403. Engineering Micro-Dystrophin AAV Vector for Clinical Translation

Abstract

Duchenne muscular dystrophy (DMD) is the most severe muscular dystrophy affecting boys. It is caused by the loss of a cellular structural protein called dystrophin. Dystrophin preserves the integrity of the muscle cell membrane by providing a molecular link between the cytoskeleton and the extracellular matrix. The absence of dystrophin destabilizes the muscle membrane and promotes muscle death. AAV micro-dystrophin gene therapy has transformative potentials to significantly ameliorate DMD. Unfortunately, AAV therapy is challenged by destructive cellular immunity to newly expressed dystrophin. To overcome this obstacle, we engineered a double restrictive AAV microgene vector. Specifically, we limited off-target expression in non-muscle tissue with a muscle specific promoter and removed untoward expression in antigen-presenting cells using miR142-3p, a hematopoietic specific microRNA target sequence. Compared with the original AAV microgene vector, the newly engineered vector significantly reduced infiltration of CD4+ and CD8+ T cells in the canine DMD model. Our results suggest that the dual restriction strategy may greatly enhance clinical translation of AAV micro-dystrophin gene therapy.