A two-pronged approach to generating novel models of photoreceptor degeneration for regenerative cell therapy

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PROJECT SUMMARY Inherited retinal diseases (IRD), such as retinitis pigmentosa (RP), Leber congenital amaurosis (LCA), are a common cause of visual impairment worldwide. Recent studies show that cell replacement therapy is a promising therapeutic option for IRD patients with severe retinal degeneration that destroys photoreceptors. We and others recently reported long-term survival and integration of human embryonic stem cells (hESCs) or human induced pluripotent stem cells (hiPSCs) derived retinas in a novel laser-induced photoreceptor degeneration model. While the laser ablation model can be generated rapidly as an acute degeneration model, it does not have the progressive pathophysiological nature (e.g., disease time course, retinal remodeling, etc.) of IRD. To address this issue, we developed a highly efficient genome-editing technology, Platinum Talen, which leads to gene disruption in multiple organisms with high efficacy and specificity. By applying Platinum Talen to this novel model, we achieved one-step generation of biallelic gene modifications with high efficiency (40-50%) in preliminary studies. Since substantial information regarding the survival and integration of transplanted photoreceptor cells or retinal sheets have been obtained in laser injury models, we propose a two-pronged strategy to generating novel translation-enabling models to evaluate survival and integration of regenerated photoreceptors in the visual system: 1) to use laser-induced photoreceptor degeneration model; 2) one-step generation of biallelic mutants by Platinum Talen. We will then systematically compare the survival and integration of regenerated photoreceptors in the two model systems. Aim 1 will generate novel models of photoreceptor degeneration by laser ablation and Platinum Talen. Aim 2 will perform comprehensive ophthalmic phenotyping of laser-induced and Talen-edited models of photoreceptor degeneration. Aim 3 will evaluate survival and integration of human induced pluripotent stem cell (hiPSC)-derived retina sheets in the two models of photoreceptor degeneration.