AAV2/4-RS1 gene therapy in the retinoschisin knockout mouse model of X-linked retinoschisis

Objective To evaluate efficacy of a novel adeno-associated virus (AAV) vector, AAV2/4-RS1, for retinal rescue in the retinoschisin knockout (Rs1-KO) mouse model of X-linked retinoschisis (XLRS). Brinzolamide (Azopt((R))), a carbonic anhydrase inhibitor, was tested for its ability to potentiate the effects of AAV2/4-RS1. Methods AAV2/4-RS1 with a cytomegalovirus (CMV) promoter (2x10(12) viral genomes/mL) was delivered to Rs1-KO mice via intravitreal (N = 5; 1 mu L) or subretinal (N = 21; 2 mu L) injections at postnatal day 60-90. Eleven mice treated with subretinal therapy also received topical Azopt((R)) twice a day. Serial full field electroretinography (ERG) was performed starting at day 50-60 post-injection. Mice were evaluated using a visually guided swim assay (VGSA) in light and dark conditions. The experimental groups were compared to untreated Rs1-KO (N = 11), wild-type (N = 12), and Rs1-KO mice receiving only Azopt((R)) (N = 5). Immunofluorescence staining was performed to assess RS1 protein expression following treatment. Results The ERG b/a ratio was significantly higher in the subretinal plus Azopt((R)) (p<0.0001), subretinal without Azopt((R)) (p = 0.0002), and intravitreal (p = 0.01) treated eyes compared to untreated eyes. There was a highly significant subretinal treatment effect on ERG amplitudes collectively at 7-9 months post-injection (p = 0.0003). Cones showed more effect than rods. The subretinal group showed improved time to platform in the dark VGSA compared to untreated mice (p<0.0001). RS1 protein expression was detected in the outer retina in subretinal treated mice and in the inner retina in intravitreal treated mice. Conclusions AAV2/4-RS1 shows promise for improving retinal phenotype in the Rs1-KO mouse model. Subretinal delivery was superior to intravitreal. Topical brinzolamide did not improve efficacy. AAV2/4-RS1 may be considered as a potential treatment for XLRS patients.