PROJECT SUMMARY Retinitis pigmentosa (RP) is the most common inherited retinal dystrophy (IRD), caused by more than 3,100 mutations in 80 genes that are primarily specific to rod photoreceptors. Following major rod death phase, cone death occurs regardless of the underlying gene mutations. Currently, there exists a knowledge gap in understanding how aerobic glycolysis in photoreceptors impact the delicate “metabolic coupling” between rods and the retinal pigment epithelium (RPE) in RP. The long-term goal of this project is to develop a therapy that will preserve cone function in patients with RP. The objectives of this proposal are to investigate how metabolic dysregulation due to lactate deficiency contributes to photoreceptor death in RP, test a novel metabolome reprogramming strategy, and fulfill important safety requirements for filing a Pre-Investigational New Drug application. The hypothesis is that reprogramming rod and cone aerobic glycolysis will promote cone survival in RP independent of the underlying rod-specific gene mutations. This hypothesis has been formulated based on the applicant’s strong preliminary data. The rationale for the proposed research is that by targeting a metabolic pathway common to many of the genetically heterogeneous forms of RP, cone function may be preserved for the equivalent of 10 or more human years, which would have a tremendously positive impact on the lives of patients with RP. This hypothesis will be tested by pursuing three specific aims: 1) Investigate whether photoreceptor-specific ablation of prolyl hydroxylase domain-containing protein (Phd), a metabolic enzyme that inhibits aerobic glycolysis under normoxia, preserves cones by enhancing aerobic glycolysis in an autosomal recessive RP mouse model; 2) Assess the efficacy and feasibility of enhancing cone survival and function by ablating PHD2 in photoreceptors of a dominant RP mouse model ; 3) Establish the safety of therapeutic Phd2 editing in a WT and RP mouse model as well as human cells. Specifically, Aim 1 will determine whether enhanced aerobic glycolysis in cone photoreceptors can promote their survival in a novel genetic mouse model. Aim 2 will test the potential of gene therapy to slow photoreceptor degeneration by enhancing aerobic glycolysis in a different mouse model of RP. Lastly, Aim 3 will define the pharmacokinetics and safety of the aerobic glycolysis reprogramming vector. The approach is innovative because this will be the first example of cell-specific CRISPR-mediated precision metabolic reprogramming and because the novel therapeutic-editing vectors can be redeployed in future Phase I-IIA trials without modification. The proposed research is significant as it has the potential to dramatically lower the cost of treatment, be applicable to dividing and nondividing cells, shape ongoing CRISPR research, and ultimately define aerobic glycolysis as a safe and effective therapeutic target.