Polynucleotide agents targeting Serpinc1 (AT3) and methods of use thereof
View Patent ↗The invention relates to polynucleotide agents targeting the Serpinc1 (AT3) gene, and methods of using such polynucleotide agents to inhibit expression of Serpinc1 and to treat subjects having a bleeding disorder, e.g., a hemophilia.
1. An antisense polynucleotide agent for inhibiting expression of Serpinc1 (AT3), wherein the agent comprises any one of the nucleotide sequences selected from the group consisting of SEQ ID NOs:197-380 and 565-748, wherein the agent is about 18 to about 50 nucleotides in length, wherein at least one of the nucleotides is a modified nucleotide, and wherein the nucleotide sequence of the agent is about 80% complementary over its entire length to the equivalent region of the nucleotide sequence of SEQ ID NO:1.
2. An antisense polynucleotide agent for inhibiting expression of Serpinc1 (AT3), wherein the agent comprises at least 15 contiguous nucleotides differing by no more than 3 nucleotides from any one of the nucleotide sequences selected from the group consisting of SEQ ID NOs:197-380 and 565-748, wherein the agent is about 18 to about 50 nucleotides in length, and wherein at least one of the contiguous nucleotides is a modified nucleotide.
3. The agent of claim 1 , wherein substantially all of the nucleotides of the antisense polynucleotide agent are modified nucleotides.
4. The agent of claim 1 , which is 22 to 40 nucleotides in length; 30 to 45 nucleotides in length; 18 to 30 nucleotides in length; 25 to 48 nucleotides in length; 18 to 24 nucleotides in length; or 20 to 24 nucleotides in length.
5. The agent of claim 1 , wherein the modified nucleotide comprises a modified sugar moiety selected from the group consisting of a 2′-O-methoxyethyl modified sugar moiety, a 2′-methoxy modified sugar moiety, a 2′-O-alkyl modified sugar moiety, and a bicyclic sugar moiety.
6. The agent of claim 1 , wherein the modified nucleotide is a 5-methylcytosine.
7. The agent of claim 1 , wherein the modified nucleotide comprises a modified internucleoside linkage.
8. The agent of claim 1 , comprising a plurality of 2′-deoxynucleotides flanked on each side by at least one nucleotide having a modified sugar moiety.
9. The agent of claim 8 , wherein the agent is a gapmer comprising a gap segment comprised of linked 2′-deoxynucleotides positioned between a 5′ and a 3′ wing segment.
10. An antisense polynucleotide agent for inhibiting expression of Serpinc1 (AT3), comprising at least 15 contiguous nucleotides differing by no more than 3 nucleotides from any one of the nucleotide sequences selected from the group consisting of SEQ ID NOs:197-380 and 565-748, wherein the agent is about 18 to about 50 nucleotides in length, and wherein the agent comprises
a gap segment consisting of linked deoxynucleotides;
a 5′-wing segment consisting of linked nucleotides;
a 3′-wing segment consisting of linked nucleotides;
wherein the gap segment is positioned between the 5′-wing segment and the 3′-wing segment and wherein each nucleotide of each wing segment comprises a modified sugar moiety.
11. The agent of claim 10 , wherein the gap segment is 9 to 14 2′-deoxynucleotides in length and each of the wing segments is 3 to 6 nucleotides in length.
12. The agent of claim 1 , wherein the agent further comprises a ligand at the 3′-terminus of the agent.
13. The agent of claim 12 , wherein the ligand is an N-acetylgalactosamine (GalNAc) derivative.
14. A pharmaceutical composition for inhibiting expression of a Serpinc1 (AT3) gene comprising the agent of claim 1 .
15. A pharmaceutical composition comprising the agent of claim 1 , and a lipid formulation.
16. The agent of claim 10 , wherein the modified sugar moiety is selected from the group consisting of a 2′-O-methoxyethyl modified sugar moiety, a 2′-methoxy modified sugar moiety, a 2′-O-alkyl modified sugar moiety, and a bicyclic sugar moiety.
17. The agent of claim 10 , further comprising a modified internucleoside linkage.
18. The agent of claim 11 , wherein the 5′-wing segment is 4 to 6 nucleotides in length, the 3′-wing segment is 4 to 6 nucleotides in length, and the gap segment is 9 to 13 nucleotides in length.
19. The agent of claim 1 , comprising the nucleotide sequence 5′-TTGGAATACATGGCCGGCTAA 3′ (SEQ ID NO:206).
20. The agent of claim 19 , comprising 5′-ususgsgsasdAsdTsdAs(5MdC)sdAsdTsdGsdGs(5MdC)s(5MdC)sdGsgscsusasa-3′ (SEQ ID NO:22),
wherein a, c, g, and u are 2′-O-methyl (2′-OMe) A, C, G, and U; Af, Cf, Gf, and Uf are 2′-fluoro A, C, G, and U; dA, dC, dG, and dT are 2′-deoxy (d) A, C, G, and T; (5MdC) is 5′-methyl-deoxycytidine; and s is a phosphorothioate linkage.
21. The pharmaceutical composition of claim 14 , wherein agent is present in an unbuffered solution or a buffer solution.
22. The agent of claim 1 , which is 21-40 nucleotides in length; 21-30 nucleotides in length; or 21-24 nucleotides in length.
23. A pharmaceutical composition for inhibiting expression of a Serpinc1 (AT3) gene comprising the agent of claim 2 .
24. A pharmaceutical composition for inhibiting expression of a Serpinc1 (AT3) gene comprising the agent of claim 10 .
25. A method of inhibiting Serpinc1 (AT3) expression in a cell, the method comprising:
(a) contacting the cell with the agent of any one of claim 1 , 2 , or 10 or a pharmaceutical composition of any one of claim 14 , 23 , or 24 ; and
(b) maintaining the cell produced in step (a) for a time sufficient to obtain antisense inhibition of a Serpinc1 gene, thereby inhibiting expression of Serpinc1 gene in the cell.
26. The method of claim 25 , wherein the cell is within a subject.
27. The method of claim 26 , wherein the subject is a human.