Influenza vaccine

1. A method of inducing an antigen-specific immune response in a human subject, comprising administering to the human subject messenger RNA (mRNA) in an effective amount to induce an antigen-specific immune response in the subject, wherein the mRNA comprises an open reading frame encoding an HA7 hemagglutinin antigen and is formulated in a cationic lipid nanoparticle comprising a cationic lipid of Formula (I):

or a salt or isomer thereof, wherein:

R 1 is selected from the group consisting of C 5-30 alkyl, C 5-20 alkenyl, —R*YR″, —YR″, and —R″M′R′;

R 2 and R 3 are independently selected from the group consisting of H, C 1-14 alkyl, C 2-14 alkenyl, —R*YR″, —YR″, and —R*OR″, or R 2 and R 3 , together with the atom to which they are attached, form a heterocycle or carbocycle;

R 4 is selected from the group consisting of a C 3-6 carbocycle, —(CH 2 ) n Q, —(CH 2 ) n CHQR, —CHQR, —CQ(R) 2 , and unsubstituted C 1-6 alkyl, where Q is selected from a carbocycle, heterocycle, —OR, —O(CH 2 ) n N(R) 2 , —C(O)OR, —OC(O)R, —CX 3 , —CX 2 H, —CXH 2 , —CN, —N(R) 2 , —C(O)N(R) 2 , —N(R)C(O)R, —N(R)S(O) 2 R, —N(R)C(O)N(R) 2 , —N(R)C(S)N(R) 2 , —N(R)R 8 , —O(CH 2 ) n OR, —N(R)C(═NR 9 )N(R) 2 , —N(R)C(═CHR 9 )N(R) 2 , —OC(O)N(R) 2 , —N(R)C(O)OR, —N(OR)C(O)R, —N(OR)S(O) 2 R, —N(OR)C(O)OR, —N(OR)C(O)N(R) 2 , —N(OR)C(S)N(R) 2 , —N(OR)C(═NR 9 )N(R) 2 , —N(OR)C(═CHR 9 )N(R) 2 , —C(═NR 9 )N(R) 2 , —C(═NR 9 )R, —C(O)N(R)OR, and —C(R)N(R) 2 C(O)OR, and each n is independently selected from 1, 2, 3, 4, and 5;

each R 5 is independently selected from the group consisting of C 1-3 alkyl, C 2-3 alkenyl, and H;

each R 6 is independently selected from the group consisting of C 1-3 alkyl, C 2-3 alkenyl, and H;

M and M′ are independently selected from —C(O)O—, —OC(O)—, —C(O)N(R′)—, —N(R′)C(O)—, —C(O)—, —C(S)—, —C(S)S—, —SC(S)—, —CH(OH)—, —P(O)(OR′)O—, —S(O) 2 —, —S—S—, an aryl group, and a heteroaryl group;

R 7 is selected from the group consisting of C 1-3 alkyl, C 2-3 alkenyl, and H;

R 8 is selected from the group consisting of C 3-6 carbocycle and heterocycle;

R 9 is selected from the group consisting of H, CN, NO 2 , C 1-6 alkyl, —OR, —S(O) 2 R, —S(O) 2 N(R) 2 , C 2-6 alkenyl, C 3-6 carbocycle and heterocycle;

each R is independently selected from the group consisting of C 1-3 alkyl, C 2-3 alkenyl, and H;

each R′ is independently selected from the group consisting of C 1-18 alkyl, C 2-18 alkenyl, —R*YR″, —YR″, and H;

each R″ is independently selected from the group consisting of C 3-14 alkyl and C 3-14 alkenyl;

each R* is independently selected from the group consisting of C 1-12 alkyl and C 2-12 alkenyl;

each Y is independently a C 3-6 carbocycle;

each X is independently selected from the group consisting of F, Cl, Br, and I; and

m is selected from 5, 6, 7, 8, 9, 10, 11, 12, and 13.

2. The method of claim 1 , wherein the HA7 hemagglutinin antigen is an H7N9 antigen.

3. The method of claim 1 , wherein the open reading frame encodes an HA7 antigen that comprises an amino acid sequence having at least 80% sequence identity to SEQ ID NO: 1.

4. The method of claim 3 , wherein the open reading frame encodes an HA7 antigen that comprises the amino acid sequence of SEQ ID NO: 1.

5. The method of claim 1 , wherein the mRNA has at least 80% sequence identity to SEQ ID NO: 5, 6, 7, or 9.

6. The method of claim 1 , wherein a dose of 10-100 micrograms of the mRNA is administered to the human subject.

7. The method of claim 1 further comprising administering a booster dose of the mRNA to the human subject.

8. The method of claim 1 , wherein the cationic lipid nanoparticle comprises 20-60 mol % ionizable cationic lipid, 5-25 mol % neutral lipid, 25-55 mol % cholesterol, and 0.5-5 mol % PEG-modified lipid.

9. The method of claim 1 , wherein the cationic lipid of Formula (I) is Compound 25:

10. The method of claim 1 , wherein the antigen-specific immune response comprises a T cell response and/or a B cell response.

11. The method of claim 1 , wherein the mRNA comprises a chemical modification.

12. The method of claim 11 , wherein the chemical modification is selected from the group consisting of pseudouridine, 1-methylpseudouridine, 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-methoxyuridine, and 2′-O-methyl uridine.

13. The method of claim 1 , wherein the mRNA further comprises a 5′ terminal cap and/or a 3′ polyA tail.

14. A method of inducing an antigen-specific immune response in a human subject, comprising administering to the human subject messenger RNA (mRNA) in an effective amount to induce an antigen-specific immune response in the subject, wherein the mRNA comprises an open reading frame encoding an HA7 hemagglutinin antigen that comprises an amino acid sequence having at least 80% sequence identity to SEQ ID NO: 1, and wherein the mRNA is formulated in a cationic lipid nanoparticle.

15. The method of claim 14 , wherein the HA7 antigen comprises an amino acid sequence having at least 90% sequence identity to SEQ ID NO: 1.

16. The method of claim 15 , wherein the HA7 antigen comprises the amino acid sequence of SEQ ID NO: 1.

17. The method of claim 14 , wherein a dose of 10-100 micrograms of the mRNA is administered to the human subject.

18. The method of claim 14 further comprising administering a booster dose of the mRNA to the human subject.

19. The method of claim 14 , wherein the cationic lipid nanoparticle comprises 20-60 mol % ionizable cationic lipid, 5-25 mol % neutral lipid, 25-55 mol % cholesterol, and 0.5-5 mol % PEG-modified lipid.

20. The method of claim 14 , wherein the cationic lipid nanoparticle comprises a cationic lipid of Compound 25:

21. The method of claim 14 , wherein the antigen-specific immune response comprises a T cell response and/or a B cell response.

22. The method of claim 14 , wherein the mRNA comprises a chemical modification.

23. The method of claim 22 , wherein the chemical modification is selected from the group consisting of pseudouridine, 1-methylpseudouridine, 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-methoxyuridine, and 2′-O-methyl uridine.

24. The method of claim 14 , wherein the mRNA further comprises a 5′ terminal cap and/or a 3′ polyA tail.