LIMB GIRDLE MUSCULAR DYSTROPHIES: P.137 Treatment of aged mice and long-term durability of AAV-mediated gene therapy in two mouse models of Limb girdle muscular dystrophy

Abstract

Two questions about efficacy of adeno-associated virus (AAV)-mediated gene therapy is ability to treat older more severely affected muscle and long-term durability. We used a mouse model of LGMD2D (α-sarcoglycan) treated at older age with an AAV.hSGCA vector and a model of LGMD2E (β-sarcoglycan) treated with an AAV.hSGCB vector following a long-term endpoint to investigate these questions. The goal was to demonstrate long-term durability of AAV gene therapy and provide evidence for therapeutic efficacy for aged diseased muscle. To address aged diseased muscle, we treated 12-month-old sgca−/− mice with severe muscle histopathology with an rAAVrh74.tMCK.hSGCA vector. At 6 months post-treatment, we assessed muscle for protein expression, histological rescue, and functional improvement. To investigate durability, we treated sgcb-/- mice at age 4 weeks with an rAAVrh74.MHCK7.hSGCB vector and evaluated these mice >24 months post-treatment for vector durability and biomarker protein expression. IV rAAVrh74.tMCK.hSGCA in 12-month-old sgca−/− mice resulted in widespread high-level protein expression in lower limb, upper limb, and proximal torso muscles, including diaphragm and heart. We noted reduction in fibrosis levels versus untreated controls, and functional improvement evidenced by improved force output in the tibialis anterior (TA) and diaphragm muscle and increased resistance to contraction-induced injury in TA muscle. Durability was established in sgcb−/− mice treated systemically with rAAVrh74.MHCK7.hSGCB. qPCR detected high-level vector genome copy numbers across all transduced muscles >24 months post-treatment. Immunofluorescence staining of treated muscle showed no decrease of protein expression levels in all muscles (>95%) versus earlier timepoints, with hSGCB protein remaining correctly localized at the membrane. These data show ability for AAV gene therapy to be efficacious when delivered at an older age to more severely diseased muscle, and its durability.