Functional, sustained recovery of hearing in Otoferlin-deficient mice using DB-OTO, a hair-cell-specific AAV-based gene therapy

Biallelic loss-of-function variants in the Otoferlin gene (OTOF) lead to congenital hearing loss in both humans and mice. We developed DB-OTO, a hair-cell-specific adeno-associated virus (AAV)-based dual-vector gene therapy designed to restore hearing in individuals with OTOF-related hearing loss. DB-OTO is composed of two AAV1 vectors that reconstitute a functional OTOF gene expression cassette to form the full-length human OTOF variant 5 (hOTOFv5). A synthetic promoter (mMyo15) based on the murine Myosin 15a gene was engineered to restrict the transgene expression to hair cells. The promoter construct was chosen after comparison of multiple variants using a GFP reporter in ex vivo murine explant models, followed by in vivo studies in wild-type mice and cynomolgus macaques. Comparison between the 1.0 kb mMyo15 and a ubiquitous promoter driving expression of hOTOFv5 transgene in mice showed that hair-cell-specific expression is critical for safe expression of otoferlin. DB-OTO induced dose-dependent establishment of auditory brainstem response, and OTOF expression in inner hair cells over a 10-fold dose range sustained for at least 3 months in a mouse model of OTOF deficiency. These data supported the initiation of an ongoing phase I/II clinical trial of DB-OTO in pediatric patients with OTOF-related hearing loss.