Goniodomin A (GDA) is an antifungal polyether macrolide isolated from the dinoflagellate Goniodoma pseudogoniaulax. Previous studies revealed that GDA profoundly affected cytoskeletal reorganization. We examined the effect of GDA on the angiogenic properties of vascular endothelial cells. GDA itself did not affect proliferation of, migration of, and tube formation in type I collagen gels by, bovine aortic endothelial cells (BAECs). Proliferation of BAECs stimulated by bFGF was not affected by GDA at concentrations of up to 10 nM. However, at similar concentrations, GDA significantly inhibited bFGF-induced migration and tube formation in type I collagen gels by BAECs. Actin reorganization is required for cell migration. GDA caused the perinuclear aggregation of filamentous actin and inhibited stress fiber formation in bFGF- or VEGF-stimulated BAECs and lysophosphatidic acid-stimulated HeLa cells. However, GDA did not affect stress fiber structures already formed through Gβγ expression or in constitutively active RhoA mutant HeLa cells. Finally, GDA inhibited forming of vasucular system in a chorioallantoic membrane. Our results indicated that GDA suppressed angiogenic properties of ECs at least in part through the inhibition of actin reorganization and inhibited angiogenesis in vivo.