Granted Patent B2

US 12,359,201 · App. 16/978,947 · Granted Jul 15, 2025

Inventors: Gregory Hinkle (Plymouth, MA); Frederic Tremblay (Boston, MA); Noura S. Abul-Husn (New York, NY); Omri Gottesman (Rancho Santa Fe, CA); Alexander Li (White Plains, NY); Xiping Cheng (Northvale, NJ); Yurong Xin (Ashland, MA); Jesper Gromada (Concord, MA); Frederick E. Dewey (Winchester, MA); Aris Baras (Bedford Corners, NY); Alan Shuldiner (Baltimore, MD); Stuart Milstein (Arlington, MA)

Assignee: Regeneron Pharmaceuticals, Inc.

C12N15/113C12N15/1137C12N2310/11C12N2310/14C12N2310/315C12N2310/321C12N2310/322C12N2310/3515C12N2310/3521C12N2310/3533C12N2320/32C12N2320/50

Abstract

Claims (62)

1. A method of inhibiting 17β-hydroxysteroid dehydrogenase type 13 (HSD17B13) expression in a cell, comprising contacting the cell with a double stranded ribonucleic acid (dsRNA) agent, wherein the dsRNA agent comprises a sense strand and an antisense strand, wherein the nucleotide sequence of the sense strand comprises the nucleotide sequence of SEQ ID NO: 4498 and the nucleotide sequence of the antisense strand comprises the nucleotide sequence of SEQ ID NO: 4499, thereby inhibiting expression of HSD17B13 in the cell.

2. The method of claim 1 , wherein the cell is in a subject.

3. The method of claim 2 , wherein the subject suffers from a chronic fibro-inflammatory liver disease, fatty liver disease (steatosis), inflammation of the liver, liver fibrosis, or obesity.

4. The method of claim 3 , wherein the chronic fibro-inflammatory liver disease comprises nonalcoholic steatohepatitis (NASH), nonalcoholic fatty liver disease (NAFLD), cirrhosis of the liver, alcoholic steatohepatitis (ASH), alcoholic liver diseases (ALD), drug induced liver injury, or hepatocellular necrosis.

5. The method of claim 3 , wherein the chronic fibro-inflammatory liver disease comprises a chronic fibro-inflammatory liver disease associated with the accumulation and/or expansion of lipid droplets in the liver.

6. The method of claim 2 , wherein the subject suffers from steatosis, and wherein inhibiting expression of HSD17B13 in the subject inhibits progression of steatosis to steatohepatitis in the subject.

7. The method of claim 1 , wherein the method further comprises administering to the subject a pharmaceutical composition comprising a dsRNA agent targeting a PNPLA3 gene, thereby inhibiting expression of PNPLA3 in the subject.

8. The method of claim 4 , wherein the cirrhosis of the liver comprises HCV-associated cirrhosis.

9. The method of claim 7 , wherein the subject suffers from steatosis, and wherein inhibiting the expression of HSD17B13 and PNPLA3 in the subject inhibits the progression of steatosis to steatohepatitis in the subject.

10. The method of claim 1 , wherein all of the nucleotides of the sense strand and all of the nucleotides of the antisense strand are modified.

11. The method of claim 1 , wherein the sense strand comprises no more than four 2′-fluoro modified nucleotides.

12. The method of claim 1 , wherein the antisense strand comprises no more than six 2′-fluoro modified nucleotides.

13. The method of claim 1 , wherein the antisense strand comprises at least one glycol-modified nucleotide.

14. The method of claim 13 , wherein the antisense strand comprises a glycol modified nucleotide at position 7, counting from the 5 40 -end.

15. The method of claim 1 , wherein the sense strand comprises four 2 ′-fluoro modified nucleotides at nucleotides 7 and 9-11, counting from the 5′-end, and

wherein the antisense strand comprises three 2′-fluoro modified nucleotides at nucleotides 2, 14, and 16, counting from the 5′-end, and one glycol modified nucleotide at nucleotide 7, counting from the 5′-end.

16. The method of claim 1 , wherein: (a) the sense strand comprises at least one internucleotide linkage chosen from phosphorothioate and methylphosphonate internucleotide linkage; (b) the antisense strand comprises at least one internucleotide linkage chosen from phosphorothioate and methylphosphonate internucleotide linkage; or (c) both the sense strand and the antisense strand each comprise at least one internucleotide linkage chosen from phosphorothioate and methylphosphonate internucleotide linkage.

17. The method of claim 16 , wherein the sense strand comprises two phosphorothioate or methylphosphonate internucleotide linkages at the 5 40 -terminus.

18. The method of claim 16 , wherein the antisense strand comprises two phosphorothioate or methylphosphonate internucleotide linkages at each of the 5 ′-terminus and the 3′-terminus.

19. The method of claim 16 , wherein the sense strand comprises two phosphorothioate or methylphosphonate internucleotide linkages at the 5′-terminus and the antisense strand comprises two phosphorothioate or methylphosphonate internucleotide linkages at each of the 5′-terminus and the 3′-terminus.

20. The method of claim 1 , wherein at least one strand is conjugated to a ligand.

21. The method of claim 20 , wherein the ligand is conjugated to the 3 ′-end of the sense strand.

22. The method of claim 21 , wherein the ligand comprises an N-acetylgalactosamine (GalNAc) derivative.

23. The method of claim 22 , wherein the ligand comprises one or more GalNAc derivatives attached through a monovalent linker, a bivalent branched linker, or a trivalent branched linker.

24. The method of claim 23 , wherein the ligand comprises:

25. The method of claim 24 , wherein the dsRNA agent is conjugated to the ligand as shown in the following schematic:

and, wherein X is O.

26. The method of claim 1 , wherein at least one of the nucleotides of the sense strand and at least one of nucleotides of the antisense strand comprise a modification chosen from a 2′-O-methyl modification and a 2′-fluoro modification.

27. The method of claim 26 , wherein all of the nucleotides of the sense strand and all of the nucleotides of the antisense strand are modified.

28. The method of claim 27 , wherein the sense strand and the antisense strand each contain at least one 2′-O-methyl modification and at least one 2 ′-fluoro modification.

29. The method of claim 28 , wherein the sense strand and/or the antisense strand each independently contain one or two internucleotide linkages chosen from phosphorothioate and methylphosphonate internucleotide linkage at their 5′-termini and/or 3′-termini.

30. The method of claim 29 , wherein:

(a) the sense strand comprises:

(i) four 2′-fluoro modified nucleotides at nucleotides 7 and 9-11, counting from the 5′-end;

(ii) seventeen 2′-O-methyl modified nucleotides at nucleotides 1-6, 8, and 12-21, counting from the 5′-end; and

(iii) phosphorothioate linkages between nucleotides 1 and 2 and between nucleotides 2 and 3, counting from the 5′-end; and

(b) the antisense strand comprises:

(i) three 2′-fluoro modified nucleotides at nucleotides 2, 14, and 16, counting from the 5′-end;

(ii) nineteen 2′-O-methyl modified nucleotides at nucleotides 1, 3-6, 8-13, 15, and 17-23, counting from the 5′-end;

(iii) one glycol modified nucleotide at nucleotide 7, counting from the 5′-end; and

(iv) phosphorothioate linkages between nucleotides 1 and 2, between nucleotides 2 and 3, between nucleotides 21 and 22, and between nucleotides 22 and 23, counting from the 5′-end.

31. The method of claim 30 , wherein the sense strand is conjugated to a ligand, wherein the ligand comprises a multivalent galactose, an N-acetylgalactosamine, an N-acetylgalactosamine derivative, a divalent N-acetylgalactosamine, a trivalent N-acetylgalactosamine, or

32. The method of claim 1 , wherein the dsRNA agent is provided in a pharmaceutical composition.

33. The method of claim 32 , wherein the pharmaceutical composition comprises a buffered solution.

34. The method of claim 33 , wherein the buffered solution comprises acetate, citrate, prolamine, carbonate, phosphate, phosphate buffered saline, or any combination thereof.

35. The method of claim 1 , wherein the sense strand comprises asusgcuuUfuGfCfAfuggacuaucu (SEQ ID NO: 4488) and the antisense strand comprises asGfsauag (Tgn) ccaugcAfaAfagcaususc (SEQ ID NO: 4489),

wherein a, g, c, and u are 2′-O-methyl (2′-OMe) A, G, C, and U, respectively,

wherein Af, Gf, Cf, and Uf are 2′-fluoro A, G, C, and U, respectively,

wherein (Tgn) is thymidine-glycol nucleic acid (GNA),

wherein s is a phosphorothioate linkage, and

wherein a ligand is conjugated to the 3′ end of the sense strand as shown in the following schematic:

wherein X is O.

36. A method of treating a subject suffering from a chronic fibro-inflammatory liver disease, fatty liver disease (steatosis), obesity, or accumulation of fat in the liver, comprising administering to the subject an effective dose of a dsRNA agent comprising a sense strand and an antisense, wherein the nucleotide sequence of the sense strand comprises the nucleotide sequence 5′-asusgcuuUfuGfCfAfuggacuaucu-3′ (SEQ ID NO:4488) and the nucleotide sequence of the antisense strand comprises the nucleotide sequence 5′-asGfsauag (Tgn) ccaugcAfaAfagcaususc-3′ (SEQ ID NO:4489),

wherein a, g, c and u are 2′-O-methyl (2′-OMe) A, G, C, and U, respectively;

wherein Af, Gf, Cf, and Uf are 2′-fluoro A, G, C, and U, respectively,

wherein (Tgn) is thymidine-glycol nucleic acid (GNA),

wherein s is a phosphorothioate linkage, and

wherein a ligand is conjugated to the 3′ end of the sense strand as shown in the following schematic:

wherein X is O.

37. The method of claim 36 , wherein the chronic fibro-inflammatory liver disease comprises nonalcoholic steatohepatitis (NASH), nonalcoholic fatty liver disease (NAFLD), cirrhosis of the liver, alcoholic steatohepatitis (ASH), alcoholic liver diseases (ALD), drug induced liver injury, or hepatocellular necrosis.

38. The method of claim 36 , wherein the chronic fibro-inflammatory liver disease comprises a chronic fibro-inflammatory liver disease associated with the accumulation and/or expansion of lipid droplets in the liver.

39. The method of claim 37 , wherein the cirrhosis of the liver comprises HCV-associated cirrhosis.

Assignments (6)

Continuity (4)

References Cited (308)