Influenza virus reassortment

Patent metadata

Field	Value
Publication number	US-8945904-B2
Application number	US-201113699301-A
Country	US
Kind code	B2
Filing date	May 20, 2011
Priority date	May 21, 2010
Publication date	Feb 3, 2015
Grant date	Feb 3, 2015

Abstract

Methods for producing reassortant viruses are provided wherein the transcription and/or translation of the hemagglutinin and/or neuraminidase genes are suppressed.

First claim

The invention claimed is:

1. A method of preparing a reassortant influenza virus comprising the steps of
(i) contacting a culture host infected with a vaccine strain and a backbone strain with a nucleic acid inhibitory agent wherein said inhibitory agent preferentially reduces the transcription and/or translation of the hemagglutinin and/or neuraminidase genes of the backbone strain; and,
(ii) culturing the culture host in order to produce the reassortant virus.

2. The method of claim 1 , further comprising the step (iii) of purifying the reassortant virus obtained in step (ii).

3. A method of preparing a reassortant influenza virus comprising the steps of
(i) introducing into a culture host that has been infected with a first influenza virus strain having at least one target segment one or more expression construct(s) encoding the target segment(s) from a second influenza virus strain;
(ii) contacting the culture host with a nucleic acid inhibitory agent which preferentially reduces the transcription and/or translation of the first influenza strain's target segment(s);
(iii) culturing the culture host in order to produce reassortant virus; and optionally
(iv) purifying the virus obtained in step (iii).

4. The method of claim 1 , further comprising the steps of (a) infecting a culture host with the virus obtained in step (ii) of claim 1 ; (b) culturing the host from step (a) to produce further virus; and (c) purifying the virus obtained in step (b).

5. The method of claim 3 , further comprising the steps of (a) infecting a culture host with a reassortant influenza virus obtained in step (iii) or (iv) of claim 3 ; (b) culturing the host from step (a) to produce the virus; and optionally (c) purifying the virus obtained in step (b).

6. A method of preparing a vaccine, comprising steps of (a) preparing a reassortant virus by the method of claim 1 and (b) preparing a vaccine from the reassortant virus.

7. The method of claim 1 , wherein the culture host is an embryonated hen egg.

8. The method of claim 1 , wherein the culture host is a mammalian cell.

9. The method of claim 8 , wherein the cell is an MDCK, Vero or PerC6 cell.

10. The method of claim 8 , wherein the cell grows adherently.

11. The method of claim 8 , wherein the cell grows in suspension.

12. The method of claim 11 , wherein the cell is of the cell line MDCK 33016 (DSM ACC2219).

13. The method of claim 4 , wherein step (c) involves inactivating the virus.

14. The method of claim 6 , wherein the vaccine is a whole virion vaccine.

15. The method of claim 6 , wherein the vaccine is a split virion vaccine.

16. The method of claim 6 , wherein the vaccine is a surface antigen vaccine.

17. The method of claim 6 , wherein the vaccine is a virosomal vaccine.

18. The method of claim 6 , wherein the vaccine contains less than 10 ng of residual host cell DNA per dose.

19. The method of claim 1 wherein at least one of the influenza strains is of the H1, H2, H5, H7 or H9 subtype.

20. The method of claim 19 , wherein at least one of the strains is a H1N1, H5N1, H5N3, H9N2, H2N2, H7N1 or H7N7 strain.

21. The method of claim 1 , wherein one of the influenza strains is a high-growth strain.

22. The method of claim 19 , wherein at least one influenza strain is selected from the group consisting of A/Puerto Rico/8/34, A/Ann Arbor/6/60, B/Ann Arbor/1/66, A/Chile/1/83 and A/California/04/09.

23. A cell for preparing a reassortant influenza virus comprising expression construct(s) encoding:
(i) all eight viral segments of a first influenza A or B virus genome;
(ii) at least one target segment of a second influenza A or B virus genome, wherein the second influenza strain's target segment(s) differs in sequence from the target segment of the first influenza strain; and
(iii) a nucleic acid inhibitory agent wherein said inhibitory agent preferentially reduces transcription and/or translation of the target segment(s) of the first influenza strain.

24. The method of claim 1 , wherein the nucleic acid inhibitory agent is an antisense oligonucleotide.

25. The method of claim 1 , wherein the nucleic acid inhibitory agent is a synthetic antisense oligomer.

26. The method of claim 1 , wherein the nucleic acid inhibitory agent is a phosphorothioate oligomer (PMO).

27. The method of claim 1 , wherein the nucleic acid inhibitory agent is a phosphorodiamidate morpholino oligomer (PSO).

28. The method of claim 1 , wherein the nucleic acid inhibitory agent is selected from the group consisting of short interfering RNAs (siRNA), double-stranded RNAs (dsRNA), micro-RNAs (miRNA), short hairpin RNAs (shRNA), and small interfering DNAs (siDNA).

Assignees

• Trusheim Heidi
• Mason Peter
• Franti Michael
• Keiner Bjoern
• Sackal Melissa
• Hunziker Juerg
• Natt Francois
• Morrissey David
• Novartis Ag

Inventors

• Trusheim Heidi
• Mason Peter
• Franti Michael
• Keiner Bjoern
• Sackal Melissa
• Hunziker Juerg
• Natt Francois
• Morrissey David

Classifications

• A61K39/145 Primary

  Orthomyxoviridae, e.g. influenza virus · CPC title

• C12N2320/00

  Applications; Uses · CPC title

• A61K2039/525

  Virus · CPC title

• C12N7/00 Primary

  Viruses; Bacteriophages; Compositions thereof; Preparation or purification thereof (preparing medicinal viral antigen or antibody compositions, e.g. virus vaccines, A61K39/00) · CPC title

• A61K39/12

  Viral antigens · CPC title