CRISPR/Cas9-mediated genome editing via postnatal administration of AAV vector cures haemophilia B mice

Abstract Haemophilia B, a congenital haemorrhagic disease caused by mutations in coagulation factor IX gene (F9), is considered an appropriate target for genome editing technology. Here, we describe treatment strategies for haemophilia B mice using the clustered regularly interspaced short palindrom...

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Autores principales:	Tsukasa Ohmori, Yasumitsu Nagao, Hiroaki Mizukami, Asuka Sakata, Shin-ichi Muramatsu, Keiya Ozawa, Shin-ichi Tominaga, Yutaka Hanazono, Satoshi Nishimura, Osamu Nureki, Yoichi Sakata
Formato:	article
Lenguaje:	EN
Publicado:	Nature Portfolio 2017
Materias:	Medicine R Science Q
Acceso en línea:	https://doaj.org/article/b79acbb0c9004973b1526e004ab7e566
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spelling	oai:doaj.org-article:b79acbb0c9004973b1526e004ab7e5662021-12-02T16:06:38ZCRISPR/Cas9-mediated genome editing via postnatal administration of AAV vector cures haemophilia B mice10.1038/s41598-017-04625-52045-2322https://doaj.org/article/b79acbb0c9004973b1526e004ab7e5662017-06-01T00:00:00Zhttps://doi.org/10.1038/s41598-017-04625-5https://doaj.org/toc/2045-2322Abstract Haemophilia B, a congenital haemorrhagic disease caused by mutations in coagulation factor IX gene (F9), is considered an appropriate target for genome editing technology. Here, we describe treatment strategies for haemophilia B mice using the clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9 system. Administration of adeno-associated virus (AAV) 8 vector harbouring Staphylococcus aureus Cas9 (SaCas9) and single guide RNA (sgRNA) to wild-type adult mice induced a double-strand break (DSB) at the target site of F9 in hepatocytes, sufficiently developing haemophilia B. Mutation-specific gene editing by simultaneous induction of homology-directed repair (HDR) sufficiently increased FIX levels to correct the disease phenotype. Insertion of F9 cDNA into the intron more efficiently restored haemostasis via both processes of non-homologous end-joining (NHEJ) and HDR following DSB. Notably, these therapies also cured neonate mice with haemophilia, which cannot be achieved with conventional gene therapy with AAV vector. Ongoing haemophilia therapy targeting the antithrombin gene with antisense oligonucleotide could be replaced by SaCas9/sgRNA-expressing AAV8 vector. Our results suggest that CRISPR/Cas9-mediated genome editing using an AAV8 vector provides a flexible approach to induce DSB at target genes in hepatocytes and could be a good strategy for haemophilia gene therapy.Tsukasa OhmoriYasumitsu NagaoHiroaki MizukamiAsuka SakataShin-ichi MuramatsuKeiya OzawaShin-ichi TominagaYutaka HanazonoSatoshi NishimuraOsamu NurekiYoichi SakataNature PortfolioarticleMedicineRScienceQENScientific Reports, Vol 7, Iss 1, Pp 1-11 (2017)
institution	DOAJ
collection	DOAJ
language	EN
topic	Medicine R Science Q
spellingShingle	Medicine R Science Q Tsukasa Ohmori Yasumitsu Nagao Hiroaki Mizukami Asuka Sakata Shin-ichi Muramatsu Keiya Ozawa Shin-ichi Tominaga Yutaka Hanazono Satoshi Nishimura Osamu Nureki Yoichi Sakata CRISPR/Cas9-mediated genome editing via postnatal administration of AAV vector cures haemophilia B mice
description	Abstract Haemophilia B, a congenital haemorrhagic disease caused by mutations in coagulation factor IX gene (F9), is considered an appropriate target for genome editing technology. Here, we describe treatment strategies for haemophilia B mice using the clustered regularly interspaced short palindromic repeat (CRISPR)/Cas9 system. Administration of adeno-associated virus (AAV) 8 vector harbouring Staphylococcus aureus Cas9 (SaCas9) and single guide RNA (sgRNA) to wild-type adult mice induced a double-strand break (DSB) at the target site of F9 in hepatocytes, sufficiently developing haemophilia B. Mutation-specific gene editing by simultaneous induction of homology-directed repair (HDR) sufficiently increased FIX levels to correct the disease phenotype. Insertion of F9 cDNA into the intron more efficiently restored haemostasis via both processes of non-homologous end-joining (NHEJ) and HDR following DSB. Notably, these therapies also cured neonate mice with haemophilia, which cannot be achieved with conventional gene therapy with AAV vector. Ongoing haemophilia therapy targeting the antithrombin gene with antisense oligonucleotide could be replaced by SaCas9/sgRNA-expressing AAV8 vector. Our results suggest that CRISPR/Cas9-mediated genome editing using an AAV8 vector provides a flexible approach to induce DSB at target genes in hepatocytes and could be a good strategy for haemophilia gene therapy.
format	article
author	Tsukasa Ohmori Yasumitsu Nagao Hiroaki Mizukami Asuka Sakata Shin-ichi Muramatsu Keiya Ozawa Shin-ichi Tominaga Yutaka Hanazono Satoshi Nishimura Osamu Nureki Yoichi Sakata
author_facet	Tsukasa Ohmori Yasumitsu Nagao Hiroaki Mizukami Asuka Sakata Shin-ichi Muramatsu Keiya Ozawa Shin-ichi Tominaga Yutaka Hanazono Satoshi Nishimura Osamu Nureki Yoichi Sakata
author_sort	Tsukasa Ohmori
title	CRISPR/Cas9-mediated genome editing via postnatal administration of AAV vector cures haemophilia B mice
title_short	CRISPR/Cas9-mediated genome editing via postnatal administration of AAV vector cures haemophilia B mice
title_full	CRISPR/Cas9-mediated genome editing via postnatal administration of AAV vector cures haemophilia B mice
title_fullStr	CRISPR/Cas9-mediated genome editing via postnatal administration of AAV vector cures haemophilia B mice
title_full_unstemmed	CRISPR/Cas9-mediated genome editing via postnatal administration of AAV vector cures haemophilia B mice
title_sort	crispr/cas9-mediated genome editing via postnatal administration of aav vector cures haemophilia b mice
publisher	Nature Portfolio
publishDate	2017
url	https://doaj.org/article/b79acbb0c9004973b1526e004ab7e566
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CRISPR/Cas9-mediated genome editing via postnatal administration of AAV vector cures haemophilia B mice

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