MULTIVALENT SMAC/DIABLO PROTEIN PEPTIDOMIMETIC COVALENTLY LINKED TO A TUMOR-HOMING PEPTIDE FOR TREATING CANCERS

SEQUENCE LISTING STATEMENT

A Sequence Listing conforming to the rules of WIPO Standard ST.26 is hereby incorporated by reference. The Sequence Listing has been filed as an electronic document encoded as XML in UTF-8 text. The electronic document, created on May 11, 2024, is entitled “P-629783-USP1_ST26.xml” and is 20,805 bytes in size.

FIELD OF THE INVENTION

The present invention relates to a multifunctional branched therapeutic agent comprising a target-homing moiety that is selective for cell entry, two or more target-binding moieties to engage in intracellular action, and a set of linker molecules that conjugate each of the moieties to a central core molecule. This novel therapeutic exploits the ability of a homing molecule to selectively bind to receptors of a specific cell type, promote cellular uptake, and then interact with an intracellular target through the chemical concept of avidity. In particular, the invention relates to multifunctional therapeutic peptide conjugates comprising a tumor-homing peptide, one or more target protein binding moieties, and linker molecules conjugating the tumor-homing peptide and the one or more target protein binding moieties to a branched reactive amino acid, wherein the protein binding moiety is an inhibitor of apoptosis (IAP). The invention also relates to methods for treating cancer comprising administering to a subject in need thereof a therapeutically effective amount of a multifunctional therapeutic peptide conjugate comprising a tumor-homing peptide, one or more target protein binding moieties, and linker molecules conjugating the tumor-homing peptide and the one or more target protein binding moieties to a branched reactive amino acid, wherein the protein binding moiety is an inhibitor of apoptosis (IAP).

BACKGROUND OF THE INVENTION

Targeting intracellular proteins implicated in the pathogenesis of a variety diseases, including the leading causes of mortality and morbidity, such as cancers, cardiovascular diseases and even infectious disease, has been a long-standing challenge in the field of drug discovery and development. Many of these proteins have been termed “undruggable” due to their lack of traditional small-molecule binding sites, or because of issues related to drug delivery and specificity. This problem is further exacerbated when these target proteins are overexpressed or mutated in diseased cells, contributing to disease progression and therapeutic resistance.

Many of these undruggable targets interact with other biomolecules in situ suggesting that a peptidomimetic antagonist approach may be able to invoke a therapeutic effect. However, peptides and peptide-like molecules still suffer from cellular targeting, cellular uptake, and the possibility of weak binding to the target protein. A promising strategy to overcome these issues is covalently conjugate a number of active moieties and to then conjugate those active moieties to a homing peptide. While the homing peptide promotes cellular targeting and uptake, the additional number of active binding moieties increases the avidity of the binder for the target and in turn increases the therapeutic effect.

Accordingly, there remains a critical need for developing improved anticancer therapeutic compositions and enhanced methods for treating cancers.

SUMMARY OF THE INVENTION

In still another aspect, provided herein are multifunctional therapeutic peptide conjugates represented by formula (T):

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In one aspect, provided herein are multifunctional therapeutic peptide conjugates represented by formula (II):

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In one aspect, provided herein are multifunctional therapeutic peptide conjugates represented by formula (III):

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In another aspect, provided herein are methods for treating cancer comprising administering a to a subject in need thereof a therapeutically effective amount of a multifunctional therapeutic peptide conjugate compound to a subject in need thereof, the multifunctional therapeutic peptide conjugate compound comprising: a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker and the first and second linkers are conjugated to the branched reactive amino acid, wherein the protein binding moiety is an inhibitor of apoptosis (IAP).

In still another aspect, provided herein are methods for treating cancer comprising administering to a subject in need thereof a therapeutically effective amount of a multifunctional therapeutic peptide conjugate compound comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing an RGD peptide motif consisting of the three amino acids Arg-Gly-Asp.

In one aspect, provided herein are provided herein are methods for treating cancer comprising administering to a subject in need thereof a therapeutically effective amount of a multifunctional therapeutic peptide conjugate compound comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Asp-Cys (SEQ ID NO: 1).

In another aspect, provided herein are methods for treating cancer comprising administering to a subject in need thereof a therapeutically effective amount of a multifunctional therapeutic peptide conjugate compound comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Asp-Cys (SEQ ID NO: 2).

In still another aspect, provided herein are methods for treating cancer comprising administering to a subject in need thereof a therapeutically effective amount of a multifunctional therapeutic peptide conjugate compound comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Glu-Cys (SEQ ID NO: 3).

In another aspect, provided herein are methods for treating cancer comprising administering to a subject in need thereof a therapeutically effective amount of a multifunctional therapeutic peptide conjugate compound comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a NGR motif consisting of the five amino acids Cys-Asn-Gly-Arg-Cys (SEQ ID NO: 5).

In still another aspect, provided herein are methods for treating cancer comprising administering to a subject in need thereof a therapeutically effective amount of a multifunctional therapeutic peptide conjugate compound comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein tumor homing peptide comprises a cyclic peptide sequence consisting of the nine amino acid sequence Cys-Gly-Asn-Lys-Arg-Thr-Arg-Gly-Cys (SEQ ID NO: 6).

In one aspect, provided herein are methods for treating cancer comprising administering to a subject in need thereof a therapeutically effective amount of a multifunctional therapeutic peptide conjugate compound comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein the tumor homing peptide has a peptide sequence consisting of the 29 amino acids Lys-Asp-Glu-Pro-Gln-Arg-Arg-Ser-Ala-Arg-Leu-Ser-Ala-Lys-Pro-Ala-Pro-Pro-Lys-Pro-Glu-Pro-Lys-Lys-Ala-Pro-Ala-Lys-Lys (SEQ ID NO: 7).

In another aspect, provided herein are methods for treating cancer comprising administering to a subject in need thereof a therapeutically effective amount of a multifunctional therapeutic peptide conjugate compound comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein the compounds comprise one protein binding moiety consisting of a seven amino acid sequence AVPIAQK (SEQ ID NO: 10) from the N-terminus of the apoptosis mediator Smac/DIABLO protein.

In another aspect, provided herein are methods for treating cancer comprising administering to a subject in need thereof a therapeutically effective amount of a multifunctional therapeutic peptide conjugate compound comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein the compounds comprise two protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence [AIB]VPIAQK (SEQ ID NO: 11) from the N-terminus of the apoptosis mediator Smac/DIABLO protein, wherein [AIB] is 2-Aminoisobutyric acid.

In still another aspect, provided herein are methods for treating cancer comprising administering to a subject in need thereof a therapeutically effective amount of a multifunctional therapeutic peptide conjugate compound comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein the compounds comprise two protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence [AIB]VPI[AIB]QK (SEQ ID NO: 12) from the N-terminus of the apoptosis mediator Smac/DIABLO protein, wherein [AIB] is 2-Aminoisobutyric acid.

In one aspect, provided herein are methods for treating cancer comprising administering to a subject in need thereof a therapeutically effective amount of a multifunctional therapeutic peptide conjugate compound comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein the compounds comprise two protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence X₁X₂PIAQK (SEQ ID NO: 13) and X₁is N-methylated alanine (MeA) and X₂is tert-Leucine (Tle) (i.e., MeATlePIAQK (SEQ ID NO: 13)), from the N-terminus of the apoptosis mediator Smac/DIABLO protein.

In one aspect, provided herein are methods for treating cancer comprising administering to a subject in need thereof a therapeutically effective amount of a multifunctional therapeutic peptide conjugate compound comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein the compounds comprise two protein binding moieties, wherein one of the two protein binding moiety consists of a seven amino acid sequence X₁X₂PFAQK (SEQ ID NO: 14) (i.e., MeATlePFAQK (SEQ ID NO: 14)), wherein X₁is N-methylated alanine (MeA) and X₂is tert-Leucine (Tle) and the second of the two protein binding moiety consists of a seven amino acid sequence X₁X₂PIAQK (SEQ ID NO: 13) (i.e., MeATlePIAQK (SEQ ID NO: 13)), from the N-terminus of the apoptosis mediator Smac/DIABLO protein.

Other features and advantages of the present invention will become apparent from the following detailed description examples and figures. It should be understood, however, that the detailed description and the specific examples while indicating preferred embodiments of the invention are given by way of illustration only, since various changes and modifications within the spirit and scope of the invention will become apparent to those skilled in the art from this detailed description.

BRIEF DESCRIPTION OF THE DRAWINGS

The following drawing forms part of the present specification and are included to further demonstrate certain aspects of the present disclosure, the inventions of which can be better understood by reference to one or more of these drawings in combination with the detailed description of specific embodiments presented herein.

FIG. 1 depicts an embodiment of a multifunctional therapeutic peptide conjugate provided by the present disclosure.

FIG. 2 depicts an embodiment of a multifunctional therapeutic peptide conjugate provided by the present disclosure in which each of the two XIAP binding domains, X₁X₂PIAQK (SEQ ID NO: 13), of the Smac/Diablo protein, is modified at amino acid sequence positions 1 and 2 to be N-methylated alanine and tert-leucine respectively, such that AVPIAQK (SEQ ID NO: 10) is modified to X₁X₂PIAQK (SEQ ID NO: 13), wherein X₁is N-methylated alanine (MeA) at amino acid sequence position 1 and X₂is tert-Leucine (Tle) at amino acid sequence position 2, i.e., to make MeATlePIAQK (SEQ ID NO: 13). These modifications improve proteolytic stability and increase binding affinity.

FIG. 3 depicts an embodiment of a multifunctional therapeutic peptide conjugate provided by the present disclosure in which one of the two fragment chains is modified at amino acid position 4 from isoleucine to phenylalanine (either one of the two chains may be modified): from X₁X₂PIAQK (SEQ ID NO: 13), wherein X₁is N-methyl alanine (MeA) and X₂is tert-Leucine (Tle), i.e., MeATlePIAQK (SEQ ID NO: 13) of the Smac/Diablo protein is modified at amino acid sequence position 4 from isoleucine (I) to phenylalanine (F), i.e., to X₁X₂PFAQK (SEQ ID NO: 14), wherein X₁is and X₂is N-methylated alanine (MeA) at amino acid sequence position 1 and X₂is tert-Leucine (Tle) at amino acid sequence position 2, i.e., to MeATlePFAQK (SEQ ID NO: 14). This modification specifically biases one chain to bind the Bcl-2 domain of XIAP and the other domain to bind Bcl-3 domain of XIAP.

FIG. 4 depicts an embodiment of a a multifunctional therapeutic peptide conjugate provided by the present disclosure (BXL_2023.002) comprising a multivalent XIAP antagonist branched peptide conjugated to an iRGD tumor targeting peptide.

FIG. 5 is a plot of cell viability as a function of drug concentration for BXL_2023.002 (depicted in FIG. 4; where n=8) and showing specificity for pancreatic cancer cell line ASPC-1 (circles). Cell viability for the control HEK-293 cell line is also shown (squares).

It will be appreciated that for simplicity and clarity of illustration, elements shown in the figure have not necessarily been drawn to scale. For example, the dimensions of some of the elements may be exaggerated relative to other elements for clarity. Further, where considered appropriate, reference numerals may be repeated in the figure to indicate corresponding or analogous elements.

DETAILED DESCRIPTION OF THE INVENTION

The present subject matter may be understood more readily by reference to the following detailed description which forms a part of this disclosure. It is to be understood that this invention is not limited to the specific products, methods, conditions or parameters described and/or shown herein, and that the terminology used herein is for the purpose of describing particular embodiments by way of example only and is not intended to be limiting of the claimed invention.

Numerous types of cancer occur in humans; however, each cancer cell has alterations that lead to the production of abnormal proteins responsible for uncontrolled cell growth and survival. Certain anti-cancer therapeutics bind to the such abnormal proteins, thereby impeding their activity. Nevertheless, some “undruggable” targets exist wherein the cancer-inducing proteins escape attempts to block their adverse effects.

Apoptosis is a form of programmed cell-death that occurs as a normal and controlled cellular process of an organism's growth or development to eliminate damaged or unwanted cells. Malfunctioning apoptosis is a characteristic of human cancers. Key regulators of cell death and survival are XIAP, cIAP1, and cIAP2, members of inhibitor of apoptosis (IAP) proteins.

Smac/DIABLO proteins are a family endogenous antagonists of inhibitors of apoptosis (IAP) proteins XIAP, cIAP1, and cIAP2.

The present invention provides a conjugate of a peptide moiety derived from the mature Smac/Diablo family of proteins and a tumor-homing peptide moiety known as iRGD, bridged by a reactive amino acid and polyethylene linkers, that offers the possibility to overcome the limitations of current peptide therapeutics, while delivering superior pharmacological benefits. The molecular linker may be PEG. The connection involves covalent bonding of the iRGD peptide to the α-amino group of the branched reactive amino acid via a molecular linker. In an embodiment the molecular linker is a PEG linker. The first seven amino acids from the N-terminus of the Smac/Diablo protein are then attached to both the α- and ε-amino groups of the lysine via a molecular linker creating a bifunctional therapeutic peptide conjugate. The peptide moieties are generally greater than 100 Da and less than 106.

The therapeutic conjugate described herein aim to resolve the issues of drug delivery and specificity of “undruggable” proteins.

In another aspect, the therapeutic peptide conjugate of this invention comprises an iRGD peptide, a branched reactive amino acid, and two fragments of the Smac/Diablo protein. Specifically, the conjugate consists of the iRGD peptide, a first linker, a branched reactive amino acid, a second and third linker, two fragments of the Smac/Diablo protein each consisting of the seven amino acid sequence AVPIAQK (SEQ ID NO: 10) from its N-terminus, wherein the first, second, and third linkers are each conjugated to the branched reactive amino acid by a molecular linker. A branched reactive amino acid is defined herein as an amino acid residue having two active branching sites. Each active branching site is an amine group, an azido group, a thiol group or a diamine group having two amines. In a particular embodiment, the branched reactive amino acid is a lysine containing two primary amine groups. In an embodiment, the lysine is L-lysine or D-lysine or an azido-lysine, i.e., 6-azido-L-lysine or 6-azido-D-lysine. In another embodiment, the reactive amino acid is a cysteine, which contains a reactive thiol (or “sulfhydryl”) group. In an embodiment, the reactive amino acid is a 5-azido-L-ornithine or 5-azido-d-ornithine. The each molecular linker may be polyethylene glycol (PEG). The PEG linkers may be the same PEG or different a different PEG. In an embodiment, the first linker and the second linker is an identical or different polyethylene glycol (PEG) selected from the group consisting of amino-PEG2, amino-PEG3, amino-PEG4, amino-PEG6, amino-PEG8, amino-PEG10, amino-PEG12 and amino-PEG20.

In an embodiment of the herein described multifunctional therapeutic peptide conjugates, the multifunctional therapeutic peptide conjugate comprises a tumor homing peptide, at least one protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the at least one protein binding moiety is conjugated to a second linker and the first and second linkers are conjugated to the branched reactive amino acid, wherein the at least one protein binding moiety is an inhibitor of apoptosis (IAP). In certain embodiments, the at least one protein binding moiety consists of a seven amino acid sequence AVPIAQK (SEQ ID NO: 10) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In some embodiments, the at least one protein binding moiety consists of a seven amino acid sequence 2-Aminoisobutyric acid [AIB]VPIAQK (SEQ ID NO: 11) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In an embodiment, the at least one protein binding moiety consists of a seven amino acid sequence [AIB]VPI[AIB]QK (SEQ ID NO: 12) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In a particular embodiment, the multifunctional therapeutic peptide conjugate comprises two protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence AVPIAQK (SEQ ID NO: 10) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In various embodiments, the multifunctional therapeutic peptide conjugate comprises two protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence 2-Aminoisobutyric acid [AIB]VPIAQK (SEQ ID NO: 11) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In an embodiment, the multifunctional therapeutic peptide conjugate comprises two protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence [AIB]VPI[AIB]QK (SEQ ID NO: 12) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In another embodiment, the multifunctional therapeutic peptide conjugate comprises three protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence AVPIAQK (SEQ ID NO: 10) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In some embodiments, the multifunctional therapeutic peptide conjugate comprises three protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence 2-Aminoisobutyric acid [AIB]VPIAQK (SEQ ID NO: 11) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In certain embodiments, the multifunctional therapeutic peptide conjugate comprises three protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence [AIB]VPI[AIB]QK (SEQ ID NO: 12) from the N-terminus of the apoptosis mediator Smac/DIABLO protein.

The invention relates to multifunctional therapeutic peptide conjugates comprising a tumor-homing peptide, one or more target protein binding moieties, and linker molecules conjugating the tumor-homing peptide and the one or more target protein binding moieties to a branched reactive amino acid, wherein the protein binding moiety is an inhibitor of apoptosis (IAP). The invention also relates to methods for treating cancer comprising administering to a subject in need thereof a therapeutically effective amount of a multifunctional therapeutic peptide conjugate comprising a tumor-homing peptide, one or more target protein binding moieties, and linker molecules conjugating the tumor-homing peptide and the one or more target protein binding moieties to a branched reactive amino acid, wherein the protein binding moiety is an inhibitor of apoptosis (IAP). In particular embodiments, the invention relates to a therapeutic multivalent SMAC/Diablo protein peptidomimetic covalently linked to a tumor-homing peptide, wherein the tumor-homing peptide and the N-terminal seven amino acid sequence AVPIAQK (SEQ ID NO: 10) of the SMAC/Diablo protein are each conjugated to a linker molecule and each linker molecule is connected to a branched reactive amino acid.

In one aspect, the present invention provides an improved bifunctional therapeutic peptide conjugate provided herein comprises an N-terminal seven amino acid sequence AVPIAQK (SEQ ID NO: 10) of the SMAC/Diablo protein and the iRGD peptide, connected via a central branched reactive amino acid, wherein the iRGD peptide and the N-terminal seven amino acid sequence AVPIAQK (SEQ ID NO: 10) of the SMAC/Diablo protein are each conjugated to a linker molecule and each linker molecule is connected to the central branched reactive amino acid. In an embodiment, the central branched reactive amino acid is lysine. In an embodiment, the molecular linker is polyethylene glycol (PEG).

In another aspect, the present invention provides a multifunctional therapeutic peptide conjugate comprising: a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker and the first and second linkers are conjugated to the branched reactive amino acid, wherein the protein binding moiety is an inhibitor of apoptosis (IAP). In an embodiment of the multifunctional therapeutic peptide conjugate, the tumor homing peptide is the cyclic peptide iRGD having amino acid sequence CRGD[K/R]GP[D/E]C, i.e., Cys-Arg-Gly-Asp-Lys-Gly-Pro-Asp-Cys (SEQ ID NO: 1) and Cys-Arg-Gly-Asp-Arg-Gly-Pro-Glu-Cys (SEQ ID NO: 4). In an embodiment, the tumor homing peptide is the cyclic peptide iRGD having amino acid sequence Cys-Asn-Gly-Arg-Cys (SEQ ID NO: 5). In another embodiment, the tumor homing peptide is the cyclic peptide iRGD having amino acid sequence Cys-Gly-Asn-Lys-Arg-Thr-Arg-Gly-Cys (SEQ ID NO: 6). In some embodiments, the tumor homing peptide is the cyclic peptide iRGD having amino acid sequence Lys-Asp-Glu-Pro-Gln-Arg-Arg-Ser-Ala-Arg-Leu-Ser-Ala-Lys-Pro-Ala-Pro-Pro-Lys-Pro-Glu-Pro-Lys-Lys-Ala-Pro-Ala-Lys-Lys (SEQ ID NO: 7). In an embodiment, the tumor homing peptide is the cyclic peptide iRGD having amino acid sequence Cys-Arg-Glu-Lys-Ala (SEQ ID NO: 8). In another embodiment, the tumor homing peptide is the cyclic peptide iRGD having amino acid sequence Lys-Glu-Thr-Trp-Trp-Glu-Thr-Trp-Trp-Thr-Glu-Trp-Ser-Gln-Pro-Lys-Lys-Lys-Arg-Lys-Val (SEQ ID NO: 9). In an embodiment, the branched reactive amino acid is a lysine, a cysteine, an azido-lysine or an azido-ornithine. In a particular embodiment, the branched reactive amino acid is lysine. In another particular embodiment, the branched reactive amino acid is cysteine. In an embodiment, the lysine is L-lysine or D-lysine or an azido-lysine, i.e., 6-azido-L-lysine or 6-azido-D-lysine. In an embodiment, the branched reactive amino acid is azido-Lysine. In an embodiment, the reactive amino acid is a 5-azido-L-ornithine or 5-azido-d-ornithine. In some embodiments, the multifunctional therapeutic peptide conjugate comprises one protein binding moiety consisting of a seven amino acid sequence AVPIAQK (SEQ ID NO: 10) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In an embodiment, the multifunctional therapeutic peptide conjugate comprises one protein binding moiety consisting of a seven amino acid sequence [AIB]VPIAQK (SEQ ID NO: 11), wherein [AIB] is 2-Aminoisobutyric acid. In another embodiment, the multifunctional therapeutic peptide conjugate comprises one protein binding moiety consisting of a seven amino acid sequence [AIB]VPI[AIB]QK (SEQ ID NO: 12), wherein [AIB] is 2-Aminoisobutyric acid. In some embodiments, the multifunctional therapeutic peptide conjugate comprises two protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence AVPIAQK (SEQ ID NO: 10) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In some embodiments, the first and second linker are the same. In certain embodiments, the first and second linker are different linker molecules. In an embodiment of the multifunctional therapeutic peptide conjugate, the molecular linker is polyethylene glycol (PEG). In a particular embodiment, the first and second polyethylene glycol (PEG) molecular linker, includes but is not limited to amino-PEG2, amino-PEG3, amino-PEG4, amino-PEG6, amino-PEG8, amino-PEG10, amino-PEG12, and/or amino-PEG20. In some embodiments, the first and second linker are poly(glycerols) (PGs), poly(oxazolines) (POX), poly(hydroxypropyl methacrylate) (PHPMA), poly(2-hydroxyethyl methacrylate) (PHEMA), poly(N-(2-hydroxypropyl) methacrylamide) (HPMA), poly(vinylpyrrolidone) (PVP), poly(N,N-dimethyl acrylamide) (PDMA), and/or poly(N-acryloylmorpholine) (PAcM). In an embodiment of the multifunctional therapeutic peptide conjugate, the linker enhances the stability and bioavailability of the conjugate. In certain embodiments, the multifunctional therapeutic peptide conjugate is for use in the treatment of cancer.

In another aspect, the present the invention provides an improved bifunctional therapeutic peptide conjugate comprising the N-terminus of the Smac/Diablo protein and the iRGD peptide, wherein the N-terminus of the Smac/Diablo protein and the iRGD peptide are each connected to lysine by polyethylene glycol (PEG).

In one aspect, the present invention provides an improved multifunctional therapeutic peptide conjugate comprising two N-terminal seven amino acid sequences AVPIAQK (SEQ ID NO: 10) of the SMAC/Diablo protein and the iRGD peptide, wherein each N-terminal seven amino acid sequence AVPIAQK (SEQ ID NO: 10) of the Smac/Diablo protein and the iRGD peptide are connected to a central branched reactive amino acid by a linker molecule and each linker molecule is connected to the central branched reactive amino acid. In an embodiment, the central branched reactive amino acid is lysine. In an embodiment, the molecular linker is polyethylene glycol (PEG).

In another aspect, the present invention provides an improved multifunctional therapeutic peptide conjugate comprising two N-terminal seven amino acid sequences AVPIAQK (SEQ ID NO: 10) of the SMAC/Diablo protein and the iRGD peptide, wherein each N-terminal seven amino acid sequence AVPIAQK (SEQ ID NO: 10) of the Smac/Diablo protein and the iRGD peptide are connected to a lysine by polyethylene glycol (PEG).

In a further aspect, the present the invention provides a method for treating cancer comprising administering a therapeutically effective amount of a multifunctional therapeutic peptide conjugate to a subject in need thereof, the multifunctional therapeutic peptide conjugate comprising: a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker and the first and second linkers are conjugated to the branched reactive amino acid, wherein the protein binding moiety is an inhibitor of apoptosis (IAP). In an embodiment of said method, the tumor homing peptide is the cyclic peptide iRGD having amino acid sequence CRGD[K/R]GP[D/E]C (SEQ ID NO: 1) and (SEQ ID NO: 4), i.e., Cys-Arg-Gly-Asp-Lys-Gly-Pro-Asp-Cys (SEQ ID NO: 1) and Cys-Arg-Gly-Asp-Arg-Gly-Pro-Glu-Cys (SEQ ID NO: 4).

In an embodiment, the branched reactive amino acid is lysine. In some embodiments of the herein provided method, the multifunctional therapeutic peptide conjugate comprises one protein binding moiety consisting of a seven amino acid sequence AVPIAQK (SEQ ID NO: 10) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In certain embodiments of the herein provided method, the multifunctional therapeutic peptide conjugate comprises two protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence AVPIAQK (SEQ ID NO: 10) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In some embodiments of said methods, the molecular linker is polyethylene glycol (PEG).

In one aspect, the present the invention provides a method for treating cancer comprising administering a to a subject in need thereof a therapeutically effective amount of an improved bifunctional therapeutic peptide conjugate comprising the N-terminus of the Smac/Diablo protein consisting of seven amino acid sequence AVPIAQK (SEQ ID NO: 10) and the iRGD peptide, wherein the N-terminus of the Smac/Diablo protein and the iRGD peptide are connected to a branched reactive amino acid by a molecular linker. In an embodiment of the provided methods, the branched reactive amino acid is lysine. In an embodiment, the molecular linker is polyethylene glycol (PEG).

In another aspect, the present the invention provides a method for treating cancer comprising administering to a subject in need thereof a therapeutically effective amount of an improved multifunctional therapeutic peptide conjugate comprising two N-termini of the Smac/Diablo protein, each N-terminal sequence consisting of the seven amino acid sequence AVPIAQK (SEQ ID NO: 10) of the Smac/Diablo protein and the iRGD peptide, wherein the two N-termini and the iRGD peptide are each connected to a branched reactive amino acid by a molecular linker. In an embodiment of the provided methods, the branched reactive amino acid is lysine. In an embodiment, the molecular linker is polyethylene glycol (PEG).

In an embodiment of the herein described methods for treating cancer, the multifunctional therapeutic peptide conjugate comprises a tumor homing peptide, at least one protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the at least one protein binding moiety is conjugated to a second linker and the first and second linkers are conjugated to the branched reactive amino acid, wherein the at least one protein binding moiety is an inhibitor of apoptosis (IAP). In certain embodiments, the at least one protein binding moiety consists of a seven amino acid sequence AVPIAQK (SEQ ID NO: 10) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In some embodiments, the at least one protein binding moiety consists of a seven amino acid sequence 2-Aminoisobutyric acid [AIB]VPIAQK (SEQ ID NO: 11) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In an embodiment, the at least one protein binding moiety consists of a seven amino acid sequence [AIB]VPI[AIB]QK (SEQ ID NO: 12) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In a particular embodiment, the multifunctional therapeutic peptide conjugate comprises two protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence AVPIAQK (SEQ ID NO: 10) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In various embodiments, the multifunctional therapeutic peptide conjugate comprises two protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence 2-Aminoisobutyric acid [AIB]VPIAQK (SEQ ID NO: 11) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In an embodiment, the multifunctional therapeutic peptide conjugate comprises two protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence [AIB]VPI[AIB]QK (SEQ ID NO: 12) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In another embodiment, the multifunctional therapeutic peptide conjugate comprises three protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence AVPIAQK (SEQ ID NO: 10) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In some embodiments, the multifunctional therapeutic peptide conjugate comprises three protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence 2-Aminoisobutyric acid [AIB]VPIAQK (SEQ ID NO: 11) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In certain embodiments, the multifunctional therapeutic peptide conjugate comprises three protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence [AIB]VPI[AIB]QK (SEQ ID NO: 12) from the N-terminus of the apoptosis mediator Smac/DIABLO protein.

In embodiments of the above-described multifunctional therapeutic peptide conjugate compounds comprising an active cellular targeting moiety (A) having a cyclic peptide sequence, the cyclic peptide sequence containing the cyclic iRGD peptide motif is cyclized via a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9.

In certain embodiments of the above-described multifunctional therapeutic peptide conjugate compounds comprising an active cellular targeting moiety (A) having a cyclic peptide sequence containing the NGR motif, the cyclic peptide sequence is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 5. In an embodiment of the above-described multifunctional therapeutic peptide conjugate compounds comprising an active cellular targeting moiety (A) having a cyclic peptide sequence containing the NGR motif, the active cellular targeting moiety targets the extracellular enzyme Aminopeptidase N (CD13).

In an embodiment of the above-described multifunctional therapeutic peptide conjugate compounds comprising the cyclic peptide sequence consisting of the nine amino acid sequence Cys-Gly-Asn-Lys-Arg-Thr-Arg-Gly-Cys, the cyclic peptide is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In some embodiments of said multifunctional therapeutic peptide conjugate compounds, the active cellular targeting moiety targets the cellular receptor p32/gC1qR.

In an embodiment of the above-described multifunctional therapeutic peptide conjugate compounds, the branched reactive amino acid is a lysine, a cysteine, an azido-lysine or an azido-ornithine. In a particular embodiment, the branched reactive amino acid is lysine. In another embodiment, the branched reactive amino acid is cysteine. In some embodiments, the branched reactive amino acid is azido-lysine. In an embodiment, the lysine is L-lysine or D-lysine or an azido-lysine, i.e., 6-azido-L-lysine or 6-azido-D-lysine. In an embodiment, the reactive amino acid is a 5-azido-L-ornithine or 5-azido-d-ornithine.

In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing an RGD peptide motif consisting of the three amino acids Arg-Gly-Asp. In another embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Asp-Cys (SEQ ID NO: 1). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Asp-Cys (SEQ ID NO: 2). In some embodiments, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Glu-Cys (SEQ ID NO: 3). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Glu-Cys (SEQ ID NO: 4). In another embodiment, the cyclic peptide sequence containing the cyclic iRGD peptide motif is cyclized via a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In some embodiments, the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing a NGR motif consisting of the three amino acids Asn-Gly-Arg. In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a NGR motif consisting of the five amino acids Cys-Asn-Gly-Arg-Cys (SEQ ID NO: 5). In an embodiment, the cyclic peptide sequence containing the NGR motif is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 5. In another embodiment, the tumor homing peptide comprises a cyclic peptide sequence consisting of the nine amino acid sequence Cys-Gly-Asn-Lys-Arg-Thr-Arg-Gly-Cys (SEQ ID NO: 6). In an embodiment, the cyclic peptide is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In another embodiment, the tumor homing peptide comprises a peptide sequence consisting of the 29 amino acids Lys-Asp-Glu-Pro-Gln-Arg-Arg-Ser-Ala-Arg-Leu-Ser-Ala-Lys-Pro-Ala-Pro-Pro-Lys-Pro-Glu-Pro-Lys-Lys-Ala-Pro-Ala-Lys-Lys (SEQ ID NO: 7). In some embodiments, the tumor homing peptide has a peptide sequence consisting of the amino acids Cys-Arg-Glu-Lys-Ala (SEQ ID NO: 5). In another embodiment, the tumor homing peptide comprises a peptide sequence consisting of the amino acids Cys-Arg-Glu-Lys-Ala (SEQ ID NO: 5). In an embodiment, the tumor homing peptide comprises a peptide sequence consisting of the amino acids Lys-Glu-Thr-Trp-Trp-Glu-Thr-Trp-Trp-Thr-Glu-Trp-Ser-Gln-Pro-Lys-Lys-Lys-Arg-Lys-Val (SEQ ID NO: 9). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing an RGD peptide motif consisting of the three amino acids Arg-Gly-Asp. In another embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Asp-Cys (SEQ ID NO: 1). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Asp-Cys (SEQ ID NO: 2). In some embodiments, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Glu-Cys (SEQ ID NO: 3). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Glu-Cys (SEQ ID NO: 4). In another embodiment, the cyclic peptide sequence containing the cyclic iRGD peptide motif is cyclized via a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In some embodiments, the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing a NGR motif consisting of the three amino acids Asn-Gly-Arg. In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a NGR motif consisting of the five amino acids Cys-Asn-Gly-Arg-Cys (SEQ ID NO: 5). In an embodiment, the cyclic peptide sequence containing the NGR motif is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 5. In another embodiment, the tumor homing peptide comprises a cyclic peptide sequence consisting of the nine amino acid sequence Cys-Gly-Asn-Lys-Arg-Thr-Arg-Gly-Cys (SEQ ID NO: 6). In an embodiment, the cyclic peptide is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In another embodiment, the tumor homing peptide comprises a peptide sequence consisting of the 29 amino acids Lys-Asp-Glu-Pro-Gln-Arg-Arg-Ser-Ala-Arg-Leu-Ser-Ala-Lys-Pro-Ala-Pro-Pro-Lys-Pro-Glu-Pro-Lys-Lys-Ala-Pro-Ala-Lys-Lys (SEQ ID NO: 7). In some embodiments, the tumor homing peptide has a peptide sequence consisting of the amino acids Cys-Arg-Glu-Lys-Ala (SEQ ID NO: 8). In another embodiment, the tumor homing peptide comprises a peptide sequence consisting of the amino acids Cys-Arg-Glu-Lys-Ala. In an embodiment, the tumor homing peptide comprises a peptide sequence consisting of the amino acids Lys-Glu-Thr-Trp-Trp-Glu-Thr-Trp-Trp-Thr-Glu-Trp-Ser-Gln-Pro-Lys-Lys-Lys-Arg-Lys-Val (SEQ ID NO: 9).

In another aspect, provided herein are multifunctional therapeutic peptide conjugate compounds comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein the compounds comprise one protein binding moiety consisting of a seven amino acid sequence [AIB]VPI[AIB]QK (SEQ ID NO: 12) from the N-terminus of the apoptosis mediator Smac/DIABLO protein, wherein [AIB] is 2-Aminoisobutyric acid. In an embodiment, the branched reactive amino acid is a lysine, a cysteine, an azido-lysine or an azido-ornithine. In a particular embodiment, the branched reactive amino acid is lysine. In another embodiment, the branched reactive amino acid is cysteine. In some embodiments, the branched reactive amino acid is azido-lysine. In an embodiment, the lysine is L-lysine or D-lysine or an azido-lysine, i.e., 6-azido-L-lysine or 6-azido-D-lysine. In an embodiment, the reactive amino acid is a 5-azido-L-ornithine or 5-azido-d-ornithine. In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing an RGD peptide motif consisting of the three amino acids Arg-Gly-Asp. In another embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Asp-Cys (SEQ ID NO: 1). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Asp-Cys (SEQ ID NO: 2). In some embodiments, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Glu-Cys (SEQ ID NO: 3). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Glu-Cys (SEQ ID NO: 4). In another embodiment, the cyclic peptide sequence containing the cyclic iRGD peptide motif is cyclized via a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In some embodiments, the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing a NGR motif consisting of the three amino acids Asn-Gly-Arg. In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a NGR motif consisting of the five amino acids Cys-Asn-Gly-Arg-Cys (SEQ ID NO: 5). In an embodiment, the cyclic peptide sequence containing the NGR motif is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 5. In another embodiment, the tumor homing peptide comprises a cyclic peptide sequence consisting of the nine amino acid sequence Cys-Gly-Asn-Lys-Arg-Thr-Arg-Gly-Cys (SEQ ID NO: 6). In an embodiment, the cyclic peptide is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In another embodiment, the tumor homing peptide comprises a peptide sequence consisting of the 29 amino acids Lys-Asp-Glu-Pro-Gln-Arg-Arg-Ser-Ala-Arg-Leu-Ser-Ala-Lys-Pro-Ala-Pro-Pro-Lys-Pro-Glu-Pro-Lys-Lys-Ala-Pro-Ala-Lys-Lys (SEQ ID NO: 7). In some embodiments, the tumor homing peptide has a peptide sequence consisting of the amino acids Cys-Arg-Glu-Lys-Ala (SEQ ID NO: 8). In another embodiment, the tumor homing peptide comprises a peptide sequence consisting of the amino acids Cys-Arg-Glu-Lys-Ala. In an embodiment, the tumor homing peptide comprises a peptide sequence consisting of the amino acids Lys-Glu-Thr-Trp-Trp-Glu-Thr-Trp-Trp-Thr-Glu-Trp-Ser-Gln-Pro-Lys-Lys-Lys-Arg-Lys-Val (SEQ ID NO: 9).

In an embodiment, the branched reactive amino acid is lysine, cysteine or azido-lysine. In a particular embodiment, the branched reactive amino acid is lysine. In another embodiment, the branched reactive amino acid is cysteine. In some embodiments, the branched reactive amino acid is azido-lysine. In an embodiment, the lysine is L-lysine or D-lysine or an azido-lysine, i.e., 6-azido-L-lysine or 6-azido-D-lysine. In an embodiment, the reactive amino acid is a 5-azido-L-ornithine or 5-azido-d-ornithine. In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing an RGD peptide motif consisting of the three amino acids Arg-Gly-Asp. In another embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Asp-Cys (SEQ ID NO: 1). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Asp-Cys (SEQ ID NO: 2). In some embodiments, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Glu-Cys (SEQ ID NO: 3). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Glu-Cys (SEQ ID NO: 4). In another embodiment, the cyclic peptide sequence containing the cyclic iRGD peptide motif is cyclized via a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In some embodiments, the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing a NGR motif consisting of the three amino acids Asn-Gly-Arg. In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a NGR motif consisting of the five amino acids Cys-Asn-Gly-Arg-Cys (SEQ ID NO: 5). In an embodiment, the cyclic peptide sequence containing the NGR motif is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 5. In another embodiment, the tumor homing peptide comprises a cyclic peptide sequence consisting of the nine amino acid sequence Cys-Gly-Asn-Lys-Arg-Thr-Arg-Gly-Cys (SEQ ID NO: 6). In an embodiment, the cyclic peptide is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In another embodiment, the tumor homing peptide comprises a peptide sequence consisting of the 29 amino acids Lys-Asp-Glu-Pro-Gln-Arg-Arg-Ser-Ala-Arg-Leu-Ser-Ala-Lys-Pro-Ala-Pro-Pro-Lys-Pro-Glu-Pro-Lys-Lys-Ala-Pro-Ala-Lys-Lys (SEQ ID NO: 7). In some embodiments, the tumor homing peptide has a peptide sequence consisting of the amino acids Cys-Arg-Glu-Lys-Ala. In another embodiment, the tumor homing peptide comprises a peptide sequence consisting of the amino acids Cys-Arg-Glu-Lys-Ala (SEQ ID NO: 8). In an embodiment, the tumor homing peptide comprises a peptide sequence consisting of the amino acids Lys-Glu-Thr-Trp-Trp-Glu-Thr-Trp-Trp-Thr-Glu-Trp-Ser-Gln-Pro-Lys-Lys-Lys-Arg-Lys-Val (SEQ ID NO: 9).

In still another aspect, provided herein are multifunctional therapeutic peptide conjugate compounds comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein the compounds comprise two protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence [AIB]VPI[AIB]QK (SEQ ID NO: 12) from the N-terminus of the apoptosis mediator Smac/DIABLO protein, wherein [AIB] is 2-Aminoisobutyric acid. In an embodiment, the branched reactive amino acid is a lysine, a cysteine, an azido-lysine or an azido-ornithine. In another embodiment, the branched reactive amino acid is cysteine. In some embodiments, the branched reactive amino acid is azido-lysine. In an embodiment, the lysine is L-lysine or D-lysine or an azido-lysine, i.e., 6-azido-L-lysine or 6-azido-D-lysine. In an embodiment, the reactive amino acid is a 5-azido-L-ornithine or 5-azido-d-ornithine. In some embodiments, the branched reactive amino acid is azido-lysine. In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing an RGD peptide motif consisting of the three amino acids Arg-Gly-Asp. In another embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Asp-Cys (SEQ ID NO: 1). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Asp-Cys (SEQ ID NO: 2). In some embodiments, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Glu-Cys (SEQ ID NO: 3). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Glu-Cys (SEQ ID NO: 4). In another embodiment, the cyclic peptide sequence containing the cyclic iRGD peptide motif is cyclized via a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In some embodiments, the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing a NGR motif consisting of the three amino acids Asn-Gly-Arg. In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a NGR motif consisting of the five amino acids Cys-Asn-Gly-Arg-Cys (SEQ ID NO: 5). In an embodiment, the cyclic peptide sequence containing the NGR motif is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 5. In another embodiment, the tumor homing peptide comprises a cyclic peptide sequence consisting of the nine amino acid sequence Cys-Gly-Asn-Lys-Arg-Thr-Arg-Gly-Cys (SEQ ID NO: 6). In an embodiment, the cyclic peptide is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In another embodiment, the tumor homing peptide comprises a peptide sequence consisting of the 29 amino acids Lys-Asp-Glu-Pro-Gln-Arg-Arg-Ser-Ala-Arg-Leu-Ser-Ala-Lys-Pro-Ala-Pro-Pro-Lys-Pro-Glu-Pro-Lys-Lys-Ala-Pro-Ala-Lys-Lys (SEQ ID NO: 7). In some embodiments, the tumor homing peptide has a peptide sequence consisting of the amino acids Cys-Arg-Glu-Lys-Ala (SEQ ID NO: 8). In another embodiment, the tumor homing peptide comprises a peptide sequence consisting of the amino acids Cys-Arg-Glu-Lys-Ala (SEQ ID NO: 8). In an embodiment, the tumor homing peptide comprises a peptide sequence consisting of the amino acids Lys-Glu-Thr-Trp-Trp-Glu-Thr-Trp-Trp-Thr-Glu-Trp-Ser-Gln-Pro-Lys-Lys-Lys-Arg-Lys-Val (SEQ ID NO: 9).

In an embodiment of the herein described multifunctional therapeutic peptide conjugates comprises a tumor homing peptide, at least one protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the at least one protein binding moiety is conjugated to a second linker and the first and second linkers are conjugated to the branched reactive amino acid, wherein the at least one protein binding moiety is an inhibitor of apoptosis (IAP). In certain embodiments, the at least one protein binding moiety consists of a seven amino acid sequence AVPIAQK (SEQ ID NO: 10) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In some embodiments, the at least one protein binding moiety consists of a seven amino acid sequence 2-Aminoisobutyric acid [AIB]VPIAQK (SEQ ID NO: 11) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In an embodiment, the at least one protein binding moiety consists of a seven amino acid sequence [AIB]VPI[AIB]QK (SEQ ID NO: 12) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In a particular embodiment, the multifunctional therapeutic peptide conjugate comprises two protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence AVPIAQK from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In various embodiments, the multifunctional therapeutic peptide conjugate comprises two protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence 2-Aminoisobutyric acid [AIB]VPIAQK (SEQ ID NO: 11) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In an embodiment, the multifunctional therapeutic peptide conjugate comprises two protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence [AIB]VPI[AIB]QK (SEQ ID NO: 12) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In another embodiment, the multifunctional therapeutic peptide conjugate comprises three protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence AVPIAQK (SEQ ID NO: 10) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In some embodiments, the multifunctional therapeutic peptide conjugate comprises three protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence 2-Aminoisobutyric acid [AIB]VPIAQK (SEQ ID NO: 11) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In certain embodiments, the multifunctional therapeutic peptide conjugate comprises three protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence [AIB]VPI[AIB]QK (SEQ ID NO: 12) from the N-terminus of the apoptosis mediator Smac/DIABLO protein.

In some embodiments of the herein provided multifunctional therapeutic peptide conjugate compounds, each of the first linker and the second linker is an identical linker. In certain embodiments of the herein provided multifunctional therapeutic peptide conjugate compounds each of the first linker and the second linker is a different linker. In some embodiments of the first linker and the second linker, the first linker and the second linker is polyethylene glycol (PEG). In an embodiment, wherein the first linker and the second linker is an identical polyethylene glycol (PEG) linker or a different PEG linker, the PEG is selected from the group consisting of amino-PEG2, amino-PEG3, amino-PEG4, amino-PEG6, amino-PEG8, amino-PEG10, amino-PEG12 and amino-PEG20.

In still another aspect, provided herein are multifunctional therapeutic peptide conjugates represented by formula (I):

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In one aspect, provided herein are multifunctional therapeutic peptide conjugates represented by formula (II):

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In one aspect, provided herein are multifunctional therapeutic peptide conjugates represented by formula (III):

text missing or illegible when filed

In another aspect, provided herein are methods for treating cancer comprising administering to a subject in need thereof a therapeutically effective amount of a multifunctional therapeutic peptide conjugate compound comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing an RGD peptide motif consisting of the three amino acids Arg-Gly-Asp.

In still another aspect, provided herein are methods for treating cancer comprising administering to a subject in need thereof a therapeutically effective amount of a multifunctional therapeutic peptide conjugate compound comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Glu-Cys (SEQ ID NO: 3).

In another aspect, provided herein are methods for treating cancer comprising administering to a subject in need thereof a therapeutically effective amount of a multifunctional therapeutic peptide conjugate compound comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a NGR motif consisting of the five amino acids Cys-Asn-Gly-Arg-Cys (SEQ ID NO: 5).

In still another aspect, provided herein are methods for treating cancer comprising administering to a subject in need thereof a therapeutically effective amount of a multifunctional therapeutic peptide conjugate compound comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein tumor homing peptide comprises a cyclic peptide sequence consisting of the nine amino acid sequence Cys-Gly-Asn-Lys-Arg-Thr-Arg-Gly-Cys (SEQ ID NO: 6).

In another aspect, provided herein are methods for treating cancer comprising administering to a subject in need thereof a therapeutically effective amount of a multifunctional therapeutic peptide conjugate compound comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein the compounds comprise one protein binding moiety consisting of a seven amino acid sequence AVPIAQK (SEQ ID NO: 10) from the N-terminus of the apoptosis mediator Smac/DIABLO protein.

In another aspect, provided herein are methods for treating cancer comprising administering to a subject in need thereof a therapeutically effective amount of a multifunctional therapeutic peptide conjugate compound comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein the compounds comprise two protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence [AIB]VPIAQK (SEQ ID NO: 11) from the N-terminus of the apoptosis mediator Smac/DIABLO protein, wherein [AIB] is 2-Aminoisobutyric acid.

In still another aspect, provided herein are methods for treating cancer comprising administering to a subject in need thereof a therapeutically effective amount of a multifunctional therapeutic peptide conjugate compound comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein the compounds comprise two protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence [AIB]VPI[AIB]QK (SEQ ID NO: 12) from the N-terminus of the apoptosis mediator Smac/DIABLO protein, wherein [AIB] is 2-Aminoisobutyric acid.

In embodiments of the herein provided methods for treating cancer, the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing an RGD peptide motif consisting of the three amino acids Arg-Gly-Asp. In another embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Asp-Cys (SEQ ID NO: 1). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Asp-Cys (SEQ ID NO: 2). In some embodiments, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Glu-Cys (SEQ ID NO: 3). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Glu-Cys (SEQ ID NO: 4). In another embodiment, the cyclic peptide sequence containing the cyclic iRGD peptide motif is cyclized via a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In some embodiments, the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing a NGR motif consisting of the three amino acids Asn-Gly-Arg. In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a NGR motif consisting of the five amino acids Cys-Asn-Gly-Arg-Cys (SEQ ID NO: 5). In an embodiment, the cyclic peptide sequence containing the NGR motif is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 5. In another embodiment, the tumor homing peptide comprises a cyclic peptide sequence consisting of the nine amino acid sequence Cys-Gly-Asn-Lys-Arg-Thr-Arg-Gly-Cys (SEQ ID NO: 6). In an embodiment, the cyclic peptide is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In another embodiment, the tumor homing peptide comprises a peptide sequence consisting of the 29 amino acids Lys-Asp-Glu-Pro-Gln-Arg-Arg-Ser-Ala-Arg-Leu-Ser-Ala-Lys-Pro-Ala-Pro-Pro-Lys-Pro-Glu-Pro-Lys-Lys-Ala-Pro-Ala-Lys-Lys (SEQ ID NO: 7). In some embodiments, the tumor homing peptide comprises a peptide sequence consisting of the amino acids Cys-Arg-Glu-Lys-Ala (SEQ ID NO: 8). In another embodiment, the tumor homing peptide consists of the amino acids Cys-Arg-Glu-Lys-Ala. In an embodiment, the tumor homing peptide comprises a peptide sequence consisting of the amino acids Lys-Glu-Thr-Trp-Trp-Glu-Thr-Trp-Trp-Thr-Glu-Trp-Ser-Gln-Pro-Lys-Lys-Lys-Arg-Lys-Val (SEQ ID NO: 9).

Unless otherwise defined herein, scientific and technical terms used in connection with the present application shall have the meanings that are commonly understood by those of ordinary skill in the art. Further, unless otherwise required by context, singular terms shall include pluralities and plural terms shall include the singular.

Unless defined otherwise, all technical and scientific terms used herein have the same meaning as commonly understood to one of ordinary skill in the art.

As employed above and throughout the disclosure, the following terms and abbreviations, unless otherwise indicated, shall be understood to have the following meanings.

In the present disclosure the singular forms “a,” “an,” and “the” include the plural reference, and reference to a particular numerical value includes at least that particular value, unless the context clearly indicates otherwise. Thus, for example, a reference to “a compound” is a reference to one or more of such compounds and equivalents thereof known to those skilled in the art, and so forth. The term “plurality”, as used herein, means more than one. When a range of values is expressed, another embodiment incudes from the one particular and/or to the other particular value. Similarly, when values are expressed as approximations, by use of the antecedent “about,” it is understood that the particular value forms another embodiment. All ranges are inclusive and combinable.

As used herein, the terms “component,” “composition,” “composition of compounds,” “compound,” “drug,” “pharmacologically active agent,” “active agent,” “therapeutic,” “therapy,” “treatment,” or “medicament” are used interchangeably herein to refer to a compound or compounds or composition of matter which, when administered to a subject (human or animal) induces a desired pharmacological and/or physiologic effect by local and/or systemic action.

As used herein, the terms “treatment” or “therapy” (as well as different forms thereof) include preventative (e.g., prophylactic), curative or palliative treatment. As used herein, the term “treating” includes alleviating or reducing at least one adverse or negative effect or symptom of a condition, disease or disorder.

The terms “subject,” “individual,” and “patient” are used interchangeably herein, and refer to an animal, for example a human, to whom treatment for one or more cancer, including prophylactic treatment, with the pharmaceutical compositions according to the present invention, i.e., the herein described multifunctional branched therapeutic agents, respectively, is provided. The term “subject” as used herein refers to human and non-human animals. The terms “non-human animals” and “non-human mammals” are used interchangeably herein and include all vertebrates, e.g., mammals, such as non-human primates, (particularly higher primates), sheep, dog, rodent, (e.g., mouse or rat), guinea pig, goat, pig, cat, rabbits, cows, horses and non-mammals such as reptiles, amphibians, chickens, and turkeys.

Tumor Targeting Proteins

In an embodiment of the multifunctional therapeutic peptide conjugate, the tumor homing peptide is the cyclic peptide iRGD having amino acid sequence CRGD[K/R]GP[D/E]C (SEQ ID NO: 1 and SEQ ID NO:4). In another embodiment, the tumor homing peptide is the cyclic peptide iRGD having amino acid sequence Cys-Asn-Gly-Arg-Cys (SEQ ID NO: 5). In another embodiment, the tumor homing peptide is the cyclic peptide iRGD having amino acid sequence Cys-Gly-Asn-Lys-Arg-Thr-Arg-Gly-Cys (SEQ ID NO: 6). In some embodiments, the tumor homing peptide is the cyclic peptide iRGD having amino acid sequence Lys-Asp-Glu-Pro-Gln-Arg-Arg-Ser-Ala-Arg-Leu-Ser-Ala-Lys-Pro-Ala-Pro-Pro-Lys-Pro-Glu-Pro-Lys-Lys-Ala-Pro-Ala-Lys-Lys (SEQ ID NO: 7). In an embodiment, the tumor homing peptide is the cyclic peptide iRGD having amino acid sequence Cys-Arg-Glu-Lys-Ala (SEQ ID NO: 8). In another embodiment, the tumor homing peptide is the cyclic peptide iRGD having amino acid sequence Lys-Glu-Thr-Trp-Trp-Glu-Thr-Trp-Trp-Thr-Glu-Trp-Ser-Gln-Pro-Lys-Lys-Lys-Arg-Lys-Val (SEQ ID NO: 9).

In some embodiments, the multifunctional therapeutic peptide conjugate comprises one protein binding moiety consisting of a seven amino acid sequence AVPIAQK (SEQ ID NO: 10) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In an embodiment, the multifunctional therapeutic peptide conjugate comprises one protein binding moiety consisting of a seven amino acid sequence [AIB]VPIAQK (SEQ ID NO: 11), wherein [AIB] is 2-Aminoisobutyric acid. In another embodiment, the multifunctional therapeutic peptide conjugate comprises one protein binding moiety consisting of a seven amino acid sequence [AIB]VPI[AIB]QK (SEQ ID NO: 12), wherein [AIB] is 2-Aminoisobutyric acid. In some embodiments, the multifunctional therapeutic peptide conjugate comprises two protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence AVPIAQK (SEQ ID NO: 10) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In some embodiments, the first and second linker are the same. In certain embodiments, the first and second linker are different linker molecules.

As noted herein, the tumor-homing peptide known as iRGD protein. iRGD is cyclic peptide composed of 9-amino acids including an Arg-Gly-Asp (RGD) motif, has a high binding affinity to αvβ3 and αvβ5 integrins abundant in tumor vasculatures. IRGD protein has the following chemical structure:

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In some embodiments, the tumor-homing peptide compound is an iRGD protein analog or derivative thereof. In an embodiment, the iRGD protein analog or derivative thereof comprises RGD peptide motif consisting of the three amino acids Arg-Gly-Asp. In another embodiment, the iRGD protein analog or derivative thereof is RGD peptide motif consisting of the three amino acids Arg-Gly-Asp. In an embodiment, the iRGD protein analog or derivative thereof comprises iRGD motif consisting of the nine amino acid sequence Cys-Arg-Gly-Asp-Lys-Gly-Pro-Asp-Cys (SEQ ID NO: 1). In another embodiment, the iRGD protein analog or derivative thereof is iRGD motif consisting of the nine amino acid sequence Cys-Arg-Gly-Asp-Lys-Gly-Pro-Asp-Cys (SEQ ID NO: 1). In some embodiments, the iRGD protein analog or derivative thereof comprises the iRGD motif consisting of the nine amino acid sequence Cys-Arg-Gly-Asp-Arg-Gly-Pro-Asp-Cys (SEQ ID NO: 2). In some embodiments, the iRGD protein analog or derivative thereof is the iRGD motif consisting of the nine amino acid sequence Cys-Arg-Gly-Asp-Arg-Gly-Pro-Asp-Cys (SEQ ID NO: 2). In an embodiment, the iRGD protein analog or derivative thereof comprises the iRGD motif consisting of the nine amino acid sequence Cys-Arg-Gly-Asp-Lys-Gly-Pro-Glu-Cys (SEQ ID NO: 3). In another embodiment, the iRGD protein analog or derivative thereof is the iRGD motif consisting of the nine amino acid sequence Cys-Arg-Gly-Asp-Lys-Gly-Pro-Glu-Cys (SEQ ID NO: 3). In another embodiment, the iRGD protein analog or derivative thereof comprises the iRGD motif consisting of the nine amino acid sequence Cys-Arg-Gly-Asp-Arg-Gly-Pro-Glu-Cys (SEQ ID NO: 4). In another embodiment, the iRGD protein analog or derivative thereof is iRGD motif consisting of the nine amino acid sequence Cys-Arg-Gly-Asp-Arg-Gly-Pro-Glu-Cys (SEQ ID NO: 4).

In an embodiment, the tumor-homing peptide compound comprises the NGR motif consisting of the three amino acids Asn-Gly-Arg. In another embodiment, the tumor-homing peptide compound is the NGR motif consisting of the three amino acids Asn-Gly-Arg. In an embodiment, the tumor-homing peptide compound comprises an NGR motif consisting of the five amino acids Cys-Asn-Gly-Arg-Cys (SEQ ID NO: 5). In another embodiment, the tumor-homing peptide compound is an NGR motif consisting of the five amino acids Cys-Asn-Gly-Arg-Cys (SEQ ID NO: 5).

In an embodiment, the tumor-homing peptide compound comprises a peptide sequence containing the cyclic peptide consisting of the nine amino acid sequence Cys-Gly-Asn-Lys-Arg-Thr-Arg-Gly-Cys (SEQ ID NO: 6). In another embodiment, the tumor-homing peptide compound is a peptide sequence containing the cyclic peptide consisting of the nine amino acid sequence Cys-Gly-Asn-Lys-Arg-Thr-Arg-Gly-Cys (SEQ ID NO: 6).

In an embodiment, the tumor-homing peptide compound comprises a peptide sequence consisting of the 29 amino acids Lys-Asp-Glu-Pro-Gln-Arg-Arg-Ser-Ala-Arg-Leu-Ser-Ala-Lys-Pro-Ala-Pro-Pro-Lys-Pro-Glu-Pro-Lys-Lys-Ala-Pro-Ala-Lys-Lys (SEQ ID NO: 7). In another embodiment, the tumor-homing peptide compound is an peptide sequence consisting of the 29 amino acids Lys-Asp-Glu-Pro-Gln-Arg-Arg-Ser-Ala-Arg-Leu-Ser-Ala-Lys-Pro-Ala-Pro-Pro-Lys-Pro-Glu-Pro-Lys-Lys-Ala-Pro-Ala-Lys-Lys (SEQ ID NO: 7).

In an embodiment, the tumor-homing peptide compound comprises a peptide sequence consisting of the five amino acids Cys-Arg-Glu-Lys-Ala (SEQ ID NO: 8). In another embodiment, the tumor-homing peptide compound is a peptide sequence consisting of the five amino acids Cys-Arg-Glu-Lys-Ala (SEQ ID NO: 8).

In an embodiment, the tumor-homing peptide compound comprises a peptide sequence consisting of the 21 amino acids Lys-Glu-Thr-Trp-Trp-Glu-Thr-Trp-Trp-Thr-Glu-Trp-Ser-Gln-Pro-Lys-Lys-Lys-Arg-Lys-Val (SEQ ID NO: 9). In another embodiment, the tumor-homing peptide compound is a peptide sequence consisting of the 21 amino acids Lys-Glu-Thr-Trp-Trp-Glu-Thr-Trp-Trp-Thr-Glu-Trp-Ser-Gln-Pro-Lys-Lys-Lys-Arg-Lys-Val (SEQ ID NO: 9).

In an embodiment, the tumor-homing peptide compound comprises a peptide sequence consisting of the amino acids Lys-Glu-Thr-Trp-Trp-Glu-Thr-Trp-Trp-Thr-Glu-Trp-Ser-Gln-Pro-Lys-Lys-Lys-Arg-Lys-Val-cysteamine (SEQ ID NO: 9 with a cystamine). In another embodiment, the tumor-homing peptide compound is a peptide sequence consisting of the amino acids Lys-Glu-Thr-Trp-Trp-Glu-Thr-Trp-Trp-Thr-Glu-Trp-Ser-Gln-Pro-Lys-Lys-Lys-Arg-Lys-Val-cysteamine (SEQ ID NO: 9 with a cystamine).

Combinations of substituents envisioned by this invention are preferably those that result in the formation of stable or chemically feasible compounds. The term “stable”, as used herein, refers to compounds that are not substantially altered when subjected to conditions to allow for their production, detection, and preferably their recovery, purification, and use for one or more of the purposes disclosed herein. In some embodiments, a stable compound or chemically feasible compound is one that is not substantially altered when kept at a temperature of 40° C. or less, in the absence of moisture or other chemically reactive conditions, for at least a week. In each of the foregoing and each of the following embodiments, it is to be understood that the formulas also include any and all hydrates and/or solvates of the compound formulas. It is appreciated that certain functional groups, such as the hydroxy, amino, and like groups form complexes and/or coordination compounds with water and/or various solvents, in the various physical forms of the compounds. Accordingly, the above formulas are to be understood to include and represent those various hydrates and/or solvates.

As used herein, the term “solvate” refers to compounds that further include a stoichiometric or non-stoichiometric amount of a solvent bound by non-covalent intermolecular forces. If the solvent is water, the solvate is referred to as “hydrate.” Pharmaceutically acceptable solvates and hydrates are complexes that, for example, may include from 1 to about 100, or from 1 to about 10, or from one to about 2.3 or 4 molecules of water or a solvent. In some embodiments, the hydrate may be a channel hydrate. It should be understood that the term “compound” in this application covers the compound and solvates of the compound, as well as mixtures thereof.

In some embodiments, the phrase “pharmaceutically acceptable” is employed herein to refer to those compounds, materials, compositions, and/or dosage forms which are, within the scope of sound medical judgment, suitable for use in contact with the tissues of human beings and animals without excessive toxicity, irritation, allergic response, or other problem or complication, commensurate with a reasonable benefit/risk ratio.

The present invention also includes “pharmaceutically acceptable salts” of the compounds described herein. As used herein, “pharmaceutically acceptable salts” refers to derivatives of the disclosed compounds wherein the parent compound is modified by converting an existing acid or base moiety to its salt form. Examples of pharmaceutically acceptable salts include, but are not limited to, mineral or organic acid salts of basic residues such as amines; alkali or organic salts of acidic residues such as carboxylic acids; and the like. The pharmaceutically acceptable salts of the compound of the invention include the conventional non-toxic salts of the parent compound formed, for example, from non-toxic inorganic or organic acids. The pharmaceutically acceptable salts of the compound of the invention can be synthesized from the parent compound which contains a basic or acidic moiety by conventional chemical methods. Generally, such salts can be prepared by reacting the free acid or base forms of these compounds with a stoichiometric amount of the appropriate base or acid in water or in an organic solvent, or in a mixture of the two. In some embodiments, the solvent is a nonaqueous media like ether, ethyl acetate, ethanol, isopropanol, or acetonitrile.

The pharmaceutically acceptable salts of the compound of the invention can be formed by conventional means, such as by reacting the free base or free acid form of the product with one or more equivalents of the appropriate acid or base in a solvent or medium in which the salt is insoluble or in a solvent such as water, which is removed in vacuo or by freeze drying or by exchanging the ions of an existing salt for another ion or suitable ion-exchange resin.

Possible pharmaceutically acceptable salts are well known in the art. For example, S. M. Berge, et al. describe pharmaceutically acceptable salts in detail in J. Pharmaceutical Sciences, 66:1-19, 1977; which is incorporated herein by reference in its entirety.

The invention further includes analogs of the compound of the invention. The term “derivatives” includes but is not limited to ether derivatives, acid derivatives, amide derivatives, ester derivatives and the like.

The invention further includes pharmaceutical products of the compound of the invention. The term “pharmaceutical product” means a composition suitable for pharmaceutical use (pharmaceutical composition), as defined herein.

Multifunctional Therapeutic Agent

In an embodiment of the above-described multifunctional therapeutic peptide conjugate compounds comprising the cyclic peptide sequence consisting of the nine amino acid sequence Cys-Gly-Asn-Lys-Arg-Thr-Arg-Gly-Cys (SEQ ID NO: 6), the cyclic peptide is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In some embodiments of said multifunctional therapeutic peptide conjugate compounds, the active cellular targeting moiety targets the cellular receptor p32/gC1qR.

In another aspect, provided herein are multifunctional therapeutic peptide conjugate compounds comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein the compounds comprise one protein binding moiety consisting of a seven amino acid sequence AVPIAQK (SEQ ID NO: 10) from the N-terminus of the apoptosis mediator Smac/DIABLO protein. In an embodiment, the branched reactive amino acid is a lysine, a cysteine, an azido-lysine or an azido-ornithine. In a particular embodiment, the branched reactive amino acid is lysine. In another embodiment, the branched reactive amino acid is cysteine. In some embodiments, the branched reactive amino acid is azido-lysine. In an embodiment, the lysine is L-lysine or D-lysine or an azido-lysine, i.e., 6-azido-L-lysine or 6-azido-D-lysine. In an embodiment, the reactive amino acid is a 5-azido-L-ornithine or 5-azido-d-ornithine. In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing an RGD peptide motif consisting of the three amino acids Arg-Gly-Asp. In another embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Asp-Cys (SEQ ID NO: 1). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Asp-Cys (SEQ ID NO: 2). In some embodiments, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Glu-Cys (SEQ ID NO: 3). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Glu-Cys (SEQ ID NO: 4). In another embodiment, the cyclic peptide sequence containing the cyclic iRGD peptide motif is cyclized via a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In some embodiments, the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing a NGR motif consisting of the three amino acids Asn-Gly-Arg. In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a NGR motif consisting of the five amino acids Cys-Asn-Gly-Arg-Cys (SEQ ID NO: 5). In an embodiment, the cyclic peptide sequence containing the NGR motif is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 5. In another embodiment, the tumor homing peptide comprises a cyclic peptide sequence consisting of the nine amino acid sequence Cys-Gly-Asn-Lys-Arg-Thr-Arg-Gly-Cys (SEQ ID NO: 6). In an embodiment, the cyclic peptide is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In another embodiment, the tumor homing peptide comprises a peptide sequence consisting of the 29 amino acids Lys-Asp-Glu-Pro-Gln-Arg-Arg-Ser-Ala-Arg-Leu-Ser-Ala-Lys-Pro-Ala-Pro-Pro-Lys-Pro-Glu-Pro-Lys-Lys-Ala-Pro-Ala-Lys-Lys (SEQ ID NO: 7). In some embodiments, the tumor homing peptide has a peptide sequence consisting of the amino acids Cys-Arg-Glu-Lys-Ala (SEQ ID NO: 8). In another embodiment, the tumor homing peptide comprises a peptide sequence consisting of the amino acids Cys-Arg-Glu-Lys-Ala (SEQ ID NO: 8). In an embodiment, the tumor homing peptide comprises a peptide sequence consisting of the amino acids Lys-Glu-Thr-Trp-Trp-Glu-Thr-Trp-Trp-Thr-Glu-Trp-Ser-Gln-Pro-Lys-Lys-Lys-Arg-Lys-Val (SEQ ID NO: 9).

In one aspect, provided herein are multifunctional therapeutic peptide conjugate compounds comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein the compounds comprise one protein binding moiety consisting of a seven amino acid sequence consists of a seven amino acid sequence [AIB]VPIAQK (SEQ ID NO: 11) from the N-terminus of the apoptosis mediator Smac/DIABLO protein, wherein [AIB] is 2-Aminoisobutyric acid. In an embodiment, the branched reactive amino acid is a lysine, a cysteine, an azido-lysine or an azido-ornithine. In a particular embodiment, the branched reactive amino acid is lysine. In another embodiment, the branched reactive amino acid is cysteine. In some embodiments, the branched reactive amino acid is azido-lysine. In an embodiment, the lysine is L-lysine or D-lysine or an azido-lysine, i.e., 6-azido-L-lysine or 6-azido-D-lysine. In an embodiment, the reactive amino acid is a 5-azido-L-ornithine or 5-azido-d-ornithine. In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing an RGD peptide motif consisting of the three amino acids Arg-Gly-Asp. In another embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Asp-Cys (SEQ ID NO: 1). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Asp-Cys (SEQ ID NO: 2). In some embodiments, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Glu-Cys (SEQ ID NO: 3). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Glu-Cys (SEQ ID NO: 4). In another embodiment, the cyclic peptide sequence containing the cyclic iRGD peptide motif is cyclized via a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In some embodiments, the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing a NGR motif consisting of the three amino acids Asn-Gly-Arg. In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a NGR motif consisting of the five amino acids Cys-Asn-Gly-Arg-Cys (SEQ ID NO: 5). In an embodiment, the cyclic peptide sequence containing the NGR motif is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 5. In another embodiment, the tumor homing peptide comprises a cyclic peptide sequence consisting of the nine amino acid sequence Cys-Gly-Asn-Lys-Arg-Thr-Arg-Gly-Cys (SEQ ID NO: 6). In an embodiment, the cyclic peptide is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In another embodiment, the tumor homing peptide comprises a peptide sequence consisting of the 29 amino acids Lys-Asp-Glu-Pro-Gln-Arg-Arg-Ser-Ala-Arg-Leu-Ser-Ala-Lys-Pro-Ala-Pro-Pro-Lys-Pro-Glu-Pro-Lys-Lys-Ala-Pro-Ala-Lys-Lys (SEQ ID NO: 7). In some embodiments, the tumor homing peptide has a peptide sequence consisting of the amino acids Cys-Arg-Glu-Lys-Ala (SEQ ID NO: 8). In another embodiment, the tumor homing peptide comprises a peptide sequence consisting of the amino acids Cys-Arg-Glu-Lys-Ala (SEQ ID NO: 8). In an embodiment, the tumor homing peptide comprises a peptide sequence consisting of the amino acids Lys-Glu-Thr-Trp-Trp-Glu-Thr-Trp-Trp-Thr-Glu-Trp-Ser-Gln-Pro-Lys-Lys-Lys-Arg-Lys-Val (SEQ ID NO: 9). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing an RGD peptide motif consisting of the three amino acids Arg-Gly-Asp. In another embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Asp-Cys (SEQ ID NO: 1). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Asp-Cys (SEQ ID NO: 2). In some embodiments, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Glu-Cys (SEQ ID NO: 3). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Glu-Cys (SEQ ID NO: 4). In another embodiment, the cyclic peptide sequence containing the cyclic iRGD peptide motif is cyclized via a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In some embodiments, the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing a NGR motif consisting of the three amino acids Asn-Gly-Arg. In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a NGR motif consisting of the five amino acids Cys-Asn-Gly-Arg-Cys (SEQ ID NO: 5). In an embodiment, the cyclic peptide sequence containing the NGR motif is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 5. In another embodiment, the tumor homing peptide comprises a cyclic peptide sequence consisting of the nine amino acid sequence Cys-Gly-Asn-Lys-Arg-Thr-Arg-Gly-Cys (SEQ ID NO: 6). In an embodiment, the cyclic peptide is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In another embodiment, the tumor homing peptide comprises a peptide sequence consisting of the 29 amino acids Lys-Asp-Glu-Pro-Gln-Arg-Arg-Ser-Ala-Arg-Leu-Ser-Ala-Lys-Pro-Ala-Pro-Pro-Lys-Pro-Glu-Pro-Lys-Lys-Ala-Pro-Ala-Lys-Lys (SEQ ID NO: 7). In some embodiments, the tumor homing peptide has a peptide sequence consisting of the amino acids Cys-Arg-Glu-Lys-Ala (SEQ ID NO: 8). In another embodiment, the tumor homing peptide comprises a peptide sequence consisting of the amino acids Cys-Arg-Glu-Lys-Ala. In an embodiment, the tumor homing peptide comprises a peptide sequence consisting of the amino acids Lys-Glu-Thr-Trp-Trp-Glu-Thr-Trp-Trp-Thr-Glu-Trp-Ser-Gln-Pro-Lys-Lys-Lys-Arg-Lys-Val (SEQ ID NO: 9).

In another aspect, provided herein are multifunctional therapeutic peptide conjugate compounds comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein the compounds comprise one protein binding moiety consisting of a seven amino acid sequence [AIB]VPI[AIB]QK (SEQ ID NO: 11) from the N-terminus of the apoptosis mediator Smac/DIABLO protein, wherein [AIB] is 2-Aminoisobutyric acid. In an embodiment, the branched reactive amino acid is a lysine, a cysteine, an azido-lysine or an azido-ornithine. In a particular embodiment, the branched reactive amino acid is lysine. In another embodiment, the branched reactive amino acid is cysteine. In some embodiments, the branched reactive amino acid is azido-lysine. In some embodiments, the branched reactive amino acid is azido-lysine. In an embodiment, the lysine is L-lysine or D-lysine or an azido-lysine, i.e., 6-azido-L-lysine or 6-azido-D-lysine. In an embodiment, the reactive amino acid is a 5-azido-L-ornithine or 5-azido-d-ornithine. In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing an RGD peptide motif consisting of the three amino acids Arg-Gly-Asp. In another embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Asp-Cys (SEQ ID NO: 1). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Asp-Cys (SEQ ID NO: 2). In some embodiments, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Glu-Cys (SEQ ID NO: 3). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Glu-Cys (SEQ ID NO: 4). In another embodiment, the cyclic peptide sequence containing the cyclic iRGD peptide motif is cyclized via a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In some embodiments, the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing a NGR motif consisting of the three amino acids Asn-Gly-Arg. In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a NGR motif consisting of the five amino acids Cys-Asn-Gly-Arg-Cys (SEQ ID NO: 5). In an embodiment, the cyclic peptide sequence containing the NGR motif is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 5. In another embodiment, the tumor homing peptide comprises a cyclic peptide sequence consisting of the nine amino acid sequence Cys-Gly-Asn-Lys-Arg-Thr-Arg-Gly-Cys (SEQ ID NO: 6). In an embodiment, the cyclic peptide is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In another embodiment, the tumor homing peptide comprises a peptide sequence consisting of the 29 amino acids Lys-Asp-Glu-Pro-Gln-Arg-Arg-Ser-Ala-Arg-Leu-Ser-Ala-Lys-Pro-Ala-Pro-Pro-Lys-Pro-Glu-Pro-Lys-Lys-Ala-Pro-Ala-Lys-Lys (SEQ ID NO: 7). In some embodiments, the tumor homing peptide has a peptide sequence consisting of the amino acids Cys-Arg-Glu-Lys-Ala (SEQ ID NO: 8). In another embodiment, the tumor homing peptide comprises a peptide sequence consisting of the amino acids Cys-Arg-Glu-Lys-Ala. In an embodiment, the tumor homing peptide comprises a peptide sequence consisting of the amino acids Lys-Glu-Thr-Trp-Trp-Glu-Thr-Trp-Trp-Thr-Glu-Trp-Ser-Gln-Pro-Lys-Lys-Lys-Arg-Lys-Val (SEQ ID NO: 3).

In another aspect, provided herein are multifunctional therapeutic peptide conjugate compounds comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein the compounds comprise two protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence [AIB]VPIAQK (SEQ ID NO: 11) from the N-terminus of the apoptosis mediator Smac/DIABLO protein, wherein [AIB] is 2-Aminoisobutyric acid. In an embodiment, the branched reactive amino acid is a lysine, a cysteine, an azido-lysine or an azido-ornithine. In a particular embodiment, the branched reactive amino acid is lysine. In another embodiment, the branched reactive amino acid is cysteine. In some embodiments, the branched reactive amino acid is azido-lysine. In an embodiment, the lysine is L-lysine or D-lysine or an azido-lysine, i.e., 6-azido-L-lysine or 6-azido-D-lysine. In an embodiment, the reactive amino acid is a 5-azido-L-ornithine or 5-azido-d-ornithine. In an embodiment, the branched reactive amino acid is lysine, cysteine or azido-lysine. In a particular embodiment, the branched reactive amino acid is lysine. In another embodiment, the branched reactive amino acid is cysteine. In some embodiments, the branched reactive amino acid is azido-lysine. In an embodiment, the lysine is L-lysine or D-lysine or an azido-lysine, i.e., 6-azido-L-lysine or 6-azido-D-lysine. In an embodiment, the reactive amino acid is a 5-azido-L-ornithine or 5-azido-d-ornithine. In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing an RGD peptide motif consisting of the three amino acids Arg-Gly-Asp. In another embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Asp-Cys (SEQ ID NO: 1). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Asp-Cys (SEQ ID NO: 2). In some embodiments, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Glu-Cys (SEQ ID NO: 3). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Glu-Cys (SEQ ID NO: 4). In another embodiment, the cyclic peptide sequence containing the cyclic iRGD peptide motif is cyclized via a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In some embodiments, the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing a NGR motif consisting of the three amino acids Asn-Gly-Arg. In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a NGR motif consisting of the five amino acids Cys-Asn-Gly-Arg-Cys (SEQ ID NO: 5). In an embodiment, the cyclic peptide sequence containing the NGR motif is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 5. In another embodiment, the tumor homing peptide comprises a cyclic peptide sequence consisting of the nine amino acid sequence Cys-Gly-Asn-Lys-Arg-Thr-Arg-Gly-Cys (SEQ ID NO: 6). In an embodiment, the cyclic peptide is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In another embodiment, the tumor homing peptide comprises a peptide sequence consisting of the 29 amino acids Lys-Asp-Glu-Pro-Gln-Arg-Arg-Ser-Ala-Arg-Leu-Ser-Ala-Lys-Pro-Ala-Pro-Pro-Lys-Pro-Glu-Pro-Lys-Lys-Ala-Pro-Ala-Lys-Lys (SEQ ID NO: 7). In some embodiments, the tumor homing peptide has a peptide sequence consisting of the amino acids Cys-Arg-Glu-Lys-Ala. In another embodiment, the tumor homing peptide comprises a peptide sequence consisting of the amino acids Cys-Arg-Glu-Lys-Ala (SEQ ID NO: 8). In an embodiment, the tumor homing peptide comprises a peptide sequence consisting of the amino acids Lys-Glu-Thr-Trp-Trp-Glu-Thr-Trp-Trp-Thr-Glu-Trp-Ser-Gln-Pro-Lys-Lys-Lys-Arg-Lys-Val (SEQ ID NO: 9).

In still another aspect, provided herein are multifunctional therapeutic peptide conjugate compounds comprising a tumor homing peptide, a protein binding moiety and a branched reactive amino acid, wherein the tumor homing peptide is conjugated to a first linker, the protein binding moiety is conjugated to a second linker, and the first and second linkers are conjugated to the branched reactive amino acid, wherein the compounds comprise two protein binding moieties, wherein each protein binding moiety consists of a seven amino acid sequence [AIB]VPI[AIB]QK (SEQ ID NO: 12) from the N-terminus of the apoptosis mediator Smac/DIABLO protein, wherein [AIB] is 2-Aminoisobutyric acid. In an embodiment, the branched reactive amino acid is a lysine, a cysteine, an azido-lysine or an azido-ornithine. In a particular embodiment, the branched reactive amino acid is lysine. In another embodiment, the branched reactive amino acid is cysteine. In some embodiments, the branched reactive amino acid is azido-lysine. In an embodiment, the lysine is L-lysine or D-lysine or an azido-lysine, i.e., 6-azido-L-lysine or 6-azido-D-lysine. In an embodiment, the reactive amino acid is a 5-azido-L-ornithine or 5-azido-d-ornithine. In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing an RGD peptide motif consisting of the three amino acids Arg-Gly-Asp. In another embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Asp-Cys (SEQ ID NO: 1). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Asp-Cys (SEQ ID NO: 2). In some embodiments, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Lys-Gly-Pro-Glu-Cys (SEQ ID NO: 3). In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a cyclic iRGD peptide motif consisting of the nine amino acids Cys-Arg-Gly-Asp-Arg-Gly-Pro-Glu-Cys (SEQ ID NO: 4). In another embodiment, the cyclic peptide sequence containing the cyclic iRGD peptide motif is cyclized via a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In some embodiments, the tumor homing peptide comprises an active cellular targeting moiety (A) having a peptide sequence containing a NGR motif consisting of the three amino acids Asn-Gly-Arg. In an embodiment, the tumor homing peptide comprises an active cellular targeting moiety (A) having a cyclic peptide sequence containing a NGR motif consisting of the five amino acids Cys-Asn-Gly-Arg-Cys (SEQ ID NO: 5). In an embodiment, the cyclic peptide sequence containing the NGR motif is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 5. In another embodiment, the tumor homing peptide comprises a cyclic peptide sequence consisting of the nine amino acid sequence Cys-Gly-Asn-Lys-Arg-Thr-Arg-Gly-Cys (SEQ ID NO: 6). In an embodiment, the cyclic peptide is cyclized with a disulfide bond between cysteine at residue position 1 and cysteine at residue position 9. In another embodiment, the tumor homing peptide comprises a peptide sequence consisting of the 29 amino acids Lys-Asp-Glu-Pro-Gln-Arg-Arg-Ser-Ala-Arg-Leu-Ser-Ala-Lys-Pro-Ala-Pro-Pro-Lys-Pro-Glu-Pro-Lys-Lys-Ala-Pro-Ala-Lys-Lys (SEQ ID NO: 7). In some embodiments, the tumor homing peptide has a peptide sequence consisting of the amino acids Cys-Arg-Glu-Lys-Ala (SEQ ID NO: 8). In another embodiment, the tumor homing peptide comprises a peptide sequence consisting of the amino acids Cys-Arg-Glu-Lys-Ala (SEQ ID NO: 8). In an embodiment, the tumor homing peptide comprises a peptide sequence consisting of the amino acids Lys-Glu-Thr-Trp-Trp-Glu-Thr-Trp-Trp-Thr-Glu-Trp-Ser-Gln-Pro-Lys-Lys-Lys-Arg-Lys-Val (SEQ ID NO: 9).

In one aspect, the present the invention provides a multifunctional branched therapeutic agent represented by formula (I):

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In an embodiment of the herein provided multifunctional therapeutic peptide conjugate compounds, each of the first linker and the second linker is an identical linker. In certain embodiments of the herein provided multifunctional therapeutic peptide conjugate compounds each of the first linker and the second linker is a different linker. In some embodiments of the of the first linker and the second linker, the first linker and the second linker is polyethylene glycol (PEG). In an embodiment, wherein the first linker and the second linker is an identical polyethylene glycol (PEG) linker or a different PEG linker, the PEG is selected from the group consisting of amino-PEG2, amino-PEG3, amino-PEG4, amino-PEG6, amino-PEG8, amino-PEG10, amino-PEG12 and amino-PEG20.

Therapeutic Methods

In one aspect, the present the invention provides a method for treating cancer comprising administering to a subject in need thereof a therapeutically effective amount of any of the bifunctional and/or multifunctional therapeutic peptide conjugates described herein.

In a particular aspect, the present the invention provides a method for treating cancer comprising administering a to a subject in need thereof a therapeutically effective amount of an improved bifunctional therapeutic peptide conjugate comprising the N-terminus of the Smac/Diablo protein consisting of seven amino acid sequence AVPIAQK (SEQ ID NO: 10) and the iRGD peptide, wherein the N-terminus of the Smac/Diablo protein and the iRGD peptide are connected to a branched reactive amino acid by a molecular linker. In an embodiment of the provided methods, the branched reactive amino acid is lysine. In an embodiment, the molecular linker is polyethylene glycol (PEG).