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Full-Length Dystrophin Restoration via Targeted Exon Addition in DMD-Patient Specific iPSCs and Cardiomyocytes

Duchenne muscular dystrophy (DMD) is the most common fatal muscle disease, with an estimated incidence of 1/3500–1/5000 male births, and it is associated with mutations in the X-linked DMD gene encoding dystrophin, the largest known human gene. There is currently no cure for DMD. The large size of t...

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Autores principales: Xiao, Rou, Zhou, Miaojin, Wang, Peiyun, Zeng, Baitao, Wu, Lingqian, Hu, Zhiqing, Liang, Desheng
Formato: Online Artículo Texto
Lenguaje:English
Publicado: MDPI 2022
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9409156/
https://www.ncbi.nlm.nih.gov/pubmed/36012442
http://dx.doi.org/10.3390/ijms23169176
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author Xiao, Rou
Zhou, Miaojin
Wang, Peiyun
Zeng, Baitao
Wu, Lingqian
Hu, Zhiqing
Liang, Desheng
author_facet Xiao, Rou
Zhou, Miaojin
Wang, Peiyun
Zeng, Baitao
Wu, Lingqian
Hu, Zhiqing
Liang, Desheng
author_sort Xiao, Rou
collection PubMed
description Duchenne muscular dystrophy (DMD) is the most common fatal muscle disease, with an estimated incidence of 1/3500–1/5000 male births, and it is associated with mutations in the X-linked DMD gene encoding dystrophin, the largest known human gene. There is currently no cure for DMD. The large size of the DMD gene hampers exogenous gene addition and delivery. The genetic correction of DMD patient-derived induced pluripotent stem cells (DMD-iPSCs) and differentiation into suitable cells for transplantation is a promising autologous therapeutic strategy for DMD. In this study, using CRISPR/Cas9, the full-length dystrophin coding sequence was reconstructed in an exon-50-deleted DMD-iPSCs by the targeted addition of exon 50 at the junction of exon 49 and intron 49 via homologous-directed recombination (HDR), with a high targeting efficiency of 5/15, and the genetically corrected iPSCs were differentiated into cardiomyocytes (iCMs). Importantly, the full-length dystrophin expression and membrane localization were restored in genetically corrected iPSCs and iCMs. Thus, this is the first study demonstrating that full-length dystrophin can be restored in iPSCs and iCMs via targeted exon addition, indicating potential clinical prospects for DMD gene therapy.
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spelling pubmed-94091562022-08-26 Full-Length Dystrophin Restoration via Targeted Exon Addition in DMD-Patient Specific iPSCs and Cardiomyocytes Xiao, Rou Zhou, Miaojin Wang, Peiyun Zeng, Baitao Wu, Lingqian Hu, Zhiqing Liang, Desheng Int J Mol Sci Article Duchenne muscular dystrophy (DMD) is the most common fatal muscle disease, with an estimated incidence of 1/3500–1/5000 male births, and it is associated with mutations in the X-linked DMD gene encoding dystrophin, the largest known human gene. There is currently no cure for DMD. The large size of the DMD gene hampers exogenous gene addition and delivery. The genetic correction of DMD patient-derived induced pluripotent stem cells (DMD-iPSCs) and differentiation into suitable cells for transplantation is a promising autologous therapeutic strategy for DMD. In this study, using CRISPR/Cas9, the full-length dystrophin coding sequence was reconstructed in an exon-50-deleted DMD-iPSCs by the targeted addition of exon 50 at the junction of exon 49 and intron 49 via homologous-directed recombination (HDR), with a high targeting efficiency of 5/15, and the genetically corrected iPSCs were differentiated into cardiomyocytes (iCMs). Importantly, the full-length dystrophin expression and membrane localization were restored in genetically corrected iPSCs and iCMs. Thus, this is the first study demonstrating that full-length dystrophin can be restored in iPSCs and iCMs via targeted exon addition, indicating potential clinical prospects for DMD gene therapy. MDPI 2022-08-16 /pmc/articles/PMC9409156/ /pubmed/36012442 http://dx.doi.org/10.3390/ijms23169176 Text en © 2022 by the authors. https://creativecommons.org/licenses/by/4.0/Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https://creativecommons.org/licenses/by/4.0/).
spellingShingle Article
Xiao, Rou
Zhou, Miaojin
Wang, Peiyun
Zeng, Baitao
Wu, Lingqian
Hu, Zhiqing
Liang, Desheng
Full-Length Dystrophin Restoration via Targeted Exon Addition in DMD-Patient Specific iPSCs and Cardiomyocytes
title Full-Length Dystrophin Restoration via Targeted Exon Addition in DMD-Patient Specific iPSCs and Cardiomyocytes
title_full Full-Length Dystrophin Restoration via Targeted Exon Addition in DMD-Patient Specific iPSCs and Cardiomyocytes
title_fullStr Full-Length Dystrophin Restoration via Targeted Exon Addition in DMD-Patient Specific iPSCs and Cardiomyocytes
title_full_unstemmed Full-Length Dystrophin Restoration via Targeted Exon Addition in DMD-Patient Specific iPSCs and Cardiomyocytes
title_short Full-Length Dystrophin Restoration via Targeted Exon Addition in DMD-Patient Specific iPSCs and Cardiomyocytes
title_sort full-length dystrophin restoration via targeted exon addition in dmd-patient specific ipscs and cardiomyocytes
topic Article
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9409156/
https://www.ncbi.nlm.nih.gov/pubmed/36012442
http://dx.doi.org/10.3390/ijms23169176
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