Gene therapy vectors based on adeno-associated viruses (AAVs) show promise for the treatment of retinal degenerative diseases. In prior work, subretinal injections of AAV2, AAV5, and AAV2 pseudotyped with AAV5 capsids (AAV2/5) showed variable retinal pigmented epithelium (RPE) and photoreceptor cell transduction, while AAV2/1 predominantly transduced the RPE. To more thoroughly compare the efficiencies of gene transfer of AAV2, AAV3, AAV5, and AAV6, we quantified, using stereological methods, the kinetics and efficiency of AAV transduction to mouse photoreceptor cells. We observed persistent photoreceptor and RPE transduction by AAV5 and AAV2 up to 31 weeks and found that AAV5 transduced a greater volume than AAV2. AAV5 containing full-length or half-length genomes and AAV2/5 transduced comparable numbers of photoreceptor cells with similar rates of onset of expression. Compared to AAV2, AAV5 transduced significantly greater numbers of photoreceptor cells at 5 and 15 weeks after surgery (greater than 1,000 times and up to 400 times more, respectively). Also, there were 30 times more genome copies in eyes injected with AAV2/5 than in eyes injected with AAV2. Comparing AAVs with half-length genomes, AAV5 transduced only four times more photoreceptor cells than AAV2 at 5 weeks and nearly equivalent numbers at 15 weeks. The enhancement of transduction was seen at the DNA level, with 50 times more viral genome copies in retinas injected with AAV having short genomes than in retinas injected with AAV containing full-length ones. Subretinal injection of AAV2/6 showed only RPE transduction at 5 and 15 weeks, while AAV2/3 did not transduce retinal cells. We conclude that varying genome length and AAV capsids may allow for improved expression and/or gene transfer to specific cell types in the retina.

Gene transfer using adeno-associated viruses (AAVs) has been effective for treating inherited retinal diseases in animal models. Further evaluation in primates must be performed prior to clinical application, however, because of the difference between the retina of the primate and those of other animals. Prior work has shown that AAV2 can transduce rod-photoreceptor and RPE cells in the non-human primate retina and that AAV5 is more efficient at transducing photoreceptor cells than AAV2 in the rodent retina. In this study, we evaluated the efficiency of AAV5 in the non-human primate retina after subretinal injections of the vector to distinct anatomic retinal regions (superior, inferior, nasal, macula, temporal). rAAV5 led to a rapid onset of transgene expression (within 2 weeks), with expression persisting up to 10 months. Postoperative electrophysiology studies showed that global retinal function was preserved following gene transfer. Quantitative analysis of gene transfer demonstrated a maximum transduction efficiency of 22% in the injected areas. Evaluation of cell types using confocal microscopy and cone-specific antibodies revealed that AAV5, expressing reporter genes from the cytomegalovirus (CMV) promoter/enhancer, preferentially transduced rods. No significant differences were found in the regional tropism of AAV5 among the five areas injected despite variation in retinal topography. Immunohistochemical studies revealed that the AAV5 receptor, PDGFR-A, is localized to the outer segments of rods but not cones providing a basis for the observed tropism. Our results support the utility of AAV5 for rod photoreceptor degeneration therapies.