Sensitization of human K562 leukemic cells to TRAIL-induced apoptosis by inhibiting the DNA-PKcs/Akt-mediated cell survival pathway

The mRNA levels of both DR4 and DR5 were significantly increased in K562 cells transfected with DNA-PKcs siRNA compared to the cells transfected with scrambled siRNA. We also found that suppression of DNA-PKcs using siRNA down-regulated c-FLIP and sensitized K562 cells to TRAIL-induced apoptosis through activation of caspase-8, -9 and -3. Moreover, the growth inhibitory effect of TRAIL in K562 cells was significantly increased after transfection with DNA-PKcs siRNA as compared with scrambled siRNA. This result was followed by increased susceptibility to TRAIL-induced apoptosis in K562 cells transfected with DNA-PKcs siRNA compared with that in the cells transfected with scrambled siRNA. Despite the fact that many cancer cells are sensitive to TNF-related apoptosis-inducing ligand (TRAIL)-induced apoptosis, human K562 leukemic cells showed resistance to TRAIL-induced apoptosis. Interestingly, K562/R3 cells, a stable TRAIL-sensitive variant isolated from K562 cells, showed down-regulation of DNA-PK/Akt pathway and a high responsiveness to TRAIL-mediated growth inhibition and apoptosis. We revealed that siRNA-mediated suppression of DNA-PKcs led to decreased phosphorylation of Akt and Bad, a target molecule of Akt, and increased expression of DR4/DR5. Also, we found that suppression of DNA-PKcs using siRNA down-regulated c-FLIP and sensitized K562 cells to TRAIL-induced apoptosis through activation of caspase-8, -9 and -3. In addition, we revealed that treatment with DMNB, a specific inhibitor of DNA-PK, resulted in an increase of DR4/DR5 mRNA levels and their surface expression and a decrease of c-FLIP mRNA levels in K562 cells. DMNB potentiated TRAIL-induced cytotoxicity and apoptosis through inhibition of DNA-PK/Akt pathway and activation of caspase-8, -9 and -3 in K562 cells. This study is the first to show that a protective role of DNA-PK/Akt pathway against TRAIL-induced apoptosis and thus TRAIL in combination with agents that inhibit DNA-PK/Akt pathway would have clinical applicability in treating TRAIL-insensitive human leukemic cells. This model may provide a novel framework for overcoming TRAIL resistance of other cancer cells with agents that inhibit DNA-PK/Akt pathway.