynucleotide of the invention and the corresponding gene between affected and unaffected individuals can be examined. First, visible structural alterations in the chromosomes, such as deletions or translocations, are examined in chromosome spreads or by PCR. If no structural alterations exist, the presence of point mutations are ascertained. Mutations observed in some or all affected individuals, but not in normal individuals, indicates that the mutation may cause the disease. However, complete sequencing of the polypeptide and the corresponding gene from several normal individuals is required to distinguish the mutation from a polymorphism. If a new polymorphism is identified, this polymorphic polypeptide can be used for further linkage analysis.

[0356] Furthermore, increased or decreased expression of the gene in affected individuals as compared to unaffected individuals can be assessed using the polynucleotides of the invention. Any of these alterations (altered expression, chromosomal rearrangement, or mutation) can be used as a diagnostic or prognostic marker. Diagnostic and prognostic methods, kits and reagents encompassed by the present invention are briefly described below and more thoroughly elsewhere herein (see e.g., the sections labeled “Antibodies”, “Diagnostic Assays”, and “Methods for Detecting Diseases”).

[0357] Thus, the invention also provides a diagnostic method useful during diagnosis of a disorder, involving measuring the expression level of polynucleotides of the present invention in cells or body fluid from an individual and comparing the measured gene expression level with a standard level of polynucleotide expression level, whereby an increase or decrease in the gene expression level compared to the standard is indicative of a disorder. Additional non-limiting examples of diagnostic methods encompassed by the present invention are more thoroughly described elsewhere herein (see, e.g., Example 12).

[0358] In still another embodiment, the invention includes a kit for analyzing samples for the presence of proliferative and/or cancerous polynucleotides derived from a test subject. In a general embodiment, the kit includes at least one polynucleotide probe containing a nucleotide sequence that will specifically hybridize with a polynucleotide of the invention and a suitable container. In a specific embodiment, the kit includes two polynucleotide probes defining an internal region of the polynucleotide of the invention, where each probe has one strand containing a 31′mer-end internal to the region. In a further embodiment, the probes may be useful as primers for polymerase chain reaction amplification.

[0359] Where a diagnosis of a related disorder, including, for example, diagnosis of a tumor, has already been made according to conventional methods, the present invention is useful as a prognostic indicator, whereby patients exhibiting enhanced or depressed polynucleotide of the invention expression will experience a worse clinical outcome relative to patients expressing the gene at a level nearer the standard level.

[0360] By “measuring the expression level of polynucleotides of the invention” is intended qualitatively or quantitatively measuring or estimating the level of the polypeptide of the invention or the level of the mRNA encoding the polypeptide of the invention in a first biological sample either directly (e.g., by determining or estimating absolute protein level or mRNA level) or relatively (e.g., by comparing to the polypeptide level or mRNA level in a second biological sample). Preferably, the polypeptide level or mRNA level in the first biological sample is measured or estimated and compared to a standard polypeptide level or mRNA level, the standard being taken from a second biological sample obtained from an individual not having the related disorder or being determined by averaging levels from a population of individuals not having a related disorder. As will be appreciated in the art, once a standard polypeptide level or mRNA level is known, it can be used repeatedly as a standard for comparison.

[0361] By “biological sample” is intended any biological sample obtained from an individual, body fluid, cell line, tissue culture, or other source which contains polypeptide of the present invention or the corresponding mRNA. As indicated, biological samples include body fluids (such as semen, lymph, vaginal pool, sera, plasma, urine, synovial fluid and spinal fluid) which contain the polypeptide of the present invention, and tissue sources found to express the polypeptide of the present invention. Methods for obtaining tissue biopsies and body fluids from mammals are well known in the art. Where the biological sample is to include mRNA, a tissue biopsy is the preferred source.

[0362] The method(s) provided above may preferably be applied in a diagnostic method and/or kits in which polynucleotides and/or polypeptides of the invention are attached to a solid support. In one exemplary method, the support may be a “gene chip” or a “biological chip” as described in U.S. Pat. Nos. 5,837,832, 5,874,219, and 5,856,174. Further, such a gene chip with polynucleotides of the invention attached may be used to identify polymorphisms between the isolated polynucleotide sequences of the invention, with polynucleotides isolated from a test subject. The knowledge of such polymorphisms (i.e. their location, as well as, their existence) would be beneficial in identifying disease loci for many disorders, such as for example, in neural disorders, immune system disorders, muscular disorders, reproductive disorders, gastrointestinal disorders, pulmonary disorders, digestive disorders, metabolic disorders, cardiovascular disorders, renal disorders, proliferative disorders, and/or cancerous diseases and conditions. Such a method is described in U.S. Pat. Nos. 5,858,659 and 5,856,104. The US Patents referenced supra are hereby incorporated by reference in their entirety herein.

[0363] The present invention encompasses polynucleotides of the present invention that are chemically synthesized, or reproduced as peptide nucleic acids (PNA), or according to other methods known in the art. The use of PNAs would serve as the preferred form if the polynucleotides of the invention are incorporated onto a solid support, or gene chip. For the purposes of the present invention, a peptide nucleic acid (PNA) is a polyamide type of DNA analog and the monomeric units for adenine, guanine, thymine and cytosine are available commercially (Perceptive Biosystems). Certain components of DNA, such as phosphorus, phosphorus oxides, or deoxyribose derivatives, are not present in PNAs. As disclosed by Nielsen et al., Science 254, 1497 (1991); and Egholm et al., Nature 365, 666 (1993), PNAs bind specifically and tightly to complementary DNA strands and are not degraded by nucleases. In fact, PNA binds more strongly to DNA than DNA itself does. This is probably because there is no electrostatic repulsion between the two strands, and also the polyamide backbone is more flexible. Because of this, PNA/DNA duplexes bind under a wider range of stringency conditions than DNA/DNA duplexes, making it easier to perform multiplex hybridization. Smaller probes can be used than with DNA due to the strong binding. In addition, it is more likely that single base mismatches can be determined with PNA/DNA hybridization because a single mismatch in a PNA/DNA 15-mer lowers the melting point (T.sub.m) by 8°-20° C., vs. 4°-16° C. for the DNA/DNA 15-mer duplex. Also, the absence of charge groups in PNA means that hybridization can be done at low ionic strengths and reduce possible interference by salt during the analysis.

[0364] The compounds of the present invention have uses which include, but are not limited to, detecting cancer in mammals. In particular the invention is useful during diagnosis of pathological cell proliferative neoplasias which include, but are not limited to: acute myelogenous leukemias including acute monocytic leukemia, acute myeloblastic leukemia, acute promyelocytic leukemia, acute myelomonocytic leukemia, acute erythroleukemia, acute megakaryocytic leukemia, and acute undifferentiated leukemia, etc.; and chronic myelogenous leukemias including chronic myelomonocytic leukemia, chronic granulocytic leukemia, etc. Preferred mammals include monkeys, apes, cats, dogs, cows, pigs, horses, rabbits and humans. Particularly preferred are humans.

[0365] Pathological cell proliferative disorders are often associated with inappropriate activation of proto-oncogenes. (Gelmann, E. P. et al., “The Etiology of Acute Leukemia: Molecular Genetics and Viral Oncology,” in Neoplastic Diseases of the Blood, Vol 1., Wiernik, P. H. et al. eds., 161-182 (1985)). Neoplasias are now believed to result from the qualitative alteration of a normal cellular gene product, or from the quantitative modification of gene expression by insertion into the chromosome of a viral sequence, by chromosomal translocation of a gene to a more actively transcribed region, or by some other mechanism. (Gelmann et al., supra) It is likely that mutated or altered expression of specific genes is involved in the pathogenesis of some leukemias, among other tissues and cell types. (Gelmann et al., supra) Indeed, the human counterparts of the oncogenes involved in some animal neoplasias have been amplified or translocated in some cases of human leukemia and carcinoma. (Gelmann et al., supra)

[0366] For example, c-myc expression is highly amplified in the non-lymphocytic leukemia cell line HL-60. When HL-60 cells are chemically induced to stop proliferation, the level of c-myc is found to be downregulated. (International Publication Number WO 91/15580). However, it has been shown that exposure of HL-60 cells to a DNA construct that is complementary to the 5′ end of c-myc or c-myb blocks translation of the corresponding mRNAs which downregulates expression of the c-myc or c-myb proteins and causes arrest of cell proliferation and differentiation of the treated cells. (International Publication Number WO 91/15580; Wickstrom et al., Proc. Natl. Acad. Sci. 85:1028 (1988); Anfossi et al., Proc. Natl. Acad. Sci. 86:3379 (1989)). However, the skilled artisan would appreciate the present invention's usefulness is not be limited to treatment, prevention, and/or prognosis of proliferative disorders of cells and tissues of hematopoietic origin, in light of the numerous cells and cell types of varying origins which are known to exhibit proliferative phenotypes.

[0367] In addition to the foregoing, a polynucleotide of the present invention can be used to control gene expression through triple helix formation or through antisense DNA or RNA. Antisense techniques are discussed, for example, in Okano, J. Neurochem. 56: 560 (1991); “Oligodeoxynucleotides as Antisense Inhibitors of Gene Expression, CRC Press, Boca Raton, Fla. (1988). Triple helix formation is discussed in, for instance Lee et al., Nucleic Acids Research 6: 3073 (1979); Cooney et al., Science 241: 456 (1988); and Dervan et al., Science 251: 1360 (1991). Both methods rely on binding of the polynucleotide to a complementary DNA or RNA. For these techniques, preferred polynucleotides are usually oligonucleotides 20 to 40 bases in length and complementary to either the region of the gene involved in transcription (triple helix—see Lee et al., Nucl. Acids Res. 6:3073 (1979); Cooney et al., Science 241:456 (1988); and Dervan et al., Science 251:1360 (1991)) or to the mRNA itself (antisense—Okano, J. Neurochem. 56:560 (1991); Oligodeoxy-nucleotides as Antisense Inhibitors of Gene Expression, CRC Press, Boca Raton, Fla. (1988)). Triple helix formation optimally results in a shut-off of RNA transcription from DNA, while antisense RNA hybridization blocks translation of an mRNA molecule into polypeptide. The oligonucleotide described above can also be delivered to cells such that the antisense RNA or DNA may be expressed in vivo to inhibit production of polypeptide of the present invention antigens. Both techniques are effective in model systems, and the information disclosed herein can be used to design antisense or triple helix polynucleotides in an effort to treat disease, and in particular, for the treatment of proliferative diseases and/or conditions. Non-limiting antisense and triple helix methods encompassed by the present invention are more thoroughly described elsewhere herein (see, e.g., the section labeled “Antisense and Ribozyrne (Antagonists)”).

[0368] Polynucleotides of the present invention are also useful in gene therapy. One goal of gene therapy is to insert a normal gene into an organism having a defective gene, in an effort to correct the genetic defect. The polynucleotides disclosed in the present invention offer a means of targeting such genetic defects in a highly accurate manner. Another goal is to insert a new gene that was not present in the host genome, thereby producing a new trait in the host cell. Additional non-limiting examples of gene therapy methods encompassed by the present invention are more thoroughly described elsewhere herein (see, e.g., the sections labeled “Gene Therapy Methods”, and Examples 16, 17 and 18).

[0369] The polynucleotides are also useful for identifying individuals from minute biological samples. The United States military, for example, is considering the use of restriction fragment length polymorphism (RFLP) for identification of its personnel. In this technique, an individual's genomic DNA is digested with one or more restriction enzymes, and probed on a Southern blot to yield unique bands for identifying personnel. This method does not suffer from the current limitations of “Dog Tags” which can be lost, switched, or stolen, making positive identification difficult. The polynucleotides of the present invention can be used as additional DNA markers for RFLP.

[0370] The polynucleotides of the present invention can also be used as an alternative to RFLP, by determining the actual base-by-base DNA sequence of selected portions of an individual's genome. These sequences can be used to prepare PCR primers for amplifying and isolating such selected DNA, which can then be sequenced. Using this technique, individuals can be identified because each individual will have a unique set of DNA sequences. Once an unique ID database is established for an individual, positive identification of that individual, living or dead, can be made from extremely small tissue samples.

[0371] Forensic biology also benefits from using DNA-based identification techniques as disclosed herein. DNA sequences taken from very small biological samples such as tissues, e.g., hair or skin, or body fluids, e.g., blood, saliva, semen, synovial fluid, amniotic fluid, breast milk, lymph, pulmonary sputum or surfactant, urine, fecal matter, etc., can be amplified using PCR. In one prior art technique, gene sequences amplified from polymorphic loci, such as DQa class II HLA gene, are used in forensic biology to identify individuals. (Erlich, H., PCR Technology, Freeman and Co. (1992)). Once these specific polymorphic loci are amplified, they are digested with one or more restriction enzymes, yielding an identifying set of bands on a Southern blot probed with DNA corresponding to the DQa class II HLA gene. Similarly, polynucleotides of the present invention can be used as polymorphic markers for forensic purposes.

[0372] There is also a need for reagents capable of identifying the source of a particular tissue. Such need arises, for example, in forensics when presented with tissue of unknown origin. Appropriate reagents can comprise, for example, DNA probes or primers prepared from the sequences of the present invention, specific to tissues, including but not limited to those shown in Table 1A. Panels of such reagents can identify tissue by species and/or by organ type. In a similar fashion, these reagents can be used to screen tissue cultures for contamination. Additional non-limiting examples of such uses are further described herein.

[0373] The polynucleotides of the present invention are also useful as hybridization probes for differential identification of the tissue(s) or cell type(s) present in a biological sample. Similarly, polypeptides and antibodies directed to polypeptides of the present invention are useful to provide immunological probes for differential identification of the tissue(s) (e.g., immunohistochemistry assays) or cell type(s) (e.g., immunocytochemistry assays). In addition, for a number of disorders of the above tissues or cells, significantly higher or lower levels of gene expression of the polynucleotides/polypeptides of the present invention may be detected in certain tissues (e.g., tissues expressing polypeptides and/or polynucleotides of the present invention, for example, those disclosed in column 8 of Table A, and/or cancerous and/or wounded tissues) or bodily fluids (e.g., semen, lymph, vaginal pool, serum, plasma, urine, synovial fluid or spinal fluid) taken from an individual having such a disorder, relative to a “standard” gene expression level, i.e., the expression level in healthy tissue from an individual not having the disorder.

[0374] Thus, the invention provides a diagnostic method of a disorder, which involves: (a) assaying gene expression level in cells or body fluid of an individual; (b) comparing the gene expression level with a standard gene expression level, whereby an increase or decrease in the assayed gene expression level compared to the standard expression level is indicative of a disorder.

[0375] In the very least, the polynucleotides of the present invention can be used as molecular weight markers on Southern gels, as diagnostic probes for the presence of a specific mRNA in a particular cell type, as a probe to “subtract-out” known sequences in the process of discovering novel polynucleotides, for selecting and making oligomers for attachment to a “gene chip” or other support, to raise anti-DNA antibodies using DNA immunization techniques, and as an antigen to elicit an immune response.

[0376] Uses of the Polypeptides

[0377] Each of the polypeptides identified herein can be used in numerous ways. The following description should be considered exemplary and utilizes known techniques.

[0378] Polypeptides and antibodies directed to polypeptides of the present invention are useful to provide immunological probes for differential identification of the tissue(s) (e.g., immunohistochemistry assays such as, for example, ABC immunoperoxidase (Hsu et al., J. Histochem. Cytochem. 29:577-580 (1981)) or cell type(s) (e.g., immunocytochemistry assays).

[0379] Antibodies can be used to assay levels of polypeptides encoded by polynucleotides of the invention in a biological sample using classical immunohistological methods known to those of skill in the art (e.g., see Jalkanen, et al., J. Cell. Biol. 101:976-985 (1985); Jalkanen, et al., J. Cell. Biol. 105:3087-3096 (1987)). Other antibody-based methods useful for detecting protein gene expression include immunoassays, such as the enzyme linked immunosorbent assay (ELISA) and the radioimmunoassay (RIA). Suitable antibody assay labels are known in the art and include enzyme labels, such as, glucose oxidase; radioisotopes, such as iodine (131I, 125I, 123I, 121I), carbon (14C), sulfur (35S), tritium (3H), indium (115mIn, 113mIn, 112In, 111In), and technetium (99Tc, 99mTc), thallium (201Ti), gallium (68Ga, 67Ga), palladium (103Pd), molybdenum (99Mo), xenon (133Xe), fluorine (18F), 153Sm, 177Lu, 159Gd, 149Pm, 140La, 175Yb, 166Ho, 90Y, 47Sc, 186Re, 188Re 142Pr, 105Rh, 97Ru; luminescent labels, such as luminol; and fluorescent labels, such as fluorescein and rhodamine, and biotin.

[0380] In addition to assaying levels of polypeptide of the present invention in a biological sample, proteins can also be detected in vivo by imaging. Antibody labels or markers for in vivo imaging of protein include those detectable by X-radiography, NMR or ESR. For X-radiography, suitable labels include radioisotopes such as barium or cesium, which emit detectable radiation but are not overtly harmful to the subject. Suitable markers for NMR and ESR include those with a detectable characteristic spin, such as deuterium, which may be incorporated into the antibody by labeling of nutrients for the relevant hybridoma.

[0381] A protein-specific antibody or antibody fragment which has been labeled with an appropriate detectable imaging moiety, such as a radioisotope (for example, 131I, 112In, 99mTc, (131I, 125I, 123I, 121I), carbon (14C), sulfur (35S), tritium (3H), indium (115mIn, 113mIn, 112In, 111In), and technetium (99Tc, 99mTc), thallium (201Ti), gallium (68Ga, 67Ga), palladium (103Pd), molybdenum (99Mo), xenon (133Xe), fluorine (18F, 153Sm, 177Lu, 159Gd, 149Pm, 140La, 175Yb, 166Ho, 90Y, 47Sc, 18Re, 88Re, 42Pr, 105Rh, 97Ru), a radio-opaque substance, or a material detectable by nuclear magnetic resonance, is introduced (for example, parenterally, subcutaneously or intraperitoneally) into the mammal to be examined for immune system disorder. It will be understood in the art that the size of the subject and the imaging system used will determine the quantity of imaging moiety needed to produce diagnostic images. In the case of a radioisotope moiety, for a human subject, the quantity of radioactivity injected will normally range from about 5 to 20 millicuries of 99mTc. The labeled antibody or antibody fragment will then preferentially accumulate at the location of cells which express the polypeptide encoded by a polynucleotide of the invention. In vivo tumor imaging is described in S. W. Burchiel et al., “Immunopharmacokinetics of Radiolabeled Antibodies and Their Fragments” (Chapter 13 in Tumor Imaging: The Radiochemical Detection of Cancer, S. W. Burchiel and B. A. Rhodes, eds., Masson Publishing Inc. (1982)).

[0382] In one embodiment, the invention provides a method for the specific delivery of compositions of the invention to cells by administering polypeptides of the invention (e.g., polypeptides encoded by polynucleotides of the invention and/or antibodies) that are associated with heterologous polypeptides or nucleic acids. In one example, the invention provides a method for delivering a therapeutic protein into the targeted cell. In another example, the invention provides a method for delivering a single stranded nucleic acid (e.g., antisense or ribozymes) or double stranded nucleic acid (e.g., DNA that can integrate into the cell's genome or replicate episomally and that can be transcribed) into the targeted cell.

[0383] In another embodiment, the invention provides a method for the specific destruction of cells (e.g., the destruction of tumor cells) by administering polypeptides of the invention in association with toxins or cytotoxic prodrugs.

[0384] By “toxin” is meant one or more compounds that bind and activate endogenous cytotoxic effector systems, radioisotopes, holotoxins, modified toxins, catalytic subunits of toxins, or any molecules or enzymes not normally present in or on the surface of a cell that under defined conditions cause the cell's death. Toxins that may be used according to the methods of the invention include, but are not limited to, radioisotopes known in the art, compounds such as, for example, antibodies (or complement fixing containing portions thereof) that bind an inherent or induced endogenous cytotoxic effector system, thymidine kinase, endonuclease, RNAse, alpha toxin, ricin, abrin, Pseudomonas exotoxin A, diphtheria toxin, saporin, momordin, gelonin, pokeweed antiviral protein, alpha-sarcin and cholera toxin. “Toxin” also includes a cytostatic or cytocidal agent, a therapeutic agent or a radioactive metal ion, e.g., alpha-emitters such as, for example, 213Bi, or other radioisotopes such as, for example, 103Pd, 133Xe, 131I, 68Ge, 57Co, 65Zn, 85Sr, 32P, 35S, 90Y, 153Sm, 153Gd, 169Yb, 51Cr, 54Mn, 75Se, 113Sn, 90Yttrium, 117Tin, 186Rhenium, 166Holmium, and 188Rhenium; luminescent labels, such as luminol; and fluorescent labels, such as fluorescein and rhodamine, and biotin. In a specific embodiment, the invention provides a method for the specific destruction of cells (e.g., the destruction of tumor cells) by administering polypeptides of the invention or antibodies of the invention in association with the radioisotope 90Y. In another specific embodiment, the invention provides a method for the specific destruction of cells (e.g., the destruction of tumor cells) by administering polypeptides of the invention or antibodies of the invention in association with the radioisotope 111In. In a further specific embodiment, the invention provides a method for the specific destruction of cells (e.g., the destruction of tumor cells) by administering polypeptides of the invention or antibodies of the invention in association with the radioisotope 131I.

[0385] Techniques known in the art may be applied to label polypeptides of the invention (including antibodies). Such techniques include, but are not limited to, the use of bifunctional conjugating agents (see e.g., U.S. Pat. Nos. 5,756,065; 5,714,631; 5,696,239; 5,652,361; 5,505,931; 5,489,425; 5,435,990; 5,428,139; 5,342,604; 5,274,119; 4,994,560; and 5,808,003; the contents of each of which are hereby incorporated by reference in its entirety).

[0386] Thus, the invention provides a diagnostic method of a disorder, which involves (a) assaying the expression level of a polypeptide of the present invention in cells or body fluid of an individual; and (b) comparing the assayed polypeptide expression level with a standard polypeptide expression level, whereby an increase or decrease in the assayed polypeptide expression level compared to the standard expression level is indicative of a disorder. With respect to cancer, the presence of a relatively high amount of transcript in biopsied tissue from an individual may indicate a predisposition for the development of the disease, or may provide a means for detecting the disease prior to the appearance of actual clinical symptoms. A more definitive diagnosis of this type may allow health professionals to employ preventative measures or aggressive treatment earlier thereby preventing the development or further progression of the cancer.

[0387] Moreover, polypeptides of the present invention can be used to treat or prevent diseases or conditions such as, for example, neural disorders, immune system disorders, muscular disorders, reproductive disorders, gastrointestinal disorders, pulmonary disorders, cardiovascular disorders, renal disorders, proliferative disorders, and/or cancerous diseases and conditions. For example, patients can be administered a polypeptide of the present invention in an effort to replace absent or decreased levels of the polypeptide (e.g., insulin), to supplement absent or decreased levels of a different polypeptide (e.g., hemoglobin S for hemoglobin B, SOD, catalase, DNA repair proteins), to inhibit the activity of a polypeptide (e.g., an oncogene or tumor supressor), to activate the activity of a polypeptide (e.g., by binding to a receptor), to reduce the activity of a membrane bound receptor by competing with it for free ligand (e.g., soluble TNF receptors used in reducing inflammation), or to bring about a desired response (e.g., blood vessel growth inhibition, enhancement of the immune response to proliferative cells or tissues).

[0388] Similarly, antibodies directed to a polypeptide of the present invention can also be used to treat disease (as described supra, and elsewhere herein). For example, administration of an antibody directed to a polypeptide of the present invention can bind, and/or neutralize the polypeptide, and/or reduce overproduction of the polypeptide. Similarly, administration of an antibody can activate the polypeptide, such as by binding to a polypeptide bound to a membrane (receptor).

[0389] At the very least, the polypeptides of the present invention can be used as molecular weight markers on SDS-PAGE gels or on molecular sieve gel filtration columns using methods well known to those of skill in the art. Polypeptides can also be used to raise antibodies, which in turn are used to measure protein expression from a recombinant cell, as a way of assessing transformation of the host cell. Moreover, the polypeptides of the present invention can be used to test the biological activities described herein.

[0390] Diagnostic Assays

[0391] The compounds of the present invention are useful for diagnosis, treatment, prevention and/or prognosis of various disorders in mammals, preferably humans. Such disorders include, but are not limited to, those described herein under the section heading “Biological Activities”.

[0392] For a number of disorders, substantially altered (increased or decreased) levels of gene expression can be detected in tissues, cells or bodily fluids (e.g., sera, plasma, urine, semen, synovial fluid or spinal fluid) taken from an individual having such a disorder, relative to a “standard” gene expression level, that is, the expression level in tissues or bodily fluids from an individual not having the disorder. Thus, the invention provides a diagnostic method useful during diagnosis of a disorder, which involves measuring the expression level of the gene encoding the polypeptide in tissues, cells or body fluid from an individual and comparing the measured gene expression level with a standard gene expression level, whereby an increase or decrease in the gene expression level(s) compared to the standard is indicative of a disorder. These diagnostic assays may be performed in vivo or in vitro, such as, for example, on blood samples, biopsy tissue or autopsy tissue.

[0393] The present invention is also useful as a prognostic indicator, whereby patients exhibiting enhanced or depressed gene expression will experience a worse clinical outcome relative to patients expressing the gene at a level nearer the standard level.

[0394] In certain embodiments, a polypeptide of the invention, or polynucleotides, antibodies, agonists, or antagonists corresponding to that polypeptide, may be used to diagnose and/or prognose diseases and/or disorders associated with the tissue(s) in which the polypeptide of the invention is expressed, including one, two, three, four, five, or more tissues disclosed in Table 1A, column 8 (Tissue Distribution Library Code).

[0395] By “assaying the expression level of the gene encoding the polypeptide” is intended qualitatively or quantitatively measuring or estimating the level of the polypeptide of the invention or the level of the mRNA encoding the polypeptide of the invention in a first biological sample either directly (e.g., by determining or estimating absolute protein level or mRNA level) or relatively (e.g., by comparing to the polypeptide level or mRNA level in a second biological sample). Preferably, the polypeptide expression level or mRNA level in the first biological sample is measured or estimated and compared to a standard polypeptide level or mRNA level, the standard being taken from a second biological sample obtained from an individual not having the disorder or being determined by averaging levels from a population of individuals not having the disorder. As will be appreciated in the art, once a standard polypeptide level or mRNA level is known, it can be used repeatedly as a standard for comparison.

[0396] By “biological sample” is intended any biological sample obtained from an individual, cell line, tissue culture, or other source containing polypeptides of the invention (including portions thereof) or mRNA. As indicated, biological samples include body fluids (such as sera, plasma, urine, synovial fluid and spinal fluid) and tissue sources found to express the full length or fragments thereof of a polypeptide or mRNA. Methods for obtaining tissue biopsies and body fluids from mammals are well known in the art. Where the biological sample is to include mRNA, a tissue biopsy is the preferred source.

[0397] Total cellular RNA can be isolated from a biological sample using any suitable technique such as the single-step guanidinium-thiocyanate-phenol-chloroform method described in Chomczynski and Sacchi, Anal. Biochem. 162:156-159 (1987). Levels of mRNA encoding the polypeptides of the invention are then assayed using any appropriate method. These include Northern blot analysis, S1 nuclease mapping, the polymerase chain reaction (PCR), reverse transcription in combination with the polymerase chain reaction (RT-PCR), and reverse transcription in combination with the ligase chain reaction (RT-LCR).

[0398] The present invention also relates to diagnostic assays such as quantitative and diagnostic assays for detecting levels of polypeptides of the invention, in a biological sample (e.g., cells and tissues), including determination of normal and abnormal levels of polypeptides. Thus, for instance, a diagnostic assay in accordance with the invention for detecting over-expression of polypeptides of the invention compared to normal control tissue samples may be used to detect the presence of tumors. Assay techniques that can be used to determine levels of a polypeptide, such as a polypeptide of the present invention in a sample derived from a host are well-known to those of skill in the art. Such assay methods include radioimmunoassays, competitive-binding assays, Western Blot analysis and ELISA assays. Assaying polypeptide levels in a biological sample can occur using any art-known method.

[0399] Assaying polypeptide levels in a biological sample can occur using antibody-based techniques. For example, polypeptide expression in tissues can be studied with classical immunohistological methods (Jalkanen et al., J. Cell. Biol. 101:976-985 (1985); Jalkanen, M., et al., J. Cell . Biol. 105:3087-3096 (1987)). Other antibody-based methods useful for detecting polypeptide gene expression include immunoassays, such as the enzyme linked immunosorbent assay (ELISA) and the radioimmunoassay (RIA). Suitable antibody assay labels are known in the art and include enzyme labels, such as, glucose oxidase, and radioisotopes, such as iodine (125I, 121I), carbon (14C), sulfur (35S), tritium (3H), indium (112In), and technetium (99mTc), and fluorescent labels, such as fluorescein and rhodamine, and biotin.

[0400] The tissue or cell type to be analyzed will generally include those which are known, or suspected, to express the gene of inteest (such as, for example, cancer). The protein isolation methods employed herein may, for example, be such as those described in Harlow and Lane (Harlow, E. and Lane, D., 1988, “Antibodies: A Laboratory Manual”, Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y.), which is incorporated herein by reference in its entirety. The isolated cells can be derived from cell culture or from a patient. The analysis of cells taken from culture may be a necessary step in the assessment of cells that could be used as part of a cell-based gene therapy technique or, alternatively, to test the effect of compounds on the expression of the gene.

[0401] For example, antibodies, or fragments of antibodies, such as those described herein, may be used to quantitatively or qualitatively detect the presence of gene products or conserved variants or peptide fragments thereof. This can be accomplished, for example, by immunofluorescence techniques employing a fluorescently labeled antibody coupled with light microscopic, flow cytometric, or fluorimetric detection.

[0402] In a preferred embodiment, antibodies, or fragments of antibodies directed to any one or all of the predicted epitope domains of the polypeptides of the invention (shown in column 7 of Table 1A) may be used to quantitatively or qualitatively detect the presence of gene products or conserved variants or peptide fragments thereof. This can be accomplished, for example, by immunofluorescence techniques employing a fluorescently labeled antibody coupled with light microscopic, flow cytometric, or fluorimetric detection.

[0403] In an additional preferred embodiment, antibodies, or fragments of antibodies directed to a conformational epitope of a polypeptide of the invention may be used to quantitatively or qualitatively detect the presence of gene products or conserved variants or peptide fragments thereof. This can be accomplished, for example, by immunofluorescence techniques employing a fluorescently labeled antibody coupled with light microscopic, flow cytometric, or fluorimetric detection.

[0404] The antibodies (or fragments thereof), and/or polypeptides of the present invention may, additionally, be employed histologically, as in immunofluorescence, immunoelectron microscopy or non-immunological assays, for in situ detection of gene products or conserved variants or peptide fragments thereof. In situ detection may be accomplished by removing a histological specimen from a patient, and applying thereto a labeled antibody or polypeptide of the present invention. The antibody (or fragment thereof) or polypeptide is preferably applied by overlaying the labeled antibody (or fragment) onto a biological sample. Through the use of such a procedure, it is possible to determine not only the presence of the gene product, or conserved variants or peptide fragments, or polypeptide binding, but also its distribution in the examined tissue. Using the present invention, those of ordinary skill will readily perceive that any of a wide variety of histological methods (such as staining procedures) can be modified in order to achieve such in situ detection.

[0405] Immunoassays and non-immunoassays for gene products or conserved variants or peptide fragments thereof will typically comprise incubating a sample, such as a biological fluid, a tissue extract, freshly harvested cells, or lysates of cells which have been incubated in cell culture, in the presence of a detectably labeled antibody capable of binding gene products or conserved variants or peptide fragments thereof, and detecting the bound antibody by any of a number of techniques well-known in the art.

[0406] The biological sample may be brought in contact with and immobilized onto a solid phase support or carrier such as nitrocellulose, or other solid support which is capable of immobilizing cells, cell particles or soluble proteins. The support may then be washed with suitable buffers followed by treatment with the detectably labeled antibody or detectable polypeptide of the invention. The solid phase support may then be washed with the buffer a second time to remove unbound antibody or polypeptide. Optionally the antibody is subsequently labeled. The amount of bound label on solid support may then be detected by conventional means.

[0407] By “solid phase support or carrier” is intended any support capable of binding an antigen or an antibody. Well-known supports or carriers include glass, polystyrene, polypropylene, polyethylene, dextran, nylon, amylases, natural and modified celluloses, polyacrylamides, gabbros, and magnetite. The nature of the carrier can be either soluble to some extent or insoluble for the purposes of the present invention. The support material may have virtually any possible structural configuration so long as the coupled molecule is capable of binding to an antigen or antibody. Thus, the support configuration may be spherical, as in a bead, or cylindrical, as in the inside surface of a test tube, or the external surface of a rod. Alternatively, the surface may be flat such as a sheet, test strip, etc. Preferred supports include polystyrene beads. Those skilled in the art will know many other suitable carriers for binding antibody or antigen, or will be able to ascertain the same by use of routine experimentation.

[0408] The binding activity of a given lot of antibody or antigen polypeptide may be determined according to well known methods. Those skilled in the art will be able to determine operative and optimal assay conditions for each determination by employing routine experimentation.

[0409] In addition to assaying polypeptide levels or polynucleotide levels in a biological sample obtained from an individual, polypeptide or polynucleotide can also be detected in vivo by imaging. For example, in one embodiment of the invention, polypeptides and/or antibodies of the invention are used to image diseased cells, such as neoplasms. In another embodiment, polynucleotides of the invention (e.g., polynucleotides complementary to all or a portion of an mRNA) and/or antibodies (e.g., antibodies directed to any one or a combination of the epitopes of a polypeptide of the invention, antibodies directed to a conformational epitope of a polypeptide of the invention, or antibodies directed to the full length polypeptide expressed on the cell surface of a mammalian cell) are used to image diseased or neoplastic cells.

[0410] Antibody labels or markers for in vivo imaging of polypeptides of the invention include those detectable by X-radiography, NMR, MRI, CAT-scans or ESR. For X-radiography, suitable labels include radioisotopes such as barium or cesium, which emit detectable radiation but are not overtly harmful to the subject. Suitable markers for NMR and ESR include those with a detectable characteristic spin, such as deuterium, which may be incorporated into the antibody by labeling of nutrients for the relevant hybridoma. Where in vivo imaging is used to detect enhanced levels of polypeptides for diagnosis in humans, it may be preferable to use human antibodies or “humanized” chimeric monoclonal antibodies. Such antibodies can be produced using techniques described herein or otherwise known in the art. For example methods for producing chimeric antibodies are known in the art. See, for review, Morrison, Science 229:1202 (1985); Oi et al., BioTechniques 4:214 (1986); Cabilly et al., U.S. Pat. No. 4,816,567; Taniguchi et al., EP 171496; Morrison et al., EP 173494; Neuberger et al., WO 8601533; Robinson et al., WO 8702671; Boulianne et al., Nature 312:643 (1984); Neuberger et al., Nature 314:268 (1985).

[0411] Additionally, any polypeptides of the invention whose presence can be detected, can be administered. For example, polypeptides of the invention labeled with a radio-opaque or other appropriate compound can be administered and visualized in vivo, as discussed, above for labeled antibodies. Further, such polypeptides can be utilized for in vitro diagnostic procedures.

[0412] A polypeptide-specific antibody or antibody fragment which has been labeled with an appropriate detectable imaging moiety, such as a radioisotope (for example, 131I, 112In, 99mTc), a radio-opaque substance, or a material detectable by nuclear magnetic resonance, is introduced (for example, parenterally, subcutaneously or intraperitoneally) into the mammal to be examined for a disorder. It will be understood in the art that the size of the subject and the imaging system used will determine the quantity of imaging moiety needed to produce diagnostic images. In the case of a radioisotope moiety, for a human subject, the quantity of radioactivity injected will normally range from about 5 to 20 millicuries of 99mTc. The labeled antibody or antibody fragment will then preferentially accumulate at the location of cells which contain the antigenic protein. In vivo tumor imaging is described in S. W. Burchiel et al., “Immunopharmacokinetics of Radiolabeled Antibodies and Their Fragments” (Chapter 13 in Tumor Imaging: The Radiochemical Detection of Cancer, S. W. Burchiel and B. A. Rhodes, eds., Masson Publishing Inc. (1982)).

[0413] With respect to antibodies, one of the ways in which an antibody of the present invention can be detectably labeled is by linking the same to a reporter enzyme and using the linked product in an enzyme immunoassay (EIA) (Voller, A., “The Enzyme Linked Immunosorbent Assay (ELISA)”, 1978, Diagnostic Horizons 2:1-7, Microbiological Associates Quarterly Publication, Walkersville, Md.); Voller et al., J. Clin. Pathol. 31:507-520 (1978); Butler, J. E., Meth. Enzymol. 73:482-523 (1981); Maggio, E. (ed.), 1980, Enzyme Immunoassay, CRC Press, Boca Raton, Fla.,; Ishikawa, E. et al., (eds.), 1981, Enzyme Immunoassay, Kgaku Shoin, Tokyo). The reporter enzyme which is bound to the antibody will react with an appropriate substrate, preferably a chromogenic substrate, in such a manner as to produce a chemical moiety which can be detected, for example, by spectrophotometric, fluorimetric or by visual means. Reporter enzymes which can be used to detectably label the antibody include, but are not limited to, malate dehydrogenase, staphylococcal nuclease, delta-5-steroid isomerase, yeast alcohol dehydrogenase, alpha-glycerophosphate, dehydrogenase, triose phosphate isomerase, horseradish peroxidase, alkaline phosphatase, asparaginase, glucose oxidase, beta-galactosidase, ribonuclease, urease, catalase, glucose-6-phosphate dehydrogenase, glucoamylase and acetylcholinesterase. Additionally, the detection can be accomplished by colorimetric methods which employ a chromogenic substrate for the reporter enzyme. Detection may also be accomplished by visual comparison of the extent of enzymatic reaction of a substrate in comparison with similarly prepared standards.

[0414] Detection may also be accomplished using any of a variety of other immunoassays. For example, by radioactively labeling the antibodies or antibody fragments, it is possible to detect polypeptides through the use of a radioimmunoassay (RIA) (see, for example, Weintraub, B., Principles of Radioimmunoassays, Seventh Training Course on Radioligand Assay Techniques, The Endocrine Society, March, 1986, which is incorporated by reference herein). The radioactive isotope can be detected by means including, but not limited to, a gamma counter, a scintillation counter, or autoradiography.

[0415] It is also possible to label the antibody with a fluorescent compound. When the fluorescently labeled antibody is exposed to light of the proper wave length, its presence can then be detected due to fluorescence. Among the most commonly used fluorescent labeling compounds are fluorescein isothiocyanate, rhodamine, phycoerythrin, phycocyanin, allophycocyanin, ophthaldehyde and fluorescamine.

[0416] The antibody can also be detectably labeled using fluorescence emitting metals such as 152Eu, or others of the lanthamide series. These metals can be attached to the antibody using such metal chelating groups as diethylenetriaminepentacetic acid (DTPA) or ethylenediaminetetraacetic acid (EDTA).

[0417] The antibody also can be detectably labeled by coupling it to a chemiluminescent compound. The presence of the chemiluminescent-tagged antibody is then determined by detecting the presence of luminescence that arises during the course of a chemical reaction. Examples of particularly useful chemiluminescent labeling compounds are luminol, isoluminol, theromatic acridinium ester, imidazole, acridinium salt and oxalate ester.

[0418] Likewise, a bioluminescent compound may be used to label the antibody of the present invention. Bioluminescence is a type of chemiluminescence found in biological systems in, which a catalytic protein increases the efficiency of the chemiluminescent reaction. The presence of a bioluminescent protein is determined by detecting the presence of luminescence. Important bioluminescent compounds for purposes of labeling are luciferin, luciferase and aequorin.

[0419] Methods for Detecting Diseases

[0420] In general, a disease may be detected in a patient based on the presence of one or more proteins of the invention and/or polynucleotides encoding such proteins in a biological sample (for example, blood, sera, urine, and/or tumor biopsies) obtained from the patient. In other words, such proteins may be used as markers to indicate the presence or absence of a disease or disorder, including cancer and/or as described elsewhere herein. In addition, such proteins may be useful for the detection of other diseases and cancers. The binding agents provided herein generally permit detection of the level of antigen that binds to the agent in the biological sample. Polynucleotide primers and probes may be used to detect the level of mRNA encoding polypeptides of the invention, which is also indicative of the presence or absence of a disease or disorder, including cancer. In general, polypeptides of the invention should be present at a level that is at least three fold higher in diseased tissue than in normal tissue.

[0421] There are a variety of assay formats known to those of ordinary skill in the art for using a binding agent to detect polypeptide markers in a sample. See, e.g., Harlow and Lane, supra. In general, the presence or absence of a disease in a patient may be determined by (a) contacting a biological sample obtained from a patient with a binding agent; (b) detecting in the sample a level of polypeptide that binds to the binding agent; and (c) comparing the level of polypeptide with a predetermined cut-off value.

[0422] In a preferred embodiment, the assay involves the use of a binding agent(s) immobilized on a solid support to bind to and remove the polypeptide of the invention from the remainder of the sample. The bound polypeptide may then be detected using a detection reagent that contains a reporter group and specifically binds to the binding agent/polypeptide complex. Such detection reagents may comprise, for example, a binding agent that specifically binds to the polypeptide or an antibody or other agent that specifically binds to the binding agent, such as an anti-immunoglobulin, protein G, protein A or a lectin. Alternatively, a competitive assay may be utilized, in which a polypeptide is labeled with a reporter group and allowed to bind to the immobilized binding agent after incubation of the binding agent with the sample. The extent to which components of the sample inhibit the binding of the labeled polypeptide to the binding agent is indicative of the reactivity of the sample with the immobilized binding agent. Suitable polypeptides for use within such assays include polypeptides of the invention and portions thereof, or antibodies, to which the binding agent binds, as described above.

[0423] The solid support may be any material known to those of skill in the art to which polypeptides of the invention may be attached. For example, the solid support may be a test well in a microtiter plate or a nitrocellulose or other suitable membrane. Alternatively, the support may be a bead or disc, such as glass fiberglass, latex or a plastic material such as polystyrene or polyvinylchloride. The support may also be a magnetic particle or a fiber optic sensor, such as those disclosed, for example, in U.S. Pat. No. 5,359,681. The binding agent may be immobilized on the solid support using a variety of techniques known to those of skill in the art, which are amply described in the patent and scientific literature. In the context of the present invention, the term “immobilization” refers to both noncovalent association, such as adsorption, and covalent attachment (which may be a direct linkage between the agent and functional groups on the support or may be a linkage by way of a cross-linking agent). Immobilization by adsorption to a well in a microtiter plate or to a membrane is preferred. In such cases, adsorption may be achieved by contacting the binding agent, in a suitable buffer, with the solid support for the suitable amount of time. The contact time varies with temperature, but is typically between about 1 hour and about 1 day. In general, contacting a well of plastic microtiter plate (such as polystyrene or polyvinylchloride) with an amount of binding agent ranging from about 10 ng to about 10 ug, and preferably about 100 ng to about 1 ug, is sufficient to immobilize an adequate amount of binding agent.

[0424] Covalent attachment of binding agent to a solid support may generally be achieved by first reacting the support with a bifunctional reagent that will react with both the support and a functional group, such as a hydroxyl or amino group, on the binding agent. For example, the binding agent may be covalently attached to supports having an appropriate polymer coating using benzoquinone or by condensation of an aldehyde group on the support with an amine and an active hydrogen on the binding partner (see, e.g., Pierce Immunotechnology Catalog and Handbook, 1991, at A12-A13).

[0425] Gene Therapy Methods

[0426] Also encompassed by the invention are gene therapy methods for treating or preventing disorders, diseases and conditions. The gene therapy methods relate to the introduction of nucleic acid (DNA, RNA and antisense DNA or RNA) sequences into an animal to achieve expression of the polypeptide of the present invention. This method requires a polynucleotide which codes for a polypeptide of the present invention operatively linked to a promoter and any other genetic elements necessary for the expression of the polypeptide by the target tissue. Such gene therapy and delivery techniques are known in the art, see, for example, WO90/11092, which is herein incorporated by reference.

[0427] Thus, for example, cells from a patient may be engineered with a polynucleotide (DNA or RNA) comprising a promoter operably linked to a polynucleotide of the present invention ex vivo, with the engineered cells then being provided to a patient to be treated with the polypeptide of the present invention. Such methods are well-known in the art. For example, see Belldegrun, A., et al., J. Natl. Cancer Inst. 85: 207-216 (1993); Ferrantini, M. et al., Cancer Research 53: 1107-1112 (1993); Ferrantini, M. et al., J. Immunology 153: 4604-4615 (1994); Kaido, T., et al., Int. J. Cancer 60: 221-229 (1995); Ogura, H., et al., Cancer Research 50: 5102-5106 (1990); Santodonato, L., et al., Human Gene Therapy 7:1-10 (1996); Santodonato, L., et al., Gene Therapy 4:1246-1255 (1997); and Zbang, J.-F. et al., Cancer Gene Therapy 3: 31-38 (1996)), which are herein incorporated by reference. In one embodiment, the cells which are engineered are arterial cells. The arterial cells may be reintroduced into the patient through direct injection to the artery, the tissues surrounding the artery, or through catheter injection.

[0428] As discussed in more detail below, the polynucleotide constructs can be delivered by any method that delivers injectable materials to the cells of an animal, such as, injection into the interstitial space of tissues (heart, muscle, skin, lung, liver, and the like). The polynucleotide constructs may be delivered in a pharmaceutically acceptable liquid or aqueous carrier.

[0429] In one embodiment, the polynucleotide of the present invention is delivered as a naked polynucleotide. The term “naked” polynucleotide, DNA or RNA refers to sequences that are free from any delivery vehicle that acts to assist, promote or facilitate entry into the cell, including viral sequences, viral particles, liposome formulations, lipofectin or precipitating agents and the like. However, the polynucleotide of the present invention can also be delivered in liposome formulations and lipofectin formulations and the like can be prepared by methods well known to those skilled in the art. Such methods are described, for example, in U.S. Pat. Nos. 5,593,972, 5,589,466, and 5,580,859, which are herein incorporated by reference.

[0430] The polynucleotide vector constructs used in the gene therapy method are preferably constructs that will not integrate into the host genome nor will they contain sequences that allow for replication. Appropriate vectors include pWLNEO, pSV2CAT, pOG44, pXT1 and pSG available from Stratagene; pSVK3, pBPV, pMSG and pSVL available from Pharmacia; and pEF1/V5, pcDNA3.1, and pRc/CMV2 available from Invitrogen. Other suitable vectors will be readily apparent to the skilled artisan.

[0431] Any strong promoter known to those skilled in the art can be used for driving the expression of the polynucleotide sequence. Suitable promoters include adenoviral promoters, such as the adenoviral major late promoter; or heterologous promoters, such as the cytomegalovirus (CMV) promoter; the respiratory syncytial virus (RSV) promoter; inducible promoters, such as the MMT promoter, the metallothionein promoter; heat shock promoters; the albumin promoter; the ApoAI promoter; human globin promoters; viral thymidine kinase promoters, such as the Herpes Simplex thymidine kinase promoter; retroviral LTRs; the b-actin promoter; and human growth hormone promoters. The promoter also may be the native promoter for the polynucleotide of the present invention.

[0432] Unlike other gene therapy techniques, one major advantage of introducing naked nucleic acid sequences into target cells is the transitory nature of the polynucleotide synthesis in the cells. Studies have shown that non-replicating DNA sequences can be introduced into cells to provide production of the desired polypeptide for periods of up to six months.

[0433] The polynucleotide construct can be delivered to the interstitial space of tissues within the an animal, including of muscle, skin, brain, lung, liver, spleen, bone marrow, thymus, heart, lymph, blood, bone, cartilage, pancreas, kidney, gall bladder, stomach, intestine, testis, ovary, uterus, rectum, nervous system, eye, gland, and connective tissue. Interstitial space of the tissues comprises the intercellular, fluid, mucopolysaccharide matrix among the reticular fibers of organ tissues, elastic fibers in the walls of vessels or chambers, collagen fibers of fibrous tissues, or that same matrix within connective tissue ensheathing muscle cells or in the lacunae of bone. It is similarly the space occupied by the plasma of the circulation and the lymph fluid of the lymphatic channels. Delivery to the interstitial space of muscle tissue is preferred for the reasons discussed below. They may be conveniently delivered by injection into the tissues comprising these cells. They are preferably delivered to and expressed in persistent, non-dividing cells which are differentiated, although delivery and expression may be achieved in non-differentiated or less completely differentiated cells, such as, for example, stem cells of blood or skin fibroblasts. In vivo muscle cells are particularly competent in their ability to take up and express polynucleotides.

[0434] For the naked nucleic acid sequence injection, an effective dosage amount of DNA or RNA will be in the range of from about 0.05 mg/kg body weight to about 50 mg/kg body weight. Preferably the dosage will be from about 0.005 mg/kg to about 20 mg/kg and more preferably from about 0.05 mg/kg to about 5 mg/kg. Of course, as the artisan of ordinary skill will appreciate, this dosage will vary according to the tissue site of injection. The appropriate and effective dosage of nucleic acid sequence can readily be determined by those of ordinary skill in the art and may depend on the condition being treated and the route of administration.

[0435] The preferred route of administration is by the parenteral route of injection into the interstitial space of tissues. However, other parenteral routes may also be used, such as, inhalation of an aerosol formulation particularly for delivery to lungs or bronchial tissues, throat or mucous membranes of the nose. In addition, naked DNA constructs can be delivered to arteries during angioplasty by the catheter used in the procedure.

[0436] The naked polynucleotides are delivered by any method known in the art, including, but not limited to, direct needle injection at the delivery site, intravenous injection, topical administration, catheter infusion, and so-called “gene guns”. These delivery methods are known in the art.

[0437] The constructs may also be delivered with delivery vehicles such as viral sequences, viral particles, liposome formulations, lipofectin, precipitating agents, etc. Such methods of delivery are known in the art.

[0438] In certain embodiments, the polynucleotide constructs are complexed in a liposome preparation. Liposomal preparations for use in the instant invention include cationic (positively charged), anionic (negatively charged) and neutral preparations. However, cationic liposomes are particularly preferred because a tight charge complex can be formed between the cationic liposome and the polyanionic nucleic acid. Cationic liposomes have been shown to mediate intracellular delivery of plasmid DNA (Felgner et al., Proc. Natl. Acad. Sci. USA (1987) 84:7413-7416, which is herein incorporated by reference); mRNA (Malone et al., Proc. Natl. Acad. Sci. USA (1989) 86:6077-6081, which is herein incorporated by reference); and purified transcription factors (Debs et al., J. Biol. Chem. (1990) 265:10189-10192, which is herein incorporated by reference), in functional form.

[0439] Cationic liposomes are readily available. For example, N[1-2,3-dioleyloxy)propyl]-N,N,N-triethylammonium (DOTMA) liposomes are particularly useful and are available under the trademark Lipofectin, from GIBCO BRL, Grand Island, N.Y. (See, also, Felgner et al., Proc. Natl. Acad. Sci. USA (1987) 84:7413-7416, which is herein incorporated by reference). Other commercially available liposomes include transfectace (DDAB/DOPE) and DOTAP/DOPE (Boehringer).

[0440] Other cationic liposomes can be prepared from readily available materials using techniques well known in the art. See, e.g. PCT Publication No. WO 90/11092 (which is herein incorporated by reference) for a description of the synthesis of DOTAP (1,2-bis(oleoyloxy)-3-(trimethylammonio)propane) liposomes. Preparation of DOTMA liposomes is explained in the literature, see, e.g., P. Felgner et al., Proc. Natl. Acad. Sci. USA 84:7413-7417, which is herein incorporated by reference. Similar methods can be used to prepare liposomes from other cationic lipid materials.

[0441] Similarly, anionic and neutral liposomes are readily available, such as from Avanti Polar Lipids (Birmingham, Ala.), or can be easily prepared using readily available materials. Such materials include phosphatidyl, choline, cholesterol, phosphatidyl ethanolamine, dioleoylphosphatidyl choline (DOPC), dioleoylphosphatidyl glycerol (DOPG), dioleoylphoshatidyl ethanolamine (DOPE), among others. These materials can also be mixed with the DOTMA and DOTAP starting materials in appropriate ratios. Methods for making liposomes using these materials are well known in the art.

[0442] For example, commercially dioleoylphosphatidyl choline (DOPC), dioleoylphosphatidyl glycerol (DOPG), and dioleoylphosphatidyl ethanolamine (DOPE) can be used in various combinations to make conventional liposomes, with or without the addition of cholesterol. Thus, for example, DOPG/DOPC vesicles can be prepared by drying 50 mg each of DOPG and DOPC under a stream of nitrogen gas into a sonication vial. The sample is placed under a vacuum pump overnight and is hydrated the following day with deionized water. The sample is then sonicated for 2 hours in a capped vial, using a Heat Systems model 350 sonicator equipped with an inverted cup (bath type) probe at the maximum setting while the bath is circulated at 15EC. Alternatively, negatively charged vesicles can be prepared without sonication to produce multilamellar vesicles or by extrusion through nucleopore membranes to produce unilamellar vesicles of discrete size. Other methods are known and available to those of skill in the art.

[0443] The liposomes can comprise multilamellar vesicles (MLVs), small unilamellar vesicles (SUVs), or large unilamellar vesicles (LUVs), with SUVs being preferred. The various liposome-nucleic acid complexes are prepared using methods well known in the art. See, e.g., Straubinger et al., Methods of Immunology (1983), 101:512-527, which is herein incorporated by reference. For example, MLVs containing nucleic acid can be prepared by depositing a thin film of phospholipid on the walls of a glass tube and subsequently hydrating with a solution of the material to be encapsulated. SUVs are prepared by extended sonication of MLVs to produce a homogeneous population of unilamellar liposomes. The material to be entrapped is added to a suspension of preformed MLVs and then sonicated. When using liposomes containing cationic lipids, the dried lipid film is resuspended in an appropriate solution such as sterile water or an isotonic buffer solution such as 10 mM Tris/NaCl, sonicated, and then the preformed liposomes are mixed directly with the DNA. The liposome and DNA form a very stable complex due to binding of the positively charged liposomes to the cationic DNA. SUVs find use with small nucleic acid fragments. LUVs are prepared by a number of methods, well known in the art. Commonly used methods include Ca2+-EDTA chelation (Papahadjopoulos et al., Biochim. Biophys. Acta (1975) 394:483; Wilson et al., Cell 17:77 (1979)); ether injection (Deamer, D. and Bangham, A., Biochim. Biophys. Acta 443:629 (1976); Ostro et al., Biochem. Biophys. Res. Commun. 76:836 (1977); Fraley et al., Proc. Natl. Acad. Sci. USA 76:3348 (1979)); detergent dialysis (Enoch, H. and Strittmatter, P., Proc. Natl. Acad. Sci. USA 76:145 (1979)); and reverse-phase evaporation (REV) (Fraley et al., J. Biol. Chem. 255:10431 (1980); Szoka, F. and Papahadjopoulos, D., Proc. Natl. Acad. Sci. USA 75:145 (1978); Schaefer-Ridder et al., Science 215:166 (1982)), which are herein incorporated by reference.

[0444] Generally, the ratio of DNA to liposomes will be from about 10:1 to about 1:10. Preferably, the ration will be from about 5:1 to about 1:5. More preferably, the ration will be about 3:1 to about 1:3. Still more preferably, the ratio will be about 1:1.

[0445] U.S. Pat. No. 5,676,954 (which is herein incorporated by reference) reports on the injection of genetic material, complexed with cationic liposomes carriers, into mice. U.S. Pat. Nos. 4,897,355, 4,946,787, 5,049,386, 5,459,127, 5,589,466, 5,693,622, 5,580,859, 5,703,055, and international publication no. WO 94/9469 (which are herein incorporated by reference) provide cationic lipids for use in transfecting DNA into cells and mammals. U.S. Pat. Nos. 5,589,466, 5,693,622, 5,580,859, 5,703,055, and international publication no. WO 94/9469 provide methods for delivering DNA-cationic lipid complexes to mammals.

[0446] In certain embodiments, cells are engineered, ex vivo or in vivo, using a retroviral particle containing RNA which comprises a sequence encoding a polypeptide of the present invention. Retroviruses from which the retroviral plasmid vectors may be derived include, but are not limited to, Moloney Murine Leukemia Virus, spleen necrosis virus, Rous sarcoma Virus, Harvey Sarcoma Virus, avian leukosis virus, gibbon ape leukemia virus, human immunodeficiency virus, Myeloproliferative Sarcoma Virus, and mammary tumor virus.

[0447] The retroviral plasmid vector is employed to transduce packaging cell lines to form producer cell lines. Examples of packaging cells which may be transfected include, but are not limited to, the PE501, PA317, R-2, R-AM, PA12, T19-14×, VT-19-17-H2, RCRE, RCRIP, GP+E-86, GP+envAm12, and DAN cell lines as described in Miller, Human Gene Therapy 1:5-14 (1990), which is incorporated herein by reference in its entirety. The vector may transduce the packaging cells through any means known in the art. Such means include, but are not limited to, electroporation, the use of liposomes, and CaPO4 precipitation. In one alternative, the retroviral plasmid vector may be encapsulated into a liposome, or coupled to a lipid, and then administered to a host.

[0448] The producer cell line generates infectious retroviral vector particles which include polynucleotide encoding a polypeptide of the present invention. Such retroviral vector particles then may be employed, to transduce eukaryotic cells, either in vitro or in vivo. The transduced eukaryotic cells will express a polypeptide of the present invention.

[0449] In certain other embodiments, cells are engineered, ex vivo or in vivo, with polynucleotide contained in an adenovirus vector. Adenovirus can be manipulated such that it encodes and expresses a polypeptide of the present invention, and at the same time is inactivated in terms of its ability to replicate in a normal lytic viral life cycle. Adenovirus expression is achieved without integration of the viral DNA into the host cell chromosome, thereby alleviating concerns about insertional mutagenesis. Furthermore, adenoviruses have been used as live enteric vaccines for many years with an excellent safety profile (Schwartz et al. Am. Rev. Respir. Dis.109:233-238 (1974)). Finally, adenovirus mediated gene transfer has been demonstrated in a number of instances including transfer of alpha-1-antitrypsin and CFTR to the lungs of cotton rats (Rosenfeld, M. A. et al. (1991) Science 252:431-434; Rosenfeld et al., (1992) Cell 68:143-155). Furthermore, extensive studies to attempt to establish adenovirus as a causative agent in human cancer were uniformly negative (Green, M. et al. (1979) Proc. Natl. Acad. Sci. USA 76:6606).

[0450] Suitable adenoviral vectors useful in the present invention are described, for example, in Kozarsky and Wilson, Curr. Opin. Genet. Devel. 3:499-503 (1993); Rosenfeld et al., Cell 68:143-155 (1992); Engelhardt et al., Human Genet. Ther. 4:759-769 (1993); Yang et al., Nature Genet. 7:362-369 (1994); Wilson et al., Nature 365:691-692 (1993); and U.S. Pat. No. 5,652,224, which are herein incorporated by reference. For example, the adenovirus vector Ad2 is useful and can be grown in human 293 cells. These cells contain the E1 region of adenovirus and constitutively express E1a and E1b, which complement the defective adenoviruses by providing the products of the genes deleted from the vector. In addition to Ad2, other varieties of adenovirus (e.g., Ad3, Ad5, and Ad7) are also useful in the present invention.

[0451] Preferably, the adenoviruses used in the present invention are replication deficient. Replication deficient adenoviruses require the aid of a helper virus and/or packaging cell line to form infectious particles. The resulting virus is capable of infecting cells and can express a polynucleotide of interest which is operably linked to a promoter, but cannot replicate in most cells. Replication deficient adenoviruses may be deleted in one or more of all or a portion of the following genes: E1a, E1b, E3, E4, E2a, or L1 through L5.

[0452] In certain other embodiments, the cells are engineered, ex vivo or in vivo, using an adeno-associated virus (AAV). AAVs are naturally occurring defective viruses that require helper viruses to produce infectious particles (Muzyczka, N., Curr. Topics in Microbiol. immunol. 158:97 (1992)). It is also one of the few viruses that may integrate its DNA into non-dividing cells. Vectors containing as little as 300 base pairs of AAV can be packaged and can integrate, but space for exogenous DNA is limited to about 4.5 kb. Methods for producing and using such AAVs are known in the art. See, for example, U.S. Pat. Nos. 5,139,941, 5,173,414, 5,354,678, 5,436,146, 5,474,935, 5,478,745, and 5,589,377.

[0453] For example, an appropriate AAV vector for use in the present invention will include all the sequences necessary for DNA replication, encapsidation, and host-cell integration. The polynucleotide construct is inserted into the AAV vector using standard cloning methods, such as those found in Sambrook et al., Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Press (1989). The recombinant AAV vector is then transfected into packaging cells which are infected with a helper virus, using any standard technique, including lipofection, electroporation, calcium phosphate precipitation, etc. Appropriate helper viruses include adenoviruses, cytomegaloviruses, vaccinia viruses, or herpes viruses. Once the packaging cells are transfected and infected, they will produce infectious AAV viral particles which contain the polynucleotide construct. These viral particles are then used to transduce eukaryotic cells, either ex vivo or in vivo. The transduced cells will contain the polynucleotide construct integrated into its genome, and will express a polypeptide of the invention.

[0454] Another method of gene therapy involves operably associating heterologous control regions and endogenous polynucleotide sequences (e.g. encoding a polypeptide of the present invention) via homologous recombination (see, e.g., U.S. Pat. No. 5,641,670, issued Jun. 24, 1997; International Publication No. WO 96/29411, published Sep. 26, 1996; International Publication No. WO 94/12650, published Aug. 4, 1994; Koller et al., Proc. Natl. Acad. Sci. USA 86:8932-8935 (1989); and Zijlstra et al., Nature 342:435-438 (1989), which are herein encorporated by reference. This method involves the activation of a gene which is present in the target cells, but which is not normally expressed in the cells, or is expressed at a lower level than desired.

[0455] Polynucleotide constructs are made, using standard techniques known in the art, which contain the promoter with targeting sequences flanking the promoter. Suitable promoters are described herein. The targeting sequence is sufficiently complementary to an endogenous sequence to permit homologous recombination of the promoter-targeting sequence with the endogenous sequence. The targeting sequence will be sufficiently near the 5′ end of the desired endogenous polynucleotide sequence so the promoter will be operably linked to the endogenous sequence upon homologous recombination.

[0456] The promoter and the targeting sequences can be amplified using PCR. Preferably, the amplified promoter contains distinct restriction enzyme sites on the 5′ and 3′ ends. Preferably, the 3′ end of the first targeting sequence contains the same restriction enzyme site as the 5′ end of the amplified promoter and the 5′ end of the second targeting sequence contains the same restriction site as the 3′ end of the amplified promoter. The amplified promoter and targeting sequences are digested and ligated together.

[0457] The promoter-targeting sequence construct is delivered to the cells, either as naked polynucleotide, or in conjunction with transfection-facilitating agents, such as liposomes, viral sequences, viral particles, whole viruses, lipofection, precipitating agents, etc., described in more detail above. The P promoter-targeting sequence can be delivered by any method, included direct needle injection, intravenous injection, topical administration, catheter infusion, particle accelerators, etc. The methods are described in more detail below.

[0458] The promoter-targeting sequence construct is taken up by cells. Homologous recombination between the construct and the endogenous sequence takes place, such that an endogenous sequence is placed under the control of the promoter. The promoter then drives the expression of the endogenous sequence.

[0459] The polynucleotide encoding a polypeptide of the present invention may contain a secretory signal sequence that facilitates secretion of the protein. Typically, the signal sequence is positioned in the coding region of the polynucleotide to be expressed towards or at the 5′ end of the coding region. The signal sequence may be homologous or heterologous to the polynucleotide of interest and may be homologous or heterologous to the cells to be transfected. Additionally, the signal sequence may be chemically synthesized using methods known in the art.

[0460] Any mode of administration of any of the above-described polynucleotides constructs can be used so long as the mode results in the expression of one or more molecules in an amount sufficient to provide a therapeutic effect. This includes direct needle injection, systemic injection, catheter infusion, biolistic injectors, particle accelerators (i.e., “gene guns”), gelfoam sponge depots, other commercially available depot materials, osmotic pumps (e.g., Alza minipumps), oral or suppositorial solid (tablet or pill) pharmaceutical formulations, and decanting or topical applications during surgery. For example, direct injection of naked calcium phosphate-precipitated plasmid into rat liver and rat spleen or a protein-coated plasmid into the portal vein has resulted in gene expression of the foreign gene in the rat livers (Kaneda et al., Science 243:375 (1989)).

[0461] A preferred method of local administration is by direct injection. Preferably, a recombinant molecule of the present invention complexed with a delivery vehicle is administered by direct injection into or locally within the area of arteries. Administration of a composition locally within the area of arteries refers to injecting the composition centimeters and preferably, millimeters within arteries.

[0462] Another method of local administration is to contact a polynucleotide construct of the present invention in or around a surgical wound. For example, a patient can undergo surgery and the polynucleotide construct can be coated on the surface of tissue inside the wound or the construct can be injected into areas of tissue inside the wound.

[0463] Therapeutic compositions useful in systemic administration, include recombinant molecules of the present invention complexed to a targeted delivery vehicle of the present invention. Suitable delivery vehicles for use with systemic administration comprise liposomes comprising ligands for targeting the vehicle to a particular site. In specific embodiments, suitable delivery vehicles for use with systemic administration comprise liposomes comprising polypeptides of the invention for targeting the vehicle to a particular site.

[0464] Preferred methods of systemic administration, include intravenous injection, aerosol, oral and percutaneous (topical) delivery. Intravenous injections can be performed using methods standard in the art. Aerosol delivery can also be performed using methods standard in the art (see, for example, Stribling et al., Proc. Natl. Acad. Sci. USA 189:11277-11281, 1992, which is incorporated herein by reference). Oral delivery can be performed by complexing a polynucleotide construct of the present invention to a carrier capable of withstanding degradation by digestive enzymes in the gut of an animal. Examples of such carriers, include plastic capsules or tablets, such as those known in the art. Topical delivery can be performed by mixing a polynucleotide construct of the present invention with a lipophilic reagent (e.g., DMSO) that is capable of passing into the skin.

[0465] Determining an effective amount of substance to be delivered can depend upon a number of factors including, for example, the chemical structure and biological activity of the substance, the age and weight of the animal, the precise condition requiring treatment and its severity, and the route of administration. The frequency of treatments depends upon a number of factors, such as the amount of polynucleotide constructs administered per dose, as well as the health and history of the subject. The precise amount, number of doses, and timing of doses will be determined by the attending physician or veterinarian.

[0466] Therapeutic compositions of the present invention can be administered to any animal, preferably to mammals and birds. Preferred mammals include humans, dogs, cats, mice, rats, rabbits sheep, cattle, horses and pigs, with humans being particularly preferred.

[0467] Biological Activities

[0468] Polynucleotides or polypeptides, or agonists or antagonists of the present invention, can be used in assays to test for one or more biological activities. If these polynucleotides or polypeptides, or agonists or antagonists of the present invention, do exhibit activity in a particular assay, it is likely that these molecules may be involved in the diseases associated with the biological activity. Thus, the polynucleotides and polypeptides, and agonists or antagonists could be used to treat the associated disease.

[0469] Nervous system-associated proteins are believed to be involved in biological activities associated with nervous system functions. Accordingly, compositions of the invention (including polynucleotides, polypeptides and antibodies of the invention, and fragments and variants thereof) may be used in the diagnosis, prognosis, prevention, and/or treatment of diseases and/or disorders associated with inappropriate nervous system activity.

[0470] In preferred embodiments, compositions of the invention (including polynucleotides, polypeptides and antibodies of the invention, and fragments and variants thereof) may be used in the diagnosis, prognosis, prevention, and/or treatment of diseases and/or disorders relating to the nervous system, including, for example, cerebrovascular disorders (e.g., stroke, cerebral aneurysm, and/or as described below under “Cardiovascular Disorders” and “Neural Activity and Neurological Diseases”), metabolic brain diseases (e.g., Hartnup disease, hepatic encephalopathy, phenylketonuria, pyruvate carboxylase deficiency, and/or as described below in the section entitled “Neural Activity and Neurological Diseases”), neuromuscular disorders (e.g., amyotrophic lateral sclerosis, muscular dystrophy, myotonia atrophica, familial periodic paralysis, and/or as described below in the section entitled “Neural Activity and Neurological Diseases”), nervous system infections (e.g., AIDS dementia complex, encephalitis, meningitis, cerebral malaria, and/or as described below in the section entitled “Neural Activity and Neurological Diseases”), demyelinating diseases (e.g., multiple sclerosis, Canavan disease, metachromatic eukodystrophy, and/or as described below in the section entitled “Neural Activity and Neurological Diseases”), nervous system neoplasms (e.g., cerebellar neoplasms, infratenorial neoplasms, supratentorial neoplasms, and/or as described below in the section entitled “Neural Activity and Neurological Diseases”), spinal cord diseases (e.g., spinal muscular atrophy, Werdnig-Hoffmann disease, myelitis, and/or as described below in the section entitled “Neural Activity and Neurological Diseases”), developmental nervous system abnormalities (e.g., spinal dysraphism, neural tube defects, holoprosencephaly, and/or as described below in the section entitled “Neural Activity and Neurological Diseases”), neurologic manifestations (agnosia, dyslexia, hallucinations, ataxia, and/or as described below in the section entitled “Neural Activity and Neurological Diseases”), diseases of the peripheral nervous system (e.g., cranial nerve diseases, facial nerve disease, optic nerve diseases, and/or as described below in the section entitled “Neural Activity and Neurological Diseases”), inflammatory disorders of the nervous system (e.g., encephalitis, meningitis, and/or those described under “Immune Activity” and “Neural Activity and Neurological Diseases” below), and others described below in the section entitled “Neural Activity and Neurological Diseases”.

[0471] In another preferred embodiment, compositions of the invention (including polynucleotides, polypeptides and antibodies of the invention, and fragments and variants thereof) may be used in the diagnosis, prognosis, prevention, and/or treatment of traumatic injury to the nervous system (e.g., traumatic head injury, spinal cord injury, peripheral nerve damage, and/or as described in the section entitled “Neural Activity and Neurological Diseases”).

[0472] In another embodiment, a polypeptide of the invention, or polynucleotides, antibodies, agonists, or antagonists corresponding to that polypeptide, may be used to diagnose, prognose, prevent, and/or treat neurological disorders or injuries associated with the tissue(s) in which the polypeptide of the invention is expressed, including one, two, three, four, five, or more tissues disclosed in Table 1A, column 8 (Tissue Distribution Library Code).

[0473] Thus, polynucleotides, translation products and antibodies of the invention are useful in the diagnosis, detection and/or treatment of diseases and/or disorders associated with activities that include, but are not limited to, neuromuscular disorders, infections of the nervous system, demyelinating diseases, cerebrovascular disorders, nervous system neoplasms, spinal cord diseases, inflammatory disorders of the nervous system, traumatic injury, nervous system abnormalities, neurologic manifestations, diseases of the peripheral nervous system, and metabolic brain diseases.

[0474] More generally, polynucleotides, translation products and antibodies corresponding to this gene may be useful for the diagnosis, detection and/or treatment of diseases and/or disorders associated with the following systems.

[0475] Neural Activity and Neurological Diseases

[0476] The polynucleotides, polypeptides and agonists or antagonists of the invention may be used for the diagnosis and/or treatment of diseases, disorders, damage or injury of the brain and/or nervous system. Nervous system disorders that can be treated with the compositions of the invention (e.g., polypeptides, polynucleotides, and/or agonists or antagonists), include, but are not limited to, nervous system injuries, and diseases or disorders which result in either a disconnection of axons, a diminution or degeneration of neurons, or demyelination. Nervous system lesions which may be treated in a patient (including human and non-human mammalian patients) according to the methods of the invention, include but are not limited to, the following lesions of either the central (including spinal cord, brain) or peripheral nervous systems: (1) ischemic lesions, in which a lack of oxygen in a portion of the nervous system results in neuronal injury or death, including cerebral infarction or ischemia, or spinal cord infarction or ischemia; (2) traumatic lesions, including lesions caused by physical injury or associated with surgery, for example, lesions which sever a portion of the nervous system, or compression injuries; (3) malignant lesions, in which a portion of the nervous system is destroyed or injured by malignant tissue which is either a nervous system associated malignancy or a malignancy derived from non-nervous system tissue; (4) infectious lesions, in which a portion of the nervous system is destroyed or injured as a result of infection, for example, by an abscess or associated with infection by human immunodeficiency virus, herpes zoster, or herpes simplex virus or with Lyme disease, tuberculosis, or syphilis; (5) degenerative lesions, in which a portion of the nervous system is destroyed or injured as a result of a degenerative process including but not limited to, degeneration associated with Parkinson's disease, Alzheimer's disease, Huntington's chorea, or amyotrophic lateral sclerosis (ALS); (6) lesions associated with nutritional diseases or disorders, in which a portion of the nervous system is destroyed or injured by a nutritional disorder or disorder of metabolism including, but not limited to, vitamin B12 deficiency, folic acid deficiency, Wernicke disease, tobacco-alcohol amblyopia, Marchiafava-Bignami disease (primary degeneration of the corpus callosum), and alcoholic cerebellar degeneration; (7) neurological lesions associated with systemic diseases including, but not limited to, diabetes (diabetic neuropathy, Bell's palsy), systemic lupus erythematosus, carcinoma, or sarcoidosis; (8) lesions caused by toxic substances including alcohol, lead, or particular neurotoxins; and (9) demyelinated lesions in which a portion of the nervous system is destroyed or injured by a demyelinating disease including, but not limited to, multiple sclerosis, human immunodeficiency virus-associated myelopathy, transverse myclopathy or various etiologies, progressive multifocal leukoencephalopathy, and central pontine myelinolysis.

[0477] In one embodiment, the polypeptides, polynucleotides, or agonists or antagonists of the invention are used to protect neural cells from the damaging effects of hypoxia. In a further preferred embodiment, the polypeptides, polynucleotides, or agonists or antagonists of the invention are used to protect neural cells from the damaging effects of cerebral hypoxia. According to this embodiment, the compositions of the invention are used to treat or prevent neural cell injury associated with cerebral hypoxia. In one non-exclusive aspect of this embodiment, the polypeptides, polynucleotides, or agonists or antagonists of the invention, are used to treat or prevent neural cell injury associated with cerebral ischemia. In another non-exclusive aspect of this embodiment, the polypeptides, polynucleotides, or agonists or antagonists of the invention are used to treat or prevent neural cell injury associated with cerebral infarction.

[0478] In another preferred embodiment, the polypeptides, polynucleotides, or agonists or antagonists of the invention are used to treat or prevent neural cell injury associated with a stroke. In a specific embodiment, the polypeptides, polynucleotides, or agonists or antagonists of the invention are used to treat or prevent cerebral neural cell injury associated with a stroke.

[0479] In another preferred embodiment, the polypeptides, polynucleotides, or agonists or antagonists of the invention are used to treat or prevent neural cell injury associated with a heart attack. In a specific embodiment, the polypeptides, polynucleotides, or agonists or antagonists of the invention are used to treat or prevent cerebral neural cell injury associated with a heart attack.

[0480] The compositions of the invention which are useful for treating or preventing a nervous system disorder may be selected by testing for biological activity in promoting the survival or differentiation of neurons. For example, and not by way of limitation, compositions of the invention which elicit any of the following effects may be useful according to the invention: (1) increased survival time of neurons in culture either in the presence or absence of hypoxia or hypoxic conditions; (2) increased sprouting of neurons in culture or in vivo; (3) increased production of a neuron-associated molecule in culture or in vivo, e.g., choline acetyltransferase or acetylcholinesterase with respect to motor neurons; or (4) decreased symptoms of neuron dysfunction in vivo. Such effects may be measured by any method known in the art. In preferred, non-limiting embodiments, increased survival of neurons may routinely be measured using a method set forth herein or otherwise known in the art, such as, for example, in Zhang et al., Proc Natl Acad Sci USA 97:3637-42 (2000) or in Arakawa et al., J. Neurosci., 10:3507-15 (1990); increased sprouting of neurons may be detected by methods known in the art, such as, for example, the methods set forth in Pestronk et al., Exp. Neurol., 70:65-82 (1980), or Brown et al, Ann. Rev. Neurosci., 4:17-42 (1981); increased production of neuron-associated molecules may be measured by bioassay, enzymatic assay, antibody binding, Northern blot assay, etc., using techniques known in the art and depending on the molecule to be measured; and motor neuron dysfunction may be measured by assessing the physical manifestation of motor neuron disorder, e.g., weakness, motor neuron conduction velocity, or functional disability.

[0481] In specific embodiments, motor neuron disorders that may be treated according to the invention include, but are not limited to, disorders such as infarction, infection, exposure to toxin, trauma, surgical damage, degenerative disease or malignancy that may affect motor neurons as well as other components of the nervous system, as well as disorders that selectively affect neurons such as amyotrophic lateral sclerosis, and including, but not limited to, progressive spinal muscular atrophy, progressive bulbar palsy, primary lateral sclerosis, infantile and juvenile muscular atrophy, progressive bulbar paralysis of childhood (Fazio-Londe syndrome), poliomyelitis and the post polio syndrome, and Hereditary Motorsensory Neuropathy (Charcot-Marie-Tooth Disease).

[0482] Further, polypeptides or polynucleotides of the invention may play a role in neuronal survival; synapse formation; conductance; neural differentiation, etc. Thus, compositions of the invention (including polynucleotides, polypeptides, and agonists or antagonists) may be used to diagnose and/or treat or prevent diseases or disorders associated with these roles, including, but not limited to, learning and/or cognition disorders. The compositions of the invention may also be useful in the treatment or prevention of neurodegenerative disease states and/or behavioural disorders. Such neurodegenerative disease states and/or behavioral disorders include, but are not limited to, Alzheimer's Disease, Parkinson's Disease, Huntington's Disease, Tourette Syndrome, schizophrenia, mania, dementia, paranoia, obsessive compulsive disorder, panic disorder, learning disabilities, ALS, psychoses, autism, and altered behaviors, including disorders in feeding, sleep patterns, balance, and perception. In addition, compositions of the invention may also play a role in the treatment, prevention and/or detection of developmental disorders associated with the developing embryo, or sexually-linked disorders.

[0483] Additionally, polypeptides, polynucleotides and/or agonists or antagonists of the invention, may be useful in protecting neural cells from diseases, damage, disorders, or injury, associated with cerebrovascular disorders including, but not limited to, carotid artery diseases (e.g., carotid artery thrombosis, carotid stenosis, or Moyamoya Disease), cerebral amyloid angiopathy, cerebral aneurysm, cerebral anoxia, cerebral arteriosclerosis, cerebral arterioyenous malformations, cerebral artery diseases, cerebral embolism and thrombosis (e.g., carotid artery thrombosis, sinus thrombosis, or Wallenberg's Syndrome), cerebral hemorrhage (e.g., epidural or subdural hematoma, or subarachnoid hemorrhage), cerebral infarction, cerebral ischemia (e.g., transient cerebral ischemia, Subclavian Steal Syndrome, or vertebrobasilar insufficiency), vascular dementia (e.g., multi-infarct), leukomalacia, periventricular, and vascular headache (e.g., cluster headache or migraines).

[0484] In accordance with yet a further aspect of the present invention, there is provided a process for utilizing polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, for therapeutic purposes, for example, to stimulate neurological cell proliferation and/or differentiation. Therefore, polynucleotides, polypeptides, agonists and/or antagonists of the invention may be used to treat and/or detect neurologic diseases. Moreover, polynucleotides or polypeptides, or agonists or antagonists of the invention, can be used as a marker or detector of a particular nervous system disease or disorder.

[0485] Examples of neurologic diseases which can be treated or detected with polynucleotides, polypeptides, agonists, and/or antagonists of the present invention include brain diseases, such as metabolic brain diseases which includes phenylketonuria such as maternal phenylketonuria, pyruvate carboxylase deficiency, pyruvate dehydrogenase complex deficiency, Wernicke's Encephalopathy, brain edema, brain neoplasms such as cerebellar neoplasms which include infratentorial neoplasms, cerebral ventricle neoplasms such as choroid plexus neoplasms, hypothalamic neoplasms, supratentorial neoplasms, canavan disease, cerebellar diseases such as cerebellar ataxia which include spinocerebellar degeneration such as ataxia telangiectasia, cerebellar dyssynergia, Friederich's Ataxia, Machado-Joseph Disease, olivopontocerebellar atrophy, cerebellar neoplasms such as infratentorial neoplasms, diffuse cerebral sclerosis such as encephalitis periaxialis, globoid cell leukodystrophy, metachromatic leukodystrophy and subacute sclerosing panencephalitis.

[0486] Additional neurologic diseases which can be treated or detected with polynucleotides, polypeptides, agonists, and/or antagonists of the present invention include cerebrovascular disorders (such as carotid artery diseases which include carotid artery thrombosis, carotid stenosis and Moyamoya Disease), cerebral amyloid angiopathy, cerebral aneurysm, cerebral anoxia, cerebral arteriosclerosis, cerebral arterioyenous malformations, cerebral artery diseases, cerebral embolism and thrombosis such as carotid artery thrombosis, sinus thrombosis and Wallenberg's Syndrome, cerebral hemorrhage such as epidural hematoma, subdural hematoma and subarachnoid hemorrhage, cerebral infarction, cerebral ischemia such as transient cerebral ischemia, Subclavian Steal Syndrome and vertebrobasilar insufficiency, vascular dementia such as multi-infarct dementia, periventricular leukomalacia, vascular headache such as cluster headache and migraine.

[0487] Additional neurologic diseases which can be treated or detected with polynucleotides, polypeptides, agonists, and/or antagonists of the present invention include dementia such as AIDS Dementia Complex, presenile dementia such as Alzheimer's Disease and Creutzfeldt-Jakob Syndrome, senile dementia such as Alzheimer's Disease and progressive supranuclear palsy, vascular dementia such as multi-infarct dementia, encephalitis which include encephalitis periaxialis, viral encephalitis such as epidemic encephalitis, Japanese Encephalitis, St. Louis Encephalitis, tick-borne encephalitis and West Nile Fever, acute disseminated encephalomyelitis, meningoencephalitis such as uveomeningoencephalitic syndrome, Postencephalitic Parkinson Disease and subacute sclerosing panencephalitis, encephalomalacia such as periventricular leukomalacia, epilepsy such as generalized epilepsy which includes infantile spasms, absence epilepsy, myoclonic epilepsy which includes MERRF Syndrome, tonic-clonic epilepsy, partial epilepsy such as complex partial epilepsy, frontal lobe epilepsy and temporal lobe epilepsy, post-traumatic epilepsy, status epilepticus such as Epilepsia Partialis Continua, and Hallervorden-Spatz Syndrome.

[0488] Additional neurologic diseases which can be treated or detected with polynucleotides, polypeptides, agonists, and/or antagonists of the present invention include hydrocephalus such as Dandy-Walker Syndrome and normal pressure hydrocephalus, hypothalamic diseases such as hypothalamic neoplasms, cerebral malaria, narcolepsy which includes cataplexy, bulbar poliomyelitis, cerebri pseudotumor, Rett Syndrome, Reye's Syndrome, thalamic diseases, cerebral toxoplasmosis, intracranial tuberculoma and Zellweger Syndrome, central nervous system infections such as AIDS Dementia Complex, Brain Abscess, subdural empyema, encephalomyelitis such as Equine Encephalomyelitis, Venezuelan Equine Encephalomyelitis, Necrotizing Hemorrhagic Encephalomyelitis, Visna, and cerebral malaria.

[0489] Additional neurologic diseases which can be treated or detected with polynucleotides, polypeptides, agonists, and/or antagonists of the present invention include meningitis such as arachnoiditis, aseptic meningtitis such as viral meningtitis which includes lymphocytic choriomeningitis, Bacterial meningtitis which includes Haemophilus Meningtitis, Listeria Meningtitis, Meningococcal Meningtitis such as Waterhouse-Friderichsen Syndrome, Pneumococcal Meningtitis and meningeal tuberculosis, fungal meningitis such as Cryptococcal Meningtitis, subdural effusion, meningoencephalitis such as uvemeningoencephalitic syndrome, myelitis such as transverse myelitis, neurosyphilis such as tabes dorsalis, poliomyelitis which includes bulbar poliomyelitis and postpoliomyelitis syndrome, prion diseases (such as Creutzfeldt-Jakob Syndrome, Bovine Spongiform Encephalopathy, Gerstmann-Straussler Syndrome, Kuru, Scrapie), and cerebral toxoplasmosis.

[0490] Additional neurologic diseases which can be treated or detected with polynucleotides, polypeptides, agonists, and/or antagonists of the present invention include central nervous system neoplasms such as brain neoplasms that include cerebellar neoplasms such as infratentorial neoplasms, cerebral ventricle neoplasms such as choroid plexus neoplasms, hypothalamic neoplasms and supratentorial neoplasms, meningeal neoplasms, spinal cord neoplasms which include epidural neoplasms, demyelinating diseases such as Canavan Diseases, diffuse cerebral sceloris which includes adrenoleukodystrophy, encephalitis periaxialis, globoid cell leukodystrophy, diffuse cerebral sclerosis such as metachromatic leukodystrophy, allergic encephalomyelitis, necrotizing hemorrhagic encephalomyelitis, progressive multifocal leukoencephalopathy, multiple sclerosis, central pontine myelinolysis, transverse myelitis, neuromyelitis optica, Scrapie, Swayback, Chronic Fatigue Syndrome, Visna, High Pressure Nervous Syndrome, Meningism, spinal cord diseases such as amyotonia congenita, amyotrophic lateral sclerosis, spinal muscular atrophy such as Werdnig-Hoffmann Disease, spinal cord compression, spinal cord neoplasms such as epidural neoplasms, syringomyelia, Tabes Dorsalis, Stiff-Man Syndrome, mental retardation such as Angelman Syndrome, Cri-du-Chat Syndrome, De Lange's Syndrome, Down Syndrome, Gangliosidoses such as gangliosidoses G(M1), Sandhoff Disease, Tay-Sachs Disease, Hartnup Disease, homocystinuria, Laurence-Moon-Biedl Syndrome, Lesch-Nyhan Syndrome, Maple Syrup Urine Disease, mucolipidosis such as fucosidosis, neuronal ceroid-lipofuscinosis, oculocerebrorenal syndrome, phenylketonuria such as maternal phenylketonuria, Prader-Willi Syndrome, Rett Syndrome, Rubinstein-Taybi Syndrome, Tuberous Sclerosis, WAGR Syndrome, nervous system abnormalities such as holoprosencephaly, neural tube defects such as anencephaly which includes hydrangencephaly, Arnold-Chairi Deformity, encephalocele, meningocele, meningomyelocele, spinal dysraphism such as spina bifida cystica and spina bifida occulta.

[0491] Additional neurologic diseases which can be treated or detected with polynucleotides, polypeptides, agonists, and/or antagonists of the present invention include hereditary motor and sensory neuropathies which include Charcot-Marie Disease, Hereditary optic atrophy, Refsum's Disease, hereditary spastic paraplegia, Werdnig-Hoffmann Disease, Hereditary Sensory and Autonomic Neuropathies such as Congenital Analgesia and Familial Dysautonomia, Neurologic manifestations (such as agnosia that include Gerstmann's Syndrome, Amnesia such as retrograde amnesia, apraxia, neurogenic bladder, cataplexy, communicative disorders such as hearing disorders that includes deafness, partial hearing loss, loudness recruitment and tinnitus, language disorders such as aphasia which include agraphia, anomia, broca aphasia, and Wernicke Aphasia, Dyslexia such as Acquired Dyslexia, language development disorders, speech disorders such as aphasia which includes anomia, broca aphasia and Wernicke Aphasia, articulation disorders, communicative disorders such as speech disorders which include dysarthria, echolalia, mutism and stuttering, voice disorders such as aphonia and hoarseness, decerebrate state, delirium, fasciculation, hallucinations, meningism, movement disorders such as angelman syndrome, ataxia, athetosis, chorea, dystonia, hypokinesia, muscle hypotonia, myoclonus, tic, torticollis and tremor, muscle hypertonia such as muscle rigidity such as stiff-man syndrome, muscle spasticity, paralysis such as facial paralysis which includes Herpes Zoster Oticus, Gastroparesis, Hemiplegia, ophthalmoplegia such as diplopia, Duane's Syndrome, Horner's Syndrome, Chronic progressive external ophthalmoplegia such as Kearns Syndrome, Bulbar Paralysis, Tropical Spastic Paraparesis, Paraplegia such as Brown-Sequard Syndrome, quadriplegia, respiratory paralysis and vocal cord paralysis, paresis, phantom limb, taste disorders such as ageusia and dysgeusia, vision disorders such as amblyopia, blindness, color vision defects, diplopia, hemianopsia, scotoma and subnormal vision, sleep disorders such as hypersomnia which includes Kleine-Levin Syndrome, insomnia, and somnambulism, spasm such as trismus, unconsciousness such as coma, persistent vegetative state and syncope and vertigo, neuromuscular diseases such as amyotonia congenita, amyotrophic lateral sclerosis, Lambert-Eaton Myasthenic Syndrome, motor neuron disease, muscular atrophy such as spinal muscular atrophy, Charcot-Marie Disease and Werdnig-Hoffmann Disease, Postpoliomyelitis Syndrome, Muscular Dystrophy, Myasthenia Gravis, Myotonia Atrophica, Myotonia Confenita, Nemaline Myopathy, Familial Periodic Paralysis, Multiplex Paramyloclonus, Tropical Spastic Paraparesis and Stiff-Man Syndrome, peripheral nervous system diseases such as acrodynia, amyloid neuropathies, autonomic nervous system diseases such as Adie's Syndrome, Barre-Lieou Syndrome, Familial Dysautonomia, Homer's Syndrome, Reflex Sympathetic Dystrophy and Shy-Drager Syndrome, Cranial Nerve Diseases such as Acoustic Nerve Diseases such as Acoustic Neuroma which includes Neurofibromatosis 2, Facial Nerve Diseases such as Facial Neuralgia, Melkersson-Rosenthal Syndrome, ocular motility disorders which includes amblyopia, nystagmus, oculomotor nerve paralysis, ophthalmoplegia such as Duane's Syndrome, Homer's Syndrome, Chronic Progressive External Ophthalmoplegia which includes Kearns Syndrome, Strabismus such as Esotropia and Exotropia, Oculomotor Nerve Paralysis, Optic Nerve Diseases such as Optic Atrophy which includes Hereditary Optic Atrophy, Optic Disk Drusen, Optic Neuritis such as Neuromyclitis Optica, Papilledema, Trigeminal Neuralgia, Vocal Cord Paralysis, Demyelinating Diseases such as Neuromyelitis Optica and Swayback, and Diabetic neuropathies such as diabetic foot.

[0492] Additional neurologic diseases which can be treated or detected with polynucleotides, polypeptides, agonists, and/or antagonists of the present invention include nerve compression syndromes such as carpal tunnel syndrome, tarsal tunnel syndrome, thoracic outlet syndrome such as cervical rib syndrome, ulnar nerve compression syndrome, neuralgia such as causalgia, cervico-brachial neuralgia, facial neuralgia and trigeminal neuralgia, neuritis such as experimental allergic neuritis, optic neuritis, polyneuritis, polyradiculoneuritis and radiculities such as polyradiculitis, hereditary motor and sensory neuropathies such as Charcot-Marie Disease, Hereditary Optic Atrophy, Refsum's Disease, Hereditary Spastic Paraplegia and Werdnig-Hoffmann Disease, Hereditary Sensory and Autonomic Neuropathies which include Congenital Analgesia and Familial Dysautonomia, POEMS Syndrome, Sciatica, Gustatory Sweating and Tetany).

[0493] Immune Activity

[0494] Polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in treating, preventing, diagnosing and/or prognosing diseases, disorders, and/or conditions of the immune system, by, for example, activating or inhibiting the proliferation, differentiation, or mobilization (chemotaxis) of immune cells. Immune cells develop through a process called hematopoiesis, producing myeloid (platelets, red blood cells, neutrophils, and macrophages) and lymphoid (B and T lymphocytes) cells from pluripotent stem cells. The etiology of these immune diseases, disorders, and/or conditions may be genetic, somatic, such as cancer and some autoimmune diseases, acquired (e.g., by chemotherapy or toxins), or infectious. Moreover, polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention can be used as a marker or detector of a particular immune system disease or disorder.

[0495] In another embodiment, a polypeptide of the invention, or polynucleotides, antibodies, agonists, or antagonists corresponding to that polypeptide, may be used to treat diseases and disorders of the immune system and/or to inhibit or enhance an immune response generated by cells associated with the tissue(s) in which the polypeptide of the invention is expressed, including one, two, three, four, five, or more tissues disclosed in Table 1A, column 8 (Tissue Distribution Library Code).

[0496] Polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in treating, preventing, diagnosing, and/or prognosing immunodeficiencies, including both congenital and acquired immunodeficiencies. Examples of B cell immunodeficiencies in which immunoglobulin levels B cell function and/or B cell numbers are decreased include: X-linked agammaglobulinemia (Bruton's disease), X-linked infantile agammaglobulinemia, X-linked immunodeficiency with hyper IgM, non X-linked immunodeficiency with hyper IgM, X-linked lymphoproliferative syndrome (XLP), agammaglobulinemia including congenital and acquired agammaglobulinemia, adult onset agammaglobulinemia, late-onset agammaglobulinemia, dysgammaglobulinemia, hypogammaglobulinemia, unspecified hypogammaglobulinemia, recessive agammaglobulinemia (Swiss type), Selective IgM deficiency, selective IgA deficiency, selective IgG subclass deficiencies, IgG subclass deficiency (with or without IgA deficiency), Ig deficiency with increased IgM, IgG and IgA deficiency with increased IgM, antibody deficiency with normal or elevated Igs, Ig heavy chain deletions, kappa chain deficiency, B cell lymphoproliferative disorder (BLPD), common variable immunodeficiency (CVID), common variable immunodeficiency (CVI) (acquired), and transient hypogammaglobulinemia of infancy.

[0497] In specific embodiments, ataxia-telangiectasia or conditions associated with ataxia-telangiectasia are treated, prevented, diagnosed, and/or prognosing using the polypeptides or polynucleotides of the invention, and/or agonists or antagonists thereof.

[0498] Examples of congenital immunodeficiencies in which T cell and/or B cell function and/or number is decreased include, but are not limited to: DiGeorge anomaly, severe combined immunodeficiencies (SCID) (including, but not limited to, X-linked SCID, autosomal recessive SCID, adenosine deaminase deficiency, purine nucleoside phosphorylase (PNP) deficiency, Class II MHC deficiency (Bare lymphocyte syndrome), Wiskott-Aldrich syndrome, and ataxia telangiectasia), thymic hypoplasia, third and fourth pharyngeal pouch syndrome, 22q11.2 deletion, chronic mucocutaneous candidiasis, natural killer cell deficiency (NK), idiopathic CD4+ T-lymphocytopenia, immunodeficiency with predominant T cell defect (unspecified), and unspecified immunodeficiency of cell mediated immunity.

[0499] In specific embodiments, DiGeorge anomaly or conditions associated with DiGeorge anomaly are treated, prevented, diagnosed, and/or prognosed using polypeptides or polynucleotides of the invention, or antagonists or agonists thereof.

[0500] Other immunodeficiencies that may be treated, prevented, diagnosed, and/or prognosed using polypeptides or polynucleotides of the invention, and/or agonists or antagonists thereof, include, but are not limited to, chronic granulomatous disease, Chédiak-Higashi syndrome, myeloperoxidase deficiency, leukocyte glucose-6-phosphate dehydrogenase deficiency, X-linked lymphoproliferative syndrome (XLP), leukocyte adhesion deficiency, complement component deficiencies (including C1, C2, C3, C4, C5, C6, C7, C8 and/or C9 deficiencies), reticular dysgenesis, thymic alymphoplasia-aplasia, immunodeficiency with thymoma, severe congenital leukopenia, dysplasia with immunodeficiency, neonatal neutropenia, short limbed dwarfism, and Nezelof syndrome-combined immunodeficiency with Igs.

[0501] In a preferred embodiment, the immunodeficiencies and/or conditions associated with the immunodeficiencies recited above are treated, prevented, diagnosed and/or prognosed using polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention.

[0502] In a preferred embodiment polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention could be used as an agent to boost immunoresponsiveness among immunodeficient individuals. In specific embodiments, polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention could be used as an agent to boost immunoresponsiveness among B cell and/or T cell immunodeficient individuals.

[0503] The polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in treating, preventing, diagnosing and/or prognosing autoimmune disorders. Many autoimmune disorders result from inappropriate recognition of self as foreign material by immune cells. This inappropriate recognition results in an immune response leading to the destruction of the host tissue. Therefore, the administration of polynucleotides and polypeptides of the invention that can inhibit an immune response, particularly the proliferation, differentiation, or chemotaxis of T-cells, may be an effective therapy in preventing autoimmune disorders.

[0504] Autoimmune diseases or disorders that may be treated, prevented, diagnosed and/or prognosed by polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention include, but are not limited to, one or more of the following: systemic lupus erythematosus, rheumatoid arthritis, ankylosing spondylitis, multiple sclerosis, autoimmune thyroiditis, Hashimoto's thyroiditis, autoimmune hemolytic anemia, hemolytic anemia, thrombocytopenia, autoimmune thrombocytopenia purpura, autoimmune neonatal thrombocytopenia, idiopathic thrombocytopenia purpura, purpura (e.g., Henloch-Scoenlein purpura), autoimmunocytopenia, Goodpasture's syndrome, Pemphigus vulgaris, myasthenia gravis, Grave's disease (hyperthyroidism), and insulin-resistant diabetes mellitus.

[0505] Additional disorders that are likely to have an autoimmune component that may be treated, prevented, and/or diagnosed with the compositions of the invention include, but are not limited to, type II collagen-induced arthritis, antiphospholipid syndrome, dermatitis, allergic encephalomyelitis, myocarditis, relapsing polychondritis, rheumatic heart disease, neuritis, uveitis ophthalmia, polyendocrinopathies, Reiter's Disease, Stiff-Man Syndrome, autoimmune pulmonary inflammation, autism, Guillain-Barre Syndrome, insulin dependent diabetes mellitus, and autoimmune inflammatory eye disorders.

[0506] Additional disorders that are likely to have an autoimmune component that may be treated, prevented, diagnosed and/or prognosed with the compositions of the invention include, but are not limited to, scleroderma with anti-collagen antibodies (often characterized, e.g., by nucleolar and other nuclear antibodies), mixed connective tissue disease (often characterized, e.g., by antibodies to extractable nuclear antigens (e.g., ribonucleoprotein)), polymyositis (often characterized, e.g., by nonhistone ANA), pernicious anemia (often characterized, e.g., by antiparietal cell, microsomes, and intrinsic factor antibodies), idiopathic Addison's disease (often characterized, e.g., by humoral and cell-mediated adrenal cytotoxicity, infertility (often characterized, e.g., by antispermatozoal antibodies), glomerulonephritis (often characterized, e.g., by glomerular basement membrane antibodies or immune complexes), bullous pemphigoid (often characterized, e.g., by IgG and complement in basement membrane), Sjogren's syndrome (often characterized, e.g., by multiple tissue antibodies, and/or a specific nonhistone ANA (SS-B)), diabetes mellitus (often characterized, e.g., by cell-mediated and humoral islet cell antibodies), and adrenergic drug resistance (including adrenergic drug resistance with asthma or cystic fibrosis) (often characterized, e.g., by beta-adrenergic receptor antibodies).

[0507] Additional disorders that may have an autoimmune component that may be treated, prevented, diagnosed and/or prognosed with the compositions of the invention include, but are not limited to, chronic active hepatitis (often characterized, e.g., by smooth muscle antibodies), primary biliary cirrhosis (often characterized, e.g., by mitochondria antibodies), other endocrine gland failure (often characterized, e.g., by specific tissue antibodies in some cases), vitiligo (often characterized, e.g., by melanocyte antibodies), vasculitis (often characterized, e.g., by Ig and complement in vessel walls and/or low serum complement), post-MI (often characterized, e.g., by myocardial antibodies), cardiotomy syndrome (often characterized, e.g., by myocardial antibodies), urticaria (often characterized, e.g., by IgG and IgM antibodies to IgE), atopic dermatitis (often characterized, e.g., by IgG and IgM antibodies to IgE), asthma (often characterized, e.g., by IgG and IgM antibodies to IgE), and many other inflammatory, granulomatous, degenerative, and atrophic disorders.

[0508] In a preferred embodiment, the autoimmune diseases and disorders and/or conditions associated with the diseases and disorders recited above are treated, prevented, diagnosed and/or prognosed using for example, antagonists or agonists, polypeptides or polynucleotides, or antibodies of the present invention. In a specific preferred embodiment, rheumatoid arthritis is treated, prevented, and/or diagnosed using polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention.

[0509] In another specific preferred embodiment, systemic lupus erythematosus is treated, prevented, and/or diagnosed using polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention. In another specific preferred embodiment, idiopathic thrombocytopenia purpura is treated, prevented, and/or diagnosed using polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention.

[0510] In another specific preferred embodiment IgA nephropathy is treated, prevented, and/or diagnosed using polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention.

[0511] In a preferred embodiment, the autoimmune diseases and disorders and/or conditions associated with the diseases and disorders recited above are treated, prevented, diagnosed and/or prognosed using polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention

[0512] In preferred embodiments, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a immunosuppressive agent(s).

[0513] Polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in treating, preventing, prognosing, and/or diagnosing diseases, disorders, and/or conditions of hematopoietic cells. Polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention could be used to increase differentiation and proliferation of hematopoietic cells, including the pluripotent stem cells, in an effort to treat or prevent those diseases, disorders, and/or conditions associated with a decrease in certain (or many) types hematopoietic cells, including but not limited to, leukopenia, neutropenia, anemia, and thrombocytopenia. Alternatively, Polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention could be used to increase differentiation and proliferation of hematopoietic cells, including the pluripotent stem cells, in an effort to treat or prevent those diseases, disorders, and/or conditions associated with an increase in certain (or many) types of hematopoietic cells, including but not limited to, histiocytosis.

[0514] Allergic reactions and conditions, such as asthma (particularly allergic asthma) or other respiratory problems, may also be treated, prevented, diagnosed and/or prognosed using polypeptides, antibodies, or polynucleotides of the invention, and/or agonists or antagonists thereof. Moreover, these molecules can be used to treat, prevent, prognose, and/or diagnose anaphylaxis, hypersensitivity to an antigenic molecule, or blood group incompatibility.

[0515] Additionally, polypeptides or polynucleotides of the invention, and/or agonists or antagonists thereof, may be used to treat, prevent, diagnose and/or prognose IgE-mediated allergic reactions. Such allergic reactions include, but are not limited to, asthma, rhinitis, and eczema. In specific embodiments, polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be used to modulate IgE concentrations in vitro or in vivo.

[0516] Moreover, polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention have uses in the diagnosis, prognosis, prevention, and/or treatment of inflammatory conditions. For example, since polypeptides, antibodies, or polynucleotides of the invention, and/or agonists or antagonists of the invention may inhibit the activation, proliferation and/or differentiation of cells involved in an inflammatory response, these molecules can be used to prevent and/or treat chronic and acute inflammatory conditions. Such inflammatory conditions include, but are not limited to, for example, inflammation associated with infection (e.g., septic shock, sepsis, or systemic inflammatory response syndrome), ischemia-reperfusion injury, endotoxin lethality, complement-mediated hyperacute rejection, nephritis, cytokine or chemokine induced lung injury, inflammatory bowel disease, Crohn's disease, over production of cytokines (e.g., TNF or IL-1.), respiratory disorders (e.g., asthma and allergy); gastrointestinal disorders (e.g., inflammatory bowel disease); cancers (e.g., gastric, ovarian, lung, bladder, liver, and breast); CNS disorders (e.g., multiple sclerosis; ischemic brain injury and/or stroke, traumatic brain injury, neurodegenerative disorders (e.g., Parkinson's disease and Alzheimer's disease); AIDS-related dementia; and prion disease); cardiovascular disorders (e.g., atherosclerosis, myocarditis, cardiovascular disease, and cardiopulmonary bypass complications); as well as many additional diseases, conditions, and disorders that are characterized by inflammation (e.g., hepatitis, rheumatoid arthritis, gout, trauma, pancreatitis, sarcoidosis, dermatitis, renal ischemia-reperfusion injury, Grave's disease, systemic lupus erythematosus, diabetes mellitus, and allogenic transplant rejection).

[0517] Because inflammation is a fundamental defense mechanism, inflammatory disorders can effect virtually any tissue of the body. Accordingly, polynucleotides, polypeptides, and antibodies of the invention, as well as agonists or antagonists thereof, have uses in the treatment of tissue-specific inflammatory disorders, including, but not limited to, adrenalitis, alveolitis, angiocholecystitis, appendicitis, balanitis, blepharitis, bronchitis, bursitis, carditis, cellulitis, cervicitis, cholecystitis, chorditis, cochlitis, colitis, conjunctivitis, cystitis, dermatitis, diverticulitis, encephalitis, endocarditis, esophagitis, eustachitis, fibrositis, folliculitis, gastritis, gastroenteritis, gingivitis, glossitis, hepatosplenitis, keratitis, labyrinthitis, laryngitis, lymphangitis, mastitis, media otitis, meningitis, metritis, mucitis, myocarditis, myosititis, myringitis, nephritis, neuritis, orchitis, osteochondritis, otitis, pericarditis, peritendonitis, peritonitis, pharyngitis, phlebitis, poliomyelitis, prostatitis, pulpitis, retinitis, rhinitis, salpingitis, scleritis, sclerochoroiditis, scrotitis, sinusitis, spondylitis, steatitis, stomatitis, synovitis, syringitis, tendonitis, tonsillitis, urethritis, and vaginitis.

[0518] In specific embodiments, polypeptides, antibodies, or polynucleotides of the invention, and/or agonists or antagonists thereof, are useful to diagnose, prognose, prevent, and/or treat organ transplant rejections and graft-versus-host disease. Organ rejection occurs by host immune cell destruction of the transplanted tissue through an immune response. Similarly, an immune response is also involved in GVHD, but, in this case, the foreign transplanted immune cells destroy the host tissues. Polypeptides, antibodies, or polynucleotides of the invention, and/or agonists or antagonists thereof, that inhibit an immune response, particularly the activation, proliferation, differentiation, or chemotaxis of T-cells, may be an effective therapy in preventing organ rejection or GVHD. In specific embodiments, polypeptides, antibodies, or polynucleotides of the invention, and/or agonists or antagonists thereof, that inhibit an immune response, particularly the activation, proliferation, differentiation, or chemotaxis of T-cells, may be an effective therapy in preventing experimental allergic and hyperacute xenograft rejection.

[0519] In other embodiments, polypeptides, antibodies, or polynucleotides of the invention, and/or agonists or antagonists thereof, are useful to diagnose, prognose, prevent, and/or treat immune complex diseases, including, but not limited to, serum sickness, post streptococcal glomerulonephritis, polyarteritis nodosa, and immune complex-induced vasculitis.

[0520] Polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the invention can be used to treat, detect, and/or prevent infectious agents. For example, by increasing the immune response, particularly increasing the proliferation activation and/or differentiation of B and/or T cells, infectious diseases may be treated, detected, and/or prevented. The immune response may be increased by either enhancing an existing immune response, or by initiating a new immune response. Alternatively, polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may also directly inhibit the infectious agent (refer to section of application listing infectious agents, etc), without necessarily eliciting an immune response.

[0521] In another embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a vaccine adjuvant that enhances immune responsiveness to an antigen. In a specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an adjuvant to enhance tumor-specific immune responses.

[0522] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an adjuvant to enhance anti-viral immune responses. Anti-viral immune responses that may be enhanced using the compositions of the invention as an adjuvant, include virus and virus associated diseases or symptoms described herein or otherwise known in the art. In specific embodiments, the compositions of the invention are used as an adjuvant to enhance an immune response to a virus, disease, or symptom selected from the group consisting of: AIDS, meningitis, Dengue, EBV, and hepatitis (e.g., hepatitis B). In another specific embodiment, the compositions of the invention are used as an adjuvant to enhance an immune response to a virus, disease, or symptom selected from the group consisting of: HIV/AIDS, respiratory syncytial virus, Dengue, rotavirus, Japanese B encephalitis, influenza A and B, parainfluenza, measles, cytomegalovirus, rabies, Junin, Chikungunya, Rift Valley Fever, herpes simplex, and yellow fever.

[0523] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an adjuvant to enhance anti-bacterial or anti-fungal immune responses. Anti-bacterial or anti-fungal immune responses that may be enhanced using the compositions of the invention as an adjuvant, include bacteria or fungus and bacteria or fungus associated diseases or symptoms described herein or otherwise known in the art. In specific embodiments, the compositions of the invention are used as an adjuvant to enhance an immune response to a bacteria or fungus, disease, or symptom selected from the group consisting of: tetanus, Diphtheria, botulism, and meningitis type B.

[0524] In another specific embodiment, the compositions of the invention are used as an adjuvant to enhance an immune response to a bacteria or fungus, disease, or symptom selected from the group consisting of: Vibrio cholerae, Mycobacterium leprae, Salmonella typhi, Salmonella paratyphi, Meisseria meningitidis, Streptococcus pneumoniae, Group B streptococcus, Shigella spp., Enterotoxigenic Escherichia coli, Enterohemorrhagic E. coli, and Borrelia burgdorferi.

[0525] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an adjuvant to enhance anti-parasitic immune responses. Anti-parasitic immune responses that may be enhanced using the compositions of the invention as an adjuvant, include parasite and parasite associated diseases or symptoms described herein or otherwise known in the art. In specific embodiments, the compositions of the invention are used as an adjuvant to enhance an immune response to a parasite. In another specific embodiment, the compositions of the invention are used as an adjuvant to enhance an immune response to Plasmodium (malaria) or Leishmania.

[0526] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention may also be employed to treat infectious diseases including silicosis, sarcoidosis, and idiopathic pulmonary fibrosis; for example, by preventing the recruitment and activation of mononuclear phagocytes.

[0527] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an antigen for the generation of antibodies to inhibit or enhance immune mediated responses against polypeptides of the invention.

[0528] In one embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are administered to an animal (e.g., mouse, rat, rabbit, hamster, guinea pig, pigs, micro-pig, chicken, camel, goat, horse, cow, sheep, dog, cat, non-human primate, and human, most preferably human) to boost the immune system to produce increased quantities of one or more antibodies (e.g., IgG, IgA, IgM, and IgE), to induce higher affinity antibody production and immunoglobulin class switching (e.g., IgG, IgA, IgM, and IgE), and/or to increase an immune response.

[0529] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a stimulator of B cell responsiveness to pathogens.

[0530] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an activator of T cells.

[0531] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an agent that elevates the immune status of an individual prior to their receipt of immunosuppressive therapies.

[0532] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an agent to induce higher affinity antibodies.

[0533] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an agent to increase serum immunoglobulin concentrations.

[0534] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an agent to accelerate recovery of immunocompromised individuals.

[0535] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an agent to boost immunoresponsiveness among aged populations and/or neonates.

[0536] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an immune system enhancer prior to, during, or after bone marrow transplant and/or other transplants (e.g., allogeneic or xenogeneic organ transplantation). With respect to transplantation, compositions of the invention may be administered prior to, concomitant with, and/or after transplantation. In a specific embodiment, compositions of the invention are administered after transplantation, prior to the beginning of recovery of T-cell populations. In another specific embodiment, compositions of the invention are first administered after transplantation after the beginning of recovery of T cell populations, but prior to full recovery of B cell populations.

[0537] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an agent to boost immunoresponsiveness among individuals having an acquired loss of B cell function. Conditions resulting in an acquired loss of B cell function that may be ameliorated or treated by administering the polypeptides, antibodies, polynucleotides and/or agonists or antagonists thereof, include, but are not limited to, HIV Infection, AIDS, bone marrow transplant, and B cell chronic lymphocytic leukemia (CLL).

[0538] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an agent to boost immunoresponsiveness among individuals having a temporary immune deficiency. Conditions resulting in a temporary immune deficiency that may be ameliorated or treated by administering the polypeptides, antibodies, polynucleotides and/or agonists or antagonists thereof, include, but are not limited to, recovery from viral infections (e.g., influenza), conditions associated with malnutrition, recovery from infectious mononucleosis, or conditions associated with stress, recovery from measles, recovery from blood transfusion, and recovery from surgery.

[0539] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a regulator of antigen presentation by monocytes, dendritic cells, and/or B-cells. In one embodiment, polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention enhance antigen presentation or antagonizes antigen presentation in vitro or in vivo. Moreover, in related embodiments, said enhancement or antagonism of antigen presentation may be useful as an anti-tumor treatment or to modulate the immune system.

[0540] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as an agent to direct an individual's immune system towards development of a humoral response (i.e. TH2) as opposed to a TH1 cellular response.

[0541] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a means to induce tumor proliferation and thus make it more susceptible to anti-neoplastic agents. For example, multiple myeloma is a slowly dividing disease and is thus refractory to virtually all anti-neoplastic regimens. If these cells were forced to proliferate more rapidly their susceptibility profile would likely change.

[0542] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a stimulator of B cell production in pathologies such as AIDS, chronic lymphocyte disorder and/or Common Variable Immunodificiency.

[0543] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a therapy for generation and/or regeneration of lymphoid tissues following surgery, trauma or genetic defect. In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used in the pretreatment of bone marrow samples prior to transplant.

[0544] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a gene-based therapy for genetically inherited disorders resulting in immuno-incompetence/immunodeficiency such as observed among SCID patients.

[0545] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a means of activating monocytes/macrophages to defend against parasitic diseases that effect monocytes such as Leishmania.

[0546] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a means of regulating secreted cytokines that are elicited by polypeptides of the invention.

[0547] In another embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used in one or more of the applications decribed herein, as they may apply to veterinary medicine.

[0548] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a means of blocking various aspects of immune responses to foreign agents or self. Examples of diseases or conditions in which blocking of certain aspects of immune responses may be desired include autoimmune disorders such as lupus, and arthritis, as well as immunoresponsiveness to skin allergies, inflammation, bowel disease, injury and diseases/disorders associated with pathogens.

[0549] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a therapy for preventing the B cell proliferation and Ig secretion associated with autoimmune diseases such as idiopathic thrombocytopenic purpura, systemic lupus erythematosus and multiple sclerosis.

[0550] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a inhibitor of B and/or T cell migration in endothelial cells. This activity disrupts tissue architecture or cognate responses and is useful, for example in disrupting immune responses, and blocking sepsis.

[0551] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a therapy for chronic hypergammaglobulinemia evident in such diseases as monoclonal gammopathy of undetermined significance (MGUS), Waldenstrom's disease, related idiopathic monoclonal gammopathies, and plasmacytomas.

[0552] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention may be employed for instance to inhibit polypeptide chemotaxis and activation of macrophages and their precursors, and of neutrophils, basophils, B lymphocytes and some T-cell subsets, e.g., activated and CD8 cytotoxic T cells and natural killer cells, in certain autoimmune and chronic inflammatory and infective diseases. Examples of autoimmune diseases are described herein and include multiple sclerosis, and insulin-dependent diabetes.

[0553] The polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention may also be employed to treat idiopathic hyper-eosinophilic syndrome by, for example, preventing eosinophil production and migration.

[0554] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used to enhance or inhibit complement mediated cell lysis.

[0555] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used to enhance or inhibit antibody dependent cellular cytotoxicity.

[0556] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention may also be employed for treating atherosclerosis, for example, by preventing monocyte infiltration in the artery wall.

[0557] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention may be employed to treat adult respiratory distress syndrome (ARDS).

[0558] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention may be useful for stimulating wound and tissue repair, stimulating angiogenesis, and/or stimulating the repair of vascular or lymphatic diseases or disorders. Additionally, agonists and antagonists of the invention may be used to stimulate the regeneration of mucosal surfaces.

[0559] In a specific embodiment, polynucleotides or polypeptides, and/or agonists thereof are used to diagnose, prognose, treat, and/or prevent a disorder characterized by primary or acquired immunodeficiency, deficient serum immunoglobulin production, recurrent infections, and/or immune system dysfunction. Moreover, polynucleotides or polypeptides, and/or agonists thereof may be used to treat or prevent infections of the joints, bones, skin, and/or parotid glands, blood-borne infections (e.g., sepsis, meningitis, septic arthritis, and/or osteomyelitis), autoimmune diseases (e.g., those disclosed herein), inflammatory disorders, and malignancies, and/or any disease or disorder or condition associated with these infections, diseases, disorders and/or malignancies) including, but not limited to, CVID, other primary immune deficiencies, HIV disease, CLL, recurrent bronchitis, sinusitis, otitis media, conjunctivitis, pneumonia, hepatitis, meningitis, herpes zoster (e.g., severe herpes zoster), and/or pneumocystis carnii. Other diseases and disorders that may be prevented, diagnosed, prognosed, and/or treated with polynucleotides or polypeptides, and/or agonists of the present invention include, but are not limited to, HIV infection, HTLV-BLV infection, lymphopenia, phagocyte bactericidal dysfunction anemia, thrombocytopenia, and hemoglobinuria.

[0560] In another embodiment, polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention are used to treat, and/or diagnose an individual having common variable immunodeficiency disease (“CVID”; also known as “acquired agammaglobulinemia” and “acquired hypogammaglobulinemia”) or a subset of this disease.

[0561] In a specific embodiment, polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be used to diagnose, prognose, prevent, and/or treat cancers or neoplasms including immune cell or immune tissue-related cancers or neoplasms. Examples of cancers or neoplasms that may be prevented, diagnosed, or treated by polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention include, but are not limited to, acute myelogenous leukemia, chronic myelogenous leukemia, Hodgkin's disease, non-Hodgkin's lymphoma, acute lymphocytic anemia (ALL) Chronic lymphocyte leukemia, plasmacytomas, multiple myeloma, Burkitt's lymphoma, EBV-transformed diseases, and/or diseases and disorders described in the section entitled “Hyperproliferative Disorders” elsewhere herein.

[0562] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a therapy for decreasing cellular proliferation of Large B-cell Lymphomas.

[0563] In another specific embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are used as a means of decreasing the involvement of B cells and Ig associated with Chronic Myelogenous Leukemia.

[0564] In specific embodiments, the compositions of the invention are used as an agent to boost immunoresponsiveness among B cell immunodeficient individuals, such as, for example, an individual who has undergone a partial or complete splenectomy.

[0565] Antagonists of the invention include, for example, binding and/or inhibitory antibodies, antisense nucleic acids, ribozymes or soluble forms of the polypeptides of the present invention (e.g., Fc fusion protein; see, e.g., Example 9). Agonists of the invention include, for example, binding or stimulatory antibodies, and soluble forms of the polypeptides (e.g., Fc fusion proteins; see, e.g., Example 9). polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention may be employed in a composition with a pharmaceutically acceptable carrier, e.g., as described herein.

[0566] In another embodiment, polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention are administered to an animal (including, but not limited to, those listed above, and also including transgenic animals) incapable of producing functional endogenous antibody molecules or having an otherwise compromised endogenous immune system, but which is capable of producing human immunoglobulin molecules by means of a reconstituted or partially reconstituted immune system from another animal (see, e.g., published PCT Application Nos. WO98/24893, WO/9634096, WO/9633735, and WO/9110741). Administration of polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention to such animals is useful for the generation of monoclonal antibodies against the polypeptides, antibodies, polynucleotides and/or agonists or antagonists of the present invention.

[0567] Blood-Related Disorders

[0568] The polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be used to modulate hemostatic (the stopping of bleeding) or thrombolytic (clot dissolving) activity. For example, by increasing hemostatic or thrombolytic activity, polynucleotides or polypeptides, and/or agonists or antagonists of the present invention could be used to treat or prevent blood coagulation diseases, disorders, and/or conditions (e.g., afibrinogenemia, factor deficiencies, hemophilia), blood platelet diseases, disorders, and/or conditions (e.g., thrombocytopenia), or wounds resulting from trauma, surgery, or other causes. Alternatively, polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention that can decrease hemostatic or thrombolytic activity could be used to inhibit or dissolve clotting. These molecules could be important in the treatment or prevention of heart attacks (infarction), strokes, or scarring.

[0569] In specific embodiments, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be used to prevent, diagnose, prognose, and/or treat thrombosis, arterial thrombosis, venous thrombosis, thromboembolism, pulmonary embolism, atherosclerosis, myocardial infarction, transient ischemic attack, unstable angina. In specific embodiments, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be used for the prevention of occulsion of saphenous grafts, for reducing the risk of periprocedural thrombosis as might accompany angioplasty procedures, for reducing the risk of stroke in patients with atrial fibrillation including nonrheumatic atrial fibrillation, for reducing the risk of embolism associated with mechanical heart valves and or mitral valves disease. Other uses for the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention, include, but are not limited to, the prevention of occlusions in extrcorporeal devices (e.g., intravascular canulas, vascular access shunts in hemodialysis patients, hemodialysis machines, and cardiopulmonary bypass machines).

[0570] In another embodiment, a polypeptide of the invention, or polynucleotides, antibodies, agonists, or antagonists corresponding to that polypeptide, may be used to prevent, diagnose, prognose, and/or treat diseases and disorders of the blood and/or blood forming organs associated with the tissue(s) in which the polypeptide of the invention is expressed, including one, two, three, four, five, or more tissues disclosed in Table 1A, column 8 (Tissue Distribution Library Code).

[0571] The polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be used to modulate hematopoietic activity (the formation of blood cells). For example, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be used to increase the quantity of all or subsets of blood cells, such as, for example, erythrocytes, lymphocytes (B or T cells), myeloid cells (e.g., basophils, eosinophils, neutrophils, mast cells, macrophages) and platelets. The ability to decrease the quantity of blood cells or subsets of blood cells may be useful in the prevention, detection, diagnosis and/or treatment of anemias and leukopenias described below. Alternatively, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be used to decrease the quantity of all or subsets of blood cells, such as, for example, erythrocytes, lymphocytes (B or T cells), myeloid cells (e.g., basophils, eosinophils, neutrophils, mast cells, macrophages) and platelets. The ability to decrease the quantity of blood cells or subsets of blood cells may be useful in the prevention, detection, diagnosis and/or treatment of leukocytoses, such as, for example eosinophilia.

[0572] The polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be used to prevent, treat, or diagnose blood dyscrasia.

[0573] Anemias are conditions in which the number of red blood cells or amount of hemoglobin (the protein that carries oxygen) in them is below normal. Anemia may be caused by excessive bleeding, decreased red blood cell production, or increased red blood cell destruction (hemolysis). The polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in treating, preventing, and/or diagnosing anemias. Anemias that may be treated prevented or diagnosed by the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention include iron deficiency anemia, hypochromic anemia, microcytic anemia, chlorosis, hereditary siderob;astic anemia, idiopathic acquired sideroblastic anemia, red cell aplasia, megaloblastic anemia (e.g., pernicious anemia, (vitamin B12 deficiency) and folic acid deficiency anemia), aplastic anemia, hemolytic anemias (e.g., autoimmune helolytic anemia, microangiopathic hemolytic anemia, and paroxysmal nocturnal hemoglobinuria). The polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in treating, preventing, and/or diagnosing anemias associated with diseases including but not limited to, anemias associated with systemic lupus erythematosus, cancers, lymphomas, chronic renal disease, and enlarged spleens. The polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in treating, preventing, and/or diagnosing anemias arising from drug treatments such as anemias associated with methyldopa, dapsone, and/or sulfadrugs. Additionally, rhe polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in treating, preventing, and/or diagnosing anemias associated with abnormal red blood cell architecture including but not limited to, hereditary spherocytosis, hereditary elliptocytosis, glucose-6-phosphate dehydrogenase deficiency, and sickle cell anemia.

[0574] The polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in treating, preventing, and/or diagnosing hemoglobin abnormalities, (e.g., those associated with sickle cell anemia, hemoglobin C disease, hemoglobin S-C disease, and hemoglobin E disease). Additionally, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in diagnosing, prognosing, preventing, and/or treating thalassemias, including, but not limited to major and minor forms of alpha-thalassemia and beta-thalassemia.

[0575] In another embodiment, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in diagnosing, prognosing, preventing, and/or treating bleeding disorders including, but not limited to, thrombocytopenia (e.g., idiopathic thrombocytopenic purpura, and thrombotic thrombocytopenic purpura), Von Willebrand's disease, hereditary platelet disorders (e.g., storage pool disease such as Chediak-Higashi and Hermansky-Pudlak syndromes, thromboxane A2 dysfunction, thromboasthenia, and Bemard-Soulier syndrome), hemolytic-uremic syndrome, hemophelias such as hemophelia A or Factor VII deficiency and Christmas disease or Factor IX deficiency, Hereditary Hemorhhagic Telangiectsia, also known as Rendu-Osler-Weber syndrome, allergic purpura (Henoch Schonlein purpura) and disseminated intravascular coagulation.

[0576] The effect of the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention on the clotting time of blood may be monitored using any of the clotting tests known in the art including, but not limited to, whole blood partial thromboplastin time (PTT), the activated partial thromboplastin time (aPTT), the activated clotting time (ACT), the recalcified activated clotting time, or the Lee-White Clotting time.

[0577] Several diseases and a variety of drugs can cause platelet dysfunction. Thus, in a specific embodiment, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in diagnosing, prognosing, preventing, and/or treating acquired platelet dysfunction such as platelet dysfunction accompanying kidney failure, leukemia, multiple myeloma, cirrhosis of the liver, and systemic lupus erythematosus as well as platelet dysfunction associated with drug treatments, including treatment with aspirin, ticlopidine, nonsteroidal anti-inflammatory drugs (used for arthritis, pain, and sprains), and penicillin in high doses.

[0578] In another embodiment, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in diagnosing, prognosing, preventing, and/or treating diseases and disorders characterized by or associated with increased or decreased numbers of white blood cells. Leukopenia occurs when the number of white blood cells decreases below normal. Leukopenias include, but are not limited to, neutropenia and lymphocytopenia. An increase in the number of white blood cells compared to normal is known as leukocytosis. The body generates increased numbers of white blood cells during infection. Thus, leukocytosis may simply be a normal physiological parameter that reflects infection. Alternatively, leukocytosis may be an indicator of injury or other disease such as cancer. Leokocytoses, include but are not limited to, eosinophilia, and accumulations of macrophages. In specific embodiments, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in diagnosing, prognosing, preventing, and/or treating leukopenia. In other specific embodiments, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in diagnosing, prognosing, preventing, and/or treating leukocytosis.

[0579] Leukopenia may be a generalized decreased in all types of white blood cells, or may be a specific depletion of particular types of white blood cells. Thus, in specific embodiments, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in diagnosing, prognosing, preventing, and/or treating decreases in neutrophil numbers, known as neutropenia. Neutropenias that may be diagnosed, prognosed, prevented, and/or treated by the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention include, but are not limited to, infantile genetic agranulocytosis, familial neutropenia, cyclic neutropenia, neutropenias resulting from or associated with dietary deficiencies (e.g., vitamin B 12 deficiency or folic acid deficiency), neutropenias resulting from or associated with drug treatments (e.g., antibiotic regimens such as penicillin treatment, sulfonamide treatment, anticoagulant treatment, anticonvulsant drugs, anti-thyroid drugs, and cancer chemotherapy), and neutropenias resulting from increased neutrophil destruction that may occur in association with some bacterial or viral infections, allergic disorders, autoimmune diseases, conditions in which an individual has an enlarged spleen (e.g., Felty syndrome, malaria and sarcoidosis), and some drug treatment regimens.

[0580] The polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in diagnosing, prognosing, preventing, and/or treating lymphocytopenias (decreased numbers of B and/or T lymphocytes), including, but not limited lymphocytopenias resulting from or associated with stress, drug treatments (e.g., drug treatment with corticosteroids, cancer chemotherapies, and/or radiation therapies), AIDS infection and/or other diseases such as, for example, cancer, rheumatoid arthritis, systemic lupus erythematosus, chronic infections, some viral infections and/or hereditary disorders (e.g., DiGeorge syndrome, Wiskott-Aldrich Syndome, severe combined immunodeficiency, ataxia telangiectsia).

[0581] The polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in diagnosing, prognosing, preventing, and/or treating diseases and disorders associated with macrophage numbers and/or macrophage function including, but not limited to, Gaucher's disease, Niemann-Pick disease, Letterer-Siwe disease and Hand-Schuller-Christian disease.

[0582] In another embodiment, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in diagnosing, prognosing, preventing, and/or treating diseases and disorders associated with eosinophil numbers and/or eosinophil function including, but not limited to, idiopathic hypereosinophilic syndrome, eosinophilia-myalgia syndrome, and Hand-Schuller-Christian disease.

[0583] In yet another embodiment, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in diagnosing, prognosing, preventing, and/or treating leukemias and lymphomas including, but not limited to, acute lymphocytic (lymphpblastic) leukemia (ALL), acute myeloid (myelocytic, myelogenous, myeloblastic, or myelomonocytic) leukemia, chronic lymphocytic leukemia (e.g., B cell leukemias, T cell leukemias, Sezary syndrome, and Hairy cell leukenia), chronic myelocytic (myeloid, myelogenous, or granulocytic) leukemia, Hodgkin's lymphoma, non-hodgkin's lymphoma, Burkitt's lymphoma, and mycosis fungoides.

[0584] In other embodiments, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in diagnosing, prognosing, preventing, and/or treating diseases and disorders of plasma cells including, but not limited to, plasma cell dyscrasias, monoclonal gammaopathies, monoclonal gammopathies of undetermined significance, multiple myeloma, macroglobulinemia, Waldenstrom's macroglobulinemia, cryoglobulinemia, and Raynaud's phenomenon.

[0585] In other embodiments, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in treating, preventing, and/or diagnosing myeloproliferative disorders, including but not limited to, polycythemia vera, relative polycythemia, secondary polycythemia, myelofibrosis, acute myelofibrosis, agnogenic myelod metaplasia, thrombocythemia, (including both primary and seconday thrombocythemia) and chronic myelocytic leukemia.

[0586] In other embodiments, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful as a treatment prior to surgery, to increase blood cell production.

[0587] In other embodiments, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful as an agent to enhance the migration, phagocytosis, superoxide production, antibody dependent cellular cytotoxicity of neutrophils, eosionophils and macrophages.

[0588] In other embodiments, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful as an agent to increase the number of stem cells in circulation prior to stem cells pheresis. In another specific embodiment, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful as an agent to increase the number of stem cells in circulation prior to platelet pheresis.

[0589] In other embodiments, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful as an agent to increase cytokine production.

[0590] In other embodiments, the polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention may be useful in preventing, diagnosing, and/or treating primary hematopoietic disorders.

[0591] Hyperproliferative Disorders

[0592] In certain embodiments, polynucleotides or polypeptides, or agonists or antagonists of the present invention can be used to treat or detect hyperproliferative disorders, including neoplasms. Polynucleotides or polypeptides, or agonists or antagonists of the present invention may inhibit the proliferation of the disorder through direct or indirect interactions. Alternatively, Polynucleotides or polypeptides, or agonists or antagonists of the present invention may proliferate other cells which can inhibit the hyperproliferative disorder.

[0593] For example, by increasing an immune response, particularly increasing antigenic qualities of the hyperproliferative disorder or by proliferating, differentiating, or mobilizing T-cells, hyperproliferative disorders can be treated. This immune response may be increased by either enhancing an existing immune response, or by initiating a new immune response. Alternatively, decreasing an immune response may also be a method of treating hyperproliferative disorders, such as a chemotherapeutic agent.

[0594] Examples of hyperproliferative disorders that can be treated or detected by polynucleotides or polypeptides, or agonists or antagonists of the present invention include, but are not limited to neoplasms located in the: colon, abdomen, bone, breast, digestive system, liver, pancreas, peritoneum, endocrine glands (adrenal, parathyroid, pituitary, testicles, ovary, thymus, thyroid), eye, head and neck, nervous (central and peripheral), lymphatic system, pelvis, skin, soft tissue, spleen, thorax, and urogenital tract.

[0595] Similarly, other hyperproliferative disorders can also be treated or detected by polynucleotides or polypeptides, or agonists or antagonists of the present invention. Examples of such hyperproliferative disorders include, but are not limited to: Acute Childhood Lymphoblastic Leukemia, Acute Lymphoblastic Leukemia, Acute Lymphocytic Leukemia, Acute Myeloid Leukemia, Adrenocortical Carcinoma, Adult (Primary) Hepatocellular Cancer, Adult (Primary) Liver Cancer, Adult Acute Lymphocytic Leukemia, Adult Acute Myeloid Leukemia, Adult Hodgkin's Disease, Adult Hodgkin's Lymphoma, Adult Lymphocytic Leukemia, Adult Non-Hodgkin's Lymphoma, Adult Primary Liver Cancer, Adult Soft Tissue Sarcoma, AIDS-Related Lymphoma, AIDS-Related Malignancies, Anal Cancer, Astrocytoma, Bile Duct Cancer, Bladder Cancer, Bone Cancer, Brain Stem Glioma, Brain Tumors, Breast Cancer, Cancer of the Renal Pelvis and Ureter, Central Nervous System (Primary) Lymphoma, Central Nervous System Lymphoma, Cerebellar Astrocytoma, Cerebral Astrocytoma, Cervical Cancer, Childhood (Primary) Hepatocellular Cancer, Childhood (Primary) Liver Cancer, Childhood Acute Lymphoblastic Leukemia, Childhood Acute Myeloid Leukemia, Childhood Brain Stem Glioma, Childhood Cerebellar Astrocytoma, Childhood Cerebral Astrocytoma, Childhood Extracranial Germ Cell Tumors, Childhood Hodgkin's Disease, Childhood Hodgkin's Lymphoma, Childhood Hypothalamic and Visual Pathway Glioma, Childhood Lymphoblastic Leukemia, Childhood Medulloblastoma, Childhood Non-Hodgkin's Lymphoma, Childhood Pineal and Supratentorial Primitive Neuroectodermal Tumors, Childhood Primary Liver Cancer, Childhood Rhabdomyosarcoma, Childhood Soft Tissue Sarcoma, Childhood Visual Pathway and Hypothalamic Glioma, Chronic Lymphocytic Leukemia, Chronic Myelogenous Leukemia, Colon Cancer, Cutaneous T-Cell Lymphoma, Endocrine Pancreas Islet Cell Carcinoma, Endometrial Cancer, Ependymoma, Epithelial Cancer, Esophageal Cancer, Ewing's Sarcoma and Related Tumors, Exocrine Pancreatic Cancer, Extracranial Germ Cell Tumor, Extragonadal Germ Cell Tumor, Extrahepatic Bile Duct Cancer, Eye Cancer, Female Breast Cancer, Gaucher's Disease, Gallbladder Cancer, Gastric Cancer, Gastrointestinal Carcinoid Tumor, Gastrointestinal Tumors, Germ Cell Tumors, Gestational Trophoblastic Tumor, Hairy Cell Leukemia, Head and Neck Cancer, Hepatocellular Cancer, Hodgkin's Disease, Hodgkin's Lymphoma, Hypergammaglobulinemia, Hypopharyngeal Cancer, Intestinal Cancers, Intraocular Melanoma, Islet Cell Carcinoma, Islet Cell Pancreatic Cancer, Kaposi's Sarcoma, Kidney Cancer, Laryngeal Cancer, Lip and Oral Cavity Cancer, Liver Cancer, Lung Cancer, Lymphoproliferative Disorders, Macroglobulinemia, Male Breast Cancer, Malignant Mesothelioma, Malignant Thymoma, Medulloblastoma, Melanoma, Mesothelioma, Metastatic Occult Primary Squamous Neck Cancer, Metastatic Primary Squamous Neck Cancer, Metastatic Squamous Neck Cancer, Multiple Myeloma, Multiple Myeloma/Plasma Cell Neoplasm, Myelodysplastic Syndrome, Myelogenous Leukemia, Myeloid Leukemia, Mycloproliferative Disorders, Nasal Cavity and Paranasal Sinus Cancer, Nasopharyngeal Cancer, Neuroblastoma, Non-Hodgkin's Lymphoma During Pregnancy, Nonmelanoma Skin Cancer, Non-Small Cell Lung Cancer, Occult Primary Metastatic Squamous Neck Cancer, Oropharyngeal Cancer, Osteo-/Malignant Fibrous Sarcoma, Osteosarcoma/Malignant Fibrous Histiocytoma, Osteosarcoma/Malignant Fibrous Histiocytoma of Bone, Ovarian Epithelial Cancer, Ovarian Germ Cell Tumor, Ovarian Low Malignant Potential Tumor, Pancreatic Cancer, Paraproteinemias, Purpura, Parathyroid Cancer, Penile Cancer, Pheochromocytoma, Pituitary Tumor, Plasma Cell Neoplasmi/Multiple Myeloma, Primary Central Nervous System Lymphoma, Primary Liver Cancer, Prostate Cancer, Rectal Cancer, Renal Cell Cancer, Renal Pelvis and Ureter Cancer, Retinoblastoma, Rhabdomyosarcoma, Salivary Gland Cancer, Sarcoidosis Sarcomas, Sezary Syndrome, Skin Cancer, Small Cell Lung Cancer, Small Intestine Cancer, Soft Tissue Sarcoma, Squamous Neck Cancer, Stomach Cancer, Supratentorial Primitive Neuroectodernal and Pineal Tumors, T-Cell Lymphoma, Testicular Cancer, Thymoma, Thyroid Cancer, Transitional Cell Cancer of the Renal Pelvis and Ureter, Transitional Renal Pelvis and Ureter Cancer, Trophoblastic Tumors, Ureter and Renal Pelvis Cell Cancer, Urethral Cancer, Uterine Cancer, Uterine Sarcoma, Vaginal Cancer, Visual Pathway and Hypothalamic Glioma, Vulvar Cancer, Waldenstrom's Macroglobulinemia, Wilms' Tumor, and any other hyperproliferative disease, besides neoplasia, located in an organ system listed above.

[0596] In another preferred embodiment, polynucleotides or polypeptides, or agonists or antagonists of the present invention are used to diagnose, prognose, prevent, and/or treat premalignant conditions and to prevent progression to a neoplastic or malignant state, including but not limited to those disorders described above. Such uses are indicated in conditions known or suspected of preceding progression to neoplasia or cancer, in particular, where non-neoplastic cell growth consisting of hyperplasia, metaplasia, or most particularly, dysplasia has occurred (for review of such abnormal growth conditions, see Robbins and Angell, 1976, Basic Pathology, 2d Ed., W. B. Saunders Co., Philadelphia, pp. 68-79.)

[0597] Hyperplasia is a form of controlled cell proliferation, involving an increase in cell number in a tissue or organ, without significant alteration in structure or function. Hyperplastic disorders which can be diagnosed, prognosed, prevented, and/or treated with compositions of the invention (including polynucleotides, polypeptides, agonists or antagonists) include, but are not limited to, angiofollicular mediastinal lymph node hyperplasia, angiolymphoid hyperplasia with eosinophilia, a typical melanocytic hyperplasia, basal cell hyperplasia, benign giant lymph node hyperplasia, cementum hyperplasia, congenital adrenal hyperplasia, congenital sebaceous hyperplasia, cystic hyperplasia, cystic hyperplasia of the breast, denture hyperplasia, ductal hyperplasia, endometrial hyperplasia, fibromuscular hyperplasia, focal epithelial hyperplasia, gingival hyperplasia, inflammatory fibrous hyperplasia, inflammatory papillary hyperplasia, intravascular papillary endothelial hyperplasia, nodular hyperplasia of prostate, nodular regenerative hyperplasia, pseudoepitheliomatous hyperplasia, senile sebaceous hyperplasia, and verrucous hyperplasia.

[0598] Metaplasia is a form of controlled cell growth in which one type of adult or fully differentiated cell substitutes for another type of adult cell. Metaplastic disorders which can be diagnosed, prognosed, prevented, and/or treated with compositions of the invention (including polynucleotides, polypeptides, agonists or antagonists) include, but are not limited to, agnogenic myeloid metaplasia, apocrine metaplasia, a typical metaplasia, autoparenchymatous metaplasia, connective tissue metaplasia, epithelial metaplasia, intestinal metaplasia, metaplastic anemia, metaplastic ossification, metaplastic polyps, myeloid metaplasia, primary myeloid metaplasia, secondary myeloid metaplasia, squamous metaplasia, squamous metaplasia of amnion, and symptomatic myeloid metaplasia.

[0599] Dysplasia is frequently a forerunner of cancer, and is found mainly in the epithelia; it is the most disorderly form of non-neoplastic cell growth, involving a loss in individual cell uniformity and in the architectural orientation of cells. Dysplastic cells often have abnormally large, deeply stained nuclei, and exhibit pleomorphism. Dysplasia characteristically occurs where there exists chronic irritation or inflammation. Dysplastic disorders which can be diagnosed, prognosed, prevented, and/or treated with compositions of the invention (including polynucleotides, polypeptides, agonists or antagonists) include, but are not limited to, anhidrotic ectodermal dysplasia, anterofacial dysplasia, asphyxiating thoracic dysplasia, atriodigital dysplasia, bronchopulmonary dysplasia, cerebral dysplasia, cervical dysplasia, chondroectodermal dysplasia, cleidocranial dysplasia, congenital ectodermal dysplasia, craniodiaphysial dysplasia, craniocarpotarsal dysplasia, craniometaphysial dysplasia, dentin dysplasia, diaphysial dysplasia, ectodermal dysplasia, enamel dysplasia, encephalo-ophthalmic dysplasia, dysplasia epiphysialis hemimelia, dysplasia epiphysialis multiplex, dysplasia epiphysialis punctata, epithelial dysplasia, faciodigitogenital dysplasia, familial fibrous dysplasia of jaws, familial white folded dysplasia, fibromuscular dysplasia, fibrous dysplasia of bone, florid osseous dysplasia, hereditary renal-retinal dysplasia, hidrotic ectodermal dysplasia, hypohidrotic ectodermal dysplasia, lymphopenic thymic dysplasia, mammary dysplasia, mandibulofacial dysplasia, metaphysial dysplasia, Mondini dysplasia, monostotic fibrous dysplasia, mucoepithelial dysplasia, multiple epiphysial dysplasia, oculoauriculovertebral dysplasia, oculodentodigital dysplasia, oculovertebral dysplasia, odontogenic dysplasia, ophthalmomandibulomelic dysplasia, periapical cemental dysplasia, polyostotic fibrous dysplasia, pseudoachondroplastic spondyloepiphysial dysplasia, retinal dysplasia, septo-optic dysplasia, spondyloepiphysial dysplasia, and ventriculoradial dysplasia.

[0600] Additional pre-neoplastic disorders which can be diagnosed, prognosed, prevented, and/or treated with compositions of the invention (including polynucleotides, polypeptides, agonists or antagonists) include, but are not limited to, benign dysproliferative disorders (e.g., benign tumors, fibrocystic conditions, tissue hypertrophy, intestinal polyps, colon polyps, and esophageal dysplasia), leukoplakia, keratoses, Bowen's disease, Farmer's Skin, solar cheilitis, and solar keratosis.

[0601] In another embodiment, a polypeptide of the invention, or polynucleotides, antibodies, agonists, or antagonists corresponding to that polypeptide, may be used to diagnose and/or prognose disorders associated with the tissue(s) in which the polypeptide of the invention is expressed, including one, two, three, four, five, or more tissues disclosed in Table 1A, column 8 (Tissue Distribution Library Code).

[0602] In another embodiment, polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention conjugated to a toxin or a radioactive isotope, as described herein, may be used to treat cancers and neoplasms, including, but not limited to those described herein. In a further preferred embodiment, polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention conjugated to a toxin or a radioactive isotope, as described herein, may be used to treat acute myelogenous leukemia.

[0603] Additionally, polynucleotides, polypeptides, and/or agonists or antagonists of the invention may affect apoptosis, and therefore, would be useful in treating a number of diseases associated with increased cell survival or the inhibition of apoptosis. For example, diseases associated with increased cell survival or the inhibition of apoptosis that could be diagnosed, prognosed, prevented, and/or treated by polynucleotides, polypeptides, and/or agonists or antagonists of the invention, include cancers (such as follicular lymphomas, carcinomas with p53 mutations, and hormone-dependent tumors, including, but not limited to colon cancer, cardiac tumors, pancreatic cancer, melanoma, retinoblastoma, glioblastoma, lung cancer, intestinal cancer, testicular cancer, stomach cancer, neuroblastoma, myxoma, myoma, lymphoma, endothelioma, osteoblastoma, osteoclastoma, osteosarcoma, chondrosarcoma, adenoma, breast cancer, prostate cancer, Kaposi's sarcoma and ovarian cancer); autoimmune disorders such as, multiple sclerosis, Sjogren's syndrome, Hashimoto's thyroiditis, biliary cirrhosis, Behcet's disease, Crohn's disease, polymyositis, systemic lupus erythematosus and immune-related glomerulonephritis and rheumatoid arthritis) and viral infections (such as herpes viruses, pox viruses and adenoviruses), inflammation, graft v. host disease, acute graft rejection, and chronic graft rejection.

[0604] In preferred embodiments, polynucleotides, polypeptides, and/or agonists or antagonists of the invention are used to inhibit growth, progression, and/or metastasis of cancers, in particular those listed above.

[0605] Additional diseases or conditions associated with increased cell survival that could be diagnosed, prognosed, prevented, and/or treated by polynucleotides, polypeptides, and/or agonists or antagonists of the invention, include, but are not limited to, progression, and/or metastases of malignancies and related disorders such as leukemia (including acute leukemias (e.g., acute lymphocytic leukemia, acute myelocytic leukemia (including myeloblastic, promyelocytic, myelomonocytic, monocytic, and erythroleukemia)) and chronic leukemias (e.g., chronic myelocytic (granulocytic) leukemia and chronic lymphocytic leukemia)), polycythemia vera, lymphomas (e.g., Hodgkin's disease and non-Hodgkin's disease), multiple myeloma, Waldenstrom's macroglobulinemia, heavy chain disease, and solid tumors including, but not limited to, sarcomas and carcinomas such as fibrosarcoma, myxosarcoma, liposarcoma, chondrosarcoma, osteogenic sarcoma, chordoma, angiosarcoma, endotheliosarcoma, lymphangiosarcoma, lymphangioendotheliosarcoma, synovioma, mesothelioma, Ewing's tumor, leiomyosarcoma, rhabdomyosarcoma, colon carcinoma, pancreatic cancer, breast cancer, ovarian cancer, prostate cancer, squamous cell carcinoma, basal cell carcinoma, adenocarcinoma, sweat gland carcinoma, sebaceous gland carcinoma, papillary carcinoma, papillary adenocarcinomas, cystadenocarcinoma, medullary carcinoma, bronchogenic carcinoma, renal cell carcinoma, hepatoma, bile duct carcinoma, choriocarcinoma, seminoma, embryonal carcinoma, Wilm's tumor, cervical cancer, testicular tumor, lung carcinoma, small cell lung carcinoma, bladder carcinoma, epithelial carcinoma, glioma, astrocytoma, medulloblastoma, craniopharyngioma, ependymoma, pinealoma, emangioblastoma, acoustic neuroma, oligodendroglioma, menangioma, melanoma, neuroblastoma, and retinoblastoma.

[0606] Diseases associated with increased apoptosis that could be diagnosed, prognosed, prevented, and/or treated by polynucleotides, polypeptides, and/or agonists or antagonists of the invention, include AIDS; neurodegenerative disorders (such as Alzheimer's disease, Parkinson's disease, amyotrophic lateral sclerosis, retinitis pigmentosa, cerebellar degeneration and brain tumor or prior associated disease); autoimmune disorders (such as, multiple sclerosis, Sjogren's syndrome, Hashimoto's thyroiditis, biliary cirrhosis, Behcet's disease, Crohn's disease, polymyositis, systemic lupus erythematosus and immune-related glomerulonephritis and rheumatoid arthritis) myelodysplastic syndromes (such as aplastic anemia), graft v. host disease, ischemic injury (such as that caused by myocardial infarction, stroke and reperfusion injury), liver injury (e.g., hepatitis related liver injury, ischemia/reperfusion injury, cholestosis (bile duct injury) and liver cancer); toxin-induced liver disease (such as that caused by alcohol), septic shock, cachexia and anorexia.

[0607] Hyperproliferative diseases and/or disorders that could be diagnosed, prognosed, prevented, and/or treated by polynucleotides, polypeptides, and/or agonists or antagonists of the invention, include, but are not limited to, neoplasms located in the liver, abdomen, bone, breast, digestive system, pancreas, peritoneum, endocrine glands (adrenal, parathyroid, pituitary, testicles, ovary, thymus, thyroid), eye, head and neck, nervous system (central and peripheral), lymphatic system, pelvis, skin, soft tissue, spleen, thorax, and urogenital tract.

[0608] Similarly, other hyperproliferative disorders can also be diagnosed, prognosed, prevented, and/or treated by polynucleotides, polypeptides, and/or agonists or antagonists of the invention. Examples of such hyperproliferative disorders include, but are not limited to: hypergammaglobulinemia, lymphoproliferative disorders, paraproteinemias, purpura, sarcoidosis, Sezary Syndrome, Waldenstron's macroglobulinemia, Gaucher's Disease, histiocytosis, and any other hyperproliferative disease, besides neoplasia, located in an organ system listed above.

[0609] Another preferred embodiment utilizes polynucleotides of the present invention to inhibit aberrant cellular division, by gene therapy using the present invention, and/or protein fusions or fragments thereof.

[0610] Thus, the present invention provides a method for treating cell proliferative disorders by inserting into an abnormally proliferating cell a polynucleotide of the present invention, wherein said polynucleotide represses said expression.

[0611] Another embodiment of the present invention provides a method of treating cell-proliferative disorders in individuals comprising administration of one or more active gene copies of the present invention to an abnormally proliferating cell or cells. In a preferred embodiment, polynucleotides of the present invention is a DNA construct comprising a recombinant expression vector effective in expressing a DNA sequence encoding said polynucleotides. In another preferred embodiment of the present invention, the DNA construct encoding the poynucleotides of the present invention is inserted into cells to be treated utilizing a retrovirus, or more preferably an adenoviral vector (See G J. Nabel, et. al., PNAS 1999 96: 324-326, which is hereby incorporated by reference). In a most preferred embodiment, the viral vector is defective and will not transform non-proliferating cells, only proliferating cells. Moreover, in a preferred embodiment, the polynucleotides of the present invention inserted into proliferating cells either alone, or in combination with or fused to other polynucleotides, can then be modulated via an external stimulus (i.e. magnetic, specific small molecule, chemical, or drug administration, etc.), which acts upon the promoter upstream of said polynucleotides to induce expression of the encoded protein product. As such the beneficial therapeutic affect of the present invention may be expressly modulated (i.e. to increase, decrease, or inhibit expression of the present invention) based upon said external stimulus.

[0612] Polynucleotides of the present invention may be useful in repressing expression of oncogenic genes or antigens. By “repressing expression of the oncogenic genes” is intended the suppression of the transcription of the gene, the degradation of the gene transcript (pre-message RNA), the inhibition of splicing, the destruction of the messenger RNA, the prevention of the post-translational modifications of the protein, the destruction of the protein, or the inhibition of the normal function of the protein.

[0613] For local administration to abnormally proliferating cells, polynucleotides of the present invention may be administered by any method known to those of skill in the art including, but not limited to transfection, electroporation, microinjection of cells, or in vehicles such as liposomes, lipofectin, or as naked polynucleotides, or any other method described throughout the specification. The polynucleotide of the present invention may be delivered by known gene delivery systems such as, but not limited to, retroviral vectors (Gilboa, J. Virology 44:845 (1982); Hocke, Nature 320:275 (1986); Wilson, et al., Proc. Natl. Acad. Sci. U.S.A. 85:3014), vaccinia virus system (Chakrabarty et al., Mol. Cell Biol. 5:3403 (1985) or other efficient DNA delivery systems (Yates et al., Nature 313:812 (1985)) known to those skilled in the art. These references are exemplary only and are hereby incorporated by reference. In order to specifically deliver or transfect cells which are abnormally proliferating and spare non-dividing cells, it is preferable to utilize a retrovirus, or adenoviral (as described in the art and elsewhere herein) delivery system known to those of skill in the art. Since host DNA replication is required for retroviral DNA to integrate and the retrovirus will be unable to self replicate due to the lack of the retrovirus genes needed for its life cycle. Utilizing such a retroviral delivery system for polynucleotides of the present invention will target said gene and constructs to abnormally proliferating cells and will spare the non-dividing normal cells.

[0614] The polynucleotides of the present invention may be delivered directly to cell proliferative disorder/disease sites in internal organs, body cavities and the like by use of imaging devices used to guide an injecting needle directly to the disease site. The polynucleotides of the present invention may also be administered to disease sites at the time of surgical intervention.

[0615] By “cell proliferative disease” is meant any human or animal disease or disorder, affecting any one or any combination of organs, cavities, or body parts, which is characterized by single or multiple local abnormal proliferations of cells, groups of cells, or tissues, whether benign or malignant.

[0616] Any amount of the polynucleotides of the present invention may be administered as long as it has a biologically inhibiting effect on the proliferation of the treated cells. Moreover, it is possible to administer more than one of the polynucleotide of the present invention simultaneously to the same site. By “biologically inhibiting” is meant partial or total growth inhibition as well as decreases in the rate of proliferation or growth of the cells. The biologically inhibitory dose may be determined by assessing the effects of the polynucleotides of the present invention on target malignant or abnormally proliferating cell growth in tissue culture, tumor growth in animals and cell cultures, or any other method known to one of ordinary skill in the art.

[0617] The present invention is further directed to antibody-based therapies which involve administering of anti-polypeptides and anti-polynucleotide antibodies to a mammalian, preferably human, patient for treating one or more of the described disorders. Methods for producing anti-polypeptides and anti-polynucleotide antibodies polyclonal and monoclonal antibodies are described in detail elsewhere herein. Such antibodies may be provided in pharmaceutically acceptable compositions as known in the art or as described herein.

[0618] A summary of the ways in which the antibodies of the present invention may be used therapeutically includes binding polynucleotides or polypeptides of the present invention locally or systemically in the body or by direct cytotoxicity of the antibody, e.g. as mediated by complement (CDC) or by effector cells (ADCC). Some of these approaches are described in more detail below. Armed with the teachings provided herein, one of ordinary skill in the art will know how to use the antibodies of the present invention for diagnostic, monitoring or therapeutic purposes without undue experimentation.

[0619] In particular, the antibodies, fragments and derivatives of the present invention are useful for treating a subject having or developing cell proliferative and/or differentiation disorders as described herein. Such treatment comprises administering a single or multiple doses of the antibody, or a fragment, derivative, or a conjugate thereof.

[0620] The antibodies of this invention may be advantageously utilized in combination with other monoclonal or chimeric antibodies, or with lymphokines or hematopoietic growth factors, for example., which serve to increase the number or activity of effector cells which interact with the antibodies.

[0621] It is preferred to use high affinity and/or potent in vivo inhibiting and/or neutralizing antibodies against polypeptides or polynucleotides of the present invention, fragments or regions thereof, for both immunoassays directed to and therapy of disorders related to polynucleotides or polypeptides, including fragements thereof, of the present invention. Such antibodies, fragments, or regions, will preferably have an affinity for polynucleotides or polypeptides, including fragements thereof Preferred binding affinities include those with a dissociation constant or Kd less than 5×10−6M, 10−6M, 5×10−7M, 10−7M, 5×10−8 M, 10−8M, 5×10−9M, 10−9M, 5×10−10M, 10−10M, 5×10−11M, 10−11M, 5×10−12M, 10−12M, 5×10−13M, 10−13M, 5×10−14M, 10−14M, 5×10−15M, and 10−15M.

[0622] Moreover, polypeptides of the present invention are useful in inhibiting the angiogenesis of proliferative cells or tissues, either alone, as a protein fusion, or in combination with other polypeptides directly or indirectly, as described elsewhere herein. In a most preferred embodiment, said anti-angiogenesis effect may be achieved indirectly, for example, through the inhibition of hematopoietic, tumor-specific cells, such as tumor-associated macrophages (See Joseph I B, et al. J Natl Cancer Inst, 90(21):1648-53 (1998), which is hereby incorporated by reference). Antibodies directed to polypeptides or polynucleotides of the present invention may also result in inhibition of angiogenesis directly, or indirectly (See Witte L, et al., Cancer Metastasis Rev. 17(2):155-61 (1998), which is hereby incorporated by reference)).

[0623] Polypeptides, including protein fusions, of the present invention, or fragments thereof may be useful in inhibiting proliferative cells or tissues through the induction of apoptosis. Said polypeptides may act either directly, or indirectly to induce apoptosis of proliferative cells and tissues, for example in the activation of a death-domain receptor, such as tumor necrosis factor (TNF) receptor-1, CD95 (Fas/APO-1), TNF-receptor-related apoptosis-mediated protein (TRAMP) and TNF-related apoptosis-inducing ligand (TRAIL) receptor-1 and -2 (See Schulze-Osthoff K, et.al., Eur J Biochem 254(3):439-59 (1998), which is hereby incorporated by reference). Moreover, in another preferred embodiment of the present invention, said polypeptides may induce apoptosis through other mechanisms, such as in the activation of other proteins which will activate apoptosis, or through stimulating the expression of said proteins, either alone or in combination with small molecule drugs or adjuviants, such as apoptonin, galectins, thioredoxins, anti-inflammatory proteins (See for example, Mutat Res 400(1-2):447-55 (1998), Med Hypotheses.50(5):423-33 (1998), Chem Biol Interact. Apr 24;111-112:23-34 (1998), J Mol Med.76(6):402-12 (1998), Int J Tissue React;20(1):3-15 (1998), which are all hereby incorporated by reference).

[0624] Polypeptides, including protein fusions to, or fragments thereof, of the present invention are useful in inhibiting the metastasis of proliferative cells or tissues. Inhibition may occur as a direct result of administering polypeptides, or antibodies directed to said polypeptides as described elsewere herein, or indirectly, such as activating the expression of proteins known to inhibit metastasis, for example alpha 4,integrins, (See, e.g., Curr Top Microbiol Immunol 1998;231:125-41, which is hereby incorporated by reference). Such thereapeutic affects of the present invention may be achieved either alone, or in combination with small molecule drugs or adjuvants.

[0625] In another embodiment, the invention provides a method of delivering compositions containing the polypeptides of the invention (e.g., compositions containing polypeptides or polypeptide antibodes associated with heterologous polypeptides, heterologous nucleic acids, toxins, or prodrugs) to targeted cells expressing the polypeptide of the present invention. Polypeptides or polypeptide antibodes of the invention may be associated with with heterologous polypeptides, heterologous nucleic acids, toxins, or prodrugs via hydrophobic, hydrophilic, ionic and/or covalent interactions.

[0626] Polypeptides, protein fusions to, or fragments thereof, of the present invention are useful in enhancing the immunogenicity and/or antigenicity of proliferating cells or tissues, either directly, such as would occur if the polypeptides of the present invention ‘vaccinated’ the immune response to respond to proliferative antigens and immunogens, or indirectly, such as in activating the expression of proteins known to enhance the immune response (e.g. chemokines), to said antigens and immunogens.

[0627] Renal Disorders

[0628] Polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention, may be used to treat, prevent, diagnose, and/or prognose disorders of the renal system. Renal disorders which can be diagnosed, prognosed, prevented, and/or treated with compositions of the invention include, but are not limited to, kidney failure, nephritis, blood vessel disorders of kidney, metabolic and congenital kidney disorders, urinary disorders of the kidney, autoimmune disorders, sclerosis and necrosis, electrolyte imbalance, and kidney cancers.

[0629] Kidney diseases which can be diagnosed, prognosed, prevented, and/or treated with compositions of the invention include, but are not limited to, acute kidney failure, chronic kidney failure, atheroembolic renal failure, end-stage renal disease, inflammatory diseases of the kidney (e.g., acute glomerulonephritis, postinfectious glomerulonephritis, rapidly progressive glomerulonephritis, nephrotic syndrome, membranous glomerulonephritis, familial nephrotic syndrome, membranoproliferative glomerulonephritis I and II, mesangial proliferative glomerulonephritis, chronic glomerulonephritis, acute tubulointerstitial nephritis, chronic tubulointerstitial nephritis, acute post-streptococcal glomerulonephritis (PSGN), pyelonephritis, lupus nephritis, chronic nephritis, interstitial nephritis, and post-streptococcal glomerulonephritis), blood vessel disorders of the kidneys (e.g., kidney infarction, atheroembolic kidney disease, cortical necrosis, malignant nephrosclerosis, renal vein thrombosis, renal underperfusion, renal retinopathy, renal ischemia-reperfusion, renal artery embolism, and renal artery stenosis), and kidney disorders resulting form urinary tract disease (e.g., pyelonephritis, hydronephrosis, urolithiasis (renal lithiasis, nephrolithiasis), reflux nephropathy, urinary tract infections, urinary retention, and acute or chronic unilateral obstructive uropathy.)

[0630] In addition, compositions of the invention can be used to diagnose, prognose, prevent, and/or treat metabolic and congenital disorders of the kidney (e.g., uremia, renal amyloidosis, renal osteodystrophy, renal tubular acidosis, renal glycosuria, nephrogenic diabetes insipidus, cystinuria, Fanconi's syndrome, renal fibrocystic osteosis (renal rickets), Hartnup disease, Bartter's syndrome, Liddle's syndrome, polycystic kidney disease, medullary cystic disease, medullary sponge kidney, Alport's syndrome, nail-patella syndrome, congenital nephrotic syndrome, CRUSH syndrome, horseshoe kidney, diabetic nephropathy, nephrogenic diabetes insipidus, analgesic nephropathy, kidney stones, and membranous nephropathy), and autoimmune disorders of the kidney (e.g., systemic lupus erythematosus (SLE), Goodpasture syndrome, IgA nephropathy, and IgM mesangial proliferative glomerulonephritis).

[0631] Compositions of the invention can also be used to diagnose, prognose, prevent, and/or treat sclerotic or necrotic disorders of the kidney (e.g., glomerulosclerosis, diabetic nephropathy, focal segmental glomerulosclerosis (FSGS), necrotizing glomerulonephritis, and renal papillary necrosis), cancers of the kidney (e.g., nephroma, hypemephroma, nephroblastoma, renal cell cancer, transitional cell cancer, renal adenocarcinoma, squamous cell cancer, and Wilm's tumor), and electrolyte imbalances (e.g., nephrocalcinosis, pyuria, edema, hydronephritis, proteinuria, hyponatremia, hypematremia, hypokalernia, hyperkalemia, hypocalcemia, hypercalcemia, hypophosphatemia, and hyperphosphatemia).

[0632] Polypeptides may be administered using any method known in the art, including, but not limited to, direct needle injection at the delivery site, intravenous injection, topical administration, catheter infusion, biolistic injectors, particle accelerators, gelfoam sponge depots, other commercially available depot materials, osmotic pumps, oral or suppositorial solid pharmaceutical formulations, decanting or topical applications during surgery, aerosol delivery. Such methods are known in the art. Polypeptides may be administered as part of a Therapeutic, described in more detail below. Methods of delivering polynucleotides are described in more detail herein.

[0633] Cardiovascular Disorders

[0634] Polynucleotides or polypeptides, or agonists or antagonists of the present invention, may be used to treat, prevent, diagnose, and/or prognose cardiovascular disorders, including, but not limited to, peripheral artery disease, such as limb ischemia.

[0635] Cardiovascular disorders include, but are not limited to, cardiovascular abnormalities, such as arterio-arterial fistula, arterioyenous fistula, cerebral arterioyenous malformations, congenital heart defects, pulmonary atresia, and Scimitar Syndrome. Congenital heart defects include, but are not limited to, aortic coarctation, cor triatriatum, coronary vessel anomalies, crisscross heart, dextrocardia, patent ductus arteriosus, Ebstein's anomaly, Eisenmenger complex, hypoplastic left heart syndrome, levocardia, tetralogy of fallot, transposition of great vessels, double outlet right ventricle, tricuspid atresia, persistent truncus arteriosus, and heart septal defects, such as aortopulmonary septal defect, endocardial cushion defects, Lutembacher's Syndrome, trilogy of Fallot, ventricular heart septal defects.

[0636] Cardiovascular disorders also include, but are not limited to, heart disease, such as arrhythmias, carcinoid heart disease, high cardiac output, low cardiac output, cardiac tamponade, endocarditis (including bacterial), heart aneurysm, cardiac arrest, congestive heart failure, congestive cardiomyopathy, paroxysmal dyspnea, cardiac edema, heart hypertrophy, congestive cardiomyopathy, left ventricular hypertrophy, right ventricular hypertrophy, post-infarction heart rupture, ventricular septal rupture, heart valve diseases, myocardial diseases, myocardial ischemia, pericardial effusion, pericarditis (including constrictive and tuberculous), pneumopericardium, postpericardiotomy syndrome, pulmonary heart disease, rheumatic heart disease, ventricular dysfunction, hyperemia, cardiovascular pregnancy complications, Scimitar Syndrome, cardiovascular syphilis, and cardiovascular tuberculosis.

[0637] Arrhythmias include, but are not limited to, sinus arrhythmia, atrial fibrillation, atrial flutter, bradycardia, extrasystole, Adams-Stokes Syndrome, bundle-branch block, sinoatrial block, long QT syndrome, parasystole, Lown-Ganong-Levine Syndrome, Mahaim-type pre-excitation syndrome, Wolff-Parkinson-White syndrome, sick sinus syndrome, tachycardias, and ventricular fibrillation. Tachycardias include paroxysmal tachycardia, supraventricular tachycardia, accelerated idioventricular rhythm, atrioventricular nodal reentry tachycardia, ectopic atrial tachycardia, ectopic junctional tachycardia, sinoatrial nodal reentry tachycardia, sinus tachycardia, Torsades de Pointes, and ventricular tachycardia.

[0638] Heart valve diseases include, but are not limited to, aortic valve insufficiency, aortic valve stenosis, hear murmurs, aortic valve prolapse, mitral valve prolapse, tricuspid valve prolapse, mitral valve insufficiency, mitral valve stenosis, pulmonary atresia, pulmonary valve insufficiency, pulmonary valve stenosis, tricuspid atresia, tricuspid valve insufficiency, and tricuspid valve stenosis.

[0639] Myocardial diseases include, but are not limited to, alcoholic cardiomyopathy, congestive cardiomyopathy, hypertrophic cardiomyopathy, aortic subvalvular stenosis, pulmonary subvalvular stenosis, restrictive cardiomyopathy, Chagas cardiomyopathy, endocardial fibroelastosis, endomyocardial fibrosis, Kearns Syndrome, myocardial reperfusion injury, and myocarditis.

[0640] Myocardial ischemias include, but are not limited to, coronary disease, such as angina pectoris, coronary aneurysm, coronary arteriosclerosis, coronary thrombosis, coronary vasospasm, myocardial infarction and myocardial stunning.

[0641] Cardiovascular diseases also include vascular diseases such as aneurysms, angiodysplasia, angiomatosis, bacillary angiomatosis, Hippel-Lindau Disease, Klippel-Trenaunay-Weber Syndrome, Sturge-Weber Syndrome, angioneurotic edema, aortic diseases, Takayasu's Arteritis, aortitis, Leriche's Syndrome, arterial occlusive diseases, arteritis, enarteritis, polyarteritis nodosa, cerebrovascular disorders, diabetic angiopathies, diabetic retinopathy, embolisms, thrombosis, erythromelalgia, hemorrhoids, hepatic veno-occlusive disease, hypertension, hypotension, ischemia, peripheral vascular diseases, phlebitis, pulmonary veno-occlusive disease, Raynaud's disease, CREST syndrome, retinal vein occlusion, Scimitar syndrome, superior vena cava syndrome, telangiectasia, atacia telangiectasia, hereditary hemorrhagic telangiectasia, varicocele, varicose veins, varicose ulcer, vasculitis, and venous insufficiency.

[0642] Aneurysms include, but are not limited to, dissecting aneurysms, false aneurysms, infected aneurysms, ruptured aneurysms, aortic aneurysms, cerebral aneurysms, coronary aneurysms, heart aneurysms, and iliac aneurysms.

[0643] Arterial occlusive diseases include, but are not limited to, arteriosclerosis, intermittent claudication, carotid stenosis, fibromuscular dysplasias, mesenteric vascular occlusion, Moyamoya disease, renal artery obstruction, retinal artery occlusion, and thromboangiitis obliterans.

[0644] Cerebrovascular disorders include, but are not limited to, carotid artery diseases, cerebral amyloid angiopathy, cerebral aneurysm, cerebral anoxia, cerebral arteriosclerosis, cerebral arterioyenous malformation, cerebral artery diseases, cerebral embolism and thrombosis, carotid artery thrombosis, sinus thrombosis, Wallenberg's syndrome, cerebral hemorrhage, epidural hematoma, subdural hematoma, subaraxhnoid hemorrhage, cerebral infarction, cerebral ischemia (including transient), subclavian steal syndrome, periventricular leukomalacia, vascular headache, cluster headache, migraine, and vertebrobasilar insufficiency.

[0645] Embolisms include, but are not limited to, air embolisms, amniotic fluid embolisms, cholesterol embolisms, blue toe syndrome, fat embolisms, pulmonary embolisms, and thromoboembolisms. Thrombosis include, but are not limited to, coronary thrombosis, hepatic vein thrombosis, retinal vein occlusion, carotid artery thrombosis, sinus thrombosis, Wallenberg's syndrome, and thrombophlebitis.

[0646] Ischemic disorders include, but are not limited to, cerebral ischemia, ischemic colitis, compartment syndromes, anterior compartment syndrome, myocardial ischemia, reperfusion injuries, and peripheral limb ischemia. Vasculitis includes, but is not limited to, aortitis, arteritis, Behcet's Syndrome, Churg-Strauss Syndrome, mucocutaneous lymph node syndrome, thromboangiitis obliterans, hypersensitivity vasculitis, Schoenlein-Henoch purpura, allergic cutaneous vasculitis, and Wegener's granulomatosis.

[0647] Polypeptides may be administered using any method known in the art, including, but not limited to, direct needle injection at the delivery site, intravenous injection, topical administration, catheter infusion, biolistic injectors, particle accelerators, gelfoam sponge depots, other commercially available depot materials, osmotic pumps, oral or suppositorial solid pharmaceutical formulations, decanting or topical applications during surgery, aerosol delivery. Such methods are known in the art. Polypeptides may be administered as part of a Therapeutic, described in more detail below. Methods of delivering polynucleotides are described in more detail herein.

[0648] Respiratory Disorders

[0649] Polynucleotides or polypeptides, or agonists or antagonists of the present invention may be used to treat, prevent, diagnose, and/or prognose diseases and/or disorders of the respiratory system.

[0650] Diseases and disorders of the respiratory system include, but are not limited to, nasal vestibulitis, nonallergic rhinitis (e.g., acute rhinitis, chronic rhinitis, atrophic rhinitis, vasomotor rhinitis), nasal polyps, and sinusitis, juvenile angiofibromas, cancer of the nose and juvenile papillomas, vocal cord polyps, nodules (singer's nodules), contact ulcers, vocal cord paralysis, laryngoceles, pharyngitis (e.g., viral and bacterial), tonsillitis, tonsillar cellulitis, parapharyngeal abscess, laryngitis, laryngoceles, and throat cancers (e.g., cancer of the nasopharynx, tonsil cancer, larynx cancer), lung cancer (e.g., squamous cell carcinoma, small cell (oat cell) carcinoma, large cell carcinoma, and adenocarcinoma), allergic disorders (eosinophilic pneumonia, hypersensitivity pneumonitis (e.g., extrinsic allergic alveolitis, allergic interstitial pneumonitis, organic dust pneumoconiosis, allergic bronchopulmonary aspergillosis, asthma, Wegener's granulomatosis (granulomatous vasculitis), Goodpasture's syndrome)), pneumonia (e.g., bacterial pneumonia (e.g., Streptococcus pneumoniae (pneumoncoccal pneumonia), Staphylococcus aureus (staphylococcal pneumonia), Gram-negative bacterial pneumonia (caused by, e.g., Klebsiella and Pseudomas spp.), Mycoplasma pneumoniae pneumonia, Hemophilus influenzae pneumonia, Legionella pneumophila (Legionnaires' disease), and Chlamydia psittaci (Psittacosis)), and viral pneumonia (e.g., influenza, chickenpox (varicella).

[0651] Additional diseases and disorders of the respiratory system include, but are not limited to bronchiolitis, polio (poliomyelitis), croup, respiratory syncytial viral infection, mumps, erythema infectiosum (fifth disease), roseola infantum, progressive rubella panencephalitis, german measles, and subacute sclerosing panencephalitis), fungal pneumonia (e.g., Histoplasmosis, Coccidioidomycosis, Blastomycosis, fungal infections in people with severely suppressed immune systems (e.g., cryptococcosis, caused by Cryptococcus neoformans; aspergillosis, caused by Aspergillus spp.; candidiasis, caused by Candida; and mucormycosis)), Pneumocystis carinii (pneumocystis pneumonia), a typical pneumonias (e.g., Mycoplasma and Chlamydia spp.), opportunistic infection pneumonia, nosocomial pneumonia, chemical pneumonitis, and aspiration pneumonia, pleural disorders (e.g., pleurisy, pleural effusion, and pneumothorax (e.g., simple spontaneous pneumothorax, complicated spontaneous pneumothorax, tension pneumothorax)), obstructive airway diseases (e.g., asthma, chronic obstructive pulmonary disease (COPD), emphysema, chronic or acute bronchitis), occupational lung diseases (e.g., silicosis, black lung (coal workers' pneumoconiosis), asbestosis, berylliosis, occupational asthsma, byssinosis, and benign pneumoconioses), Infiltrative Lung Disease (e.g., pulmonary fibrosis (e.g., fibrosing alveolitis, usual interstitial pneumonia), idiopathic pulmonary fibrosis, desquamative interstitial pneumonia, lymphoid interstitial pneumonia, histiocytosis X (e.g., Letterer-Siwe disease, Hand-Schüller-Christian disease, eosinophilic granuloma), idiopathic pulmonary hemosiderosis, sarcoidosis and pulmonary alveolar proteinosis), Acute respiratory distress syndrome (also called, e.g., adult respiratory distress syndrome), edema, pulmonary embolism, bronchitis (e.g., viral, bacterial), bronchiectasis, atelectasis, lung abscess (caused by, e.g., Staphylococcus aureus or Legionella pneumophila), and cystic fibrosis.

[0652] Anti-Angiogenesis Activity

[0653] The naturally occurring balance between endogenous stimulators and inhibitors of angiogenesis is one in which inhibitory influences predominate. Rastinejad et al., Cell 56:345-355 (1989). In those rare instances in which neovascularization occurs under normal physiological conditions, such as wound healing, organ regeneration, embryonic development, and female reproductive processes, angiogenesis is stringently regulated and spatially and temporally delimited. Under conditions of pathological angiogenesis such as that characterizing solid tumor growth, these regulatory controls fail. Unregulated angiogenesis becomes pathologic and sustains progression of many neoplastic and non-neoplastic diseases. A number of serious diseases are dominated by abnormal neovascularization including solid tumor growth and metastases, arthritis, some types of eye disorders, and psoriasis. See, e.g., reviews by Moses et al., Biotech. 9:630-634 (1991); Folkman et al., N. Engl. J. Med., 333:1757-1763 (1995); Auerbach et al., J Microvasc. Res. 29:401-411 (1985); Folkman, Advances in Cancer Research, eds. Klein and Weinhouse, Academic Press, New York, pp. 175-203 (1985); Patz, Am. J. Opthalmol. 94:715-743 (1982); and Folkman et al., Science 221:719-725 (1983). In a number of pathological conditions, the process of angiogenesis contributes to the disease state. For example, significant data have accumulated which suggest that the growth of solid tumors is dependent on angiogenesis. Folkman and Klagsbrun, Science 235:442-447 (1987).

[0654] The present invention provides for treatment of diseases or disorders associated with neovascularization by administration of the polynucleotides and/or polypeptides of the invention, as well as agonists or antagonists of the present invention. Malignant and metastatic conditions which can be treated with the polynucleotides and polypeptides, or agonists or antagonists of the invention include, but are not limited to, malignancies, solid tumors, and cancers described herein and otherwise known in the art (for a review of such disorders, see Fishman et al., Medicine, 2d Ed., J. B. Lippincott Co., Philadelphia (1985)). Thus, the present invention provides a method of treating an angiogenesis-related disease and/or disorder, comprising administering to an individual in need thereof a therapeutically effective amount of a polynucleotide, polypeptide, antagonist and/or agonist of the invention. For example, polynucleotides, polypeptides, antagonists and/or agonists may be utilized in a variety of additional methods in order to therapeutically treat a cancer or tumor. Cancers which may be treated with polynucleotides, polypeptides, antagonists and/or agonists include, but are not limited to solid tumors, including prostate, lung, breast, ovarian, stomach, pancreas, larynx, esophagus, testes, liver, parotid, biliary tract, colon, rectum, cervix, uterus, endometrium, kidney, bladder, thyroid cancer; primary tumors and metastases; melanomas; glioblastoma; Kaposi's sarcoma; leiomyosarcoma; non-small cell lung cancer; colorectal cancer; advanced malignancies; and blood born tumors such as leukemias. For example, polynucleotides, polypeptides, antagonists and/or agonists may be delivered topically, in order to treat cancers such as skin cancer, head and neck tumors, breast tumors, and Kaposi's sarcoma.

[0655] Within yet other aspects, polynucleotides, polypeptides, antagonists and/or agonists may be utilized to treat superficial forms of bladder cancer by, for example, intravesical administration. Polynucleotides, polypeptides, antagonists and/or agonists may be delivered directly into the tumor, or near the tumor site, via injection or a catheter. Of course, as the artisan of ordinary skill will appreciate, the appropriate mode of administration will vary according to the cancer to be treated. Other modes of delivery are discussed herein.

[0656] Polynucleotides, polypeptides, antagonists and/or agonists may be useful in treating other disorders, besides cancers, which involve angiogenesis. These disorders include, but are not limited to: benign tumors, for example hemangiomas, acoustic neuromas, neurofibromas, trachomas, and pyogenic granulomas; artheroscleric plaques; ocular angiogenic diseases, for example, diabetic retinopathy, retinopathy of prematurity, macular degeneration, corneal graft rejection, neovascular glaucoma, retrolental fibroplasia, rubeosis, retinoblastoma, uvietis and Pterygia (abnormal blood vessel growth) of the eye; rheumatoid arthritis; psoriasis; delayed wound healing; endometriosis; vasculogenesis; granulations; hypertrophic scars (keloids); nonunion fractures; scleroderma; trachoma; vascular adhesions; myocardial angiogenesis; coronary collaterals; cerebral collaterals; arterioyenous malformations; ischemic limb angiogenesis; Osler-Webber Syndrome; plaque neovascularization; telangiectasia; hemophiliac joints; angiofibroma; fibromuscular dysplasia; wound granulation; Crohn's disease; and atherosclerosis.

[0657] For example, within one aspect of the present invention methods are provided for treating hypertrophic scars and keloids, comprising the step of administering a polynucleotide, polypeptide, antagonist and/or agonist of the invention to a hypertrophic scar or keloid.

[0658] Within one embodiment of the present invention polynucleotides, polypeptides, antagonists and/or agonists of the invention are directly injected into a hypertrophic scar or keloid, in order to prevent the progression of these lesions. This therapy is of particular value in the prophylactic treatment of conditions which are known to result in the development of hypertrophic scars and keloids (e.g., bums), and is preferably initiated after the proliferative phase has had time to progress (approximately 14 days after the initial injury), but before hypertrophic scar or keloid development. As noted above, the present invention also provides methods for treating neovascular diseases of the eye, including for example, comeal neovascularization, neovascular glaucoma, proliferative diabetic retinopathy, retrolental fibroplasia and macular degeneration.

[0659] Moreover, Ocular disorders associated with neovascularization which can be treated with the polynucleotides and polypeptides of the present invention (including agonists and/or antagonists) include, but are not limited to: neovascular glaucoma, diabetic retinopathy, retinoblastoma, retrolental fibroplasia, uveitis, retinopathy of prematurity macular degeneration, comeal graft neovascularization, as well as other eye inflammatory diseases, ocular tumors and diseases associated with choroidal or iris neovascularization. See, e.g., reviews by Waltman et al., Am. J. Ophthal. 85:704-710 (1978) and Gartner et al, Surv. Ophthal. 22:291-312 (1978).

[0660] Thus, within one aspect of the present invention methods are provided for treating neovascular diseases of the eye such as corneal neovascularization (including corneal graft neovascularization), comprising the step of administering to a patient a therapeutically effective amount of a compound (as described above) to the cornea, such that the formation of blood vessels is inhibited. Briefly, the cornea is a tissue which normally lacks blood vessels. In certain pathological conditions however, capillaries may extend into the cornea from the pericorneal vascular plexus of the limbus. When the cornea becomes vascularized, it also becomes clouded, resulting in a decline in the patient's visual acuity. Visual loss may become complete if the cornea completely opacitates. A wide variety of disorders can result in corneal neovascularization, including for example, corneal infections (e.g., trachoma, herpes simplex keratitis, leishmaniasis and onchocerciasis), immunological processes (e.g., graft rejection and Stevens-Johnson's syndrome), alkali bums, trauma, inflammation (of any cause), toxic and nutritional deficiency states, and as a complication of wearing contact lenses.

[0661] Within particularly preferred embodiments of the invention, may be prepared for topical administration in saline (combined with any of the preservatives and antimicrobial agents commonly used in ocular preparations), and administered in eyedrop form. The solution or suspension may be prepared in its pure form and administered several times daily. Alternatively, anti-angiogenic compositions, prepared as described above, may also be administered directly to the cornea. Within preferred embodiments, the anti-angiogenic composition is prepared with a muco-adhesive polymer which binds to cornea. Within further embodiments, the anti-angiogenic factors or anti-angiogenic compositions may be utilized as an adjunct to conventional steroid therapy. Topical therapy may also be useful prophylactically in corneal lesions which are known to have a high probability of inducing an angiogenic response (such as chemical burns). In these instances the treatment, likely in combination with steroids, may be instituted immediately to help prevent subsequent complications.

[0662] Within other embodiments, the compounds described above may be injected directly into the corneal stroma by an ophthalmologist under microscopic guidance. The preferred site of injection may vary with the morphology of the individual lesion, but the goal of the administration would be to place the composition at the advancing front of the vasculature (i.e., interspersed between the blood vessels and the normal cornea). In most cases this would involve perilimbic corneal injection to “protect” the cornea from the advancing blood vessels. This method may also be utilized shortly after a corneal insult in order to prophylactically prevent corneal neovascularization. In this situation the material could be injected in the perilimbic cornea interspersed between the corneal lesion and its undesired potential limbic blood supply. Such methods may also be utilized in a similar fashion to prevent capillary invasion of transplanted corneas. In a sustained-release form injections might only be required 2-3 times per year. A steroid could also be added to the injection solution to reduce inflammation resulting from the injection itself.

[0663] Within another aspect of the present invention, methods are provided for treating neovascular glaucoma, comprising the step of administering to a patient a therapeutically effective amount of a polynucleotide, polypeptide, antagonist and/or agonist to the eye, such that the formation of blood vessels is inhibited. In one embodiment, the compound may be administered topically to the eye in order to treat early forms of neovascular glaucoma. Within other embodiments, the compound may be implanted by injection into the region of the anterior chamber angle. Within other embodiments, the compound may also be placed in any location such that the compound is continuously released into the aqueous humor. Within another aspect of the present invention, methods are provided for treating proliferative diabetic retinopathy, comprising the step of administering to a patient a therapeutically effective amount of a polynucleotide, polypeptide, antagonist and/or agonist to the eyes, such that the formation of blood vessels is inhibited.

[0664] Within particularly preferred embodiments of the invention, proliferative diabetic retinopathy may be treated by injection into the aqueous humor or the vitreous, in order to increase the local concentration of the polynucleotide, polypeptide, antagonist and/or agonist in the retina. Preferably, this treatment should be initiated prior to the acquisition of severe disease requiring photocoagulation.

[0665] Within another aspect of the present invention, methods are provided for treating retrolental fibroplasia, comprising the step of administering to a patient a therapeutically effective amount of a polynucleotide, polypeptide, antagonist and/or agonist to the eye, such that the formation of blood vessels is inhibited. The compound may be administered topically, via intravitreous injection and/or via intraocular implants.

[0666] Additionally, disorders which can be treated with the polynucleotides, polypeptides, agonists and/or agonists include, but are not limited to, hemangioma, arthritis, psoriasis, angiofibroma, atherosclerotic plaques, delayed wound healing, granulations, hemophilic joints, hypertrophic scars, nonunion fractures, Osler-Weber syndrome, pyogenic granuloma, scleroderma, trachoma, and vascular adhesions.

[0667] Moreover, disorders and/or states, which can be treated, prevented, diagnosed, and/or prognosed with the the polynucleotides, polypeptides, agonists and/or agonists of the invention include, but are not limited to, solid tumors, blood born tumors such as leukemias, tumor metastasis, Kaposi's sarcoma, benign tumors, for example hemangiomas, acoustic neuromas, neurofibromas, trachomas, and pyogenic granulomas, rheumatoid arthritis, psoriasis, ocular angiogenic diseases, for example, diabetic retinopathy, retinopathy of prematurity, macular degeneration, corneal graft rejection, neovascular glaucoma, retrolental fibroplasia, rubeosis, retinoblastoma, and uvietis, delayed wound healing, endometriosis, vascluogenesis, granulations, hypertrophic scars (keloids), nonunion fractures, scleroderma, trachoma, vascular adhesions, myocardial angiogenesis, coronary collaterals, cerebral collaterals, arterioyenous malformations, ischemic limb angiogenesis, Osler-Webber Syndrome, plaque neovascularization, telangiectasia, hemophiliac joints, angiofibroma fibromuscular dysplasia, wound granulation, Crohn's disease, atherosclerosis, birth control agent by preventing vascularization required for embryo implantation controlling menstruation, diseases that have angiogenesis as a pathologic consequence such as cat scratch disease (Rochele minalia quintosa), ulcers (Helicobacter pylori), Bartonellosis and bacillary angiomatosis.

[0668] In one aspect of the birth control method, an amount of the compound sufficient to block embryo implantation is administered before or after intercourse and fertilization have occurred, thus providing an effective method of birth control, possibly a “morning after” method. Polynucleotides, polypeptides, agonists and/or agonists may also be used in controlling menstruation or administered as either a peritoneal lavage fluid or for peritoneal implantation in the treatment of endometriosis.

[0669] Polynucleotides, polypeptides, agonists and/or agonists of the present invention may be incorporated into surgical sutures in order to prevent stitch granulomas.

[0670] Polynucleotides, polypeptides, agonists and/or agonists may be utilized in a wide variety of surgical procedures. For example, within one aspect of the present invention a compositions (in the form of, for example, a spray or film) may be utilized to coat or spray an area prior to removal of a tumor, in order to isolate normal surrounding tissues from malignant tissue, and/or to prevent the spread of disease to surrounding tissues. Within other aspects of the present invention, compositions (e.g., in the form of a spray) may be delivered via endoscopic procedures in order to coat tumors, or inhibit angiogenesis in a desired locale. Within yet other aspects of the present invention, surgical meshes which have been coated with anti-angiogenic compositions of the present invention may be utilized in any procedure wherein a surgical mesh might be utilized. For example, within one embodiment of the invention a surgical mesh laden with an anti-angiogenic composition may be utilized during abdominal cancer resection surgery (e.g., subsequent to colon resection) in order to provide support to the structure, and to release an amount of the anti-angiogenic factor.

[0671] Within further aspects of the present invention, methods are provided for treating tumor excision sites, comprising administering a polynucleotide, polypeptide, agonist and/or agonist to the resection margins of a tumor subsequent to excision, such that the local recurrence of cancer and the formation of new blood vessels at the site is inhibited. Within one embodiment of the invention, the anti-angiogenic compound is administered directly to the tumor excision site (e.g., applied by swabbing, brushing or otherwise coating the resection margins of the tumor with the anti-angiogenic compound). Alternatively, the anti-angiogenic compounds may be incorporated into known surgical pastes prior to administration. Within particularly preferred embodiments of the invention, the anti-angiogenic compounds are applied after hepatic resections for malignancy, and after neurosurgical operations.

[0672] Within one aspect of the present invention, polynucleotides, polypeptides, agonists and/or agonists may be administered to the resection margin of a wide variety of tumors, including for example, breast, colon, brain and hepatic tumors. For example, within one embodiment of the invention, anti-angiogenic compounds may be administered to the site of a neurological tumor subsequent to excision, such that the formation of new blood vessels at the site are inhibited.

[0673] The polynucleotides, polypeptides, agonists and/or agonists of the present invention may also be administered along with other anti-angiogenic factors. Representative examples of other anti-angiogenic factors include: Anti-Invasive Factor, retinoic acid and derivatives thereof, paclitaxel, Suramin, Tissue Inhibitor of Metalloproteinase-1, Tissue Inhibitor of Metalloproteinase-2, Plasminogen Activator Inhibitor-1, Plasminogen Activator Inhibitor-2, and various forms of the lighter “d group” transition metals.

[0674] Lighter “d group” transition metals include, for example, vanadium, molybdenum, tungsten, titanium, niobium, and tantalum species. Such transition metal species may form transition metal complexes. Suitable complexes of the above-mentioned transition metal species include oxo transition metal complexes.

[0675] Representative examples of vanadium complexes include oxo vanadium complexes such as vanadate and vanadyl complexes. Suitable vanadate complexes include metavanadate and orthovanadate complexes such as, for example, ammonium metavanadate, sodium metavanadate, and sodium orthovanadate. Suitable vanadyl complexes include, for example, vanadyl acetylacetonate and vanadyl sulfate including vanadyl sulfate hydrates such as vanadyl sulfate mono- and trihydrates.

[0676] Representative examples of tungsten and molybdenum complexes also include oxo complexes. Suitable oxo tungsten complexes include tungstate and tungsten oxide complexes. Suitable tungstate complexes include ammonium tungstate, calcium tungstate, sodium tungstate dihydrate, and tungstic acid. Suitable tungsten oxides include tungsten (IV) oxide and tungsten (VI) oxide. Suitable oxo molybdenum complexes include molybdate, molybdenum oxide, and molybdenyl complexes. Suitable molybdate complexes include ammonium molybdate and its hydrates, sodium molybdate and its hydrates, and potassium molybdate and its hydrates. Suitable molybdenum oxides include molybdenum (VI) oxide, molybdenum (VI) oxide, and molybdic acid. Suitable molybdenyl complexes include, for example, molybdenyl acetylacetonate. Other suitable tungsten and molybdenum complexes include hydroxo derivatives derived from, for example, glycerol, tartaric acid, and sugars.

[0677] A wide variety of other anti-angiogenic factors may also be utilized within the context of the present invention. Representative examples include platelet factor 4; protamine sulphate; sulphated chitin derivatives (prepared from queen crab shells), (Murata et al., Cancer Res. 51:22-26, 1991); Sulphated Polysaccharide Peptidoglycan Complex (SP-PG) (the function of this compound may be enhanced by the presence of steroids such as estrogen, and tamoxifen citrate); Staurosporine; modulators of matrix metabolism, including for example, proline analogs, cishydroxyproline, d,L-3,4-dehydroproline, Thiaproline, alpha,alpha-dipyridyl, aminopropionitrile fumarate; 4-propyl-5-(4-pyridinyl)-2(3H)-oxazolone; Methotrexate; Mitoxantrone; Heparin; Interferons; 2 Macroglobulin-serum; ChIMP-3 (Pavloff et al., J. Bio. Chem. 267:17321-17326, 1992); Chymostatin (Tomkinson et al., Biochem J. 286:475-480, 1992); Cyclodextrin Tetradecasulfate; Eponemycin; Camptothecin; Fumagillin (Ingber et al., Nature 348:555-557, 1990); Gold Sodium Thiomalate (“GST”; Matsubara and Ziff, J. Clin. Invest. 79:1440-1446, 1987); anticollagenase-serum; alpha2-antiplasmin (Holmes et al., J. Biol. Chem. 262(4):1659-1664, 1987); Bisantrene (National Cancer Institute); Lobenzarit disodium (N-(2)-carboxyphenyl-4-chloroanthronilic acid disodium or “CCA”; Takeuchi et al., Agents Actions 36:312-316, 1992); Thalidomide; Angostatic steroid; AGM-1470; carboxynaminolmidazole; and metalloproteinase inhibitors such as BB94.

[0678] Diseases at the Cellular Level

[0679] Diseases associated with increased cell survival or the inhibition of apoptosis that could be treated, prevented, diagnosed, and/or prognosed using polynucleotides or polypeptides, as well as antagonists or agonists of the present invention, include cancers (such as follicular lymphomas, carcinomas with p53 mutations, and hormone-dependent tumors, including, but not limited to colon cancer, cardiac tumors, pancreatic cancer, melanoma, retinoblastoma, glioblastoma, lung cancer, intestinal cancer, testicular cancer, stomach cancer, neuroblastoma, myxoma, myoma, lymphoma, endothelioma, osteoblastoma, osteoclastoma, osteosarcoma, chondrosarcoma, adenoma, breast cancer, prostate cancer, Kaposi's sarcoma and ovarian cancer); autoimmune disorders (such as, multiple sclerosis, Sjogren's syndrome, Hashimoto's thyroiditis, biliary cirrhosis, Behcet's disease, Crohn's disease, polymyositis, systemic lupus erythematosus and immune-related glomerulonephritis and rheumatoid arthritis) and viral infections (such as herpes viruses, pox viruses and adenoviruses), inflammation, graft v. host disease, acute graft rejection, and chronic graft rejection.

[0680] In preferred embodiments, polynucleotides, polypeptides, and/or antagonists of the invention are used to inhibit growth, progression, and/or metasis of cancers, in particular those listed above.

[0681] Additional diseases or conditions associated with increased cell survival that could be treated or detected by polynucleotides or polypeptides, or agonists or antagonists of the present invention include, but are not limited to, progression, and/or metastases of malignancies and related disorders such as leukemia (including acute leukemias (e.g., acute lymphocytic leukemia, acute myelocytic leukemia (including myeloblastic, promyelocytic, myelomonocytic, monocytic, and erythroleukemia)) and chronic leukemias (e.g., chronic myelocytic (granulocytic) leukemia and chronic lymphocytic leukemia)), polycythemia vera, lymphomas (e.g., Hodgkin's disease and non-Hodgkin's disease), multiple myeloma, Waldenstrom's macroglobulinemia, heavy chain disease, and solid tumors including, but not limited to, sarcomas and carcinomas such as fibrosarcoma, myxosarcoma, liposarcoma, chondrosarcoma, osteogenic sarcoma, chordoma, angiosarcoma, endotheliosarcoma, lymphangiosarcoma, lymphangioendotheliosarcoma, synovioma, mesothelioma, Ewing's tumor, leiomyosarcoma, rhabdomyosarcoma, colon carcinoma, pancreatic cancer, breast cancer, ovarian cancer, prostate cancer, squamous cell carcinoma, basal cell carcinoma, adenocarcinoma, sweat gland carcinoma, sebaceous gland carcinoma, papillary carcinoma, papillary adenocarcinomas, cystadenocarcinoma, medullary carcinoma, bronchogenic carcinoma, renal cell carcinoma, hepatoma, bile duct carcinoma, choriocarcinoma, seminoma, embryonal carcinoma, Wilm's tumor, cervical cancer, testicular tumor, lung carcinoma, small cell lung carcinoma, bladder carcinoma, epithelial carcinoma, glioma, astrocytoma, medulloblastoma, craniopharyngioma, ependymoma, pinealoma, hemangioblastoma, acoustic neuroma, oligodendroglioma, menangioma, melanoma, neuroblastoma, and retinoblastoma.

[0682] Diseases associated with increased apoptosis that could be treated, prevented, diagnosed, and/or prognesed using polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, include, but are not limited to, AIDS; neurodegenerative disorders (such as Alzheimer's disease, Parkinson's disease, Amyotrophic lateral sclerosis, Retinitis pigmentosa, Cerebellar degeneration and brain tumor or prior associated disease); autoimmune disorders (such as, multiple sclerosis, Sjogren's syndrome, Hashimoto's thyroiditis, biliary cirrhosis, Behcet's disease, Crohn's disease, polymyositis, systemic lupus erythematosus and immune-related glomerulonephritis and rheumatoid arthritis) myelodysplastic syndromes (such as aplastic anemia), graft v. host disease, ischemic injury (such as that caused by myocardial infarction, stroke and reperfusion injury), liver injury (e.g., hepatitis related liver injury, ischemia/reperfusion injury, cholestosis (bile duct injury) and liver cancer); toxin-induced liver disease (such as that caused by alcohol), septic shock, cachexia and anorexia.

[0683] Wound Healing and Epithelial Cell Proliferation

[0684] In accordance with yet a further aspect of the present invention, there is provided a process for utilizing polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, for therapeutic purposes, for example, to stimulate epithelial cell proliferation and basal keratinocytes for the purpose of wound healing, and to stimulate hair follicle production and healing of dermal wounds. Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, may be clinically useful in stimulating wound healing including surgical wounds, excisional wounds, deep wounds involving damage of the dermis and epidermis, eye tissue wounds, dental tissue wounds, oral cavity wounds, diabetic ulcers, dermal ulcers, cubitus ulcers, arterial ulcers, venous stasis ulcers, burns resulting from heat exposure or chemicals, and other abnormal wound healing conditions such as uremia, malnutrition, vitamin deficiencies and complications associated with systemic treatment with steroids, radiation therapy and antineoplastic drugs and antimetabolites. Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could be used to promote dermal reestablishment subsequent to dermal loss

[0685] Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could be used to increase the adherence of skin grafts to a wound bed and to stimulate re-epithelialization from the wound bed. The following are types of grafts that polynucleotides or polypeptides, agonists or antagonists of the present invention, could be used to increase adherence to a wound bed: autografts, artificial skin, allografts, autodermic graft, autoepdermic grafts, avacular grafts, Blair-Brown grafts, bone graft, brephoplastic grafts, cutis graft, delayed graft, dermic graft, epidermic graft, fascia graft, full thickness graft, heterologous graft, xenograft, homologous graft, hyperplastic graft, lamellar graft, mesh graft, mucosal graft, Ollier-Thiersch graft, omenpal graft, patch graft, pedicle graft, penetrating graft, split skin graft, thick split graft. Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, can be used to promote skin strength and to improve the appearance of aged skin.

[0686] It is believed that polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, will also produce changes in hepatocyte proliferation, and epithelial cell proliferation in the lung, breast, pancreas, stomach, small intestine, and large intestine. Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could promote proliferation of epithelial cells such as sebocytes, hair follicles, hepatocytes, type II pneumocytes, mucin-producing goblet cells, and other epithelial cells and their progenitors contained within the skin, lung, liver, and gastrointestinal tract. Polynucleotides or polypeptides, agonists or antagonists of the present invention, may promote proliferation of endothelial cells, keratinocytes, and basal keratinocytes.

[0687] Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could also be used to reduce the side effects of gut toxicity that result from radiation, chemotherapy treatments or viral infections. Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, may have a cytoprotective effect on the small intestine mucosa. Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, may also stimulate healing of mucositis (mouth ulcers) that result from chemotherapy and viral infections.

[0688] Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could further be used in full regeneration of skin in full and partial thickness skin defects, including burns, (i.e., repopulation of hair follicles, sweat glands, and sebaceous glands), treatment of other skin defects such as psoriasis. Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could be used to treat epidermolysis bullosa, a defect in adherence of the epidermis to the underlying dermis which results in frequent, open and painful blisters by accelerating reepithelialization of these lesions. Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could also be used to treat gastric and doudenal ulcers and help heal by scar formation of the mucosal lining and regeneration of glandular mucosa and duodenal mucosal lining more rapidly. Inflammatory bowel diseases, such as Crohn's disease and ulcerative colitis, are diseases which result in destruction of the mucosal surface of the small or large intestine, respectively. Thus, polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could be used to promote the resurfacing of the mucosal surface to aid more rapid healing and to prevent progression of inflammatory bowel disease. Treatment with polynucleotides or polypeptides, agonists or antagonists of the present invention, is expected to have a significant effect on the production of mucus throughout the gastrointestinal tract and could be used to protect the intestinal mucosa from injurious substances that are ingested or following surgery. Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could be used to treat diseases associate with the under expression.

[0689] Moreover, polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could be used to prevent and heal damage to the lungs due to various pathological states. Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, which could stimulate proliferation and differentiation and promote the repair of alveoli and brochiolar epithelium to prevent or treat acute or chronic lung damage. For example, emphysema, which results in the progressive loss of aveoli, and inhalation injuries, i.e., resulting from smoke inhalation and burns, that cause necrosis of the bronchiolar epithelium and alveoli could be effectively treated using polynucleotides or polypeptides, agonists or antagonists of the present invention. Also, polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could be used to stimulate the proliferation of and differentiation of type II pneumocytes, which may help treat or prevent disease such as hyaline membrane diseases, such as infant respiratory distress syndrome and bronchopulmonary displasia, in premature infants.

[0690] Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could stimulate the proliferation and differentiation of hepatocytes and, thus, could be used to alleviate or treat liver diseases and pathologies such as fulminant liver failure caused by cirrhosis, liver damage caused by viral hepatitis and toxic substances (i.e., acetaminophen, carbon tetraholoride and other hepatotoxins known in the art).

[0691] In addition, polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could be used treat or prevent the onset of diabetes mellitus. In patients with newly diagnosed Types I and II diabetes, where some islet cell function remains, polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could be used to maintain the islet function so as to alleviate, delay or prevent permanent manifestation of the disease. Also, polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, could be used as an auxiliary in islet cell transplantation to improve or promote islet cell function.

[0692] Endocrine Disorders

[0693] Polynucleotides or polypeptides, or agonists or antagonists of the present invention, may be used to treat, prevent, diagnose, and/or prognose disorders and/or diseases related to hormone imbalance, and/or disorders or diseases of the endocrine system.

[0694] Hormones secreted by the glands of the endocrine system control physical growth, sexual function, metabolism, and other functions. Disorders may be classified in two ways: disturbances in the production of hormones, and the inability of tissues to respond to hormones. The etiology of these hormone imbalance or endocrine system diseases, disorders or conditions may be genetic, somatic, such as cancer and some autoimmune diseases, acquired (e.g., by chemotherapy, injury or toxins), or infectious. Moreover, polynucleotides, polypeptides, antibodies, and/or agonists or antagonists of the present invention can be used as a marker or detector of a particular disease or disorder related to the endocrine system and/or hormone imbalance.

[0695] Endocrine system and/or hormone imbalance and/or diseases encompass disorders of uterine motility including, but not limited to: complications with pregnancy and labor (e.g., pre-term labor, post-term pregnancy, spontaneous abortion, and slow or stopped labor); and disorders and/or diseases of the menstrual cycle (e.g., dysmenorrhea and endometriosis).

[0696] Endocrine system and/or hormone imbalance disorders and/or diseases include disorders and/or diseases of the pancreas, such as, for example, diabetes mellitus, diabetes insipidus, congenital pancreatic agenesis, pheochromocytoma—islet cell tumor syndrome; disorders and/or diseases of the adrenal glands such as, for example, Addison's Disease, corticosteroid deficiency, virilizing disease, hirsutism, Cushing's Syndrome, hyperaldosteronism, pheochromocytoma; disorders and/or diseases of the pituitary gland, such as, for example, hyperpituitarism, hypopituitarism, pituitary dwarfism, pituitary adenoma, panhypopituitarism, acromegaly, gigantism; disorders and/or diseases of the thyroid, including but not limited to, hyperthyroidism, hypothyroidism, Plummer's disease, Graves' disease (toxic diffuse goiter), toxic nodular goiter, thyroiditis (Hashimoto's thyroiditis, subacute granulomatous thyroiditis, and silent lymphocytic thyroiditis), Pendred's syndrome, myxedema, cretinism, thyrotoxicosis, thyroid hormone coupling defect, thymic aplasia, Hurthle cell tumours of the thyroid, thyroid cancer, thyroid carcinoma, Medullary thyroid carcinoma; disorders and/or diseases of the parathyroid, such as, for example, hyperparathyroidism, hypoparathyroidism; disorders and/or diseases of the hypothalamus.

[0697] In addition, endocrine system and/or hormone imbalance disorders and/or diseases may also include disorders and/or diseases of the testes or ovaries, including cancer. Other disorders and/or diseases of the testes or ovaries further include, for example, ovarian cancer, polycystic ovary syndrome, Klinefelter's syndrome, vanishing testes syndrome (bilateral anorchia), congenital absence of Leydig's cells, cryptorchidism, Noonan's syndrome, myotonic dystrophy, capillary haemangioma of the testis (benign), neoplasias of the testis and neo-testis.

[0698] Moreover, endocrine system and/or hormone imbalance disorders and/or diseases may also include disorders and/or diseases such as, for example, polyglandular deficiency syndromes, pheochromocytoma, neuroblastoma, multiple Endocrine neoplasia, and disorders and/or cancers of endocrine tissues.

[0699] In another embodiment, a polypeptide of the invention, or polynucleotides, antibodies, agonists, or antagonists corresponding to that polypeptide, may be used to diagnose, prognose, prevent, and/or treat endocrine diseases and/or disorders associated with the tissue(s) in which the polypeptide of the invention is expressed, including one, two, three, four, five, or more tissues disclosed in Table 1A, column 8 (Tissue Distribution Library Code).

[0700] Reproductive System Disorders

[0701] The polynucleotides or polypeptides, or agonists or antagonists of the invention may be used for the diagnosis, treatment, or prevention of diseases and/or disorders of the reproductive system. Reproductive system disorders that can be treated by the compositions of the invention, include, but are not limited to, reproductive system injuries, infections, neoplastic disorders, congenital defects, and diseases or disorders which result in infertility, complications with pregnancy, labor, or parturition, and postpartum difficulties.

[0702] Reproductive system disorders and/or diseases include diseases and/or disorders of the testes, including testicular atrophy, testicular feminization, cryptorchism (unilateral and bilateral), anorchia, ectopic testis, epididymitis and orchitis (typically resulting from infections such as, for example, gonorrhea, mumps, tuberculosis, and syphilis), testicular torsion, vasitis nodosa, germ cell tumors (e.g., seminomas, embryonal cell carcinomas, teratocarcinomas, choriocarcinomas, yolk sac tumors, and teratomas), stromal tumors (e.g., Leydig cell tumors), hydrocele, hematocele, varicocele, spermatocele, inguinal hernia, and disorders of sperm production (e.g., immotile cilia syndrome, aspermia, asthenozoospermia, azoospermia, oligospermia, and teratozoospermia).

[0703] Reproductive system disorders also include disorders of the prostate gland, such as acute non-bacterial prostatitis, chronic non-bacterial prostatitis, acute bacterial prostatitis, chronic bacterial prostatitis, prostatodystonia, prostatosis, granulomatous prostatitis, malacoplakia, benign prostatic hypertrophy or hyperplasia, and prostate neoplastic disorders, including adenocarcinomas, transitional cell carcinomas, ductal carcinomas, and squamous cell carcinomas.

[0704] Additionally, the compositions of the invention may be useful in the diagnosis, treatment, and/or prevention of disorders or diseases of the penis and urethra, including inflammatory disorders, such as balanoposthitis, balanitis xerotica obliterans, phimosis, paraphimosis, syphilis, herpes simplex virus, gonorrhea, non-gonococcal urethritis, chlamydia, mycoplasma, trichomonas, HIV, AIDS, Reiter's syndrome, condyloma acuminatum, condyloma latum, and pearly penile papules; urethral abnormalities, such as hypospadias, epispadias, and phimosis; premalignant lesions, including Erythroplasia of Queyrat, Bowen's disease, Bowenoid paplosis, giant condyloma of Buscke-Lowenstein, and varrucous carcinoma; penile cancers, including squamous cell carcinomas, carcinoma in situ, verrucous carcinoma, and disseminated penile carcinoma; urethral neoplastic disorders, including penile urethral carcinoma, bulbomembranous urethral carcinoma, and prostatic urethral carcinoma; and erectile disorders, such as priapism, Peyronie's disease, erectile dysfunction, and impotence.

[0705] Moreover, diseases and/or disorders of the vas deferens include vasculititis and CBAVD (congenital bilateral absence of the vas deferens); additionally, the polynucleotides, polypeptides, and agonists or antagonists of the present invention may be used in the diagnosis, treatment, and/or prevention of diseases and/or disorders of the seminal vesicles, including hydatid disease, congenital chloride diarrhea, and polycystic kidney disease.

[0706] Other disorders and/or diseases of the male reproductive system include, for example, Klinefelter's syndrome, Young's syndrome, premature ejaculation, diabetes mellitus, cystic fibrosis, Kartagener's syndrome, high fever, multiple sclerosis, and gynecomastia.

[0707] Further, the polynucleotides, polypeptides, and agonists or antagonists of the present invention may be used in the diagnosis, treatment, and/or prevention of diseases and/or disorders of the vagina and vulva, including bacterial vaginosis, candida vaginitis, herpes simplex virus, chancroid, granuloma inguinale, lymphogranuloma venereum, scabies, human papillomavirus, vaginal trauma, vulvar trauma, adenosis, chlamydia vaginitis, gonorrhea, trichomonas vaginitis, condyloma acuminatum, syphilis, molluscum contagiosum, atrophic vaginitis, Paget's disease, lichen sclerosus, lichen planus, vulvodynia, toxic shock syndrome, vaginismus, vulvovaginitis, vulvar vestibulitis, and neoplastic disorders, such as squamous cell hyperplasia, clear cell carcinoma, basal cell carcinoma, melanomas, cancer of Bartholin's gland, and vulvar intraepithelial neoplasia.

[0708] Disorders and/or diseases of the uterus include dysmenorrhea, retroverted uterus, endometriosis, fibroids, adenomyosis, anovulatory bleeding, amenorrhea, Cushing's syndrome, hydatidiform moles, Asherman's syndrome, premature menopause, precocious puberty, uterine polyps, dysfunctional uterine bleeding (e.g., due to aberrant hormonal signals), and neoplastic disorders, such as adenocarcinomas, keiomyosarcomas, and sarcomas. Additionally, the polypeptides, polynucleotides, or agonists or antagonists of the invention may be useful as a marker or detector of, as well as in the diagnosis, treatment, and/or prevention of congenital uterine abnormalities, such as bicornuate uterus, septate uterus, simple unicornuate uterus, unicornuate uterus with a noncavitary rudimentary horn, unicornuate uterus with a non-communicating cavitary rudimentary horn, unicornuate uterus with a communicating cavitary horn, arcuate uterus, uterine didelfus, and T-shaped uterus.

[0709] Ovarian diseases and/or disorders include anovulation, polycystic ovary syndrome (Stein-Leventhal syndrome), ovarian cysts, ovarian hypofunction, ovarian insensitivity to gonadotropins, ovarian overproduction of androgens, right ovarian vein syndrome, amenorrhea, hirutism, and ovarian cancer (including, but not limited to, primary and secondary cancerous growth, Sertoli-Leydig tumors, endometriod carcinoma of the ovary, ovarian papillary serous adenocarcinoma, ovarian mucinous adenocarcinoma, and Ovarian Krukenberg tumors).

[0710] Cervical diseases and/or disorders include cervicitis, chronic cervicitis, mucopurulent cervicitis, cervical dysplasia, cervical polyps, Nabothian cysts, cervical erosion, cervical incompetence, and cervical neoplasms (including, for example, cervical carcinoma, squamous metaplasia, squamous cell carcinoma, adenosquamous cell neoplasia, and columnar cell neoplasia).

[0711] Additionally, diseases and/or disorders of the reproductive system include disorders and/or diseases of pregnancy, including miscarriage and stillbirth, such as early abortion, late abortion, spontaneous abortion, induced abortion, therapeutic abortion, threatened abortion, missed abortion, incomplete abortion, complete abortion, habitual abortion, missed abortion, and septic abortion; ectopic pregnancy, anemia, Rh incompatibility, vaginal bleeding during pregnancy, gestational diabetes, intrauterine growth retardation, polyhydramnios, HELLP syndrome, abruptio placentae, placenta previa, hyperemesis, preeclampsia, eclampsia, herpes gestationis, and urticaria of pregnancy. Additionally, the polynucleotides, polypeptides, and agonists or antagonists of the present invention may be used in the diagnosis, treatment, and/or prevention of diseases that can complicate pregnancy, including heart disease, heart failure, rheumatic heart disease, congenital heart disease, mitral valve prolapse, high blood pressure, anemia, kidney disease, infectious disease (e.g., rubella, cytomegalovirus, toxoplasmosis, infectious hepatitis, chlamydia, HIV, AIDS, and genital herpes), diabetes mellitus, Graves' disease, thyroiditis, hypothyroidism, Hashimoto's thyroiditis, chronic active hepatitis, cirrhosis of the liver, primary biliary cirrhosis, asthma, systemic lupus eryematosis, rheumatoid arthritis, myasthenia gravis, idiopathic thrombocytopenic purpura, appendicitis, ovarian cysts, gallbladder disorders, and obstruction of the intestine.

[0712] Complications associated with labor and parturition include premature rupture of the membranes, pre-term labor, post-tern pregnancy, postmaturity, labor that progresses too slowly, fetal distress (e.g., abnormal heart rate (fetal or maternal), breathing problems, and abnormal fetal position), shoulder dystocia, prolapsed umbilical cord, amniotic fluid embolism, and aberrant uterine bleeding.

[0713] Further, diseases and/or disorders of the postdelivery period, including endometritis, myometritis, parametritis, peritonitis, pelvic thrombophlebitis, pulmonary embolism, endotoxemia, pyelonephritis, saphenous thrombophlebitis, mastitis, cystitis, postpartum hemorrhage, and inverted uterus.

[0714] Other disorders and/or diseases of the female reproductive system that may be diagnosed, treated, and/or prevented by the polynucleotides, polypeptides, and agonists or antagonists of the present invention include, for example, Turner's syndrome, pseudohermaphroditism, premenstrual syndrome, pelvic inflammatory disease, pelvic congestion (vascular engorgement), frigidity, anorgasmia, dyspareunia, ruptured fallopian tube, and Mittelschmerz.

[0715] Infectious Disease

[0716] Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention can be used to treat or detect infectious agents. For example, by increasing the immune response, particularly increasing the proliferation and differentiation of B and/or T cells, infectious diseases may be treated. The immune response may be increased by either enhancing an existing immune response, or by initiating a new immune response. Alternatively, polynucleotides or polypeptides, as well as agonists or antagonists of the present invention may also directly inhibit the infectious agent, without necessarily eliciting an immune response.

[0717] Viruses are one example of an infectious agent that can cause disease or symptoms that can be treated or detected by a polynucleotide or polypeptide and/or agonist or antagonist of the present invention. Examples of viruses, include, but are not limited to Examples of viruses, include, but are not limited to the following DNA and RNA viruses and viral families: Arbovirus, Adenoviridae, Arenaviridae, Arterivirus, Birnaviridae, Bunyaviridae, Caliciviridae, Circoviridae, Coronaviridae, Dengue, EBV, HIV, Flaviviridae, Hepadnaviridae (Hepatitis), Herpesviridae (such as, Cytomegalovirus, Herpes Simplex, Herpes Zoster), Mononegavirus (e.g., Paramyxoviridae, Morbillivirus, Rhabdoviridae), Orthomyxoviridae (e.g., Influenza A, Influenza B, and parainfluenza), Papiloma virus, Papovaviridae, Parvoviridae, Picomaviridae, Poxyiridae (such as Smallpox or Vaccinia), Reoviridae (e.g., Rotavirus), Retroviridae (HTLV-I, HTLV-II, Lentivirus), and Togaviridae (e.g., Rubivirus). Viruses falling within these families can cause a variety of diseases or symptoms, including, but not limited to: arthritis, bronchiollitis, respiratory syncytial virus, encephalitis, eye infections (e.g., conjunctivitis, keratitis), chronic fatigue syndrome, hepatitis (A, B, C, E, Chronic Active, Delta), Japanese B encephalitis, Junin, Chikungunya, Rift Valley fever, yellow fever, meningitis, opportunistic infections (e.g., AIDS), pneumonia, Burkitt's Lymphoma, chickenpox, hemorrhagic fever, Measles, Mumps, Parainfluenza, Rabies, the common cold, Polio, leukemia, Rubella, sexually transmitted diseases, skin diseases (e.g., Kaposi's, warts), and viremia. polynucleotides or polypeptides, or agonists or antagonists of the invention, can be used to treat or detect any of these symptoms or diseases. In specific embodiments, polynucleotides, polypeptides, or agonists or antagonists of the invention are used to treat: meningitis, Dengue, EBV, and/or hepatitis (e.g., hepatitis B). In an additional specific embodiment polynucleotides, polypeptides, or agonists or antagonists of the invention are used to treat patients nonresponsive to one or more other commercially available hepatitis vaccines. In a further specific embodiment polynucleotides, polypeptides, or agonists or antagonists of the invention are used to treat AIDS.

[0718] Similarly, bacterial and fungal agents that can cause disease or symptoms and that can be treated or detected by a polynucleotide or polypeptide and/or agonist or antagonist of the present invention include, but not limited to, the following Gram-Negative and Gram-positive bacteria, bacterial families, and fungi: Actinomyces (e.g., Norcardia), Acinetobacter, Cryptococcus neoformans, Aspergillus, Bacillaceae (e.g., Bacillus anthrasis), Bacteroides (e.g., Bacteroides fragilis), Blastomycosis, Bordetella, Borrelia (e.g., Borrelia burgdorferi), Brucella, Candidia, Campylobacter, Chlamydia, Clostridium (e.g., Clostridium botulinum, Clostridium dificile, Clostridium perfringens, Clostridium tetani), Coccidioides, Corynebacterium (e.g., Corynebacterium diptheriae), Cryptococcus, Dermatocycoses, E. coli (e.g., Enterotoxigenic E. coli and Enterohemorrhagic E. coli), Enterobacter (e.g. Enterobacter aerogenes), Enterobacteriaceae (Klebsiella, Salmonella (e.g., Salmonella typhi, Salmonella enteritidis, Salmonella typhi), Serratia, Yersinia, Shigella), Erysipelothrix, Haemophilus (e.g., Haemophilus influenza type B), Helicobacter, Legionella (e.g., Legionella pneumophila), Leptospira, Listeria (e.g., Listeria monocytogenes), Mycoplasma, Mycobacterium (e.g., Mycobacterium leprae and Mycobacterium tuberculosis), Vibrio (e.g., Vibrio cholerae), Neisseriaceae (e.g., Neisseria gonorrhea, Neisseria meningitidis), Pasteurellacea, Proteus, Pseudomonas (e.g., Pseudomonas aeruginosa), Rickettsiaceae, Spirochetes (e.g., Treponema spp., Leptospira spp., Borrelia spp.), Shigella spp., Staphylococcus (e.g., Staphylococcus aureus), Meningiococcus, Pneumococcus and Streptococcus (e.g., Streptococcus pneumoniae and Groups A, B, and C Streptococci), and Ureaplasmas. These bacterial, parasitic, and fungal families can cause diseases or symptoms, including, but not limited to: antibiotic-resistant infections, bacteremia, endocarditis, septicemia, eye infections (e.g., conjunctivitis), uveitis, tuberculosis, gingivitis, bacterial diarrhea, opportunistic infections (e.g., AIDS related infections), paronychia, prosthesis-related infections, dental caries, Reiter's Disease, respiratory tract infections, such as Whooping Cough or Empyema, sepsis, Lyme Disease, Cat-Scratch Disease, dysentery, paratyphoid fever, food poisoning, Legionella disease, chronic and acute inflammation, erythema, yeast infections, typhoid, pneumonia, gonorrhea, meningitis (e.g., mengitis types A and B), chlamydia, syphillis, diphtheria, leprosy, brucellosis, peptic ulcers, anthrax, spontaneous abortions, birth defects, pneumonia, lung infections, ear infections, deafness, blindness, lethargy, malaise, vomiting, chronic diarrhea, Crohn's disease, colitis, vaginosis, sterility, pelvic inflammatory diseases, candidiasis, paratuberculosis, tuberculosis, lupus, botulism, gangrene, tetanus, impetigo, Rheumatic Fever, Scarlet Fever, sexually transmitted diseases, skin diseases (e.g., cellulitis, dermatocycoses), toxemia, urinary tract infections, wound infections, noscomial infections. Polynucleotides or polypeptides, agonists or antagonists of the invention, can be used to treat or detect any of these symptoms or diseases. In specific embodiments, polynucleotides, polypeptides, agonists or antagonists of the invention are used to treat: tetanus, diptheria, botulism, and/or meningitis type B.

[0719] Moreover, parasitic agents causing disease or symptoms that can be treated, prevented, and/or diagnosed by a polynucleotide or polypeptide and/or agonist or antagonist of the present invention include, but not limited to, the following families or class: Amebiasis, Babesiosis, Coccidiosis, Cryptosporidiosis, Dientamoebiasis, Dourine, Ectoparasitic, Giardias, Helminthiasis, Leishmaniasis, Schistisoma, Theileriasis, Toxoplasmosis, Trypanosomiasis, and Trichomonas and Sporozoans (e.g., Plasmodium virax, Plasmodium falciparium, Plasmodium malariae and Plasmodium ovale). These parasites can cause a variety of diseases or symptoms, including, but not limited to: Scabies, Trombiculiasis, eye infections, intestinal disease (e.g., dyseritery, giardiasis), liver disease, lung disease, opportunistic infections (e.g., AIDS related), malaria, pregnancy complications, and toxoplasmosis. polynucleotides or polypeptides, or agonists or antagonists of the invention, can be used to treat, prevent, and/or diagnose any of these symptoms or diseases. In specific embodiments, polynucleotides, polypeptides, or agonists or antagonists of the invention are used to treat, prevent, and/or diagnose malaria.

[0720] Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention of the present invention could either be by administering an effective amount of a polypeptide to the patient, or by removing cells from the patient, supplying the cells with a polynucleotide of the present invention, and returning the engineered cells to the patient (ex vivo therapy). Moreover, the polypeptide or polynucleotide of the present invention can be used as an antigen in a vaccine to raise an immune response against infectious disease.

[0721] Regeneration

[0722] Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention can be used to differentiate, proliferate, and attract cells, leading to the regeneration of tissues. (See, Science 276:59-87 (1997)). The regeneration of tissues could be used to repair, replace, or protect tissue damaged by congenital defects, trauma (wounds, bums, incisions, or ulcers), age, disease (e.g. osteoporosis, osteocarthritis, periodontal disease, liver failure), surgery, including cosmetic plastic surgery, fibrosis, reperfusion injury, or systemic cytokine damage.

[0723] Tissues that could be regenerated using the present invention include organs (e.g., pancreas, liver, intestine, kidney, skin, endothelium), muscle (smooth, skeletal or cardiac), vasculature (including vascular and lymphatics), nervous, hematopoietic, and skeletal (bone, cartilage, tendon, and ligament) tissue. Preferably, regeneration occurs without or decreased scarring. Regeneration also may include angiogenesis.

[0724] Moreover, polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, may increase regeneration of tissues difficult to heal. For example, increased tendon/ligament regeneration would quicken recovery time after damage. Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention could also be used prophylactically in an effort to avoid damage. Specific diseases that could be treated include of tendinitis, carpal tunnel syndrome, and other tendon or ligament defects. A further example of tissue regeneration of non-healing wounds includes pressure ulcers, ulcers associated with vascular insufficiency, surgical, and traumatic wounds.

[0725] Similarly, nerve and brain tissue could also be regenerated by using polynucleotides or polypeptides, as well as agonists or antagonists of the present invention, to proliferate and differentiate nerve cells. Diseases that could be treated using this method include central and peripheral nervous system diseases, neuropathies, or mechanical and traumatic disorders (e.g., spinal cord disorders, head trauma, cerebrovascular disease, and stoke). Specifically, diseases associated with peripheral nerve injuries, peripheral neuropathy (e.g., resulting from chemotherapy or other medical therapies), localized neuropathies, and central nervous system diseases (e.g., Alzheimer's disease, Parkinson's disease, Huntington's disease, amyotrophic lateral sclerosis, and Shy-Drager syndrome), could all be treated using the polynucleotides or polypeptides, as well as agonists or antagonists of the present invention.

[0726] Gastrointestinal Disorders

[0727] Polynucleotides or polypeptides, or agonists or antagonists of the present invention, may be used to treat, prevent, diagnose, and/or prognose gastrointestinal disorders, including inflammatory diseases and/or conditions, infections, cancers (e.g., intestinal neoplasms (carcinoid tumor of the small intestine, non-Hodgkin's lymphoma of the small intestine, small bowl lymphoma)), and ulcers, such as peptic ulcers.

[0728] Gastrointestinal disorders include dysphagia, odynophagia, inflammation of the esophagus, peptic esophagitis, gastric reflux, submucosal fibrosis and stricturing, Mallory-Weiss lesions, leiomyomas, lipomas, epidermal cancers, adeoncarcinomas, gastric retention disorders, gastroenteritis, gastric atrophy, gastric/stomach cancers, polyps of the stomach, autoimmune disorders such as pernicious anemia, pyloric stenosis, gastritis (bacterial, viral, eosinophilic, stress-induced, chronic erosive, atrophic, plasma cell, and Ménétrier's), and peritoneal diseases (e.g., chyloperioneum, hemoperitoneum, mesenteric cyst, mesenteric lymphadenitis, mesenteric vascular occlusion, panniculitis, neoplasms, peritonitis, pneumoperitoneum, bubphrenic abscess,).

[0729] Gastrointestinal disorders also include disorders associated with the small intestine, such as malabsorption syndromes, distension, irritable bowel syndrome, sugar intolerance, celiac disease, duodenal ulcers, duodenitis, tropical sprue, Whipple's disease, intestinal lymphangiectasia, Crohn's disease, appendicitis, obstructions of the ileum, Meckel's diverticulum, multiple diverticula, failure of complete rotation of the small and large intestine, lymphoma, and bacterial and parasitic diseases (such as Traveler's diarrhea, typhoid and paratyphoid, cholera, infection by Roundworms (Ascariasis lumbricoides), Hookworms (Ancylostoma duodenale), Threadworms (Enterobius vermicularis), Tapeworms (Taenia saginata, Echinococcus granulosus, Diphyllobothrium spp., and T. solium).

[0730] Liver diseases and/or disorders include intrahepatic cholestasis (alagille syndrome, biliary liver cirrhosis), fatty liver (alcoholic fatty liver, reye syndrome), hepatic vein thrombosis, hepatolentricular degeneration, hepatomegaly, hepatopulmonary syndrome, hepatorenal syndrome, portal hypertension (esophageal and gastric varices), liver abscess (amebic liver abscess), liver cirrhosis (alcoholic, biliary and experimental), alcoholic liver diseases (fatty liver, hepatitis, cirrhosis), parasitic (hepatic echinococcosis, fascioliasis, amebic liver abscess), jaundice (hemolytic, hepatocellular, and cholestatic), cholestasis, portal hypertension, liver enlargement, ascites, hepatitis (alcoholic hepatitis, animal hepatitis, chronic hepatitis (autoimmune, hepatitis B, hepatitis C, hepatitis D, drug induced), toxic hepatitis, viral human hepatitis (hepatitis A, hepatitis B, hepatitis C, hepatitis D, hepatitis E), Wilson's disease, granulomatous hepatitis, secondary biliary cirrhosis, hepatic encephalopathy, portal hypertension, varices, hepatic encephalopathy, primary biliary cirrhosis, primary sclerosing cholangitis, hepatocellular adenoma, hemangiomas, bile stones, liver failure (hepatic encephalopathy, acute liver failure), and liver neoplasms (angiomyolipoma, calcified liver metastases, cystic liver metastases, epithelial tumors, fibrolamellar hepatocarcinoma, focal nodular hyperplasia, hepatic adenoma, hepatobiliary cystadenoma, hepatoblastoma, hepatocellular carcinoma, hepatoma, liver cancer, liver hemangioendothelioma, mesenchymal hamartoma, mesenchymal tumors of liver, nodular regenerative hyperplasia, benign liver tumors (Hepatic cysts [Simple cysts, Polycystic liver disease, Hepatobiliary cystadenoma, Choledochal cyst], Mesenchymal tumors [Mesenchymal hamartoma, Infantile hemangioendothelioma, Hemangioma, Peliosis hepatis, Lipomas, Inflammatory pseudotumor, Miscellaneous], Epithelial tumors [Bile duct epithelium (Bile duct hamartoma, Bile duct adenoma), Hepatocyte (Adenoma, Focal nodular hyperplasia, Nodular regenerative hyperplasia)], malignant liver tumors [hepatocellular, hepatoblastoma, hepatocellular carcinoma, cholangiocellular, cholangiocarcinoma, cystadenocarcinoma, tumors of blood vessels, angiosarcoma, Karposi's sarcoma, hemangioendothelioma, other tumors, embryonal sarcoma, fibrosarcoma, leiomyosarcoma, rhabdomyosarcoma, carcinosarcoma, teratoma, carcinoid, squamous carcinoma, primary lymphoma]), peliosis hepatis, erythrohepatic porphyria, hepatic porphyria (acute intermittent porphyria, porphyria cutanea tarda), Zellweger syndrome).

[0731] Pancreatic diseases and/or disorders include acute pancreatitis, chronic pancreatitis (acute necrotizing pancreatitis, alcoholic pancreatitis), neoplasms (adenocarcinoma of the pancreas, cystadenocarcinoma, insulinoma, gastrinoma, and glucagonoma, cystic neoplasms, islet-cell tumors, pancreoblastoma), and other pancreatic diseases (e.g., cystic fibrosis, cyst (pancreatic pseudocyst, pancreatic fistula, insufficiency)).

[0732] Gallbladder diseases include gallstones (cholelithiasis and choledocholithiasis), postcholecystectomy syndrome, diverticulosis of the gallbladder, acute cholecystitis, chronic cholecystitis, bile duct tumors, and mucocele.

[0733] Diseases and/or disorders of the large intestine include antibiotic-associated colitis, diverticulitis, ulcerative colitis, acquired megacolon, abscesses, fungal and bacterial infections, anorectal disorders (e.g., fissures, hemorrhoids), colonic diseases (colitis, colonic neoplasms [colon cancer, adenomatous colon polyps (e.g., villous adenoma), colon carcinoma, colorectal cancer], colonic diverticulitis, colonic diverticulosis, megacolon [Hirschsprung disease, toxic megacolon]; sigmoid diseases [proctocolitis, sigmoin neoplasms]), constipation, Crohn's disease, diarrhea (infantile diarrhea, dysentery), duodenal diseases (duodenal neoplasms, duodenal obstruction, duodenal ulcer, duodenitis), enteritis (enterocolitis), HIV enteropathy, ileal diseases (ileal neoplasms, ileitis), immunoproliferative small intestinal disease, inflammatory bowel disease (ulcerative colitis, Crohn's disease), intestinal atresia, parasitic diseases (anisakiasis, balantidiasis, blastocystis infections, cryptosporidiosis, dientamoebiasis, amebic dysentery, giardiasis), intestinal fistula (rectal fistula), intestinal neoplasms (cecal neoplasms, colonic neoplasms, duodenal neoplasms, ileal neoplasms, intestinal polyps, jejunal neoplasms, rectal neoplasms), intestinal obstruction (afferent loop syndrome, duodenal obstruction, impacted feces, intestinal pseudo-obstruction [cecal volvulus], intussusception), intestinal perforation, intestinal polyps (colonic polyps, gardner syndrome, peutz-jeghers syndrome), jejunal diseases (jejunal neoplasms), malabsorption syndromes (blind loop syndrome, celiac disease, lactose intolerance, short bowl syndrome, tropical sprue, whipple's disease), mesenteric vascular occlusion, pneumatosis cystoides intestinalis, protein-losing enteropathies (intestinal lymphagiectasis), rectal diseases (anus diseases, fecal incontinence, hemorrhoids, proctitis, rectal fistula, rectal prolapse, rectocele), peptic ulcer (duodenal ulcer, peptic esophagitis, hemorrhage, perforation, stomach ulcer, Zollinger-Ellison syndrome), postgastrectomy syndromes (dumping syndrome), stomach diseases (e.g., achlorhydria, duodenogastric reflux (bile reflux), gastric antral vascular ectasia, gastric fistula, gastric outlet obstruction, gastritis (atrophic or hypertrophic), gastroparesis, stomach dilatation, stomach diverticulum, stomach neoplasms (gastric cancer, gastric polyps, gastric adenocarcinoma, hyperplastic gastric polyp), stomach rupture, stomach ulcer, stomach volvulus), tuberculosis, visceroptosis, vomiting (e.g., hematemesis, hyperemesis gravidarum, postoperative nausea and vomiting) and hemorrhagic colitis.

[0734] Further diseases and/or disorders of the gastrointestinal system include biliary tract diseases, such as, gastroschisis, fistula (e.g., biliary fistula, esophageal fistula, gastric fistula, intestinal fistula, pancreatic fistula), neoplasms (e.g., biliary tract neoplasms, esophageal neoplasms, such as adenocarcinoma of the esophagus, esophageal squamous cell carcinoma, gastrointestinal neoplasms, pancreatic neoplasms, such as adenocarcinoma of the pancreas, mucinous cystic neoplasm of the pancreas, pancreatic cystic neoplasms, pancreatoblastoma, and peritoneal neoplasms), esophageal disease (e.g., bullous diseases, candidiasis, glycogenic acanthosis, ulceration, barrett esophagus varices, atresia, cyst, diverticulum (e.g., Zenker's diverticulum), fistula (e.g., tracheoesophageal fistula), motility disorders (e.g., CREST syndrome, deglutition disorders, achalasia, spasm, gastroesophageal reflux), neoplasms, perforation (e.g., Boerhaave syndrome, Mallory-Weiss syndrome), stenosis, esophagitis, diaphragmatic hernia (e.g., hiatal hernia); gastrointestinal diseases, such as, gastroenteritis (e.g., cholera morbus, norwalk virus infection), hemorrhage (e.g., hematemesis, melena, peptic ulcer hemorrhage), stomach neoplasms (gastric cancer, gastric polyps, gastric adenocarcinoma, stomach cancer)), hernia (e.g., congenital diaphragmatic hernia, femoral hernia, inguinal hernia, obturator hernia, umbilical hernia, ventral hernia), and intestinal diseases (e.g., cecal diseases (appendicitis, cecal neoplasms)).

[0735] Chemotaxis

[0736] Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention may have chemotaxis activity. A chemotaxic molecule attracts or mobilizes cells (e.g., monocytes, fibroblasts, neutrophils, T-cells, mast cells, eosinophils, epithelial and/or endothelial cells) to a particular site in the body, such as inflammation, infection, or site of hyperproliferation. The mobilized cells can then fight off and/or heal the particular trauma or abnormality.

[0737] Polynucleotides or polypeptides, as well as agonists or antagonists of the present invention may increase chemotaxic activity of particular cells. These chemotactic molecules can then be used to treat inflammation, infection, hyperproliferative disorders, or any immune system disorder by increasing the number of cells targeted to a particular location in the body. For example, chemotaxic molecules can be used to treat wounds and other trauma to tissues by attracting immune cells to the injured location. Chemotactic molecules of the present invention can also attract fibroblasts, which can be used to treat wounds.

[0738] It is also contemplated that polynucleotides or polypeptides, as well as agonists or antagonists of the present invention may inhibit chemotactic activity. These molecules could also be used to treat disorders. Thus, polynucleotides or polypeptides, as well as agonists or antagonists of the present invention could be used as an inhibitor of chemotaxis.

[0739] Binding Activity

[0740] A polypeptide of the present invention may be used to screen for molecules that bind to the polypeptide or for molecules to which the polypeptide binds. The binding of the polypeptide and the molecule may activate (agonist), increase, inhibit (antagonist), or decrease activity of the polypeptide or the molecule bound. Examples of such molecules include antibodies, oligonucleotides, proteins (e.g., receptors), or small molecules.

[0741] Preferably, the molecule is closely related to the natural ligand of the polypeptide, e.g., a fragment of the ligand, or a natural substrate, a ligand, a structural or functional mimetic. (See, Coligan et al., Current Protocols in Immunology 1(2):Chapter 5 (1991)). Similarly, the molecule can be closely related to the natural receptor to which the polypeptide binds, or at least, a fragment of the receptor capable of being bound by the polypeptide (e.g., active site). In either case, the molecule can be rationally designed using known techniques.

[0742] Preferably, the screening for these molecules involves producing appropriate cells which express the polypeptide. Preferred cells include cells from mammals, yeast, Drosophila, or E. Coli. Cells expressing the polypeptide (or cell membrane containing the expressed polypeptide) are then preferably contacted with a test compound potentially containing the molecule to observe binding, stimulation, or inhibition of activity of either the polypeptide or the molecule.

[0743] The assay may simply test binding of a candidate compound to the polypeptide, wherein binding is detected by a label, or in an assay involving competition with a labeled competitor. Further, the assay may test whether the candidate compound results in a signal generated by binding to the polypeptide.

[0744] Alternatively, the assay can be carried out using cell-free preparations, polypeptide/molecule affixed to a solid support, chemical libraries, or natural product mixtures. The assay may also simply comprise the steps of mixing a candidate compound with a solution containing a polypeptide, measuring polypeptide/molecule activity or binding, and comparing the polypeptide/molecule activity or binding to a standard.

[0745] Preferably, an ELISA assay can measure polypeptide level or activity in a sample (e.g., biological sample) using a monoclonal or polyclonal antibody. The antibody can measure polypeptide level or activity by either binding, directly or indirectly, to the polypeptide or by competing with the polypeptide for a substrate.

[0746] Additionally, the receptor to which the polypeptide of the present invention binds can be identified by numerous methods known to those of skill in the art, for example, ligand panning and FACS sorting (Coligan, et al., Current Protocols in Immun., 1(2), Chapter 5, (1991)). For example, expression cloning is employed wherein polyadenylated RNA is prepared from a cell responsive to the polypeptides, for example, NIH3T3 cells which are known to contain multiple receptors for the FGF family proteins, and SC-3 cells, and a cDNA library created from this RNA is divided into pools and used to transfect COS cells or other cells that are not responsive to the polypeptides. Transfected cells which are grown on glass slides are exposed to the polypeptide of the present invention, after they have been labeled. The polypeptides can be labeled by a variety of means including iodination or inclusion of a recognition site for a site-specific protein kinase.

[0747] Following fixation and incubation, the slides are subjected to auto-radiographic analysis. Positive pools are identified and sub-pools are prepared and re-transfected using an iterative sub-pooling and re-screening process, eventually yielding a single clones that encodes the putative receptor.

[0748] As an alternative approach for receptor identification, the labeled polypeptides can be photoaffinity linked with cell membrane or extract preparations that express the receptor molecule. Cross-linked material is resolved by PAGE analysis and exposed to X-ray film. The labeled complex containing the receptors of the polypeptides can be excised, resolved into peptide fragments, and subjected to protein microsequencing. The amino acid sequence obtained from microsequencing would be used to design a set of degenerate oligonucleotide probes to screen a cDNA library to identify the genes encoding the putative receptors.

[0749] Moreover, the techniques of gene-shuffling, motif-shuffling, exon-shuffling, and/or codon-shuffling (collectively referred to as “DNA shuffling”) may be employed to modulate the activities of the polypeptide of the present invention thereby effectively generating agonists and antagonists of the polypeptide of the present invention. See generally, U.S. Pat. Nos. 5,605,793, 5,811,238, 5,830,721, 5,834,252, and 5,837,458, and Patten, P. A., et al., Curr. Opinion Biotechnol. 8:724-33 (1997); Harayama, S. Trends Biotechnol. 16(2):76-82 (1998); Hansson, L. O., et al., J. Mol. Biol. 287:265-76 (1999); and Lorenzo, M. M. and Blasco, R. Biotechniques 24(2):308-13 (1998); each of these patents and publications are hereby incorporated by reference). In one embodiment, alteration of polynucleotides and corresponding polypeptides may be achieved by DNA shuffling. DNA shuffling involves the assembly of two or more DNA segments into a desired molecule by homologous, or site-specific, recombination. In another embodiment, polynucleotides and corresponding polypeptides may be altered by being subjected to random mutagenesis by error-prone PCR, random nucleotide insertion or other methods prior to recombination. In another embodiment, one or more components, motifs, sections, parts, domains, fragments, etc., of the polypeptide of the present invention may be recombined with one or more components, motifs, sections, parts, domains, fragments, etc. of one or more heterologous molecules. In preferred embodiments, the heterologous molecules are family members. In further preferred embodiments, the heterologous molecule is a growth factor such as, for example, platelet-derived growth factor (PDGF), insulin-like growth factor (IGF-I), transforming growth factor (TGF)-alpha, epidermal growth factor (EGF), fibroblast growth factor (FGF), TGF-beta, bone morphogenetic protein (BMP)-2, BMP-4, BMP-5, BMP-6, BMP-7, activins A and B, decapentaplegic(dpp), 60A, OP-2, dorsalin, growth differentiation factors (GDFs), nodal, MIS, inhibin-alpha, TGF-beta1, TGF-beta2, TGF-beta3, TGF-beta5, and glial-derived neurotrophic factor (GDNF).

[0750] Other preferred fragments are biologically active fragments of the polypeptide of the present invention. Biologically active fragments are those exhibiting activity similar, but not necessarily identical, to an activity of the polypeptide of the present invention. The biological activity of the fragments may include an improved desired activity, or a decreased undesirable activity.

[0751] Additionally, this invention provides a method of screening compounds to identify those which modulate the action of the polypeptide of the present invention. An example of such an assay comprises combining a mammalian fibroblast cell, a the polypeptide of the present invention, the compound to be screened and 3[H] thymidine under cell culture conditions where the fibroblast cell would normally proliferate. A control assay may be performed in the absence of the compound to be screened and compared to the amount of fibroblast proliferation in the presence of the compound to determine if the compound stimulates proliferation by determining the uptake of 3[H] thymidine in each case. The amount of fibroblast cell proliferation is measured by liquid scintillation chromatography which measures the incorporation of 3[H] thymidine. Both agonist and antagonist compounds may be identified by this procedure.

[0752] In another method, a mammalian cell or membrane preparation expressing a receptor for a polypeptide of the present invention is incubated with a labeled polypeptide of the present invention in the presence of the compound. The ability of the compound to enhance or block this interaction could then be measured. Alternatively, the response of a known second messenger system following interaction of a compound to be screened and the receptor is measured and the ability of the compound to bind to the receptor and elicit a second messenger response is measured to determine if the compound is a potential agonist or antagonist. Such second messenger systems include but are not limited to, cAMP guanylate cyclase, ion channels or phosphoinositide hydrolysis.

[0753] All of these above assays can be used as diagnostic or prognostic markers. The molecules discovered using these assays can be used to treat disease or to bring about a particular result in a patient (e.g., blood vessel growth) by activating or inhibiting the polypeptide/molecule. Moreover, the assays can discover agents which may inhibit or enhance the production of the polypeptides of the invention from suitably manipulated cells or tissues.

[0754] Therefore, the invention includes a method of identifying compounds which bind to a polypeptide of the invention comprising the steps of: (a) incubating a candidate binding compound with a polypeptide of the present invention; and (b) determining if binding has occurred. Moreover, the invention includes a method of identifying agonists/antagonists comprising the steps of: (a) incubating a candidate compound with a polypeptide of the present invention, (b) assaying a biological activity, and (b) determining if a biological activity of the polypeptide has been altered.

[0755] Targeted Delivery

[0756] In another embodiment, the invention provides a method of delivering compositions to targeted cells expressing a receptor for a polypeptide of the invention, or cells expressing a cell bound form of a polypeptide of the invention.

[0757] As discussed herein, polypeptides or antibodies of the invention may be associated with heterologous polypeptides, heterologous nucleic acids, toxins, or prodrugs via hydrophobic, hydrophilic, ionic and/or covalent interactions. In one embodiment, the invention provides a method for the specific delivery of compositions of the invention to cells by administering polypeptides of the invention (including antibodies) that are associated with heterologous polypeptides or nucleic acids. In one example, the invention provides a method for delivering a therapeutic protein into the targeted cell. In another example, the invention provides a method for delivering a single stranded nucleic acid (e.g., antisense or ribozymes) or double stranded nucleic acid (e.g., DNA that can integrate into the cell's genome or replicate episomally and that can be transcribed) into the targeted cell.

[0758] In another embodiment, the invention provides a method for the specific destruction of cells (e.g., the destruction of tumor cells) by administering polypeptides of the invention (e.g., polypeptides of the invention or antibodies of the invention) in association with toxins or cytotoxic prodrugs.

[0759] By “toxin” is meant compounds that bind and activate endogenous cytotoxic effector systems, radioisotopes, holotoxins, modified toxins, catalytic subunits of toxins, or any molecules or enzymes not normally present in or on the surface of a cell that under defined conditions cause the cell's death. Toxins that may be used according to the methods of the invention include, but are not limited to, radioisotopes known in the art, compounds such as, for example, antibodies (or complement fixing containing portions thereof) that bind an inherent or induced endogenous cytotoxic effector system, thymidine kinase, endonuclease, RNAse, alpha toxin, ricin, abrin, Pseudomonas exotoxin A, diphtheria toxin, saporin, momordin, gelonin, pokeweed antiviral protein, alpha-sarcin and cholera toxin. By “cytotoxic prodrug” is meant a non-toxic compound that is converted by an enzyme, normally present in the cell, into a cytotoxic compound. Cytotoxic prodrugs that may be used according to the methods of the invention include, but are not limited to, glutamyl derivatives of benzoic acid mustard alkylating agent, phosphate derivatives of etoposide or mitomycin C, cytosine arabinoside, daunorubisin, and phenoxyacetamide derivatives of doxorubicin.

[0760] Drug Screening

[0761] Further contemplated is the use of the polypeptides of the present invention, or the polynucleotides encoding these polypeptides, to screen for molecules which modify the activities of the polypeptides of the present invention. Such a method would include contacting the polypeptide of the present invention with a selected compound(s) suspected of having antagonist or agonist activity, and assaying the activity of these polypeptides following binding.

[0762] This invention is particularly useful for screening therapeutic compounds by using the polypeptides of the present invention, or binding fragments thereof, in any of a variety of drug screening techniques. The polypeptide or fragment employed in such a test may be affixed to a solid support, expressed on a cell surface, free in solution, or located intracellularly. One method of drug screening utilizes eukaryotic or prokaryotic host cells which are stably transformed with recombinant nucleic acids expressing the polypeptide or fragment. Drugs are screened against such transformed cells in competitive binding assays. One may measure, for example, the formulation of complexes between the agent being tested and a polypeptide of the present invention.

[0763] Thus, the present invention provides methods of screening for drugs or any other agents which affect activities mediated by the polypeptides of the present invention. These methods comprise contacting such an agent with a polypeptide of the present invention or a fragment thereof and assaying for the presence of a complex between the agent and the polypeptide or a fragment thereof, by methods well known in the art. In such a competitive binding assay, the agents to screen are typically labeled. Following incubation, free agent is separated from that present in bound form, and the amount of free or uncomplexed label is a measure of the ability of a particular agent to bind to the polypeptides of the present invention.

[0764] Another technique for drug screening provides high throughput screening for compounds having suitable binding affinity to the polypeptides of the present invention, and is described in great detail in European Patent Application 84/03564, published on Sep. 13, 1984, which is incorporated herein by reference herein. Briefly stated, large numbers of different small peptide test compounds are synthesized on a solid substrate, such as plastic pins or some other surface. The peptide test compounds are reacted with polypeptides of the present invention and washed. Bound polypeptides are then detected by methods well known in the art. Purified polypeptides are coated directly onto plates for use in the aforementioned drug screening techniques. In addition, non-neutralizing antibodies may be used to capture the peptide and immobilize it on the solid support.

[0765] This invention also contemplates the use of competitive drug screening assays in which neutralizing antibodies capable of binding polypeptides of the present invention specifically compete with a test compound for binding to the polypeptides or fragments thereof. In this manner, the antibodies are used to detect the presence of any peptide which shares one or more antigenic epitopes with a polypeptide of the invention.

[0766] Antisense and Ribozyme (Antagonists)

[0767] In specific embodiments, antagonists according to the present invention are nucleic acids corresponding to the sequences contained in SEQ ID NO:X, or the complementary strand thereof, and/or to cDNA sequences contained in cDNA Clone ID NO:Z identified for example, in Table 1A. In one embodiment, antisense sequence is generated internally, by the organism, in another embodiment, the antisense sequence is separately administered (see, for example, O'Connor, J., Neurochem. 56:560 (1991). Oligodeoxynucleotides as Antisense Inhibitors of Gene Expression, CRC Press, Boca Raton, Fla. (1988). Antisense technology can be used to control gene expression through antisense DNA or RNA, or through triple-helix formation. Antisense techniques are discussed for example, in Okano, J., Neurochem. 56:560 (1991); Oligodeoxynucleotides as Antisense Inhibitors of Gene Expression, CRC Press, Boca Raton, Fla. (1988). Triple helix formation is discussed in, for instance, Lee et al., Nucleic Acids Research 6:3073 (1979); Cooney et al., Science 241:456 (1988); and Dervan et al., Science 251:1300 (1991). The methods are based on binding of a polynucleotide to a complementary DNA or RNA.

[0768] For example, the use of c-myc and c-myb antisense RNA constructs to inhibit the growth of the non-lymphocytic leukemia cell line HL-60 and other cell lines was previously described. (Wickstrom et al. (1988); Anfossi et al. (1989)). These experiments were performed in vitro by incubating cells with the oligoribonucleotide. A similar procedure for in vivo use is described in WO 91/15580. Briefly, a pair of oligonucleotides for a given antisense RNA is produced as follows: A sequence complimentary to the first 15 bases of the open reading frame is flanked by an EcoRI site on the 5 end and a HindIII site on the 3 end. Next, the pair of oligonucleotides is heated at 90° C. for one minute and then annealed in 2× ligation buffer (20 mM TRIS HCl pH 7.5, 10 mM MgCl2, 10MM dithiothreitol (DTT) and 0.2 mM ATP) and then ligated to the EcoRI/Hind III site of the retroviral vector PMV7 (WO 91/15580).

[0769] For example, the 5′ coding portion of a polynucleotide that encodes the polypeptide of the present invention may be used to design an antisense RNA oligonucleotide of from about 10 to 40 base pairs in length. A DNA oligonucleotide is designed to be complementary to a region of the gene involved in transcription thereby preventing transcription and the production of the receptor. The antisense RNA oligonucleotide hybridizes to the mRNA in vivo and blocks translation of the mRNA molecule into receptor polypeptide.

[0770] In one embodiment, the antisense nucleic acid of the invention is produced intracellularly by transcription from an exogenous sequence. For example, a vector or a portion thereof, is transcribed, producing an antisense nucleic acid (RNA) of the invention. Such a vector would contain a sequence encoding the antisense nucleic acid. Such a vector can remain episomal or become chromosomally integrated, as long as it can be transcribed to produce the desired antisense RNA. Such vectors can be constructed by recombinant DNA technology methods standard in the art. Vectors can be plasmid, viral, or others known in the art, used for replication and expression in vertebrate cells. Expression of the sequence encoding the polypeptide of the present invention or fragments thereof, can be by any promoter known in the art to act in vertebrate, preferably human cells. Such promoters can be inducible or constitutive. Such promoters include, but are not limited to, the SV40 early promoter region (Bernoist and Chambon, Nature 29:304-310 (1981), the promoter contained in the 3′ long terminal repeat of Rous sarcoma virus (Yamamoto et al., Cell 22:787-797 (1980), the herpes thymidine promoter (Wagner et al., Proc. Natl. Acad. Sci. U.S.A. 78:1441-1445 (1981), the regulatory sequences of the metallothionein gene (Brinster, et al., Nature 296:39-42 (1982)), etc.

[0771] The antisense nucleic acids of the invention comprise a sequence complementary to at least a portion of an RNA transcript of a gene of the present invention. However, absolute complementarity, although preferred, is not required. A sequence “complementary to at least a portion of an RNA,” referred to herein, means a sequence having sufficient complementarity to be able to hybridize with the RNA, forming a stable duplex; in the case of double stranded antisense nucleic acids, a single strand of the duplex DNA may thus be tested, or triplex formation may be assayed. The ability to hybridize will depend on both the degree of complementarity and the length of the antisense nucleic acid. Generally, the larger the hybridizing nucleic acid, the more base mismatches with a RNA it may contain and still form a stable duplex (or triplex as the case may be). One skilled in the art can ascertain a tolerable degree of mismatch by use of standard procedures to determine the melting point of the hybridized complex.

[0772] Oligonucleotides that are complementary to the 5′ end of the message, e.g., the 5′ untranslated sequence up to and including the AUG initiation codon, should work most efficiently at inhibiting translation. However, sequences complementary to the 3′ untranslated sequences of mRNAs have been shown to be effective at inhibiting translation of mRNAs as well. See generally, Wagner, R., 1994, Nature 372:333-335. Thus, oligonucleotides complementary to either the 5′- or 3′-non-translated, non-coding regions of polynucleotide sequences described herein could be used in an antisense approach to inhibit translation of endogenous mRNA. Oligonucleotides complementary to the 5′ untranslated region of the mRNA should include the complement of the AUG start codon. Antisense oligonucleotides complementary to mRNA coding regions are less efficient inhibitors of translation but could be used in accordance with the invention. Whether designed to hybridize to the 5′-, 3′- or coding region of mRNA of the present invention, antisense nucleic acids should be at least six nucleotides in length, and are preferably oligonucleotides ranging from 6 to about 50 nucleotides in length. In specific aspects the oligonucleotide is at least 10 nucleotides, at least 17 nucleotides, at least 25 nucleotides or at least 50 nucleotides.

[0773] The polynucleotides of the invention can be DNA or RNA or chimeric mixtures or derivatives or modified versions thereof, single-stranded or double-stranded. The oligonucleotide can be modified at the base moiety, sugar moiety, or phosphate backbone, for example, to improve stability of the molecule, hybridization, etc. The oligonucleotide may include other appended groups such as peptides (e.g., for targeting host cell receptors i[vivo), or agents facilitating transport across the cell membrane (see, e.g., Letsinger et al., 1989, Proc. Natl. Acad. Sci. U.S.A. 86:6553-6556; Lemaitre et al., 1987, Proc. Natl. Acad. Sci. 84:648-652; PCT Publication No. WO88/09810, published Dec. 15, 1988) or the blood-brain barrier (see, e.g., PCT Publication No. WO89/10134, published Apr. 25, 1988), hybridization-triggered cleavage agents. (See, e.g., Krol et al., 1988, BioTechniques 6:958-976) or intercalating agents. (See, e.g., Zon, 1988, Pharm. Res. 5:539-549). To this end, the oligonucleotide may be conjugated to another molecule, e.g., a peptide, hybridization triggered cross-linking agent, transport agent, hybridization-triggered cleavage agent, etc.

[0774] The antisense oligonucleotide may comprise at least one modified base moiety which is selected from the group including, but not limited to, 5-fluorouracil, 5-bromouracil, 5-chlorouracil, 5-iodouracil, hypoxanthine, xantine, 4-acetylcytosine, 5-(carboxyhydroxylmethyl) uracil, 5-carboxymethylaminomethyl-2-thiouridine, 5-carboxymethylaminomethyluracil, dihydrouracil, beta-D-galactosylqueosine, inosine, N6-isopentenyladenine, 1-methylguanine, 1-methylinosine, 2,2-dimethylguanine, 2-methyladenine, 2-methylguanine, 3-methylcytosine, 5-methylcytosine, N6-adenine, 7-methylguanine, 5-methylaminomethyluracil, 5-methoxyaminomethyl-2-thiouracil, beta-D-mannosylqueosine, 5′-methoxycarboxymethyluracil, 5-methoxyuracil, 2-methylthio-N-6-isopentenyladenine, uracil-5-oxyacetic acid (v), wybutoxosine, pseudouracil, queosine, 2-thiocytosine, 5-methyl-2-thiouracil, 2-thiouracil, 4-thiouracil, 5-methyluracil, uracil-5-oxyacetic acid methylester, uracil-5-oxyacetic acid (v), 5-methyl-2-thiouracil, 3-(3-amino-3-N-2-carboxypropyl) uracil, (acp3)w, and 2,6-diaminopurine.

[0775] The antisense oligonucleotide may also comprise at least one modified sugar moiety selected from the group including, but not limited to, arabinose, 2-fluoroarabinose, xylulose, and hexose.

[0776] In yet another embodiment, the antisense oligonucleotide comprises at least one modified phosphate backbone selected from the group including, but not limited to, a phosphorothioate, a phosphorodithioate, a phosphoramidothioate, a phosphoramidate, a phosphordiamidate, a methylphosphonate, an alkyl phosphotriester, and a formacetal or analog thereof.

[0777] In yet another embodiment, the antisense oligonucleotide is an a-anomeric oligonucleotide. An a-anomeric oligonucleotide forms specific double-stranded hybrids with complementary RNA in which, contrary to the usual b-units, the strands run parallel to each other (Gautier et al., 1987, Nucl. Acids Res. 15:6625-6641). The oligonucleotide is a 2′-O-methylribonucleotide (Inoue et al., 1987, Nucl. Acids Res. 15:6131-6148), or a chimeric RNA-DNA analogue (Inoue et al., 1987, FEBS Lett. 215:327-330).

[0778] Polynucleotides of the invention may be synthesized by standard methods known in the art, e.g. by use of an automated DNA synthesizer (such as are commercially available from Biosearch, Applied Biosystems, etc.). As examples, phosphorothioate oligonucleotides may be synthesized by the method of Stein et al. (1988, Nucl. Acids Res. 16:3209), methylphosphonate oligonucleotides can be prepared by use of controlled pore glass polymer supports (Sarin et al., 1988, Proc. Natl. Acad. Sci. U.S.A. 85:7448-7451), etc.

[0779] While antisense nucleotides complementary to the coding region sequence could be used, those complementary to the transcribed untranslated region are most preferred.

[0780] Potential antagonists according to the invention also include catalytic RNA, or a ribozyme (See, e.g., PCT International Publication WO 90/11364, published Oct. 4, 1990; Sarver et al, Science 247:1222-1225 (1990). While ribozymes that cleave mRNA at site specific recognition sequences can be used to destroy mRNAs, the use of hammerhead ribozymes is preferred. Hammerhead ribozymes cleave mRNAs at locations dictated by flanking regions that form complementary base pairs with the target mRNA. The sole requirement is that the target mRNA have the following sequence of two bases: 5′-UG-3′. The construction and production of hammerhead ribozymes is well known in the art and is described more fully in Haseloff and Gerlach, Nature 334:585-591 (1988). There are numerous potential hammerhead ribozyme cleavage sites within the nucleotide sequence of SEQ ID NO:X. Preferably, the ribozyme is engineered so that the cleavage recognition site is located near the 5′ end of the mRNA; i.e., to increase efficiency and minimize the intracellular accumulation of non-functional mRNA transcripts.

[0781] As in the antisense approach, the ribozymes of the invention can be composed of modified oligonucleotides (e.g., for improved stability, targeting, etc.) and should be delivered to cells which express in vivo. DNA constructs encoding the ribozyme may be introduced into the cell in the same manner as described above for the introduction of antisense encoding DNA. A preferred method of delivery involves using a DNA construct “encoding” the ribozyme under the control of a strong constitutive promoter, such as, for example, pol III or pol II promoter, so that transfected cells will produce sufficient quantities of the ribozyme to destroy endogenous messages and inhibit translation. Since ribozymes unlike antisense molecules, are catalytic, a lower intracellular concentration is required for efficiency.

[0782] Antagonist/agonist compounds may be employed to inhibit the cell growth and proliferation effects of the polypeptides of the present invention on neoplastic cells and tissues, i.e. stimulation of angiogenesis of tumors, and, therefore, retard or prevent abnormal cellular growth and proliferation, for example, in tumor formation or growth.

[0783] The antagonist/agonist may also be employed to prevent hyper-vascular diseases, and prevent the proliferation of epithelial lens cells after extracapsular cataract surgery. Prevention of the mitogenic activity of the polypeptides of the present invention may also be desirous in cases such as restenosis after balloon angioplasty.

[0784] The antagonist/agonist may also be employed to prevent the growth of scar tissue during wound healing.

[0785] The antagonist/agonist may also be employed to treat the diseases described herein.

[0786] Thus, the invention provides a method of treating disorders or diseases, including but not limited to the disorders or diseases listed throughout this application, associated with overexpression of a polynucleotide of the present invention by administering to a patient (a) an antisense molecule directed to the polynucleotide of the present invention, and/or (b) a ribozyme directed to the polynucleotide of the present invention.

[0787] Binding Peptides and Other Molecules

[0788] The invention also encompasses screening methods for identifying polypeptides and nonpolypeptides that bind polypeptides of the invention, and the binding molecules identified thereby. These binding molecules are useful, for example, as agonists and antagonists of the polypeptides of the invention. Such agonists and antagonists can be used, in accordance with the invention, in the therapeutic embodiments described in detail, below.

[0789] This method comprises the steps of:

[0790] a. contacting polypeptides of the invention with a plurality of molecules; and

[0791] b. identifying a molecule that binds the polypeptides of the invention.

[0792] The step of contacting the polypeptides of the invention with the plurality of molecules may be effected in a number of ways. For example, one may contemplate immobilizing the polypeptides on a solid support and bringing a solution of the plurality of molecules in contact with the immobilized polypeptides. Such a procedure would be akin to an affinity chromatographic process, with the affinity matrix being comprised of the immobilized polypeptides of the invention. The molecules having a selective affinity for the polypeptides can then be purified by affinity selection. The nature of the solid support, process for attachment of the polypeptides to the solid support, solvent, and conditions of the affinity isolation or selection are largely conventional and well known to those of ordinary skill in the art.

[0793] Alternatively, one may also separate a plurality of polypeptides into substantially separate fractions comprising a subset of or individual polypeptides. For instance, one can separate the plurality of polypeptides by gel electrophoresis, column chromatography, or like method known to those of ordinary skill for the separation of polypeptides. The individual polypeptides can also be produced by a transformed host cell in such a way as to be expressed on or about its outer surface (e.g., a recombinant phage). Individual isolates can then be “probed” by the polypeptides of the invention, optionally in the presence of an inducer should one be required for expression, to determine if any selective affinity interaction takes place between the polypeptides and the individual clone. Prior to contacting the polypeptides with each fraction comprising individual polypeptides, the polypeptides could first be transferred to a solid support for additional convenience. Such a solid support may simply be a piece of filter membrane, such as one made of nitrocellulose or nylon. In this manner, positive clones could be identified from a collection of transformed host cells of an expression library, which harbor a DNA construct encoding a polypeptide having a selective affinity for polypeptides of the invention. Furthermore, the amino acid sequence of the polypeptide having a selective affinity for the polypeptides of the invention can be determined directly by conventional means or the coding sequence of the DNA encoding the polypeptide can frequently be determined more conveniently. The primary sequence can then be deduced from the corresponding DNA sequence. If the amino acid sequence is to be determined from the polypeptide itself, one may use microsequencing techniques. The sequencing technique may include mass spectroscopy.

[0794] In certain situations, it may be desirable to wash away any unbound polypeptides from a mixture of the polypeptides of the invention and the plurality of polypeptides prior to attempting to determine or to detect the presence of a selective affinity interaction. Such a wash step may be particularly desirable when the polypeptides of the invention or the plurality of polypeptides are bound to a solid support.

[0795] The plurality of molecules provided according to this method may be provided by way of diversity libraries, such as random or combinatorial peptide or nonpeptide libraries which can be screened for molecules that specifically bind polypeptides of the invention. Many libraries are known in the art that can be used, e.g., chemically synthesized libraries, recombinant (e.g., phage display libraries), and in vitro translation-based libraries. Examples of chemically synthesized libraries are described in Fodor et al., 1991, Science 251:767-773; Houghten et al., 1991, Nature 354:84-86; Lam et al., 1991, Nature 354:82-84; Medynski, 1994, Bio/Technology 12:709-710;Gallop et al., 1994, J. Medicinal Chemistry 37(9):1233-1251; Ohlmeyer et al., 1993, Proc. Natl. Acad. Sci. USA 90:10922-10926; Erb et al., 1994, Proc. Natl. Acad. Sci. USA 91:11422-11426; Houghten et al., 1992, Biotechniques 13:412; Jayawickreme et al., 1994, Proc. Natl. Acad. Sci. USA 91:1614-1618; Salmon et al., 1993, Proc. Natl. Acad. Sci. USA 90:11708-11712; PCT Publication No. WO 93/20242; and Brenner and Lerner, 1992, Proc. Natl. Acad. Sci. USA 89:5381-5383.

[0796] Examples of phage display libraries are described in Scott and Smith, 1990, Science 249:386-390; Devlin et al., 1990, Science, 249:404-406; Christian, R. B., et al., 1992, J. Mol. Biol. 227:711-718); Lenstra, 1992, J. Immunol. Meth. 152:149-157; Kay et al., 1993, Gene 128:59-65; and PCT Publication No. WO 94/18318 dated Aug. 18, 1994. 1746] In vitro translation-based libraries include but are not limited to those described in PCT Publication No. WO 91/05058 dated Apr. 18, 1991; and Mattheakis et al., 1994, Proc. Natl. Acad. Sci. USA 91:9022-9026.

[0797] By way of examples of nonpeptide libraries, a benzodiazepine library (see e.g., Bunin et al., 1994, Proc. Natl. Acad. Sci. USA 91:4708-4712) can be adapted for use. Peptoid libraries (Simon et al., 1992, Proc. Natl. Acad. Sci. USA 89:9367-9371) can also be used. Another example of a library that can be used, in which the amide functionalities in peptides have been permethylated to generate a chemically transformed combinatorial library, is described by Ostresh et al. (1994, Proc. Natl. Acad. Sci. USA 91:11138-11142).

[0798] The variety of non-peptide libraries that are useful in the present invention is great. For example, Ecker and Crooke, 1995, Bio/Technology 13:351-360 list benzodiazepines, hydantoins, piperazinediones, biphenyls, sugar analogs, beta-mercaptoketones, arylacetic acids, acylpiperidines, benzopyrans, cubanes, xanthines, aminimides, and oxazolones as among the chemical species that form the basis of various libraries.

[0799] Non-peptide libraries can be classified broadly into two types: decorated monomers and oligomers. Decorated monomer libraries employ a relatively simple scaffold structure upon which a variety functional groups is added. Often the scaffold will be a molecule with a known useful pharmacological activity. For example, the scaffold might be the benzodiazepine structure.

[0800] Non-peptide oligomer libraries utilize a large number of monomers that are assembled together in ways that create new shapes that depend on the order of the monomers. Among the monomer units that have been used are carbamates, pyrrolinones, and morpholinos. Peptoids, peptide-like oligomers in which the side chain is attached to the alpha amino group rather than the alpha carbon, form the basis of another version of non-peptide oligomer libraries. The first non-peptide oligomer libraries utilized a single type of monomer and thus contained a repeating backbone. Recent libraries have utilized more than one monomer, giving the libraries added flexibility.

[0801] Screening the libraries can be accomplished by any of a variety of commonly known methods. See, e.g., the following references, which disclose screening of peptide libraries: Parmley and Smith, 1989, Adv. Exp. Med. Biol. 251:215-218; Scott and Smith, 1990, Science 249:386-390; Fowlkes et al., 1992; BioTechniques 13:422-427; Oldenburg et al., 1992, Proc. Natl. Acad. Sci. USA 89:5393-5397; Yu et al., 1994, Cell 76:933-945; Staudt et al., 1988, Science 241:577-580; Bock et al., 1992, Nature 355:564-566; Tuerk et al., 1992, Proc. Natl. Acad. Sci. USA 89:6988-6992; Ellington et al., 1992, Nature 355:850-852; U.S. Pat. No. 5,096,815, U.S. Pat. No. 5,223,409, and U.S. Pat. No. 5,198,346, all to Ladner et al.; Rebar and Pabo, 1993, Science 263:671-673; and CT Publication No. WO 94/18318.

[0802] In a specific embodiment, screening to identify a molecule that binds polypeptides of the invention can be carried out by contacting the library members with polypeptides of the invention immobilized on a solid phase and harvesting those library members that bind to the polypeptides of the invention. Examples of such screening methods, termed “panning” techniques are described by way of example in Parmley and Smith, 1988, Gene 73:305-318; Fowlkes et al., 1992, BioTechniques 13:422-427; PCT Publication No. WO 94/18318; and in references cited herein.

[0803] In another embodiment, the two-hybrid system for selecting interacting proteins in yeast (Fields and Song, 1989, Nature 340:245-246; Chien et al., 1991, Proc. Natl. Acad. Sci. USA 88:9578-9582) can be used to identify molecules that specifically bind to polypeptides of the invention.

[0804] Where the binding molecule is a polypeptide, the polypeptide can be conveniently selected from any peptide library, including random peptide libraries, combinatorial peptide libraries, or biased peptide libraries. The term “biased” is used herein to mean that the method of generating the library is manipulated so as to restrict one or more parameters that govern the diversity of the resulting collection of molecules, in this case peptides.

[0805] Thus, a truly random peptide library would generate a collection of peptides in which the probability of finding a particular amino acid at a given position of the peptide is the same for all 20 amino acids. A bias can be introduced into the library, however, by specifying, for example, that a lysine occur every fifth amino acid or that positions 4, 8, and 9 of a decapeptide library be fixed to include only arginine. Clearly, many types of biases can be contemplated, and the present invention is not restricted to any particular bias. Furthermore, the present invention contemplates specific types of peptide libraries, such as phage displayed peptide libraries and those that utilize a DNA construct comprising a lambda phage vector with a DNA insert.

[0806] As mentioned above, in the case of a binding molecule that is a polypeptide, the polypeptide may have about 6 to less than about 60 amino acid residues, preferably about 6 to about 10 amino acid residues, and most preferably, about 6 to about 22 amino acids. In another embodiment, a binding polypeptide has in the range of 15-100 amino acids, or 20-50 amino acids.

[0807] The selected binding polypeptide can be obtained by chemical synthesis or recombinant expression.

[0808] Other Activities

[0809] A polypeptide, polynucleotide, agonist, or antagonist of the present invention, as a result of the ability to stimulate vascular endothelial cell growth, may be employed in treatment for stimulating re-vascularization of ischemic tissues due to various disease conditions such as thrombosis, arteriosclerosis, and other cardiovascular conditions. The polypeptide, polynucleotide, agonist, or antagonist of the present invention may also be employed to stimulate angiogenesis and limb regeneration, as discussed above.

[0810] A polypeptide, polynucleotide, agonist, or antagonist of the present invention may also be employed for treating wounds due to injuries, burns, post-operative tissue repair, and ulcers since they are mitogenic to various cells of different origins, such as fibroblast cells and skeletal muscle cells, and therefore, facilitate the repair or replacement of damaged or diseased tissue.

[0811] A polypeptide, polynucleotide, agonist, or antagonist of the present invention may also be employed stimulate neuronal growth and to treat and prevent neuronal damage which occurs in certain neuronal disorders or neuro-degenerative conditions such as Alzheimer's disease, Parkinson's disease, and AIDS-related complex. A polypeptide, polynucleotide, agonist, or antagonist of the present invention may have the ability to stimulate chondrocyte growth, therefore, they may be employed to enhance bone and periodontal regeneration and aid in tissue transplants or bone grafts.

[0812] A polypeptide, polynucleotide, agonist, or antagonist of the present invention may be also be employed to prevent skin aging due to sunburn by stimulating keratinocyte growth.

[0813] A polypeptide, polynucleotide, agonist, or antagonist of the present invention may also be employed for preventing hair loss, since FGF family members activate hair-forming cells and promotes melanocyte growth. Along the same lines, a polypeptide, polynucleotide, agonist, or antagonist of the present invention may be employed to stimulate growth and differentiation of hematopoietic cells and bone marrow cells when used in combination with other cytokines.

[0814] A polypeptide, polynucleotide, agonist, or antagonist of the present invention may also be employed to maintain organs before transplantation or for supporting cell culture of primary tissues. A polypeptide, polynucleotide, agonist, or antagonist of the present invention may also be employed for inducing tissue of mesodermal origin to differentiate in early embryos.

[0815] A polypeptide, polynucleotide, agonist, or antagonist of the present invention may also increase or decrease the differentiation or proliferation of embryonic stem cells, besides, as discussed above, hematopoietic lineage.

[0816] A polypeptide, polynucleotide, agonist, or antagonist of the present invention may also be used to modulate mammalian characteristics, such as body height, weight, hair color, eye color, skin, percentage of adipose tissue, pigmentation, size, and shape (e.g., cosmetic surgery). Similarly, a polypeptide, polynucleotide, agonist, or antagonist of the present invention may be used to modulate mammalian metabolism affecting catabolism, anabolism, processing, utilization, and storage of energy.

[0817] A polypeptide, polynucleotide, agonist, or antagonist of the present invention may be used to change a mammal's mental state or physical state by influencing biorhythms, caricadic rhythms, depression (including depressive disorders), tendency for violence, tolerance for pain, reproductive capabilities (preferably by Activin or Inhibin-like activity), hormonal or endocrine levels, appetite, libido, memory, stress, or other cognitive qualities.

[0818] A polypeptide, polynucleotide, agonist, or antagonist of the present invention may also be used as a food additive or preservative, such as to increase or decrease storage capabilities, fat content, lipid, protein, carbohydrate, vitamins, minerals, cofactors or other nutritional components.

[0819] The above-recited applications have uses in a wide variety of hosts. Such hosts include, but are not limited to, human, murine, rabbit, goat, guinea pig, camel, horse, mouse, rat, hamster, pig, micro-pig, chicken, goat, cow, sheep, dog, cat, non-human primate, and human. In specific embodiments, the host is a mouse, rabbit, goat, guinea pig, chicken, rat, hamster, pig, sheep, dog or cat. In preferred embodiments, the host is a mammal. In most preferred embodiments, the host is a human.

[0820] Other Preferred Embodiments

[0821] Other preferred embodiments of the claimed invention include an isolated nucleic acid molecule comprising a nucleotide sequence which is at least 95% identical to a sequence of at least about 50 contiguous nucleotides in the nucleotide sequence of SEQ ID NO:X or the complementary strand thereto, the nucleotide sequence as defined in column 5 of Table 1A or columns 8 and 9 of Table 2 or the complementary strand thereto, and/or cDNA contained in Clone ID NO:Z.

[0822] Also preferred is a nucleic acid molecule wherein said sequence of contiguous nucleotides is included in the nucleotide sequence of the portion of SEQ ID NO:X as defined in column 5, “ORF (From-To)”, in Table 1A.

[0823] Also preferred is a nucleic acid molecule wherein said sequence of contiguous nucleotides is included in the nucleotide sequence of the portion of SEQ ID NO:X as defined in columns 8 and 9, “NT From” and “NT To” respectively, in Table 2.

[0824] Also preferred is an isolated nucleic acid molecule comprising a nucleotide sequence which is at least 95% identical to a sequence of at least about 150 contiguous nucleotides in the nucleotide sequence of SEQ ID NO:X or the complementary strand thereto, the nucleotide sequence as defined in column 5 of Table 1A or columns 8 and 9 of Table 2 or the complementary strand thereto, and/or cDNA contained in Clone ID NO:Z.

[0825] Further preferred is an isolated nucleic acid molecule comprising a nucleotide sequence which is at least 95% identical to a sequence of at least about 500 contiguous nucleotides in the nucleotide sequence of SEQ ID NO:X or the complementary strand thereto, the nucleotide sequence as defined in column 5 of Table 1A or columns 8 and 9 of Table 2 or the complementary strand thereto, and/or cDNA contained in Clone ID NO:Z.

[0826] A further preferred embodiment is a nucleic acid molecule comprising a nucleotide sequence which is at least 95% identical to the nucleotide sequence of the portion of SEQ ID NO:X defined in column 5, “ORF (From-To)”, in Table 1A.

[0827] A further preferred embodiment is a nucleic acid molecule comprising a nucleotide sequence which is at least 95% identical to the nucleotide sequence of the portion of SEQ ID NO:X defined in columns 8 and 9, “NT From” and “NT To”, respectively, in Table 2.

[0828] A further preferred embodiment is an isolated nucleic acid molecule comprising a nucleotide sequence which is at least 95% identical to the complete nucleotide sequence of SEQ ID NO:X or the complementary strand thereto, the nucleotide sequence as defined in column 5 of Table 1A or columns 8 and 9 of Table 2 or the complementary strand thereto, and/or cDNA contained in Clone ID NO:Z.

[0829] Also preferred is an isolated nucleic acid molecule which hybridizes under stringent hybridization conditions to a nucleic acid molecule comprising a nucleotide sequence of SEQ ID NO:X or the complementary strand thereto, the nucleotide sequence as defined in column 5 of Table 1A or columns 8 and 9 of Table 2 or the complementary strand thereto, and/or cDNA contained in Clone ID NO:Z, wherein said nucleic acid molecule which hybridizes does not hybridize under stringent hybridization conditions to a nucleic acid molecule having a nucleotide sequence consisting of only A residues or of only T residues.

[0830] Also preferred is a composition of matter comprising a DNA molecule which comprises the cDNA contained in Clone ID NO:Z.

[0831] Also preferred is an isolated nucleic acid molecule comprising a nucleotide sequence which is at least 95% identical to a sequence of at least 50 contiguous nucleotides of the cDNA sequence contained in Clone ID NO:Z.

[0832] Also preferred is an isolated nucleic acid molecule, wherein said sequence of at least 50 contiguous nucleotides is included in the nucleotide sequence of an open reading frame sequence encoded by cDNA contained in Clone ID NO:Z.

[0833] Also preferred is an isolated nucleic acid molecule comprising a nucleotide sequence which is at least 95% identical to sequence of at least 150 contiguous nucleotides in the nucleotide sequence encoded by cDNA contained in Clone ID NO:Z.

[0834] A further preferred embodiment is an isolated nucleic acid molecule comprising a nucleotide sequence which is at least 95% identical to sequence of at least 500 contiguous nucleotides in the nucleotide sequence encoded by cDNA contained in Clone ID NO:Z.

[0835] A further preferred embodiment is an isolated nucleic acid molecule comprising a nucleotide sequence which is at least 95% identical to the complete nucleotide sequence encoded by cDNA contained in Clone ID NO:Z.

[0836] A further preferred embodiment is a method for detecting in a biological sample a nucleic acid molecule comprising a nucleotide sequence which is at least 95% identical to a sequence of at least 50 contiguous nucleotides in a sequence selected from the group consisting of: a nucleotide sequence of SEQ ID NO:X or the complementary strand thereto; the nucleotide sequence as defined in column 5 of Table 1A or columns 8 and 9 of Table 2 or the complementary strand thereto; and a nucleotide sequence encoded by cDNA contained in Clone ID NO:Z; which method comprises a step of comparing a nucleotide sequence of at least one nucleic acid molecule in said sample with a sequence selected from said group and determining whether the sequence of said nucleic acid molecule in said sample is at least 95% identical to said selected sequence.

[0837] Also preferred is the above method wherein said step of comparing sequences comprises determining the extent of nucleic acid hybridization between nucleic acid molecules in said sample and a nucleic acid molecule comprising said sequence selected from said group. Similarly, also preferred is the above method wherein said step of comparing sequences is performed by comparing the nucleotide sequence determined from a nucleic acid molecule in said sample with said sequence selected from said group. The nucleic acid molecules can comprise DNA molecules or RNA molecules.

[0838] A further preferred embodiment is a method for identifying the species, tissue or cell type of a biological sample which method comprises a step of detecting nucleic acid molecules in said sample, if any, comprising a nucleotide sequence that is at least 95% identical to a sequence of at least 50 contiguous nucleotides in a sequence selected from the group consisting of: a nucleotide sequence of SEQ ID NO:X or the complementary strand thereto; the nucleotide sequence as defined in column 5 of Table 1A or columns 8 and 9 of Table 2 or the complementary strand thereto; and a nucleotide sequence of the cDNA contained in Clone ID NO:Z.

[0839] The method for identifying the species, tissue or cell type of a biological sample can comprise a step of detecting nucleic acid molecules comprising a nucleotide sequence in a panel of at least two nucleotide sequences, wherein at least one sequence in said panel is at least 95% identical to a sequence of at least 50 contiguous nucleotides in a sequence selected from said group.

[0840] Also preferred is a method for diagnosing in a subject a pathological condition associated with abnormal structure or expression of a nucleotide sequence of SEQ ID NO:X or the complementary strand thereto; the nucleotide sequence as defined in column 5 of Table 1A or columns 8 and 9 of Table 2 or the complementary strand thereto; or the cDNA contained in Clone ID NO:Z which encodes a protein, wherein the method comprises a step of detecting in a biological sample obtained from said subject nucleic acid molecules, if any, comprising a nucleotide sequence that is at least 95% identical to a sequence of at least 50 contiguous nucleotides in a sequence selected from the group consisting of: a nucleotide sequence of SEQ ID NO:X or the complementary strand thereto; the nucleotide sequence as defined in column 5 of Table 1A or columns 8 and 9 of Table 2 or the complementary strand thereto; and a nucleotide sequence of cDNA contained in Clone ID NO:Z.

[0841] The method for diagnosing a pathological condition can comprise a step of detecting nucleic acid molecules comprising a nucleotide sequence in a panel of at least two nucleotide sequences, wherein at least one sequence in said panel is at least 95% identical to a sequence of at least 50 contiguous nucleotides in a sequence selected from said group.

[0842] Also preferred is a composition of matter comprising isolated nucleic acid molecules wherein the nucleotide sequences of said nucleic acid molecules comprise a panel of at least two nucleotide sequences, wherein at least one sequence in said panel is at least 95% identical to a sequence of at least 50 contiguous nucleotides in a sequence selected from the group consisting of: a nucleotide sequence of SEQ ID NO:X or the complementary strand thereto; the nucleotide sequence as defined in column 5 of Table 1A or columns 8 and 9 of Table 2 or the complementary strand thereto; and a nucleotide sequence encoded by cDNA contained in Clone ID NO:Z. The nucleic acid molecules can comprise DNA molecules or RNA molecules.

[0843] Also preferred is a composition of matter comprising isolated nucleic acid molecules wherein the nucleotide sequences of said nucleic acid molecules comprise a DNA microarray or “chip” of at least 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 15, 20, 25, 30, 40, 50, 100, 150, 200, 250, 300, 500, 1000, 2000, 3000, or 4000 nucleotide sequences, wherein at least one sequence in said DNA microarray or “chip” is at least 95% identical to a sequence of at least 50 contiguous nucleotides in a sequence selected from the group consisting of: a nucleotide sequence of SEQ ID NO:X wherein X is any integer as defined in Table 1A; and a nucleotide sequence encoded by a human cDNA clone identified by a cDNA “Clone ID” in Table 1A.

[0844] Also preferred is an isolated polypeptide comprising an amino acid sequence at least 90% identical to a sequence of at least about 10 contiguous amino acids in the polypeptide sequence of SEQ ID NO:Y; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and/or a polypeptide encoded by cDNA contained in Clone ID NO:Z.

[0845] Also preferred is an isolated polypeptide comprising an amino acid sequence at least 95% identical to a sequence of at least about 30 contiguous amino acids in the amino acid sequence of SEQ ID NO:Y; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and/or a polypeptide encoded by cDNA contained in Clone ID NO:Z.

[0846] Further preferred is an isolated polypeptide comprising an amino acid sequence at least 95% identical to a sequence of at least about 100 contiguous amino acids in the amino acid sequence of SEQ ID NO:Y; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and/or a polypeptide encoded by cDNA contained in Clone ID NO:Z.

[0847] Further preferred is an isolated polypeptide comprising an amino acid sequence at least 95% identical to the complete amino acid sequence of SEQ ID NO:Y; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and/or a polypeptide encoded by cDNA contained in Clone ID NO:Z.

[0848] Further preferred is an isolated polypeptide comprising an amino acid sequence at least 90% identical to a sequence of at least about 10 contiguous amino acids in the complete amino acid sequence of a polypeptide encoded by contained in Clone ID NO:Z

[0849] Also preferred is a polypeptide wherein said sequence of contiguous amino acids is included in the amino acid sequence of a portion of said polypeptide encoded by cDNA contained in Clone ID NO:Z; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and/or the polypeptide sequence of SEQ ID NO:Y.

[0850] Also preferred is an isolated polypeptide comprising an amino acid sequence at least 95% identical to a sequence of at least about 30 contiguous amino acids in the amino acid sequence of a polypeptide encoded by the cDNA contained in Clone ID NO:Z.

[0851] Also preferred is an isolated polypeptide comprising an amino acid sequence at least 95% identical to a sequence of at least about 100 contiguous amino acids in the amino acid sequence of a polypeptide encoded by cDNA contained in Clone ID NO:Z.

[0852] Also preferred is an isolated polypeptide comprising an amino acid sequence at least 95% identical to the amino acid sequence of a polypeptide encoded by the cDNA contained in Clone ID NO:Z.

[0853] Further preferred is an isolated antibody which binds specifically to a polypeptide comprising an amino acid sequence that is at least 90% identical to a sequence of at least 10 contiguous amino acids in a sequence selected from the group consisting of: a polypeptide sequence of SEQ ID NO:Y; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and a polypeptide encoded by the cDNA contained in Clone ID NO:Z.

[0854] Further preferred is a method for detecting in a biological sample a polypeptide comprising an amino acid sequence which is at least 90% identical to a sequence of at least 10 contiguous amino acids in a sequence selected from the group consisting of: a polypeptide sequence of SEQ ID NO:Y; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and a polypeptide encoded by the cDNA contained in Clone ID NO:Z; which method comprises a step of comparing an amino acid sequence of at least one polypeptide molecule in said sample with a sequence selected from said group and determining whether the sequence of said polypeptide molecule in said sample is at least 90% identical to said sequence of at least 10 contiguous amino acids.

[0855] Also preferred is the above method wherein said step of comparing an amino acid sequence of at least one polypeptide molecule in said sample with a sequence selected from said group comprises determining the extent of specific binding of polypeptides in said sample to an antibody which binds specifically to a polypeptide comprising an amino acid sequence that is at least 90% identical to a sequence of at least 10 contiguous amino acids in a sequence selected from the group consisting of: a polypeptide sequence of SEQ ID NO:Y; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and a polypeptide encoded by the cDNA contained in Clone ID NO:Z.

[0856] Also preferred is the above method wherein said step of comparing sequences is performed by comparing the amino acid sequence determined from a polypeptide molecule in said sample with said sequence selected from said group.

[0857] Also preferred is a method for identifying the species, tissue or cell type of a biological sample which method comprises a step of detecting polypeptide molecules in said sample, if any, comprising an amino acid sequence that is at least 90% identical to a sequence of at least 10 contiguous amino acids in a sequence selected from the group consisting of: polypeptide sequence of SEQ ID NO:Y; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and a polypeptide encoded by the cDNA contained in Clone ID NO:Z.

[0858] Also preferred is the above method for identifying the species, tissue or cell type of a biological sample, which method comprises a step of detecting polypeptide molecules comprising an amino acid sequence in a panel of at least two amino acid sequences, wherein at least one sequence in said panel is at least 90% identical to a sequence of at least 10 contiguous amino acids in a sequence selected from the above group.

[0859] Also preferred is a method for diagnosing in a subject a pathological condition associated with abnormal structure or expression of a nucleic acid sequence identified in Table 1A or Table 2 encoding a polypeptide, which method comprises a step of detecting in a biological sample obtained from said subject polypeptide molecules comprising an amino acid sequence in a panel of at least two amino acid sequences, wherein at least one sequence in said panel is at least 90% identical to a sequence of at least 10 contiguous amino acids in a sequence selected from the group consisting of: polypeptide sequence of SEQ ID NO:Y; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and a polypeptide encoded by the cDNA contained in Clone ID NO:Z.

[0860] In any of these methods, the step of detecting said polypeptide molecules includes using an antibody.

[0861] Also preferred is an isolated nucleic acid molecule comprising a nucleotide sequence which is at least 95% identical to a nucleotide sequence encoding a polypeptide wherein said polypeptide comprises an amino acid sequence that is at least 90% identical to a sequence of at least 10 contiguous amino acids in a sequence selected from the group consisting of: polypeptide sequence of SEQ ID NO:Y; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and a polypeptide encoded by the cDNA contained in Clone ID NO:Z.

[0862] Also preferred is an isolated nucleic acid molecule, wherein said nucleotide sequence encoding a polypeptide has been optimized for expression of said polypeptide in a prokaryotic host.

[0863] Also preferred is a polypeptide molecule, wherein said polypeptide comprises an amino acid sequence selected from the group consisting of: polypeptide sequence of SEQ ID NO:Y; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and a polypeptide encoded by the cDNA contained in Clone ID NO:Z.

[0864] Further preferred is a method of making a recombinant vector comprising inserting any of the above isolated nucleic acid molecule into a vector. Also preferred is the recombinant vector produced by this method. Also preferred is a method of making a recombinant host cell comprising introducing the vector into a host cell, as well as the recombinant host cell produced by this method.

[0865] Also preferred is a method of making an isolated polypeptide comprising culturing this recombinant host cell under conditions such that said polypeptide is expressed and recovering said polypeptide. Also preferred is this method of making an isolated polypeptide, wherein said recombinant host cell is a eukaryotic cell and said polypeptide is a human protein comprising an amino acid sequence selected from the group consisting of: polypeptide sequence of SEQ ID NO:Y; a polypeptide encoded by SEQ ID NO:X or the complementary strand thereto; the polypeptide encoded by the nucleotide sequence as defined in columns 8 and 9 of Table 2; and a polypeptide encoded by the cDNA contained in Clone ID NO:Z. The isolated polypeptide produced by this method is also preferred.

[0866] Also preferred is a method of treatment of an individual in need of an increased level of a protein activity, which method comprises administering to such an individual a Therapeutic comprising an amount of an isolated polypeptide, polynucleotide, immunogenic fragment or analogue thereof, binding agent, antibody, or antigen binding fragment of the claimed invention effective to increase the level of said protein activity in said individual.

[0867] Also preferred is a method of treatment of an individual in need of a decreased level of a protein activity, which method comprised administering to such an individual a Therapeutic comprising an amount of an isolated polypeptide, polynucleotide, immunogenic fragment or analogue thereof, binding agent, antibody, or antigen binding fragment of the claimed invention effective to decrease the level of said protein activity in said individual.

[0868] Also preferred is a method of treatment of an individual in need of a specific delivery of toxic compositions to diseased cells (e.g., tumors, leukemias or lymphomas), which method comprises administering to such an individual a Therapeutic comprising an amount of an isolated polypeptide of the invention, including, but not limited to a binding agent, or antibody of the claimed invention that are associated with toxin or cytotoxic prodrugs.

[0869] Having generally described the invention, the same will be more readily understood by reference to the following examples, which are provided by way of illustration and are not intended as limiting.

8TABLE 6ATCC DepositsDeposit DateATCC Designation NumberLP01, LP02, LP03,May-20-97209059, 209060, 209061,LP04, LP05, LP06,209062, 209063, 209064,LP07, LP08, LP09,209065, 209066, 209067,LP10, LP11,209068, 209069LP12Jan-12-98209579LP13Jan-12-98209578LP14Jul-16-98203067LP15Jul-16-98203068LP16Feb-1-99203609LP17Feb-1-99203610LP20Nov-17-98203485LP21Jun-18-99PTA-252LP22Jun-18-99PTA-253LP23Dec-22-99PTA-1081

EXAMPLES

Example 1

Isolation of a Selected cDNA Clone from the Deposited Sample

[0870] Each Clone ID NO:Z is contained in a plasmid vector. Table 7 identifies the vectors used to construct the cDNA library from which each clone was isolated. In many cases, the vector used to construct the library is a phage vector from which a plasmid has been excised. The following correlates the related plasmid for each phage vector used in constructing the cDNA library. For example, where a particular clone is identified in Table 7 as being isolated in the vector “Lambda Zap,” the corresponding deposited clone is in “pBluescript.”

9Vector Used to Construct LibraryCorresponding Deposited PlasmidLambda ZappBluescript (pBS)Uni-Zap XRpBluescript (pBS)Zap ExpresspBKlafmid BAplafMid BApSport1pSport1pCMVSport 2.0pCMVSport 2.0pCMVSport 3.0pCMVSport 3.0pCR ® 2.1pCR ® 2.1

[0871] Vectors Lambda Zap (U.S. Pat. Nos. 5,128,256 and 5,286,636), Uni-Zap XR (U.S. Pat. Nos. 5,128, 256 and 5,286,636), Zap Express (U.S. Pat. Nos. 5,128,256 and 5,286,636), pBluescript (pBS) (Short, J. M. et al., Nucleic Acids Res. 16:7583-7600 (1988); Alting-Mees, M. A. and Short, J. M., Nucleic Acids Res. 17:9494 (1989)) and pBK (Alting-Mees, M. A. et al., Strategies 5:58-61 (1992)) are commercially available from Stratagene Cloning Systems, Inc., 11011 N. Torrey Pines Road, La Jolla, Calif., 92037. pBS contains an ampicillin resistance gene and pBK contains a neomycin resistance gene. Both can be transformed into E. coli strain XL-1 Blue, also available from Stratagene. pBS comes in 4 forms SK+, SK−, KS+ and KS. The S and K refers to the orientation of the polylinker to the T7 and T3 primer sequences which flank the polylinker region (“S” is for SacI and “K” is for KpnI which are the first sites on each respective end of the linker). “+” or “−” refer to the orientation of the fl origin of replication (“ori”), such that in one orientation, single stranded rescue initiated from the fl ori generates sense strand DNA and in the other, antisense.

[0872] Vectors pSport1, pCMVSport 2.0 and pCMVSport 3.0, were obtained from Life Technologies, Inc., P. O. Box 6009, Gaithersburg, Md. 20897. All Sport vectors contain an ampicillin resistance gene and may be transformed into E. coli strain DH10B, also available from Life Technologies. (See, for instance, Gruber, C. E., et al., Focus 15:59 (1993)). Vector lafmid BA (Bento Soares, Columbia University, NY) contains an ampicillin resistance gene and can be transformed into E. coli strain XL-1 Blue. Vector pCR®2.1, which is available from Invitrogen, 1600 Faraday Avenue, Carlsbad, Calif. 92008, contains an ampicillin resistance gene and may be transformed into E. coli strain DH10B, available from Life Technologies. (See, for instance, Clark, J. M., Nuc. Acids Res. 16:9677-9686 (1988) and Mead, D. et al., Bio/Technology 9: (1991)). Preferably, a polynucleotide of the present invention does not comprise the phage vector sequences identified for the particular clone in Table 7, as well as the corresponding plasmid vector sequences designated above.

[0873] The deposited material in the sample assigned the ATCC Deposit Number cited by reference to Tables 1, 2, 6 and 7 for any given cDNA clone also may contain one or more additional plasmids, each comprising a cDNA clone different from that given clone. Thus, deposits sharing the same ATCC Deposit Number contain at least a plasmid for each Clone ID NO:Z.

10TABLE 7ATCCLibraries owned by CatalogCatalog DescriptionVectorDepositHUKA HUKB HUKC HUKD HUKEHuman Uterine CancerLambda ZAP IILP01HUKF HUKGHCNA HCNBHuman ColonLambda Zap IILP01HFFAHuman Fetal Brain, random primedLambda Zap IILP01HTWAResting T-CellLambda ZAP IILP01HBQAEarly Stage Human Brain, randomLambda ZAP IILP01primedHLMB HLMF HLMG HLMH HLMIbreast lymph node CDNA libraryLambda ZAP IILP01HLMJ HLMM HLMNHCQA HCQBhuman colon cancerLamda ZAP IILP01HMEA HMEC HMED HMEE HMEFHuman Microvascular EndothelialLambda ZAP IILP01HMEG HMEI HMEJ HMEK HMELCells, fract. AHUSA HUSCHuman Umbilical Vein EndothelialLambda ZAP IILP01Cells, fract. AHLQA HLQBHepatocellular TumorLambda ZAP IILP01HHGA HHGB HHGC HHGDHemangiopericytomaLambda ZAP IILP01HSDMHuman Striatum Depression, re-Lambda ZAP IILP01rescueHUSHH Umbilical Vein Endothelial Cells,Lambda ZAP IILP01frac A, re-excisionHSGSSalivary gland, subtractedLambda ZAP IILP01HFXA HFXB HFXC HFXD HFXEBrain frontal cortexLambda ZAP IILP01HFXF HFXG HFXHHPQA HPQB HPQCPERM TF274Lambda ZAP IILP01HFXJ HFXKBrain Frontal Cortex, re-excisionLambda ZAP IILP01HCWA HCWB HCWC HCWD HCWECD34 positive cells (Cord Blood)ZAP ExpressLP02HCWF HCWG HCWH HCWI HCWJHCWKHCUA HCUB HCUCCD34 depleted Buffy Coat (CordZAP ExpressLP02Blood)HRSMA-14 cell lineZAP ExpressLP02HRSAA1-CELL LINEZAP ExpressLP02HCUD HCUE HCUF HCUG HCUHCD34 depleted Buffy Coat (CordZAP ExpressLP02HCUIBlood), re-excisionHBXE HBXF HBXGH. Whole Brain #2, re-excisionZAP ExpressLP02HRLML8 cell lineZAP ExpressLP02HBXA HBXB HBXC HBXDHuman Whole Brain #2 - Oligo dT >ZAP ExpressLP021.5 KbHUDA HUDB HUDCTestesZAP ExpressLP02HHTM HHTN HHTOH. hypothalamus, frac A, re-excisionZAP ExpressLP02HHTLH. hypothalamus, frac AZAP ExpressLP02HASA HASDHuman Adult SpleenUni-ZAP XRLP03HFKC HFKD HFKE HFKF HFKGHuman Fetal KidneyUni-ZAP XRLP03HE8A HE8B HE8C HE8D HE8E HE8FHuman 8 Week Whole EmbryoUni-ZAP XRLP03HE8M HE8NHGBA HGBD HGBE HGBF HGBGHuman Gall BladderUni-ZAP XRLP03HGBH HGBIHLHA HLHB HLHC HLHD HLHEHuman Fetal Lung IIIUni-ZAP XRLP03HLHF HLHG HLHH HLHQHPMA HPMB HPMC HPMD HPMEHuman PlacentaUni-ZAP XRLP03HPMF HPMG HPMHHPRA HPRB HPRC HPRDHuman ProstateUni-ZAP XRLP03HSIA HSIC HSID HSIEHuman Adult Small IntestineUni-ZAP XRLP03HTEA HTEB HTEC HTED HTEEHuman TestesUni-ZAP XRLP03HTEF HTEG HTEH HTEI HTEJ HTEKHTPA HTPB HTPC HTPD HTPEHuman Pancreas TumorUni-ZAP XRLP03HTTA HTTB HTTC HTTD HTTEHuman Testes TumorUni-ZAP XRLP03HTTFHAPA HAPB HAPC HAPMHuman Adult PulmonaryUni-ZAP XRLP03HETA HETB HETC HETD HETEHuman Endometrial TumorUni-ZAP XRLP03HETF HETG HETH HETIHHFB HHFC HHFD HHFE HHFFHuman Fetal HeartUni-ZAP XRLP03HHFG HHFH HHFIHHPB HHPC HHPD HHPE HHPFHuman HippocampusUni-ZAP XRLP03HHPG HHPHHCE1 HCE2 HCE3 HCE4 HCE5 HCEBHuman CerebellumUni-ZAP XRLP03HCEC HCED HCEE HCEF HCEGHUVB HUVC HUVD HUVEHuman Umbilical Vein, Endo.Uni-ZAP XRLP03remakeHSTA HSTB HSTC HSTDHuman Skin TumorUni-ZAP XRLP03HTAA HTAB HTAC HTAD HTAEHuman Activated T-CellsUni-ZAP XRLP03HFEA HFEB HFECHuman Fetal Epithelium (Skin)Uni-ZAP XRLP03HJPA HJPB HJPC HJPDHUMAN JURKAT MEMBRANEUni-ZAP XRLP03BOUND POLYSOMESHESAHuman epithelioid sarcomaUni-Zap XRLP03HLTA HLTB HLTC HLTD HLTEHuman T-Cell LymphomaUni-ZAP XRLP03HLTFHFTA HFTB HFTC HFTDHuman Fetal Dura MaterUni-ZAP XRLP03HRDA HRDB HRDC HRDD HRDEHuman RhabdomyosarcomaUni-ZAP XRLP03HRDFHCAA HCAB HCACCem cells cyclohexamide treatedUni-ZAP XRLP03HRGA HRGB HRGC HRGDRaji Cells, cyclohexamide treatedUni-ZAP XRLP03HSUA HSUB HSUC HSUMSupt Cells, cyclohexamide treatedUni-ZAP XRLP03HT4A HT4C HT4DActivated T-Cells, 12 hrs.Uni-ZAP XRLP03HE9A HE9B HE9C HE9D HE9E HE9FNine Week Old Early Stage HumanUni-ZAP XRLP03HE9G HE9H HE9M HE9NHATA HATB HATC HATD HATEHuman Adrenal Gland TumorUni-ZAP XRLP03HT5AActivated T-Cells, 24 hrs.Uni-ZAP XRLP03HFGA HFGMHuman Fetal BrainUni-ZAP XRLP03HNEA HNEB HNEC HNED HNEEHuman NeutrophilUni-ZAP XRLP03HBGB HBGDHuman Primary Breast CancerUni-ZAP XRLP03HBNA HBNBHuman Normal BreastUni-ZAP XRLP03HCASCem Cells, cyclohexamide treated,Uni-ZAP XRLP03subtraHHPSHuman Hippocampus, subtractedpBSLP03HKCS HKCUHuman Colon Cancer, subtractedpBSLP03HRGSRaji cells, cyclohexamide treated,pBSLP03subtractedHSUTSupt cells, cyclohexamide treated,pBSLP03differentially expressedHT4SActivated T-Cells, 12 hrs, subtractedUni-ZAP XRLP03HCDA HCDB HCDC HCDD HCDEHuman ChondrosarcomaUni-ZAP XRLP03HOAA HOAB HOACHuman OsteosarcomaUni-ZAP XRLP03HTLA HTLB HTLC HTLD HTLEHuman adult testis, large insertsUni-ZAP XRLP03HTLFHLMA HLMC HLMDBreast Lymph node cDNA libraryUni-ZAP XRLP03H6EA H6EB H6ECHL-60, PMA 4HUni-ZAP XRLP03HTXA HTXB HTXC HTXD HTXEActivated T-Cell (12 hs)/ThiouridineUni-ZAP XRLP03HTXF HTXG HTXHlabelledEcoHNFA HNFB HNFC HNFD HNFEHuman Neutrophil, ActivatedUni-ZAP XRLP03HNFF HNFG HNFH HNFJHTOB HTOCHUMAN TONSILS, FRACTION 2Uni-ZAP XRLP03HMGBHuman OB MG63 control fraction IUni-ZAP XRLP03HOPBHuman OB HOS control fraction IUni-ZAP XRLP03HORBHuman OB HOS treated (10 nM E2)Uni-ZAP XRLP03fraction IHSVA HSVB HSVCHuman Chronic SynovitisUni-ZAP XRLP03HROAHUMAN STOMACHUni-ZAP XRLP03HBJA HBJB HBJC HBJD HBJE HBJFHUMAN B CELL LYMPHOMAUni-ZAP XRLP03HBJG HBJH HBJI HBJJ HBJKHCRA HCRB HCRChuman corpus colosumUni-ZAP XRLP03HODA HODB HODC HODDhuman ovarian cancerUni-ZAP XRLP03HDSADermatofibrosarcoma ProtuberanceUni-ZAP XRLP03HMWA HMWB HMWC HMWDBone Marrow Cell Line (RS4, 11)Uni-ZAP XRLP03HMWE HMWF HMWG HMWHHMWI HMWJHSOAstomach cancer (human)Uni-ZAP XRLP03HERASKINUni-ZAP XRLP03HMDABrain-medulloblastomaUni-ZAP XRLP03HGLA HGLB HGLDGlioblastomaUni-ZAP XRLP03HEAAH. Atrophic EndometriumUni-ZAP XRLP03HBCA HBCBH. Lymph node breast CancerUni-ZAP XRLP03HPWTHuman Prostate BPH, re-excisionUni-ZAP XRLP03HFVG HFVH HFVIFetal Liver, subtraction IIpBSLP03HNFIHuman Neutrophils, Activated, re-pBSLP03excisionHBMB HBMC HBMDHuman Bone Marrow, re-excisionpBSLP03HKML HKMM HKMNH. Kidney Medulla, re-excisionpBSLP03HKIX HKIYH. Kidney Cortex, subtractedpBSLP03HADTH. Amygdala Depression, subtractedpBSLP03H6ASHI-60, untreated, subtractedUni-ZAP XRLP03H6ESHL-60, PMA 4 H, subtractedUni-ZAP XRLP03H6BSHL-60, RA 4 h, SubtractedUni-ZAP XRLP03H6CSHL-60, PMA 1d, subtractedUni-ZAP XRLP03HTXJ HTXKActivated T-cell(12 h)/Thiouridine-re-Uni-ZAP XRLP03excisionHMSA HMSB HMSC HMSD HMSEMonocyte activatedUni-ZAP XRLP03HMSF HMSG HMSH HMSI HMSJHMSKHAGA HAGB HAGC HAGD HAGEHuman AmygdalaUni-ZAP XRLP03HAGFHSRA HSRB HSRESTROMAL-OSTEOCLASTOMAUni-ZAP XRLP03HSRD HSRF HSRG HSRHHuman Osteoclastoma Stromal Cells -Uni-ZAP XRLP03unamplifiedHSQA HSQB HSQC HSQD HSQEStromal cell TF274Uni-ZAP XRLP03HSQF HSQGHSKA HSKB HSKC HSKD HSKESmooth muscle, serum treatedUni-ZAP XRLP03HSKF HSKZHSLA HSLB HSLC HSLD HSLE HSLFSmooth muscle, controlUni-ZAP XRLP03HSLGHSDA HSDD HSDE HSDF HSDGSpinal cordUni-ZAP XRLP03HSDHHPWSProstate-BPH subtracted IIpBSLP03HSKW HSKX HSKYSmooth Muscle-HASTE normalizedpBSLP03HFPB HFPC HFPDH. Frontal cortex, epileptic; re-excisionUni-ZAP XRLP03HSDI HSDJ HSDKSpinal Cord, re-excisionUni-ZAP XRLP03HSKN HSKOSmooth Muscle Serum Treated, NormpBSLP03HSKG HSKH HSKISmooth muscle, serum induced, re-excpBSLP03HFCA HFCB HFCC HFCD HFCEHuman Fetal BrainUni-ZAP XRLP04HFCFHPTA HPTB HPTDHuman PituitaryUni-ZAP XRLP04HTHB HTHC HTHDHuman ThymusUni-ZAP XRLP04HE6B HE6C HE6D HE6E HE6F HE6GHuman Whole Six Week Old EmbryoUni-ZAP XRLP04HE6SHSSA HSSB HSSC HSSD HSSE HSSFHuman Synovial SarcomaUni-ZAP XRLP04HSSG HSSH HSSI HSSJ HSSKHE7T7 Week Old Early Stage Human,Uni-ZAP XRLP04subtractedHEPA HEPB HEPCHuman EpididymusUni-ZAP XRLP04HSNA HSNB HSNC HSNM HSNNHuman SynoviumUni-ZAP XRLP04HPFB HPFC HPFD HPFEHuman Prostate Cancer, Stage CUni-ZAP XRLP04fractionHE2A HE2D HE2E HE2H HE2I HE2M12 Week Old Early Stage HumanUni-ZAP XRLP04HE2N HE2OHE2B HE2C HE2F HE2G HE2P HE2Q12 Week Old Early Stage Human, IIUni-ZAP XRLP04HPTS HPTT HPTUHuman Pituitary, subtractedUni-ZAP XRLP04HAUA HAUB HAUCAmniotic Cells - TNF inducedUni-ZAP XRLP04HAQA HAQB HAQC HAQDAmniotic Cells - Primary CultureUni-ZAP XRLP04HWTA HWTB HWTCwilm's tumorUni-ZAP XRLP04HBSDBone Cancer, re-excisionUni-ZAP XRLP04HSGBSalivary gland, re-excisionUni-ZAP XRLP04HSJA HSJB HSJCSmooth muscle-ILb inducedUni-ZAP XRLP04HSXA HSXB HSXC HSXDHuman Substantia NigraUni-ZAP XRLP04HSHA HSHB HSHCSmooth muscle, IL1b inducedUni-ZAP XRLP04HOUA HOUB HOUC HOUD HOUEAdipocytesUni-ZAP XRLP04HPWA HPWB HPWC HPWD HPWEProstate BPHUni-ZAP XRLP04HELA HELB HELC HELD HELEEndothelial cells-controlUni-ZAP XRLP04HELF HELG HELHHEMA HEMB HEMC HEMD HEMEEndothelial-inducedUni-ZAP XRLP04HEMF HEMG HEMHHBIA HBIB HBICHuman Brain, StriatumUni-ZAP XRLP04HHSA HHSB HHSC HHSD HHSEHuman Hypothalmus, SchizophreniaUni-ZAP XRLP04HNGA HNGB HNGC HNGD HNGEneutrophils controlUni-ZAP XRLP04HNGF HNGG HNGH HNGI HNGJHNHA HNHB HNHC HNHD HNHENeutrophils IL-1 and LPS inducedUni-ZAP XRLP04HNHF HNHG HNHH HNHI HNHJHSDB HSDCSTRIATUM DEPRESSIONUni-ZAP XRLP04HHPTHypothalamusUni-ZAP XRLP04HSAT HSAU HSAV HSAW HSAXAnergic T-cellUni-ZAP XRLP04HSAY HSAZHBMS HBMT HBMU HBMV HBMWBone marrowUni-ZAP XRLP04HBMXHOEA HOEB HOEC HOED HOEEOsteoblastsUni-ZAP XRLP04HOEF HOEJHAIA HAIB HAIC HAID HAIE HAIFEpithelial-TNFa and INF inducedUni-ZAP XRLP04HTGA HTGB HTGC HTGDApoptotic T-cellUni-ZAP XRLP04HMCA HMCB HMCC HMCD HMCEMacrophage-oxLDLUni-ZAP XRLP04HMAA HMAB HMAC HMAD HMAEMacrophage (GM-CSF treated)Uni-ZAP XRLP04HMAF HMAGHPHANormal ProstateUni-ZAP XRLP04HPIA HPIB HPICLNCAP prostate cell lineUni-ZAP XRLP04HPJA HPJB HPJCPC3 Prostate cell lineUni-ZAP XRLP04HOSE HOSF HOSGHuman Osteoclastoma, re-excisionUni-ZAP XRLP04HTGE HTGFApoptotic T-cell, re-excisionUni-ZAP XRLP04HMAJ HMAKH Macrophage (GM-CSF treated), re-Uni-ZAP XRLP04excisionHACB HACC HACDHuman Adipose Tissue, re-excisionUni-ZAP XRLP04HFPAH. Frontal Cortex, EpilepticUni-ZAP XRLP04HFAA HFAB HFAC HFAD HFAEAlzheimer's, spongy changeUni-ZAP XRLP04HFAMFrontal Lobe, DementiaUni-ZAP XRLP04HMIA HMIB HMICHuman Manic Depression TissueUni-ZAP XRLP04HTSA HTSE HTSF HTSG HTSHHuman ThymuspBSLP05HPBA HPBB HPBC HPBD HPBEHuman Pineal GlandpBSLP05HSAA HSAB HSACHSA 172 CellspBSLP05HSBA HSBB HSBC HSBMHSC172 cellspBSLP05HJAA HJAB HJAC HJADJurkat T-cell G1 phasepBSLP05HJBA HJBB HJBC HJBDJurkat T-Cell, S phasepBSLP05HAFA HAFBAorta endothelial cells + TNF-apBSLP05HAWA HAWB HAWCHuman White AdiposepBSLP05HTNA HTNBHuman ThyroidpBSLP05HONANormal Ovary, PremenopausalpBSLP05HARA HARBHuman Adult RetinapBSLP05HLJA HLJBHuman LungpCMVSport 1LP06HOFM HOFN HOFOH. Ovarian Tumor, II, OV5232pCMVSport 2.0LP07HOGA HOGB HOGCOV 10-3-95pCMVSport 2.0LP07HCGLCD34+cells, IIpCMVSport 2.0LP07HDLAHodgkin's Lymphoma IpCMVSport 2.0LP07HDTA HDTB HDTC HDTD HDTEHodgkin's Lymphoma IIpCMVSport 2.0LP07HKAA HKAB HKAC HKAD HKAEKeratinocytepCMVSport 2.0LP07HKAF HKAG HKAHHCIMCAPFINDER, Crohn's Disease, lib 2pCMVSport 2.0LP07HKALKeratinocyte, lib 2pCMVSport 2.0LP07HKATKeratinocyte, lib 3pCMVSport 2.0LP07HNDANasal polypspCMVSport 2.0LP07HDRAH. Primary Dendritic Cells, lib 3pCMVSport 2.0LP07HOHA HOHB HOHCHuman Osteoblasts IIpCMVSport 2.0LP07HLDA HLDB HLDCLiver, HepatomapCMVSport 3.0LP08HLDN HLDO HLDPHuman Liver, normalpCMVSport 3.0LP08HMTApBMC stimulated w/poly I/CpCMVSport 3.0LP08HNTANTERA2, controlpCMVSport 3.0LP08HDPA HDPB HDPC HDPD HDPFPrimary Dendritic Cells, lib 1pCMVSport 3.0LP08HDPG HDPH HDPI HDPJ HDPKHDPM HDPN HDPO HDPPPrimary Dendritic cells, frac 2pCMVSport 3.0LP08HMUA HMUB HMUCMyoloid Progenitor Cell LinepCMVSport 3.0LP08HHEA HHEB HHEC HHEDT Cell helper IpCMVSport 3.0LP08HHEM HHEN HHEO HHEPT cell helper IIpCMVSport 3.0LP08HEQA HEQB HEQCHuman endometrial stromal cellspCMVSport 3.0LP08HJMA HJMBHuman endometrial stromal cells-pCMVSport 3.0LP08treated with progesteroneHSWA HSWB HSWCHuman endometrial stromal cells-pCMVSport 3.0LP08treated with estradiolHSYA HSYB HSYCHuman Thymus Stromal CellspCMVSport 3.0LP08HLWA HLWB HLWCHuman PlacentapCMVSport 3.0LP08HRAA HRAB HRACRejected Kidney, lib 4pCMVSport 3.0LP08HMTMPCR, pBMC I/C treatedPCRIILP09HMJAH. Meniingima, M6pSport 1LP10HMKA HMKB HMKC HMKD HMKEH Meningima, M1pSport 1LP10HUSG HUSIHuman umbilical vein endothelialpSport 1LP10cells, IL-4 inducedHUSX HUSYHuman Umbilical Vein EndothelialpSport 1LP10Cells, uninducedHOFAOvarian Tumor I, OV5232pSport 1LP10HCFA HCFB HCFC HCFDT-Cell PHA 16 hrspSport 1LP10HCFL HCFM HCFN HCFOT-Cell PHA 24 hrspSport 1LP10HADA HADC HADD HADE HADFHuman AdiposepSport 1LP10HADGHOVA HOVB HOVCHuman OvarypSport 1LP10HTWB HTWC HTWD HTWE HTWFResting T-Cell Library, IIpSport 1LP10HMMASpleen metastic melanomapSport 1LP10HLYA HLYB HLYC HLYD HLYESpleen, Chronic lymphocyticpSport 1LP10leukemiaHCGACD34+cell, IpSport 1LP10HEOM HEONHuman EosinophilspSport 1LP10HTDAHuman Tonsil, Lib 3pSport 1LP10HSPASalivary Gland, Lib 2pSport 1LP10HCHA HCHB HCHCBreast Cancer cell line, MDA 36pSport 1LP10HCHM HCHNBreast Cancer Cell line, angiogenicpSport 1LP10HCIACrohn's DiseasepSport 1LP10HDAA HDAB HDACHEL cell linepSport 1LP10HABAHuman AstrocytepSport 1LP10HUFA HUFB HUFCUlcerative ColitispSport 1LP10HNTMNTERA2 + retinoic acid, 14 dayspSport 1LP10HDQAPrimary Dendritic cells, CapFinder2,pSport 1LP10frac 1HDQMPrimary Dendritic Cells, CapFinder,pSport 1LP10frac 2HLDXHuman Liver, normal, CapFinderpSport 1LP10HULA HULB HULCHuman Dermal EndothelialpSport 1LP10Cells, untreatedHUMAHuman Dermal EndothelialpSport 1LP10cells, treatedHCJAHuman Stromal EndometrialpSport 1LP10fibroblasts, untreatedHCJMHuman Stromal endometrialpSport 1LP10fibroblasts, treated w/estradiolHEDAHuman Stromal endometrialpSport 1LP10fibroblasts, treated with progesteroneHFNAHuman ovary tumor cell OV350721pSport 1LP10HKGA HKGB HKGC HKGDMerkel CellspSport 1LP10HISA HISB HISCPancreas Islet Cell TumorpSport 1LP10HLSASkin, burnedpSport 1LP10HBZAProstate, BPH, Lib 2pSport 1LP10HBZSProstate BPH, Lib 2, subtractedpSport 1LP10HFIA HFIB HFICSynovial Fibroblasts (control)pSport 1LP10HFIH HFII HFIJSynovial hypoxiapSport 1LP10HFIT HFIU HFIVSynovial IL-1/TNF stimulatedpSport 1LP10HGCAMessangial cell, frac 1pSport 1LP10HMVA HMVB HMVCBone Marrow Stromal Cell, untreatedpSport 1LP10HFIX HFIY HFIZSynovial Fibroblasts (III/TNF), subtpSport 1LP10HFOX HFOY HFOZSynovial hypoxia-RSF subtractedpSport 1LP10HMQA HMQB HMQC HMQDHuman Activated MonocytesUni-ZAP XRLP11HLIA HLIB HLICHuman LiverpCMVSport 1LP012HHBA HHBB HHBC HHBD HHBEHuman HeartpCMVSport 1LP012HBBA HBBBHuman BrainpCMVSport 1LP012HLJA HLJB HLJC HLJD HLJEHuman LungpCMVSport 1LP012HOGA HOGB HOGCOvarian TumorpCMVSport 2.0LP012HTJMHuman Tonsils, Lib 2pCMVSport 2.0LP012HAMF HAMGKMH2pCMVSport 3.0LP012HAJA HAJB HAJCL428pCMVSport 3.0LP012HWBA HWBB HWBC HWBD HWBEDendritic cells, pooledpCMVSport 3.0LP012HWAA HWAB HWAC HWAD HWAEHuman Bone Marrow, treatedpCMVSport 3.0LP012HYAA HYAB HYACB Cell lymphomapCMVSport 3.0LP012HWHG HWHH HWHIHealing groin wound, 6.5 hours postpCMVSport 3.0LP012incisionHWHP HWHQ HWHRHealing groin wound, 7.5 hours postpCMVSport 3.0LP012incisionHARMHealing groin wound - zero hr post-pCMVSport 3.0LP012incision (control)HBIMOlfactory epithelium, nasalcavitypCMVSport 3.0LP012HWDAHealing Abdomen wound; 70&90pCMVSport 3.0LP012min post incisionHWEAHealing Abdomen Wound; 15 dayspCMVSport 3.0LP012post incisionHWJAHealing Abdomen Wound; 21&29pCMVSport 3.0LP012daysHNALHuman Tongue, frac 2pSport 1LP012HMJAH. Meniingima, M6pSport 1LP012HMKA HMKB HMKC HMKD HMKEH. Meningima, M1pSport 1LP012HOFAOvarian Tumor I, OV5232pSport 1LP012HCFA HCFB HCFC HCFDT-Cell PHA 16 hrspSport 1LP012HCFL HCFM HCFN HCFOT-Cell PHA 24 hrspSport 1LP012HMMA HMMB HMMCSpleen metastic melanomapSport 1LP012HTDAHuman Tonsil, Lib 3pSport 1LP012HDBAHuman Fetal ThymuspSport 1LP012HDUAPericardiumpSport 1LP012HBZAProstate, BPH, Lib 2pSport 1LP012HWCALarynx tumorpSport 1LP012HWKANormal lungpSport 1LP012HSMBBone marrow stroma, treatedpSport 1LP012HBHMNormal tracheapSport 1LP012HLFCHuman LarynxpSport 1LP012HLRBSiebben PolyposispSport 1LP012HNIAMammary GlandpSport 1LP012HNJBPalate carcinomapSport 1LP012HNKAPalate normalpSport 1LP012HMZAPharynx carcinomapSport 1LP012HABGCheek CarcinomapSport 1LP012HMZMPharynx CarcinomapSport 1LP012HDRMLarynx CarcinomapSport 1LP012HVAAPancreas normal PCA4 NopSport 1LP012HICATongue carcinomapSport 1LP012HUKA HUKB HUKC HUKD HUKEHuman Uterine CancerLambda ZAP IILP013HFFAHuman Fetal Brain, random primedLambda ZAP IILP013HTUAActivated T-cell labeled with 4-Lambda ZAP IILP013thioluriHBQAEarly Stage Human Brain, randomLambda ZAP IILP013primedHMEBHuman microvascular EndothelialLambda ZAP IILP013cells, fract. BHUSHHuman Umbilical Vein EndothelialLambda ZAP IILP013cells, fract. A, re-excisionHLQC HLQDHepatocellular tumor, re-excisionLambda ZAP IILP013HTWJ HTWK HTWLResting T-cell, re-excisionLambda ZAP IILP013HF6SHuman Whole 6 week Old EmbryopBluescriptLP013(II), subtHHPSHuman Hippocampus, subtractedpBluescriptLP013HLISLNCAP, differential expressionpBluescriptLP013HLHS HLHTEarly Stage Human Lung, SubtractedpBluescriptLP013HSUSSupt cells, cyclohexamide treated,pBluescriptLP013subtractedHSUTSupt cells, cyclohexamide treated,pBluescriptLP013differentially expressedHSDSH. Striatum Depression, subtractedpBluescriptLP013HPTZHuman Pituitary, Subtracted VIIpBluescriptLP013HSDXH. Striatum Depression, subt IIpBluescriptLP013HSDZH. Striatum Depression, subtpBluescriptLP013HPBA HPBB HPBC HPBD HPBEHuman Pineal GlandpBluescript SK-LP013HRTAColorectal TumorpBluescript SK-LP013HSBA HSBB HSBC HSBMHSC172 cellspBluescript SK-LP013HJAA HJAB HJAC HJADJurkat T-cell G1 phasepBluescript SK-LP013HJBA HJBB HJBC HJBDJurkat T-cell, S1 phasepBluescript SK-LP013HTNA HTNBHuman ThyroidpBluescript SK-LP013HAHA HAHBHuman Adult HeartUni-ZAP XRLP013HE6AWhole 6 week Old EmbryoUni-ZAP XRLP013HFCA HFCB HFCC HFCD HFCEHuman Fetal BrainUni-ZAP XRLP013HFKC HFKD HFKE HFKF HFKGHuman Fetal KidneyUni-ZAP XRLP013HGBA HGBD HGBE HGBF HGBGHuman Gall BladderUni-ZAP XRLP013HPRA HPRB HPRC HPRDHuman ProstateUni-ZAP XRLP013HTEA HTEB HTEC HTED HTEEHuman TestesUni-ZAP XRLP013HTTA HTTB HTTC HTTD HTTEHuman Testes TumorUni-ZAP XRLP013HYBA HYBBHuman Fetal BoneUni-ZAP XRLP013HFLAHuman Fetal LiverUni-ZAP XRLP013HHFB HHFC HHFD HHFE HHFFHuman Fetal HeartUni-ZAP XRLP013HUVB HUVC HUVD HUVEHuman Umbilical Vein, End. remakeUni-ZAP XRLP013HTHB HTHC HTHDHuman ThymusUni-ZAP XRLP013HSTA HSTB HSTC HSTDHuman Skin TumorUni-ZAP XRLP013HTAA HTAB HTAC HTAD HTAEHuman Activated T-cellsUni-ZAP XRLP013HFEA HFEB HFECHuman Fetal Epithelium (skin)Uni-ZAP XRLP013HJPA HJPB HJPC HJPDHuman Jurkat Membrane BoundUni-ZAP XRLP013PolysomesHESAHuman Epithelioid SarcomaUni-ZAP XRLP013HALSHuman Adult Liver, SubtractedUni-ZAP XRLP013HFTA HFTB HFTC HFTDHuman Fetal Dura MaterUni-ZAP XRLP013HCAA HCAB HCACCem cells, cyclohexamide treatedUni-ZAP XRLP013HRGA HRGB HRGC HRGDRaji Cells, cyclohexamide treatedUni-ZAP XRLP013HE9A HE9B HE9C HE9D HE9ENine Week Old Early Stage HumanUni-ZAP XRLP013HSFAHuman FibrosarcomaUni-ZAP XRLP013HATA HATB HATC HATD HATEHuman Adrenal Gland TumorUni-ZAP XRLP013HTRAHuman Trachea TumorUni-ZAP XRLP013HE2A HE2D HE2E HE2H HE2I12 Week Old Early Stage HumanUni-ZAP XRLP013HE2B HE2C HE2F HE2G HE2P12 Week Old Early Stage Human, IIUni-ZAP XRLP013HNEA HNEB HNEC HNED HNEEHuman NeutrophilUni-ZAP XRLP013HBGAHuman Primary Breast CancerUni-ZAP XRLP013HPTS HPTT HPTUHuman Pituitary, subtractedUni-ZAP XRLP013HMQA HMQB HMQC HMQDHuman Activated MonocytesUni-ZAP XRLP013HOAA HOAB HOACHuman OsteosarcomaUni-ZAP XRLP013HTOA HTOD HTOE HTOF HTOGhuman tonsilsUni-ZAP XRLP013HMGBHuman OB MG63 control fraction IUni-ZAP XRLP013HOPBHuman OB HOS control fraction IUni-ZAP XRLP013HOQBHuman OB HOS treated (1 nM E2)Uni-ZAP XRLP013fraction IHAUA HAUB HAUCAmniotic Cells - TNF inducedUni-ZAP XRLP013HAQA HAQB HAQC HAQDAmniotic Cells - Primary CultureUni-ZAP XRLP013HROA HROCHUMAN STOMACHUni-ZAP XRLP013HBJA HBJB HBJC HBJD HBJEHUMAN B CELL LYMPHOMAUni-ZAP XRLP013HODA HODB HODC HODDhuman ovarian cancerUni-ZAP XRLP013HCPACorpus CallosumUni-ZAP XRLP013HSOAstomach cancer (human)Uni-ZAP XRLP013HERASKINUni-ZAP XRLP013HMDABrain-medulloblastomaUni-ZAP XRLP013HGLA HGLB HGLDGlioblastomaUni-ZAP XRLP013HWTA HWTB HWTCwilm's tumorUni-ZAP XRLP013HEAAH. Atrophic EndometriumUni-ZAP XRLP013HAPN HAPO HAPP HAPQ HAPRHuman Adult Pulmonary; re-excisionUni-ZAP XRLP013HLTG HLTHHuman T-cell lymphoma; re-excisionUni-ZAP XRLP013HAHC HAHD HAHEHuman Adult Heart; re-excisionUni-ZAP XRLP013HAGA HAGB HAGC HAGD HAGEHuman AmygdalaUni-ZAP XRLP013HSJA HSJB HSJCSmooth muscle-ILb inducedUni-ZAP XRLP013HSHA HSHB HSHCSmooth muscle, IL1b inducedUni-ZAP XRLP013HPWA HPWB HPWC HPWD HPWEProstate BPHUni-ZAP XRLP013HPIA HPIB HPICLNCAP prostate cell lineUni-ZAP XRLP013HPJA HPJB HPJCPC3 Prostate cell lineUni-ZAP XRLP013HBTABone Marrow Stroma, TNF&LPS indUni-ZAP XRLP013HMCF HMCG HMCH HMCI HMCJMacrophage-oxLDL; re-excisionUni-ZAP XRLP013HAGG HAGH HAGIHuman Amygdala; re-excisionUni-ZAP XRLP013HACAH Adipose TissueUni-ZAP XRLP013HKFBK562 + PMA (36 hrs), re-excisionZAP ExpressLP013HCWT HCWU HCWVCD34 positive cells (cord blood), re-ZAP ExpressLP013exHBWAWhole brainZAP ExpressLP013HBXA HBXB HBXC HBXDHuman Whole Brain #2 - Oligo dT >ZAP ExpressLP0131.5 KbHAVMTemporal cortex-AlzheizmerpT-AdvLP014HAVTHippocampus, Alzheimer SubtractedpT-AdvLP014HHASCHME Cell LineUni-ZAP XRLP014HAJRLarynx normalpSport 1LP014HWLE HWLF HWLG HWLHColon NormalpSport 1LP014HCRM HCRN HCROColon CarcinomapSport 1LP014HWLI HWLJ HWLKColon NormalpSport 1LP014HWLQ HWLR HWLS HWLTColon TumorpSport 1LP014HBFMGastrocnemius MusclepSport 1LP014HBOD HBOEQuadriceps MusclepSport 1LP014HBKD HBKESoleus MusclepSport 1LP014HCCMPancreatic LangerhanspSport 1LP014HWGALarynx carcinomapSport 1LP014HWGM HWGNLarynx carcinomapSport 1LP014HWLA HWLB HWLCNormal colonpSport 1LP014HWLM HWLNColon TumorpSport 1LP014HVAM HVAN HVAOPancreas TumorpSport 1LP014HWGQLarynx carcinomapSport 1LP014HAQM HAQNSalivary GlandpSport 1LP014HASMStomach; normalpSport 1LP014HBCMUterus; normalpSport 1LP014HCDMTestis, normalpSport 1LP014HDJMBrain; normalpSport 1LP014HEFMAdrenal Gland, normalpSport 1LP014HBAARectum normalpSport 1LP014HFDMRectum tumorpSport 1LP014HGAMColon, normalpSport 1LP014HHMMColon, tumorpSport 1LP014HCLB HCLCHuman Lung CancerLambda Zap IILP015HRLAL1 Cell lineZAP ExpressLP015HHAMHypothalamus, Alzheimer'spCMVSport 3.0LP015HKBAKu 812F Basophils LinepSport 1LP015HS2SSaos2, Dexamethosome TreatedpSport 1LP016HA5ALung Carcinoma A549 TNF alphapSport 1LP016activatedHTFMTF-1 Cell Line GM-CSF TreatedpSport 1LP016HYASThyroid TumorpSport 1LP016HUTSLarynx NormalpSport 1LP016HXOALarynx TumorpSport 1LP016HEAHEa.hy.926 cell linepSport 1LP016HINAAdenocarcinoma HumanpSport 1LP016HRMALung MesotheliumpSport 1LP016HLCLHuman Pre-Differentiated AdipocytesUni-Zap XRLP017HS2ASaos2 CellspSport 1LP020HS2ISaos2 Cells; Vitamin D3 TreatedpSport 1LP020HUCMCHME Cell Line, untreatedpSport 1LP020HEPNAryepiglottis NormalpSport 1LP020HPSNSinus Piniformis TumorpSport 1LP020HNSAStomach NormalpSport 1LP020HNSMStomach TumorpSport 1LP020HNLALiver Normal Met5NopSport 1LP020HUTALiver Tumor Met 5 TupSport 1LP020HOCNColon NormalpSport 1LP020HOCTColon TumorpSport 1LP020HTNTTongue TumourpSport 1LP020HLXNLarynx NormalpSport 1LP020HLXTLarynx TumourpSport 1LP020HTYNThymuspSport 1LP020HPLNPlacentapSport 1LP020HTNGTongue NormalpSport 1LP020HZAAThyroid Normal (SDCA2 No)pSport 1LP020HWESThyroid ThyroiditispSport 1LP020HFHDFicolled Human Stromal Cells, 5FupTrip1Ex2LP021treatedHFHM, HFHNFicolled Human Stromal Cells,pTrip1Ex2LP021UntreatedHPCIHep G2 Cells, lambda librarylambda Zap-CMV XRLP021HBCA, HBCB, HBCCH. Lymph node breast CancerUni-ZAP XRLP021HCOKChondrocytespSPORT1LP022HDCA, HDCB, HDCCDendritic Cells From CD34 CellspSPORT1LP022HDMA, HDMBCD40 activated monocyte dendriticpSPORT1LP022cellsHDDM, HDDN, HDDOLPS activated derived dendritic cellspSPORT1LP022HPCRHep G2 Cells, PCR librarylambda Zap-CMV XRLP022HAAA, HAAB, HAACLung, Cancer (4005313A3): InvasivepSPORT1LP022Poorly Differentiated LungAdenocarcinomaHIPA, HIPB, HIPCLung, Cancer (4005163 B7):pSPORT1LP022Invasive, Poorly DiffAdenocarcinoma, MetastaticHOOH, HOOIOvary, Cancer (4004562 B6)pSPORT1LP022Papillary Serous Cystic Neoplasm,Low Malignant PotHIDALung, Normal: (4005313 B1)pSPORT1LP022HUJA, HUJB, HUJC, HUJD, HUJEB-CellspCMVSport 3.0LP022HNOA, HNOB, HNOC, HNODOvary, Normal: (9805C040R)pSPORT1LP022HNLMLung, Normal: (4005313 B1)pSPORT1LP022HSCLStromal CellspSPORT1LP022HAAXLung, Cancer: (4005313 A3) InvasivepSPORT1LP022Poorly-differentiated Metastatic lungadenocarcinomaHUUA, HUUB, HUUC, HUUDB-cells (unstimulated)pTrip1Ex2LP022HWWA, HWWB, HWWC, HWWD, HWWE,B-cells (stimulated)pSPORT1LP022HWWF, HWWGHCCCColon, Cancer: (9808C064R)pCMVSport 3.0LP023HPDO HPDP HPDQ HPDR HPDOvary, Cancer (9809C332): PoorlypSport 1LP023differentiated adenocarcinomaHPCO HPCP HPCQ HPCTOvary, Cancer (15395A1F): Grade IIpSport 1LP023Papillary CarcinomaHOCM HOCO HOCP HOCQOvary, Cancer: (15799A1F) PoorlypSport 1LP023differentiated carcinomaHCBM HCBN HCBOBreast, Cancer: (4004943 A5)pSport 1LP023HNBT HNBU HNBVBreast, Normal: (4005522B2)pSport 1LP023HBCP HBCQBreast, Cancer: (4005522 A2)pSport 1LP023HBCJBreast, Cancer: (9806C012R)pSport 1LP023HSAM HSANStromal cells 3.88pSport 1LP023HVCA HVCB HVCC HVCDOvary, Cancer: (4004332 A2)pSport 1LP023HSCK HSEN HSEOStromal cells (HBM3 18)pSport 1LP023HSCP HSCQstromal cell clone 2.5pSport 1LP023HUXABreast Cancer: (4005385 A2)pSport 1LP023HCOM HCON HCOO HCOP HCOQOvary, Cancer (4004650 A3): Well-pSport 1LP023Differentiated Micropapillary SerousCarcinomaHBNMBreast, Cancer: (9802C020E)pSport 1LP023HVVA HVVB HVVC HVVD HVVEHuman Bone Marrow, treatedpSport 1LP023

[0874] Two nonlimiting examples are provided below for isolating a particular clone from the deposited sample of plasmid cDNAs cited for that clone in Table 7. First, a plasmid is directly isolated by screening the clones using a polynucleotide probe corresponding to the nucleotide sequence of SEQ ID NO:X.

[0875] Particularly, a specific polynucleotide with 30-40 nucleotides is synthesized using an Applied Biosystems DNA synthesizer according to the sequence reported. The oligonucleotide is labeled, for instance, with 32P-γ-ATP using T4 polynucleotide kinase and purified according to routine methods. (E.g., Maniatis et al., Molecular Cloning: A Laboratory Manual, Cold Spring Harbor Press, Cold Spring, N.Y. (1982)). The plasmid mixture is transformed into a suitable host, as indicated above (such as XL-1 Blue (Stratagene)) using techniques known to those of skill in the art, such as those provided by the vector supplier or in related publications or patents cited above. The transformants are plated on 1.5% agar plates (containing the appropriate selection agent, e.g., ampicillin) to a density of about 150 transformants (colonies) per plate. These plates are screened using Nylon membranes according to routine methods for bacterial colony screening (e.g., Sambrook et al., Molecular Cloning: A Laboratory Manual, 2nd Edit., (1989), Cold Spring Harbor Laboratory Press, pages 1.93 to 1.104), or other techniques known to those of skill in the art.

[0876] Alternatively, two primers of 17-20 nucleotides derived from both ends of the nucleotide sequence of SEQ ID NO:X are synthesized and used to amplify the desired cDNA using the deposited cDNA plasmid as a template. The polymerase chain reaction is carried out under routine conditions, for instance, in 25 l of reaction mixture with 0.5 ug of the above cDNA template. A convenient reaction mixture is 1.5-5 mM MgCl2, 0.01% (w/v) gelatin, 20 μM each of dATP, dCTP, dGTP, dTTP, 25 pmol of each primer and 0.25 Unit of Taq polymerase. Thirty five cycles of PCR (denaturation at 94° C. for 1 min; annealing at 55° C. for 1 min; elongation at 72° C. for 1 min) are performed with a Perkin-Elmer Cetus automated thermal cycler. The amplified product is analyzed by agarose gel electrophoresis and the DNA band with expected molecular weight is excised and purified. The PCR product is verified to be the selected sequence by subcloning and sequencing the DNA product.

[0877] Several methods are available for the identification of the 5′ or 3′ non-coding portions of a gene which may not be present in the deposited clone. These methods include but are not limited to, filter probing, clone enrichment using specific probes, and protocols similar or identical to 5′ and 3′ “RACE” protocols which are well known in the art. For instance, a method similar to 5′ RACE is available for generating the missing 5′ end of a desired full-length transcript. (Fromont-Racine et al., Nucleic Acids Res. 21(7):1683-1684 (1993)).

[0878] Briefly, a specific RNA oligonucleotide is ligated to the 5′ ends of a population of RNA presumably containing full-length gene RNA transcripts. A primer set containing a primer specific to the ligated RNA oligonucleotide and a primer specific to a known sequence of the gene of interest is used to PCR amplify the 5′ portion of the desired full-length gene. This amplified product may then be sequenced and used to generate the full length gene.

[0879] This above method starts with total RNA isolated from the desired source, although poly-A+ RNA can be used. The RNA preparation can then be treated with phosphatase if necessary to eliminate 5′ phosphate groups on degraded or damaged RNA which may interfere with the later RNA ligase step. The phosphatase should then be inactivated and the RNA treated with tobacco acid pyrophosphatase in order to remove the cap structure present at the 5′ ends of messenger RNAs. This reaction leaves a 5′ phosphate group at the 5′ end of the cap cleaved RNA which can then be ligated to an RNA oligonucleotide using T4 RNA ligase.

[0880] This modified RNA preparation is used as a template for first strand cDNA synthesis using a gene specific oligonucleotide. The first strand synthesis reaction is used as a template for PCR amplification of the desired 5′ end using a primer specific to the ligated RNA oligonucleotide and a primer specific to the known sequence of the gene of interest. The resultant product is then sequenced and analyzed to confirm that the 5′ end sequence belongs to the desired gene.

Example 2

Isolation of Genomic Clones Corresponding to a Polynucleotide

[0881] A human genomic P1 library (Genomic Systems, Inc.) is screened by PCR using primers selected for the sequence corresponding to SEQ ID NO:X according to the method described in Example 1. (See also, Sambrook.)

Example 3

Tissue Specific Expression Analysis

[0882] The Human Genome Sciences, Inc. (HGS) database is derived from sequencing tissue and/or disease specific cDNA libraries. Libraries generated from a particular tissue are selected and the specific tissue expression pattern of EST groups or assembled contigs within these libraries is determined by comparison of the expression patterns of those groups or contigs within the entire database. ESTs and assembled contigs which show tissue specific expression are selected.

[0883] The original clone from which the specific EST sequence was generated, or in the case of an assembled contig, the clone from which the 5′ most EST sequence was generated, is obtained from the catalogued library of clones and the insert amplified by PCR using methods known in the art. The PCR product is denatured and then transferred in 96 or 384 well format to a nylon membrane (Schleicher and Scheull) generating an array filter of tissue specific clones. Housekeeping genes, maize genes, and known tissue specific genes are included on the filters. These targets can be used in signal normalization and to validate assay sensitivity. Additional targets are included to monitor probe length and specificity of hybridization.

[0884] Radioactively labeled hybridization probes are generated by first strand cDNA synthesis per the manufacturer's instructions (Life Technologies) from mRNA/RNA samples prepared from the specific tissue being analyzed (e.g., prostate, prostate cancer, ovarian, ovarian cancer, etc.). The hybridization probes are purified by gel exclusion chromatography, quantitated, and hybridized with the array filters in hybridization bottles at 65° C. overnight. The filters are washed under stringent conditions and signals are captured using a Fuji phosphorimager.

[0885] Data is extracted using AIS software and following background subtraction, signal normalization is performed. This includes a normalization of filter-wide expression levels between different experimental runs. Genes that are differentially expressed in the tissue of interest are identified.

Example 4

Chromosomal Mapping of the Polynucleotides

[0886] An oligonucleotide primer set is designed according to the sequence at the 5′ end of SEQ ID NO:X. This primer preferably spans about 100 nucleotides. This primer set is then used in a polymerase chain reaction under the following set of conditions: 30 seconds, 95° C.; 1 minute, 56° C.; 1 minute, 70° C. This cycle is repeated 32 times followed by one 5 minute cycle at 70° C. Human, mouse, and hamster DNA is used as template in addition to a somatic cell hybrid panel containing individual chromosomes or chromosome fragments (Bios, Inc). The reactions are analyzed on either 8% polyacrylamide gels or 3.5% agarose gels. Chromosome mapping is determined by the presence of an approximately 100 bp PCR fragment in the particular somatic cell hybrid.

Example 5

Bacterial Expression of a Polypeptide

[0887] A polynucleotide encoding a polypeptide of the present invention is amplified using PCR oligonucleotide primers corresponding to the 5′ and 3′ ends of the DNA sequence, as outlined in Example 1, to synthesize insertion fragments. The primers used to amplify the cDNA insert should preferably contain restriction sites, such as BamHI and XbaI, at the 5′ end of the primers in order to clone the amplified product into the expression vector. For example, BamHI and XbaI correspond to the restriction enzyme sites on the bacterial expression vector pQE-9. (Qiagen, Inc., Chatsworth, Calif.). This plasmid vector encodes antibiotic resistance (Ampr), a bacterial origin of replication (ori), an IPTG-regulatable promoter/operator (P/O), a ribosome binding site (RBS), a 6-histidine tag (6-His), and restriction enzyme cloning sites.

[0888] The pQE-9 vector is digested with BamHI and XbaI and the amplified fragment is ligated into the pQE-9 vector maintaining the reading frame initiated at the bacterial RBS. The ligation mixture is then used to transform the E. coli strain M15/rep4 (Qiagen, Inc.) which contains multiple copies of the plasmid pREP4, which expresses the lad repressor and also confers kanamycin resistance (Kanr). Transformants are identified by their ability to grow on LB plates and ampicillin/kanamycin resistant colonies are selected. Plasmid DNA is isolated and confirmed by restriction analysis.

[0889] Clones containing the desired constructs are grown overnight (O/N) in liquid culture in LB media supplemented with both Amp (100 ug/ml) and Kan (25 ug/ml). The O/N culture is used to inoculate a large culture at a ratio of 1:100 to 1:250. The cells are grown to an optical density 600 (O.D.600) of between 0.4 and 0.6. IPTG (Isopropyl-B-D-thiogalacto pyranoside) is then added to a final concentration of 1 mM. IPTG induces by inactivating the lacI repressor, clearing the P/O leading to increased gene expression.

[0890] Cells are grown for an extra 3 to 4 hours. Cells are then harvested by centrifugation (20 mins at 6000×g). The cell pellet is solubilized in the chaotropic agent 6 Molar Guanidine HCl by stirring for 3-4 hours at 4° C. The cell debris is removed by centrifugation, and the supernatant containing the polypeptide is loaded onto a nickel-nitrilo-tri-acetic acid (“Ni-NTA”) affinity resin column (available from QIAGEN, Inc., supra). Proteins with a 6×His tag bind to the Ni-NTA resin with high affinity and can be purified in a simple one-step procedure (for details see: The QIAexpressionist (1995) QIAGEN, Inc., supra).

[0891] Briefly, the supernatant is loaded onto the column in 6 M guanidine-HCl, pH 8. The column is first washed with 10 volumes of 6 M guanidine-HCl, pH 8, then washed with 10 volumes of 6 M guanidine-HCl pH 6, and finally the polypeptide is eluted with 6 M guanidine-HCl, pH 5.

[0892] The purified protein is then renatured by dialyzing it against phosphate-buffered saline (PBS) or 50 mM Na-acetate, pH 6 buffer plus 200 mM NaCl. Alternatively, the protein can be successfully refolded while immobilized on the Ni-NTA column. The recommended conditions are as follows: renature using a linear 6M-1M urea gradient in 500 mM NaCl, 20% glycerol, 20 mM Tris/HCl pH 7.4, containing protease inhibitors. The renaturation should be performed over a period of 1.5 hours or more. After renaturation the proteins are eluted by the addition of 250 mM immidazole. Immidazole is removed by a final dialyzing step against PBS or 50 mM sodium acetate pH 6 buffer plus 200 mM NaCl. The purified protein is stored at 4° C. or frozen at −80° C.

[0893] In addition to the above expression vector, the present invention further includes an expression vector, called pHE4a (ATCC Accession Number 209645, deposited on Feb. 25, 1998) which contains phage operator and promoter elements operatively linked to a polynucleotide of the present invention, called pHE4a. (ATCC Accession Number 209645, deposited on Feb. 25, 1998.) This vector contains: 1) a neomycinphosphotransferase gene as a selection marker, 2) an E. coli origin of replication, 3) a T5 phage promoter sequence, 4) two lac operator sequences, 5) a Shine-Delgarno sequence, and 6) the lactose operon repressor gene (lacIq). The origin of replication (oriC) is derived from pUC19 (LTI, Gaithersburg, Md.). The promoter and operator sequences are made synthetically.

[0894] DNA can be inserted into the pHE4a by restricting the vector with NdeI and XbaI, BamHI, XhoI, or Asp718, running the restricted product on a gel, and isolating the larger fragment (the stuffer fragment should be about 310 base pairs). The DNA insert is generated according to the PCR protocol described in Example 1, using PCR primers having restriction sites for NdeI (5′ primer) and XbaI, BamHI, XhoI, or Asp718 (3′ primer). The PCR insert is gel purified and restricted with compatible enzymes. The insert and vector are ligated according to standard protocols.

[0895] The engineered vector could easily be substituted in the above protocol to express protein in a bacterial system.

Example 6

Purification of a Polypeptide from an Inclusion Body

[0896] The following alternative method can be used to purify a polypeptide expressed in E coli when it is present in the form of inclusion bodies. Unless otherwise specified, all of the following steps are conducted at 4-10° C.

[0897] Upon completion of the production phase of the E. coli fermentation, the cell culture is cooled to 4-10° C. and the cells harvested by continuous centrifugation at 15,000 rpm (Heraeus Sepatech). On the basis of the expected yield of protein per unit weight of cell paste and the amount of purified protein required, an appropriate amount of cell paste, by weight, is suspended in a buffer solution containing 100 mM Tris, 50 mM EDTA, pH 7.4. The cells are dispersed to a homogeneous suspension using a high shear mixer.

[0898] The cells are then lysed by passing the solution through a microfluidizer (Microfuidics, Corp. or APV Gaulin, Inc.) twice at 4000-6000 psi. The homogenate is then mixed with NaCl solution to a final concentration of 0.5 M NaCl, followed by centrifugation at 7000×g for 15 min. The resultant pellet is washed again using 0.5M NaCl, 100 mM Tris, 50 mM EDTA, pH 7.4.

[0899] The resulting washed inclusion bodies are solubilized with 1.5 M guanidine hydrochloride (GuHCl) for 2-4 hours. After 7000×g centrifugation for 15 min., the pellet is discarded and the polypeptide containing supernatant is incubated at 4° C. overnight to allow further GuHCl extraction.

[0900] Following high speed centrifugation (30,000×g) to remove insoluble particles, the GuHCl solubilized protein is refolded by quickly mixing the GuHCl extract with 20 volumes of buffer containing 50 mM sodium, pH 4.5, 150 mM NaCl, 2 mM EDTA by vigorous stirring. The refolded diluted protein solution is kept at 4° C. without mixing for 12 hours prior to further purification steps.

[0901] To clarify the refolded polypeptide solution, a previously prepared tangential filtration unit equipped with 0.16 μm membrane filter with appropriate surface area (e.g., Filtron), equilibrated with 40 mM sodium acetate, pH 6.0 is employed. The filtered sample is loaded onto a cation exchange resin (e.g., Poros HS-50, Perseptive Biosystems). The column is washed with 40 mM sodium acetate, pH 6.0 and eluted with 250 mM, 500 mM, 1000 mM, and 1500 mM NaCl in the same buffer, in a stepwise manner. The absorbance at 280 nm of the effluent is continuously monitored. Fractions are collected and further analyzed by SDS-PAGE.

[0902] Fractions containing the polypeptide are then pooled and mixed with 4 volumes of water. The diluted sample is then loaded onto a previously prepared set of tandem columns of strong anion (Poros HQ-50, Perseptive Biosystems) and weak anion (Poros CM-20, Perseptive Biosystems) exchange resins. The columns are equilibrated with 40 mM sodium acetate, pH 6.0. Both columns are washed with 40 mM sodium acetate, pH 6.0, 200 mM NaCl. The CM-20 column is then eluted using a 10 column volume linear gradient ranging from 0.2 M NaCl, 50 mM sodium acetate, pH 6.0 to 1.0 M NaCl, 50 mM sodium acetate, pH 6.5. Fractions are collected under constant A280 monitoring of the effluent. Fractions containing the polypeptide (determined, for instance, by 16% SDS-PAGE) are then pooled.

[0903] The resultant polypeptide should exhibit greater than 95% purity after the above refolding and purification steps. No major contaminant bands should be observed from Commassie blue stained 16% SDS-PAGE gel when 5 μg of purified protein is loaded. The purified protein can also be tested for endotoxin/LPS contamination, and typically the LPS content is less than 0.1 ng/ml according to LAL assays.

Example 7

Cloning and Expression of a Polypeptide in a Baculovirus Expression System

[0904] In this example, the plasmid shuttle vector pA2 is used to insert a polynucleotide into a baculovirus to express a polypeptide. This expression vector contains the strong polyhedrin promoter of the Autographa californica nuclear polyhedrosis virus (AcMNPV) followed by convenient restriction sites such as BamHI, Xba I and Asp718. The polyadenylation site of the simian virus 40 (“SV40”) is used for efficient polyadenylation. For easy selection of recombinant virus, the plasmid contains the beta-galactosidase gene from E. coli under control of a weak Drosophila promoter in the same orientation, followed by the polyadenylation signal of the polyhedrin gene. The inserted genes are flanked on both sides by viral sequences for cell-mediated homologous recombination with wild-type viral DNA to generate a viable virus that express the cloned polynucleotide.

[0905] Many other baculovirus vectors can be used in place of the vector above, such as pAc373, pVL941, and pAcIM1, as one skilled in the art would readily appreciate, as long as the construct provides appropriately located signals for transcription, translation, secretion and the like, including a signal peptide and an in-frame AUG as required. Such vectors are described, for instance, in Luckow et al., Virology 170:31-39 (1989).

[0906] Specifically, the cDNA sequence contained in the deposited clone, including the AUG initiation codon, is amplified using the PCR protocol described in Example 1. If a naturally occurring signal sequence is used to produce the polypeptide of the present invention, the pA2 vector does not need a second signal peptide. Alternatively, the vector can be modified (pA2 GP) to include a baculovirus leader sequence, using the standard methods described in Summers et al., “A Manual of Methods for Baculovirus Vectors and Insect Cell Culture Procedures,” Texas Agricultural Experimental Station Bulletin No. 1555 (1987).

[0907] The amplified fragment is isolated from a 1% agarose gel using a commercially available kit (“Geneclean,” BIO 101 Inc., La Jolla, Calif.). The fragment then is digested with appropriate restriction enzymes and again purified on a 1% agarose gel.

[0908] The plasmid is digested with the corresponding restriction enzymes and optionally, can be dephosphorylated using calf intestinal phosphatase, using routine procedures known in the art. The DNA is then isolated from a 1% agarose gel using a commercially available kit (“Geneclean” BIO 101 Inc., La Jolla, Calif.).

[0909] The fragment and the dephosphorylated plasmid are ligated together with T4 DNA ligase. E. coli HB101 or other suitable E. coli hosts such as XL-1 Blue (Stratagene Cloning Systems, La Jolla, Calif.) cells are transformed with the ligation mixture and spread on culture plates. Bacteria containing the plasmid are identified by digesting DNA from individual colonies and analyzing the digestion product by gel electrophoresis. The sequence of the cloned fragment is confirmed by DNA sequencing.

[0910] Five μg of a plasmid containing the polynucleotide is co-transfected with 1.0 μg of a commercially available linearized baculovirus DNA (“BaculoGold™ baculovirus DNA, Pharmingen, San Diego, Calif.), using the lipofection method described by Felgner et al., Proc. Natl. Acad. Sci. USA 84:7413-7417 (1987). One μg of BaculoGold™ virus DNA and 5 μg of the plasmid are mixed in a sterile well of a microtiter plate containing 50 μl of serum-free Grace's medium (Life Technologies Inc., Gaithersburg, Md.). Afterwards, 10 μl Lipofectin plus 90 μl Grace's medium are added, mixed and incubated for 15 minutes at room temperature. Then the transfection mixture is added drop-wise to Sf9 insect cells (ATCC CRL 1711) seeded in a 35 mm tissue culture plate with 1 ml Grace's medium without serum. The plate is then incubated for 5 hours at 27° C. The transfection solution is then removed from the plate and 1 ml of Grace's insect medium supplemented with 10% fetal calf serum is added. Cultivation is then continued at 27° C. for four days.

[0911] After four days the supernatant is collected and a plaque assay is performed, as described by Summers and Smith, supra. An agarose gel with “Blue Gal” (Life Technologies Inc., Gaithersburg) is used to allow easy identification and isolation of gal-expressing clones, which produce blue-stained plaques. (A detailed description of a “plaque assay” of this type can also be found in the user's guide for insect cell culture and baculovirology distributed by Life Technologies Inc., Gaithersburg, page 9-10.) After appropriate incubation, blue stained plaques are picked with the tip of a micropipettor (e.g., Eppendorf). The agar containing the recombinant viruses is then resuspended in a microcentrifuge tube containing 200 μl of Grace's medium and the suspension containing the recombinant baculovirus is used to infect Sf9 cells seeded in 35 mm dishes. Four days later the supernatants of these culture dishes are harvested and then they are stored at 4° C.

[0912] To verify the expression of the polypeptide, Sf9 cells are grown in Grace's medium supplemented with 10% heat-inactivated FBS. The cells are infected with the recombinant baculovirus containing the polynucleotide at a multiplicity of infection (“MOI”) of about 2. If radiolabeled proteins are desired, 6 hours later the medium is removed and is replaced with SF900 II medium minus methionine and cysteine (available from Life Technologies Inc., Rockville, Md.). After 42 hours, 5 μCi of 35S-methionine and 5 μCi 35S-cysteine (available from Amersham) are added. The cells are further incubated for 16 hours and then are harvested by centrifugation. The proteins in the supernatant as well as the intracellular proteins are analyzed by SDS-PAGE followed by autoradiography (if radiolabeled).

[0913] Microsequencing of the amino acid sequence of the amino terminus of purified protein may be used to determine the amino terminal sequence of the produced protein.

Example 8

Expression of a Polypeptide in Mammalian Cells

[0914] The polypeptide of the present invention can be expressed in a mammalian cell. A typical mammalian expression vector contains a promoter element, which mediates the initiation of transcription of mRNA, a protein coding sequence, and signals required for the termination of transcription and polyadenylation of the transcript. Additional elements include enhancers, Kozak sequences and intervening sequences flanked by donor and acceptor sites for RNA splicing. Highly efficient transcription is achieved with the early and late promoters from SV40, the long terminal repeats (LTRs) from Retroviruses, e.g., RSV, HTLVI, HIVI and the early promoter of the cytomegalovirus (CMV). However, cellular elements can also be used (e.g., the human actin promoter).

[0915] Suitable expression vectors for use in practicing the present invention include, for example, vectors such as pSVL and pMSG (Pharmacia, Uppsala, Sweden), pRSVcat (ATCC 37152), pSV2dhfr (ATCC 37146), pBC12MI (ATCC 67109), pCMVSport 2.0, and pCMVSport 3.0. Mammalian host cells that could be used include, human Hela, 293, H9 and Jurkat cells, mouse NIH3T3 and C127 cells, Cos 1, Cos 7 and CV1, quail QC1-3 cells, mouse L cells and Chinese hamster ovary (CHO) cells.

[0916] Alternatively, the polypeptide can be expressed in stable cell lines containing the polynucleotide integrated into a chromosome. The co-transfection with a selectable marker such as DHFR, gpt, neomycin, or hygromycin allows the identification and isolation of the transfected cells.

[0917] The transfected gene can also be amplified to express large amounts of the encoded protein. The DHFR (dihydrofolate reductase) marker is useful in developing cell lines that carry several hundred or even several thousand copies of the gene of interest. (See, e.g., Alt, F. W., et al., J. Biol. Chem. 253:1357-1370 (1978); Hamlin, J. L. and Ma, C., Biochem. et Biophys. Acta, 1097:107-143 (1990); Page, M. J. and Sydenham, M. A., Biotechnology 9:64-68 (1991)). Another useful selection marker is the enzyme glutamine synthase (GS) (Murphy et al., Biochem J. 227:277-279 (1991); Bebbington et al., Bio/Technology 10:169-175 (1992). Using these markers, the mammalian cells are grown in selective medium and the cells with the highest resistance are selected. These cell lines contain the amplified gene(s) integrated into a chromosome. Chinese hamster ovary (CHO) and NSO cells are often used for the production of proteins.

[0918] Derivatives of the plasmid pSV2-dhfr (ATCC Accession No. 37146), the expression vectors pC4 (ATCC Accession No. 209646) and pC6 (ATCC Accession No.209647) contain the strong promoter (LTR) of the Rous Sarcoma Virus (Cullen et al., Molecular and Cellular Biology, 438-447 (March, 1985)) plus a fragment of the CMV-enhancer (Boshart et al., Cell 41:521-530 (1985)). Multiple cloning sites, e.g., with the restriction enzyme cleavage sites BamHI, XbaI and Asp718, facilitate the cloning of the gene of interest. The vectors also contain the 3′ intron, the polyadenylation and termination signal of the rat preproinsulin gene, and the mouse DHFR gene under control of the SV40 early promoter.

[0919] Specifically, the plasmid pC6, for example, is digested with appropriate restriction enzymes and then dephosphorylated using calf intestinal phosphates by procedures known in the art. The vector is then isolated from a 1% agarose gel.

[0920] A polynucleotide of the present invention is amplified according to the protocol outlined in Example 1. If a naturally occurring signal sequence is used to produce the polypeptide of the present invention, the vector does not need a second signal peptide. Alternatively, if a naturally occurring signal sequence is not used, the vector can be modified to include a heterologous signal sequence. (See, e.g., International Publication No. WO 96/34891.)

[0921] The amplified fragment is isolated from a 1% agarose gel using a commercially available kit (“Geneclean,” BIO 101 Inc., La Jolla, Calif.). The fragment then is digested with appropriate restriction enzymes and again purified on a 1% agarose gel.

[0922] The amplified fragment is then digested with the same restriction enzyme and purified on a 1% agarose gel. The isolated fragment and the dephosphorylated vector are then ligated with T4 DNA ligase. E. coli HB101 or XL-1 Blue cells are then transformed and bacteria are identified that contain the fragment inserted into plasmid pC6 using, for instance, restriction enzyme analysis.

[0923] Chinese hamster ovary cells lacking an active DHFR gene is used for transfection. Five μg of the expression plasmid pC6 or pC4 is cotransfected with 0.5 μg of the plasmid pSVneo using lipofectin (Felgner et al., supra). The plasmid pSV2-neo contains a dominant selectable marker, the neo gene from Tn5 encoding an enzyme that confers resistance to a group of antibiotics including G418. The cells are seeded in alpha minus MEM supplemented with 1 mg/ml G418. After 2 days, the cells are trypsinized and seeded in hybridoma cloning plates (Greiner, Germany) in alpha minus MEM supplemented with 10, 25, or 50 ng/ml of methotrexate plus 1 mg/ml G418. After about 10-14 days single clones are trypsinized and then seeded in 6-well petri dishes or 10 ml flasks using different concentrations of methotrexate (50 nM, 100 nM, 200 nM, 400 nM, 800 nM). Clones growing at the highest concentrations of methotrexate are then transferred to new 6-well plates containing even higher concentrations of methotrexate (1 μM, 2 μM, 5 μM, 10 mM, 20 mM). The same procedure is repeated until clones are obtained which grow at a concentration of 100-200 μM. Expression of the desired gene product is analyzed, for instance, by SDS-PAGE and Western blot or by reversed phase HPLC analysis.

Example 9

Protein Fusions

[0924] The polypeptides of the present invention are preferably fused to other proteins. These fusion proteins can be used for a variety of applications. For example, fusion of the present polypeptides to His-tag, HA-tag, protein A, IgG domains, and maltose binding protein facilitates purification. (See Example 5; see also EP A 394,827; Traunecker, et al., Nature 331:84-86 (1988)). Similarly, fusion to IgG-1, IgG-3, and albumin increases the halflife time in vivo. Nuclear localization signals fused to the polypeptides of the present invention can target the protein to a specific subcellular localization, while covalent heterodimer or homodimers can increase or decrease the activity of a fusion protein. Fusion proteins can also create chimeric molecules having more than one function. Finally, fusion proteins can increase solubility and/or stability of the fused protein compared to the non-fused protein. All of the types of fusion proteins described above can be made by modifying the following protocol, which outlines the fusion of a polypeptide to an IgG molecule, or the protocol described in Example 5.

[0925] Briefly, the human Fc portion of the IgG molecule can be PCR amplified, using primers that span the 5′ and 3′ ends of the sequence described below. These primers also should have convenient restriction enzyme sites that will facilitate cloning into an expression vector, preferably a mammalian expression vector.

[0926] For example, if pC4 (ATCC Accession No. 209646) is used, the human Fc portion can be ligated into the BamHI cloning site. Note that the 3′ BamHI site should be destroyed. Next, the vector containing the human Fc portion is re-restricted with BamHI, linearizing the vector, and a polynucleotide of the present invention, isolated by the PCR protocol described in Example 1, is ligated into this BamHI site. Note that the polynucleotide is cloned without a stop codon, otherwise a fusion protein will not be produced.

[0927] If the naturally occurring signal sequence is used to produce the polypeptide of the present invention, pC4 does not need a second signal peptide. Alternatively, if the naturally occurring signal sequence is not used, the vector can be modified to include a heterologous signal sequence. (See, e.g., International Publication No. WO 96/34891.)

11Human IgG Fc region(SEQ ID NO: 1)GGGATCCGGAGCCCAAATCTTCTGACAAAACTCACACATGCCCACCGTGCCCAGCACCTGAATTCGAGGGTGCACCGTCAGTCTTCCTCTTCCCCCCAAAACCCAAGGACACCCTCATGATCTCCCGGACTCCTGAGGTCACATGCGTGGTGGTGGACGTAAGCCACGAAGACCCTGAGGTCAAGTTCAACTGGTACGTGGACGGCGTGGAGGTGCATAATGCCAAGACAAAGCCGCGGGAGGAGCAGTACAACAGCACGTACCGTGTGGTCAGCGTCCTCACCGTCCTGCACCAGGACTGGCTGAATGGCAAGGAGTACAAGTGCAAGGTCTCCAACAAAGCCCTCCCAACCCCCATCGAGAAAACCATCTCCAAAGCCAAAGGGCAGCCCCGAGAACCACAGGTGTACACCCTGCCCCCATCCCGGGATGAGCTGACCAAGAACCAGGTCAGCCTGACCTGCCTGGTCAAAGGCTTCTATCCAAGCGACATCGCCGTGGAGTGGGAGAGCAATGGGCAGCCGGAGAACAACTACAAGACCACGCCTCCCGTGCTGGACTCCGACGGCTCCTTCTTCCTCTACAGCAAGCTCACCGTGGACAAGAGCAGGTGGCAGCAGGGGAACGTCTTCTCATGCTCCGTGATGCATGAGGCTCTGCACAACCACTACACGCAGAAGAGCCTCTCCCTGTCTCCGGGTAAATGAGTGCGACGGCCGCGACTCTAGAGGAT

Example 10

Production of an Antibody from a Polypeptide

[0928] a) Hybridoma Technology

[0929] The antibodies of the present invention can be prepared by a variety of methods. (See, Current Protocols, Chapter 2.) As one example of such methods, cells expressing a polypeptide of the present invention are administered to an animal to induce the production of sera containing polyclonal antibodies. In a preferred method, a preparation of a a polypeptide of the present invention is prepared and purified to render it substantially free of natural contaminants. Such a preparation is then introduced into an animal in order to produce polyclonal antisera of greater specific activity.

[0930] Monoclonal antibodies specific for a polypeptide of the present invention are prepared using hybridoma technology (Kohler et al., Nature 256:495 (1975); Kohler et al., Eur. J. Immunol. 6:511 (1976); Kohler et al., Eur. J. Immunol. 6:292 (1976); Hammerling et al., in: Monoclonal Antibodies and T-Cell Hybridomas, Elsevier, N.Y., pp. 563-681 (1981)). In general, an animal (preferably a mouse) is immunized with a polypeptide of the present invention or, more preferably, with a secreted polypeptide of the present invention-expressing cell. Such polypeptide-expressing cells are cultured in any suitable tissue culture medium, preferably in Earle's modified Eagle's medium supplemented with 10% fetal bovine serum (inactivated at about 56° C.), and supplemented with about 10 g/l of nonessential amino acids, about 1,000 U/ml of penicillin, and about 100 μg/ml of streptomycin.

[0931] The splenocytes of such mice are extracted and fused with a suitable myeloma cell line. Any suitable myeloma cell line may be employed in accordance with the present invention; however, it is preferable to employ the parent myeloma cell line (SP20), available from the ATCC. After fusion, the resulting hybridoma cells are selectively maintained in HAT medium, and then cloned by limiting dilution as described by Wands et al. (Gastroenterology 80:225-232 (1981)). The hybridoma cells obtained through such a selection are then assayed to identify clones which secrete antibodies capable of binding the polypeptide of the present invention.

[0932] Alternatively, additional antibodies capable of binding to polypeptide of the present invention can be produced in a two-step procedure using anti-idiotypic antibodies. Such a method makes use of the fact that antibodies are themselves antigens, and therefore, it is possible to obtain an antibody which binds to a second antibody. In accordance with this method, protein specific antibodies are used to immunize an animal, preferably a mouse. The splenocytes of such an animal are then used to produce hybridoma cells, and the hybridoma cells are screened to identify clones which produce an antibody whose ability to bind to the polypeptide of the present invention-specific antibody can be blocked by polypeptide of the present invention. Such antibodies comprise anti-idiotypic antibodies to the polypeptide of the present invention-specific antibody and are used to immunize an animal to induce formation of further polypeptide of the present invention-specific antibodies.

[0933] For in vivo use of antibodies in humans, an antibody is “humanized”. Such antibodies can be produced using genetic constructs derived from hybridoma cells producing the monoclonal antibodies described above. Methods for producing chimeric and humanized antibodies are known in the art and are discussed herein. (See, for review, Morrison, Science 229:1202 (1985); Oi et al., BioTechniques 4:214 (1986); Cabilly et al., U.S. Pat. No. 4,816,567; Taniguchi et al., EP 171496; Morrison et al., EP 173494; Neuberger et al., WO 8601533; Robinson et al., International Publication No. WO 8702671; Boulianne et al., Nature 312:643 (1984); Neuberger et al., Nature 314:268 (1985)).

[0934] b) Isolation of Antibody Fragments Directed Against Polypeptide of the Present Invention from a Library of scFvs

[0935] Naturally occurring V-genes isolated from human PBLs are constructed into a library of antibody fragments which contain reactivities against polypeptide of the present invention to which the donor may or may not have been exposed (see e.g., U.S. Pat. No. 5,885,793 incorporated herein by reference in its entirety).

[0936] Rescue of the Library. A library of scFvs is constructed from the RNA of human PBLs as described in International Publication No. WO 92/01047. To rescue phage displaying antibody fragments, approximately 109 E. coli harboring the phagemid are used to inoculate 50 ml of 2×TY containing 1% glucose and 100 μg/ml of ampicillin (2×TY-AMP-GLU) and grown to an O.D. of 0.8 with shaking. Five ml of this culture is used to inoculate 50 ml of 2×TY-AMP-GLU, 2×108 TU of delta gene 3 helper (M13 delta gene III, see International Publication No. WO 92/01047) are added and the culture incubated at 37° C. for 45 minutes without shaking and then at 37° C. for 45 minutes with shaking. The culture is centrifuged at 4000 r.p.m. for 10 min. and the pellet resuspended in 2 liters of 2×TY containing 100 μg/ml ampicillin and 50 ug/ml kanamycin and grown overnight. Phage are prepared as described in International Publication No. WO 92/01047.

[0937] M13 delta gene III is prepared as follows: M13 delta gene III helper phage does not encode gene III protein, hence the phage(mid) displaying antibody fragments have a greater avidity of binding to antigen. Infectious M13 delta gene III particles are made by growing the helper phage in cells harboring a pUC19 derivative supplying the wild type gene III protein during phage morphogenesis. The culture is incubated for 1 hour at 37° C. without shaking and then for a further hour at 37° C. with shaking. Cells are spun down (IEC-Centra 8,400 r.p.m. for 10 min), resuspended in 300 ml 2×TY broth containing 100 μg ampicillin/ml and 25 μg kanamycin/ml (2×TY-AMP-KAN) and grown overnight, shaking at 37° C. Phage particles are purified and concentrated from the culture medium by two PEG-precipitations (Sambrook et al., 1990), resuspended in 2 ml PBS and passed through a 0.45 μm filter (Minisart NML; Sartorius) to give a final concentration of approximately 1013 transducing units/ml (ampicillin-resistant clones).

[0938] Panning of the Library. Immunotubes (Nunc) are coated overnight in PBS with 4 ml of either 100 μg/ml or 10 μg/ml of a polypeptide of the present invention. Tubes are blocked with 2% Marvel-PBS for 2 hours at 37° C. and then washed 3 times in PBS. Approximately 1013 TU of phage is applied to the tube and incubated for 30 minutes at room temperature tumbling on an over and under turntable and then left to stand for another 1.5 hours. Tubes are washed 10 times with PBS 0.1% Tween-20 and 10 times with PBS. Phage are eluted by adding 1 ml of 100 mM triethylamine and rotating 15 minutes on an under and over turntable after which the solution is immediately neutralized with 0.5 ml of 1.0M Tris-HCl, pH 7.4. Phage are then used to infect 10 ml of mid-log E. coli TG1 by incubating eluted phage with bacteria for 30 minutes at 37° C. The E. coli are then plated on TYE plates containing 1% glucose and 100 μg/ml ampicillin. The resulting bacterial library is then rescued with delta gene 3 helper phage as described above to prepare phage for a subsequent round of selection. This process is then repeated for a total of 4 rounds of affinity purification with tube-washing increased to 20 times with PBS, 0.1% Tween-20 and 20 times with PBS for rounds 3 and 4.

[0939] Characterization of Binders. Eluted phage from the 3rd and 4th rounds of selection are used to infect E. coli HB 2151 and soluble scFv is produced (Marks, et al., 1991) from single colonies for assay. ELISAs are performed with microtitre plates coated with either 10 pg/ml of the polypeptide of the present invention in 50 mM bicarbonate pH 9.6. Clones positive in ELISA are further characterized by PCR fingerprinting (see, e.g., International Publication No. WO 92/01047) and then by sequencing. These ELISA positive clones may also be further characterized by techniques known in the art, such as, for example, epitope mapping, binding affinity, receptor signal transduction, ability to block or competitively inhibit antibody/antigen binding, and competitive agonistic or antagonistic activity.

Example 11

Method of Determining Alterations in a Gene Corresponding to a Polynucleotide

[0940] RNA isolated from entire families or individual patients presenting with a phenotype of interest (such as a disease) is isolated. cDNA is then generated from these RNA samples using protocols known in the art. (See, Sambrook.) The cDNA is then used as a template for PCR, employing primers surrounding regions of interest in SEQ ID NO:X; and/or the nucleotide sequence of the cDNA contained in Clone ID NO:Z. Suggested PCR conditions consist of 35 cycles at 95 degrees C. for 30 seconds; 60-120 seconds at 52-58 degrees C.; and 60-120 seconds at 70 degrees C., using buffer solutions described in Sidransky et al., Science 252:706 (1991).

[0941] PCR products are then sequenced using primers labeled at their 5′ end with T4 polynucleotide kinase, employing SequiTherm Polymerase (Epicentre Technologies). The intron-exon boundaries of selected exons is also determined and genomic PCR products analyzed to confirm the results. PCR products harboring suspected mutations are then cloned and sequenced to validate the results of the direct sequencing.

[0942] PCR products are cloned into T-tailed vectors as described in Holton et al., Nucleic Acids Research, 19:1156 (1991) and sequenced with T7 polymerase (United States Biochemical). Affected individuals are identified by mutations not present in unaffected individuals.

[0943] Genomic rearrangements are also observed as a method of determining alterations in a gene corresponding to a polynucleotide. Genomic clones isolated according to Example 2 are nick-translated with digoxigenindeoxy-uridine 5′-triphosphate (Boehringer Manheim), and FISH performed as described in Johnson et al., Methods Cell Biol. 35:73-99 (1991). Hybridization with the labeled probe is carried out using a vast excess of human cot-1 DNA for specific hybridization to the corresponding genomic locus.

[0944] Chromosomes are counterstained with 4,6-diamino-2-phenylidole and propidium iodide, producing a combination of C- and R-bands. Aligned images for precise mapping are obtained using a triple-band filter set (Chroma Technology, Brattleboro, Vt.) in combination with a cooled charge-coupled device camera (Photometrics, Tucson, Ariz.) and variable excitation wavelength filters. (Johnson et al., Genet. Anal. Tech. Appl., 8:75 (1991)). Image collection, analysis and chromosomal fractional length measurements are performed using the ISee Graphical Program System. (Inovision Corporation, Durham, N.C.) Chromosome alterations of the genomic region hybridized by the probe are identified as insertions, deletions, and translocations. These alterations are used as a diagnostic marker for an associated disease.

Example 12

Method of Detecting Abnormal Levels of a Polypeptide in a Biological Sample

[0945] A polypeptide of the present invention can be detected in a biological sample, and if an increased or decreased level of the polypeptide is detected, this polypeptide is a marker for a particular phenotype. Methods of detection are numerous, and thus, it is understood that one skilled in the art can modify the following assay to fit their particular needs.

[0946] For example, antibody-sandwich ELISAs are used to detect polypeptides in a sample, preferably a biological sample. Wells of a microtiter plate are coated with specific antibodies, at a final concentration of 0.2 to 10 ug/ml. The antibodies are either monoclonal or polyclonal and are produced by the method described in Example 10. The wells are blocked so that non-specific binding of the polypeptide to the well is reduced.

[0947] The coated wells are then incubated for >2 hours at RT with a sample containing the polypeptide. Preferably, serial dilutions of the sample should be used to validate results. The plates are then washed three times with deionized or distilled water to remove unbound polypeptide.

[0948] Next, 50 ul of specific antibody-alkaline phosphatase conjugate, at a concentration of 25-400 ng, is added and incubated for 2 hours at room temperature. The plates are again washed three times with deionized or distilled water to remove unbound conjugate.

[0949] Add 75 ul of 4-methylumbelliferyl phosphate (MUP) or p-nitrophenyl phosphate (NPP) substrate solution to each well and incubate 1 hour at room temperature. Measure the reaction by a microtiter plate reader. Prepare a standard curve, using serial dilutions of a control sample, and plot polypeptide concentration on the X-axis (log scale) and fluorescence or absorbance of the Y-axis (linear scale). Interpolate the concentration of the polypeptide in the sample using the standard curve.

Example 13

Formulation

[0950] The invention also provides methods of treatment and/or prevention of diseases or disorders (such as, for example, any one or more of the diseases or disorders disclosed herein) by administration to a subject of an effective amount of a Therapeutic. By therapeutic is meant polynucleotides or polypeptides of the invention (including fragments and variants), agonists or antagonists thereof, and/or antibodies thereto, in combination with a pharmaceutically acceptable carrier type (e.g., a sterile carrier).

[0951] The Therapeutic will be formulated and dosed in a fashion consistent with good medical practice, taking into account the clinical condition of the individual patient (especially the side effects of treatment with the Therapeutic alone), the site of delivery, the method of administration, the scheduling of administration, and other factors known to practitioners. The “effective amount” for purposes herein is thus determined by such considerations.

[0952] As a general proposition, the total pharmaceutically effective amount of the Therapeutic administered parenterally per dose will be in the range of about 1 ug/kg/day to 10 mg/kg/day of patient body weight, although, as noted above, this will be subject to therapeutic discretion. More preferably, this dose is at least 0.01 mg/kg/day, and most preferably for humans between about 0.01 and 1 mg/kg/day for the hormone. If given continuously, the Therapeutic is typically administered at a dose rate of about 1 ug/kg/hour to about 50 ug/kg/hour, either by 1-4 injections per day or by continuous subcutaneous infusions, for example, using a mini-pump. An intravenous bag solution may also be employed. The length of treatment needed to observe changes and the interval following treatment for responses to occur appears to vary depending on the desired effect.

[0953] Therapeutics can be are administered orally, rectally, parenterally, intracistemally, intravaginally, intraperitoneally, topically (as by powders, ointments, gels, drops or transdermal patch), bucally, or as an oral or nasal spray. “Pharmaceutically acceptable carrier” refers to a non-toxic solid, semisolid or liquid filler, diluent, encapsulating material or formulation auxiliary of any. The term “parenteral” as used herein refers to modes of administration which include intravenous, intramuscular, intraperitoneal, intrastemal, subcutaneous and intraarticular injection and infusion.

[0954] Therapeutics of the invention are also suitably administered by sustained-release systems. Suitable examples of sustained-release Therapeutics are administered orally, rectally, parenterally, intracistemally, intravaginally, intraperitoneally, topically (as by powders, ointments, gels, drops or transdermal patch), bucally, or as an oral or nasal spray. “Pharmaceutically acceptable carrier” refers to a non-toxic solid, semisolid or liquid filler, diluent, encapsulating material or formulation auxiliary of any type. The term “parenteral” as used herein refers to modes of administration which include intravenous, intramuscular, intraperitoneal, intrastemal, subcutaneous and intraarticular injection and infusion.

[0955] Therapeutics of the invention are also suitably administered by sustained-release systems. Suitable examples of sustained-release Therapeutics include suitable polymeric materials (such as, for example, semi-permeable polymer matrices in the form of shaped articles, e.g., films, or mirocapsules), suitable hydrophobic materials (for example as an emulsion in an acceptable oil) or ion exchange resins, and sparingly soluble derivatives (such as, for example, a sparingly soluble salt).

[0956] Sustained-release matrices include polylactides (U.S. Pat. No. 3,773,919, EP 58,481), copolymers of L-glutamic acid and gamma-ethyl-L-glutamate (Sidman et al., Biopolymers 22:547-556 (1983)), poly (2-hydroxyethyl methacrylate) (Langer et al., J. Biomed. Mater. Res. 15:167-277 (1981), and Langer, Chem. Tech. 12:98-105 (1982)), ethylene vinyl acetate (Langer et al., Id.) or poly-D-(−)-3-hydroxybutyric acid (EP 133,988).

[0957] Sustained-release Therapeutics also include liposomally entrapped Therapeutics of the invention (see generally, Langer, Science 249:1527-1533 (1990); Treat et al., in Liposomes in the Therapy of Infectious Disease and Cancer, Lopez-Berestein and Fidler (eds.), Liss, New York, pp. 317-327 and 353-365 (1989)). Liposomes containing the Therapeutic are prepared by methods known per se: DE 3,218,121; Epstein et al., Proc. Natl. Acad. Sci. (USA) 82:3688-3692 (1985); Hwang et al., Proc. Natl. Acad. Sci.(USA) 77:4030-4034 (1980); EP 52,322; EP 36,676; EP 88,046; EP 143,949; EP 142,641; Japanese Pat. Appl. 83-118008; U.S. Pat. Nos. 4,485,045 and 4,544,545; and EP 102,324. Ordinarily, the liposomes are of the small (about 200-800 Angstroms) unilamellar type in which the lipid content is greater than about 30 mol. percent cholesterol, the selected proportion being adjusted for the optimal Therapeutic.

[0958] In yet an additional embodiment, the Therapeutics of the invention are delivered by way of a pump (see Langer, supra; Sefton, CRC Crit. Ref. Biomed. Eng. 14:201 (1987); Buchwald et al., Surgery 88:507 (1980); Saudek et al., N. Engl. J. Med. 321:574 (1989)).

[0959] Other controlled release systems are discussed in the review by Langer (Science 249:1527-1533 (1990)).

[0960] For parenteral administration, in one embodiment, the Therapeutic is formulated generally by mixing it at the desired degree of purity, in a unit dosage injectable form (solution, suspension, or emulsion), with a pharmaceutically acceptable carrier, i.e., one that is non-toxic to recipients at the dosages and concentrations employed and is compatible with other ingredients of the formulation. For example, the formulation preferably does not include oxidizing agents and other compounds that are known to be deleterious to the Therapeutic.

[0961] Generally, the formulations are prepared by contacting the Therapeutic uniformly and intimately with liquid carriers or finely divided solid carriers or both. Then, if necessary, the product is shaped into the desired formulation. Preferably the carrier is a parenteral carrier, more preferably a solution that is isotonic with the blood of the recipient. Examples of such carrier vehicles include water, saline, Ringer's solution, and dextrose solution. Non-aqueous vehicles such as fixed oils and ethyl oleate are also useful herein, as well as liposomes.

[0962] The carrier suitably contains minor amounts of additives such as substances that enhance isotonicity and chemical stability. Such materials are non-toxic to recipients at the dosages and concentrations employed, and include buffers such as phosphate, citrate, succinate, acetic acid, and other organic acids or their salts; antioxidants such as ascorbic acid; low molecular weight (less than about ten residues) polypeptides, e.g., polyarginine or tripeptides; proteins, such as serum albumin, gelatin, or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids, such as glycine, glutamic acid, aspartic acid, or arginine; monosaccharides, disaccharides, and other carbohydrates including cellulose or its derivatives, glucose, manose, or dextrins; chelating agents such as EDTA; sugar alcohols such as mannitol or sorbitol; counterions such as sodium; and/or nonionic surfactants such as polysorbates, poloxamers, or PEG.

[0963] The Therapeutic is typically formulated in such vehicles at a concentration of about 0.1 mg/ml to 100 mg/ml, preferably 1-10 mg/ml, at a pH of about 3 to 8. It will be understood that the use of certain of the foregoing excipients, carriers, or stabilizers will result in the formation of polypeptide salts.

[0964] Any pharmaceutical used for therapeutic administration can be sterile. Sterility is readily accomplished by filtration through sterile filtration membranes (e.g., 0.2 micron membranes). Therapeutics generally are placed into a container having a sterile access port, for example, an intravenous solution bag or vial having a stopper pierceable by a hypodermic injection needle.

[0965] Therapeutics ordinarily will be stored in unit or multi-dose containers, for example, sealed ampoules or vials, as an aqueous solution or as a lyophilized formulation for reconstitution. As an example of a lyophilized formulation, 10-ml vials are filled with 5 ml of sterile-filtered 1% (w/v) aqueous Therapeutic solution, and the resulting mixture is lyophilized. The infusion solution is prepared by reconstituting the lyophilized Therapeutic using bacteriostatic Water-for-Injection.

[0966] The invention also provides a pharmaceutical pack or kit comprising one or more containers filled with one or more of the ingredients of the Therapeutics of the invention. Associated with such container(s) can be a notice in the form prescribed by a governmental agency regulating the manufacture, use or sale of pharmaceuticals or biological products, which notice reflects approval by the agency of manufacture, use or sale for human administration. In addition, the Therapeutics may be employed in conjunction with other therapeutic compounds.

[0967] The Therapeutics of the invention may be administered alone or in combination with adjuvants. Adjuvants that may be administered with the Therapeutics of the invention include, but are not limited to, alum, alum plus deoxycholate (ImmunoAg), MTP-PE (Biocine Corp.), QS21 (Genentech, Inc.), BCG (e.g., THERACYS®), MPL and nonviable prepartions of Corynebacterium parvum. In a specific embodiment, Therapeutics of the invention are administered in combination with alum. In another specific embodiment, Therapeutics of the invention are administered in combination with QS-21. Further adjuvants that may be administered with the Therapeutics of the invention include, but are not limited to, Monophosphoryl lipid immunomodulator, AdjuVax lOa, QS-21, QS-18, CRL1005, Aluminum salts, MF-59, and Virosomal adjuvant technology. Vaccines that may be administered with the Therapeutics of the invention include, but are not limited to, vaccines directed toward protection against MMR (measles, mumps, rubella), polio, varicella, tetanus/diptheria, hepatitis A, hepatitis B, haemophilus influenzae B, whooping cough, pneumonia, influenza, Lyme's Disease, rotavirus, cholera, yellow fever, Japanese encephalitis, poliomyelitis, rabies, typhoid fever, and pertussis. Combinations may be administered either concomitantly, e.g., as an admixture, separately but simultaneously or concurrently; or sequentially. This includes presentations in which the combined agents are administered together as a therapeutic mixture, and also procedures in which the combined agents are administered separately but simultaneously, e.g., as through separate intravenous lines into the same individual. Administration “in combination” further includes the separate administration of one of the compounds or agents given first, followed by the second.

[0968] The Therapeutics of the invention may be administered alone or in combination with other therapeutic agents. Therapeutic agents that may be administered in combination with the Therapeutics of the invention, include but are not limited to psychotherapeutic agents (e.g., antipsychotic agents, antimanic agents, antidepressants, antianxiety agents, and stimulants), anti-epileptics, antiparkinsonian agents, ALS therapeutics, steroidal and non-steroidal anti-inflammatory agents, anticaogulation and thrombolytic agents, chemotherapeutic agents, cytokines and/or growth factors, antiviral agents, members of the TNF family, antibiotics, and/or conventional immunotherapeutic agents. Combinations may be administered either concomitantly, e.g., as an admixture, separately but simultaneously or concurrently; or sequetially. This includes presentations in which the combined agents are administered together as a therapeutic mixture, and also procedures in which the combined agents are administered separately but simultaneously, e.g., as through separate intravenous lines into the same individual. Administration “in combination” further includes the separate administration of one of the compounds or agents given first, followed by the second.

[0969] In one embodiment, the Therapeutics of the invention are administered in combination with an anticoagulant. Anticoagulants that may be administered with the compositions of the invention include, but are not limited to, heparin, low molecular weight heparin, warfarin sodium (e.g., COUMADIN®), dicumarol, 4-hydroxycoumarin, anisindione (e.g., MIRADON™), acenocoumarol (e.g., nicoumalone, SINTHROME™), indan-1,3-dione, phenprocoumon (e.g., MARCUMAR™), ethyl biscoumacetate (e.g., TROMEXAN™), and aspirin. In a specific embodiment, compositions of the invention are administered in combination with heparin and/or warfarin. In another specific embodiment, compositions of the invention are administered in combination with warfarin. In another specific embodiment, compositions of the invention are administered in combination with warfarin and aspirin. In another specific embodiment, compositions of the invention are administered in combination with heparin. In another specific embodiment, compositions of the invention are administered in combination with heparin and aspirin.

[0970] In another embodiment, the Therapeutics of the invention are administered in combination with thrombolytic drugs. Thrombolytic drugs that may be administered with the compositions of the invention include, but are not limited to, plasminogen, lys-plasminogen, alpha2-antiplasmin, streptokinae (e.g., KABIKINASE™), antiresplace (e.g., EMINASE™), tissue plasminogen activator (t-PA, altevase, ACTIVASE™), urokinase (e.g., ABBOKINASE™), sauruplase, (Prourokinase, single chain urokinase), and aminocaproic acid (e.g., AMICAR™). In a specific embodiment, compositions of the invention are administered in combination with tissue plasminogen activator and aspirin.

[0971] In another embodiment, the Therapeutics of the invention are administered in combination with antiplatelet drugs. Antiplatelet drugs that may be administered with the compositions of the invention include, but are not limited to, aspirin, dipyridamole (e.g., PERSANTINE™), and ticlopidine (e.g., TICLID™).

[0972] In specific embodiments, the use of anti-coagulants, thrombolytic and/or antiplatelet drugs in combination with Therapeutics of the invention is contemplated for the prevention, diagnosis, and/or treatment of thrombosis, arterial thrombosis, venous thrombosis, thromboembolism, pulmonary embolism, atherosclerosis, myocardial infarction, transient ischemic attack, unstable angina. In specific embodiments, the use of anticoagulants, thrombolytic drugs and/or antiplatelet drugs in combination with Therapeutics of the invention is contemplated for the prevention of occulsion of saphenous grafts, for reducing the risk of periprocedural thrombosis as might accompany angioplasty procedures, for reducing the risk of stroke in patients with atrial fibrillation including nonrheumatic atrial fibrillation, for reducing the risk of embolism associated with mechanical heart valves and or mitral valves disease. Other uses for the therapeutics of the invention, alone or in combination with antiplatelet, anticoagulant, and/or thrombolytic drugs, include, but are not limited to, the prevention of occlusions in extracorporeal devices (e.g., intravascular canulas, vascular access shunts in hemodialysis patients, hemodialysis machines, and cardiopulmonary bypass machines).

[0973] In certain embodiments, Therapeutics of the invention are administered in combination with antiretroviral agents, nucleoside/nucleotide reverse transcriptase inhibitors (NRTIs), non-nucleoside reverse transcriptase inhibitors (NNRTIs), and/or protease inhibitors (PIs). NRTIs that may be administered in combination with the Therapeutics of the invention, include, but are not limited to, RETROVIR™ (zidovudine/AZT), VIDEX™ (didanosine/ddI), HIVID™ (zalcitabine/ddC), ZERIT™ (stavudine/d4T), EPIVIR™ (lamivudine/3TC), and COMBIVIR™ (zidovudine/lamivudine). NNRTIs that may be administered in combination with the Therapeutics of the invention, include, but are not limited to, VIRAMUNE™ (nevirapine), RESCRIPTOR™ (delavirdine), and SUSTIVA™ (efavirenz). Protease inhibitors that may be administered in combination with the Therapeutics of the invention, include, but are not limited to, CRIXIVAN™ (indinavir), NORVIR™ (ritonavir), INVIRASE™ (saquinavir), and VIRACEPT™ (nelfinavir). In a specific embodiment, antiretroviral agents, nucleoside reverse transcriptase inhibitors, non-nucleoside reverse transcriptase inhibitors, and/or protease inhibitors may be used in any combination with Therapeutics of the invention to treat AIDS and/or to prevent or treat HIV infection.

[0974] Additional NRTIs include LODENOSINE™ (F-ddA; an acid-stable adenosine NRTI; Triangle/Abbott; COVIRACIL™ (emtricitabine/FTC; structurally related to lamivudine (3TC) but with 3- to 10-fold greater activity in vitro; Triangle/Abbott); dOTC (BCH-10652, also structurally related to lamivudine but retains activity against a substantial proportion of lamivudine-resistant isolates; Biochem Pharma); Adefovir (refused approval for anti-HIV therapy by FDA; Gilead Sciences); PREVEON® (Adefovir Dipivoxil, the active prodrug of adefovir; its active form is PMEA-pp); TENOFOVIR™ (bis-POC PMPA, a PMPA prodrug; Gilead); DAPD/DXG (active metabolite of DAPD; Triangle/Abbott); D-D4FC (related to 3TC, with activity against AZT/3TC-resistant virus); GW420867X (Glaxo Wellcome); ZIAGEN™ (abacavir/159U89; Glaxo Wellcome Inc.); CS-87 (3′azido-2′,3′-dideoxyuridine; WO 99/66936); and S-acyl-2-thioethyl (SATE)-bearing prodrug forms of β-L-FD4C and β-L-FddC (WO 98/17281).

[0975] Additional NNRTIs include COACTINON™ (Emivirine/MKC-442, potent NNRTI of the HEPT class; Triangle/Abbott); CAPRAVIRINE™ (AG-1549/S-1153, a next generation NNRTI with activity against viruses containing the K103N mutation; Agouron); PNU-142721 (has 20- to 50-fold greater activity than its predecessor delavirdine and is active against K103N mutants; Pharmacia & Upjohn); DPC-961 and DPC-963 (second-generation derivatives of efavirenz, designed to be active against viruses with the K103N mutation; DuPont); GW-420867×(has 25-fold greater activity than HBY097 and is active against K103N mutants; Glaxo Wellcome); CALANOLIDE A (naturally occurring agent from the latex tree; active against viruses containing either or both the Y181 C and K103N mutations); and Propolis (WO 99/49830).

[0976] Additional protease inhibitors include LOPINAVIR™ (ABT378/r; Abbott Laboratories); BMS-232632 (an azapeptide; Bristol-Myres Squibb); TIPRANAVIR™ (PNU-140690, a non-peptic dihydropyrone; Pharmacia & Upjohn); PD-178390 (a nonpeptidic dihydropyrone; Parke-Davis); BMS 232632 (an azapeptide; Bristol-Myers Squibb); L-756,423 (an indinavir analog; Merck); DMP-450 (a cyclic urea compound; Avid & DuPont); AG-1776 (a peptidomimetic with in vitro activity against protease inhibitor-resistant viruses; Agouron); VX-175/GW-433908 (phosphate prodrug of amprenavir; Vertex & Glaxo Welcome); CGP61755 (Ciba); and AGENERASE™ (amprenavir; Glaxo Wellcome Inc.).

[0977] Additional antiretroviral agents include fusion inhibitors/gp41 binders. Fusion inhibitors/gp41 binders include T-20 (a peptide from residues 643-678 of the HIV gp41 transmembrane protein ectodomain which binds to gp41 in its resting state and prevents transformation to the fusogenic state; Trimeris) and T-1249 (a second-generation fusion inhibitor; Trimeris).

[0978] Additional antiretroviral agents include fusion inhibitors/chemokine receptor antagonists. Fusion inhibitors/chemokine receptor antagonists include CXCR4 antagonists such as AMD 3100 (a bicyclam), SDF-1 and its analogs, and ALX40-4C (a cationic peptide), T22 (an 18 amino acid peptide; Trimeris) and the T22 analogs T134 and T140; CCR5 antagonists such as RANTES (9-68), AOP-RANTES, NNY-RANTES, and TAK-779; and CCR5/CXCR4 antagonists such as NSC 651016 (a distamycin analog). Also included are CCR2B, CCR3, and CCR6 antagonists. Chemokine recpetor agonists such as RANTES, SDF-1, MIP-1α, MIP-1β, etc., may also inhibit fusion.

[0979] Additional antiretroviral agents include integrase inhibitors. Integrase inhibitors include dicaffeoylquinic (DFQA) acids; L-chicoric acid (a dicaffeoyltartaric (DCTA) acid); quinalizarin (QLC) and related anthraquinones; ZINTEVIR™ (AR 177, an oligonucleotide that probably acts at cell surface rather than being a true integrase inhibitor; Arondex); and naphthols such as those disclosed in WO 98150347.

[0980] Additional antiretroviral agents include hydroxyurea-like compunds such as BCX-34 (a purine nucleoside phosphorylase inhibitor; Biocryst); ribonucleotide reductase inhibitors such as DIDOX™ (Molecules for Health); inosine monophosphate dehydrogenase (IMPDH) inhibitors sucha as VX-497 (Vertex); and mycopholic acids such as CellCept (mycophenolate mofetil; Roche).

[0981] Additional antiretroviral agents include inhibitors of viral integrase, inhibitors of viral genome nuclear translocation such as arylene bis(methylketone) compounds; inhibitors of HIV entry such as AOP-RANTES, NNY-RANTES, RANTES-IgG fusion protein, soluble complexes of RANTES and glycosaminoglycans (GAG), and AMD-3100; nucleocapsid zinc finger inhibitors such as dithiane compounds; targets of HIV Tat and Rev; and pharmacoenhancers such as ABT-378.

[0982] Other antiretroviral therapies and adjunct therapies include cytokines and lymphokines such as MIP-1α, MIP-1β, SDF-1α, IL-2, PROLEUKIN™ (aldesleukin/L2-7001; Chiron), IL-4, IL-10, IL-12, and IL-13; interferons such as IFN-α2a; antagonists of TNFs, NFκB, GM-CSF, M-CSF, and IL-10; agents that modulate immune activation such as cyclosporin and prednisone; vaccines such as Remune™ (HIV Immunogen), APL 400-003 (Apollon), recombinant gp120 and fragments, bivalent (B/E) recombinant envelope glycoprotein, rgp120CM235, MN rgp120, SF-2 rgp120, gp120/soluble CD4 complex, Delta JR-FL protein, branched synthetic peptide derived from discontinuous gp120 C3/C4 domain, fusion-competent immunogens, and Gag, Pol, Nef, and Tat vaccines; gene-based therapies such as genetic suppressor elements (GSEs; WO 98/54366), and intrakines (genetically modified CC chemokines targetted to the ER to block surface expression of newly synthesized CCR5 (Yang et al., PNAS 94:11567-72 (1997); Chen et al., Nat. Med. 3:1110-16 (1997)); antibodies such as the anti-CXCR4 antibody 12G5, the anti-CCR5 antibodies 2D7, 5C7, PA8, PA9, PA10, PA11, PA12, and PA14, the anti-CD4 antibodies Q4120 and RPA-T4, the anti-CCR3 antibody 7B11, the anti-gp120 antibodies 17b, 48d, 447-52D, 257-D, 268-D and 50.1, anti-Tat antibodies, anti-TNF-α antibodies, and monoclonal antibody 33A; aryl hydrocarbon (AH) receptor agonists and antagonists such as TCDD, 3,3′,4,4′,5-pentachlorobiphenyl, 3,3′,4,4′-tetrachlorobiphenyl, and α-naphthoflavone (WO 98/30213); and antioxidants such as γ-L-glutamyl-L-cysteine ethyl ester (γ-GCE; WO 99/56764).

[0983] In a further embodiment, the Therapeutics of the invention are administered in combination with an antiviral agent. Antiviral agents that may be administered with the Therapeutics of the invention include, but are not limited to, acyclovir, ribavirin, amantadine, and remantidine.

[0984] In other embodiments, Therapeutics of the invention may be administered in combination with anti-opportunistic infection agents. Anti-opportunistic agents that may be administered in combination with the Therapeutics of the invention, include, but are not limited to, TRIMETHOPRIM-SULFAMETHOXAZOLE™, DAPSONE™, PENTAMIDINE™, ATOVAQUONE™, ISONIAZID™ RIFAMPIN™, PYRAZINAMIDE™, ETHAMBUTOL™, RIFABUTIN™, CLARITHROMYCIN™, AZITHROMYCIN™, GANCICLOVIR™, FOSCARNET™, CIDOFOVIR™, FLUCONAZOLE™, ITRACONAZOLE™, KETOCONAZOLE™, ACYCLOVIR™, FAMCICOLVIR™, PYRIMETHAMINE™, LEUCOVORIN™, NEUPOGEN™ (filgrastim/G-CSF), and LEUKINE™ (sargramostim/GM-CSF). In a specific embodiment, Therapeutics of the invention are used in any combination with TRIMETHOPRIM-SULFAMETHOXAZOLE™, DAPSONE™, PENTAMIDINE™, and/or ATOVAQUONE™ to prophylactically treat or prevent an opportunistic Pneumocystis carinii pneumonia infection. In another specific embodiment, Therapeutics of the invention are used in any combination with ISONLAZID™, RIFAMPIN™, PYRAZINAMIDE™, and/or ETHAMBUTOL™ to prophylactically treat or prevent an opportunistic Mycobacterium avium complex infection. In another specific embodiment, Therapeutics of the invention are used in any combination with RIFABUTIN™, CLARITHROMYCIN™, and/or AZITHROMYCIN™ to prophylactically treat or prevent an opportunistic Mycobacterium tuberculosis infection. In another specific embodiment, Therapeutics of the invention are used in any combination with GANCICLOVIR™, FOSCARNET™, and/or CIDOFOVIR™ to prophylactically treat or prevent an opportunistic cytomegalovirus infection. In another specific embodiment, Therapeutics of the invention are used in any combination with FLUCONAZOLE™, ITRACONAZOLE™, and/or KETOCONAZOLE™ to prophylactically treat or prevent an opportunistic fungal infection. In another specific embodiment, Therapeutics of the invention are used in any combination with ACYCLOVIR™ and/or FAMCICOLVIR™ to prophylactically treat or prevent an opportunistic herpes simplex virus type I and/or type II infection. In another specific embodiment, Therapeutics of the invention are used in any combination with PYRIMETHAMINE™ and/or LEUCOVORIN™ to prophylactically treat or prevent an opportunistic Toxoplasma gondii infection. In another specific embodiment, Therapeutics of the invention are used in any combination with LEUCOVORIN™ and/or NEUPOGEN™ to prophylactically treat or prevent an opportunistic bacterial infection.

[0985] In a further embodiment, the Therapeutics of the invention are administered in combination with an antibiotic agent. Antibiotic agents that may be administered with the Therapeutics of the invention include, but are not limited to, amoxicillin, beta-lactamases, aminoglycosides, beta-lactam (glycopeptide), beta-lactamases, Clindamycin, chloramphenicol, cephalosporins, ciprofloxacin, erythromycin, fluoroquinolones, macrolides, metronidazole, penicillins, quinolones, rapamycin, rifampin, streptomycin, sulfonamide, tetracyclines, trimethoprim, trimethoprim-sulfamethoxazole, and vancomycin.

[0986] In other embodiments, the Therapeutics of the invention are administered in combination with immunestimulants. Immunostimulants that may be administered in combination with the Therapeutics of the invention include, but are not limited to, levamisole (e.g., ERGAMISOL™), isoprinosine (e.g. INOSIPLEX™), interferons (e.g. interferon alpha), and interleukins (e.g., IL-2).

[0987] In other embodiments, Therapeutics of the invention are administered in combination with immunosuppressive agents. Immunosuppressive agents that may be administered in combination with the Therapeutics of the invention include, but are not limited to, steroids, cyclosporine, cyclosporine analogs, cyclophosphamide methylprednisone, prednisone, azathioprine, FK-506, 15-deoxyspergualin, and other immunosuppressive agents that act by suppressing the function of responding T cells. Other immunosuppressive agents that may be administered in combination with the Therapeutics of the invention include, but are not limited to, prednisolone, methotrexate, thalidomide, methoxsalen, rapamycin, leflunomide, mizoribine (BREDININ™), brequinar, deoxyspergualin, and azaspirane (SKF 105685), ORTHOCLONE OKT® 3 (muromonab-CD3), SANDIMMUNE™, NEORAL™, SANGDYA™ (cyclosporine), PROGRAF® (FK506, tacrolimus), CELLCEPT® (mycophenolate motefil, of which the active metabolite is mycophenolic acid), IMURAN™ (azathioprine), glucocorticosteroids, adrenocortical steroids such as DELTASONE™ (prednisone) and HYDELTRASOL™ (prednisolone), FOLEX™ and MEXATE™ (methotrxate), OXSORALEN-ULTRA™ (methoxsalen) and RAPAMUNE™ (sirolimus). In a specific embodiment, immunosuppressants may be used to prevent rejection of organ or bone marrow transplantation.

[0988] In an additional embodiment, Therapeutics of the invention are administered alone or in combination with one or more intravenous immune globulin preparations. Intravenous immune globulin preparations that may be administered with the Therapeutics of the invention include, but not limited to, GAMMAR™, IVEEGAM™, SANDOGLOBULIN™, GAMMAGARD S/D™, ATGAM™ (antithymocyte glubulin), and GAMIMUNE™. In a specific embodiment, Therapeutics of the invention are administered in combination with intravenous immune globulin preparations in transplantation therapy (e.g., bone marrow transplant).

[0989] In certain embodiments, the Therapeutics of the invention are administered alone or in combination with an anti-inflammatory agent. Anti-inflammatory agents that may be administered with the Therapeutics of the invention include, but are not limited to, corticosteroids (e.g. betamethasone, budesonide, cortisone, dexamethasone, hydrocortisone, methylprednisolone, prednisolone, prednisone, and triamcinolone), nonsteroidal anti-inflammatory drugs (e.g., diclofenac, diflunisal, etodolac, fenoprofen, floctafenine, flurbiprofen, ibuprofen, indomethacin, ketoprofen, meclofenamate, mefenamic acid, meloxicam, nabumetone, naproxen, oxaprozin, phenylbutazone, piroxicam, sulindac, tenoxicam, tiaprofenic acid, and tolmetin.), as well as antihistamines, aminoarylcarboxylic acid derivatives, arylacetic acid derivatives, arylbutyric acid derivatives, arylcarboxylic acids, arylpropionic acid derivatives, pyrazoles, pyrazolones, salicylic acid derivatives, thiazinecarboxamides, e-acetamidocaproic acid, S-adenosylmethionine, 3-amino-4-hydroxybutyric acid, amixetrine, bendazac, benzydamine, bucolome, difenpiramide, ditazol, emorfazone, guaiazulene, nabumetone, nimesulide, orgotein, oxaceprol, paranyline, perisoxal, pifoxime, proquazone, proxazole, and tenidap.

[0990] In an additional embodiment, the compositions of the invention are administered alone or in combination with an anti-angiogenic agent. Anti-angiogenic agents that may be administered with the compositions of the invention include, but are not limited to, Angiostatin (Entremed, Rockville, Md.), Troponin-1 (Boston Life Sciences, Boston, Mass.), anti-Invasive Factor, retinoic acid and derivatives thereof, paclitaxel (Taxol), Suramin, Tissue Inhibitor of Metalloproteinase-1, Tissue Inhibitor of Metalloproteinase-2, VEGI, Plasminogen Activator Inhibitor-1, Plasminogen Activator Inhibitor-2, and various forms of the lighter “d group” transition metals.

[0991] Lighter “d group” transition metals include, for example, vanadium, molybdenum, tungsten, titanium, niobium, and tantalum species. Such transition metal species may form transition metal complexes. Suitable complexes of the above-mentioned transition metal species include oxo transition metal complexes.

[0992] Representative examples of vanadium complexes include oxo vanadium complexes such as vanadate and vanadyl complexes. Suitable vanadate complexes include metavanadate and orthovanadate complexes such as, for example, ammonium metavanadate, sodium metavanadate, and sodium orthovanadate. Suitable vanadyl complexes include, for example, vanadyl acetylacetonate and vanadyl sulfate including vanadyl sulfate hydrates such as vanadyl sulfate mono- and trihydrates.

[0993] Representative examples of tungsten and molybdenum complexes also include oxo complexes. Suitable oxo tungsten complexes include tungstate and tungsten oxide complexes. Suitable tungstate complexes include ammonium tungstate, calcium tungstate, sodium tungstate dihydrate, and tungstic acid. Suitable tungsten oxides include tungsten (IV) oxide and tungsten (VI) oxide. Suitable oxo molybdenum complexes include molybdate, molybdenum oxide, and molybdenyl complexes. Suitable molybdate complexes include ammonium molybdate and its hydrates, sodium molybdate and its hydrates, and potassium molybdate and its hydrates. Suitable molybdenum oxides include molybdenum (VI) oxide, molybdenum (VI) oxide, and molybdic acid. Suitable molybdenyl complexes include, for example, molybdenyl acetylacetonate. Other suitable tungsten and molybdenum complexes include hydroxo derivatives derived from, for example, glycerol, tartaric acid, and sugars.

[0994] A wide variety of other anti-angiogenic factors may also be utilized within the context of the present invention. Representative examples include, but are not limited to, platelet factor 4; protamine sulphate; sulphated chitin derivatives (prepared from queen crab shells), (Murata et al., Cancer Res. 51:22-26, (1991)); Sulphated Polysaccharide Peptidoglycan Complex (SP-PG) (the function of this compound may be enhanced by the presence of steroids such as estrogen, and tamoxifen citrate); Staurosporine; modulators of matrix metabolism, including for example, proline analogs, cishydroxyproline, d,L-3,4-dehydroproline, Thiaproline, alpha,alpha-dipyridyl, aminopropionitrile fumarate; 4-propyl-5-(4-pyridinyl)-2(3H)-oxazolone; Methotrexate; Mitoxantrone; Heparin; Interferons; 2 Macroglobulin-serum; ChIMP-3 (Pavloff et al., J. Bio. Chem. 267:17321-17326, (1992)); Chymostatin (Tomkinson et al., Biochem J. 286:475-480, (1992)); Cyclodextrin Tetradecasulfate; Eponemycin; Camptothecin; Fumagillin (Ingber et al., Nature 348:555-557, (1990)); Gold Sodium Thiomalate (“GST”; Matsubara and Ziff, J. Clin. Invest. 79:1440-1446, (1987)); anticollagenase-serum; alpha2-antiplasmin (Holmes et al., J. Biol. Chem. 262(4):1659-1664, (1987)); Bisantrene (National Cancer Institute); Lobenzarit disodium (N-(2)-carboxyphenyl-4-chloroanthronilic acid disodium or “CCA”; (Takeuchi et al., Agents Actions 36:312-316, (1992)); and metalloproteinase inhibitors such as BB94.

[0995] Additional anti-angiogenic factors that may also be utilized within the context of the present invention include Thalidomide, (Celgene, Warren, N.J.); Angiostatic steroid; AGM-1470 (H. Brem and J. Folkman J Pediatr. Surg. 28:445-51 (1993)); an integrin alpha v beta 3 antagonist (C. Storgard et al., J. Clin. Invest. 103:47-54 (1999)); carboxynaminolmidazole; Carboxyamidotriazole (CAI) (National Cancer Institute, Bethesda, Md.); Conbretastatin A-4 (CA4P) (OXiGENE, Boston, Mass.); Squalamine (Magainin Pharmaceuticals, Plymouth Meeting, Pa.); TNP-470, (Tap Pharmaceuticals, Deerfield, Ill.); ZD-0101 AstraZeneca (London, UK); APRA (CT2584); Benefin, Byrostatin-1 (SC339555); CGP-41251 (PKC 412); CM101; Dexrazoxane (ICRF187); DMXAA; Endostatin; Flavopridiol; Genestein; GTE; lInmTher; Iressa (ZD1839); Octreotide (Somatostatin); Panretin; Penacillamine; Photopoint; PI-88; Prinomastat (AG-3340) Purlytin; Suradista (FCE26644); Tamoxifen (Nolvadex); Tazarotene; Tetrathiomolybdate; Xeloda (Capecitabine); and 5-Fluorouracil.

[0996] Anti-angiogenic agents that may be administed in combination with the compounds of the invention may work through a variety of mechanisms including, but not limited to, inhibiting proteolysis of the extracellular matrix, blocking the function of endothelial cell-extracellular matrix adhesion molecules, by antagonizing the function of angiogenesis inducers such as growth factors, and inhibiting integrin receptors expressed on proliferating endothelial cells. Examples of anti-angiogenic inhibitors that interfere with extracellular matrix proteolysis and which may be administered in combination with the compositons of the invention include, but are not lmited to, AG-3340 (Agouron, La Jolla, Calif.), BAY-12-9566 (Bayer, West Haven, Conn.), BMS-275291 (Bristol Myers Squibb, Princeton, N.J.), CGS-27032A (Novartis, East Hanover, N.J.), Marimastat (British Biotech, Oxford, UK), and Metastat (Aeterna, St-Foy, Quebec). Examples of anti-angiogenic inhibitors that act by blocking the function of endothelial cell-extracellular matrix adhesion molecules and which may be administered in combination with the compositons of the invention include, but are not Imited to, EMD-121974 (Merck KcgaA Darmstadt, Germany) and Vitaxin (Ixsys, La Jolla, Calif./Medimmune, Gaithersburg, Md.). Examples of anti-angiogenic agents that act by directly antagonizing or inhibiting angiogenesis inducers and which may be administered in combination with the compositons of the invention include, but are not lmited to, Angiozyme (Ribozyme, Boulder, Colo.), Anti-VEGF antibody (Genentech, S. San Francisco, Calif.), PTK-787/ZK-225846 (Novartis, Basel, Switzerland), SU-101 (Sugen, S. San Francisco, Calif.), SU-5416 (Sugen/Pharmacia Upjohn, Bridgewater, N.J.), and SU-6668 (Sugen). Other anti-angiogenic agents act to indirectly inhibit angiogenesis. Examples of indirect inhibitors of angiogenesis which may be administered in combination with the compositons of the invention include, but are not limited to, IM-862 (Cytran, Kirkland, Wash.), Interferon-alpha, IL-12 (Roche, Nutley, N.J.), and Pentosan polysulfate (Georgetown University, Washington, D.C.).

[0997] In particular embodiments, the use of compositions of the invention in combination with anti-angiogenic agents is contemplated for the treatment, prevention, and/or amelioration of an autoimmune disease, such as for example, an autoimmune disease described herein.

[0998] In a particular embodiment, the use of compositions of the invention in combination with anti-angiogenic agents is contemplated for the treatment, prevention, and/or amelioration of arthritis. In a more particular embodiment, the use of compositions of the invention in combination with anti-angiogenic agents is contemplated for the treatment, prevention, and/or amelioration of rheumatoid arthritis.

[0999] In another embodiment, the polynucleotides encoding a polypeptide of the present invention are administered in combination with an angiogenic protein, or polynucleotides encoding an angiogenic protein. Examples of angiogenic proteins that may be administered with the compositions of the invention include, but are not limited to, acidic and basic fibroblast growth factors, VEGF-1, VEGF-2, VEGF-3, epidermal growth factor alpha and beta, platelet-derived endothelial cell growth factor, platelet-derived growth factor, tumor necrosis factor alpha, hepatocyte growth factor, insulin-like growth factor, colony stimulating factor, macrophage colony stimulating factor, granulocyte/macrophage colony stimulating factor, and nitric oxide synthase.

[1000] In additional embodiments, compositions of the invention are administered in combination with a chemotherapeutic agent. Chemotherapeutic agents that may be administered with the Therapeutics of the invention include, but are not limited to alkylating agents such as nitrogen mustards (for example, Mechlorethamine, cyclophosphamide, Cyclophosphamide Ifosfamide, Melphalan (L-sarcolysin), and Chlorambucil), ethylenimines and methylmelamines (for example, Hexamethylmelamine and Thiotepa), alkyl sulfonates (for example, Busulfan), nitrosoureas (for example, Carmustine (BCNU), Lomustine (CCNU), Semustine (methyl-CCNU), and Streptozocin (streptozotocin)), triazenes (for example, Dacarbazine (DTIC; dimethyltriazenoimidazolecarboxamide)), folic acid analogs (for example, Methotrexate (amethopterin)), pyrimidine analogs (for example, Fluorouacil (5-fluorouracil; 5-FU), Floxuridine (fluorodeoxyunridine; FudR), and Cytarabine (cytosine arabinoside)), purine analogs and related inhibitors (for example, Mercaptopurine (6-mercaptopurine; 6-MP), Thioguanine (6-thioguanine; TG), and Pentostatin (2′-deoxycoformycin)), vinca alkaloids (for example, Vinblastine (VLB, vinblastine sulfate)) and Vincristine (vincristine sulfate)), epipodophyllotoxins (for example, Etoposide and Teniposide), antibiotics (for example, Dactinomycin (actinomycin D), Daunorubicin (daunomycin; rubidomycin), Doxorubicin, Bleomycin, Plicamycin (mithramycin), and Mitomycin (mitomycin C), enzymes (for example, L-Asparaginase), biological response modifiers (for example, Interferon-alpha and interferon-alpha-2b), platinum coordination compounds (for example, Cisplatin (cis-DDP) and Carboplatin), anthracenedione (Mitoxantrone), substituted ureas (for example, Hydroxyurea), methylhydrazine derivatives (for example, Procarbazine (N-methylhydrazine; MIH), adrenocorticosteroids (for example, Prednisone), progestins (for example, Hydroxyprogesterone caproate, Medroxyprogesterone, Medroxyprogesterone acetate, and Megestrol acetate), estrogens (for example, Diethylstilbestrol (DES), Diethylstilbestrol diphosphate, Estradiol, and Ethinyl estradiol), antiestrogens (for example, Tamoxifen), androgens (Testosterone proprionate, and Fluoxymesterone), antiandrogens (for example, Flutamide), gonadotropin-releasing horomone analogs (for example, Leuprolide), other hormones and hormone analogs (for example, methyltestosterone, estramustine, estramustine phosphate sodium, chlorotrianisene, and testolactone), and others (for example, dicarbazine, glutamic acid, and mitotane).

[1001] In one embodiment, the compositions of the invention are administered in combination with one or more of the following drugs: infliximab (also known as Remicade™ Centocor, Inc.), Trocade (Roche, RO-32-3555), Leflunomide (also known as Arava™ from Hoechst Marion Roussel), Kineret™ (an IL-1 Receptor antagonist also known as Anakinra from Amgen, Inc.)

[1002] In a specific embodiment, compositions of the invention are administered in combination with CHOP (cyclophosphamide, doxorubicin, vincristine, and prednisone) or combination of one or more of the components of CHOP. In one embodiment, the compositions of the invention are administered in combination with anti-CD20 antibodies, human monoclonal anti-CD20 antibodies. In another embodiment, the compositions of the invention are administered in combination with anti-CD20 antibodies and CHOP, or anti-CD20 antibodies and any combination of one or more of the components of CHOP, particularly cyclophosphamide and/or prednisone. In a specific embodiment, compositions of the invention are administered in combination with Rituximab. In a further embodiment, compositions of the invention are administered with Rituximab and CHOP, or Rituximab and any combination of one or more of the components of CHOP, particularly cyclophosphamide and/or prednisone. In a specific embodiment, compositions of the invention are administered in combination with tositumomab. In a further embodiment, compositions of the invention are administered with tositumomab and CHOP, or tositumomab and any combination of one or more of the components of CHOP, particularly cyclophosphamide and/or prednisone. The anti-CD20 antibodies may optionally be associated with radioisotopes, toxins or cytotoxic prodrugs.

[1003] In another specific embodiment, the compositions of the invention are administered in combination Zevalin™. In a further embodiment, compositions of the invention are administered with Zevalin™ and CHOP, or Zevalin™ and any combination of one or more of the components of CHOP, particularly cyclophosphamide and/or prednisone. Zevalin™ may be associated with one or more radisotopes. Particularly preferred isotopes are 90Y and 111In.

[1004] In an additional embodiment, the Therapeutics of the invention are administered in combination with cytokines. Cytokines that may be administered with the Therapeutics of the invention include, but are not limited to, IL2, IL3, IL4, IL5, IL6, IL7, IL10, IL12, IL13, IL15, anti-CD40, CD40L, IFN-gamma and TNF-alpha. In another embodiment, Therapeutics of the invention may be administered with any interleukin, including, but not limited to, IL-1alpha, IL-1beta, IL-2, IL-3, IL-4, IL-5, IL-6, IL-7, IL-8, IL-9, IL-10, IL-11, IL-12, IL-13, IL-14, IL-15, IL-16, IL-17, IL-18, IL-19, IL-20, and IL-21.

[1005] In one embodiment, the Therapeutics of the invention are administered in combination with members of the TNF family. TNF, TNF-related or TNF-like molecules that may be administered with the Therapeutics of the invention include, but are not limited to, soluble forms of TNF-alpha, lymphotoxin-alpha (LT-alpha, also known as TNF-beta), LT-beta (found in complex heterotrimer LT-alpha2-beta), OPGL, FasL, CD27L, CD30L, CD40L, 4-1BBL, DcR3, OX40L, TNF-gamma (International Publication No. WO 96/14328), AIM-I (International Publication No. WO 97/33899), endokine-alpha (International Publication No. WO 98/07880), OPG, and neutrokine-alpha (International Publication No. WO 98/18921, OX40, and nerve growth factor (NGF), and soluble forms of Fas, CD30, CD27, CD40 and 4-IBB, TR2 (International Publication No. WO 96/34095), DR3 (International Publication No. WO 97/33904), DR4 (International Publication No. WO 98/32856), TR5 (International Publication No. WO 98/30693), TRANK, TR9 (International Publication No. WO 98/56892), TR10 (International Publication No. WO 98/54202), 312C2 (International Publication No. WO 98/06842), and TR12, and soluble forms CD154, CD70, and CD153.

[1006] In an additional embodiment, the Therapeutics of the invention are administered in combination with angiogenic proteins. Angiogenic proteins that may be administered with the Therapeutics of the invention include, but are not limited to, Glioma Derived Growth Factor (GDGF), as disclosed in European Patent Number EP-399816; Platelet Derived Growth Factor-A (PDGF-A), as disclosed in European Patent Number EP-682110; Platelet Derived Growth Factor-B (PDGF-B), as disclosed in European Patent Number EP-282317; Placental Growth Factor (PlGF), as disclosed in International Publication Number WO 92/06194; Placental Growth Factor-2 (PlGF-2), as disclosed in Hauser et al., Growth Factors, 4:259-268 (1993); Vascular Endothelial Growth Factor (VEGF), as disclosed in International Publication Number WO 90/13649; Vascular Endothelial Growth Factor-A (VEGF-A), as disclosed in European Patent Number EP-506477; Vascular Endothelial Growth Factor-2 (VEGF-2), as disclosed in International Publication Number WO 96/39515; Vascular Endothelial Growth Factor B (VEGF-3); Vascular Endothelial Growth Factor B-186 (VEGF-B186), as disclosed in International Publication Number WO 96/26736; Vascular Endothelial Growth Factor-D (VEGF-D), as disclosed in International Publication Number WO 98/02543; Vascular Endothelial Growth Factor-D (VEGF-D), as disclosed in International Publication Number WO 98/07832; and Vascular Endothelial Growth Factor-E (VEGF-E), as disclosed in German Patent Number DE19639601. The above mentioned references are herein incorporated by reference in their entireties.

[1007] In an additional embodiment, the Therapeutics of the invention are administered in combination with Fibroblast Growth Factors. Fibroblast Growth Factors that may be administered with the Therapeutics of the invention include, but are not limited to, FGF-1, FGF-2, FGF-3, FGF-4, FGF-5, FGF-6, FGF-7, FGF-8, FGF-9, FGF-10, FGF-11, FGF-12, FGF-13, FGF-14, and FGF-15.

[1008] In an additional embodiment, the Therapeutics of the invention are administered in combination with hematopoietic growth factors. Hematopoietic growth factors that may be administered with the Therapeutics of the invention include, but are not limited to, granulocyte macrophage colony stimulating factor (GM-CSF) (sargramostim, LEUKINE™, PROKINE™), granulocyte colony stimulating factor (G-CSF) (filgrastim, NEUPOGEN™), macrophage colony stimulating factor (M-CSF, CSF-1) erythropoietin (epoetin alfa, EPOGEN™, PROCRIT™), stem cell factor (SCF, c-kit ligand, steel factor), megakaryocyte colony stimulating factor, PIXY321 (a GMCSF/IL-3 fusion protein), interleukins, especially any one or more of IL-1 through IL-12, interferon-gamma, or thrombopoietin.

[1009] In certain embodiments, Therapeutics of the present invention are administered in combination with adrenergic blockers, such as, for example, acebutolol, atenolol, betaxolol, bisoprolol, carteolol, labetalol, metoprolol, nadolol, oxprenolol, penbutolol, pindolol, propranolol, sotalol, and timolol.

[1010] In another embodiment, the Therapeutics of the invention are administered in combination with an antiarrhythmic drug (e.g., adenosine, amidoarone, bretylium, digitalis, digoxin, digitoxin, diliazem, disopyramide, esmolol, flecainide, lidocaine, mexiletine, moricizine, phenytoin, procainamide, N-acetyl procainamide, propafenone, propranolol, quini dine, sotalol, tocainide, and verapamil).

[1011] In another embodiment, the Therapeutics of the invention are administered in combination with diuretic agents, such as carbonic anhydrase-inhibiting agents (e.g., acetazolamide, dichlorphenamide, and methazolamide), osmotic diuretics (e.g., glycerin, isosorbide, mannitol, and urea), diuretics that inhibit Na+-K+-2Cl− symport (e.g., furosemide, bumetamide, azosemide, piretamide, tripamide, ethacrynic acid, muzolimine, and torsemide), thiazide and thiazide-like diuretics (e.g., bendroflumethiazide, benzthiazide, chlorothiazide, hydrochlorothiazide, hydroflumethiazide, methyclothiazide, polythiazide, trichormethiazide, chlorthalidone, indapamide, metolazone, and quinethazone), potassium sparing diuretics (e.g., amiloride and triamterene), and mineralcorticoid receptor antagonists (e.g., spironolactone, canrenone, and potassium canrenoate).

[1012] In one embodiment, the Therapeutics of the invention are administered in combination with treatments for endocrine and/or hormone imbalance disorders. Treatments for endocrine and/or hormone imbalance disorders include, but are not limited to, 127I, radioactive isotopes of iodine such as 131I and 123I; recombinant growth hormone, such as HUMATROPE™ (recombinant somatropin); growth hormone analogs such as PROTROPIN™ (somatrem); dopamine agonists such as PARLODEL™ (bromocriptine); somatostatin analogs such as SANDOSTATIN™ (octreotide); gonadotropin preparations such as PREGNYL™, A.P.L. ™ and PROFASI™ (chorionic gonadotropin (CG)), PERGONAL™ (menotropins), and METRODIN™ (urofollitropin (uFSH)); synthetic human gonadotropin releasing hormone preparations such as FACTREL™ and LUTREPULSE™ (gonadorelin hydrochloride); synthetic gonadotropin agonists such as LUPRON™ (leuprolide acetate), SUPPRELIN™ (histrelin acetate), SYNAREL™ (nafarelin acetate), and ZOLADEX™ (goserelin acetate); synthetic preparations of thyrotropin-releasing hormone such as RELEFACT TRH™ and THYPINONE™ (protirelin); recombinant human TSH such as THYROGEN™; synthetic preparations of the sodium salts of the natural isomers of thyroid hormones such as L-T4™, SYNTHROID™ and LEVOTHROID™ (levothyroxine sodium), L-T3™, CYTOMEL™ and TRIOSTAT™ (liothyroine sodium), and THYROLAR™ (liotrix); antithyroid compounds such as 6-n-propylthiouracil (propylthiouracil), 1-methyl-2-mercaptoimidazole and TAPAZOLE™ (methimazole), NEO-MERCAZOLE™ (carbimazole); beta-adrenergic receptor antagonists such as propranolol and esmolol; Ca2+ channel blockers; dexamethasone and iodinated radiological contrast agents such as TELEPAQUE™ (iopanoic acid) and ORAGRAFIN™ (sodium ipodate).

[1013] Additional treatments for endocrine and/or hormone imbalance disorders include, but are not limited to, estrogens or congugated estrogens such as ESTRACE™ (estradiol), ESTINYL™ (ethinyl estradiol), PREMARIN™, ESTRATAB™, ORTHO-EST™, OGEN™ and estropipate (estrone), ESTROVIS™ (quinestrol), ESTRADERM™ (estradiol), DELESTROGEN™ and VALERGEN™ (estradiol valerate), DEPO-ESTRADIOL CYPIONATE™ and ESTROJECT LA™ (estradiol cypionate); antiestrogens such as NOLVADEX™ (tamoxifen), SEROPHENE™ and CLOMID™ (clomiphene); progestins such as DURALUTIN™ (hydroxyprogesterone caproate), MPA™ and DEPO-PROVERA™ (medroxyprogesterone acetate), PROVERAT™ and CYCRIN™ (MPA), MEGACE™ (megestrol acetate), NORLUTIN™ (norethindrone), and NORLUTATE™ and AYGESTIN™ (norethindrone acetate); progesterone implants such as NORPLANT SYSTEM™ (subdermal implants of norgestrel); antiprogestins such as RU 486™ (mifepristone); hormonal contraceptives such as ENOVID™ (norethynodrel plus mestranol), PROGESTASERT™ (intrauterine device that releases progesterone), LOESTRIN™, BREVICON™, MODICON™, GENORA™, NELONA™, NORINYL™, OVACON-35 ™ and OVACON-50™ (ethinyl estradiol/norethindrone), LEVLEN™, NORDETTE™, TR1-LEVLEN™ and TRIPHASIL-21™ (ethinyl estradiol/levonorgestrel) LO/OVRAL™ and OVRAL™ (ethinyl estradiol/norgestrel), DEMULEN™ (ethinyl estradiol/ethynodiol diacetate), NORINYL™, ORTHO-NOVUM™, NORETHIN™, GENORA™, and NELOVA™ (norethindrone/mestranol), DESOGEN™ and ORTHO-CEPT™ (ethinyl estradiol/desogestrel), ORTHO-CYCLEN™ and ORTHO-TRICYCLEN™ (ethinyl estradiol/norgestimate), MICRONOR™ and NOR-QD™ (norethindrone), and OVRETTE™ (norgestrel).

[1014] Additional treatments for endocrine and/or hormone imbalance disorders include, but are not limited to, testosterone esters such as methenolone acetate and testosterone undecanoate; parenteral and oral androgens such as TESTOJECT-50™ (testosterone), TESTEX™ (testosterone propionate), DELATESTRYL™ (testosterone enanthate), DEPO-TESTOSTERONE™ (testosterone cypionate), DANOCRINE™ (danazol), HALOTESTIN™ (fluoxymesterone), ORETON METHYL™, TESTRED™ and VIRILON™ (methyltestosterone), and OXANDRIN™ (oxandrolone); testosterone transdermal systems such as TESTODERM™; androgen receptor antagonist and 5-alpha-reductase inhibitors such as ANDROCUR™ (cyproterone acetate), EULEXIN™ (flutamide), and PROSCAR™ (finasteride); adrenocorticotropic hormone preparations such as CORTROSYN™ (cosyntropin); adrenocortical steroids and their synthetic analogs such as ACLOVATE™ (alclometasone dipropionate), CYCLOCORT™ (amcinonide), BECLOVENT™ and VANCERIL™ (beclomethasone dipropionate), CELESTONE™ (betamethasone), BENISONE™ and UTICORT™ (betamethasone benzoate), DIPROSONE™ (betamethasone dipropionate), CELESTONE PHOSPHATE™ (betamethasone sodium phosphate), CELESTONE SOLUSPAN™ (betamethasone sodium phosphate and acetate), BETA-VAL™ and VALISONE™ (betamethasone valerate), TEMOVATE™ (clobetasol propionate), CLODERM™ (clocortolone pivalate), CORTEF™ and HYDROCORTONE™ (cortisol (hydrocortisone)), HYDROCORTONE ACETATE™ (cortisol (hydrocortisone) acetate), LOCOID™ (cortisol (hydrocortisone) butyrate), HYDROCORTONE PHOSPHATE™ (cortisol (hydrocortisone) sodium phosphate), A-HYDROCORT™ and SOLU CORTEF™ (cortisol (hydrocortisone) sodium succinate), WESTCORT™ (cortisol (hydrocortisone) valerate), CORTISONE ACETATET™ (cortisone acetate), DESOWEN™ and TRIDESILON™ (desonide), TOPICORT™ (desoximetasone), DECADRON™ (dexamethasone), DECADRON LA™ (dexamethasone acetate), DECADRON PHOSPHATE™ and HEXADROL PHOSPHATE™ (dexamethasone sodium phosphate), FLORONE™ and MAXIFLOR™ (diflorasone diacetate), FLORINEF ACETATE™ (fludrocortisone acetate), AEROBID™ and NASALIDE™ (flunisolide), FLUONID™ and SYNALAR™ (fluocinolone acetonide), LIDEX™ (fluocinonide), FLUOR-OP™ and FML™ (fluorometholone), CORDRAN™ (flurandrenolide), HALOG™ (halcinonide), HMS LIZUIFILM™ (medrysone), MEDROL™ (methylprednisolone), DEPO-MEDROL™ and MEDROL ACETATE™ (methylprednisone acetate), A-METHAPRED™ and SOLUMEDROL™ (methylprednisolone sodium succinate), ELOCON™ (mometasone furoate), HALDRONE™ (paramethasone acetate), DELTA-CORTEF™ (prednisolone), ECONOPRED™ (prednisolone acetate), HYDELTRASOL™ (prednisolone sodium phosphate), HYDELTRA-T.B.A™ (prednisolone tebutate), DELTASONE™ (prednisone), ARISTOCORT™ and KENACORT™ (triamcinolone), KENALOG™ (triamcinolone acetonide), ARISTOCORT™ and KENACORT DIACETATE™ (triamcinolone diacetate), and ARISTOSPAN™ (triamcinolone hexacetonide); inhibitors of biosynthesis and action of adrenocortical steroids such as CYTADREN™ (aminoglutethimide), NIZORAL™ (ketoconazole), MODRASTANE™ (trilostane), and METOPIRONE™ (metyrapone); bovine, porcine or human insulin or mixtures thereof; insulin analogs; recombinant human insulin such as HUMULIN™ and NOVOLIN™; oral hypoglycemic agents such as ORAMIDE™ and ORINASE™ (tolbutamide), DIABINESE™ (chlorpropamide), TOLAMIDE™ and TOLINASE™ (tolazamide), DYMELOR™ (acetohexamide), glibenclamide, MICRONASE™, DIBETA™ and GLYNASE™ (glyburide), GLUCOTROL™ (glipizide), and DIAMICRON™ (gliclazide), GLUCOPHAGE™ (metformin), ciglitazone, pioglitazone, and alpha-glucosidase inhibitors; bovine or porcine glucagon; somatostatins such as SANDOSTATIN™ (octreotide); and diazoxides such as PROGLYCEM™ (diazoxide).

[1015] In one embodiment, the Therapeutics of the invention are administered in combination with treatments for uterine motility disorders. Treatments for uterine motility disorders include, but are not limited to, estrogen drugs such as conjugated estrogens (e.g., PREMARIN® and ESTRATAB®), estradiols (e.g., CLIMARA® and ALORA®), estropipate, and chlorotrianisene; progestin drugs (e.g., AMEN® (medroxyprogesterone), MICRONOR® (norethidrone acetate), PROMETRIUM® progesterone, and megestrol acetate); and estrogen/progesterone combination therapies such as, for example, conjugated estrogenslmedroxyprogesterone (e.g., PREMPRO™ and PREMPHASE®) and norethindrone acetate/ethinyl estsradiol (e.g., FEMHRT™).

[1016] In an additional embodiment, the Therapeutics of the invention are administered in combination with drugs effective in treating iron deficiency and hypochromic anemias, including but not limited to, ferrous sulfate (iron sulfate, FEOSOL™), ferrous fumarate (e.g., FEOSTAT™), ferrous gluconate (e.g., FERGON™), polysaccharide-iron complex (e.g., NIFEREX™), iron dextran injection (e.g., INFED™), cupric sulfate, pyroxidine, riboflavin, Vitamin B12, cyancobalamin injection (e.g., REDISOL™, RTUBRAMIN PC™), hydroxocobalamin, folic acid (e.g., FOLVITE™), leucovorin (folinic acid, 5-CHOH4PteGlu, citrovorum factor) or WELLCOVORIN (Calcium salt of leucovorin), transferrin or ferritin.

[1017] In certain embodiments, the Therapeutics of the invention are administered in combination with agents used to treat psychiatric disorders. Psychiatric drugs that may be administered with the Therapeutics of the invention include, but are not limited to, antipsychotic agents (e.g., chlorpromazine, chlorprothixene, clozapine, fluphenazine, haloperidol, loxapine, mesoridazine, molindone, olanzapine, perphenazine, pimozide, quetiapine, risperidone, thioridazine, thiothixene, trifluoperazine, and triflupromazine), antimanic agents (e.g., carbamazepine, divalproex sodium, lithium carbonate, and lithium citrate), antidepressants (e.g., amitriptyline, amoxapine, bupropion, citalopram, clomipramine, desipramine, doxepin, fluvoxamine, fluoxetine, imipramine, isocarboxazid, maprotiline, mirtazapine, nefazodone, nortriptyline, paroxetine, phenelzine, protriptyline, sertraline, tranylcypromine, trazodone, trimipramine, and venlafaxine), antianxiety agents (e.g., alprazolam, buspirone, chlordiazepoxide, clorazepate, diazepam, halazepam, lorazepam, oxazepam, and prazepam), and stimulants (e.g., d-amphetamine, methylphenidate, and pemoline).

[1018] In other embodiments, the Therapeutics of the invention are administered in combination with agents used to treat neurological disorders. Neurological agents that may be administered with the Therapeutics of the invention include, but are not limited to, antiepileptic agents (e.g., carbamazepine, clonazepam, ethosuximide, phenobarbital, phenytoin, primidone, valproic acid, divalproex sodium, felbamate, gabapentin, lamotrigine, levetiracetam, oxcarbazepine, tiagabine, topiramate, zonisamide, diazepam, lorazepam, and clonazepam), antiparkinsonian agents (e.g., levodopa/carbidopa, selegiline, amantidine, bromocriptine, pergolide, ropinirole, pramipexole, benztropine; biperiden; ethopropazine; procyclidine; trihexyphenidyl, tolcapone), and ALS therapeutics (e.g. riluzole).

[1019] In another embodiment, Therapeutics of the invention are administered in combination with vasodilating agents and/or calcium channel blocking agents. Vasodilating agents that may be administered with the Therapeutics of the invention include, but are not limited to, Angiotensin Converting Enzyme (ACE) inhibitors (e.g., papaverine, isoxsuprine, benazepril, captopril, cilazapril, enalapril, enalaprilat, fosinopril, lisinopril, moexipril, perindopril, quinapril, ramipril, spirapril, trandolapril, and nylidrin), and nitrates (e.g., isosorbide dinitrate, isosorbide mononitrate, and nitroglycerin). Examples of calcium channel blocking agents that may be administered in combination with the Therapeutics of the invention include, but are not limited to amlodipine, bepridil, diltiazem, felodipine, flunarizine, isradipine, nicardipine, nifedipine, nimodipine, and verapamil.

[1020] In certain embodiments, the Therapeutics of the invention are administered in combination with treatments for gastrointestinal disorders. Treatments for gastrointestinal disorders that may be administered with the Therapeutic of the invention include, but are not limited to, H2 histamine receptor antagonists (e.g., TAGAMET™ (cimetidine), ZANTAC™ (ranitidine), PEPCID™ (famotidine), and AXID™ (nizatidine)); inhibitors of H+, K+ ATPase (e.g., PREVACID™ (lansoprazole) and PRILOSEC™ (omeprazole)); Bismuth compounds (e.g., PEPTO-BISMOL™ (bismuth subsalicylate) and DE-NOL™ (bismuth subeitrate)); various antacids; sucralfate; prostaglandin analogs (e.g. CYTOTEC™ (misoprostol)); muscarinic cholinergic antagonists; laxatives (e.g., surfactant laxatives, stimulant laxatives, saline and osmotic laxatives); antidiarrheal agents (e.g., LOMOTIL™ (diphenoxylate), MOTOFEN™ (diphenoxin), and IMODIUM™ (loperamide hydrochloride)), synthetic analogs of somatostatin such as SANDOSTATIN™ (octreotide), antiemetic agents (e.g., ZOFRAN™ (ondansetron), KYTRIL™ (granisetron hydrochloride), tropisetron, dolasetron, metoclopramide, chlorpromazine, perphenazine, prochlorperazine, promethazine, thiethylperazine, triflupromazine, domperidone, haloperidol, droperidol, trimethobenzamide, dexamethasone, methylprednisolone, dronabinol, and nabilone); D2 antagonists (e.g., metoclopramide, trimethobenzamide and chlorpromazine); bile salts; chenodeoxycholic acid; ursodeoxycholic acid; and pancreatic enzyme preparations such as pancreatin and pancrelipase.

[1021] In additional embodiments, the Therapeutics of the invention are administered in combination with other therapeutic or prophylactic regimens, such as, for example, radiation therapy.

Example 14

Method of Treating Decreased Levels of the Polypeptide

[1022] The present invention relates to a method for treating an individual in need of an increased level of a polypeptide of the invention in the body comprising administering to such an individual a composition comprising a therapeutically effective amount of an agonist of the invention (including polypeptides of the invention). Moreover, it will be appreciated that conditions caused by a decrease in the standard or normal expression level of a polypeptide of the present invention in an individual can be treated by administering the agonist or antagonist of the present invention. Thus, the invention also provides a method of treatment of an individual in need of an increased level of the polypeptide comprising administering to such an individual a Therapeutic comprising an amount of the agonist or antagonist to increase the activity level of the polypeptide in such an individual.

[1023] For example, a patient with decreased levels of a polypeptide receives a daily dose 0.1-100 ug/kg of the agonist or antagonist for six consecutive days. The exact details of the dosing scheme, based on administration and formulation, are provided in Example 13.

Example 15

Method of Treating Increased Levels of the Polypeptide

[1024] The present invention also relates to a method of treating an individual in need of a decreased level of a polypeptide of the invention in the body comprising administering to such an individual a composition comprising a therapeutically effective amount of an antagonist of the invention (including polypeptides and antibodies of the invention).

[1025] In one example, antisense technology is used to inhibit production of a polypeptide of the present invention. This technology is one example of a method of decreasing levels of a polypeptide, due to a variety of etiologies, such as cancer.

[1026] For example, a patient diagnosed with abnormally increased levels of a polypeptide is administered intravenously antisense polynucleotides at 0.5, 1.0, 1.5, 2.0 and 3.0 mg/kg day for 21 days. This treatment is repeated after a 7-day rest period if the treatment was well tolerated. The antisense polynucleotides of the present invention can be formulated using techniques and formulations described herein (e.g. see Example 13), or otherwise known in the art.

Example 16

Method of Treatment Using Gene Therapy—Ex Vivo

[1027] One method of gene therapy transplants fibroblasts, which are capable of expressing a polypeptide, onto a patient. Generally, fibroblasts are obtained from a subject by skin biopsy. The resulting tissue is placed in tissue-culture medium and separated into small pieces. Small chunks of the tissue are placed on a wet surface of a tissue culture flask, approximately ten pieces are placed in each flask. The flask is turned upside down, closed tight and left at room temperature over night. After 24 hours at room temperature, the flask is inverted and the chunks of tissue remain fixed to the bottom of the flask and fresh media (e.g., Ham's F12 media, with 10% FBS, penicillin and streptomycin) is added. The flasks are then incubated at 37 degree C. for approximately one week.

[1028] At this time, fresh media is added and subsequently changed every several days. After an additional two weeks in culture, a monolayer of fibroblasts emerge. The monolayer is trypsinized and scaled into larger flasks.

[1029] pMV-7 (Kirschmeier, P. T. et al., DNA, 7:219-25 (1988)), flanked by the long terminal repeats of the Moloney murine sarcoma virus, is digested with EcoRI and HindIII and subsequently treated with calf intestinal phosphatase. The linear vector is fractionated on agarose gel and purified, using glass beads.

[1030] The cDNA encoding a polypeptide of the present invention can be amplified using PCR primers which correspond to the 5′ and 3′ end sequences respectively as set forth in Example 1 using primers and having appropriate restriction sites and initiation/stop codons, if necessary. Preferably, the 5′ primer contains an EcoRI site and the 3′ primer includes a HindIII site. Equal quantities of the Moloney murine sarcoma virus linear backbone and the amplified EcoRI and HindIII fragment are added together, in the presence of T4 DNA ligase. The resulting mixture is maintained under conditions appropriate for ligation of the two fragments. The ligation mixture is then used to transform bacteria HB101, which are then plated onto agar containing kanamycin for the purpose of confirming that the vector has the gene of interest properly inserted.

[1031] The amphotropic pA317 or GP+am12 packaging cells are grown in tissue culture to confluent density in Dulbecco's Modified Eagles Medium (DMEM) with 10% calf serum (CS), penicillin and streptomycin. The MSV vector containing the gene is then added to the media and the packaging cells transduced with the vector. The packaging cells now produce infectious viral particles containing the gene (the packaging cells are now referred to as producer cells).

[1032] Fresh media is added to the transduced producer cells, and subsequently, the media is harvested from a 10 cm plate of confluent producer cells. The spent media, containing the infectious viral particles, is filtered through a millipore filter to remove detached producer cells and this media is then used to infect fibroblast cells. Media is removed from a sub-confluent plate of fibroblasts and quickly replaced with the media from the producer cells. This media is removed and replaced with fresh media. If the titer of virus is high, then virtually all fibroblasts will be infected and no selection is required. If the titer is very low, then it is necessary to use a retroviral vector that has a selectable marker, such as neo or his. Once the fibroblasts have been efficiently infected, the fibroblasts are analyzed to determine whether protein is produced.

[1033] The engineered fibroblasts are then transplanted onto the host, either alone or after having been grown to confluence on cytodex 3 microcarrier beads.

Example 17

Gene Therapy Using Endogenous Genes Corresponding to Polynucleotides of the Invention

[1034] Another method of gene therapy according to the present invention involves operably associating the endogenous polynucleotide sequence of the invention with a promoter via homologous recombination as described, for example, in U.S. Pat. No. 5,641,670, issued Jun. 24, 1997; International Publication NO: WO 96/29411, published Sep. 26, 1996; International Publication NO: WO 94/12650, published Aug. 4, 1994; Koller et al., Proc. Natl. Acad. Sci. USA, 86:8932-8935 (1989); and Zijlstra et al., Nature, 342:435-438 (1989). This method involves the activation of a gene which is present in the target cells, but which is not expressed in the cells, or is expressed at a lower level than desired.

[1035] Polynucleotide constructs are made which contain a promoter and targeting sequences, which are homologous to the 5′ non-coding sequence of endogenous polynucleotide sequence, flanking the promoter. The targeting sequence will be sufficiently near the 5′ end of the polynucleotide sequence so the promoter will be operably linked to the endogenous sequence upon homologous recombination. The promoter and the targeting sequences can be amplified using PCR. Preferably, the amplified promoter contains distinct restriction enzyme sites on the 5′ and 3′ ends. Preferably, the 3′ end of the first targeting sequence contains the same restriction enzyme site as the 5′ end of the amplified promoter and the 5′ end of the second targeting sequence contains the same restriction site as the 3′ end of the amplified promoter.

[1036] The amplified promoter and the amplified targeting sequences are digested with the appropriate restriction enzymes and subsequently treated with calf intestinal phosphatase. The digested promoter and digested targeting sequences are added together in the presence of T4 DNA ligase. The resulting mixture is maintained under conditions appropriate for ligation of the two fragments. The construct is size fractionated on an agarose gel, then purified by phenol extraction and ethanol precipitation.

[1037] In this Example, the polynucleotide constructs are administered as naked polynucleotides via electroporation. However, the polynucleotide constructs may also be administered with transfection-facilitating agents, such as liposomes, viral sequences, viral particles, precipitating agents, etc. Such methods of delivery are known in the art.

[1038] Once the cells are transfected, homologous recombination will take place which results in the promoter being operably linked to the endogenous polynucleotide sequence. This results in the expression of polynucleotide corresponding to the polynucleotide in the cell. Expression may be detected by immunological staining, or any other method known in the art.

[1039] Fibroblasts are obtained from a subject by skin biopsy. The resulting tissue is placed in DMEM+10% fetal calf serum. Exponentially growing or early stationary phase fibroblasts are trypsinized and rinsed from the plastic surface with nutrient medium. An aliquot of the cell suspension is removed for counting, and the remaining cells are subjected to centrifugation. The supernatant is aspirated and the pellet is resuspended in 5 ml of electroporation buffer (20 mM HEPES pH 7.3, 137 mM NaCl, 5 mM KCl, 0.7 mM Na2 HPO4, 6 mM dextrose). The cells are recentrifuged, the supernatant aspirated, and the cells resuspended in electroporation buffer containing 1 mg/ml acetylated bovine serum albumin. The final cell suspension contains approximately 3×106 cells/ml. Electroporation should be performed immediately following resuspension.

[1040] Plasmid DNA is prepared according to standard techniques. For example, to construct a plasmid for targeting to the locus corresponding to the polynucleotide of the invention, plasmid pUC18 (MBI Fermentas, Amherst, N.Y.) is digested with HindIII. The CMV promoter is amplified by PCR with an XbaI site on the 5′ end and a BamHI site on the 3′ end. Two non-coding sequences are amplified via PCR: one non-coding sequence (fragment 1) is amplified with a HindIII site at the 5′ end and an Xba site at the 3′end; the other non-coding sequence (fragment 2) is amplified with a BamHI site at the 5′end and a HindIII site at the 3′end. The CMV promoter and the fragments (1 and 2) are digested with the appropriate enzymes (CMV promoter—XbaI and BamHI; fragment 1—XbaI; fragment 2—BamHI) and ligated together. The resulting ligation product is digested with HindIII, and ligated with the HindIII-digested pUC18 plasmid.

[1041] Plasmid DNA is added to a sterile cuvette with a 0.4 cm electrode gap (Bio-Rad). The final DNA concentration is generally at least 120 μg/ml. 0.5 ml of the cell suspension (containing approximately 1.5.×106 cells) is then added to the cuvette, and the cell suspension and DNA solutions are gently mixed. Electroporation is performed with a Gene-Pulser apparatus (Bio-Rad). Capacitance and voltage are set at 960 μF and 250-300 V, respectively. As voltage increases, cell survival decreases, but the percentage of surviving cells that stably incorporate the introduced DNA into their genome increases dramatically. Given these parameters, a pulse time of approximately 14-20 mSec should be observed.

[1042] Electroporated cells are maintained at room temperature for approximately 5 min, and the contents of the cuvette are then gently removed with a sterile transfer pipette. The cells are added directly to 10 ml of prewarmed nutrient media (DMEM with 15% calf serum) in a 10 cm dish and incubated at 37 degree C. The following day, the media is aspirated and replaced with 10 ml of fresh media and incubated for a further 16-24 hours.

[1043] The engineered fibroblasts are then injected into the host, either alone or after having been grown to confluence on cytodex 3 microcarrier beads. The fibroblasts now produce the protein product. The fibroblasts can then be introduced into a patient as described above.

Example 18

Method of Treatment Using Gene Therapy—In Vivo

[1044] Another aspect of the present invention is using in vivo gene therapy methods to treat disorders, diseases and conditions. The gene therapy method relates to the introduction of naked nucleic acid (DNA, RNA, and antisense DNA or RNA) sequences into an animal to increase or decrease the expression of the polypeptide. The polynucleotide of the present invention may be operatively linked to (i.e., associated with) a promoter or any other genetic elements necessary for the expression of the polypeptide by the target tissue. Such gene therapy and delivery techniques and methods are known in the art, see, for example, WO90/11092, WO98/11779; U.S. Pat. No. 5,693,622, 5705151, 5580859; Tabata et al., Cardiovasc. Res. 35(3):470-479 (1997); Chao et al., Pharmacol. Res. 35(6):517-522 (1997); Wolff, Neuromuscul. Disord. 7(5):314-318 (1997); Schwartz et al., Gene Ther. 3(5):405-411 (1996); Tsurumi et al., Circulation 94(12):3281-3290 (1996) (incorporated herein by reference).

[1045] The polynucleotide constructs may be delivered by any method that delivers injectable materials to the cells of an animal, such as, injection into the interstitial space of tissues (heart, muscle, skin, lung, liver, intestine and the like). The polynucleotide constructs can be delivered in a pharmaceutically acceptable liquid or aqueous carrier.

[1046] The term “naked” polynucleotide, DNA or RNA, refers to sequences that are free from any delivery vehicle that acts to assist, promote, or facilitate entry into the cell, including viral sequences, viral particles, liposome formulations, lipofectin or precipitating agents and the like. However, the polynucleotides of the present invention may also be delivered in liposome formulations (such as those taught in Felgner P. L. et al. (1995) Ann. NY Acad. Sci. 772:126-139 and Abdallah B. et al. (1995) Biol. Cell 85(1):1-7) which can be prepared by methods well known to those skilled in the art.

[1047] The polynucleotide vector constructs used in the gene therapy method are preferably constructs that will not integrate into the host genome nor will they contain sequences that allow for replication. Any strong promoter known to those skilled in the art can be used for driving the expression of DNA. Unlike other gene therapy techniques, one major advantage of introducing naked nucleic acid sequences into target cells is the transitory nature of the polynucleotide synthesis in the cells. Studies have shown that non-replicating DNA sequences can be introduced into cells to provide production of the desired polypeptide for periods of up to six months.

[1048] The polynucleotide construct can be delivered to the interstitial space of tissues within an animal, including muscle, skin, brain, lung, liver, spleen, bone marrow, thymus, heart, lymph, blood, bone, cartilage, pancreas, kidney, gall bladder, stomach, intestine, testis, ovary, uterus, rectum, nervous system, eye, gland, and connective tissue. Interstitial space of the tissues comprises the intercellular fluid, mucopolysaccharide matrix among the reticular fibers of organ tissues, elastic fibers in the walls of vessels or chambers, collagen fibers of fibrous tissues, or that same matrix within connective tissue ensheathing muscle cells or in the lacunae of bone. It is similarly the space occupied by the plasma of the circulation and the lymph fluid of the lymphatic channels. Delivery to the interstitial space of muscle tissue is preferred for the reasons discussed below. They may be conveniently delivered by injection into the tissues comprising these cells. They are preferably delivered to and expressed in persistent, non-dividing cells which are differentiated, although delivery and expression may be achieved in non-differentiated or less completely differentiated cells, such as, for example, stem cells of blood or skin fibroblasts. In vivo muscle cells are particularly competent in their ability to take up and express polynucleotides.

[1049] For the naked polynucleotide injection, an effective dosage amount of DNA or RNA will be in the range of from about 0.05 g/kg body weight to about 50 mg/kg body weight. Preferably the dosage will be from about 0.005 mg/kg to about 20 mg/kg and more preferably from about 0.05 mg/kg to about 5 mg/kg. Of course, as the artisan of ordinary skill will appreciate, this dosage will vary according to the tissue site of injection. The appropriate and effective dosage of nucleic acid sequence can readily be determined by those of ordinary skill in the art and may depend on the condition being treated and the route of administration. The preferred route of administration is by the parenteral route of injection into the interstitial space of tissues. However, other parenteral routes may also be used, such as, inhalation of an aerosol formulation particularly for delivery to lungs or bronchial tissues, throat or mucous membranes of the nose. In addition, naked polynucleotide constructs can be delivered to arteries during angioplasty by the catheter used in the procedure.

[1050] The dose response effects of injected polynucleotide in muscle in vivo is determined as follows. Suitable template DNA for production of mRNA coding for polypeptide of the present invention is prepared in accordance with a standard recombinant DNA methodology. The template DNA, which may be either circular or linear, is either used as naked DNA or complexed with liposomes. The quadriceps muscles of mice are then injected with various amounts of the template DNA.

[1051] Five to six week old female and male Balb/C mice are anesthetized by intraperitoneal injection with 0.3 ml of 2.5% Avertin. A 1.5 cm incision is made on the anterior thigh, and the quadriceps muscle is directly visualized. The template DNA is injected in 0.1 ml of carrier in a 1 cc syringe through a 27 gauge needle over one minute, approximately 0.5 cm from the distal insertion site of the muscle into the knee and about 0.2 cm deep. A suture is placed over the injection site for future localization, and the skin is closed with stainless steel clips.

[1052] After an appropriate incubation time (e.g., 7 days) muscle extracts are prepared by excising the entire quadriceps. Every fifth 15 um cross-section of the individual quadriceps muscles is histochemically stained for protein expression. A time course for protein expression may be done in a similar fashion except that quadriceps from different mice are harvested at different times. Persistence of DNA in muscle following injection may be determined by Southern blot analysis after preparing total cellular DNA and HIRT supernatants from injected and control mice. The results of the above experimentation in mice can be used to extrapolate proper dosages and other treatment parameters in humans and other animals using naked DNA.

Example 19

Transgenic Animals

[1053] The polypeptides of the invention can also be expressed in transgenic animals. Animals of any species, including, but not limited to, mice, rats, rabbits, hamsters, guinea pigs, pigs, micro-pigs, goats, sheep, cows and non-human primates, e.g., baboons, monkeys, and chimpanzees may be used to generate transgenic animals. In a specific embodiment, techniques described herein or otherwise known in the art, are used to express polypeptides of the invention in humans, as part of a gene therapy protocol.

[1054] Any technique known in the art may be used to introduce the transgene (i.e., polynucleotides of the invention) into animals to produce the founder lines of transgenic animals. Such techniques include, but are not limited to, pronuclear microinjection (Paterson et al., Appl. Microbiol. Biotechnol. 40:691-698 (1994); Carver et al., Biotechnology (NY) 11:1263-1270 (1993); Wright et al., Biotechnology (NY) 9:830-834 (1991); and Hoppe et al., U.S. Pat. No. 4,873,191 (1989)); retrovirus mediated gene transfer into germ lines (Van der Putten et al., Proc. Natl. Acad. Sci., USA 82:6148-6152 (1985)), blastocysts or embryos; gene targeting in embryonic stem cells (Thompson et al., Cell 56:313-321 (1989)); electroporation of cells or embryos (Lo, 1983, Mol Cell. Biol. 3:1803-1814 (1983)); introduction of the polynucleotides of the invention using a gene gun (see, e.g., Ulmer et al., Science 259:1745 (1993); introducing nucleic acid constructs into embryonic pleuripotent stem cells and transferring the stem cells back into the blastocyst; and sperm-mediated gene transfer (Lavitrano et al., Cell 57:717-723 (1989); etc. For a review of such techniques, see Gordon, “Transgenic Animals,” Intl. Rev. Cytol. 115:171-229 (1989), which is incorporated by reference herein in its entirety.

[1055] Any technique known in the art may be used to produce transgenic clones containing polynucleotides of the invention, for example, nuclear transfer into enucleated oocytes of nuclei from cultured embryonic, fetal, or adult cells induced to quiescence (Campell et al., Nature 380:64-66 (1996); Wilmut et al., Nature 385:810-813 (1997)).

[1056] The present invention provides for transgenic animals that carry the transgene in all their cells, as well as animals which carry the transgene in some, but not all their cells, i.e., mosaic animals or chimeric. The transgene may be integrated as a single transgene or as multiple copies such as in concatamers, e.g., head-to-head tandems or head-to-tail tandems. The transgene may also be selectively introduced into and activated in a particular cell type by following, for example, the teaching of Lasko et al. (Lasko et al., Proc. Natl. Acad. Sci. USA 89:6232-6236 (1992)). The regulatory sequences required for such a cell-type specific activation will depend upon the particular cell type of interest, and will be apparent to those of skill in the art. When it is desired that the polynucleotide transgene be integrated into the chromosomal site of the endogenous gene, gene targeting is preferred. Briefly, when such a technique is to be utilized, vectors containing some nucleotide sequences homologous to the endogenous gene are designed for the purpose of integrating, via homologous recombination with chromosomal sequences, into and disrupting the function of the nucleotide sequence of the endogenous gene. The transgene may also be selectively introduced into a particular cell type, thus inactivating the endogenous gene in only that cell type, by following, for example, the teaching of Gu et al. (Gu et al., Science 265:103-106 (1994)). The regulatory sequences required for such a cell-type specific inactivation will depend upon the particular cell type of interest, and will be apparent to those of skill in the art.

[1057] Once transgenic animals have been generated, the expression of the recombinant gene may be assayed utilizing standard techniques. Initial screening may be accomplished by Southern blot analysis or PCR techniques to analyze animal tissues to verify that integration of the transgene has taken place. The level of mRNA expression of the transgene in the tissues of the transgenic animals may also be assessed using techniques which include, but are not limited to, Northern blot analysis of tissue samples obtained from the animal, in situ hybridization analysis, and reverse transcriptase-PCR (rt-PCR). Samples of transgenic gene-expressing tissue may also be evaluated immunocytochemically or immunohistochemically using antibodies specific for the transgene product.

[1058] Once the founder animals are produced, they may be bred, inbred, outbred, or crossbred to produce colonies of the particular animal. Examples of such breeding strategies include, but are not limited to: outbreeding of founder animals with more than one integration site in order to establish separate lines; inbreeding of separate lines in order to produce compound transgenics that express the transgene at higher levels because of the effects of additive expression of each transgene; crossing of heterozygous transgenic animals to produce animals homozygous for a given integration site in order to both augment expression and eliminate the need for screening of animals by DNA analysis; crossing of separate homozygous lines to produce compound heterozygous or homozygous lines; and breeding to place the transgene on a distinct background that is appropriate for an experimental model of interest.

[1059] Transgenic animals of the invention have uses which include, but are not limited to, animal model systems useful in elaborating the biological function of polypeptides of the present invention, studying conditions and/or disorders associated with aberrant expression, and in screening for compounds effective in ameliorating such conditions and/or disorders.

Example 20

Knock-Out Animals

[1060] Endogenous gene expression can also be reduced by inactivating or “knocking out” the gene and/or its promoter using targeted homologous recombination. (e.g., see Smithies et al., Nature 317:230-234 (1985); Thomas & Capecchi, Cell 51:503-512 (1987); Thompson et al., Cell 5:313-321 (1989); each of which is incorporated by reference herein in its entirety). For example, a mutant, non-functional polynucleotide of the invention (or a completely unrelated DNA sequence) flanked by DNA homologous to the endogenous polynucleotide sequence (either the coding regions or regulatory regions of the gene) can be used, with or without a selectable marker and/or a negative selectable marker, to transfect cells that express polypeptides of the invention in vivo. In another embodiment, techniques known in the art are used to generate knockouts in cells that contain, but do not express the gene of interest. Insertion of the DNA construct, via targeted homologous recombination, results in inactivation of the targeted gene. Such approaches are particularly suited in research and agricultural fields where modifications to embryonic stem cells can be used to generate animal offspring with an inactive targeted gene (e.g., see Thomas & Capecchi 1987 and Thompson 1989, supra). However this approach can be routinely adapted for use in humans provided the recombinant DNA constructs are directly administered or targeted to the required site in vivo using appropriate viral vectors that will be apparent to those of skill in the art.

[1061] In further embodiments of the invention, cells that are genetically engineered to express the polypeptides of the invention, or alternatively, that are genetically engineered not to express the polypeptides of the invention (e.g., knockouts) are administered to a patient in vivo. Such cells may be obtained from the patient (i.e., animal, including human) or an MHC compatible donor and can include, but are not limited to fibroblasts, bone marrow cells, blood cells (e.g., lymphocytes), adipocytes, muscle cells, endothelial cells etc. The cells are genetically engineered in vitro using recombinant DNA techniques to introduce the coding sequence of polypeptides of the invention into the cells, or alternatively, to disrupt the coding sequence and/or endogenous regulatory sequence associated with the polypeptides of the invention, e.g., by transduction (using viral vectors, and preferably vectors that integrate the transgene into the cell genome) or transfection procedures, including, but not limited to, the use of plasmids, cosmids, YACs, naked DNA, electroporation, liposomes, etc. The coding sequence of the polypeptides of the invention can be placed under the control of a strong constitutive or inducible promoter or promoter/enhancer to achieve expression, and preferably secretion, of the polypeptides of the invention. The engineered cells which express and preferably secrete the polypeptides of the invention can be introduced into the patient systemically, e.g., in the circulation, or intraperitoneally.

[1062] Alternatively, the cells can be incorporated into a matrix and implanted in the body, e.g., genetically engineered fibroblasts can be implanted as part of a skin graft; genetically engineered endothelial cells can be implanted as part of a lymphatic or vascular graft. (See, for example, Anderson et al. U.S. Pat. No. 5,399,349; and Mulligan & Wilson, U.S. Pat. No. 5,460,959 each of which is incorporated by reference herein in its entirety).

[1063] When the cells to be administered are non-autologous or non-MHC compatible cells, they can be administered using well known techniques which prevent the development of a host immune response against the introduced cells. For example, the cells may be introduced in an encapsulated form which, while allowing for an exchange of components with the immediate extracellular environment, does not allow the introduced cells to be recognized by the host immune system.

[1064] Transgenic and “knock-out” animals of the invention have uses which include, but are not limited to, animal model systems useful in elaborating the biological function of polypeptides of the present invention, studying conditions and/or disorders associated with aberrant expression, and in screening for compounds effective in ameliorating such conditions and/or disorders.

Example 21

Assays Detecting Stimulation or Inhibition of B cell Proliferation and Differentiation

[1065] Generation of functional humoral immune responses requires both soluble and cognate signaling between B-lineage cells and their microenvironment. Signals may impart a positive stimulus that allows a B-lineage cell to continue its programmed development, or a negative stimulus that instructs the cell to arrest its current developmental pathway. To date, numerous stimulatory and inhibitory signals have been found to influence B cell responsiveness including IL-2, IL-4, IL-5, IL-6, IL-7, IL10, IL-13, IL-14 and IL-15. Interestingly, these signals are by themselves weak effectors but can, in combination with various co-stimulatory proteins, induce activation, proliferation, differentiation, homing, tolerance and death among B cell populations.

[1066] One of the best studied classes of B-cell co-stimulatory proteins is the TNF-superfamily. Within this family CD40, CD27, and CD30 along with their respective ligands CD154, CD70, and CD153 have been found to regulate a variety of immune responses. Assays which allow for the detection and/or observation of the proliferation and differentiation of these B-cell populations and their precursors are valuable tools in determining the effects various proteins may have on these B-cell populations in terms of proliferation and differentiation. Listed below are two assays designed to allow for the detection of the differentiation, proliferation, or inhibition of B-cell populations and their precursors.

[1067] In Vitro Assay-Agonists or antagonists of the invention can be assessed for its ability to induce activation, proliferation, differentiation or inhibition and/or death in B-cell populations and their precursors. The activity of the agonists or antagonists of the invention on purified human tonsillar B cells, measured qualitatively over the dose range from 0.1 to 10,000 ng/mL, is assessed in a standard B-lymphocyte co-stimulation assay in which purified tonsillar B cells are cultured in the presence of either formalin-fixed Staphylococcus aureus Cowan I (SAC) or immobilized anti-human IgM antibody as the priming agent. Second signals such as IL-2 and IL-15 synergize with SAC and IgM crosslinking to elicit B cell proliferation as measured by tritiated-thymidine incorporation. Novel synergizing agents can be readily identified using this assay. The assay involves isolating human tonsillar B cells by magnetic bead (MACS) depletion of CD3-positive cells. The resulting cell population is greater than 95% B cells as assessed by expression of CD45R(B220).

[1068] Various dilutions of each sample are placed into individual wells of a 96-well plate to which are added 105 B-cells suspended in culture medium (RPMI 1640 containing 10% FBS, 5×10−5M 2ME, 100U/ml penicillin, 10 ug/ml streptomycin, and 10−5 dilution of SAC) in a total volume of 150 ul. Proliferation or inhibition is quantitated by a 20 h pulse (1 uCi/well) with 3H-thymidine (6.7 Ci/mM) beginning 72 h post factor addition. The positive and negative controls are IL2 and medium respectively.

[1069] In vivo Assay—BALB/c mice are injected (i.p.) twice per day with buffer only, or 2 mg/Kg of agonists or antagonists of the invention, or truncated forms thereof. Mice receive this treatment for 4 consecutive days, at which time they are sacrificed and various tissues and serum collected for analyses. Comparison of H&E sections from normal spleens and spleens treated with agonists or antagonists of the invention identify the results of the activity of the agonists or antagonists on spleen cells, such as the diffusion of peri-arterial lymphatic sheaths, and/or significant increases in the nucleated cellularity of the red pulp regions, which may indicate the activation of the differentiation and proliferation of B-cell populations. Immunohistochemical studies using a B cell marker, anti-CD45R(B220), are used to determine whether any physiological changes to splenic cells, such as splenic disorganization, are due to increased B-cell representation within loosely defined B-cell zones that infiltrate established T-cell regions.

[1070] Flow cytometric analyses of the spleens from mice treated with agonist or antagonist is used to indicate whether the agonists or antagonists specifically increases the proportion of ThB+, CD45R(B220)dull B cells over that which is observed in control mice.

[1071] Likewise, a predicted consequence of increased mature B-cell representation in vivo is a relative increase in serum Ig titers. Accordingly, serum IgM and IgA levels are compared between buffer and agonists or antagonists-treated mice.

[1072] The studies described in this example tested activity of agonists or antagonists of the invention. However, one skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides or polypeptides of the invention (e.g., gene therapy).

Example 22

T Cell Proliferation Assay

[1073] A CD3-induced proliferation assay is performed on PBMCs and is measured by the uptake of 3H-thymidine. The assay is performed as follows. Ninety-six well plates are coated with 100 μl/well of mAb to CD3 (HIT3a, Pharningen) or isotype-matched control mAb (B33.1) overnight at 4 degrees C. (1 μg/ml in 0.05M bicarbonate buffer, pH 9.5), then washed three times with PBS. PBMC are isolated by F/H gradient centrifugation from human peripheral blood and added to quadruplicate wells (5×104/well) of mAb coated plates in RPMI containing 10% FCS and P/S in the presence of varying concentrations of agonists or antagonists of the invention (total volume 200 ul). Relevant protein buffer and medium alone are controls. After 48 hr. culture at 37 degrees C., plates are spun for 2 min. at 1000 rpm and 100 μl of supernatant is removed and stored −20 degrees C. for measurement of IL-2 (or other cytokines) if effect on proliferation is observed. Wells are supplemented with 100 ul of medium containing 0.5 uCi of 3H-thymidine and cultured at 37 degrees C. for 18-24 hr. Wells are harvested and incorporation of 3H-thymidine used as a measure of proliferation. Anti-CD3 alone is the positive control for proliferation. IL-2 (100 U/ml) is also used as a control which enhances proliferation. Control antibody which does not induce proliferation of T cells is used as the negative control for the effects of agonists or antagonists of the invention.

[1074] The studies described in this example tested activity of agonists or antagonists of the invention. However, one skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides or polypeptides of the invention (e.g., gene therapy).

Example 23

Effect of Agonists or Antagonists of the Invention on the Expression of MHC Class II, Costimulatory and Adhesion Molecules and Cell Differentiation of Monocytes and Monocyte-Derived Human Dendritic Cells

[1075] Dendritic cells are generated by the expansion of proliferating precursors found in the peripheral blood; adherent PBMC or elutriated monocytic fractions are cultured for 7-10 days with GM-CSF (50 ng/ml) and IL-4 (20 ng/ml). These dendritic cells have the characteristic phenotype of immature cells (expression of CD1, CD80, CD86, CD40 and MHC class II antigens). Treatment with activating factors, such as TNF-α, causes a rapid change in surface phenotype (increased expression of MHC class I and II, costimulatory and adhesion molecules, downregulation of FCγ RII, upregulation of CD83). These changes correlate with increased antigen-presenting capacity and with functional maturation of the dendritic cells.

[1076] FACS analysis of surface antigens is performed as follows. Cells are treated 1-3 days with increasing concentrations of agonist or antagonist of the invention or LPS (positive control), washed with PBS containing 1% BSA and 0.02 mM sodium azide, and then incubated with 1:20 dilution of appropriate FITC- or PE-labeled monoclonal antibodies for 30 minutes at 4 degrees C. After an additional wash, the labeled cells are analyzed by flow cytometry on a FACScan (Becton Dickinson).

[1077] Effect on the production of cytokines. Cytokines generated by dendritic cells, in particular IL-12, are important in the initiation of T-cell dependent immune responses. IL-12 strongly influences the development of Th1 helper T-cell immune response, and induces cytotoxic T and NK cell function. An ELISA is used to measure the IL-12 release as follows. Dendritic cells (106/ml) are treated with increasing concentrations of agonists or antagonists of the invention for 24 hours. LPS (100 ng/ml) is added to the cell culture as positive control. Supernatants from the cell cultures are then collected and analyzed for IL-12 content using commercial ELISA kit (e.g., R & D Systems (Minneapolis, Minn.)). The standard protocols provided with the kits are used.

[1078] Effect on the expression of MHC Class II, costimulatory and adhesion molecules. Three major families of cell surface antigens can be identified on monocytes: adhesion molecules, molecules involved in antigen presentation, and Fc receptor. Modulation of the expression of MHC class II antigens and other costimulatory molecules, such as B7 and ICAM-1, may result in changes in the antigen presenting capacity of monocytes and ability to induce T cell activation. Increased expression of Fc receptors may correlate with improved monocyte cytotoxic activity, cytokine release and phagocytosis.

[1079] FACS analysis is used to examine the surface antigens as follows. Monocytes are treated 1-5 days with increasing concentrations of agonists or antagonists of the invention or LPS (positive control), washed with PBS containing 1% BSA and 0.02 mM sodium azide, and then incubated with 1:20 dilution of appropriate FITC— or PE-labeled monoclonal antibodies for 30 minutes at 4 degrees C. After an additional wash, the labeled cells are analyzed by flow cytometry on a FACScan (Becton Dickinson).

[1080] Monocyte activation and/or increased survival. Assays for molecules that activate (or alternatively, inactivate) monocytes and/or increase monocyte survival (or alternatively, decrease monocyte survival) are known in the art and may routinely be applied to determine whether a molecule of the invention functions as an inhibitor or activator of monocytes. Agonists or antagonists of the invention can be screened using the three assays described below. For each of these assays, Peripheral blood mononuclear cells (PBMC) are purified from single donor leukopacks (American Red Cross, Baltimore, Md.) by centrifugation through a Histopaque gradient (Sigma). Monocytes are isolated from PBMC by counterflow centrifugal elutriation.

[1081] Monocyte Survival Assay. Human peripheral blood monocytes progressively lose viability when cultured in absence of serum or other stimuli. Their death results from internally regulated processes (apoptosis). Addition to the culture of activating factors, such as TNF-alpha dramatically improves cell survival and prevents DNA fragmentation. Propidium iodide (PI) staining is used to measure apoptosis as follows. Monocytes are cultured for 48 hours in polypropylene tubes in serum-free medium (positive control), in the presence of 100 ng/ml TNF-alpha (negative control), and in the presence of varying concentrations of the compound to be tested. Cells are suspended at a concentration of 2×106/ml in PBS containing PI at a final concentration of 5 μg/ml, and then incubated at room temperature for 5 minutes before FACScan analysis. PI uptake has been demonstrated to correlate with DNA fragmentation in this experimental paradigm.

[1082] Effect on cytokine release. An important function of monocytes/macrophages is their regulatory activity on other cellular populations of the immune system through the release of cytokines after stimulation. An ELISA to measure cytokine release is performed as follows. Human monocytes are incubated at a density of 5×105 cells/ml with increasing concentrations of agonists or antagonists of the invention and under the same conditions, but in the absence of agonists or antagonists. For IL-12 production, the cells are primed overnight with IFN (100 U/ml) in the presence of agonist or antagonist of the invention. LPS (10 ng/ml) is then added. Conditioned media are collected after 24 h and kept frozen until use. Measurement of TNF-alpha, IL-10, MCP-1 and IL-8 is then performed using a commercially available ELISA kit (e.g., R & D Systems (Minneapolis, Minn.)) and applying the standard protocols provided with the kit.

[1083] Oxidative burst. Purified monocytes are plated in 96-w plate at 2-1×105 cell/well. Increasing concentrations of agonists or antagonists of the invention are added to the wells in a total volume of 0.2 ml culture medium (RPMI 1640+10% FCS, glutamine and antibiotics). After 3 days incubation, the plates are centrifuged and the medium is removed from the wells. To the macrophage monolayers, 0.2 ml per well of phenol red solution (140 mM NaCl, 10 mM potassium phosphate buffer pH 7.0, 5.5 mM dextrose, 0.56 mM phenol red and 19 U/ml of HRPO) is added, together with the stimulant (200 nM PMA). The plates are incubated at 37° C. for 2 hours and the reaction is stopped by adding 20 μl 1N NaOH per well. The absorbance is read at 610 nm. To calculate the amount of H2O2 produced by the macrophages, a standard curve of a H2O2 solution of known molarity is performed for each experiment.

[1084] The studies described in this example tested activity of agonists or antagonists of the invention. However, one skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides or polypeptides of the invention (e.g., gene therapy).

Example 24

Biological Effects of Agonists or Antagonists of the Invention

[1085] Astrocyte and Neuronal Assays

[1086] Agonists or antagonists of the invention, expressed in Escherichia coli and purified as described above, can be tested for activity in promoting the survival, neurite outgrowth, or phenotypic differentiation of cortical neuronal cells and for inducing the proliferation of glial fibrillary acidic protein immunopositive cells, astrocytes. The selection of cortical cells for the bioassay is based on the prevalent expression of FGF-1 and FGF-2 in cortical structures and on the previously reported enhancement of cortical neuronal survival resulting from FGF-2 treatment. A thymidine incorporation assay, for example, can be used to elucidate an agonist or antagonist of the invention's activity on these cells.

[1087] Moreover, previous reports describing the biological effects of FGF-2 (basic FGF) on cortical or hippocampal neurons in vitro have demonstrated increases in both neuron survival and neurite outgrowth (Walicke et al., “Fibroblast growth factor promotes survival of dissociated hippocampal neurons and enhances neurite extension.” Proc. Natl. Acad. Sci. USA 83:3012-3016. (1986), assay herein incorporated by reference in its entirety). However, reports from experiments done on PC-12 cells suggest that these two responses are not necessarily synonymous and may depend on not only which FGF is being tested but also on which receptor(s) are expressed on the target cells. Using the primary cortical neuronal culture paradigm, the ability of an agonist or antagonist of the invention to induce neurite outgrowth can be compared to the response achieved with FGF-2 using, for example, a thymidine incorporation assay.

[1088] Fibroblast and Endothelial Cell Assays

[1089] Human lung fibroblasts are obtained from Clonetics (San Diego, Calif.) and maintained in growth media from Clonetics. Dermal microvascular endothelial cells are obtained from Cell Applications (San Diego, Calif.). For proliferation assays, the human lung fibroblasts and dermal microvascular endothelial cells can be cultured at 5,000 cells/well in a 96-well plate for one day in growth medium. The cells are then incubated for one day in 0.1% BSA basal medium. After replacing the medium with fresh 0.1% BSA medium, the cells are incubated with the test proteins for 3 days. Alamar Blue (Alamar Biosciences, Sacramento, Calif.) is added to each well to a final concentration of 10%. The cells are incubated for 4 hr. Cell viability is measured by reading in a CytoFluor fluorescence reader. For the PGE2 assays, the human lung fibroblasts are cultured at 5,000 cells/well in a 96-well plate for one day. After a medium change to 0.1% BSA basal medium, the cells are incubated with FGF-2 or agonists or antagonists of the invention with or without IL-1α for 24 hours. The supernatants are collected and assayed for PGE2 by EIA kit (Cayman, Ann Arbor, Mich.). For the IL-6 assays, the human lung fibroblasts are cultured at 5,000 cells/well in a 96-well plate for one day. After a medium change to 0.1% BSA basal medium, the cells are incubated with FGF-2 or with or without agonists or antagonists of the invention IL-1α for 24 hours. The supernatants are collected and assayed for IL-6 by ELISA kit (Endogen, Cambridge, Mass.).

[1090] Human lung fibroblasts are cultured with FGF-2 or agonists or antagonists of the invention for 3 days in basal medium before the addition of Alamar Blue to assess effects on growth of the fibroblasts. FGF-2 should show a stimulation at 10-2500 ng/ml which can be used to compare stimulation with agonists or antagonists of the invention.

[1091] Parkinson Models.

[1092] The loss of motor function in Parkinson's disease is attributed to a deficiency of striatal dopamine resulting from the degeneration of the nigrostriatal dopaminergic projection neurons. An animal model for Parkinson's that has been extensively characterized involves the systemic administration of 1-methyl-4 phenyl 1,2,3,6-tetrahydropyridine (MPTP). In the CNS, MPTP is taken-up by astrocytes and catabolized by monoamine oxidase B to 1-methyl-4-phenyl pyridine (MPP+) and released. Subsequently, MPP+ is actively accumulated in dopaminergic neurons by the high-affinity reuptake transporter for dopamine. MPP+ is then concentrated in mitochondria by the electrochemical gradient and selectively inhibits nicotidamide adenine disphosphate: ubiquinone oxidoreductionase (complex I), thereby interfering with electron transport and eventually generating oxygen radicals.

[1093] It has been demonstrated in tissue culture paradigms that FGF-2 (basic FGF) has trophic activity towards nigral dopaminergic neurons (Ferrari et al., Dev. Biol. 1989). Recently, Dr. Unsicker's group has demonstrated that administering FGF-2 in gel foam implants in the striatum results in the near complete protection of nigral dopaminergic neurons from the toxicity associated with MPTP exposure (Otto and Unsicker, J. Neuroscience, 1990).

[1094] Based on the data with FGF-2, agonists or antagonists of the invention call be evaluated to determine whether it has an action similar to that of FGF-2 in enhancing dopaminergic neuronal survival in vitro and it can also be tested in vivo for protection of dopaminergic neurons in the striatum from the damage associated with MPTP treatment. The potential effect of an agonist or antagonist of the invention is first examined in vitro in a dopaminergic neuronal cell culture paradigm. The cultures are prepared by dissecting the midbrain floor plate from gestation day 14 Wistar rat embryos. The tissue is dissociated with trypsin and seeded at a density of 200,000 cells/cm2 on polyorthinine-laminin coated glass coverslips. The cells are maintained in Dulbecco's Modified Eagle's medium and F12 medium containing hormonal supplements (N1). The cultures are fixed with paraformaldehyde after 8 days in vitro and are processed for tyrosine hydroxylase, a specific marker for dopaminergic neurons, immunohistochemical staining. Dissociated cell cultures are prepared from embryonic rats. The culture medium is changed every third day and the factors are also added at that time.

[1095] Since the dopaminergic neurons are isolated from animals at gestation day 14, a developmental time which is past the stage when the dopaminergic precursor cells are proliferating, an increase in the number of tyrosine hydroxylase immunopositive neurons would represent an increase in the number of dopaminergic neurons surviving in vitro. Therefore, if an agonist or antagonist of the invention acts to prolong the survival of dopaminergic neurons, it would suggest that the agonist or antagonist may be involved in Parkinson's Disease.

[1096] The studies described in this example tested activity of agonists or antagonists of the invention. However, one skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides or polypeptides of the invention (e.g., gene therapy).

Example 25

The Effect of Agonists or Antagonists of the Invention on the Growth of Vascular Endothelial Cells

[1097] On day 1, human umbilical vein endothelial cells (HUVEC) are seeded at 2-5×104 cells/35 mm dish density in M199 medium containing 4% fetal bovine serum (FBS), 16 units/ml heparin, and 50 units/ml endothelial cell growth supplements (ECGS, Biotechnique, Inc.). On day 2, the medium is replaced with Ml 99 containing 10% FBS, 8 units/ml heparin. An agonist or antagonist of the invention, and positive controls, such as VEGF and basic FGF (bFGF) are added, at varying concentrations. On days 4 and 6, the medium is replaced. On day 8, cell number is determined with a Coulter Counter.

[1098] An increase in the number of HUVEC cells indicates that the compound of the invention may proliferate vascular endothelial cells, while a decrease in the number of HUVEC cells indicates that the compound of the invention inhibits vascular endothelial cells.

[1099] The studies described in this example tested activity of a polypeptide of the invention. However, one skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides (e.g., gene therapy), agonists, and/or antagonists of the invention.

Example 26

Rat Corneal Wound Healing Model

[1100] This animal model shows the effect of an agonist or antagonist of the invention on neovascularization. The experimental protocol includes:

[1101] a) Making a 1-1.5 mm long incision from the center of cornea into the stromal layer.

[1102] b) Inserting a spatula below the lip of the incision facing the outer corner of the eye.

[1103] c) Making a pocket (its base is 1-1.5 mm form the edge of the eye).

[1104] d) Positioning a pellet, containing 50 ng-5 ug of an agonist or antagonist of the invention, within the pocket.

[1105] e) Treatment with an agonist or antagonist of the invention can also be applied topically to the corneal wounds in a dosage range of 20 mg-500 mg (daily treatment for five days).

[1106] The studies described in this example tested activity of agonists or antagonists of the invention. However, one skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides or polypeptides of the invention (e.g., gene therapy).

Example 27

Diabetic Mouse and Glucocorticoid-Impaired Wound Healing Models

[1107] Diabetic db+/db+ Mouse Model.

[1108] To demonstrate that an agonist or antagonist of the invention accelerates the healing process, the genetically diabetic mouse model of wound healing is used. The full thickness wound healing model in the db+/db+ mouse is a well characterized, clinically relevant and reproducible model of impaired wound healing. Healing of the diabetic wound is dependent on formation of granulation tissue and re-epithelialization rather than contraction (Gartner, M. H. et al., J. Surg. Res. 52:389 (1992); Greenhalgh, D. G. et al., Am. J. Pathol. 136:1235 (1990)).

[1109] The diabetic animals have many of the characteristic features observed in Type II diabetes mellitus. Homozygous (db+/db+) mice are obese in comparison to their normal heterozygous (db+/+m) littermates. Mutant diabetic (db+/db+) mice have a single autosomal recessive mutation on chromosome 4 (db+) (Coleman et al. Proc. Nat?. Acad. Sci. USA 77:283-293 (1982)). Animals show polyphagia, polydipsia and polyuria. Mutant diabetic mice (db+/db+) have elevated blood glucose, increased or normal insulin levels, and suppressed cell-mediated immunity (Mandel et al., J. Immunol. 120:1375 (1978); Debray-Sachs, M. et al., Clin. Exp. Immunol. 51(1):1-7 (1983); Leiter et al., Am. J. of Pathol. 114:46-55 (1985)). Peripheral neuropathy, myocardial complications, and microvascular lesions, basement membrane thickening and glomerular filtration abnormalities have been described in these animals (Norido, F. et al., Exp. Neurol. 83(2):221-232 (1984); Robertson et al., Diabetes 29(1):60-67 (1980); Giacomelli et al., Lab Invest. 40(4):460-473 (1979); Coleman, D. L., Diabetes 31 (Suppl):1-6 (1982)). These homozygous diabetic mice develop hyperglycemia that is resistant to insulin analogous to human type II diabetes (Mandel et al., J. Immunol. 120:1375-1377 (1978)).

[1110] The characteristics observed in these animals suggests that healing in this model may be similar to the healing observed in human diabetes (Greenhalgh, et al., Am. J. of Pathol. 136:1235-1246 (1990)).

[1111] Genetically diabetic female C57BL/KsJ (db+/db+) mice and their non-diabetic (db+/+m) heterozygous littermates are used in this study (Jackson Laboratories). The animals are purchased at 6 weeks of age and are 8 weeks old at the beginning of the study. Animals are individually housed and received food and water ad libitum. All manipulations are performed using aseptic techniques. The experiments are conducted according to the rules and guidelines of Human Genome Sciences, Inc. Institutional Animal Care and Use Committee and the Guidelines for the Care and Use of Laboratory Animals.

[1112] Wounding protocol is performed according to previously reported methods (Tsuboi, R. and Rifkin, D. B., J. Exp. Med. 172:245-251 (1990)). Briefly, on the day of wounding, animals are anesthetized with an intraperitoneal injection of Avertin (0.01 mg/mL), 2,2,2-tribromoethanol and 2-methyl-2-butanol dissolved in deionized water. The dorsal region of the animal is shaved and the skin washed with 70% ethanol solution and iodine. The surgical area is dried with sterile gauze prior to wounding. An 8 mm full-thickness wound is then created using a Keyes tissue punch. Immediately following wounding, the surrounding skin is gently stretched to eliminate wound expansion. The wounds are left open for the duration of the experiment. Application of the treatment is given topically for 5 consecutive days commencing on the day of wounding. Prior to treatment, wounds are gently cleansed with sterile saline and gauze sponges.

[1113] Wounds are visually examined and photographed at a fixed distance at the day of surgery and at two day intervals thereafter. Wound closure is determined by daily measurement on days 1-5 and on day 8. Wounds are measured horizontally and vertically using a calibrated Jameson caliper. Wounds are considered healed if granulation tissue is no longer visible and the wound is covered by a continuous epithelium.

[1114] An agonist or antagonist of the invention is administered using at a range different doses, from 4 mg to 500 mg per wound per day for 8 days in vehicle. Vehicle control groups received 50 mL of vehicle solution.

[1115] Animals are euthanized on day 8 with an intraperitoneal injection of sodium pentobarbital (300 mg/kg). The wounds and surrounding skin are then harvested for histology and immunohistochemistry. Tissue specimens are placed in 10% neutral buffered formalin in tissue cassettes between biopsy sponges for further processing.

[1116] Three groups of 10 animals each (5 diabetic and 5 non-diabetic controls) are evaluated: 1) Vehicle placebo control, 2) untreated group, and 3) treated group.

[1117] Wound closure is analyzed by measuring the area in the vertical and horizontal axis and obtaining the total square area of the wound. Contraction is then estimated by establishing the differences between the initial wound area (day 0) and that of post treatment (day 8). The wound area on day 1 is 64 mm2, the corresponding size of the dermal punch. Calculations are made using the following formula:

[Open area on day 8]−[Open area on day 1]/[Open area on day 1]

[1118] Specimens are fixed in 10% buffered formalin and paraffin embedded blocks are sectioned perpendicular to the wound surface (5 mm) and cut using a Reichert-Jung microtome. Routine hematoxylin-eosin (H&E) staining is performed on cross-sections of bisected wounds. Histologic examination of the wounds are used to assess whether the healing process and the morphologic appearance of the repaired skin is altered by treatment with an agonist or antagonist of the invention. This assessment included verification of the presence of cell accumulation, inflammatory cells, capillaries, fibroblasts, re-epithelialization and epidermal maturity (Greenhalgh, D. G. et al., Am. J. Pathol. 136:1235 (1990)). A calibrated lens micrometer is used by a blinded observer.

[1119] Tissue sections are also stained immunohistochemically with a polyclonal rabbit anti-human keratin antibody using ABC Elite detection system. Human skin is used as a positive tissue control while non-immune IgG is used as a negative control. Keratinocyte growth is determined by evaluating the extent of reepithelialization of the wound using a calibrated lens micrometer.

[1120] Proliferating cell nuclear antigen/cyclin (PCNA) in skin specimens is demonstrated by using anti-PCNA antibody (1:50) with an ABC Elite detection system. Human colon cancer served as a positive tissue control and human brain tissue is used as a negative tissue control. Each specimen included a section with omission of the primary antibody and substitution with non-immune mouse IgG. Ranking of these sections is based on the extent of proliferation on a scale of 0-8, the lower side of the scale reflecting slight proliferation to the higher side reflecting intense proliferation.

[1121] Experimental data are analyzed using an unpaired t test. A p value of <0.05 is considered significant.

[1122] Steroid Impaired Rat Model

[1123] The inhibition of wound healing by steroids has been well documented in various in vitro and in vivo systems (Wahl, Glucocorticoids and Wound healing. In: Anti-Inflammatory Steroid Action: Basic and Clinical Aspects. 280-302 (1989); Wahlet al., J. Immunol. 115: 476-481 (1975); Werb et al., J. Exp. Med. 147:1684-1694 (1978)). Glucocorticoids retard wound healing by inhibiting angiogenesis, decreasing vascular permeability (Ebert et al., An. Intern. Med. 37:701-705 (1952)), fibroblast proliferation, and collagen synthesis (Beck et al., Growth Factors. 5: 295-304 (1991); Haynes et al., J. Clin. Invest. 61: 703-797 (1978)) and producing a transient reduction of circulating monocytes (Haynes et al., J. Clin. Invest. 61: 703-797 (1978); Wahl, “Glucocorticoids and wound healing”, In: Antiinflammatory Steroid Action: Basic and Clinical Aspects, Academic Press, New York, pp. 280-302 (1989)). The systemic administration of steroids to impaired wound healing is a well establish phenomenon in rats (Beck et al., Growth Factors. 5: 295-304 (1991); Haynes et al., J. Clin. Invest. 61: 703-797 (1978); Wahl, “Glucocorticoids and wound healing”, In: Antiinflammatory Steroid Action: Basic and Clinical Aspects, Academic Press, New York, pp. 280-302 (1989); Pierce et al., Proc. Natl. Acad. Sci. USA 86: 2229-2233 (1989)).

[1124] To demonstrate that an agonist or antagonist of the invention can accelerate the healing process, the effects of multiple topical applications of the agonist or antagonist on full thickness excisional skin wounds in rats in which healing has been impaired by the systemic administration of methylprednisolone is assessed.

[1125] Young adult male Sprague Dawley rats weighing 250-300 g (Charles River Laboratories) are used in this example. The animals are purchased at 8 weeks of age and are 9 weeks old at the beginning of the study. The healing response of rats is impaired by the systemic administration of methylprednisolone (17 mg/kg/rat intramuscularly) at the time of wounding. Animals are individually housed and received food and water ad libitum. All manipulations are performed using aseptic techniques. This study is conducted according to the rules and guidelines of Human Genome Sciences, Inc. Institutional Animal Care and Use Committee and the Guidelines for the Care and Use of Laboratory Animals.

[1126] The wounding protocol is followed according to section A, above. On the day of wounding, animals are anesthetized with an intramuscular injection of ketamine (50 mg/kg) and xylazine (5 mg/kg). The dorsal region of the animal is shaved and the skin washed with 70% ethanol and iodine solutions. The surgical area is dried with sterile gauze prior to wounding. An 8 mm full-thickness wound is created using a Keyes tissue punch. The wounds are left open for the duration of the experiment. Applications of the testing materials are given topically once a day for 7 consecutive days commencing on the day of wounding and subsequent to methylprednisolone administration. Prior to treatment, wounds are gently cleansed with sterile saline and gauze sponges.

[1127] Wounds are visually examined and photographed at a fixed distance at the day of wounding and at the end of treatment. Wound closure is determined by daily measurement on days 1-5 and on day 8. Wounds are measured horizontally and vertically using a calibrated Jameson caliper. Wounds are considered healed if granulation tissue is no longer visible and the wound is covered by a continuous epithelium.

[1128] The agonist or antagonist of the invention is administered using at a range different doses, from 4 mg to 500 mg per wound per day for 8 days in vehicle. Vehicle control groups received 50 mL of vehicle solution.

[1129] Animals are euthanized on day 8 with an intraperitoneal injection of sodium pentobarbital (300 mg/kg). The wounds and surrounding skin are then harvested for histology. Tissue specimens are placed in 10% neutral buffered formalin in tissue cassettes between biopsy sponges for further processing.

[1130] Three groups of 10 animals each (5 with methylprednisolone and 5 without glucocorticoid) are evaluated: 1) Untreated group 2) Vehicle placebo control 3) treated groups.

[1131] Wound closure is analyzed by measuring the area in the vertical and horizontal axis and obtaining the total area of the wound. Closure is then estimated by establishing the differences between the initial wound area (day 0) and that of post treatment (day 8). The wound area on day 1 is 64 mm2, the corresponding size of the dermal punch. Calculations are made using the following formula:

[Open area on day 8]−[Open area on day 1]/[Open area on day 1]

[1132] Specimens are fixed in 10% buffered formalin and paraffin embedded blocks are sectioned perpendicular to the wound surface (5 mm) and cut using an Olympus microtome. Routine hematoxylin-eosin (H&E) staining is performed on cross-sections of bisected wounds. Histologic examination of the wounds allows assessment of whether the healing process and the morphologic appearance of the repaired skin is improved by treatment with an agonist or antagonist of the invention. A calibrated lens micrometer is used by a blinded observer to determine the distance of the wound gap.

[1133] Experimental data are analyzed using an unpaired t test. A p value of <0.05 is considered significant.

[1134] The studies described in this example tested activity of agonists or antagonists of the invention. However, one skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides or polypeptides of the invention (e.g., gene therapy).

Example 28

Lymphadema Animal Model

[1135] The purpose of this experimental approach is to create an appropriate and consistent lymphedema model for testing the therapeutic effects of an agonist or antagonist of the invention in lymphangiogenesis and re-establishment of the lymphatic circulatory system in the rat hind limb. Effectiveness is measured by swelling volume of the affected limb, quantification of the amount of lymphatic vasculature, total blood plasma protein, and histopathology. Acute lymphedema is observed for 7-10 days. Perhaps more importantly, the chronic progress of the edema is followed for up to 3-4 weeks.

[1136] Prior to beginning surgery, blood sample is drawn for protein concentration analysis. Male rats weighing approximately ˜350 g are dosed with Pentobarbital. Subsequently, the right legs are shaved from knee to hip. The shaved area is swabbed with gauze soaked in 70% EtOH. Blood is drawn for serum total protein testing. Circumference and volumetric measurements are made prior to injecting dye into paws after marking 2 measurement levels (0.5 cm above heel, at mid-pt of dorsal paw). The intradermal dorsum of both right and left paws are injected with 0.05 ml of 1% Evan's Blue. Circumference and volumetric measurements are then made following injection of dye into paws.

[1137] Using the knee joint as a landmark, a mid-leg inguinal incision is made circumferentially allowing the femoral vessels to be located. Forceps and hemostats are used to dissect and separate the skin flaps. After locating the femoral vessels, the lymphatic vessel that runs along side and underneath the vessel(s) is located. The main lymphatic vessels in this area are then electrically coagulated or suture ligated.

[1138] Using a microscope, muscles in back of the leg (near the semitendinosis and adductors) are bluntly dissected. The popliteal lymph node is then located. The 2 proximal and 2 distal lymphatic vessels and distal blood supply of the popliteal node are then ligated by suturing. The popliteal lymph node, and any accompanying adipose tissue, is then removed by cutting connective tissues.

[1139] Care is taken to control any mild bleeding resulting from this procedure. After lymphatics are occluded, the skin flaps are sealed by using liquid skin (Vetbond) (AJ Buck). The separated skin edges are sealed to the underlying muscle tissue while leaving a gap of ˜0.5 cm around the leg. Skin also may be anchored by suturing to underlying muscle when necessary.

[1140] To avoid infection, animals are housed individually with mesh (no bedding). Recovering animals are checked daily through the optimal edematous peak, which typically occurred by day 5-7. The plateau edematous peak are then observed. To evaluate the intensity of the lymphedema, the circumference and volumes of 2 designated places on each paw before operation and daily for 7 days are measured. The effect of plasma proteins on lymphedema is determined and whether protein analysis is a useful testing perimeter is also investigated. The weights of both control and edematous limbs are evaluated at 2 places. Analysis is performed in a blind manner.

[1141] Circumference Measurements: Under brief gas anesthetic to prevent limb movement, a cloth tape is used to measure limb circumference. Measurements are done at the ankle bone and dorsal paw by 2 different people and those 2 readings are averaged. Readings are taken from both control and edematous limbs.

[1142] Volumetric Measurements: On the day of surgery, animals are anesthetized with Pentobarbital and are tested prior to surgery. For daily volumetrics animals are under brief halothane anesthetic (rapid immobilization and quick recovery), and both legs are shaved and equally marked using waterproof marker on legs. Legs are first dipped in water, then dipped into instrument to each marked level then measured by Buxco edema software(Chen/Victor). Data is recorded by one person, while the other is dipping the limb to marked area.

[1143] Blood-plasma protein measurements: Blood is drawn, spun, and serum separated prior to surgery and then at conclusion for total protein and Ca2+ comparison.

[1144] Limb Weight Comparison: After drawing blood, the animal is prepared for tissue collection. The limbs are amputated using a quillitine, then both experimental and control legs are cut at the ligature and weighed. A second weighing is done as the tibio-cacaneal joint is disarticulated and the foot is weighed.

[1145] Histological Preparations: The transverse muscle located behind the knee (popliteal) area is dissected and arranged in a metal mold, filled with freezeGel, dipped into cold methylbutane, placed into labeled sample bags at −80EC until sectioning. Upon sectioning, the muscle is observed under fluorescent microscopy for lymphatics.

[1146] The studies described in this example tested activity of agonists or antagonists of the invention. However, one skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides or polypeptides of the invention (e.g., gene therapy).

Example 29

Suppression of TNF Alpha-Induced Adhesion Molecule Expression by an Agonist or Antagonist of the Invention

[1147] The recruitment of lymphocytes to areas of inflammation and angiogenesis involves specific receptor-ligand interactions between cell surface adhesion molecules (CAMs) on lymphocytes and the vascular endothelium. The adhesion process, in both normal and pathological settings, follows a multi-step cascade that involves intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and endothelial leukocyte adhesion molecule-1 (E-selectin) expression on endothelial cells (EC). The expression of these molecules and others on the vascular endothelium determines the efficiency with which leukocytes may adhere to the local vasculature and extravasate into the local tissue during the development of an inflammatory response. The local concentration of cytokines and growth factor participate in the modulation of the expression of these CAMs.

[1148] Tumor necrosis factor alpha (TNF-a), a potent proinflammatory cytokine, is a stimulator of all three CAMs on endothelial cells and may be involved in a wide variety of inflammatory responses, often resulting in a pathological outcome.

[1149] The potential of an agonist or antagonist of the invention to mediate a suppression of TNF-a induced CAM expression can be examined. A modified ELISA assay which uses ECs as a solid phase absorbent is employed to measure the amount of CAM expression on TNF-a treated ECs when co-stimulated with a member of the FGF family of proteins.

[1150] To perform the experiment, human umbilical vein endothelial cell (HUVEC) cultures are obtained from pooled cord harvests and maintained in growth medium (EGM-2; Clonetics, San Diego, Calif.) supplemented with 10% FCS and 1% penicillin/streptomycin in a 37 degree C. humidified incubator containing 5% CO2. HUVECs are seeded in 96-well plates at concentrations of 1×104 cells/well in EGM medium at 37 degree C. for 18-24 hrs or until confluent. The monolayers are subsequently washed 3 times with a serum-free solution of RPMI-1640 supplemented with 100 U/ml penicillin and 100 mg/ml streptomycin, and treated with a given cytokine and/or growth factor(s) for 24 h at 37 degree C. Following incubation, the cells are then evaluated for CAM expression.

[1151] Human Umbilical Vein Endothelial cells (HUVECs) are grown in a standard 96 well plate to confluence. Growth medium is removed from the cells and replaced with 90 ul of 199 Medium (10% FBS). Samples for testing and positive or negative controls are added to the plate in triplicate (in 10 ul volumes). Plates are incubated at 37 degree C. for either 5 h (selectin and integrin expression) or 24 h (integrin expression only). Plates are aspirated to remove medium and 100 μl of 0.1% paraformaldehyde-PBS(with Ca++ and Mg++) is added to each well. Plates are held at 4° C. for 30 min.

[1152] Fixative is then removed from the wells and wells are washed 1× with PBS(+Ca,Mg)+0.5% BSA and drained. Do not allow the wells to dry. Add 10 μl of diluted primary antibody to the test and control wells. Anti-ICAM-1-Biotin, Anti-VCAM-1-Biotin and Anti-E-selectin-Biotin are used at a concentration of 10 μg/ml (1:10 dilution of 0.1 mg/ml stock antibody). Cells are incubated at 37° C. for 30 min. in a humidified environment. Wells are washed ×3 with PBS(+Ca,Mg)+0.5% BSA.

[1153] Then add 20 μl of diluted ExtrAvidin-Alkaline Phosphotase (1:5,000 dilution) to each well and incubated at 37° C. for 30 min. Wells are washed X3 with PBS(+Ca,Mg)+0.5% BSA. 1 tablet of p-Nitrophenol Phosphate pNPP is dissolved in 5 ml of glycine buffer (pH 10.4). 100 μl of pNPP substrate in glycine buffer is added to each test well. Standard wells in triplicate are prepared from the working dilution of the ExtrAvidin-Alkaline Phosphotase in glycine buffer: 1:5,000 (100)>10−0.5>10−1>10−1.5. 5 μl of each dilution is added to triplicate wells and the resulting AP content in each well is 5.50 ng, 1.74 ng, 0.55 ng, 0.18 ng. 100 μl of pNNP reagent must then be added to each of the standard wells. The plate must be incubated at 37° C. for 4 h. A volume of 50 μl of 3M NaOH is added to all wells. The results are quantified on a plate reader at 405 nm. The background subtraction option is used on blank wells filled with glycine buffer only. The template is set up to indicate the concentration of AP-conjugate in each standard well [5.50 ng; 1.74 ng; 0.55 ng; 0.18 ng]. Results are indicated as amount of bound AP-conjugate in each sample.

[1154] The studies described in this example tested activity of agonists or antagonists of the invention. However, one skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides or polypeptides of the invention (e.g., gene therapy).

Example 30

Production of Polypeptide of the Invention For High-Throughput Screening Assays

[1155] The following protocol produces a supernatant containing polypeptide of the present invention to be tested. This supernatant can then be used in the Screening Assays described in Examples 32-41.

[1156] First, dilute Poly-D-Lysine (644 587 Boehringer-Mannheim) stock solution (1 mg/ml in PBS) 1:20 in PBS (w/o calcium or magnesium 17-516F Biowhittaker) for a working solution of 50 ug/ml. Add 200 ul of this solution to each well (24 well plates) and incubate at RT for 20 minutes. Be sure to distribute the solution over each well (note: a 12-channel pipetter may be used with tips on every other channel). Aspirate off the Poly-D-Lysine solution and rinse with 1 ml PBS (Phosphate Buffered Saline). The PBS should remain in the well until just prior to plating the cells and plates may be poly-lysine coated in advance for up to two weeks.

[1157] Plate 293T cells (do not carry cells past P+20) at 2×105 cells/well in 0.5 ml DMEM(Dulbecco's Modified Eagle Medium)(with 4.5 G/L glucose and L-glutamine (12-604F Biowhittaker))/10% heat inactivated FBS(14-503F Biowhittaker)/1×Penstrep(17-602E Biowhittaker). Let the cells grow overnight.

[1158] The next day, mix together in a sterile solution basin: 300 ul Lipofectamine (18324-012 Gibco/BRL) and 5 ml Optimem 1 (31985070 Gibco/BRL)/96-well plate. With a small volume multi-channel pipetter, aliquot approximately 2 ug of an expression vector containing a polynucleotide insert, produced by the methods described in Examples 8-10, into an appropriately labeled 96-well round bottom plate. With a multi-channel pipetter, add 50 ul of the Lipofectamine/Optimem I mixture to each well. Pipette up and down gently to mix. Incubate at RT 15-45 minutes. After about 20 minutes, use a multi-channel pipetter to add 150 ul Optimem I to each well. As a control, one plate of vector DNA lacking an insert should be transfected with each set of transfections.

[1159] Preferably, the transfection should be performed by tag-teaming the following tasks. By tag-teaming, hands on time is cut in half, and the cells do not spend too much time on PBS. First, person A aspirates off the media from four 24-well plates of cells, and then person B rinses each well with 0.5-1 ml PBS. Person A then aspirates off PBS rinse, and person B, using al2-channel pipetter with tips on every other channel, adds the 200 ul of DNA/Lipofectamine/Optimem I complex to the odd wells first, then to the even wells, to each row on the 24-well plates. Incubate at 37 degree C. for 6 hours.

[1160] While cells are incubating, prepare appropriate media, either 1%BSA in DMEM with 1× penstrep, or HGS CHO-5 media (116.6 mg/L of CaCl2 (anhyd); 0.00130 mg/L CuSO4-5H2O; 0.050 mg/L of Fe(NO3)3-9H2O; 0.417 mg/L of FeSO4-7H2O; 311.80 mg/L of Kcl; 28.64 mg/L of MgCl2; 48.84 mg/L of MgSO4; 6995.50 mg/L of NaCl; 2400.0 mg/L of NaHCO3; 62.50 mg/L of NaH2PO4—H2O; 71.02 mg/L of Na2HPO4; 0.4320 mg/L of ZnSO4-7H2O; 0.002 mg/L of Arachidonic Acid; 1.022 mg/L of Cholesterol; 0.070 mg/L of DL-alpha-Tocopherol-Acetate; 0.0520 mg/L of Linoleic Acid; 0.010 mg/L of Linolenic Acid; 0.010 mg/L of Myristic Acid; 0.010 mg/L of Oleic Acid; 0.010 mg/L of Palmitric Acid; 0.010 mg/L of Palmitic Acid; 100 mg/L of Pluronic F-68; 0.010 mg/L of Stearic Acid; 2.20 mg/L of Tween 80; 4551 mg/L of D-Glucose; 130.85 mg/ml of L-Alanine; 147.50 mg/ml of L-Arginine-HCL; 7.50 mg/ml of L-Asparagine-H2O; 6.65 mg/ml of L-Aspartic Acid; 29.56 mg/ml of L-Cystine-2HCL-H2O; 31.29 mg/ml of L-Cystine-2HCL; 7.35 mg/ml of L-Glutamic Acid; 365.0 mg/ml of L-Glutamine; 18.75 mg/ml of Glycine; 52.48 mg/ml of L-Histidine-HCL-H2O; 106.97 mg/ml of L-Isoleucine; 111.45 mg/ml of L-Leucine; 163.75 mg/ml of L-Lysine HCL; 32.34 mg/ml of L-Methionine; 68.48 mg/ml of L-Phenylalainine; 40.0 mg/ml of L-Proline; 26.25 mg/ml of L-Serine; 101.05 mg/ml of L-Threonine; 19.22 mg/ml of L-Tryptophan; 91.79 mg/ml of L-Tryrosine-2Na-2H2O; and 99.65 mg/ml of L-Valine; 0.0035 mg/L of Biotin; 3.24 mg/L of D-Ca Pantothenate; 11.78 mg/L of Choline Chloride; 4.65 mg/L of Folic Acid; 15.60 mg/L of i-Inositol; 3.02 mg/L of Niacinamide; 3.00 mg/L of Pyridoxal HCL; 0.031 mg/L of Pyridoxine HCL; 0.319 mg/L of Riboflavin; 3.17 mg/L of Thiamine HCL; 0.365 mg/L of Thymidine; 0.680 mg/L of Vitamin B12; 25 mM of HEPES Buffer; 2.39 mg/L of Na Hypoxanthine; 0.105 mg/L of Lipoic Acid; 0.081 mg/L of Sodium Putrescine-2HCL; 55.0 mg/L of Sodium Pyruvate; 0.0067 mg/L of Sodium Selenite; 20 uM of Ethanolamine; 0.122 mg/L of Ferric Citrate; 41.70 mg/L of Methyl-B-Cyclodextrin complexed with Linoleic Acid; 33.33 mg/L of Methyl-B-Cyclodextrin complexed with Oleic Acid; 10 mg/L of Methyl-B-Cyclodextrin complexed with Retinal Acetate. Adjust osmolarity to 327 mOsm) with 2 mm glutamine and 1× penstrep. (BSA (81-068-3 Bayer) 100 gm dissolved in IL DMEM for a 10% BSA stock solution). Filter the media and collect 50 ul for endotoxin assay in 15 ml polystyrene conical.

[1161] The transfection reaction is terminated, preferably by tag-teaming, at the end of the incubation period. Person A aspirates off the transfection media, while person B adds 1.5 ml appropriate media to each well. Incubate at 37 degree C. for 45 or 72 hours depending on the media used: 1%BSA for 45 hours or CHO-5 for 72 hours.

[1162] On day four, using a 300 ul multichannel pipetter, aliquot 600 ul in one 1 ml deep well plate and the remaining supernatant into a 2 ml deep well. The supernatants from each well can then be used in the assays described in Examples 32-39.

[1163] It is specifically understood that when activity is obtained in any of the assays described below using a supernatant, the activity originates from either the polypeptide of the present invention directly (e.g., as a secreted protein) or by polypeptide of the present invention inducing expression of other proteins, which are then secreted into the supernatant. Thus, the invention further provides a method of identifying the protein in the supernatant characterized by an activity in a particular assay.

Example 31

Construction of GAS Reporter Construct

[1164] One signal transduction pathway involved in the differentiation and proliferation of cells is called the Jaks-STATs pathway. Activated proteins in the Jaks-STATs pathway bind to gamma activation site “GAS” elements or interferon-sensitive responsive element (“ISRE”), located in the promoter of many genes. The binding of a protein to these elements alter the expression of the associated gene.

[1165] GAS and ISRE elements are recognized by a class of transcription factors called Signal Transducers and Activators of Transcription, or “STATs.” There are six members of the STATs family. Stat1 and Stat3 are present in many cell types, as is Stat2 (as response to IFN-alpha is widespread). Stat4 is more restricted and is not in many cell types though it has been found in T helper class I, cells after treatment with IL-12. Stat5 was originally called mammary growth factor, but has been found at higher concentrations in other cells including myeloid cells. It can be activated in tissue culture cells by many cytokines.

[1166] The STATs are activated to translocate from the cytoplasm to the nucleus upon tyrosine phosphorylation by a set of kinases known as the Janus Kinase (“Jaks”) family. Jaks represent a distinct family of soluble tyrosine kinases and include Tyk2, Jak1, Jak2, and Jak3. These kinases display significant sequence similarity and are generally catalytically inactive in resting cells.

[1167] The Jaks are activated by a wide range of receptors summarized in the Table below. (Adapted from review by Schidler and Darnell, Ann. Rev. Biochem. 64:621-51 (1995)). A cytokine receptor family, capable of activating Jaks, is divided into two groups: (a) Class I includes receptors for IL-2, IL-3, IL-4, IL-6, IL-7, IL-9, IL-11, IL-12, IL-15, Epo, PRL, GH, G-CSF, GM-CSF, LIF, CNTF, and thrombopoietin; and (b) Class 2 includes IFN-a, IFN-g, and IL-10. The Class 1 receptors share a conserved cysteine motif (a set of four conserved cysteines and one tryptophan) and a WSXWS motif (a membrane proximal region encoding Trp-Ser-Xaa-Trp-Ser (SEQ ID NO: 2)).

[1168] Thus, on binding of a ligand to a receptor, Jaks are activated, which in turn activate STATs, which then translocate and bind to GAS elements. This entire process is encompassed in the Jaks-STATs signal transduction pathway. Therefore, activation of the Jaks-STATs pathway, reflected by the binding of the GAS or the ISRE element, can be used to indicate proteins involved in the proliferation and differentiation of cells. For example, growth factors and cytokines are known to activate the Jaks-STATs pathway (See Table below). Thus, by using GAS elements linked to reporter molecules, activators of the Jaks-STATs pathway can be identified.

12JAKsLigandtyk2Jak1Jak2Jak3STATSGAS (elements) or ISREIFN familyIFN-a/B++−−1, 2, 3ISREIFN-g++−1GAS (IRF1 > Lys6 > IFP)Il-10+??−1, 3gp130 familyIL-6 (Pleiotropic)+++?1, 3GAS (IRF1 > Lys6 > IFP)Il-11 (Pleiotropic)?+??1, 3OnM (Pleiotropic)?++?1, 3LIF (Pleiotropic)?++?1, 3CNTF (Pleiotropic)−/++++1, 3G-CSF (Pleiotropic)?+??1, 3IL-12 (Pleiotropic)+−++1, 3g-C familyIL-2 (lymphocytes)−+−+1, 3, 5GASIL-4 (lymph/myeloid)−+−+6GAS (IRF1 = IFP >> Ly6)(IgH)IL-7 (lymphocytes)−+−+5GASIL-9 (lymphocytes)−+−+5GASIL-13 (lymphocyte)−+??6GASIL-15?+?+5GASgp140 familyIL-3 (myeloid)−−+−5GAS (IRF1 > IFP >> Ly6)IL-5 (myeloid)−−+−5GASGM-CSF (myeloid)−−+−5GASGrowth hormone familyGH?−+−5PRL?+/−+−1, 3, 5EPO?−+−5GAS (B-CAS > IRF1 = IFP >> Ly6)Receptor Tyrosine KinasesEGF?++−1, 3GAS(IRF1)PDGF?++−1, 3CSF-1?++−1, 3GAS (not IRF1)

[1169] To construct a synthetic GAS containing promoter element, which is used in the Biological Assays described in Examples 32-33, a PCR based strategy is employed to generate a GAS-SV40 promoter sequence. The 5′ primer contains four tandem copies of the GAS binding site found in the IRFI promoter and previously demonstrated to bind STATs upon induction with a range of cytokines (Rothman et al., Immunity 1:457-468 (1994).), although other GAS or ISRE elements can be used instead. The 5′ primer also contains 18 bp of sequence complementary to the SV40 early promoter sequence and is flanked with an XhoI site. The sequence of the 5′ primer is:

13(SEQ ID NO: 3)5′:GCGCCTCGAGATTTCCCCGAAATCTAGATTTCCCCGAAATGATTTCCCCGAAATGATTTCCCCGAAATATCTGCCATCTCAATTAG:3′

[1170] The downstream primer is complementary to the SV40 promoter and is flanked with a Hind III site: 5′:GCGGCAAGCTTTTTGCAAAGCCTAGGC:3′ (SEQ ID NO: 4)

[1171] PCR amplification is performed using the SV40 promoter template present in the B-gal:promoter plasmid obtained from Clontech. The resulting PCR fragment is digested with XhoI/Hind III and subcloned into BLSK2-. (Stratagene.) Sequencing with forward and reverse primers confirms that the insert contains the following sequence:

14(SEQ ID NO: 5)5′:CTCGAGATTTCCCCGAAATCTAGATTTCCCCGAAATGATTTCCCCGAAATGATTTCCCCGAAATATCTGCCATCTCAATTAGTCAGCAACCATAGTCCCGCCCCTAACTCCGCCCATCCCGCCCCTAACTCCGCCCAGTTCCGCCCATTCTCCGCCCCATGGCTGACTAATTTTTTTTATTTATGCAGAGGCCGAGGCCGCCTCGGCCTCTGAGCTATTCCAGAAGTAGTGAGGAGGCTTTTTTGGAGGCCTAGGCTTTTGCAAAAAGCTT:3′

[1172] With this GAS promoter element linked to the SV40 promoter, a GAS:SEAP2 reporter construct is next engineered. Here, the reporter molecule is a secreted alkaline phosphatase, or “SEAP.” Clearly, however, any reporter molecule can be instead of SEAP, in this or in any of the other Examples. Well known reporter molecules that can be used instead of SEAP include chloramphenicol acetyltransferase (CAT), luciferase, alkaline phosphatase, B-galactosidase, green fluorescent protein (GFP), or any protein detectable by an antibody.

[1173] The above sequence confirmed synthetic GAS-SV40 promoter element is subcloned into the pSEAP-Promoter vector obtained from Clontech using HindIII and XhoI, effectively replacing the SV40 promoter with the amplified GAS:SV40 promoter element, to create the GAS-SEAP vector. However, this vector does not contain a neomycin resistance gene, and therefore, is not preferred for mammalian expression systems.

[1174] Thus, in order to generate mammalian stable cell lines expressing the GAS-SEAP reporter, the GAS-SEAP cassette is removed from the GAS-SEAP vector using SalI and NotI, and inserted into a backbone vector containing the neomycin resistance gene, such as pGFP-1 (Clontech), using these restriction sites in the multiple cloning site, to create the GAS-SEAP/Neo vector. Once this vector is transfected into mammalian cells, this vector can then be used as a reporter molecule for GAS binding as described in Examples 32-33.

[1175] Other constructs can be made using the above description and replacing GAS with a different promoter sequence. For example, construction of reporter molecules containing EGR and NF-KB promoter sequences are described in Examples 34 and 35. However, many other promoters can be substituted using the protocols described in these Examples. For instance, SRE, IL-2, NFAT, or Osteocalcin promoters can be substituted, alone or in combination (e.g., GAS/NF-KB/EGR, GAS/NF-KB, Il-2/NFAT, or NF-KB/GAS). Similarly, other cell lines can be used to test reporter construct activity, such as HELA (epithelial), HUVEC (endothelial), Reh (B-cell), Saos-2 (osteoblast), HUVAC (aortic), or Cardiomyocyte.

Example 32

High-Throughput Screening Assay for T-cell Activity

[1176] The following protocol is used to assess T-cell activity by identifying factors, and determining whether sup emate containing a polypeptide of the invention proliferates and/or differentiates T-cells. T-cell activity is assessed using the GAS/SEAP/Neo construct produced in Example 31. Thus, factors that increase SEAP activity indicate the ability to activate the Jaks-STATS signal transduction pathway. The T-cell used in this assay is Jurkat T-cells (ATCC Accession No. TIB-152), although Molt-3 cells (ATCC Accession No. CRL-1552) and Molt-4 cells (ATCC Accession No. CRL-1582) cells can also be used.

[1177] Jurkat T-cells are lymphoblastic CD4+ Th1 helper cells. In order to generate stable cell lines, approximately 2 million Jurkat cells are transfected with the GAS-SEAP/neo vector using DMRIE-C (Life Technologies)(transfection procedure described below). The transfected cells are seeded to a density of approximately 20,000 cells per well and transfectants resistant to 1 mg/ml genticin selected. Resistant colonies are expanded and then tested for their response to increasing concentrations of interferon gamma. The dose response of a selected clone is demonstrated.

[1178] Specifically, the following protocol will yield sufficient cells for 75 wells containing 200 ul of cells. Thus, it is either scaled up, or performed in multiple to generate sufficient cells for multiple 96 well plates. Jurkat cells are maintained in RPMI+10% serum with 1%Pen-Strep. Combine 2.5 mls of OPTI-MEM (Life Technologies) with 10 ug of plasmid DNA in a T25 flask. Add 2.5 ml OPTI-MEM containing 50 ul of DMRIE-C and incubate at room temperature for 15-45 mins.

[1179] During the incubation period, count cell concentration, spin down the required number of cells (107 per transfection), and resuspend in OPTI-MEM to a final concentration of 107 cells/ml. Then add 1 ml of 1×107 cells in OPTI-MEM to T25 flask and incubate at 37 degree C. for 6 hrs. After the incubation, add 10 ml of RPMI+15% serum.

[1180] The Jurkat:GAS-SEAP stable reporter lines are maintained in RPMI+10% serum, 1 mg/ml Genticin, and 1% Pen-Strep. These cells are treated with supernatants containing polypeptide of the present invention or polypeptide of the present invention induced polypeptides as produced by the protocol described in Example 30.

[1181] On the day of treatment with the supernatant, the cells should be washed and resuspended in fresh RPMI+10% serum to a density of 500,000 cells per ml. The exact number of cells required will depend on the number of supernatants being screened. For one 96 well plate, approximately 10 million cells (for 10 plates, 100 million cells) are required.

[1182] Transfer the cells to a triangular reservoir boat, in order to dispense the cells into a 96 well dish, using a 12 channel pipette. Using a 12 channel pipette, transfer 200 ul of cells into each well (therefore adding 100, 000 cells per well).

[1183] After all the plates have been seeded, 50 ul of the supernatants are transferred directly from the 96 well plate containing the supernatants into each well using a 12 channel pipette. In addition, a dose of exogenous interferon gamma (0.1, 1.0, 10 ng) is added to wells H9, H10, and H11 to serve as additional positive controls for the assay.

[1184] The 96 well dishes containing Jurkat cells treated with supernatants are placed in an incubator for 48 hrs (note: this time is variable between 48-72 hrs). 35 ul samples from each well are then transferred to an opaque 96 well plate using a 12 channel pipette. The opaque plates should be covered (using sellophene covers) and stored at −20 degree C. until SEAP assays are performed according to Example 36. The plates containing the remaining treated cells are placed at 4 degree C. and serve as a source of material for repeating the assay on a specific well if desired.

[1185] As a positive control, 100 Unit/ml interferon gamma can be used which is known to activate Jurkat T cells. Over 30 fold induction is typically observed in the positive control wells.

[1186] The above protocol may be used in the generation of both transient, as well as, stable transfected cells, which would be apparent to those of skill in the art.

Example 33

High-Throughput Screening Assay Identifying Myeloid Activity

[1187] The following protocol is used to assess myeloid activity of polypeptide of the present invention by determining whether polypeptide of the present invention proliferates and/or differentiates myeloid cells. Myeloid cell activity is assessed using the GAS/SEAPINeo construct produced in Example 31. Thus, factors that increase SEAP activity indicate the ability to activate the Jaks-STATS signal transduction pathway. The myeloid cell used in this assay is U937, a pre-monocyte cell line, although TF-1, HL60, or KG1 can be used.

[1188] To transiently transfect U937 cells with the GAS/SEAP/Neo construct produced in Example 31, a DEAE-Dextran method (Kharbanda et. al., 1994, Cell Growth & Differentiation, 5:259-265) is used. First, harvest 2×107 U937 cells and wash with PBS. The U937 cells are usually grown in RPMI 1640 medium containing 10% heat-inactivated fetal bovine serum (FBS) supplemented with 100 units/ml penicillin and 100 mg/ml streptomycin.

[1189] Next, suspend the cells in 1 ml of 20 mM Tris-HCl (pH 7.4) buffer containing 0.5 mg/ml DEAE-Dextran, 8 ug GAS-SEAP2 plasmid DNA, 140 mM NaCl, 5 mM KCl, 375 uM Na2HPO4.7H2O, 1 mM MgCl2, and 675 uM CaCl2. Incubate at 37 degrees C. for 45 min.

[1190] Wash the cells with RPMI 1640 medium containing 10% FBS and then resuspend in 10 ml complete medium and incubate at 37 degree C. for 36 hr.

[1191] The GAS-SEAP/U937 stable cells are obtained by growing the cells in 400 ug/ml G418. The G418-free medium is used for routine growth but every one to two months, the cells should be re-grown in 400 ug/ml G418 for couple of passages.

[1192] These cells are tested by harvesting 1×108 cells (this is enough for ten 96-well plates assay) and wash with PBS. Suspend the cells in 200 ml above described growth medium, with a final density of 5×105 cells/ml. Plate 200 ul cells per well in the 96-well plate (or 1×005 cells/well).

[1193] Add 50 ul of the supernatant prepared by the protocol described in Example 30. Incubate at 37 degee C for 48 to 72 hr. As a positive control, 100 Unit/ml interferon gamma can be used which is known to activate U937 cells. Over 30 fold induction is typically observed in the positive control wells. SEAP assay the supernatant according to the protocol described in Example 36.

Example 34

High-Throughput Screening Assay Identifying Neuronal Activity

[1194] When cells undergo differentiation and proliferation, a group of genes are activated through many different signal transduction pathways. One of these genes, EGR1 (early growth response gene 1), is induced in various tissues and cell types upon activation. The promoter of EGR1 is responsible for such induction. Using the EGR1 promoter linked to reporter molecules, activation of cells can be assessed by polypeptide of the present invention.

[1195] Particularly, the following protocol is used to assess neuronal activity in PC12 cell lines. PC12 cells (rat phenochromocytoma cells) are known to proliferate and/or differentiate by activation with a number of mitogens, such as TPA (tetradecanoyl phorbol acetate), NGF (nerve growth factor), and EGF (epidermal growth factor). The EGR1 gene expression is activated during this treatment. Thus, by stably transfecting PC12 cells with a construct containing an EGR promoter linked to SEAP reporter, activation of PC12 cells by polypeptide of the present invention can be assessed.

[1196] The EGR/SEAP reporter construct can be assembled by the following protocol. The EGR-1 promoter sequence (−633 to +1)(Sakamoto K et al., Oncogene 6:867-871 (1991)) can be PCR amplified from human genomic DNA using the following primers:

15(SEQ ID NO: 6)5′ GCGCTCGAGGGATGACAGCGATAGAACCCCGG-3′(SEQ ID NO: 7)5′ GCGAAGCTTCGCGACTCCCCGGATCCGCCTC-3′

[1197] Using the GAS:SEAP/Neo vector produced in Example 31, EGR1 amplified product can then be inserted into this vector. Linearize the GAS:SEAP/Neo vector using restriction enzymes XhoI/HindIII, removing the GAS/SV40 stuffer. Restrict the EGR1 amplified product with these same enzymes. Ligate the vector and the EGR1 promoter.

[1198] To prepare 96 well-plates for cell culture, two mls of a coating solution (1:30 dilution of collagen type I (Upstate Biotech Inc. Cat#08-115) in 30% ethanol (filter sterilized)) is added per one 10 cm plate or 50 ml per well of the 96-well plate, and allowed to air dry for 2 hr.

[1199] PC12 cells are routinely grown in RPMI-1640 medium (Bio Whittaker) containing 10% horse serum (JRH BIOSCIENCES, Cat. # 12449-78P), 5% heat-inactivated fetal bovine serum (FBS) supplemented with 100 units/ml penicillin and 100 ug/ml streptomycin on a precoated 10 cm tissue culture dish. One to four split is done every three to four days. Cells are removed from the plates by scraping and resuspended with pipetting up and down for more than 15 times.

[1200] Transfect the EGR/SEAP/Neo construct into PCl2 using the Lipofectamine protocol described in Example 30. EGR-SEAP/PC12 stable cells are obtained by growing the cells in 300 ug/ml G418. The G418-free medium is used for routine growth but every one to two months, the cells should be re-grown in 300 ug/ml G418 for couple of passages.

[1201] To assay for neuronal activity, a 10 cm plate with cells around 70 to 80% confluent is screened by removing the old medium. Wash the cells once with PBS (Phosphate buffered saline). Then starve the cells in low serum medium (RPMI-1640 containing 1% horse serum and 0.5% FBS with antibiotics) overnight.

[1202] The next morning, remove the medium and wash the cells with PBS. Scrape off the cells from the plate, suspend the cells well in 2 ml low serum medium. Count the cell number and add more low serum medium to reach final cell density as 5×105 cells/ml.

[1203] Add 200 ul of the cell suspension to each well of 96-well plate (equivalent to 1×105 cells/well). Add 50 ul supernatant produced by Example 30, 37 degree C. for 48 to 72 hr. As a positive control, a growth factor known to activate PC12 cells through EGR can be used, such as 50 ng/ul of Neuronal Growth Factor (NGF). Over fifty-fold induction of SEAP is typically seen in the positive control wells. SEAP assay the supernatant according to Example 36.

Example 35

High-Throughput Screening Assay for T-cell Activity

[1204] NF-KB (Nuclear Factor KB) is a transcription factor activated by a wide variety of agents including the inflammatory cytokines IL-1 and TNF, CD30 and CD40, lymphotoxin-alpha and lymphotoxin-beta, by exposure to LPS or thrombin, and by expression of certain viral gene products. As a transcription factor, NF-KB regulates the expression of genes involved in immune cell activation, control of apoptosis (NF-KB appears to shield cells from apoptosis), B and T-cell development, anti-viral and antimicrobial responses, and multiple stress responses.

[1205] In non-stimulated conditions, NF-KB is retained in the cytoplasm with I-KB (Inhibitor KB). However, upon stimulation, I-KB is phosphorylated and degraded, causing NF-KB to shuttle to the nucleus, thereby activating transcription of target genes. Target genes activated by NF-KB include IL-2, IL-6, GM-CSF, ICAM-1 and class I MHC.

[1206] Due to its central role and ability to respond to a range of stimuli, reporter constructs utilizing the NF-KB promoter element are used to screen the supernatants produced in Example 30. Activators or inhibitors of NF-KB would be useful in treating, preventing, and/or diagnosing diseases. For example, inhibitors of NF-KB could be used to treat those diseases related to the acute or chronic activation of NF-KB, such as rheumatoid arthritis.

[1207] To construct a vector containing the NF-KB promoter element, a PCR based strategy is employed. The upstream primer contains four tandem copies of the NF-KB binding site (GGGGACTTTCCC) (SEQ ID NO: 8), 18 bp of sequence complementary to the 5′ end of the SV40 early promoter sequence, and is flanked with an XhoI site:

16(SEQ ID NO: 9)5′:GCGGCCTCGAGGGGACTTTCCCGGGGACTTTCCGGGGACTTTCCGGGACTTTCCATCCTGCCATCTCAATTAG:3′

[1208] The downstream primer is complementary to the 3′ end of the SV40 promoter and is flanked with a Hind III site: 5′:GCGGCAAGCTTTTTGCAAAGCCTAGGC:3′(SEQ ID NO: 4)

[1209] PCR amplification is performed using the SV40 promoter template present in the pB-gal:promoter plasmid obtained from Clontech. The resulting PCR fragment is digested with XhoI and Hind III and subcloned into BLSK2-. (Stratagene) Sequencing with the T7 and T3 primers confirms the insert contains the following sequence:

17(SEQ ID NO: 10)5′:CTCGAGGGGACTTTCCCGGGGACTTTCCGGGGACTTTCCGGGACTTTCCATCTGCCATCTCAATTAGTCAGCAACCATAGTCCCGCCCCTAACTCCGCCCATCCCGCCCCTAACTCCGCCCAGTTCCGCCCATTCTCCGCCCCATGGCTGACTAATTTTTTTTATTTATGCAGAGGCCGAGGCCGCCTCGGCCTCTGAGCTATTCCAGAAGTAGTGAGGAGGCTTTTTTGGAGGCCTAGGCTTTTGCAAAAAGCTT:3′

[1210] Next, replace the SV40 minimal promoter element present in the pSEAP2-promoter plasmid (Clontech) with this NF-KB/SV40 fragment using XhoI and HindIII. However, this vector does not contain a neomycin resistance gene, and therefore, is not preferred for mammalian expression systems.

[1211] In order to generate stable mammalian cell lines, the NF-KB/SV40/SEAP cassette is removed from the above NF-KB/SEAP vector using restriction enzymes SalI and NotI, and inserted into a vector containing neomycin resistance. Particularly, the NF-KB/SV40/SEAP cassette was inserted into pGFP-1 (Clontech), replacing the GFP gene, after restricting pGFP-1 with SalI and NotI.

[1212] Once NF-KB/SV40/SEAP/Neo vector is created, stable Jurkat T-cells are created and maintained according to the protocol described in Example 32. Similarly, the method for assaying supernatants with these stable Jurkat T-cells is also described in Example 32. As a positive control, exogenous TNF alpha (0.1,1, 10 ng) is added to wells H9, H10, and H11, with a 5-10 fold activation typically observed.

Example 36

Assay for SEAP Activity

[1213] As a reporter molecule for the assays described in Examples 32-35, SEAP activity is assayed using the Tropix Phospho-light Kit (Cat. BP-400) according to the following general procedure. The Tropix Phospho-light Kit supplies the Dilution, Assay, and Reaction Buffers used below.

[1214] Prime a dispenser with the 2.5× Dilution Buffer and dispense 15 ul of 2.5× dilution buffer into Optiplates containing 35 ul of a supernatant. Seal the plates with a plastic sealer and incubate at 65 degree C. for 30 min. Separate the Optiplates to avoid uneven heating.

[1215] Cool the samples to room temperature for 15 minutes. Empty the dispenser and prime with the Assay Buffer. Add 50 ml Assay Buffer and incubate at room temperature 5 min. Empty the dispenser and prime with the Reaction Buffer (see the Table below). Add 50 ul Reaction Buffer and incubate at room temperature for 20 minutes. Since the intensity of the chemiluminescent signal is time dependent, and it takes about 10 minutes to read 5 plates on a luminometer, thus one should treat 5 plates at each time and start the second set 10 minutes later.

[1216] Read the relative light unit in the luminometer. Set H12 as blank, and print the results. An increase in chemiluminescence indicates reporter activity.

18Reaction Buffer Formulation:# of platesRxn buffer diluent (ml)CSPD (ml)1060311653.2512703.513753.751480415854.2516904.517954.75181005191055.25201105.5211155.75221206231256.25241306.5251356.75261407271457.25281507.5291557.75301608311658.25321708.5331758.75341809351859.25361909.5371959.7538200103920510.254021010.54121510.7542220114322511.254423011.54523511.7546240124724512.254825012.54925512.755026013

Example 37

High-Throughput Screening Assay Identifying Changes in Small Molecule Concentration and Membrane Permeability

[1217] Binding of a ligand to a receptor is known to alter intracellular levels of small molecules, such as calcium, potassium, sodium, and pH, as well as alter membrane potential. These alterations can be measured in an assay to identify supernatants which bind to receptors of a particular cell. Although the following protocol describes an assay for calcium, this protocol can easily be modified to detect changes in potassium, sodium, pH, membrane potential, or any other small molecule which is detectable by a fluorescent probe.

[1218] The following assay uses Fluorometric Imaging Plate Reader (“FLIPR”) to measure changes in fluorescent molecules (Molecular Probes) that bind small molecules. Clearly, any fluorescent molecule detecting a small molecule can be used instead of the calcium fluorescent molecule, fluo-4 (Molecular Probes, Inc.; catalog no. F-14202), used here. RPTKs for which the corresponding ligand is unknown. Ligands for RPTKs include mainly secreted small proteins, but also membrane-bound and extracellular matrix proteins.

[1219] Activation of RPTK by ligands involves ligand-mediated receptor dimerization, resulting in transphosphorylation of the receptor subunits and activation of the cytoplasmic tyrosine kinases. The cytoplasmic tyrosine kinases include receptor associated tyrosine kinases of the src-family (e.g., src, yes, lck, lyn, fyn) and non-receptor linked and cytosolic protein tyrosine kinases, such as the Jak family, members of which mediate signal transduction triggered by the cytokine superfamily of receptors (e.g., the Interleukins, Interferons, GM-CSF, and Leptin).

[1220] Because of the wide range of known factors capable of stimulating tyrosine kinase activity, identifying whether polypeptide of the present invention or a molecule induced by polypeptide of the present invention is capable of activating tyrosine kinase signal transduction pathways is of interest. Therefore, the following protocol is designed to identify such molecules capable of activating the tyrosine kinase signal transduction pathways.

[1221] Seed target cells (e.g., primary keratinocytes) at a density of approximately 25,000 cells per well in a 96 well Loprodyne Silent Screen Plates purchased from Nalge Nunc (Naperville, Ill.). The plates are sterilized with two 30 minute rinses with 100% ethanol, rinsed with water and dried overnight. Some plates are coated for 2 hr with 100 ml of cell culture grade type I collagen (50 mg/ml), gelatin (2%) or polylysine (50 mg/ml), all of which can be purchased from Sigma Chemicals (St. Louis, Mo.) or 10% Matri gel purchased from Becton Dickinson (Bedford,Mass.), or calf serum, rinsed with PBS and stored at 4 degree C. Cell growth on these plates is assayed by seeding 5,000 cells/well in growth medium and indirect quantitation of cell number through use of alamarBlue as described by the manufacturer Alamar Biosciences, Inc. (Sacramento, Calif.) after 48 hr. Falcon plate covers #3071 from Becton Dickinson (Bedford,Mass.) are used to cover the Loprodyne Silent Screen Plates. Falcon Microtest III cell culture plates can also be used in some proliferation experiments.

[1222] To prepare extracts, A431 cells are seeded onto the nylon membranes of Loprodyne plates (20,000/200 ml/well) and cultured overnight in complete medium. Cells are quiesced by incubation in serum-free basal medium for 24 hr. After 5-20 minutes treatment with EGF (60 ng/ml) or 50 ul of the supernatant produced in Example 30, the medium was removed and 100 ml of extraction buffer ((20 mM HEPES pH 7.5, 0.15 M NaCl, 1% Triton X-100, 0.1% SDS, 2 mM Na3VO4, 2 mM Na4P2O7 and a cocktail of protease inhibitors (# 1836170) obtained from Boeheringer Mannheim (Indianapolis, Ind.)) is added to each well and the plate is shaken on a rotating shaker for 5 minutes at 4° C. The plate is then placed in a vacuum transfer manifold and the extract filtered through the 0.45 mm membrane bottoms of each well using house vacuum. Extracts are collected in a 96-well catch/assay plate in the bottom of the vacuum manifold and immediately placed on ice. To obtain extracts clarified by centrifugation, the content of each well, after detergent solubilization for 5 minutes, is removed and centrifuged for 15 minutes at 4 degree C. at 16,000×g.

[1223] Test the filtered extracts for levels of tyrosine kinase activity. Although many methods of detecting tyrosine kinase activity are known, one method is described here.

[1224] Generally, the tyrosine kinase activity of a supernatant is evaluated by determining its ability to phosphorylate a tyrosine residue on a specific substrate (a biotinylated peptide). Biotinylated peptides that can be used for this purpose include PSK1 (corresponding to amino acids 6-20 of the cell division kinase cdc2-p34) and PSK2 (corresponding to amino acids 1-17 of gastrin). Both peptides are substrates for a range of tyrosine kinases and are available from Boehringer Mannheim.

[1225] The tyrosine kinase reaction is set up by adding the following components in order. First, add 10 ul of 5 uM Biotinylated Peptide, then 10 ul ATP/Mg2+(5 mM ATP/50 mM MgCl2), then 10 ul of 5× Assay Buffer (40 mM imidazole hydrochloride, pH 7.3, 40 mM beta-glycerophosphate, 1 mM EGTA, 100 mM MgCl2, 5 mM MnCl2, 0.5 mg/ml BSA), then 5 ul of Sodium Vanadate(1 mM), and then 5 ul of water. Mix the components gently and preincubate the reaction mix at 30 degree C. for 2 min. Initial the reaction by adding 10 ul of the control enzyme or the filtered supernatant.

[1226] The tyrosine kinase assay reaction is then terminated by adding 10 ul of 120 mm EDTA and place the reactions on ice.

[1227] Tyrosine kinase activity is determined by transferring 50 ul aliquot of reaction mixture to a microtiter plate (MTP) module and incubating at 37 degree C. for 20 min. This allows the streptavidin coated 96 well plate to associate with the biotinylated peptide. Wash the MTP module with 300 ul/well of PBS four times. Next add 75 ul of anti-phospolyrosine antibody conjugated to horse radish peroxidase(anti-P-Tyr-POD(0.5u/ml)) to each well and incubate at 37 degree C. for one hour. Wash the well as above.

[1228] Next add 100 ul of peroxidase substrate solution (Boehringer Mannheim) and incubate at room temperature for at least 5 mins (up to 30 min). Measure the absorbance of the sample at 405 nm by using ELISA reader. The level of bound peroxidase activity is quantitated using an ELISA reader and reflects the level of tyrosine kinase activity.

Example 39

High-Throughput Screening Assay Identifying Phosphorylation Activity

[1229] As a potential alternative and/or complement to the assay of protein tyrosine kinase activity described in Example 38, an assay which detects activation (phosphorylation) of major intracellular signal transduction intermediates can also be used. For example, as described below one particular assay can detect tyrosine phosphorylation of the Erk-1 and Erk-2 kinases. However, phosphorylation of other molecules, such as Raf, JNK, p38 MAP, Map kinase kinase (MEK), MEK kinase, Src, Muscle specific kinase (MuSK), IRAK, Tec, and Janus, as well as any other phosphoserine, phosphotyrosine, or phosphothreonine molecule, can be detected by substituting these molecules for Erk-1 or Erk-2 in the following assay.

[1230] Specifically, assay plates are made by coating the wells of a 96-well ELISA plate with 0.1 ml of protein G (lug/ml) for 2 hr at room temp, (RT). The plates are then rinsed with PBS and blocked with 3% BSA/PBS for 1 hr at RT. The protein G plates are then treated with 2 commercial monoclonal antibodies (100 ng/well) against Erk-1 and Erk-2 (1 hr at RT) (Santa Cruz Biotechnology). (To detect other molecules, this step can easily be modified by substituting a monoclonal antibody detecting any of the above described molecules.) After 3-5 rinses with PBS, the plates are stored at 4 degree C. until use.

[1231] A431 cells are seeded at 20,000/well in a 96-well Loprodyne filterplate and cultured overnight in growth medium. The cells are then starved for 48 hr in basal medium (DMEM) and then treated with EGF (6 ng/well) or 50 ul of the supernatants obtained in Example 30 for 5-20 minutes. The cells are then solubilized and extracts filtered directly into the assay plate.

[1232] After incubation with the extract for 1 hr at RT, the wells are again rinsed. As a positive control, a commercial preparation of MAP kinase (10 ng/well) is used in place of A431 extract. Plates are then treated with a commercial polyclonal (rabbit) antibody (1 ug/ml) which specifically recognizes the phosphorylated epitope of the Erk-1 and Erk-2 kinases (1 hr at RT). This antibody is biotinylated by standard procedures. The bound polyclonal antibody is then quantitated by successive incubations with Europium-streptavidin and Europium fluorescence enhancing reagent in the Wallac DELFIA instrument (time-resolved fluorescence). An increased fluorescent signal over background indicates a phosphorylation by polypeptide of the present invention or a molecule induced by polypeptide of the present invention.

Example 40

Assay for the Stimulation of Bone Marrow CD34+ Cell Proliferation

[1233] This assay is based on the ability of human CD34+ to proliferate in the presence of hematopoietic growth factors and evaluates the ability of isolated polypeptides expressed in mammalian cells to stimulate proliferation of CD34+ cells.

[1234] It has been previously shown that most mature precursors will respond to only a single signal. More immature precursors require at least two signals to respond. Therefore, to test the effect of polypeptides on hematopoietic activity of a wide range of progenitor cells, the assay contains a given polypeptide in the presence or absence of other hematopoietic growth factors. Isolated cells are cultured for 5 days in the presence of Stem Cell Factor (SCF) in combination with tested sample. SCF alone has a very limited effect on the proliferation of bone marrow (BM) cells, acting in such conditions only as a “survival” factor. However, combined with any factor exhibiting stimulatory effect on these cells (e.g., IL-3), SCF will cause a synergistic effect. Therefore, if the tested polypeptide has a stimulatory effect on hematopoietic progenitors, such activity can be easily detected. Since normal BM cells have a low level of cycling cells, it is likely that any inhibitory effect of a given polypeptide, or agonists or antagonists thereof, might not be detected. Accordingly, assays for an inhibitory effect on progenitors is preferably tested in cells that are first subjected to in vitro stimulation with SCF+IL+3, and then contacted with the compound that is being evaluated for inhibition of such induced proliferation.

[1235] Briefly, CD34+ cells are isolated using methods known in the art. The cells are thawed and resuspended in medium (QBSF 60 serum-free medium with 1% L-glutamine (SOOml) Quality Biological, Inc., Gaithersburg, Md. Cat# 160-204-101). After several gentle centrifugation steps at 200×g, cells are allowed to rest for one hour. The cell count is adjusted to 2.5×105 cells/ml. During this time, 100 μl of sterile water is added to the peripheral wells of a 96-well plate. The cytokines that can be tested with a given polypeptide in this assay is rhSCF (R&D Systems, Minneapolis, Minn., Cat# 255-SC) at 50 ng/ml alone and in combination with rhSCF and rhIL-3 (R&D Systems, Minneapolis, Minn., Cat# 203-ML) at 30 ng/ml. After one hour, 10 μl of prepared cytokines, 50 μl of the supernatants prepared in Example 30 (supernatants at 1:2 dilution=50 μl) and 20 μl of diluted cells are added to the media which is already present in the wells to allow for a final total volume of 100 μl. The plates are then placed in a 37° C./5% CO2 incubator for five days.

[1236] Eighteen hours before the assay is harvested, 0.5 μCi/well of [3H] Thymidine is added in a 10 μl volume to each well to determine the proliferation rate. The experiment is terminated by harvesting the cells from each 96-well plate to a filtermat using the Tomtec Harvester 96. After harvesting, the filtermats are dried, trimmed and placed into OmniFilter assemblies consisting of one OmniFilter plate and one OmniFilter Tray. 60 μl Microscint is added to each well and the plate sealed with TopSeal-A press-on sealing film A bar code 15 sticker is affixed to the first plate for counting. The sealed plates are then loaded and the level of radioactivity determined via the Packard Top Count and the printed data collected for analysis. The level of radioactivity reflects the amount of cell proliferation.

[1237] The studies described in this example test the activity of a given polypeptide to stimulate bone marrow CD34+ cell proliferation. One skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides (e.g., gene therapy), antibodies, agonists, and/or antagonists and fragments and variants thereof. As a nonlimiting example, potential antagonists tested in this assay would be expected to inhibit cell proliferation in the presence of cytokines and/or to increase the inhibition of cell proliferation in the presence of cytokines and a given polypeptide. In contrast, potential agonists tested in this assay would be expected to enhance cell proliferation and/or to decrease the inhibition of cell proliferation in the presence of cytokines and a given polypeptide.

[1238] The ability of a gene to stimulate the proliferation of bone marrow CD34+ cells indicates that polynucleotides and polypeptides corresponding to the gene are useful for the diagnosis and treatment of disorders affecting the immune system and hematopoiesis. Representative uses are described in the “Immune Activity” and “Infectious Disease” sections above, and elsewhere herein.

Example 41

Assay for Extracellular Matrix Enhanced Cell Response (EMECR)

[1239] The objective of the Extracellular Matrix Enhanced Cell Response (EMECR) assay is to identify gene products (e.g., isolated polypeptides) that act on the hematopoietic stem cells in the context of the extracellular matrix (ECM) induced signal.

[1240] Cells respond to the regulatory factors in the context of signal(s) received from the surrounding microenvironment. For example, fibroblasts, and endothelial and epithelial stem cells fail to replicate in the absence of signals from the ECM. Hematopoietic stem cells can undergo self-renewal in the bone marrow, but not in in vitro suspension culture. The ability of stem cells to undergo self-renewal in vitro is dependent upon their interaction with the stromal cells and the ECM protein fibronectin (fn). Adhesion of cells to fn is mediated by the α5.β1 and α4.β1 integrin receptors, which are expressed by human and mouse hematopoietic stem cells. The factor(s) which integrate with the ECM environment and are responsible for stimulating stem cell self-renewal havea not yet been identified. Discovery of such factors should be of great interest in gene therapy and bone marrow transplant applications

[1241] Briefly, polystyrene, non tissue culture treated, 96-well plates are coated with fn fragment at a coating concentration of 0.2 μg/cm2. Mouse bone marrow cells are plated (1,000 cells/well) in 0.2 ml of serum-free medium. Cells cultured in the presence of IL-3 (5 ng/ml)+SCF (50 ng/ml) would serve as the positive control, conditions under which little self-renewal but pronounced differentiation of the stem cells is to be expected. Gene products of the invention (e.g., including, but not limited to, polynucleotides and polypeptides of the present invention, and supernatants produced in Example 30), are tested with appropriate negative controls in the presence and absence of SCF(5.0 ng/ml), where test factor supernatants represent 10% of the total assay volume. The plated cells are then allowed to grow by incubating in a low oxygen environment (5% CO2, 7% O2, and 88% N2) tissue culture incubator for 7 days. The number of proliferating cells within the wells is then quantitated by measuring thymidine incorporation into cellular DNA. Verification of the positive hits in the assay will require phenotypic characterization of the cells, which can be accomplished by scaling up of the culture system and using appropriate antibody reagents against cell surface antigens and FACScan.

[1242] One skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides (e.g., gene therapy), antibodies, agonists, and/or antagonists and fragments and variants thereof.

[1243] If a particular polypeptide of the present invention is found to be a stimulator of hematopoietic progenitors, polynucleotides and polypeptides corresponding to the gene encoding said polypeptide may be useful for the diagnosis and treatment of disorders affecting the immune system and hematopoiesis. Representative uses are described in the “Immune Activity” and “Infectious Disease” sections above, and elsewhere herein. The gene product may also be useful in the expansion of stem cells and committed progenitors of various blood lineages, and in the differentiation and/or proliferation of various cell types.

[1244] Additionally, the polynucleotides and/or polypeptides of the gene of interest and/or agonists and/or antagonists thereof, may also be employed to inhibit the proliferation and differentiation of hematopoietic cells and therefore may be employed to protect bone marrow stem cells from chemotherapeutic agents during chemotherapy. This antiproliferative effect may allow administration of higher doses of chemotherapeutic agents and, therefore, more effective chemotherapeutic treatment.

[1245] Moreover, polynucleotides and polypeptides corresponding to the gene of interest may also be useful for the treatment and diagnosis of hematopoietic related disorders such as, for example, anemia, pancytopenia, leukopenia, thrombocytopenia or leukemia since stromal cells are important in the production of cells of hematopoietic lineages. The uses include bone marrow cell ex-vivo culture, bone marrow transplantation, bone marrow reconstitution, radiotherapy or chemotherapy of neoplasia.

Example 42

Human Dermal Fibroblast and Aortic Smooth Muscle Cell Proliferation

[1246] The polypeptide of interest is added to cultures of normal human dermal fibroblasts (NHDF) and human aortic smooth muscle cells (AoSMC) and two co-assays are performed with each sample. The first assay examines the effect of the polypeptide of interest on the proliferation of normal human dermal fibroblasts (NHDF) or aortic smooth muscle cells (AoSMC). Aberrant growth of fibroblasts or smooth muscle cells is a part of several pathological processes, including fibrosis, and restenosis. The second assay examines IL6 production by both NHDF and SMC. IL6 production is an indication of functional activation. Activated cells will have increased production of a number of cytokines and other factors, which can result in a proinflammatory or immunomodulatory outcome. Assays are run with and without co-TNFa stimulation, in order to check for costimulatory or inhibitory activity.

[1247] Briefly, on day 1, 96-well black plates are set up with 1000 cells/well (NHDF) or 2000 cells/well (AOSMC) in 100 μl culture media. NHDF culture media contains: Clonetics FB basal media, 1 mg/ml hFGF, 5 mg/ml insulin, 50 mg/ml gentamycin, 2%FBS, while AoSMC culture media contains Clonetics SM basal media, 0.5 μg/ml HEGF, 5 mg/ml insulin, 1 μg/ml hFGF, 50 mg/ml gentamycin, 50 μg/ml Amphotericin B, 5%FBS. After incubation at 37° C. for at least 4-5 hours culture media is aspirated and replaced with growth arrest media. Growth arrest media for NHDF contains fibroblast basal media, 50 mg/ml gentamycin, 2% FBS, while growth arrest media for AoSMC contains SM basal media, 50 mg/ml gentamycin, 50 μg/ml Amphotericin B, 0.4% FBS. Incubate at 37° C. until day 2.

[1248] On day 2, serial dilutions and templates of the polypeptide of interest are designed such that they always include media controls and known-protein controls. For both stimulation and inhibition experiments, proteins are diluted in growth arrest media. For inhibition experiments, TNFa is added to a final concentration of 2 ng/ml (NHDF) or 5 ng/ml (AoSMC). Add ⅓ vol media containing controls or polypeptides of the present invention and incubate at 37 degrees C./5% CO2 until day 5.

[1249] Transfer 60 μl from each well to another labeled 96-well plate, cover with a plate-sealer, and store at 4 degrees C. until Day 6 (for IL6 ELISA). To the remaining 100 μl in the cell culture plate, aseptically add Alamar Blue in an amount equal to 10% of the culture volume (10 μl). Return plates to incubator for 3 to 4 hours. Then measure fluorescence with excitation at 530 nm and emission at 590 nm using the CytoFluor. This yields the growth stimulation/inhibition data.

[1250] On day 5, the IL6 ELISA is performed by coating a 96 well plate with 50-100 ul/well of Anti-Human IL6 Monoclonal antibody diluted in PBS, pH 7.4, incubate ON at room temperature.

[1251] On day 6, empty the plates into the sink and blot on paper towels. Prepare Assay Buffer containing PBS with 4% BSA. Block the plates with 200 μl/well of Pierce Super Block blocking buffer in PBS for 1-2 hr and then wash plates with wash buffer (PBS, 0.05% Tween-20). Blot plates on paper towels. Then add 50 μl/well of diluted Anti-Human IL-6 Monoclonal, Biotin-labeled antibody at 0.50 mg/ml. Make dilutions of IL-6 stock in media (30, 10, 3, 1, 0.3, 0 ng/ml). Add duplicate samples to top row of plate. Cover the plates and incubate for 2 hours at RT on shaker.

[1252] Plates are washed with wash buffer and blotted on paper towels. Dilute EU-labeled Streptavidin 1:1000 in Assay buffer, and add 100 μl/well. Cover the plate and incubate 1 h at RT. Plates are again washed with wash buffer and blotted on paper towels.

[1253] Add 100 μl/well of Enhancement Solution. Shake for 5 minutes. Read the plate on the Wallac DELFIA Fluorometer. Readings from triplicate samples in each assay were tabulated and averaged.

[1254] A positive result in this assay suggests AoSMC cell proliferation and that the polypeptide of the present invention may be involved in dermal fibroblast proliferation and/or smooth muscle cell proliferation. A positive result also suggests many potential uses of polypeptides, polynucleotides, agonists and/or antagonists of the polynucleotide/polypeptide of the present invention which gives a positive result. For example, inflammation and immune responses, wound healing, and angiogenesis, as detailed throughout this specification. Particularly, polypeptides of the present invention and polynucleotides of the present invention may be used in wound healing and dermal regeneration, as well as the promotion of vasculogenesis, both of the blood vessels and lymphatics. The growth of vessels can be used in the treatment of, for example, cardiovascular diseases. Additionally, antagonists of polypeptides and polynucleotides of the invention may be useful in treating diseases, disorders, and/or conditions which involve angiogenesis by acting as an anti-vascular agent (e.g., anti-angiogenesis). These diseases, disorders, and/or conditions are known in the art and/or are described herein, such as, for example, malignancies, solid tumors, benign tumors, for example hemangiomas, acoustic neuromas, neurofibromas, trachomas, and pyogenic granulomas; artheroscleric plaques; ocular angiogenic diseases, for example, diabetic retinopathy, retinopathy of prematurity, macular degeneration, corneal graft rejection, neovascular glaucoma, retrolental fibroplasia, rubeosis, retinoblastoma, uvietis and Pterygia (abnormal blood vessel growth) of the eye; rheumatoid arthritis; psoriasis; delayed wound healing; endometriosis; vasculogenesis; granulations; hypertrophic scars (keloids); nonunion fractures; scleroderma; trachoma; vascular adhesions; myocardial angiogenesis; coronary collaterals; cerebral collaterals; arterioyenous malformations; ischemic limb angiogenesis; Osler-Webber Syndrome; plaque neovascularization; telangiectasia; hemophiliac joints; angiofibroma; fibromuscular dysplasia; wound granulation; Crohn's disease; and atherosclerosis. Moreover, antagonists of polypeptides and polynucleotides of the invention may be useful in treating anti-hyperproliferative diseases and/or anti-inflammatory known in the art and/or described herein.

[1255] One skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides (e.g., gene therapy), antibodies, agonists, and/or antagonists and fragments and variants thereof.

Example 43

Cellular Adhesion Molecule (CAM) Expression on Endothelial Cells

[1256] The recruitment of lymphocytes to areas of inflammation and angiogenesis involves specific receptor-ligand interactions between cell surface adhesion molecules (CAMs) on lymphocytes and the vascular endothelium. The adhesion process, in both normal and pathological settings, follows a multi-step cascade that involves intercellular adhesion molecule-1 (ICAM-1), vascular cell adhesion molecule-1 (VCAM-1), and endothelial leukocyte adhesion molecule-1 (E-selectin) expression on endothelial cells (EC). The expression of these molecules and others on the vascular endothelium determines the efficiency with which leukocytes may adhere to the local vasculature and extravasate into the local tissue during the development of an inflammatory response. The local concentration of cytokines and growth factor participate in the modulation of the expression of these CAMs.

[1257] Briefly, endothelial cells (e.g., Human Umbilical Vein Endothelial cells (HUVECs)) are grown in a standard 96 well plate to confluence, growth medium is removed from the cells and replaced with 100 μl of 199 Medium (10% fetal bovine serum (FBS)). Samples for testing and positive or negative controls are added to the plate in triplicate (in 10 μl volumes). Plates are then incubated at 37° C. for either 5 h (selectin and integrin expression) or 24 h (integrin expression only). Plates are aspirated to remove medium and 100 μl of 0.1% paraformaldehyde-PBS(with Ca++ and Mg++) is added to each well. Plates are held at 4° C. for 30 min. Fixative is removed from the wells and wells are washed 1× with PBS(+Ca,Mg)+0.5% BSA and drained. 10 μl of diluted primary antibody is added to the test and control wells. Anti-ICAM-1-Biotin, Anti-VCAM-1-Biotin and Anti-E-selectin-Biotin are used at a concentration of 10 μg/ml (1:10 dilution of 0.1 mg/ml stock antibody). Cells are incubated at 37° C. for 30 min. in a humidified environment. Wells are washed three times with PBS(+Ca,Mg)+0.5% BSA. 20 μl of diluted ExtrAvidin-Alkaline Phosphatase (1:5,000 dilution, referred to herein as the working dilution) are added to each well and incubated at 37° C. for 30 min. Wells are washed three times with PBS(+Ca,Mg)+0.5% BSA. Dissolve 1 tablet of p-Nitrophenol Phosphate pNPP per 5 ml of glycine buffer (pH 10.4). 100 Ill of pNPP substrate in glycine buffer is added to each test well. Standard wells in triplicate are prepared from the working dilution of the ExtrAvidin-Alkaline Phosphotase in glycine buffer: 1:5,000 (100)>10−0.5>10−1>10−1.5. 5 μl of each dilution is added to triplicate wells and the resulting AP content in each well is 5.50 ng, 1.74 ng, 0.55 ng, 0.18 ng. 100 μl of pNNP reagent is then added to each of the standard wells. The plate is incubated at 37° C. for 4 h. A volume of 50 ill of 3M NaOH is added to all wells. The plate is read on a plate reader at 405 nm using the background subtraction option on blank wells filled with glycine buffer only. Additionally, the template is set up to indicate the concentration of AP-conjugate in each standard well [5.50 ng; 1.74 ng; 0.55 ng; 0.18 ng]. Results are indicated as amount of bound AP-conjugate in each sample.

Example 44

Alamar Blue Endothelial Cells Proliferation Assay

[1258] This assay may be used to quantitatively determine protein mediated inhibition of bFGF-induced proliferation of Bovine Lymphatic Endothelial Cells (LECs), Bovine Aortic Endothelial Cells (BAECs) or Human Microvascular Uterine Myometrial Cells (UTMECs). This assay incorporates a fluorometric growth indicator based on detection of metabolic activity. A standard Alamar Blue Proliferation Assay is prepared in EGM-2MV with 10 ng/ml of bFGF added as a source of endothelial cell stimulation. This assay may be used with a variety of endothelial cells with slight changes in growth medium and cell concentration. Dilutions of the protein batches to be tested are diluted as appropriate. Serum-free medium (GIBCO SFM) without bFGF is used as a non-stimulated control and Angiostatin or TSP-1 are included as a known inhibitory controls.

[1259] Briefly, LEC, BAECs or UTMECs are seeded in growth media at a density of 5000 to 2000 cells/well in a 96 well plate and placed at 37 degreesC overnight. After the overnight incubation of the cells, the growth media is removed and replaced with GIBCO EC-SFM. The cells are treated with the appropriate dilutions of the protein of interest or control protein sample(s) (prepared in SFM) in triplicate wells with additional bFGF to a concentration of 10 ng/ml. Once the cells have been treated with the samples, the plate(s) is/are placed back in the 37° C. incubator for three days. After three days 10 ml of stock alamar blue (Biosource Cat# DALI 100) is added to each well and the plate(s) is/are placed back in the 37° C. incubator for four hours. The plate(s) are then read at 530 nm excitation and 590 nm emission using the CytoFluor fluorescence reader. Direct output is recorded in relative fluorescence units.

[1260] Alamar blue is an oxidation-reduction indicator that both fluoresces and changes color in response to chemical reduction of growth medium resulting from cell growth. As cells grow in culture, innate metabolic activity results in a chemical reduction of the immediate surrounding environment. Reduction related to growth causes the indicator to change from oxidized (non-fluorescent blue) form to reduced (fluorescent red) form (i.e., stimulated proliferation will produce a stronger signal and inhibited proliferation will produce a weaker signal and the total signal is proportional to the total number of cells as well as their metabolic activity). The background level of activity is observed with the starvation medium alone. This is compared to the output observed from the positive control samples (bFGF in growth medium) and protein dilutions.

Example 45

Detection of Inhibition of a Mixed Lymphocyte Reaction

[1261] This assay can be used to detect and evaluate inhibition of a Mixed Lymphocyte Reaction (MLR) by gene products (e.g., isolated polypeptides). Inhibition of a MLR may be due to a direct effect on cell proliferation and viability, modulation of costimulatory molecules on interacting cells, modulation of adhesiveness between lymphocytes and accessory cells, or modulation of cytokine production by accessory cells. Multiple cells may be targeted by these polypeptides since the peripheral blood mononuclear fraction used in this assay includes T, B and natural killer lymphocytes, as well as monocytes and dendritic cells.

[1262] Polypeptides of interest found to inhibit the MLR may find application in diseases associated with lymphocyte and monocyte activation or proliferation. These include, but are not limited to, diseases such as asthma, arthritis, diabetes, inflammatory skin conditions, psoriasis, eczema, systemic lupus erythematosus, multiple sclerosis, glomerulonephritis, inflammatory bowel disease, crohn's disease, ulcerative colitis, arteriosclerosis, cirrhosis, graft vs. host disease, host vs. graft disease, hepatitis, leukemia and lymphoma.

[1263] Briefly, PBMCs from human donors are purified by density gradient centrifugation using Lymphocyte Separation Medium (LSM®, density 1.0770 g/ml, Organon Teknika Corporation, West Chester, Pa.). PBMCs from two donors are adjusted to 2×106 cells/ml in RPMI-1640 (Life Technologies, Grand Island, N.Y.) supplemented with 10% FCS and 2 mM glutamine. PBMCs from a third donor is adjusted to 2×105 cells/ml. Fifty microliters of PBMCs from each donor is added to wells of a 96-well round bottom microtiter plate. Dilutions of test materials (50 μl) is added in triplicate to microtiter wells. Test samples (of the protein of interest) are added for final dilution of 1:4; rhuIL-2 (R&D Systems, Minneapolis, Minn., catalog number 202-IL) is added to a final concentration of 1 μg/ml; anti-CD4 mAb (R&D Systems, clone 34930.11, catalog number MAB379) is added to a final concentration of 10 μg/ml. Cells are cultured for 7-8 days at 37° C. in 5% CO2, and 1 μC of [3H] thymidine is added to wells for the last 16 hrs of culture. Cells are harvested and thymidine incorporation determined using a Packard TopCount. Data is expressed as the mean and standard deviation of triplicate determinations.

[1264] Samples of the protein of interest are screened in separate experiments and compared to the negative control treatment, anti-CD4 mAb, which inhibits proliferation of lymphocytes and the positive control treatment, IL-2 (either as recombinant material or supernatant), which enhances proliferation of lymphocytes.

[1265] One skilled in the art could easily modify the exemplified studies to test the activity of polynucleotides (e.g., gene therapy), antibodies, agonists, and/or antagonists and fragments and variants thereof.

Example 46

Assays for Protease Activity

[1266] The following assay may be used to assess protease activity of the polypeptides of the invention.

[1267] Gelatin and casein zymography are performed essentially as described (Heusen et al., Anal. Biochem., 102:196-202 (1980); Wilson et al., Journal of Urology, 149:653-658 (1993)). Samples are run on 10% polyacryamide/0.1% SDS gels containing 1% gelain orcasein, soaked in 2.5% triton at room temperature for 1 hour, and in 0.1M glycine, pH 8.3 at 37° C. 5 to 16 hours. After staining in amido black areas of proteolysis apear as clear areas agains the blue-black background. Trypsin (Sigma T8642) is used as a positive control.

[1268] Protease activity is also determined by monitoring the cleavage of n-a-benzoyl-L-arginine ethyl ester (BAEE) (Sigma B-4500. Reactions are set up in (25 mMNaPO4, 1 mM EDTA, and 1 mM BAEE), pH 7.5. Samples are added and the change in adsorbance at 260 nm is monitored on the Beckman DU-6 spectrophotometer in the time-drive mode. Trypsin is used as a positive control.

[1269] Additional assays based upon the release of acid-soluble peptides from casein or hemoglobin measured as adsorbance at 280 nm or calorimetrically using the Folin method are performed as described in Bergmeyer, et al., Methods of Enzymatic Analysis, 5 (1984). Other assays involve the solubilization of chromogenic substrates (Ward, Applied Science, 251-317 (1983)).

Example 47

Identifying Serine Protease Substrate Specificity

[1270] Methods known in the art or described herein may be used to determine the substrate specificity of the polypeptides of the present invention having serine protease activity. A preferred method of determining substrate specificity is by the use of positional scanning synthetic combinatorial libraries as described in GB 2 324 529 (incorporated herein in its entirety).

Example 48

Ligand Binding Assays

[1271] The following assay may be used to assess ligand binding activity of the polypeptides of the invention.

[1272] Ligand binding assays provide a direct method for ascertaining receptor pharmacology and are adaptable to a high throughput format. The purified ligand for a polypeptide is radiolabeled to high specific activity (50-2000 Ci/mmol) for binding studies. A determination is then made that the process of radiolabeling does not diminish the activity of the ligand towards its polypeptide. Assay conditions for buffers, ions, pH and other modulators such as nucleotides are optimized to establish a workable signal to noise ratio for both membrane and whole cell polypeptide sources. For these assays, specific polypeptide binding is defined as total associated radioactivity minus the radioactivity measured in the presence of an excess of unlabeled competing ligand. Where possible, more than one competing ligand is used to define residual nonspecific binding.

Example 49

Functional Assay in Xenopus Oocytes

[1273] Capped RNA transcripts from linearized plasmid templates encoding the polypeptides of the invention are synthesized in vitro with RNA polymerases in accordance with standard procedures. In vitro transcripts are suspended in water at a final concentration of 0.2 mg/ml. Ovarian lobes are removed from adult female toads, Stage V defolliculated oocytes are obtained, and RNA transcripts (10 ng/oocytc) are injected in a 50 nl bolus using a microinjection apparatus. Two electrode voltage clamps are used to measure the currents from individual Xenopus oocytes in response polypeptides and polypeptide agonist exposure. Recordings are made in Ca2+ free Barth's medium at room temperature. The Xenopus system can be used to screen known ligands and tissue/cell extracts for activating ligands.

Example 50

Microphysiometric Assays

[1274] Activation of a wide variety of secondary messenger systems results in extrusion of small amounts of acid from a cell. The acid formed is largely as a result of the increased metabolic activity required to fuel the intracellular signaling process. The pH changes in the media surrounding the cell are very small but are detectable by the CYTOSENSOR microphysiometer (Molecular Devices Ltd., Menlo Park, Calif). The CYTOSENSOR is thus capable of detecting the activation of polypeptide which is coupled to an energy utilizing intracellular signaling pathway.

Example 51

Extract/Cell Supernatant Screening

[1275] A large number of mammalian receptors exist for which there remains, as yet, no cognate activating ligand (agonist). Thus, active ligands for these receptors may not be included within the ligands banks as identified to date. Accordingly, the polypeptides of the invention can also be functionally screened (using calcium, cAMP, microphysiometer, oocyte electrophysiology, etc., functional screens) against tissue extracts to identify its natural ligands. Extracts that produce positive functional responses can be sequentially subfractionated until an activating ligand is isolated and identified.

Example 52

Calcium and cAMP Functional Assays

[1276] Seven transmembrane receptors which are expressed in HEK 293 cells have been shown to be coupled functionally to activation of PLC and calcium mobilization and/or cAMP stimulation or inhibition. Basal calcium levels in the HEK 293 cells in receptor-transfected or vector control cells were observed to be in the normal, 100 nM to 200 nM, range. HEK 293 cells expressing recombinant receptors are loaded with fura 2 and in a single day >150 selected ligands or tissue/cell extracts are evaluated for agonist induced calcium mobilization. Similarly, HEK 293 cells expressing recombinant receptors are evaluated for the stimulation or inhibition of cAMP production using standard cAMP quantitation assays. Agonists presenting a calcium transient or cAMP fluctuation are tested in vector control cells to determine if the response is unique to the transfected cells expressing receptor.

Example 53

ATP-Binding Assay

[1277] The following assay may be used to assess ATP-binding activity of polypeptides of the invention.

[1278] ATP-binding activity of the polypeptides of the invention may be detected using the ATP-binding assay described in U.S. Pat. No. 5,858,719, which is herein incorporated by reference in its entirety. Briefly, ATP-binding to polypeptides of the invention is measured via photoaffinity labeling with 8-azido-ATP in a competition assay. Reaction mixtures containing 1 mg/ml of the ABC transport protein of the present invention are incubated with varying concentrations of ATP, or the non-hydrolyzable ATP analog adenyl-5′-imidodiphosphate for 10 minutes at 4° C. A mixture of 8-azido-ATP (Sigma Chem. Corp., St. Louis, Mo.) plus 8-azido-ATP (32P-ATP) (5 mCi/μmol, ICN, Irvine Calif.) is added to a final concentration of 100 μM and 0.5 ml aliquots are placed in the wells of a porcelain spot plate on ice. The plate is irradiated using a short wave 254 nm UV lamp at a distance of 2.5 cm from the plate for two one-minute intervals with a one-minute cooling interval in between. The reaction is stopped by addition of dithiothreitol to a final concentration of 2 mM. The incubations are subjected to SDS-PAGE electrophoresis, dried, and autoradiographed. Protein bands corresponding to the particular polypeptides of the invention are excised, and the radioactivity quantified. A decrease in radioactivity with increasing ATP or adenly-5′-imidodiphosphate provides a measure of ATP affinity to the polypeptides.

Example 54

Small Molecule Screening

[1279] This invention is particularly useful for screening therapeutic compounds by using the polypeptides of the invention, or binding fragments thereof, in any of a variety of drug screening techniques. The polypeptide or fragment employed in such a test may be affixed to a solid support, expressed on a cell surface, free in solution, or located intracellularly. One method of drug screening utilizes eukaryotic or prokaryotic host cells which are stably transformed with recombinant nucleic acids expressing the polypeptide or fragment. Drugs are screened against such transformed cells in competitive binding assays. One may measure, for example, the formulation of complexes between the agent being tested and polypeptide of the invention.

[1280] Thus, the present invention provides methods of screening for drugs or any other agents which affect activities mediated by the polypeptides of the invention. These methods comprise contacting such an agent with a polypeptide of the invention or fragment thereof and assaying for the presence of a complex between the agent and the polypeptide or fragment thereof, by methods well known in the art. In such a competitive binding assay, the agents to screen are typically labeled. Following incubation, free agent is separated from that present in bound form, and the amount of free or uncomplexed label is a measure of the ability of a particular agent to bind to the polypeptides of the invention.

[1281] Another technique for drug screening provides high throughput screening for compounds having suitable binding affinity to the polypeptides of the invention, and is described in great detail in European Patent Application 84/03564, published on Sep. 13, 1984, which is herein incorporated by reference in its entirety. Briefly stated, large numbers of different small molecule test compounds are synthesized on a solid substrate, such as plastic pins or some other surface. The test compounds are reacted with polypeptides of the invention and washed. Bound polypeptides are then detected by methods well known in the art. Purified polypeptides are coated directly onto plates for use in the aforementioned drug screening techniques. In addition, non-neutralizing antibodies may be used to capture the peptide and immobilize it on the solid support.

[1282] This invention also contemplates the use of competitive drug screening assays in which neutralizing antibodies capable of binding polypeptides of the invention specifically compete with a test compound for binding to the polypeptides or fragments thereof. In this manner, the antibodies are used to detect the presence of any peptide which shares one or more antigenic epitopes with a polypeptide of the invention.

Example 55

Phosphorylation Assay

[1283] In order to assay for phosphorylation activity of the polypeptides of the invention, a phosphorylation assay as described in U.S. Pat. No. 5,958,405 (which is herein incorporated by reference) is utilized. Briefly, phosphorylation activity may be measured by phosphorylation of a protein substrate using gamma-labeled 32P-ATP and quantitation of the incorporated radioactivity using a gamma radioisotope counter. The polypeptides of the invention are incubated with the protein substrate, 32P-ATP, and a kinase buffer. The 32P incorporated into the substrate is then separated from free 32P-ATP by electrophoresis, and the incorporated 32P is counted and compared to a negative control. Radioactivity counts above the negative control are indicative of phosphorylation activity of the polypeptides of the invention.

Example 56

Detection of Phosphorylation Activity (Activation) of the Polypeptides of the Invention in the Presence of Polypeptide Ligands

[1284] Methods known in the art or described herein may be used to determine the phosphorylation activity of the polypeptides of the invention. A preferred method of determining phosphorylation activity is by the use of the tyrosine phosphorylation assay as described in U.S. Pat. No. 5,817,471 (incorporated herein by reference).

Example 57

Identification of Signal Transduction Proteins that Interact with Polypeptides of the Present Invention

[1285] The purified polypeptides of the invention are research tools for the identification, characterization and purification of additional signal transduction pathway proteins or receptor proteins. Briefly, labeled polypeptides of the invention are useful as reagents for the purification of molecules with which it interacts. In one embodiment of affinity purification, polypeptides of the invention are covalently coupled to a chromatography column. Cell-free extract derived from putative target cells, such as carcinoma tissues, is passed over the column, and molecules with appropriate affinity bind to the polypeptides of the invention. The protein complex is recovered from the column, dissociated, and the recovered molecule subjected to N-terminal protein sequencing. This amino acid sequence is then used to identify the captured molecule or to design degenerate oligonucleotide probes for cloning the relevant gene from an appropriate cDNA library.

Example 58

IL-6 Bioassay

[1286] To test the proliferative effects of the polypeptides of the invention, the IL-6 Bioassay as described by Marz et al. is utilized (Proc. Natl. Acad. Sci., U.S.A., 95:3251-56 (1998), which is herein incorporated by reference). Briefly, IL-6 dependent B9 murine cells are washed three times in IL-6 free medium and plated at a concentration of 5,000 cells per well in 50 μl, and 50 μl of the IL-6-like polypeptide is added. After 68 hrs. at 37° C., the number of viable cells is measured by adding the tetrazolium salt thiazolyl blue (MTT) and incubating for a further 4 hrs. at 37° C. B9 cells are lysed by SDS and optical density is measured at 570 nm. Controls containing IL-6 positive) and no cytokine (negative) are utilized. Enhanced proliferation in the test sample(s) relative to the negative control is indicative of proliferative effects mediated by polypeptides of the invention.

Example 59

Support of Chicken Embryo Neuron Survival

[1287] To test whether sympathetic neuronal cell viability is supported by polypeptides of the invention, the chicken embryo neuronal survival assay of Senaldi et al is utilized (Proc. Natl. Acad. Sci., U.S.A., 96:11458-63 (1998), which is herein incorporated by reference). Briefly, motor and sympathetic neurons are isolated from chicken embryos, resuspended in L15 medium (with 10% FCS, glucose, sodium selenite, progesterone, conalbumin, putrescine, and insulin; Life Technologies, Rockville, Md.) and Dulbecco's modified Eagles medium [with 10% FCS, glutamine, penicillin, and 25 mM Hepes buffer (pH 7.2); Life Technologies, Rockville, Md.], respectively, and incubated at 37° C. in 5% CO2 in the presence of different concentrations of the purified IL-6-like polypeptide, as well as a negative control lacking any cytokine. After 3 days, neuron survival is determined by evaluation of cellular morphology, and through the use of the colorimetric assay of Mosmann (Mosmann, T., J. Immunol. Methods, 65:55-63 (1983)). Enhanced neuronal cell viability as compared to the controls lacking cytokine is indicative of the ability of the inventive purified IL-6-like polypeptide(s) to enhance the survival of neuronal cells.

Example 60

Assay for Phosphatase Activity

[1288] The following assay may be used to assess serine/threonine phosphatase (PTPase) activity of the polypeptides of the invention.

[1289] In order to assay for serine/threonine phosphatase (PTPase) activity, assays can be utilized which are widely known to those skilled in the art. For example, the serine/threonine phosphatase (PSPase) activity is measured using a PSPase assay kit from New England Biolabs, Inc. Myelin basic protein (MyBP), a substrate for PSPase, is phosphorylated on serine and threonine residues with cAMP-dependent Protein Kinase in the presence of [32P]ATP. Protein serine/threonine phosphatase activity is then determined by measuring the release of inorganic phosphate from 32P-labeled MyBP.

Example 61

Interaction of Serine/Threonine Phosphatases with other Proteins

[1290] The polypeptides of the invention with serine/threonine phosphatase activity as determined in Example 60 are research tools for the identification, characterization and purification of additional interacting proteins or receptor proteins, or other signal transduction pathway proteins. Briefly, labeled polypeptide(s) of the invention is useful as a reagent for the purification of molecules with which it interacts. In one embodiment of affinity purification, polypeptide of the invention is covalently coupled to a chromatography column. Cell-free extract derived from putative target cells, such as neural or liver cells, is passed over the column, and molecules with appropriate affinity bind to the polypeptides of the invention. The polypeptides of the invention-complex is recovered from the column, dissociated, and the recovered molecule subjected to N-terminal protein sequencing. This amino acid sequence is then used to identify the captured molecule or to design degenerate oligonucleotide probes for cloning the relevant gene from an appropriate cDNA library.

Example 62

Assaying for Heparanase Activity

[1291] In order to assay for heparanase activity of the polypeptides of the invention, the heparanase assay described by Vlodavsky et al is utilized (Vlodavsky, I., et al., Nat. Med., 5:793-802 (1999)). Briefly, cell lysates, conditioned media or intact cells (1×106 cells per 35-mm dish) are incubated for 18 hrs at 37° C., pH 6.2-6.6, with 35S-labeled ECM or soluble ECM derived peak I proteoglycans. The incubation medium is centrifuged and the supernatant is analyzed by gel filtration on a Sepharose CL-6B column (0.9×30 cm). Fractions are eluted with PBS and their radioactivity is measured. Degradation fragments of heparan sulfate side chains are eluted from Sepharose 6B at 0.5<Kav <0.8 (peak II). Each experiment is done at least three times. Degradation fragments corresponding to “peak II,” as described by Vlodavsky et al., is indicative of the activity of the polypeptides of the invention in cleaving heparan sulfate.

Example 63

Immobilization of Biomolecules

[1292] This example provides a method for the stabilization of polypeptides of the invention in non-host cell lipid bilayer constucts (see, e.g., Bieri et al., Nature Biotech 17:1105-1108 (1999), hereby incorporated by reference in its entirety herein) which can be adapted for the study of polypeptides of the invention in the various functional assays described above. Briefly, carbohydrate-specific chemistry for biotinylation is used to confine a biotin tag to the extracellular domain of the polypeptides of the invention, thus allowing uniform orientation upon immobilization. A 50 uM solution of polypeptides of the invention in washed membranes is incubated with 20 mM NaIO4 and 1.5 mg/ml (4 mM) BACH or 2 mg/ml (7.5 mM) biotin-hydrazide for 1 hr at room temperature (reaction volume, 150 ul). Then the sample is dialyzed (Pierce Slidealizer Cassett, 10 kDa cutoff; Pierce Chemical Co., Rockford Ill.) at 4C first for 5 h, exchanging the buffer after each hour, and finally for 12 h against 500 ml buffer R (0.15 M NaCl, 1 mM MgCl2, 10 mM sodium phosphate, pH 7). Just before addition into a cuvette, the sample is diluted 1:5 in buffer ROG50 (Buffer R supplemented with 50 mM octylglucoside).

Example 64

TAQMAN

[1293] Quantitative PCR (QPCR). Total RNA from cells in culture are extracted by Trizol separation as recommended by the supplier (LifeTechnologies). (Total RNA is treated with DNase I (Life Technologies) to remove any contaminating genomic DNA before reverse transcription.) Total RNA (50 ng) is used in a one-step, 50 ul, RT-QPCR, consisting of Taqman Buffer A (Perkin-Elmer; 50 mM KCl/10 mM Tris, pH 8.3), 5.5 mM MgCl2, 240 μM each dNTP, 0.4 units RNase inhibitor(Promega), 8%glycerol, 0.012% Tween-20, 0.05% gelatin, 0.3 uM primers, O. 1 uM probe, 0.025units Amplitaq Gold (Perkin-Elmer) and 2.5 units Superscript II reverse transcriptase (Life Technologies). As a control for genomic contamination, parallel reactions are setup without reverse transcriptase. The relative abundance of (unknown) and 18S RNAs are assessed by using the Applied Biosystems Prism 7700 Sequence Detection System (Livak, K. J., Flood, S. J., Marmaro, J., Giusti, W. & Deetz, K. (1995) PCR Methods Appl. 4, 357-362). Reactions are carried out at 48° C. for 30 min, 95° C. for 10 min, followed by 40 cycles of 95° C. for 15s, 60° C. for 1 min. Reactions are performed in triplicate.

[1294] Primers (f & r) and FRET probes sets are designed using Primer Express Software (Perkin-Elmer). Probes are labeled at the 5′-end with the reporter dye 6-FAM and on the 3′-end with the quencher dye TAMRA (Biosource International, Camarillo, Calif. or Perkin-Elmer).

Example 65

Assays for Metalloproteinase Activity

[1295] Metalloproteinases (EC 3.4.24.-) are peptide hydrolases which use metal ions, such as Zn2+, as the catalytic mechanism. Metalloproteinase activity of polypeptides of the present invention can be assayed according to the following methods.

[1296] Proteolysis of alpha-2-macroglobulin

[1297] To confirm protease activity, purified polypeptides of the invention are mixed with the substrate alpha-2-macroglobulin (0.2 unit/ml; Boehringer Mannheim, Germany) in 1× assay buffer (50 mM HEPES, pH 7.5, 0.2 M NaCl, 10 mM CaCl2, 25 μM ZnCl2 and 0.05% Brij-35) and incubated at 37° C. for 1-5 days. Trypsin is used as positive control. Negative controls contain only alpha-2-macroglobulin in assay buffer. The samples are collected and boiled in SDS-PAGE sample buffer containing 5% 2-mercaptoethanol for 5-min, then loaded onto 8% SDS-polyacrylamide gel. After electrophoresis the proteins are visualized by silver staining. Proteolysis is evident by the appearance of lower molecular weight bands as compared to the negative control.

[1298] Inhibition of alpha-2-macroglobulin Proteolysis by Inhibitors of Metalloproteinases

[1299] Known metalloproteinase inhibitors (metal chelators (EDTA, EGTA, AND HgCl2), peptide metalloproteinase inhibitors (TIMP-1 and TIMP-2), and commercial small molecule MMP inhibitors) are used to characterize the proteolytic activity of polypeptides of the invention. The three synthetic MMP inhibitors used are: MMP inhibitor I, [IC50=1.0 μM against MMP-1 and MMP-8; IC50=30 μM against MMP-9; IC50=150 μM against MMP-3]; MMP-3 (stromelysin-1) inhibitor I [IC50=5 μM against MMP-3], and MMP-3 inhibitor II [Ki=130 nM against MMP-3]; inhibitors available through Calbiochem, catalog # 444250, 444218, and 444225, respectively). Briefly, different concentrations of the small molecule MMP inhibitors are mixed with purified polypeptides of the invention (50 μg/ml) in 22.9 μl of 1×HEPES buffer (50 mM HEPES, pH 7.5, 0.2 M NaCl, 10 mM CaCl2, 25 μM ZnCl2 and 0.05%Brij-35) and incubated at room temperature (24° C.) for 2-hr, then 7.1 μl of substrate alpha-2-macroglobulin (0.2 unit/ml) is added and incubated at 37° C. for 20-hr. The reactions are stopped by adding 4× sample buffer and boiled immediately for 5 minutes. After SDS-PAGE, the protein bands are visualized by silver stain.

[1300] Synthetic Fluorogenic Peptide Substrates Cleavage Assay

[1301] The substrate specificity for polypeptides of the invention with demonstrated metalloproteinase activity can be determined using synthetic fluorogenic peptide substrates (purchased from BACHEM Bioscience Inc). Test substrates include, M-1985, M-2225, M-2105, M-2110, and M-2255. The first four are MMP substrates and the last one is a substrate of tumor necrosis factor-α (TNF-α) converting enzyme (TACE). All the substrates are prepared in 1:1 dimethyl sulfoxide (DMSO) and water. The stock solutions are 50-500 μM. Fluorescent assays are performed by using a Perkin Elmer LS 50B luminescence spectrometer equipped with a constant temperature water bath. The excitation λ is 328 nm and the emission λ is 393 nm. Briefly, the assay is carried out by incubating 176 μl 1× HEPES buffer (0.2 M NaCl, 10 mM CaCl2, 0.05% Brij-35 and 50 mM HEPES, pH 7.5) with 4 μM of substrate solution (50 μM) at 25<C for 15 minutes, and then adding 20 μl of a purified polypeptide of the invention into the assay cuvett. The final concentration of substrate is 1 μM. Initial hydrolysis rates are monitored for 30-min.

Example 66

Characterization of the cDNA Contained in a Deposited Plasmid

[1302] The size of the cDNA insert contained in a deposited plasmid may be routinely determined using techniques known in the art, such as PCR amplification using synthetic primers hybridizable to the 3′ and 5′ ends of the cDNA sequence. For example, two primers of 17-30 nucleotides derived from each end of the cDNA (i.e., hybridizable to the absolute 5′ nucleotide or the 3′ nucleotide end of the sequence of SEQ ID NO:X, respectively) are synthesized and used to amplify the cDNA using the deposited cDNA plasmid as a template. The polymerase chain reaction is carried out under routine conditions, for instance, in 25 ul of reaction mixture with 0.5 ug of the above cDNA template. A convenient reaction mixture is 1.5-5 mM MgCl2, 0.01% (w/v) gelatin, 20 uM each of DATP, dCTP, dGTP, dTTP, 25 pmol of each primer and 0.25 Unit of Taq polymerase. Thirty five cycles of PCR (denaturation at 94 degree C. for 1 min; annealing at 55 degree C. for 1 min; elongation at 72 degree C. for 1 min) are performed with a Perkin-Elmer Cetus automated thermal cycler. The amplified product is analyzed by agarose gel electrophoresis. The PCR product is verified to be the selected sequence by subcloning and sequencing the DNA product.

[1303] Use of the above methodologies and/or other methodologies known in the art generates fragments from the clone corresponding to the approximate fragments described in Table 8, below. Accordingly, Table 8 provides a physical characterization of certain clones encompassed by the invention. The first column provides the unique clone identifier, “Clone ID NO:Z”, for cDNA clones of the invention, as described in Table 1A. The second column provides the approximate size of the cDNA insert contained in the corresponding cDNA clone.

19TABLE 8cDNAClone IDInsertNO: ZSize: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

[1304] It will be clear that the invention may be practiced otherwise than as particularly described in the foregoing description and examples. Numerous modifications and variations of the present invention are possible in light of the above teachings and, therefore, are within the scope of the appended claims.

[1305] The entire disclosure of each document cited (including patents, patent applications, journal articles, abstracts, laboratory manuals, books, or other disclosures) in the Background of the Invention, Detailed Description, and Examples is hereby incorporated herein by reference. In addition, the CD-R copy of the sequence listing submitted herewith and the corresponding computer readable form are both incorporated herein by reference in their entireties. The specification and sequence listing of each of the following U.S. applications are herein incorporated by reference in their entirety: Application No. 60/179,065, filed on Jan. 31, 2000; Application No. 60/180,628, filed on Feb. 4, 2000; Application No. 60/214,886, filed on Jun. 28, 2000; Application No. 60/217,487, filed on Jul. 11, 2000; Application No. 60/225,758, filed on Aug. 14, 2000; Application No. 60/220,963, filed on Jul. 26, 2000; Application No. 60/217,496, filed on Jul. 11, 2000; Application No. 60/225,447, filed on Aug. 14, 2000; Application No. 60/218,290, filed on Jul. 14, 2000; Application No. 60/225,757, filed on Aug. 14, 2000; Application No. 60/226,868, filed on Aug. 22, 2000; Application No. 60/216,647, filed on Jul. 7, 2000; Application No. 60/225,267, filed on Aug. 14, 2000; Application No. 60/216,880, filed on Jul. 7, 2000; Application No. 60/225,270, filed on Aug. 14, 2000; Application No. 60/251,869, filed on Dec. 8, 2000; Application No. 60/235,834, filed on Sep. 27, 2000; Application No. 60/234,274, filed on Sep. 21, 2000; Application No. 60/234,223, filed on Sep. 21, 2000; Application No. 60/228,924, filed on Aug. 30, 2000; Application No. 60/224,518, filed on Aug. 14, 2000; Application No. 60/236,369, filed on Sep. 29, 2000; Application No. 60/224,519, filed on Aug. 14, 2000; Application No. 60/220,964, filed on Jul. 26, 2000; Application No. 60/241,809, filed on Oct. 20, 2000; Application No. 60/249,299, filed on Nov. 17, 2000; Application No. 60/236,327, filed on Sep. 29, 2000; Application No. 60/241,785, filed on Oct. 20, 2000; Application No. 60/244,617, filed on Nov. 1, 2000; Application No. 60/225,268, filed on Aug. 14, 2000; Application No. 60/236,368, filed on Sep. 29, 2000; Application No. 60/251,856, filed on Dec. 8, 2000; Application No. 60/251,868, filed on Dec. 8, 2000; Application No. 60/229,344, filed on Sep. 1, 2000; Application No. 60/234,997, filed on Sep. 25, 2000; Application No. 60/229,343, filed on Sep. 1, 2000; Application No. 60/229,345, filed on Sep. 1, 2000; Application No. 60/229,287, filed on Sep. 1, 2000; Application No. 60/229,513, filed on Sep. 5, 2000; Application No. 60/231,413, filed on Sep. 8, 2000; Application No. 60/229,509, filed on Sep. 5, 2000; Application No. 60/236,367, filed on Sep. 29, 2000; Application No. 60/237,039, filed on Oct. 2, 2000; Application No. 60/237,038, filed on Oct. 2, 2000; Application No. 60/236,370, filed on Sep. 29, 2000; Application No. 60/236,802, filed on Oct. 2, 2000; Application No. 60/237,037, filed on Oct. 2, 2000; Application No. 60/237,040, filed on Oct. 2, 2000; Application No. 60/240,960, filed on Oct. 20, 2000; Application No. 60/239,935, filed on Oct. 13, 2000; Application No. 60/239,937, filed on Oct. 13, 2000; Application No. 60/241,787, filed on Oct. 20, 2000; Application No. 60/246,474, filed on Nov. 8, 2000; Application No. 60/246,532, filed on Nov. 8, 2000; Application No. 60/249,216, filed on Nov. 17, 2000; Application No. 60/249,210, filed on Nov. 17, 2000; Application No. 60/226,681, filed on Aug. 22, 2000; Application No. 60/225,759, filed on Aug. 14, 2000; Application No. 60/225,213, filed on Aug. 14, 2000; Application No. 60/227,182, filed on Aug. 22, 2000; Application No. 60/225,214, filed on Aug. 14, 2000; Application No. 60/235,836, filed on Sep. 27, 2000; Application No. 60/230,438, filed on Sep. 6, 2000; Application No. 60/215,135, filed on Jun. 30, 2000; Application No. 60/225,266, filed on Aug. 14, 2000; Application No. 60/249,218, filed on Nov. 17, 2000; Application No. 60/249,208, filed on Nov. 17, 2000; Application No. 60/249,213, filed on Nov. 17, 2000; Application No. 60/249,212, filed on Nov. 17, 2000; Application No. 60/249,207, filed on Nov. 17, 2000; Application No. 60/249,245, filed on Nov. 17, 2000; Application No. 60/249,244, filed on Nov. 17, 2000; Application No. 60/249,217, filed on Nov. 17, 2000; Application No. 60/249,211, filed on Nov. 17, 2000; Application No. 60/249,215, filed on Nov. 17, 2000; Application No. 60/249,264, filed on Nov. 17, 2000; Application No. 60/249,214, filed on Nov. 17, 2000; Application No. 60/249,297, filed on Nov. 17, 2000; Application No. 60/232,400, filed on Sep. 14, 2000; Application No. 60/231,242, filed on Sep. 8, 2000; Application No. 60/232,081, filed on Sep. 8, 2000; Application No. 60/232,080, filed on Sep. 8, 2000; Application No. 60/231,414, filed on Sep. 8, 2000; Application No. 60/231,244, filed on Sep. 8, 2000; Application No. 60/233,064, filed on Sep. 14, 2000; Application No. 60/233,063, filed on Sep. 14, 2000; Application No. 60/232,397, filed on Sep. 14, 2000; Application No. 60/232,399, filed on Sep. 14, 2000; Application No. 60/232,401, filed on Sep. 14, 2000; Application No. 60/241,808, filed on Oct. 20, 2000; Application No. 60/241,826, filed on Oct. 20, 2000; Application No. 60/241,786, filed on Oct. 20, 2000; Application No. 60/241,221, filed on Oct. 20, 2000; Application No. 60/246,475, filed on Nov. 8, 2000; Application No. 60/231,243, filed on Sep. 8, 2000; Application No. 60/233,065, filed on Sep. 14, 2000; Application No. 60/232,398, filed on Sep. 14, 2000; Application No. 60/234,998, filed on Sep. 25, 2000; Application No. 60/246,477, filed on Nov. 8, 2000; Application No. 60/246,528, filed on Nov. 8, 2000; Application No. 60/246,525, filed on Nov. 8, 2000; Application No. 60/246,476, filed on Nov. 8, 2000; Application No. 60/246,526, filed on Nov. 8, 2000; Application No. PT172, filed on Nov. 17, 2000; Application No. 60/246,527, filed on Nov. 8, 2000; Application No. 60/246,523, filed on Nov. 8, 2000; Application No. 60/246,524, filed on Nov. 8, 2000; Application No. 60/246,478, filed on Nov. 8, 2000; Application No. 60/246,609, filed on Nov. 8, 2000; Application No. 60/246,613, filed on Nov. 8, 2000; Application No. 60/249,300, filed on Nov. 17, 2000; Application No. 60/249,265, filed on Nov. 17, 2000; Application No. 60/246,610, filed on Nov. 8, 2000; Application No. 60/246,611, filed on Nov. 8, 2000; Application No. 60/230,437, filed on Sep. 6, 2000; Application No. 60/251,990, filed on Dec. 8, 2000; Application No. 60/251,988, filed on Dec. 5, 2000; Application No. 60/251,030, filed on Dec. 5, 2000; Application No. 60/251,479, filed on Dec. 6, 2000; Application No. PJ005, filed on Dec. 5, 2000; Application No. PJ006, filed on Dec. 6, 2000; Application No. 60/251,989, filed on Dec. 8, 2000; Application No. 60/250,391, filed on Dec. 1, 2000; and Application No. 60/254,097, filed on Dec. 11, 2000.

[1306] Moreover, the microfiche copy and the corresponding computer readable form of the Sequence Listing of U.S. Application Serial No. 60/179,065, and the hard copy of and the corresponding computer readable form of the Sequence Listing of U.S. Application Serial No. 60/180,628 are also incorporated herein by reference in their entireties.

Number	Date	Country
60179065	Jan 2000	US
60180628	Feb 2000	US
60214886	Jun 2000	US
60217487	Jul 2000	US
60225758	Aug 2000	US
60220963	Jul 2000	US
60217496	Jul 2000	US
60225447	Aug 2000	US
60218290	Jul 2000	US
60225757	Aug 2000	US
60226868	Aug 2000	US
60216647	Jul 2000	US
60225267	Aug 2000	US
60216880	Jul 2000	US
60225270	Aug 2000	US
60251869	Dec 2000	US
60235834	Sep 2000	US
60234274	Sep 2000	US
60234223	Sep 2000	US
60228924	Aug 2000	US
60224518	Aug 2000	US
60236369	Sep 2000	US
60224519	Aug 2000	US
60220964	Jul 2000	US
60241809	Oct 2000	US
60249299	Nov 2000	US
60236327	Sep 2000	US
60241785	Oct 2000	US
60244617	Nov 2000	US
60225268	Aug 2000	US
60236368	Sep 2000	US
60251856	Dec 2000	US
60251868	Dec 2000	US
60229344	Sep 2000	US
60234997	Sep 2000	US
60229343	Sep 2000	US
60229345	Sep 2000	US
60229287	Sep 2000	US
60229513	Sep 2000	US
60231413	Sep 2000	US
60229509	Sep 2000	US
60236367	Sep 2000	US
60237039	Oct 2000	US
60237038	Oct 2000	US
60236370	Sep 2000	US
60236802	Oct 2000	US
60237037	Oct 2000	US
60237040	Oct 2000	US
60240960	Oct 2000	US
60239935	Oct 2000	US
60239937	Oct 2000	US
60241787	Oct 2000	US
60246474	Nov 2000	US
60246532	Nov 2000	US
60249216	Nov 2000	US
60249210	Nov 2000	US
60226681	Aug 2000	US
60225759	Aug 2000	US
60225213	Aug 2000	US
60227182	Aug 2000	US
60225214	Aug 2000	US
60235836	Sep 2000	US
60230438	Sep 2000	US
60215135	Jun 2000	US
60225266	Aug 2000	US
60249218	Nov 2000	US
60249208	Nov 2000	US
60249213	Nov 2000	US
60249212	Nov 2000	US
60249207	Nov 2000	US
60249245	Nov 2000	US
60249244	Nov 2000	US
60249217	Nov 2000	US
60249211	Nov 2000	US
60249215	Nov 2000	US
60249264	Nov 2000	US
60249214	Nov 2000	US
60249297	Nov 2000	US
60232400	Sep 2000	US
60231242	Sep 2000	US
60232081	Sep 2000	US
60232080	Sep 2000	US
60231414	Sep 2000	US
60231244	Sep 2000	US
60233064	Sep 2000	US
60233063	Sep 2000	US
60232397	Sep 2000	US
60232399	Sep 2000	US
60232401	Sep 2000	US
60241808	Oct 2000	US
60241826	Oct 2000	US
60241786	Oct 2000	US
60241221	Oct 2000	US
60246475	Nov 2000	US
60231243	Sep 2000	US
60233065	Sep 2000	US
60232398	Sep 2000	US
60234998	Sep 2000	US
60246477	Nov 2000	US
60246528	Nov 2000	US
60246525	Nov 2000	US
60246476	Nov 2000	US
60246526	Nov 2000	US
60249209	Nov 2000	US
60246527	Nov 2000	US
60246523	Nov 2000	US
60246524	Nov 2000	US
60246478	Nov 2000	US
60246609	Nov 2000	US
60246613	Nov 2000	US
60249300	Nov 2000	US
60249265	Nov 2000	US
60246610	Nov 2000	US
60246611	Nov 2000	US
60230437	Sep 2000	US
60251990	Dec 2000	US
60251988	Dec 2000	US
60251030	Dec 2000	US
60251479	Dec 2000	US
60256719	Dec 2000	US
60250160	Dec 2000	US
60251989	Dec 2000	US
60250391	Dec 2000	US
60254097	Dec 2000	US
60231968	Sep 2000	US
60226279	Aug 2000	US
60186350	Mar 2000	US
60184664	Feb 2000	US
60189874	Mar 2000	US
60198123	Apr 2000	US
60227009	Aug 2000	US
60235484	Sep 2000	US
60190076	Mar 2000	US
60209467	Jun 2000	US
60205515	May 2000	US
60259678	Jan 2001	US

Nucleic acids, proteins, and antibodies

Information

Publication Number

Date Filed

Date Published

Inventors

CPC

US Classifications

International Classifications

Abstract

Description

Claims

Provisional Applications (136)