Compositions and methods for modulation of SMN2 splicing

• US 8,361,977 B2
• Filed: 06/23/2006
• Issued: 01/29/2013
• Est. Priority Date: 06/23/2005
• Status: Active Grant

• Current Assignee: Biogen MA Inc. (Biogen, Inc.), Cold Spring Harbor Laboratory
• Original Assignee: ISIS Pharmaceuticals Incorporated
• Inventors: Baker, Brenda F., Krainer, Adrian R., Hua, Yimin
• Primary Examiner(s): McGarry, Sean
• Application Number:
  US11/993,609
• Publication Number:
  US 20100216238A1
• Time in Patent Office: 2,412 Days
• Field of Search: None
• US Class Current: 514/44.A
• CPC Class Codes: A61K 48/00 Medicinal preparations containing genetic material which is inserted into cells of the living body to treat genetic diseases; Gene therapy
• CPC Class Codes: C07H 21/02 with ribosyl as saccharide radical
• 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 2310/11 Antisense
• CPC Class Codes: C12N 2310/321 2'-O-R Modification
• CPC Class Codes: C12N 2310/3341 5-Methylcytosine
• CPC Class Codes: C12N 2310/3525 MOE, methoxyethoxy
• CPC Class Codes: C12N 2320/33 Alteration of splicing

Abstract

Disclosed herein are compounds, compositions and methods for modulating splicing of SMN2 mRNA in a cell, tissue or animal. Also provided are uses of disclosed compounds and compositions in the manufacture of a medicament for treatment of diseases and disorders, including spinal muscular atrophy.

Claims

1. An antisense oligonucleotide having a nucleotide sequence at least 90% complementary to a target region of SEQ ID NO:

1 as measured over the entirety of the antisense oligonucleotide, wherein;

the 5′

-most nucleotide of the target region is nucleotide 121, 122, 123, 124, 125, 126, 127, 128 or 129 of SEQ ID NO;

1;

the antisense oligonucleotide is 12 to 20 nucleotides in length; and

each nucleoside of the antisense oligonucleotide comprises a 2′

-O-methoxyethyl sugar modification.

2. The antisense oligonucleotide of claim 1 that is 12 nucleotides in length.

3. The antisense oligonucleotide of claim 1 that is 15 nucleotides in length.

4. The antisense oligonucleotide of claim 1 that is 18 nucleotides in length.

5. The antisense oligonucleotide of claim 1 that is 20 nucleotides in length.

6. The antisense oligonucleotide of claim 1 comprising at least one phosphorothioate linkage.

7. The antisense oligonucleotide of claim 1 that is 100% complementary to the target region of SEQ ID NO:

1.

8. The antisense oligonucleotide of claim 7 that is 12 nucleotides in length.

9. The antisense oligonucleotide of claim 7 that is 15 nucleotides in length.

10. The antisense oligonucleotide of claim 7 that is 18 nucleotides in length.

11. The antisense oligonucleotide of claim 7 that is 20 nucleotides in length.

12. The antisense oligonucleotide of claim 7 comprising at least one phosphorothioate linkage.

41. A pharmaceutical composition comprising an antisense oligonucleotide of claim 1.

13. An antisense oligonucleotide having a nucleotide sequence 100% complementary to a target region of SEQ ID NO:

1 as measured over the entirety of the antisense oligonucleotide, wherein;

the 5′

-most nucleotide of the target region is nucleotide 123 of SEQ ID NO;

1;

the antisense oligonucleotide is 12 to 20 nucleotides in length; and

each nucleoside of the antisense oligonucleotide comprises a 2′

-O-methoxyethyl sugar modification.

14. The antisense oligonucleotide of claim 13 that is 12 nucleotides in length.

15. The antisense oligonucleotide of claim 13 that is 15 nucleotides in length.

16. The antisense oligonucleotide of claim 13 that is 18 nucleotides in length.

17. The antisense oligonucleotide of claim 13 that is 20 nucleotides in length.

18. The antisense oligonucleotide of claim 13 comprising at least one phosphorothioate linkage.

42. A pharmaceutical composition comprising an antisense oligonucleotide of claim 13.

19. An antisense oligonucleotide having a nucleotide sequence 100% complementary to a target region of SEQ ID NO:

1 as measured over the entirety of the antisense oligonucleotide, wherein;

the 5′

-most nucleotide of the target region is nucleotide 124 of SEQ ID NO;

1;

the antisense oligonucleotide is 12 to 20 nucleotides in length; and

each nucleoside of the antisense oligonucleotide comprises a 2′

-O-methoxyethyl sugar modification.

20. The antisense oligonucleotide of claim 19 that is 12 nucleotides in length.

21. The antisense oligonucleotide of claim 19 that is 15 nucleotides in length.

22. The antisense oligonucleotide of claim 19 that is 18 nucleotides in length.

23. The antisense oligonucleotide of claim 19 that is 20 nucleotides in length.

24. The antisense oligonucleotide of claim 19 comprising at least one phosphorothioate linkage.

25. The antisense oligonucleotide of claim 22 comprising at least one phosphorothioate linkage.

26. The antisense oligonucleotide of claim 25 wherein each internucleoside linkage is a phosphorothioate linkage.

43. A pharmaceutical composition comprising an antisense oligonucleotide of claim 19.

44. A pharmaceutical composition comprising an antisense oligonucleotide of claim 22.

45. A pharmaceutical composition comprising an antisense oligonucleotide of claim 25.

46. A pharmaceutical composition comprising an antisense oligonucleotide of claim 26.

27. An antisense oligonucleotide having a nucleotide sequence 100% complementary to a target region of SEQ ID NO:

1 as measured over the entirety of the antisense oligonucleotide, wherein;

the 5′

-most nucleotide of the target region is nucleotide 125 of SEQ ID NO;

1;

the antisense oligonucleotide is 12 to 20 nucleotides in length; and

each nucleoside of the antisense oligonucleotide comprises a 2′

-O-methoxyethyl sugar modification.

28. The antisense oligonucleotide of claim 27 that is 12 nucleotides in length.

29. The antisense oligonucleotide of claim 27 that is 15 nucleotides in length.

30. The antisense oligonucleotide of claim 27 that is 18 nucleotides in length.

31. The antisense oligonucleotide of claim 27 that is 20 nucleotides in length.

32. The antisense oligonucleotide of claim 27 comprising at least one phosphorothioate linkage.

47. A pharmaceutical composition comprising an antisense oligonucleotide of claim 27.

33. An antisense oligonucleotide targeted to intron 7 of a nucleic acid molecule encoding SMN2, wherein:

the antisense oligonucleotide comprises SEQ ID NO;

84 and is at least 90% complementary to the nucleic acid molecule encoding SMN2 as measured over the entirety of the antisense oligonucleotide;

the antisense oligonucleotide is 12 to 20 nucleotides in length; and

each nucleoside of the antisense oligonucleotide comprises a 2′

-O-methoxyethyl sugar modification.

34. The antisense oligonucleotide of claim 33 that is 12 nucleotides in length.

35. The antisense oligonucleotide of claim 33 that is 15 nucleotides in length.

36. The antisense oligonucleotide of claim 33 that is 18 nucleotides in length.

37. The antisense oligonucleotide of claim 33 that is 20 nucleotides in length.

38. The antisense oligonucleotide of claim 33 comprising at least one phosphorothioate linkage.

39. The antisense oligonucleotide of claim 38 wherein each internucleoside linkage is a phosphorothioate linkage.

40. The antisense oligonucleotide of claim 35, wherein the antisense oligonucleotide comprises SEQ ID NO:

83.

48. A pharmaceutical composition comprising an antisense oligonucleotide of claim 33.