A Novel Sphingosine-l-Phosphate Receptor 1 Antagonist Prevents the Proliferation and Relaxation of Vascular Endothelial Cells by Sphingosine-l-Phosphate

摘要

A sphingosine-l-phosphate receptor 1 (S1Р_1) antagonist is expected to be an anti-angiogenic compound; however, there are few reports that demonstrated that a SIP, inhibitor improved the disease state in an angio-genic animal model. Since we determined that a prototype S1P_1, antagonist was an in vivo angiogenesis inhib-itor, we developed the derivatives to acquire more effective compounds. In this report, we show the S1P_1 antago-nistic activity of some representatives, especially compound 5 {sodium 4-[(4-butoxyphenyl)thio]-2'-[{4-((hep-tylthio)methyl]-2-hydroxyphenyl}(hydroxy)methyl]biphenyl-3-sulfonate}. The IC_(50) values calculated from an intracellular cyclic AMP measurement assay and a (~(33)P]sphingosine-1-phosphate (Sph-1-P)/511'1 binding assay were 38 and 200 nM, respectively. A subtype specificity test for the other Sph-1-P receptors showed that compound 5 was the SIP_1-directional antagonist. It also inhibited the proliferation, migration, and tube formation of human umbilical vein endothelial cells stimulated by Sph-1-P with the IC_(50) values of 18, 650, and 230 пм, respectively. A cytotoxicity assay concurrently performed with a tube formation assay supported the hypothesis that these biological effects were not due to its cytotoxicity. Furthermore, administration (10 mg/kg, intravenously) to anesthetized Sprague-Dawley rats inhibited Sph-1-P-induced hypotension by 100-90% for 30 min. This is pre-sumably through the inhibition of Sph-1-P-induced vasorelaxation, mainly by the blocking of 511'1 and/or S1P3. Taken together, these results show that compound 5 is an inhibitor of in vitro and in vivo Sph-1-P signaling, and that it will be useful to elucidate the in vivo effect of Sph-1-P on vascular endothelial cells.