Preclinical evaluation of rAAV8.PDE6A in the Pde6a^D670G mutant mouse model of RP43

Abstract

Purpose : Mutations in the phosphodiesterase 6 alpha (PDE6A) gene are known to cause retinitis pigmentosa type 43 (RP43) – A currently cureless blinding disease. Here, a novel recombinant adeno-associated virus (AAV) vector for gene supplementation therapy of PDE6A-linked RP was developed and tested for efficacy in the preclinical Pde6a^D670G mouse model of RP43.

Methods : An AAV-based gene supplementation vector coding for full length human PDE6A under control of the human, rod-specific rhodopsin promoter was generated and used for packaging of AAV8-pseudotyped vectors. The resulting vector (rAAV8.PDE6A) was tested for efficacy by subretinal injections in two-week old Pde6a^D670G mutant mice with short-term and long-term analysis at 4 weeks and 6 months post-injection, respectively. AAV vector-mediated expression of human PDE6A protein was assessed by immunohistochemistry using a PDE6A-specific antibody combined with confocal microscopy. Efficacy was assessed by measuring morphological preservation in vivo using optical coherence tomography (OCT) and ex vivo by analyzing histological cross-section of treated and untreated retinas.

Results : We designed and generated rAAV8.PDE6A, a novel AAV vector optimized for efficient PDE6A gene expression in human rod photoreceptors. Transgene expression assay of rAAV8.PDE6A confirmed efficient and specific human PDE6A protein expression that localized to rod photoreceptor outer segments. A biological activity assay for rAAV8.PDE6A based on analysis of morphological preservation confirmed a beneficial effect of the treatment at both observation time points. Additionally, we found a preservation of rod outer segment structure and prolonged survival of cone photoreceptors up to 6 months after treatment.

Conclusions : The novel rAAV8.PDE6A vector supports efficient and specific transgene expression and biological activity in the preclinical Pde6a^D670G mouse model of RP43.

This is an abstract that was submitted for the 2017 ARVO Annual Meeting, held in Baltimore, MD, May 7-11, 2017.