Targeting Soluble Epoxide Hydrolase to Treat Choroidal Neovascularization

Abstract

Neovascular or “wet” age-related macular degeneration (nvAMD) is a leading cause of blindness among older adults, affecting millions of people worldwide. Choroidal neovascularization (CNV) is a major pathological feature of nvAMD, in which abnormal new blood vessel growth from the choroid leads to irreversible loss of vision. Currently, the effort to treat nvAMD is hampered by resistance and refractory responses to the current standard of anti-angiogenic care, anti-vascular endothelial growth factor biologics. Thus, there is a critical need to develop novel therapeutic strategies. Previously, we discovered an anti-angiogenic small molecule SH-11037, and identified soluble epoxide hydrolase (sEH) as a target of SH-11037 through a forward chemical genetics approach. sEH, encoded by the EPHX2 gene, is a lipid-metabolizing enzyme that hydrolyzes epoxy fatty acids into corresponding diols. I hypothesized that sEH is a key mediator of CNV. Given that the kinetic mechanism of sEH inhibition by SH-11037 and the cellular role of sEH in CNV are poorly understood, the objectives of my thesis project were to elucidate drug-target interactions through enzyme kinetics, investigate sEH mediated mechanisms that regulate CNV, and preclinically validate sEH as a therapeutic target. I discovered that SH-11037 is a mixed inhibitor of sEH with a binding affinity for both the enzyme and enzyme-substrate complex. I examined retinal spatial expression of sEH at both the protein and mRNA levels through immunohistochemistry and RNAscope in situ hybridization and investigated the efficacy of adeno-associated virus (AAV) serotype 8 vector expressing shRNA against Ephx2, in the mouse laser-induced (L-) CNV model with features of nvAMD. My study revealed sEH protein and mRNA overexpression in the retinal pigment epithelium (RPE), vasculature and photoreceptors under the disease state. The delivery of AAV8-Ephx2 shRNA, which has tropism towards RPE and photoreceptor cells, significantly reduced CNV. In addition, gene expression analysis showed normalized Vegfc and CNV-related inflammatory markers upon sEH knockdown. Thus, my study demonstrated sEH overexpression in disease-relevant cell types, highlighted a functional role of sEH in AMD pathophysiology, and provided a novel context to target these cell types for developing pharmacotherapies.