treated with STK295900 were arrested in G2 phase without activation of DNA damage checkpoint. These findings may therefore suggest a potential development of symmetric bibenzimidazole as a chemotherapeutic agent. In the search for new chemotherapeutic agents from the small molecule library, we identified STK295900 that exhibited efficient antiproliferative activity against various cancer cell lines of different origin, especially MCF7 and HepG2. Furthermore, analyzing the DprE1-IN-1 effect of STK295900 on HeLa cells demonstrated that it induced G2 cell cycle arrest. This is due to STK295900-induced accumulation of 4N DNA content with no significant change in mitotic index. In addition, STK295900-induced G2 arrest was confirmed by investigating the cell cycle regulatory proteins. Progression through the eukaryotic cell cycle is driven in part by a subfamily of Cdks whose activities are modulated by NSC 601980 distributor forming bipartite complexes with different cyclins. Levels of cyclins oscillate throughout the cell cycle whereas Cdk protein levels remain stable. Therefore, the activity of Cdks is regulated by the presence of different cyclins. In mitotic cells, cyclin B1 level was relatively high whereas cyclin A was undetectable. In contrast, STK295900 showed similar effect as camptothecin and etoposide did on cyclin A and cyclin B1 accumulation without induction of Histone H3 phosphorylation at S10, which is crucial for chromosome condensation and cell-cycle progression during mitosis. STK295900 belongs to a class of symmetric bibenzimidazole group. Compounds containing benzimidazole ring have been used extensively for pharmacological purposes such as antimicrobial and anticancer agents. Several asymmetric, head-to-tail bibenzimidazole derivatives, such as Hoechst 33258 and Hoechst 33342, exhibited antitumor activity by binding to minor groove of DNA at three consecutive A:T base pairs, leading to the inhibition of Top 1 activity. In addition, the symmetric bibenzimidazole derivatives, containing two groups of benzimidazole linked in head-to-head fashion, have been reported that they bind DNA minor groove with extending the binding site to four A:T base pairs and exhibit antitumor activity. However, there is no report on the mechanism of action for their antitumor activity. Here, we showe