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Here, capitalizing on exonic protospacer adjacent motif\u2013altering (PAM-altering) SNP (PAS), we developed an allele-specific CRISPR\/Cas9 strategy to permanently inactivate mutant\n                    <jats:italic>HTT<\/jats:italic>\n                    through nonsense-mediated decay (NMD). Comprehensive sequence\/haplotype analysis identified SNP-generated NGG PAM sites on exons of common\n                    <jats:italic>HTT<\/jats:italic>\n                    haplotypes in HD subjects, revealing a clinically relevant PAS-based mutant-specific CRISPR\/Cas9 strategy. Alternative allele of rs363099 (29th exon) eliminates the NGG PAM site on the most frequent normal\n                    <jats:italic>HTT<\/jats:italic>\n                    haplotype in HD, permitting mutant\n                    <jats:italic>-<\/jats:italic>\n                    specific CRISPR\/Cas9 therapeutics in a predicted ~20% of HD subjects with European ancestry. Our rs363099-based CRISPR\/Cas9 showed perfect allele specificity and good targeting efficiencies in patient-derived cells. Dramatically reduced mutant\n                    <jats:italic>HTT<\/jats:italic>\n                    mRNA and complete loss of mutant protein suggest that our allele-specific CRISPR\/Cas9 strategy inactivates mutant\n                    <jats:italic>HTT<\/jats:italic>\n                    through NMD. In addition, GUIDE-Seq analysis and subsequent validation experiments support high levels of on-target gene specificity. Our data demonstrate a significant target population, complete mutant specificity, decent targeting efficiency in patient-derived cells, and minimal off-target effects on protein-coding genes, proving the concept of PAS-based allele\n                    <jats:italic>-<\/jats:italic>\n                    specific NMD-CRISPR\/Cas9 and supporting its therapeutic potential in HD.\n                  <\/jats:p>","DOI":"10.1172\/jci.insight.141042","type":"journal-article","created":{"date-parts":[[2022,8,30]],"date-time":"2022-08-30T12:01:58Z","timestamp":1661860918000},"source":"Crossref","is-referenced-by-count":35,"title":["Allele-specific silencing of the gain-of-function mutation in Huntington\u2019s disease using CRISPR\/Cas9"],"prefix":"10.1172","volume":"7","author":[{"given":"Jun Wan","family":"Shin","sequence":"first","affiliation":[{"name":"Center for Genomic Medicine, Massachusetts General Hospital, Boston, Massachusetts, USA."},{"name":"Department of Neurology, Harvard Medical School, Boston, Massachusetts, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7789-686X","authenticated-orcid":true,"given":"Eun Pyo","family":"Hong","sequence":"additional","affiliation":[{"name":"Center for Genomic Medicine, Massachusetts General Hospital, Boston, Massachusetts, USA."},{"name":"Department of Neurology, Harvard Medical School, Boston, Massachusetts, USA."},{"name":"Medical and Population Genetics Program, The Broad Institute of M.I.T. and Harvard, Cambridge, Massachusetts, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Seri S.","family":"Park","sequence":"additional","affiliation":[{"name":"Center for Genomic Medicine, Massachusetts General Hospital, Boston, Massachusetts, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Doo Eun","family":"Choi","sequence":"additional","affiliation":[{"name":"Center for Genomic Medicine, Massachusetts General Hospital, Boston, Massachusetts, USA."},{"name":"Department of Neurology, Harvard Medical School, Boston, Massachusetts, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Ihn Sik","family":"Seong","sequence":"additional","affiliation":[{"name":"Center for Genomic Medicine, Massachusetts General Hospital, Boston, Massachusetts, USA."},{"name":"Department of Neurology, Harvard Medical School, Boston, Massachusetts, USA."},{"name":"Medical and Population Genetics Program, The Broad Institute of M.I.T. and Harvard, Cambridge, Massachusetts, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-3382-1134","authenticated-orcid":true,"given":"Madelynn N.","family":"Whittaker","sequence":"additional","affiliation":[{"name":"Center for Genomic Medicine, Massachusetts General Hospital, Boston, Massachusetts, USA."},{"name":"Department of Pathology, Massachusetts General Hospital, Boston, Massachusetts, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0002-5469-0655","authenticated-orcid":true,"given":"Benjamin P.","family":"Kleinstiver","sequence":"additional","affiliation":[{"name":"Center for Genomic Medicine, Massachusetts General Hospital, Boston, Massachusetts, USA."},{"name":"Department of Pathology, Massachusetts General Hospital, Boston, Massachusetts, USA."},{"name":"Department of Pathology, Harvard Medical School, Boston, Massachusetts, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"ORCID":"https:\/\/orcid.org\/0000-0001-7104-9555","authenticated-orcid":true,"given":"Richard Z.","family":"Chen","sequence":"additional","affiliation":[{"name":"CHDI Foundation, Princeton, New Jersey, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jong-Min","family":"Lee","sequence":"additional","affiliation":[{"name":"Center for Genomic Medicine, Massachusetts General Hospital, Boston, Massachusetts, USA."},{"name":"Department of Neurology, Harvard Medical School, Boston, Massachusetts, USA."},{"name":"Medical and Population Genetics Program, The Broad Institute of M.I.T. and Harvard, Cambridge, Massachusetts, USA."}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"232","published-online":{"date-parts":[[2022,10,10]]},"reference":[{"key":"B1","doi-asserted-by":"publisher","DOI":"10.1016\/0092-8674(93)90585-E"},{"key":"B2","doi-asserted-by":"publisher","DOI":"10.1038\/nrdp.2015.5"},{"key":"B3","doi-asserted-by":"publisher","DOI":"10.1212\/WNL.0b013e318249f683"},{"key":"B4","doi-asserted-by":"publisher","DOI":"10.1016\/j.cell.2015.07.003"},{"key":"B5","doi-asserted-by":"crossref","unstructured":"Genetic Modifiers of Huntington\u2019s Disease (GeM-HD) Consortium, et al. 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