Carboxyamido-triazole inhibits angiogenesis by blocking the calcium-mediated nitric-oxide synthase-vascular endothelial growth factor pathway

The induction of angiogenesis is known to play a critical role in the succe ssful growth, invasion, and metastasis of a tumor. A tumor will not grow be yond a few cubic millimeters without the formation of its own capillary net work. Several antiangiogenic agents are under investigation in the clinic s etting for the treatment of cancer. Carboxyamido-triazole (CAI), an inhibit or of Ca2+-mediated signal transduction, has been previously shown to inhib it angiogenesis in vitro and in vivo and to downregulate matrix metalloprot einase-2 in vitro. Diminished levels of intracellular Ca2+ result in decrea sed nitric-oxide synthase (NOS) activity and thereby inhibit the production and release of NO. The antiangiogenic activity of CAI was investigated by assessing microvessel growth from rat aortic segments and in cell culture u sing human aortic endothelial cells (HAECs). With these models, vascular en dothelial growth factor (VEGF) and NOS production and secretion were evalua ted. CAI concentrations ranging from 0.25 to 12.0 mu g/ml inhibited new mic rovessel formation in rat aortic cultures and HAEC proliferation in a dose- dependent manner. Additionally, HAECs treated with CAI showed a dose-depend ent decrease of NOS expression and a decrease in both VEGF expression and s ecretion. Rat aortic segments demonstrated decreased VEGF expression in sit u on immunostaining. These data suggest that modulation of the NOS-NO-VEGF pathway through Ca2+-mediated signaling by CAI inhibits angiogenesis in vit ro.