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JCI Insight. 2018 Dec 6;3(23). pii: 124297. doi: 10.1172/jci.insight.124297.

AAV CRISPR editing rescues cardiac and muscle function for 18 months in dystrophic mice.

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Department of Molecular Microbiology and Immunology, School of Medicine, University of Missouri, Columbia, Missouri, USA.
National Center for Advancing Translational Sciences, NIH, Rockville, Maryland, USA.
Department of Biomedical Engineering and.
Center for Genomic and Computational Biology, Duke University, Durham, North Carolina, USA.
Department of Physics and.
Department of Biochemistry, University of Missouri, Columbia, Missouri, USA.
Department of Orthopaedic Surgery, Duke University Medical Center, Durham, North Carolina, USA.
Department of Neurology, School of Medicine.
Department of Bioengineering, and.
Department of Biomedical Sciences, College of Veterinary Medicine, University of Missouri, Columbia, Missouri, USA.


Adeno-associated virus-mediated (AAV-mediated) CRISPR editing is a revolutionary approach for treating inherited diseases. Sustained, often life-long mutation correction is required for treating these diseases. Unfortunately, this has never been demonstrated with AAV CRISPR therapy. We addressed this question in the mdx model of Duchenne muscular dystrophy (DMD). DMD is caused by dystrophin gene mutation. Dystrophin deficiency leads to ambulation loss and cardiomyopathy. We treated 6-week-old mice intravenously and evaluated disease rescue at 18 months. Surprisingly, nominal dystrophin was restored in skeletal muscle. Cardiac dystrophin was restored, but histology and hemodynamics were not improved. To determine the underlying mechanism, we evaluated components of the CRISPR-editing machinery. Intriguingly, we found disproportional guide RNA (gRNA) vector depletion. To test whether this is responsible for the poor outcome, we increased the gRNA vector dose and repeated the study. This strategy significantly increased dystrophin restoration and reduced fibrosis in all striated muscles at 18 months. Importantly, skeletal muscle function and cardiac hemodynamics were significantly enhanced. Interestingly, we did not see selective depletion of the gRNA vector after intramuscular injection. Our results suggest that gRNA vector loss is a unique barrier for systemic AAV CRISPR therapy. This can be circumvented by vector dose optimization.


Gene therapy; Neuromuscular disease; Therapeutics

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