Recent studies have shown that
integrin alpha v beta 3, a receptor for
vitronectin, plays an important role in
tumor-induced angiogenesis and
tumor growth and that antagonists of alpha v beta 3 inhibit angiogenic processes including endothelial cell adhesion and migration. On the other hand, most inhibitors of
integrin alpha v beta 3 are
peptide antagonists that include the
Arg-Gly-Asp (RGD) motif. We therefore reasoned that non-
peptide inhibitors of endothelial cell adhesion to
vitronectin might be useful for inhibition of
tumor angiogenesis in vivo. We screened for low-molecular-weight natural products able to inhibit adhesion of human umbilical vein endothelial cells (HUVECs) to
vitronectin, and pyrrothine group compounds including
aureothricin, thioaurin and
thiolutin were isolated from microbial culture broths. Of these compounds,
thiolutin inhibited adhesion of HUVECs to
vitronectin the most effectively (IC(50), 0.83 microM). In vivo experiments showed that
thiolutin significantly suppressed angiogenesis induced by
tumor cells (S-180), a pathological form of neovascularization, in a mouse dorsal air sac assay system. To explore the mechanism of inhibition of HUVEC adhesion to
vitronectin by
thiolutin, we examined the effect of this agent on intracellular cell adhesion signaling. We found that the amount of
paxillin in HUVECs was significantly reduced by
thiolutin treatment, while those of other focal adhesion
proteins including
vinculin and
focal adhesion kinase (FAK) were not. Metabolic labeling experiments showed that
thiolutin enhanced degradation of
paxillin in HUVECs.
Protease inhibitors (
MG115 and E64-D) decreased the rate of degradation of the
paxillin induced by
thiolutin and partially restored
thiolutin-induced inhibition of HUVEC adhesion to
vitronectin. Based on these findings, we concluded that
thiolutin, an inhibitor of HUVEC adhesion to
vitronectin, reduces the
paxillin level in HUVECs and suppresses
tumor cell-induced angiogenesis in vivo.