MT-21 is a synthetic apoptosis inducer that directly induces cytochrome c release from mitochondria

We reported previously that a synthetic compound, MT-21, induced Apoptosis by activating c-Jun-NH2-terminal kinase via the Krs/MST protein, which is activated by Caspase-3 cleavage dependent on Reactive Oxygen Species production. Here we examine the activation mechanism of Caspase-3, an important cysteine aspartic protease, during MT-21-induced Apoptosis. We found that MT-21 activated Caspase-3 via caspase-9, but not via Caspase-8. In addition, MT-21 induced the release of cytochrome c from the mitochondria that is necessary to activate caspase-9, and this release occurred before a change in membrane potential. This initiation process of MT-21-induced Apoptosis was suppressed by overexpression of Bcl-2, which is known to prevent cells from undergoing Apoptosis in response to a variety of stimuli. Moreover, when we treated mitochondria isolated from the cells with MT-21, the direct release of cytochrome c from the mitochondria was observed, whereas this effect was not observed in the mitochondria isolated from cells that overexpressed Bcl-2. Other apoptosis-inducing agents known to induce Apoptosis via cytochrome c release from the mitochondria failed to release cytochrome c directly from isolated mitochondria. These findings indicate that MT-21 is a possible candidate antitumor agent that is able to induce Apoptosis via the direct release of cytochrome c from the mitochondria.