RNA CANCER VACCINES

1 . A method of treating a cancer in a human subject comprising:

administering to the human subject multiple doses of a mRNA cancer vaccine composition at a dosage of 0.04-0.13 mg, 0.13-0.39 mg, or 0.39-1.0 mg, 1.0-5.0 mg wherein the mRNA cancer vaccine composition comprises one or more mRNAs each having one or more open reading frames encoding 3-50 peptide epitopes, wherein each of the peptide epitopes are portions of personalized cancer antigens or portions of cancer hotspot antigens, formulated in a lipid nanoparticle formulation, thereby treating the cancer in the human subject.

2 . The method of claim 1 , wherein at least 3 doses of the cancer vaccine composition are administered to the subject.

3 . The method of claim 1 , wherein at least 5 doses of the cancer vaccine composition are administered to the subject.

4 . The method of claim 1 , wherein at least 9 doses of the cancer vaccine composition are administered to the subject.

5 . The method of any one of claims 1 - 4 , wherein the multiple doses of the cancer vaccine composition are administered 20-22 days apart.

6 . The method of any one of claims 1 - 4 , wherein the multiple doses of the cancer vaccine composition are administered 21 days apart.

7 . The method of any one of claims 1 - 6 , wherein the cancer vaccine composition comprises one mRNA having one open reading frame encoding 30-35 peptide epitopes.

8 . The method of any one of claims 1 - 6 , wherein the cancer vaccine composition comprises one mRNA having one open reading frame encoding 34 peptide epitopes.

9 . The method of any one of claims 1 - 6 , wherein the cancer vaccine composition comprises a first and second mRNA, wherein the first mRNA has one open reading frame encoding 10-20 peptide epitopes which are portions of personalized cancer antigens and wherein the second mRNA has one open reading frame encoding peptide epitopes that are portions of cancer hotspot antigens.

10 . The method of claim 9 , wherein the cancer hotspot antigens comprise a KRAS G12 mutation or a KRAS G13 mutation or both mutations.

11 . The method of any one of claims 1 - 10 , wherein the cancer vaccine composition is administered at a dosage of 0.04 mg to 2 mg.

12 . The method of any one of claims 1 - 10 , wherein the cancer vaccine composition is administered at a dosage of 0.04 mg to 1 mg.

13 . The method of any one of claims 1 - 10 , wherein the cancer vaccine composition is administered at a dosage of 0.39 mg to 2 mg.

14 . The method of any one of claims 1 - 10 , wherein the cancer vaccine composition is administered at a dosage of 0.39 mg to 1 mg.

15 . The method of any one of claims 1 - 14 , wherein the minimum length of any peptide epitope is 8 amino acids.

16 . The method of any one of claims 1 - 15 , wherein the maximum length of any peptide epitope is 31 amino acids.

17 . The method of claim 1 , wherein the one or more mRNA each comprise a 5′ UTR and/or a 3′ UTR.

18 . The method of any one of claims 1 - 17 , wherein the one or more mRNA comprise at least one chemical modification.

19 . The method of claim 18 , wherein the chemical modification is selected from the group consisting of pseudouridine, N1-methylpseudouridine, N1-ethylpseudouridine, 2-thiouridine, 4′-thiouridine, 5-methylcytosine, 2-thio-1-methyl-1-deaza-pseudouridine, 2-thio-1-methyl-pseudouridine, 2-thio-5-aza-uridine, 2-thio-dihydropseudouridine, 2-thio-dihydrouridine, 2-thio-pseudouridine, 4-methoxy-2-thio-pseudouridine, 4-methoxy-pseudouridine, 4-thio-1-methyl-pseudouridine, 4-thio-pseudouridine, 5-aza-uridine, dihydropseudouridine, 5-methyluridine, 5-methyluridine, 5-methoxyuridine, and 2′-O-methyl uridine.

20 . The method of claim 18 or claim 19 , wherein the one or more mRNA is fully modified.

21 . The method of any one of claims 1 - 6 and 8 - 20 , wherein the peptide epitopes are in the form of a concatemeric cancer antigen comprised of 30-40 peptide epitopes.

22 . The method of claim 21 , wherein the peptide epitopes are interspersed by cleavage sensitive sites.

23 . The method of claim 21 , wherein each peptide epitope is linked directly to one another without a linker.

24 . The method of claim 21 , wherein each peptide epitope is linked to one another with a single amino acid linker.

25 . The method of claim 21 , wherein each peptide epitope is linked to one another with a short peptide linker.

26 . The method of claim 21 , wherein each peptide epitope includes one or more SNP mutations; and/or a mutation causing a unique expressed peptide sequence.

27 . The method of claim 21 , wherein the mRNA encoding the peptide epitopes is arranged such that the peptide epitopes are ordered to minimize pseudo-epitopes.

28 . The method of claim 21 , wherein the ratio of class I MHC molecule peptide epitopes to class II MHC molecule peptide epitopes is at least 1:1, 2:1, 3:1, 4:1, or 5:1.

29 . The method of any one of claims 1 - 28 , wherein the lipid nanoparticle comprises an ionizable amino lipid.

30 . The method of claim 29 , wherein the lipid nanoparticle formulation comprises a molar ratio of about 20-60% ionizable amino lipid, about 5-25% non-cationic lipid, about 25-55% sterol, and about 0.5-15% PEG-modified lipid.

31 . The method of claim 29 , wherein the lipid nanoparticle formulation comprises an ionizable amino lipid of Compound 1, wherein the non-cationic lipid is DSPC, the structural lipid is cholesterol, and the PEG lipid is PEG-DMG.

32 . The method of any one of claims 1 - 31 , wherein the cancer vaccine composition prevents the human subject's cancer from recurring by provoking the immune system to develop a memory immune response against a cancerous tissue derived from an original lesion of the cancer.

33 . The method of any one of claims 1 - 32 , wherein the vaccine composition is administered via intramuscular injection.

34 . The method of claim 33 , wherein the vaccine composition is administered in 2 or more injections.

35 . The method of claim 33 , wherein the vaccine composition is administered in a single injection.

36 . The method of any one of claims 1 - 35 , wherein the human subject has a favorable response to the treatment method based on RECIST (Response Evaluation Criteria In Solid Tumors).

37 . The method of any one of claims 1 - 35 , wherein the human subject has a favorable response to the treatment method based on irRECIST (Immune-related Response Evaluation Criteria In Solid Tumors).

38 . The method of any one of claims 1 - 35 , wherein the personalized cancer antigens are selected based on a next generation sequencing (NGS) analysis of the human subjects DNA from a tumor sample, relative to DNA from a blood sample.

39 . A method of treating a cancer in a human subject comprising:

administering to the human subject a combination therapy comprising 1) multiple doses of a mRNA cancer vaccine composition at a dosage of 0.04-0.13 mg, 0.13-0.39 mg, or 0.39-1.0 mg, 1.0-5.0 mg, wherein the cancer vaccine composition comprises one or more mRNAs each having one or more open reading frames encoding 3-50 peptide epitopes, wherein each of the peptide epitopes are portions of personalized cancer antigens or portions of cancer hotspot antigens, formulated in a lipid nanoparticle formulation, and 2) multiple doses of an anti-cancer immunotherapy thereby treating the cancer in the human subject with the combination therapy.

40 . The method of claim 39 , wherein the human subject is administered at least 0.04 mg of the vaccine composition.

41 . The method of claim 39 , wherein the human subject is administered at least 0.13 mg of the vaccine composition.

42 . The method of claim 39 , wherein the human subject is administered at least 0.39 mg of the vaccine composition.

43 . The method of claim 39 , wherein the human subject is administered at least 1.0 mg of the vaccine composition.

44 . The method of any one of claims 39 - 43 , wherein the human subject is administered less than 2.0 mg of the vaccine composition.

45 . The method of any one of claims 39 - 44 , wherein the lipid nanoparticle formulation comprises an ionizable amino lipid nanoparticle.

46 . The method of claim 45 , wherein the lipid nanoparticle formulation comprises a molar ratio of about 20-60% ionizable amino lipid, about 5-25% non-cationic lipid, about 25-55% sterol, and about 0.5-15% PEG-modified lipid.

47 . The method of claim 46 , wherein the lipid nanoparticle formulation comprises an ionizable amino lipid of Compound 1, wherein the non-cationic lipid is DSPC, the structural lipid is cholesterol, and the PEG lipid is PEG2000-DMG.

48 . The method of any one of claims 39 - 47 , wherein the anti-cancer immunotherapy is a checkpoint inhibitor.

49 . The method of claim 48 , wherein the checkpoint inhibitor is pembrolizumab.

50 . The method of any one of claims 39 - 49 , wherein the cancer vaccine composition is administered to the subject twice, thrice, four times, five times, six times, seven times, eight times, nine times, or 10 times.

51 . The method of any one of claims 39 - 50 , wherein the cancer vaccine composition is administered via intramuscular injection.

52 . The method of any one of claims 39 - 51 , wherein the anti-cancer immunotherapy is administered to the human subject at least 2 times.

53 . The method of any one of claims 39 - 51 , wherein the anti-cancer immunotherapy is administered to the human subject at least 3 times.

54 . The method of any one of claims 39 - 51 , wherein the anti-cancer immunotherapy is administered to the human subject at least 9 times.

55 . The method of claim 52 , wherein the anti-cancer immunotherapy is administered to the human subject at least 2 times prior to the administration of the cancer vaccine composition.

56 . The method of claim 53 , wherein the third, fourth or fifth dose of anti-cancer immunotherapy is administered to the human subject on the same day as a first dose of the cancer vaccine composition.

57 . The method of claim 56 , wherein subsequent doses of the anti-cancer immunotherapy and the cancer vaccine composition are administered to the human subject on the same day.

58 . The method of claim 57 , wherein the anti-cancer immunotherapy is administered after the cancer vaccine composition.

59 . The method of any one of claims 39 - 58 , wherein the cancer is selected from the group consisting of small cell lung cancer, urothelial cancer, colorectal cancer, endometrial cancer, stomach cancer, gastro-esophageal junction cancer melanoma, bladder carcinoma, HPV negative HNSCC, NSCLC, SCLC, MSI (microsatellite)-High tumors, or TMB (tumor mutational burden)-High cancers.

60 . The method of any one of claims 39 - 58 , wherein the anti-cancer immunotherapy is administered every 20-24 days.

61 . The method of any one of claims 39 - 58 , wherein the anti-cancer immunotherapy is administered every 21 days.

62 . The method of any one of claims 39 - 61 , wherein the anti-cancer immunotherapy is administered at a dose of 150-250 mgs.

63 . The method of any one of claims 39 - 61 , wherein the anti-cancer immunotherapy is administered at a dose of 200 mgs.

64 . The method of any one of claims 39 - 63 , wherein the cancer is a PD-L1 negative tumor.

65 . The method of any one of claims 39 - 55 , wherein the anti-cancer immunotherapy is administered within 7 days of the mRNA cancer vaccine.

66 . The method of any one of claims 39 - 55 , wherein the anti-cancer immunotherapy is administered within 2-3 weeks of the mRNA cancer vaccine.

67 . The method of any one of claims 39 - 66 , wherein the cancer has a high tumor mutational burden.

68 . The method of any one of claims 39 - 66 , wherein the subject is selected for treatment based on a threshold tumor mutational burden.

69 . The method of any one of claims 39 - 66 , wherein the subject is selected for treatment based on a threshold microsatellite instability (MSI) value.

70 . The method of any one of claims 39 - 66 , wherein the subject is selected for treatment based on a threshold T-cell inflamed gene expression profile (GEP).

71 . The method of claim 70 , wherein the GEP comprises PD-1, PD-L1 and PD-L2.

72 . The method of claim 71 , wherein the subject is selected for treatment based on high levels of PD-L1 and PD-L2.

73 . The method of claim 71 , wherein the subject is selected for treatment based high levels of PD-L1 and PD-1.

74 . The method of any one of claims 39 - 66 , wherein the subject is selected for treatment based on a threshold GEP and TMB.

75 . The method of claim 48 , wherein the checkpoint inhibitor targets PD1, PD-L1, CTLA4, TIM-3, VISTA, A2AR, B7-H3, B7-H4, BTLA, IDO, KIR, LAG3, or a combination thereof.

76 . The method of claim 48 , wherein the checkpoint inhibitor is an antibody.

77 . The method of claim 76 , wherein the checkpoint inhibitor is an antibody selected from an anti-CTLA4 antibody or antigen-binding fragment thereof that specifically binds CTLA4, an anti-PD1 antibody or antigen-binding fragment thereof that specifically binds PD1, an anti-PD-L1 antibody or antigen-binding fragment thereof that specifically binds PD-L1, and a combination thereof.

78 . The method of claim 48 , wherein the checkpoint inhibitor is an anti-PD-L1 antibody selected from atezolizumab, avelumab, or durvalumab.

79 . The method of claim 48 , wherein the checkpoint inhibitor is an anti-CTLA-4 antibody selected from tremelimumab or ipilimumab.

80 . The method of claim 48 , wherein the checkpoint inhibitor is an anti-PD1 antibody selected from nivolumab or pembrolizumab.