Patatin-like phospholipase domain containing 3 (PNPLA3) iRNA compositions and methods of use thereof

Patent metadata

Field	Value
Publication number	US-11149276-B2
Application number	US-202117182348-A
Country	US
Kind code	B2
Filing date	Feb 23, 2021
Priority date	Dec 16, 2019
Publication date	Oct 19, 2021
Grant date	Oct 19, 2021

Abstract

The present invention relates to RNAi agents, e.g., double stranded RNA (dsRNA) agents, targeting the Patatin-Like Phospholipase Domain Containing 3 (PNPLA3) gene. The invention also relates to methods of using such RNAi agents to inhibit expression of a PNPLA3 gene and to methods of preventing and treating an PNPLA3-associated disorder, e.g., Nonalcoholic Fatty Liver Disease (NAFLD).

First claim

We claim:

1. A double stranded ribonucleic acid (dsRNA) agent for inhibiting expression of Patatin-Like Phospholipase Domain Containing 3 (PNPLA3) in a cell, wherein the dsRNA agent comprises a sense strand and an antisense strand forming a double stranded region, wherein the sense strand comprises at least 15 contiguous nucleotides differing by no more than 3 nucleotides from the nucleotide sequence of nucleotides 1214-1234 of SEQ ID NO:1, and the antisense strand comprises at least 15 contiguous nucleotides from the nucleotide sequence of nucleotides 1572-1592 of SEQ ID NO:2, wherein all of the nucleotides of the sense strand and all of the nucleotides of the antisense strand comprise a modification selected from the group consisting of a 2′-O-methyl modification and a 2′-fluoro modification, wherein both the sense strand and the antisense strand independently comprise at least one phosphorothioate or methylphosphonate internucleotide linkage, and wherein at least one strand is conjugated to a ligand.

2. The dsRNA agent of claim 1 , wherein the sense strand comprises 2-6 2′-fluoro modified nucleotides.

3. The dsRNA agent of claim 2 , wherein the sense strand comprises 4 2′-fluoro modified nucleotides.

4. The dsRNA agent of claim 1 , wherein the antisense strand comprises 2-8 2′-fluoro modified nucleotides.

5. The dsRNA agent of claim 4 , wherein the antisense strand comprises 6 2′-fluoro modified nucleotides.

6. The dsRNA agent of claim 1 , wherein the sense strand comprises 4 2′-fluoro modified nucleotides at nucleotides 7 and 9-11, counting from the 5′-end, and the antisense strand comprises 6 2′-fluoro modified nucleotides at nucleotides 2, 6, 8, 9, 14 and 16, counting from the 5′-end.

7. The dsRNA agent of claim 1 , wherein the sense strand comprises two phosphorothioate or methylphosphonate internucleotide linkages at the 5′-terminus.

8. The dsRNA agent of claim 1 , wherein the antisense strand comprises two phosphorothioate or methylphosphonate internucleotide linkages at both the 5′- and the 3′-terminus.

9. The dsRNA agent of claim 1 , 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 both the 5′- and the 3′-terminus.

10. The dsRNA agent of claim 1 , wherein the ligand is conjugated to the 3′-end of the sense strand.

11. The dsRNA agent of claim 1 , wherein the ligand is an N-acetylgalactosamine (GalNAc) derivative.

12. The dsRNA agent of claim 11 , wherein the ligand is one or more GalNAc derivatives attached through a monovalent, bivalent, or trivalent branched linker.

13. The dsRNA agent of claim 12 , wherein the ligand is

14. The dsRNA agent of claim 13 , wherein the dsRNA agent is conjugated to the ligand as shown in the following schematic

and, wherein X is O.

15. A double stranded ribonucleic acid (dsRNA) agent for inhibiting expression of Patatin-Like Phospholipase Domain Containing 3 (PNPLA3) in a cell, wherein the dsRNA agent comprises a sense strand and an antisense strand forming a double stranded region, wherein the sense strand differs by no more than 3 modified nucleotides from the nucleotide sequence of 5′-csasuuagGfaUfAfAfugucuuaugu-3′ (SEQ ID NO:2648) and wherein the antisense strand differs by no more than 3 modified nucleotides from the nucleotide sequence of 5′-asCfsauaAfgAfCfauuaUfcCfuaaugsgsg-3′ (SEQ ID NO:2732), wherein a, g, c and u are 2′-O-methyl (2′-OMe) A, G, C, and U respectively; Af, Gf, Cf and Uf are 2′-fluoro A, G, C and U respectively; and s is a phosphorothioate linkage.

16. The dsRNA agent of claim 15 , wherein the sense strand differs by no more than 2 modified nucleotides from the nucleotide sequence of 5′-csasuuagGfaUfAfAfugucuuaugu-3′ (SEQ ID NO:2648) and wherein the antisense strand differs by no more than 2 modified nucleotides from the nucleotide sequence of 5′-asCfsauaAfgAfCfauuaUfcCfuaaugsgsg-3′ (SEQ ID NO:2732).

17. The dsRNA agent of claim 15 , wherein the sense strand differs by no more than 1 modified nucleotide from the nucleotide sequence of 5′-csasuuagGfaUfAfAfugucuuaugu-3′ (SEQ ID NO:2648) and wherein the antisense strand differs by no more than 1 modified nucleotides from the nucleotide sequence of 5′-asCfsauaAfgAfCfauuaUfcCfuaaugsgsg-3′ (SEQ ID NO:2732).

18. The dsRNA agent of claim 15 , wherein the dsRNA agent is conjugated to the ligand as shown in the following schematic

and, wherein X is O.

19. A double stranded ribonucleic acid (dsRNA) agent for inhibiting expression of Patatin-Like Phospholipase Domain Containing 3 (PNPLA3) in a cell, wherein the dsRNA agent comprises a sense strand and an antisense strand forming a double stranded region, wherein the sense strand comprises the nucleotide sequence of 5′-csasuuagGfaUfAfAfugucuuaugu-3′ (SEQ ID NO:2648) and wherein the antisense strand comprises the nucleotide sequence of 5′-asCfsauaAfgAfCfauuaUfcCfuaaugsgsg-3′ (SEQ ID NO:2732), wherein a, g, c and u are 2′-O-methyl (2′-OMe) A, G, C, and U respectively; Af, Gf, Cf and Uf are 2′-fluoro A, G, C and U respectively; and s is a phosphorothioate linkage.

20. The dsRNA agent of claim 19 , wherein the sense strand comprises the nucleotide sequence of 5′-csasuuagGfaUfAfAfugucuuauguL96-3′ (SEQ ID NO:2648) and wherein the antisense strand comprises the nucleotide sequence of 5′-asCfsauaAfgAfCfauuaUfcCfuaaugsgsg-3′ (SEQ ID NO:2732), wherein a, g, c and u are 2′-O-methyl (2′-OMe) A, G, C, and U respectively; Af, Gf, Cf and Uf are 2′-fluoro A, G, C and U respectively; s is a phosphorothioate linkage, and L96 is N-[tris(GalNAc-alkyl)-amidodecanoyl)]-4-hydroxyprolinol.

21. The dsRNA agent of claim 19 , wherein the 3′-end of the sense strand is conjugated to a ligand as shown in the following schematic:

wherein X is O.

22. A double stranded ribonucleic acid (dsRNA) agent for inhibiting expression of Patatin-Like Phospholipase Domain Containing 3 (PNPLA3) in a cell, wherein the dsRNA agent comprises a sense strand and an antisense strand forming a double stranded region, wherein the sense strand consists of the nucleotide sequence of 5′-csasuuagGfaUfAfAfugucuuaugu-3′ (SEQ ID NO:2648) and wherein the antisense strand consists of the nucleotide sequence of 5′-asCfsauaAfgAfCfauuaUfcCfuaaugsgsg-3′ (SEQ ID NO:2732), wherein a, g, c and u are 2′-O-methyl (2′-OMe) A, G, C, and U respectively; Af, Gf, Cf and Uf are 2′-fluoro A, G, C and U respectively; and s is a phosphorothioate linkage; and wherein the 3′-end of the sense strand is conjugated to a ligand as shown in the following schematic:

wherein X is O.

23. A pharmaceutical composition for inhibiting expression of a gene encoding Patatin-Like Phospholipase Domain Containing 3 (PNPLA3) comprising the dsRNA agent of claim 1 .

24. The pharmaceutical composition of claim 23 , wherein dsRNA agent is in an unbuffered solution.

25. The ph

Assignees

• Alnylam Pharmaceuticals Inc

Inventors

• Tremblay Frederic
• Mcininch James D
• Castoreno Adam
• Schlegel Mark K
• Kaittanis Charalambos

Classifications

• C12N15/1137 Primary

  against enzymes (viral enzymes C12N15/1131; receptors C12N15/1138) · CPC title

• C12N2310/351

  Conjugate · CPC title

• C12N2310/3183

  Diol linkers, e.g. glycols or propanediols · CPC title

• C12N2310/322

  2'-R Modification · CPC title

• C12N2310/321

  2'-O-R Modification · CPC title