Abstract
Purpose: :
Inherited RPE65 deficiency causes a severe reduction in rod and cone photoreceptor sensitivity and progressive photoreceptor degeneration in mice, dogs and humans. Gene therapy can improve retinal function when performed prior to photoreceptor loss. The RPE65-deficient dog is an important large animal model of human disease, and rescue of rod and cone photoreceptor function can be achieved in young RPE65-deficient dogs by gene replacement therapy. The aims of this study were to investigate whether gene replacement therapy promotes survival of cone photoreceptors in younger RPE65-deficient dogs and if it can rescue cone function in older dogs.
Methods: :
To investigate the effect of gene replacement therapy on cone survival, we injected AAV2/2.hRPE65p.hRPE65 subretinally in 8 young animals (6 months - 2.5 years; 15 eyes). We monitored ERGs at regular intervals for 1-3 years post-injection. Following sacrifice we evaluated cone survival in matched treated and untreated retinal regions using immunohistochemistry for human cone arrestin, short- and long/medium-wavelength cone opsins and by staining with peanut agglutinin. We also injected 5 eyes of older animals (3-5 years old), evaluated cone function at 4 months post-injection then investigated cone density following sacrifice. To investigate age-related changes in cone photoreceptors we examined cone ERGs and cone density of untreated young and middle-aged RPE65-deficient dogs.
Results: :
Studies of the untreated dogs showed a loss of cones with age. This was predominantly due to loss of short-wavelength opsin positive cones (S-cones). Gene therapy in the younger dogs improved cone function as assessed by ERG and also promoted S-cone survival only in the injected areas. The extent of rescue of cone function was more variable in older animals, and there was evidence of cone degeneration, particularly of S-cones, throughout the retina compared with both the young uninjected animals and those injected at an earlier age.
Conclusions: :
RPE65 gene replacement therapy promotes rescue of cone photoreceptor function and survival of S-cones in the RPE65-deficient dog when administered at a young age. Sufficient cones remain in middle-aged dogs to achieve functional rescue that is detectable by ERG, although the extent of rescue is less predictable. S-cones are particularly susceptible to loss in the RPE65-deficient dog.
Keywords: photoreceptors • gene transfer/gene therapy • retinal degenerations: hereditary