882. Non-Viral Ocular Gene Transfer for Hereditary Retinal Degeneration

Abstract

Top of pageAbstract The present study was undertaken to determine the efficacy of DNA nanotechnology as an effective therapeutic strategy for eye diseases; particularly those associated with loss-of-function mutations in the retina and retinal pigment epithelium (RPE). Since many hereditary blinding disorders have an onset from birth, we first evaluated a method of delivery to the newborn mouse eye. Subretinal injections of saline were delivered into one eye at post-natal day 2 (P2) and the contra-lateral eye was left uninjected to serve as a control. These mice were then left in normal cyclic light to develop into adults. At P21, retinal function was assessed with electroretinography (ERG) and ocular structure was examined with histology. Eyes injected at P2 demonstrated normal scotopic and photopic ERG waveforms and displayed a normal cellular organization and morphology, as compared to contra-lateral controls. As this route of delivery was safe, we then sought to determine whether this method could be used to deliver compacted DNA nanoparticles for gene transfer. The Rds+/|[minus]| mouse has only one functional copy of the photoreceptor specific gene Rds, and presents with a progressive retinal degeneration which causes perturbation of retinal function and structure, eventually leading to death of photoreceptors. Rds+/|[minus]| mice at P2 were injected with compacted nanoparticles of plasmid DNA carrying the wild-type Rds cDNA directed by the chicken b-actin promoter (Rds-particles), or the non-compacted naked plasmid (Rds-plasmid). Similar to experiments described above, one eye was injected, and the contra-lateral eye served as a non-injected control. At P30 and P45 these animals were subjected to ERG to assess photoreceptor viability. Eyes receiving Rds-nanoparticles showed an increased scotopic a- and b-wave, in addition to an increased photopic b-wave. However, eyes receiving naked Rds-plasmid failed to demonstrate an increase in ERG function at the timepoints examined. In contrast, Rds+/|[minus]| mice that received a subretinal delivery of Rds-nanoparticles at P21 failed to show any rescue of photoreceptor function when assessed by ERG at 7, 14, and 21 days post-injection. These studies demonstrate the feasibility of newborn treatment strategies for retinal degenerative disorders and provide a rationale for future therapeutic interventions. Further studies are underway to determine the longevity of rescue in animals treated at P2 and whether nanoparticle treatment resulted in amelioration of the structural abnormalities associated with the Rds+/|[minus]| retina.