Attenuated influenza vaccines and uses thereof
Provided herein are attenuated influenza viruses and methods of making attenuated influenza viruses.
1. A modified influenza A virus comprising a PB1 polymerase, wherein the PB1 polymerase comprises a leucine to glutamine substitution (L319Q) at an amino acid corresponding to amino acid 319 of SEQ ID NO: 2.
2. The virus of claim 1 , wherein the PB1 polymerase further comprises one or more mutations selected from the group consisting of a lysine to glutamic acid substitution at position 391 (K391E), a glutamic acid to glycine substitution at position 581 (E581G) and an alanine to threonine substitution at position 661 (A661T).
3. The virus of claim 1 , wherein the virus further comprises a PB2 polymerase comprising an asparagine to serine substitution at position 265 (N265S).
4. The virus of claim 1 , wherein the virus further comprises an influenza virus nucleoprotein (NP) comprising an aspartic acid to glycine substitution at position 35 (D35G).
5. The virus of claim 1 , wherein the influenza A virus is selected from the group consisting of an H2N2 virus, an H3N2 virus, an H1N1 virus, an H9N2 virus and an H5N1 virus.
6. The virus of claim 1 , wherein the influenza A virus is A/Ann Arbor/6/60 (H2N2).
7. The virus of claim 1 , wherein the virus is a live attenuated influenza A virus with reduced growth from about 37° C. to about 39° C., as compared to an influenza A virus comprising a PB1 polymerase lacking a leucine to glutamine substitution (L319Q) at an amino acid corresponding to amino acid 319 of SEQ ID NO: 2.
8. An immunogenic composition comprising the virus of claim 1 and a pharmaceutically acceptable carrier.
9. A method for eliciting an immune response against an influenza A virus in a subject, comprising administering an effective dose of the immunogenic composition of claim 8 to the subject.
10. A method of producing an influenza A virus, comprising:
a) transfecting a population of host cells with one or more vectors comprising
i) nucleic acid sequences encoding the internal genome segments of an influenza A virus and;
ii) a nucleic acid encoding a PB1 polymerase with a leucine to glutamine substitution (L319Q) at an amino acid corresponding to amino acid 319 of SEQ ID NO: 2;
b) culturing the host cells; and
c) recovering the modified influenza A virus.
11. The method of claim 10 , wherein the nucleic acid encoding the PB1 polymerase further comprises one or more mutations selected from the group consisting of a lysine to glutamic acid substitution at position 391 (K391E), a glutamic acid to glycine substitution at position 581 (E581G) and an alanine to threonine substitution at position 661 (A661T).
12. The method of claim 10 , further comprising transforming the cells with a nucleic acid encoding a PB2 polymerase comprising a N265S mutation.
13. The method of claim 10 , further comprising transforming the cells with a nucleic acid encoding an NP protein comprising a D35G mutation.
14. The method of claim 10 , wherein the influenza A virus is selected from the group consisting of an H2N2 virus, an H3N2 virus, an H1N1 virus, an H9N2 virus and an H5N1 virus.
15. The method of claim 10 , wherein the cells are Vero cells, MDCK cells or CEK cells.
16. A method for producing an influenza A virus comprising:
a) infecting a population of cells with the virus of claim 1 ;
b) culturing the cells;
c) harvesting the virus from the culture of step b).
17. The method of claim 16 , wherein the cell is a mammalian cell or an avian cell.
18. A population of isolated cells comprising the virus of claim 1 .