Dual VEGF-Targeting Strategy Via AAV2-Delivered sFLT-1 and shVEGF for Retinal Neovascularization Therapy.

PURPOSE: Retinal neovascular diseases, often associated with elevated vascular endothelial growth factor (VEGF) levels, are major causes of blindness. Current therapies lack sustained efficacy and safety. This study aimed to evaluate the effectiveness of an adeno-associated virus 2 (AAV2)-based gene therapy vector carrying soluble fms-like tyrosine kinase-1 (sFLT-1) and short hairpin RNA targeting VEGF (shVEGF) in treating retinal neovascularization. METHODS: P12 mice were divided into six groups: normal group, control group, AAV2-EGFP group, AAV2-sFLT group, AAV2-shVEGF group, and AAV2-sFLT-shVEGF group. AAV2 vectors encoding sFLT-1 alone, shVEGF alone, or a combination of both sFLT-1 and shVEGF were administered to oxygen-induced retinopathy (OIR) mice via intravitreal injection. Fundus photography and fluorescein fundus angiography (FFA) were used to evaluate retinal vascular morphology and density. Hematoxylin and eosin (H&E) staining was used to observe the growth characteristics of retinal neovascularization. The expression levels of angiogenic factors, inflammatory mediators, and blood-retina barrier-associated markers were assessed using immunofluorescence and quantitative PCR (qPCR). Electroretinography (ERG) was performed to evaluate retinal function. RESULTS: All three gene therapy groups effectively suppressed pathological neovascularization in OIR mice, although their therapeutic outcomes showed subtle differences. The AAV2-shVEGF group demonstrated the strongest efficacy in promoting the regression of the central non-perfusion area. The AAV2-sFLT-shVEGF group was more effective in suppressing neovascular growth and inflammatory infiltration. All three groups exhibited comparable effects in reducing angiogenic factor expression and enhancing blood-retina barrier-related markers. Notably, only the AAV2-sFLT-shVEGF group showed a pronounced ability to restore retinal function. CONCLUSIONS: Dual inhibition with AAV2-sFLT-shVEGF offers a promising therapeutic approach for retinal neovascular diseases, demonstrating efficacy in reducing neovascularization and improving retinal health.