499. Retinal Gene Therapy for Peroxisome Biogenesis Disorders

Abstract

Peroxisome biogenesis disorders (PBDs) are a group of autosomal recessive disorders most frequently caused by inherited defects in the PEX1 gene, which is required for normal peroxisome assembly and biochemical functions. Approximately 80% of patients fall within the category of Zellweger spectrum disorder (PBD-ZSD), which has an overall incidence of 1:50,000 births in the United States. The majority of patients have milder forms of disease compatible with survival through adulthood; however, they typically show mild to moderate developmental delays, and progressive vision and hearing loss. Thus, therapeutic interventions that prevent or slow down visual loss could have a profound impact on the lives of these individualsHere, we present a retinal gene therapy approach that addresses visual deterioration in patients with milder forms of disease. Optical coherence tomography (OCT) has demonstrated that the cone photoreceptor cells are most significantly affected by loss of peroxisome functions in such patients. These visual phenotypes are recapitulated in a new mouse model of disease that expresses the murine equivalent of most common PEX1 mutation found in patients (PEX1-p.G843D). Electroretinogram (ERG) analyses indicated severe impairment of the cone visual pathway in these homozygous Pex1-mutant mice by 4 months of age with the rod visual system being relatively preserved. Staining retinal sections with peanut agglutinin showed that some cone photoreceptors were retained in the homozygote murine Pex1 -mutant retina at 3 weeks, but were completely degenerated in the adult. Finally, genome-wide expression studies showed specific reduction of photoreceptor cone-specific genes with no differential expression of other cell lineage-specific genes or others that indicate cell death or stress responses.We have developed AAV vectors to deliver normal copies of the 3.85-kb human PEX1 gene to the mammalian retina. These vectors use the rhodopsin kinase 1 (RK1) or truncated cytomegalovirus (CMV) promoters, respectively. Subretinal injections of RK1.PEX1. rAAV9 in healthy mice resulted in robust expression of human PEX1 mRNA. Furthermore, transduction of cultured Pex1 -mutant murine skin fibroblasts with CMV.PEX1.rAAV9 results in the rescue of peroxisome assembly defects in these cells, as determine by the intracellular localization of a modified GFP reporter protein with a peroxisome targeting signal. This indicates that the human PEX1 transgene can complement the genetic defect in the murine Pex1 gene. We are currently testing the ability of the AAV9-PEX1 gene delivery system to complement the retinal gene defect in our Pex1 -mutant mouse model. If successful, this could provide the preliminary data required to begin initial planning for eventual clinical trials in patients.