Abstract
Limb girdle muscular dystrophy (LGMD) 2A/R1, caused by mutations in the CAPN3 gene and CAPN3 loss of function, is known to play a role in disease pathogenicity. In this study, AAVrh74.tMCK.CAPN3 was delivered systemically to two different age groups of CAPN3 knockout (KO) mice; each group included two treatment cohorts receiving low (1.17 × 10(14) vg/kg) and high (2.35 × 10(14) vg/kg) doses of the vector and untreated controls. Treatment efficacy was tested 20 weeks after gene delivery using functional (treadmill), physiological (in vivo muscle contractility assay), and histopathological outcomes. AAV.CAPN3 gene therapy resulted in significant, robust improvements in functional outcomes and muscle physiology at low and high doses in both age groups. Histological analyses of skeletal muscle showed remodeling of muscle, a switch to fatigue-resistant oxidative fibers in females, and fiber size increases in both sexes. Safety studies revealed no organ tissue abnormalities; specifically, there was no histopathological evidence of cardiotoxicity. These results show that CAPN3 gene replacement therapy improved the phenotype in the CAPN3 KO mouse model at both doses independent of age at the time of vector administration. The improvements were supported by an absence of cardiotoxicity, showing the efficacy and safety of the AAV.CAPN3 vector as a potential gene therapy for LGMDR1.