Shared neoantigens

Abstract

Disclosed herein in one aspect is a pharmaceutical composition comprising a plurality of neoantigen peptides and a pharmaceutically acceptable carrier, each neoantigen peptide comprising a tumor-specific neoepitope capable of binding to an HLA protein in a subject, each tumor-specific neoepitope comprising a tumor-specific mutation present in a tumor, wherein (a) the composition comprises neoantigen peptides comprising tumor-specific mutations present in at least 1% of subjects in a population of subjects having cancer; (b) the composition comprises neoantigen peptides comprising tumor-specific neoepitopes that bind to an HLA protein present in at least 5% of subjects in the population; and (c) the composition comprises at least one neoantigen peptide capable of eliciting an immune response against a tumor present in at least 5% of subjects in the population of subjects having cancer.

Description

[0001] Cross-reference to related applications

[0002] This application is a divisional application of Chinese Patent Application No. 201680042436.4, filed on May 20, 2016, entitled "Common Neoantigen". The original application was the national phase application PCT / US2016 / 033452, which claims priority and benefit to U.S. Provisional Application Serial No. 62 / 179,877, filed on May 20, 2015, and U.S. Provisional Application Serial No. 62 / 389,377, filed on February 23, 2016.

[0003] All documents cited in or during the examination of the foregoing applications (“References”) and all documents referenced or cited in those References, and all documents referenced or cited herein (“Documents Referenced herein”), together with any manufacturer’s specifications, descriptions, product specifications, and product tables for any product mentioned herein or incorporated herein by reference, are hereby incorporated by reference and may be used in the practice of the invention. More specifically, all referenced documents are incorporated herein by reference to the extent that each individual document is explicitly and individually indicated to be incorporated herein by reference. Technical Field

[0004] The present invention relates to methods and compositions for treating tumor formation (e.g., tumors), particularly using at least one neoantigen peptide suitable for treating a significant proportion of subjects in a population suffering from cancer. Background Technology

[0005] Approximately 1.6 million Americans are diagnosed with neoplasia each year, and about 580,000 people were expected to die from the disease in the United States in 2013. Significant improvements have been made in the detection, diagnosis, and treatment of neoplasia over the past few decades, which have significantly improved survival rates for many types of neoplasia. However, only about 60% of those diagnosed with neoplasia survive five years after treatment begins, making neoplasia the second leading cause of death in the United States.

[0006] Currently, there are many different existing cancer therapies, including resection techniques (e.g., surgery, cryo / thermal therapy, ultrasound, radiofrequency ablation, and radiation) and chemotherapeutic techniques (e.g., pharmaceuticals, cytotoxic agents / chemotherapy agents, monoclonal antibodies, and different combinations thereof). Unfortunately, such therapies are frequently associated with serious risks, toxic side effects, extremely high costs, and uncertain efficacy.

[0007] There is growing interest in cancer therapies that target cancerous cells using the patient's own immune system (e.g., cancer vaccines) because such therapies can mitigate / eliminate some of the drawbacks described herein. Cancer vaccines typically consist of tumor antigens and immunostimulatory molecules (e.g., cytokines or TLR ligands) that work together to induce antigen-specific cytotoxic T cells that target and destroy tumor cells. Current cancer vaccines may contain common tumor antigens, which are naturally occurring proteins (i.e., proteins encoded by the DNA of all normal cells in an individual) selectively expressed or overexpressed in tumors found in many individuals. While such common tumor antigens are useful in identifying specific types of tumors, they are undesirable as immunogens for targeting T-cell responses against specific tumor types because they are susceptible to self-suppressive immunosuppression. Vaccines containing both tumor-specific and patient-specific neoantigens can overcome some of the drawbacks of vaccines containing common tumor antigens. However, using patient-specific neoantigens requires sequencing the genome of an individual subject and generating a personalized composition that includes a combination of neoantigens present in that individual subject. Therefore, improved methods and compositions for delivering cancer vaccines remain needed.

[0008] The citation or designation of any reference in this application does not imply an admission that such reference can be obtained as prior art to this invention. Summary of the Invention

[0009] Preferred statements (features) and embodiments of the invention are set forth below. Unless expressly indicated otherwise, each statement and embodiment of the invention thus defined may be combined with any other statement and / or embodiment. Specifically, any feature indicated as preferred or advantageous may be combined with one or more other features or statements indicated as preferred or advantageous. Thus, the invention is captured particularly by one or more of the following statements and embodiments in combination with any or any other statement and / or embodiment.

[0010] The object of this invention is to provide methods and compositions for treating a population of cancer patients by inducing an immune response targeting cancer. In one aspect, the invention relates to a pharmaceutical composition comprising at least one neoantigen peptide and a pharmaceutically acceptable carrier, each of the at least one neoantigen peptide comprising a tumor-specific neoepitope capable of binding to an HLA protein in a subject, each tumor-specific neoepitope comprising a tumor-specific mutation present in the tumor. The composition may comprise one neoantigen peptide. In other embodiments, the composition may comprise more than 100 neoantigen peptides. Preferably, the composition comprises about 20 neoantigen peptides. The at least one neoantigen peptide may comprise a tumor-specific mutation. The mutation may be frequent. Preferably, the mutation is present in a large proportion of the population. Frequent mutations may be based on mutations present in the tumors of at least 1% of subjects in a population of subjects with cancer. The composition may comprise at least one neoantigen peptide containing a tumor-specific neoepitope that binds to an HLA protein present in at least 5% of subjects in a population of subjects with cancer. Additionally, the composition may contain at least one neoantigen peptide capable of inducing an immune response against tumors present in at least 5% of subjects in a population of subjects with cancer. The ability to elicit an immune response refers to the immune system's capacity to present antigens to lymphocytes. For the immune system to present an antigen, it needs to be presented by the subject's HLA proteins. For an immune response to be elicited against a tumor, the tumor needs to contain a mutation that leads to the expression of that antigen. For this composition to benefit a population in need, the population must include subjects expressing HLA alleles capable of binding to at least one neoantigen peptide present in the composition, and the population must include subjects with tumors containing mutations that lead to the presence of the neoantigen epitope in the neoantigen peptide.

[0011] This composition can be specific to a group of subjects with cancer who share a common characteristic. This group may have cancer or may have a specific type of cancer. This group may share a common set of HLA subtypes. Based on ethnicity, they may share HLA subtypes. Without being bound by theory, the percentage of HLA types in the group can be predicted based on ethnicity without testing. Without being bound by theory, different groups express different HLA types capable of binding different neoantigen peptides. Therefore, compositions can be formulated to provide a large proportion of benefit to said group, while not providing benefit to another group. Without being bound by theory, different cancers contain different mutations, and therefore, compositions tailored for a specific cancer can be used to provide greater benefit to a group with only one type of cancer compared to a group with more than one type of cancer. In one embodiment, this group has adrenocortical carcinoma (ACC), bladder urothelial carcinoma (BLCA), invasive breast carcinoma (BRCA), cervical squamous cell carcinoma and cervical adenocarcinoma (CESC), colonic adenocarcinoma (COAD), chronic lymphocytic leukemia (CLL), colorectal cancer (CRC), diffuse large B-cell lymphoma (DLBCL), glioblastoma multiforme (GBM), head and neck squamous cell carcinoma (HNSC), chromophobe renal cell carcinoma (KICH), clear cell renal cell carcinoma (KIRC), and papillary renal cell carcinoma. Cancer (KIRP), acute myeloid leukemia (LAML), hepatocellular carcinoma (LIHC), lung adenocarcinoma (LUAD), lung squamous cell carcinoma (LUSC), multiple myeloma (MM), ovarian serous cystadenocarcinoma (OV), pancreatic cancer (PAAD), prostate adenocarcinoma (PRAD), rectal adenocarcinoma (READ), skin melanoma (SKCM), gastric adenocarcinoma (STAD), testicular germ cell tumor (TGCT), thyroid adenocarcinoma (THCA), endometrioid carcinoma of the uterine corpus (UCEC) or uterine carcinosarcoma (UCS).

[0012] In one embodiment, the subject population has CLL; the at least one tumor-specific mutation includes any combination of mutations in Table 8 with the example disease “CLL”; and at least one of a set of six of the at least one tumor-specific mutation will be found in 17.49% of the subjects in the CLL population. The subject population may have BLCA; the at least one tumor-specific mutation includes any combination of mutations in Table 8 with the example disease “BLCA”; and at least one of a set of six of the at least one tumor-specific mutation will be found in 26.92% of the subjects in this population. The subject population may have BRCA; the at least one tumor-specific mutation includes any combination of mutations in Table 8 with the example disease “BRCA”; and at least one of a set of 18 of the at least one tumor-specific mutation will be found in 36.04% of the subjects in this population. The subject population may have COAD; the at least one tumor-specific mutation includes any combination of mutations in Table 8 with the example disease “COAD”; and at least one of a set of three of the at least one tumor-specific mutation will be found in 27.14% of the subjects in this population. This group of subjects may have GBM; the at least one tumor-specific mutation includes any combination of the mutations in Table 8 with the example disease “GBM”; and at least one of a set of 14 of the at least one tumor-specific mutations will be found in 34.36% of the subjects in this group. This group of subjects may have HNSC; the at least one tumor-specific mutation includes any combination of the mutations in Table 8 with the example disease “HNSC”; and at least one of a set of 10 of the at least one tumor-specific mutations will be found in 21.61% of the subjects in this group. This group of subjects may have KIRC; the at least one tumor-specific mutation includes any combination of the mutations in Table 8 with the example disease “KIRC”; and at least one of a set of four of the at least one tumor-specific mutations will be found in 6% of the subjects in this group. This group of subjects may have LAML; the at least one tumor-specific mutation includes any combination of the mutations in Table 8 with the example disease “LAML”; and at least one of a set of 11 of the at least one tumor-specific mutations will be found in 47.45% of the subjects in this group. This group of subjects may have LUAD; the at least one tumor-specific mutation includes any combination of the mutations in Table 8 with the example disease “LUAD”; and at least one of a set of 11 of the at least one tumor-specific mutations will be found in 33.42% of the subjects in this group.This group of subjects may have LUSC; the at least one tumor-specific mutation includes any combination of the mutations in Table 8 with the example disease “LUSC”; and at least one of a set of two of the at least one tumor-specific mutation will be found in 7.87% of the subjects in this group. This group of subjects may have OV; the at least one tumor-specific mutation includes any combination of the mutations in Table 8 with the example disease “OV”; and at least one of a set of ten of the at least one tumor-specific mutation will be found in 22.78% of the subjects in this group. This group of subjects may have READ; the at least one tumor-specific mutation includes any combination of the mutations in Table 8 with the example disease “READ”; and at least one of a set of two of the at least one tumor-specific mutation will be found in 20.51% of the subjects in this group. This group of subjects may have SKCM; the at least one tumor-specific mutation includes any combination of the mutations in Table 8 with the example disease “SKCM”; and at least one of a set of 64 of the at least one tumor-specific mutation will be found in 90.91% of the subjects in this group. This group of participants may have UCEC; the at least one tumor-specific mutation includes any combination of the mutations in Table 8 with the example disease “UCEC”; and at least one of a set of 30 of the at least one tumor-specific mutations will be found in 67.74% of the participants in this group. This group of participants may have ACC; the at least one tumor-specific mutation includes any combination of the mutations in Table 8 with the example disease “ACC”; and at least one of a set of 161 of the at least one tumor-specific mutations will be found in 50% of the participants in this group. This group of participants may have CESC; the at least one tumor-specific mutation includes any combination of the mutations in Table 8 with the example disease “CESC”; and at least one of a set of four of the at least one tumor-specific mutations will be found in 23.71% of the participants in this group. This group of participants may have CRC; the at least one tumor-specific mutation includes any combination of the mutations in Table 8 with the example disease “CRC”; and at least one of a set of 15 of the at least one tumor-specific mutations will be found in 56.65% of the participants in this group. This group of subjects may have DLBCL; the at least one tumor-specific mutation includes any combination of the mutations in Table 8 with the example disease “DLBCL”; and at least one of a set of 2 of the at least one tumor-specific mutation will be found in 13.79% of the subjects in this group. This group of subjects may have KICH; the at least one tumor-specific mutation includes any combination of the mutations in Table 8 with the example disease “KICH”; and at least one of a set of 24 of the at least one tumor-specific mutation will be found in 50% of the subjects in this group.This group of subjects may have KIRP; the at least one tumor-specific mutation includes any combination of mutations in Table 8 with the example disease “KIRP”; and at least one of a set of nine of the at least one tumor-specific mutations will be found in 42.24% of the subjects in this group. This group of subjects may have LIHC; the at least one tumor-specific mutation includes any combination of mutations in Table 8 with the example disease “LIHC”; and at least one of a set of two of the at least one tumor-specific mutations will be found in 6.57% of the subjects in this group. This group of subjects may have MM; the at least one tumor-specific mutation includes any combination of mutations in Table 8 with the example disease “MM”; and at least one of a set of six of the at least one tumor-specific mutations will be found in 23.9% of the subjects in this group. This group of subjects may have PRAD; the at least one tumor-specific mutation includes any combination of mutations in Table 8 with the example disease “PRAD”; and at least one of a set of 24 of the at least one tumor-specific mutations will be found in 39.85% of the subjects in this group. This group of subjects may have STAD; the at least one tumor-specific mutation includes any combination of the mutations in Table 8 with the example disease “STAD”; and at least one of a set of 150 of the at least one tumor-specific mutations will be found in 48.79% of the subjects in this group. This group of subjects may have TGCT; the at least one tumor-specific mutation includes any combination of the mutations in Table 8 with the example disease “TGCT”; and at least one of a set of 14 of the at least one tumor-specific mutations will be found in 51.61% of the subjects in this group. This group of subjects may have THCA; the at least one tumor-specific mutation includes any combination of the mutations in Table 8 with the example disease “THCA”; and at least one of a set of five of the at least one tumor-specific mutations will be found in 69.88% of the subjects in this group. This group of subjects may have UCS; the at least one tumor-specific mutation includes any combination of the mutations in Table 8 with the example disease “UCS”; and at least one of a set of two of the at least one tumor-specific mutations will be found in 16.07% of the subjects in this group. This group of subjects may have PAAD; the at least one tumor-specific mutation includes any combination of mutations in Table 8 with the example diseases of “PAAD”; and at least one of the 53 groups of the at least one tumor-specific mutation will be found in 50% of the subjects in this group. This group of subjects may also have solid tumors. Solid tumors can be clear cell renal cell carcinoma (ccRCC), melanoma, sarcoma, or bladder cancer, colon cancer, brain cancer, breast cancer, head and neck cancer, endometrial cancer, lung cancer, ovarian cancer, pancreatic cancer, or prostate cancer. This group of subjects may have liquid tumors.The liquid tumor could be non-Hodgkin's lymphoma or leukemia.

[0013] In another embodiment, the at least one tumor-specific mutation occurs in at least 500 patients per year in a subject population with cancer, and the at least one mutation may be a mutation listed for the population in Table 9. The at least one neoantigen peptide may be at least one peptide listed in Table 9.

[0014] In another embodiment, the group of people with cancer is being treated with a drug or therapy. The group of people with cancer may have previously, are currently, or have been selected to be treated with ilotinib, erlotinib, imatinib, gefitinib, crizotinib, trastuzumab, vemurafenib, RAF / MEK, or anti-estrogenic therapy.

[0015] In another embodiment, the composition includes at least one neoantigen peptide capable of eliciting an immune response against tumors present in at least 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or 99% of subjects in a subject population with cancer.

[0016] In another embodiment, at least 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or 99% of the subjects in the population have at least one tumor-specific mutation present in the composition; and at least 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or 99% of the subjects in the population have at least one HLA protein that binds to a tumor-specific neoepitope present in the composition.

[0017] In one embodiment, the tumor-specific mutation includes splice variant mutations, point mutations, and / or frameshift mutations. In another embodiment, the tumor-specific mutation includes drug resistance mutations. In one embodiment, the neoantigen peptide includes not only the resulting mutated neoantigen protein sequence but also a long peptide region surrounding and including the mutation, and including all contiguous segments therein (see Tables 1-4). In one embodiment, the tumor-specific mutation is present in one or more genes encoding proteins selected from the group consisting of: programmed death-ligand 1 (PD-L1), androgen receptor (AR), Bruton's tyrosine kinase (BTK), epidermal growth factor receptor (EGFR), BCR-Abl, c-kit, PIK3CA, HER2, EML4-ALK, KRAS, ALK, ROS1, AKT1, BRAF, MEK1, MEK2, NRAS, RAC1, and ESR1. In one embodiment, the tumor-specific mutation is present in one or more genes listed in any of the tables presented herein. In one embodiment, the at least one tumor-specific mutation originates from alternative splicing of PD-L1 or AR. In one embodiment, the at least one tumor-specific mutation originates from splice variants sPD-L1, AR-V1, or AR-V7. In one embodiment, the at least one tumor-specific mutation is a drug resistance mutation selected from the group consisting of: BTK / C481S, EGFR / T790M, BCR-Abl / T315I, BCR-Abl / Y253H, BCR-Abl / E255K, BCR-Abl / E255V, c-kit / T670I, PIK3CA / E545K, PIK3CA / E542K, HER2 / G776(YVMA), HER2 / E545K, EML4-ALK / G1269A, KRAS / G12V / D, ALK / L1196M, ALK / G1202R, ALK / S1206Y, ALK / 1151T(ins), ALK / F1174C, ROS1 / G2032R, AKT1 / E17K, BRAF / V600E, MEK1 / Q56P, MEK1 / E203K, MEK1 / C121S, MEK1 / V60E, MEK1 / G128V, MEK1 / V154I, MEK1 / P124S, MEK1 / P124L, NRAS / Q61K / L / R, NRAS / T58I, MEK2 / C125S, RAC1 / P29S, ESR1 / S463P, AR / V534E, AR / P53 5H, AR / L536Q, AR / L536R, AR / Y537C, AR / Y537S, AR / Y537N, AR / D538G and AR / F876L.In one embodiment, the drug resistance mutation is induced by treatment with erlotinib, imatinib, gefitinib, crizotinib, trastuzumab, vemurafenib, RAF / MEK, or anti-estrogen therapy. In another embodiment, the subject has the drug resistance mutation prior to treatment.

[0018] In another embodiment, the composition comprises at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 neoantigen peptides. The composition may include 15 to 20 neoantigen peptides. The composition may include more than 100, 200, or 300 neoantigen peptides. The length of each neoantigen peptide may be from about 5 to about 50 amino acids.

[0019] In another embodiment, the pharmaceutical composition is an immunogenic composition or a vaccine composition. The pharmaceutical composition may further include an immunomodulator or adjuvant. The immunomodulator or adjuvant may be selected from the group consisting of: poly-ICLC, 1018ISS, aluminum salts, Amplivax, AS15, BCG, CP-870, 893, CpG7909, CyaA, cyclic dinucleotides such as STING, dSLIM, GM-CSF, IC30, IC31, imiquimod, ImuFact IMP321, IS Patch, ISS, ISCMATRIX, Juvlmmune, LipoVac, MF59, monophospholipase A, Montanide IMS1312, Montanide ISA 206, Montanide ISA 50V, Montanide ISA-51, OK-432, OM-174, OM-197-MP-EC, ONTAK, Vector systems, PLGA microparticles, remiquimod, SRL172, virions and other virus-like particles, YF-17D, VEGF traps, R848, β-glucan, Pam3Cys and Aquila's QS21stimulon.

[0020] In another embodiment, the pharmaceutical composition comprises one or more neoantigen peptides as defined in Tables 1, 2, 3 or 4.

[0021] In one embodiment, each tumor-specific neoepitope is at a K0.050 nM. D Combined with HLA-A, HLA-B, or HLA-C, or HLA-ADRB, HLA-ADBM XXXXX.

[0022] In another aspect, the present invention relates to a method of treating or preventing tumors in a subject of need by administering to the subject any pharmaceutical composition as described herein.

[0023] In one embodiment, a method of treating or preventing tumors in a patient in need is provided, the method comprising administering to the patient a composition comprising at least one neoantigen peptide and a pharmaceutically acceptable carrier, each of the at least one neoantigen peptide comprising a tumor-specific neoepitope capable of binding to an HLA protein in the subject's body, each tumor-specific neoepitope comprising a tumor-specific mutation present in the tumor, wherein the composition comprises at least one neoantigen peptide comprising a tumor-specific mutation present in the tumors of at least 1% of subjects in a subject population with cancer; the composition comprises at least one neoantigen peptide containing a tumor-specific neoepitope that binds to an HLA protein present in at least 5% of subjects in the subject population with cancer; and the composition comprises at least one neoantigen peptide capable of eliciting an immune response against tumors present in at least 5% of subjects in the subject population with cancer.

[0024] In one embodiment, the subject population has adrenocortical carcinoma (ACC), bladder urothelial carcinoma (BLCA), invasive breast carcinoma (BRCA), cervical squamous cell carcinoma and cervical adenocarcinoma (CESC), colonic adenocarcinoma (COAD), chronic lymphocytic leukemia (CLL), colorectal cancer (CRC), diffuse large B-cell lymphoma (DLBCL), glioblastoma multiforme (GBM), head and neck squamous cell carcinoma (HNSC), chromophobe renal cell carcinoma (KICH), clear cell renal carcinoma (KIRC), and papillary renal cell carcinoma. The subjects may be diagnosed with the following cancers: Kidney cell carcinoma (KIRP), acute myeloid leukemia (LAML), hepatocellular carcinoma (LIHC), lung adenocarcinoma (LUAD), lung squamous cell carcinoma (LUSC), multiple myeloma (MM), ovarian serous cystadenocarcinoma (OV), pancreatic cancer (PAAD), prostate adenocarcinoma (PRAD), rectal adenocarcinoma (READ), cutaneous melanoma (SKCM), gastric adenocarcinoma (STAD), testicular germ cell tumor (TGCT), thyroid adenocarcinoma (THCA), endometrioid carcinoma of the uterine corpus (UCEC), or uterine carcinosarcoma (UCS). In one embodiment, the subject population may have a solid tumor. Solid tumors may be clear cell renal cell carcinoma (ccRCC), melanoma, sarcoma, or bladder cancer, colon cancer, brain cancer, breast cancer, head and neck cancer, endometrial cancer, lung cancer, ovarian cancer, pancreatic cancer, or prostate cancer. In another embodiment, the subject population may have a liquid tumor. The liquid tumor may be non-Hodgkin's lymphoma or leukemia.

[0025] In one embodiment, the group of people with cancer has been, is, or has been selected to be treated with ilotinib, erlotinib, imatinib, gefitinib, crizotinib, trastuzumab, vemurafenib, RAF / MEK, or anti-estrogenic therapy.

[0026] In one embodiment, the at least one neoantigen peptide is capable of eliciting an immune response against tumors present in at least 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or 99% of subjects in a subject population with cancer. In one embodiment, at least 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or 99% of subjects in the population have at least one tumor-specific mutation present in the composition, and at least 5%, 10%, 20%, 30%, 40%, 50%, 60%, 70%, 80%, 90%, 95%, or 99% of subjects in the population have at least one HLA protein that binds to a tumor-specific neoepitope present in the composition.

[0027] In another embodiment, tumor-specific mutations include splice variant mutations, point mutations, and / or frameshift mutations. Tumor-specific mutations may be drug resistance mutations. Tumor-specific mutations may be present in one or more genes encoding proteins selected from the group consisting of: programmed death-ligand 1 (PD-L1), androgen receptor (AR), Bruton's tyrosine kinase (BTK), epidermal growth factor receptor (EGFR), BCR-Abl, c-kit, PIK3CA, HER2, EML4-ALK, KRAS, ALK, ROS1, AKT1, BRAF, MEK1, MEK2, NRAS, RAC1, and ESR1. Tumor-specific mutations may be present in one or more genes listed in any of these tables. The at least one tumor-specific mutation may originate from alternative splicing of PD-L1 or AR. The at least one tumor-specific mutation may originate from splice variants sPD-L1, AR-V1, or AR-V7.

[0028] In one embodiment, the at least one tumor-specific mutation is a drug resistance mutation selected from the group consisting of: BTK / C481S, EGFR / T790M, BCR-Abl / T315I, BCR-Abl / Y253H, BCR-Abl / E255K, BCR-Abl / E255V, c-kit / T670I, PIK3CA / E545K, PIK3CA / E542K, HER2 / G776(YVMA), HER2 / E545K, EML4-ALK / G1269A, KRAS / G12V / D, ALK / L1196M, ALK / G1202R, ALK / S1206Y, ALK / 1151T(ins), ALK / F1174C, ROS1 / G2032R, AKT1 / E17K, BRAF / V600E, MEK1 / Q56P, MEK1 / E203K, MEK1 / C121S, MEK1 / V60E, MEK1 / G128V, MEK1 / V154I, MEK1 / P124S, MEK1 / P124L, NRAS / Q61K / L / R, NRAS / T58I, MEK2 / C125S, RAC1 / P29S, ESR1 / S463P, AR / V534E, AR / P53 5H, AR / L536Q, AR / L536R, AR / Y537C, AR / Y537S, AR / Y537N, AR / D538G and AR / F876L. Drug resistance mutations can be induced by treatment with ilotinib, erlotinib, imatinib, gefitinib, crizotinib, trastuzumab, vemurafenib, RAF / MEK, or anti-estrogenic therapy.

[0029] In another embodiment, the composition comprises at least 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, 16, 17, 18, 19, or 20 neoantigen peptides. In a preferred embodiment, the composition comprises 15 to 20 neoantigen peptides.

[0030] In another embodiment, the length of each neoantigen peptide ranges from about 5 to about 50 amino acids.

[0031] In another embodiment, the composition is an immunogenic composition or a vaccine composition. For example, the immunogenic or vaccine composition may include an immunomodulator or adjuvant. The immunomodulator or adjuvant may be selected from the group consisting of: poly-ICLC, 1018ISS, aluminum salts, Amplivax, AS15, BCG, CP-870, 893, CpG7909, CyaA, cyclic dinucleotides such as STING, dSLIM, GM-CSF, IC30, IC31, imiquimod, ImuFact IMP321, IS Patch, ISS, ISCMATRIX, Juvlmmune, LipoVac, MF59, monophospholipase A, Montanide IMS1312, Montanide ISA 206, Montanide ISA 50V, Montanide ISA-51, OK-432, OM-174, OM-197-MP-EC, ONTAK, Vector systems, PLGA microparticles, remiquimod, SRL172, virions and other virus-like particles, YF-17D, VEGF traps, R848, β-glucan, Pam3Cys and Aquila's QS21 stimulon.

[0032] In one embodiment, the composition comprises one or more neoantigen peptides as defined in Tables 1, 2, 3 or 4.

[0033] In one embodiment, each tumor-specific neoepitope is at a K0.050 nM. D Combined with HLA-A, HLA-B, or HLA-C, or HLA-ADRB, HLA-ADBM XXXXX.

[0034] In another aspect, the present invention provides a method of cancer preventive treatment, the method comprising selecting a cancer drug for a patient in need, the drug being selected from the group consisting of: erlotinib, imatinib, gefitinib, crizotinib, trastuzumab, vemurafenib, RAF / MEK, and anti-estrogen therapy; and prophylactically administering to the subject a pharmaceutical composition comprising a neoantigen peptide derived from a drug resistance mutation associated with the selected cancer drug before a drug resistance mutation can be detected.

[0035] The common neoantigen immunogenic composition can be administered via sub-compositions, each containing a portion of the neoantigen, and these sub-compositions can be administered to different sites on the subject or patient; for example, a composition containing 20 different neoantigens can be divided into four (4) sub-compositions, each containing 5 of the 20 different neoantigens, and these four (4) sub-compositions can be administered to deliver each sub-composition to a separate group of draining lymph nodes in the patient, such as to each arm and each leg (e.g., the thigh or upper thigh on each side of the patient or near the buttock or lower back), in an effort to deliver as few neoantigens as possible to each group of draining lymph nodes in the patient or subject, and thus limit competition between neoantigens. Of course, the number of sites and thus the number of sub-compositions can vary; for example, a skilled practitioner can consider administration at or near the spleen with a fifth point, and a skilled practitioner can change the sites so that only one, two, or three of the limbs are used (e.g., each arm and one leg, each leg and one arm, each leg and no arm, or only two arms). The shared neoantigen immunogenic compositions administered at the aforementioned intervals can be different formulations, and the sub-compositions administered at different sites on the subject or patient during a single administration can be different compositions. For example, the shared neoantigen immunogenic composition can be administered first, followed by or later by a vector (e.g., a viral vector or plasmid) expressing one or more antigens in vivo. Similarly, in the administration of different sub-compositions at different sites on the patient or subject, some sub-compositions may include the whole antigen, and some sub-compositions may include a vector (e.g., a viral vector or plasmid) expressing one or more antigens in vivo. Furthermore, some compositions and sub-compositions may include both one or more vectors (e.g., viral vectors or plasmids) expressing one or more antigens in vivo and the whole antigen. Vectors expressing one or more antigens in vivo (e.g., poxviruses) can have an immunostimulatory or adjuvant effect, and therefore compositions or sub-compositions containing such vectors can be adjuvants themselves. Moreover, by demonstrating the nature of how the antigen is presented to the immune system, these administrations can be "priming" and then "boosting" the immune system. Furthermore, when “vaccine” is mentioned in this document, it is intended to refer to the invention comprising an immunogenic composition, and when “patient” or “subject” is mentioned, it is intended to refer to an individual who is a patient or subject who needs the treatment, administration, composition and general subject matter invention disclosed herein.

[0036] Furthermore, this invention is applicable to the use of any type of expression vector, such as viral expression vectors, such as poxviruses (e.g., orthopoxvirus or avipoxvirus, such as cowpoxvirus, including modified Ankara vaccinia vaccine or MVA, MVA-BN, NYVAC (according to WO-A-92 / 15672), fowlpox, such as TROVAX, canarypox, such as ALVAC (WO-A-95 / 27780 and WO-A-92 / 15672), pigeonpox virus, swinepox, and the like), adenoviruses, AAV herpesviruses, and lentiviruses; or plasmid or DNA or nucleic acid molecular vectors. Some vectors that are cytoplasmic, such as poxvirus vectors, will be advantageous. However, in the practice of this invention, the use of adenoviruses, AAVs, and lentiviruses will also be advantageous.

[0037] In readily available, especially reconstructed, common neoantigen immunogenic compositions, the vector, such as a viral vector, is present in an amount that is within the skill of a person skilled in the art, as disclosed herein and known in the art (e.g., in the patents and scientific literature cited herein).

[0038] Whole antigens or vectors, such as recombinant live vaccines, may exist in a freeze-dried form that allows them to be stored and reconstituted immediately before use in a solvent or excipient, which may include adjuvants as discussed herein.

[0039] Therefore, the subject of this invention is still a vaccination or immunization device or kit comprising separately packaged, freeze-dried vaccines and solutions, advantageously including adjuvant compounds for reconstructing freeze-dried vaccines as discussed herein.

[0040] The subject of this invention is also a method of vaccination or immunization, which includes or consists essentially of: administering, for example, via a parenteral, preferably subcutaneous, intramuscular, or intradermal route, or via a mucosal route, at one or more administration ratios, a vaccine or immunogenic composition according to the invention. Optionally, this method includes a preparative step of reconstructing a lyophilized common neoantigen immunogenic composition (e.g., if lyophilized whole antigen or carrier) in a solution (advantageously also including an adjuvant).

[0041] In one embodiment, the common neoantigen immunogenic composition is administered at a dose of about 10 μg to 1 mg per 70 kg individual for each neoantigen peptide. In another embodiment, the common neoantigen immunogenic composition is administered at an average weekly dose level of about 10 μg to 2000 μg per 70 kg individual for each neoantigen peptide. In another related embodiment, administration is intravenous. In one embodiment, the common neoantigen immunogenic composition is administered intravenously or subcutaneously.

[0042] In another embodiment, the method further includes (a) obtaining a tumor tissue sample from each subject; (b) detecting one or more of these tumor-specific mutations in the sample; and (c) if at least one of these tumor-specific mutations is detected in the sample from the subject, selecting a subject from the subject population to be treated with the at least one neoantigen peptide.

[0043] In another embodiment, the method further includes (a) identifying HLA allotypes present in each subject; and (b) selecting subjects from the subject population to be treated with the at least one neoantigen peptide if one or more HLA allotypes present in the subject bind to one or more tumor-specific neoepitopes present in the at least one neoantigen peptide.

[0044] Embodiments of the present invention relate to compositions and methods using common neoantigens that, unlike common natural (non-mutated) antigens derived from genes differentially expressed in tumors, possess desirable properties such as immunity to centrally resistant immunosuppression and high tumor specificity. This is because these neoantigens are expressed only in tumor tissues, for example, by tumor-specific mutations or splicing defects. Such splicing variants or mutations can generate immunogenic epitopes across multiple HLA alleles, thus covering a significant proportion of the population. Furthermore, because these mutations can be present in a significant proportion of subjects with cancer, the compositions described herein do not require whole-genome sequencing of the subject and can be used as “off-the-shelf” products to treat multiple subjects. For example, the method could simply involve detecting one or more specific mutations present in the composition in a tumor sample from the subject and administering the composition to a subject with at least one mutation present in vivo. This contrasts with methods using patient-specific neoantigen mixtures, which require whole-genome or whole-exome sequencing for each subject and the production of personalized therapeutic compositions.

[0045] Other embodiments relate to combination therapies in which a treatment using the common neoantigen composition of the present invention is used in conjunction with a current pharmaceutical regimen. The common neoantigen composition may be administered prophylactically. In one embodiment, patients in need are treated with chemotherapy and / or targeted therapy in combination with the common neoantigen immunogenic composition before resistance mutations can be detected. The common neoantigen immunogenic composition may be tailored to include neoantigen peptides specific for resistance mutations associated with the selected therapy. In another embodiment, the common neoantigen composition is administered to a subject prior to treatment with chemotherapy and / or targeted therapy to induce an immune response against cells carrying drug resistance mutations prior to such cell development. Administration may be sequential, sequential, or substantially at the same time or substantially simultaneously. For example, administration of the common neoantigen immunogenic composition and administration of cancer therapy may be performed at approximately the same time or substantially simultaneously. Alternatively, the common neoantigen immunogenic composition can be administered on a schedule, such as weekly, bi-weekly, bi-weekly, monthly, bi-monthly, quarterly (every three months), every third of a year (every four months), every five months, twice a year (every six months), every seven months, every eight months, every nine months, every ten months, every eleven months, annually, etc., and cancer therapy can be administered according to different schedules typical for that therapy. This allows the subject or patient to have two different treatment regimens being administered simultaneously, and the administration of the common neoantigen immunogenic composition and cancer therapy can be carried out sequentially or consecutively. In a preferred embodiment, the subject can be treated with ilotinib, erlotinib, imatinib, gefitinib, crizotinib, trastuzumab, vemurafenib, RAF / MEK, or anti-estrogen therapy.

[0046] In another aspect, the present invention provides a diagnostic method for the early detection and tracking of cancer progression by determining the presence of at least one neoantigen peptide of the present invention in a patient sample. The patient sample may be derived from blood, sputum, saliva, urine, tumor tissue, lymph, semen, or feces.

[0047] In one embodiment, the diagnostic method is used prior to administration of the common neoantigen composition described herein. The diagnostic method may include comparing the amount of common neoantigen mutations in a series of at least two samples obtained during treatment with cancer therapy and / or the common neoantigen composition. Without being bound by theory, an increase or decrease in common neoantigen mutations may be used to determine treatment efficacy.

[0048] In one embodiment, PCR-based methods or sequencing can be used to detect the mutated gene. Reverse transcription PCR (RT-PCR) can be used to detect mutations in transcribed neoantigen genes. Alternatively, any sequencing technology can be used to determine the presence of a mutation. In a preferred embodiment, pyrosequencing is used. The present invention also provides a kit comprising primers specific to sequences containing these neoantigen mutations.

[0049] In another embodiment, the mutated gene is detected by an immunological assay. Antibodies specific to these common neoantigen mutations can be used to detect these mutations. These antibodies can be bound to an array. The array can include antibodies for detecting more than one common neoantigen mutation of the present invention. These antibodies can be configured for use in ELISA assays. Thus, compositions or kits comprising antibodies that specifically recognize the common neoantigens of the present invention can be provided.

[0050] In another aspect, the present invention provides a method for treating or preventing tumors in a population of subjects in need, the method comprising administering to the subject an agent comprising an extracellular ligand-binding domain that recognizes a tumor-specific neoepitope comprising a tumor-specific mutation occurring in at least 1% of the population. The agent may be an antibody, antibody fragment, antibody-drug conjugate, aptamer, CAR, or T-cell receptor. The antibody or antibody fragment may be humanized, fully humanized, or chimeric. The antibody fragment may be a nanobody, Fab, Fab', (Fab')2, Fv, ScFv, bisomatic antibody, trisomatic antibody, tetrasomatic antibody, bi-scFv, mini-antibody, Fab2, or Fab3 fragment. The tumor-specific mutation may be any of the mutations listed in Table 9 for any population. The tumor-specific mutation may be within a gene containing an extracellular domain. Tumor-specific mutations can be FGFR3 S249C, ERBB3 V104M, EGFR L858R, MUC4H4205Q, PDGFRA R483fs, TMEM52 23_26LLPL>L, or PODXL 28_30PSP>P. Tumor-specific mutations can occur within extracellular domains. Tumor-specific mutations include FGFR3 S249C or ERBB3V104M. Unbound by theory, the presence of novel epitopes in proteins with extracellular domains allows these epitopes to exist on the cell surface. Unbound by theory, the presence of novel epitopes in extracellular domains allows these epitopes to exist on the cell surface.

[0051] The present invention is further described in the following numbered paragraphs.

[0052] 1. An isolated neoantigen peptide comprising a tumor-specific neoepitope as defined in Tables 1-9, wherein the isolated neoantigen peptide is not a natural polypeptide.

[0053] 2. A separate neoantigen peptide of 100 amino acids or less in length, comprising tumor-specific neoepitopes as defined in Tables 1-9.

[0054] 3. The isolated neoantigen peptide as described in paragraph 1 or 2, having a length between about 5 and about 50 amino acids.

[0055] 4. The isolated neoantigen peptide as described in any one of paragraphs 1-3, having a length between about 15 and about 35 amino acids.

[0056] 5. The isolated neoantigen peptide as described in paragraph 4, having a length of about 15 amino acids or less.

[0057] 6. The isolated neoantigen peptide as described in paragraph 5, having a length between about 8 and about 11 amino acids.

[0058] 7. The isolated neoantigen peptide as described in paragraph 6 has a length of 9 or 10 amino acids.

[0059] 8. The isolated neoantigen peptide as described in any one of paragraphs 1-7, which binds to major histocompatibility complex (MHC) class I.

[0060] 9. The isolated neoantigen peptide as described in paragraph 8, which binds to MHCI class with a binding affinity of less than about 500 nM.

[0061] 10. The isolated neoantigen peptide as described in any one of paragraphs 1-3, having a length of about 30 amino acids or less.

[0062] 11. The isolated neoantigen peptide as described in paragraph 10, having a length between about 6 and about 25 amino acids.

[0063] 12. The isolated neoantigen peptide as described in paragraph 11 has a length between about 15 and about 24 amino acids.

[0064] 13. The isolated neoantigen peptide as described in paragraph 11 has a length between about 9 and about 15 amino acids.

[0065] 14. The isolated neoantigen peptide as described in any one of paragraphs 1-3 and 10-13, which binds to MHC class II.

[0066] 15. The isolated neoantigen peptide as described in paragraph 14, which binds to MHC class II with a binding affinity of less than about 1000 nM.

[0067] 16. The isolated neoantigen peptide as described in any one of paragraphs 1-15, further comprising flanking amino acids.

[0068] 17. The isolated neoantigen peptides as described in paragraph 16, wherein these flanking amino acids are not natural flanking amino acids.

[0069] 18. The isolated neoantigen peptide as described in any one of paragraphs 1-17, which is linked to at least a second neoantigen peptide.

[0070] 19. The isolated neoantigen peptide as described in paragraph 18, wherein the peptide is linked using a poly-glycine or poly-serine linker.

[0071] 20. The isolated neoantigen peptide as described in paragraph 18 or 19, wherein the second neoantigen peptide binds to MHCI or II classes with a binding affinity of less than about 1000 nM.

[0072] 21. The isolated neoantigen peptide as described in paragraph 20, wherein the second neoantigen peptide binds to class MH1 or class II with a binding affinity of less than about 500 nM.

[0073] 22. The isolated neoantigen peptide as described in paragraph 20 or 21, wherein both neoepois bind to human leukocyte antigen (HLA)-A,-B,-C,-DP,-DQ, or-DR.

[0074] 23. The isolated neoantigen peptide as described in any one of paragraphs 20-22, wherein the isolated neoantigen peptide and the second neoantigen peptide bind to class I HLA, or the isolated neoantigen peptide and the second neoantigen peptide bind to class II HLA.

[0075] 24. The isolated neoantigen peptide as described in any one of paragraphs 20-22, wherein the isolated neoantigen peptide binds to class II HLA and the second neoantigen peptide binds to class I HLA, or the isolated neoantigen peptide binds to class I HLA and the second neoantigen peptide binds to class II HLA.

[0076] 25. The isolated neoantigen peptide as described in any one of paragraphs 1-24, further comprising modifications that increase in vivo half-life, cell targeting, antigen uptake, antigen processing, MHC affinity, MHC stability, or antigen presentation.

[0077] 26. The isolated neoantigen peptide as described in paragraph 25, wherein the modification is conjugation to a carrier protein, conjugation to a ligand, conjugation to an antibody, PEGylation, polysialylation / HES-ization, recombinant PEG mimicry, Fc fusion, albumin fusion, nanoparticle attachment, nanoparticle encapsulation, cholesterol fusion, iron fusion, acylation, amidation, glycosylation, side chain oxidation, phosphorylation, biotinylation, addition of a surfactant, addition of an amino acid mimicry, or addition of a non-natural amino acid.

[0078] 27. The isolated neoantigen peptide as described in paragraph 25, wherein the targeted cells are antigen-presenting cells.

[0079] 28. Isolated neoantigen peptides as described in paragraph 27, wherein the antigen-presenting cells are dendritic cells.

[0080] 29. The isolated neoantigen peptides as described in paragraph 29, wherein these dendritic cells are targeted using CD141, DEC205, or XCR1 markers.

[0081] 30. A pharmaceutical composition comprising at least one neoantigen peptide and a pharmaceutically acceptable carrier, wherein each of the at least one neoantigen peptide comprises a tumor-specific neoepitope capable of binding to an HLA protein in a subject, and each tumor-specific neoepitope comprises a tumor-specific mutation present in a tumor, wherein:

[0082] (a) The composition includes at least one neoantigen peptide, which includes a tumor-specific mutation present in the tumors of at least 1% of subjects in a subject population with cancer;

[0083] (b) The composition comprises at least one neoantigen peptide containing a tumor-specific neoepitope that binds to an HLA protein present in at least 5% of subjects in a subject population with cancer; or

[0084] (c) The composition includes at least one neoantigen peptide capable of eliciting an immune response against tumors present in at least 5% of subjects in a subject population with cancer.

[0085] 31. The pharmaceutical composition as described in paragraph 30, wherein the subject population has adrenocortical carcinoma (ACC), bladder urothelial carcinoma (BLCA), invasive breast carcinoma (BRCA), cervical squamous cell carcinoma and cervical adenocarcinoma (CESC), colonic adenocarcinoma (COAD), chronic lymphocytic leukemia (CLL), colorectal cancer (CRC), diffuse large B-cell lymphoma (DLBCL), glioblastoma multiforme (GBM), head and neck squamous cell carcinoma (HNSC), renal chromophobe carcinoma (KICH), renal clear cell carcinoma (KIRC), and kidney... The following are considered cancers: renal papillary cell carcinoma (KIRP), acute myeloid leukemia (LAML), hepatocellular carcinoma (LIHC), lung adenocarcinoma (LUAD), lung squamous cell carcinoma (LUSC), multiple myeloma (MM), ovarian serous cystadenocarcinoma (OV), pancreatic cancer (PAAD), prostate adenocarcinoma (PRAD), rectal adenocarcinoma (READ), skin melanoma (SKCM), gastric adenocarcinoma (STAD), testicular germ cell tumor (TGCT), thyroid adenocarcinoma (THCA), endometrioid carcinoma of the uterine corpus (UCEC) or uterine carcinosarcoma (UCS).

[0086] 32. The pharmaceutical composition as described in paragraph 30 or 31, wherein the population with cancer has been, is, or has been selected as a candidate cancer treatment agent, optionally treated with erlotinib, erlotinib, imatinib, gefitinib, crizotinib, trastuzumab, vemurafenib, RAF / MEK inhibitors, or anti-estrogenic therapy.

[0087] 33. The pharmaceutical composition as described in any one of paragraphs 30-33, wherein the tumor-specific mutation includes splice variant mutations, point mutations, and / or frameshift mutations.

[0088] 34. The pharmaceutical composition of any one of paragraphs 30-33, wherein the at least one neoantigen peptide comprises at least one neoantigen peptide derived from a long peptide region flanking and including the tumor-specific mutation, and including all consecutive segments within the long peptide.

[0089] 35. The pharmaceutical composition as described in any one of paragraphs 30-34, wherein a tumor-specific mutation is present in one or more genes listed in Tables 1-9.

[0090] 36. The pharmaceutical composition as described in any one of paragraphs 30-35, wherein the composition comprises at least one neoantigen peptide as defined in any one of Tables 1-9.

[0091] 37. The pharmaceutical composition of any one of paragraphs 30-36, wherein a tumor-specific mutation is present in one or more genes encoding proteins selected from the group consisting of: programmed death-ligand 1 (PD-L1), androgen receptor (AR), Bruton's tyrosine kinase (BTK), epidermal growth factor receptor (EGFR), BCR-Abl, c-kit, PIK3CA, HER2, EML4-ALK, KRAS, ALK, ROS1, AKT1, BRAF, MEK1, MEK2, NRAS, RAC1, and ESR1.

[0092] 38. The pharmaceutical composition as described in paragraph 37, wherein at least one tumor-specific mutation originates from alternative splicing of PD-L1 or AR.

[0093] 39. The pharmaceutical composition as described in paragraph 38, wherein at least one tumor-specific mutation originates from a splice variant sPD-L1, AR-V1, or AR-V7.

[0094] 40. The pharmaceutical composition as described in any one of paragraphs 30-39, wherein the tumor-specific mutation includes a drug resistance mutation.

[0095] 41. The pharmaceutical composition as described in paragraph 40, wherein at least one tumor-specific mutation is a drug resistance mutation selected from the group consisting of: BTK / C481S, EGFR / T790M, BCR-Abl / T315I, BCR-Abl / Y253H, BCR-Abl / E255K, BCR-Abl / E255V, c-kit / T670I, PIK3CA / E545K, PIK3CA / E542K, HER2 / G776(YVMA), HER2 / E545K, EML4-ALK / G1269A, KRAS / G12V / D, ALK / L1196M, ALK / G1202R, ALK / S1206Y, ALK / 1151T(ins ), ALK / F1174C, ROS1 / G2032R, AKT1 / E17K, BRAF / V600E, MEK1 / Q56P, MEK1 / E203K, MEK1 / C121S, MEK1 / V60E, MEK1 / G128V, MEK1 / V154I, MEK1 / P124S, MEK1 / P124L, NR AS / Q61K / L / R, NRAS / T58I, MEK2 / C125S, RAC1 / P29S, ESR1 / S463P, AR / V534E, AR / P 535H, AR / L536Q, AR / L536R, AR / Y537C, AR / Y537S, AR / Y537N, AR / D538G and AR / F876L.

[0096] 42. The pharmaceutical composition of any one of paragraphs 30-41, wherein the at least one tumor-specific mutation occurs in at least 500 patients per year in a subject population with cancer, and wherein the at least one mutation includes mutations listed for the populations in Table 9.

[0097] 43. The pharmaceutical composition as described in paragraph 42, wherein the at least one neoantigen peptide comprises at least one peptide listed in Table 9.

[0098] 44. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0099] (a) This subject population has CLL; and

[0100] (b) The at least one tumor-specific mutation comprises any combination of mutations selected from the group consisting of: SF3B1: p.K700E, MYD88: p.L273P, NOTCH1: p.P2514fs, ABCA11P: p.E901D, AHNAK: p.D3823E, ZNF814: p.E348D, AHNAK: p.V1220I, AHNAK: p.H1203N, ANKRD30A: p.A232V, APOOL: p.I138L, EGR2: p.H397N, MKI67: p.H2213D, NRAS: p.Q61R, PLIN4: p.M691V, XPO 1:p.E571K、ZCRB1:p.L76F、ZNF700:p.N652H、ZNF700:p.Q654R、ZNF844:p.D458H、AHNAK:p.A4046V、ANKRD36:p.P337R、C1orf170:p.T203I、CAST:p.D 639E, EGR2:p.E369K, GPR123:p.L630P, IKZF3:p.L162R, MUC4:p.P4224R, OR9Q1:p.M34L, PKD2:p.Y486F, PRAMEF11:p.R104Q, SYNJ1:p.I681F, TP53:p. R248Q、TP53:p.R248W、TRPV2:p.L627del、ZNF254:p.S498A、ZNF732:p.A459T、ZNF749:p.E530Q、ZNF845:p.M423I、ABCA11P:p.G900E、ACRC:p.E243D、 ACRC: p.A244V, ACSL3: p.T188S, ADAMTS2: p.D948N, AGAP6: p.S127I, AHNAK: p.A2114G, ANKRD36: p.D1014Y, ARID3A: p.G550fs, ARID4A: p.D1154E, ATP2 B4:p.R183H, ATRNL1:p.L1244F, BNC1:p.Y937N, BRAF:p.K601N, BTLA:p.Q86K, C14orf177:p.G90V, C2orf44:p.N456K, C3orf15:p.R552Q, CACNA2D1:p. Y376N, CALD1:p.E340K, CCDC15:p.P488H, CCDC79:p.N440T, CCNB3:p.A932T, CD109:p.L470Q, CD209:p.Q189L, CKAP2:p.*684K, CMA1:p.I81K, CMIP:p.A230T、CNTNAP4:p.I12F、CRYM:p.*315K、DICER1:p.E1705K、DPCR1:p.L716P、EIF3A:p.M1093L、EIF4G3:p.R8H、ETFDH:p.I281F、EWSR1:p.Y656C、F5:p.L1332P、F5:p.L1253F、FAM50A:p.H317R、FBXL13:p.S102R、FBXW7:p.R465H、FHL1:p.D184E、FILIP1:p.I522K、FRG1B:p.Q39K、GNB1:p.I80T、GPR110:p.R443G、GPR98:p.Y6152F、HDGFL1:p.188_189insA、IGF2BP2:p.T186S、IL1R2:p.L364fs、KIAA1109:p.L4680P、KRAS:p.G13D、KRTAP19-1:p.G61S、MAF:p.G53fs、MAGEC1:p.L609H、MAP2K1:p.K57N、MED12:p.L36R、MED12:p.G44S、METAP2:p.Y137N、METTL9:p.Y57F、MGP:p.V15L、MKI67:p.R2222K、MUC16:p.T11005I、MUC4:p.S3941N、MUC4:p.S3941G、MUC4:p.V3091L、MUC4:p.S2951Y、MUC4:p.A2841S、MUC4:p.S2760A、MUC4:p.T2335M、MUC4:p.T1627K、MUC4:p.T1547S、MUC4:p.H1133Q、MYD88:p.M240T、NEDD4L:p.P194del、NEFH:p.S704T、NRG4:p.G21fs、OR2A25:p.S105C、OR4C16:p.Y63F、OR4N4:p.L150fs、PABPC1:p.K254fs、PIWIL1:p.P372fs、PLCD3:p.E499fs、PLEKHB1:p.S146P、PPIL4:p.S382R、PRDM4:p.*802K、PRG4:p.N675H、PRKAB1:p.P104H、R3HDM2:p.S592G、R3HDM2:p.S588N、R3HDM2:p.R206W、RPS2:p.R200G、RPTN:p.G364S、SF3B1:p.K666E、SF3B1:p.N626Y、SF3B1:p.Y623C、SIX3:p.I27L, SLC39A7: p.L456fs, SLC6A9: p.R94K, TFG: p.A382V, TGOLN2: p.K83R, TGOLN2: p.T80S, TLR2: p.D327V, TNKS2:p.T619fs, TP53:p.R273H, TP53:p.C242F, TP53:p.R175H, TWISTNB:p.H306Q, UBXN 7: p.A276V, WDR78: p.N110K, XIRP2: p.V3008E, ZNF382: p.H186Q, ZNF578: p.R306H, ZNF578: p.G311S, ZNF578: p.H334R, ZNF700: p.S649C, ZNF705A: p.D298N, ZNF836: p.K608Q, and ZNF836: p.I571N; and...

[0101] 45. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0102] (a) This subject population has BLCA; and

[0103] (b) The at least one tumor-specific mutation comprises any combination of mutations selected from the group consisting of: PIK3CA:p.E545K, FGFR3:p.S249C, TP53:p.R248Q, PIK3CA:p.E542K, RXRA:p.S427F, ZNF814:p.D404E, FBXW7:p.R505G, NOTCH2:p.P6fs, TP53:p.E285K, ANKRD30A:p.A353P, C3orf70:p.S6L, EFCAB6:p.R379K, ERCC2:p.N238S, FAM47C:p.Q225E, FOXQ1: p.S135L, HLA-A: p.Q78R, MUC4: p.H4205Q, OTUD4: p.T909I, SLAMF1: p.S277fs, SPRED3: p.S128del, TMCO2: p.S15fs, TP53: p.R280T, TP53: p.E271K, T P53:p.A159V、ZNF706:p.I8N、ZNF706:p.R3P、ACACB:p.E2318Q、ACPP:p.E321K、ACRC:p.A264V、ADAMTS2:p.23_24insL、AFF3:p.E919K、AHNAK:p.S415 0F, AHNAK:p.D2889H, AHNAK:p.V1940A, ALX4:p.R126Q, ANKRD12:p.E627K, ANKRD32:p.T999N, ARID1A:p.S614L, ASXL2:p.117_118SS>S, ATP12A:p.R 858C, ATP9A:p.R519Q, BCAS3:p.T214M, BPI:p.M255I, CACNG8:p.V146G, CAMSAP1:p.T466fs, CDC27:p.I91fs, CDKN1A:p.E44fs, CEP192:p.S2058L, CG B8: p.T18A, CHRNA3: p.L23del, CHST4: p.D352N, CLIP1: p.S1018fs, COX6A1: p.S8L, CREBBP: p.D1435H, CRIPAK: p.M48fs, CSPG5: p.D119N, CUL1: p.E48 5K, DLC1:p.S741T, DLL3:p.D318H, DOPEY2:p.E1196K, ECM1:p.E266K, EEF1A2:p.Y418S, EEF2K:p.E673K, EMILIN1:p.R27G, ERBB2:p.S310F, ERBB3:p.M91I、ERBB3:p.V104L、ERBB3:p.D297Y、ERCC2:p.Y14C、FAM155A:p.Q86del、FAM43B:p.E272del、FASTKD3:p.Q625E、FBXW7:p.S546L、FGFR3:p.R248C、 FGFR3:p.G380R、FGFRL1:p.H479fs、GBE1:p.M587I、GIMAP1-GIMAP5:p.S311C、GNA13:p.R200G、H1FOO:p.A214fs、HEATR7B2:p.E1109K、HIST1H1D:p.I 81M、HRAS:p.G12D、HRCT1:p.H92P、ILF3:p.E484K、KCNK2:p.S6W、KIAA0907:p.Q446P、KIF23:p.E350K、KLF5:p.S118L、KLHL15:p.D185G、LAMA4:p.E63 9K、LILRA1:p.H410Y、LILRB1:p.L479del、LLGL2:p.P955fs、LPIN1:p.S974L、LRRC16A:p.D227N、LRTM2:p.S139L、LURAP1L:p.55_56insGGG、MAGEC1:p .P553del、MCL1:p.E171del、MN1:p.S472L、MUC7:p.A191V、MVP:p.E412K、NBPF10:p.E3455K、NFE2L2:p.E79K、NFE2L2:p.R34G、NOS1AP:p.Q306del、OR 2T35:p.V319fs、OR4N2:p.L150fs、PABPC3:p.K333fs、PAX3:p.S197L、PBX2:p.E70K、PBXIP1:p.H729del、PCDP1:p.E537K、PEX1:p.I370fs、PHLDA3:p. E82K, PLEKHM2:p.S459L, PLVAP:p.A321V, POLR3B:p.L372F, POTEC:p.R477Q, PPL:p.H326Y, PPP1R15A:p.E196K, PRDM16:p.E271Q, PRIC285:p.E1289Q, PRMT8:p.S31P, PUF60:p.S396L, RAB11FIP4:p.S596L, RAD51C:p.D167N, RAD51C:p.Y224H, RALGPS1:p.R381Q, RARS2:p.R6C, RBM26:p.P644A, RERE:p.K176N, RXRA:p.S427Y, SERPINA12:p.R211G, SF3B1:p.E902K, SLC6A9:p.R243W, SLC9A5:p.L447F, SPESP 1:p.F121L, SRPRB:p.G14S, SYN2:p.A34del, SYTL2:p.I440M, TAB3:p.R211T, TAF1B:p.R292C, TAOK2:p.L 981del, TAS1R3:p.E525K, TAS2R9:p.E163Q, TBC1D1:p.S71F, TBC1D2B:p.R920Q, TFPI2:p.R222C, TM6SF 1:p.S15W, TMEM131:p.K640fs, TMEM19:p.G331fs, TP53:p.R273C, TP53:p.R248W, TP53:p.R175H, TP53:p .K132N、TRAM1:p.E41Q、TSKS:p.E513K、TTN:p.C20935G、UBOX5:p.S417L、UGP2:p.D262H、VGF:p.E433K、 XAB2:p.E782K, 749:p.Q457E、ZNF761:p.H373R、ZNF799:p.T43A、ZNF799:p.W41G、ZNF799:p.E589G、ZNF844:p.P503R、ZN F845:p.M423T, ZNF845:p.T479M, ZNF860:p.H464R, ZNF878:p.S181R, ZNF91:p.R333H, and ZNF91:p.H305R. .

[0104] 46. ​​The pharmaceutical composition as described in any one of paragraphs 30-43, wherein:

[0105] (a) This subject population has BRCA; and

[0106] (b) The at least one tumor-specific mutation includes any combination of frameshift mutations selected from the group consisting of: GATA3:p.L328fs, GATA3:p.N334fs, GATA3:p.L344fs, GATA3:p.H400fs, GATA3:p.S408fs, GATA3:p.S430fs, GATA3:p.H434fs, GATA3:p.H435fs, and GATA3:p.S408fs.

[0107] 47. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0108] (a) This subject population has BRCA; and

[0109] (b) The at least one tumor-specific mutation comprises any combination of mutations selected from the group consisting of: PIK3CA:p.H1047R, PIK3CA:p.E545K, PIK3CA:p.E542K, AKT1:p.E17K, TP53:p.R175H, PIK3CA:p.N345K, PIK3CA:p.H1047L, SF3B1:p.K700E, GATA3:p.S408fs, PIK3CA:p.E726K, TP53:p.Y220C, TP53:p.H193R, PIK3CA:p.Q546R, TP53:p.R273C, TP53:p.R 248W, TP53:p.R273H, TP53:p.I195T, TP53:p.H179R, FGFR2:p.N549K, NUP93:p.E14K, PIK3CA:p.C420R, PIK3CA:p.E453K, PIK3CA:p.Q546K, TP53:p.V 216M, TP53:p.C176F, CDH1:p.E243K, ERBB2:p.L755S, KRAS:p.G12V, PIK3CA:p.E545A, TBL1XR1:p.I141fs, TP53:p.G266E, TP53:p.R248Q, TP53:p.Y1 63C, TP53:p.C141Y, TP53:p.G108fs, ACPP:p.R43W, AKT2:p.I289M, ARHGAP9:p.R137C, C9orf174:p.R136W, CDC42BPA:p.P675T, COL12A1:p.S395L, CR ISPLD1:p.R222W、CT47B1:p.234_243EKLTEEATEE>E、CYP1A2:p.V483M、DAB2IP:p.E161K、DGKB:p.S13L、DMD:p.K1772N、DPEP1:p.V11L、ERBB2:p.S310 F. ERBB2:p.D769Y, ERBB3:p.E928G, ESYT1:p.R816W, FAM179A:p.A831T, FAM58BP:p.A70T, FMN2:p.S751F, GALNTL6:p.K567del, GATA3:p.L328fs, GAT A3:p.N334fs、GATA3:p.L344fs、GATA3:p.H400fs、GATA3:p.S408fs、GATA3:p.S430fs、GATA3:p.H434fs、GATA3:p.H435fs、GDAP1:p.T307A、GRB14:p.A300T、GUCY2C:p.G549C、IL17B:p.R34W、KCNB2:p.R231H、KIF1B:p.R1320W、KIF26B:p.V1113M、KLF4:p.K434Q、LY9:p.I69L、MAP2K4:p.S184L、MAP2K4:p.S251I、MAP2K4:p.T261fs、MAP3K1:p.L318fs、MAP3K1:p.I761fs、MAP3K1:p.V1346del、MAP3K1:p.L1384fs、MAPK13:p.E315K、MAPK4:p.V100M、MARCH5:p.R170C、MBP:p.E120K、MEFV:p.R377H、METTL15:p.Q53E、MS4A4A:p.V99M、MUC17:p.R4415H、MYH6:p.T847M、MYO5B:p.A405V、NARS2:p.P240R、NLGN4X:p.D382N、NLRC4:p.R288W、OR13G1:p.R258H、OR2AK2:p.V45I、OTOF:p.T388M、PACSIN2:p.Q331H、PALM2-AKAP2:p.A299T、PCDH19:p.R286C、PCDHGC5:p.D664N、PIK3CA:p.R88Q、PIK3CA:p.E110del、PIK3CA:p.K111del、PIK3CA:p.PVPHGLEDL447del、PIK3CA:p.L455fs、PIK3CA:p.M1004I、PIK3CA:p.M1043I、PIK3CA:p.N1044Y、PIK3R1:p.KPDL567del、PREX2:p.R363Q、PRRX1:p.A196V、PTEN:p.V317fs、RGSL1:p.V222I、RUNX1:p.R142fs、RUNX1:p.D96fs、SCN2A:p.R36K、SLC25A32:p.Q83E、SLC25A45:p.G106C、STRA6:p.Q68R、STX6:p.H153D、TBX3:p.H187Y、TFPT:p.S252C、TINAG:p.R332W、TMEM71:p.R63Q、TP53:p.E286K、TP53:p.R282W、TP53:p.V272M、TP53:p.S241fs、TP53:p.C238fs、TP53:p.C238F、TP53:p.C238Y、TP53:p.Y234C、TP53:p.Y220S, TP53:p.R209fs, TP53:p.G199V, TP53:p.L194R, TP53:p.H193L, TP53:p.H193Y, TP53:p.V173L, TP53:p.V173M, TP53:p. K132N, TP53:p.R110fs, TUBD1:p.A200V, VLDLR:p.R231H, VWA3A:p.V955I, VWF:p.K1720N, XPO1:p.E571K, and ZNF268:p.F901del. .

[0110] 48. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0111] (a) This group of subjects had COAD; and

[0112] (b) The at least one tumor-specific mutation includes any combination of mutations selected from the group consisting of: KRAS: p.G12D, BRAF: p.V600E, KRAS: p.G12V, ACVR2A: p.K435fs, GRB14: p.KKK295del, SEC63: p.L532fs, TGFBR2: p.E125fs, ATR: p.K771fs, ICA1: p.N204fs, KRAS: p.G12C, TP53: p.R175H, ABCA8: p.R842Q, ACTL7B: p.R354H, ACVR2A: p.K435fs, AIM2: p. K340fs, ALG2:p.S302Y, ANKIB1:p.K144fs, ARSG:p.V131I, ATP10D:p.R311H, AXIN2:p.W663fs, C5orf30:p.D4N, CACNG3:p.V134I, CASP5:p.K78fs, C C2D2A:p.R1284C, CDH10:p.E349K, DNMT1:p.E432K, DOCK2:p.G170R, DOCK5:p.E177K, EGR2:p.R390H, ERBB3:p.V104M, FAM135B:p.R884H, FBXW7:p.R 505C, FBXW7: p.R465H, FHDC1: p.R254W, FOXL1: p.N89K, HCN4: p.R525H, HLA-DMA: p.E84K, HTR3B: p.R236C, ITGA4: p.T673M, KIF18A: p.R17C, KIF20B: p.E991K、KLHL5:p.R326C、KRAS:p.A146T、KRAS:p.G13D、LPHN3:p.R1183Q、MAP2K4:p.R287H、MAPK8IP1:p.L217fs、MFSD5:p.R280Q、MUC16:p.R8606H , MYO6: p.D1180N, NAA25: p.S807Y, NBPF14: p.V44L, NRAS: p.Q61K, NRAS: p.G13R, PAX3: p.T424M, PGAM1: p.R240H, PHF3: p.R1410I, PIK3CA: p.R88Q, P IK3CA:p.E545K, PIK3CA:p.H1047R, PLXNA3:p.V14fs, POSTN:p.R508C, PTPRU:p.D1434N, PYGO2:p.Q150fs, RBBP7:p.E274K, SFPQ:p.R611Q, SGSM1:p.F1117L, SLC25A40:p.R96Q, SLC8A1:p.R431H, SLITRK3:p.S298L, SPATA22:p.S150L, ​​SUN3:p.E128K, TGFBR1:p.S241L, TP53:p.R273H, TP53:p.R273C, TP53 :p.R248W、TRPV5:p.R492H、USP40:p.S851L、VPS13C:p.D1359Y、ZBTB24:p.L6 07I, ZNF434:p.R306C, ZNF443:p.R301I, ZNF484:p.R138C, and ZNF770:p.S441P. .

[0113] 49. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0114] (a) This subject population has GBM; and

[0115] (b) The at least one tumor-specific mutation includes any combination of mutations selected from the group consisting of: HSD17B7P2: p.N175S, IDH1: p.R132H, EGFR: p.A289V, EGFR: p.G598V, WASH3P: p.G175S, ZNF814: p.D404E, RPSA: p.Q111E, NBPF10: p.E3455K, TP53: p.R248Q, BRAF: p.V600E, EGFR: p.A289T, PRB2: p.N230del, RGPD5: p.P1760A, TP53: p.R175H, CHEK2: p.K37 3E, EGFR: p.R108K, EGFR: p.R222C, PIK3CA: p.E545K, PIK3R1: p.G376R, POTEC: p.K507E, SDHAP2: p.V195E, SLC6A10P: p.K88N, TP53: p.R282W, TP53:p. R273H, CD3EAP:p.K219del, DST:p.R146C, EGFR:p.A289D, EGFR:p.H304Y, FRG1B:p.S71N, GOLGA8DP:p.A116E, KRTAP4-11:p.R121K, KRTAP4-11:p.S48R , MAP3K1:p.P324L, OGDH:p.I78fs, PODXL:p.S162fs, PSPH:p.V145I, SPINT1:p.A316V, TP53:p.R248W, TP53:p.G245S, TP53:p.Y220C, TP53:p.R158H, TSHZ2:p.A222T、UBC:p.L149R、ZDHHC4:p.R300H、ZNF844:p.R447P、AASS:p.T878fs、ABCC10:p.R570W、ADAM29:p.V205I、ADAMTS8:p.V524M、AGAP3:p.R 766W, AICDA:p.Y144F, AK7:p.A159V, AK8:p.D243A, ANO2:p.R334C, AOX1:p.A507V, ARHGAP5:p.M691L, CALN1:p.V231I, CARM1:p.A202V, CD163L1:p.V7 21M, CD1D:p.L25fs, CD209:p.A283T, CDH18:p.A195T, CILP2:p.V553M, CIZ1:p.L89P, CLOCK:p.L123fs, COL6A5:p.T2224M, CSF2RB:p.G298S, CSMD3:p.E171K、CYP2D6:p.H352R、DCAF12L1:p.R335H、DCAF12L2:p.R246H、DPP10:p.V183I、DPY19L2P1:p.R378Q、DQX1:p.R505H、DRD5:p.S275R、DVL2:p.V66G、EFCAB6:p.R379K、EGFR:p.L62R、EGFR:p.R252C、EGFR:p.P596S、EGFR:p.P596L、EGFR:p.G598A、EGFR:p.E709K、EPHA1:p.A184T、ERC2:p.R20H、ESPNP:p.R627Q、FAM126B:p.R382H、FBN3:p.V886I、FGF14:p.T229M、FLG2:p.H1901fs、FLG:p.R2886H、FLNA:p.V1240M、FOXG1:p.H57del、FPR2:p.R54Q、FRG1B:p.K13N、FRG1B:p.A53T、GABRA6:p.V314I、GJB3:p.R160H、GLT8D2:p.A178V、GRM3:p.R183C、HERC1:p.R2330H、HNF1B:p.T417M、HTRA3:p.Q403R、IDH1:p.R132G、IFNA10:p.L80F、IFNA10:p.V79A、JHDM1D:p.R313H、JPH1:p.A395T、KEL:p.V411M、KIAA0907:p.R516fs、KIAA1704:p.D88del、KLK6:p.R120H、KRAS:p.G12D、KRTAP4-7:p.L121V、KRTAP4-7:p.L148V、KRTAP5-4:p.S131C、LAT2:p.L18W、LIMK2:p.R203H、LUM:p.R330C、MCOLN3:p.V141I、MGAT4B:p.T444P、MUC17:p.V77M、MUC17:p.3204_3205insP、MYO1D:p.T109M、MYO6:p.Q914fs、NAP1L5:p.140_141EE>E、NF1:p.F1658fs、NHP2L1:p.R84C、NLRP5:p.R737W、NPTX1:p.A263T、NUFIP2:p.Q29del、ODF4:p.R61C、OR11H12:p.H154P、OR2A7:p.V18I、OR2H1:p.V287I、OR2T12:p.R184H、OR5D13:p.R236C、OR5P2:p.A100V、OR6N2:p.R293C、PASD1:p.A236del、PCDH11X:p.T486M、PCDHB13:p.P221L、PDGFRA:p.E229K、PDGFRB:p.S650L、PHC3:p.T35del、PIK3C2B:p.R287fs、PIK3CA:p.M1V、PIK3CA:p.R88Q、PIK3CA:p.M1043V、PIK3CA:p.H1047R、PIK3R1:p.K379N、PODNL1:p.A150V、POTEE:p.V166M、POTEG:p.R136H、PRKCD:p.G432fs、PROKR2:p.V297I、PTEN:p.C136Y、PTEN:p.S170N、PTEN:p.R173H、PTEN:p.T277I、PTEN:p.V317fs、PTPN14:p.E716del、R3HDM2:p.412_413QQ>Q、RAB11FIP5:p.R170H、RASAL3:p.R82H、RB1:p.N316fs、RDH8:p.A198V、REN:p.15_16LL>L、RIMBP2:p.R830H、SCAF11:p.E926fs、SCN7A:p.R1358H、SCNN1G:p.R564H、SDHAP2:p.R31C、SDHAP3:p.A66T、SEMG2:p.R292C、SH3RF2:p.R318C、SHB:p.A460T、SIGLEC10:p.T250M、SLC13A5:p.Q273P、SLC17A9:p.V324I、SLC22A9:p.R407Q、SLC26A3:p.V88I、SLC5A3:p.A302fs、SLC9A4:p.R631H、SPAM1:p.R346Q、SPEN:p.E803fs、SPTA1:p.A2011V、SUSD5:p.T513M、SYNE1:p.R8468H、TARSL2:p.G366D、TAS2R41:p.A255T、TAT:p.R367H、TFPI2:p.R206C、THSD7B:p.R90C、TMEM147:p.A92V、TMEM156:p.R81C、TMPRSS6:p.V302I、TNFSF9:p.A232T、TP53:p.C238F、TP53:p.C238Y、TP53:p.Y234C、TP53:p.V216M、TP53:p.H179R、TP53:p.T155N, TRAPPC10:p.K133fs, TTN:p.R21402W, TTN:p.V16403M, TUBBP5:p.V102M, TYRP1:p.T352fs, UBC:p.R73L, UGT2B28:p.P289H, USH2A:p.R3719H, WASH6 P:p.L211V、ZFP42:p.V227I、ZFP42:p.T264M、ZNF181:p.V305G、ZNF280B:p.E 400K, ZNF534:p.N583K, ZNF563:p.W208fs, ZNF844:p.F487L, and ZPBP:p.R154C. .

[0116] 50. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0117] (a) This subject population has HNSC; and

[0118] (b) The at least one tumor-specific mutation comprises any combination of mutations selected from the group consisting of: PIK3CA:p.E545K, PIK3CA:p.E542K, TP53:p.R175H, PIK3CA:p.H1047R, TP53:p.R282W, TP53:p.R248Q, TP53:p.R273H, TP53:p.R248W, TP53:p.G245S, RHOA:p.E40Q, EP300:p.D1399N, HRAS:p.G13V, MB21D2:p.Q311E, NFE2L2:p.E79Q, TP53:p.H179Y, FBXW 7:p.R505G, HIST1H2BF:p.E77K, HRAS:p.G12D, MAPK1:p.E322K, NFE2L2:p.D29H, TP53:p.P278S, TP53:p.C242F, TP53:p.Y220C, TP53:p.H193L, TP53:p .H179R, TP53:p.V157F, TP53:p.R110L, AKNAD1:p.K620R, ANXA6:p.R231Q, AP1G2:p.D243N, ATAD5:p.D441N, ATP6AP2:p.E119Q, B2M:p.M1I, BCL11A:p. E579K, C1orf172:p.Y30fs, C7orf57:p.E30K, CCDC135:p.E313K, CDH12:p.P706T, CDH7:p.Q225K, CDK11B:p.E79del, CDKN2A:p.H83Y, CHCHD4:p.T79M, CIRH1A: p.S250I, CLSTN2: p.P759L, CRB1: p.L628fs, DENND5B: p.G1023E, DNAH5: p.Q1797E, DSP: p.R160G, EDA: p.L58F, EFCAB6: p.E1002K, ELF4: p.S41 5L, EP300:p.C1164Y, EPHA3:p.T802R, EPHA6:p.D952H, ERBB2:p.M916I, ESRRA:p.D219N, FAM101A:p.I89del, FBXO24:p.M553V, FCAR:p.V233M, GPANK1 ;K37M、HIST1H4C:p.R68P、HLX:p.S12T、HOXD10:p.Y151C、HPS3:p.K812N、HRAS:p.G12A、HRAS:p.G12S、IFT140:p.E664K、INPPL1:p.T493M、ITGA10:p.R669Q、ITGB1:p.D158N、KIAA1429:p.D1526N、KIAA1429:p.S138F、KPRP:p.E553fs、KSR2:p.T555M、LINGO2:p.P410T、LPCAT1:p.V187del、MAGEB3:p.V75A、MAP3K7:p.E524Q、MAP4K3:p.P657fs、MAP9:p.K485N、MARS2:p.R481Q、MBOAT7:p.R424W、MUC16:p.R12774H、MUC5B:p.T4388M、MYH11:p.E993K、MYOCD:p.T493M、MYOM1:p.R63Q、NANOS3:p.S183L、NCOR1:p.R1561Q、NCOR1:p.Q169E、NCR1:p.D213N、NFE2L2:p.E79K、ODZ1:p.R366M、OPN1MW:p.A285T、OR2M2:p.A95fs、OR2M3:p.M273I、OR2T33:p.R120S、OR6V1:p.I248fs、PABPC5:p.P58L、PACSIN1:p.E359K、PIK3CA:p.M1043V、PIK3CA:p.H1047L、PIWIL1:p.V699M、PLIN5:p.430_431insNG、PLXNA3:p.P58S、PRB1:p.R274fs、PRSS1:p.D107N、RAC1:p.A159V、RGS7:p.L21fs、RPA1:p.R31H、RPL18:p.R178fs、SFI1:p.R821Q、SLC35D3:p.*417S、SLC5A7:p.G336C、SMARCA4:p.P913L、STAT3:p.D661V、SYCP2:p.K474N、SYT6:p.R249H、TBX21:p.E494K、THSD7A:p.R1046C、THSD7A:p.C728F、TMC3:p.R934S、TMTC2:p.T409R、TP53:p.E285K、TP53:p.C275F、TP53:p.R273C、TP53:p.G266E、TP53:p.G262V、TP53:p.R249S, TP53:p.G245V, TP53:p.C238F, TP53:p.M237I, TP53:p.Y236C, TP53:p.Y236D, TP53:p.R196P, TP53:p.PHHERC177del, TP53:p.V173L, T P53:p.V173M, TP53:p.Y163C, TP53:p.P151T, TP53:p.V143M, TP53:p.P58fs, URI1:p.S13fs, ZNF177:p.K384N, ZNF750:p.S96fs, and ZZZ3:p.R5Q. .

[0119] 51. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0120] (a) This subject group has KIRC; and

[0121] (b) The at least one tumor-specific mutation comprises any combination of mutations selected from the group consisting of: WASH3P: p.G175S, VHL: p.L89H, VHL: p.S111N, WDR52: p.V1227G, KRT1: p.552_559YGSGGSSY>Y, KRTAP1-1: p.S34C, PALM2-AKAP2: p.1075_1076insEA, ZNF814: p.D404E, DOPEY2: p.Y2048S, KAT2B: p.W111fs, PABPC1: p.E156fs, PCDHGC5: p.G599V, PIK3CA: p.E545K, RRAD: p.A278E, SIRPA: p.D131del, UQCRFS1: p.I83V, VHL: p.P45L, VHL: p.V74D, VHL: p.R82P, VHL: p.L116fs, VHL: p.L158V, VHL: p.L169P, WD R73: p.DGTRSQ315del, ABCA3: p.E95D, ABCC5: p.L1090fs, ACADS: p.R330H, ACAN: p.G952E, ACSM2A: p.L402fs, ADAM23: p.K380M, ADH1A: p.D154V, AFF3: p.SA620del、AGAP6:p.D69fs、AGAP7:p.E71fs、AHNAK:p.5_6insE、AIDA:p.K247M、ALAS1:p.G302R、ANAPC16:p.R95fs、ANK2:p.N453S、ANKRD36:p.K37 8R, ARHGEF5:p.E487G, ARSD:p.AGV234del, ARSD:p.A234G, ATP2A1:p.G704C, ATP7A:p.Q990fs, AVIL:p.G299fs, AXDND1:p.EQ991del, BAP1:p.N78S, BA P1:p.M1I、BLM:p.H660Q、BMPER:p.RIAL444del、BRK1:p.K70Q、BTRC:p.I416M、C16orf55:p.D118A、C19orf33:p.K102E、C20orf132:p.E382D、C2orf71 :p.1225_1226insS, C6orf132:p.173_182PPPLLLEPPP>P, CASP5:p.R23fs, CATSPER4:p.T425M, CCDC120:p.I8V, CCR5:p.S185I, CCZ1:p.E214D, CD7:p.P174fs、CDAN1:p.L646fs、CDH23:p.F1132Y、CDK5RAP2:p.H1592Q、CENPB:p.E410V、CERCAM:p.A85fs、CHEK2:p.K373E、CHIT1:p.P284fs、CLCN2:p.645_645R>RR、CLUL1:p.G463R、CNTNAP4:p.Y436S、CUL9:p.D1726E、CWC25:p.K364E、CXorf51B:p.V43I、DDX39B:p.F149fs、DIRAS1:p.G79C、DISP2:p.F1021S、DNMBP:p.T78P、DOCK8:p.A177fs、DPCR1:p.H383N、DPCR1:p.L768del、EGFR:p.L838M、ENPEP:p.F289C、ESPNP:p.W122fs、FAM105A:p.H126N、FAM186A:p.IPPQAQELEIPL1556del、FAM194B:p.EEEEYL135del、FAM22F:p.S691del、FAM22F:p.P690fs、FAM47A:p.LRPEPPETGVSH235del、FAM47C:p.P388S、FAM78A:p.W192L、FBXO34:p.Q294fs、FGFR3:p.R571fs、FGFR3:p.P716H、FMN2:p.AIPPPPPLPGA956del、FOXD4L4:p.C405fs、FUT6:p.S140fs、GJA1:p.A311fs、GOLGA5:p.L492I、GPM6A:p.A50V、GPRIN1:p.231_239RKEDPGSLR>R、GRAMD1B:p.P356H、GREB1:p.S344Y、GRM6:p.A718fs、GUSB:p.L501V、GUSB:p.C500R、HBB:p.F86C、HDAC6:p.G977D、HEXDC:p.T482P、HNF1B:p.N302K、HNRPLL:p.M327V、HRC:p.P439fs、HSFX2:p.D92E、IL1RAP:p.F50C、IVL:p.EQQEGQLKHP167del、KANK4:p.S253P、KCNJ18:p.E378K、KIAA1751:p.K97N、KRT1:p.SSYGSGG557del、KRT2:p.L299W、KRT4:p.F154fs、KRTAP10-6:p.49_49P>PSCCAP、KRTAP5-7:p.C120Y、KRTAP9-2:p.CCQP140del、LARS:p.P185fs、LCP1:p.P445fs、LOC338651:p.PHRSHSPPWS102del、LRCH2:p.D717G、LTA4H:p.F107L、LYST:p.Q710H、MAFA:p.207_208HH>H、MAGEC1:p.P239del、MAP2K5:p.Q445R、MAPKAPK2:p.T214fs、MARCKS:p.K152fs、MED12L:p.P2071S、MEGF6:p.A582fs、MGST3:p.G143fs、MLXIPL:p.S790R、MOCOS:p.S849P、MST1R:p.M464V、MTOR:p.C1483F、MTOR:p.L1460P、MUC16:p.P11260A、MUC17:p.R1227fs、MUC17:p.H1228fs、MUC2:p.1480_1481insI、MUC6:p.P1569fs、MYO3A:p.N525S、NBPF3:p.D491V、NCOR1P1:p.L52P、NDUFA4L2:p.G3fs、NEFH:p.651_651K>KAKSPEK、NES:p.V611L、NFAT5:p.Q906E、NOXO1:p.G3fs、NR2C1:p.S270I、NSMCE2:p.Q31fs、NUDT21:p.W13fs、ODZ2:p.W628fs、ONECUT1:p.L424M、OR10A3:p.F73V、OR4F4:p.E15G、OR4N2:p.L150fs、OR51B5:p.A66fs、OR7C1:p.F104fs、PABPC1:p.Y408F、PABPC1:p.K333fs、PABPC1:p.A181T、PABPC3:p.P191T、PALLD:p.A996T、PALM2-AKAP2:p.G1118fs、PARD6A:p.G84fs、PASK:p.T62I、PCDH15:p.C1713F、PCNT:p.G136S、PGM5:p.G426fs、PGPEP1L:p.R164fs、PIK3C2B:p.F1473L、PIK3CA:p.N1044K、PIK3R5:p.L371R、PITRM1:p.P816T、PLIN4:p.T347I、PODXL:p.28_30PSP>P、POLR1C:p.K332Q、POTED:p.I214V、PPM1E:p.R311W、PRKCE:p.Q157fs、PROX1:p.V225D PRRC2C:p.P1883T, PRX:p.P549L, PSD3:p.T563P, PTCH1:p.P689H, RANBP3 :p.L386W、RASGEF1C:p.A188T、RGPD6:p.F946L、RHEB:p.Y35N、RIMBP3:p.A 396del, RIN3:p.L449V, RLIM:p.S501L, RNF17:p.S351C, RUNX2:p.P466H,S CAF1:p.P208fs, SDK1:p.K508fs, SECISBP2:p.D608E, SERPINB3:p.S209C. SESTD1:p.I306M, SFRP4:p.P325fs, SH3KBP1:p.P563fs, SIPA1L3:p.G777A 、SLC13A2:p.L493fs、SLC16A9:p.CVLLGG470del、SLC25A5:p.A118T、SLC44 A5: p.V70F, SLC4A8:p.N229K, SLC52A1:p.G370del, SLC52A2:p.G399fs, SLC 6A10P:p.K88N、SLC6A14:p.A85fs、SLC9B1:p.V446fs、SON:p.VLESSAVT135 9del, SP8:p.G165del, SPAG1:p.353_354insD, SPATA9:p.C189F, SPEG:p.A 992fs、SPTB:p.T1864I、SRA1:p.V110L、STAT6:p.P354fs、STK11IP:p.A155 E, STXBP3:p.E279G, SVIL:p.M93T, SYNE1:p.R8468S, SYNJ2:p.K832T, SYNPO :p.G619fs、TAOK2:p.Q899fs、TAS2R38:p.I311T、TBC1D12:p.F608Y、TBC1D 1:p.H277R、TBC1D3:p.A556fs、TBC1D3C:p.A495fs、TBC1D3F:p.A556fs、TC F7:p.H140P, TDRD10:p.W276C, THRAP3:p.K551R,TMEM102:p.A110P,TMEM1 61B:p.L142P、TMEM230:p.D140G、TMEM47:p.G87S、TRDN:p.*730Y、TTBK1:p.T1065S, UBE2O: p.R1118fs, UBR5: p.T1306fs, UPK3A: p.G272fs, VHL: p.G39S, V HL:p.S65L, VHL:p.N78D, VHL:p.R79P, VHL:p.W88L, VHL:p.L89P, VHL:p.R107P , VHL:p.S111R, VHL:p.H115N, VHL:p.D121Y, VHL:p.G123fs, VHL:p.D126fs, VH L:p.L128H, VHL:p.L135F, VHL:p.I151T, VHL:p.L153P, VHL:p.L158P, VHL:p.Q 164fs、VHL:p.L184P、VHL:p.L188P、WASH6P:p.315_316insAPP、WASH6P:p.T201M、WWP2:p.G458A、ZCCHC6:p.K937N、ZFAND2B:p.I149T、ZFR2:p.Y107N、ZNF27 3:p.N319K, ZNF462:p.S650T, ZNF516:p.A256D, ZNF519:p.H431Y, ZNF687:p.F 858C, ZNF732:p.E227Q, ZNF880:p.Q406R, ZP3:p.V362fs, and ZRANB1:p.*735fs. .

[0122] 52. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0123] (a) This subject group has LAML; and

[0124] (b) The at least one tumor-specific mutation comprises any combination of mutations selected from the group consisting of: NPM1: p.W288fs, DNMT3A: p.R882H, NPM1: p.L287fs, IDH2: p.R140Q, IDH1: p.R132C, FLT3: p.D835Y, DNMT3A: p.R882C, FLT3: p.600_601insFREYEYD, IDH1: p.R132H, NRAS: p.G13D, U2AF1: p.S34F, KIT: p.D 816V, FLT3:p.D835E, IDH2:p.R172K, NRAS:p.G12D, WT1:p.S381fs, ABTB1:p.L249fs, DNMT3A:p.R736H, FLT3:p.D835H, KRAS: p.G12D, NPM1:p.L287fs, NRAS:p.Q61H, NRAS:p.Q61K, PHACTR1:p.V251fs, RBBP4:p.E330K, RUNX1:p.R135G, and U2AF1:p.S34Y.

[0125] 53. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0126] (a) This subject group suffers from LUAD; and

[0127] (b) The at least one tumor-specific mutation comprises any combination of mutations selected from the group consisting of: KRAS: p.G12C, KRAS: p.G12V, EGFR: p.L858R, U2AF1: p.S34F, KRAS: p.G12A, TP53: p.R158L, KRAS: p.G12D, PIK3CA: p.E545K, TP53: p.R273L, EGFR: p.ELREA746del, KRAS: p.G13D, A2ML1: p.S654fs, BRAF: p.G469V, CTNNB1: p.S37F, EGFR: p.G719A, KRAS: p.G 13C, MYOF: p.G165fs, EGFR: p.S768I, FAM47C: p.G948W, KRAS: p.Q61L, MYH10: p.L1091fs, NRAS: p.Q61L, OR4C3: p.H130fs, PI15: p.V22F, RAD50: p.D69Y , RIT1:p.M90I, TP53:p.C275F, TP53:p.R249M, TP53:p.R249G, TP53:p.R248P, TP53:p.R175H, TP53:p.Y163C, TP53:p.A159P, TP53:p.V157F, TP53:p.G 154V, ABCB1:p.R467L, ACBD3:p.R224L, ACTA1:p.G275C, ACTN2:p.D893Y, ADAM30:p.Q741H, ADAMTS14:p.G238C, ADAMTS20:p.R1251S, ADAMTS20:p.R54 1L, ADAMTS5:p.L549M, ADAMTS9:p.G659W, ADCY2:p.P1016T, ADCY5:p.G623C, AFP:p.A182G, AHDC1:p.P155Q, AKAP1:p.LDRNEEG317del, ALKBH1:p.K137 E. ANK2:p.Q3076L, ANKRD44:p.G339C, ANO3:p.A41S, AP1G1:p.R723L, APBB2:p.T243fs, APOB:p.L973M, APOBR:p.R840L, AQP10:p.Q261L, ARAP3:p.R12 26L, ARFIP2:p.R86L, ARHGAP36:p.P16H, ARL13B:p.R358L, ASCC2:p.R365L, ASPM:p.S240F, ASXL3:p.P1470Q, ATRN:p.P197Q, AVIL:p.G64W, AXDND1:p.W101R、B3GAT1:p.R125L、BARX2:p.R68P、BCL9L:p.G980C、BCOR:p.N1459S、 BEND2:p.P536Q、BMS1:p.G455V、BRAF:p.V600E、BRAF:p.G466V、BRD9:p.G3 30W、BRF1:p.V469L、BRWD3:p.H160N、BTRC:p.G260W、C11orf68:p.V135L、C 15orf2:p.V753F、C15orf2:p.G906W、C18orf8:p.M61I、C1GALT1:p.G299V、C 1orf173:p.G1454S、C1orf173:p.S688Y、C1orf87:p.R541L、C2orf53:p.P2 72H、C3orf20:p.R740L、C7:p.R687S、C7orf58:p.G140W、C7orf58:p.R238L 、CACNA1A:p.S772Y、CACNA1D:p.R1073L、CACNA1E:p.R2089Q、CACNA2D1:p. A352E、CACNG3:p.R232W、CADPS:p.R959S、CALB2:p.R258C、CAMK2B:p.G131V 、CARD11:p.I1065M、CCDC111:p.R417L、CCDC141:p.E1204V、CCDC19:p.R27 9L、CCDC19:p.R207L、CCKAR:p.L271M、CD1B:p.W41L、CDH10:p.S577R、CDH1 0:p.R472C、CDH10:p.R128S、CDH18:p.A721S、CDH20:p.P433H、CDH6:p.Q23 7K、CDK13:p.R880S、CDK4:p.R24L、CELF4:p.A309P、CFDP1:p.P129fs、CHN1: p.K264N、CHRNA4:p.S396R、CHRNA9:p.P361Q、CLCNKA:p.P124Q、CLEC12B:p .W217L、CLK4:p.R68L、CNTFR:p.D252Y、CNTN6:p.R807M、CNTNAP2:p.F395L 、COL19A1:p.P538Q、COL5A2:p.G612W、COL5A2:p.G516W、COL9A1:p.P211Q、 CPE:p.P290Q、CPNE8:p.Q127H、CPSF4:p.P219Q、CRIPAK:p.S180fs、CROT:p.Q580H、CRTC3:p.S363L、CSMD2:p.P1855Q、CSMD3:p.T2810N、CSMD3:p.P2727T、CSMD3:p.Q174H、CUBN:p.G596C、CUL4B:p.R91S、CUL7:p.L371F、CXCL9:p.K122N、CXCR4:p.E345Q、CXorf59:p.R198M、CYP11B1:p.R498G、CYP27A1:p.P112Q、CYP2B6:p.A444E、DACH2:p.R539L、DCC:p.R446H、DDX56:p.R329L、DEFA1:p.W90C、DENND2A:p.R688Q、DENND2A:p.R499L、DMBT1:p.R1521L、DNAH5:p.R3822L、DNAH9:p.S2993R、DNAI2:p.V231L、DPP6:p.L757F、DSG4:p.R128L、DST:p.A4410S、DZIP3:p.M322L、EBF3:p.R231S、EFCAB4B:p.E265Q、EHHADH:p.Q704H、ELAVL2:p.L263F、EMR1:p.R493H、ENAH:p.R514L、ENPP1:p.G738E、EPB41L3:p.A896S、EPG5:p.R2289L、EPHA1:p.G111V、EPHB6:p.R337H、EPRS:p.V1151L、ERBB2:p.S310Y、ERBB2:p.774_775insAYVM、ERBB2:p.776_776G>VC、ERN2:p.T295K、FAM120B:p.P467H、FAM127C:p.F52L、FAM135B:p.W240C、FAM210B:p.L112F、FAM47A:p.R690L、FAM47B:p.W163C、FAM47B:p.L567F、FAM5C:p.R457G、FAM70B:p.P277T、FAM71B:p.L583M、FAM75A6:p.R304S、FAM75A6:p.P54L、FAM75D1:p.R1265S、FARP1:p.R299L、FAT1:p.R4359L、FAT3:p.R1266H、FAT3:p.G1899V、FAT3:p.H3574N、FBXO18:p.M144I、FBXO31:p.G443fs、FCGBP:p.A1022S、FCRL2:p.V505L、FERD3L:p.P92H、FGB:p.E339Q、FGFR2:p.E116K、FGFRL1:p.R243L、FGFRL1:p.V274L、FKBPL:p.R320L、FLG2:p.G1545V、FLG2:p.L572F、FLG:p.P3254H、FLG:p.P2466Q、FMN2:p.P992T、FOLH1:p.A643S、FOXRED1:p.R136L、FRAS1:p.C382F、FRG2B:p.D142Y、FRMPD1:p.E1093Q、FSHB:p.T43N、GABRA5:p.Q224K、GADL1:p.L352I、GAL3ST3:p.A271S、GALNT14:p.D234E、GAS8:p.R313S、GATA3:p.M443I、GCDH:p.R82C、GEM:p.R268L、GFRAL:p.Q308K、GIT2:p.R123L、GJB4:p.R22S、GLB1L2:p.I407M、GLOD4:p.Q223fs、GNAO1:p.P283Q、GPNMB:p.I174M、GPR137B:p.G240C、GPR158:p.P762T、GPR98:p.G4307W、GRB7:p.R239L、GRHL1:p.G608W、GRID1:p.R683L、GRIK1:p.R368Q、GRM5:p.P895fs、GTF2E1:p.R192L、H3F3C:p.R131L、HAO2:p.H12N、HCN1:p.P231Q、HECW1:p.A183S、HGF:p.M686T、HIP1:p.R940L、HIST1H1E:p.R25P、HLA-DMA:p.A236fs、HOXA5:p.G11C、HS3ST3A1:p.G399W、HSD17B6:p.F209L、HSPA13:p.V85L、HSPBAP1:p.R282L、HTR5A:p.W298C、IGHMBP2:p.R615S、IL2:p.R103M、IL2RA:p.G61W、IL32:p.P215T、ING1:p.A220S、INMT:p.G56V、ITGA8:p.G616C、ITGAD:p.L528fs、ITGAX:p.R283H、ITIH1:p.G254W、ITIH2:p.L842V、ITK:p.R29L、ITPR2:p.P358Q、JMJD1C:p.R1198S、KCNA1:p.G376C、KCNH8:p.M455I、KCNJ3:p.L430F、KCNK18:p.G23V、KCNK2:p.R166L、KEAP1:p.G603W、KEAP1:p.R260L、KEAP1:p.S144F、KHDRBS2:p.S203L、KIAA1211:p.P1203Q、KIAA1549:p.L1272F、KIAA1755:p.Q108H、KIF15:p.E252Q、KIF9:p.G480R、KIRREL:p.G604C、KLF5:p.E419Q、KRAS:p.Q61H、KRTAP10-12:p.R64P、KRTAP27-1:p.M124I、KRTAP4-5:p.C91F、KRTAP5-1:p.S193Y、L1CAM:p.R632S、L3MBTL4:p.W162L、LAMA1:p.D1030Y、LAMB1:p.T1610fs、LAMB4:p.G1239W、LAMB4:p.G588W、LEF1:p.I53V、LEKR1:p.Q450K、LIM2:p.S150T、LIPJ:p.P236Q、LPHN3:p.E740D、LPPR4:p.R527S、LRFN5:p.N132K、LRP1B:p.G3563C、LRP2:p.M4039I、LRRC4C:p.Q10L、LRRIQ1:p.W792L、LRRTM4:p.S243Y、MAGEA10:p.R7H、MAGEC2:p.W109C、MAGI1:p.G1156V、MAGI2:p.P1044T、MAK:p.P373Q、MAP2K1:p.K57N、MARCH11:p.R193L、MEPE:p.G142C、MKI67:p.R1081S、MKRN3:p.P448H、MLL3:p.N393K、MLL3:p.Q356K、MMRN1:p.A1013S、MOGAT2:p.Q66fs、MXRA5:p.D324Y、MYH4:p.T790M、MYH8:p.R1117C、MYH8:p.H1006N、MYO5B:p.R708L、MYO7B:p.P2040H、MYO9B:p.R94L、MYT1L:p.P351Q、NAA11:p.T184K、NAB1:p.L72F、NAV1:p.R938L、NBPF15:p.G665E、NCAM2:p.G698C、NCAPD2:p.R220L、NDST3:p.V427I、NEK2:p.R239S、NFIA:p.L294F、NLRP3:p.R157C、NOTCH2:p.R2105L、NR4A2:p.R314L、NRG1:p.V481L、NRXN1:p.R813S、NRXN1:p.A660S、NRXN3:p.P23H、NRXN3:p.R103C、NTM:p.G333C、NUAK1:p.G173C、NYAP2:p.P437L、ODZ3:p.P218Q、OIT3:p.R508S、OOEP:p.R101C、OPN1LW:p.P283H、OR10H4:p.M199I、OR10J1:p.L157Q、OR10X1:p.L298I、OR10Z1:p.L205F、OR14A16:p.G160C、OR2A25:p.M80I、OR2AG2:p.G249W、OR2AK2:p.W37C、OR2H2:p.L205F、OR2J2:p.G234W、OR2L13:p.M106I、OR2L13:p.T242A、OR2L3:p.M1I、OR2L3:p.L67I、OR2L8:p.R121C、OR2L8:p.R171S、OR2M2:p.F177L、OR2M2:p.F323L、OR2M5:p.V205L、OR2T12:p.M258L、OR2T27:p.D11Y、OR2T33:p.P165Q、OR2T34:p.C246F、OR2T6:p.V213L、OR4C12:p.D309Y、OR4C12:p.M279I、OR4C16:p.L162M、OR4M2:p.A119S、OR4M2:p.A161S、OR51V1:p.P298T、OR5AS1:p.M39I、OR5B12:p.S289C、OR5B17:p.M266I、OR5D14:p.H246N、OR5D16:p.P264T、OR5D18:p.R123H、OR5F1:p.G44V、OR5J2:p.A36S、OR5L1:p.T275N、OR6C65:p.I154fs、OR6C75:p.G94W、OR6K2:p.P79Q、OR8D2:p.R306M、OR9A2:p.R289W、OR9G9:p.R169L、P2RX7:p.P142Q、P2RY10:p.T10K、P2RY10:p.V196L、PABPC5:p.R99S、PAPPA2:p.P917T、PAPPA2:p.P1706H、PBLD:p.P55Q、PCDH10:p.R587S、PCDH10:p.V986L、PCDH11X:p.R1010I、PCDHAC2:p.A742V、PCDHB5:p.P649S、PCDHGC5:p.K12N、PCDHGC5:p.P684H、PCLO:p.P3946T、PCMTD1:p.R271M、PDPR:p.G793W、PDYN:p.G191W、PDZD2:p.R565S、PDZD8:p.S980G、PFKM:p.R118S、PIGM:p.R225L、PIK3CA:p.E542K、PIK3CG:p.V165I、PILRA:p.S291fs、PLCE1:p.G564C、PLCL1:p.M564I、PLEKHA6:p.R110L、PNKP:p.G174W、POGZ:p.G75W、POLE:p.R573L、POM121L12:p.P231T、POM121L12:p.P242H、POTEE:p.V288M、POTEM:p.S78R、POU3F3:p.D321Y、PPT2:p.R265L、PRDM16:p.P1036L、PRELP:p.D201Y、PRPF40B:p.R160S、PRPF6:p.R763L、PTEN:p.R234L、PTPN11:p.G503V、PTPN13:p.E2067K、PTPRJ:p.G334W、PTPRT:p.R928L、PTPRU:p.P559S、PXDNL:p.P1456T、QSOX1:p.R401L、QSOX2:p.R683L、RAB13:p.R167L、RAB8A:p.G20W、RAPGEFL1:p.R356L、RBM19:p.G390W、RCL1:p.P112Q、REG1B:p.W57L、REG3A:p.S150L、REG4:p.G110V、RIMS2:p.R55L、RIT2:p.R85L、RLN2:p.S138C、RNF20:p.P529Q、RORB:p.G94W、RPL10L:p.K187T、RPRD2:p.R97S、RTN1:p.S103W、RUNX2:p.R337M、RYR2:p.K2413N、RYR2:p.M4334I、RYR3:p.P1670T、S100PBP:p.R5L、S1PR1:p.L104F、SAGE1:p.H298Q、SALL1:p.E965K、SALL1:p.R898W、SALL4:p.R187L、SBSPON:p.G133W、SCAF8:p.G740C、SCG2:p.P252Q、SCML4:p.L261F、SCN2A:p.T155K、SEC24D:p.A50fs、SEC61A2:p.G126V、SERPINA12:p.D253Y、SERPINA9:p.M414I、SERPINC1:p.R45L、SGIP1:p.R502L、SH3GL3:p.R174L、SH3PXD2A:p.S759L、SI:p.V1217F、SKOR1:p.Y883C、SLC1A2:p.F348fs、SLC24A5:p.R35S、SLC25A48:p.R101S、SLC35E2:p.R201L、SLC39A12:p.C628S、SLC39A6:p.R53L、SLC4A5:p.I533V、SLC5A1:p.G53W、SLC5A7:p.G442V、SLC6A11:p.W299L、SLC6A2:p.S354C、SLC8A1:p.G433C、SLIT1:p.R1460L、SLITRK5:p.R68L、SLITRK5:p.R468M、SLITRK6:p.N741K、SORL1:p.R205L、SOS1:p.N233Y、SOX9:p.E75K、SPAG16:p.V439L、SPIN4:p.Y171C、SPRR2D:p.P30fs、SPTA1:p.G2367C、SPTA1:p.D2243Y、SSX3:p.P127T、ST18:p.H778Q、STAC3:p.G117W、STOML3:p.D86Y、STX2:p.R107L、SUMF2:p.G110E、SUN3:p.P339Q、SV2C:p.P60Q、SYNDIG1:p.D135Y、SYNE1:p.K8632E、TARS2:p.E199K、TAS2R16:p.Q177H、TCOF1:p.K264R、TCTE1:p.S127I、TDO2:p.Q197H、THSD7A:p.G810W、THSD7A:p.R801L、TIFAB:p.D43E、TIGD4:p.S312F、TLL1:p.P53Q、TMPRSS11E:p.G259C、TMTC1:p.A864D、TMTC1:p.G212V、TMX3:p.R151C、TNNI1:p.R67L、TNR:p.L692I、TOP2A:p.R736L、TP53:p.R337L、TP53:p.E285K、TP53:p.R283P、TP53:p.D281N、TP53:p.C277F、TP53:p.V274F、TP53:p.R273H、TP53:p.I255F、TP53:p.R249S, TP53:p.M237I, TP53:p.S215I, TP53:p.C176F, TP53:p.R110L, TP53:p.G105C, TP53:p.P72fs, TPO:p.E558K, TRAF6:p. R502S, TRIM42:p.Q127K, TRIM48:p.A93D, TRIM4:p.R398L, TRIM51:p.W131C, TRIM9:p.R337S, TRIML1:p.H399Q, TRPM3:p.G29 8W, TSC1:p.G378C, TSG101:p.R276S, TSHZ1:p.K501N, TSHZ3:p.G677V, TTF2:p.R761S, TUBA3C:p.Q176fs, UBAC1:p.K330N, UB E2J2:p.G193W、UBR1:p.G1647W、UGT2B7:p.M214I、VMP1:p.E369Q、VPS13B:p.G2575W、VSTM2A:p.G75V、VWA3B:p.R557L、WBP11: p.P227fs、WDR52:p.G612C、WDR59:p.R837S、WDR75:p.P287Q、WDR88:p.G100W、ZCCHC5:p.G335W、ZFHX4:p.L811F、ZFHX4:p.T1 663N, ZFHX4:p.H2511Q, ZFP14:p.Q17L, ZIC1:p.A112E, ZNF154:p.T408N, ZNF223:p.G23W, ZNF295:p.S732C, ZNF322:p.K106N, ZNF385D:p.T226S、ZNF454:p.S190I、ZNF492:p.P392H、ZNF521:p.G640C、ZNF521:p.P270H、ZNF536:p.G186C、ZNF536:p.G663 W, ZNF644:p.G21W, ZNF716:p.H263L, ZNF71:p.V411L, ZNF782:p.G484W, ZNF831:p.Q617K, ZNF98:p.C492F, and ZSWIM2:p.S214Y. .

[0128] 54. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0129] (a) This subject group has LUSC; and

[0130] (b) The at least one tumor-specific mutation comprises any combination of mutations selected from the group consisting of: PIK3CA: p.E545K, TP53: p.R158L, KRTAP5-5: p.GCG47del, NFE2L2: p.E79Q, CDKN2A: p.D108Y, DHX9: p.V40G, MAFA: p.207_208HH>H, NFE2L2: p.R34Q, PBX2: p.Y262F, PIK3CA: p.E542K, TP53: p.R273L, TP53: p.C242F, TP53: p.R175G, TP53: p.Y163C, TP53: p.V 157F, AICDA:p.R131G, ALPK2:p.D53N, ANKFN1:p.M280I, ARPC1A:p.F212L, ASXL2:p.S1081L, C1orf74:p.D254N, C3orf30:p.D227E, CCDC121:p.W397L, CHN2:p.I43M, CLEC4C:p.R179L, CLN3:p.G206S, CNTN5:p.T178N, COL12A1:p.G2753C, CPS1:p.T855K, CSMD3:p.T1094K, CSMD3:p.Q691K, DDX11:p.R167 T, EGFR: p.L861Q, EME1: p.D570H, EP300: p.D1399N, ESYT3: p.S574F, FAM135B: p.L648M, FAM135B: p.Q285H, FAM47A: p.G372W, FBXW7: p.R505G, FGFR3:p .S249C, GALNT13:p.G358C, GNL3L:p.K20N, GPC5:p.R347L, HCN1:p.A714S, HCN1:p.R659L, HCN1:p.G499V, HCN1:p.P326T, HERC2P3:p.A803V, HEXDC:p. T482P, HIST1H3B:p.E74K, HIST2H2BE:p.G54D, IFNA10:p.V79A, IL7R:p.S54L, INADL:p.P1340A, ISX:p.C2F, ITGAX:p.R685H, ITPR1:p.E1883Q, KCNN3: p.80_81insQQ, KEAP1:p.G480W, KEAP1:p.R470C, KEAP1:p.V155F, KIAA1751:p.L63F, KIAA2022:p.C345F, KIR3DL2:p.K229E, KLF5:p.E419Q, LAMA4:p.M1293I, LMLN: p.G199C, LRP2: p.A516V, LRRC66: p.F458L, LSG1: p.R517L, LUM: p.R310L, MB21D2: p.Q311E, MCHR1: p.S306F, MKRN3: p.G 270V, MUC16:p.N11594K, NFE2L2:p.G81S, NFE2L2:p.G31A, NFE2L2:p.L30F, NFE2L2:p.D29H, OR2B11:p.G10V, OR2T2:p.F13V, OR4K2:p .C254F, OR51F2:p.R67P, OR51S1:p.R159Q, OR5D18:p.T271K, OR8H2:p.L166F, OR8J3:p.S160L, OR8K3:p.K235N, PCDHB1:p.N568K, PHI P:p.I1681M, PIK3CA:p.E726K, PIK3CA:p.H1047R, PLCE1:p.G439C, PRSS57:p.E39Q, PYHIN1:p.G148A, RANBP6:p.I984L, RBMXL1:p.G3 05C, REG1B:p.M67I, RGS6:p.W366L, RNF5:p.T136I, RP1:p.S1771L, RRP15:p.L214F, RYR2:p.E711K, SAMD3:p.Q206H, SLITRK3:p.R214 L, SON: p.S908L, SP4: p.E11del, STK11: p.G279fs, TARBP1: p.L782V, TBCD: p.R476C, TMPRSS11F: p.R274Q, TP53: p.R337L, TP53: p.E27 1K、TP53:p.R267P、TP53:p.G245V、TP53:p.Y234C、TP53:p.Y220C、TP53:p.H214R、TP53:p.H193L、TP53:p.H179L、TPTE:p.M541I、TRIM 7:p.L332I, TTN:p.T32425M, ZFP36L2:p.D240N, ZNF208:p.H883Q, ZNF48:p.R235H, ZNF626:p.K473R, ZNF676:p.P43T, ZZZ3:p.R162Q. .

[0131] 55. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0132] (a) This subject group has OV; and

[0133] (b) The at least one tumor-specific mutation includes any combination of mutations selected from the group consisting of: TP53:p.R273H, TP53:p.Y220C, TP53:p.R248Q, TP53:p.R175H, TP53:p.R273C, TP53:p.I195T, TP53:p.R248W, TP53:p.R282W, TP53:p.C176Y, TP53:p.V157F, TP53:p.S241F, TP53:p.H179R. TP53:p.G245S, TP53:p.H193R, ADCY2:p.V888I, B2M:p.M1V, BAP1:p.R227C, CYP4A11:p.V185F, DNAH5:p.R3197Q, GART :p.K807fs、GRIN2B:p.R519Q、HRNR:p.M1fs、KLHL29:p.L716fs、KRAS:p.G12V、MGA:p.R2435Q、MYO3A:p.N525S、NPAS2:p .Q201R、NRAS:p.Q61R、PDAP1:p.K55fs、PGAP1:p.F565C、TP53:p.S315fs、TP53:p.C275Y、TP53:p.R273L、TP53:p.V272 M, TP53:p.G266V, TP53:p.G266R, TP53:p.D259Y, TP53:p.P250L, TP53:p.G245D, TP53:p.G245V, TP53:p.G244C, TP53:p .C238fs、TP53:p.Y236C、TP53:p.Y234C、TP53:p.V216M、TP53:p.S215R、TP53:p.Y205C、TP53:p.L194R、TP53:p.P191d el, TP53:p.Y163C, TP53:p.A159V, TP53:p.K132N, TRPC7:p.D210V, UXS1:p.V100L, WNT11:p.C344Y, and ZNF295:p.E885A.

[0134] 56. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0135] (a) This subject group has READ; and

[0136] (b) The at least one tumor-specific mutation comprises any combination of mutations selected from the group consisting of: KRAS: p.G12V, TP53: p.R273H, KRAS: p.A146T, KRAS: p.G12D, TP53: p.R175H, AKAP9: p.L3482I, APBA1: p.E624K, BAG5: p.D439N, C17orf97: p.E230D, CDH23: p.F177L, CERS3: p.E95D, DNAH5: p.R982H, ERBB2: p.V842I, GABRB3: p.D500N, KRAS: p.G13D, KR AS: p.G12C, KRAS: p.G12S, LRP6: p.R675Q, MACF1: p.F722L, MBOAT2: p.R43Q, MYO1D: p.E246K, NLRC4: p.E409K, NRAP: p.E327K, NRAS: p.Q61K, PCDH15: p.R1552I, PIK3CA:p.N345K, PIK3CA:p.E545K, POLE:p.S459F, PPP2R2B:p.P326L, SMAD4:p.R361H, TP53:p.R248W, ZFP2:p.R150I, and ZNF563:p.K26N.

[0137] 57. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0138] (a) This subject group suffers from SKCM; and

[0139] (b) The at least one tumor-specific mutation comprises any combination of mutations selected from the group consisting of: BRAF: p.V600E, NRAS: p.Q61R, NRAS: p.Q61K, HSD17B7P2: p.N175S, BRAF: p.V600K, DISP1: p.G732L, IDH1: p.R132C, NRAS: p.Q61L, MUC16: p.P5119S, RAC1: p.P29S, WASH3P: p.G175S, AGAP9: p.M248V, C15orf23: p.S24F, DNAH5: p.D3236N , SPTLC3: p.R97K, TMC5: p.R276C, CFB: p.R314M, FRG1B: p.A50P, INMT: p.S212F, LOC649330: p.G93E, MAP2K1: p.P124S, RGS7: p.R44C, STK19:p .D89N, ADAM30:p.G97L, ARL16:p.G6R, ARMC4:p.E22K, BRAF:p.K601E, CAPN13:p.P405S, CD1C:p.R89C, CLCC1:p.P406Q, CNTN5:p.S379F, DNAH 5:p.R742Q, EEF1B2:p.S43G, FRG1B:p.I59V, GABRG1:p.E205K, IARS2:p.R832C, IL32:p.D218fs, ISX:p.R86C, KLHDC7A:p.E635K, NAP1L4:p. P285Q, NBPF10:p.Q908E, OR2A5:p.S71L, OR4E2:p.R226Q, OR4M1:p.G41E, OR4M2:p.S268F, OR4N2:p.G41E, OR51B2:p.S163L, PCDHGC5:p.R293 C. PCLO: p.R4133C, PHGDH: p.G173L, POTEG: p.D51N, PPP6C: p.R301C, PRAMEF11: p.C84S, PSG9: p.E404K, PTPRB: p.D1560N, RNF152: p.P95S, SP AG16:p.P488S, SPATA8:p.E18K, TAF1A:p.R172M, TCEB3C:p.E308K, THSD7B:p.E126K, TTN:p.E12129K, XIRP2:p.D2439N, and ZNF831:p.R1393Q.

[0140] 58. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0141] (a) This group of participants has UCEC; and

[0142] (b) The at least one tumor-specific mutation comprises any combination of mutations selected from the group consisting of: RPL22:p.K15fs, PTEN:p.R130G, PTEN:p.R130Q, KRAS:p.G12D, KRAS:p.G12V, PIK3CA:p.H1047R, PIK3CA:p.R88Q, PIK3CA:p.E545K, PTEN:p.V317fs, FGFR2:p.S252W, PIK3CA:p.E542K, CTNNB1:p.S37F, POLE:p.P286R, PPP2R1A:p.P179R, CTNNB1:p.S37C , KRAS:p.G13D, CTNNB1:p.D32N, CTNNB1:p.S33F, CTNNB1:p.G34R, KIAA2026:p.R574C, LIMCH1:p.R806fs, PIK3CA:p.H1047L, ALPK2:p.K523fs, CTNNB 1:p.S33C, FBXW7: p.R505C, HPD: p.R284fs, KRAS: p.G12A, PIK3CA: p.R93Q, POLE: p.V411L, TP53: p.R248W, ABCA11P: p.R385I, ABI1: p.K445N, ACSM2B: p.K195N、APOB:p.F3102L、ASCC3:p.R136Q、C12orf4:p.R335Q、CCDC132:p.R838C、CHD4:p.R975H、CSDE1:p.R220C、CTNNB1:p.D32Y、CTNNB1:p.S33Y、C TNNB1:p.T41I, EXOC1:p.R588C, FBXW7:p.R465H, FGFR2:p.N549K, FUBP1:p.R430C, GEN1:p.S509L, IK:p.E90fs, KIF20B:p.E54K, MAX:p.H28R, MBOAT2 :p.R43Q、METTL14:p.R298P、MFGE8:p.D170N、MS4A8B:p.S3L、NSMCE1:p.D244N、OXR1:p.E122K、PCDH19:p.E530K、PIK3CA:p.R108H、PIK3CA:p.N345K、 PIK3CA:p.C420R, PIK3CA:p.Q546P, PIK3CA:p.Q546R, PTEN:p.R130L, RBL2:p.E127K, RXFP1:p.S223Y, SF3B1:p.R957Q, SLC20A1:p.P328fs, SOX17:p.S403I、TNS1:p.Q659del、TP53:p.R273H、TP53:p.R273C、TP53:p.R248Q、TTN:p.D16823N、TXNL1:p.R234C、ZFHX3:p.R1893fs、ZNF180:p.R625I、ZNF257:p.R392I、ZNF354B:p.D609N、ZNF43:p.R280C、ZNF709:p.R468I、ZNF765:p.S254L、ABCA5:p.R1476Q、ACVR1:p.R206H、ADAD1:p.S11L、ADAM9:p.R256Q、ADD3:p.E570K、ADGB:p.S1124L、AGXT2:p.R502C、AMBN:p.S225Y、ANKDD1A:p.R24H、ARHGEF33:p.R46I、ATP10B:p.L1304I、ATP2C1:p.E724K、ATP9A:p.R290Q、ATR:p.R1814fs、AVL9:p.F34L、BMPER:p.R241Q、BTN3A2:p.E153K、C14orf118:p.R279I、C14orf166B:p.F230L、C3orf23:p.R217C、C3orf62:p.R185Q、CACNA1C:p.S710L、CAGE1:p.E539K、CARD10:p.KE272del、CCDC144A:p.S1264L、CCDC168:p.D5020Y、CCDC36:p.R209I、CD55:p.E156K、CEP44:p.S253L、CIITA:p.E728K、CREBBP:p.P2094L、CTNNB1:p.S37A、CTTNBP2:p.S420L、DCT:p.R532Q、DIAPH2:p.E121K、DLG2:p.S624L、DNAH10:p.R1888Q、DNAH14:p.R1367C、DNAH7:p.R2961Q、DNAH8:p.R1347H、DNAJC13:p.E1248K、DNMT1:p.E51K、DST:p.S1767Y、DYNC2H1:p.E883D、EMR1:p.R631Q、EPHX4:p.R282Q、ERCC6L2:p.L445I、F10:p.E117K、FAM155B:p.E158K、FAM83B:p.R206Q、FARP1:p.S383L、FAT3:p.A4159T、FBXW7:p.R689W、FBXW7:p.R465C、FBXW7:p.G423V、FN1:p.R290C、FZD6:p.R416Q、GABRA3:p.R73H、GABRA4:p.R460Q、GALNTL2:p.E395K、GFAP:p.A233T、GGA2:p.A63V、GIGYF2:p.R227H、GNPTAB:p.R1189Q、GPR112:p.S1283Y、GPR98:p.R4142W、GRIA3:p.S646Y、GRM6:p.E363D、HMCN1:p.S133Y、HSPA4L:p.R483C、HTR2A:p.S219L、INTS7:p.R940C、INTS7:p.R106I、ITM2C:p.E167K、JAKMIP2:p.R283I、KCND3:p.S438L、KCNS2:p.D211N、KDM1B:p.F361L、KIAA0556:p.L330I、KIAA1147:p.A149V、KIF23:p.R150Q、KIF27:p.K925N、KIF9:p.R594Q、KLHL13:p.E213K、KLHL28:p.E33K、LIN9:p.R183W、LRBA:p.E2103K、LRP2:p.R2432I、MAGI2:p.L450M、MC5R:p.A109T、MEGF10:p.S1053L、MKI67:p.T1664fs、MKLN1:p.F485L、MMRN1:p.F917L、MSH4:p.E730K、MTOR:p.S2215Y、MUC7:p.S336L、MYBPC2:p.R646H、N4BP2L2:p.R506C、NAPSA:p.R121Q、NCOA7:p.E369D、NCR1:p.R258W、NEK11:p.R374Q、NHEJ1:p.R109Q、NNMT:p.E233K、NOTCH4:p.15_16LL>L、NPY1R:p.A371T、NRAS:p.Q61R、OGDHL:p.R57C、OMA1:p.R445Q、OPRM1:p.R462C、OR4C12:p.F248L、OR5AK2:p.K89N、OSBPL6:p.R577Q、PCDHAC2:p.K138N、PCDHB12:p.R289C、PCDHGC5:p.A70T、PIK3CA:p.R38H、PIK3CA:p.E39K、PIK3CA:p.E110del、PIK3CA:p.K111E、PIK3CA:p.Q546K、PIK3CA:p.M1043V、PIK3CA:p.M1043I、PLA2G3:p.R201Q、PLXNA1:p.E1295K、PON1:p.R306Q、POTEE:p.R303I、POTEF:p.K674N、PPP2R1A:p.S256F、PPP2R3B:p.F310L、PRAM1:p.A268T、PREX1:p.E1246K、PRKCQ:p.A324V、PTEN:p.R130P、PVRL4:p.A358T、RAI2:p.S385Y、RBM39:p.T353I、RELN:p.F2722L、RFPL1:p.R148Q、ROBO2:p.D1018N、ROS1:p.R245I、RPS6KA6:p.S394Y、RSBN1:p.E572K、RYR1:p.A2576T、SACS:p.R2906Q、SCAPER:p.R366Q、SELP:p.R429W、SENP7:p.S673Y、SEPHS1:p.E13K、SFRP4:p.R232Q、SGK1:p.K367del、SIX1:p.E191K、SLC10A7:p.S261L、SLC12A2:p.R828Q、SLC16A14:p.R495Q、SLC7A2:p.R322W、SMCR8:p.E175K、SOS1:p.N233Y、SPOP:p.E50K、STRN3:p.K218N、STXBP6:p.D92N、SULT1E1:p.R77Q、SUN3:p.L124I、SUSD1:p.R343C、SYNM:p.R516Q、TAF1:p.R843W、TDRD3:p.R322Q、THADA:p.S1941L、TLN2:p.S208L、TMEM161B:p.R315Q、TMPRSS3:p.R16Q、TP53:p.Y220C、TPTE:p.S423L、TRANK1:p.E846K、TRPC5:p.S490L、TRPM3:p.R429W、TSSK1B:p.E301K、TTLL7:p.R751H、TTN:p.S20317L、TTN:p.E6404K、TTN:p.R4434Q、TTN:p.R2506Q、UGT8:p.E102K、USF1:p.R52Q、USP16:p.R455Q、USP25:p.R873H、USP33:p.R36Q、VPRBP:p.R802Q、VPS13B:p.R692Q、WDR65:p.F110C、YTHDC2:p.E185K、ZFYVE1:p.R266Q、ZKSCAN1:p.R541fs、ZNF117:p.R157I、ZNF180:p.R569I、ZNF195:p.R59Q、ZNF254:p.K179N、ZNF263:p.R510I、ZNF333:p.R554Q、ZNF354B :p.R402I、ZNF442:p.R309Q、ZNF454:p.R376I、ZNF485:p.R374I、ZNF488:p.R206Q、ZNF559:p.E284K、ZNF594:p.R287I、ZNF6 11:p.R390I、ZNF645:p.R154C、ZNF649:p.R338Q、ZNF649:p.R198I、ZNF674:p.R405I、ZNF675:p.R220I、ZNF678:p.R564I、ZN F732:p.R354I, ZNF780A:p.R466Q, ZNF823:p.R547I, ZNF836:p.R854I, ZNF836:p.R630I, ZNF841:p.R757I, and ZNF98:p.R370I. .

[0143] 59. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0144] (a) This subject group has ACC; and

[0145] (b) The at least one tumor-specific mutation includes any combination of mutations selected from the group consisting of: ZFPM1: p.EPL444del, GARS: p.P42A, ZNF517: p.V349A, LRIG1: ​​p.L24V, CCDC102A: p.R96W, OPRD1: p.C27F, SOWAHA: p.R124P, LACTB: p.M5L, TOR3A: p.F13L, ZFPM1: p.E444fs, ZNF787: p.D367del, LRIG1: ​​p.L26V, IRX3: p.L422P, TRIOBP: p.H1300R, TUBA1C :p.L146F、ZFPM1:p.P445fs、ZFPM1:p.446_447LA>P、TPO:p.S398T、USP42:p.R779P、ERCC2:p.D312N、GLTPD2:p.D209E、OTOP1:p.LLW104del、RINL:p P402L, AMDHD1:p.S3G, ASPDH:p.Q266R, KCNK17:p.S21G, TMEM247:p.Q128E, MUC5B:p.D682G, OBSCN:p.R4516W, FAM184B:p.R784W, SEMA5B:p.V840D, Z NF598:p.E25G, ADAD2:p.G44E, C1orf106:p.R538C, ZAR1:p.Q42H, PANK2:p.G126A, PODXL:p.28_30PSP>P, SALL3:p.L593V, THEM4:p.L17R, C2orf81:p .T315P、CLDN23:p.V210M、FAM109A:p.GGG156del、FPGS:p.I22V、HHIPL1:p.V692A、MUC5B:p.M2869T、PLEC:p.R1386Q、SYT8:p.R373W、TAF5:p.S130A、 TMEM189-UBE2V1:p.N6D, UQCRFS1:p.S6A, B3GNT6:p.L316fs, CCDC105:p.P499T, CLIC6:p.Q298E, IDUA:p.T374P, NOTCH2:p.C19W, RGS9BP:p.A96S, RR EB1:p.G783V, SP8:p.G165del, WDR34:p.W60G, C19orf10:p.G12R, CELSR2:p.16_17insP, FAM75C1:p.71_71H>HLVSQRH, GPRIN2:p.R446H, KBTBD13:p.A81V、OGFR:p.S557T、PODXL:p.30_30P>PSP、BHLHE22:p.L62Q、C4orf32:p.G32E、C5orf65:p.Q245R、KNDC1:p.V806D、KRTAP10-6:p.49_49P>PSCCAP、LRP11:p.P92R、MAP1S:p.S411C、NOL9:p.S58A、RASIP1:p.R601C、RGMB:p.S63R、SARM1:p.R23P、TSC22D2:p.A419T、ZNF628:p.T230A、ZNF814:p.A337V、AATK:p.A541T、BTBD11:p.G265A、CRIPAK:p.C143R、KCTD3:p.F9V、KRT8:p.S59A、MUC5B:p.S681G、NCOR2:p.1846_1847insSSG、OGFR:p.E556K、APOE:p.C130R、C10orf95:p.A85S、C13orf33:p.R59G、CRIPAK:p.C174R、FAM18B2:p.C51Y、GLI3:p.P998L、GLTSCR2:p.Q389R、HECTD2:p.P19A、IRF2BPL:p.123_125QQQ>Q、MEX3C:p.179_182AAAA>A、NEFH:p.EE658del、RNF149:p.S9G、RNF222:p.A133T、SEZ6L2:p.R74P、TNIP2:p.R73G、ARRDC4:p.T79A、B3GNT6:p.P330fs、BAG1:p.G45R、C22orf26:p.P28L、CHDH:p.E40A、COQ2:p.V66L、CTGF:p.H83D、DLEU7:p.A83V、EPPK1:p.D2378H、FAM86C1:p.R30P、FZD1:p.93_94insP、GPRIN2:p.V241M、GPX1:p.11_13AAA>A、HES3:p.P96T、JMJD4:p.A11V、KANK3:p.R359H、LPPR2:p.A186S、NEFH:p.665_666insEE、NOM1:p.R24G、RNF39:p.G263C、SCRT1:p.S133A、SNED1:p.L1228P、TTLL11:p.122_123insKA、ZCCHC3:p.A159del、ZNF219:p.QP233del、ASB16:p.T249A、ASB2:p.H515P, ATP9B:p.S39G, AVL9:p.G7fs, C17orf96:p.L63V, C19orf29:p.A499V, CRB2:p.T1110M, CRIPAK:p.P173R, CRIPAK:p.I190L, CSGALNACT 2:p.L362F, CTBS:p.LAL31del, CTNNB1:p.S45P, DMRT1:p.S45T, DOK7:p.G461D, FBRSL1:p.A836V, FEZ2:p.P50L, FRG1:p.S169N, HSD17B1:p.G3 13S, IBA57: p.S130R, KIF1A: p.E917D, KRTAP9-1: p.160_160Q>QPSCGSSCCQ, LURAP1L: p.55_56insGGG, NMU: p.A19E, NMU: p.A18E, NOXA1: p.D6E , NPTX1:p.G100D, PLIN5:p.R306W, TBP:p.95_96insQ, TMEM200C:p.S498G, TNXB:p.V706fs, VARS:p.P51S, ZC3H12D:p.P405S, and ZZEF1:p.V30A. .

[0146] 60. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0147] (a) This subject population has CESC; and

[0148] (b) The at least one tumor-specific mutation comprises any combination of mutations selected from the group consisting of: PIK3CA:p.E545K, PIK3CA:p.E542K, MAPK1:p.E322K, EP300:p.D1399N, ERBB2:p.S310F, ERBB3:p.V104M, KRAS:p.G12D, ANKRD12:p.E721Q, ANKRD36:p.M1144T, MICA:p.G318fs, PIK3CA:p.E726K, PTEN:p.R130Q, ABCD1:p.S606P, ACTL7B:p.E211K, ADAM2 1:p.F129C, ADAMTS12:p.P1053A, AKT1:p.E17K, ANKLE1:p.V643L, ANO3:p.M956I, AAOAH:p.R326T, APOD:p.S115L, ASCC1:p.H207Y, ATM:p.S800F, AURK A:p.S387L、BAG5:p.M286I、C12orf43:p.E28Q、C16orf3:p.G65S、C3orf70:p.S6L、C4orf21:p.E800Q、CALB2:p.K60N、CALCB:p.R81T、CCDC152:p.E153Q , CCDC53:p.R58C, CDC27:p.P242S, CFHR5:p.R441H, CLOCK:p.L123fs, CMYA5:p.E2733K, CNTRL:p.P185S, CSHL1:p.R117Q, CSMD3:p.H952Y, CTNNB1:p. D32G, CTSH: p.E254Q, DHPS: p.F49L, DMPK: p.R44H, DNAH14: p.F622fs, DNAH3: p.E3367Q, DNAH8: p.E587D, DNASE1L1: p.D212N, ECE2: p.D254N, FAM71B: p.H445D、FAM73A:p.G23V、FAS:p.E261K、FBXW7:p.R505G、FBXW7:p.R465C、FEZF2:p.E82K、FKBPL:p.E161Q、FMNL1:p.E927Q、GPATCH3:p.E275Q、GPR14 2:p.R304T, GPRIN2: p.T100P, GRAMD2: p.I123M, HERC2: p.S329F, HGF: p.G229A, HIF3A: p.A72T, HIST1H1B: p.K188N, HIST1H2AL: p.R30P, HIST2H2AC: p.R30P、HLA-C:p.N104K、HLA-DPB1:p.G114fs、HRNR:p.G2539S、INVS:p.R799 K、JPH3:p.Q433H、JUP:p.S627L、KIAA1211:p.R308fs、KIAA1211:p.E309fs 、KLK2:p.E161K、KRAS:p.G13D、KRAS:p.G12V、LIN9:p.E231K、LOC151174:p .P90S、LRRC37A3:p.A406D、LRTM2:p.L176V、MEPE:p.S30T、MUC12:p.R2634 C, MUC4:p.S2936L, MYOM2:p.D988N, NFE2L2:p.D29H, NOTCH2:p.R2298W, NP IPL1: p.P250L, NR5A2: p. E80K, NYAP2: p. R197Q, OBSL1: p. E1642K, OR13C2: p.L9V、OSBP:p.Q721H、PAOX:p.H107Y、PDILT:p.E500K、PIAS3:p.D460N、PL EKHO2:p.E351Q, PNRC1:p.R73C, PPP4R1:p.L597F, PREP:p.F469L, PRKDC:p .Q3568E、PSME3:p.R231W、RANBP6:p.R915W、RCAN2:p.D440N、RNPC3:p.E11 6fs, SDHAP1:p.H66Y, SDHAP2:p.S37fs, SERPIN3:p.K158N, SERPIN4:p.R 98C、SF1:p.R255W、SGSM1:p.E818K、SIM1:p.V213M、SLC10A4:p.F281L、SLC 25A5:p.I79F, SLC35G2:p.K62fs, SLC4A9:p.R617C, SLCO2A1:p.M479I, SND 1:p.Q38E、SPATA17:p.R72K、SRSF12:p.S150C、TADA2B:p.E67K、TCTEX1D2: p.S74L, TEDDM1:p.M166I, TEX15:p.E1652Q, TMC2:p.E92D, TMEM131:p.E13 19Q, TNKS2:p.T619fs, TNS1:p.Q659del, TP53:p.E285K, TRAF3:p.S9F, TRI M61:p.K98N、TRPM1:p.M996I、TUFT1:p.L101F、U2AF1:p.S34F、UNC93B1:p.V498M, USP4:p.L259V, VCAN:p.S1308C, WDR17:p.P278S, ZBED4:p.S385L, ZEB2:p. E1094K, ZFYVE9:p.M1147I, ZNF16:p.R452W, ZNF677:p.R131T, and ZSWIM4:p.E407K. .

[0149] 61. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0150] (a) This group of participants has CRC; and

[0151] (b) The at least one tumor-specific mutation includes any combination of mutations selected from the group consisting of: KRAS: p.G12D, KRAS: p.G12V, BRAF: p.V600E, KRAS: p.G13D, TP53: p.R175H, PIK3CA: p.E545K, FBXW7: p.R465H, KRAS: p.A146T, PIK3CA: p.H1047R, TP53: p.R248W, CDC27: p.D555E, SMAD4: p.R361H, TP53: p.R273H, KRAS: p.G12C, NRAS: p.Q61K, ERBB2: p.V8 42I, ERBB3:p.V104M, FBXW7:p.R465C, PIK3CA:p.R88Q, PIK3CA:p.E542K, TP53:p.R273C, TP53:p.G245S, AXIN2:p.G665fs, C16orf45:p.T106N, C20or f26:p.R1088Q、DNMT1:p.E432K、FBXW7:p.R505C、HLCS:p.E362K、HPSE2:p.K58N、KIF14:p.R598Q、KIF18A:p.R17C、KIF20B:p.E991K、KLHL5:p.R326C、K LK2:p.P57T、KRAS:p.G12A、KRAS:p.G12S、LPHN3:p.R1183Q、LRP6:p.R675Q、MYH8:p.R1048Q、NRAP:p.E327K、NRAS:p.G12C、PIK3CA:p.N345K、POSTN:p .R508C, PPP2R1A:p.R183W, PTEN:p.R130Q, RAF1:p.S257L, SDK1:p.T1181M, SGSM1:p.F1117L, TCF7L2:p.R482fs, TP53:p.R282W, TRIM23:p.R289Q, UG T8:p.E102K、ZNF491:p.R343Q、A2M:p.R732Q、AADACL4:p.A266T、ABCA8:p.E1158K、ABCA8:p.R842Q、ABCA8:p.A696T、ABCB8:p.R345H、ACACA:p.R1731 C. ACADM:p.F48C, ACOT9:p.R50Q, ACPP:p.R105Q, ACTL7B:p.R354H, ACTL9:p.R331H, ACVR1:p.S290L, ADAM30:p.S314Y, ADAM32:p.R559Q, ADAMTS16:p.D817N、ADAMTS4:p.R156W、ADCY5:p.R661H、AGMAT:p.V313M、AGPAT4:p.A212T、AKAP12:p.E1282K、AKAP9:p.L3482I、ALB:p.S294L、ALDH1L1:p.A870T、ALG2:p.S302Y、AMOTL1:p.R676Q、AMPD1:p.K502N、AMPH:p.R292W、ANKRD6:p.R479C、APBA1:p.K730N、APBA1:p.E624K、APC:p.E847fs、APC:p.F1354fs、APC:p.M1413fs、APOB:p.R3136C、APOB:p.A43V、APPL1:p.R668W、AQPEP:p.A309T、ARF4:p.R149H、ARFGEF1:p.D1632N、ARHGAP32:p.E1253K、ARHGAP36:p.R128C、ARHGAP36:p.A147V、ARHGAP5:p.D890fs、ARNTL:p.T395M、ARPP21:p.R338H、ARSG:p.V131I、ASCC3:p.R1197Q、ATP10D:p.R311H、ATP6V0A4:p.R191Q、ATP9B:p.R265Q、AXDND1:p.E930D、AXIN2:p.W663fs、B2M:p.L13fs、B3GALNT1:p.R145Q、BACH1:p.R538Q、BAG5:p.D439N、BBOX1:p.F176V、BCL2L11:p.R91Q、BCL7A:p.T52M、BCLAF1:p.R37fs、BEND5:p.R198C、BICD2:p.R162H、BLVRA:p.S44L、BMP3:p.R344W、BNC2:p.R512W、BRPF1:p.R66C、BRWD3:p.R787C、BTBD7:p.S436L、BUB1B:p.F996L、BZRAP1:p.V1627I、C11orf30:p.R1111C、C14orf101:p.E295K、C14orf102:p.D115N、C14orf105:p.R100I、C15orf2:p.V488I、C15orf33:p.D340N、C16orf87:p.R151I、C1RL:p.L351fs、C22orf40:p.P32fs、C3orf39:p.R333W、C5orf30:p.D4N、C5orf4:p.R114Q、C6orf170:p.K724T、C7orf63:p.A10T、CACHD1:p.S720Y、CACNA1A:p.T665M、CACNA2D3:p.A332T、CACNB2:p.R608H、CACNG3:p.V134I、CACNG3:p.A138V、CACNG5:p.G121R、CADM1:p.S190L、CADPS:p.A1073T、CAPRIN2:p.E13K、CARD11:p.R423Q、CASC1:p.R54Q、CASP14:p.R5W、CBFB:p.E152K、CC2D2A:p.R1284C、CCDC18:p.K615N、CCDC60:p.R230H、CCDC81:p.R259I、CCDC88C:p.P1851fs、CCKBR:p.V236M、CD101:p.D283Y、CD101:p.R594Q、CD180:p.N228T、CDC14B:p.R375C、CDCA7L:p.P405fs、CDH10:p.E349K、CDH12:p.D674N、CDH20:p.A134V、CDH23:p.F177L、CDH2:p.D547Y、CDH9:p.F523L、CDK16:p.R108C、CEACAM5:p.L640I、CEP152:p.E21K、CERS3:p.E95D、CHD4:p.R975H、CHD5:p.A801T、CIZ1:p.V668A、CLEC18A:p.R423H、CLTCL1:p.R481W、CMAS:p.R110Q、CNRIP1:p.R102W、COBLL1:p.K732N、COL14A1:p.R1082I、COL17A1:p.P1004L、COL4A6:p.L550I、COL6A3:p.D2792N、COPB1:p.R425C、CORO2A:p.*526R、COX15:p.L86I、CSMD1:p.S781Y、CTCFL:p.E423K、CTDNEP1:p.E126K、CTTNBP2:p.R164C、CYP4B1:p.E434D、DACH2:p.R539C、DBC1:p.V216I、DBF4B:p.S254Y、DCHS2:p.F2149L、DCLK2:p.S549Y、DDI1:p.R275Q、DENND4A:p.P357H、DENND4C:p.R1081Q、DHTKD1:p.R410Q、DISP1:p.R763C、DKK2:p.R230H、DKK4:p.R203Q、DLC1:p.A350V、DLC1:p.E222D、DMD:p.R3195H、DNAH5:p.R982H、DNAH5:p.R224Q、DNAH9:p.D1547N、DNAJC24:p.E61K、DNM1:p.A251T、DNMT1:p.E1531Q、DNMT3B:p.R92W、DOCK10:p.A1830V、DOCK1:p.E864K、DOCK2:p.G170R、DOCK3:p.R1183C、DOCK5:p.E177K、DOK5:p.R274W、DPP8:p.G165R、DPY19L1:p.F378L、DUOX2:p.F880L、DVL2:p.A601fs、EBAG9:p.E187K、EBF3:p.G255fs、EDNRB:p.L450R、EGR2:p.R390H、EHD3:p.E44K、EIF2C1:p.R139Q、ELF3:p.F305fs、ELMOD2:p.T141M、EMR2:p.S75L、ENAM:p.R373H、ENOX2:p.R356W、ENTPD7:p.E327K、EPG5:p.D369N、EPHB2:p.R392H、ERCC6:p.V780I、ERCC6L:p.R505Q、ERRFI1:p.A421T、ESCO1:p.R300Q、ETV6:p.R369W、F8:p.S2269Y、FAM123B:p.F173fs、FAM135B:p.R884H、FAM169B:p.K165N、FAM170A:p.E56K、FAM171B:p.D459N、FAM181A:p.R109H、FAM5B:p.R402C、FBXO11:p.A432V、FBXW7:p.R689W、FBXW7:p.S582L、FBXW7:p.R14Q、FGF14:p.A236V、FHDC1:p.R254W、FHOD3:p.A225T、FHOD3:p.E813K、FMO3:p.F510L、FNDC1:p.R652H、FOXK1:p.R354W、FOXN3:p.P96fs、FPGT-TNNI3K:p.R455H、FZD3:p.D367N、GABRA4:p.R460Q、GABRA5:p.S126N、GABRB3:p.D500N、GALNTL5:p.R262I、GJA1:p.R362Q、GLRA3:p.L454I、GLRA3:p.F132L、GOLGA4:p.Q1536H、GP2:p.S41L、GPC6:p.A214T、GPLD1:p.R717Q、GPR125:p.R113Q、GPR156:p.F754L、GPR158:p.D566N、GPR21:p.R216H、GPR61:p.A62T、GPR98:p.R4142W、GPRC5A:p.V30I、GRAP2:p.E69D、GRIA1:p.R218C、GRIA2:p.R845Q、GRM7:p.R679Q、GTF3A:p.K306N、HAO1:p.R172C、HARS2:p.R168H、HBB:p.F42L、HCN4:p.R525H、HDAC5:p.A1044T、HGF:p.S467Y、HIPK4:p.R280H、HLA-DMA:p.E84K、HMG20A:p.E248D、HPS3:p.S468L、HRSP12:p.R120Q、HS3ST1:p.E287K、HTR3B:p.R236C、HTR5A:p.R152C、HTT:p.D1548N、HYDIN:p.R1187C、HYDIN:p.R939Q、HYDIN:p.R451Q、HYOU1:p.R158C、IFT172:p.A944V、IGJ:p.R77Q、IL17RA:p.Q803fs、IL1RAPL2:p.T647M、IL3:p.A90T、IL5RA:p.L47I、INPP5D:p.R523Q、INPP5K:p.R263C、IRAK3:p.R267Q、IREB2:p.R419Q、ITGA4:p.T673M、ITGA4:p.F900L、ITIH5:p.A912T、ITK:p.E196K、JAG1:p.A462T、JAK1:p.V310I、KAL1:p.V303I、KBTBD8:p.V549I、KCNA3:p.A415V、KCND3:p.S438L、KCNMB4:p.F209L、KCTD20:p.L314fs、KDELC1:p.L447I、KIAA0528:p.R181Q、KIAA0556:p.R1082W、KIAA1109:p.S4937Y、KIAA1804:p.V474M、KIAA1804:p.R477W、KIF16B:p.R145Q、KIF26B:p.A1114V、KPNA4:p.R29Q、KRAS:p.K117N、KRAS:p.Q61L、KRAS:p.Q61K、KRT6B:p.L197P、L1CAM:p.T186M、LALBA:p.A41T、LAMA4:p.A558V、LBX1:p.R176W、LPAR4:p.R145Q、LR P1B: p.K2623N, LRP2: p. R3043C, LRP2: p. S737L, LRRC18: p. R218W, LRRC31: p.K23T, LRRC7:p.R1389H, LZTS2:p.P100fs, MACF1:p.S292L, MACF1:p.F72 2L, MAEL:p.R345C, MAGEE1:p.V380M, MAGI1:p.R1198C, MAP1B:p.E2046D,M AP2:p.K530N、MAP2K4:p.R287H、MAP3K4:p.R275Q、MAP7D2:p.R487C、MAPK8 IP1:p.L217fs、MBOAT2:p.R43Q、MCF2L2:p.R926Q、MECOM:p.R969C、METTL1 6:p.R200Q、METTL21A:p.R174Q、METTL6:p.F56L、MFF:p.R162C、MFSD5:p.R 280Q、MIA3:p.Q356H、MMAA:p.R326C、MORC1:p.D113Y、MORC2:p.R740H、MPDZ :p.L804I, MR1:p.S46L, MRPL47:p.L234I, MS4A8B:p.S3L,MSH4:p.K464N,M SH6:p.T1085fs、MSH6:p.R1095H、MUC16:p.R8606H、MYH13:p.D311N、MYH7: p.R1689C, MYO1D:p.E246K, MYO3A:p.N525H, MYO6:p.D1180N, MYO9A:p.R21 79Q, MYO9A:p.R167Q, MYOZ2:p.E251K, MYT1:p.E226K, NAA25:p.S807Y, NCA M1:p.R474W、NCOA4:p.R562Q、NEB:p.D5434N、NEB:p.L1591I、NEB:p.E1214 K、NEDD9:p.A798T、NEDD9:p.A316T、NEK1:p.R608C、NFASC:p.V256I、NINL: p.R1366C、NLRC4:p.D593N、NLRC4:p.E409K、NLRP4:p.V229I、NLRP5:p.R39 2H、NME9:p.E75K、NOLC1:p.T428M、NPC1:p.E451K、NPSR1:p.R235Q、NRAS:p.Q61L, NRAS:p.G13R, NRAS:p.G12D, NRG2:p.T246M, NTN4:p.E59K, NUB1:p.R 373Q, NUDT15:p.S83Y, NUF2:p.S340L, NUP88:p.A302V, ODZ1:p.R2556W, OGD HL:p.A427T、OGFRL1:p.E427K、OLFM4:p.K132N、OPRM1:p.R353H、OR10A3:p .S93Y、OR2M3:p.R235H、OR52W1:p.R133C、OR5AU1:p.R312H、OR5B17:p.R163 H、OR8S1:p.A99V、OSTN:p.R115Q、OTOL1:p.V431I、OTUD3:p.R277I、PAN3:p .S580N PANK3:p.R260I 、PAX3:p.T424M 、PCBP1:p.L102Q 、PCDH10:p.V477M . PCDH15:p.R1552I, PCDHAC2:p.A519T, PCDHAC2:p.E190K, PCDHAC2:p.A266 T, PCDHAC2:p.A156V, PCDHAC2:p.E271K, PCDHAC2:p.A736V, PCDHB5:p.D51Y PCDHB8:p.D235N, PCDHGC5:p.S289L, PCDHGC5:p.V662M, PCNXL2:p.R135Q PCOLCE2:p.A348V, PCOLCE2:p.R87H, PDE4B:p.S417L, PGAM1:p.R240H, PHF 3:p.R1410I, PIAS2:p.S519L, PIGR:p.A580T, PICK3CA:p.D350G, PICK3CA:p. E545A, PIK3CA:p.E545G, PIK3CA:p.Q546K, PIP4K2C:p.R204H, PKHD1L1:p.F 1856L, PLA2G4A:p.E443K, PLCG2:p.E544K, PLCG2:p.D973N, PLEKHA6:p.V3 28fs, PLEKHG4B:p.E384K, PLK1:p.D233G, PLOD3:p.R297fs, PLSCR3:p.E77K PLXNC1:p.S462L, PLXNC1:p.R819C, POLA1:p.E603D, POLE:p.S459F, POLE: p.V411L, POLQ:p.R860Q, PPP2R2B:p.P326L, PPP2R5C:p.S259Y, PRAMEF4:p.R248H、PREX1:p.V731I、PRKAA2:p.R407Q、PRKAR2B:p.S309L、PRKCI:p.R480C、PRKRA:p.K122N、PSG8:p.R397C、PSG8:p.R320C、PSMD12:p.R201Q、PTPDC1:p.R430W、PTPN12:p.R765Q、PTPN13:p.S887L、PTPRD:p.L1053I、PTPRU:p.D1434N、PXDN:p.P856fs、PXDNL:p.T1312M、QRSL1:p.S226L、RAB7L1:p.R79W、RALGAPA1:p.R398C、RANBP2:p.R1231C、RBBP7:p.E313K、RBBP7:p.E274K、RBFOX2:p.A340T、RBMXL1:p.R331Q、RHOBTB1:p.T464M、RIMS2:p.R599Q、RIN3:p.S708L、RLBP1:p.D281N、RLBP1:p.A72V、RNASET2:p.A127V、RNF113B:p.A172V、RNF150:p.R236Q、RNF150:p.S208L、RNF43:p.S216L、ROR2:p.D672N、RPL6:p.F193C、RPS6KA5:p.E166K、RSPO2:p.R28C、RUVBL1:p.E431K、RUVBL1:p.R117C、RWDD2B:p.R254H、RXFP3:p.R113C、RYR3:p.R2705Q、SAGE1:p.R229C、SCFD2:p.R545W、SCML4:p.R194Q、SCN10A:p.T1570M、SCN11A:p.A1688T、SCN11A:p.V1289I、SCN11A:p.V566I、SCUBE2:p.V342M、SEMA3A:p.D81N、SEMA4D:p.R252Q、SEPHS1:p.R371Q、SEZ6L:p.S207L、SFPQ:p.R611Q、SFSWAP:p.S617Y、SGCG:p.A220V、SGCZ:p.I41M、SH3TC2:p.R89C、SIGLEC11:p.S363F、SIPA1L1:p.R1063Q、SIPA1L1:p.S1227Y、SLC12A1:p.S292L、SLC22A15:p.S201L、SLC24A2:p.A134V、SLC25A40:p.R96Q、SLC2A7:p.A65T、SLC30A9:p.R194H、SLC33A1:p.S542L、SLC35F3:p.A280T、SLC39A7:p.R382C、SLC43A1:p.P133L、SLC43A3:p.R216H、SLC44A5:p.R185H、SLC6A2:p.A562T、SLC8A1:p.R431H、SLFN12L:p.F232fs、SLITRK1:p.R52H、SLITRK3:p.S298L、SMAD2:p.R321Q、SMARCA4:p.R381Q、SOCS5:p.S464L、SORBS1:p.V1156M、SORBS1:p.F570L、SORCS2:p.R320W、SOX6:p.R719W、SPATA22:p.S150L、SPEG:p.A944V、SPTB:p.R86C、SPTBN4:p.A1993V、STIM2:p.R572Q、STT3B:p.D583Y、SULT1C4:p.R85Q、SUN3:p.E128K、SUPT6H:p.A957T、SYNE1:p.I1249L、SYNE1:p.R170W、SYNE2:p.K3103N、SYNGR4:p.R169Q、SYT7:p.T349M、TANK:p.S380L、TAS1R2:p.R270C、TAS2R1:p.F183L、TCF7L2:p.R488C、TDRD10:p.S322L、TECTB:p.L29I、TEKT5:p.R401H、TGFBR1:p.S241L、THAP5:p.S287Y、THSD7B:p.R90H、TLL1:p.T153M、TLL2:p.S872L、TM9SF2:p.R91H、TMCC3:p.R110H、TMEM132A:p.R481C、TMEM132D:p.R578W、TMEM55A:p.R189Q、TMEM74:p.R125Q、TMPRSS11A:p.S288L、TNIP2:p.A139T、TOP2B:p.R656H、TOX:p.S354L、TP53:p.G244D、TP53:p.R175C、TPO:p.A826T、TPR:p.S2155L、TPTE2:p.R258Q、TPTE:p.S423L、TRAK1:p.D627N、TRAPPC11:p.R568Q、TRIM23:p.R396Q、TRIM44:p.D331N、TRIO:p.R661W、TRPA1:p.K54N、TRPC5:p.S490L、TRPM6:p.R995H、TRPM7:p.R1862C、TRPM7:p.R843Q、TRPS1:p.R1125W、TRPV5:p.R492H、TRRAP:p.R3515W、TSHZ1:p.R881M、TTC21A:p.S270Y、TTN:p.R22795C、TTN:p.R3193Q、TTN:p.R328H、TUBA3D:p.R243Q、TUFT1:p.A340T、TXNDC15:p.R343Q、UBE2NL:p.R86I、UBIAD1:p.A97T、UGT2A1:p.N97fs、USH2A:p.F2369L、USP11:p.A286T、USP25:p.R1119Q、USP26:p.R861Q、USP29:p.F81L、USP31:p.D391N、USP40:p.S851L、UTP14A:p.V148I、VAV3:p.E685K、VCAN:p.R1125H、VPS13C:p.D1359Y、WBSCR17:p.R228C、WDR3:p.E841K、WDR52:p.A157T、XKR6:p.R268Q、XPOT:p.R541W、YTHDC1:p.R267Q、YTHDC2:p.E634K、ZBBX:p.R596I、ZBTB24:p.L607I、ZC3H13:p.R103Q、ZCWPW2:p.D144N、ZEB2:p.R156H、ZFHX4:p.E237D、ZFP14:p.R386C、ZFP28:p.R525I、ZFP2:p.R150I、ZFP3:p.R273I、ZFP90:p.R330Q、ZHX2:p.V790I、ZIC4:p.S305L、ZIM3:p.D352N、ZKSCAN4:p.R319Q、ZMYM4:p.R1446Q、ZNF117:p.R185I、ZNF167:p.R683I、ZNF180:p.R401I、ZNF19:p.R349I、ZNF205:p.R384C、ZNF236:p.S1480L、ZNF248:p.R568I、ZNF259:p.R174I、ZNF266:p.R512Q、ZNF266:p.R344Q、ZNF280B:p.E363K、ZNF283:p.R392Q、ZNF32:p.S62L、ZNF345:p.R82Q、ZNF345:p.R334I、ZNF350:p.R310Q、ZNF434:p.R306C、ZNF439:p.E239D, ZNF439:p.R262I, ZNF443:p.R301I, ZNF445:p.L682M, ZNF470:p.R641I, ZNF471:p.R282I, ZNF484 :p.R138C、ZNF528:p.R279Q、ZNF563:p.K26N、ZNF573:p.R350I、ZNF583:p.R344I、ZNF585A:p.E638K、ZNF 585A:p.E491D、ZNF625:p.R235Q、ZNF652:p.K327N、ZNF677:p.R451I、ZNF678:p.R368I、ZNF699:p.R41I、 ZNF70:p.R244I, ZNF770:p.S441P, ZNF774:p.R423Q, ZNF782:p.K247T, ZNF7:p.R337I, and ZNF831:p.E949D. .

[0152] 62. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0153] (a) This subject population has DLBCL; and

[0154] (b) The at least one tumor-specific mutation includes any combination of mutations selected from the group consisting of: EZH2: p.Y641F, MYD88: p.L273P, BCL2: p.G33R, CARD11: p.E626K, ADCY2: p.A87V, BCL2: p.N172S, BCL2: p.H20Q, BRAF: p.K601E, BTG1: p.L31F, CACNA1E: p.R1458C, CARD11: p.E93D, CD79B: p.Y197D, CD79B: p.Y197H, CREBBP: p.R1446 H. GRID1:p.E622K, HIST1H1C:p.A65V, HIST1H1E:p.G133A, HIST1H3B:p.A48S, KRAS:p.G13D, MYD88:p.S251N, PABPC1:p.R94C, PIM1:p.L16 4F, PIM1:p.L184F, POU2F2:p.T239A, POU2F2:p.T239S, RELN:p.R2971Q, SLC25A48:p.A67T, STAT6:p.D468H, TNF:p.L47F, and TRAF7:p.R11H.

[0155] 63. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0156] (a) This subject group has KICH; and

[0157] (b) The at least one tumor-specific mutation includes any combination of mutations selected from the group consisting of: ACR: p.W279C, AGRN: p.1284_1285VT>A, C7orf25: p.R384fs, CAMSAP1: p.T466fs, CBWD6: p.E102fs, DOCK8: p.L1111fs, EBPL: p.Q196P, EBPL: p.L189V, GFM1: p.A17fs, GOLGA6L6: p.D570E, ITGA5: p. A48D, LUZP2: p.S154fs, MTMR9: p.K193fs, MUC16: p.P10452fs, MUC4: p.S2832P, ODF2L: p.K407fs, RHBDD3: p.G34fs, RILPL 1:p.S358R, TAS2R30:p.L236fs, TRRAP:p.A973S, UBR5:p.K2120fs, URGCP:p.G639fs, ZNF98:p.A222T, and ZSWIM6:p.Q610fs.

[0158] 64. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0159] (a) This subject group has KIRP; and

[0160] (b) The at least one tumor-specific mutation comprises any combination of mutations selected from the group consisting of: FAM18B2: p.C51Y, ZNF598: p.E25G, NEFH: p.E645K, EEF1B2: p.S43G, NEFH: p.AKSPEKEE652del, OBP2B: p.K61N, SKI: p.A62G, C14orf126: p.R6W, KRT8: p.S59A, ACSBG2: p.I250M, ASIC2: p.R46L, CSGALNACT2: p.L362F, FRG1B: p.A50P, IDUA: p.H33Q, KRTAP 4-5: p.S74C, SCAF11: p.E926fs, SYN2: p.A34del, ZNF814: p.R322K, BMS1: p.E878D, JMY: p.P822T, KIF1A: p.E917D, KRTAP4-7: p.S57P, LAMA5: p.L2223 R, LRP1: p.P1058T, MED16: p.H449Q, MUC2: p.T1488P, MUC5B: p.D682G, NACA2: p.R75K, NEFH: p.665_666insEE, OR2L8: p.S201fs, RGPD5: p.P1760A, RRN 3:p.P11S, RRN3:p.R9C, STAG3L2:p.L81fs, ZNF814:p.G320E, ACP6:p.V29G, AHNAK2:p.S2166F, AHNAK2:p.P1215S, AP1G1:p.I782fs, AQP2:p.N68T, BA IAP2L2:p.V396M, BMP6:p.Q118L, BST1:p.G36A, CDR1:p.V31A, CLDN7:p.S172A, CLIP1:p.S1018fs, COL18A1:p.G884fs, CROCC:p.A355P, CTAGE15P:p. A364V, CUBN: p.I2816M, DMRT2: p.T106S, DPY19L1: p.V249L, DSPP: p.D1047N, EBPL: p.L189V, EIF4G1: p.E465del, EXOSC2: p.R11P, FAM216A: p.P36S, F CGR2A:p.V222G, FMOD:p.S331R, FOLR2:p.Q112R, FRG1B:p.L20P, GAGE2B:p.9_10insY, GDPD5:p.G593fs, GIMAP8:p.A544S, GLUD2:p.R300G, GLUD2:p.S496R, GPR135:p.Q5P, HOXD8:p.Q67H, IER5:p.R194G, IL25:p.C168fs, JSRP1:p.V92A, KRAS:p.G12D, KRTAP1-1 :p.Y86C, KRTAP4-11: p.L161V, LTBP1: p.L163P, MAML2: p.Q591K, MAPK7: p.A501D, MEF2A: p.P99S, MET: p.H1094Y , MET:p.M1250T, MST1:p.N435fs, MUC2:p.T1582R, MUC2:p.T1722I, MUC4:p.A4222T, MUC4:p.T2335M, MUC4:p.P 1138L, MUC5B:p.S1098A, MUC5B:p.S3431N, MYH7:p.A1487T, NBPF10:p.R39fs, NBPF10:p.Y638S, NEFH:p.654_65 4S>SPEKAKS、PARG:p.A584T、PBX2:p.Y262F、PIP4K2A:p.R219K、RLIM:p.S471P、RUNX2:p.Q71E、SGK223:p.R63S , SMARCB1:p.L365fs, SRCAP:p.Q1875fs, TBC1D2B:p.R920Q, TCF7L2:p.R482fs, TMEM131:p.K640fs, TMEM60:p.K 77fs, TPPP:p.R30K, TRPV3:p.A218E, TTBK2:p.C83W, UBXN11:p.S510G, UGT1A1:p.T4A, UTS2R:p.A289E, YBX1:p. P250L, ZNF514:p.V81G, ZNF516:p.A256D, ZNF681:p.K405Q, ZNF814:p.D404E, ZNF814:p.P323H, ZXDB:p.G206R. .

[0161] 65. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0162] (a) This subject population has LIHC; and

[0163] (b) The at least one tumor-specific mutation comprises any combination of mutations selected from the group consisting of: TP53:p.R249S, CTNNB1:p.D32V, CTNNB1:p.D32G, CTNNB1:p.S33P, CTNNB1:p.K335I, CTNNB1:p.H36P, EEF1A1:p.T432L, GNAS:p.R844C, OR2T4:p.V137L, TP53:p.H193R, ATXN1:p.Q217H, CSMD3:p.F2383fs, CTNNB1:p.D32N, CTNNB1:p.S33C, CTNNB1:p.G34V, C TNNB1:p.S45P, CTNNB1:p.N387K, DHRS4:p.I218T, DNM2:p.E378D, F5:p.Q426L, GALNTL5:p.A45T, GPX1:p.P77R, GRM8:p.R852C, IDH1:p.R132C, KIF26B: p.A2033T, KRT8:p.S59A, LOC100132247:p.T532P, NEB:p.D3854H, PIK3CA:p.H1047R, SOLH:p.R714H, TP53:p.R158H, TP53:p.V157F, and ZNF638:p.D400N.

[0164] 66. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0165] (a) This subject group has MM; and

[0166] (b) The at least one tumor-specific mutation comprises any combination of mutations selected from the group consisting of: NRAS: p.Q61R, KRAS: p.Q61H, KRAS: p.G13D, NRAS: p.Q61K, BRAF: p.V600E, NRAS: p.Q61H, NRAS: p.G13R, ZNF717: p.W315C, ATP13A4: p.V431G, DNAJC12: p.R135K, IRF4: p.K12 3R, KRAS:p.A146T, KRAS:p.Q61R, KRAS:p.G12A, KRAS:p.G12D, ZNF717:p.N594I, ACTG1:p.A22P, ARL6IP1:p.M 75L, BEND2:p.E630K, BRAF:p.G469A, CDHR1:p.R218G, DIS3:p.R780K, DMXL2:p.D2412E, DNAJC10:p.I80K, EGR 1:p.Q9H, FGFR3:p.*807S, IDH1:p.R132C, IL6ST:p.P216H, INTS12:p.M1V, KRAS:p.K117N, KRAS:p.A59G, KRAS :p.G12R、MAX:p.R36W、MLL5:p.G492E、NBPF1:p.E810K、NRAS:p.Q61L、NRAS:p.G12D、ODF2L:p.E294K、PADI2:p .T114P, PNLIP:p.T37M, PRDM1:p.S588C, PTPN11:p.E76K, PTPN14:p.E286K, RBM6:p.V675G, SCN10A:p.R1142H , SRGAP1:p.T61M, SUSD1:p.T168P, TAS2R16:p.V231I, TINAG:p.E403K, TRIP12:p.L1775P, and ZNF717:p.C844S.

[0167] 67. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0168] (a) This group of subjects suffers from PRAD; and

[0169] (b) The at least one tumor-specific mutation comprises any combination of mutations selected from the group consisting of: HSD17B7P2: p.N175S, RGPD5: p.P1760A, FRG1B: p.L52S, EEF1B2: p.S43G, FRG1B: p.I10T, FRG1B: p.A53T, LRRC37A2: p.T102S, NBPF10: p.E3455K, PTH2: p.L22V, CYP2D7P1: p.S32A, FAM47C: p.N648D, MAP3K9: p.E38del, MUC4: p.H4205Q, CHEK2: p.K373E , FRG1B:p.A11T, FRG1B:p.A50P, HLA-J:p.R124W, KRTAP1-5:p.I88T, KRTAP4-9:p.D18V, NPIP:p.A271V, PDGFRA:p.R483fs, ZNF780A:p.Q600H, ZNF84 5:p.R925H, ZNF91:p.R333H, ARFGAP3:p.N299fs, BTN2A3P:p.P3S, FNBP4:p.TT58del, HLA-A:p.Q78R, LOC554223:p.RAPWMEQ147del, PODXL:p.28_30 PSP>P, POLI:p.D17del, SPOP:p.F133L, SYN2:p.A34del, TMEM52:p.23_26LLPL>L, UBC:p.L149R, ZNF208:p.I647S, ZNF799:p.E589G, ZNF814:p.D404 E. ASTN2:p.L221del, B4GALNT1:p.G88fs, C16orf74:p.S21del, CCDC15:p.H458P, CD209:p.R129W, CNTNAP1:p.S1029I, DBR1:p.541_542DD>D, FAM22 F:p.S691del、FRG1B:p.D32V、FRG1B:p.I34T、FRG1B:p.N55D、FRG1B:p.I59V、FRG1B:p.S71N、KIF25:p.W3R、KRTAP4-11:p.L161V、KRTAP4-11:p.M93V , KRTAP4-11:p.R51K, KRTAP4-6:p.S153Y, LILRB5:p.S598P, LMOD2:p.E124del, LOC645752:p.L40P, LRP1:p.P1058T, LRRIQ3:p.K244fs, LURAP1L:p.55_56insGGG、MLLT10:p.V463E、MYOCD:p.Q310del、NBPF10:p.N1369D、OT UD4:p.T909I、PARG:p.A584T、PEX1:p.I370fs、POTEC:p.K507E、POTEC:p.R 477Q, POU4F2:p.68_69insG, PRG4:p.T417P, SDHAP2:p.R31C, SPOP:p.F133 C, SPOP: p.W131G, TIMD4: p.T152del, TMEM121:p.P299del, TP53:p.G245S. UBC: p.R73L, UBC: p.I191T, WASH3P: p.G175S, ZMIZ1: p.D1048fs, ZNF709: p.T413I、ACADS:p.R330H、ADAMTS7:p.K1357fs、AFF2:p.R597H、AGAP6:p.S 127I, AK302238:p.A44T, AK302879:p.Q191R, ALDH1A2:p.R85C, ANAPC1:p. T537A, ANKRD36C:p.H438R, AP4B1:p.R276W, ARFGAP2:p.S38N, BBS9:p.F26 8fs, BC139719:p.L133R, BRAF:p.G469A, C22orf43:p.D171del, CANT1:p. K131R, CHD3:p.E35del, CLEC4A:p.R209H, CNOT3:p.E20K, CNPY3:p.17_18L L>L、CNTNAP3B:p.S317T、CNTNAP3B:p.M1247I、CTNNB1:p.T41A、DDX10:p.D 788part, DLC1:p.S741T, DPY19L2:p.M210V, EDC4:p.S617part, EFCAB6:p.R3 79K、ERC2:p.927_928HH>H、FAM111B:p.S269fs、FEM1A:p.L620M、FHOD3:p. A632fs、FLJ43860:p.L850fs、FMN2:p.G59del、FNBP4:p.914_915PP>P、FRG 1:p.E86del, FRG1B:p.K13N, FRG1B:p.P42Q, GABRB1:p.R416C, GABRR2:p.A 368V、GAGE2B:p.9_10insY、GOLGA8DP:p.N84H、GOT2:p.R355W、GPATCH4:p.K210fs、HDGFL1:p.188_189insA、HLA-DQB2:p.G250S、HLA-DQB2:p.R247H、 IDH1:p.R132H、IL27:p.E176del、IRF2BPL:p.123_125QQQ>Q、KANK3:p.DGD S489del、KIAA1462:p.858_859SS>S、KRTAP4-11:p.S48R、KRTAP4-7:p.S57 P、KRTAP4-8:p.C95S、LPHN3:p.R826H、LRP10:p.L11del、LRP5:p.S1609P、L RRC16B:p.R787W、MAS1L:p.R324G、MECOM:p.R915Q、MED12:p.L1224F、MED1 2L:p.Q2115part、MESP2:p.GQGQGQGQ195part、MGAT4C:p.T345M、MLEC:p.E23 8del, MSLNL:p.T68P, MUC7:p.S173P, MYC:p.Q37del, NBPF10:p.N440D, NLR P6:p.E611del, NOX3:p.C404fs, OR1M1:p.V69I, OR7E24:p.L7fs, OTUD4:p. A153del, PANK2:p.T417fs, PCLO:p.S496P, PCNT:p.S162G, PCSK9:p.23_24insL, PHOSPHO1:p.S32del, POU4F1:p.H108del, PRAMEF8:p.R319H, PRDM7:1. p.M387L、PRG4:p.T597P、PTPRD:p.R1323C、PTPRF:p.R1174Q、ROBO3:p.RS1 367del、ROCK1:p.T518S、RPTN:p.G296S、RTL1:p.152_152E>EE、SIRPA:p.V 233I, SLC2A6:p.A230D, SLC8A2:p.E710del, SMG7:p.E846fs, SNAPC4:p.S5 42part, SP8:p.G165part, SPOP:p.F133I, SPOP:p.F133V, SPOP:p.F102C, SPO P:p.F102V、SRSF11:p.G17fs、SRSF4:p.K396del、SSPO:p.S4198fs、STAG3L 2:p.L81fs、STK19:p.R18fs、TBC1D2B:p.R920Q、TBC1D9:p.P1233T、TCHH:p.P1158R, TCOF1:p.K1366del, TNRC18:p.2664_2665SS>S, TP53:p.R248Q, TP53:p.R175H, TP53:p.C141G, TSPAN4:p.L92V, UBXN11:p.GP GPGPSP504del, UTP3:p.E81del, WASH3P:p.L187V, ZAN:p.P717L, ZAN:p.L878P, ZFP90:p.R591fs, ZNF761:p.H373R, and ZNF91:p.H305R. .

[0170] 68. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0171] (a) This subject group has STAD; and

[0172] (b) The at least one tumor-specific mutation includes any combination of mutations selected from the group consisting of: RNF43: p.G659fs, BZRAP1: p.P1416fs, XYLT2: p.Y526fs, LARP4B: p.T163fs, PGM5: p.I98V, ZBTB20: p.P692fs, ARID1A: p.G1848fs, FHOD3: p.P334fs, KIAA0182: p.T120fs, ATP6V1B1: p.Y383fs, PIK3CA: p.H1047R, FRMD4A: p.P1005fs, PIK3CA: p.E545K, CD C14A:p.N123fs、KRAS:p.G13D、MLL2:p.T172fs、BCORL1:p.S1679fs、PLEKHA6:p.V328fs、C9orf131:p.P342fs、CD4:p.Q164fs、FBXW7:p.R465C、GNG12 :p.T68fs、IRS4:p.G591fs、JARID2:p.V422fs、KIAA0195:p.I902fs、MBD6:p.P732fs、MVK:p.P138fs、PAMR1:p.G101fs、WNT16:p.W165fs、ZNF43:p.N25 1fs、ABCA6:p.L306fs、ADAM28:p.K73fs、AOC3:p.L79fs、ATP2A1:p.R819fs、B2M:p.L13fs、C6orf89:p.P58fs、CNTLN:p.K1305fs、CR2:p.V206fs、DYRK 4:p.K468fs、ERBB3:p.V104M、GLI1:p.W272fs、KRAS:p.G12D、MLL2:p.T172fs、MSH6:p.T1085fs、NLK:p.C190fs、OR5M3:p.T89fs、PAX6:p.P375fs、PTEN :p.L265fs、RABGAP1:p.K928fs、RAD51AP2:p.T316fs、SVIL:p.G1862fs、TP53:p.R273H、WNK4:p.G606fs、ARID1A:p.P2139fs、AXIN2:p.G665fs、C13orf 33:p.R67fs, C1QTNF5:p.P308fs, CELSR1:p.G614fs, CRYGD:p.G159fs, DCHS1:p.R235fs, DDC:p.I433fs, EDNRB:p.Y383fs, EPHA2:p.P460fs, FOXN3:p.P96fs、HDAC4:p.P901fs、INF2:p.S527fs、KIRREL2:p.V649fs、KLF3:p.I104fs、KLHL14:p.P231fs、MAP7D3:p.Q308fs、OTX2:p.R44fs、PAFAH1B1:p.K302fs、PLAGL2:p.P10fs、POLM:p.P97fs、PRPF40B:p.I31fs、RALGAPB:p.T379fs、SBNO1:p.N1139fs、SERPINI1:p.L81fs、SH3KBP1:p.L574fs、SLC12A7:p.H686fs、SLC27A3:p.P643fs、TBX4:p.S370fs、TP53:p.R273C、TP53:p.R175H、TRAM1L1:p.R345fs、WBP1:p.P138fs、ABCC4:p.L883fs、AKAP13:p.K2785fs、ALDH3A1:p.P562fs、ALPK2:p.L356fs、ARFGEF1:p.P1552fs、ARID1A:p.G1848fs、AVPR1A:p.F351fs、BAX:p.M38fs、C14orf43:p.P313fs、C1QTNF5:p.G194fs、C7orf50:p.L179fs、CDC25C:p.K322fs、CETN3:p.K63fs、CHD3:p.P597fs、CTCF:p.K202fs、CTSC:p.F105fs、DDX17:p.G163fs、DLGAP3:p.G377fs、EBF3:p.G255fs、FHDC1:p.F100fs、FILIP1L:p.K749fs、FLNB:p.W529fs、GBP7:p.G431fs、GCC2:p.E700fs、GPR161:p.G517fs、IWS1:p.S802fs、KIAA0240:p.K895fs、KIAA1967:p.P415fs、LRRC43:p.D558fs、MACF1:p.R707fs、MBD6:p.G780fs、MLL3:p.F4496fs、MPRIP:p.A351fs、MUC6:p.2129_2130SS>S、NOX5:p.P467fs、OPTN:p.P24fs、OR4K5:p.F177fs、PIK3CA:p.N345K、PIK3CA:p.E542K、PLXNA1:p.P1016fs、PNPLA7:p.P1199fs、PODN:p.I301fs、PPP2R3B:p.T389fs、PRSS36:p.L680fs、RGL2:p.G203fs、RHOQ:p.V190fs、RNF111:p.R771fs、RTN2:p.P313fs、SALL4:p.V995fs、SBF1:p.P1076fs、SETDB2:p.R715fs、SNAPC2:p.T292fs、SPG20:p.F232fs、SRCAP:p.P1876fs、STAT2:p.P489fs、TCHP:p.E172fs、TP53:p.R282W、TP53:p.R248Q、USP21:p.K474fs、WDR7:p.G262fs、ZBTB7C:p.E157fs、ZFC3H1:p.K385fs、ZNF124:p.T339fs、ZNF626:p.K115fs、ADNP2:p.S322fs、AGAP1:p.G127fs、ALDH2:p.L286fs、ARHGAP5:p.D890fs、ARHGEF17:p.A615fs、ARID1A:p.Y1324fs、ART1:p.I243fs、ASCL4:p.D35fs、ATXN2L:p.G998fs、B3GNT5:p.F30fs、BCKDHA:p.H37fs、BCL9L:p.P1127fs、BEND3:p.D265fs、BNC2:p.S575R、BRD3:p.P24fs、C12orf51:p.P4235fs、C1R:p.P216fs、C7orf49:p.G130fs、CA2:p.I145fs、CABP5:p.R145fs、CASD1:p.F781fs、CASP8:p.R471fs、CCDC153:p.P200fs、CD93:p.D280fs、CROT:p.L32fs、CSF3R:p.P468fs、CTCF:p.K202fs、ERBB2:p.S310F、FAM46D:p.S69R、FBN3:p.G601fs、FBXO21:p.F144fs、GAS6:p.G150fs、GLYR1:p.G380fs、GXYLT1:p.L223fs、HAUS6:p.S530fs、IGF2R:p.T1314fs、ITGB1:p.L378I、KDM3B:p.P1316fs、KIF13A:p.K1115fs、KLF3:p.S224fs、LARP1:p.A223fs、LRP1:p.G1488fs、LRP1:p.G1488fs、MAGEE2:p.Q45fs、MAMSTR:p.P162fs、MAPK15:p.Q511fs、MLL2:p.P647fs、MOCS2:p.P22fs、MTG1:p.L105fs、MTG1:p.H327fs、MTIF2:p.N109fs、NID2:p.R1035fs、PAX2:p.P395fs、PCCA:p.R230H、PDZD2:p.R101fs、PFKP:p.M593fs、PIK3CA:p.R88Q、PLA2G1B:p.L53fs、PLAU:p.R201fs、PMEPA1:p.P208fs、POP1:p.K750fs、PTCH1:p.P1307fs、PTPRT:p.P1075fs、RDBP:p.P6fs、RNMT:p.K392fs、ROBO2:p.P1080fs、RUNDC3B:p.L6fs、SDAD1:p.K275fs、SLC10A6:p.G109fs、SNAPC1:p.D211fs、SPATA5L1:p.C685fs、SPTA1:p.K1732T、STAT5B:p.P367fs、SYT4:p.M1fs、TAF1L:p.K851fs、TAP2:p.L75fs、TBL1XR1:p.N126fs、THEMIS:p.K406fs、TMEM79:p.P161fs、TP53:p.C176F、TP53BP2:p.K69fs、TP53RK:p.L174fs、UBQLN2:p.A523fs、UHRF1BP1:p.I1330fs、VPRBP:p.K939fs、VPS13B:p.T56fs、WASF3:p.P305fs、YLPM1:p.E1178fs、ZC3H13:p.K1006fs、ZC3H18:p.P825fs、ZC3H4:p.E779Q、ZNF48:p.P247fs、ZNF608:p.A465fs、ZNF878:p.S238fs、ZSCAN18:p.P225fs、ABCB1:p.R527fs、ABCB6:p.G318fs、ACACB:p.G255fs、ACP1:p.Q123fs、ACTL6A:p.L88fs、ADAMTSL4:p.G778fs、AGBL5:p.I420fs、AHI1:p.K303fs、AKAP9:p.M3743fs、AKD1:p.R1209fs、ANKRD40:p.D99E、ARHGEF5:p.S1512fs、ARID1A:p.K1071fs、ARID3A:p.S557G、ARPP21:p.I130fs、ASPN:p.F67fs、ASXL3:p.E873fs、ATP6V1C2:p.R312fs、BEST3:p.P444fs、BRAF:p.P403fs、BRMS1:p.G107fs、BTBD11:p.T451fs、BTBD11:p.A561V、C11orf9:p.S261fs、C14orf102:p.R90fs、C14orf43:p.Q36fs、C15orf52:p.G98fs、C19orf21:p.R262C、C19orf70:p.P50fs、C20orf160:p.P46fs、C3:p.P890fs、CADPS2:p.N468fs、CASC3:p.S232F、CASC3:p.P603L、CASC3:p.P645L、CASC3:p.S658L、CASKIN2:p.P727fs、CBLL1:p.E138fs、CBLN3:p.P69fs、CCDC108:p.P1164fs、CCDC148:p.K420fs、CCDC153:p.P200fs、CCDC169-SOHLH2:p.K162R、CCDC88A:p.K677fs、CD1E:p.F85V、CD3EAP:p.K218fs、CDH11:p.K357T、CDH1:p.D254Y、CDH23:p.V403I、CFI:p.K37fs、CHPF2:p.D645fs、CIC:p.R507fs、CIC:p.A1114fs、CIC:p.A1114fs、CLSTN1:p.T615M、CNBD1:p.L396P、CNGA4:p.K510T、CNOT6:p.S248fs、CNTROB:p.R920fs、COL9A1:p.P283fs、CPAMD8:p.P784fs、CR1L:p.L79fs、CRB1:p.F630V、CSMD1:p.L3410V、CTNNA3:p.K856fs、CTNND1:p.I447fs、CTSD:p.P89fs、CUX1:p.A439fs、CYP7B1:p.K332T、DAB2IP:p.D994fs、DNAH11:p.T871fs、DNAH8:p.K1688fs、DNAJC1:p.K193fs、DNM2:p.P791fs、DSTN:p.F101fs、DYRK1B:p.Q545fs、EAF2:p.V109fs、EDNRB:p.A104V、EEA1:p.N570fs, EFHA1:p.F290fs, EGR1:p.P332fs, EIF4G3:p.K563fs, ELK3:p.S173fs, ENTPD2:p.G204fs, EOMES:p.G332fs, EPHA10:p.P868fs, EPHB6:p.G54fs, EPHX1:p.P132fs, EPPK1:p.G2015fs, ERBB4:p.M1fs, ESF1:p.T99fs, EXOSC8:p.L160fs, FAM113B:p.R51fs, FAM116A:p.L441fs, FAM135B:p.S645R, FAM151A:p.P117fs、FAM193A:p.D428fs、FAM193A:p.D428fs、FAM214B:p.A42fs、FAM40B:p.R740C、FAM70B:p.S19L、FASTKD1:p.K3fs、FBXW7:p.R479Q 、FBXW9:p.G298fs、FER:p.L474fs、FERMT2:p.K152fs、FGGY:p.G138fs、FIGNL1:p.K309fs、FLG:p.K159fs、FLNB:p.W529fs、FOLH1:p.S501fs、FYB:p.G3 24fs、GABRD:p.Q412fs、GALNTL1:p.W317fs、GANAB:p.L23fs、GCDH:p.L389fs、GIMAP7:p.V276fs、GIPC3:p.G227fs、GLI3:p.P1033fs、GLIPR1L2:p.G9 2fs、GNPNAT1:p.F54fs、GON4L:p.M134fs、GPATCH4:p.K210fs、GRK4:p.K22fs、GTF3C1:p.S767fs、GTF3C4:p.E562fs、H2AFY2:p.K144fs、HCFC1R1:p.P8 3fs、HCRTR2:p.S9fs、HCRTR2:p.S9fs、HDLBP:p.G747fs、HECA:p.R333fs、HIVEP3:p.H554fs、HIVEP3:p.P534fs、HLA-C:p.P209fs、HOOK1:p.L361fs、HO XD8:p.P122fs、HTT:p.G697fs、IBTK:p.K1213fs、IDE:p.K37fs、IFT172:p.A837T、INPPL1:p.A974fs、INPPL1:p.P1154fs、INSM2:p.T533fs、INTS12:p.L14fs、INVS:p.R815fs、IPO11:p.S844fs、IRX6:p.A425V、ISG20L2:p.P288fs、ITGB8:p.A7fs、JARID2:p.G394fs、JHDM1D:p.R97fs、KBTBD6:p.G442fs、KCNC1:p.K455fs、KCNH2:p.G149A、KCNJ10:p.P102fs、KCNMB2:p.N151K、KCTD21:p.T6M、KIAA0586:p.A1592fs、KIAA1009:p.F406fs、KIAA1109:p.E1588fs、KIAA2026:p.K690fs、KIF26B:p.S1065fs、KIF6:p.L204fs、KIRREL:p.P335fs、KLC2:p.T568fs、KRAS:p.Q61H、KRAS:p.G12S、MAN1C1:p.G431fs、MAP1A:p.P2063fs、MAP2:p.K1472fs、MAP3K12:p.R449del、MAP7D1:p.A80fs、MGST2:p.K102fs、MKI67:p.T1664fs、MKL1:p.P307fs、MLL2:p.P2354fs、MLL2:p.L656fs、MLL2:p.P647fs、MLL2:p.L1877fs、MMP3:p.I64fs、MPDZ:p.K1582fs、MTUS2:p.R1005W、MUC16:p.A6156T、MYB:p.R481fs、MYEOV:p.L269fs、MYH11:p.K1263del、MYO18A:p.P209fs、MYO7A:p.I539fs、MYOCD:p.G226fs、NAA16:p.H514fs、NBEA:p.V2247fs、NCAPD3:p.Q909fs、NCAPH:p.T466fs、NCOR2:p.P1308fs、NEFM:p.A213V、NEK8:p.V690fs、NF1:p.T676fs、NHLRC1:p.F204fs、NKD1:p.P286fs、NPR3:p.Y138H、NT5M:p.P206fs、NUFIP2:p.R224fs、NUP210:p.L135fs、NYNRIN:p.G113fs、OBSCN:p.G997fs、OGDH:p.Y948fs、OR4C16:p.S135R、OR51A7:p.L124R、OR7C1:p.C179fs、OSBP2:p.H627fs、OTOF:p.E1304K、P2RX1:p.R20fs、PALB2:p.M296fs、PALB2:p.N280fs、PANK1:p.K400fs、PAPD4:p.C225fs、PAPPA2:p.I1683fs、PARP15:p.K461fs、PARP4:p.K847fs、PCDH10:p.N118fs、PCDH10:p.P225fs、PCGF3:p.H63fs、PELI2:p.G197fs、PHACTR1:p.V251fs、PHACTR2:p.S237fs、PHACTR4:p.S354fs、PHKB:p.K642fs、PIAS3:p.H116fs、PIGO:p.P787fs、PIGT:p.A346fs、PIK3R3:p.M341fs、PITPNM1:p.P295fs、PKN2:p.K76fs、PLA2G15:p.W230fs、PLAG1:p.K184fs、PLEKHO1:p.T254fs、PLOD3:p.R297fs、PLOD3:p.P296fs、PLXNA2:p.P464fs、POLQ:p.L1430fs、PPARGC1B:p.P135fs、PPL:p.P454fs、PPM1H:p.P226fs、PPP1R12C:p.P372fs、PREX2:p.R562fs、PRICKLE4:p.Q109fs、PRKAR1B:p.P87fs、PRKCG:p.R345C、PRMT8:p.S28fs、PROX1:p.F592fs、PRRG3:p.R163fs、PSD2:p.G256fs、PTCHD3:p.F588fs、PTPN4:p.N319fs、PTPRC:p.Q895H、PWWP2B:p.S84fs、PYGO2:p.Q150fs、RABGAP1:p.K928fs、RB1CC1:p.N1171fs、RBM6:p.R96fs、RHOA:p.Y42C、RIMS1:p.R71G、RIMS2:p.V401fs、RING1:p.G171fs、RINT1:p.L107fs、RNF43:p.P116fs、ROBO2:p.K1293fs、RPS6KA6:p.K109fs、RRS1:p.N45fs、RSF1:p.K386fs、RUSC2:p.P486fs、RXFP3:p.A60V、SAFB:p.W798fs、SCARF1:p.R614Q、SCLT1:p.K109fs、SERPINB12:p.Q168fs、SGK3:p.L61fs、SGOL2:p.E407fs、SIGLEC1:p.P318fs、SIK1:p.Q678fs、SLC16A6:p.G98fs、SLC25A17:p.F28fs、SLC26A7:p.I629fs、SLC32A1:p.V494I、SLC4A3:p.L1061fs、SLC7A10:p.P157fs、SLC9A2:p.T746fs、SLITRK1:p.K45fs、SND1:p.H721fs、SOAT1:p.F64fs、SORBS2:p.E1158fs、SOX7:p.L309fs、SPAG17:p.Q1264fs、SPTY2D1:p.P485fs、SRCIN1:p.P865fs、SREBF2:p.H763fs、SRRT:p.G102fs、STAB1:p.P1120fs、STRADA:p.R333fs、STX2:p.K252fs、SV2A:p.E138fs、SYCP2:p.M176fs、SYNJ2:p.P1111fs、TAS2R10:p.L196fs、TBC1D22B:p.A175fs、TEAD2:p.P298fs、TFE3:p.G482fs、TGM6:p.T358fs、TIMM44:p.K83fs、TIMP3:p.A199fs、TLR4:p.L498V、TMEM132D:p.P206fs、TMEM41A:p.F156fs、TMEM41B:p.F230fs、TMTC4:p.R611C、TNK2:p.P632fs、TOPBP1:p.I1381fs、TP53:p.E286K、TP53:p.P152fs、TRIP11:p.K541fs、TRPA1:p.T673fs、TRPM8:p.H765fs、TTF1:p.K336fs、TTI1:p.R707H、TTN:p.E15192D、U2AF2:p.L175fs、UBC:p.G684fs、UBR4:p.P2802fs、UPF2:p.E1033D、UPK2:p.P49fs、USP13:p.I116fs、USP15:p.K782fs、VASH1:p.G3fs、VEZF1:p.355_356insN、VPS13A:p.F2883fs、WAPAL:p.R522fs、WDFY3:p.L1842fs、WDR59:p.N160fs、WDR5:p.N214fs、WDR60:p.Q412fs, WDTC1:p.M287fs, WHSC1L1:p.K418fs, WNT1:p.W167fs, XIRP2:p.E1007D, YBX2:p.P226fs, YIF1A:p.R131fs, ZBBX:p.E1 51del、ZBTB40:p.L262fs、ZBTB7C:p.G342fs、ZBTB7C:p.D154fs、ZC3H18:p.T701fs、ZDHHC5:p.E651del、ZDHHC7:p.P316fs、ZFH X3:p.R1893fs、ZFHX3:p.E763fs、ZFHX4:p.L408fs、ZHX3:p.N249K、ZIM3:p.I384fs、ZKSCAN5:p.D13fs、ZMYM4:p.K345fs、ZNF23 6: p.T1410M, ZNF23: p.F122fs, ZNF334: p.K426fs, ZNF358: p.T130fs, ZNF701: p.L296fs, ZNF711: p.L737fs, and ZNF831: p.A49fs. .

[0173] 69. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0174] (a) This subject population has TGCT; and

[0175] (b) The at least one tumor-specific mutation includes any combination of mutations selected from the group consisting of: FAM18B2: p.C51Y, BTN2A3P: p.P3S, MUC2: p.G1715S, NBPF10: p.L44V, SP8: p.G156S, DCP1B: p.Q252H, DEK: p.E41D, ERC1: p.K692R, FAM104B: p.D75H, FRG1B: p.M49V, KRTAP10-10: p.V234M, LRRCC1: p.A6V, NRAS: p.Q61R, PNPLA4: p.L223P, ANKLE1: p.C644 fs. 814:p.Y324H, ADAMTS17:p.N572T, ATRX:p.K1936R, BCL11B:p.E535D, BMP2K:p.Q460H, BMP2K:p.H487Q, C12orf32:p.D60V, C22orf43:p.K19E, CDC27: p.N571I, CDC27:p.P242S, DDX11:p.K208fs, EBPL:p.L189V, EZH2:p.K510R, FAM86A:p.A141T, GAS2L2:p.D189A, GRID2IP:p.LS754del, HGC6.3:p.E17 1G, KIT: p.D816V, KIT: p.N822Y, KIT: p.N822K, KRAS: p.Q61R, KRAS: p.G12V, KRTAP1-1: p.I116V, LRRC37BP1: p.Y166D, MEF2A: p.R127Q, MFF: p.S7F, MS T1:p.R347W、MUC4:p.S3048L、MUC6:p.H2000Q、MUC6:p.P1977H、NAT10:p.I393T、OPLAH:p.A900D、PIEZO1:p.Q749E、PRAMEF4:p.F300V、RBM10:p.E184 D. SERINC2:p.T121P, SPIN2A:p.M150V, SRRM2:p.A2257S, SSBP3:p.K6R, ZNF680:p.R501W, ABCC8:p.Y512C, ABCC9:p.L466P, ABCD1:p.H169Q, ABL2:p.P19T、ACVR2B:p.R48C、AHDC1:p.P33fs、AHNAK2:p.L1640M、ALPPL2:p.W31S、AMMECR1:p.G77C、ANK3:p.D1322E、ANKHD1-EIF4EBP3:p.G60S、ANKRD11:p.Y2015S、ANKRD11:p.K369R、ANKRD50:p.V637M、APBB3:p.L450P、ARHGAP24:p.T35A、ARID4B:p.G1076A、ARMC3:p.A514T、ARRB2:p.T99P、ATAD5:p.I305V、.

[0176] ATXN3:p.305_306insQQQQQQQ、AVPR1B:p.G39R、AXDND1:p.E994Q、BAI2:p.A231G、BEST3:p.P383L、BIRC6:p.V414L、BIRC8:p.A225M、BRWD1:p.K1319R、BTN2A2:p.L15F、C12orf51:p.A2644T、C12orf65:p.K143T、C16orf62:p.L244I、C1QBP:p.T225I、C1orf167:p.S123G、C5orf25:p.Y4F、CACNA1E:p.G2080S、CAPNS1:p.LV303del、CCDC159:p.A332S、CDKAL1:p.P409L、CDYL:p.V48A、CDYL:p.A60G、CELSR2:p.L17P、CHD4:p.E138D、CKAP5:p.G576A、CLCC1:p.K52R、CMTM8:p.S26T、CNKSR2:p.P249L、CNTN5:p.I501T、COG5:p.H617R、COL15A1:p.K708R、COL6A3:p.A2378D、CRYGB:p.R143G、CSGALNACT2:p.L362F、CUL4A:p.I438F、CXXC1:p.Q156H、CYP19A1:p.F406L、DCLRE1B:p.F28I、DDX11:p.A376T、DDX11:p.E680D、DEPDC5:p.R1525Q、DLC1:p.S741T、DNMT1:p.R995Q、DOCK11:p.Q169E、DSPP:p.D1047N、E2F7:p.I91S、EBF1:p.D353G、ECI2:p.K55R、EEF1A2:p.Y418S、EIF3J:p.A8G、EML6:p.K805R、EPAS1:p.S474T、EPRS:p.L1335I、ERICH1:p.E327K、FAM101B:p.L5P、FAM104A:p.M1R、FAM110D:p.R71H、FAM155A:p.Q95R、FAM186A:p.G1492E、FAM194B:p.Y139H、FAM21B:p.P1231S、FAM32A:p.K9R、FAM46B:p.H416R、FAM48B1:p.I499V、FAM48B1:p.A516P、FAM5C:p.S425W、FAM86C2P:p.C120Y、FBXL14:p.V48G、FRMPD3:p.Q832del、FRS2:p.L47S、GDF5:p.E105fs、GPNMB:p.C3fs、GPT2:p.R10P、H2AFV:p.Q125R、HDLBP:p.R503C、HERC2:p.R2129C、HIST1H2BJ:p.K13R、HLX:p.N231K、HMGB3:p.E198D、HSF4:p.R169W、HSF4:p.S491P、HYAL4:p.D222N、INO80E:p.P206fs、INTS4:p.S460A、IQCF6:p.R3H、ITPR1:p.M1569I、ITPR3:p.R1698G、KANSL3:p.G376E、KCNA4:p.E627del、KDM5A:p.P423S、KDM6A:p.Y362fs、KIAA0020:p.K63R、KIDINS220:p.N851S、KIT:p.W557G、KLHDC2:p.W321S、KRAS:p.A146T、KRAS:p.Q61H、KRAS:p.Q61L、KRAS:p.G12A、KRAS:p.G12R、KRBA1:p.R839G、KRTAP4-8:p.T63S、L2HGDH:p.P441del、LAMC3:p.P174Q、LHCGR:p.L16Q、LOC401296:p.L144M、LPHN2:p.F906I、LRP12:p.G310C、LTB4R:p.F73L、LTBP3:p.L35del、LUC7L3:p.S148T、LYPD4:p.T64K、MAMLD1:p.Q572L、MAP4K2:p.R341G、MAPK7:p.A501D、MAT2A:p.E166G、MED12L:p.C1292Y、MESP2:p.Q182E、MEX3C:p.R534S、MIER2:p.L131F、MLL5:p.Y66C、MLLT3:p.177_178SS>S、MMS19:p.D1005N、MRPS25:p.E119del、MSH6:p.D576A、MTIF3:p.G65E、MUC17:p.M1807T、MUC17:p.T2279N、MUC17:p.G2474S、MUC2:p.TTPSPP1475del、MUC2:p.T1568M、MUC2:p.T1580N、MUC2:p.T1704I、MUC2:p.T1706M、MUC4:p.H1117D、MUC5B:p.R1097H、MYEF2:p.K323E、MYEOV:p.L302H、MYH8:p.A785V、MYO1A:p.N584K、NAP1L3:p.P353R、NAV1:p.I1433M、NCAM1:p.E131G、NEB:p.D3107N、NEFH:p.V670E、NELL2:p.G170D、NHS:p.D1561N、NKD2:p.H447del、NSD1:p.T461R、NT5C3:p.A3P、NYAP1:p.P480S、OBSCN:p.A908T、OR10J1:p.R244Q、OR1S2:p.M298I、OR2L3:p.K294R、OR6K6:p.F311L、PABPC3:p.V325fs、PBX2:p.Y262F、PCDHB4:p.P255F、PCMTD1:p.V281A、PCP4L1:p.K64R、PDE3A:p.A98E、PDIA6:p.N56K、PDS5A:p.L1309F、PHLDA2:p.R28S、PIGR:p.V183G、PIK3CA:p.E545K、PIK3CD:p.C381R、PKD1:p.T938M、PLEKHM1:p.A895V、PLEKHN1:p.A600D、PLXND1:p.R367L、PMS2:p.K651R、PNMA3:p.E200G、POTEF:p.S112G、PRAMEF8:p.I448V、PRDM2:p.E278D、PRODH:p.L527V、PRPF31:p.R289W、PSME4:p.N495D、PTGR1:p.E40A、PTPRB:p.Q726H、RABGEF1:p.N207D、RAC1:p.P34R、RANBP17:p.M900I、REV3L:p.A30S、RFC3:p.I82N、RFC3:p.K296N、RIMBP3:p.Q1154R、RPL19:p.R151C、RPL5:p.R58fs、RPTN:p.M538I、RRAD:p.A278E、RYR1:p.D668Y、RYR2:p.L2023F、SAFB:p.G799V、SCRIB:p.G332V、SDK1:p.Y2146C、SEC16A:p.T443K、SEC31B:p.P905S、SELO:p.R565Q、SELP:p.A297T、SI:p.I1681K、SLC2A7:p.H268Q、SLC37A1:p.V528I、SLC38A1:p.G100R、SMARCA2:p.D1158A、SMARCA5:p.T156fs、SMC3:p.E970Q, SMG1:p.P2696H, SNRNP200:p.A2129G, SPIN2B:p.M150V, ST6GALNAC1:p.S354N, STAMBPL1:p.Y14 3H, STARD8:p.G662A, STON1-GTF2A1L:p.N451S, SYMPK:p.A336G, TAS2R8:p.W98C, TCHH:p.W1016R, TET1 :p.T1472S、TIAM1:p.G247M、TNS1:p.P183S、TOR1AIP2:p.G146R、TPRX1:p.S216P、TPRX1:p.S200P、TRMT 61A:p.S244I, TSPAN4:p.L92V, TTF1:p.Q530R, UBE2M:p.G131D, UBR5:p.R2517S, UGT2B11:p.R447I, UMO DL1:p.M559I, UNC93A:p.V445A, USP46:p.Q137R, VWA2:p.G317D, VWA7:p.V792G, WASH3P:p.L187V, WNT5 B:p.K327E、WRN:p.E510D、XDH:p.P410S、ZAN:p.S755P、ZC3H11A:p.I777T、ZC3H7A:p.C575S、ZDHHC11:p .H250Q、ZFHX4:p.D3239N、ZKSCAN3:p.K200A、ZMYM4:p.T367I、ZNF174:p.P353T、ZNF322:p.Y353C、ZNF5 92:p.K324Q, ZNF592:p.P500T, ZNF782:p.C145F, ZNF799:p.C453R, ZNF804B:p.P644S, and ZNRF3:p.R889W. .

[0177] 70. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0178] (a) This group of subjects has THCA; and

[0179] (b) The at least one tumor-specific mutation includes any combination of mutations selected from the group consisting of: BRAF: p.V600E, NRAS: p.Q61R, HRAS: p.Q61R, NRAS: p.Q61K, OTUD4: p.T909I, HRAS: p.Q61K, NLRP6: p.E611G, AKT1: p.E17K, ANKMY1: p.N302I, ATP6V1A: p.L237P, CYP19A1: p.S113I, DCUN1D4: p.L275P, DGCR8: p.E518K, DLC1: ​​p.S741T, DNAH1 0:p.C1853F, EIF1AX:p.G9D, FAM75D5:p.L222P, FCGRT:p.P40A, KRAS:p.Q61K, LMX1B:p.Q285del, MAS1L:p.R324G, MED15:p.S35I, MEGF6:p.Y3 93C, ODZ2:p.A1529V, OR5L1:p.R122H, OR6K6:p.F311L, OTX1:p.D315N, POTEE:p.S75G, SCN5A:p.D1978H, TOP2A:p.K1199E, and TSG101:p.K265R.

[0180] 71. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0181] (a) This subject population has UCS; and

[0182] (b) The at least one tumor-specific mutation includes any combination of mutations selected from the group consisting of: TP53:p.R248Q, ZNF814:p.D404E, BTN2A3P:p.P3S, FBXW7:p.R465C, FRG1B:p.G65E, MUC4:p.H4205Q, NBPF10:p.V99F, PIK3CA:p.E545K, PIK3CA:p.H1047R, PPP2R1A:p.P179R, DDX11L2:p.*128Q, FBXW7:p.R479Q , FRG1B:p.K13N, FRG1B:p.L52S, HSD17B7P2:p.N175S, KRAS:p.G12V, LOC283788:p.S37G, TP53:p.R273H, TP53:p.S241Y, ADAMTS 12:p.E359K、BCL2L11:p.L187fs、CDC27:p.L460fs、CHEK2:p.K373E、ESPNP:p.W122fs、FBXW7:p.R689W、FBXW7:p.R505G、FBXW7:p .R465H, FCGBP:p.V4019M, FRG1B:p.I10T, FRG1B:p.D32V, FRG1B:p.R37K, KRAS:p.G12D, LOC100233156:p.R21C, LOC283788:p.I 46M, LRP1B:p.L1392F, MAMLD1:p.Q572L, MST1P9:p.L319P, MUC4:p.A2390T, MUC4:p.G2172S, NBPF10:p.E3455K, PIK3CA:p.G106V , PODXL:p.28_30PSP>P, POTEC:p.R477Q, PPP2R1A:p.R183W, PPP2R1A:p.S219L, PTPN18:p.TG378del, RGPD3:p.N756D, RPL13AP20 :p.G107R, SAMD4B:p.R477W, SMAP1:p.E169fs, TP53:p.H193R, TP53:p.H179R, TP53:p.R175H, TUBBP5:p.R119H, and U2AF1:p.S34F.

[0183] 72. The pharmaceutical composition as described in any one of paragraphs 30-36, wherein:

[0184] (a) This subject group suffers from PAAD; and

[0185] (b) The at least one tumor-specific mutation includes any combination of mutations selected from the group consisting of: RBM14: p.AAAAAAA286del, KRAS: p.G12D, JMY: p.PPPPPPPPPPPP811del, RIOK1: p.D69del, LCE2A: p.SSGGCCGSSSGGCC47del, KRAS: p.G12V, C1QB: p.GPKGPMGPKGGPGAPGAP90del, ZFHX3: p.V777del, DBR1: p.541_542DD>D, AEBP1: p.K1133del, KRAS: p.G12 R、RBM47:p.495_502AAAAAAAAA>A、AP3S1:p.K41fs、MLL2:p.AEGPHLSPQPEELHLSPQ792del、RFX1:p.386_401GGGGGGGGGGGGGGSG>G、AXDND1:p.EQ991del , HERC2P3:p.A803V, RGPD3:p.N756D, FNDC1:p.D1180del, ANAPC1:p.T537A, IRS4:p.21_22AA>A, GIGYF2:p.Q1005del, NCOA3:p.Q1253fs, SIK3:p.950 _951QQ>Q、GPR6:p.AAAAATAAGGPDTGEWGPPA36del、NBPF12:p.D1323fs、SHROOM4:p.1156_1157EE>E、ZMIZ2:p.VAAAAATATATATAT153del、DGKK:p.PAP P41del, LZTS1: p.RTQDLEGALRTKGLEL432del, CASQ2: p.395_396DD>D, DCP1B: p.251_252insH, ESPNP: p.296_317PPPPSFPPPPPPPGTQLPPPPP>P, KBTBD6 :p.T403K, NBPF16: p.D449fs, ANKRD36C: p.H438R, ESPN: p.PPPPPPSFPPPPPPPGTQLPP430del, FCGBP: p.A2493V, KRAS: p.Q61H, NCOA3: p.Q1276del, OR2 T2: p.C203fs, TMCC1: p.Q565L, BCKDHA: p.G129fs, ESPNP: p.H64fs, GNAS: p.R844H, NBPF14: p.R25C, OGFOD1: p.G477fs, RBM12: p.P693S, SLC38A10: p.1071_1072II>I, SORBS2:p.P866S, TP53:p.R248W, TP53:p.R175H, and UBAC1:p.E269del. .

[0186] 73. The pharmaceutical composition as described in any one of paragraphs 30-72, wherein the composition comprises at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 26, at least 27, at least 28, at least 29, or at least 30 neoantigen peptides.

[0187] 74. The pharmaceutical composition as described in paragraph 73, wherein the composition comprises 15 to 20 neoantigen peptides.

[0188] 75. The pharmaceutical composition as described in paragraph 73 or 74, further comprising at least one additional neoantigen peptide that is specific to the tumor of an individual patient.

[0189] 76. The pharmaceutical composition as described in paragraph 75, wherein a patient-specific neoantigen peptide is selected by identifying sequence differences between the genome, exome, and / or transcriptome of a tumor sample from the patient and the genome, exome, and / or transcriptome of a non-tumor sample.

[0190] 77. The pharmaceutical composition as described in paragraph 75, wherein the samples are fresh or formalin-fixed paraffin-embedded tumor tissue, freshly isolated cells, or circulating tumor cells.

[0191] 78. The pharmaceutical composition as described in paragraph 75, wherein these sequence differences are determined by next-generation sequencing.

[0192] 79. The pharmaceutical composition as described in any one of paragraphs 30-78, wherein the length of each neoantigen peptide is from about 5 to about 50 amino acids.

[0193] 80. The pharmaceutical composition as described in paragraph 79, wherein each neoantigen peptide is of a length between about 15 and about 35 amino acids; of a length of about 15 amino acids or less; of a length of about 8 and about 11 amino acids; or of a length of 9 or 10 amino acids.

[0194] 81. The pharmaceutical composition as described in paragraph 79 or 80, wherein each neoantigen peptide binds to major histocompatibility complex (MHC) class I.

[0195] 82. The pharmaceutical composition as described in any one of paragraphs 30-81, wherein each neoantigen peptide binds to MHC class I with a binding affinity of less than about 500 nM, or optionally each neoantigen peptide binds to MHC class I with a binding affinity of less than 500 nM. D Combined with HLA-A, HLA-B, or HLA-C.

[0196] 83. The pharmaceutical composition as described in paragraph 79, wherein each neoantigen peptide is about 30 amino acids or less in length; between about 6 and about 25 amino acids in length; between about 15 and about 24 amino acids in length; or between about 9 and about 15 amino acids in length.

[0197] 84. The pharmaceutical composition as described in paragraphs 79, 82 or 83, wherein each neoantigen peptide binds to major histocompatibility complex (MHC) class II.

[0198] 85. The pharmaceutical composition as described in paragraph 84, wherein each neoantigen peptide binds to MHC class I with a binding affinity of less than about 500 nM, or optionally each neoantigen peptide binds to MHC class I with a binding affinity of less than 500 nM. D Combined with HLA-A, HLA-B, or HLA-C.

[0199] 86. The pharmaceutical composition as described in any one of paragraphs 30-85, wherein at least one neoantigen peptide further comprises flanking amino acids.

[0200] 87. The pharmaceutical composition as described in paragraph 86, wherein these flanking amino acids are not naturally occurring flanking amino acids.

[0201] 88. The pharmaceutical composition of any one of paragraphs 30-87, wherein at least one neoantigen peptide is linked to at least one second neoantigen peptide.

[0202] 89. The pharmaceutical composition as described in paragraph 88, wherein the peptide is linked using a poly-glycine or poly-serine linker.

[0203] 90. The pharmaceutical composition as described in paragraph 88 or 89, wherein the second neoantigen peptide binds to MHC class I or II with a binding affinity of less than about 1000 nM.

[0204] 91. The pharmaceutical composition as described in any one of paragraphs 88-90, wherein the second neoantigen peptide binds to MHC class I or II with a binding affinity of less than about 500 nM.

[0205] 92. The pharmaceutical composition as described in any one of paragraphs 88-91, wherein both novel epitopes are bound to human leukocyte antigen (HLA)-A,-B,-C,-DP,-DQ, or-DR.

[0206] 93. The pharmaceutical composition of any one of paragraphs 88-92, wherein the isolated neoantigen peptide and the second neoantigen peptide bind to class I HLA, or the isolated neoantigen peptide and the second neoantigen peptide bind to class II HLA.

[0207] 94. The pharmaceutical composition of any one of paragraphs 88-92, wherein the isolated neoantigen peptide binds to class II HLA and the second neoantigen peptide binds to class I HLA, or the isolated neoantigen peptide binds to class I HLA and the second neoantigen peptide binds to class II HLA.

[0208] 95. The pharmaceutical composition of any one of paragraphs 30-94, wherein at least one neoantigen peptide further comprises modifications that increase in vivo half-life, cell targeting, antigen uptake, antigen processing, MHC affinity, MHC stability, or antigen presentation.

[0209] 96. The pharmaceutical composition as described in paragraph 95, wherein the modification is conjugation to a carrier protein, conjugation to a ligand, conjugation to an antibody, PEGylation, polysialylation, HES-enzyme, recombinant PEG mimicry, Fc fusion, albumin fusion, nanoparticle attachment, nanoparticle encapsulation, cholesterol fusion, iron fusion, acylation, amidation, glycosylation, side chain oxidation, phosphorylation, biotinylation, addition of a surfactant, addition of an amino acid mimicry, or addition of a non-natural amino acid.

[0210] 97. The pharmaceutical composition as described in paragraph 95, wherein the targeted cells are antigen-presenting cells.

[0211] 98. The pharmaceutical composition as described in paragraph 97, wherein the antigen-presenting cells are dendritic cells.

[0212] 99. The pharmaceutical composition as described in paragraph 98, wherein these dendritic cells are targeted using DEC205, XCR1, CD197, CD80, CD86, CD123, CD209, CD273, CD283, CD289, CD184, CD85h, CD85j, CD85k, CD85d, CD85g, CD85a, CD141, CD11c, CD83, TSLP receptor, or CD1a marker.

[0213] 100. The pharmaceutical composition as described in paragraph 99, wherein these dendritic cells are targeted using a CD141, DEC205, or XCR1 marker.

[0214] 101. The pharmaceutical composition as described in any one of paragraphs 30-100, wherein it is an immunogenic composition or a vaccine composition.

[0215] 102. The pharmaceutical composition as described in paragraph 101, further comprising an immunomodulator or adjuvant.

[0216] 103. The pharmaceutical composition as described in paragraph 102, wherein the immunomodulator or adjuvant is selected from the group consisting of: poly(I:C), poly-ICLC, STING agonist, 1018ISS, aluminum salt, Amplivax, AS15, BCG, CP-870, 893, CpG7909, CyaA, dSLIM, GM-CSF, IC30, IC31, imiquimod, ImuFact IMP321, IS Patch, ISS, ISCMATRIX, JuvImmune, LipoVac, MF59, monophospholipase A, Montanide IMS1312VG, Montanide ISA 206VG, ​​Montanide ISA 50V2, Montanide ISA 51VG, OK-432, OM-174, OM-197-MP-EC, ISA-TLR2 agonist, ONTAK, Vector systems, PLG microparticles, remiquimod, SRL172, virions and other virus-like particles, YF-17D, VEGF traps, R848, β-glucan, Pam3Cys, Pam3CSK4, acrylic or methacrylic acid polymers, maleic anhydride copolymers, and QS21 stimulon.

[0217] 104. An isolated polynucleotide encoding an isolated neoantigen peptide as described in any one of paragraphs 1-24.

[0218] 105. The isolated polynucleotide as described in paragraph 104 is RNA.

[0219] 106. The isolated polynucleotide as described in paragraph 105, wherein the RNA is modified to increase stability, increase cell targeting, increase translation efficiency, adjuvant activity, cytosol accessibility, and / or reduce cytotoxicity.

[0220] 107. The isolated polynucleotide as described in paragraph 106, wherein the modification is conjugation to a carrier protein, conjugation to a ligand, conjugation to an antibody, codon optimization, increased GC content, incorporation of a modified nucleoside, incorporation of a 5'-cap or cap analogue, and / or incorporation of an unmasked poly-A sequence.

[0221] 108. A cell comprising any of the polynucleotides described in paragraphs 104-107.

[0222] 109. A vector comprising any of the polynucleotides described in paragraphs 104-107.

[0223] 110. The vector as described in paragraph 110, wherein the polynucleotide is operatively linked to a promoter.

[0224] 111. The vector as described in paragraph 109 or 110, wherein the vector is a plasmid, bacteriophage, transposon, spore, virus, or viral particle.

[0225] 112. The vector as described in paragraph 111 is an adeno-associated virus, herpesvirus, lentivirus, or a pseudotype thereof.

[0226] 113. An in vivo delivery system comprising isolated polynucleotides as described in any one of paragraphs 104-107.

[0227] 114. The delivery system as described in paragraph 113, wherein the delivery system comprises spherical nucleic acids, viruses, virus-like particles, plasmids, bacterial plasmids, or nanoparticles.

[0228] 115. A cell comprising a carrier or delivery system as described in any one of paragraphs 109-114.

[0229] 116. The cell described in paragraph 115 is an antigen-presenting cell.

[0230] 117. The cell described in paragraph 116 is a dendritic cell.

[0231] 118. The cell described in paragraph 117 is an immature dendritic cell.

[0232] 119. A composition comprising at least one polynucleotide as described in any one of paragraphs 104-107.

[0233] 120. The composition as described in paragraph 119, wherein the composition comprises at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 26, at least 27, at least 28, at least 29, or at least 30 isolated polynucleotides.

[0234] 121. The composition as described in paragraph 120, wherein the composition comprises between about 2 and about 20 polynucleotides.

[0235] 122. The composition of any one of paragraphs 119-121, wherein the composition further comprises at least 1, at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, at least 8, at least 9, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 26, at least 27, at least 28, at least 29 or at least 30 other neoantigen polynucleotides encoding other neoantigen peptides.

[0236] 123. The composition as described in paragraph 122, wherein the composition comprises between about 4 and about 20 additional neoantigen polynucleotides.

[0237] 124. The composition as described in paragraph 122, wherein these isolated polynucleotides are linked to these additional neoantigen polynucleotides.

[0238] 125. The composition as described in paragraph 124, wherein these polynucleotides are linked using nucleic acids encoding poly-glycine or poly-serine linkers.

[0239] 126. The composition as described in any one of paragraphs 122-125, wherein at least one of the additional neoantigen peptides is specific to the tumor of the individual patient.

[0240] 127. The composition as described in paragraph 126, wherein patient-specific neoantigen peptides are selected by identifying sequence differences between the genome, exome, and / or transcriptome of a tumor sample from the patient and the genome, exome, and / or transcriptome of a non-tumor sample.

[0241] 128. The composition as described in paragraph 127, wherein the samples are fresh or formalin-fixed paraffin-embedded tumor tissue, freshly isolated cells, or circulating tumor cells.

[0242] 129. The composition as described in paragraphs 127 or 128, wherein these sequence differences are determined by next-generation sequencing.

[0243] 130. A T-cell receptor (TCR) capable of binding at least one neoantigen peptide listed in any one of paragraphs 1-27, optionally including FGFR3 S249C, ERBB3 V104M, EGFR L858R, MUC4 H4205Q, PDGFRAR483fs, TMEM52 23_26LLPL>L, or PODXL 28_30PSP>P.

[0244] 131. The TCR as described in paragraph 130, which is capable of binding to isolated neoantigen peptides in a class MHCI or class II background.

[0245] 132. A chimeric antigen receptor comprising: (i) a T cell activating molecule; (ii) a transmembrane region; and (iii) an antigen recognition portion of a separated neoantigen peptide as described in any one of paragraphs 1-27.

[0246] 133. The chimeric antigen receptor as described in paragraph 132, wherein CD3-ζ is a T cell activating molecule.

[0247] 134. The chimeric antigen receptor as described in paragraphs 132 or 133, further comprising at least one co-stimulatory signal transduction domain.

[0248] 135. The chimeric antigen receptor as described in any one of paragraphs 132-134, wherein the signal transduction domain is CD28, 4-1BB, ICOS, OX40, ITAM, or FcεRI-γ.

[0249] 136. The chimeric antigen receptor as described in any one of paragraphs 132-135, wherein the antigen recognition portion is capable of binding to isolated neoantigen peptides in a class MHCI or class II background.

[0250] 137. The chimeric antigen receptor as described in any one of paragraphs 132-136, comprising a transmembrane region of CD3-ζ, CD28, CTLA-4, ICOS, BTLA, KIR, LAG3, CD137, OX40, CD27, CD40L, Tim-3, A2aR, or PD-1.

[0251] 138. The chimeric antigen receptor as described in any one of paragraphs 132-137, wherein the tumor-specific epitope is located in the extracellular domain of a tumor-associated polypeptide, optionally including FGFR3S249C, ERBB3V104M, EGFR L858R, MUC4 H4205Q, PDGFRA R483fs, TMEM5223_26LLPL>L or PODXL 28_30PSP>P.

[0252] 139. A T cell comprising a T cell receptor or chimeric antigen receptor as described in any one of paragraphs 130-138.

[0253] 140. T cells as described in paragraph 139, wherein the T cells are helper T cells or cytotoxic T cells.

[0254] 141. A nucleic acid comprising a promoter operatively linked to a polynucleotide encoding a T-cell receptor as described in paragraphs 130 or 131.

[0255] 142. The nucleic acid as described in paragraph 141, wherein the TCR is capable of binding at least one neoantigen peptide in a major histocompatibility complex (MHC) class I or II background.

[0256] 143. A nucleic acid comprising a promoter operatively linked to a polynucleotide encoding a chimeric antigen receptor as described in any one of paragraphs 132-138.

[0257] 144. The nucleic acid as described in paragraph 143, wherein the antigen recognition portion is capable of binding to at least one neoantigen peptide in a major histocompatibility complex (MHC) class I or II context.

[0258] 145. The nucleic acid as described in paragraph 143 or 144, wherein the tumor-specific epitope is located in the extracellular domain of the tumor-associated polypeptide.

[0259] 146. The nucleic acid as described in any one of paragraphs 143-145, comprising a transmembrane region of CD3-ζ, CD28, CTLA-4, ICOS, BTLA, KIR, LAG3, CD137, OX40, CD27, CD40L, Tim-3, A2aR, or PD-1.

[0260] 147. An antibody capable of binding to at least one neoantigen peptide listed in Tables 1-9.

[0261] 148. A modified cell that is transfected or transduced with a nucleic acid as described in any one of paragraphs 141-146.

[0262] 149. The modified cell as described in paragraph 148, wherein the modified cell is a T cell, a tumor-infiltrating lymphocyte, an NK-T cell, a TCR-expressing cell, a CD4+ T cell, a CD8+ T cell, or an NK cell.

[0263] 150. A composition comprising a T-cell receptor or chimeric antigen receptor as described in any one of paragraphs 130-138.

[0264] 151. A composition comprising patient-derived autologous T cells, wherein the patient-derived autologous T cells contain a T cell receptor or chimeric antigen receptor as described in any one of paragraphs 130-138.

[0265] 152. The composition as described in paragraphs 150 or 151, further comprising an immune checkpoint inhibitor.

[0266] 153. The composition as described in paragraphs 150 or 151, further comprising at least two immune checkpoint inhibitors.

[0267] 154. The composition as described in paragraph 152 or 153, wherein the immune checkpoint inhibitor inhibits checkpoint proteins selected from the group consisting of: CTLA-4, PDL1, PDL2, PD1, B7-H3, B7-H4, BTLA, HVEM, TIM3, GAL9, LAG3, VISTA, KIR, 2B4, CD160, CGEN-15049, CHK1, CHK2, A2aR, and B-7 family ligands, or combinations thereof.

[0268] 155. The composition as described in paragraph 154, wherein the immune checkpoint inhibitor interacts with a ligand selected from the group consisting of: CTLA-4, PDL1, PDL2, PD1, B7-H3, B7-H4, BTLA, HVEM, TIM3, GAL9, LAG3, VISTA, KIR, 2B4, CD160, CGEN-15049, CHK1, CHK2, A2aR, and B-7 family ligands, or combinations thereof.

[0269] 156. The composition as described in any one of paragraphs 119-129 or 150-156, further comprising an immunomodulator or adjuvant.

[0270] 157. The composition as described in paragraph 156, wherein the immunomodulator is a co-stimulatory ligand, a TNF ligand, an Ig superfamily ligand, CD28, CD80, CD86, ICOS, CD40L, OX40, CD27, GITR, CD30, DR3, CD69, or 4-1BB.

[0271] 158. The composition as described in paragraph 156, wherein the immunomodulator is at least one cancer cell or cancer cell extract.

[0272] 159. The composition as described in paragraph 158, wherein the cancer cells are autologous to the subject who requires the composition.

[0273] 160. The composition as described in paragraph 159, wherein the cancer cells have undergone lysis or been exposed to UV radiation.

[0274] 161. The composition as described in paragraph 156, wherein the composition further comprises an adjuvant.

[0275] 162. The composition as described in paragraph 161, wherein the adjuvant is selected from the group consisting of: poly(I:C), poly-ICLC, STING agonist, 1018ISS, aluminum salt, Amplivax, AS15, BCG, CP-870, 893, CpG7909, CyaA, dSLIM, GM-CSF, IC30, IC31, imiquimod, ImuFact IMP321, IS Patch, ISS, ISCMATRIX, JuvImmune, LipoVac, MF59, monophospholipase A, Montanide IMS1312VG, Montanide ISA206VG, ​​Montanide ISA 50V2, Montanide ISA 51VG, OK-432, OM-174, OM-197-MP-EC, ISA-TLR2 agonist, ONTAK, Vector systems, PLG microparticles, remiquimod, SRL172, virions and other virus-like particles, YF-17D, VEGF traps, R848, β-glucan, Pam3Cys, Pam3CSK4, acrylic or methacrylic acid polymers, maleic anhydride copolymers, and QS21 stimulon.

[0276] 163. The composition as described in paragraph 161 or 162, wherein the adjuvant induces a humoral immune response when administered to a subject.

[0277] 164. The composition as described in paragraph 162, wherein the adjuvant induces a type 1 T helper cell response when administered to a subject.

[0278] 165. An in vivo delivery system comprising a pharmaceutical composition as described in any one of paragraphs 30-103.

[0279] 166. The delivery system as described in paragraph 165, wherein the delivery system comprises cell-penetrating peptides, nanoparticle capsules, virus-like particles, or liposomes.

[0280] 167. The delivery system as described in paragraph 166, wherein the cell-penetrating peptide is a TAT peptide, herpes simplex virus VP22, a transportin, or an antp.

[0281] 168. A cell comprising an isolated neoantigen peptide as described in any one of paragraphs 1-29.

[0282] 169. The cell described in paragraph 168 is an antigen-presenting cell.

[0283] 170. The cell described in paragraph 169 is a dendritic cell.

[0284] 171. A method for treating cancer or initiating, enhancing or prolonging an antitumor response in a subject in need, the method comprising administering to the subject a peptide, polynucleotide, carrier, composition, antibody or cell as described in any one of paragraphs 1-164.

[0285] 172. A method for cancer preventive treatment, the method comprising:

[0286] (a) Selecting cancer drugs for patients in need, from the following group, which consists of: ilotinib, erlotinib, imatinib, gefitinib, crizotinib, trastuzumab, vemurafenib, RAF / MEK inhibitors, and anti-estrogen therapy; and

[0287] (b) Prophylactic administration to a subject of a pharmaceutical composition according to any one of paragraphs 30-103, wherein the at least one neoantigen peptide is derived from a resistance mutation associated with a selected cancer drug.

[0288] 173. A method for treating or preventing tumors in a population of subjects in need, the method comprising administering to the subjects an agent comprising an extracellular ligand-binding domain that recognizes a tumor-specific neoepitope comprising a tumor-specific mutation occurring in at least 1% of the population.

[0289] 174. The method as described in any one of paragraphs 171-173, wherein the tumor-specific mutation includes mutations listed for any population in Table 9.

[0290] 175. The method according to any one of paragraphs 171-173, wherein the tumor-specific mutation is located within a gene containing an extracellular domain.

[0291] 176. The method according to paragraph 175, wherein the tumor-specific mutations include FGFR3 S249C, ERBB3V104M, EGFR L858R, MUC4 H4205Q, PDGFRA R483fs, and TMEM52.

[0292] 23_26LLPL>L or PODXL 28_30PSP>P.

[0293] 177. The method described in paragraph 176, wherein the tumor-specific mutation is located within an extracellular domain.

[0294] 178. The method according to paragraph 177, wherein the tumor-specific mutation includes FGFR3 S249C or ERBB3V104M.

[0295] 179. The method as described in any one of paragraphs 171-178, wherein the subject is a human being.

[0296] 180. The method as described in paragraph 179, wherein the subject has cancer.

[0297] 181. The method as described in paragraph 180, wherein the cancer is selected from the group consisting of: cancers of the genitourinary system, gynecological cancers, lung cancer, gastrointestinal cancer, head and neck cancer, malignant glioblastoma, malignant mesothelioma, non-metastatic or metastatic breast cancer, malignant melanoma, Merkel cell carcinoma or bone and soft tissue sarcoma, hematologic malignancies, multiple myeloma, acute myeloid leukemia, chronic myeloid leukemia, myelodysplastic syndrome and acute lymphoblastic leukemia, non-small cell lung cancer (NSCLC), breast cancer, metastatic colorectal cancer, hormone-sensitive or hormone-refractory prostate cancer, colorectal cancer, ovarian cancer, hepatocellular carcinoma, renal cell carcinoma, pancreatic cancer, gastric cancer, esophageal cancer, hepatocellular carcinoma, cholangiocarcinoma, squamous cell carcinoma of the head and neck, soft tissue sarcoma and small cell lung cancer.

[0298] 182. The method as described in any one of paragraphs 171-181, wherein the subject has undergone surgical removal of the tumor.

[0299] 183. The method of any one of paragraphs 171-182, wherein the peptide, polynucleotide, carrier, composition or cell is administered intravenously, intraperitoneally, intratumorally, intradermally or subcutaneously.

[0300] 184. The method as described in paragraph 183, wherein the peptide, polynucleotide, carrier, composition, or cell is administered to an anatomical site that drains into a lymph node basin.

[0301] 185. The method described in paragraph 184, wherein the administration is to multiple lymph nodes in the pelvis.

[0302] 186. The method as described in any one of paragraphs 183-185, wherein administration is carried out via a subcutaneous or intradermal route.

[0303] 187. The method as described in paragraph 183, wherein a peptide is given.

[0304] 188. The method described in paragraph 187, wherein the administration is performed intratumorally.

[0305] 189. The method as described in paragraph 183, wherein a polynucleotide, optionally RNA, is given.

[0306] 190. The method as described in paragraph 189, wherein the polynucleotide is administered intravenously.

[0307] 191. The method described in paragraph 183, wherein the cell is a T cell or a dendritic cell.

[0308] 192. The method as described in paragraph 191, wherein the peptide or polynucleotide includes an antigen-presenting cell-targeting portion.

[0309] 193. The method as described in any one of paragraphs 171-192, further comprising administering at least one immune checkpoint inhibitor to the subject.

[0310] 194. The method as described in paragraph 193, wherein the checkpoint inhibitor is a biotherapeutic agent or a small molecule.

[0311] 195. The method as described in paragraph 193 or 194, wherein the checkpoint inhibitor is selected from the group consisting of monoclonal antibodies, humanized antibodies, fully human antibodies, and fusion proteins or combinations thereof.

[0312] 196. The method of any one of paragraphs 193-195, wherein the checkpoint inhibitor inhibits checkpoint proteins selected from the group consisting of: CTLA-4, PDL1, PDL2, PD1, B7-H3, B7-H4, BTLA, HVEM, TIM3, GAL9, LAG3, VISTA, KIR, 2B4, CD160, CGEN-15049, CHK1, CHK2, A2aR and B-7 family ligands, or combinations thereof.

[0313] 197. The method of any one of paragraphs 193-196, wherein the checkpoint inhibitor interacts with a ligand selected from the group consisting of: CTLA-4, PDL1, PDL2, PD1, B7-H3, B7-H4, BTLA, HVEM, TIM3, GAL9, LAG3, VISTA, KIR, 2B4, CD160, CGEN-15049, CHK1, CHK2, A2aR and B-7 family ligands, or combinations thereof.

[0314] 198. The method as described in any one of paragraphs 193-197, wherein two or more checkpoint inhibitors are administered.

[0315] 199. The method as described in paragraph 198, wherein the checkpoint inhibitor is: (i) ipilimumab or trimemumab, and (ii) nivolumab.

[0316] 200. The method as described in any one of paragraphs 193-199, wherein the checkpoint inhibitor and the composition are administered simultaneously or sequentially in any order.

[0317] 201. The method as described in paragraph 200, wherein the peptide, polynucleotide, carrier, composition, or cell is administered prior to the checkpoint inhibitor.

[0318] 202. The method as described in paragraph 200, wherein the peptide, polynucleotide, carrier, composition, or cell is administered after the checkpoint inhibitor.

[0319] 203. The method as described in paragraph 200, wherein the checkpoint inhibitor is continued to be administered throughout the neoantigen peptide, polynucleotide, carrier, composition, or cell therapy.

[0320] 204. The method of any one of paragraphs 193-203, wherein the neoantigen peptide, polynucleotide, carrier, composition or cell therapy is given to a subject who has only a partial or no response to checkpoint inhibitor therapy.

[0321] 205. The method as described in any one of paragraphs 193-204, wherein the checkpoint inhibitor is administered intravenously or subcutaneously.

[0322] 206. The method as described in paragraph 205, wherein the checkpoint inhibitor is administered subendothelially at approximately 2 cm of the site of administration of the composition.

[0323] 207. The method as described in paragraph 206, wherein the composition is administered to the same draining lymph node as the checkpoint inhibitor.

[0324] 208. The method of any one of paragraphs 171-207, further comprising administering an additional therapeutic agent to the subject before, during, or after treatment with the peptide, polynucleotide, carrier, composition, or cell.

[0325] 209. The method as described in paragraph 208, wherein the additional agent is a chemotherapeutic agent, an immunomodulatory agent, an immunometabolite modifier, a targeted therapy, radiation, an antiangiogenic agent, or an agent that reduces immunosuppression.

[0326] 210. The method as described in paragraph 209, wherein the chemotherapeutic agent is an alkylating agent, a topoisomerase inhibitor, an antimetabolite, or an antimitotic agent.

[0327] 211. The method as described in paragraph 208, wherein the additional agent is an anti-glucocorticoid-induced tumor necrosis factor family receptor (GITR) agonist antibody or antibody fragment, ilotinib, docetaxel, cisplatin, or cyclophosphamide.

[0328] 212. The method as described in any one of paragraphs 171-211, which induces a CD4+ T cell immune response.

[0329] 213. The method as described in any one of paragraphs 171-212, which induces CD4+ T cell immune responses and CD8+ T cell immune responses.

[0330] 214. A method for stimulating an immune response in a subject, the method comprising administering an effective amount of any of the modified cells or compositions described in paragraphs 30-103, 108, 115-129, 139, 140, 148-164, and 168-170.

[0331] 215. The method as described in paragraph 214, wherein the immune response is a cytotoxic and / or humoral immune response.

[0332] 216. The method as described in paragraph 214, wherein the method stimulates a T-cell-mediated immune response in the subject.

[0333] 217. The method as described in paragraph 216, wherein the T cell-mediated immune response is directed against target cells.

[0334] 218. The method described in paragraph 217, wherein the target cell is a tumor cell.

[0335] 219. The method as described in any one of paragraphs 214-218, wherein the modified cells are transfected or transduced in vivo.

[0336] 220. The method as described in any one of paragraphs 214-219, wherein the modified cells are transfected or transduced in vitro.

[0337] 221. The method as described in any one of paragraphs 214-220, wherein the modified cells are the patient's own T cells.

[0338] 222. The method as described in paragraph 221, wherein these patient autologous T cells are obtained from patients who have already received a neoantigen peptide or nucleic acid vaccine.

[0339] 223. The method as described in paragraph 222, wherein the neoantigen peptide or nucleic acid vaccine comprises at least one personalized neoantigen.

[0340] 224. The method as described in paragraph 223, wherein the neoantigen peptide or nucleic acid vaccine includes at least one additional neoantigen peptide listed in Tables 1-9.

[0341] 225. The method as described in paragraph 224, wherein the patient received chemotherapy, immunomodulatory drugs, immunometabolite-modifying drugs, targeted therapy, or radiation before and / or during the administration of the neoantigen peptide or nucleic acid vaccine.

[0342] 226. The method as described in any one of paragraphs 222-225, wherein the patient receives treatment with at least one checkpoint inhibitor.

[0343] 227. The method of any one of paragraphs 222-226, wherein the autologous T cells are obtained from a patient who has received at least one round of T-cell therapy containing a neoantigen.

[0344] 228. The method as described in any one of paragraphs 222-227, wherein the method further comprises adoptive T-cell therapy.

[0345] 229. The method as described in paragraph 228, wherein the adoptive T-cell therapy comprises autologous T cells.

[0346] 230. The method as described in paragraph 229, wherein these autologous T cells target tumor antigens.

[0347] 231. The method as described in paragraphs 228 or 229, wherein the adoptive T-cell therapy further comprises allogeneic T cells.

[0348] 232. The method as described in paragraph 231, wherein these allogeneic T cells target tumor antigens.

[0349] 233. The method as described in any one of paragraphs 227-231, wherein the adoptive T-cell therapy is administered prior to the checkpoint inhibitor.

[0350] 234. A method for evaluating efficacy as described in any one of paragraphs 171-213, the method comprising: (i) measuring the number or concentration of target cells in a first sample obtained from the subject before administering the modified cells; (ii) measuring the number or concentration of target cells in a second sample obtained from the subject after administering the modified cells; and (iii) determining an increase or decrease in the number or concentration of target cells in the second sample compared to the number or concentration of target cells in the first sample.

[0351] 235. The method as described in paragraph 234, wherein therapeutic efficacy is determined by monitoring the following: clinical outcomes; an increase, enhancement, or prolongation of antitumor activity of T cells; an increase in the number of antitumor T cells or activated T cells compared to the pre-treatment number; B cell activity; CD4 T cell activity; or a combination thereof.

[0352] 236. The method described in paragraph 235, wherein therapeutic efficacy is determined by monitoring biomarkers.

[0353] 237. The method as described in paragraph 236, wherein the biomarker is selected from the group consisting of: CEA, Her-2 / neu, bladder tumor antigen, thyroglobulin, alpha-fetoprotein, PSA, CA 125, CA 19.9, CA 15.3, leptin, prolactin, osteopontin, IGF-II, CD98, myofascitis, sPIgR, 14-3-3eta, troponin I, and b-type natriuretic peptide.

[0354] 238. The method as described in paragraph 235, wherein the clinical outcome is selected from the group consisting of: tumor regression; tumor shrinkage; tumor necrosis; antitumor response of the immune system; tumor expansion, recurrence or spread; or a combination thereof.

[0355] 239. The method as described in paragraph 235, wherein the therapeutic effect is predicted by the presence of T cells or the presence of gene markers indicating T cell inflammation, or a combination thereof.

[0356] 240. A kit comprising a neoantigen therapeutic agent as described in any one of paragraphs 1-164.

[0357] Therefore, the object of this invention is to exclude any previously known products, processes for manufacturing such products, or methods of using such products, thereby reserving and hereby disclosing the waiver of any rights to such previously known products, processes, or methods. Furthermore, within the scope of this invention, it is not intended to cover any product, process, or method of manufacturing or using such product that does not meet the written description and implementability requirements of the USPTO (35 U.S. SC § 112, paragraph 1) or the EPO (Section 83 of the EPC), thereby reserving and hereby disclosing the waiver of any previously described product, process for manufacturing such product, or method of using such product. It may be advantageous to comply with Clause 53(c) of the EPC and Regulations 28(b) and (c) of the EPC in the practice of this invention. All rights expressly reserved in respect of any embodiment of the subject matter of any one or more of the applicant's granted patents in any previously filed application that is part of the lineage of this application or any other lineage or any third party shall not be construed as an undertaking.

[0358] It should be noted that in this disclosure, and particularly in the claims and / or paragraphs, terms such as “comprises,” “comprised,” “comprising,” etc., may have the meanings that fall under their jurisdiction in U.S. patent law; for example, they may mean “includes,” “included,” “including,” etc.; and these terms such as “consisting essentially of” and “consists essentially of” have the meanings that fall under their jurisdiction in U.S. patent law, for example, they allow elements not expressly stated but exclude elements found in the prior art or affecting the essential or novel features of the invention. No commitment is intended herein.

[0359] These and other embodiments are disclosed in the following detailed description, or are obvious from thereto and are covered therein. Detailed Implementation

[0360] To aid in understanding this invention, several terms and phrases are defined herein:

[0361] Unless explicitly stated or obvious from the context, as used herein, the term "about" is understood to mean within the normal tolerance range in the field, for example, within 2 standard deviations of the mean. "About" can be understood as within 50%, 45%, 40%, 35%, 30%, 25%, 20%, 15%, 10%, 9%, 8%, 7%, 6%, 5%, 4%, 3%, 2%, 1%, 0.5%, 0.1%, 0.05%, or 0.01% of the declared value. All numerical values ​​provided herein are modified by the term "about" unless otherwise clearly apparent from the context.

[0362] Unless expressly stated or obvious from the context, the term "or" as used herein is to be understood as inclusive. Unless expressly stated or obvious from the context, the terms "a," "an," and "the" as used herein are to be understood as singular or plural.

[0363] All gene name symbols refer to genes generally known in the field. Gene symbols may be those mentioned by the HUGO Genetic Nomenclature Committee (HGNC). Any reference to a gene symbol is a reference to the entire gene or a variant of that gene. The HUGO Genetic Nomenclature Committee is responsible for providing guidelines for human gene nomenclature and approving new unique human gene names and symbols. All human gene names and symbols can be searched at www.genenames.org (the HGNC website), and their formation guidelines can be found there (www.genenames.org / guidelines).

[0364] The term "agent" refers to any small molecule chemical compound, antibody, nucleic acid molecule, or polypeptide or fragment thereof.

[0365] The term "ameliorate" refers to reducing, inhibiting, weakening, diminishing, blocking, or stabilizing the development or progression of a disease (e.g., tumor formation, tumors, etc.).

[0366] The term "change" means a change (increase or decrease) in the expression level or activity of a gene or polypeptide, as detected by methods known by standard techniques (such as those described herein). As used herein, a change includes a 10% change in expression level, preferably a 25% change, more preferably a 40% change, and most preferably a 50% or greater change.

[0367] The term "analog" refers to molecules that are not identical but have similar functions or structural features. For example, tumor-specific neoantigen peptide analogs retain the biological activity of the corresponding naturally occurring tumor-specific neoantigen peptide while possessing certain biochemical modifications that enhance the function of the analog compared to the naturally occurring peptide. Such biochemical modifications can increase its protease resistance, membrane permeability, or half-life without altering, for example, ligand binding. Analogs may include non-natural amino acids.

[0368] "Combination therapy" is intended to include the sequential administration of therapeutic agents (e.g., neoantigen peptides described herein), that is, the administration of each therapeutic agent at different times, and the administration of these therapeutic agents or at least two of these therapeutic agents in a substantially simultaneous manner. Substantially simultaneous administration can be accomplished, for example, by administering to a subject a single capsule of each therapeutic agent in a fixed ratio or by administering multiple single capsules targeting each of these therapeutic agents. For example, one combination of the invention may include pooled samples of neoantigen peptides administered at the same or different times, or they may be formulated as a single co-formulated pharmaceutical composition comprising these peptides. As another example, one combination of the invention (e.g., pooled samples of tumor-specific neoantigens) may be formulated as separate pharmaceutical compositions that can be administered at the same or different times. As used herein, the term "simultaneously" is intended to mean the administration of one or more agents at the same time. For example, in some embodiments, these neoantigen peptides are administered simultaneously. Simultaneously includes administration during the same period (i.e., within the same time period). In some embodiments, the one or more agents are administered simultaneously within the same hour or within the same day. Each therapeutic agent may be administered sequentially or substantially simultaneously via any suitable route, including but not limited to oral, intravenous, subcutaneous, intramuscular, direct absorption through mucosal tissues (e.g., nose, mouth, vagina, and rectum), and ocular routes (e.g., intravitreal, intraocular, etc.). These therapeutic agents may be administered via the same or different routes. For example, one component of a particular combination may be administered intravenously while one or more other components of the combination may be administered orally. These components may be administered in any therapeutically effective order. The phrase "combination" includes groups of compounds or non-pharmacological therapies that are useful as part of a combination therapy.

[0369] The term “neoantigen” or “neoantigenic” refers to a class of tumor antigens that arise from one or more tumor-specific mutations that alter the amino acid sequence of proteins encoded by the genome.

[0370] The term "neoplasia" refers to any disease caused by an inappropriately high level of cell division, an inappropriately low level of apoptosis, or both. Cancer, for example, is an instance of neoplasia. Examples of cancer include, but are not limited to, leukemia (e.g., acute leukemia, acute lymphoblastic leukemia, acute myeloid leukemia, acute myeloblastic leukemia, acute promyelocytic leukemia, acute myelomonocytic leukemia, acute monocytic leukemia, acute erythroleukemia, chronic leukemia, chronic myeloid leukemia, chronic lymphocytic leukemia), polycythemia vera, lymphoma (e.g., Hodgkin's disease, non-Hodgkin's disease), Waldenstrom's macroglobulinemia, heavy chain disease, and solid tumors such as sarcomas and carcinomas (e.g., fibrosarcoma, myxosarcoma, liposarcoma, chondrosarcoma, osteosarcoma, chordoma, angiosarcoma, endothelial sarcoma, lymphangiosarcoma, lymphangioendothelial cell carcinoma). Sarcoma, synovial malformation, mesothelioma, Ewing's tumor, leiomyosarcoma, rhabdomyosarcoma, colon cancer, pancreatic cancer, breast cancer, ovarian cancer, prostate cancer, squamous cell carcinoma, basal cell carcinoma, adenocarcinoma, sweat gland carcinoma, sebaceous gland carcinoma, papillary carcinoma, papillary gland carcinoma, cystadenocarcinoma, medullary carcinoma, bronchial carcinoma, renal cell carcinoma, liver cancer, bile duct carcinoma, choriocarcinoma, seminoma, embryonal carcinoma, renal cell carcinoma, cervical cancer, uterine cancer, testicular cancer, lung cancer, small cell lung cancer, bladder cancer, epithelial carcinoma, glioma, astrocytoma, medulloblastoma, craniopharyngioma, ependymocytoma, pineal tumor, hemangioblastoma, acoustic neuroma, oligodendroglioma, schwannoma, meningioma, melanoma, neuroblastoma, and retinoblastoma. Lymphoproliferative disorders are also considered proliferative diseases.

[0371] In this context, the term "vaccine" means a pooled sample of tumor-specific neoantigen peptides, such as at least two, three, four, five, or more neoantigen peptides. "Vaccine" should be understood to mean a composition used to generate immunity for the prevention and / or treatment of a disease (e.g., tumor formation / tumor). Accordingly, a vaccine is a pharmaceutical agent containing antigens and intended to be used in humans or animals to generate specific defensive and protective substances through vaccination. A "vaccine composition" may include a pharmaceutically acceptable excipient, carrier, or diluent.

[0372] The term "pharmaceutical acceptable" means that a substance approved or permitted for use in animals (including humans) by a regulatory agency of the U.S. federal or state government, or listed in the United States Pharmacopeia or other recognized pharmacopoeia.

[0373] "Pharmaceutical acceptable excipients, carriers or diluents" means excipients, carriers or diluents that can be administered to a subject together with a reagent and that do not impair the pharmacological activity of the reagent and are non-toxic when administered at a dose that effectively delivers a therapeutic amount of the reagent.

[0374] As listed herein, a “pharmaceutically acceptable salt” of a pooled tumor-specific neoantigen can be an acid salt or base salt that is generally considered in the art to be suitable for contact with human or animal tissues without excessive toxicity, irritation, allergic reactions or other problems or complications. Such salts include mineral salts and organic acid salts of basic residues (such as amines) and base or organic salts of acidic residues (such as carboxylic acids). Specific pharmaceutical salts include, but are not limited to, salts of acids, such as hydrochloric acid, phosphoric acid, hydrobromic acid, malic acid, glycolic acid, fumaric acid, sulfuric acid, sulfamic acid, formic acid, toluenesulfonic acid, methanesulfonic acid, benzenesulfonic acid, ethanedisulfonic acid, 2-hydroxyethylsulfonic acid, nitric acid, benzoic acid, 2-acetoxybenzoic acid, citric acid, tartaric acid, lactic acid, stearic acid, salicylic acid, glutamic acid, ascorbic acid, pyric acid, succinic acid, fumaric acid, maleic acid, propionic acid, hydroxymaleic acid, hydroiodic acid, phenylacetic acid, alkanonic acids such as acetic acid, and HOOC-(CH2)n-COOH (where n is 0-4). Similarly, pharmaceutically acceptable cations include, but are not limited to, sodium, potassium, calcium, aluminum, lithium, and ammonium. Those skilled in the art will recognize from this disclosure and the knowledge of the art that other pharmaceutically acceptable salts of the pooled tumor-specific neoantigens provided herein include those listed in Remington's Pharmaceutical Sciences, 17th ed., Mack Publishing Company, Easton, PA, p. 1418 (1985). Generally, pharmaceutically acceptable acid or base salts can be synthesized by any conventional chemical method from a parent compound containing a basic or acidic moiety. In short, such salts can be prepared by reacting the free acidic or basic form of these compounds with a stoichiometric amount of an appropriate base or acid in a suitable solvent.

[0375] The term "polypeptide" or "peptide" refers to a polypeptide that has been isolated from its naturally occurring components. Typically, when the polypeptide comprises at least 60% by weight, it is isolated from its naturally associated proteins and naturally occurring organic molecules. Preferably, the formulation comprises at least 75% by weight, more preferably at least 90%, and most preferably at least 99% polypeptide by weight. The isolated polypeptide can be obtained, for example, by extraction from a natural source, by expression of a recombinant nucleic acid encoding such a polypeptide, or by chemical synthesis of the protein. Purity can be measured by any suitable method, such as column chromatography, polyacrylamide gel electrophoresis, or analysis by HPLC.

[0376] As used herein, the terms “prevent, preventing, prevention”, “prophylactic treatment”, etc., refer to reducing the likelihood that a subject who does not have a disease or condition but is at risk of developing or is susceptible to a disease or condition will develop a disease or condition.

[0377] The terms "primer / booster" or "primer / booster dosage regimen" refer to the continuous administration of a vaccine or immunogenic or immunological composition. A primary immunization (primer) is the administration of a first type of vaccine or immunogenic or immunological composition and may include one, two, or more administrations. A booster immunization is a second administration of a type of vaccine or immunogenic or immunological composition and may include one, two, or more administrations, and may, for example, include or consist substantially of an annual administration. In some embodiments, the administration of a tumorigenesis vaccine or immunogenic composition is performed according to a primary / booster dosage regimen.

[0378] The ranges provided herein are to be understood as abbreviations of all values ​​within that range. For example, the range 1 to 50 is to be understood as including any number, combination of numbers, or subrange from the following group, which consists of: 1, 2, 3, 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, 40, 41, 42, 43, 44, 45, 46, 47, 48, 49, or 50, and all intermediate decimal values ​​between the integers mentioned above, such as 1.1, 1.2, 1.3, 1.4, 1.5, 1.6, 1.7, 1.8, and 1.9. Regarding subranges, “nested subranges” extending from any endpoint of the range are specifically considered. For example, nested subranges of the exemplary range of 1 to 50 may include 1 to 10, 1 to 20, 1 to 30, and 1 to 40 in one direction, or 50 to 40, 50 to 30, 50 to 20, and 50 to 10 in another direction.

[0379] "Receptor" should be understood to mean a biomolecule or group of molecules capable of binding a ligand. Receptors can be used to transmit information within cells, cell formation, or organisms. A receptor comprises at least one receptor unit and often contains two or more receptor units, each of which may be composed of a protein molecule, particularly a glycoprotein molecule. Receptors have a structure complementary to that of a ligand and can complex the ligand into a binding partner. Signal transduction information can be transmitted through conformational changes in the receptor following binding to a ligand on the cell surface. According to the invention, a receptor can refer to specific MHC class I and II proteins capable of forming receptor / ligand complexes with a ligand, particularly peptides or peptide fragments of suitable length.

[0380] The term "subject" refers to an animal used as a subject of treatment, observation, or experimentation. By way of example only, subjects include, but are not limited to, mammals, including, but not limited to, humans or non-human mammals such as non-human primates, cattle, horses, dogs, sheep, or cats.

[0381] The term "treatment" (treat, treated, treating, etc.) means reducing or improving a disorder and / or symptoms associated with it (e.g., tumor formation or tumor). "Treatment" can refer to administering the therapy to a subject after the onset or suspected onset of cancer. "Treatment" includes the concept of "relief," which refers to reducing the frequency or severity of the occurrence or recurrence of any symptoms or other adverse effects related to cancer and / or cancer treatments. The term "treatment" also encompasses the concept of "management," which refers to reducing the severity of a specific disease or disorder in a patient, or delaying its recurrence, such as prolonging remission in a patient with the disease. It should be understood that, although not excluded, treating a disorder or condition does not require the complete elimination of the disorder, condition, or associated symptoms.

[0382] The term "therapeutic effect" refers to the reduction to a certain extent of one or more symptoms of an impairment (e.g., tumor formation or tumor) or its associated lesions. As used herein, "therapeutic effective amount" refers to the amount of agent that, when administered to cells or a subject in a single or multiple doses, is effective in prolonging the survival of a patient with such an impairment, reducing one or more signs or symptoms of the impairment, preventing or delaying, and exceeding the expected outcome without such treatment. "Therapeutic effective amount" is intended to define the amount required to achieve the therapeutic effect. A physician or veterinarian with ordinary skill in the art can readily determine and prescribe the desired "therapeutic effective amount" (e.g., ED50) of a pharmaceutical composition. For example, a physician or veterinarian may begin by administering the composition of the present invention used in a pharmaceutical composition at a level below that required to achieve the desired therapeutic effect and gradually increase the dose until the desired effect is achieved.

[0383] As used with respect to fusion proteins, the terms "spacer region" or "connector" refer to the peptides that link proteins (including fusion proteins). Typically, spacer regions do not have specific biological activity other than being added to or preserving some minimum distance or other spatial relationship between protein or RNA sequences. However, in some embodiments, the constituent amino acids of the spacer region may be selected to affect certain properties of the molecule, such as folding, net charge, or hydrophobicity.

[0384] Suitable linkers for use in embodiments of the present invention are well known to those skilled in the art and include, but are not limited to, straight-chain or branched-chain carbon linkers, heterocyclic carbon linkers, or peptide linkers. The linker is used to separate two neoantigen peptides by a sufficient distance to ensure that, in a preferred embodiment, each neoantigen peptide folds properly. Preferred peptide linker sequences employ a flexible, extended conformation and do not exhibit a tendency to develop ordered secondary structures. Typical amino acids in flexible protein regions include Gly, Asn, and Ser. In fact, any substitution of amino acid sequences containing Gly, Asn, and Ser is expected to satisfy the aforementioned criteria for the linker sequence. Other near-neutral amino acids, such as Thr and Ala, may also be used in the linker sequence. Further amino acid sequences that can be used as linkers are disclosed in Maratea et al. (1985), Gene 40:39-46; Murphy et al. (1986), Proc. Nat'l. Acad. Sci. USA 83:8258-62; U.S. Patent No. 4,935,233; and U.S. Patent No. 4,751,180.

[0385] In any definition of the variables herein, the statement of the list of chemical groups includes the definition of the variable as any single group or a combination of the listed groups. Statements of embodiments of the variables or aspects herein include the embodiment as any single embodiment or a combination with any other embodiment or part thereof.

[0386] Any composition or method provided herein may be combined with one or more other compositions and methods provided herein.

[0387] The therapies disclosed herein constitute novel approaches for treating various types of cancer. The therapies described herein also offer treatment methods that achieve clinical benefits without unacceptable levels of side effects.

[0388] In one aspect, the present invention relates to methods of treating tumorigenesis and, more specifically, tumors by administering to a subject a vaccine or immunogenic composition comprising a plurality of tumor-specific neoantigen peptides. As described in more detail herein, in some embodiments, the composition provides a specifically optimized subset of tumor-specific neoantigens suitable for treating tumors in a high proportion of subjects with cancer. In some embodiments, these tumor-specific neoantigens may together bind to a high total proportion of HLA allotypes present in the subject population.

[0389] The immune system can be divided into two functional subsystems: the innate and the adaptive immune system. The innate immune system is the first line of defense against infection, and most potential pathogens are rapidly neutralized by this system before they can cause, for example, significant infection. The adaptive immune system reacts with molecular structures called antigens that invade an organism. There are two types of adaptive immune responses: humoral immune responses and cell-mediated immune responses. In humoral immune responses, antibodies secreted into the body fluids by B cells bind to pathogen-derived antigens, leading to the elimination of the pathogen through various mechanisms, such as complement-mediated cleavage. In cell-mediated immune responses, T cells capable of destroying other cells are activated. For example, if disease-associated proteins are present in a cell, they are proteasically cleaved into peptides within that cell. Specific cellular proteins then attach themselves to the antigens or peptides formed in this way and transport them to the cell surface, where they are presented to the body's molecular defense mechanisms, particularly T cells. Cytotoxic T cells recognize these antigens and kill cells containing them.

[0390] The molecules that transport and present peptides on the cell surface are called proteins of the major histocompatibility complex (MHC). MHC proteins are divided into two types, called MHC class I and MHC class II. The proteins of the two MHC classes are structurally very similar; however, they have very different functions. MHC class I proteins are present on the surface of almost all cells in the body, including most tumor cells. MHC class I proteins are loaded with antigens, which are usually derived from endogenous proteins or from pathogens present within the cell, and are subsequently presented to naïve or cytotoxic T lymphocytes (CTLs). MHC class II proteins are present on dendritic cells, B lymphocytes, macrophages, and other antigen-presenting cells. They primarily present peptides processed from in vitro antigen sources (i.e., extracellular) to T helper (Th) cells. Most peptides bound by MHC class I proteins originate from cytoplasmic proteins produced in the organism's own healthy host and generally do not stimulate an immune response. Accordingly, cytotoxic T lymphocytes that recognize class I MHC molecules that present their own peptides are absent in the thymus (central tolerance) or are absent or inactivated after release from the thymus, i.e., tolerance (peripheral tolerance). MHC molecules can stimulate an immune response when they present peptides to non-tolerant T lymphocytes. Cytotoxic T lymphocytes possess both T cell receptors (TCRs) and CD8 molecules on their surface. T cell receptors can recognize and bind peptides complexed with class I MHC molecules. Each cytotoxic T lymphocyte expresses a unique T cell receptor that can bind to a specific MHC / peptide complex.

[0391] Before being presented on the cell surface, peptide antigens attach themselves to MHCI-class molecules via competitive affinity binding within the endoplasmic reticulum. Here, the affinity of a single peptide antigen is directly related to its amino acid sequence and the presence of a specific binding motif at a defined position within that amino acid sequence. If the sequence of such a peptide is known, the immune system can be manipulated to fight diseased cells, for example, using peptide vaccines.

[0392] One of the key obstacles to developing curative and tumor-specific immunotherapies is the identification and selection of highly specific and restrictive tumor antigens to avoid autoimmunity. Tumor neoantigens, which arise as a result of genetic changes within malignant cells (e.g., inversions, translocations, deletions, missense mutations, splice site mutations, etc.), represent the most tumor-specific classes of antigens. Neoantigens are rarely used in cancer vaccines or immunogenic compositions due to technical challenges in identifying them, selecting optimized neoantigens, and generating neoantigens for use in vaccines or immunogenic compositions. These challenges can be addressed through the following:

[0393] • Identify tumor-forming / tumor-related mutations present at the tumor DNA level that are not present in matched germline samples from high-proportion cancer subjects;

[0394] • These identified mutations were analyzed using one or more peptide-MHC binding prediction algorithms to generate multiple neoantigen T-cell epitopes that are expressed within the tumor and bind to a high proportion of patient HLA alleles; and

[0395] • Synthesize the multiple neoantigen peptides selected from all neoantigen peptides in these groups and predicted binding peptides for use in cancer vaccines or immunogenic compositions suitable for treating high proportions of cancer subjects.

[0396] For example, converting sequencing information into therapeutic vaccines can include:

[0397] (1) Predicting mutant peptides that can bind to HLA molecules in a high proportion of individuals. Effective selection of which specific mutations to use as immunogens requires the ability to predict which mutant peptides will effectively bind to HLA alleles in a high proportion of patients. Recently, neural network-based learning methods using validated binding and non-binding peptides have improved the accuracy of prediction algorithms for major HLA-A and -B alleles.

[0398] (2) Formulating drugs as multi-epitope vaccines using long peptides. Practically targeting as many mutant epitopes as possible leverages the immense power of the immune system, preventing immune escape through downregulation of specific immune-targeting gene products and compensating for known inaccuracies in epitope prediction methods. Synthetic peptides provide a particularly useful means for the efficient preparation of multiple immunogens and the rapid conversion of mutant epitope identification into effective vaccines. Peptides can be readily synthesized chemically and easily purified using reagents free from contaminating bacteria or animal matter. Small size allows for precise targeting of mutant regions of the protein and also reduces competition from irrelevant antigens from other components (unmutated proteins or viral vector antigens).

[0399] (3) Combination with strong vaccine adjuvants. Effective vaccines require strong adjuvants to initiate an immune response. As described below, poly-ICLCs have shown several promising properties as vaccine adjuvants, acting as agonists of the TLR3 and RNA helicase-MDA5 and RIG3 domains. These properties include inducing local and systemic activation of immune cells in vivo, producing stimulating chemokines and cytokines, and stimulating antigen presentation via dendritic cells. Furthermore, poly-ICLCs can induce durable CD4+ and CD8+ responses in humans. Importantly, striking similarities were observed in the upregulation of transcriptional and signal transduction pathways in subjects vaccinated with poly-ICLCs and in volunteers who had received a highly effective, replicating yellow fever vaccine. In addition, in a recent phase 1 study, >90% of ovarian cancer patients immunized with poly-ICLCs in combination with NYES0-1 peptide vaccines (excluding Montaned) showed induction of CD4+ and CD8+ T cells, as well as antibody responses to the peptide. Meanwhile, poly-ICLC has been extensively tested in more than 25 clinical trials to date and has shown relatively benign toxicity characteristics.

[0400] The advantages of the present invention are further described herein.

[0401] As described here, there is substantial evidence in both animals and humans that mutated epitopes are effective in inducing immune responses, and cases of spontaneous tumor regression or long-term survival are associated with CD8+ T cell responses against mutated epitopes (Buckwalter and Srivastava PK. "It is the antigen(s), stupid" and other lessons from over a decade of vaccitherapy of human cancer). Seminars in Immunology 20:296-300 (2008); Karanikas et al., High frequency of cytolytic T lymphocytes directed against a tumor-specific mutated antigen detectable with HLA tetramers in the blood of a lung carcinoma patient with long survival. Cancer Res. [Cancer Research] 61:3718-3724 (2001); Lennerz et al., The response of autologous T cells to a human melanoma is dominated by mutated neoantigens. Proc Natl Acad Sci US A.[Proceedings of the National Academy of Sciences of the United States of America] 102:16013 (2005) and “immunoediting” can be traced back to changes in the expression of dominant mutant antigens in mice and humans (Matsushita et al., Cancer exome analysis reveals a T-cell-dependent mechanism of cancer immunoediting, Nature 482:400 (2012); DuPage et al., Expression of tumor-specific antigens underlies cancer immunoediting, Nature 482:405 (2012); and Sampson et al., Immunologic escape after prolonged progression-free survival with epidermal growth factor receptor variant III peptide vaccination in patients with newly diagnosed [Immune escape from epidermal growth factor receptor variant III peptide vaccination in newly diagnosed glioblastoma patients after prolonged progression-free survival] J Clin Oncol. [Journal of Clinical Oncology] 28:4722-4729 (2010)

[0402] Sequencing technology has revealed that each tumor contains multiple patient-specific mutations that alter the protein-coding content of genes. These mutations produce altered proteins ranging from single-amino acid changes (caused by missense mutations) to novel amino acid sequences that add long regions (novel open reading frame mutations; new ORFs) attributable to frameshifts, read linking, or translation of intron regions of stop codons. These mutant proteins are valuable targets for the host's immune response to the tumor because, unlike natural proteins, they are not affected by the host's own immunosuppressive effects. Therefore, mutant proteins are more likely to be immunogenic and more specific to tumor cells than to the patient's normal cells.

[0403] In one embodiment, the neoantigen peptides in the composition together have affinity for multiple MHC molecules, for example, they together cover a large proportion of the target population. Effective selection of which specific mutations to utilize as immunogens requires the ability to predict which mutant peptides will effectively bind to HLA alleles present in the patient population. Recently, neural network-based learning methods using validated binding and non-binding peptides have improved the accuracy of prediction algorithms targeting major HLA-A and -B alleles. Using recently improved algorithms for predicting which missense mutations produce strong-binding peptides to homologous MHC molecules, a set of peptides representing the best mutant epitopes (both novel ORFs and mistranslations) of a patient population can be identified and prioritized (Zhang et al., Machine learning competition in immunology – Prediction of HLA class I binding peptides, J Immunol Methods 374:1 (2011); Lundegaard et al., Prediction of epitopes using neural network based methods, J Immunol Methods 374:26 (2011)).

[0404] Practically targeting as many mutant epitopes as possible leverages the immense power of the immune system, preventing immune escape through the downregulation of specific immune-targeting gene products and compensating for known inaccuracies in epitope prediction methods. Synthetic peptides offer a particularly useful means of efficiently preparing multiple immunogens and rapidly converting the identification of mutant epitopes into effective vaccines or immunogenic compositions. Peptides can be readily synthesized chemically and easily purified using reagents free from contaminating bacteria or animal matter. Their small size allows for precise targeting of mutant regions of the protein and also reduces competition from irrelevant antigens from other components (unmutated proteins or viral vector antigens).

[0405] In one embodiment, the pharmaceutical formulation is a long peptide multi-epitope vaccine or immunogenic composition. Such “long” peptides undergo efficient internalization, processing, and cross-presentation in professional antigen-presenting cells (such as dendritic cells) and have been shown to induce CTLs in humans (Melief and van der Burg, Immunotherapy of established (pre)malignant disease by synthetic long peptide vaccines, Nature Rev Cancer 8:351 (2008)). In one embodiment, at least two peptides are prepared for immunization. In some embodiments, 20 or more peptides are prepared for immunization. In one embodiment, the neoantigen peptide is of length ranging from about 5 to about 50 amino acids. In another embodiment, a peptide of length ranging from about 15 to about 35 amino acids is synthesized. In a preferred embodiment, the neoantigen peptide is of length ranging from about 20 to about 35 amino acids.

[0406] Production of tumor-specific neoantigens

[0407] This invention is based, at least in part, on the ability to provide a pool of tumor-specific neoantigens to a patient's immune system. Those skilled in the art will recognize from this disclosure and the knowledge of the art that there are various ways to generate such tumor-specific neoantigens. Typically, such tumor-specific neoantigens can be generated in vitro or in vivo. Tumor-specific neoantigens can be generated in vitro as peptides or polypeptides, which can then be formulated into tumor-forming vaccines or immunogenic compositions and administered to a subject. As further detailed herein, such in vitro generation can occur by a variety of methods known to those skilled in the art, such as, for example, synthesizing peptides or expressing peptides / polypeptides from DNA or RNA molecules in any of a variety of bacterial, eukaryotic, or viral recombinant expression systems, followed by purification of the expressed peptides / polypeptides. Alternatively, tumor-specific neoantigens can be generated in vivo by introducing a molecule encoding a tumor-specific neoantigen (e.g., DNA, RNA, viral expression systems, etc.) into a subject, in which the encoded tumor-specific neoantigen is expressed. Furthermore, methods for the production of neoantigens in vitro and in vivo are described herein in relation to pharmaceutical compositions and methods of delivery of the therapy.

[0408] In some embodiments, the present invention includes modified neoantigen peptides. As used herein, the terms “modified,” “modified,” etc., with respect to neoantigen peptides refer to one or more changes that enhance the desired properties of the neoantigen peptide, wherein such changes do not alter the primary amino acid sequence of the neoantigen peptide. “Modification” includes covalent chemical modifications that do not alter the primary amino acid sequence of the neoantigen peptide itself. Such desired properties include, for example, prolonged in vivo half-life, increased stability, reduced clearance, altered immunogenicity or allergenicity, and enhanced specific antibody, cell targeting, antigen uptake, antigen processing, MHC affinity, MHC stability, or antigen presentation. Possible modifications to the neoantigen peptide include, but are not limited to, conjugation with carrier proteins, conjugation with ligands, conjugation with antibodies, PEGylation, polysialylation-HES, recombinant PEG mimicry, Fc fusion, albumin fusion, nanoparticle attachment, nanoparticle encapsulation, cholesterol fusion, iron fusion, acylation, amidation, glycosylation, side-chain oxidation, phosphorylation, biotinylation, addition of surfactants, addition of amino acid mimicry, or addition of non-natural amino acids.

[0409] The clinical efficacy of protein therapeutics is often limited by short plasma half-lives and susceptibility to protease degradation. Studies of various therapeutic proteins (e.g., filgrastim) have shown that these difficulties can be overcome through various modifications, including conjugating or linking polypeptide sequences to any of a variety of non-protein polymers such as polyethylene glycol (PEG), polypropylene glycol, or polyoxyethylene (see, for example, typically via a linker covalently bound to the protein and the non-protein polymer (e.g., PEG)). Such PEG-conjugated biomolecules have shown clinically useful properties, including better physical and thermal stability, protection against susceptibility to enzymatic degradation, increased solubility, longer in vivo circulating half-life and reduced clearance, reduced immunogenicity and antigenicity, and reduced toxicity.

[0410] PEGs suitable for conjugating peptide sequences are generally soluble in water at room temperature and have the general formula R(0-CH2-CH2). n OR, where R is hydrogen or a protecting group such as an alkyl or alkanol group, and where n is an integer from 1 to 1000. When R is a protecting group, it typically has from 1 to 8 carbons. PEG conjugated to a polypeptide sequence can be linear or branched. This disclosure covers branched PEG derivatives, “star-PEG”, and multi-arm PEG. The molecular weight of PEG used in this disclosure is not limited to any particular range, but some examples have molecular weights between 500 and 20,000, while other examples have molecular weights between 4,000 and 10,000.

[0411] This disclosure also covers compositions of conjugates in which PEG has different n values ​​and therefore various different PEGs are present in specific proportions. For example, some compositions comprise mixtures of conjugates in which n = 1, 2, 3, and 4. In some compositions, the percentage of the conjugate in which n = 1 is 18%-25%, the percentage of the conjugate in which n = 2 is 50%-66%, the percentage of the conjugate in which n = 3 is 12%-16%, and the percentage of the conjugate in which n = 4 is up to 5%. Such compositions can be produced by reaction conditions and purification methods known in the art. For example, cation exchange chromatography can be used to separate the conjugates and then identify the moiety containing the conjugate, for example having the desired number of attached PEGs, and purified from unmodified protein sequences (purified free form) and from conjugates having other numbers of attached PEGs.

[0412] PEG can be bound to the disclosed peptides via terminal reactive groups (“spacer regions”). A spacer region is, for example, a terminal reactive group that mediates a bond between a free amino or carboxyl group of one or more peptide sequences and polyethylene glycol. PEGs having spacer regions that can bind to free amino groups include N-hydroxysuccinimide polyethylene glycol, which can be prepared by activating a succinate of polyethylene glycol with N-hydroxysuccinimide. Another activated polyethylene glycol that can bind to free amino groups is 2,4-bis(O-methoxy polyethylene glycol)-6-chloro-s-triazine, which can be prepared by reacting polyethylene glycol monomethyl ether with cyanuric chloride. Activated polyethylene glycols that bind to free carboxyl groups include polyethylene oxide diamine.

[0413] The conjugation of one or more polypeptide sequences disclosed herein with PEG having spacer regions can be carried out by a variety of conventional methods. For example, the conjugation reaction can be carried out in solution with a protein in a molar ratio of 4:1 to 30:1, at a pH of 5 to 10, and at a temperature from 4°C to room temperature for 30 minutes to 20 hours. Reaction conditions can be selected to guide the reaction to produce the desired degree of substitution. Generally, low temperatures, low pH (e.g., pH = 5), and short reaction times tend to reduce the number of PEGs attached, while high temperatures, medium to high pH (e.g., pH > 7), and longer reaction times tend to increase the number of PEGs attached. Various methods known in the art can be used to terminate the reaction. In some embodiments, the reaction is terminated by acidifying the reaction mixture and freezing it at, for example, -20°C.

[0414] This disclosure also covers the use of PEG mimics. Recombinant PEG mimics have been developed that retain the properties of PEG (e.g., enhanced serum half-life) while conferring several additional advantageous properties. By way of example, simple polypeptide chains capable of forming extended conformations similar to PEG (including, for example, Ala, Glu, Gly, Pro, Ser, and Thr) can be recombinantly generated and fused with peptide or protein drugs of interest (e.g., Amunix's XTEN technology; Mountain View, CA). This eliminates the need for additional conjugation steps in the manufacturing process. Furthermore, established molecular biology techniques enable control over the side-chain composition of the polypeptide chains, thereby allowing for optimization of immunogenicity and manufacturing properties.

[0415] For the purposes of this disclosure, "glycosylation" is intended to broadly refer to the enzymatic process of attaching glycans to proteins, lipids, or other organic molecules. When used in conjunction with this disclosure, the term "glycosylation" generally refers to the addition or deletion of one or more carbohydrate moieties (by removing potential glycosylation sites or by deleting glycosylation using chemical and / or enzymatic means), and / or the addition of one or more glycosylation sites that may or may not be present in the natural sequence. Additionally, the phrase includes qualitative changes in the glycosylation of native proteins, involving variations in the nature and proportions of the present carbohydrate moieties. Glycosylation can significantly affect the physical properties of proteins and is also important in protein stability, secretion, and subcellular localization. Proper glycosylation can be crucial for biological activity. In fact, some genes from eukaryotes, when expressed in bacteria (e.g., *E. coli*) lacking glycosylated proteins in cellular processes, produce proteins that are little or no activity due to the lack of glycosylation.

[0416] The addition of glycosylation sites can be accomplished by altering the amino acid sequence. For example, the peptide can be modified by adding or substituting one or more serine or threonine residues (for O-linked glycosylation sites) or asparagine residues (for N-linked glycosylation sites). The structures of N-linked and O-linked oligosaccharides and the sugar residues found in each type may differ. One type of sugar commonly found on both is N-acetylneuraminic acid (hereinafter referred to as sialic acid). Sialic acid is often a terminal residue of both N-linked and O-linked oligosaccharides and, due to its negative charge, can impart acidic properties to glycoproteins. Specific embodiments disclosed herein include the generation and use of N-glycosylated variants.

[0417] The disclosed polypeptide sequence can optionally be altered by changes at the DNA level, particularly by mutating the DNA encoding the polypeptide at a preselected base, thus generating a codon that will be translated into the desired amino acid. Another means of increasing the number of carbohydrate moieties on the polypeptide is through chemical or enzymatic coupling with a glycoside.

[0418] Carbohydrate removal can be accomplished chemically or enzymatically, or by substituting codons encoding glycosylated amino acid residues. Chemical deglycosylation is known and can be achieved by using various endoglucosidases and exoglucosidases to cleave carbohydrate moieties from peptides.

[0419] Dihydrofolate reductase (DHFR) deficient Chinese hamster ovary (CHO) cells are commonly used host cells for the production of recombinant glycoproteins. These cells do not express the enzyme β-galactosidase α-2,6-sialylate transferase, and therefore do not add sialic acid to the α-2,6 linker of the N-linked oligosaccharide in the glycoproteins produced in these cells.

[0420] To improve their stability and in vivo pharmacokinetics, this disclosure also covers the use of polysialylation, the conjugation of peptides and proteins to naturally occurring, biodegradable α-(2→8)-linked polysialic acid (“PSA”). PSA is a biodegradable, non-toxic natural polymer that is highly hydrophilic, giving it a high apparent molecular weight in the blood, which increases its serum half-life. In addition, polysialylation of a range of peptide and protein therapeutics has resulted in significant reductions in proteolysis, retention of in vivo activity, and decreased immunogenicity and antigenicity (see, for example, G. Gregoriadis et al., Int. J. Pharmaceutics [International Journal of Pharmacy] 300(1-2):125-30). As with modifications using other conjugates (e.g., PEG), various techniques for site-specific polysialylation are available (see, for example, T. Lindhout et al., PNAS 108(18)7397-7402(2011)).

[0421] Other suitable components and molecules for conjugation include, for example, thyroglobulin; albumins such as human serum albumin (HAS); tetanus toxoid; diphtheria toxoid; polyamino acids such as poly(D-lysine:D-glutamic acid); rotavirus VP6 polypeptide; influenza virus hemagglutinin, influenza virus nucleoprotein; keyfora hemocyanin (KLH); and hepatitis B virus core protein and surface antigen; or any combination of the foregoing.

[0422] The fusion of albumin with one or more polypeptides disclosed herein can be achieved, for example, through genetic manipulation, such that DNA encoding HSA or a fragment thereof is linked to DNA encoding the one or more polypeptide sequences. Subsequently, a suitable host can be transformed or transfected with the fused nucleotide sequence, for example in the form of a suitable plasmid, to express the fusion polypeptide. Expression can be achieved in vitro from, for example, prokaryotic or eukaryotic cells, or in vivo from, for example, transgenic organisms. In some embodiments of this disclosure, the expression of the fusion protein is performed in mammalian cell lines such as CHO cell lines. Transformation is used broadly herein to refer to cellular genetic alterations resulting from the direct uptake, incorporation, and expression of exogenous genetic material (exogenous DNA) from its surroundings and its uptake through one or more cell membranes. Transformation is naturally occurring in some bacterial species, but can also be achieved artificially in other cells.

[0423] Furthermore, albumin itself can be modified to prolong its cycling half-life. The fusion of modified albumin with one or more polypeptides can be obtained through the aforementioned genetic manipulation techniques or through chemical conjugation; the resulting fusion molecule has a longer half-life than the fusion with unmodified albumin. (See WO)

[0424] 2011 / 051489).

[0425] Several albumin-binding strategies have been developed as alternatives to direct fusion, including albumin binding via conjugation of fatty acid chains (acylation). Since serum albumin is a fatty acid transporter, these natural ligands with albumin-binding activity have been used to extend the half-life of small protein therapeutics. For example, insulin detemir (LEVEMIR)—an approved product for diabetes—involves a nutmeg chain conjugated to genetically modified insulin, producing a long-acting insulin analog.

[0426] Another type of modification is the conjugation (e.g., linking) of one or more other components or molecules, such as another protein (e.g., a protein having an amino acid sequence heterologous to the subject protein) or a carrier molecule, to the N- and / or C-terminus of the polypeptide sequence. Thus, exemplary polypeptide sequences can be provided as conjugates with another component or molecule.

[0427] Conjugation modification can result in the retention of an active peptide sequence that possesses additional or complementary functions or activities in a second molecule. For example, the peptide sequence can be conjugated to a molecule to improve solubility, storage, in vivo or preservation half-life or stability, reduce immunogenicity, delay or control release in vivo, etc. Other functions or activities include reduced toxicity of the conjugation relative to the unconjugated peptide sequence, more effective targeting of a type of cell or organ than the unconjugated peptide sequence, or further resistance of the drug to causes or effects associated with the disorder or disease described herein (e.g., diabetes).

[0428] Peptides can also be conjugated to large, slowly metabolizing macromolecules, such as proteins; polysaccharides, such as cross-linked agarose, agarose, cellulose, cellulose beads; polymeric amino acids, such as polyglutamic acid, polylysine; amino acid copolymers; inactivated viral particles; inactivated bacterial toxins, such as toxoids derived from diphtheria, tetanus, cholera, and leukocyte toxin molecules; inactivated bacteria; and dendritic cells.

[0429] Other candidate components and molecules for conjugation include those suitable for isolation or purification. Specific, non-limiting examples include conjugating molecules such as biotin (a biotin-avidin specific binding pair), antibodies, receptors, ligands, lectins, or molecules comprising a solid support, such as plastic or polystyrene beads, plates or beads, magnetic beads, test strips, and membranes.

[0430] Purification methods such as cation exchange chromatography can be used to separate conjugates by charge difference, effectively separating the conjugates into their individual molecular weights. For example, a cation exchange column can be loaded and washed with -20 mM sodium acetate (pH-4), followed by elution with a linear (0 M to 0.5 M) NaCl gradient (in a pH buffer from about 3 to 5.5, e.g., pH-4.5). The fractions obtained by cation exchange chromatography can be identified by molecular weight using conventional methods, such as mass spectrometry, SDS-PAGE, or other known methods for separating molecular entities by molecular weight.

[0431] In some embodiments, the amino or carboxyl terminus of the disclosed polypeptide sequence may be fused to an immunoglobulin Fc region (e.g., human Fc) to form a fusion conjugate (or fusion molecule). Fc fusion conjugates have been shown to increase the systemic half-life of the biologic, and therefore the biologic product may not need to be administered as frequently.

[0432] Fc binds to the neonatal Fc receptor (FcRn) in the endothelial cells lining blood vessels, and upon binding, the Fc fusion molecule is protected from degradation and re-released into circulation, thus extending molecular circulation. This Fc binding is considered the mechanism underlying the long plasma half-life of endogenous IgG. Compared to traditional Fc-fusion conjugates, recent Fc-fusion technologies link a single copy of a biologic to the Fc region of an antibody to optimize the pharmacokinetic and pharmacodynamic properties of the biologic.

[0433] This disclosure covers the use of other modifications, currently known or to be developed in the future, to improve one or more properties of the polypeptide. One such method for extending the cyclic half-life of the polypeptide disclosed herein, increasing stability, reducing clearance, or altering immunogenicity or allergenicity involves modifying the polypeptide sequence via hesylation, which utilizes a hydroxyethyl starch derivative linked to other molecules to modify the molecule's characteristics. Aspects of hesylation are described, for example, in U.S. Patent Applications Nos. 2007 / 0134197 and 2006 / 0258607.

[0434] In vitro peptide / peptide synthesis

[0435] Proteins or peptides can be prepared using any technique known to those skilled in the art, including expressing proteins, polypeptides, or peptides using standard molecular biotechnology, isolating proteins or peptides from natural sources, through in vitro translation, or through chemical synthesis. Nucleotide and protein, polypeptide, and peptide sequences corresponding to various genes have previously been disclosed and can be found in computerized databases known to those skilled in the art. One such database is the Genbank and GenPept databases of the National Center for Biotechnology Information, located on the website of the National Institutes of Health. The coding regions of known genes can be amplified and / or expressed using techniques disclosed herein or that should be known to those skilled in the art. Alternatively, various commercial formulations of proteins, polypeptides, and peptides are known to those skilled in the art.

[0436] Peptides can be readily synthesized chemically using reagents free from contaminating bacteria or animal matter (Merrifield RB: Solid phase peptide synthesis. I. The synthesis of atetrapeptide. J. Am. Chem. Soc. 85: 2149-54, 1963). In some embodiments, neoantigen peptides are prepared by the following steps: (1) parallel solid-phase synthesis on a multichannel instrument using uniform synthesis and cleavage conditions; (2) purification on an RP-HPLC column using column stripping; and rewashing, but without substitution between peptides; followed by (3) analysis using a limited set of assays with the highest information content. A standard manufacturing practice (GMP) footprint can be defined around a set of peptides for individual patients, thus requiring only a set of conversion procedures between the synthesis of peptides for different patients.

[0437] Alternatively, a nucleic acid (e.g., a polynucleotide) encoding the neoantigenic peptide of the present invention can be used to produce the neoantigenic peptide in vitro. The polynucleotide can be, for example, single-stranded and / or double-stranded DNA, cDNA, PNA, CNA, RNA, or a natural or stable form of a polynucleotide (such as a polynucleotide having a phosphate-thiocyanate backbone) or a combination thereof, and it may or may not contain introns, as long as it encodes the peptide. In one embodiment, in vitro translation is used to produce the peptide. Many exemplary systems exist that can be utilized by those skilled in the art (e.g., the Retic Lysis IVT Kit, Life Technologies, Waltham, MA).

[0438] Expression vectors capable of expressing peptides have also been prepared. Expression vectors for different cell types are well known in the art and can be selected without excessive experimentation. Typically, DNA is inserted into an expression vector (such as a plasmid) in the appropriate orientation and within the correct reading frame for expression. If necessary, the DNA can be ligated to an appropriate transcriptional and translational regulatory nucleotide sequence that is recognized by the desired host (e.g., bacteria), although such controls are usually available in the expression vector. The vector is then introduced into the host bacteria for cloning using standard techniques (see, for example, Sambrook et al. (1989) Molecular Cloning, A Laboratory Manual, Cold Spring Harbor Laboratory, Cold Spring Harbor, NY).

[0439] Expression vectors containing isolated polynucleotides, along with host cells containing the expression vectors, are also considered. Neoantigen peptides can be provided in the form of RNA or cDNA molecules encoding the desired neoantigen peptide. One or more neoantigen peptides of the present invention can be encoded by a single expression vector.

[0440] The term "polynucleotide encoding a polypeptide" encompasses polynucleotides that include only the coding sequence for the polypeptide, as well as polynucleotides that include additional coding and / or non-coding sequences. Polynucleotides can be in RNA or DNA form. DNA includes cDNA, genomic DNA, and synthetic DNA; and can be double-stranded or single-stranded, and if single-stranded, can be a coding strand or a non-coding (antisense) strand.

[0441] In embodiments, these polynucleotides may include coding sequences for tumor-specific neoantigen peptides fused to a polynucleotide within the same reading frame, such as an aid to the expression and / or secretion of a polypeptide by a host cell (e.g., acting as a leader sequence for controlling the transport of the polypeptide from the cell). The polypeptide having the leader sequence is a pre-protein and the leader sequence can be cleaved by the host cell to form the mature form of the polypeptide.

[0442] In embodiments, these polynucleotides may include a coding sequence for a tumor-specific neoantigen peptide fused to a marker sequence within the same reading frame. This marker sequence, for example, allows for the purification of the encoded polypeptide, which can then be incorporated into a personalized tumorigenesis vaccine or immunogenic composition. For instance, in the case of a bacterial host, the marker sequence may be a hexahistine tag provided by the pQE-9 vector to prepare for the purification of the mature polypeptide fused to the marker, or when using a mammalian host (e.g., COS-7 cells), the marker sequence may be a hemagglutinin (HA) tag derived from the influenza hemagglutinin protein. Other tags include, but are not limited to, calmodulin tag, FLAG tag, Myc tag, S tag, SBP tag, Softag 1, Softag 3, V5 tag, Xpress tag, Isopeptag, SpyTag biotin carboxyl carrier protein (BCCP) tag, GST tag, fluorescent protein tag (e.g., green fluorescent protein tag), maltose-binding protein tag, Nus tag, Strep- tag, thioredoxin tag, TC tag, Ty tag, etc.

[0443] In embodiments, these polynucleotides may include coding sequences for one or more of these tumor-specific neoantigen peptides, the coding sequences being concatenated in the same reading frame to produce a single concatamerized neoantigen peptide construct capable of generating multiple neoantigen peptides.

[0444] In some embodiments, isolated nucleic acid molecules may be provided having a nucleotide sequence that is at least 60%, at least 65%, at least 70%, at least 75%, at least 80%, at least 85%, at least 90%, at least 95%, or at least 96%, 97%, 98%, or 99% identical to a polynucleotide encoding the tumor-specific neoantigen peptide of the present invention.

[0445] A polynucleotide having at least, for example, 95% "identical" nucleotide sequence to a reference nucleotide sequence means that the polynucleotide's nucleotide sequence is identical to the reference sequence, except that the polynucleotide sequence may include up to five point mutations per 100 nucleotides of the reference nucleotide sequence. In other words, to obtain a polynucleotide having at least 95% identical nucleotide sequence to a reference nucleotide sequence, up to 5% of the nucleotides in the reference sequence may be deleted or substituted with another nucleotide, or up to 5% of the total nucleotides in the reference sequence may be inserted into the reference sequence. These mutations in the reference sequence may occur at the amino-terminal or carboxyl-terminal position of the reference nucleotide sequence, or at any position between those positions, and they may be scattered individually among the nucleotides in the reference sequence or scattered in one or more consecutive groups within the reference sequence.

[0446] As a practical problem, known computer programs can be used to routinely determine whether any specific nucleic acid molecule is at least 80%, at least 85%, at least 90%, and in some embodiments at least 95%, 96%, 97%, 98%, or 99% identical to a reference sequence. These computer programs include Bestfit (Wisconsin Sequence Analysis Package, Unix-based version 8, Genetics Computer Group, University Research Park, 575 Science Drive, Madison, Wisconsin 53711). Bestfit uses a local homology algorithm (Smith and Waterman, Advances in Applied Mathematics 2:482-489 (1981)) to find the optimal homology region between two sequences. When using Bestfit or any other sequence alignment program to determine whether a specific sequence is 95% identical to a reference sequence according to the invention, these parameters are set such that the percentage of similarity is calculated over the full length of the reference nucleotide sequence and allows for homology vacancies of up to 5% of the total number of nucleotides in the reference sequence.

[0447] The isolated tumor-specific neoantigen peptides described herein can be generated in vitro (e.g., in a laboratory) by any suitable method known in the art. Such methods range from direct protein synthesis methods to constructing DNA sequences encoding isolated polypeptide sequences and expressing those sequences in a suitable transforming host. In some embodiments, the DNA sequence is constructed by isolating or synthesizing a DNA sequence encoding a wild-type protein of interest using recombinant techniques. Optionally, the sequence can be mutagenized by site-specific mutagenesis to provide its functional analogue. See, for example, Zoeller et al., Proc. Nat'l. Acad. Sci. USA [Proceedings of the National Academy of Sciences] 81:5662-5066 (1984) and U.S. Patent No. 4,588,585.

[0448] In this embodiment, an oligonucleotide synthesizer is used to construct DNA sequences encoding polypeptides of interest through chemical synthesis. Such oligonucleotides can be designed based on the amino acid sequence of the desired polypeptide and by selecting codons preferred in the host cell that produces the recombinant polypeptide of interest. Standard methods can be used to synthesize isolated polynucleotide sequences encoding isolated polypeptides of interest. For example, complete amino acid sequences can be used to construct back-translated genes. Furthermore, DNA oligomers containing nucleotide sequences encoding specific isolated polypeptides can be synthesized. For example, several small oligonucleotides encoding portions of the desired polypeptide can be synthesized and then ligated together. Individual oligonucleotides typically contain 5' or 3' overhangs for complementary components.

[0449] Once assembled (e.g., by synthesis, site-directed mutagenesis, or another method), the polynucleotide sequence encoding the specific isolated polypeptide of interest is inserted into the expression vector and optionally operably ligated to an expression control sequence adapted to express the protein in a desired host. Proper assembly can be confirmed by nucleotide sequencing, restriction enzyme mapping, and expression of the bioactive polypeptide in a suitable host. As is well known in the art, to obtain high expression levels of the transfected gene in a host, the gene can be operably ligated to transcriptional and translational expression control sequences that are functional in the selected expression host.

[0450] Recombinant expression vectors can be used to amplify and express DNA encoding tumor-specific neoantigen peptides. Recombinant expression vectors are reproducible DNA constructs containing synthetic or cDNA-derived DNA fragments encoding tumor-specific neoantigen peptides or bioequivalent analogs, operatively linked to suitable transcriptional or translational regulatory elements derived from mammalian, microbial, viral, or insect genes. Transcriptional units typically comprise a set of: (1) one or more genetic elements that play a regulatory role in gene expression, such as transcription promoters or enhancers; (2) structural or coding sequences that are transcribed into mRNA and translated into protein; and (3) appropriate transcriptional and translational initiation and termination sequences, as detailed herein. Such regulatory elements may include operon sequences for controlling transcription. An origin of replication, which typically confers the ability to replicate in the host, and a selection gene that helps identify the transformant may also be incorporated. DNA regions are operatively linked when they are functionally related to each other. For example, the DNA of a signal peptide (secretory precursor) can be operatively linked to the DNA of a polypeptide if it is expressed as a precursor involved in the secretion of the polypeptide; a promoter can be operatively linked to a coding sequence if it controls the transcription of that sequence; or a ribosome-binding site can be operatively linked to a coding sequence if it is positionally permitted for translation. Generally, operatively linked means sequential, and in the case of a secretory precursor, means sequential and within the reading frame. Structural elements intended for use in yeast expression systems include a precursor sequence that allows the host cell to secrete the translated protein extracellularly. Alternatively, in cases where recombinant protein expression is not performed without a precursor or transport sequence, it may include an N-terminal methionine residue. This residue may then optionally be cleaved from the expressed recombinant protein to provide the final product.

[0451] Useful expression vectors for eukaryotic hosts, especially mammals or humans, include, for example, vectors containing expression control sequences from SV40, bovine papillomavirus, adenovirus, and cytomegalovirus. Useful expression vectors for bacterial hosts include known bacterial plasmids, such as plasmids from Escherichia coli (including pCR1, pBR322, pMB9, and their derivatives), broader host-range plasmids such as M13, and filamentous single-stranded DNA bacteriophages.

[0452] Suitable host cells for expressing peptides include prokaryotes, yeast, insects, or higher eukaryotic cells under the control of appropriate promoters. Prokaryotes include Gram-negative or Gram-positive organisms such as *Escherichia coli* or *Bacillus*. Higher eukaryotic cells include established mammalian-derived cell lines. Cell-free translation systems can also be used. Suitable cloning and expression vectors for use with bacterial, fungal, yeast, and mammalian cell hosts are well known in the art (see Pouwels et al., Cloning Vectors: A Laboratory Manual, Elsevier, NY, 1985).

[0453] Recombinant proteins can also be expressed using a variety of mammalian or insect cell culture systems. Recombinant proteins can be expressed in mammalian cells because such proteins are typically correctly folded, appropriately modified, and fully functional. Examples of suitable mammalian host cells include the COS-7 monkey kidney cell line described by Gluzman (Cell 23:175, 1981), as well as other cell lines capable of expressing appropriate vectors, including, for example, L cells, C127, 3T3, Chinese hamster ovary (CHO), 293, HeLa, and BHK cell lines. Mammalian expression vectors may include non-transcriptional elements (such as origin of replication), suitable promoters and enhancers linked to the gene to be expressed, and other 5' or 3' flanking non-transcriptional and 5' or 3' untranslated sequences, such as necessary ribosome binding sites, polyadenylation sites, splice donor and acceptor sites, and transcription termination sequences. Baculovirus systems for producing heterologous proteins in insect cells were outlined by Luckow and Summers, Bio / Technology 6:47 (1988).

[0454] Proteins produced by a transformed host can be purified using any suitable method. Standard methods include chromatography (e.g., ion exchange, affinity and size fractionation column chromatography, etc.), centrifugation, differential solubility, or any other standard technique used for protein purification. Affinity tags (such as hexahistidine, maltose-binding domains, influenza coating sequences, glutathione S-transferases, etc.) can be attached to proteins to allow for easy purification by passing them through a suitable affinity column. Separated proteins can also be characterized physically using techniques such as proteolysis, nuclear magnetic resonance, and X-ray crystallography.

[0455] For example, the supernatant from the system that secretes the recombinant protein into the culture medium can be first concentrated using a commercially available protein concentrator filter (e.g., an Amicon or Millipore Pellicon ultrafiltration unit). Following the concentration step, the concentrate can be applied to a suitable purification matrix. Alternatively, anion exchange resins, such as matrices or substrates with pendant diethylaminoethyl (DEAE) groups, can be used. The matrix can be acrylamide, agarose, dextran, cellulose, or other types commonly used in protein purification. Alternatively, a cation exchange step can be used. Suitable cation exchangers include various insoluble matrices containing sulfopropyl or carboxymethyl groups. Finally, one or more reversed-phase high-performance liquid chromatography (RP-HPLC) steps using a hydrophobic RP-HPLC medium (e.g., silica gel with pendant methyl or other aliphatic groups) can be used for further purification of the cancer stem cell protein-Fc composition. Some or all of the foregoing purification steps, in different combinations, can also be used to provide homogeneous recombinant protein.

[0456] Recombinant proteins produced in bacterial cultures can be separated, for example, by initial extraction from cell pellets followed by one or more concentration, salting-out, aqueous ion exchange, or size exclusion chromatography steps. High-performance liquid chromatography (HPLC) can be used for the final purification step. The microbial cells used in expressing the recombinant protein can be destroyed by any conventional method, including freeze-thaw cycles, sonication, mechanical destruction, or the use of cell lysis agents.

[0457] In vivo peptide / polypeptide synthesis

[0458] The present invention also contemplates the use of nucleic acid molecules as a carrier for delivering neoantigen peptides / polypeptides, in the form of, for example, DNA / RNA vaccines, to subjects in need (see, for example, WO 2012 / 159643 and WO 2012 / 159754, which are incorporated herein by reference in their entirety).

[0459] In one embodiment, a neoantigen can be administered to a patient in need by using plasmids. These are plasmids that typically consist of strong viral promoters to drive in vivo transcription and translation of the gene of interest (or complementary DNA) (Mor et al., (1995). The Journal of Immunology 155(4):2039–204). Sometimes introns A can be included to improve mRNA stability and thus increase protein expression (Leitner et al., (1997). The Journal of Immunology 159(12):6112–6119). Plasmids also include strong polyadenylation / transcription termination signals, such as bovine growth hormone or rabbit β-globulin polyadenylation sequences (Alarcon et al., (1999). Adv. Parasitol. Advances in Parasitology 42:343–410; Robinson et al. (2000) Adv. Virus Res. Advances in Virus Research 55:1–74; Lewis et al., (1996) Journal of Immunological Methods, 193(1):29–40. Sometimes polycistronic vectors are constructed to express more than one immunogen, or to express immunogens and immunostimulatory proteins (Lewis et al., (1999) Advances in Virus Research (Academic Press), 54:129–88).

[0460] Because plasmids are the “vectors” from which they express immunogens, optimized vector design for maximizing protein expression is crucial (Lewis et al., (1999). Advances in Virus Research (Academic Press) 54:129–88). One way to enhance protein expression is by optimizing the codon usage of pathogenic mRNAs targeting eukaryotic cells. Another consideration is promoter selection. Such promoters could be the SV40 promoter or Rous sarcoma virus (RSV).

[0461] Plasmids can be introduced into animal tissues using a variety of different methods. The two most common routes are injection of DNA in saline using a standard hypodermic needle and gene gun delivery. A schematic diagram illustrating the construction of DNA vaccine plasmids and their subsequent delivery into the host via these two methods is illustrated in Scientific American (Weiner et al., (1999) Scientific American 281(1):34–41). Injection in saline is typically performed in the intramuscular (IM) or intradermal (ID) space of skeletal muscle, where DNA is delivered into the extracellular space. This can be achieved by using muscle toxins, such as bupivacaine; or by temporarily damaging muscle fibers using a hypertonic solution of saline or sucrose, assisted by electroporation (Alarcon et al., ...).

[0462] (1999). Adv. Parasitol. Advances in Parasitology [Advances in Parasitology Research] 42:343–410). Immune responses to this delivery method can be influenced by many factors, including the type of needle, needle alignment, injection speed, injection volume, muscle type, and the age, sex, and physiological condition of the animal being injected (Alarcon et al., (1999). Adv. Parasitol. Advances in Parasitology [Advances in Parasitology Research] 42:343–410).

[0463] Gene gun delivery (another commonly used delivery method) uses compressed helium as a catalytic agent to ballistically accelerate plasmid DNA (pDNA) already adsorbed onto gold or tungsten microparticles into target cells (Alarcon et al.).

[0464] (1999). Adv. Parasitol. Advances in Parasitology 42:343–410; Lewis et al., (1999). Advances in Virus Research (Academic Press) 54:129–88.

[0465] Alternative delivery methods may include aerosol instillation of naked DNA onto mucosal surfaces, such as the nasal and lung mucosa (Lewis et al., (1999). Advances in Virus Research (Academic Press) 54:129–88), and topical administration of pDNA to the eye and vaginal mucosa (Lewis et al., (1999) Advances in Virus Research (Academic Press) 54:129–88). Mucosal surface delivery has been achieved using positively charged liposome-DNA formulations, biodegradable microspheres, attenuated Shigella or Listeria vectors, and recombinant adenovirus vectors for oral administration to the intestinal mucosa. DNA or RNA may also be delivered into cells after mild mechanical disruption of the cell membrane (temporary permeation of the cell). Such mild mechanical disruption of the membrane can be accomplished by gently forcing cells through a small pore (Ex Vivo Cytosolic Delivery of Functional Macromolecules to Immune Cells, Sharei et al, PLOS ONE|DOI:10.1371 / journal.pone.0118803April 13,2015).

[0466] The delivery method determines the dosage of DNA required to elicit an effective immune response. Saline injection requires a variable amount of DNA, ranging from 10 μg to 1 mg, while gene gun delivery requires 100 to 1000 times less DNA than intramuscular saline injection to elicit an effective immune response. Typically, 0.2 μg–20 μg is required, although amounts as low as 16 ng have been reported. These amounts vary between species; for example, mice require approximately 10 times less DNA than primates. Saline injection requires more DNA because the DNA is delivered to the extracellular space of the target tissue (usually muscle), where it must overcome physical barriers (such as the substrate and large amounts of connective tissue, to name a few) before it can be taken up by the cell. In contrast, gene gun delivery bombards DNA directly into the cell, resulting in less “loss” (see, for example, Sedegah et al., (1994). Proceedings of the National Academy of Sciences of the United States of America 91(21):9866–9870; Daheshiaet et al., (1997). The Journal of Immunology 59(4):1945–1952; Chen et al., (1998). The Journal of Immunology 160(5):2425–2432; Sizemore (1995) Science 270(5234):299–302; Fynan et al. (1993) Proc. Natl. Acad. Sci. USA 90(24):11478–82.

[0467] In one embodiment, the tumor-forming vaccine or immunogenic composition may comprise a separate DNA plasmid encoding, for example, one or more neoantigen peptides / peptides as identified according to the invention. As discussed herein, the exact choice of expression vector will depend on the peptide / peptide to be expressed and is entirely within the skill of a person of ordinary skill. The anticipated persistence of the DNA construct (e.g., in an appendage-type, non-replicating, non-integrating form in muscle cells) is expected to provide an increased duration of protection.

[0468] One or more of the novel antigenic peptides of the present invention can be encoded and expressed in vivo using virus-based systems (e.g., adenovirus systems, adeno-associated virus (AAV) vectors, poxviruses, or lentiviruses). In one embodiment, the tumorigenesis vaccine or immunogenic composition may include a virus-based vector for use in human patients in need of it, such as an adenovirus (see, for example, Baden et al., First-in-human evaluation of the safety and immunogenicity of a recombinant adenovirus serotype 26HIV-1Env vaccine (IPCAVD001). J Infect Dis. 2013 Jan 15; 207(2):240-7, which is incorporated herein by reference in its entirety). Plasmids that can be used for delivery of adeno-associated virus, adenovirus, and lentivirus have been previously described (see, for example, U.S. Patent Nos. 6,955,808 and 6,943,019, and U.S. Patent Application No. 20080254008, which are incorporated herein by reference).

[0469] The peptides and polypeptides of the present invention can also be expressed via vectors, such as nucleic acid molecules as discussed herein, such as RNA or DNA plasmids, viral vectors such as poxviruses, such as orthopoxvirus, fowlpoxvirus, or adenovirus, AAV, or lentivirus. This method involves using a vector to express the nucleotide sequence encoding the peptide of the present invention. Upon introduction into an acutely or chronically infected host or into an uninfected host, the vector expresses the immunogenic peptide and thereby elicits a host CTL response.

[0470] In vectors suitable for use in the practice of this invention, integration into the host genome of a cell using retroviral gene transfer methods is possible, which typically leads to long-term expression of the inserted transgene. In a preferred embodiment, the retrovirus is a lentivirus. Furthermore, high transduction efficiency has been observed in various cell types and target tissues. The tropism of the retrovirus can be altered by incorporating exogenous envelope proteins to expand the potential target population of the target cells. Retroviruses can also be engineered to allow conditional expression of the inserted transgene, thus enabling certain cell types to be infected by the lentivirus. Cell type-specific promoters can be used for targeted expression in specific cell types. Lentiviral vectors are retroviral vectors (and therefore both lentiviral and retroviral vectors can be used in the practice of this invention). Furthermore, lentiviral vectors are preferred because they are capable of transducing or infecting non-dividing cells and typically produce high viral titers. Therefore, the choice of retroviral gene transfer system depends on the target tissue. Retroviral vectors consist of cis-acting long terminal repeats that have the ability to package up to 6-10 kb of exogenous sequences. The minimum amount of cis-acting LTR is sufficient for vector replication and packaging, and then these vectors are used to integrate the desired nucleic acid into target cells to provide permanent expression. Widely used retroviral vectors that can be used in the practice of this invention include those based on murine leukemia virus (MuLV), gibberish leukemia virus (GaLV), simian immunodeficiency virus (SIV), human immunodeficiency virus (HIV), and combinations thereof (see, for example, Buchscher et al., (1992) J. Virol. [Journal of Virology] 66:2731-2739; Johann et al., (1992) J. Virol. [Journal of Virology] 66:1635-1640; Sommnerfelt et al., (1990) Virol. [Journal of Virology] 176:58-59; Wilson et al., (1998) J. Virol. [Journal of Virology] 63:2374-2378; Miller et al., (1991) J. Virol. [Journal of Virology] 65:2220-2224; PCT / US 94 / 05700).

[0471] Also useful in the practice of this invention are minimal nonprimate lentiviral vectors, such as lentiviral vectors based on equine infectious anemia virus (see, for example, Balagaan. (2006) J Gene Med; 8:275-285, published online on 21 November 2005 in the Wiley InterScience database (www.interscience.wiley.com) DOI:10.1002 / jgm.845). These vectors may have cytomegalovirus (CMV) promoters that drive the expression of target genes. Accordingly, the present invention contemplates one or more vectors useful in the practice of this invention: viral vectors, including retroviral vectors and lentiviral vectors.

[0472] Lentiviral vectors have also been disclosed for use in the treatment of Parkinson's disease, see, for example, U.S. Patent Publication No. 20120295960 and U.S. Patent Nos. 7303910 and 7351585. Lentiviral vectors have also been disclosed for delivery to the brain, see, for example, U.S. Patent Publication Nos. US20110293571; US20040013648, US...

[0473] US20070025970, US20090111106, and US Patent No. 7259015. In another embodiment, a lentiviral vector is used to deliver the vector to the brain of a subject undergoing treatment for a disease.

[0474] Regarding lentivirus vector systems useful in the practice of this invention, references are made to U.S. Patent Nos. 6,428,953, 6,165,782, 6,013,516, 5,994,136, 6,312,682, and 7,198,784, and the documents referenced therein.

[0475] In one embodiment described herein, the delivery is performed via lentivirus. Zou et al. administered approximately 10 μl of lentivirus with a 1x10 nucleotide count via an intrathecal catheter. 9 Recombinant lentivirus at titers of transduction units (TU) / ml. These types of doses can be adapted or extrapolated to allow the use of retroviral or lentiviral vectors in this invention. For transduction in tissues such as the brain, very small volumes are required; therefore, the viral preparation is concentrated by ultracentrifugation. The resulting preparation should have a titer of at least 10... 8 TU / ml, preferably 10 8 Up to 10 9 TU / ml, more preferably at least 10 9 TU / ml. Other concentration methods can be used, such as ultrafiltration or binding to and eluting from a matrix.

[0476] In other embodiments, the amount of lentivirus administered may be 1.x.10. 5 Or approximately 1.x.10 5 5.x.10 phage plaque-forming units (PFU) 5 Or approximately 5.x.10 5 PFU, 1.x.10 6 Or approximately 1.x10 6 PFU, 5.x.10 6 Or approximately 5.x.10 6 PFU, 1.x.10 7 Or approximately 1.x.10 7 PFU, 5.x.10 7 Or approximately 5.x.10 7 PFU, 1.x.10 8 Or approximately 1.x.10 8 PFU, 5.x.10 8 Or approximately 5.x.10 8 PFU, 1.x.10 9 Or approximately 1.x.10 9 PFU, 5.x.10 9 Or approximately 5.x.10 9 PFU, 1.x.10 10 Or approximately 1.x.10 10 PFU or 5.x.10 10 Or approximately 5.x.10 10 PFU may be administered as a single total dose to a typical individual weighing 75 kg, or adjusted for the subject's weight, size, and species. Those skilled in the art can determine the appropriate dose. The appropriate dose of the virus can be determined empirically.

[0477] Adenoviral vectors are also useful in the practice of this invention. One advantage is the ability of recombinant adenoviruses to efficiently transfer and express recombinant genes, resulting in high expression of the transferred nucleic acids, both in vitro and in vivo in a variety of mammalian cells and tissues. Additionally, the ability to efficiently infect dormant cells extends the utility of the recombinant adenoviral vector. Furthermore, the high expression level ensures that the nucleic acid product will be expressed at a sufficient level to generate an immune response (see, for example, U.S. Patent No. 7,029,848, which is incorporated herein by reference).

[0478] Regarding adenovirus vectors useful in practicing this invention, U.S. Patent No. 6,955,808 is cited. The adenovirus vectors used may be selected from the group consisting of Ad5, Ad35, Ad11, C6, and C7 vectors. The sequence of the adenovirus 5 (“Ad5”) genome has been disclosed. (Chroboczek, J., Bieber, F., and Jacrot, B. (1992) The Sequence of the Genome of Adenovirus Type 5 and Its Comparison with the Genome of Adenovirus Type 5) 2. Virology [Sequences of the genome of adenovirus type 5 and comparisons with the genome of adenovirus type 2, Virology] 186, 280-285; the contents of which are incorporated herein by reference. The Ad35 vector is described in U.S. Patent Nos. 6,974,695, 6,913,922, and 6,869,794. The Ad11 vector is described in U.S. Patent No. 6,913,922. The C6 adenovirus vector is described in U.S. Patent Nos. 6,780,407; 6,537,594; 6,309,647; 6,265,18 9; 6,156,567; 6,090,393; 5,942,235 and 5,833,975. The C7 vector is described in U.S. Patent No. 6,277,558. Also usable are E1-deficient or deleted, E3-deficient or deleted, and / or E4-deficient or deleted adenovirus vectors. Certain adenoviruses with mutations in the E1 region have improved safety limits because, in non-permissive cells, E1-deficient adenovirus mutants are replication-deficient, or minimally attenuated, and highly attenuated. By disrupting this mechanism, the adenovirus thereby downregulates MHC. Class I molecules, adenoviruses with mutations in the E3 region may have enhanced immunogenicity. Adenoviruses with E4 mutations may have reduced immunogenicity compared to adenovirus vectors due to suppression of late gene expression. Such vectors are particularly useful when repeated reinoculations using the same vector are desired. According to the invention, E1, E3, E4, E1 and E3, and E1 and E4 deletions or mutations can be used. Furthermore, according to the invention, "gutless" adenovirus vectors (where all viral genes are deleted) can also be used. For replication, such vectors require helper viruses and specific human 293 cell lines expressing both E1a and Cre, conditions not present in the natural environment. Such "gutless" vectors are non-immunogenic, and therefore can be inoculated multiple times for reinoculation. "Gutless" adenovirus vectors can be used for inserting heterologous inserts / genes (e.g., the transgenes of the present invention) and even for co-delivery of large quantities of heterologous inserts / genes.

[0479] In one embodiment of this document, delivery is carried out via adenovirus, which may contain at least 1 x 10⁻⁶ ions. 5 A single booster dose of one adenovirus vector particle (also known as a particle unit, pu). In one embodiment herein, this dose is preferably at least about 1 x 10⁻⁶ of the adenovirus vector. 6 1 x 103 particles (e.g., approximately 1 x 10<s 6 -1x 10 12 (particles), more preferably at least about 1 x 103 7 One particle, more preferably at least about 1 x 10⁻⁶ 8 1 x 103 particles (e.g., approximately 1 x 10<s 8 -1x 10 11 One particle or approximately 1 x 10 8 -1x 10 12 (number of particles), and preferably at least about 1 x 10^6 particles. 9 1 x 103 particles (e.g., approximately 1 x 10<s 9 -1x 10 10 One particle or approximately 1x10 9 -1x 10 12 (a few particles), or even at least about 1 x 10^6 particles. 10 1 x 103 particles (e.g., approximately 1 x 10<s 10 -1x 10 12 (particles). Alternatively, the dose contains no more than about 1 x 10⁻⁶ particles. 14 Particles, preferably no more than about 1 x 10^6 13 One particle, or even more preferably no more than about 1 x 10⁻⁶. 12 One particle, or even more preferably no more than about 1 x 10⁻⁶. 11 The number of particles is approximately 1 x 10^6, and the optimal value is no more than approximately 1 x 10^ 10 10 particles (e.g., no more than about 1 x 10^12) 9 (particles). Therefore, this dose may contain a single dose of adenovirus vector, having, for example, approximately 1 x 103 particles. 6 Particle unit (pu), approximately 2 x 10-1 6 PU, approximately 4 x 10 6 PU, approximately 1x10 7 PU, approximately 2 x 10 7 PU, approximately 4x10 7 PU, approximately 1 x 10 8 PU, approximately 2 x 10 8 PU, approximately 4 x 10 8 PU, approximately 1 x 10 9 PU, approximately 2 x 10 9 PU, approximately 4 x 10 9PU, approximately 1x10 10 PU, approximately 2 x 10 10 PU, approximately 4 x 10 10 PU, approximately 1 x 10 11 PU, approximately 2 x 10 11 PU, approximately 4 x 10 11 PU, approximately 1 x 10 12 PU, approximately 2 x 10 12 pu, or approximately 4 x 10 12 The adenovirus vector of pu. See, for example, the adenovirus vector (incorporated herein by reference) in U.S. Patent No. 8,454,972B2 to Nabel et al., issued June 4, 2013, and its dosage form in column 29, lines 36-58. In one embodiment herein, the adenovirus is delivered via multiple doses.

[0480] In terms of in vivo delivery, AAV is superior to other viral vectors due to its low toxicity and low likelihood of inducing insertional mutagenesis, as it does not integrate into the host genome. AAV has a packaging limit of 4.5 or 4.75 kb. Constructs larger than 4.5 or 4.75 kb result in a significant reduction in viral production. Many promoters exist that can be used to drive nucleic acid molecule expression. AAV ITR can be used as a promoter and is advantageous for eliminating the need for additional promoter elements. For widespread expression, the following promoters can be used: CMV, CAG, CBh, PGK, SV40, ferritin heavy or light chains, etc. For brain expression, the following promoters can be used: synaptic I for all neurons, CaMKIIα for excitatory neurons, GAD67 or GAD65 or VGAT for GABAergic neurons, etc. Promoters for driving RNA synthesis can include: Pol III promoters, such as U6 or H1. Pol II promoters and intron cassettes can be used to express guide RNA (gRNA).

[0481] Regarding AAV carriers useful in the practice of this invention, references are made to U.S. Patent Nos. 6,58785, 7,115,391, 7,172,893, 6,953,690, 6,936,466, 6,924,128, 6,893,865, 6,793,926, 6,537,540, 6,475,769, and 6,258,595, and the documents referenced therein.

[0482] Regarding AAV, the AAV can be AAV1, AAV2, AAV5, or any combination thereof. The AAV can be selected relative to the cells to be targeted; for example, AAV serotypes 1, 2, 5, or hybrid capsids AAV1, AAV2, AAV5, or any combination thereof can be selected for targeting brain or neuronal cells; and AAV4 can be selected for targeting cardiac tissue. AAV8 can be used for delivery to the liver. The above promoters and vectors are individually preferred.

[0483] In one embodiment of this document, the delivery is performed via AAV. Therapeuticly effective doses for in vivo delivery of AAV to humans are considered to be those containing approximately 1 x 10⁻⁶ mg / L. 10 To approximately 1 x 10 50 The dosage of AAV / ml solution ranges from about 20 to about 50 ml of saline solution. This dosage can be adjusted to balance the therapeutic benefit with any side effects. In one embodiment described herein, the AAV dosage is approximately from about 1 x 10⁻⁶. 5 To 1x 10 50 One genome AAV, from approximately 1 x 10 8 To 1x 10 20 One genome AAV, from approximately 1 x 10 10 To approximately 1 x 10 16 One genome, or approximately 1 x 10^6 11 To approximately 1 x 10 16 The concentration range of AAV in each genome. The human dose can be approximately 1 x 10^6. 13 One genomic AAV. Such concentrations can be delivered in vector solutions ranging from about 0.001 ml to about 100 ml, about 0.05 ml to about 50 ml, or about 10 ml to about 25 ml. In a preferred embodiment, about 2 x 10⁻⁶ genomic AAVs are used. 13 The viral genome / mL titer was measured using AAV, and each striatal hemisphere of the mouse received a single 500 nm injection. Other effective doses can be readily determined by those skilled in the art through routine experiments that establish dose-response curves. See, for example, U.S. Patent No. 8,404,658B2 to Hajjar et al., issued March 26, 2013, column 27, lines 45-60.

[0484] In another embodiment, effective activation of a cellular immune response to a neoantigen vaccine or immunogenic composition can be achieved by expressing the relevant neoantigen in a vaccine or immunogenic composition in a non-pathogenic microorganism. Well-known examples of such microorganisms are Mycobacterium bovis (BCG), Salmonella spp., and Pseudomonas spp. (see U.S. Patent No. 6,991,797, which is incorporated herein by reference in its entirety).

[0485] In another embodiment, poxviruses are used in tumorigenesis vaccines or immunogenic compositions. These include orthopoxvirus, fowlpox, cowpox, MVA, NYVAC, canarypox, ALVAC, fowlpox, TROVAC, etc. (see, for example, Verardiet et al., Hum Vaccin Immunother. [Human Vaccines and Immunotherapy] 2012 July; 8(7):961-70; and Moss, Vaccine. 2013; 31(39):4220–4222). Poxvirus expression vectors were described in 1982 and quickly became widely used in vaccine development along with research in multiple fields. The advantages of the vectors include simple construction, the ability to adapt to large amounts of exogenous DNA, and high expression levels.

[0486] Regarding poxviruses that can be used in practicing this invention, such as vertebrate poxviruses (vertebrate poxviruses), such as orthopoxvirus and fowlpoxvirus, such as cowpoxvirus (e.g., Wyeth Strain, WR strain (e.g.) VR-1354), Copenhagen strain, NYVAC, NYVAC.1, NYVAC.2, MVA, MVA-BN), canarypox virus (e.g., Wheatley C93 strain, ALVAC), fowlpox virus (e.g., FP9 strain, Webster strain, TROVAC), pigeonpox, pigeonpox virus, quailpox, and raccoon pox, especially their synthetic or non-naturally occurring recombinants, their uses, and methods for manufacturing and using such recombinants can be found in scientific and patent literature, for example:

[0487] U.S. Patent Nos. 4,603,112, 4,769,330, 5,110,587, 5,174,993, 5,364,773, 5,762,938, 5,494,807, 5,766,597, 7,767,449, 6,780,407, 6,537,594, 6,265,189, 6,214,353, 6,130,066, 6,004,777, 5,990,091, 5,942,235, 5,833,975, 5,766,597, 5,756,101, 7,045,313, 6,780,417, 8,470,598, 8,372,622 8,268,329, 8,268,325, 8,236,560, 8,163,293, 7,964,398, 7,964,396, 7,964,395, 7,939,086, 7,923,017, 7,897,156, 7,892,533, 7,628,980, 7,459,270, 7,445,924, 7,384,644, 7,335,364, 7,189,536, 7,097,842, 6,913,752, 6,761,893, 6,682,743, 5,770,212, 5,766,882, and 5,989,562, and

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[0523] Each reference is cited here.

[0524] In another embodiment, vaccinia virus is used in tumorigenesis vaccines or immunogenic compositions to express neoantigens. (Rolph et al., Recombinant viruses as vaccines and immunological tools. Curr Opin Immunol)

[0525] 9:517-524, 1997). Recombinant vaccinia virus can replicate within the cytoplasm of infected host cells, and therefore the polypeptides of interest can induce an immune response. Furthermore, poxviruses have been widely used as vectors for vaccines or immunogenic compositions due to their ability to target antigens encoded by the major histocompatibility complex class I pathway through direct infection of immune cells, specifically antigen-presenting cells, but also due to their adjuvant capabilities.

[0526] In another embodiment, ALVAC is used as a carrier in tumorigenesis vaccines or immunogenic compositions. ALVAC is a canarypox virus that can be modified to express exogenous transgenes and has been used as a method for inoculation against both prokaryotic and eukaryotic antigens (Horig H, Lee DS, Conkright W et al., Phase I clinical trial of a recombinant canarypoxvirus (ALVAC) vaccine expressing human carcinoembryonic antigen and the B7.1 co-stimulatory molecule. Cancer Immunol Immunother 2000; 49:504–14; von Mehren M, Arlen P, Tsang Ky et al., Pilot study of a dual gene recombinant avipox vaccine containing both carcinoembryonic antigen (CEA) and B7.1 transgenes in patients with recurrent CEA-expressing Adenocarcinoma [A pilot study of a double-gene recombinant fowlpox containing both carcinoembryonic antigen (CEA) and B7.1 transgenes in patients with recurrent CEA-expressing adenocarcinoma]. Clin Cancer Res 2000; 6:2219–28; Musey L, Ding Y, Elizaga M et al. HIV-1 vaccination administered intramuscularly can induce both systemic and mucosal T cell immunity in HIV-1-uninfected individuals. J Immunol 2003; 171:1094–101; Paoletti E. Applications of pox virus vectors to vaccination: an update. Proc. Natl. Acad. Sci.USA [Proceedings of the National Academy of Sciences of the United States of America] 1996; 93:11349–53; US Patent No. 7,255,862). In a Phase I clinical trial, ALVAC virus expressing the tumor antigen CEA showed excellent safety and resulted in increased CEA-specific T-cell responses in selected patients; however, the target clinical response was not observed (Marshall JL, Hawkins MJ, Tsang KY et al., Phase I study in cancer patients of a replication-defective avipox recombinant vaccine that expresses human carcinoembryonic antigen. J Clin Oncol [Phase I study in cancer patients of a replication-defective avipox recombinant vaccine expressing human carcinoembryonic antigen, Journal of Clinical Oncology] 1999; 17:332–7).

[0527] In another embodiment, the modified Ankara vaccinia virus (MVA) can be used as a viral vector for neoantigen vaccines or immunogenic compositions. MVA is a member of the orthopoxvirus family and has been propagated for approximately 570 consecutive passages in chicken embryo fibroblasts of the Ankara strain of vaccinia virus (for review, see Mayr, A. et al., Infection 3, 6-14, 1975). As a result of these passages, the resulting MVA virus contains 31 kilobases less genomic information than CVA and is highly host-cell restricted (Meyer, H. et al., J. Gen. Virol. 72, 1031-1038, 1991). MVA is characterized by its extreme attenuation, i.e., reduced virulence or infectivity, but still retains excellent immunogenicity. When tested in various animal models, MVA has demonstrated to be non-virulent, even in immunosuppressed individuals. Furthermore, MVA- -HER2 is a candidate immunotherapy designed for the treatment of HER-2 positive breast cancer and is currently in clinical trials. (Mandl et al., Cancer Immunol Immunother. [Cancer Immunology and Immunotherapy] 2012 Jan; 61(1):19–29). Methods for preparing and using recombinant MVA have been described (see, for example, U.S. Patent Nos. 8,309,098 and 5,185,146, which are incorporated herein by reference in their entirety).

[0528] In another embodiment, modified Copenhagen strains of vaccinia virus, NYVAC and NYVAC variants were used as vectors (see U.S. Patent Nos. 7,255,862; PCT WO 95 / 30018; U.S. Patent Nos. 5,364,773 and 5,494,807, which are incorporated herein by reference in their entirety).

[0529] In one embodiment, recombinant viral particles of a vaccine or immunogenic composition are administered to a patient in need. The dose range of the expressed neoantigen can be from a few micrograms to hundreds of micrograms, for example, 5 to 500 μg. The vaccine or immunogenic composition can be administered in any suitable amount to achieve expression at these dose levels. It can be administered at a dose of at least about 10 μg. 3.5 The amount of PFU is used to administer viral particles to patients who need them or to transfect cells; therefore, preferably, at least about 10 4 pfu to approximately 10 6 PFU (Prophylactic Fuel) delivers viral particles to patients who need them or infects or transfects cells; however, at least approximately 10 PFU can be given to patients who need them. 8 pfu, thus allowing for a more preferred amount to be at least 10 7 pfu to approximately 10 9 PFU. The dosage for NYVAC is applicable to ALVAC, MVA, MVA-BN, and fowlpox, such as canarypox and chickenpox.

[0530] Vaccine or immunogenic composition adjuvant

[0531] Effective vaccines or immunogenic compositions advantageously include strong adjuvants to initiate immune responses. As described herein, poly-ICLCs have demonstrated several promising properties for use as adjuvants in vaccines or immunogenic compositions; poly-ICLCs are agonists of the TLR3 and RNA helicase-MDA5 and RIG3 domains. These properties include inducing local and systemic activation of immune cells in vivo, producing stimulating chemokines and cytokines, and stimulating antigen presentation via dendritic cells (DCs). Furthermore, poly-ICLCs can induce durable CD4+ and CD8+ responses in humans. Importantly, striking similarities were observed in the upregulation of transcriptional and signal transduction pathways in subjects vaccinated with poly-ICLCs and in volunteers who had received a highly effective, replicating yellow fever vaccine. Moreover, in a recent phase 1 study, >90% of ovarian cancer patients immunized with poly-ICLCs in combination with NY-ESO-1 peptide vaccines (excluding Montaned) showed induction of CD4+ and CD8+ T cells, as well as antibody responses to the peptide. Meanwhile, poly-ICLCs have been extensively tested in more than 25 clinical trials to date and have demonstrated relatively benign toxicity profiles. In addition to being potent and specific immunogens, these neoantigen peptides can be combined with adjuvants (e.g., poly-ICLCs) or other antitumor agents. Unbound by theory, these neoantigens are expected to circumvent central thymic tolerance (thus allowing for stronger antitumor T-cell responses) while reducing the likelihood of autoimmunity (e.g., by avoiding targeting normal autoantigens). Effective immune responses favorably include strong adjuvants to activate the immune system (Speiser and Romero, Molecularly defined vaccines for cancer immunotherapy, and protective T-cell immunity, Seminarsin Immunol, 22:144 (2010)). For example, Toll-like receptors (TLRs) have been shown to be powerful sensors of “danger signals” from microbial and viral pathogens, effectively inducing the innate immune system and consequently, the adaptive immune system (Bhardwaj and Gnjatic, TLR AGONISTS: Are They Good Adjuvants? [TCR Agonists: Are They Good Adjuvants?] Cancer J. 16:382-391 (2010)). Among TLR agonists, poly-ICLC (a synthetic double-stranded RNA mimic) is one of the most potent activators of bone marrow-derived dendritic cells.In a human volunteer study, poly-ICLC has been shown to be safe and to induce gene expression profiles in peripheral blood cells comparable to those induced by yellow fever vaccine YF-17D, one of the most effective live attenuated viral vaccines (Caskey et al., Synthetic double-stranded RNA induces innate immune responses similar to alive viral vaccine in humans J Exp Med [Synthetic double-stranded RNA induces innate immune responses similar to alive viral vaccine in humans J Exp Med [Journal of Experimental Medicine] 208:2357 (2011)]). In a preferred embodiment, ... (A GMP formulation of poly-ICLC prepared by Oncovir) is used as an adjuvant. In other embodiments, other adjuvants described herein are contemplated. For example, oil-in-water, water-in-oil, or multiphase W / O / W; see, for example, US 7,608,279 and Aucouturier et al., Vaccine 19 (2001), 2666-2672, and the documents cited herein.

[0532] Indications

[0533] Examples of cancers and cancer conditions for which the therapies in this document can be used include, but are not limited to, patients who have been diagnosed with cancer or are at risk of developing cancer and who require treatment. Subjects may have solid tumors such as breast cancer, ovarian cancer, prostate cancer, lung cancer, kidney cancer, stomach cancer, colon cancer, testicular cancer, head and neck cancer, pancreatic cancer, brain cancer, melanoma, and other tumors of tissues and organs, as well as hematologic malignancies such as lymphoma and leukemia, including acute myeloid leukemia, chronic myeloid leukemia, chronic lymphocytic lymphoma, T-cell lymphocytic leukemia, and B-cell lymphoma; tumors of the brain and central nervous system (e.g., tumors of the meninges, brain, spinal cord, cranial nerves, and other parts of the CNS, such as malignant gliomas or medulloblastomas); head and / or neck cancer; breast tumors; circulatory system tumors (e.g., heart, mediastinum and pleura, and other intrathoracic organs, blood vessels); and other tumors of the circulatory system. Tumors (tumors and tumor-associated vascular tissues); tumors of the blood and lymphatic system (e.g., Hodgkin's disease, non-Hodgkin's lymphoma, Burkitt lymphoma, S-related lymphoma, malignant immunoproliferative disorders, multiple myeloma, and malignant plasma cell tumors, lymphoid leukemia, myeloid leukemia, acute or chronic lymphocytic leukemia, monocytic leukemia, other leukemias of specific cell types, leukemias of unspecified cell types, unspecified malignancies of lymphoid, hematopoietic, and related tissues, such as diffuse large cell lymphoma, T-cell lymphoma, or T-cell lymphoma of the skin); tumors of the excretory system (e.g., kidneys, renal pelvis, ureters, bladder, and other urinary organs). Tumors; tumors of the gastrointestinal tract (e.g., esophagus, stomach, small intestine, colon, colorectal, rectosigmoid junction, rectum, anus, and anal canal); tumors involving the liver and intrahepatic bile ducts, gallbladder, and other parts of the bile ducts, pancreas, and other digestive organs; tumors of the oral cavity (e.g., lips, tongue, gums, floor of mouth, palate, parotid glands, salivary glands, tonsils, oropharynx, nasopharynx, piriform sinus, hypopharynx, and other parts of the oral cavity); tumors of the reproductive system (e.g., vulva, vagina, cervix, uterus, ovaries, and other parts associated with female genitalia, placenta, penis, prostate, testes, and other parts associated with male genitalia); tumors of the respiratory tract (e.g., nasal cavity, middle ear, paranasal sinuses, larynx, trachea, bronchi). Tumors of the tubes and lungs (e.g., small cell lung cancer and non-small cell lung cancer); tumors of the skeletal system (e.g., bones and articular cartilage of the limbs, joint cartilage, and other sites); tumors of the skin (e.g., malignant melanoma of the skin, non-melanoma skin cancer, basal cell carcinoma of the skin, squamous cell carcinoma of the skin, mesothelioma, Kaposi's sarcoma); and tumors involving other tissues, including peripheral nerves and the autonomic nervous system, connective tissue and soft tissue, retroperitoneum and peritoneum, eyes, thyroid gland, adrenal glands and other endocrine glands and related structures, secondary and unspecified malignancies of lymph nodes, secondary malignancies of the respiratory and digestive systems, and secondary malignancies of other sites.Therefore, the subject population described herein may have one of the above-mentioned cancer types. In other embodiments, the subject population may be all subjects with solid tumors or all subjects with liquid tumors.

[0534] Of particular significance is the treatment of non-Hodgkin's lymphoma (NHL), clear cell renal cell carcinoma (ccRCC), metastatic melanoma, sarcoma, leukemia, or bladder cancer, colon cancer, brain cancer, breast cancer, head and neck cancer, endometrial cancer, lung cancer, ovarian cancer, pancreatic cancer, or prostate cancer. In some embodiments, the melanoma is high-risk melanoma.

[0535] Among other things, cancers that can be treated with the therapies described herein may include cases that are refractory to treatment with other chemotherapeutic agents. As used herein, the term “refractory” means cancer (and / or its metastases) that, after treatment with another chemotherapeutic agent, shows no or only a weak antiproliferative response (e.g., no or only a weak inhibition of tumor growth). These are cancers that cannot be satisfactorily treated with other chemotherapeutic agents. Refractory cancers encompass not only (i) cancers in which one or more chemotherapeutic agents have failed during the patient’s treatment, but also (ii) cancers that may be refractory by other means, such as biopsy cultures in the presence of a chemotherapeutic agent.

[0536] The therapy described herein is also applicable to the treatment of previously untreated patients who require it.

[0537] The therapy described herein is also applicable in cases where the subject does not have detectable tumor formation but is at high risk of disease recurrence.

[0538] Of particular interest is the treatment of patients in need who have undergone autologous hematopoietic stem cell transplantation (AHSCT), and specifically, patients who have demonstrated residual disease after undergoing AHSCT. The post-AHSCT background is characterized by low levels of residual disease, immune cell infusion leading to a self-balancing expansion, and the absence of any standard relapse-delaying treatments. These characteristics offer a rare opportunity to delay disease relapse using claimed tumorigenesis vaccines or immunogenic compositions.

[0539] Pharmaceutical Composition / Delivery Method

[0540] The present invention also relates to pharmaceutical compositions comprising an effective amount of one or more neoantigen peptides described herein (including pharmaceutically acceptable salts thereof) optionally combined with a pharmaceutically acceptable carrier, excipient or additive.

[0541] When administered in combination, the therapeutic agents (i.e., neoantigen peptides) can be formulated into separate compositions that are administered simultaneously or at different times, or these therapeutic agents can be administered as a single composition.

[0542] These compositions can be administered once daily, twice daily, every two days, every three days, every four days, every five days, every six days, every seven days, every two weeks, every three weeks, every four weeks, every two months, every six months, or once a year. The dosing interval can be adjusted according to the individual patient's needs. For longer dosing intervals, extended-release or long-acting formulations can be used.

[0543] The compositions of the present invention can be used to treat acute diseases and symptoms, and can also be used to treat chronic conditions. Specifically, the compositions of the present invention can be used in methods for treating or preventing tumor formation. In some embodiments, the compounds of the present invention are administered for a period of time exceeding two weeks, three weeks, one month, two months, three months, four months, five months, six months, one year, two years, three years, four years, or five years, ten years, or fifteen years; or, for example, any range of time periods in days, months, or years, wherein the lower limit of the range is any period between 14 days and 15 years and the upper limit of the range is between 15 days and 20 years (e.g., between 4 weeks and 15 years, between 6 months and 20 years). In some cases, it may be advantageous to administer the compounds of the present invention to the patient for the remainder of their life. In a preferred embodiment, the patient is monitored to examine the progression of the disease or disorder and the dosage is adjusted accordingly. In a preferred embodiment, treatment according to the present invention is effectively sustained for at least two weeks, three weeks, one month, two months, three months, four months, five months, six months, one year, two years, three years, four years, or five years, ten years, fifteen years, twenty years, or for the remainder of the subject's life.

[0544] Surgical resection uses surgical procedures to remove abnormal tissue from cancerous tissue, such as mediastinal, neurogenic, or germ cell tumors, or thymomas. In some embodiments, administration of the composition is initiated after tumor resection. In other embodiments, administration of the tumor-forming vaccine or immunogenic composition is initiated 1, 2, 3, 4, 5, 6, 7, 8, 9, 10, 11, 12, 13, 14, 15, or more weeks after tumor resection. Preferably, administration of the tumor-forming vaccine or immunogenic composition is initiated 4, 5, 6, 7, 8, 9, 10, 11, or 12 weeks after tumor resection.

[0545] A prime / boost regimen refers to the continuous administration of a vaccine or immunogenic or immunological composition. In some embodiments, the tumorigenesis vaccine or immunogenic composition is administered in a prime / boost dose regimen, for example, at weeks 1, 2, 3, or 4 as a prime and at months 2, 3, or 4 as a booster. In another embodiment, a heterologous prime strategy is used to induce a greater cytotoxic T cell response (see Schneider et al., Induction of CD8+ T cells using heterologous prime-boost immunisation strategies, Immunological Reviews Volume 170, Issue 1, pages 29–38, August 1999). In yet another embodiment, DNA encoding a neoantigen is used for the prime, followed by a protein booster. In another embodiment, the protein is used for primary immunization, followed by a booster with a virus encoding a neoantigen. In another embodiment, a virus encoding a neoantigen is used for primary immunization, and another virus is used for booster. In another embodiment, the protein is used for primary immunization, and DNA is used for booster. In a preferred embodiment, a DNA vaccine or immunogenic composition is used for the primary T-cell response, and a recombinant viral vaccine or immunogenic composition is used to boost that response. In another preferred embodiment, the viral vaccine or immunogenic composition is co-administered with a protein or DNA vaccine or immunogenic composition used as an adjuvant for the protein or DNA vaccine or immunogenic composition. The patient can then be enhanced with a viral vaccine or immunogenic composition, a protein, or a DNA vaccine or immunogenic composition (see Hutchings et al., Combination of protein and viral vaccines induces potent cellular and humoral immune responses and enhanced protection from murinemalaria challenge. Infect Immun. 2007 Dec; 75(12):5819-26. Electronic publication 1 October 2007).

[0546] Pharmaceutical compositions can be processed using conventional pharmaceutical methods to produce a medicament for administration to patients (including humans and other mammals) in need of it.

[0547] Modification of neoantigen peptides can affect their solubility, bioavailability, and metabolic rate, thereby providing control over the delivery of these active species. Solubility can be assessed by preparing and testing the neoantigen peptides using known methods entirely within the art of those skilled in the art.

[0548] In some embodiments of the pharmaceutical composition, the pharmaceutically acceptable carrier comprises water. In some embodiments, the pharmaceutically acceptable carrier further comprises dextran. In some embodiments, the pharmaceutically acceptable carrier further comprises dimethyl sulfoxide. In some embodiments, the pharmaceutical composition further comprises an immunomodulator or adjuvant. In some embodiments, the immunomodulator or adjuvant is selected from the group consisting of: poly-ICLC, STING agonist, 1018ISS, aluminum salt, Amplivax, AS15, BCG, CP-870, 893, CpG7909, CyaA, dSLIM, GM-CSF, IC30, IC31, imiquimod, ImuFact IMP321, IS Patch, ISS, ISCMATRIX, Juvlmmune, LipoVac, MF59, monophospholipase A, Montanide IMS1312, Montanide ISA 206, Montanide ISA 50V, Montanid ISA-51, OK-432, OM-174, OM-197-MP-EC, ONTAK, PEPTEL, vector system, PLGA microparticles, Remiquimod, SRL172, virions and other virus-like particles, YF-17D, VEGF trap, R848, β-glucan, Pam3Cys, and Aquila's QS21 stimulon. In some embodiments, the immunomodulator or adjuvant includes poly-ICLC.

[0549] Cortexyl derivatives (such as, for example, Vadimezan or AsA404 (also known as 5,6-dimethylcortexyl-4-acetic acid (DMXAA))) can also be used as adjuvants according to embodiments of the invention. Alternatively, such derivatives can also be administered, for example, in parallel with the vaccine or immunogenic composition of the invention via systemic or intratumoral delivery to stimulate immunity at the tumor site. Unbound by theory, these oxanthenone derivatives are believed to act by stimulating interferon (IFN) via IFN gene-stimulating factor (ISTING) receptors (see, for example, Conlon et al. (2013) Mouse, but not Human STING, Binds and Signals in Response to the Vascular Disrupting Agent 5,6-Dimethylxanthenone-4-Acetic Acid, Journal of Immunology, 190:5216-25 and Kim et al. (2013) Anticancer Flavonoids are Mouse-Selective STING Agonists, 8:1396-1401).

[0550] The vaccine or immunological composition may also include adjuvant compounds selected from polymers of acrylic acid or methacrylic acid and copolymers of maleic anhydride and alkenyl derivatives. Specifically, it is a polymer (carbomer) crosslinked with a polyalkenyl ether of acrylic acid or methacrylic acid and a sugar or polyol, specifically crosslinked with allyl sucrose or with allyl pentaerythritol. For example, it may also be a copolymer of maleic anhydride and ethylene crosslinked with divinyl ether (see U.S. Patent No. 6,713,068, which is incorporated herein by reference in its entirety).

[0551] In some embodiments, the pH modifier can stabilize adjuvants or immunomodulators as described herein.

[0552] In some embodiments, the pharmaceutical composition comprises: one to five peptides, dimethyl sulfoxide (DMSO), dextran, water, succinate, poly(I:polyC), poly(L-lysine), carboxymethyl cellulose, and chloride. In some embodiments, each of the one to five peptides is present at a concentration of 300 μg / ml. In some embodiments, the pharmaceutical composition comprises ≤3% DMSO by volume. In some embodiments, the pharmaceutical composition comprises 3.6–3.7% aqueous solution of dextran. In some embodiments, the pharmaceutical composition comprises 3.6–3.7 mM succinate (e.g., sodium succinate) or a salt thereof. In some embodiments, the pharmaceutical composition comprises 0.5 mg / ml poly(I:polyC). In some embodiments, the pharmaceutical composition comprises 0.375 mg / ml poly(L-lysine). In some embodiments, the pharmaceutical composition comprises 1.25 mg / ml sodium carboxymethyl cellulose. In some embodiments, the pharmaceutical composition comprises 0.225% sodium chloride.

[0553] The pharmaceutical composition comprises, optionally in combination with pharmaceutically acceptable additives, carriers, and / or excipients, a therapeutically effective amount of the tumor-specific neoantigen peptide described herein for the treatment of diseases and conditions already described herein (e.g., tumor formation / tumor). Those skilled in the art will recognize from this disclosure and the knowledge of the art that the therapeutically effective amount of one or more compounds according to the invention can vary depending on the condition to be treated, its severity, the treatment regimen to be utilized, the pharmacokinetics of the reagents used, and the patient (animal or human) being treated.

[0554] To prepare the pharmaceutical composition according to the invention, preferably, a therapeutically effective amount of one or more compounds according to the invention is closely mixed with a pharmaceutically acceptable carrier to produce a dose, according to conventional pharmaceutical compounding techniques. The carrier can take a variety of forms depending on the desired formulation form (e.g., ocular, oral, topical, or parenteral), including gels, creams, ointments, lotions, and delayed-release implantable formulations. In preparing the pharmaceutical composition for oral dosage forms, any of the common pharmaceutical media can be used. Thus, for liquid oral formulations (e.g., suspensions, elixirs, and solutions), suitable carriers and additives can be used, including water, glycerin, oils, alcohols, flavoring agents, preservatives, coloring agents, etc. For solid oral formulations (e.g., powders, tablets, capsules) and for solid formulations (e.g., suppositories), suitable carriers and additives can be used, including starch, sugar carriers (e.g., glucose, mannitol, lactose) and related carriers, diluents, granulating agents, lubricants, binders, disintegrants, etc. If desired, tablets or capsules can be enteric-coated or sustained-released using standard techniques.

[0555] The active compound is included in a pharmaceutically acceptable carrier or diluent in an amount sufficient to deliver to a patient a therapeutically effective amount for the desired indication without causing serious toxicity in the patient being treated.

[0556] Oral compositions typically include an inert diluent or an edible carrier. They may be encapsulated in gelatin capsules or compressed into tablets. For oral therapeutic administration, the active compound or its prodrug derivative may be incorporated with excipients and administered in tablet, lozenge, or capsule form. Pharmaceutically compatible binders and / or adjuvant materials may be included as part of the composition.

[0557] Tablets, pills, capsules, lozenges, etc., may contain any of the following ingredients or compounds with similar properties: binders, such as microcrystalline cellulose, astragalus gum, or gelatin; excipients, such as starch or lactose; dispersants, such as alginate or corn starch; lubricants, such as magnesium stearate; glidants, such as colloidal silica; sweeteners, such as sucrose or saccharin; or flavorings, such as peppermint, methyl salicylate, or orange flavoring. When the unit dosage form is a capsule, it may also contain a liquid carrier, such as fatty oil, in addition to the materials discussed herein. Furthermore, the unit dosage form may contain a variety of other materials in physical form that modify the dosage unit, such as coatings of sugar, shellac, or enteric solvents.

[0558] The formulations of the present invention suitable for oral administration can be presented in discrete units, such as capsules, flat capsules or tablets each containing a predetermined amount of active ingredient; in powder or granule form; in solution or suspension in aqueous or non-aqueous liquid; or in oil-in-water or water-in-oil liquid emulsions and in pellet form, etc.

[0559] Tablets can be prepared by compression or molding, optionally with one or more excipients. Compressed tablets can be prepared by compressing the active ingredient in a free-flowing form (such as powder or granules) in a suitable machine, optionally mixed with a binder, lubricant, inert diluent, preservative, surfactant, or dispersant. Molded tablets can be prepared by molding a mixture of powdered compounds wetted with an inert liquid diluent in a suitable machine. Tablets can optionally be coated or scored and can be formulated to provide a slow or controlled release of the active ingredient therein.

[0560] Methods for formulating such slow- or controlled-release compositions of pharmaceutical active ingredients are known in the art and described in several published U.S. patents, some of which include, but are not limi...

Claims

1. A composition for manufacturing a medicament for treating GATA-3-related cancers, said composition comprising: (i) An isolated neoantigen polypeptide comprising at least one tumor-specific neoepitope, wherein the tumor-specific neoepitope is an amino acid sequence of the following: CLQCLWALL (SEQ ID NO: 13809), QWGPCLQCL (SEQ ID NO: 13810), TTTTTLWRL (SEQ ID NO: 13811), WALLQASQY (SEQ ID NO: 13812), EECQWGPCL (SEQ ID NO: 13813), QWGPCLQCLW (SEQ ID NO: 13814), LWALLQASQY (SEQ ID NO: 13815), QTTTTTLWRL (SEQ ID NO: 13816), CQWGPCLQCL (SEQ ID NO: 13817) or LQCLWALLQA (SEQ ID NO: 13818), wherein the neoantigen polypeptide is 8 to 50 amino acids in length. (ii) The nucleic acid encoding (i), (iii) Antigen-presenting cells containing (i) or (ii), or Use T cells stimulated by (iii).

2. The composition of claim 1, wherein the tumor-specific neoepitope binds to a protein encoded by an HLA allele.

3. The composition according to claim 2, wherein the tumor-specific neoepitope is at a K0.05 of less than 500 nM. D It binds to proteins encoded by HLA-A, HLA-B, or HLA-C alleles.

4. The composition according to claim 2, wherein the tumor-specific neoepitope is at a K0 of less than 1000 nM. D It binds to proteins encoded by class II HLA alleles.

5. The composition of claim 1, wherein the tumor-specific neoepitope is derived from a tumor-specific GATA3 frameshift mutation.

6. The composition according to claim 1, wherein the composition comprises (ii) a nucleic acid encoding (i).

7. The composition of claim 1, wherein the composition comprises (iv) T cells excited by antigen-presenting cells comprising (i) or (ii).

8. The composition according to claim 1, further comprising an adjuvant or an immunomodulator.

9. The composition according to claim 8, wherein the immunomodulator or adjuvant is poly(I:C).

10. The composition according to claim 1, wherein the tumor-specific novel epitope is the amino acid sequence CLQCLWALL (SEQ ID NO: 13809).

11. The composition according to claim 1, wherein the tumor-specific novel epitope is the amino acid sequence QWGPCLQCL (SEQ ID NO: 13810).

12. The composition according to claim 1, wherein the tumor-specific novel epitope is the amino acid sequence TTTTTLWRL (SEQ ID NO: 13811).

13. The composition according to claim 1, wherein the tumor-specific novel epitope is the amino acid sequence WALLQASQY (SEQ ID NO: 13812).

14. The composition according to claim 1, wherein the tumor-specific novel epitope is the amino acid sequence EECQWGPCL (SEQ ID NO: 13813).

15. The composition according to claim 1, wherein the tumor-specific novel epitope is the amino acid sequence QWGPCLQCLW (SEQ ID NO: 13814).

16. The composition according to claim 1, wherein the tumor-specific novel epitope is the amino acid sequence LWALLQASQY (SEQ ID NO: 13815).

17. The composition according to claim 1, wherein the tumor-specific novel epitope is the amino acid sequence QTTTTTLWRL (SEQ ID NO: 13816).

18. The composition according to claim 1, wherein the tumor-specific novel epitope is the amino acid sequence CQWGPCLQCL (SEQ ID NO: 13817).

19. The composition according to claim 1, wherein the tumor-specific novel epitope is the amino acid sequence LQCLWALLQA (SEQ ID NO: 13818).

20. Use of the composition according to any one of claims 1 to 19 in the preparation of a medicament for treating or preventing breast cancer in a subject of need.

21. The use according to claim 20, wherein the subject has a GATA3 frameshift mutation.

22. The use according to claim 21, wherein the GATA frameshift mutation is the GATA3:p.L328 frameshift mutation.

23. The use according to claim 20, wherein the breast cancer is non-metastatic breast cancer, metastatic breast cancer, HER-2 positive breast cancer, stage I breast cancer, stage II breast cancer, stage IIA breast cancer, or operable stage IIC breast cancer.

24. The use according to claim 20, wherein the composition is administered in combination with an additional cancer therapy, wherein the additional cancer therapy is a checkpoint inhibitor or surgery, chemotherapy, or targeted therapy.

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