A dose escalation and safety study of AAVrh10-mediated Schwann cell-targeted gene therapy for CMT1X.

X-linked Charcot-Marie-Tooth disease (CMT1X) is an inherited demyelinating neuropathy caused by loss-of-function mutations in the GJB1 gene, encoding the gap junction protein connexin32 (Cx32). Cx32 plays a critical role in Schwann cell function and myelin formation in the peripheral nervous system. We have developed a gene replacement therapeutic approach using a humanized AAVrh10 vector construct expressing GJB1 under the control of the Schwann cell-specific human myelin protein zero (MPZ) promoter. Lumbar intrathecal injection of increasing AAVrh10-hMPZ.GJB1 doses (low: 1 ​× ​10 vg, standard: 2 ​× ​10 vg and high: 1 ​× ​10 vg) into Gjb1-null mice resulted in adequate, dose-dependent biodistribution of the vector in anterior lumbar roots and peripheral nerves, as well as high rates of Schwann cell-specific Cx32 expression in the standard- and high-dose groups. Both standard and high vector doses provided significant therapeutic benefit evaluated by behavioural, electrophysiological and morphological outcomes. Intrathecal delivery of AAVrh10-hMPZ.GJB1 induced the production of anti-AAVrh10 antibodies at 6 weeks post-injection. However, no histopathological or inflammatory changes were observed in neural or peripheral tissues, besides a mild increase in inflammatory cell numbers in sciatic nerves of mice treated with the high dose only. This study provides proof of concept for a clinically translatable AAVrh10-mediated gene therapy approach for CMT1X.