Adeno_Associated Virus (AAV)-Mediated Transduction of the Macula in Non-Human Primates (NHPs): Cell-Specificity, Stability and Safety of Transduction After Subretinal Delivery of AAV2 and AAV2/8 Vectors

Abstract

Purpose:: This study aimed to compare transduction patterns, dose effects, onset and stability of transgene expression and safety of subretinal delivery of transgenes through AAV2 and 2/8 to the NHP retina.

Methods:: Subretinal injections of 150 µl of AAV2 or 2/8 carrying an eGFP transgene were performed at doses ranging from 1E8 to 1E11 genome copies in a total of 28 eyes of 14 cynomolgus monkeys. These animals were seronegative for AAV2 and AAV8 capsids. Animal’s behavior, vital signs, blood counts and chemistries, and immune responses to capsid and transgene were evaluated. EGFP expression was evaluated non-invasively and histologically both quantitatively and qualitatively.

Results:: Delivery of the vector was well tolerated locally and systemically and the animals showed no alteration in behavior or general well-being. The time of onset of ophthalmoscopically detectable eGFP differed between the two AAV vectors, with the onset of transgene expression being 21 and 14 days for AAV2 and AAV2/8 respectively. AAV2/8 was 10-fold more efficient than AAV2 in retinal transduction. Both AAVs efficiently transduced retinal pigment epithelium (RPE) at all doses. AAV2/8 displayed a high tropism for retinal photoreceptors and occasional Muller and ganglion cell transduction. Expression was localized to the region exposed to vector but was not always uniform. Some of the both AAV2 or AAV2/8-injected retinas, dropout of eGFP signal correlated with localized retinal histopathology. ELISPOT analyses of one of those animals, revealed T-cell populations positive for AAV caspsid as well as the EGFP.

Conclusions:: AAV based-vectors efficiently transduce neuronal and pigment epithelial cells of the macular retina, thus providing delivery vehicles relevant to the treatment of retinal diseases. Dose for dose, AAV2/8 delivers transgene more efficiently and more quickly than AAV2. While further studies are warranted to identify and avoid cellular and immunological factors that could lead to localized retinal toxicity, both AAV2 and 2/8 vectors hold great promise for treatment of retinal disease.

Keywords: gene transfer/gene therapy • retina • degenerations/dystrophies