CRISPR-targeted genome editing of human induced pluripotent stem cell-derived hepatocytes for the treatment of Wilson’s disease

Background & Aims: Wilson’s disease (WD) is an autosomal recessive disorder of copper metabolism caused by loss-of-function mutations in ATP7B, which encodes a copper-transporting protein. It is characterized by excessive copper deposition in tissues, predominantly in the liver and brain. We...

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Autores principales:	Rui Wei, Jiayin Yang, Chi-Wa Cheng, Wai-In Ho, Na Li, Yang Hu, Xueyu Hong, Jian Fu, Bo Yang, Yuqing Liu, Lixiang Jiang, Wing-Hon Lai, Ka-Wing Au, Wai-Ling Tsang, Yiu-Lam Tse, Kwong-Man Ng, Miguel A. Esteban, Hung-Fat Tse
Formato:	article
Lenguaje:	EN
Publicado:	Elsevier 2022
Materias:	Wilson’s disease induced pluripotent stem cell (iPSC) iPSC-derived hepatocytes (iHeps) ATPase copper transporting beta polypeptide (ATP7B) Clustered regularly interspaced palindromic repeats (CRISPR)/Cas9 Single-stranded Oligodeoxynucleotide (ssODN) Diseases of the digestive system. Gastroenterology RC799-869
Acceso en línea:	https://doaj.org/article/7251fd1424cb470896b40e7153068479
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spelling	oai:doaj.org-article:7251fd1424cb470896b40e71530684792021-11-28T04:37:18ZCRISPR-targeted genome editing of human induced pluripotent stem cell-derived hepatocytes for the treatment of Wilson’s disease2589-555910.1016/j.jhepr.2021.100389https://doaj.org/article/7251fd1424cb470896b40e71530684792022-01-01T00:00:00Zhttp://www.sciencedirect.com/science/article/pii/S2589555921001658https://doaj.org/toc/2589-5559Background & Aims: Wilson’s disease (WD) is an autosomal recessive disorder of copper metabolism caused by loss-of-function mutations in ATP7B, which encodes a copper-transporting protein. It is characterized by excessive copper deposition in tissues, predominantly in the liver and brain. We sought to investigate whether gene-corrected patient-specific induced pluripotent stem cell (iPSC)-derived hepatocytes (iHeps) could serve as an autologous cell source for cellular transplantation therapy in WD. Methods: We first compared the in vitro phenotype and cellular function of ATP7B before and after gene correction using CRISPR/Cas9 and single-stranded oligodeoxynucleotides (ssODNs) in iHeps (derived from patients with WD) which were homozygous for the ATP7B R778L mutation (ATP7BR778L/R778L). Next, we evaluated the in vivo therapeutic potential of cellular transplantation of WD gene-corrected iHeps in an immunodeficient WD mouse model (Atp7b-/- / Rag2-/- / Il2rg-/-; ARG). Results: We successfully created iPSCs with heterozygous gene correction carrying 1 allele of the wild-type ATP7B gene (ATP7BWT/-) using CRISPR/Cas9 and ssODNs. Compared with ATP7BR778L/R778L iHeps, gene-corrected ATP7BWT/- iHeps restored in vitro ATP7B subcellular localization, its subcellular trafficking in response to copper overload and its copper exportation function. Moreover, in vivo cellular transplantation of ATP7BWT/- iHeps into ARG mice via intra-splenic injection significantly attenuated the hepatic manifestations of WD. Liver function improved and liver fibrosis decreased due to reductions in hepatic copper accumulation and consequently copper-induced hepatocyte toxicity. Conclusions: Our findings demonstrate that gene-corrected patient-specific iPSC-derived iHeps can rescue the in vitro and in vivo disease phenotypes of WD. These proof-of-principle data suggest that iHeps derived from gene-corrected WD iPSCs have potential use as an autologous ex vivo cell source for in vivo therapy of WD as well as other inherited liver disorders. Lay summary: Gene correction restored ATP7B function in hepatocytes derived from induced pluripotent stem cells that originated from a patient with Wilson’s disease. These gene-corrected hepatocytes are potential cell sources for autologous cell therapy in patients with Wilson’s disease.Rui WeiJiayin YangChi-Wa ChengWai-In HoNa LiYang HuXueyu HongJian FuBo YangYuqing LiuLixiang JiangWing-Hon LaiKa-Wing AuWai-Ling TsangYiu-Lam TseKwong-Man NgMiguel A. EstebanHung-Fat TseElsevierarticleWilson’s diseaseinduced pluripotent stem cell (iPSC)iPSC-derived hepatocytes (iHeps)ATPase copper transporting beta polypeptide (ATP7B)Clustered regularly interspaced palindromic repeats (CRISPR)/Cas9Single-stranded Oligodeoxynucleotide (ssODN)Diseases of the digestive system. GastroenterologyRC799-869ENJHEP Reports, Vol 4, Iss 1, Pp 100389- (2022)
institution	DOAJ
collection	DOAJ
language	EN
topic	Wilson’s disease induced pluripotent stem cell (iPSC) iPSC-derived hepatocytes (iHeps) ATPase copper transporting beta polypeptide (ATP7B) Clustered regularly interspaced palindromic repeats (CRISPR)/Cas9 Single-stranded Oligodeoxynucleotide (ssODN) Diseases of the digestive system. Gastroenterology RC799-869
spellingShingle	Wilson’s disease induced pluripotent stem cell (iPSC) iPSC-derived hepatocytes (iHeps) ATPase copper transporting beta polypeptide (ATP7B) Clustered regularly interspaced palindromic repeats (CRISPR)/Cas9 Single-stranded Oligodeoxynucleotide (ssODN) Diseases of the digestive system. Gastroenterology RC799-869 Rui Wei Jiayin Yang Chi-Wa Cheng Wai-In Ho Na Li Yang Hu Xueyu Hong Jian Fu Bo Yang Yuqing Liu Lixiang Jiang Wing-Hon Lai Ka-Wing Au Wai-Ling Tsang Yiu-Lam Tse Kwong-Man Ng Miguel A. Esteban Hung-Fat Tse CRISPR-targeted genome editing of human induced pluripotent stem cell-derived hepatocytes for the treatment of Wilson’s disease
description	Background & Aims: Wilson’s disease (WD) is an autosomal recessive disorder of copper metabolism caused by loss-of-function mutations in ATP7B, which encodes a copper-transporting protein. It is characterized by excessive copper deposition in tissues, predominantly in the liver and brain. We sought to investigate whether gene-corrected patient-specific induced pluripotent stem cell (iPSC)-derived hepatocytes (iHeps) could serve as an autologous cell source for cellular transplantation therapy in WD. Methods: We first compared the in vitro phenotype and cellular function of ATP7B before and after gene correction using CRISPR/Cas9 and single-stranded oligodeoxynucleotides (ssODNs) in iHeps (derived from patients with WD) which were homozygous for the ATP7B R778L mutation (ATP7BR778L/R778L). Next, we evaluated the in vivo therapeutic potential of cellular transplantation of WD gene-corrected iHeps in an immunodeficient WD mouse model (Atp7b-/- / Rag2-/- / Il2rg-/-; ARG). Results: We successfully created iPSCs with heterozygous gene correction carrying 1 allele of the wild-type ATP7B gene (ATP7BWT/-) using CRISPR/Cas9 and ssODNs. Compared with ATP7BR778L/R778L iHeps, gene-corrected ATP7BWT/- iHeps restored in vitro ATP7B subcellular localization, its subcellular trafficking in response to copper overload and its copper exportation function. Moreover, in vivo cellular transplantation of ATP7BWT/- iHeps into ARG mice via intra-splenic injection significantly attenuated the hepatic manifestations of WD. Liver function improved and liver fibrosis decreased due to reductions in hepatic copper accumulation and consequently copper-induced hepatocyte toxicity. Conclusions: Our findings demonstrate that gene-corrected patient-specific iPSC-derived iHeps can rescue the in vitro and in vivo disease phenotypes of WD. These proof-of-principle data suggest that iHeps derived from gene-corrected WD iPSCs have potential use as an autologous ex vivo cell source for in vivo therapy of WD as well as other inherited liver disorders. Lay summary: Gene correction restored ATP7B function in hepatocytes derived from induced pluripotent stem cells that originated from a patient with Wilson’s disease. These gene-corrected hepatocytes are potential cell sources for autologous cell therapy in patients with Wilson’s disease.
format	article
author	Rui Wei Jiayin Yang Chi-Wa Cheng Wai-In Ho Na Li Yang Hu Xueyu Hong Jian Fu Bo Yang Yuqing Liu Lixiang Jiang Wing-Hon Lai Ka-Wing Au Wai-Ling Tsang Yiu-Lam Tse Kwong-Man Ng Miguel A. Esteban Hung-Fat Tse
author_facet	Rui Wei Jiayin Yang Chi-Wa Cheng Wai-In Ho Na Li Yang Hu Xueyu Hong Jian Fu Bo Yang Yuqing Liu Lixiang Jiang Wing-Hon Lai Ka-Wing Au Wai-Ling Tsang Yiu-Lam Tse Kwong-Man Ng Miguel A. Esteban Hung-Fat Tse
author_sort	Rui Wei
title	CRISPR-targeted genome editing of human induced pluripotent stem cell-derived hepatocytes for the treatment of Wilson’s disease
title_short	CRISPR-targeted genome editing of human induced pluripotent stem cell-derived hepatocytes for the treatment of Wilson’s disease
title_full	CRISPR-targeted genome editing of human induced pluripotent stem cell-derived hepatocytes for the treatment of Wilson’s disease
title_fullStr	CRISPR-targeted genome editing of human induced pluripotent stem cell-derived hepatocytes for the treatment of Wilson’s disease
title_full_unstemmed	CRISPR-targeted genome editing of human induced pluripotent stem cell-derived hepatocytes for the treatment of Wilson’s disease
title_sort	crispr-targeted genome editing of human induced pluripotent stem cell-derived hepatocytes for the treatment of wilson’s disease
publisher	Elsevier
publishDate	2022
url	https://doaj.org/article/7251fd1424cb470896b40e7153068479
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CRISPR-targeted genome editing of human induced pluripotent stem cell-derived hepatocytes for the treatment of Wilson’s disease

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