The Novel Antitubulin Agent TR-764 Strongly Reduces Tumor Vasculature and Inhibits HIF-1α Activation.

Tubulin binding agents (TBAs) are commonly used in cancer therapy as antimitotics. It has been described that TBAs, like combretastatin A-4 (CA-4), present also antivascular activity and among its derivatives we identified TR-764 as a new inhibitor of tubulin polymerization, based on the 2-(alkoxycarbonyl)-3-(3',4',5'-trimethoxyanilino)benzo[b]thiophene molecular skeleton. The antiangiogenic activity of TR-764 (1-10 nM) was tested in vitro on human umbilical endothelial cells (HUVECs), and in vivo, on the chick embryo chorioallantoic membrane (CAM) and two murine tumor models. TR-764 binding to tubulin triggers cytoskeleton rearrangement without affecting cell cycle and viability. It leads to capillary tube disruption, increased cell permeability, and cell motility reduction. Moreover it disrupts adherens junctions and focal adhesions, through mechanisms involving VE-cadherin/β-catenin and FAK/Src. Importantly, TR-764 is active in hypoxic conditions significantly reducing HIF-1α. In vivo TR-764 (1-100 pmol/egg) remarkably blocks the bFGF proangiogenic activity on CAM and shows a stronger reduction of tumor mass and microvascular density both in murine syngeneic and xenograft tumor models, compared to the lead compound CA-4P. Altogether, our results indicate that TR-764 is a novel TBA with strong potential as both antivascular and antitumor molecule that could improve the common anticancer therapies, by overcoming hypoxia-induced resistance mechanisms.