Serpina1 iRNA compositions and methods of use thereof

Patent metadata

Field	Value
Publication number	US-10995336-B2
Application number	US-201916420263-A
Country	US
Kind code	B2
Filing date	May 23, 2019
Priority date	Nov 23, 2016
Publication date	May 4, 2021
Grant date	May 4, 2021

Abstract

The invention relates to RNAi agents, e.g., double stranded RNAi agents, targeting the Serpina1 gene, and methods of using such RNAi agents to inhibit expression of Serpina1 and methods of treating subjects having a Serpina1 associated disease, such as a liver disorder.

First claim

We claim:

1. A double stranded RNA (dsRNA) agent that inhibits expression of a serine peptidase inhibitor, clade A, member 1 (Serpina1) gene, comprising a sense strand and an antisense strand forming a double stranded region,
wherein the sense strand comprises at least 19 contiguous nucleotides differing by no more than 3 nucleotides from the nucleotide sequence of 5′-CUUCUUAAUGAUUGAACAAAA-3′ (SEQ ID NO: 417), and the antisense strand comprises at least 19 contiguous nucleotides differing by no more than 3 nucleotides from the nucleotide sequence of 5′-UUUUGUUCAAUCAUUAAGAAGAC-3′ (SEQ ID NO: 419),
wherein substantially all of the nucleotides of the antisense strand comprise a nucleotide modification,
wherein the antisense strand comprises at least one thermally destabilizing modification selected from the group consisting of an abasic modification, a mismatch with the opposing nucleotide in the opposing strand, a 2′-deoxy modification, and an acyclic nucleotide modification, at position 4, 5, 6, 7 and/or 8 from the 5′-end,
wherein the nucleotides at positions 2, 6, 8, 9, 14, and 16, or at positions 2, 6, 14, and 16, or at positions 2, 14, and 16 from the 5′-end of the antisense strand are 2′-F modified nucleotides,
wherein substantially all of the nucleotides of the sense strand comprise a nucleotide modification,
wherein the nucleotides at positions 7, 9, 10, and 11, or at positions 7, 10, and 11 from the 5′-end of the sense strand are 2′-F modified nucleotides,
wherein the sense strand comprises an ASGPR ligand,
wherein each of the sense strand and the antisense strand are independently 19 to 25 nucleotides in length.

2. The dsRNA agent of claim 1 , wherein the thermally destabilizing modification is selected from the group consisting of

wherein B is nucleobase.

3. The dsRNA agent of claim 1 , wherein the ASGPR ligand is one or more GalNAc derivatives attached through a bivalent or trivalent branched linker.

4. A double stranded RNA (dsRNA) agent that inhibits expression of a serine peptidase inhibitor, clade A, member 1 (Serpina1) target gene sequence, comprising a sense strand and an antisense strand forming a double stranded region,
wherein the antisense strand comprises at least 19 contiguous nucleotides differing by no more than 3 nucleotides from the nucleotide sequence of 5′-UUUUGUUCAAUCAUUAAGAAGAC-3′ (SEQ ID NO: 419),
wherein substantially all of the nucleotides of the antisense strand comprise a nucleotide modification,
wherein the antisense strand comprises at least one thermally destabilizing modification selected from the group consisting of an abasic modification, a mismatch with the opposing nucleotide in the opposing strand, a 2′-deoxy modification, and an acyclic nucleotide modification at position 4, 5, 6, 7 and/or 8 from the 5′-end,
wherein the nucleotides at positions 2, 6, 8, 9, 14, and 16, or at positions 2, 6, 14, and 16, or at positions 2, 14, and 16 from the 5′-end of the antisense strand are 2′-F modified nucleotides,
wherein substantially all of the nucleotides of the sense strand comprise a nucleotide modification,
wherein the nucleotides at positions 7, 9, 10, and 11, or at positions 7, 10, and 11 from the 5′-end of the sense strand are 2′-F modified nucleotides,
wherein the sense strand comprises an ASGPR ligand, and
wherein each of the sense strand and the antisense strand are independently 19 to 25 nucleotides in length.

5. The dsRNA agent of claim 4 , wherein the ASGPR ligand is one or more GalNAc derivatives attached through a bivalent or trivalent branched linker.

6. The dsRNA agent of claim 4 , wherein the antisense strand comprises the nucleotide sequence 5′-UUUUGUUCAAUCAUUAAGAAGAC-3′ (SEQ ID NO: 419).

7. An isolated cell containing the dsRNA agent of claim 1 or 4 .

8. A pharmaceutical composition comprising the dsRNA agent of claim 1 or 4 .

9. The dsRNA agent of claim 1 or 4 , wherein the destabilizing modification is located in position 7 of the antisense strand.

10. The dsRNA of claim 1 or 4 , wherein all of the nucleotides of the sense strand and all of the nucleotides of the antisense strand are modified nucleotides.

11. The dsRNA agent of claim 1 or 4 , wherein the ASGPR ligand is:

12. The dsRNA agent of claim 11 , wherein the dsRNA agent is conjugated to the ligand as shown in the following schematic

wherein X is O or S.

13. A double stranded RNA (dsRNA) agent that inhibits expression of a serine peptidase inhibitor, clade A, member 1 (Serpina1) target gene sequence, comprising a sense strand and an antisense strand forming a double stranded region,
wherein the sense strand comprises at least 19 contiguous nucleotides differing by no more than 3 nucleotides from the nucleotide sequence of 5′-CUUCUUAAUGAUUGAACAAAA-3′ (SEQ ID NO: 417), and the antisense strand comprises at least 19 contiguous nucleotides differing by no more than 3 nucleotides from the nucleotide sequence of 5′-UUUUGUUCAAUCAUUAAGAAGAC-3′ (SEQ ID NO: 419),
wherein all of the nucleotides of the antisense strand comprise a nucleotide modification selected from the group consisting of a thermally destabilizing modification, a 2′-F modification and a 2′-OMe modification,
wherein the thermally destabilizing modification is located in position 4 to 8 from the 5′-end of the antisense strand and is selected from the group consisting of an abasic modification, a mismatch with the opposing nucleotide in the opposing strand, a 2′-deoxy modification, and an acyclic nucleotide modification,
wherein the nucleotides at positions 2, 6, 8, 9, 14, and 16, or at positions 2, 6, 14, and 16, or at positions 2, 14, and 16 from the 5′-end of the antisense strand are 2′-F modified nucleotides,
wherein all of the nucleotides of the sense strand comprise a nucleotide modification,
wherein the nucleotides at positions 7, 9, 10, and 11, or at positions 7, 10, and 11 from the 5′-end of the sense strand are 2′-F modified nucleotides,
wherein each of the sense strand and the antisense strand are independently 19 to 25 nucleotides in length, and
wherein the dsRNA agent has a melting temperature of from about 40° C. to about 80° C.

14. A double stranded RNA (dsRNA) agent that inhibits expression of a serine peptidase inhibitor, clade A, member 1 (Serpina1) target gene sequence, comprising a sense strand and an antisense strand forming a double stranded region,
wherein the antisense strand comprises at least 19 contiguous nucleotides differing by no more than 3 nucleotides from the nucleotide sequence of 5′-UUUUGUUCAAUCAUUAAGAAGAC-3′ (SEQ ID NO: 419),
wherein all of the nucleotides of the antisense strand comprise a nucleotide modification selected from the group consisting of a thermally destabilizing modification, a 2′-F modification and a 2′-OMe modification,
wherein the thermally destabilizing modification is located in position 4 to 8 from the 5′-end of the antisense strand and is selected from the group consisting of an abasic modification, a mismatch with the opposing nucleotide in the opposing strand, a 2′-deoxy modification, and an acyclic nucleotide modification,
wherein the nucleotides at positions 2, 6, 8, 9, 14, and 16, or at positions 2, 6, 14, and 16, or at positions 2, 14, and 16 from the 5′-end of the antisense strand are 2′-F modified nucleotides,
wherein all of the nucleotides of the sen

Assignees

• Alnylam Pharmaceuticals Inc

Inventors

• Schlegel Mark K
• Janas Maja
• Jadhav Vasant R
• Foster Donald
• Manoharan Muthiah
• Rajeev Kallanthottathil G
• Kel'In Alexander V
• Charisse Klaus
• Nair Jayaprakash K
• Maier Martin A
• Matsuda Shigeo
• Jayaraman Muthusamy
• Sehgal Alfica
• Brown Christopher
• Fitzgerald Kevin
• Milstein Stuart

Classifications

• C12N15/85

  for animal cells · CPC title

• C12N15/113 Primary

  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 title

• C12N5/10

  Cells modified by introduction of foreign genetic material · CPC title

• A61P35/00

  Antineoplastic agents · CPC title

• C12N2310/351

  Conjugate · CPC title