Modulation of HSP47 expression
View Patent ↗Provided herein are compositions, methods and kits for modulating expression of target genes, particularly heat shock protein 47 (hsp47). The compositions, methods and kits may include nucleic acid molecules (for example, short interfering nucleic acid (siNA), short interfering RNA (siRNA), double-stranded RNA (dsRNA), micro-RNA (miRNA) or short hairpin RNA (shRNA)) that modulate a gene encoding hsp47, for example, the gene encoding human hsp47. The composition and methods disclosed herein may also be used in treating conditions and disorders associated with hsp47 such as liver fibrosis, pulmonary fibrosis, peritoneal fibrosis and kidney fibrosis.
1. A double stranded nucleic acid molecule having a structure (A2) set forth below:
wherein each of N 2 , N and N′ is independently an unmodified or modified ribonucleotide, or an unconventional moiety;
wherein each of (N)x and (N′)y is an oligonucleotide in which each consecutive N or N′ is joined to the adjacent N or N′ by a covalent bond;
wherein each of x and y is 18;
wherein the sequence of (N′)y has complementarity to the sequence of (N)x and the sequence of (N)x has complementarity to a consecutive sequence in a mRNA encoding hsp47;
wherein N 1 is covalently bound to (N)x and is mismatched to the mRNA encoding hsp47;
or is a complementary DNA moiety to the mRNA encoding hsp47;
wherein N 1 is a moiety selected from natural or modified uridine or deoxyribouridine;
wherein z″ may be present or absent, but if present is a capping moiety covalently attached at the 5′ terminus of N 2 —(N′)y;
wherein each of Z and Z′ is independently present or absent, but if present is independently 1-5 consecutive nucleotides, consecutive non-nucleotide moieties or a combination thereof covalently attached at the 3′ terminus of the strand in which it is present; and
wherein the sense strand and the antisense strand comprise the oligonucleotides described as SERPINH1 — 2 (SEQ ID NOS: 60 and 127).
2. A composition comprising the double stranded nucleic acid molecule of claim 1 ; and a pharmaceutically acceptable carrier.
3. The double stranded nucleic acid molecule of claim 1 , having the structure
wherein each “|” represents base pairing between the ribonucleotides;
wherein each of A, C, G and U is independently an unmodified or modified ribonucleotide, or an unconventional moiety;
wherein each A, C, G and U is joined to an adjacent A, C, G or U by a covalent bond;
wherein each of Z and Z′ is independently present or absent, but if present is independently 1-5 consecutive nucleotides or non-nucleotide moieties or a combination thereof covalently attached at the 3′ terminus of the strand in which it is present; and
wherein z″ may be present or absent, but if present is a capping moiety covalently attached at the 5′ terminus of the sense strand.
4. The double stranded nucleic acid molecule of claim 3 , wherein each of A, C, G and U is an unmodified ribonucleotide, and wherein z″, Z and Z′ are absent.
5. The double stranded nucleic acid molecule of claim 3 , wherein each of Z and Z′ is present.
6. The double stranded nucleic acid molecule of claim 5 , wherein each of Z and Z′ independently comprises a nucleotide overhang or a non-nucleotide overhang.
7. The double stranded nucleic acid molecule of claim 6 , wherein each of Z and Z′ comprises a non-nucleotide overhang.
8. The double stranded nucleic acid molecule of claim 7 , wherein Z′ comprises C3Pi and wherein Z comprises C3Pi-C3OH.
9. The double stranded nucleic acid molecule of claim 3 , wherein the antisense strand comprises 2′-O-methyl sugar modified ribonucleotides.
10. The double stranded nucleic acid molecule of claim 9 , wherein the 2′-O-methyl sugar modified ribonucleotides are present in positions (5′>3′) 3, 5, 9, 11, 13, 15, 17 and 19.
11. The double stranded nucleic acid molecule of claim 10 , wherein the antisense strand further comprises a 2′-O-methyl sugar modified ribonucleotide or a 2′5′ ribonucleotide in position (5′>3′) 1.
12. The double stranded nucleic acid molecule of claim 11 , wherein the antisense strand comprises a 2′-O-methyl sugar modified ribonucleotide in position (5′>3′) 1.
13. The double stranded nucleic acid molecule of claim 11 , wherein the antisense strand comprises a 2′5′ ribonucleotide in position (5′>3′) 1.
14. The double stranded nucleic acid molecule of claim 11 , wherein the antisense strand comprises a 2′5′ ribonucleotide in any one of positions (5′>3′) 6, 7 or 8.
15. The double stranded nucleic acid molecule of claim 14 , wherein the 2′5′ ribonucleotide is present in position 7.
16. The double stranded nucleic acid molecule of claim 3 , wherein the sense strand comprises 4 or 5 consecutive 2′5′ nucleotides at the 3′ terminal or penultimate positions.
17. The double stranded nucleic acid molecule of claim 16 , wherein the sense strand comprises 2′5′ ribonucleotides in positions (5′>3′) 15, 16, 17, and 18.
18. The double stranded nucleic acid molecule of claim 16 , wherein the sense strand comprises 2′5′ ribonucleotides in positions (5′>3′) 15, 16, 17, 18 and 19.
19. The double stranded nucleic acid molecule of claim 3 , wherein the sense strand comprises 2′-O-methyl sugar modified ribonucleotides in positions (5′>3′) 7, 13, 16 and 18.
20. The double stranded nucleic acid molecule of claim 19 , wherein the sense strand further comprises a 2′5′ nucleotide in position (5′>3′) 9.
21. The double stranded nucleic acid molecule of claim 3 , wherein z″ is present and comprises a capping moiety covalently attached to the 5′ terminus of the sense strand.
22. The double stranded nucleic acid molecule of claim 21 , wherein the capping moiety is selected from an abasic ribose moiety, an inverted abasic ribose moiety, an inverted abasic deoxyribose moiety, an abasic deoxyribose moiety and modifications thereof; C6-imino-Pi; a mirror nucleotide; 5′OMe nucleotide; 4′,5′-methylene nucleotide; 1-(β-D-erythrofuranosyl)nucleotide; 4′-thio nucleotide, carbocyclic nucleotide; 5′-amino-alkyl phosphate; 1,3-diamino-2-propyl phosphate, 3-aminopropyl phosphate; 6-aminohexyl phosphate; 12-aminododecyl phosphate; hydroxypropyl phosphate; 1,5-anhydrohexitol nucleotide; alpha-nucleotide; threo-pentofuranosyl nucleotide; acyclic 3′,4′-seco nucleotide; 3,4-dihydroxybutyl nucleotide; 3,5-dihydroxypentyl nucleotide, 5′-5′-inverted abasic moiety; 1,4-butanediol phosphate; 5′-amino; and bridging or non bridging methylphosphonate and 5′-mercapto moieties.
23. The double stranded nucleic acid molecule of claim 22 , wherein the capping moiety comprises an inverted abasic deoxyribose moiety.
24. The double stranded nucleic acid molecule of claim 1 , wherein the antisense strand comprises 2′-O-methyl sugar modified ribonucleotides, a 2′5′ ribonucleotide in at least one of positions (5′>3′) 1, 5, 6 or 7, and a 3′ terminal non-nucleotide moiety covalently attached to the 3′ terminus; and
wherein the sense strand comprises at least one 2′5′ ribonucleotide and/or a 2′-O-methyl modified ribonucleotide, a non-nucleotide moiety covalently attached at the 3′ terminus and a capping moiety covalently attached at the 5′ terminus.
25. The double stranded nucleic acid molecule of claim 24 , wherein the antisense strand comprises 2′-O-methyl sugar modified ribonucleotides at positions (5′>3′) 3, 5, 9, 11, 13, 15, 17 and 19, a 2′5′ ribonucleotide in position 7, and a non-nucleotide moiety covalently attached at the 3′ terminus; and
wherein the sense strand comprises five consecutive 2′5′ ribonucleotides in the 3′ terminal positions (5′>3′) 15, 16, 17, 18 and 19, a non-nucleotide moiety covalently attached at the 3′ terminus and an inverted abasic capping moiety covalently attached at the 5′ terminus.
26. The double stranded nucleic acid molecule of claim 25 , wherein the antisense strand comprises 2′-O-methyl sugar modified ribonucleotides at positions (5′>3′) 1, 3, 5, 9, 11, 13, 15, 17 and 19, a 2′5′ ribonucleotide in position 7, and a C3Pi-C3OH non-nucleotide moiety covalently attached at the 3′ terminus; and
wherein the sense strand comprises five consecutive 2′5′ ribonucleotides in the 3′ terminal positions (5′>3′) 15, 16, 17, 18 and 19, a C3Pi non-nucleotide moiety covalently attached at the 3′ terminus and an inverted abasic capping moiety covalently attached at the 5′ terminus.
27. The double stranded nucleic acid molecule of claim 25 , wherein the antisense strand comprises 2′-O-methyl sugar modified ribonucleotides at positions (5′>3′) 3, 5, 9, 11, 13, 15, 17 and 19, a 2′5′ ribonucleotide in positions 1 and 7, and a C3Pi-C3OH non-nucleotide moiety covalently attached at the 3′ terminus; and
wherein the sense strand comprises five consecutive 2′5′ ribonucleotides in the 3′ terminal positions (5′>3′) 15, 16, 17, 18 and 19, a C3Pi non-nucleotide moiety covalently attached at the 3′ terminus and an inverted abasic capping moiety covalently attached at the 5′ terminus.
28. The double stranded nucleic acid molecule of claim 24 , wherein the antisense strand comprises 2′-O-methyl sugar modified ribonucleotides in positions (5′>3′) 1, 3, 5, 9, 11, 13, 15, 17 and 19, a 2′5′ ribonucleotide in position 7, and a C3Pi-C3OH non-nucleotide moiety covalently attached at the 3′ terminus; and
wherein the sense strand comprises 2′-O-methyl sugar modified ribonucleotides in positions (5′>3′) 7, 13, 16 and 18, a 2′5′ ribonucleotide at position 9, a C3 non-nucleotide moiety covalently attached at the 3′ terminus and an inverted abasic capping moiety covalently attached at the 5′ terminus.
29. A method for treating an individual suffering from a disease associated with hsp47 comprising administering to said individual the double stranded nucleic acid molecule of claim 1 in an amount sufficient to reduce expression of hsp47, wherein the disease associated with hsp47 is fibrosis.
30. A method for treating an individual suffering from a disease associated with hsp47 comprising administering to said individual the double stranded nucleic acid molecule of claim 26 in an amount sufficient to reduce expression of hsp47, wherein the disease associated with hsp47 is fibrosis.
31. The method of claim 30 wherein the fibrosis is selected from the group consisting of liver fibrosis, liver cirrhosis, pulmonary fibrosis, kidney fibrosis, peritoneal fibrosis, chronic hepatic damage, cardiofibrosis, retinal fibrosis, retro-orbital fibrosis, lacrimal gland fibrosis, myelofibrosis, intestinal fibrosis, vocal cord mucosal fibrosis, laryngeal fibrosis, nephrogenic systemic fibrosis, congenital hepatic fibrosis and oral submucosal fibrosis.
32. The method of claim 31 , wherein the fibrosis comprises liver cirrhosis, liver fibrosis, pulmonary fibrosis, kidney fibrosis, myelofibrosis and intestinal fibrosis.
33. The double stranded nucleic acid molecule of claim 3 , wherein each covalent bond joining each A, C, G and U to the adjacent A, C, G or U is a phosphodiester bond.
34. The method of claim 30 wherein the fibrosis is a fibrosis of skin, peritoneum, liver, pancreas, kidney, heart, lung, bone marrow, eye, intestine, vocal cord and/or vasculature.
35. The method of claim 29 , wherein the fibrosis is selected from the group consisting of liver fibrosis, liver cirrhosis, pulmonary fibrosis, kidney fibrosis, peritoneal fibrosis, chronic hepatic damage, cardiofibrosis, retinal fibrosis, retro-orbital fibrosis, lacrimal gland fibrosis, myelofibrosis, intestinal fibrosis, vocal cord mucosal fibrosis, laryngeal fibrosis, nephrogenic systemic fibrosis, congenital hepatic fibrosis and oral submucosal fibrosis.
36. The method of claim 35 , wherein the fibrosis comprises liver fibrosis, liver cirrhosis, pulmonary fibrosis, kidney fibrosis, myelofibrosis and intestinal fibrosis.
37. The method of claim 29 wherein the fibrosis is a fibrosis of skin, peritoneum, liver, pancreas, kidney, heart, lung, bone marrow, eye, intestine, vocal cord and/or vasculature.