Merbarone, a catalytic inhibitor of DNA topoisomerase II, induces apoptosis in CEM cells through activation of ICE/CED-3-like protease

Merbarone (5-[N-phenyl carboxamido]-2-thiobarbituric acid) is an anticancer drug that inhibits the catalytic activity of DNA topoisomerase II (topo II ) without damaging DNA or stabilizing DNA-topo II cleavable complexes. Alth ough the cytotoxicity of the complex-stabilizing DNA-topo II inhibitors suc h as VP-16 (etoposide) has been partially elucidated, the cytotoxicity of m erbarone is poorly understood. Here, we report that merbarone induces progr ammed cell death or apoptosis in human leukemic CEM cells, characterized by internucleosomal DNA cleavage and nuclear condensation. Treatment of CEM c ells with apoptosis-inducing concentrations of merbarone caused activation of c-Jun NH2-terminal kinase/stress-activated protein kinase, c-jun gene in duction, activation of caspase-3/CPP32-like protease but not caspase-1, and the proteolytic cleavage of poly(ADP-ribose) polymerase. Treatment of CEM cells with a potent inhibitor of caspases, Z-Asp-2.6-dichlorobenzoyloxymeth yl-ketone, inhibited merbarone-induced caspase-3/CPP32-like activity and ap optosis in a dose-dependent manner. These results indicate that the catalyt ic inhibition of topo II by merbarone leads to apoptotic cell death through a caspase-3-like protease-dependent mechanism. These results further sugge st that c-Jun and c-Jun NH2-terminal kinase/ stress-activated protein kinas e signaling may be involved in the cytotoxicity of merbarone.