Compositions and methods for inhibiting expression of the PCSK9 gene
View Patent ↗The invention relates to a double-stranded ribonucleic acid (dsRNA) for inhibiting the expression of the PCSK9 gene (PCSK9 gene), comprising an antisense strand having a nucleotide sequence which is less that 30 nucleotides in length, generally 19-25 nucleotides in length, and which is substantially complementary to at least a part of the PCSK9 gene. The invention also relates to a pharmaceutical composition comprising the dsRNA together with a pharmaceutically acceptable carrier and method for treating diseases caused by PCSK9 gene expression.
1. A double-stranded ribonucleic acid (dsRNA) for inhibiting expression of a human proprotein convertase subtilisin kexin 9 (PCSK9) gene in a cell, wherein the dsRNA comprises a sense strand and an antisense strand complementary to at least 15 contiguous nucleotides of a PCSK9 gene and comprises a duplex structure between 15 and 30 base pairs in length, wherein the sense strand comprises at least 15 contiguous nucleotides of the nucleotide sequence of SEQ ID NO: 455.
2. The dsRNA of claim 1 comprising a duplex structure between 19 and 21 base pairs in length.
3. The dsRNA of claim 1 , consisting of a sense strand comprising the nucleotide sequence of SEQ ID NO:455 and an antisense strand comprising the nucleotide sequence of SEQ ID NO:456.
4. The dsRNA of claim 1 , consisting of a modified dsRNA, wherein the sense stand and antisense comprises a terminal 3′ sequence TsT, wherein the terminal thymine comprises a 3′-O-5′-O phosphorothiodiester as indicated by a lower case “s”.
5. The dsRNA of claim 1 , wherein the dsRNA comprises at least one modified nucleotide.
6. The dsRNA of claim 1 , wherein the dsRNA comprises at least one 2′-O-methyl modified nucleotide and at least one nucleotide comprising a 5′-phosphorothioate group.
7. The dsRNA claim 1 , wherein the dsRNA comprises at least one modified nucleotide, wherein the modified nucleotide is chosen from the group of: a 2′-O-methyl modified nucleotide, a 2′-deoxy-2′-fluoro modified nucleotide, a 2′-deoxy-modified nucleotide, a locked nucleotide, an abasic nucleotide, 2′-amino-modified nucleotide, 2′-alkyl-modified nucleotide, a morpholino nucleotide, a phosphoramidate, a nucleotide comprising a 5′-phosphorothioate group, a terminal nucleotide linked to a cholesteryl derivative or dodecanoic acid bisdecylamide group, and a non-natural base comprising nucleotide.
8. An isolated cell comprising the dsRNA of claim 1 .
9. A pharmaceutical composition comprising the dsRNA of claim 1 and a pharmaceutically acceptable carrier.
10. A composition comprising the dsRNA of claim 1 and a lipid formulation.
11. A vector comprising a regulatory sequence operably linked to a nucleotide sequence that encodes at least one strand of the dsRNA of claim 1 .
12. An isolated cell comprising the vector of claim 11 .
13. The dsRNA of claim 1 , wherein contacting a cell in vitro with 30 nM or less of the dsRNA and maintaining the cell for a time sufficient to obtain degradation of a mRNA transcript of a PCSK9 gene, inhibits expression of the PCSK9 gene in the cell.
14. The dsRNA of claim 1 , wherein contacting HepG2 cells expressing the PCSK9 gene in vitro with the dsRNA and maintaining the cells for a time sufficient to obtain degradation of a mRNA transcript of a PCSK9 gene, inhibits expression of the PCSK9 gene in the cell by at least 20%.
15. The dsRNA of claim 1 , wherein administering the dsRNA to an animal decreases total serum cholesterol in the animal.
16. A method for inhibiting expression of a proprotein convertase subtilisin kexin 9 (PCSK9) gene in a cell, comprising contacting the cell with the dsRNA of claim 1 and maintaining the cell for a time sufficient to obtain degradation of a mRNA transcript of a PCSK9 gene, thereby inhibiting expression of the PCSK9 gene in the cell.
17. A method of treating or managing pathological processes which can be mediated by down regulating expression of a proprotein convertase subtilisin kexin 9 (PCSK9) gene comprising administering to a patient in need of such treatment or management a therapeutically effective amount of the dsRNA of claim 1 .
18. A method of treating a proprotein convertase subtilisin kexin 9 (PCSK9) gene-associated disorder comprising administering to a patient in need of such treatment, a therapeutically effective amount of the dsRNA of claim 1 .
19. The dsRNA of claim 1 , wherein the sense strand comprises the nucleotide sequence of SEQ ID NO:455, wherein the dsRNA is selected from the group consisting of the dsRNA comprising sense strand SEQ ID NO 935 (GfgCfcUfgGfaGfuUfuAfuUfcGfgAfTsT) and antisense strand SEQ ID NO 936 (p-uCfcGfaAfuAfaAfcUfcCfaGfgCfcTsT), or the dsRNA comprising sense strand SEQ ID NO 937 (GGCfCfUfGGAGUfUfUfAUfUfCfGGATsT) and antisense strand SEQ ID NO 938 (UfCfCfGAAUfAAACfUfCfCfAGGCfCfTsT), or the dsRNA comprising sense strand SEQ ID NO 939 (GgCcUgGaGuUuAuUcGgATsT) and antisense strand SEQ ID NO 940 (p-uCfcGfaAfuAfaAfcUfcCfaGfgCfcTsT), or the dsRNA comprising sense strand SEQ ID NO 941 (GgCcUgGaGuUuAuUcGgATsT) and antisense strand SEQ ID NO 942 (UfCfCfGAAUfAAACfUfCfCfAGGCfCfTsT), or the dsRNA comprising sense strand SEQ ID NO 943 (GfgCfcUfgGfaGfuUfuAfuUfcGfgAfTsT) and antisense strand SEQ ID NO 944 (UCCGAauAAacUCCAGgccTsT), or the dsRNA comprising sense strand SEQ ID NO 945 (GGCfCfUfGGAGUfUfUfUfAUfUfCfGGATsT) and antisense strand SEQ ID NO 946 (UCCGAauAAacUCCAGgccTsT), or the dsRNA comprising sense strand SEQ ID NO 947 (GgCcUgGaGuUuAuUcGgATsT) and antisense strand SEQ ID NO 948 (UCCGAauAAacUCCAGgccTsT), wherein a strand is modified to include a 2′-O-methyl ribonucleotide as indicated by a lower case letter “a”, “u”, “c” or “g” and a 3′-O-5′-O phosphorothiodiester as indicated by a lower case letter “s,” optionally wherein a strand is modified to include 2′-deoxy-2′-fluoro ribonucleotides as indicated with “Uf,” “Cf,” “Af” or “Gf,” and optionally wherein a strand is modified to include a 5′-phosphate group on the 5′-most nucleotide as indicated by a prefix “p-”.