Dose-response effect of RPE65 gene therapy on retinoid levels and correlation with clinical rescue in Rpe65-deficient dogs.

Abstract

 

Purpose

 

Although the mouse and dog models of Rpe65 deficiency have been stably rescued and the retinas preserved following AAV vector-mediated gene therapy delivered before degeneration ensues, young human patients treated with identical vectors and doses have incomplete vision recovery, and suffer continued retinal degeneration despite treatment. The purpose of this study was to examine the physiological and clinical effects of different doses of gene therapy in the Rpe65-deficient dog to investigate the effect on visual cycle activity.

 

Methods

 

We treated 12 eyes with 4 different doses of gene therapy, the lowest dose was 8x10^8 vg, the highest dose was 1x10^11 vg. We evaluated electroretinographic responses, vision outcomes, and following euthanasia, we removed retinas under dark-adapted conditions for histology and 11-cis retinal quantification. 11-cis retinal was used as a marker for enzyme activity, as it is a product of normalized RPE65 enzymatic activity.

 

Results

 

Rod and cone rescue was present in the highest two dose groups. Significant improvements in vision were noted in the top 3 dose groups. We detected RPE65 protein expression in the RPE using western blotting and IHC in the highest dose group, and in patchy regions using IHC in the next highest dose group. Significantly higher levels of 11-cis retinal were detected in the highest dose group compared to all other groups. Eyes of the highest dose group contained less than 25% of the 11-cis retinal present in untreated normal dogs' eyes collected in the same manner.

 

Conclusions

 

These results demonstrate that AAV-mediated expression of RPE65 protein can improve vision and electrophysiological responses despite only partial restoration of visual cycle activity.