S1P receptors control endothelial cell proliferation, migration, and survival. Evidence of the ability of S1P receptor modulators to influence multiple endothelial cell functions suggests their potential use for antiangiogenic effect. The main purpose of our study was to investigate the potential of siponimod for the inhibition of ocular angiogenesis in vitro and in vivo. We investigated the effects of siponimod on the metabolic activity (MTT assay), cell toxicity (LDH release), basal proliferation and growth factor induced proliferation (BrdU assay), and migration (transwell migration assay) of human umbilical vein endothelial cells (HUVEC) and retinal microvascular endothelial cells (HRMEC). The effects of siponimod on HRMEC monolayer integrity, and barrier function under basal conditions and TNF-α induced disruption were assessed using the trans-endothelial electrical resistance (TEER) and FITC-dextran permeability assays. Siponimod's effect on TNF-α induced distribution of barrier proteins in HRMEC was investigated using immunofluorescence. Finally, the effect of siponimod on ocular neovascularization in vivo was assessed using suture-induced corneal neovascularization in albino rabbits.Our results show that siponimod did not affect endothelial cell proliferation or metabolic activity, but significantly inhibited endothelial cell migration, increased HRMEC barrier integrity, and reduced TNF-α induced barrier disruption. Siponimod also protected against TNF-α induced disruption of claudin-5, ZO-1, and VE-cadherin in HRMEC. These actions are mainly mediated by S1PR 1 modulation. Finally, siponimod prevented the progression of suture-induced corneal neovascularization in albino rabbits. In conclusion, the effects of siponimod on various processes known to be involved in angiogenesis support its therapeutic potential in disorders associated with ocular neovascularization.

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