Dose-Escalation Study of Systemically Delivered rAAVrh74.MHCK7.micro-dystrophin in the mdx Mouse Model of Duchenne Muscular Dystrophy

Overview

Duchenne muscular dystrophy (DMD) is a rare neuromuscular disease caused by mutations in the DMD gene. These mutations disrupt the expression of dystrophin, a protein important for muscle fiber stability. DMD is characterized by a progressive loss of skeletal and cardiac muscle strength. This study focuses on characterizing the effects of rAAVrh74.MHCK7.micro-dystrophin, an adeno-associated virus vector containing a codon-optimized human micro-dystrophin transgene, on DMD mice. The authors administered systemic injections of the dystrophin-restoring vector to mdx mice at low, intermediate, and high doses. Three months post-treatment, they analyzed micro-dystrophin positive fibers via immunofluorescent staining. They observed mean expression percentages of 46.7%, 66.8%, and 78.3% for low, intermediate, and high doses, respectively, across all muscles assessed (TA, GAS, QD, PSO, TRI, GLUT and DIA). Fiber diameter and fibrosis were then assessed, where low and intermediate doses reduced fibrosis and normalized fiber size in the diaphragm, similar to that of wild type. Force production was analyzed using Aurora’s 1200A isolated muscle system. In vitro analysis of the dystrophic diaphragm revealed improvement in specific force output following high dose treatment similar to that of wild type. To determine if restoring dystrophin would have a functional effect on hindlimb muscle, the authors further assessed the tibialis anterior (TA) in situ using the 1300A Whole Animal System. rAAVrh74.MHCK7.micro-dystrophin not only improved force output in the TA but rescued the muscle from eccentric contraction-induced damage. Lastly, western blot analysis of tissues from injected mice showed micro-dystrophin protein expression across all skeletal muscles. No micro-dystrophin was detected in off-target organs at low and intermediate doses, with high dose exhibiting faint expression in only the liver. These findings demonstrate safety and efficacy of systemic delivery of rAAVrh74.MHCK7.micro-dystrophin, supporting the use of this vector in Phase I/II safety study in boys with DMD.