SCGE Disease Models Studies Supplement: Correction of RHO P23H adRP

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Project Summary The broad, long-term objective of this work is to establish a rubric to evaluate safety and efficacy of somatic cell genome editing technologies. These goals will be addressed in the context of an experimental therapy for retinitis pigmentosa caused by the RHO P23H variant. The specific aims of this proposal address refinement of the therapeutic window for this therapy, the minimum number of cells that must be edited to save vision, and the general safety of the therapeutic strategy. These aims will be met through the use of AAV5 to deliver an engineered meganuclease that specifically targets the dominant P23H allele while saving the wildtype allele that differs by only one base. This reagent will be evaluated in a validated, rapidly progressing swine model that harbors a human RHO P23H transgene. A previously optimized dose that produces a durable response will be delivered via subretinal injection and the animals will be followed for 32 weeks to repeatedly measure effectiveness over time via clinical eye exam, Full-Field Electroretinogram (ffERG), and Optical Coherence Tomography (OCT). At the end of the experiment, eyes will be examined via immunohistochemistry (IHC) to determine the regional, minimum number of rods that must be edited to save cone function and therefore to save vision. These experiments will be performed at two different stages in disease progression that are hypothesized to represent the last treatable state and the first non-treatable state so that guidance can be provided for criteria of enrollment of patients in a future clinical trial. In parallel, animals will be similarly treated and then sacrificed at two weeks post injection to determine the range of adjacent tissues to which the AAV5 vector will migrate. These animals will also be used to determine the level of editing that occurs in non-intended tissues, including the germline. All animals will be used to obtain data on general physiological response to the therapy.