Methods of treatment using antisense oligomers and conjugates targeting PCSK9
The present invention relates to oligomeric compounds and conjugates thereof that target Proprotein Convertase Subtilisin/Kexin type 9 (PCSK9) PCSK9 mRNA in a cell, leading to reduced expression of PCSK9. Reduction of PCSK9 expression is beneficial for a range of medical disorders, such as hypercholesterolemia and related disorders.
1. A method of reducing cholesterol levels in a subject in need thereof comprising administering to said subject an effective amount of a compound consisting of
an antisense oligomer that is 16 contiguous nucleotides in length, wherein the sequence of the antisense oligomer is 100% complementary to the sequence of SEQ ID NO: 31, wherein the antisense oligomer is a gapmer comprising at least one LNA unit; and
wherein the antisense oligomer targets mRNA encoding PCSK9.
2. The method of claim 1 , wherein the cholesterol is serum LDL cholesterol.
3. The method of claim 1 , further comprising the administration of a second therapeutic agent.
4. The method of claim 3 , wherein the second therapeutic agent is a statin.
5. The method of claim 4 , wherein the statin is selected from the group consisting of lovastatin, cerivastatin, pravastatin, atorvastatin, simvastatin, rosuvastatin, and fluvastatin.
6. The method of claim 4 , wherein the second therapeutic agent is a bile sequestering resin, nicotinic acid, a fibric acid derivative, probucol, neomycin, dextrothyroxine, a plant stanol ester, a cholesterol absorption inhibitor, implitapide, an inhibitor of bile acid transporters, a regulator of hepatic CYP7a, an estrogen replacement therapeutic, or an anti-inflammatory.
7. The method of claim 1 , wherein the compound is administered intravenously or subcutaneously.
8. The method of claim 1 , wherein the compound is administered as a single dose or as multiple doses.
9. The method of claim 1 , wherein the sequence of the antisense oligomer is SEQ ID NO: 26.
10. The method of claim 1 , wherein the LNA is oxy-LNA, thio-LNA, amino-5 LNA, 5′-methyl-LNA, ENA, cET, cMOE or a combination thereof.
11. The method of claim 1 , wherein the LNA is an stereoisomer in the beta-D-configuration or the alpha-L configuration.
12. The method of claim 1 , wherein the antisense oligomer comprises at least one cET unit.
13. The method of claim 1 , wherein the antisense oligomer comprises six or seven LNA units.
14. The method of claim 13 , wherein every LNA unit in the antisense oligomer is a beta-D-oxy LNA unit.
15. The method of claim 13 , wherein every LNA unit in the antisense oligomer is a stereoisomer in the same configuration.
16. The method of claim 15 , wherein every the LNA unit in the antisense oligomer is an stereoisomer in the R configuration.
17. The method of claim 15 , wherein every the LNA unit in the antisense oligomer is an stereoisomer in the S configuration.
18. The method of claim 15 , wherein every the LNA unit in the antisense oligomer is an stereoisomer in the D configuration.
19. The method of claim 15 , wherein every the LNA unit in the antisense oligomer is an stereoisomer in the L configuration.
20. The method of claim 1 , wherein the LNA is an (R,S) racemic mixture.
21. The method of claim 1 , wherein the sequence of the antisense oligomer comprises at least one phosphorothioate internucleoside linkage.
22. The method of claim 1 , wherein the sequence of the antisense oligomer is SEQ ID NO: 2.
23. The method of claim 1 , wherein the sequence of the antisense oligomer is SEQ ID NO: 3.
24. The method of claim 1 , wherein the compound and an oligomer of SEQ ID NO:31 can form a duplex with increased thermal stability with respect to a corresponding duplex comprising the corresponding antisense oligomer without LNA.
25. A method of reducing cholesterol levels in a subject in need thereof comprising administering to said subject an effective amount of an antisense oligonucleotide conjugate comprising:
an antisense oligomer that is 16 contiguous nucleotides in length, wherein the sequence of the antisense oligomer is 100% complementary to the sequence of SEQ ID NO: 31, and wherein the antisense oligomer is a gapmer comprising at least one LNA unit; and
at least one non-nucleotide or non-polynucleotide moiety covalently attached to said antisense oligomer directly or via a linker positioned between the contiguous oligomer sequence and the non-nucleotide or non-polynucleotide moiety,
wherein the antisense oligomer targets mRNA encoding PCSK9.
26. The method of claim 25 , wherein the non-nucleotide or non-polynucleotide moiety is a liver targeting moiety that is attached to the 5′-end or to the 3′-end of the antisense oligomer.
27. The method of claim 26 , wherein the liver targeting moiety is linked to the antisense oligomer via a linker.
28. The method of claim 27 , wherein the linker is a biocleavable linker.
29. The method of claim 26 , wherein the liver targeting moiety comprises a carbohydrate conjugate moiety comprising a carbohydrate or a carbohydrate group.
30. The method of claim 29 , wherein the carbohydrate is selected from the group consisting of galactose, lactose, N-acetylgalactosamine (GalNAc), mannose, mannose-6-phosphate, and combinations thereof.
31. The method of claim 29 , wherein the carbohydrate conjugate moiety is not a linear carbohydrate polymer.
32. The method of claim 29 , wherein the carbohydrate conjugate moiety is a carbohydrate group comprising 1 2, 3, or 4 carbohydrate moieties.
33. The method of claim 32 , wherein the carbohydrate moieties are identical or non-identical.
34. The method of claim 32 , wherein the carbohydrate moieties are non-identical.
35. The method of claim 29 , wherein the carbohydrate conjugate moiety comprises at least one asialoglycoprotein receptor targeting conjugate moiety.
36. The method of claim 35 , wherein the asialoglycoprotein receptor targeting conjugate moiety comprises a GalNAc cluster.
37. The method of claim 35 , wherein the asialoglycoprotein receptor targeting conjugate moiety comprises a monovalent, divalent, trivalent, or tetravalent GalNAc cluster.
38. The method of claim 36 , wherein the GalNAc cluster is a trivalent GalNAc cluster.
39. The method of claim 38 , wherein each GalNAc in the trivalent GalNAc cluster is attached to a branch point group via a spacer.
40. The method of claim 39 , wherein the branch point group comprises di-lysine.
41. The method of claim 39 , wherein the spacer comprises a PEG spacer.
42. The method of claim 27 , wherein the linker comprises a C6 to C12 amino alkyl group or a biocleavable phosphate nucleotide linker comprising between 1 to 6 nucleotides.
43. The method of claim 38 , wherein the trivalent GalNAc cluster comprises Conj 2a:
44. The method of claim 25 , wherein the non-nucleotide or non-polynucleotide moiety is covalently attached to the antisense oligomer via a covalent bond.
45. The method of claim 25 , wherein the antisense oligonucleotide conjugate is SEQ ID NO: 18.
46. The method of claim 25 , wherein the antisense oligonucleotide conjugate is SEQ ID NO: 19.
47. The method of claim 25 , wherein the cholesterol is serum LDL cholesterol.
48. The method of claim 25 , further comprising the administration of a second therapeutic agent.
49. The method of claim 48 , wherein the second therapeutic agent is a statin.
50. The method of claim 49 , wherein the statin is selected from the group consisting of lovastatin, cerivastatin, pravastatin, atorvastatin, simvastatin, rosuvastatin, and fluvastatin.
51. The method of claim 48 , wherein the second therapeutic agent is a bile sequestering resin, nicotinic acid, a fibric acid derivative, probucol, neomycin, dextrothyroxine, a plant stanol ester, a cholesterol absorption inhibitor, implitapide, an inhibitor of bile acid transporters, a regulator of hepatic CYP7a, an estrogen replacement therapeutic, or an anti-inflammatory.
52. The method of claim 25 , wherein the antisense oligonucleotide conjugate is administered intravenously or subcutaneously.
53. The method of claim 25 , wherein the antisense oligonucleotide conjugate is administered as a single dose or as multiple doses.
54. A method of reducing cholesterol levels in a subject in need thereof comprising administering to said subject an effective amount of an antisense oligonucleotide conjugate of SEQ ID NO: 18
wherein
a superscript L identifies a beta-D-oxy LNA unit,
a Me C identifies a 5-methylcytosine unit,
a subscript s identifies a phosphorothioate internucleoside linkage,
and wherein
is a Conj 2a asialoglycoprotein receptor targeting conjugate moiety
and wherein the antisense oligonucleotide conjugate targets mRNA encoding PCSK9.
55. A method of reducing cholesterol levels in a subject in need thereof comprising administering to said subject an effective amount of an antisense oligonucleotide conjugate of SEQ ID NO: 19
wherein
a superscript L identifies a beta-D-oxy LNA unit,
a Me C identifies a 5-methylcytosine unit,
a subscript s identifies a phosphorothioate internucleoside linkage,
and wherein
is a Conj 2a asialoglycoprotein receptor targeting conjugate moiety
and wherein the antisense oligonucleotide conjugate targets mRNA encoding PCSK9.
56. The method of claim 25 , wherein the antisense oligomer comprises six or seven LNA units.
57. The method of claim 56 , wherein every LNA unit in the antisense oligomer is a beta-D-oxy LNA unit.
58. The method of claim 56 , wherein every LNA unit in the antisense oligomer is a stereoisomer in the same configuration.
59. The method of claim 58 , wherein every the LNA unit in the antisense oligomer is an stereoisomer in the R configuration.
60. The method of claim 58 , wherein every the LNA unit in the antisense oligomer is an stereoisomer in the S configuration.
61. The method of claim 58 , wherein every the LNA unit in the antisense oligomer is an stereoisomer in the D configuration.
62. The method of claim 58 , wherein every the LNA unit in the antisense oligomer is an stereoisomer in the L configuration.
63. The method of claim 25 , wherein the LNA is an (R,S) racemic mixture.