PMID- 31327755 OWN - NLM STAT- MEDLINE DCOM- 20200616 LR - 20240104 IS - 1525-0024 (Electronic) IS - 1525-0016 (Print) IS - 1525-0016 (Linking) VI - 27 IP - 9 DP - 2019 Sep 4 TI - AAV9 Edits Muscle Stem Cells in Normal and Dystrophic Adult Mice. PG - 1568-1585 LID - S1525-0016(19)30307-7 [pii] LID - 10.1016/j.ymthe.2019.06.012 [doi] AB - CRISPR editing of muscle stem cells (MuSCs) with adeno-associated virus serotype-9 (AAV9) holds promise for sustained gene repair therapy for muscular dystrophies. However, conflicting evidence exists on whether AAV9 transduces MuSCs. To rigorously address this question, we used a muscle graft model. The grafted muscle underwent complete necrosis before regenerating from its MuSCs. We injected AAV9.Cre into Ai14 mice. These mice express tdTomato upon Cre-mediated removal of a floxed stop codon. About 28%-47% and 24%-89% of Pax7(+) MuSCs expressed tdTomato in pre-grafts and regenerated grafts (p > 0.05), respectively, suggesting AAV9 efficiently transduced MuSCs, and AAV9-edited MuSCs renewed successfully. Robust MuSC transduction was further confirmed by delivering AAV9.Cre to Pax7-ZsGreen-Ai14 mice in which Pax7(+) MuSCs are genetically labeled by ZsGreen. Next, we co-injected AAV9.Cas9 and AAV9.gRNA to dystrophic mdx mice to repair the mutated dystrophin gene. CRISPR-treated and untreated muscles were grafted to immune-deficient, dystrophin-null NSG.mdx4cv mice. Grafts regenerated from CRISPR-treated muscle contained the edited genome and yielded 2.7-fold more dystrophin(+) cells (p = 0.015). Importantly, increased dystrophin expression was not due to enhanced formation of revertant fibers or de novo transduction by residual CRISPR vectors in the graft. We conclude that AAV9 effectively transduces MuSCs. AAV9 CRISPR editing of MuSCs may provide enduring therapy. CI - Copyright (c) 2019 The American Society of Gene and Cell Therapy. All rights reserved. FAU - Nance, Michael E AU - Nance ME AD - Department of Molecular Microbiology and Immunology, School of Medicine, University of Missouri, Columbia, MO 65212, USA. FAU - Shi, Ruicheng AU - Shi R AD - Department of Biomedical, Biological and Chemical Engineering, College of Engineering, University of Missouri, Columbia, MO 65212, USA. FAU - Hakim, Chady H AU - Hakim CH AD - Department of Molecular Microbiology and Immunology, School of Medicine, University of Missouri, Columbia, MO 65212, USA; National Center for Advancing Translational Sciences, NIH, Rockville, MD 20850, USA. FAU - Wasala, Nalinda B AU - Wasala NB AD - Department of Molecular Microbiology and Immunology, School of Medicine, University of Missouri, Columbia, MO 65212, USA. FAU - Yue, Yongping AU - Yue Y AD - Department of Molecular Microbiology and Immunology, School of Medicine, University of Missouri, Columbia, MO 65212, USA. FAU - Pan, Xiufang AU - Pan X AD - Department of Molecular Microbiology and Immunology, School of Medicine, University of Missouri, Columbia, MO 65212, USA. FAU - Zhang, Tracy AU - Zhang T AD - The Hugo W. Moser Research Institute, Kennedy Krieger Institute, Baltimore, MD 21205, USA; Department of Neurology and Neuroscience, Johns Hopkins School of Medicine, Baltimore, MD 21205, USA. FAU - Robinson, Carolyn A AU - Robinson CA AD - Department of Molecular Microbiology and Immunology, School of Medicine, University of Missouri, Columbia, MO 65212, USA. FAU - Duan, Sean X AU - Duan SX AD - Department of Molecular Microbiology and Immunology, School of Medicine, University of Missouri, Columbia, MO 65212, USA. FAU - Yao, Gang AU - Yao G AD - Department of Biomedical, Biological and Chemical Engineering, College of Engineering, University of Missouri, Columbia, MO 65212, USA. FAU - Yang, N Nora AU - Yang NN AD - National Center for Advancing Translational Sciences, NIH, Rockville, MD 20850, USA. FAU - Chen, Shi-Jie AU - Chen SJ AD - Department of Physics, University of Missouri, Columbia, MO 65212, USA. FAU - Wagner, Kathryn R AU - Wagner KR AD - The Hugo W. Moser Research Institute, Kennedy Krieger Institute, Baltimore, MD 21205, USA; Department of Neurology and Neuroscience, Johns Hopkins School of Medicine, Baltimore, MD 21205, USA. FAU - Gersbach, Charles A AU - Gersbach CA AD - Department of Biomedical Engineering, Duke University, Durham, NC 27708, USA. FAU - Duan, Dongsheng AU - Duan D AD - Department of Molecular Microbiology and Immunology, School of Medicine, University of Missouri, Columbia, MO 65212, USA; Department of Biomedical, Biological and Chemical Engineering, College of Engineering, University of Missouri, Columbia, MO 65212, USA; Department of Biomedical Sciences, College of Veterinary Medicine, University of Missouri, Columbia, MO 65211, USA; Department of Neurology, School of Medicine, University of Missouri, Columbia, MO 65212, USA. Electronic address: duand@missouri.edu. LA - eng GR - R01 AR069085/AR/NIAMS NIH HHS/United States GR - R01 GM063732/GM/NIGMS NIH HHS/United States GR - R01 GM117059/GM/NIGMS NIH HHS/United States PT - Journal Article PT - Research Support, N.I.H., Extramural PT - Research Support, U.S. Gov't, Non-P.H.S. DEP - 20190703 PL - United States TA - Mol Ther JT - Molecular therapy : the journal of the American Society of Gene Therapy JID - 100890581 RN - 0 (Dystrophin) RN - 0 (RNA, Guide, CRISPR-Cas Systems) SB - IM MH - Animals MH - Clustered Regularly Interspaced Short Palindromic Repeats MH - Dependovirus/*genetics MH - Disease Models, Animal MH - Dystrophin/chemistry/*genetics MH - *Gene Editing MH - Gene Expression MH - Gene Transfer Techniques MH - Genes, Reporter MH - Genetic Vectors/*genetics MH - Mice MH - Mice, Knockout MH - Muscle, Skeletal/metabolism/pathology MH - Muscular Dystrophy, Duchenne/genetics/therapy MH - Myoblasts/*metabolism MH - RNA, Guide, CRISPR-Cas Systems/genetics MH - Regeneration MH - Satellite Cells, Skeletal Muscle/metabolism MH - Transduction, Genetic PMC - PMC6731180 OTO - NOTNLM OT - AAV OT - Ai14 OT - CRISPR OT - Cas9 OT - Cre OT - DMD OT - MuSC OT - Pax7 OT - Pax7-ZsGreen OT - dystrophin OT - gRNA OT - gene editing OT - mdx OT - muscle OT - muscle graft OT - muscle stem cell OT - regeneration OT - satellite cell OT - stem cell OT - stem cell renewal EDAT- 2019/07/23 06:00 MHDA- 2020/06/17 06:00 PMCR- 2020/09/04 CRDT- 2019/07/23 06:00 PHST- 2019/01/27 00:00 [received] PHST- 2019/06/07 00:00 [revised] PHST- 2019/06/19 00:00 [accepted] PHST- 2019/07/23 06:00 [pubmed] PHST- 2020/06/17 06:00 [medline] PHST- 2019/07/23 06:00 [entrez] PHST- 2020/09/04 00:00 [pmc-release] AID - S1525-0016(19)30307-7 [pii] AID - 10.1016/j.ymthe.2019.06.012 [doi] PST - ppublish SO - Mol Ther. 2019 Sep 4;27(9):1568-1585. doi: 10.1016/j.ymthe.2019.06.012. Epub 2019 Jul 3.