Antisense oligonucleotides for inducing exon skipping and methods of use thereof

• US RE47,769 E1
• Filed: 11/11/2016
• Issued: 12/17/2019
• Est. Priority Date: 06/28/2004
• Status: Active Grant

• Current Assignee: University of Western Australia
• Original Assignee: University of Western Australia
• Inventors: Wilton, Stephen Donald, Fletcher, Sue, McClorey, Graham
• Primary Examiner(s): Ponnaluri, Padmashri
• Application Number:
  US15/349,535
• Time in Patent Office: 1,131 Days
• Field of Search: 536 245
• CPC Class Codes: N/A

Abstract

Antisense molecules capable of binding to a selected target site in the dystrophin gene to induce exon skipping are described.

Claims

1. An isolated antisense oligonucleotide of 30 to 50 nucleotides in length comprising SEQ ID NO:

181, wherein the uracil bases are optionally thymine bases.

2. The antisense oligonucleotide of claim 1, wherein the antisense oligonucleotide comprises a non-natural backbone.

3. The antisense oligonucleotide of claim 1, wherein the antisense oligonucleotide is chemically linked to one or more moieties or conjugates that enhance the activity, cellular distribution, or cellular uptake of the antisense oligonucleotide.

4. The antisense oligonucleotide of claim 1, wherein the antisense oligonucleotide does not activate RNase H.

5. The antisense oligonucleotide of claim 2, wherein the sugar moieties of the oligonucleotide backbone are replaced with non-natural moieties.

6. The antisense oligonucleotide of claim 5, wherein the non-natural moieties are morpholinos.

7. The antisense oligonucleotide of claim 6, wherein the uracil bases are thymine bases.

8. The antisense oligonucleotide of claim 1, wherein the inter-nucleotide linkages of the oligonucleotide backbone are replaced with non-natural inter-nucleotide linkages.

9. The antisense oligonucleotide of claim 8, wherein the non-natural inter-nucleotide linkages are modified phosphates.

10. The antisense oligonucleotide of claim 9, wherein the modified phosphates are methyl phosphonates, methyl phosphorothioates, phosphoromorpholidates, phosphoropiperazidates or phosphoroamidates.

11. The antisense oligonucleotide of claim 10, wherein the modified phosphates are phosphoroamidates.

12. The antisense oligonucleotide of claim 1, wherein the sugar moieties of the oligonucleotide backbone are replaced with non-natural moieties and the inter-nucleotide linkages of the oligonucleotide backbone are replaced with non-natural inter-nucleotide linkages.

13. The antisense oligonucleotide of claim 12, wherein the non-natural moieties are morpholinos and the non-natural internucleotide linkages are modified phosphates.

14. The antisense oligonucleotide of claim 13, wherein the modified phosphates are methyl phosphonates, methyl phosphorothioates, phosphoromorpholidates, phosphoropiperazidates or phosphoroamidates.

15. The antisense oligonucleotide of claim 14, wherein the modified phosphates are phosphoroamidates.

16. The antisense oligonucleotide of claim 15, wherein the uracil bases are thymine bases.

17. The antisense oligonucleotide of claim 16, wherein the antisense oligonucleotide is chemically linked to one or more moieties or conjugates that enhance the activity, cellular distribution, or cellular uptake of the antisense oligonucleotide.

18. The antisense oligonucleotide of claim 10, wherein the modified phosphates are phosphoromorpholidates.

19. The antisense oligonucleotide of claim 14, wherein the modified phosphates are phosphoromorpholidates.

20. The antisense oligonucleotide of claim 1, wherein the antisense oligonucleotide is 30 nucleotides in length.

21. The antisense oligonucleotide of claim 20, wherein the uracil bases are thymine bases.

22. An injectable solution comprising:

an antisense oligonucleotide of 30 nucleotides in length comprising the base sequence 5′

-CUCCAACAUCAAGGAAGAUGGCAUUUCUAG-3′

(SEQ ID NO;

     181), in which the uracil bases are thymine bases, wherein the antisense oligonucleotide is a morpholino antisense oligonucleotide, and wherein the antisense oligonucleotide is chemically linked to a polyethylene glycol chain; and

a pharmaceutically acceptable carrier or diluent;

wherein the injectable solution is formulated for intravenous administration.

23. The injectable solution of claim 22, wherein the pharmaceutically acceptable carrier or diluent comprises an isotonic saline solution.

24. An injectable solution comprising:

an antisense oligonucleotide of 30 nucleotides in length comprising the base sequence 5′

-CUCCAACAUCAAGGAAGAUGGCAUUUCUAG-3′

(SEQ ID NO;

     181), in which the uracil bases are thymine bases, wherein the antisense oligonucleotide is a morpholino antisense oligonucleotide, and wherein the antisense oligonucleotide is chemically linked to a polyethylene glycol chain; and

phosphate-buffered saline;

wherein the injectable solution is formulated for intravenous administration.

25. An injectable solution comprising:

an antisense oligonucleotide of 30 nucleotides in length comprising the base sequence 5′

-CUCCAACAUCAAGGAAGAUGGCAUUUCUAG-3′

(SEQ ID NO;

     181), in which the uracil bases are thymine bases, wherein the antisense oligonucleotide is a morpholino antisense oligonucleotide, and wherein the antisense oligonucleotide is chemically linked to a polyethylene glycol chain; and

a pharmaceutically acceptable carrier or diluent;

wherein the injectable solution is formulated for parenteral administration.

26. The injectable solution of claim 25, wherein the pharmaceutically acceptable carrier or diluent comprises an isotonic saline solution.

27. An injectable solution comprising:

an antisense oligonucleotide of 30 nucleotides in length comprising the base sequence 5′

-CUCCAACAUCAAGGAAGAUGGCAUUUCUAG-3′

(SEQ ID NO;

     181), in which the uracil bases are thymine bases, wherein the antisense oligonucleotide is a morpholino antisense oligonucleotide, and wherein the antisense oligonucleotide is chemically linked to a polyethylene glycol chain; and

phosphate-buffered saline;

wherein the injectable solution is formulated for parenteral administration.

28. An injectable solution comprising:

an antisense oligonucleotide of 30 nucleotides in length comprising the base sequence 5′

-CUCCAACAUCAAGGAAGAUGGCAUUUCUAG-3′

(SEQ ID NO;

     181), in which the uracil bases are thymine bases, wherein the antisense oligonucleotide is a morpholino antisense oligonucleotide, and wherein the antisense oligonucleotide is chemically linked to a polyethylene glycol chain; and

a pharmaceutically acceptable carrier or diluent;

wherein the injectable solution is formulated for intramuscular administration.

29. The injectable solution of claim 28, wherein the pharmaceutically acceptable carrier or diluent comprises an isotonic saline solution.

30. An injectable solution comprising:

an antisense oligonucleotide of 30 nucleotides in length comprising the base sequence 5′

-CUCCAACAUCAAGGAAGAUGGCAUUUCUAG-3′

(SEQ ID NO;

     181), in which the uracil bases are thymine bases, wherein the antisense oligonucleotide is a morpholino antisense oligonucleotide, and wherein the antisense oligonucleotide is chemically linked to a polyethylene glycol chain; and

phosphate-buffered saline;

wherein the injectable solution is formulated for intramuscular administration.

31. An injectable solution comprising:

an antisense oligonucleotide of 30 nucleotides in length comprising the base sequence 5′

-CUCCAACAUCAAGGAAGAUGGCAUUUCUAG-3′

(SEQ ID NO;

     181), in which the uracil bases are thymine bases, wherein the antisense oligonucleotide is a morpholino antisense oligonucleotide, and wherein the antisense oligonucleotide is chemically linked to a polyethylene glycol chain; and

a pharmaceutically acceptable carrier or diluent;

wherein the injectable solution is formulated for subcutaneous administration.

32. The injectable solution of claim 31, wherein the pharmaceutically acceptable carrier or diluent comprises an isotonic saline solution.

33. An injectable solution comprising:

an antisense oligonucleotide of 30 nucleotides in length comprising the base sequence 5′

-CUCCAACAUCAAGGAAGAUGGCAUUUCUAG-3′

(SEQ ID NO;

     181), in which the uracil bases are thymine bases, wherein the antisense oligonucleotide is a morpholino antisense oligonucleotide, and wherein the antisense oligonucleotide is chemically linked to a polyethylene glycol chain; and

phosphate-buffered saline;

wherein the injectable solution is formulated for subcutaneous administration.