Disulfiram/copper induces BAK-mediated caspase-independent apoptosis in MCF-7 cells

Disulfiram (DSF) and copper (Cu^2 +) in combination exhibit powerful anti-cancer effect on a variety of cancer cell lines. Here, we found that DSF/Cu²⁺ facilitated the accumulation of intracellular reactive oxygen species (ROS), and induced ROS-dependent apoptosis accompanied by chromatin condensation and phosphatidylserine externalization in MCF-7 cells. DSF/Cu²⁺ caused caspase-independent apoptosis by promoting the AIF translocation from mitochondria to nucleus. Most importantly, the cytotoxicity of DSF/Cu²⁺ was markedly inhibited by knocking out AIF, suggesting the indispensability of AIF in DSF/Cu²⁺-induced apoptosis. The pro-apoptotic protein BAK instead of BAX was upregulated and activated upon DSF/Cu²⁺ treatment, and the BAK knockout cells exhibited high resistance to DSF/Cu²⁺, indicating the importance of BAK in DSF/Cu²⁺-induced apoptosis. Additionally, both co-immunoprecipitation and live-cell quantitative fluorescence resonance energy transfer (FRET) analysis revealed that DSF/Cu²⁺ unlocked the binding of MCL-1 to BAK, which resulted in subsequent BAK homo-oligomerization. Overall, our data demonstrate for the first time that DSF/Cu²⁺ unlocks the binding of MCL-1 to BAK, thus leading BAK oligomerization and subsequent AIF nucleus translocation to mediate caspase-independent apoptosis in MCF-7 cells.