RNA interference mediated inhibition of hepatitis B virus (HBV) using short interfering nucleic acid (siNA)

• US 20030206887A1
• Filed: 09/16/2002
• Published: 11/06/2003
• Est. Priority Date: 05/14/1992
• Status: Abandoned Application

• Current Assignee: Sirna Therapeutics, Inc. (Alnylam Pharmaceuticals Incorporated)
• Original Assignee: Sirna Therapeutics, Inc. (Alnylam Pharmaceuticals Incorporated)
• Inventors: Beigelman, Leonid, Morrissey, David, McSwiggen, James A.
• Application Number:
  US10/244,647
• Publication Number:
  US 20030206887A1
• Time in Patent Office: Days
• US Class Current: 424/93.2
• CPC Class Codes: A61K 2300/00 Mixtures or combinations of active ingredients, wherein at least one active ingredient is fully defined in groups A61K31/00 - A61K41/00
• CPC Class Codes: A61K 38/21 Interferons [IFN]
• CPC Class Codes: A61K 47/54 the modifying agent being an organic compound
• CPC Class Codes: C07H 19/10 with the saccharide radical esterified by phosphoric or polyphosphoric acids
• CPC Class Codes: C07H 19/20 with the saccharide radical esterified by phosphoric or polyphosphoric acids
• CPC Class Codes: C07H 21/00 Compounds containing two or more mononucleotide units having separate phosphate or polyphosphate groups linked by saccharide radicals of nucleoside groups, e.g. nucleic acids
• CPC Class Codes: C07K 14/005 from viruses
• CPC Class Codes: C12N 15/85 for animal cells
• CPC Class Codes: C12N 15/86 Viral vectors
• CPC Class Codes: C12N 2730/10122 New viral proteins or individual genes, new structural or functional aspects of known viral proteins or genes
• CPC Class Codes: C12N 2830/32 being an silencer not forming part of the promoter region
• CPC Class Codes: C12P 19/30 Nucleotides
• CPC Class Codes: C12P 19/305 Pyrimidine nucleotides
• CPC Class Codes: C12Q 1/6876 Nucleic acid products used in the analysis of nucleic acids, e.g. primers or probes
• CPC Class Codes: C12Q 1/706 for hepatitis

Abstract

The present invention concerns methods and reagents useful in modulating hepatitis B virus (HBV) gene expression in a variety of applications, including use in therapeutic, diagnostic, target validation, and genomic discovery applications. Specifically, the invention relates to short interfering nucleic acid (siNA) or short interfering RNA (siRNA) molecules capable of mediating RNA interference (RNAi) against against hepatitis B virus (HBV).

Claims

1. A short interfering nucleic acid (siNA) molecule that down-regulates expression of a HBV gene by RNA interference.

3. The siNA molecule of claim 1, wherein the HBV gene encodes sequence comprising Genbank Accession number AB073834.

4. The siNA molecule of claim 1, wherein said siNA molecule is adapted for use to treat HBV infection.

5. The siNA molecule of claim 1, wherein said siNA molecule comprises a sense region and an antisense region and wherein said antisense region comprises sequence complementary to an RNA sequence encoding HBV and the sense region comprises sequence complementary to the antisense region.

6. The siNA molecule of claim 5, wherein said siNA molecule is assembled from two nucleic acid fragments wherein one fragment comprises the sense region and the second fragment comprises the antisense region of said siNA molecule.

7. The siNA molecule of claim 6, wherein said sense region and said antisense region comprise separate oligonucleotides.

8. The siNA molecule of claim 6, wherein said sense region and said antisense region are covalently connected via a linker molecule.

9. The siNA molecule of claim 8, wherein said linker molecule is a polynucleotide linker.

10. The siNA molecule of claim 8, wherein said linker molecule is a non-nucleotide linker.

11. The siNA molecule of claim 1, wherein the siNA molecule comprises sequence having any of SEQ ID NOs.:

1-1524.

12. The siNA molecule of claim 5, wherein said sense region comprises a 3′

-terminal overhang and said antisense region comprises a 3′

-terminal overhang.

13. The siNA molecule of claim 12, wherein said 3′

-terminal overhangs each comprise about 2 nucleotides.

14. The siNA molecule of claim 12, wherein said antisense region 3′

-terminal overhang is complementary to RNA encoding HBV.

15. The siNA molecule of claim 5, wherein said sense region comprises one or more 2′

-O-methyl pyrimidine nucleotides and one or more 2′

-deoxy purine nucleotides.

16. The siNA molecule of claim 5, wherein any pyrimidine nucleotides present in said sense region comprise 2′

-deoxy-2′

-fluoro pyrimidine nucleotides and wherein any purine nucleotides present in said sense region comprise 2′

-deoxy purine nucleotides.

17. The siNA molecule of claim 16, wherein any nucleotides comprising a 3′

-terminal nucleotide overhang that are present in said sense region are 2′

-deoxy nucleotides.

18. The siNA molecule of claim 5, wherein said sense region comprises a 3′

-end, a 5′

-end, and a terminal cap moiety at 3′

-end, the 5′

-end, or both of the 5′

- and 3′

-ends of said sense region.

19. The siNA molecule of claim 18, wherein said terminal cap moiety is an inverted deoxy abasic moiety.

20. The siNA molecule of claim 5, wherein said antisense region comprises one or more 2′

-deoxy-2′

-fluoro pyrimidine nucleotides and one or more 2′

-O-methyl purine nucleotides.

21. The siNA molecule of claim 5, wherein any pyrimidine nucleotides present in said antisense region comprise 2′

-deoxy-2′

-fluoro pyrimidine nucleotides and wherein any purine nucleotides present in said antisense region comprise 2′

-O-methyl purine nucleotides.

22. The siNA molecule of claim 21, wherein any nucleotides comprising a 3′

-terminal nucleotide overhang that are present in said antisense region are 2′

-deoxy nucleotides.

23. The siNA molecule of claim 5, wherein said antisense region comprises a phosphorothioate internucleotide linkage at the 3′

-end of said antisense region.

24. The siNA molecule of claim 5, wherein said antisense region comprises a glyceryl modification at the 3′

-end of said antisense region.

25. The siNA molecule of claim 12, wherein said 3′

-terminal nucleotide overhangs comprise deoxyribonucleotides.

26. An expression vector comprising a nucleic acid sequence encoding at least one siNA molecule of claim 1 in a manner that allows expression of the nucleic acid molecule.

27. A mammalian cell comprising an expression vector of claim 26.

28. The mammalian cell of claim 27, wherein said mammalian cell is a human cell.

29. The expression vector of claim 26, wherein said at least one siNA molecule comprises a sense region and an antisense region and wherein said antisense region comprises sequence complementary to an RNA sequence encoding HBV and the sense region comprises sequence complementary to the antisense region.

30. The expression vector of claim 26, wherein said at least one siNA molecule comprises two distinct strands having complementary sense and antisense regions.

31. The expression vector of claim 26, wherein said siNA molecule comprises a single strand having complementary sense and antisense regions.

2. A short interfering nucleic acid (siNA) molecule that inhibits HBV replication.