Compositions and methods for treating disorders of genomic imprinting
Inventors
Chamberlain, Stormy • Cotney, Justin • Langouët, Maéva • Lalande, Marc
Assignees
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Publication Number
US-12018255-B2
Publication Date
2024-06-25
Expiration Date
Abstract
Disclosed herein are compositions, kits, and methods for treating a disorder of genomic imprinting in a subject. The method may include modifying a zinc-finger protein 274 (ZNF274) binding site on maternal chromosome 15 at position 15q11-q13 of the subject, such that the binding of a ZNF274 protein to the ZNF274 binding site is reduced relative to a control. The ZNF274 binding site comprises a polynucleotide having at least 90% sequence identify to SEQ ID NO: 1 or SEQ ID NO: 42. Further provided are DNA targeting systems that bind to a ZNF274 binding site or to a gene encoding a ZNF274 protein.
Core Innovation
The disclosure relates to treating a disorder of genomic imprinting, including Prader-Willi syndrome, by reducing binding of zinc-finger protein 274 (ZNF274) to a ZNF274 binding site on a maternal nucleotide sequence in the PWS critical region (PWSCR) on chromosome 15q11-q13. The ZNF274 binding site is defined by sequence identity to SEQ ID NOs and is used as a target in DNA targeting systems.
The disclosure provides DNA targeting systems comprising at least one guide RNA (gRNA) that binds and targets a polynucleotide sequence corresponding to specified ZNF274 binding site sequences together with a CRISPR-associated (Cas) protein. A DNA targeting system is further described as binding to a gene encoding ZNF274 using at least one gRNA and a Cas protein.
Mechanistic findings using PWS-specific iPSC-derived cells show that ZNF274 knockout reduces H3K9me3 at ZNF274 binding sites in the SNORD116 cluster without altering PWS-IC DNA methylation. The disclosure reports restoration of maternal transcript expression across 15q11-q13/within PWSCR during neuronal differentiation, including 116HGGI/SNORD116, IPW, SNORD115, and SNRPN upstream exons.
Claims Coverage
The provided claim set includes three independent claims covering DNA targeting systems for ZNF274 binding site sequences, DNA targeting systems for a gene encoding ZNF274, and a formulation for treating a genomic imprinting disorder using an agent that reduces maternal ZNF274 binding via a DNA targeting system including specified gRNAs and a Cas protein.
DNA targeting system for ZNF274 binding site sequences
A DNA targeting system that binds to a ZNF274 binding site, comprising at least one gRNA that binds and targets a polynucleotide sequence comprising a nucleotide sequence corresponding to at least one of SEQ ID NO: 1, SEQ ID NO: 42, SEQ ID NO: 43, SEQ ID NO: 44, SEQ ID NO: 45, SEQ ID NO: 46; and a Cas protein.
DNA targeting system for a gene encoding ZNF274
A DNA targeting system that binds to a gene encoding a ZNF274 protein, comprising at least one gRNA that binds and targets a polynucleotide sequence comprising a nucleotide sequence corresponding to at least one of SEQ ID NO: 13, SEQ ID NO: 16, SEQ ID NO: 17, SEQ ID NO: 47, SEQ ID NO: 48; and a Cas protein.
Formulation to reduce maternal ZNF274 binding for genomic imprinting disorder treatment
A formulation for treating a disorder of genomic imprinting in a subject, comprising an agent that reduces relative to a control the binding of a ZNF274 protein to a ZNF274 binding site on a maternal nucleotide sequence, wherein the agent is a DNA targeting system comprising at least one gRNA that binds and targets a polynucleotide sequence comprising a nucleotide sequence corresponding to at least one of SEQ ID NO: 1, SEQ ID NO: 42, SEQ ID NO: 43, SEQ ID NO: 44, SEQ ID NO: 45, SEQ ID NO: 46; and a Cas protein.
Across the independent claims, the core coverage is directed to DNA targeting systems that use gRNAs and a Cas protein to target ZNF274 binding site sequence sets or a gene encoding ZNF274, and to formulations that treat a disorder of genomic imprinting by reducing maternal ZNF274 binding through the specified system.
Stated Advantages
Reduces binding of ZNF274 protein to a ZNF274 binding site on a maternal nucleotide sequence relative to a control.
Restores maternal transcript expression across 15q11-q13/within PWSCR during neuronal differentiation.
Reduces H3K9me3 at ZNF274 binding sites in the SNORD116 cluster without altering PWS-IC DNA methylation.
Documented Applications
Treating a disorder of genomic imprinting in a subject, including Prader-Willi syndrome (PWS).
Treating Prader-Willi syndrome (PWS) by using an agent that reduces relative to a control the binding of ZNF274 protein to a ZNF274 binding site on a maternal nucleotide sequence within 15q11-q13/PWSCR.
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