Prevention of early-onset cardiomyopathy in Dmd exon 52–54 deletion mice by CRISPR-Cas9-mediated exon skipping

Abstract

Duchenne muscular dystrophy (DMD) is a disease with a life-threatening trajectory resulting from mutations in the dystrophin gene, leading to degeneration of skeletal muscle and fibrosis of cardiac muscle. The overwhelming majority of mutations are multiexonic deletions. We previously established a dystrophic mouse model with deletion of exons 52–54 in Dmd that develops an early-onset cardiac phenotype similar to DMD patients. Here we employed CRISPR-Cas9 delivered intravenously by adeno-associated virus (AAV) vectors to restore functional dystrophin expression via excision or skipping of exon 55. Exon skipping with a solitary guide significantly improved editing outcomes and dystrophin recovery over dual guide excision. Some improvements to genomic and transcript editing levels were observed when the guide dose was enhanced, but dystrophin restoration did not improve considerably. Editing and dystrophin recovery were restricted primarily to cardiac tissue. Remarkably, our exon skipping approach completely prevented onset of the cardiac phenotype in treated mice up to 12 weeks. Thus, our results demonstrate that intravenous delivery of a single-cut CRISPR-Cas9-mediated exon skipping therapy can prevent heart dysfunction in DMD in vivo.

Graphical abstract

Abstract

Rok and colleagues treated a novel Dmd deletion mouse with three unique CRISPR-Cas9 exon skipping strategies delivered by a therapeutically translatable route. Single-cut exon skipping was shown to be effective primarily at restoring dystrophin in the heart, completely preventing onset of cardiomyopathy in treated mice.