non-viral medicated gene targeting to the retina

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DESCRIPTION (provided by applicant): Inherited retinal diseases are a major cause of blindness. Mutations in over 140 different causative genes have been identified, of which over 60% have been cloned. Several animal models, both spontaneous mutants and genetically engineered animals have been identified for inherited retinal diseases. The vast majority of gene therapy protocols for ocular disease involve the use of viral mediated gene transfer. Although efficient in transduction and potent transgene expression, viral mediated gene therapy can cause potentially lethal random integration, immunological reaction, and has a high cost in both funding and time. Non-viral gene therapy approaches reduce many of these unwanted effects, but exhibits poor integration, limited transgene expression and decreased transduction efficiency. Recently, an irreversible recombinase, phage FC31 integrase has been shown to have stable, site-specific integration that can promote stable, long-term transgene expression, with the ability to carry large DMA sequences. It can be delivered through non-viral techniques, such as electroporation or lipofection, which have few side effects. This technology has not been applied to ocular disease, and in particular hereditary retinal degenerative disease. The research proposed here aims to apply this technology to a murine model for Leber's Congenital Amaurosis, caused by mutations in the RPE65 gene, the Rpe65-/- mouse. Viral mediated gene therapy has achieved function recovery in this model, subsequently the model has been well characterized, and functional testing parameters have been established. The overall aim of this proposal to utilize phage phiC31 integrase technology to demonstrate functional recovery in this animal model, and thereby offer an alternative or complementary therapy for this disease. Further to demonstrate possible application of this technology to other inherited ocular diseases that may benefit from gene therapy intervention.