Base editing of PCSK9 and methods of using same for treatment of disease
Inventors
Chadwick, Alexandra • Rajeev, Kallanthottathil G. • Rohde, Ellen • Cheng, Christopher • Reiss, Caroline
Assignees
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Publication Number
US-12029795-B2
Publication Date
2024-07-09
Expiration Date
Abstract
Provided herein are compositions for gene modification or editing and methods of using same to treat or prevent certain conditions. Specific compositions and methods capable of safely and effectively editing gene targets expressed in the liver to durably lower LDL-C thereby treating a leading cause of cardiovascular disease are disclosed.
Core Innovation
The disclosure provides compositions for editing a gene target comprising an mRNA encoding a base editor protein and a guide RNA. The base editor protein comprises a DNA binding domain and a deaminase, and the mRNA comprises a sequence having at least 95% sequence identity to SEQ ID NO: 2192 or to the coding sequence of SEQ ID NO: 2192. The guide RNA comprises a tracr sequence that serves as a binding scaffold for the base editor protein and a spacer sequence corresponding to a protospacer on a target gene, where the target gene is PCSK9.
The disclosure further describes engineering of adenosine deaminase variants, programmable DNA-binding protein configurations including CRISPR/Cas systems, and guide RNA architectures using tracrRNA as a binding scaffold together with a spacer sequence corresponding to a protospacer. It also describes chemical modifications for nucleic-acid molecules and sequence-identity or GC-content constraints for mRNA and guide RNA components. The disclosed embodiments include splice-site targeting, including splice donor or splice acceptor sites in PCSK9, with positional detail referenced by sequence identifiers.
The disclosure also presents preclinical and therapeutic context for PCSK9 and ANGPTL3 base editing, including in vivo liver base editing, reductions in circulating PCSK9 and ANGPTL3 protein, and reductions in LDL-C and triglycerides. The disclosed compositions are further associated with lipid-related biomarkers and therapeutic use concepts in liver cells or hepatocytes.
Claims Coverage
Two independent claims are identified across the provided items. Both are directed to a PCSK9 gene-editing composition comprising an mRNA encoding a base editor protein with a DNA binding domain and a deaminase, together with a guide RNA comprising a tracr sequence and a spacer sequence corresponding to a PCSK9 protospacer; dependent features add sequence identity, GC-content, guide RNA modification, ratio, and splice-site constraints.
Pcsk9 base editing composition with base editor mRNA and tracr-spacer guide RNA
A composition for editing a gene target comprising an mRNA encoding a base editor protein comprising a DNA binding domain and a deaminase, wherein the mRNA comprises a sequence having at least 95% sequence identity to SEQ ID NO: 2192; and a guide RNA comprising a tracr sequence that serves as a binding scaffold for the base editor protein and a spacer sequence that corresponds to a protospacer on a target gene, wherein the target gene is PCSK9.
Pcsk9 base editing composition with coding-sequence identity base editor mRNA and tracr-spacer guide RNA
A composition for editing a gene target comprising an mRNA encoding a base editor protein comprising a DNA binding domain and a deaminase, wherein the mRNA comprises a sequence having at least 95% sequence identity to the coding sequence of SEQ ID NO: 2192; and a guide RNA comprising a tracr sequence that serves as a binding scaffold for the base editor protein and a spacer sequence that corresponds to a protospacer on a target gene, wherein the target gene is PCSK9.
Across the independent claims, the inventive coverage centers on a PCSK9-editing composition defined by an mRNA encoding a base editor protein with a DNA binding domain and a deaminase, meeting a 95% sequence-identity requirement to SEQ ID NO: 2192 or its coding sequence, together with a tracr-scaffolded guide RNA whose spacer corresponds to a PCSK9 protospacer.
Stated Advantages
Increases stability of the guide RNA alone and of base-editor/Cas complexes, including base-editor fusion complexes, with quantified stability enhancements measured in vitro, ex vivo, and/or in vivo.
Reductions in PCSK9 protein and plasma lipid-related biomarkers, including LDL-C, triglycerides, and lipoprotein(a), after in vivo liver base editing.
Documented Applications
Gene editing of the PCSK9 gene using a composition comprising an mRNA encoding a base editor protein and a guide RNA with a tracr sequence and spacer corresponding to a PCSK9 protospacer.
Therapeutic use for directing nucleobase alterations in PCSK9 and ANGPTL3 in liver cells or hepatocytes, reducing circulating PCSK9 and ANGPTL3 protein, LDL-C, and triglycerides.
In vivo liver base editing in cynomolgus monkeys with reported reductions of PCSK9 protein and plasma biomarkers including LDL-C, triglycerides, and lipoprotein(a).
Preclinical evaluation of ABE/LNP base editing in non-human primates and related in vitro work, including ANGPTL3 editing and dual-target PCSK9+ANGPTL3 editing using a single lipid nanoparticle.
Use framed for cardiovascular outcomes related to atherosclerotic cardiovascular disease, metabolic syndrome, and diabetes-related rationale.
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