Vascular endothelial growth factor (VEGF) is a critical stimulus for both retinal and choroidal neovascularization, and for diabetic macular edema. We used mouse models for these diseases to explore the potential of gene transfer of soluble VEGF receptor-1 (sFlt-1) as a treatment. Intravitreous or periocular injection of an adenoviral vector encoding sFlt-1 (AdsFlt-1.10) markedly suppressed choroidal neovascularization at rupture sites in Bruch's membrane. Periocular injection of AdsFlt-1.10 also caused significant reduction in VEGF-induced breakdown of the blood-retinal barrier, but failed to significantly inhibit ischemia-induced retinal neovascularization. Periocular delivery of an adenoviral vector encoding pigment epithelium-derived factor (PEDF), another secreted protein, resulted in high levels of PEDF in the retinal pigmented epithelium and choroid, but not in the retina. This may explain why periocular injection of AdsFlt-1.10 inhibited choroidal, but not retinal neovascularization. Periocular delivery offers potential advantages over other routes of delivery and the demonstration that sFlt-1 enters the eye from the periocular space in sufficient levels to achieve efficacy in treating choroidal neovascularization and retinal vascular permeability is a novel finding that has important clinical implications. These data suggest that periocular gene transfer of sFlt-1 should be considered for treatment of choroidal neovascularization and diabetic macular edema.