877877-35-5

TW-37 (TW37) is a novel nonpeptide small-molecule inhibitor designed using a structure-based design strategy. TW-37 targets the BH3-binding groove in Bcl-2 where proapoptotic Bcl-2 proteins, such as Bak, Bax, and Bid bind. In fluorescence polarization-based binding assays using recombinant Bcl-2 and Bcl-xL proteins, TW-37 binds to Bcl-2 and Bcl-xL with K _i values of 290 and 1110 nM, respectively. TW-37 has an IC ₅₀ of 1.8 μM for endothelial cells but shows no cytotoxic effects for fibroblasts at concentrations up to 50 μM. The mechanism of TW-37-induced endothelial cell death is apoptosis, in a process mediated by mitochondrial depolarization and activation of caspase-9 and caspase-3. The effect of TW-37 on endothelial cell apoptosis is not prevented by coexposure to the growth factor milieu secreted by tumor cells. Inhibition of the angiogenic potential of endothelial cells (i.e., migration and capillary sprouting assays) and expression of the angiogenic chemokines CXCL1 and CXCL8 are accomplished at subapoptotic TW-37 concentrations (0.005-0.05 μM). TW-37 is a potent Bcl-2 and Mcl-1 inhibitor. In fluorescence polarization-based binding assays using recombinant Bcl-2, Bcl-xL, and Mcl-1 proteins, TW-37 binds to Bcl-2, Bcl-xL, and Mcl-1 with K _i values of 290, 1,110 and 260 nM, respectively.

A murine model of humanized vasculature is used to investigate the biological effect of TW-37 (TW37) on human microvascular endothelial cell in vivo. Using this model, a significant decrease is observed in total blood vessel number (P<0.05) comparing both 3 and 30 mg/kg TW-37 against vehicle control. In addition to reduction in total number of blood vessels, an unusual number of occluded vessels are occurring in the treated groups. The levels of vessel occlusion are assessed by counting completely blocked vessels and determining their number as a percentage of total vessel number. TW-37 concentration mediates a significant increase in the number of occluded vessels when compared with control.