Attenuated influenza vaccines and uses thereof

Patent metadata

Field	Value
Publication number	US-10323231-B2
Application number	US-201615568836-A
Country	US
Kind code	B2
Filing date	Apr 22, 2016
Priority date	Apr 24, 2015
Publication date	Jun 18, 2019
Grant date	Jun 18, 2019

Abstract

Provided herein are attenuated influenza viruses and methods of making attenuated influenza viruses.

First claim

What is claimed is:

1. A modified influenza A virus comprising a PB1 polymerase comprising one or more mutations selected from the group consisting of a leucine to glutamic acid, aspartic acid or asparagine substitution at an amino acid corresponding to position 319 (L319E/D/N) of SEQ ID NO: 1, a threonine to glutamic acid, aspartic acid, glutamine or asparagine substitution at an amino acid corresponding to position 323 (T323E/D/Q/N) of SEQ ID NO: 1; and an isoleucine to glutamic acid, aspartic acid, glutamine or asparagine substitution at an amino acid corresponding to position 342 (I342E/D/Q/N) of SEQ ID NO: 1.

2. The virus of claim 1 , wherein the PB1 polymerase comprises a leucine to glutamic acid substitution (L319E).

3. The virus of claim 1 , wherein the PB1 polymerase comprises a leucine to asparagine substitution (L319N).

4. 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 an amino acid corresponding to position 391 (K391E) of SEQ ID NO: 1, a glutamic acid to glycine substitution at an amino acid corresponding to position 581 (E581G) of SEQ ID NO: 1 and an alanine to threonine substitution at an amino acid corresponding to position 661 (A661T) of SEQ ID NO: 1.

5. The virus of claim 1 , wherein the virus further comprises a PB2 polymerase comprising an asparagine to serine substitution at position 265 (N265S).

6. The virus of claim 1 , wherein the virus further comprises an influenza virus nucleoprotein (NP) comprising an aspartic acid to glycine substitution at position 34 (D34G).

7. 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.

8. The virus of claim 1 , wherein the influenza A virus is (A/Puerto Rico/8/34/H1 N1)(PR8), (A/California/04/2007 H1N1) or (A/Ann Arbor/6/60 H2N2).

9. 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 one or more mutations selected from the group consisting of a leucine to glutamic acid, aspartic acid or asparagine substitution at an amino acid corresponding to position 319 (L319E/D/N) of SEQ ID NO: 1, a threonine to glutamic acid, aspartic acid, glutamine or asparagine substitution at an amino acid corresponding to position 323 (T323E/D/Q/N) of SEQ ID NO: 1; and an isoleucine to glutamic acid, aspartic acid, glutamine or asparagine substitution at an amino acid corresponding to position 342 (I342 E/D/Q/N) of SEQ ID NO: 1.

10. An immunogenic composition comprising the virus of claim 1 and a pharmaceutically acceptable carrier.

11. A method for eliciting an immune response against an influenza virus in a subject, comprising administering an effective dose of the immunogenic composition of claim 10 to the subject.

12. A method of treating or reducing the risk of influenza infection in a subject, comprising administering to a subject with an influenza infection or at risk of exposure to an influenza infection an effective dose of the immunogenic composition of claim 10 .

13. A recombinant nucleic acid encoding a PB1 polymerase of an influenza A virus, wherein the nucleic acid encodes a PB1 polymerase having one or more mutations selected from the group consisting of a leucine to glutamic acid, aspartic acid or asparagine substitution at an amino acid corresponding to position 319 (L319E/D/N) of SEQ ID NO: 1, a threonine to glutamic acid, aspartic acid, glutamine or asparagine substitution at an amino acid corresponding to position 323 (T323E/D/Q/N) of SEQ ID NO: 1; and an isoleucine to glutamic acid, aspartic acid, glutamine or asparagine substitution at an amino acid corresponding to position 342 (I342E/D/Q/N) of SEQ ID NO: 1.

14. A recombinant nucleic acid encoding a PB1 polymerase of an influenza A virus wherein the nucleic acid encodes a PB1 polymerase comprising one or more mutations selected from the group consisting of a leucine to glutamic acid, aspartic acid or asparagine substitution at an amino acid corresponding to position 319 (L319E/D/N) of SEQ ID NO: 1, a threonine to glutamic acid, aspartic acid, glutamine or asparagine substitution at amino acid corresponding to position 323 (T323E/D/Q/N) of SEQ ID NO: 1; a serine to glutamic acid, aspartic acid, glutamine or asparagine substitution at an amino acid corresponding to position 338 (S338E/D/Q/N) and an isoleucine to glutamic acid, aspartic acid, glutamine or asparagine substitution at an amino acid corresponding to position 342 (I342E/D/Q/N) of SEQ ID NO: 1; and one or more mutations selected from the group consisting of a lysine to glutamic acid substitution at an amino acid corresponding to position 391 (K391E) of SEQ ID NO: 1, a glutamic acid to glycine substitution at an amino acid corresponding to position 581 (E581G) of SEQ ID NO: 1 and an alanine to threonine substitution at an amino acid corresponding to position 661 (A661T) of SEQ ID NO: 1.

15. A vector comprising the nucleic acid of claim 13 .

16. A method of producing the influenza virus of claim 1 , 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 comprising one or more mutations selected from the group consisting of a leucine to glutamic acid, aspartic acid or asparagine substitution at an amino acid corresponding to position 319 (L319E/D/N) of SEQ ID NO: 1, a threonine to glutamic acid, aspartic acid, glutamine or asparagine substitution at an amino acid corresponding to position 323 (T323E/D/Q/N) of SEQ ID NO: 1; and an isoleucine to glutamic acid, aspartic acid, glutamine or asparagine substitution at an amino acid corresponding to position 342 (I342 E/D/Q/N) of SEQ ID NO: 1;

b) culturing the host cells; and
c) recovering the influenza A virus.

17. The method of claim 16 , wherein the nucleic acid encoding the PB1 polymerase encodes a PB1 polymerase comprising one or more mutations selected from the group consisting of a leucine to glutamic acid, aspartic acid or asparagine substitution at an amino acid corresponding to position 319 (L319E/D/N) of SEQ ID NO: 1, a threonine to glutamic acid, aspartic acid, glutamine or asparagine substitution at an amino acid corresponding to position 323 (T323E/D/Q/N) of SEQ ID NO: 1; a serine to glutamic acid, aspartic acid, glutamine or asparagine substitution at an amino acid corresponding to position 338 (S338 E/D/Q/N) of SEQ ID NO: 1 and an isoleucine to glutamic acid, aspartic acid, glutamine or asparagine substitution at an amino acid corresponding to position 342 (I342 E/D/Q/N) of SEQ ID NO: 1; and one or mutations selected from the group consisting of a glutamic acid to glycine substitution at an amino acid corresponding to position 581 (E581G) of SEQ ID NO: 1 and an alanine to threonine substitution at an amino acid corresponding to position 661 (A661T) of SEQ ID NO: 1.

18. The method of claim 16 , wherein the one or more vectors further comprise a nucleic acid encoding a PB2 polymerase comprising a N265S mutation.

19. The method of claim 16 , further comprising transforming the cells with a nucleic acid encoding an influenza virus nucleoprotein (NP) comprising an aspartic acid to glycine substitution at position 34 (D34G).

20. The method of claim 16 , wh

Assignees

• Univ Rochester

Inventors

• Cox Andrew
• Dewhurst Stephen
• Treanor John

Classifications

• A61K39/12

  Viral antigens · CPC title

• C12N2760/16151

  Methods of production or purification of viral material · CPC title

• C12N2760/16162

  by genetic engineering · CPC title

• C12N2760/16122

  New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes · CPC title

• C12N2760/16134

  Use of virus or viral component as vaccine, e.g. live-attenuated or inactivated virus, VLP, viral protein · CPC title