Extracellular vesicles-associated AAVs for the treatment of Machado-Joseph disease.

Abstract

Machado-Joseph disease (MJD) is the most common dominant autosomal inherited ataxia worldwide, caused by the overrepetition of the trinucleotide CAG in the ATXN3 gene. This leads to the accumulation of mutant ataxin-3 protein and neurodegeneration. Currently, treatment remains symptomatic, although gene therapy has emerged as a promising approach. However, efficient and minimally invasive gene delivery to the brain remains a challenge. Extracellular vesicle-associated adeno-associated virus (EV-AAV) vectors are a novel delivery system, combining the ability of AAV vectors to deliver genes with the capacity of extracellular vesicles to bypass the immune system and cross the blood-brain barrier (BBB). Previous studies, however, have only combined AAV serotypes known to efficiently cross the BBB with EVs as a non-invasive delivery system to the brain. Thus, the ability of EV-AAVs to cross the BBB remained inconclusive. In this study, we evaluated whether AAV1/2 serotype, combined with rabies virus glycoprotein (RVg)-coated EVs, could effectively target the brain. Two isolation methods, differential ultracentrifugation and size-exclusion chromatography (SEC) were compared, with SEC yielding higher EV recovery. Moreover, RVg-EV-AAV1/2 successfully crossed the BBB and transduced mouse brains, leading to motor and neuropathologic improvements in an MJD mouse model. This study demonstrates that RVg-EV-AAVs are promising non-invasive delivery systems for MJD gene therapy.