title: Muscle-specific CRISPR/Cas9 dystrophin gene editing ameliorates pathophysiology in a mouse model for Duchenne muscular dystrophy.
abstract: Gene replacement therapies utilizing adeno-associated viral (AAV) vectors hold great promise for treating Duchenne muscular dystrophy (DMD). A related approach uses AAV vectors to edit specific regions of the DMD gene using CRISPR/Cas9. Here we develop multiple approaches for editing the mutation in dystrophic mdx4cvmice using single and dual AAV vector delivery of a muscle-specific Cas9 cassette together with single-guide RNA cassettes and, in one approach, a dystrophin homology region to fully correct the mutation. Muscle-restricted Cas9 expression enables direct editing of the mutation, multi-exon deletion or complete gene correction via homologous recombination in myogenic cells. Treated muscles express dystrophin in up to 70% of the myogenic area and increased force generation following intramuscular delivery. Furthermore, systemic administration of the vectors results in widespread expression of dystrophin in both skeletal and cardiac muscles. Our results demonstrate that AAV-mediated muscle-specific gene editing has significant potential for therapy of neuromuscular disorders.
authors.affiliation: Department of Neurology, University of Washington, Seattle, Washington 98195-7720, USA.|Senator Paul D. Wellstone Muscular Dystrophy Cooperative Research Center, University of Washington, Seattle, Washington 98195-7720, USA.
authors.forename: Niclas E
authors.initials: NE
authors.lastname: Bengtsson
authors.affiliation: Department of Neurology, University of Washington, Seattle, Washington 98195-7720, USA.|Senator Paul D. Wellstone Muscular Dystrophy Cooperative Research Center, University of Washington, Seattle, Washington 98195-7720, USA.
authors.forename: John K
authors.initials: JK
authors.lastname: Hall
authors.affiliation: Department of Neurology, University of Washington, Seattle, Washington 98195-7720, USA.|Senator Paul D. Wellstone Muscular Dystrophy Cooperative Research Center, University of Washington, Seattle, Washington 98195-7720, USA.
authors.forename: Guy L
authors.initials: GL
authors.lastname: Odom
authors.affiliation: Department of Pathology, University of Washington, Seattle, Washington 98195-7720, USA.
authors.forename: Michael P
authors.initials: MP
authors.lastname: Phelps
authors.affiliation: Department of Medicine, University of Washington, Seattle, Washington 98195-7720, USA.|Department of Genome Sciences, University of Washington, Seattle, Washington 98195-7720, USA.
authors.forename: Colin R
authors.initials: CR
authors.lastname: Andrus
authors.affiliation: Department of Medicine, University of Washington, Seattle, Washington 98195-7720, USA.|Department of Genome Sciences, University of Washington, Seattle, Washington 98195-7720, USA.
authors.forename: R David
authors.initials: RD
authors.lastname: Hawkins
authors.affiliation: Senator Paul D. Wellstone Muscular Dystrophy Cooperative Research Center, University of Washington, Seattle, Washington 98195-7720, USA.|Department of Biochemistry, University of Washington, Seattle, Washington 98195-7720, USA.
authors.forename: Stephen D
authors.initials: SD
authors.lastname: Hauschka
authors.affiliation: Senator Paul D. Wellstone Muscular Dystrophy Cooperative Research Center, University of Washington, Seattle, Washington 98195-7720, USA.|Department of Medicine, University of Washington, Seattle, Washington 98195-7720, USA.
authors.forename: Joel R
authors.initials: JR
authors.lastname: Chamberlain
authors.affiliation: Department of Neurology, University of Washington, Seattle, Washington 98195-7720, USA.|Senator Paul D. Wellstone Muscular Dystrophy Cooperative Research Center, University of Washington, Seattle, Washington 98195-7720, USA.|Department of Medicine, University of Washington, Seattle, Washington 98195-7720, USA.|Department of Biochemistry, University of Washington, Seattle, Washington 98195-7720, USA.
authors.forename: Jeffrey S
authors.initials: JS
authors.lastname: Chamberlain
mesh_terms: D000818:Animals; D001426:Bacterial Proteins / Q000235:genetics; D000076987:CRISPR-Associated Protein 9; D064113:CRISPR-Cas Systems / Q000235:genetics*; D064112:Clustered Regularly Interspaced Short Palindromic Repeats / Q000235:genetics; D000229:Dependovirus / Q000235:genetics; D004195:Disease Models, Animal; D016189:Dystrophin / Q000235:genetics*; D004720:Endonucleases / Q000235:genetics; D000072669:Gene Editing / Q000379:methods*; D015316:Genetic Therapy / Q000379:methods; D005822:Genetic Vectors; D059014:High-Throughput Nucleotide Sequencing; D008297:Male; D051379:Mice; D018101:Mice, Inbred mdx; D018482:Muscle, Skeletal / Q000503:physiopathology; D020388:Muscular Dystrophy, Duchenne / Q000235:genetics* / Q000378:metabolism* / Q000503:physiopathology* / Q000628:therapy; D009154:Mutation; D009206:Myocardium; D009468:Neuromuscular Diseases / Q000628:therapy; D000094704:RNA, Guide, CRISPR-Cas Systems; D017384:Sequence Deletion
chemical_list: D001426:Bacterial Proteins; D016189:Dystrophin; D000094704:RNA, Guide, CRISPR-Cas Systems; D000076987:CRISPR-Associated Protein 9; C000606107:Cas9 endonuclease Streptococcus pyogenes; D004720:Endonucleases