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Quinacrine(QU), a phospholipase-A-2(PLA-2) inhibitor has been used clinically as a chemotherapeutic adjuvant. To understand the mechanisms leading to its chemotherapeutic effect , we have investigated QU-induced apoptotic signaling pathways in human cervical squamous carcinoma HeLa cells. In this study, we found that QU induced c was QU concentration- andtime-dependent, and preceded activation of caspase-9 and-3. Flow cytometric FACScan analysis using fluorescence intensities of DiOC6 demonstrated that QU-induced cytochrome c release was independent of mitochondrial permeability transtion(MPT), since the concentrations of QU that induces cytochrome c release did not alter mitochondrial membrane potential(▲Ψm). Moreover, kinetic analysis of caspase activities showed that cytochrome c release led to the activation of caspase-9 and downstream death effector, caspase-3. Caspase-3 inhibitor (Ac-DEVD-CHO) partially blocked QU-induced apoptosis, suggesting the importance of caspase-3 in this apoptotic signaling mechanism. Supplementation with arachidonic acid(AA) sustained caspase-3 activation induced by QU. Using inhibitors against celleular arachidonate metabolism of lipooxygenase(Nordihydrozyguaiaretic Acid,NDGA) and cyclooxygenase(5,8,11,14-Eicosateraynois Acid, ETYA) demonstrated that QU-induced apoptoptic signaling may be dependent on its role as a PLA-2 inhibitor. Interestingly, NDGA attenuated QU-induced cytochrome c release, caspase activity as well as apoptotic cell death. The blockade of cytochrome c release by NDGA was much more effective than that attained with cyclosporin A (CsA), a MPT inhibitor. ETYA was not effective in blocking cytochrome c release, except under very high concentrations. Caspase inhibitor z-VAD blocked the release of cytochrome c suggesting that this signaling events. In summary, we report caspase-8 activation may be upstream of the mitlchondrial events. In summary, we report that QU induced cytochrome c-dependent apoptotic signaling cascade, which may be dependent on its role as a PLA-2 inhibitor. This apoptotic mechanism induced by QU may contribute to its known chemotherapeutic effects.
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