Calculation method for selecting individualized neoantigen vaccines

Abstract

A system and method for identifying personalized tumor vaccines are provided. A target somatic variant is determined, and RNA sequence reads are obtained from the target tumor. From the RNA sequence reads, fusion proteins are determined, each fusion protein being a fusion of a portion of a first protein and a portion of a second protein. Candidate nascent antigens are selected, including a first subset of candidate nascent antigens encoding a somatic variant, and a second subset of candidate nascent antigens encoding residues from the first and second protein portions. A score is determined for each candidate nascent antigen using a scoring function that includes a first scoring term that increases the weighting of candidate nascent antigens with relatively high class I MHC affinity, taking into account the target class I HLA type. Based on their respective scores, two or more candidate nascent antigens are selected as the final set of nascent antigens for the tumor vaccine.

Claims

1. A method for identifying tumor vaccines personalized for humans with cancer, wherein the method is (A) Determining the first set of somatic cell variants of the subject, (B) Obtaining a first set of sequence reads from RNA molecules in a tumor sample obtained from the subject, (C) Determining one or more fusion proteins encoded by the first plurality of sequence reads from the first plurality of sequence reads, wherein each of the one or more fusion proteins is a fusion of a portion of a first human protein and a portion of a second human protein. (D) Selecting a plurality of candidate neonatal antigens, including a first subset of candidate neonatal antigens and a second subset of candidate neonatal antigens, Each nascent antigen in the first subset of candidate nascent antigens encodes a somatic variant in the first plurality of somatic variants, The selection is such that each nascent antigen in the second subset of the candidate nascent antigens codes for one or more residues from the portion of the respective first human protein and one or more residues from the portion of the respective second human protein. (E) Determining the respective scores for each of the multiple candidate nascent antigens using a scoring function that includes a first scoring term that increases the weighting of each candidate nascent antigen having a higher class I major histocompatibility complex (MHC) affinity to each candidate nascent antigen having a lower class I major histocompatibility complex (MHC) affinity, taking into account the class I human leukocyte antigen (HLA) type of the human subject, (F) A method for the tumor vaccine comprising selecting two or more candidate neonatal antigens from a plurality of candidate neonatal antigens as the final set of neonatal antigens based on the respective scores of each candidate neonatal antigen in the plurality of candidate neonatal antigens.

2. The method described above is The method further includes determining the allele-specific expression of each somatic variant in the first plurality of variants using the first plurality of sequence reads, The method according to claim 1, further comprising a second scoring term which increases the weighting of each candidate nascent antigen representing an allele that is more abundant in the first sequence reads than each candidate nascent antigen representing an allele that is less abundant in the first sequence reads.

3. Determining the first plurality of somatic cell variants of the subject is To obtain a second set of sequence reads from DNA molecules in the tumor sample obtained from the subject, To obtain a third set of sequence reads from DNA molecules in a normal sample obtained from the aforementioned subject, The method according to claim 1, comprising identifying the first plurality of somatic cell variants of interest by using the second plurality of sequence reads and the third plurality of sequence reads to include in the first plurality of somatic cell variants those somatic cell variants observed in the second plurality of sequence reads but not observed in the third plurality of sequence reads.

4. The method described above is The first exome sequencing is performed with at least 100x coverage to obtain the second set of sequence reads, The method according to claim 3, further comprising performing a second exome sequencing with at least 100 times coverage to obtain the second plurality of sequence reads.

5. The method according to claim 3 or 4, wherein the normal sample is a tissue sample located close to the original location of the tumor in the subject.

6. The method according to claim 3 or 4, wherein the normal sample is a blood sample.

7. The method according to any one of claims 1 to 6, wherein the cancer is glioblastoma or prostate cancer.

8. The method according to any one of claims 1 to 7, wherein the cancer is a carcinoma, melanoma, lymphoma / leukemia, sarcoma, or neuroglial tumor.

9. The method according to any one of claims 1 to 8, wherein the cancer is lung cancer, pancreatic cancer, colon cancer, stomach cancer or esophageal cancer, breast cancer, ovarian cancer, prostate cancer, or liver cancer.

10. The method according to any one of claims 1 to 9, wherein each somatic cell variant in the first plurality of somatic cell variants is a single nucleotide variant or an indel mutation.

11. The method according to any one of claims 1 to 10, wherein the sample of the tumor is a fresh frozen sample.

12. The method according to any one of claims 1 to 11, wherein each of the candidate nascent antigens in the plurality of candidate nascent antigens has the length of an N residue, and N is 8, 9, 10, 11, 12, 13, or 14.

13. The method according to any one of claims 1 to 11, wherein each of the candidate nascent antigens in the plurality of candidate nascent antigens has a length of 9 residues.

14. The method according to any one of claims 1 to 13, further comprising excluding from the plurality of candidate nascent antigens, or from the final set of candidate nascent antigens, any candidate nascent antigens that include a mutation from the wild-type sequence at the N-terminal residue position.

15. The method according to any one of claims 1 to 14, further comprising determining the class I HLA type of the human subject using the first plurality of sequencing reads.

16. The method according to any one of claims 1 to 14, further comprising determining the class I HLA type of the human subject using a polymerase chain reaction with respect to a biological sample from the subject of cancer.

17. The method described above is The process further includes determining the hydrophobicity of each of the candidate nascent antigens in the aforementioned plurality of candidate nascent antigens, The method according to any one of claims 1 to 16, wherein the scoring function further comprises a scoring term for the hydrophobicity of each of the candidate nascent antigens.

18. The method according to claim 17, wherein determining the hydrophobicity of each of the plurality of candidate nascent antigens includes assigning to each of the candidate nascent antigens the maximum hydrophobic score of any 7-mer within each of the candidate nascent antigens.

19. The method according to claim 17 or 18, wherein the scoring term for the hydrophobicity of each candidate nascent antigen increases the weighting of a more hydrophobic candidate nascent antigen relative to a less hydrophobic candidate nascent antigen.

20. The method according to any one of claims 1 to 19, wherein the scoring function further includes a term for the class II MHC affinity of each candidate nascent antigen, taking into account the class II HLA type of the human subject, the term increasing the weighting of each candidate nascent antigen having a higher class II MHC affinity than each candidate nascent antigen having a lower class II MHC affinity.

21. The method described above is The process further includes verifying multiple indel mutations present in the tumor sample using the first set of sequence reads, The plurality of candidate neonatal antigens include a third subset of candidate neonatal antigens, The method according to any one of claims 1 to 20, wherein each nascent antigen in the third subset of the plurality of candidate nascent antigens encodes all or part of the indel mutations in the plurality of indel mutations.

22. The first plurality of array reads are 1 × 10 ６ The method according to any one of claims 1 to 21, comprising a sequence read.

23. The method according to any one of claims 1 to 22, wherein the plurality of candidate nascent antigens comprises five candidate nascent antigens.

24. The method according to any one of claims 1 to 23, wherein the final set of the nascent antigens consists of 2 to 20 nascent antigens from the plurality of candidate nascent antigens.

25. The method according to any one of claims 1 to 24, wherein the final set of nascent antigens comprises one or more nascent antigens from the first subset of the plurality of candidate nascent antigens and one or more nascent antigens from the second subset of the plurality of candidate nascent antigens.

26. The method according to claim 21, wherein the final set of nascent antigens comprises one or more nascent antigens from the first subset of the plurality of candidate nascent antigens, one or more nascent antigens from the second subset of the plurality of candidate nascent antigens, and one or more nascent antigens from the third subset of the plurality of candidate nascent antigens.

27. The method described above is The tumor vaccine is formed by solubilizing the final set of the newly generated antigens in a carrier, The method according to any one of claims 1 to 26, further comprising administering the vaccine to the human subject.

28. The method according to claim 27, further comprising forming the tumor vaccine with polyICLC.

29. The method according to claim 27 or 28, wherein the administration is repeated multiple times over a period of several months.

30. A system for identifying tumor vaccines that are personalized for individuals with cancer, Memory and One or more processors, The system comprises one or more modules, which are stored in the memory and configured for execution by one or more processors, and the one or more modules are (A) Determining the first set of somatic cell variants of the subject, (B) Obtaining a first set of sequence reads from RNA molecules in a tumor sample obtained from the subject in electronic form, (C) Determining one or more fusion proteins encoded by the first plurality of sequence reads from the first plurality of sequence reads, wherein each of the one or more fusion proteins is a fusion of a portion of a first human protein and a portion of a second human protein. (D) Selecting a plurality of candidate neonatal antigens, including a first subset of candidate neonatal antigens and a second subset of candidate neonatal antigens, Each nascent antigen in the first subset of candidate nascent antigens encodes a somatic variant in the first plurality of somatic variants, The selection is such that each nascent antigen in the second subset of the candidate nascent antigens codes for one or more residues from the portion of the respective first human protein and one or more residues from the portion of the respective second human protein. (E) Determining the respective scores for each of the multiple candidate nascent antigens using a scoring function that includes a first scoring term that increases the weighting of each candidate nascent antigen having a higher class I major histocompatibility complex (MHC) affinity to each candidate nascent antigen having a lower class I major histocompatibility complex (MHC) affinity, taking into account the class I human leukocyte antigen (HLA) type of the human subject, (F) A system including instructions for performing the following: selecting two or more candidate neonatal antigens from a plurality of candidate neonatal antigens as the final set of neonatal antigens based on the respective scores of each candidate neonatal antigen in the plurality of candidate neonatal antigens for the tumor vaccine.

31. A non-temporary computer-readable storage medium for identifying tumor vaccines personalized for humans with cancer, wherein the non-temporary computer-readable storage medium stores one or more programs for execution by one or more processors of a computer system, and the one or more computer programs (A) Determining the first set of somatic cell variants of the subject, (B) Obtaining a first set of sequence reads from RNA molecules in a tumor sample obtained from the subject in electronic form, (C) Determining one or more fusion proteins encoded by the first plurality of sequence reads from the first plurality of sequence reads, wherein each of the one or more fusion proteins is a fusion of a portion of a first human protein and a portion of a second human protein. (D) Selecting a plurality of candidate neonatal antigens, including a first subset of candidate neonatal antigens and a second subset of candidate neonatal antigens, Each nascent antigen in the first subset of candidate nascent antigens encodes a somatic variant in the first plurality of somatic variants, The selection is such that each nascent antigen in the second subset of the candidate nascent antigens codes for one or more residues from the portion of the respective first human protein and one or more residues from the portion of the respective second human protein. (E) Determining the respective scores for each of the multiple candidate nascent antigens using a scoring function that includes a first scoring term that increases the weighting of each candidate nascent antigen having a higher class I major histocompatibility complex (MHC) affinity to each candidate nascent antigen having a lower class I major histocompatibility complex (MHC) affinity, taking into account the class I human leukocyte antigen (HLA) type of the human subject, (F) A non-temporary computer-readable storage medium containing instructions for selecting two or more candidate neonatal antigens from a plurality of candidate neonatal antigens as the final set of neonatal antigens, based on the respective scores of each candidate neonatal antigen in the plurality of candidate neonatal antigens for the tumor vaccine.

32. A tumor vaccine comprising multiple newly synthesized antigen peptides and adjuvants, which are individualized for subjects suffering from cancer, The first neonatal antigen in the tumor vaccine encodes a single nucleotide polymorphism present in the RNA molecule in the tumor biopsy obtained from the subject. A tumor vaccine in which a second neonatal antigen encodes one or more residues from a portion of a first human protein and one or more residues from a portion of a second human protein, and the tumor biopsy comprises an RNA molecule which is a fusion of the first human protein and the second human protein.

33. The tumor vaccine according to claim 32, wherein the cancer is prostate cancer, and the second neogenic antigen encodes a fusion product shown in Table 1.

34. The tumor vaccine according to claim 32 or 33, wherein the cancer is prostate cancer and the single nucleotide polymorphism is SPOP, TP53, FOXA1, or PTEN.

35. The tumor vaccine according to any one of claims 32 to 34, wherein the cancer is a carcinoma, melanoma, lymphoma / leukemia, sarcoma, or neuroglial tumor.

36. The tumor vaccine according to any one of claims 32 to 35, wherein the cancer is lung cancer, pancreatic cancer, colon cancer, stomach cancer or esophageal cancer, breast cancer, ovarian cancer, prostate cancer, or liver cancer.

37. The tumor vaccine according to any one of claims 32 to 36, wherein the tumor biopsy is a fresh frozen sample.

38. The tumor vaccine according to any one of claims 32 to 37, wherein each of the multiple nascent antigen peptides has the length of an N residue, and N is 8, 9, 10, 11, 12, 13, or 14.

39. The tumor vaccine according to any one of claims 32 to 38, wherein each of the multiple nascent antigen peptides has a length of 9 residues.

40. The tumor vaccine according to any one of claims 32 to 39, wherein each of the plurality of newly synthesized antigen peptides does not contain a mutation from the wild-type sequence at the N-terminal residue position.

41. The tumor vaccine according to any one of claims 32 to 40, wherein the plurality of newly synthesized antigen peptides are solubilized in a carrier.

42. The tumor vaccine according to any one of claims 32 to 41, wherein the adjuvant is poly-ICLC.

43. A method comprising administering the tumor vaccine described in any one of claims 32 to 42 to the subject.

44. The method according to claim 43, wherein the administration is repeated multiple times over a period of several months.

45. A method for identifying tumor vaccines personalized for humans with cancer, wherein the method is (A) Determining the first set of somatic cell variants of the subject, (B) Selecting a plurality of candidate nascent antigens such that each nascent antigen in at least a subset of the candidate nascent antigens encodes a somatic variant in the first plurality of somatic variants. (C) Using a scoring function to determine the score for each of the candidate neonatal antigens in the plurality of candidate neonatal antigens, wherein the scoring function is A first scoring term that, taking into account the class I human leukocyte antigen (HLA) type of the human subject, increases the weighting of candidate nascent antigens with higher class I major histocompatibility complex (MHC) affinity compared to candidate nascent antigens with lower class I major histocompatibility complex (MHC) affinity, and The determination includes a second scoring term that increases the weighting of candidate nascent antigens with higher hydrophobicity compared to candidate nascent antigens with lower hydrophobicity, (D) A method for the tumor vaccine, comprising selecting two or more candidate neonatal antigens from the plurality of candidate neonatal antigens as the final set of neonatal antigens based on the respective scores of each candidate neonatal antigen in the plurality of candidate neonatal antigens.

46. After the above decision (A), To obtain a first set of sequence reads from RNA molecules in tumor samples obtained from the aforementioned subject, The method further includes determining the allele-specific expression of each somatic variant in the first plurality of variants using the first plurality of sequence reads, The method according to claim 45, wherein the scoring function further includes a third scoring term that increases the weighting of each candidate nascent antigen representing an allele that is more abundant in the first sequence reads than each candidate nascent antigen representing an allele that is less abundant in the first sequence reads.

47. Determining one or more fusion proteins encoded by the first plurality of sequence reads from the first plurality of sequence reads, further comprising determining that each of the one or more fusion proteins is a fusion of a portion of a first human protein and a portion of a second human protein, The method according to claim 46, wherein the plurality of candidate nascent antigens further comprises a second subset of candidate nascent antigens, each nascent antigen in the second subset of candidate nascent antigens codes for one or more residues from the respective portion of the first human protein and one or more residues from the respective portion of the second human protein.

48. The method according to any one of claims 45 to 47, wherein the cancer is glioblastoma or prostate cancer.

49. The method according to any one of claims 45 to 48, wherein the cancer is a carcinoma, melanoma, lymphoma / leukemia, sarcoma, or neuroglial tumor.

50. The method according to any one of claims 45 to 49, wherein the cancer is lung cancer, pancreatic cancer, colon cancer, stomach cancer or esophageal cancer, breast cancer, ovarian cancer, prostate cancer, or liver cancer.

51. The method according to any one of claims 45 to 50, wherein each somatic cell variant in the first plurality of somatic cell variants is a single nucleotide variant or an indel mutation.

52. The method according to claim 46 or 47, wherein the tumor biopsy is a fresh frozen sample.

53. The method according to any one of claims 45 to 52, wherein each candidate nascent antigen in the plurality of candidate nascent antigens has the length of an N residue, and N is 8, 9, 10, 11, 12, 13, or 14.

54. The method according to any one of claims 45 to 53, wherein each candidate nascent antigen in the plurality of candidate nascent antigens has a length of 9 residues.

55. The method according to any one of claims 45 to 54, further comprising excluding from the plurality of candidate nascent antigens, or from the final set of candidate nascent antigens, those candidate nascent antigens having a mutation from the wild-type sequence at the N-terminal residue position.

56. The method described above is The tumor vaccine is formed by solubilizing the final set of the newly generated antigens in a carrier, The method according to any one of claims 45 to 55, further comprising administering the vaccine to the human subject.

57. The method according to claim 56, further comprising forming the tumor vaccine with polyICLC.

58. The method according to claim 56 or 57, wherein the administration is repeated multiple times over a period of several months.

59. A method for identifying tumor vaccines personalized for humans with cancer, wherein the method is (A) Obtaining a first set of sequence reads from RNA molecules in a tumor sample obtained from the subject, (B) Obtaining a second set of sequence reads from the DNA molecules in the tumor sample obtained from the subject, (C) Obtaining a third set of sequence reads from DNA molecules in a normal tissue sample adjacent to the original location of the tumor in the subject, (D) Identifying the target plurality of somatic cell variants by using the second plurality of sequence reads and the third plurality of sequence reads to include somatic cell variants that are observed in the second plurality of sequence reads but not in the third plurality of sequence reads, (E) Selecting a plurality of candidate nascent antigens such that each nascent antigen in at least a subset of the candidate nascent antigens encodes a somatic variant in the plurality of somatic variants. (F) Determining the respective scores for each of the multiple candidate nascent antigens using a scoring function that includes a first scoring term that increases the weighting of candidate nascent antigens having a higher class I major histocompatibility complex (MHC) affinity compared to candidate nascent antigens having a lower class I major histocompatibility complex (MHC) affinity, taking into account the class I human leukocyte antigen (HLA) type of the human subject, (G) A method for the tumor vaccine, comprising selecting two or more candidate neonatal antigens from the plurality of candidate neonatal antigens as the final set of neonatal antigens based on the respective scores of each candidate neonatal antigen in the plurality of candidate neonatal antigens.

60. Determining one or more fusion proteins encoded by the first plurality of sequence reads from the first plurality of sequence reads, further comprising determining that each of the one or more fusion proteins is a fusion of a portion of a first human protein and a portion of a second human protein, The method according to claim 59, wherein the plurality of candidate nascent antigens further comprises a second subset of candidate nascent antigens, each nascent antigen in the second subset of candidate nascent antigens codes for one or more residues from the respective portion of the first human protein and one or more residues from the respective portion of the second human protein.

61. The method further includes determining the allele-specific expression of each somatic variant in the first plurality of variants using the first plurality of sequence reads, The method according to claim 59 or 60, wherein the scoring function further includes a second scoring term that increases the weighting of each candidate nascent antigen representing an allele that is more abundant in the first sequence reads than each candidate nascent antigen representing an allele that is less abundant in the first sequence reads.

62. The method according to any one of claims 59 to 61, wherein the cancer is glioblastoma or prostate cancer.

63. The method according to any one of claims 59 to 62, wherein the cancer is a carcinoma, melanoma, lymphoma / leukemia, sarcoma, or neuroglial tumor.

64. The method according to any one of claims 59 to 63, wherein the cancer is lung cancer, pancreatic cancer, colon cancer, stomach cancer or esophageal cancer, breast cancer, ovarian cancer, prostate cancer, or liver cancer.

65. The method according to any one of claims 59 to 64, wherein each somatic cell variant in the first plurality of somatic cell variants is a single nucleotide variant or an indel mutation.

66. The method according to any one of claims 59 to 65, wherein the sample of the tumor is a fresh frozen sample.

67. The method according to any one of claims 59 to 66, wherein each candidate nascent antigen in the plurality of candidate nascent antigens has the length of an N residue, and N is 8, 9, 10, 11, 12, 13, or 14.

68. The method according to any one of claims 59 to 67, wherein each candidate nascent antigen in the plurality of candidate nascent antigens has a length of 9 residues.

69. The method according to any one of claims 59 to 68, further comprising excluding from the plurality of candidate nascent antigens, or from the final set of candidate nascent antigens, any candidate nascent antigens having a mutation from the wild-type sequence at the N-terminal residue position.

70. The method described above is The tumor vaccine is formed by solubilizing the final set of the newly generated antigens in a carrier, The method according to any one of claims 59 to 69, further comprising administering the vaccine to the human subject.

71. The method according to claim 70, further comprising forming the tumor vaccine with polyICLC.

72. The method according to claim 70 or 71, wherein the administration is repeated multiple times over a period of several months.

73. A method for identifying tumor vaccines personalized for humans with cancer, wherein the method is (A) Determining the first set of somatic cell variants of the subject, (B) Selecting a plurality of candidate nascent antigens such that each nascent antigen in at least a subset of the candidate nascent antigens encodes a somatic variant in the plurality of somatic variants, (C) Using a scoring function to determine the score for each of the candidate neonatal antigens in the plurality of candidate neonatal antigens, wherein the scoring function is A first scoring term that, taking into account the class I human leukocyte antigen (HLA) type of the human subject, increases the weighting of each candidate nascent antigen having a higher class I major histocompatibility complex (MHC) affinity compared to each candidate nascent antigen having a lower class I major histocompatibility complex (MHC) affinity, and The determination includes a second scoring term that, taking into account the class II HLA types of the human subject, increases the weighting of each candidate nascent antigen with higher class II MHC affinity compared to each candidate nascent antigen with lower class II MHC affinity, (D) A method for the tumor vaccine, comprising selecting two or more candidate neonatal antigens from the plurality of candidate neonatal antigens as the final set of neonatal antigens based on the respective scores of each candidate neonatal antigen in the plurality of candidate neonatal antigens.

74. After the above decision (A), To obtain a first set of sequence reads from RNA molecules in tumor samples obtained from the aforementioned subject, The method further includes determining the allele-specific expression of each somatic variant in the first plurality of variants using the first plurality of sequence reads, The method according to claim 73, further comprising a third scoring term that increases the weighting of each candidate nascent antigen representing an allele that is more abundant in the first sequence reads than each candidate nascent antigen representing an allele that is more abundant in the first sequence reads.

75. Determining one or more fusion proteins encoded by the first plurality of sequence reads from the first plurality of sequence reads, further comprising determining that each of the one or more fusion proteins is a fusion of a portion of a first human protein and a portion of a second human protein, The method according to claim 74, wherein the plurality of candidate nascent antigens further comprises a second subset of candidate nascent antigens, each nascent antigen in the second subset of candidate nascent antigens codes for one or more residues from the respective portion of the first human protein and one or more residues from the respective portion of the second human protein.

76. The method according to any one of claims 73 to 75, wherein the cancer is glioblastoma or prostate cancer.

77. The method according to any one of claims 73 to 76, wherein the cancer is a carcinoma, melanoma, lymphoma / leukemia, sarcoma, or neuroglial tumor.

78. The method according to any one of claims 73 to 77, wherein the cancer is lung cancer, pancreatic cancer, colon cancer, stomach cancer or esophageal cancer, breast cancer, ovarian cancer, prostate cancer, or liver cancer.

79. The method according to any one of claims 73 to 78, wherein each somatic cell variant in the first plurality of somatic cell variants is a single nucleotide variant or an indel mutation.

80. The method according to claim 74 or 75, wherein the sample of the tumor is a fresh frozen sample.

81. The method according to any one of claims 73 to 80, wherein each candidate nascent antigen in the plurality of candidate nascent antigens has the length of an N residue, and N is 8, 9, 10, 11, 12, 13, or 14.

82. The method according to any one of claims 73 to 81, wherein each candidate nascent antigen in the plurality of candidate nascent antigens has a length of 9 residues.

83. The method according to any one of claims 73 to 82, further comprising excluding from the plurality of candidate nascent antigens, or from the final set of candidate nascent antigens, any candidate nascent antigens having a mutation from the wild-type sequence at the N-terminal residue position.

84. The method described above is The tumor vaccine is formed by solubilizing the final set of the newly generated antigens in a carrier, The method according to any one of claims 73 to 83, further comprising administering the vaccine to the human subject.

85. The method according to claim 84, further comprising forming the tumor vaccine with polyICLC.

86. The method according to claim 84 or 85, wherein the administration is repeated multiple times over a period of several months.

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