Compositions and methods for the targeting of PCSK9
AssigneeScribe Therapeutics Inc.
InventorsJason Fernandes, Sean Higgins, Sarah Denny, Ross White, Emeric Jean Marius Charles, Addison Wright, Benjamin Demaree, Benjamin Oakes
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Abstract
Provided herein are gene repressor systems comprising fusion proteins, such as fusion proteins comprising a DNA binding domain such as a TALE, zinc finger or catalytically-dead CRISPR protein and guide nucleic acid (gRNA), which are useful in the repression of a proprotein convertase subtilisin/kexin Type 9 (PCSK9) gene. Also provided are methods of using such systems to repress transcription of PCSK9.
Claims24
A system comprising a repressor fusion protein and a guide ribonucleic acid (gRNA), wherein the repressor fusion protein comprises:
- (a) a catalytically-dead CasX protein (dCasX);
- (b) a repressor domain (RD1) comprising the sequence of any one of SEQ ID NOS:
- 130, 131, 135 and 143;
- (c) a DNA methyltransferase (DNMT) 3A ATRX-DNMT3-DNMT3L domain (ADD) comprising the sequence of SEQ ID NO:
- 125;
- (d) a DNMT3A catalytic domain comprising the sequence of SEQ ID NO:
- 126;
- and
- (e) a DNMT3L comprising the sequence of SEQ ID NO:
- 127,
- wherein the gRNA comprises a scaffold capable of binding to the dCasX and a targeting sequence complementary to a proprotein convertase subtilisin/kexin Type 9 (PCSK9) gene target nucleic acid sequence, wherein the targeting sequence comprises the sequence of any one of SEQ ID NOS:
- 1844, 1852, 1853, 1855, 1858, 1859, 1867, 1869, and 1870,
- wherein the repressor fusion protein is capable of forming a ribonucleoprotein (RNP) with the gRNA, and
- wherein the RNP is capable of repressing transcription of the PCSK9 gene upon binding to the PCSK9 gene target nucleic acid sequence.
A composition comprising:
- (a) a first nucleic acid comprising a guide ribonucleic acid (gRNA) comprising a scaffold capable of binding a catalytically dead CasX (dCasX) protein and a targeting sequence complementary to a proprotein convertase subtilisin/kexin Type 9 (PCSK9) gene target nucleic acid sequence, wherein the targeting sequence comprises the sequence of any one of SEQ ID NOS:
- 1844, 1852, 1853, 1855, 1858, 1859, 1867, 1869, and 1870, and wherein the gRNA is capable of forming a ribonucleoprotein (RNP) with a repressor fusion protein;
- and
- (b) a second nucleic acid encoding the repressor fusion protein, wherein the second nucleic acid is an mRNA encoding:
- i) the catalytically-dead CasX protein (dCasX);
- ii) a repressor domain (RD1) comprising the sequence of any one of SEQ ID NOS:
- 130, 131, 135 and 143;
- iii) a DNA methyltransferase (DNMT) 3A ATRX-DNMT3-DNMT3L domain (ADD) comprising the sequence of SEQ ID NO:
- 125;
- iv) a DNMT3A catalytic domain comprising the sequence of SEQ ID NO:
- 126;
- and
- v) a DNMT3L comprising the sequence of SEQ ID NO:
- 127,
- wherein the RNP is capable of repressing transcription of the PCSK9 gene upon binding to the PCSK9 gene target nucleic acid sequence.
A method of reducing PCSK9-expression in a subject in need thereof, comprising administering to the subject an effective dose of a composition comprising:
- (a) a first nucleic acid comprising a guide ribonucleic acid (gRNA) comprising a scaffold capable of binding a catalytically dead CasX (dCasX) protein and a targeting sequence complementary to a proprotein convertase subtilisin/kexin Type 9 (PCSK9) gene target nucleic acid sequence, wherein the targeting sequence comprises the sequence of any one of SEQ ID NOS:
- 1844, 1852, 1853, 1855, 1858, 1859, 1867, 1869, and 1870, and wherein the gRNA is capable of forming a ribonucleoprotein (RNP) with a repressor fusion protein;
- and
- (b) a second nucleic acid encoding the repressor fusion protein, wherein the second nucleic acid is an mRNA encoding:
- i) the catalytically-dead CasX protein (dCasX);
- ii) a repressor domain (RD1) comprising the sequence of any one of SEQ ID NOS:
- 130, 131, 135 and 143;
- iii) a DNA methyltransferase (DNMT) 3A ATRX-DNMT3-DNMT3L domain (ADD) comprising the sequence of SEQ ID NO:
- 125;
- iv) a DNMT3A catalytic domain comprising the sequence of SEQ ID NO:
- 126;
- and
- v) a DNMT3L comprising the sequence of SEQ ID NO:
- 127,
- wherein the RNP is capable of repressing transcription of the PCSK9 gene upon binding to the PCSK9 gene target nucleic acid sequence.
Dependencies are inferred from the claim text.