CHIMERIC OLIGOMERIC COMPOUNDS FOR MODULATION OF SPLICING

• US 20110039334A1
• Filed: 08/29/2006
• Published: 02/17/2011
• Est. Priority Date: 08/30/2005
• Status: Active Grant

• Current Assignee: Ionis Pharmaceuticals, Inc.
• Original Assignee: ISIS Pharmaceuticals Incorporated
• Inventors: Kopczynski, Casey C., Kole, Ryszard M., Dean, Nicholas M., Bennett, C. Frank
• Granted Patent: US 8,501,703 B2
• Time in Patent Office: Days
• US Class Current: 435/375
• CPC Class Codes: C12N 15/111 General methods applicable to biologically active non-coding nucleic acids
• CPC Class Codes: C12N 15/113 Non-coding nucleic acids modulating the expression of genes, e.g. antisense oligonucleotides; Antisense DNA or RNA; Triplex- forming oligonucleotides; Catalytic nucleic acids, e.g. ribozymes; Nucleic acids used in co-suppression or gene silencing when used in plants C12N15/8218
• CPC Class Codes: C12N 15/1135 against oncogenes or tumor suppressor genes
• CPC Class Codes: C12N 2310/321 2'-O-R Modification
• CPC Class Codes: C12N 2310/322 2'-R Modification
• CPC Class Codes: C12N 2310/346 having a combination of backbone and sugar modifications
• CPC Class Codes: C12N 2310/3525 MOE, methoxyethoxy
• CPC Class Codes: C12N 2310/3531 Hydrogen
• CPC Class Codes: C12N 2310/3533 Halogen
• CPC Class Codes: C12N 2320/33 Alteration of splicing

Abstract

Disclosed herein are compounds, compositions and methods for modulating splicing of a selected target mRNA. Further provided are uses of the disclosed compounds and compositions in the manufacture of a medicament for treatment of diseases and disorders. Methods of enhancing cellular uptake, modulating tissue distribution and enhancing pharmacological activity of RNase H-independent antisense oligonucleotides are also provided.

Claims

1. A method of modulating splicing of a selected target pre-mRNA in a cell comprising contacting said cell with an oligomeric compound comprising an oligonucleotide consisting of 13 to 80 linked nucleosides which is complementary to said selected target pre-mRNA, wherein the oligonucleotide comprises:

a modified sugar moiety at the 5′

-most nucleoside;

a modified sugar moiety at the 3′

-most nucleoside;

internal nucleosides comprising 2, 3, 4 or 5 separate regions of 2′

-deoxyribonucleosides, wherein each region consists of 1, 2, 3 or 4 nucleosides; and

at least one nucleoside with a modified sugar moiety separating each region of 2′

-deoxynucleosides.

2. The method of claim 1, wherein said oligonucleotide comprises 3 separate regions of 2′

-deoxyribonucleosides.

3. The method of claim 1, wherein said oligonucleotide comprises 4 separate regions of 2′

-deoxyribonucleosides.

4. The method of claim 1, wherein each region of 2′

-deoxyribonucleosides of said oligonucleotide consists of 2 or 3 nucleosides.

5. The method of claim 1, wherein the 5′

end of the oligonucleotide consists of 2, 3, 4 or 5 modified nucleosides comprising modified sugar moieties.

6. The method of claim 1, wherein the 3′

end of the oligonucleotide consists of 2, 3, 4 or 5 modified nucleosides comprising modified sugar moieties.

7. The method of claim 1, wherein the sugar modification is selected from the group consisting of 2′

-O-methoxyethyl, 2′

-fluoro, 2′

-dimethylaminooxyethoxy, 2′

-dimethylaminoethoxyethoxy, 2′

-guanidinium, 2′

-O-guanidinium ethyl, 2′

-carbamate, 2′

-aminooxy, 2′

-acetamido and locked nucleic acid.

8. The method of claim 7, wherein the sugar modification is 2′

-O-methoxyethyl.

9. The method of claim 1, wherein the oligonucleotide consists of 13 to 50 linked nucleosides.

10. The method of claim 9, wherein the oligonucleotide consists of 13 to 30 linked nucleosides.

11. The method of claim 10, wherein the oligonucleotide consists of 20 to 30 linked nucleosides.

12. The method of claim 11, wherein the oligonucleotide consists of 15 to 25 linked nucleosides.

13. The method of claim 12, wherein the oligonucleotide consists of 20 linked nucleosides.

14. The method of claim 13, wherein the oligonucleotide has 2′

-deoxyribonucleosides at positions 3, 4, 5, 7, 8, 9, 11, 12, 13, 15, 16 and 17.

15. The method of claim 13, wherein the oligonucleotide has 2′

-deoxyribonucleosides at positions 4, 5, 6, 9, 10, 11, 14, 15 and 16.

16. The method of claim 13, wherein the oligonucleotide has 2′

-deoxyribonucleosides at positions 4, 5, 8, 9, 10, 13, 14 and 15.

17. The method of claim 13, wherein the oligonucleotide has 2′

-deoxyribonucleosides at positions 5, 6, 7, 9, 10, 11, 13, 14 and 15.

18. The method of claim 13, wherein the oligonucleotide has 2′

-deoxyribonucleosides at positions 5, 6, 7, 9, 10, 13, 14 and 15.

19. The method of claim 13, wherein the oligonucleotide has 2′

-deoxyribonucleosides at positions 5, 6, 7, 10, 11, 12, 15, 16 and 17.

20. The method of claim 13, wherein the oligonucleotide has 2′

-deoxyribonucleosides at positions 6, 7, 8, 10, 11, 12, 14, 15 and 16.

21. The method of claim 13, wherein the oligonucleotide has 2′

-deoxyribonucleosides at positions 6, 7, 8, 11, 12, 13, 16 and 17.

22. The method of claim 1, wherein the cell is derived from adipose.

23. The method of claim 1, wherein the cell is derived from liver.

24. The method of claim 1, wherein the oligonucleotide is complementary to a splice site.

25. The method of claim 24, wherein the splice site is an aberrant splice site.

26. The method of claim 24, wherein the splice site is a cryptic splice site.

27. An oligomeric compound comprising an oligonucleotide consisting of 13 to 80 linked nucleosides, wherein the oligonucleotide is complementary to a selected target mRNA, and wherein the oligonucleotide comprises:

a modified sugar moiety at the 5′

-most nucleoside;

a modified sugar moiety at the 3′

-most nucleoside;

internal nucleosides comprising 2, 3, 4 or 5 separate regions of 2′

-deoxyribonucleosides, wherein each region consists of 1, 2, 3 or 4 nucleosides; and

at least one nucleoside with a modified sugar moiety separating each region of 2′

-deoxynucleosides.

28. The compound of claim 27, wherein said oligonucleotide comprises 3 separate regions of 2′

-deoxyribonucleosides.

29. The compound of claim 27, wherein said oligonucleotide comprises 4 separate regions of 2′

-deoxyribonucleosides.

30. The compound of claim 27, wherein each region of 2′

-deoxyribonucleosides of said oligonucleotide consists of 2 or 3 nucleosides.

31. The compound of claim 27, wherein the 5′

end of said oligonucleotide consists of 2, 3, 4 or 5 modified nucleosides comprising modified sugar moieties.

32. The compound of claim 27, wherein the 3′

end of said oligonucleotide consists of 2, 3, 4 or 5 modified nucleosides comprising modified sugar moieties.

33. The compound of claim 27, wherein the sugar modification is selected from the group consisting of 2′

-O-methoxyethyl, 2′

-fluoro, 2′

-dimethylaminooxyethoxy, 2′

-dimethylaminoethoxyethoxy, 2′

-guanidinium, 2′

-O-guanidinium ethyl, 2′

-carbamate, 2′

-aminooxy, 2′

-acetamido and locked nucleic acid.

34. The compound of claim 33, wherein the sugar modification is 2′

-O-methoxyethyl.

35. The compound of claim 27, wherein cellular uptake of said compound is enhanced compared to a compound of the same sequence which comprises an oligonucleotide wherein every nucleoside of said oligonucleotide comprises a modified sugar moiety.

36. The compound of claim 27, wherein said compound modulates splicing of said selected target mRNA.

37. The compound of claim 27, wherein said oligonucleotide is complementary to a splice site of the selected target mRNA.

38. The compound of claim 37, wherein the splice site is an aberrant splice site.

39. The compound of claim 37, wherein the splice site is a cryptic splice site.

40-57. -57. (canceled)