Gene therapy rescues cone function and viability in Rp2 knockout mice over a broad vector dose range and a wide therapeutic time window

Abstract

Purpose: Mutations in the RP2 gene account for ~10% of X-linked retinitis pigmentosa (XLRP). We recently generated an RP2 AAV vector (AAV8-scRK-RP2) and confirmed its ability to mediate long-term RP2 expression in Rp2 knockout (Rp2-KO) mouse model. Our initial results revealed cone function rescue in the mice receiving vector treatment at 4-6 weeks of age (ARVO 2014, poster 3310). The goal of this study was to conduct a long-term safety and efficacy investigation using a broad vector dose range. We also assessed whether older mice could benefit from the treatment.

Methods: Four to six-week old Rp2-KO mice received unilateral subretinal injections of the vector with doses of 5×10⁷, 1×10⁸, 3×10⁸ or 1×10⁹ vector genomes (vg) per eye. Fellow eyes were injected with vehicle. Another set of mice received 3×10⁸ vg/eye injection at 10 months of age. Retinal function of the treated mice was assessed by electroretinography (ERG) and optomotor test. These mice were sacrificed at 6.5 or 18 months of age for immunofluorescence analyses.

Results: A consistently higher photopic ERG response was observed in the vector-treated eyes with a wide dose range (5×10⁷ to 3×10⁸ vg/eye) during the 18-month observation. These eyes also exhibited significantly higher visual acuity by optomotor test. M-opsin was mislocalized to inner segments, cell body and synaptic regions in vehicle injected retina but was primarily localized to outer segments in the vector-treated retina. In addition, higher expressions of Grk1 and cone-Pde were observed in the photoreceptor layer of vector treated eyes. The vector-treated eyes also maintained higher number of cone cells at 18 months of age. Interestingly, prevention of loss of cone function and viability was also achieved in Rp2-KO mice treated at 10 months of age. However, reduced rod ERG amplitude and thinner photoreceptor layer were observed in mice receiving the highest vector dose (1×10⁹ vg/eye), indicating retinal toxicity caused by the over-expressed RP2 protein.

Conclusions: Administration of the AAV8-scRK-RP2 vector is able to prevent the loss of cone function and viability in Rp2-KO mice with a broad vector dose range. The wide therapeutic time window in mice suggests that patients with RP2 mutations could benefit even when treated at an advanced age. The AAV8-scRK-RP2 vector fits our criteria for further clinical development.