Long-term Visual Behavior Correlates with Functional Electroretinographic Assessment after Gene Transfer in RPE65-/- Dogs

Abstract

Abstract: : Purpose: To assess the long-term effects of subretinal rAAV.RPE65 gene transfer on functional vision in RPE65-/- dogs. Methods: Five dogs homozygous for the RPE65 null mutation and 2 unaffected, visually normal control dogs were included. Affected animals had been treated subretinally with 70 - 100 µl of rAAV.RPE65 gene construct (2 X 10¹² particles/ml) in the right eye (OD), and 0 - 100 µl of rAAV.GFP (2 X 10¹⁰ transforming units/ml) in the left eye (OS). Treatment for control dogs was 75 - 100 µl of the latter OD and 100 µl of BSS OS. Evaluations from pre-surgical and follow-up bilateral full-field electroretinograms (ERGs) 6 mo post-surgery were obtained. Objective assessment of functional vision was performed 3 and 6 mo post-operatively by counting the number of collisions made by the dogs as they negotiated a maze of obstacles in dim and day light conditions. Vision in individual eyes was also assessed 6 mo post-surgery. Results: Three mo after gene therapy there was a higher number of collisions in dim light than in day light (p<0.008) among affected dogs; a difference not evident 6 mo post-operatively. In day light conditions, no difference was noted between control and affected animals at 3 or 6 mo. In dim light, affected dogs had a higher number of collisions than control dogs at 3 (p<0.004) but not 6 mo post-operatively. Fewer collisions (p<0.1) were noted in day and dim light when only OD was exposed compared with exposure of only OS in the affected dogs. Post-surgery, high intensity scotopic stimuli and 30 Hz flicker ERG amplitudes were elevated (p<0.04) over pre-operative recordings in OD, and elevated over post-operative recordings in OS as well (p<0.032). Further, post-operative single flash photopic b-wave amplitudes for OS were increased over pre-operative values (p<0.05). Conclusions: Progressive improvement in functional binocular and monocular (OD) vision up to 6 mo after gene transfer was found by objective behavioral testing. The long-term visual improvement correlated with a significant increase in post-operative ERG amplitudes, mainly in OD but also with measurable effects in OS.

Keywords: animal model • gene transfer/gene therapy • vision and action