Bcl-2 inhibitors sensitize tumor necrosis factor-related apoptosis-inducing ligand-induced apoptosis by uncoupling of mitochondrial respiration in human leukemic CEM cells

Previous studies have shown that the lymphoblastic leukemia CEM cell line is resistant to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis because of a low expression of caspase-8. Bcl-2 inhibitors, BH3I-2' and HA14-1, are small cell-permeable nonpeptide compounds, are able to induce apoptosis by mediating cytochrome c release, and also lead to dissipation of the mitochondrial membrane potential (DeltaPsim). This study aimed to use the Bcl-2 inhibitors to sensitize CEM cells to TRAIL-induced apoptosis by switching on the mitochondrial apoptotic pathway. We found that a low dose of BH3I-2' or HA14-1, which did not induce cytochrome c release, greatly sensitized CEM cells to TRAIL-induced apoptosis. In a similar manner to the classical uncoupler carbonyl cyanide m-chlorophenylhydrazone (CCCP), both BH3I-2' and HA14-1 induced a reduction in DeltaPsim, a generation of reactive oxygen species (ROS), an increased mitochondrial respiration, and a decreased ATP synthesis. This uncoupling function of the Bcl-2 inhibitors was responsible for the synergy with TRAIL-induced apoptosis. CCCP per se did not induce apoptosis but again sensitized CEM cells to TRAIL-induced apoptosis by uncoupling mitochondrial respiration. The uncoupling effect facilitated TRAIL-induced Bax conformational change and cytochrome c release from mitochondria. Inhibition of caspases failed to block TRAIL-mediated cell death when mitochondrial respiration was uncoupled. We observed that BH3I-2', HA14-1, or CCCP can overcome resistance to TRAIL-induced apoptosis in TRAIL-resistant cell lines, such as CEM, HL-60, and U937. Our results suggest that the uncoupling of mitochondrial respiration can sensitize leukemic cells to TRAIL-induced apoptosis. However, caspase activation per se does not represent an irreversible point of commitment to TRAIL-induced cell death when mitochondrial respiration is uncoupled.