Abstract
Age-related macular degeneration (AMD) is a multifactorial chronic disease that requires long term treatment. Gene therapy is being considered as a promising tool to treat AMD. We found that increased activation of Rap1a in the retinal pigment epithelium (RPE) reduces oxidative signaling to maintain barrier integrity of the RPE and resist neural sensory retinal angiogenesis from choroidal endothelial cell invasion. To optimally deliver constitutively active Rap1a (CARap1a) into the RPE of wild type mice, self-complementary AAV2 (scAAV2) vectors driven by two different promoters, RPE65 or VMD2, were generated and tested for optimal active Rap1a expression and inhibition of choroidal neovascularization (CNV) induced by laser injury. scAAV2-VMD2, but not scAAV2-RPE65, specifically and efficiently transduced the RPE to increase active Rap1a protein in the RPE. Mice with increased Rap1a from the scAAV2-VMD2-CARap1a had a significant reduction in CNV compared to controls. Increased active Rap1a in the RPE in vivo or in vitro inhibited inflammatory and angiogenic signaling determined by decreased activation of NF-κB and expression of VEGF without causing increased cell death or autophagy measured by increased LCA3/B. Our study provides a potential future strategy to deliver active Rap1a to the RPE in order to protect against both atrophic and neovascular AMD.
Highlights
Age-related macular degeneration (AMD) remains a leading cause of legal blindness in the elderly worldwide[1,2]
Treatments targeting vascular endothelial growth factor (VEGF) have greatly improved clinical outcomes in neovascular AMD; vision improvement only occurs in less than half of patients treated for neovascular AMD20,21, and treatments remain inadequate for atrophic AMD
Using a gene therapy approach, we previously reported that the expression of exogenous active Rap1a in the retinal pigment epithelium (RPE) by a self-complementary adeno-associated virus 2 (scAAV2)-RPE65 vector significantly reduced laser-induced choroidal neovascularization (CNV) in Rap1b deficient mice but not in wild type mice[7]
Summary
Age-related macular degeneration (AMD) remains a leading cause of legal blindness in the elderly worldwide[1,2]. The barrier integrity of the RPE is compromised and stressed RPE releases growth factors at a pathologic level that lead to advanced AMD6. The insufficient reduction of CNV in wild type mice by the scAAV2-RPE65 delivered CARap1a was predicted to be related to the weak transcriptional activity of the RPE65 promoter. To test this possibility, the RPE65 promoter was compared to another specific promoter of the RPE, vitelliform macular dystrophy-2 (VMD2)[13,14]. We compared the two promoters, RPE65 and VMD2, in driving the expression of Rap1a in the RPE and in reducing experimental CNV in wild type mice. The effect of the expression of exogenous CARap1a was evaluated and compared in eyes treated with the two different promoters, RPE65 and VMD2
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