Methods for treatment of cancer with polynucleotides or antibodies that target mutated cell surface related proteins

Abstract

Methods and compositions are provided for treatment of cancer. The methods and compositions target mutations in cell surface proteins that are specific to cancer cells in an individual. Polynucleotides that encode antibodies that specifically bind to mutated cell surface proteins or that encode mutated cell surface peptides that are expressed in cancer cells in an individual and / or extrinsically produced antibodies that specifically bind to the mutated cell surface proteins that are expressed in the cancer cells may be administered to the individual, for treatment of cancer.

Description

METHODS FOR TREATMENT OF CANCER WITH POLYNUCLEOTIDES OR ANTIBODIES THAT TARGET MUTATED CELL SURFACE RELATED PROTEINSCROSS-REFERENCE TO RELATED APPLICATIONS

[0001] This application claims the benefit of U.S. Provisional Application Nos. 63 / 613,387, filed on December 21 , 2023, and 63 / 555,538, filed on February 20, 2024, both of which are incorporated by reference herein in their entireties.FIELD OF THE INVENTION

[0002] The invention relates to methods for treatment of cancer, in particular methods that target mutated cell surface proteins in cancer cells with polynucleotides that encode the mutated cell surface proteins or that encode antibodies that specifically bind to the mutated proteins, or with extrinsically produced antibodies that bind to the mutated cell surface proteins.BACKGROUND

[0003] Cancer treatment methods that target mutated proteins focus on the binding by T cells via a T cell receptor to a peptide representing the mutated section of a mutated protein. T cell binding only occurs with short peptides that are physically located in the groove of a major histocompatibility complex (MHC) molecule. The MHC and the contained peptide are located on the surface of the cell. Peptides in the groove of an MHC molecule represent a very different population of peptide targets than mutated cell surface proteins (MSPs).

[0004] Peptides that end up in the groove of an MHC molecule originate from proteins that are degraded and processed in the cytoplasm. The majority of the proteins that are processed are intracellular proteins. Thus, common understanding is that intracellular mutated proteins are the source of peptides targeted by T cells. Another distinction is that T cell targeting of mutated peptides targets a fragment of the mutated peptide that is physically far removed from the location of the original intact mutated protein. T cells only bind to a shredded portion of the original protein.

[0005] It would be desirable to target MSPs with a reagent that binds directly to the protein via a mutated epitope, rather than binding to a detached peptide that is removed from the intact protein from which it is derived as occurs with T cell targeting.BRIEF SUMMARY OF THE INVENTION

[0006] Methods and compositions for treatment of cancer are provided.

[0007] In one aspect, a method for treatment of cancer is provided, which includes: (a) sequencing DNA and / or RNA (e.g., mRNA) of cancer cells and non-cancer cells from an individual to determine mutations in the amino acid sequences of cell surface related proteins in the cancer cells in the individual in comparison to the amino acid sequences of the cell surface related proteins in the non-cancer cells in the individual; (b) administering to the individual or to a second individual, such as an animal (e.g., a rodent, such as a mouse), at least onepolynucleotide (e.g., mRNA polynucleotide) that a encodes a mutated cell surface related (MSP) peptide that is expressed in the cancer cells in the individual, wherein the MSP peptide includes an amino acid sequence of at least a portion of a cell surface related protein, which includes one or more mutation in the MSP peptide amino acid sequence relative to the amino acid sequence of the cell surface related protein in non-cancer cells in the individual, wherein the polynucleotide (e.g., mRNA polynucleotide) is expressed (e.g., translated) by the individual or the second individual to produce the MSP peptide, and wherein the MSP peptide generates an immune response in the individual or the second individual that includes production of antibodies by the individual or the second individual that specifically bind to the MSP peptide; (c) assessing the immune response by assessing antibody titer to the MSP peptide, e.g., in a blood sample that is obtained from the individual or the second individual; (d) sequencing an antibody from the individual or the second individual (e.g., an antibody in a blood sample that is obtained from the individual or the second individual) that specifically binds to the MSP peptide to determine the amino acid sequences of the antibody; (e) using the antibody amino acid sequences to produce the antibody, or a fragment or derivative thereof that retains the ability to bind to the MSP peptide, in a system that is extrinsic to the individual; and (f) administering the antibody, or fragment or derivative thereof, that is produced in the extrinsic system to the individual for treatment of the cancer.

[0008] In another aspect, a method for treatment of cancer is provided, which includes administering to an individual having cancer at least one antibody that specifically binds to an MSP peptide that is expressed in cancer cells of the individual, or a fragment or derivative thereof that retains the ability to bind to the MSP peptide, wherein the antibody, or the fragment or derivative thereof, is produced in a method that includes: (a) administering to the individual or to a second individual, such as an animal (e.g., a rodent, such as a mouse), at least one polynucleotide (e.g., mRNA polynucleotide) that encodes a MSP peptide that is expressed in the cancer cells in the individual, wherein the MSP peptide includes an amino acid sequence of at least a portion of a cell surface related protein, which includes one or more mutation in the MSP peptide amino acid sequence relative to the amino acid sequence in the cell surface protein in non-cancer cells in the individual, wherein the polynucleotide (e.g., mRNA polynucleotide) is expressed (e.g., translated) by the individual or the second individual to produce the MSP peptide, and wherein the MSP peptide generates an immune response in the individual or the second individual that includes production of antibodies by the individual or the second individual that specifically bind to the MSP peptide; (b) assessing the immune response by assessing antibody titer to the MSP peptide, e.g., in a blood sample that is obtained from the individual or the second individual; (c) sequencing an antibody from the individual or the second individual (e.g., an antibody in a blood sample that is obtained from the individual or the second individual) that specifically binds to the MSP peptide to determine the amino acid sequences of the antibody; and (d) using the antibody amino acid sequences to produce the antibody, or a fragment or derivative thereof that retains the ability to bind to the MSP peptide, in a system that is extrinsic to the individual.

[0009] In another aspect, a method for treatment of cancer is provided, which includes: (a) sequencing DNA and / or RNA (e.g., mRNA) of cancer cells and non-cancer cells from an individual to determine mutations in the amino acid sequences of cell surface related proteins in the cancer cells in the individual in comparison to the amino acid sequences of the cell surface related proteins in the non-cancer cells in the individual; (b) administering to the individual or to a second individual, such as an animal (e.g., a rodent, such as a mouse), at least one polynucleotide (e.g., mRNA polynucleotide) that a encodes a MSP peptide that is expressed in the cancer cells in the individual or the second individual, wherein the MSP peptide includes an amino acid sequence of at least a portion of a cell surface related protein, which includes one or more mutation in the MSP peptide amino acid sequence relative to the amino acid sequence of the cell surface related protein in non-cancer cells in the individual, wherein the polynucleotide (e.g., mRNA polynucleotide) is expressed (e.g., translated) by the individual or the second individual to produce the MSP peptide, and wherein the MSP peptide generates an immune response in the individual or the second individual that includes production of antibodies by the individual that specifically bind to the MSP peptide; (c) assessing said immune response by assessing antibody titer to the MSP peptide, e.g., in a blood sample that is obtained from the individual or the second individual; (d) sequencing an antibody from the individual or the second individual (e.g., an antibody in a blood sample that is obtained from the individual or the second individual) that specifically binds to the MSP peptide to determine the amino acid sequences of the antibody; and (e) administering at least one vector antibody polynucleotide (e.g., mRNA) that includes polynucleotide sequences that encode the amino acid sequences of the antibody, or a fragment or derivative thereof that retains the ability to bind to the MSP peptide, to the individual for treatment of the cancer.

[0010] In another aspect, a method is provided for treatment of cancer, which includes: administering to an individual having cancer at least one vector antibody polynucleotide (e.g., mRNA) that includes polynucleotide sequences that encode the amino acid sequences of an antibody that specifically binds to an MSP peptide that is expressed in cancer cells of the individual, or a fragment or derivative thereof that retains the ability to bind to the MSP peptide. In one embodiment of the method, the vector antibody polynucleotide (e.g., mRNA) is produced in a method that includes: (a) administering to the individual or to a second individual, such as an animal (e.g., a rodent, such as a mouse): (i) at least one polynucleotide (e.g., mRNA polynucleotide) that encodes a MSP peptide that is expressed in the cancer cells in the individual in comparison to the amino acid sequences of the cell surface related protein in non- cancer cells in the individual, wherein the polynucleotide (e.g., mRNA polynucleotide) is expressed (e.g., translated) by the individual or the second individual to produce the MSP peptide, and / or (ii) at least one MSP peptide that is expressed in the cancer cells in the individual, wherein the MSP peptide includes an amino acid sequence of at least a portion of a cell surface related protein, which includes one or more mutation in the MSP peptide amino acid sequence relative to the amino acid sequence of the cell surface related protein in non-cancer cells in the individual, and wherein the MSP peptide generates an immune response in theindividual or the second individual that includes production of antibodies by the individual or the second individual that specifically bind to the MSP peptide; (b) assessing the immune response by assessing antibody titer to the MSP peptide, e.g., in a blood sample that is obtained from the individual or the second individual; and (c) sequencing an antibody from the individual or the second individual (e.g., an antibody in a blood sample that is obtained from the individual or the second individual) that specifically binds to the MSP peptide to determine the amino acid sequences of the antibody.

[0011] In another aspect, a method is provided for treatment of cancer, which includes administering to an individual having cancer, at least one polynucleotide (e.g., mRNA polynucleotide) that encodes a MSP peptide that is expressed in cancer cells in said individual, wherein the MSP peptide an amino acid sequence of at least a portion of a cell surface related protein, which includes one or more mutation in the MSP peptide amino acid sequence relative to the amino acid sequence of the cell surface related protein in non-cancer cells in the individual, wherein the polynucleotide (e.g., mRNA polynucleotide) is expressed (e.g., translated) by the individual to produce the MSP peptide, and wherein the MSP peptide generates an immune response in the individual that includes production of antibodies by the individual that specifically bind to the MSP peptide. In one embodiment of the method, prior to administration of the at least one polynucleotide (e.g., mRNA polynucleotide) that encodes the MSP peptide, DNA and / or RNA (e.g., mRNA) of cancer cells and non-cancer cells from the individual is sequenced to determine mutations in the amino acid sequences of cell surface related proteins in the cancer cells relative to the amino acid sequences of the cell surface related proteins in non-cancer cells of the individual.

[0012] In another aspect, a method for treatment of cancer is provided, which includes: (a) sequencing DNA and / or RNA of cancer cells and non-cancer cells from an individual to determine mutations in the amino acid sequences of cell surface related proteins in the cancer cells in the individual in comparison to the amino acid sequences of the cell surface related proteins in the non-cancer cells in the individual; (b) administering to the individual, or to a second individual, such as an animal (e.g., a rodent, such as a mouse) at least one MSP peptide that is expressed in the cancer cells in the individual, wherein the MSP peptide comprises an amino acid sequence of at least a portion of a cell surface related protein, which includes one or more mutation in the MSP peptide amino acid sequence relative to the amino acid sequence in the cell surface related protein in non-cancer cells in the individual, and wherein the MSP peptide generates an immune response in the individual or the second individual that includes production of antibodies by the individual or the second individual that specifically bind to the MSP peptide; (c) assessing the immune response by assessing antibody titer to the MSP peptide in a blood sample that is obtained from the individual or the second individual; (d) sequencing an antibody from the individual or the second individual (e.g., an antibody in a blood sample that is obtained from the individual or the second individual) that specifically binds to the MSP peptide to determine the amino acid sequences of the antibody; and (e) administering a vector antibody polynucleotide (e.g., mRNA) that includespolynucleotide sequences that encode the amino acid sequences of the antibody, or a fragment or derivative thereof that retains the ability to bind to the MSP peptide, to the individual for treatment of the cancer.

[0013] In another aspect, a method for treatment of cancer is provided, which includes administering to an individual having cancer at least one polynucleotide (e.g., mRNA) that includes polynucleotide sequences that encode the amino acid sequences of an antibody that specifically binds to an MSP peptide that is expressed in cancer cells of the individual, or a fragment or derivative thereof that retains the ability to bind to the MSP peptide, wherein the MSP peptide includes an amino acid sequence of at least a portion of a cell surface related protein, which includes one or more mutation in the MSP peptide amino acid sequence relative to the amino acid sequence of the cell surface related protein in non-cancer cells in the individual, wherein the MSP peptide generates an immune response in the individual that includes production of antibodies by the individual that specifically bind to the MSP peptide, and wherein the at least one polynucleotide (e.g., mRNA) that encodes the amino acid sequences of the antibody is produced in a method that includes: (a) administering to the individual or to a second individual, such as an animal (e.g., a rodent, such as a mouse), at least one MSP peptide that is expressed in the cancer cells in said individual, wherein the MSP peptide includes an amino acid sequence of at least a portion of a cell surface related protein that includes one or more mutation in the amino acid sequence of the MSP peptide relative to the amino acid sequence of the cell surface related protein in non-cancer cells in the individual; (b) assessing the immune response by assessing antibody titer to said MSP peptide in a blood sample that is obtained from the individual or the second individual; and (c) sequencing an antibody from the individual or the second individual (e.g., an antibody in a blood sample that is obtained from the individual or the second individual) that specifically binds to the MSP peptide to determine the amino acid sequences of the antibody.

[0014] In some embodiments of the methods described herein, the cell surface related protein that includes one or more mutation from which the MSP peptide is derived is expressed on the surface of the cancer cells in the individual. In some embodiments of the methods described herein, the cell surface related protein that includes one or more mutation from which the MSP peptide is derived is secreted from the cancer cells in the individual.

[0015] In some embodiments of the methods described herein, a polynucleotide (e.g., mRNA) that encodes a MSP peptide or amino acid sequences of an antibody, or a fragment or derivative thereof, is administered to the individual in a lipid particle formulation.

[0016] In some embodiments of the methods described herein, a polynucleotide (e.g., mRNA) that encodes a MSP peptide or amino acid sequences of an antibody, or a fragment or derivative thereof, is administered to the individual intramuscularly, subcutaneously, intradermally, intravenously, or via inhalation.

[0017] In some embodiments of the methods described herein, at least one polynucleotide (e.g., mRNA) that encodes a MSP peptide includes: (i) at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten polynucleotides(e.g., mRNA polynucleotides), each of which encodes a different MSP peptide, or (ii) at least one polynucleotide (e.g., mRNA polynucleotide) that encodes at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten different MSP peptides.

[0018] In some embodiments of the methods described herein, at least one polynucleotide e.g., mRNA polynucleotide) that encodes amino acid sequences of an antibody, or a fragment or derivative thereof, that specifically binds to an MSP peptide includes: (i) at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten polynucleotides (e.g., mRNA polynucleotides), each of which encodes a different antibody, or fragment or derivative thereof, each of which specifically binds to the same or different MSP peptide(s) or (ii) at least one polynucleotide (e.g., mRNA polynucleotide) that encodes at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten different antibodies, or fragments or derivatives thereof, each of which specifically binds to the same or different MSP peptide(s).

[0019] In some embodiments of the methods described herein, immune response is determined by assessing antibody titer to an MSP peptide in a blood sample that is obtained from the individual or from a second individual, such as an animal (e.g., a rodent, such as a mouse).

[0020] In some embodiments, an antibody that specifically binds to an MSP peptide is sequenced to determine the amino acid sequences of the antibody. The antibody sequences may be used to produce the antibody, or a fragment or derivative thereof that retains the ability to bind to the MSP peptide, in a system that is extrinsic to the individual. For example, the extrinsic system may include a phage display system. In another example, the extrinsic system may include production of the antibody, or fragment or derivative thereof, in a non-human animal from a vector antibody polynucleotide (e.g., mRNA) that includes polynucleotide sequences that encode the amino acid sequences of the antibody, or fragment or derivative thereof. In some embodiments, the vector antibody polynucleotide (e.g., mRNA) further encodes an amino acid sequence that facilitates recovery of the antibody, or fragment or derivative thereof, e.g., from plasma of a non-human animal. For example, the amino acid sequence that facilitates recovery may be a His-tag sequence, such as six to nine consecutive histidine residues.

[0021] In some embodiments of the methods described herein, an antibody, or fragment or derivative thereof, that is produced in a system that is extrinsic to the individual is administered to the individual for treatment of the cancer, e.g., administered intravenously or subcutaneously. For example, antibodies, or fragments or derivatives thereof, that specifically bind at least one, at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten MSP peptides that are expressed in cancer cells in the individual are produced in the extrinsic system, and are administered to the individual for treatment of said cancer.

[0022] In some embodiments of the methods described herein, a vector antibody polynucleotide (e.g., mRNA polynucleotide) that includes polynucleotide sequences that encode the amino acid sequences of the antibody, or a fragment or derivative thereof that retains the ability to bind tothe MSP peptide, is administered to the individual for treatment of said cancer, e.g., administered intravenously, intramuscularly, or subcutaneously. For example, polynucleotides (e.g., mRNA polynucleotides) that encode at least one, at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten antibodies that specifically bind to MSP peptides that are expressed in cancer cells in the individual, e.g., antibodies, or fragments or derivatives thereof, that specifically bind at least one, at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten MSP peptides that are expressed in cancer cells in the individual, are administered to the individual for treatment of said cancer.

[0023] In certain embodiments, polynucleotides, e.g., vector antibody polynucleotides or polynucleotides that encode MSP peptides (e.g., mRNA polynucleotides) are loaded on effector cells and / or transfected into effector cells, e.g., effector cells isolated from the individual, or effector cells isolated from another individual, such as donor effector cells, or an effector cell line, such as a natural killer (NK) cell line, prior to administration to the individual, thereby producing armed effector cells. For example, the effector cells may include one or more of natural killer (NK) cells, T cells, B cells, monocytes / macrophages, neutrophils, or combinations thereof. In some embodiments, the effector cells secrete the antibodies or MSP peptides. In other embodiments, the antibodies or MSP peptides are retained in the cell and are located at the cell surface.

[0024] In some embodiments of the methods described herein, the cancer that is treated by the method is selected from basal cell carcinoma, bladder cancer, bone cancer, bowel carcinoma, breast cancer, carcinoid, anal squamous cell carcinoma, castration-resistant prostate cancer (CRPC), cervical carcinoma, colorectal cancer (CRC), colon cancer cutaneous squamous cell carcinoma, endometrial cancer, esophageal cancer, gastric carcinoma gastroesophageal junction cancer, glioblastoma / mixed glioma, glioma, head and neck cancer, hepatocellular carcinoma, hematologic malignancy, liver cancer, lung cancer, melanoma, Merkel cell carcinoma, multiple myeloma nasopharyngeal cancer, neuroblastoma, osteosarcoma, ovarian cancer, pancreatic cancer, peritoneal carcinoma, undifferentiated pleomorphic sarcoma, prostate cancer, rectal carcinoma, renal cancer, sarcoma, salivary gland carcinoma, squamous cell carcinoma, stomach cancer, testicular cancer, thymic carcinoma, thymic epithelial tumor, thymoma, thyroid cancer, urogenital cancer, urothelial cancer, uterine carcinoma, and uterine sarcoma.

[0025] In another aspect, a composition, e.g., a pharmaceutical composition, e.g., a composition for treatment of cancer, is provided. The compositions described herein may be used for treatment of cancer as described herein. The composition may include polynucleotides (e.g., mRNA polynucleotides) that encode one or more MSP peptide(s) and / or that encode one or more antibodies, or fragments or derivatives thereof, that specifically bind to one or more MSP peptide(s) that are expressed in cancer cells in the individual, wherein the MSP peptide(s) contain one or more mutation(s) relative to the cell surface related proteins in non-cancer cells in the individual. The composition may also include one or more pharmaceutically acceptableexcipient and / or other components for improved administration and / or stability of the composition.

[0026] In some embodiments of the compositions described herein, the composition includes: at least one polynucleotide (e.g., mRNA polynucleotide) that encodes a MSP peptide that is expressed in cancer cells in an individual, wherein the MSP peptide includes an amino acid sequence of at least a portion of a cell surface related protein, which includes one or more mutation in the MSP peptide amino acid sequence relative to the amino acid sequence of the cell surface related protein in non-cancer cells in the individual; and a pharmaceutically acceptable excipient. For example, the at least one polynucleotide (e.g., mRNA polynucleotide) that encodes a MSP peptide includes: (i) at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten polynucleotides (e.g., mRNA polynucleotides), wherein each of the polynucleotides (e.g., mRNA polynucleotides) encodes a different MSP peptide, and / or (ii) at least one polynucleotide (e.g., mRNA polynucleotide) that encodes at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten different MSP peptides.

[0027] In some embodiments of the compositions described herein, the composition includes: at least one polynucleotide (e.g., mRNA polynucleotide) that encodes an antibody, or a fragment or derivative thereof, that specifically binds to an MSP peptide in an individual, wherein the MSP peptide is expressed in cancer cells in the individual, wherein the MSP peptide includes an amino acid sequence of at least a portion of a cell surface related protein, which includes one or more mutation in the MSP peptide amino acid sequence relative to the amino acid sequence of the cell surface related protein in non-cancer cells in the individual; and a pharmaceutically acceptable excipient. For example, the at least one polynucleotide (e.g., mRNA polynucleotide) that comprises polynucleotide sequences that encode the amino acid sequences of the antibody, or a fragment or derivative thereof includes: (i) at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten polynucleotides (e.g., mRNA polynucleotides), each of which encodes a different antibody, or fragment or derivative thereof, and / or (ii) at least one polynucleotide (e.g., mRNA polynucleotide) that encodes at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten different antibodies, or fragments or derivatives thereof

[0028] In some embodiments of the compositions described herein, polynucleotides (e.g., mRNA polynucleotides) are loaded on effector cells and / or transfected into effector cells, e.g., effector cells isolated from the individual, effector cells isolated from another individual, such as a donor, and / or an effector cell line, such as an NK cell line, prior to administration to the individual, thereby producing armed effector cells. For example, the effector cells may include one or more of natural killer (NK) cells, T cells, B cells, monocytes / macrophages, neutrophils, or combinations thereof. The composition may include armed effector cells, and a pharmaceutically acceptable excipient.

[0029] In another aspect, a method for production of an antibody, or a fragment or derivative thereof, that specifically binds an MSP peptide that is expressed in cancer cells in an individual is provided. The MSP peptide that is recognized by the antibody, or fragment or derivative thereof, includes an amino acid sequence of at least a portion of a cell surface related protein, which includes one or more mutation in the MSP peptide amino acid sequence relative to the amino acid sequence of the cell surface related protein in non-cancer cells in the individual, wherein the MSP peptide contains an amino acid sequence of a portion of a cell surface related protein that comprises one or more mutation relative to the cell surface related protein in noncancer cells in the individual. The method includes: (i) expression of the antibody, or fragment or derivative thereof, in a phage display system, from a polynucleotide that encodes the amino acid sequences of the antibody, or fragment or derivative thereof; or (ii) expression of the antibody, or fragment or derivative thereof, in a non-human animal from a mRNA polynucleotide that encodes the amino acid sequences of the antibody, or fragment or derivative thereof. In some embodiments, the polynucleotide that encodes the amino acid sequences of the antibody, or fragment or derivative thereof, further encodes an amino acid sequence that facilitates recovery or purification of the antibody, or fragment or derivative thereof. For example, the amino acid sequence that facilitates recovery or purification may include a His-tag sequence, e.g., including six to nine consecutive histidine residues.BRIEF DESCRIPTION OF THE DRAWINGS

[0030] Figure 1 shows the effect of antibodies that target mutated intracellular tumor proteins on tumor growth, as described in Example 2.

[0031] Figure 2 shows the effect of antibodies that target mutated intracellular tumor proteins on survival, as described in Example 2.

[0032] Figures 3A - 3C show tumor growth inhibition and survival enhanced in mice treated with a customized cocktail of MSP targeting antibodies in a EMT-6 TNBC model, as described in Example 3. (3A) Overview of the timing of implantation and treatment with a 9-Ab antibody cocktail and anti-PD-1 in BALB / c mice implanted subcutaneously with 1x106EMT-6 cells. (3B) Tumor growth. (3C) Survival. Treatment with antibodies targeting the cell surface neoantigens of 9 MSPs in combination with PD1 inhibition caused tumor growth inhibition and prolonged survival. DPI: days post-implantation, PD1 i: PD-1 inhibitor, nlgG: normal rabbit control IgG, 9- Ab CT: cocktail of 9 antibodies targeting cell surface neoantigens of 9 mutated cell surface proteins. Data are means ± SEM for 6 mice per group. Tumor growth data were compared via two-way ANOVAs.

[0033] Figure 4 shows the sequences of the synthesized peptides for mutated proteins described in Example 4.

[0034] Figures 5A-5C show the effects of treatment of B16-F10 melanoma tumor-bearing mice with a cocktail of 9 antibodies targeting 9 MSP cell surface neoantigens, as described in Example 4. The antibody cocktail treatment (0.2 mg per dose) significantly increased survival ofmice compared to the control mice groups. The values represent the mean ± SEM of six or surviving animals in each group. DPI = Days post-implantation.

[0035] Figure 6 shows the results of tumor re-challenge in the mice that had a durable complete response, as described in Example 4. After 6 months, 3 mice that had durable complete response to antibody cocktail plus PD1 inhibition were reimplanted with B16-F10 melanoma cells. No additional treatment was provided after reimplantation. Tumor growth was inhibited in the challenged mice compared to untreated control mice.

[0036] Figures 7A-7B show the effect of different treatments on CT-26 tumor growth in mice, as described in Example 5. Significant tumor growth retardations were observed in the mice treated with 10-antibody cocktail. Tumor retardation was enhanced with addition of PD1 i. Two-way ANOVA analysis of the data showed significantly different curves by treatment, time, and interaction. The values represent the mean ± SEM of six mice in each group. PD1 i = P D 1 inhibitor; nrlgG=normal rabbit IgG; DPI=Days post-implantation.DETAILED DESCRIPTION

[0037] The invention provides methods and compositions for treatment of cancer.

[0038] Unless defined otherwise herein, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Singleton, et al., Dictionary of Microbiology and Molecular Biology, second ed., John Wiley and Sons, New York (1994), and Hale & Markham, The Harper Collins Dictionary of Biology, Harper Perennial, NY (1991) provide one of skill with a general dictionary of many of the terms used in this invention. Any methods and materials similar or eguivalent to those described herein can be used in the practice or testing of the present invention.

[0039] The practice of the present invention will employ, unless otherwise indicated, conventional technigues of molecular biology (including recombinant technigues), microbiology, cell biology, and biochemistry, which are within the skill of the art. Such technigues are explained fully in the literature, for example, Molecular Cloning: A Laboratory Manual, second edition (Sambrook et al., 1989); Oligonucleotide Synthesis (M. J. Gait, ed., 1984; Current Protocols in Molecular Biology (F. M. Ausubel et al., eds., 1994); PCR: The Polymerase Chain Reaction (Mullis et al., eds., 1994); and Gene Transfer and Expression: A Laboratory Manual (Kriegler, 1990).

[0040] Numeric ranges provided herein are inclusive of the numbers defining the range.

[0041] Unless otherwise indicated, nucleic acids are written left to right in 5’ to 3’ orientation; amino acid seguences are written left to right in amino to carboxy orientation, respectively.Definitions

[0042] Unless defined otherwise herein, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which this invention belongs. Singleton, et al., Dictionary of Microbiology and Molecular Biology, second ed., John Wiley and Sons, New York (1994), and Hale & Markham, The Harper Collins Dictionary ofBiology, Harper Perennial, NY (1991) provide one of skill with a general dictionary of many of the terms used in this invention. Any methods and materials similar or equivalent to those described herein can be used in the practice or testing of the present invention.

[0043] Numeric ranges provided herein are inclusive of the numbers defining the range.

[0044] “A,” “an” and “the” include plural references unless the context clearly dictates otherwise.

[0045] “About” as used herein means plus or minus ten percent (10%) of a value. For example, “about 100” refers to any number from 90 to 110.

[0046] “Acceptable” in the context of a pharmaceutically acceptable carrier means that the carrier must be compatible with the active ingredient of the composition (and preferably, capable of stabilizing the active ingredient) and not deleterious to the subject to be treated.

[0047] The phrase “and / or,” as used herein in the specification and in the claims, should be understood to mean “either or both” of the elements so conjoined, i.e., elements that are conjunctively present in some cases and disjunctively present in other cases. Additional elements may optionally be present other than the elements specifically identified by the “and / or” clause, whether related or unrelated to those elements specifically identified, unless clearly indicated to the contrary. Thus, as a non-limiting example, a reference to “A and / or B,” when used in conjunction with open ended language such as “comprising” can refer, in one embodiment, to A without B (optionally including elements other than B); in another embodiment, to B without A (optionally including elements other than A); in yet another embodiment, to both A and B (optionally including other elements); etc.

[0048] An “antibody” is an immunoglobulin molecule capable of specific binding to a target, such as a carbohydrate, polynucleotide, lipid, polypeptide, etc., through at least one antigen recognition site, located in the variable region of the immunoglobulin molecule. As used herein, the term encompasses not only intact full-length antibodies, but also fragments thereof (such as Fab, Fab’, F(ab’)2, Fv), single chain (ScFv), mutants thereof, fusion proteins comprising an antibody portion, and any other modified configuration of the immunoglobulin molecule that comprises an antigen recognition site of the required specificity. An antibody includes an antibody of any class, such as IgG, IgA, or IgM (or sub-class thereof), and the antibody need not be of any particular class. Depending on the antibody amino acid sequence of the constant domain of its heavy chains, immunoglobulins can be assigned to different classes. There are five major classes of immunoglobulins: IgA, IgD, IgE, IgG, and IgM, and several of these may be further divided into subclasses (isotypes), e.g., lgG1 , lgG2, lgG3, lgG4, lgA1 and lgA2. The heavychain constant domains that correspond to the different classes of immunoglobulins are called alpha, delta, epsilon, gamma, and mu, respectively. The subunit structures and three- dimensional configurations of different classes of immunoglobulins are well known. The antibodies described herein can be either monoclonal or polyclonal. A “monoclonal antibody” refers to a homogenous antibody population and a “polyclonal antibody” refers to a heterogeneous antibody population. These two terms do not limit the source of an antibody or the manner in which it is made. An antibody herein can include various derivatives, including but not limited to the following: bispecific, glycosylated (sugar chain-modified), Fc-modified, orADC (antibody-drug conjugate) antibody . J n one example, an antibody used in the methods described herein is a chimeric antibody. Humanized antibodies refer to forms of non-human (e.g., murine) antibodies that are specific chimeric immunoglobulins, immunoglobulin chains, or antigen-binding fragments thereof that contain minimal sequence derived from non-human immunoglobulin. In another example, an antibody described herein is a chimeric antibody, which can include a heavy constant region and a light constant region from a human antibody. Chimeric antibodies refer to antibodies having a variable region or part of variable region from a first species and a constant region from a second species. Typically, in these chimeric antibodies, the variable regions of both light and heavy chains mimic the variable regions of antibodies derived from one species of mammals (e.g., a non-human mammal such as mouse, rabbit, or rat), while the constant portions are homologous to the sequences in antibodies derived from another mammal, such as human.

[0049] As used herein, the term “anti-neoantigen antibody” refers to any antibody capable of binding to a neoantigen. In some instances, the anti-neoantigen antibody can suppress the bioactivity of the neoantigen, and by extension, tumor growth.

[0050] “Arming” effector cells (white blood cells; WBCs) as described herein refers to binding polynucleotides or antibodies to the effector cells. The antibodies bind to the effector cells via Fc receptors. Other methods for arming effector cells include chemical methods, such as polyethylene glycol, or covalent or noncovalent designer molecules, e.g., bispecific antibodies, beads, or chemical modifications of the cells surface, including but not limited to biotin, click chemistry, oligosaccharide linkages, or sortase, diphtheria toxin transmembrane domain, hybridization methods, monoclonal antibodies, or genetic modifications, including but not limited to chimeric antigen receptors (CARs) that bind to a target cell antigen, CARs that bind to a target cell antigen and have an additional loop feature that is a handle for Rituximab to manipulate CAR cells, CARs that provide a binding site for an intermediate binding element, CD16, CD16 and a tumor specific T cell receptor (TCR), or CD64 external and DC16 transmembrane domains. Effector cells that are saturated with bound polynucleotides or antibodies are “fully armed.”

[0051] “Complementarity determining region” (CDR) refers a relatively short amino acid_sequence found in the variable regions of antibody molecules. The CDRs contain amino acid residues that determine the binding specificity of antibody molecules and make contact with a specific antigen.

[0052] The term “derived from” encompasses the terms “originated from,” “obtained from,” “obtainable from,” “isolated from,” and “created from,” and generally indicates that one specified material finds its origin in another specified material or has features that can be described with reference to another specified material.

[0053] “Fv” is an antibody fragment that contains a complete antigen-recognition and binding site. In a two-chain Fv species, this region consists of a dimer of one heavy chain variable domain (VH) and one light chain variable domain (VL) in tight, non-covalent association. In a singlechain Fv species, one heavy chain variable domain and one light chain variable domain can becovalently linked by a flexible polypeptide linker such that the light and heavy chains can associate in a dimeric structure analogous to that in a two-chain Fv species. It is in this configuration that the three CDRs of each variable domain interact to define an antigen-binding specificity on the surface of the VH-VL dimer. However, even a single variable domain (or half of a Fv comprising only 3 CDRs specific for an antigen) has the ability to recognize and bind antigen, although generally at a lower affinity than the entire binding site. A “Fab” fragment also contains the constant domain of the light chain and the first constant domain (CH1) of the heavy chain. Fab' fragments differ from Fab fragments by the addition of a few residues at the carboxy terminus of the heavy chain CH1 domain including one or more cysteines from the antibody hinge regions.

[0054] “Immunological activity” of an antibody refers to any of the following activities: (a) ability to bind antigen; (b) ability to inhibit the binding of antigen, or (c) ability to elicit a specific immune response, particularly an antibody (humoral) response, and / or a T cell response, and the effector functions that result therefrom. Included in an antibody response are antibody-mediated functions such as antibody-dependent cell cytotoxicity (ADCC) and complement-dependent cytotoxicity (CDC). T cell response includes T helper cell function, cytotoxic T cell function, inflammation inducer T cells, and T cell suppression. Immunological activity is measurable by using standard methods known in the art, such as radioimmunoassay (RIA), enzyme-linked immunoabsorbant assay (ELISA), complement fixation, opsonization, detection of T cell proliferation, and various51Cr release assays. These methods are known in the art. A compound able to elicit a specific immune response according to any of these criteria is referred to as “immunogenic.” “Immunogenicity” refers to a capability to elicit a specific humoral and / or cellular immune response.

[0055] An “individual” or “subject” refers to a vertebrate, typically a mammal, such as a human. The term “individual” or “subject” also refers to non-human mammals, such as, for example, dogs, cats, rodents, farm animals, livestock, sport animals, pets, primates, horses, mice, rats etc. A subject, such as a human subject, who needs or receives a treatment may be a patient, such as a mammalian patient, e.g., a human patient, having, at risk for, or suspected of having a target disease / disorder, such as a cancer or an infectious disease.

[0056] A “monoclonal antibody” refers to a homogeneous antibody population wherein the monoclonal antibody is comprised of amino acids (naturally occurring and / or non-naturally occurring) that are involved in the selective binding of an antigen. The term “monoclonal antibody” encompasses not only intact monoclonal antibodies and full-length monoclonal antibodies, but also fragments thereof (such as Fab, Fab’, F(ab’)2, Fv), single chain (ScFv), mutants thereof, fusion proteins comprising an antibody portion, and any other modified configuration of the immunoglobulin molecule that comprises an antigen recognition site of the required specificity and the ability to bind to an antigen (see definition of antibody). It is not intended to be limited as regards to the source of the antibody or the manner in which it is made (e.g., by hybridoma, phage selection, recombinant expression, transgenic animals, etc.). Insome embodiments of the methods described herein, a plurality of monoclonal antibodies is used, each of which binds to a different neoantigen on a MSP.

[0057] A “mutated” peptide refers to a short section ( / .e., amino acid sequence) of a protein that contains a mutation, relative to a reference protein, such as the amino acid sequence of the peptide in the same protein in non-cancer cells in the same individual.

[0058] A “neoantigen” refers to a mutated protein that may induce an immune response with high specificity to cancer cells.

[0059] The term “parenteral” as used herein includes subcutaneous, intracutaneous, intravenous, intramuscular, intraarticular, intraarterial, intrasynovial, intrasternal, intrathecal, intralesional, and intracranial injection or infusion techniques.

[0060] “Pharmaceutically acceptable” means approved by a regulatory agency of the Federal or a state government or listed in the U.S. Pharmacopoeia or other generally recognized pharmacopoeia for use in animals, and more particularly in humans.

[0061] “Pharmaceutically acceptable vehicle” or “pharmaceutically acceptable excipient” refers to a diluent, adjuvant, excipient or carrier with which a polynucleotide (e.g., mRNA) or protein (e.g., antibody, or fragment or derivative thereof) as described herein is administered.

[0062] As used herein, “polypeptide” refers to a composition comprised of amino acids and recognized as a protein by those of skill in the art. The conventional one-letter or three-letter codes for amino acid residues are used herein. The terms “polypeptide” and “protein” are used interchangeably herein to refer to polymers of amino acids of any length. The polymer may be linear or branched, it may contain modified amino acids, and it may be interrupted by non-amino acids. The terms also encompass an amino acid polymer that has been modified naturally or by intervention; for example, disulfide bond formation, glycosylation, lipidation, acetylation, phosphorylation, or any other manipulation or modification, such as conjugation with a labeling component. Also included within the definition are, for example, polypeptides containing one or more analogs of an amino acid (including, for example, unnatural amino acids, etc.), as well as other modifications known in the art.

[0063] As used herein, the term “polynucleotide” refers to a polymeric form of nucleotides of any length and any three-dimensional structure and single- or multi-stranded (e.g., single-stranded, double-stranded, triple-helical, etc.), which contain deoxyribonucleotides, ribonucleotides, and / or analogs or modified forms of deoxyribonucleotides or ribonucleotides, including modified nucleotides or bases or their analogs. Because the genetic code is degenerate, more than one codon may be used to encode a particular amino acid, and the present invention encompasses polynucleotides which encode a particular amino acid sequence. Any type of modified nucleotide or nucleotide analog may be used, so long as the polynucleotide retains the desired functionality under conditions of use, including modifications that increase nuclease resistance (e.g., deoxy, 2’-O-Me, phosphorothioates, etc.). Labels may also be incorporated for purposes of detection or capture, for example, radioactive or nonradioactive labels or anchors, e.g., biotin. The term polynucleotide also includes peptide nucleic acids (PNA). Polynucleotides may be naturally occurring or non-naturally occurring. The terms “polynucleotide,” “nucleic acid,” and“oligonucleotide” are used herein interchangeably. Polynucleotides may contain RNA, DNA, or both, and / or modified forms and / or analogs thereof. A sequence of nucleotides may be interrupted by non-nucleotide components. One or more phosphodiester linkages may be replaced by alternative linking groups. These alternative linking groups include, but are not limited to, embodiments wherein phosphate is replaced by P(O)S (“thioate”), P(S)S (“dithioate”), (O)NR2(“amidate”), P(O)R, P(O)OR’, CO or CH2(“formacetal”), in which each R or R’ is independently H or substituted or unsubstituted alkyl (1-20 C) optionally containing an ether (- O-) linkage, aryl, alkenyl, cycloalkyl, cycloalkenyl or araldyl. Not all linkages in a polynucleotide need be identical. Polynucleotides may be linear or circular or contain a combination of linear and circular portions.

[0064] The terms “recovered,” “isolated,” “purified,” and “separated” as used herein refer to a material (e.g., a protein, nucleic acid, or cell) that is removed from at least one component with which it is naturally associated. For example, these terms may refer to a material which is substantially or essentially free from components which normally accompany it as found in its native state, such as, for example, an intact biological system.

[0065] An epitope that “specifically binds” or “preferentially binds” (used interchangeably herein) to an antibody is a term well understood in the art, and methods to determine such specific or preferential binding are also well known in the art. A molecule is said to exhibit “specific binding” or “preferential binding” if it reacts or associates more frequently, more rapidly, with greater duration and / or with greater affinity with a particular cell or substance than it does with alternative cells or substances. An antibody “specifically binds” or “preferentially binds” to a target if it binds with greater affinity, avidity, more readily, and / or with greater duration than it binds to other substances. For example, an antibody that specifically or preferentially binds to an epitope is an antibody that binds this epitope with greater affinity, avidity, more readily, and / or with greater duration than it binds to other epitopes. It is also understood by reading this definition that, for example, an antibody (or moiety or epitope) that specifically or preferentially binds to a first target may or may not specifically or preferentially bind to a second target. As such, “specific binding” or “preferential binding” does not necessarily require (although it can include) exclusive binding. Generally, but not necessarily, reference to binding means preferential binding.

[0066] “Treating” or “treatment” of any disease or disorder refers, in one embodiment, to ameliorating the disease or disorder ( / .e., arresting or reducing the development of the disease or at least one of the clinical symptoms thereof). In another embodiment “treating” or “treatment” refers to ameliorating at least one physical parameter, which may not be discernible by the subject. In yet another embodiment, “treating” or “treatment” refers to modulating the disease or disorder, either physically (e.g., stabilization of a discernible symptom), physiologically (e.g., stabilization of a physical parameter), or both.

[0067] “Therapeutically effective amount” means the amount of a compound that, i.e., a protein as described herein, that when administered to an individual for treating a disease or condition, is sufficient to effect such treatment for the disease or condition or to reduce severity of oreliminate at least one symptom of the disease or condition. “Therapeutically effective amount” means that amount of the compound that will elicit the biological or medical response of a subject that is being sought by a medical doctor or other clinician. The “therapeutically effective amount” can vary depending on the compound, the disease and its severity, and the age, weight, etc., of the subject to be treated.

[0068] The term “unit dosage” or “unit dosage form” refers to a physically discrete units suitable as a unitary dosages for an individual to whom administered, each unit containing a predetermined quantity of active material calculated to produce the desired therapeutic effect, in association with a suitable pharmaceutical excipient.

[0069] A “variable region” of an antibody refers to the variable region of the light chain or the variable region of the heavy chain, either alone or in combination.

[0070] A “vector” polynucleotide refers to a polynucleotide that encodes a protein. For example, a vector antibody polynucleotide (e.g., vector antibody mRNA) encodes the amino acid sequence of an antibody, or a fragment or derivative thereof that retains the binding specificity of the antibody. In another example, a vector peptide polynucleotide (e.g., vector peptide mRNA) encodes the amino acid sequence of a mutated cell surface related (MSP) peptide.Mutated cell surface related proteins

[0071] Neoantigens, or tumor-specific antigens, are protein mutations (one or more mutation in a protein amino acid sequence) that are present in one or more tumor cells, but that are not expressed or are expressed at low levels in normal non-cancerous tissue. As such, neoantigens arise from one or more tumor-specific mutations, and may induce an immune response with high specificity to cancer cells. The location at the cell surface makes mutated cell surface related protein (MSP) neoantigens accessible to antibodies, and antibodies that target these neoantigens have minimal cross-reactive binding to normal proteins. The set of MSPs collectively represents the neoantigen space available for antibody binding on a given cancer. Sequencing data identifies which MSPs harbor missense mutations, but the expression of individual MSPs is variable and unpredictable. Effective antibody-mediated cytotoxicity depends upon achieving sufficient binding of antibodies to cancer cells. In some embodiments, one MSP neoantigen may not provide sufficient antibody binding sites, and antibodies targeting only one MSP may not achieve the critical level of antibody binding required to mediate cancer cell cytotoxicity. Intentionally over-targeting multiple MSP neoantigens expands the number of antibody binding sites on target tumor cells. Antibodies are screened against target epitopes on MSPs, e.g., against MSP peptides, which contain the mutated cell surface related protein sequences.

[0072] Mutation-derived neoantigens can arise from: point mutations, non-synonymous mutations leading to substitution of different amino acids in amino acid sequence of the mutated protein versus the native, non-mutated protein; read-through mutations in which a stop codon is modified or deleted, leading to translation of a longer protein with a novel tumor-specific sequence at the C-terminus; splice site mutations that lead to the inclusion of an intron in themature mRNA and thus a unique tumor-specific protein sequence; chromosomal rearrangements that give rise to a chimeric protein with tumor-specific sequences at the junction of two proteins ( / .e., gene fusion); frameshift mutations or deletions that lead to a new open reading frame with a novel tumor-specific protein sequence; and / or translocations. In one embodiment, the mutation is a single amino acid substitution in the amino acid sequence of a protein. In other embodiments, the mutation is 2, 3, 4, 5, 6, 7, 8, 9, or 10 amino acid substitutions.

[0073] Cell surface related proteins are proteins that are expressed at the cell surface (e.g., constituents of the cell membrane) or are secreted from the cell. Mutated cell surface related proteins (MSPs) are cell surface related proteins that contain one or more mutation in cancer cells versus non-cancer cells in an individual (mutations that are present in cancer cells in the individual but not present in non-cancer cells in the individual), i.e., neoantigens that are located at the cell surface, such as within the cell membrane, or that are secreted. Unlike intracellular proteins, both membrane constitutive and secreted proteins are available for external interaction with antibodies. MSP peptides have an amino acid sequence of at least a portion of the full length MSP, and contain one or more mutation in the amino acid sequence of the peptide, relative to the amino acid sequence of the peptide in the cell surface related protein in non- cancer cells.

[0074] In some embodiments, the length of a neoantigen MSP peptide that is used to induce an immune response (i.e., induce generation of anti-neoantigen MSP antibodies) in an individual in methods described herein is 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, 20, 21 , 22, 23, 24, 25, 26, 27, 28, 29, 30, 31 , 32, 33, 34, 35, 36, 37, 38, 39, or 40, amino acids long. In some embodiments, the MSP peptide is 5-10, 5-11 , 5-12, 5-13, 5-14, 5-15, 5-16, 5-17, 5-18, 5-19, 5- 20, 6-10, 6-11 , 6-12, 6-13, 6-14, 6-15, 6-16, 6-17, 6-18, 6-19, 6-20, 7-10, 7-11 , 7-12, 7-13, 7-14, 7-15, 7-16, 7-17, 7-18, 7-19, 7-20, 8-10, 8-11 , 8-12, 8-13, 8-14, 8-15, 8-16, 8-17, 8-18, 8-19, 8- 20, 9-10, 9-11 , 9-12, 9-13, 9-14, 9-15, 9-16, 9-17, 9-18, 9-19, 9-20, 10-15, 1-16, 10-17, 10-18, 10-19, 10-20, 5-25, 10-35, or 15-40 amino acids in length, or any of about or at least about 5%, 10%, 15%, 20%, 25%, 30%, 35%, 40%, 45%, 50%, 55%, 60%, 65%, 70%, 75%, 80%, 90%, or 95% of the amino acid sequence length of the full length protein.

[0075] In some embodiments, a multimeric polypeptide is used to generate an immune response. The multimeric polypeptide may contain at least two, three, four, five, or more MSP peptide sequences, i.e., either the same or different MSP peptide sequences, e.g., having any of the amino acid lengths described herein, within a single polypeptide.Polynucleotides that encode MSP peptides

[0076] In some embodiments of the methods described herein, a polynucleotide that encodes an amino acid sequence of at least a portion of one or more MSP peptide, is administered to an individual to generate an immune response in the individual to the mutated amino acid sequence of the MSP neoantigen. The immune response may include antibodies that specifically bind to the MSP peptide. The polynucleotide may be an mRNA polynucleotide,which is translated by the individual to produce the MSP peptide. Alternatively, the polynucleotide may be a DNA polynucleotide (see, e.g., Khoshnejad, M., et al. (2019) Mol Ther 27(1):188-199, PMID: 30449662; Duperret, E.K., et al. (2018) Cancer Res 78(22):6363-6370, PMID:30287678; Hollevoet, K„ et al. (2018) Oncotarget 9(17):13623-13636, PMID: 29568382; Muthumani, K., et al. (2017) Cancer Immunol Immunother 66(12):1577-1588, PMID: 28819703) or a different form of RNA, such as self-replicating RNA (see, e.g., Morse, M.A., et al. (2023) Cancer Gene Ther 2023 10 Feb:1-9, PMID: 36765179).

[0077] A polynucleotide that encodes an MSP peptide may encode two or more different MSP peptides, such as at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten different MSP peptides. A polynucleotide that encodes an MSP peptide may encode one or more copies of the same MSP peptide, such as one, at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten copies of the same MSP peptide.

[0078] One or more polynucleotide encoding one or more MSP peptide may be administered to an individual, thereby inducing an immune response by the individual to the MSP peptide(s). The immune response may be assessed by determining titer of antibodies to the MSP peptide(s), e.g., in a blood sample that is obtained from the individual.

[0079] The polynucleotide(s) may be administered to an individual by any suitable route, including intramuscularly, subcutaneously, intradermally, intravenously, or via inhalation.Antibodies that bind MSPs

[0080] Anti-MSP antibodies that specifically bind MSPs of cancer cells of an individual may be produced by injection of MSP peptide(s) into an individual in whom mutated cell surface related protein(s) that contain the MSP peptide(s) have been identified in cancer cells of the individual or by injection of the MSP peptide(s) into another individual, i.e., a second individual, such as a non-human animal, e.g., a rodent (e.g., a mouse) or a rabbit. In some embodiments, anti-MSP antibodies that are produced by an individual in whom mutated cell surface related proteins in cancer cells of the individual are identified may be sequenced to determine the amino acid sequences of the antibody. In some embodiments, an MSP peptide is injected into another individual, i.e., a second individual, such as a non-human animal, e.g., a rodent (e.g., a mouse) or a rabbit., and antibodies that are produced by the other individual are identified and sequenced.

[0081] For example, B cells of the individual, or of another individual individual into whom MSP peptides are injected, i.e., a second individual, such as a non-human animal, e.g., a rodent (e.g., a mouse) or a rabbit, into whom an MSP peptide has been injected to produce anti-MSP antibodies that specifically bind to the MSP peptide, may be isolated by flow cytometry, or other methods that allow detection and isolation of individual B cells. Single B cell sequencing of the antibody that is produced by the B cell may be performed. Selection of B cells may be via detection of anti B-cell antibodies that are secreted by individual B cells, or by detection of the B cell receptor (anti-MSP antibodies on the cell membrane). For detection of secreted antibodies,the B cell may be immobilized or held in place in a microcontainer, a microbubble, or on a surface (e.g., a planar surface). A bead or other surface may collect and identify the secreted antibodies.

[0082] Other options for sequencing of the anti-MSP antibodies include, but are not limited to: sequencing of bulk B cells and using bioinformatics to identify the clones that have recently expanded; library formation from bulk B cells after an immune response has been induced, e.g., using phage display, yeast display, or other display technologies; and proteomics, e.g., antibodies that bind to MSPs are affinity enriched from a blood sample and then analyzed to determine the antibody protein sequences.Polynucleotides that encode anti-MSP antibodies

[0083] In some embodiments of the methods described herein, a polynucleotide that encodes an amino acid sequence of an anti-MSP antibody, or a polynucleotide that encodes an anti-MSP antibody fragment or derivative thereof that retains the ability to bind to an MSP peptide to which the antibody specifically binds, is administered to an individual for treatment of cancer. The polynucleotide may be an mRNA polynucleotide, which is translated by the individual to produce the anti-MSP antibody, or fragment or derivative thereof. Alternatively, the polynucleotide may be a DNA polynucleotide or a different form of RNA.

[0084] A polynucleotide that encodes an anti-MSP antibody, or fragment or derivative thereof, may encode two or more different anti-MSP antibodies, or fragments or derivatives thereof, such as at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten different antibodies, or fragments or derivatives thereof. A polynucleotide that encodes an anti-MSP antibody, or fragment or derivative thereof, may encode one or more copies of the same anti-MSP antibody, or fragment or derivative thereof, such as one, at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten copies of the same anti-MSP antibody, or fragment or derivative thereof. A polynucleotide that encodes an anti-MSP antibody, or fragment or derivative thereof, may encode one or more copies of two or more different anti-MSP antibodies, or fragments or derivatives thereof, such as one, at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten copies of each of at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten different antibodies, or fragments or derivatives thereof.

[0085] In various embodiments, the polynucleotide may encode a heavy chain and / or a light chain of an antibody, or fragments thereof, or may encode antigen-binding fragments of an antibody, such as, but not limited to, Fab, Fab’, F(ab’)2, Fv, single chain (ScFv), mutants thereof, fusion proteins including an antibody portion, and any other modified configuration of an immunoglobulin molecule that includes an antigen recognition site for the MSP peptide, such as a polypeptide that includes the complementarity determining regions (CDRs), (e.g., one, two, three, four, five, or six CDRs), of an antibody that specifically binds the MSP peptide.

[0086] The polynucleotide(s) may be administered to an individual by any suitable route, including intramuscularly, subcutaneously, intradermally, intravenously, or via inhalation.Production of anti-MSP antibodies

[0087] In some embodiments, anti-MSP antibodies, or MSP-binding fragments of anti-MSP antibodies, or derivatives thereof, may be produced in a system that is extrinsic to the individual in which the immune response to the MSP peptide has been generated and in whom the antibodies will be administered for treatment of cancer.

[0088] The extrinsic system may involve production of the antibodies, or fragments or derivatives thereof, in a phage display system. Phage display involves integration of a polynucleotide sequence that encodes an antibody, or fragment or derivative thereof, into a bacteriophage, e.g., filamentous bacteriophage, which results in its expression on the surface of the bacteriophage capsid. (See, e.g., Ledsgaard, et al. (2018) Toxins 10(6): 236)

[0089] Alternatively, the extrinsic system may include production of the antibody, or fragment or derivative thereof, in a non-human animal, such as, for example, a rodent (e.g., mouse, rat), a rabbit, a chicken, a goat, a pig, a horse, or a bovine. For example, a polynucleotide (e.g., mRNA) that encodes the antibody, or fragment or derivative thereof, may be administered to a non-human animal, and the antibody, or fragment or derivative thereof, is expressed (e.g., translated) and recovered in the plasma of the non-human animal.

[0090] Another extrinsic system includes expression in a cell line, such as, but not limited to, CHO cells, following transient transfection or stable transfection pools. In some embodiments, an animal, such as a mouse, may be vaccinated, hybridomas generated and then screened for production of desired antibodies, the hybridoma that produces a desired antibody sequenced, and then the binding portion of the antibody (e.g., mouse) onto a human antibody framework, thereby producing a chimeric antibody. Alternatively, humanized mice may be used for vaccination, to render human antibodies. Once the sequence is known, then production of antibodies may proceed in accordance with any of methods described above. For example, in some embodiments, an animal, such as, but not limited to, a mouse, may be vaccinated with a MSP peptide that did not induce a high titer immunological response in an individual in methods described herein, and antibodies that are produced in the animal may be sequenced and used in conjunction with antibodies that are produced by the individual to other MSP peptides in methods for treating cancer as described herein. In other embodiments, an animal, such as, but not limited to, a mouse, may be vaccinated with a MSP peptide, e.g., a MSP peptide that did not induce a high titer immunological response in an individual in methods described herein, and antibodies that are produced in the animal may be sequenced and the sequences used to produce a polynucleotide that encodes amino acid sequences of the antibody, and the polynucleotide that encodes the antibody sequences administered to the individual in methods for treating cancer as described herein.Pharmaceutical compositions

[0091] The present disclosure provides pharmaceutical compositions that include polynucleotides (e.g., mRNA polynucleotides) and / or anti-MSP antibodies, or fragments or derivatives thereof, as described herein, and uses of such for neutralizing tumor (e.g., tumor cell) bioactivity. For example, the polynucleotides and / or antibodies described herein may be used to treat cancer in a subject. Polynucleotides and / or antibodies as described herein may be formulated in a pharmaceutical composition that includes one or more pharmaceutically acceptable excipient.

[0092] Injectable compositions may contain various carriers such as vegetable oils, dimethylactamide, dimethyformamide, ethyl lactate, ethyl carbonate, isopropyl myristate, ethanol, and polyols (glycerol, propylene glycol, liquid polyethylene glycol, and the like). For intravenous injection, water soluble polynucleotides and / or antibodies can be administered by the drip method, whereby a pharmaceutical formulation containing the polynucleotide and / or anti-neoantigen cocktail, and / or effector cells armed with polynucleotides and / or anti-neoantigen antibodies and optionally at least one immune checkpoint inhibitor, and a physiologically acceptable excipient, is infused. Physiologically acceptable excipients may include, for example, 5% dextrose, 0.9% saline, Ringer’s solution or other suitable excipients.Intramuscular preparations, e.g., a sterile formulation of a suitable soluble salt form of the polynucleotide and / or anti-neoantigen antibody, polynucleotide and / or anti-neoantigen antibody cocktail, and / or effector cells armed with polynucleotides and / or anti-neoantigen antibodies, can be dissolved and administered in a pharmaceutical excipient such as Water-for-lnjection, 0.9% saline, or 5% glucose solution. “Acceptable” means that the carrier must be compatible with the active ingredient of the composition (and preferably, capable of stabilizing the active ingredient) and not deleterious to the subject to be treated. Pharmaceutically acceptable excipients (carriers) including buffers, which are well known in the art. See, e.g., Remington: The Science and Practice of Pharmacy 20thEd. (2000) Lippincott Williams and Wilkins, Ed. K. E. Hoover.

[0093] The pharmaceutical compositions to be used in the present methods can include pharmaceutically acceptable carriers, excipients, or stabilizers in the form of lyophilized formulations or aqueous solutions. (Remington: The Science and Practice of Pharmacy 20thEd. (2000) Lippincott Williams and Wilkins, Ed. K. E. Hoover). Acceptable carriers, excipients, or stabilizers are nontoxic to recipients at the dosages and concentrations used, and may include buffers such as phosphate, citrate, and other organic acids; antioxidants including ascorbic acid and methionine; preservatives (such as octadecyldimethylbenzyl ammonium chloride; hexamethonium chloride; benzalkonium chloride, benzethonium chloride; phenol, butyl or benzyl alcohol; alkyl parabens such as methyl or propyl paraben; catechol; resorcinol; cyclohexanol; 3- pentanol; and m-cresol); low molecular weight (less than about 10 residues) polypeptides; proteins, such as serum albumin, gelatin, or immunoglobulins; hydrophilic polymers such as polyvinylpyrrolidone; amino acids such as glycine, glutamine, asparagine, histidine, arginine, or lysine; monosaccharides, disaccharides, and other carbohydrates including glucose, mannose, or dextrans; chelating agents such as EDTA; sugars such as sucrose, mannitol, trehalose or sorbitol; salt-forming counter-ions such as sodium; metal complexes (e.g. Zn-proteincomplexes); and / or non-ionic surfactants such as TWEEN™, PLURONICS™ or polyethylene glycol (PEG).

[0094] In some examples, the pharmaceutical composition described herein includes liposomes containing the polynucleotides (e.g., mRNA polynucleotides) and / or anti-MSP antibodies, or fragments or derivatives thereof, which can be prepared by methods known in the art, such as described in Epstein, et al. (1985) Proc. Natl. Acad. Sci. USA 82:3688; Hwang, et al. (1980) Proc. Natl. Acad. Sci. USA 77:4030; and U.S. Pat. Nos. 4,485,045 and 4,544,545. Liposomes with enhanced circulation time are disclosed in U.S. Pat. No. 5,013,556. Particularly useful liposomes can be generated by the reverse phase evaporation method with a lipid composition including phosphatidylcholine, cholesterol and PEG-derivatized phosphatidylethanolamine (PEG-PE). Liposomes may be extruded through filters of defined pore size to yield liposomes with the desired diameter.

[0095] The polynucleotides (e.g., mRNA polynucleotides) or anti-MSP antibodies, or fragments or derivatives thereof, may also be entrapped in microcapsules prepared, for example, by coacervation techniques or by interfacial polymerization, for example, hydroxymethylcellulose or gelatin-microcapsules and poly-(methylmethacylate) microcapsules, respectively, in colloidal drug delivery systems (for example, liposomes, albumin microspheres, microemulsions, nanoparticles and nanocapsules) or in macroemulsions. Such techniques are known in the art (see, e.g., Remington, The Science and Practice of Pharmacy 20thEd. Mack Publishing (2000)).

[0096] In other examples, the pharmaceutical composition described herein can be formulated in sustained-release format. Suitable examples of sustained-release preparations include semipermeable matrices of solid hydrophobic polymers containing the antibody, which matrices are in the form of shaped articles, e.g. films, or microcapsules. Examples of sustained-release matrices include polyesters, hydrogels (for example, poly(2-hydroxyethyl-methacrylate), or poly(vinyl alcohol)), polylactides (U.S. Pat. No. 3,773,919), copolymers of L-glutamic acid and 7 ethyl-L-glutamate, non-degradable ethylene-vinyl acetate, degradable lactic acid-glycolic acid copolymers such as the LUPRON DEPOT™ (injectable microspheres composed of lactic acid- glycolic acid copolymer and leuprolide acetate), sucrose acetate isobutyrate, and poly-D-(-)-3- hydroxybutyric acid.

[0097] The pharmaceutical compositions to be used for in vivo administration to an individual must be sterile. This may be readily accomplished by, for example, filtration through sterile filtration membranes. Therapeutic compositions are generally placed into a container having a sterile access port, for example, an intravenous solution bag or vial having a stopper pierceable by a hypodermic injection needle.

[0098] Suitable emulsions may be prepared using commercially available fat emulsions, such as Intralipid™, Liposyn™, Infonutrol™, Lipofundin™, and Lipiphysan™. The active ingredient may be either dissolved in a pre-mixed emulsion composition or alternatively it may be dissolved in an oil (e.g., soybean oil, safflower oil, cottonseed oil, sesame oil, corn oil or almond oil) and an emulsion formed upon mixing with a phospholipid (e.g. egg phospholipids, soybean phospholipids or soybean lecithin) and water. It will be appreciated that other ingredients maybe added, for example glycerol or glucose, to adjust the tonicity of the emulsion. Suitable emulsions will typically contain up to 20% oil, for example, about 5% to about 20%. The fat emulsion can comprise fat droplets of about 0.1 pm to about 1 .0 pm diameter, particularly about 0.1 pm to about 0.5 pm diameter, and have a pH in the range of about 5.5 to about 8.0.

[0099] The emulsion compositions can be those prepared by mixing polynucleotides (e.g., mRNA polynucleotides) and / or anti-MSP antibodies, or fragments or derivatives thereof, with a lipid formulation, such as Intralipid™ or the components thereof (e.g., soybean oil, egg phospholipids, glycerol, and water).

[0100] In some embodiments, a composition, e.g., a pharmaceutical composition, is provided that includes one, two, or more polynucleotide(s) that encodes a MSP peptide or an anti-MSP antibody, or fragment or derivative thereof, and / or that includes one, two, or more anti- neoantigen antibodies, or fragment(s) or derivative(s) thereof (or effector cells armed with the polynucleotides and / or anti-MSP antibodies) and a pharmaceutically acceptable carrier (excipient). In some embodiments, the composition includes two or more (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, or 20) polynucleotides that encode MSP peptides or anti-MSP antibodies), or fragments or derivatives thereof, and / or includes two or more (e.g., 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, or 20) anti-MSP antibodies, or fragments or derivatives thereof. In some embodiments, the two or more polypeptides or antibodies are present in the composition in equal concentrations. In other embodiments, the two or more polypeptides or antibodies are not present in the composition in equal concentrations. In some embodiments, the at least two polypeptides or antibodies encode or are directed to to the same neoantigen. In some embodiments, the at least two polypeptides or antibodies encode or are directed to different epitopes of the same neoantigen. In other embodiments, the at least two polypeptides or antibodies do not all encode or are not all directed to directed to the same neoantigen. In another embodiment, the at least two polypeptides or antibodies all encode or are all directed to different neoantigens.Effector cells

[0101] In some embodiments, effector cells may be coated or transfected (“armed”) with polynucleotides that encode MSP peptides and / or that encode anti-MSP antibodies, or fragments or derivatives of anti-MSP antibodies thereof that retain the ability to specifically bind to the MSP peptide that was used for generation of an immune response in an individual as described herein, and / or with anti-MSP antibodies or fragments or derivatives of anti-MSP antibodies thereof that retain the ability to specifically bind to the MSP peptide that was used for generation of an immune response in an individual as described herein, ex vivo, which reduces the amount of polynucleotide or antibody required to generate an immune response or cause tumor inhibition. Previous studies have demonstrated multiple types of effector cells derived from spleen, peripheral blood, marrow, and peritoneum can mediate cytotoxicity. Multiple types of effector cells can be armed since they are readily available from peripheral blood and do notrequire ex vivo growth expansion. For example, the effector cells may include one or more of natural killer (NK) cells, T cells, B cells, monocytes, macrophages, and neutrophils.

[0102] Therefore, the polynucleotides and / or anti-neoantigen antibodies described herein may be coated onto or transfected into effector cells in vitro for administration an individual. The resulting effector cells are referred to as “armed effector cells.” Armed effector cells may include any number of polynucleotides as described herein (e.g., polynucleotides that encode MSP peptides and / or polynucleotides that encode anti-MSP antibodies, or fragments or derivatives of anti-MSP antibodies thereof that retain the ability to specifically bind to the MSP peptide that was used for generation of an immune response in an individual as described herein), such as any of 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, 20 polynucleotides as described herein, and / or may include any number of anti-neoantigen antibodies, or fragments or derivatives of anti-MSP antibodies thereof that retain the ability to specifically bind to the MSP peptide that was used for generation of an immune response in an individual as described herein, such as any of 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, 20, or more anti-neoantigen antibodies, or fragments or derivatives thereof. Examples of effector cells include, but are not limited to, white blood cells (leukocytes), such as natural killer (NK) cells, neutrophils, T cells, B cells, monocytes, macrophages, and neutrophils. In some embodiments, a single type of effector cell is used. In other embodiments, a combination of effector cells is used. The effector cells may be coated (“armed”) with the polynucleotides and / or antibodies using any method known in the art. For example, the cells may be incubated on ice with a polynucleotide and / or anti-neoantigen antibody, or fragment or derivative thereof (or cocktail of polynucleotides and / or anti-neoantigen antibodies, or fragments or derivatives thereof).

[0103] Polynucleotides (e.g., mRNA) may be loaded on isolated effector cells in vitro via lipid or other particles that contain the polynucleotides. Alternatively, physical methods such as electric current may be used.

[0104] In some embodiments, effector cells may be isolated from whole blood, e.g., from whole blood of the individual to whom the effector cells will be administered or from whole blood from a different individual. In one embodiment, effector cells are obtained from the subject using a leukocyte reduction filter, as described in U.S. Application No. 18 / 475,031 , and in Shukla, G.S., et al. (2021) Journal of Immunological Methods 499:113157, “Preparation of clinical-grade WBCs using leukocyte reduction filters,” which are incorporated herein by reference in their entireties. Other methods for obtaining effector cells include leukapheresis.

[0105] In some embodiments, effector cells secrete anti-MSP antibodies or fragments or derivatives thereof that retain the ability to bind to the MSP peptide, expressed from polynucleotides with which the effector cells are armed that encode the antibodies or binding fragments or derivatives thereof as described herein.

[0106] In some embodiments, effector cells that are armed with polynucleotides that encode anti- MSP antibodies or fragments or derivatives thereof that retain the ability to bind to the MSP peptide, express the anti-MSP antibodies or binding fragments or derivatives such that the expressed antibodies or fragments or derivatives thereof are not secreted, but are directed to bewithin cell surface membrane of the effector cell. For example, the effector cells may be isolated from whole blood as described herein, and the polynucleotide, e.g., vector polynucleotide, induces the effector cell to generate the antibodies, or fragments or derivatives thereof. Instead of secreting the antibodies or binding fragments or derivatives thereof, the polynucleotide, e.g., vector polynucleotide, guides the antibodies or binding fragments or derivatives to the surface of the effector cell.Administration of armed effector cells

[0107] Conventional methods, known to those of ordinary skill in the art of medicine, can be used to administer the armed effector cells to a subject, e.g., a subject in need of treatment for a cancer. A pharmaceutical composition that includes the effector cells can be administered via conventional routes, e.g., administered intravenously or parenterally. In one embodiment, the composition is administered via intratumoral injection.

[0108] In one embodiment, a composition is administered via site-specific or targeted local delivery techniques. Examples of site-specific or targeted local delivery techniques include various implantable depot sources of the composition or local delivery catheters, such as infusion catheters, an indwelling catheter, or a needle catheter, synthetic grafts, adventitial wraps, shunts and stents or other implantable devices, site specific carriers, direct injection, or direct application. See, e.g., PCT Publication No. WO 00 / 53211 and U.S. Pat. No. 5,981 ,568.

[0109] In some embodiments, the method further comprises administering an immune checkpoint inhibitor to the subject having cancer. In some embodiments, the armed effector cells and an immune checkpoint inhibitor are administered simultaneously. In some embodiments, the armed effector cells are administered prior to administration of an immune checkpoint inhibitor. In some embodiments, the immune checkpoint inhibitor is an antibody selected from the group consisting of anti-CTLA4 antibodies, anti-PD-1 antibodies, anti-PD-L1 antibodies, anti-PD-L2 antibodies anti-TIM-3 antibodies, anti-LAG3 antibodies, anti-B7H3 antibodies, anti-B7H4 antibodies, anti-BTLA antibodies, and anti-B7H6 antibodies.Methods for treatment of cancer

[0110] Methods are provided herein for treatment of cancer. The methods include administration of a therapeutically effective amount of one or more polynucleotide e.g., mRNA polynucleotide) that encodes an antibody that specifically binds to a MSP peptide, administration of a therapeutically effective amount of one or more polynucleotide (e.g., mRNA polynucleotide) that encodes a MSP peptide, and / or administration of a therapeutically effective amount of one or more anti-MSP antibody, or a MSP-binding fragment or derivative thereof, to an individual for treatment of cancer in the individual. The one or more polynucleotide and / or antibody may be administered in the form of a pharmaceutical composition that includes one or more pharmaceutically acceptable excipient. In some embodiments, any of 1 , 2, 3, 4, 5, 6, 7, 8, 9, 10, 11 , 12, 13, 14, 15, 16, 17, 18, 19, 20, or more MSPs (mutations in the amino acid sequences of cell surface related proteins that are specific to cancer cells in the individual, relative to theamino acid sequence of the cell surface related protein in non-cancer cells) are targeted by administration of polynucleotides (e.g., mRNA polynucleotides) and / or antibodies as described herein.

[0111] In some embodiments, the method includes administration of a therapeutically effective amount of one or more anti-MSP antibody, or MSP-binding fragment or derivative thereof, to an individual in need thereof, wherein the antibody is produced in a system that is extrinsic to the individual from polynucleotide sequences that are generated by: administering to an individual at least one polynucleotide (e.g., mRNA polynucleotide) that encodes a MSP peptide that is specifically expressed in cancer cells; assessing the immune response in the individual to the MSP peptide that is expressed (e.g., translated) from the polynucleotide; sequencing one or more antibody that is produced by the individual that specifically binds to the MSP peptide; and using the amino acid sequences of the antibody to produce the antibody, or a MSP-binding fragment or derivative thereof. The antibody thus produced may be administered to the individual for treatment of cancer (e.g., binding to MSP expressed in cancer cells in the individual for treatment of the cancer). The one or more anti-MSP antibody, or MSP-binding fragment or derivative thereof, may be administered in the form of a pharmaceutical composition that includes one or more pharmaceutically acceptable excipient. In some embodiments, the antibodies are administered to the individual in the form of armed effector cells, e.g., armed natural killer (NK) cells, T cells, B cells, monocytes / macrophages, neutrophils, or combinations thereof, for example, a pharmaceutical composition that includes armed effector cells and one or more pharmaceutically acceptable excipient.

[0112] In some embodiments, the method includes administration of a therapeutically effective amount of one or more polynucleotide (e.g., mRNA polynucleotide) that encodes an anti-MSP antibody, or MSP-binding fragment or derivative thereof, to an individual in need thereof, wherein the polynucleotide is generated in a method that includes: administering to an individual at least one polynucleotide (e.g., mRNA polynucleotide) that encodes a MSP peptide that is specifically expressed in cancer cells; assessing the immune response in the individual to the MSP peptide that is expressed (e.g., translated) from the polynucleotide; sequencing one or more antibody that is produced by the individual that specifically binds to the MSP peptide; and using the amino acid sequences of the antibody to produce a polynucleotide (e.g., mRNA polynucleotide) that encodes the antibody, or a MSP-binding fragment or derivative thereof. The polynucleotide thus produced may be administered to the individual for treatment of cancer (expression (e.g., translation) of an anti-MSP antibody, or MSP-binding fragment or derivative thereof, that specifically binds to an MSP expressed in cancer cells in the individual for treatment of the cancer). The one or more polynucleotide may be administered in the form of a pharmaceutical composition that includes one or more pharmaceutically acceptable excipient. In some embodiments, the polynucleotides are administered to in the form of armed effector cells, e.g., armed natural killer (NK) cells, T cells, B cells, monocytes / macrophages, neutrophils, or combinations thereof, for example, a pharmaceutical composition that includes armed effector cells and one or more pharmaceutically acceptable excipient.

[0113] In some embodiments, the method includes administration of a therapeutically effective amount of one or more polynucleotide (e.g., mRNA polynucleotide) that encodes an MSP peptide to an individual in need thereof. The MSP peptide is expressed (e.g., translated) from the polynucleotide and generates an immune response in the individual against the mutated cell surface protein in the cancer cells for treatment of the cancer. The one or more polynucleotide may be administered in the form of a pharmaceutical composition that includes one or more pharmaceutically acceptable excipient. In some embodiments, the polynucleotides are administered to in the form of armed effector cells, e.g., armed natural killer (NK) cells, T cells, B cells, monocytes / macrophages, neutrophils, or combinations thereof, for example, a pharmaceutical composition that includes armed effector cells and one or more pharmaceutically acceptable excipient.

[0114] In some embodiments, the cancer is selected from the group consisting of basal cell carcinoma, bladder cancer, bone cancer, bowel carcinoma, breast cancer, carcinoid, anal squamous cell carcinoma, castration-resistant prostate cancer (CRPC), cervical carcinoma, colorectal cancer (CRC), colon cancer cutaneous squamous cell carcinoma, endometrial cancer, esophageal cancer, gastric carcinoma, gastroesophageal junction cancer, glioblastoma / mixed glioma, glioma, head and neck cancer, hepatocellular carcinoma, hematologic malignancy, liver cancer, lung cancer, melanoma, Merkel cell carcinoma, multiple myeloma, nasopharyngeal cancer, neuroblastoma, osteosarcoma, ovarian cancer, pancreatic cancer, peritoneal carcinoma, undifferentiated pleomorphic sarcoma, prostate cancer, rectal carcinoma, renal cancer, sarcoma, salivary gland carcinoma, squamous cell carcinoma, stomach cancer, testicular cancer, thymic carcinoma, thymic epithelial tumor, thymoma, thyroid cancer, urogenital cancer, urothelial cancer, uterine carcinoma, or uterine sarcoma.

[0115] In some embodiments, the method further includes administering one or more anti-cancer agent, concurrently or sequentially with respect to the polynucleotides and / or antibodies described herein. In some embodiments, the anti-cancer agent is selected from a cancer vaccine, chemotherapy, radiation, and an immunotherapeutic. In one embodiment, the immunotherapeutic is a modified T cell. In some embodiments, the anti-cancer agent may be administered concurrently or sequentially with respect to armed effector cells as described herein. The therapeutic anti-cancer effect of the polynucleotides and / or antibodies and the anticancer agent may be additive or synergistic.

[0116] Conventional methods, known to those of ordinary skill in the art of medicine, can be used to administer the polynucleotides and / or antibodies (e.g., pharmaceutical composition) to the individual, depending upon the type and severity (e.g., stage) of cancer to be treated and / or the site of the disease. This composition can also be administered via other various routes, e.g., administered orally, parenterally, by inhalation spray, topically, rectally, nasally, buccally, vaginally or via an implanted reservoir. In one embodiment, the composition is administered via intratumoral injection. In a specific embodiment, the composition is administered via intraperitoneal injection. In addition, the composition can be administered to the subject via injectable depot routes of administration such as using 1-, 3-, or 6-month depot injectable orbiodegradable materials and methods. In some examples, the pharmaceutical composition is administered intraocularly or intravitreally.

[0117] In some embodiments of the methods described herein, B cells are isolated from an individual to whom MSP peptides or polynucleotides that encode MSP peptides have been administered. The B cells may be isolated from whole blood from the individual, using methods such as, but not limited to, a leukocyte reduction filter or leukapheresis. Alternatively, B cells may be isolated from excised vaccine draining lymph nodes. For example, the MSP peptide or polynucleotide that encodes the MSP peptide may be injected along with a tracer (e.g., a radioactive tracer) to aid in identification of a lymph node in which B cells that are producing antibodies to the MSP peptide will be found. (See, e.g., Pero, S.C., et al. (2017) Vaccine 2017 01 Feb., PMID: 28161423; Krag, D„ et al. (1998) N Engl J Med 339(14):941 -6, PMID:9753708.) B cells that produce antibodies to the MSP peptides that were administered or that were encoded by the polynucleotides that were administered, may then be sequenced for use in methods for treating cancer as described herein.

[0118] In some embodiments of the methods described herein, certain MSP peptides do not induce a high titer immunological response in the individual. Such MSP peptides or polynucleotides that encode the MSP peptides may be administered to another animal, such as a mouse, to produce antibodies which may be sequenced for use in conjunction with antibodies that are generated by the individual to different MSP peptides and / or to the same MSP peptide.

[0119] In some embodiments, antibodies to MSP peptides may be produced by another animal, such as a mouse. The antibodies may be sequenced and the sequences used to produce a polynucleotide that encodes the antibody, or an MSP binding fragment or derivative thereof, for administration to the individual in a method as described herein.

[0120] The following examples are intended to illustrate, but not limit, the invention.EXAMPLES Example 1

[0121] A human patient was treated for kidney cancer. A tumor specimen and normal blood were sequenced. 20 MSPs were identified. Peptides containing the missense mutation were synthesized successfully for 19 of the 20 MSPs. Wild type mice were vaccinated with the peptides under a rapid vaccination protocol. Ten days after vaccination, spleens were harvested and hybridomas generated. Hybridomas were screened for antibodies that bound the cognate vaccine peptide. Antibodies that bound to the peptide representing the mutated section of the MSP and did not bind to a peptide representing the wild type or nonmutated protein or to a panel of normal tissue were selected.

[0122] The binding portion of each selected antibody was retained and recombinantly produced after combination to human antibody framework. Eight of the mouse / human chimeric antibodies met specification and were produced in multi-liter batches. Production used short term stable pools of transfected CHO cells. After expansion of the transfected CHO cells for about 14 days,the antibodies were harvested. The material was centrifuged and the antibodies in the supernatant were collected by Protein A.

[0123] The collected antibodies were subjected to low pH hold to reduce potential virus. The buffer exchange was done following acid pH neutralization. Following prefiltration the antibodies were combined in a cocktail and subjected to nanofiltration. The rendered cocktail was vialed in 100ml glass vials. QC of the final vialed product included potency, sterility, assurance of low endotoxin, and stability. These were transferred from the CMC site across the US to the treatment location.

[0124] The patient received 100 mg and 5 days later 200mg of the antibody cocktail without evidence of infusion reaction.Example 2

[0125] Ten mutated melanoma intracellular proteins were selected. Peptides that represented the mutated sections of the proteins were synthesized and injected into rabbits. The resulting polyclonal antibodies for each peptide were affinity enriched.

[0126] The ten vials of polyclonal antibodies were mixed as a cocktail to treat mice that had melanoma cells implanted in their flank. A PD1 inhibitor was also added to attempt to maximize the potential anticancer action of the antibodies. Six animals were tested for each of the following treatments: PBS control; PD1 i only; 10 antibody cocktail + PD1 i. Five animals were tested with normal rabbit IgG + PDi (one animal that did not show any tumor was removed from the original group of six animals). The results are shown in Fig. 1 and Fig. 2.

[0127] No difference in inhibition of tumor growth or prolongation of survival was observed between the antibodies that targeted intracellular mutated proteins and negative control regular rabbit antibodies. Median survival is shown in Table 1.Table 1Example 3

[0128] The dearth of traditional drug or hormonal targets in triple-negative breast cancer (TNBC), coupled with the poor prognosis for this subset of breast cancer patients, underscores the need for an entirely novel therapeutic strategy. Antibody cocktails achieve high levels of efficacy in each patient rather than the more conventional approach of creating a single antibody that may provide limited benefit to a subset of patients. This strategy is highly adaptable. While resistance to conventional biologies as a result of target downregulation, mutation, or intrinsic tumor heterogeneity is a persistent clinical challenge, up to 10 different mutated surface proteins on tumor cells are simultaneously targeted, thereby minimizing the potential for therapeuticresistance while affording the flexibility to generate a new panel of antibodies for a given patient if they experience tumor recurrence.

[0129] Antibodies targeting neoantigens minimize the potential for cross-reactivity with nonmutated versions of these proteins on the surfaces of healthy cells. The ability of antibodies directed against the neoantigens on multiple MSPs to rapidly inhibit cancer growth was demonstrated.

[0130] As these neoantigens are generated in a largely random manner, they are almost all patient-specific. Antibodies targeting these neoantigens must be produced for each patient. Small-batch custom antibodies that target neoantigens on multiple MSPs for individual patients were produced. A phase I clinical trial using this strategy to treat stage 4 cancer patients has been approved. Patients will be enrolled with advanced-stage TNBC for whom patient-specific cocktails of antibodies can be formulated that target the patients’ unique MSPs. Through this highly personalized medicine approach, traditional barriers to the treatment of TNBC can be overcome, potentially improving the survival and quality of life of affected patients. The results of the study will inform the further development of this unique therapeutic platform for the broader treatment of TNBC and other breast cancer subtypes.Preclinical data: in vivo and in vitro studiesMurine EMT-6 TNBC Cells Harbor MSPs

[0131] In preclinical studies exploring the feasibility of treating breast cancer using MSP-specific antibodies, the murine EMT-6 TNBC tumor model was selected as it is a syngeneic immunocompetent model that has been extensively used in previous studies. 1 ,252 missense mutations in this EMT-6 cell line were identified. To identify putative MSPs, the cellular location of each missense mutation was determined by looking up each mutation site for a given gene in the UniProt database. Of these 1 ,252 missense mutations, 104 were in the extracellular domains (ECDs) of membrane proteins. These 104 mutations were present across 92 genes harboring a single mutation and 6 genes harboring two mutations. Most of these missense mutations were random and present in genes unrelated to known oncogenic pathways. Among the 98 genes, only 5 missense mutations were present in known cancer-driver genes (Al~K, FAT2, KDR, PTPRB, and TGFBT2). Unlike most targeted therapy, which focuses on changing the function of a target protein, the strategy was based on using the missense mutations as docking sites to achieve sufficient total cell antibody binding for mediating immune effector cells. In this strategy, random non-driver MSPs may function as well as driver MSPs.

[0132] Next, these candidate MSPs on EMT-6 cells were targeted by producing antibodies directed against multiple MSP neoantigens. As no reliable prediction of the expression of candidate MSPs was available, we selected a high number of MSPs (n=12) to ensure that high levels of tumor cell binding could be achieved and to obtain better activation of immune effector cells.Custom Antibodies Effectively Bind to MSPs and Mediate Tumor Cell Killing

[0133] For the preclinical experiments, polyclonal rabbit antibodies (pAbs) were produced against each of the selected 12 MSP neoantigens. Following vaccination with peptides containing theappropriate amino acid substitutions corresponding to each mutation, antibodies directed against each MSP neoantigen were affinity-enriched from the serum of vaccinated rabbits. Table 2 details the target protein name and mutation site, the peptide harboring the missense mutation used for vaccination, purity of affinity-enriched antibodies, antibody titers, and affinity levels of the resultant antibodies against their corresponding peptides. In all 12 cases, high-titer, high-affinity antibodies were successfully produced.Table 2. Characteristics of prepared pAbs targeting 12 tumor ECD neoantigens

[0134] Immunofluorescence microscopy confirmed the ability of 9 of these 12 pAbs to bind to EMT- 6 cells in vitro. The lack of any binding by 3 of the 12 pAbs and variable binding by 9 of the 12 pAbs may be related to the variable expression of the target MSP, variable accessibility to the neoantigen, or post-translational modifications of the MSP. Figure 1A shows binding by pAbs targeting individual MSPs. Figure 1B shows binding by pAbs when increasing numbers of MSPs are targeted simultaneously. As shown in Figure 1B, Cumulative pAb binding on EMT-6 cells was evident as additional MSPs are targeted, as observed by immunofluorescence. The sequential addition of each of these 9 pAbs yielded progressively stronger cell binding. When a total of 9 missense mutations were targeted simultaneously, overall strong binding of these antibodies to the EMT-6 cells was observed. In contrast, an equivalent amount of negative control IgG failed to yield a significant immunofluorescent signal.

[0135] In vitro, the antibody cocktail targeting 9 MSPs facilitated robust leukocyte-mediated killing of EMT-6 breast cancer cells. Together, the results of the binding and cytotoxicity assays thus demonstrated that targeting 9 MSPs with a cocktail of antibodies resulted in strong overall cancer cell binding and mediated robust in vitro cytotoxicity.Antibodies targeting multiple MSPs Suppress in vivo Tumor Growth

[0136] The antibody cocktail targeting 9 MSPs inhibited tumor growth and prolonged survival of immunocompetent mice implanted with EMT-6 cancer cells. Tumor growth inhibition wasobserved as a flattening of the tumor growth curve (Fig. 3B). Experiments with mice at 6 months after a durable complete response demonstrated robust resistance to reimplantation of tumor cells, indicating that a broad immune response occurred over time (data not shown).Specificity of Antibody Binding

[0137] The ability of antibodies to bind specifically to cancer cell targets is essential for their effective administration in a manner that minimizes the potential for adverse events stemming from cross-reactivity. Our 9-antibody cocktail showed clear specificity for EMT-6 tumor binding relative to normal tissues, as observed by staining of histological sections of EMT-6 and normal tissues, using 2.25 pg of the cocktail of antibodies targeting 9 MSPs, and secondary Alexa Fluor 468 goat anti rabbit IgG to detect targeting rabbit antibody binding.Validation of Specificity

[0138] Human neuroblastoma and a panel of normal tissues were used to determine the specificity of antibodies targeting neoantigens harbored by MSPs of a human malignancy. A cocktail of antibodies targeting the neoantigens of 10 neuroblastoma patient-specific MSPs was generated. The cocktail of antibodies showed striking high binding to neuroblastoma tissue but the absence of binding to a panel of normal human tissues. This confirms in human cancer the binding specificity of the antibodies that bind to MSP neoantigens and not to normal tissues.Replicability and Generalizability

[0139] To confirm that the approach described above can be replicated and generalized and is not restricted to some artifact of the EMT-6 cell line, two additional non-breast cancer tumor models were also tested. Experiments with both CT-26 colon cancer and B16 melanoma models confirmed the positive findings observed for EMT-6 breast tumors. There was no overlap in MSP neoantigens between EMT-6, B16, and CT-26. Custom antibodies were produced for each tumor type to target the unique MSP neoantigens on each cell line. The antibodies generated for each tumor confirmed that antibodies to 9-10 different MSP neoantigens resulted in cumulative increases in tumor cell binding. For each tumor, antibody binding was strikingly robust when pooled as an antibody cocktail. Prompt tumor inhibition and durable complete responses were observed in treated tumor-bearing mice, and these responses were enhanced by concomitant PD-1 inhibition.

[0140] Leukocytes from syngeneic donor mice pre-incubated with the antibody cocktail, and the resultant “armed” immune cells were able to promptly inhibit target tumor cell growth. In vivo tumor inhibition using these antibody-armed leukocytes offers further support for the role of immune cells as mediators of antibody-dependent cytotoxic tumor cell killing following treatment with customized tumor-specific MSP-targeting antibody cocktails.Validation of Monoclonal Antibodies Targeting MSPs

[0141] Human monoclonal antibodies were generated for comparison to rabbit pAbs. Both phage- displayed and yeast-displayed human antibody libraries were independently panned against the same tumor peptides as were used for rabbit pAb production. Monoclonal antibodies derived from both library systems yielded the same results as observed with rabbit pAbs, including high levels of tumor binding, in vitro cytotoxicity, in vivo tumor inhibition, and improved survival.Targeting Missense Mutations in Intracellular Proteins with Antibodies does not Suppress in vivo Tumor Growth

[0142] Polyclonal rabbit antibodies were generated against 9 missense mutations in proteins with an intracellular location. This was done in the B16 model, in which previous results demonstrated that antibodies targeting 9 missense mutations in proteins at the cell surface caused prompt tumor inhibition and prolongation of survival. The cocktail of antibodies targeting 9 intracellular neoantigens showed no difference in tumor inhibition or survival in comparison to a normal rabbit antibody negative control.Human Breast Cancer Missense Mutation Analysis

[0143] The number of unique and shared missense mutations present in 100 TNBC and 83 nontriple negative breast cancers were assessed to further explore the potential clinical applicability of targeting the landscape of breast cancer MSPs with custom-produced antibodies. In addition to considering mutations in ECDs of membrane proteins, mutations in genes encoding secreted proteins were also included. The rationale for including secretory proteins was that during the secretion process, the location of the secreted protein may be close enough to the cell membrane for bound antibodies to mediate immune cell interaction with the cancer cell.

[0144] In each cancer sample, missense mutations were confirmed by comparing sequence data of matched normal and tumor cells from the same patient. The total number of unique missense mutations identified across the 183 breast cancers was 16,758. The cell location of each missense mutation was determined by looking up the published cell location of each mutated protein in the UniProt database. In the 100 TNBC cases, 37.9% of total missense mutations (3,106 out of 8,197) were present in the MSPs. In the 83 non-TNBC cases, 35.8% of total missense mutations (3,065 out of 8,561) were present in MSPs.

[0145] In the breast cancer patients, the median number of MSPs per patient cancer was 13.5 for TNBC and 8 for non-TNBC. Preclinical animal data as described above demonstrated that targeting 9 or 10 MSPs was sufficient to achieve high levels of antibody binding to tumors and effectively inhibit tumor growth. Guided by these preliminary data and using a cutoff of 10 MSPs per tumor, 69% of the TNBC cases had > 10 MSPs and 42% of the non-TNBC cancers had >10 MSPs. Overlap or sharing of MSP neoantigens between patients was very uncommon. In the non-TNBC cohort, 99.8% of MSP neoantigens were unique, while 0.2% were shared between 2 patients. In the TNBC cohort , 97.6% of MSP neoantigens were unique while 2.4% were shared.

[0146] Based on these results, more than half of all breast cancer patients exhibited > 10 MSPs, with this percentage being even greater among TNBC patients at 69%. Almost all of these missense mutations were entirely unique and present only in the tumor of a single patient.Example 4

[0147] B16-F10 mouse melanoma is an aggressive melanoma that has hundreds of missense mutations. Antibodies were prepared against MSPs harboring tumor-specific missense mutations (neoantigens), which are ideal targets for antibodies.

[0148] Experiments were designed to target 9 MSP neoantigens as an arbitrary number for preliminary evaluation. The only selection criterion was that the neoantigen identified by sequence data was in a cell surface-related protein. The cell location of each MSP neoantigen was determined manually by looking up the location in the UniProt Database. Predicted immunogenicity of the neoantigen in the tumor-bearing mouse was not a selection criterion, since the treatment antibodies were not produced by tumor-bearing animals. The antibodies were produced in rabbits and then administered to tumor-bearing mice. This avoided the need for algorithms to predict the immunogenicity of neoantigens in the tumor-bearing animal. Immunogenicity prediction is required with other immunotherapies that rely on the patient to produce the antibodies or other elements of an immune response versus the described strategy to exogenously derive and produce antibodies to deliver to a patient. The sequences of peptides representing the neoantigens present within 9 different MSPs are shown in Fig. 4.

[0149] Rabbits were vaccinated with the mutated peptides conjugated with keyhole limpet hemocyanin. After achieving an antibody response, peptide-specific antibodies were obtained from the rabbit plasma by affinity purification. High-affinity polyclonal rabbit antibodies were obtained to all 9 mutated peptides.

[0150] Using fluorescence microscopy, binding to histologic sections of B16-F10 melanoma tumors harvested from mice was determined for each of the 9 polyclonal antibodies. This revealed the variable expression and availability of each neoantigen on the mutated proteins, indicated by the variable intensity of red fluorescence labeling of the bound antibodies. As expected, the range of expression varied for each MSP. However, when all 9 antibodies were combined in an equal mixture cocktail, there was substantially enhanced binding.Tumor growth inhibition and increased survival

[0151] Mice were implanted subcutaneously with melanoma cells. The cocktail of 9 antibodies targeting 9 different MSP neoantigens was used for experimental treatment, A single subcutaneous 0.2 mg dose of the cocktail prolonged survival (Fig. 5B). Four 0.2 mg doses of the antibody cocktail injected subcutaneously on days 3, 6, 9, and 12 nearly doubled survival time to approximately 34 days.Sustainability of the response

[0152] After 6 months, mice with complete responses were rechallenged with additional implantation of melanoma cells. These mice received no further treatment following reimplantation of the melanoma cells. Fig. 6 shows that these mice had developed immunological resistance to tumor growth.Example 5

[0153] Using the methods described in Example 4, high-affinity polyclonal antibodies were produced against 10 MSPs present in murine CT-26 colon cancer cells. As with B16-F10 cells, immunohistochemistry demonstrated variable expression of each mutated protein. Cumulative antibody binding to colon cancer cells was observed as more MSPs were targeted withprepared antibodies. When a total of 10 MSP neoantigens were targeted, there was strong overall binding by antibodies to the CT-26 cancer cells.

[0154] Tumor inhibition studies were performed with CT-26 colon cancer tumors using the methods described in Example 4. The antibody cocktail targeting 10 MSP neoantigens, was used for mouse treatments. The antibody cocktail alone inhibited CT-26 tumor growth (Fig. 7B) and prolonged survival (data not shown). PD1 inhibition alone had no significant effect on tumor growth or survival but PD1 inhibition enhanced the ability of the antibody cocktail to both inhibit tumor growth and prolong survival. These data demonstrate that targeting multiple MSP neoantigens with antibodies was highly bioactive and that PD1 inhibition enhanced the antitumor activity of the antibody cocktail.Example 6

[0155] The strategy of targeting multiple MSP neoantigens was applied to a human neuroblastoma that had been successfully introduced from a patient into an immunodeficient mouse. A cocktail of antibodies targeting 10 different MSP neoantigens was generated. Intense binding by the cocktail to the tumor was observed, and no binding was observed to a panel of normal human tissues.Example 7

[0156] A cocktail of monoclonal antibodies targeting the MSP neoantigens in B16-F10 melanoma was evaluated, and the results compared with polyclonal antibodies. Monoclonal antibodies were generated against the neoantigens of the 4 higher-expressing MSPs in B16-F10 described above. Monoclonal antibodies were generated by phage display panning against the mutated peptides using a human antibody phage library. Brightfield immunohistochemistry binding by individual monoclonal antibodies to each of the 4 mutated proteins on B16-F10 cells exhibited positive binding to MSP neoantigens like polyclonal antibodies described above.

[0157] A cocktail made of an equal mixture of the monoclonal antibodies resulted in prompt tumor inhibition and prolonged survival like the cocktail of polyclonal antibodies described above. This demonstrated that when targeting the same neoantigens, the overall effect of monoclonal antibodies was concordant with polyclonal antibodies as determined by binding, inhibition of melanoma growth, and prolongation of survival in tumor-bearing mice.

[0158] Although the foregoing invention has been described in some detail by way of illustration and examples for purposes of clarity of understanding, it will be apparent to those skilled in the art that certain changes and modifications may be practiced without departing from the spirit and scope of the invention, which is delineated in the appended claims. Therefore, the description should not be construed as limiting the scope of the invention.

[0159] All publications, patents, and patent applications cited herein are hereby incorporated by reference in their entireties for all purposes and to the same extent as if each individualpublication, patent, or patent application were specifically and individually indicated to be so incorporated by reference.

Claims

CLAIMSWe claim:1 . A method of treating cancer, the method comprising:(a) sequencing DNA and / or RNA of cancer cells from an individual to determine mutations in cell surface related proteins in said cancer cells;(b) administering to the individual or to a second individual, at least one mRNA polynucleotide that a encodes a mutated cell surface related (MSP) peptide that is expressed in the cancer cells in said individual, wherein said MSP peptide comprises an amino acid sequence of a portion of a cell surface related protein that comprises one or more mutation relative to the cell surface related protein in non-cancer cells in said individual, wherein said mRNA polynucleotide is translated by said individual or said second individual to produce said MSP peptide, and wherein said MSP peptide generates an immune response in the individual or the second individual that comprises antibodies that specifically bind to said MSP peptide;(c) assessing said immune response by assessing antibody titer to said MSP peptide in a blood sample that is obtained from the individual or the second individual;(d) sequencing an antibody from the individual or the second individual that specifically binds to the MSP peptide to determine the amino acid sequences of the antibody;(e) using said antibody amino acid sequences to produce the antibody, or a fragment or derivative thereof that retains the ability to bind to the MSP peptide, in a system that is extrinsic to the individual; and(f) administering the antibody, or fragment or derivative thereof, that is produced in the system that is extrinsic to the individual to the individual for treatment of said cancer.

2. A method of treating cancer, the method comprising administering to an individual having cancer at least one antibody that specifically binds to an MSP peptide that is expressed in cancer cells of the individual, or a fragment or derivative thereof that retains the ability to bind to the MSP peptide, wherein the antibody, or the fragment or derivative thereof, is produced in a method that comprises:(a) administering to the individual or to a second individual, at least one mRNA polynucleotide that encodes a MSP peptide that is expressed in the cancer cells in said individual, wherein said MSP peptide comprises an amino acid sequence of a portion of a cell surface related protein that comprises one or more mutation relative to the cell surface related protein in non-cancer cells in said individual, wherein said mRNA polynucleotide is translated by said individual or said second individual to produce said MSP peptide, and wherein said MSP peptide generates an immune response in the individual or the second individual that comprises antibodies that specifically bind to said MSP peptide;(b) assessing said immune response by assessing antibody titer to said MSP peptide in a blood sample that is obtained from the individual or the second individual;(c) sequencing an antibody from the individual or the second individual that specifically binds to the MSP peptide to determine the amino acid sequences of the antibody; and(d) using said antibody amino acid sequences to produce the antibody, or a fragment or derivative thereof that retains the ability to bind to the MSP peptide, in a system that is extrinsic to the individual.

3. A method of treating cancer, the method comprising:(a) sequencing DNA and / or RNA of cancer cells from an individual to determine mutations in cell surface related proteins in said cancer cells;(b) administering to the individual or to a second individual, at least one mRNA polynucleotide that a encodes a MSP peptide that is expressed in the cancer cells in said individual, wherein said MSP peptide comprises an amino acid sequence of a portion of a cell surface related protein that comprises one or more mutation relative to the cell surface related protein in non-cancer cells in said individual, wherein said mRNA polynucleotide is translated by said individual or said second individual to produce said MSP peptide, and wherein said MSP peptide generates an immune response in the individual or the second individual that comprises antibodies that specifically bind to said MSP peptide;(c) assessing said immune response by assessing antibody titer to said MSP peptide in a blood sample that is obtained from the individual or the second individual;(d) sequencing an antibody from the individual or the second individual that specifically binds to the MSP peptide to determine the amino acid sequences of the antibody; and(e) administering a vector antibody mRNA that comprises polynucleotide sequences that encode the amino acid sequences of the antibody, or a fragment or derivative thereof that retains the ability to bind to the MSP peptide, to the individual for treatment of said cancer.

4. A method of treating cancer, the method comprising administering to an individual having cancer at least one mRNA that comprises polynucleotide sequences that encode amino acid sequences of an antibody that specifically binds to an MSP peptide that is expressed in cancer cells of the individual, or a fragment or derivative thereof that retains the ability to bind to the MSP peptide.

5. The method of claim 4, wherein the mRNA is produced in a method that comprises:(a) administering to the individual or to a second individual: (i) at least one mRNA polynucleotide that a encodes a MSP peptide that is expressed in cancer cells in said individual, wherein said mRNA polynucleotide is translated by said individual or said second individual to produce said MSP peptide, or (ii) at least one MSP peptide that is expressed in the cancer cells in said individual, wherein said MSP peptide comprises an amino acid sequence of a portion of a cell surface related protein that comprises one or more mutation relative to the cell surface related protein in non-cancer cells in said individual, wherein said MSP peptide generates an immune response in the individual or the second individual that comprises antibodies that specifically bind tosaid MSP peptide;(b) assessing said immune response by assessing antibody titer to said MSP peptide in a blood sample that is obtained from the individual or the second individual; and(c) sequencing an antibody from the individual or the second individual that specifically binds to the MSP peptide to determine the amino acid sequences of the antibody.

6. A method of treating cancer, the method comprising administering to an individual having cancer, at least one mRNA polynucleotide that encodes a MSP peptide that is expressed in cancer cells in said individual, wherein said MSP peptide comprises an amino acid sequence of a portion of a cell surface related protein that comprises one or more mutation relative to the cell surface related protein in non-cancer cells in said individual, wherein said mRNA polynucleotide is translated by said individual to produce said MSP peptide, and wherein said MSP peptide generates an immune response in the individual that comprises antibodies that specifically bind to said MSP peptide.

7. The method of claim 6, wherein prior to administration of said at least one mRNA polynucleotide that encodes said MSP peptide, DNA and / or RNA of cancer cells from the individual is sequenced to determine mutations in cell surface related proteins in said cancer cells relative to non-cancer cells in said individual.

8. A method of treating cancer, the method comprising:(a) sequencing DNA and / or RNA of cancer cells from an individual to determine mutations in cell surface related proteins in said cancer cells;(b) administering to the individual or to a second individual, at least one MSP peptide that is expressed in the cancer cells in said individual, wherein said MSP peptide comprises an amino acid sequence of a portion of a cell surface related protein that comprises one or more mutation relative to the cell surface related protein in non-cancer cells in said individual, and wherein said MSP peptide generates an immune response in the individual or the second individual that comprises antibodies that specifically bind to said MSP peptide;(c) assessing said immune response by assessing antibody titer to said MSP peptide in a blood sample that is obtained from the individual or the second individual;(d) sequencing an antibody from the individual or the second individual that specifically binds to the MSP peptide to determine the amino acid sequences of the antibody; and(e) administering a vector antibody mRNA that comprises polynucleotide sequences that encode the amino acid sequences of the antibody, or a fragment or derivative thereof that retains the ability to bind to the MSP peptide, to the individual for treatment of said cancer.

9. A method of treating cancer, the method comprising administering to an individual having cancer, at least one mRNA that comprises polynucleotide sequences that encode the amino acid sequences of an antibody that specifically binds to an MSP peptide that is expressed in cancer cells of the individual, or a fragment or derivative thereof that retains the ability to bind to the MSPpeptide, wherein said MSP peptide comprises an amino acid sequence of a portion of a cell surface related protein that comprises one or more mutation relative to the cell surface related protein in non-cancer cells in said individual, wherein said MSP peptide generates an immune response in the individual that comprises antibodies that specifically bind to said MSP peptide, wherein said at least one mRNA that encodes the amino acid sequences of the antibody is produced in a method that comprises:(a) administering to the individual or to a second individual, at least one MSP peptide that is expressed in the cancer cells in said individual, wherein said MSP peptide comprises an amino acid sequence of a portion of a cell surface related protein that comprises one or more mutation relative to non-cancer cells in said individual;(b) assessing said immune response by assessing antibody titer to said MSP peptide in a blood sample that is obtained from the individual or the second individual; and(c) sequencing an antibody from the individual or the second individual that specifically binds to the MSP peptide to determine the amino acid sequences of the antibody.

10. The method of any of claims 1 to 9, wherein said cell surface related protein from which said MSP is derived is expressed on the surface of said cancer cells.11 . The method of any of claims 1 to 9, wherein said cell surface related protein from which said MSP is derived is secreted from said cancer cells.

12. The method of any of claims 1 to 9, wherein the mRNA that encodes a MSP peptide and / or the mRNA that encodes the amino acid sequences of an antibody, or fragment or derivative thereof, is administered to the individual in a lipid particle formulation.

13. The method of any of claims 1 to 9, wherein the mRNA that encodes a MSP peptide and / or the mRNA that encodes the amino acid sequences of an antibody, or fragment or derivative thereof, is administered intramuscularly, subcutaneously, intradermally, intravenously, or via inhalation.

14. The method of any of claims 1-3 or 5-7, wherein said at least one mRNA polynucleotide that encodes a MSP peptide comprises: (i) at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten mRNA polynucleotides, wherein each of said mRNA polynucleotides encodes a different MSP peptide, or (ii) at least one mRNA that encodes at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten different MSP peptides.

15. The method of any of claims 3-4 or 8-9, wherein said at least one mRNA that comprises polynucleotide sequences that encode the amino acid sequences of the antibody, or a fragment or derivative thereof, comprises: (i) at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten mRNA polynucleotides, each of whichencodes a different antibody, or fragment or derivative thereof, or (ii) at least one mRNA that encodes at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten different antibodies, or fragments or derivatives thereof.

16. The method of claim 6, wherein said immune response is determined by assessing antibody titer to said MSP peptide in a blood sample that is obtained from the individual.

17. The method according to claim 16, wherein an antibody that specifically binds to the MSP peptide is sequenced to determine the amino acid sequences of the antibody.

18. The method according to claim 17, wherein said antibody amino acid sequences are used to produce the antibody, or a fragment or derivative thereof that retains the ability to bind to the MSP peptide, in a system that is extrinsic to the individual.

19. The method of claim 18, wherein said system that is extrinsic to the individual comprises a phage display system.

20. The method of claim 18, wherein said system that is extrinsic to the individual comprises production of said antibody, or fragment or derivative thereof, in a non-human animal from a vector antibody mRNA that comprises polynucleotide sequences that encode the amino acid sequences of the antibody, or fragment or derivative thereof.21 . The method of claim 20, wherein said vector antibody mRNA further encodes an amino acid sequence that facilitates recovery of the antibody, or fragment or derivative thereof, from plasma of the non-human animal.

22. The method of claim 21 , wherein the amino acid sequence that facilitates recovery is a His- tag sequence comprising six to nine consecutive histidine residues.

23. The method of claim 18, wherein the antibody, or fragment or derivative thereof, that is produced in the system that is extrinsic to the individual is administered to the individual for treatment of said cancer.

24. The method of claim 23, wherein antibodies, or fragments or derivatives thereof, that specifically bind at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten MSP peptides that are expressed in the cancer cells in said individual are produced in said system, and are administered to the individual for treatment of said cancer.

25. The method of claim 24, wherein the antibodies, or fragments or derivatives thereof, areadministered to the individual intravenously or subcutaneously.

26. The method of claim 17, wherein a vector antibody mRNA that comprises polynucleotide sequences that encode the amino acid sequences of the antibody, or a fragment or derivative thereof that retains the ability to bind to the MSP peptide, is administered to the individual for treatment of said cancer.

27. The method of claim 17, wherein the vector mRNA is loaded on effector cells isolated from the individual prior to administration to the individual.

28. The method of claim 27, wherein the effector cells comprise natural killer (NK) cells, T cells, B cells, monocytes / macrophages, neutrophils, or combinations thereof.

29. The method of any of claims 26 to 28, wherein vector antibody mRNAs that encode antibodies, or fragments or derivatives thereof, that specifically bind to at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten MSP peptides that are expressed in the cancer cells in said individual are administered to the individual for treatment of said cancer.

30. The method of any of claims 1 to 9, wherein the cancer is selected from basal cell carcinoma, bladder cancer, bone cancer, bowel carcinoma, breast cancer, carcinoid, anal squamous cell carcinoma, castration-resistant prostate cancer (CRPC), cervical carcinoma, colorectal cancer (CRD), colon cancer cutaneous squamous cell carcinoma, endometrial cancer, esophageal cancer, gastric carcinoma gastroesophageal junction cancer, glioblastoma / mixed glioma, glioma, head and neck cancer, hepatocellular carcinoma, hematologic malignancy, liver cancer, lung cancer, melanoma, Merkel cell carcinoma, multiple myeloma nasopharyngeal cancer, neuroblastoma, osteosarcoma, ovarian cancer, pancreatic cancer, peritoneal carcinoma, undifferentiated pleomorphic sarcoma, prostate cancer, rectal carcinoma, renal cancer, sarcoma, salivary gland carcinoma, squamous cell carcinoma, stomach cancer, testicular cancer, thymic carcinoma, thymic epithelial tumor, thymoma, thyroid cancer, urogenital cancer, urothelial cancer, uterine carcinoma, and uterine sarcoma.31 . A composition, comprising: at least one mRNA polynucleotide that encodes a MSP peptide that is expressed in cancer cells in an individual, wherein said MSP peptide comprises an amino acid sequence of a portion of a cell surface related protein that comprises one or more mutation relative to the cell surface related protein in non-cancer cells in said individual; and a pharmaceutically acceptable excipient.

32. The composition of claim 31 , wherein said at least one mRNA polynucleotide that encodes a MSP peptide comprises: (i) at least two, at least three, at least four, at least five, at least six, at leastseven, at least eight, at least nine, or at least ten mRNA polynucleotides, wherein each of said mRNA polynucleotides encodes a different MSP peptide, or (ii) at least one mRNA that encodes at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten different MSP peptides.

33. A composition, comprising: at least one mRNA polynucleotide that encodes an antibody, or a fragment or derivative thereof, that specifically binds to an MSP peptide in an individual, wherein the MSP peptide is expressed in cancer cells in the individual, wherein said MSP peptide comprises an amino acid sequence of a portion of a cell surface related protein that comprises one or more mutation relative to the cell surface related protein in non-cancer cells in said individual; and a pharmaceutically acceptable excipient.

34. The composition of claim 33, wherein said at least one mRNA that comprises polynucleotide sequences that encode the amino acid sequences of the antibody, or a fragment or derivative thereof, comprises: (i) at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten mRNA polynucleotides, each of which encodes a different antibody, or fragment or derivative thereof, or (ii) at least one mRNA that encodes at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten different antibodies, or fragments or derivatives thereof35. The composition of any of claims 31 to 34, further comprising: effector cells that are isolated from the individual, wherein the mRNA is loaded on the effector cells.

36. The composition of claim 35, wherein the effector cells comprise natural killer (NK) cells, T cells, B cells, monocytes / macrophages, neutrophils, or combinations thereof.

37. A method for production of an antibody, or a fragment or derivative thereof that specifically binds a MSP peptide that is expressed in cancer cells in an individual, wherein said MSP peptide comprises an amino acid sequence of a portion of a cell surface related protein that comprises one or more mutation relative to the cell surface related protein in non-cancer cells in said individual, said method comprising: (i) expressing said antibody, or fragment or derivative thereof, in a phage display system, from a polynucleotide that encodes the amino acid sequences of the antibody, or fragment or derivative thereof; or (ii) expressing said antibody, or fragment or derivative thereof, in a non-human animal from a mRNA polynucleotide that encodes the amino acid sequences of the antibody, or fragment or derivative thereof.

38. A method according to claim 37, wherein the polynucleotide that encodes the amino acid sequences of the antibody, or fragment or derivative thereof, further encodes an amino acid sequence that facilitates recovery or purification of the antibody, or fragment or derivative thereof.

39. The method according to claim 38, wherein the amino acid sequence that facilitates recovery or purification comprises a His-tag sequence that comprises six to nine consecutive histidine residues.

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