Therapeutic Compositions

• US 20070275914A1
• Filed: 03/08/2004
• Published: 11/29/2007
• Est. Priority Date: 03/07/2003
• Status: Active Grant

• Current Assignee: Alnylam Pharmaceuticals Incorporated
• Original Assignee: Alnylam Pharmaceuticals Incorporated
• Inventors: Manoharan, Muthiah, Bumcrot, David
• Granted Patent: US 8,110,674 B2
• Time in Patent Office: Days
• US Class Current: 514/44.A
• CPC Class Codes: A01K 2217/05 Animals comprising random inserted nucleic acids (transgenic)
• CPC Class Codes: A01K 2227/105 Murine
• CPC Class Codes: A01K 2267/0362 Animal model for lipid/glucose metabolism, e.g. obesity, type-2 diabetes
• CPC Class Codes: A01K 67/0275 Genetically modified vertebrates, e.g. transgenic
• CPC Class Codes: A61K 31/713 Double-stranded nucleic acids or oligonucleotides
• CPC Class Codes: A61K 47/554 the modifying agent being a steroid plant sterol, glycyrrhetic acid, enoxolone or bile acid
• CPC Class Codes: A61P 3/10 for hyperglycaemia, e.g. antidiabetics
• CPC Class Codes: C07H 21/02 with ribosyl as saccharide radical
• CPC Class Codes: C07K 14/775 Apolipopeptides
• 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 15/1137 against enzymes viral enzymes C12N15/1131; receptors C12N15/1138
• CPC Class Codes: C12N 15/1138 against receptors or cell surface proteins
• CPC Class Codes: C12N 2310/14 interfering N.A.
• CPC Class Codes: C12N 2310/315 Phosphorothioates
• CPC Class Codes: C12N 2310/321 2'-O-R Modification
• CPC Class Codes: C12N 2310/344 Position-specific modifications, e.g. on every purine, at the 3'-end
• CPC Class Codes: C12N 2310/3515 Lipophilic moiety, e.g. cholesterol
• CPC Class Codes: C12N 2310/533 having a mismatch or nick in at least one of the strands
• CPC Class Codes: C12N 2320/32 Special delivery means, e.g. tissue-specific
• CPC Class Codes: C12N 2320/51 : modulating the chemical stability, e.g. nuclease-resistance
• CPC Class Codes: C12N 2320/53 : reducing unwanted side-effects
• CPC Class Codes: C12N 2330/30 : chemically synthesised

Abstract

Therapeutic sRNA agents and methods of making and using are enclosed.

Claims

1. An iRNA agent comprising a sense sequence and an antisense sequence, wherein the sense sequence has one or more asymmetrical 2′

-O alkyl modifications and the antisense sequence has one or more asymmetrical phosphorothioate modifications, and the antisense sequence targets a human apoB-100, glucose-6-phosphatase, or beta-catenin nucleic acid sequence.

2. The iRNA agent of claim 1, wherein at least one of said 2′

-O-alkyl modifications is a 2′

-OMe modification.

3. The iRNA agent of claim 1, wherein the sense sequence has at least 2 asymmetrical 2′

-O alkyl modifications.

10. The iRNA agent of claim 1, wherein all of the subunits of the sense sequence have an asymmetrical 2′

-O alkyl modification.

11. The iRNA agent of claim 10, wherein the asymmetrical modifications are 2′

-OMe modifications.

12. The iRNA agent of claim 1, wherein the antisense sequence has at least 2 asymmetrical phosphorothioate modifications.

16. The iRNA agent of claim 1, wherein all of the subunits of the sense sequence have an asymmetrical phosphorothioate modification.

17. The iRNA agent of claim 1, wherein the sense and antisense sequences are on different RNA strands.

18. The iRNA agent of claim 1, wherein the sense and antisense sequences are on the same RNA strand.

19. The iRNA agent of claim 1, wherein the sense and antisense sequences are fully complementary to each other.

20. The iRNA agent of claim 1, further comprising a cholesterol moiety, wherein the cholesterol moiety is coupled to the sense sequence.

28. The iRNA agent of claim 1, wherein the iRNA agent is at least 21 nucleotides in length, and the duplex region of the iRNA agent is about 19 nucleotides in length.

29. The iRNA agent of claim 1, having a duplex region of about 19 nucleotides in length and one or two 3′

overhangs of about 2 nucleotides in length.

30. A pharmaceutical preparation comprising the iRNA agent of claim 1 and a pharmaceutically acceptable carrier.

4-9. -9. (canceled)

13-15. -15. (canceled)

21-27. -27. (canceled)

31. A method for reducing apoB-100 levels in a subject comprising administering to a subject an iRNA agent comprising a sense strand sequence and an antisense sequence, wherein the sense sequence has at least 4 asymmetrical 2′

-O alkyl modifications and the antisense sequence has at least 4 asymmetrical phosphorothioate modifications, and the antisense sequence targets apoB-100.

32. The method of claim 31, wherein the subject is suffering from a disorder characterized by elevated or otherwise unwanted expression of apoB-100, elevated or otherwise unwanted levels of cholesterol, and/or disregulation of lipid metabolism.

33. (canceled)

34. A method for reducing glucose-6-phosphatase levels in a subject comprising administering to a subject an iRNA agent comprising a sense strand sequence and an antisense sequence, wherein the sense sequence has at least 4 asymmetrical 2′

-O alkyl modifications and the antisense sequence has at least 4 asymmetrical phosphorothioate modifications, and the antisense sequence targets glucose-6-phosphatase.

35. The method of claim 34, wherein the iRNA agent is administered to a subject to inhibit hepatic glucose production, or for the treatment of a glucose-metabolism-related disorder.

36. The method of claim 35, wherein said disorder is diabetes.

37. (canceled)

38. A method of making an iRNA agent, the method comprising:

providing a sense strand sequence having at least 4 asymmetrical 2′

-O alkyl modifications and an antisense sequence having at least 4 asymmetrical phosphorothioate modifications, and allowing the sense and antisense strands to hybridize.

39. A method of stabilizing an iRNA agent, comprising selecting a sequence with activity, and introducing one or more asymmetrical modifications in said sequence, wherein said modifications decrease nuclease sensitivity while not decreasing activity.