Optimized AAV CRISPR gene therapy could provide lifelong physiological benefit

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A research study investigated the long-term therapeutic benefits of AAV CRISPR therapy in a mouse model of Duchenne muscular dystrophy (DMD). The study reveals gRNA vector depletion as a major bottleneck for systemic AAV CRISPR therapy.

DMD is a progressive, X-linked degenerative disorder caused by the absence of dystrophin and is the most common inherited neuromuscular disease. Dystrophin protein levels are affected due to out-of-frame mutations in the dystrophin gene. There is no curative treatment for DMD and there has been significant progress in the development of gene replacement therapy over the last decade.

CRISPR/Cas9-mediated genome editing has emerged lately as a promising approach to treat human diseases. In this approach, Cas9 endonuclease is directed to the intended location by guide RNA (gRNA) to create a double-strand break followed by DNA repair at target site.

Delivery of CRISPR genome-editing tools by AAV has been previously tested and has provided encouraging results in restoring dystrophin expression in DMD patient cells in vitro and in mouse models in vivo. However, long-term persistence of gene expression is a mandate for diseases such as DMD.

In the present study published in JCI Insight, Dr Dongsheng Duan and team at University of Missouri investigated the long-term therapeutic benefits of AAV CRISPR therapy in a mouse DMD model. To address this, the team injected AAV-9 CRISPR vectors to the tail vein of 6-week-old mdx mice, a commonly used DMD model. Dystrophin expression and disease rescue were analysed at 18 months.

Results showed that AAV mediated CRISPR delivery yielded low CRISPR editing in skeletal muscle, limited dystrophin restoration in the heart, and no improvement in cardiac histology and hemodynamic function. Subsequent analysis showed similar levels of Cas9 expression in the heart and skeletal muscle.

Surprisingly, vector genome quantification showed a significant reduction in the gRNA vector. Increasing gRNA vector dose rescued the dystrophin expression in the heart and skeletal muscle. Importantly, it reduced fibrosis in the heart and skeletal muscle, significantly increased skeletal muscle force, and enhanced cardiac hemodynamic function.

These findings suggest that optimizing gRNA vector dose is necessary to achieve sustained skeletal muscle and heart rescue in DMD. Results from the study highlight the potential for using AAV CRISPR therapy to treat any disease that requires systemic life-long mutation correction.

Source: AAV CRISPR editing rescues cardiac and muscle function for 18 months in dystrophic mice. Hakim CH et al., JCI Insight, December 2018. DOI