Design, Synthesis and Biological Investigation of 2-Anilino Triazolopyrimidines as Tubulin Polymerization Inhibitors with Anticancer Activities.

A further investigation aiming to generate new potential antitumor agents led us to synthesize a new series of twenty-two compounds characterized by the presence of the 7-(3',4',5'-trimethoxyphenyl)-[1,2,4]triazolo[1,5- a ]pyrimidine pharmacophore modified at its 2-position. Among the synthesized compounds, three were significantly more active than the others. These bore the substituents p -toluidino ( 3d ), p -ethylanilino ( 3h ) and 3',4'-dimethylanilino ( 3f ), and these compounds had IC 50 values of 30-43, 160-240 and 67-160 nM, respectively, on HeLa, A549 and HT-29 cancer cells. The p -toluidino derivative 3d was the most potent inhibitor of tubulin polymerization (IC 50 : 0.45 µM) and strongly inhibited the binding of colchicine to tubulin (72% inhibition), with antiproliferative activity superior to CA-4 against A549 and HeLa cancer cell lines. In vitro investigation showed that compound 3d was able to block treated cells in the G2/M phase of the cell cycle and to induce apoptosis following the intrinsic pathway, further confirmed by mitochondrial depolarization and caspase-9 activation. In vivo experiments conducted on the zebrafish model showed good activity of 3d in reducing the mass of a HeLa cell xenograft. These effects occurred at nontoxic concentrations to the animal, indicating that 3d merits further developmental studies.