Molecular guide system peptides and uses thereof

Abstract

To provide compositions and methods for delivering therapeutic antibodies to a specific cell type and to specific locations within that cell.SOLUTION: Disclosed are compositions comprising an antibody conjugated to one or more molecular guidance system (MGS) peptides. Disclosed are methods of treating a subject in need thereof comprising administering to the subject in need thereof an effective amount of an antibody conjugated to one or more MGS peptides, wherein the antibody targets an intracellular target involved in the disease process. Disclosed are methods of targeting an intracellular target, comprising administering an antibody conjugated to one or more MGS peptides, wherein the antibody targets an intracellular target.SELECTED DRAWING: None

Description

Technical Field

[0001] There is a pressing need for a systematic cell targeting system that delivers therapeutic antibodies to specific cell types and specific locations within cells.

Background Art

[0002] The therapeutic monoclonal antibody (MAb) market has grown dramatically over the past decade to exceed $70 billion and is expected to grow to $125 billion by 2020. Currently, 47 therapeutic MAbs that have received FDA approval for a wide range of diseases are on the market. However, MAbs do not have cell permeability and are therefore limited to extracellular and cell surface therapeutic targets. For this reason, many intracellular therapeutic targets, especially protein-protein interactions, are not the subject of MAb therapy. Except for that point, MAbs have a unique and powerful ability to regulate protein-protein interactions. Therefore, the fact that MAbs cannot penetrate cells and regulate intracellular protein-protein interactions is a major unsolved problem in the treatment of life-threatening diseases such as infectious diseases, cancer, and other difficult-to-treat and refractory diseases.

Summary of the Invention

[0003] MAbs have a unique and powerful ability to modulate protein-protein interactions. The inability of MAbs to penetrate cells and modulate intracellular protein-protein interactions represents a major unsolved problem in the treatment of life-threatening diseases. The ability to deliver therapeutic MAbs to specific target cells and the correct compartments within those cells would have many societal health benefits. The combination of MGS-MAbs has the potential to create a new class of safe and effective intracellular-acting drugs that can treat targets previously considered "undruggable". Disclosed herein are compositions and methods using molecular guidance system (MGS) peptides that provide new treatment strategies for newly emerging infectious diseases and bioterrorism, cancer, diabetes, neurological and neurodegenerative diseases, and even genetic diseases.

[0004] Compositions comprising an antibody conjugated to one or more molecular guidance system (MGS) peptides are disclosed. Compositions comprising an antibody conjugated to one or more MGS peptides, wherein the antibody targets an intracellular target, are disclosed.

[0005] Compositions comprising an antibody conjugated to one or more MGS peptides, wherein the antibody is a monoclonal antibody, are disclosed. In some embodiments, the monoclonal antibody is an anti-Ras monoclonal antibody.

[0006] Compositions comprising an antibody conjugated to one or more MGS peptides, wherein the antibody is a monoclonal antibody, are disclosed. In some embodiments, the monoclonal antibody is an anti-Ras monoclonal antibody.

[0007] A composition comprising an antibody bound to one or more MGS peptides, wherein the one or more MGS peptides have the sequence of EHPWFNMWSWATQVQE (SEQ ID NO: 38), YPGSPTQYPSSMHEYHSSSE (SEQ ID NO: 39), AHTIDDEWASYHMQQWNSPP (SEQ ID NO: 40), FEEFYSRQSNTIPYPQQYKG (SEQ ID NO: 41), THGNKHQSWTYPSEINHKNY (SEQ ID NO: 19), NLADTWTQTQQHDFHVLRGTR (SEQ ID NO: 20), GYSWWQPNWPSSTWDT (SEQ ID NO: 21), or a combination thereof. In some embodiments, the one or more MGS peptides have the sequence of EHPWFNMWSWATQVQE (SEQ ID NO: 38). A composition comprising an antibody bound to one or more MGS peptides A composition comprising an antibody bound to one or more MGS peptides, wherein the one or more MGS peptides comprise the sequence of SEQ ID NO: 1, 2, 3, 34, 35, 36, 37, 38, 39, 40, 41, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 5, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, or 77, or a combination thereof. A composition is disclosed.

[0008] A composition comprising an antibody bound to one or more MGS peptides, wherein the one or more MGS peptides are localized to one or more intracellular targets. A composition is disclosed.

[0009] A composition comprising an antibody bound to one or more MGS peptides, wherein the one or more MGS peptides are localized to one or more intracellular targets, and the intracellular targets are lysosomes, Golgi apparatus, endoplasmic reticulum, cytoplasm, or nucleus. A composition is disclosed.

[0010] A composition comprising an antibody bound to one or more MGS peptides, wherein the antibody is bound to the one or more MGS peptides via a linker. A composition is disclosed.

[0011] A composition comprising an antibody conjugated to one or more MGS peptides, wherein the antibody conjugated to one or more MGS peptides is a fusion protein, is disclosed.

[0012] A method of targeting an intracellular target, comprising administering an antibody conjugated to one or more MGS peptides, wherein the antibody targets the intracellular target, is disclosed. In some embodiments, the intracellular target is a lysosome, Golgi apparatus, endoplasmic reticulum, cytoplasm, or nucleus.

[0013] A method of treating a subject in need of treatment, comprising administering to the subject in need of treatment an effective amount of an antibody conjugated to one or more MGS peptides, wherein the antibody targets an intracellular target involved in a disease process, is disclosed. In some embodiments, the subject in need of treatment has an infectious disease, cancer, diabetes, a neurological or neurodegenerative disease, a genetic disease, or has been exposed to a bioterrorism agent.

[0014] Further advantages of the disclosed methods and compositions are described in part in the following description, understood in part from the description, or can be understood by practicing the disclosed methods and compositions. The advantages of the disclosed methods and compositions will be realized and achieved by the elements and combinations particularly pointed out in the appended claims. It is to be understood that both the foregoing general description and the following detailed description are exemplary and explanatory only and are not restrictive of the invention as claimed.

[0015] The accompanying drawings, which are incorporated herein and constitute a part of this specification, illustrate some embodiments of the disclosed methods and compositions and, together with the description, serve to explain the principles of the disclosed methods and compositions. BRIEF DESCRIPTION OF THE DRAWINGS

[0016]

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Mode for Carrying Out the Invention

[0017] The disclosed methods and compositions can be more easily understood by referring to the detailed description and examples of specific embodiments included in the following specification, as well as the drawings and their foregoing and subsequent descriptions.

[0018] The disclosed methods and compositions are not limited to a particular synthesis method, a particular analytical technique, or a particular reagent, unless otherwise specified, and can therefore vary widely. It should also be understood that the terms used herein are for the purpose of describing particular embodiments only and are not intended to be limiting.

[0019] All publications referred to herein are hereby incorporated by reference in their entirety for the purpose of disclosing and describing the manner and / or materials by which such publications are cited. The publications discussed herein are shown only as of their disclosure prior to the filing date of the present application. Nothing in this specification is to be construed as an admission that the present invention is not entitled to antedate such publications by virtue of prior invention. Further, the dates of publication of the publications shown herein may be different from the actual publication dates, which may require independent confirmation.

[0020] Disclosed are materials, compositions, and components that can be used in, combined with, used to prepare, or are products of the disclosed methods and compositions. These materials and other materials are disclosed herein, but when combinations, subsets, interactions, groups, etc. of these materials are disclosed , although specific references to each different individual, collective combinations and permutations of these compounds may not be explicitly disclosed, each is specifically contemplated and understood to be as described herein. For example, when conjugates are disclosed and considered, and many modifications that can be made to many molecules including MGS peptides are considered, every possible combination and permutation of conjugates and possible modifications is specifically contemplated, unless specifically indicated otherwise. Thus, if a group of molecules A, B, and C, and a group of molecules D, E, and F are disclosed, and an example of the combination molecule A-D is disclosed, each is contemplated individually and collectively, even if not individually described. Thus, in this example, each of the combinations A-E, A-F, B-D, B-E, B-F, C-D, C-E, and C-F is specifically contemplated and should be considered disclosed from the disclosure of A, B, and C, D, E, and F, and the exemplary combination A-D. Similarly, any subset or combination of these is specifically contemplated and disclosed. Thus, the subgroup of A-E, B-F, and C-E should be considered specifically contemplated and disclosed from the disclosure of A, B, and C, D, E, and F, and the exemplary combination A-D. This concept applies to all aspects of this application, including but not limited to each step in the methods of making and using the disclosed compositions. If there are various additional steps that can be performed, it should be understood that each of these additional steps can be performed by any particular embodiment or combination of embodiments of the disclosed method, and that each such combination is specifically contemplated and should be considered disclosed.

[0021] A. Definitions The disclosed methods and compositions are not limited to specific methods, protocols, and reagents, as the methods, protocols, and reagents can vary widely. It should be understood that the terms used herein are for the purpose of describing particular embodiments only and are not intended to limit the scope of the invention, which is limited only by the appended claims.

[0022] As used herein and in the appended claims, the singular forms "a," "an," and "the" include plural referents unless the context clearly dictates otherwise. Thus, for example, reference to "a MGS peptide" includes a plurality of such MGS peptides, and reference to "the MGS peptide" refers to one or more MGS peptides and equivalents thereof well known to those of ordinary skill in the art.

[0023] "Optional" or "optionally" means that the subsequently described event, circumstance, or material may or may not occur or exist, and that the description includes the case where such event, circumstance, or material occurs or exists and the case where it does not occur or exist.

[0024] As used herein, the term "subject" refers to any organism to which the compositions disclosed herein can be administered, for example, for experimental, diagnostic, and / or therapeutic purposes. Common subjects include animals (e.g., non-human primates and humans; birds; domestic and farm animals such as cats, dogs, sheep, goats, cows, horses, and pigs; laboratory animals such as mice, rats, and guinea pigs; rabbits; fish; reptiles; zoo and wild animals), and / or plants. Typically, a "subject" is an animal, including mammals such as humans and non-human primates.

[0025] As used herein, the term "patient" refers to a patient suffering from a disease or disorder. refers to. The term "patient" includes humans and veterinary subjects. In some aspects of the disclosed methods, the "patient" has, for example, been diagnosed as requiring treatment for an autoimmune disease prior to the administration step.

[0026] As used herein, the term "amino acid sequence" refers to a list of abbreviations, letters, alphanumerics, or words representing amino acid residues. The abbreviations for amino acids used herein are the conventional single-letter codes for amino acids and are represented as follows. That is, A (alanine), C (cysteine), D (aspartic acid), E (glutamic acid), F (phenylalanine), G (glycine), H (histidine), I (isoleucine), K (lysine), L (leucine), M (methionine), N (asparagine), P (proline), Q (glutamine), R (arginine), S (serine), T (threonine), V (valine), W (tryptophan), Y (tyrosine).

[0027] As used herein, "polypeptide" refers to any peptide, oligopeptide, polypeptide, gene product, expression product, or protein. A polypeptide is composed of consecutive amino acids. The term "polypeptide" encompasses naturally occurring or synthetic molecules.

[0028] Furthermore, as used herein, the term "polypeptide" refers to amino acids linked to each other by peptide bonds or modified peptide bonds (e.g., peptide mimics, etc.), and may include modified amino acids other than the 20 amino acids encoded by the genes. A polypeptide may be modified by natural processes such as post-translational processing or by chemical modification methods well known in the art. Modifications may occur anywhere in the polypeptide, including the peptide backbone, amino acid side chains, and the amino or carboxyl terminus. The same type of modification may be present in the same or different degrees at multiple sites on a particular peptide. Also, a particular polypeptide may have many types. Modifications include, but are not limited to, acetylation, acylation, ADP-ribosylation, amidation, covalent cross-linking or cyclization, covalent attachment of flavin, covalent attachment of a heme moiety, covalent attachment of a nucleotide or nucleotide derivative, covalent attachment of a lipid or lipid derivative, covalent attachment of phosphatidylinositol, formation of a disulfide bond, demethylation, formation of cysteine or pyroglutamate, formylation, γ-carboxylation, glycosylation, formation of a GPI anchor, hydroxylation, iodination, methylation, myristoylation, oxidation, PEGylation, proteolytic processing, phosphorylation, prenylation, racemization, selenoylation, sulfation, and tRNA-mediated addition of amino acids to proteins such as arginylation (see Proteins-Structure and Molecular Properties 2nd Ed., T.E. Creighton, W.H. Freeman and Company, New York (1993); Posttranslational Covalent Modification of Proteins, B.C. Johnson, Ed., Academic Press, New York, pp. 1-12 (1983)).

[0029] As used herein, the term "nucleic acid" refers to a naturally occurring or synthetic oligonucleotide or polynucleotide, whether DNA, RNA, or a DNA-RNA hybrid, single-stranded or double-stranded, sense or antisense, capable of hybridizing to a complementary nucleic acid by Watson-Crick base pairing. The nucleic acids of the invention can also include nucleotide analogs (e.g., BrdU) and non-phosphodiester nucleotide linkages (e.g., peptide nucleic acids (PNA) or thiophosphate linkages). Specifically, examples of nucleic acids include, but are not limited to, DNA, RNA, cDNA, gDNA, ssDNA, dsDNA, or any combination thereof.

[0030] As used herein, the term "sample" means an animal; a tissue or organ derived from an animal; a cell (from within the subject, directly obtained from the subject, or maintained in culture or a cell line-derived cell); a cell lysate (or fraction of a lysate) or cell extract; or a solution containing one or more molecules derived from a cell or cellular material (e.g., a polypeptide or nucleic acid), which is assayed as described herein. A sample can be any body fluid or excreted product containing cells or cell components (e.g., but not limited to, blood, urine, feces, saliva, tears, bile, etc.).

[0031] As used herein, "modulate" means to change by increasing or decreasing.

[0032] As used herein, an "effective amount" of a compound means an amount of the compound sufficient to provide the desired effect. The exact amount required will vary from subject to subject, depending on the species, age, and general condition of the subject, the severity of the disease (or underlying genetic defect) being treated, the particular compound being used, its mode of administration, and the like. Accordingly, an exact "effective amount" cannot be specified. However, an appropriate "effective amount" can be determined by one of ordinary skill in the art using only routine experimentation.

[0033] As used herein, an "isolated polypeptide" or "purified polypeptide" means a polypeptide (or fragment thereof) that is substantially free of substances that normally accompany it in nature. The polypeptides of the invention, or fragments thereof, can be obtained, for example, by extraction from a natural source (e.g., mammalian cells), by expression of a recombinant nucleic acid encoding the polypeptide (e.g., in a cell or cell-free translation system), or by chemically synthesizing the polypeptide. Further, polypeptide fragments can be obtained by any of these methods or by cleaving a full-length protein and / or polypeptide.

[0034] As used herein, "isolated nucleic acid" or "purified nucleic acid" means DNA that does not contain the genes adjacent to the gene in the naturally occurring genome of the organism from which the DNA of the present invention is derived. Thus, this term includes, for example, recombinant DNA incorporated into a vector such as a self-replicating plasmid or virus, or incorporated into the genomic DNA of a prokaryote or eukaryote (e.g., a transgene), or existing as a separate molecule (e.g., cDNA or genomic or cDNA fragments produced by PCR, restriction endonuclease digestion, or in vitro synthesis). This term also includes recombinant DNA that is part of a hybrid gene encoding an additional polypeptide sequence. The term "isolated nucleic acid" also refers to RNA, e.g., mRNA molecules that are encoded by an isolated DNA molecule, or chemically synthesized, or separated from or substantially free of at least certain cellular components, such as other types of RNA molecules or polypeptide molecules.

[0035] As used herein, "prevent" means to minimize the likelihood that a subject who is highly susceptible to developing cancer will develop cancer.

[0036] As used herein, "specifically binds" means that an antibody recognizes and physically interacts with its cognate antigen or target (e.g., the disclosed synthetic MGS sequence), but recognizes and interacts little with other antigens or targets, and such an antibody may be a polyclonal or monoclonal antibody produced by methods well known in the art.

[0037] As used herein, "probe", "primer", or oligonucleotide means a single-stranded DNA or RNA molecule of a defined sequence that can form base pairs with a second DNA or RNA molecule containing a complementary sequence (the "target"). The stability of the resulting hybrid is determined by the extent of base pair formation that occurs. The extent of base pair formation is affected by parameters such as the degree of complementarity between the probe and the target molecule and the stringency of the hybridization conditions. The degree of stringency of hybridization is affected by parameters such as temperature, salt concentration, and the concentration of organic molecules such as formamide, and is determined by methods well known to those skilled in the art. Probes or primers specific for nucleic acids (e.g., genes and / or mRNAs) capable of encoding the disclosed MGS sequences have at least 80% - 90% sequence complementarity, preferably at least 91% - 95% sequence complementarity, more preferably at least 96% - 99% sequence complementarity, and most preferably 100% sequence complementarity to the region of the nucleic acid capable of encoding the disclosed MGS sequences to which they hybridize. Probes, primers, and oligonucleotides can be labeled detectably, either radioactively or non-radioactively, by methods well known to those skilled in the art. Probes, primers, and oligonucleotides are used in methods involving nucleic acid hybridization such as nucleic acid sequencing, nucleic acid amplification by reverse transcription and / or polymerase chain reaction, single-strand conformational polymorphism (SSCP) analysis, restriction fragment length polymorphism (RFLP) analysis, Southern hybridization, Northern hybridization, in situ hybridization, electrophoretic mobility shift assay (EMSA), etc.

[0038] As used herein, "specifically hybridizes" means that a probe, primer, or oligonucleotide recognizes and physically interacts with (i.e., forms base pairs with) a substantially complementary nucleic acid (e.g., a nucleic acid capable of encoding the disclosed MGS sequences) under high stringency conditions, but forms few base pairs with other nucleic acids.

[0039] As used herein, the term "high stringency conditions" means conditions that allow hybridization equivalent to that obtained by the use of a DNA probe at least 40 nucleotides in length at a temperature of 65°C in a buffer containing 0.5 M NaHPO 4 , pH 7.2, 7% SDS, 1 mM EDTA, and 1% BSA (Fraction V), or at a temperature of 42°C in a buffer containing 48% formamide, 4.8 X SSC, 0.2 M Tris-Cl, pH 7.6, 1 X Denhardt's solution, 10% dextran sulfate, and 0.1% SDS. Other conditions for high stringency hybridization such as PCR, Northern, Southern, or in situ hybridization, DNA sequencing, etc. are well known to those of ordinary skill in the art of molecular biology (see, for example, F. Ausubel et al., Current Protocols in Molecular Biology, John Wiley & Sons, New York, NY, 1998).

[0040] As used herein, the term "therapeutically effective amount" means an amount of a therapeutic, prophylactic, and / or diagnostic agent (e.g., an MGS peptide conjugate conjugated to an antibody) sufficient to treat, alleviate, ameliorate, palliate, reduce the symptoms thereof, prevent, delay the onset thereof, arrest the progression thereof, reduce the severity thereof, and / or reduce the incidence thereof when administered to a subject afflicted with or susceptible to a disease, disorder, and / or condition.

[0041] As used herein, the term "treat" means to partially or completely alleviate, ameliorate, palliate, delay the onset of, arrest the progression of, reduce the severity of, and / or reduce the incidence of one or more symptoms or characteristics of a particular disease, disorder, and / or condition Refers to reducing. For example, "treating" a microbial infection can refer to preventing the survival, growth, and / or spread of the microorganism. Treatment can be administered to a subject that does not exhibit signs of a disease, disorder, and / or condition, and / or to a subject that exhibits only the initial signs of a disease, disorder, and / or condition, for the purpose of reducing the risk of developing the medical conditions associated with such disease, disorder, and / or condition. In some embodiments, treatment includes administering to a subject one or more of the disclosed compositions.

[0042] As used herein, the term "antibody" includes full-length antibodies and antigen-binding fragments of complete antibody molecules. As used herein, the terms "antigen-binding portion", "antigen-binding fragment" of an antibody, and like terms include any naturally occurring, enzymatically obtainable, synthetic, or genetically engineered polypeptide or glycoprotein that specifically binds to an antigen to form a complex. Antigen-binding fragments of antibodies can be obtained from complete antibody molecules using any suitable standard techniques, such as proteolytic digestion, or recombinant genetic engineering techniques that involve manipulation and expression of DNA encoding the variable and optionally constant domains of the antibody. Such DNA is well known and / or can be readily obtained, for example, from commercial suppliers, DNA libraries (including, for example, phage / antibody libraries), or can be synthesized. Such DNA can be sequenced and chemically manipulated, or, for example, using molecular biology techniques, one or more variable and / or constant domains can be arranged in an appropriate configuration, or codons can be introduced, cysteine residues can be formed, amino acids can be modified, added, or deleted, etc.

[0043] Non-limiting examples of antigen-binding fragments include: (i) Fab fragments, (ii) F(ab’)2 fragments, (iii) Fd fragments, (iv) Fv fragments, (v) single-chain Fv (scFv) molecules, (vi) dAb fragments, and (vii) the minimal recognition units composed of amino acid residues that mimic the hypervariable regions of antibodies (e.g., isolated complementarity-determining regions (CDRs) such as CDR3 peptides). Domain-specific antibodies, single-domain antibodies, domain-deleted antibodies, chimeric antibodies, CDR-grafted antibodies, diabodies, triabodies, tetra-bodies, mini-bodies, nanobodies (e.g., monovalent nanobodies, bivalent nanobodies, etc.), small molecule modular immunopharmaceuticals (SMIPs), and other engineered molecules such as the variable IgNAR domains of sharks are also included within the expression “antigen-binding fragment” as used herein.

[0044] The antibodies described herein can be recombinant, chimeric, or humanized antibodies. The term “chimeric antibody” refers to an antibody that contains one or more regions from one antibody and one or more regions from one or more other antibodies. A “humanized antibody” has a sequence that differs from the sequence of an antibody derived from a non-human species in one or more amino acid substitutions, deletions, and / or additions, such that a humanized antibody is less likely to induce an immune response and / or the induced immune response is less severe when administered to a human subject compared to an antibody from a non-human species.

[0045] In this specification, ranges are expressed as from "about" (a particular value) and / or to "about" (another particular value). When such a range is expressed, the range from a particular value and / or to another particular value is also specifically contemplated and considered to be disclosed, unless the context specifically indicates otherwise. Similarly, when a value is expressed as an approximation by use of the antecedent "about", that particular value should be considered to constitute another specifically contemplated embodiment that is disclosed, unless the context specifically indicates otherwise. Each endpoint of a range is effective both in relation to the other endpoint and independently of the other endpoint, unless the context specifically indicates otherwise. Finally, all individual values and sub-ranges of values that are included within a specifically disclosed range are also specifically contemplated and should be considered to be disclosed, unless the context specifically indicates otherwise. This applies regardless of whether some or all of these embodiments are specifically disclosed in a particular case. It will be understood that the foregoing applies regardless of whether some or all of these embodiments are specifically disclosed in a particular case.

[0046] Unless otherwise defined, all technical and scientific terms used herein shall have the same meaning as commonly understood by one of ordinary skill in the art to which the disclosed methods and compositions pertain. Any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present methods and compositions, but specific useful methods, devices, and materials are as described. Publications cited herein and the materials for which such publications are cited are hereby incorporated by reference in their entirety. Nothing in this specification should be construed as an admission that the present invention is not entitled to antedate such disclosure by virtue of prior invention. No admission is made that any reference constitutes prior art. The discussion of references states what their authors assert, and applicants reserve the right to challenge the accuracy and appropriateness of the cited references. Although many publications are referred to herein, it should be clearly understood that such reference does not constitute an admission that any of these documents forms part of the common general knowledge in the art.

[0047] Throughout the description and claims of this specification, the word “comprise” and variations of the word such as “comprising” and “comprises” are meant to mean “including but not limited to,” and are not intended to (and do not) exclude other additives, components, integers, or steps. Specifically, in a method described as including one or more steps or operations, each step is specifically contemplated to include what is listed (unless that step is preceded by a limiting term such as “consisting of”), which means that each step is not intended to (and does not) exclude, for example, other additives, components, integers, or steps that may not be listed during that step.

[0048] B. Compositions Disclosed are each of the components used to prepare the disclosed compositions, and the compositions themselves used in the methods disclosed herein. Although these materials and other materials are disclosed herein, there may be cases where specific references to the different individual and collective combinations and permutations of these materials, subsets, interactions, groups, etc. are not explicitly disclosed, but each is specifically contemplated and understood to be as described herein. Disclosed are compositions comprising an antibody conjugated to one or more MGS peptides. In some embodiments, the antibody targets an intracellular target.

[0049] Disclosed is a composition comprising an antibody conjugated to one or more MGS peptides, wherein the antibody is a monoclonal antibody. In some embodiments, the monoclonal antibody may be an anti-Ras monoclonal antibody.

[0050] MGS Peptide As used herein, MGS or targeting peptides are disclosed. These peptides can selectively bind to cells. Examples of MGS peptides that can be used or modified in the disclosed compositions include, but are not limited to, one or more of the MGS peptides disclosed in McGuire et al., Sci Rep. 2014 Mar 27;4:4480. Examples of MGS peptides that can also be used in the disclosed compositions and methods include, but are not limited to, those shown in Table 1 and Figures 9 and 10 MGS sequences. Table 1. Peptide Sequences

Table 1

[0051] SEQ ID NOs: 71-77 are optimized MGS peptides. The optimized peptides were obtained by making modifications to the individual parent peptide sequences identified by the FOX-Three platform technology. These modifications were used to identify the essential amino acids within the parent sequences required for cell-specific binding and internalization. These modifications were obtained by a combination of alanine scanning of the parent peptide and cleavage of the amino-terminal and C-terminal regions. PEG11 provides protection of the C-terminus of the MGS peptide, provides a spacer between the peptide and the cargo molecule conjugated via cysteine to the C-terminus, and increases the solubility of the MGS peptide. Modification of the amino-terminus with acetylation (CH3CO-) and / or d-amino acids, e.g., d(Leu) protection against degradation by peptidases in the blood. There is no uniform length of the optimized peptides applicable to all MGS peptides, and all changes need to be tested to confirm their effect on peptide uptake and stability. In peptide 73, it is possible to monitor peptide synthesis and concentration by absorbance of light at 280 nm using YC. Without the addition of tyrosine (Y), monitoring of this peptide is significantly more difficult.

[0052] CH 3 CO-YAAWPASGAWT-PEG 11 -C-NH 2 (SEQ ID NO: 71), CH 3 CO-LQWRRNFGVWARYRL-PEG 11 -C-NH 2 (SEQ ID NO: 72), CH 3 CO-d(Leu)-RGDLATLRQL-PEG 11 -YC-NH 2 (SEQ ID NO: 74) 、CH 3 CO-LQWRRNFGVWARYRL-PEG 11 -C-NH 2 (SEQ ID NO: 75), CH 3 CO-FHAVPQSFYT-PEG 11 -C-NH 2(SEQ ID NO: 76), or CH 3 CO-FHAVPQSFYT-PEG 11 -C-NH 2 Modified peptides having the sequence of (SEQ ID NO: 77) are disclosed.

[0053] All of the MGS peptides described in Table 1 are disclosed.

[0054] In one aspect, the composition comprises one or more of the MGS peptides and an antibody. In one aspect, the composition comprises one or more of the MGS peptides disclosed herein and an antibody. In one aspect, the membrane-permeable conjugate for transport through the lipid membrane can comprise one or more MGS peptides and an antibody.

[0055] In one aspect, the one or more MGS peptides can include any of the MGS peptides disclosed herein. In one aspect, the one or more MGS peptides include SEQ ID NO: 1, 2, 3, 34, 35, 36, 37, 38, 39, 40, 41, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 5, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 81, 81, 82, 83, 84, or combinations thereof.

[0056] In one aspect, one or more MGS peptides may include SEQ ID NO: 1, 2, 3, 34, 35, 36, 37, 38, 39, 40, 41, 41, 42, 43, 44, 45, 46, 47, 48, 49, 50, 51, 52, 5, 54, 55, 56, 57, 58, 59, 60, 61, 62, 63, 64, 65, 66, 67, 68, 69, 70, 71, 72, 73, 74, 75, 76, 77, 78, 79, 81, 81, 82, 83, or 84. In one aspect, one or more MGS peptides may be SEQ ID NO: 3. In one aspect, the composition may include one or more MGS peptides, and in some aspects, the composition can include one, two, three, four, or five MGS peptides. In one aspect, one or more MGS peptides can form a tetrameric scaffold protein. In one aspect, one or more MGS peptides can be shortened.

[0057] In one aspect, one or more MGS peptides may be modified. In one aspect, the N-terminus of one or more MGS peptides may be acetylated. In one aspect, one or more MGS peptides can be chemically conjugated to an antibody. In one aspect, the chemical conjugate may be polyethylene glycol (PEG). In one aspect, the number of PEG units may be 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, or more. In each aspect, the number of PEG units can be of a length sufficient to separate one or more MGS peptides from the antibody and prevent steric hindrance between the one or more MGS peptides and the antibody. For example, disclosed herein is a composition comprising a chemical conjugate, wherein the chemical conjugate is PEG and the PEG is composed of 11 PEG units. In one aspect, one or more MGS peptides include SEQ ID NO: 3, which can be acetylated at its N-terminus and chemically conjugated to PEG, and an antibody that can be covalently conjugated to PEG.

[0058] A composition comprising an antibody conjugated to one or more MGS peptides, wherein the one or more MGS peptides have the sequence of EHPWFNMWSWATQVQE (SEQ ID NO: 38), YPGSPTQYPSSMHEYHSSSE (SEQ ID NO: 39), AHTIDDEWASYHMQQWNSPP (SEQ ID NO: 40), FEEFYSRQSNTIPYPQQYKG (SEQ ID NO: 41), THGNKHQSWTYPSEINHKNY (SEQ ID NO: 19), NLADTWTQTQQHDFHVLRGTR (SEQ ID NO: 20), GYSWWQPNWPSSTWDT (SEQ ID NO: 21), or a combination thereof, is disclosed. In some embodiments, the one or more MGS peptides have the sequence of EHPWFNMWSWATQVQE (SEQ ID NO: 38).

[0059] In some embodiments of the disclosed composition, the one or more MGS peptides localize to one or more intracellular targets. For example, the intracellular target can be, but is not limited to, a lysosome, Golgi apparatus, endoplasmic reticulum, cytoplasm, or nucleus. Any subcellular compartment can be targeted. The antibody and the one or more MGS peptides can be conjugated by any of the methods by which proteins are commonly conjugated or linked to each other. In some embodiments, the antibody can be conjugated to the one or more MGS peptides via a linker. For example, in some cases, the linker can be a peptide linker or a nucleic acid linker. In some embodiments, the linker can be a cleavable linker. In some embodiments, the antibody conjugated to the one or more MGS peptides is a fusion protein. In other words, the antibody conjugated to the one or more MGS peptides can be a single long protein consisting of several different peptides, rather than being conjugated or linked to each other after being produced as separate peptides. In some embodiments, the MGS peptide can be conjugated to the antibody using any well-known conjugation method, including but not limited to SiteClick chemistry.

[0060]

[0061] In some embodiments of the disclosed compositions, the antibody can further include a label. For example, the compositions disclosed herein can include a detectable label. Such detectable labels can include, but are not limited to, tag sequences designed for the detection (e.g., purification or localization) of the expressed polypeptide or sequence. Examples of tag sequences include, but are not limited to, green fluorescent protein, glutathione S-transferase, polyhistidine, c-myc, hemagglutinin, or Flag™ tag, which can be fused to the encoded nucleic acid. Such detectable labels can include, but are not limited to, fluorescent substances, enzyme labels, or radioisotopes.

[0062] In some embodiments, the disclosed compositions can be pharmaceutical compositions. For example, in some embodiments, pharmaceutical compositions are disclosed that include a composition comprising an antibody bound to one or more MGS peptides and a pharmaceutically acceptable carrier. As is well known to those of skill in the art, "pharmaceutically acceptable" means a material or carrier that is selected to minimize degradation of the active ingredient and to minimize adverse side effects in the subject. Examples of carriers include dimyristoyl phosphatidyl (DMPC), phosphate buffered saline, or multilamellar liposomes. For example, PG:PC:cholesterol:peptide, or PC:peptide can be used as carriers in the present invention. Other suitable pharmaceutically acceptable carriers and their formulations are described in Remington: The Science and Practice of Pharmacy (19th ed.) ed. A.R. Gennaro, Mack It is described in Publishing Company, Easton, PA 1995. Generally, an appropriate amount of pharmaceutically acceptable salts is used in the formulation to make it isotonic. Other examples of pharmaceutically acceptable carriers include, but are not limited to, physiological saline, Ringer's solution, and dextrose solution. The pH of the solution can be from about 5 to about 8, or from about 7 to about 7.5. Further, sustained-release formulations such as semipermeable matrices of solid hydrophobic polymers containing the composition are included, and this matrix is in the form of a shaped article such as, for example, a film, a stent (implanted intravascularly in angioplasty), a liposome, or microparticles. It will be apparent to those skilled in the art that certain carriers may be more preferred depending on, for example, the route of administration and the concentration of the composition being administered. These particular carriers are most commonly standard carriers for administering drugs to humans, including solutions such as sterile water, saline, and buffered solutions at physiological pH.

[0063] The pharmaceutical composition may further contain a carrier, a thickening agent, a diluent, a buffer, a preservative, etc., as long as the polypeptide, peptide, or conjugate of the present invention is not impaired. The pharmaceutical composition may further contain one or more active ingredients (other than the composition of the present invention) such as an antibacterial agent, an anti-inflammatory agent, an anesthetic agent, etc.

[0064] The pharmaceutical compositions disclosed herein can be prepared for oral or parenteral administration. Pharmaceutical compositions prepared for parenteral administration include those prepared for intravenous (or intra - arterial), intramuscular, subcutaneous, intraperitoneal, transmucosal (e.g., intranasal, intravaginal, or rectal), or transdermal (e.g., topical) administration. Delivery of the fusion protein can also be achieved using aerosol inhalation. Thus, the composition can be prepared for parenteral administration containing the fusion protein dissolved or suspended in an acceptable carrier, including but not limited to aqueous carriers such as water, buffered water, physiological saline, buffered physiological saline (e.g., PBS). One or more excipients such as pH adjusters and buffers, tonicity adjusters, wetting agents, detergents, etc. can be useful in approximating physiological conditions. If the composition contains solid components (as in the case of oral administration), one or more of the excipients can function as binders or fillers (e.g., for formulation into tablets, capsules, etc.). If the composition is formulated for application to the skin or mucosal surface, one or more of the excipients can be used as solvents or emulsifiers for formulation into creams, ointments, etc.

[0065] The pharmaceutical composition can be made sterile and can be sterilized by conventional sterilization methods or by sterile filtration. Aqueous solutions can be packaged for use as is, or can be lyophilized, and the lyophilized formulations encompassed by this disclosure can be combined with a sterile aqueous carrier prior to administration. The pH of the pharmaceutical composition is typically from 3 to 11 (e.g., about 5 to 9), or 6 to 8 (e.g., about 7 to 8). The resulting composition in solid form can be packaged as a plurality of single - dose units, such as in sealed packages of tablets or capsules, each containing a defined amount of one or more of the above - mentioned agents. The composition in solid form can also be packaged in a flexible - dosing container, such as in a squeeze tube designed for creams or ointments for topical application.

[0066] C. Method A method of targeting an intracellular target, comprising administering a therapeutically effective amount of one or more of the disclosed compositions is disclosed.

[0067] A method of targeting an intracellular target, comprising administering an antibody conjugated with one or more MGS peptides, wherein the antibody targets the intracellular target, is disclosed. In some embodiments, the intracellular target may be a lysosome, Golgi apparatus, endoplasmic reticulum, cytoplasm, or nucleus. Any subcellular compartment can be targeted.

[0068] In one embodiment, one of ordinary skill in the art can induce an intracellular target to be inactivated by determining an effective dose, effective schedule, or effective route of administration of the disclosed composition or the disclosed fusion protein so as to target the intracellular target.

[0069] A method of treating a subject in need of treatment, comprising administering to the subject in need of treatment an effective amount of an antibody conjugated with one or more MGS peptides, wherein the antibody targets an intracellular target involved in the disease process, is also disclosed.

[0070] In any one embodiment of the methods disclosed herein, the compositions, conjugates or fusion proteins described herein can be used in combination with one or more additional therapies. In one embodiment, the composition, conjugate, or fusion protein can be administered alone or in combination with an agent having other biological activity to form a composition suitable for administration to a subject. In one embodiment, a method for the treatment of a subject having cancer or at risk of developing cancer The compositions, conjugates, or fusion proteins disclosed herein can be used in combination with, for example, a therapeutically effective amount of radiotherapy, immunotherapy, or chemotherapy, or a combination thereof. The combination therapy can be administered as a co-formulation or separately. When administered separately, the combination therapy can be administered simultaneously or sequentially. The formulation can be manufactured using conventional methods in the art.

[0071] The pharmaceutical composition described above can be formulated to contain a therapeutically effective amount of the composition, conjugate, or fusion protein disclosed herein. Therapeutic administration includes prophylactic administration. Based on genetic testing and other prognostic diagnostic methods, a physician consulting a patient can select prophylactic administration if the patient has a clinically determined predisposition or high susceptibility (in some cases, significantly high susceptibility) to one or more autoimmune diseases, or if the patient has a clinically determined predisposition or high susceptibility (in some cases, significantly high susceptibility) to cancer.

[0072] The pharmaceutical compositions described herein can be administered to a subject (e.g., a human subject or patient) in an amount sufficient to delay, alleviate, or preferably prevent a clinical disease. Thus, in some embodiments, the subject is a human subject. In therapeutic use, the composition is administered to a subject (e.g., a human subject) already having or diagnosed with an autoimmune disease in an amount sufficient to at least partially improve the signs or symptoms or to arrest (and preferably stop) the progression of the disease state, its complications, and outcome. The amount suitable to achieve this is defined as a "therapeutically effective amount". The therapeutically effective amount of a pharmaceutical composition can be an amount that results in a cure, although the outcome is only one of several possible outcomes that may be achieved. As described, the therapeutically effective amount includes an amount that provides a treatment in which the onset or progression of cancer is delayed, arrested, or prevented, or in which an autoimmune disease or symptoms of an autoimmune disease are improved. One or more of the symptoms may be of low severity. Recovery may be promoted in an individual receiving the treatment.

[0073] The total effective amount of the conjugate or fusion protein in the pharmaceutical composition disclosed herein can be administered to a mammal as a bolus or as a single dose by infusion over a relatively short period of time, or multiple doses can be administered using a split treatment protocol over a longer period of time (e.g., every 4-6, 8-12, 14-16, or 18-24 hours, or every 2-4 days, 1-2 weeks, or once a month). Also included within the scope of the present disclosure is continuous intravenous infusion sufficient to maintain a therapeutically effective concentration in the blood.

[0074] The therapeutically effective amount of an antibody present in the compositions described herein and used in the methods disclosed herein for application to a mammal (e.g., a human) can be determined by one of ordinary skill in the art taking into account the individual differences in age, weight, and other general conditions (such as those described above). The compositions, conjugates, and fusion proteins of the present specification are stable in serum and blood flow, and in some cases are more specific, so the doses of the compositions, conjugates, and fusion proteins containing the individual components may be lower (or higher) than the effective dose of any of the individual components that are not added together. Thus, in some embodiments, the antibody administered, when administered as part of a conjugate or fusion protein, may have a higher effect or lower side effects compared to when the antibody is administered alone or not as part of a conjugate or fusion protein.

[0075] In some embodiments, the subject in need of treatment has an infectious disease, cancer, diabetes, a neurological or neurodegenerative disease, a genetic disease, a lysosomal disease, a mitochondrial disease, or has been exposed to a bioterrorism agent.

[0076] D. Vector Vectors are disclosed that contain nucleic acid sequences encoding one or more of the disclosed compositions. In some embodiments, the vector contains only nucleic acid sequences capable of encoding one or more of the disclosed MGS peptides.

[0077] E. Kit The materials described above and other materials can be packaged together in any suitable combination as a kit useful for carrying out or assisting in the practice of the disclosed methods. Kit elements within a particular kit are useful if they are designed and adapted to be used together in the disclosed methods. For example, kits are disclosed that include one or more of the disclosed compositions.

[0078] In some embodiments, the kit includes an MGS peptide, a monoclonal antibody, and instructions for use for performing the binding.

[0079] In some embodiments, the kit includes a cell line comprising a nucleic acid sequence encoding one or more of the MGS peptides.

Examples

[0080] A. Example 1 Systematic approaches targeting intracellular / subcellular compartments have not been reported until recently. The patented FOX-Three technology is a platform for systematically discovering MGSs that can deliver MAb to cells and subcellular targets. The FOX-Three platform identifies MGSs by three main characteristics: the ability to direct to a desired cell type, the ability to induce cellular uptake, and the ability to deliver a payload to an individual location in the target cell. An overview showing how MGS works is shown in Figure 1.

[0081] The FOX-Three platform can rapidly screen target cells and intracellular systems to identify specifically targeted MGSs (Figure 2), established MGS members based on 10 9 ~10 12 candidate peptides. It is based on a phage display library. Peptides are a well - understood class of biomolecules that can be easily synthesized by biological processes as well as by artificial chemical processes. As a result, the power of the FOX - Three platform in terms of its speed and flexibility has been demonstrated. This platform can be applied to any cell regardless of knowledge of the cell's molecular properties and generates lead MGS in 2 - 4 weeks. Subsequently, the selected peptide - based MGS is rapidly engineered to optimize its performance. Table 2 shows the MGS identified using the FOX - Three technology.

Table 2

[0082] The FOX-Three platform has already been demonstrated for targeting cells and subcellular components. Many proven MGSs for different cell targeting systems are in different stages of development (Tables 1 and 2). Methods have been established to couple various payloads to peptide MGSs without compromising their targeting ability. The selected MGSs have been demonstrated to deliver drugs, contrast agents, nanoparticles, DNA, and proteins to target cells in culture and animal models. Cell targeting is an established technology, and there are many competing antibody-based technologies for targeting the surface of target cells, but intracellular MGSs have not been known until now. The importance of intracellular targeting is shown in Figure 3, which shows the complex structure of cells. For example, the delivery of MAb intended to bind to proteins involved in DNA replication in the cell nucleus will not have a therapeutic effect if it is trapped in the Golgi apparatus or other parts of the cell. The screening process by FOX-Three has been improved in recent years to include intracellular location as a selection criterion. MGSs that accumulate in lysosomes, autophagosomes, and the Golgi apparatus in cancer cells, as well as MGSs that target the cell membrane, have been isolated. It has been demonstrated that the therapeutic effect of MGSs depends on delivery to the correct intracellular location.

[0083] Considering the cell type, pathology, subcellular organelle, and the variety of payloads that can be delivered, the potential of the FOX-Three platform is immeasurable. A stable platform can be developed that provides an optimized "toolbox" of MGSs capable of delivering a set of MAb focused on the cell type and subcellular components of the selected disease. To do this, the number of subcellular organelles that can be targeted using the FOX-Three platform can be increased (Figure 3). A group of experiments using quantitative metrics can be performed.

[0084] In some embodiments, the synthesis, characterization, and optimization of three types of MGS that target non-small cell cancer cell lines and accumulate in lysosomes, Golgi bodies, and mitochondria can be studied.

[0085] In some embodiments, three types of isolated MGS can be used to intracellularly deliver antibodies having biological activity to lysosomes, Golgi bodies, and mitochondria. Regulation of specific protein targeting in each of these subcellular components can be demonstrated.

[0086] In some embodiments, at least five types of MGS that target virus-infected cells can be identified. Furthermore, the intracellular locations can be increased to include lysosomes, Golgi bodies, mitochondria, endoplasmic reticulum, and cell nuclei.

[0087] The ability to deliver therapeutic MAb to specific target cells and the correct compartments within those cells can have many social health effects. The combination of MGS-MAb has the potential to create a new class of safe and effective intracellular-acting drugs that can treat targets previously considered "undruggable." This opens the way to new treatment strategies for newly emerging infectious diseases and bioterrorism, cancer, diabetes, neurological and neurodegenerative diseases, and even genetic diseases. The MGS-MAb conjugate drug can create an entirely new market for this new class of drugs, comparable to at least the current $70 billion market size of MAb, affecting virtually all pathologies and providing a major breakthrough in modern medicine.

[0088] FOX-Three discovers MGS and provides a highly safe and rapid response to newly emerging threats by enabling the rapid development of intracellular targeting human therapeutic antibodies at a previously unheard-of speed.

[0089] The developed FOX-Three MGS toolbox can deliver a variety of other payloads. Although the delivery of MAb is the main aim, it is important to note that MGS can deliver many different payloads and may have other clinical applications (Tables 1 and 2). MGS can deliver small molecule drugs, contrast agents, nanoparticles, DNA, radionuclides, and other proteins. MGS can be used in the early detection of diseases and also as a companion diagnosis. Early disease detection is often a major determinant of clinical outcome. Companion diagnosis can follow the response to treatment, enable rapid changes to treatment if necessary, and lead to time and cost savings. MGS-therapeutics can be used in a wide range of pathologies, in vitro and in vivo diagnostics, personalized cancer treatment, intracellular nanoparticle delivery, and targeted therapy in innovative immunotherapy (synthetic macromolecules such as small molecules, nucleic acids, polymers, and proteins).

[0090] This technology has the ability to rapidly produce MGS-MAbs that target pathogen-infected cells and improve treatments for existing and newly emerging pathogens on a weekly basis rather than annually. Current technologies are too slow to develop targeted treatments for emerging viruses.

[0091] This technology opens the way for a new class of therapeutics for the treatment of many diseases by delivering compounds that were previously cell-impermeable and represents a completely new armament for protecting combatant molecules.

[0092] This technology provides the ability to detect and neutralize latent intracellular viral infections that can be reactivated or spread / re-diffused throughout the community.

[0093] This technology can rapidly develop MGS that can be used in sensor technologies for diagnosis and biological readout.

[0094] B. Example 2 MGS can affect the treatment of many diseases and may open up new treatment options. By targeting protein interactions, a new class of intracellular-acting drugs with the potential to treat "undruggable" targets is created. Therapeutic indications include cancer, newly emerging infectious diseases and bioterrorism, heart disease, diabetes, fibrosis, and neurological and neurodegenerative diseases. MGS has broad applicability and can be used to deliver small molecule drugs, contrast agents, nucleic acids and proteins, nanoparticles, and cell therapies.

[0095] Small molecule peptide-MGS delivers large monoclonal antibodies (MAbs) intracellularly in vitro and in vivo. The small molecule peptide cannot carry an MAb that is approximately 60 times its molecular weight. When cells were treated with MAb alone, no cellular uptake occurred. However, when MAb was conjugated to one of the identified MGS peptides, a 20- to 4000-fold increase in cellular uptake was observed. The peptide not only delivers cell-impermeable MAb intracellularly, but the ability of the peptide to accumulate at the desired intracellular location is unchanged even when MAb is conjugated. Peptide-MGS delivers MAb and increases cellular uptake 20- to 380-fold (Figure 4A). Peptide-MGS can deliver MAb to individual intracellular locations determined by MGS (Figure 4B). Peptide-MGS can re-direct MAb to tumors and maintain MAb within tumors for over 72 hours (Figure 4C). Tumor uptake increases 2- to 4-fold while uptake in non-specific tissues (e.g., kidney and liver) is reduced.

[0096]

[0097] ​To show how the peptide (MGS) can deliver cargo to the correct intracellular location and the importance of this precise delivery, two different peptide - therapeutic conjugates were tested. The first peptide (MGS) is a Golgi - targeting peptide that specifically binds to NSCLC cells (Figure 9). This peptide (MGS) was conjugated to duocarmycin, a DNA - damaging agent. This conjugate has a linker between the drug and the peptide, and for it to become active and reach the cell nucleus, this linker must be cleaved by an enzyme found within lysosomes. However, when cells are treated with this conjugate, despite being a very potent drug, the conjugate is substantially non - toxic to these cells. This is because the peptide accumulates in the Golgi apparatus and is sequestered there.

[0098] Peptide - MGS can deliver cell - impermeable therapeutic agents (protein toxins, saporin, and the DNA - alkylating agent duocarmycin) to specifically kill cancer cells. Intracellular localization determines the effect of the therapeutic agent. Preliminary data on the MGS - saporin conjugate shows antitumor activity in animals.

[0099] Figure 10 shows seven MGS peptides and their intracellular localization compartments. The peptide EHPWFNMWSWATQVQE (SEQ ID NO: 38) (H2009.2) is pushed forward first. The peptide valency is optimized, the peptide binds to three different non - small cell lung cancer cells, has minimal binding to normal control cells, delivers MAb intracellularly, and effectively delivers cell - impermeable protein - toxins to induce cell death in cancer cells at an effective concentration of 80 nM.

[0100] Figure 11 shows that Ras is an important central factor in cancer proliferation and has been a target for drug therapy. The MAb - based therapeutic approach for treating Ras reduces the overall intracellular concentration of Ras, prevents activation by changing its location in the cell, and maintains Ras in an inactive state.

[0101] Figure 17 shows the reduction of tumor growth compared to the case without treatment or treatment with MAb alone. No overall toxicity was observed.

[0102] Figure 20 shows that direct binding does not prevent the ability of MGS to mediate cell uptake. The longer the linker between MGS and MAb, the greater the amount of uptake into target cells. A conjugate of MAb (NHS ester chemical reaction) and MGS-peptide (maleimide chemical reaction) was formed using a bifunctional linker with the same chemical reaction groups. While SMCC does not have a PEG spacer, PEG-12 and PEG-24 have shorter or longer PEG repeats between the chemical reaction groups, which enhances the ability of the MGS-peptide to interact with its cell receptor without being sterically hindered by the MAb protein. As demonstrated, the PEG-24 spacer achieved higher uptake of the MAb-MGS conjugate compared to two conjugates with shorter spacer arms.

[0103] Figure 23 shows how a specific MGS peptide binds to recycled cell receptors and how a specific MGS peptide binds to cell receptors that need to be newly synthesized. MGS_H2009.1 v4 rapidly recycles to the cell surface and binds to cell receptors that can be carried by another molecule of MGS there. MGS_H1299.2 v4, HCC15.1 v4, and HCC15.2 v8 bind to cell receptors that are mostly degraded upon internalization, and new receptor synthesis occurs for further internalization of MGS.

[0104] The cell receptor for MGS_H1299.2 v4 is EphA2. The following results were obtained in the study of EphA2. A decrease in EphA2 in cells abolishes the binding of MGS_H1299.2 V4. An increase in EphA2 in cells increases the binding of MGS_H1299.2 V4. MGS_H1299.2 V4 binds to EphA2 obtained from lysed cells and binds to purified EphA2 (Kd 3.2 nM). Eprhri nA1 competes with MGS_H1299.2 V4 for binding. Finally, MGS_H1299.2 V4 induces the internalization of EphaA2 and co-localizes with the receptor within lysosomes.

[0105] It will be apparent to those skilled in the art that various modifications and changes can be made without departing from the scope or spirit of the invention. Other aspects of the invention will be apparent to those skilled in the art in view of the specification and practice of the invention disclosed herein. The specification and examples are intended to be exemplary only, and the true scope and spirit of the invention are indicated by the following claims.

[0106] Those skilled in the art will recognize, or be able to ascertain using no more than routine experimentation, many equivalents to the specific embodiments of the methods and compositions described herein. Such equivalents are intended to be encompassed by the following claims.

Claims

1. FHAVPQSFYTA (SEQ ID NO: 2), CH ３ CO-FHAVPQSFYT-PEG11-C-NH ２ (SEQ ID NO: 76), CH ３ CO-FHAVPQSFYT-PEG11-C-NH ２ (SEQ ID NO: 77), FHAVPQSFY (SEQ ID NO: 78), HAVPQSFYT (SEQ ID NO: 79), LQWRRDDNVHNFGVWARYRL (SEQ ID NO: 30), CH ３ CO-LQWRRDDNVHNFGVWARYRL (SEQ ID NO: 83), THGNKHQSWTYPSSEINHKNY (SEQ ID NO: 19), LADTWTQTQQHDFHVLRGTR (SEQ ID NO: 20), GYSWWQPNWPSSTWDT (SEQ ID NO: 21), NLADTWTQTQQHDFHVLRGT (SEQ ID NO: 23), YPGSPTQYPSSMHEYHSSSE (SEQ ID NO: 39), AHTIDDEWASYHMQQWNSPP (SEQ ID NO: 40), FEEFYSRQ SNTIPYPQQYKG (SEQ ID NO: 41), LTVHGRGPEYNPSWNRRAFP (SEQ ID NO: 46), SVEYWGERMYYDVMESLGFS (SEQ ID NO: 47), FAAKRAEWWDPGQLWDAVWN (SEQ ID NO: 48), QEALEEWFWKMMPWSGPSGQ (SEQ ID NO: 49), TWTDFGQWPWPFGAEGTRAF (SEQ ID NO: 50), MDGATWWTQLDPLLVWEGET (SEQ ID NO: 51), SADWFQGPAEWLLEGW MGPL (SEQ ID NO: 52), SAKTAVSQRVWLPSHRGGEP (SEQ ID NO: 57), KSREHVNNSACPSKRITAAL (SEQ ID NO: 58), TGGETSGIKKAPYASTTRNR (SEQ ID NO: 60), SHHGVAGVDLGGGADFKSIA (SEQ ID NO: 61), SNSPLGLKDEATQRVLEQAKWLA (SEQ ID NO: 62), GPEDTSRAPENQQKTFHRRW (SEQ ID NO: 63), SGETGSNLVGHELDFRPGSP SP (SEQ ID NO: 64), RYSPAATAEGRSVSKELLRV (SEQ ID NO: 65), GQELGAWTRSKGPEVQTSVL (SEQ ID NO: 66), ASTWRGTSAGGNRLEKMEVT (SEQ ID NO: 67), LSGTPERSGQAVKVKLKAIP (SEQ ID NO: 68), GAWEAVRDRIAEWGSWGIPS (SEQ ID NO: 69), and AMDMYSIEDRYFGGYAPEVG (SEQ ID NO: 70).

2. The MGS peptide of claim 1 , wherein the MGS peptide is bound to an antibody directly or via a linker.

3. 3. The MGS peptide of claim 2, wherein the linker comprises polyethylene glycol (PEG).

4. The MGS peptide of claim 2 , wherein the antibody is a monoclonal antibody.

5. A composition comprising one or more molecular guidance system (MGS) peptides linked to an antibody directly or via a linker, wherein each of the one or more MGS peptides is selected from the group consisting of FHAVPQSFYTA (SEQ ID NO: 2), CH ３ CO-FHAVPQSFYT-PEG11-C-NH ２ (SEQ ID NO: 76), CH ３ CO-FHAVPQSFYT-PEG11-C-NH ２ (SEQ ID NO: 77), FHAVPQSFY (SEQ ID NO: 78), HAVPQSFYT (SEQ ID NO: 79), LQWRRDDNVHNFGVWARYRL (SEQ ID NO: 30), CH ３ CO-LQWRRDDNVHNFGVWARYRL (SEQ ID NO: 83), THGNKHQSWTYPSSEINHKNY (SEQ ID NO: 19), LADTWTQTQQHDFHVLRGTR (SEQ ID NO: 20), GYSWWQPNWPSSTWDT (SEQ ID NO: 21), NLADTWTQTQQHDFHVLRGT (SEQ ID NO: 23), YPGSPTQYPSSMHEYHSSSE (SEQ ID NO: 39), AHTIDDEWASYHMQQWNSPP (SEQ ID NO: 40), FEEFYS RQSNTIPYPQQYKG (SEQ ID NO: 41), LTVHGRGPEYNPSWNRRAFP (SEQ ID NO: 46), SVEYWGERMYYDVMESLGFS (SEQ ID NO: 47), FAAKRAEWWDPGQLWDAVWN (SEQ ID NO: 48), QEALEEWFWKMMPWSGPSGQ (SEQ ID NO: 49), TWTDFGQWPWPFGAEGTRAF (SEQ ID NO: 50), MDGATWWTQLDPLLVWEGET (SEQ ID NO: 51), SADWFQGPAEW LLEGWMGPL (SEQ ID NO: 52), SAKTAVSQRVWLPSHRGGEP (SEQ ID NO: 57), KSREHVNNSACPSKRITAAL (SEQ ID NO: 58), TGGETSGIKKAPYASTTRNR (SEQ ID NO: 60), SHHGVAGVDLGGGADFKSIA (SEQ ID NO: 61), SNSPLGLKDEATQRVLEQAKWLA (SEQ ID NO: 62), GPEDTSRAPENQQKTFHRRW (SEQ ID NO: 63), SGETGSNLVGHE The composition is selected from LDFRPGSPSP (SEQ ID NO: 64), RYSPAATAEGRSVSKELLRV (SEQ ID NO: 65), GQELGAWTRSKGPEVQTSVL (SEQ ID NO: 66), ASTWRGTSAGGNRLEKMEVT (SEQ ID NO: 67), LSGTPERSGQAVKVKLKAIP (SEQ ID NO: 68), GAWEAVRDRIAEWGSWGIPS (SEQ ID NO: 69), and AMDMYSIEDRYFGGYAPEVG (SEQ ID NO: 70).

6. The composition of claim 5 , wherein the antibody is a monoclonal antibody.

7. The composition of claim 5 , wherein the linker comprises polyethylene glycol (PEG).

8. The composition of claim 5, wherein the one or more MGS peptides localize to and internalize one or more intracellular targets, direct the antibody to the one or more intracellular targets, and in response, the antibody targets the one or more intracellular targets.

9. The composition of claim 8 , wherein the intracellular target is a lysosome, a Golgi apparatus, an endoplasmic reticulum, a cytoplasm, or a cell nucleus.

10. The composition of any one of claims 5 to 9, wherein the antibody conjugated to the one or more MGS peptides is a fusion protein.

11. A composition according to any one of claims 5 to 9 for targeting an intracellular target, wherein said one or more MGS peptides deliver said antibody into said cell and to said intracellular target.

12. 10. The composition of any one of claims 5 to 9 for treating a subject in need thereof, wherein the antibody targets an intracellular target involved in a disease process.

13. 13. The composition of claim 12, wherein the subject in need of treatment has an infectious disease, cancer, diabetes, a neurological or neurodegenerative disease, a genetic disease, or has been exposed to a bioterrorism agent.

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