Lonidamine triggers apoptosis via a direct, Bcl-2-inhibited effect on the mitochondrial permeability transition pore

The molecular mode of action of lonidamine, a therapeutic agent employed in cancer chemotherapy, has been elusive. Here we provide evidence that lonid amine (LND) acts on mitochondria to induce apoptosis, LND provokes a disrup tion of the mitochondrial transmembrane potential which precedes signs of n uclear apoptosis and cytolysis, The mitochondrial and cytocidal effects of LND are not prevented by inhibitors of caspases or of mRNA or protein synth esis. However, they are prevented by transfection-enforced overexpression o f Bcl-2, an oncoprotein which inhibits apoptosis by stabilizing the mitocho ndrial membrane barrier function. Accordingly, the cell death-inducing effe ct of LND is amplified by simultaneous addition of PK11195, an isoquinoline ligand of the peripheral benzodiazepine receptor which antagonizes the cyt oprotective effect of Bcl-2, When added to isolated nuclei, LND fails to pr ovoke DNA degradation unless mitochondria are added simultaneously. In isol ated mitochondria, LND causes the dissipation of the mitochondrial inner tr ansmembrane potential and the release of apoptogenic factors capable of ind ucing nuclear apoptosis in vitro. Thus the mitochondrion is the subcellular target of LND. All effects of LND on isolated mitochondria are counteracte d by cyclosporin A, an inhibitor of the mitochondrial PT pore. We therefore tested the effect of LND on the purified PT pore reconstituted into liposo mes, LND permeabilizes liposomal membranes containing the PT pore. This eff ect is prevented by addition of recombinant Bcl-2 protein but not by a muta nt Bcl-2 protein that has lost its apoptosis-inhibitory function. Altogethe r these data indicate that LND represents a novel type of anti-cancer agent which induces apoptosis via a direct effect on the mitochondrial PT pore.