Mcl-1 is a unique anti-apoptotic member of Bcl-2 family. Its expression has been demonstrated to seriously impede the most authentic BH3 mimetic ABT-737-induced apoptosis in multiple cancer cell lines. S1, which has been previously identified as a pan-Bcl-2 inhibitor and pure BH3 mimetic by our group, was investigated here to challenge Mcl-1's protection. Cell viability assay and apoptosis assay showed S1 exhibits robust anticancer effectiveness in three selected ABT-737-resistant cancer cell lines, SMMC-7721, HCT116 and K562 cells. In contract with ABT-737, S1-induced apoptosis is regardless of the baseline of Mcl-1 expression. Interestingly, it dynamically modulates Mcl-1 through a combination of proteasome and caspase-mediated pathway and this dynamical change is associated well with the resistance, apoptosis onset, and apoptosis burst out induced by S1. To investigate how Mcl-1 dynamics is related with apoptosis at molecular level, we performed time-sequence and dose-dependent analysis of cell viability, western blot, and co-immunoprecipitation experiments. We found the trafficking of Bcl-2 family members determines Mcl-1 dynamical change and in turn apoptosis. First of all S1 untethers Bax and Bim from Bcl-2, but Bim shifts to Mcl-1 which stabilizes Mcl-1 and enforces the binding with Bak. Only when Bak is released by enough dose of S1 or enough exposure time is Mcl-1 downregualted to allow apoptosis occur. Using shRNA, we identified both S1-induced apoptosis and Mcl-1 dynamical modulation depend on Bak, not Bax. As such, much distinct function of Bax and Bak in the S1-induced pathway was illustrated. While Bax can be fully compensated for by Bak, Bak could be partly compensated for by Bax. Consistently, Bax-deficient DU145 cell is sensitive to S1, whereas Bak-mutant MKN-28 cells is much more resistant. NMR analyses revealed that S1 highly mimic tBid to target the BH3-binding pocket of Mcl-1. Cytochrome c release assay in purified DU145 mitochondrial testified a nicely structural-to-functional mimic of tBid, but not Bim. The Bid-like property is also consistent with its Bak-dependent apoptosis induction. The in vitro model could be extended to an in vivo mouse xenograft model that S1 induces apoptosis through Mcl-1 dynamical modulation and released Bak is the biomarker of apoptosis. In conclusion, S1 could be a highly potent therapeutic molecule for the treatment of Mcl-1-driven cancer cells with a mechanism-based model. Bak could serve as a biomarker to specify cancer cells sensitive to S1. In addition, the highly tBid/Mcl-1 binding mimic property could be used to probe the sophisticated network of Bcl-2 family.

Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 102nd Annual Meeting of the American Association for Cancer Research; 2011 Apr 2-6; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2011;71(8 Suppl):Abstract nr 6. doi:10.1158/1538-7445.AM2011-6

©2011 American Association for Cancer Research
2011