Archives of Pharmacal Research

The Pharmaceutical Society of Korea (대한약학회)
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Quinacrin Induces Cytochrome c-dependent Apoptotic Signaling in Human Cervical Carcinoma Cells

  • Fasanmade, Adedigbo A. (Department of Pharmaceutics and Pharmacodynamics, Center for Pasrmaceutical Biotechnology, College of Pharmacy Centocor Biotechnology Inc.) ;
  • Owuor, Edward D. (Department of Pharmaceutics and Pharmacodynamics, Center for Pasrmaceutical Biotechnology, College of Pharmacy, Department of Pharmaceutics, College of Pharmacy, Rutgers University, Environmental and Occupational Health Science Institute) ;
  • Ee, Rachel P.L. (Department of Pharmaceutics and Pharmacodynamics, Center for Pasrmaceutical Biotechnology, College of Pharmacy) ;
  • Qato, Dima (Department of Pharmaceutics and Pharmacodynamics, Center for Pasrmaceutical Biotechnology, College of Pharmacy) ;
  • Heller, Mark (Centocor Biotechnologies Inc.,) ;
  • Kong, Ah Ng Tony (Department of Pharmaceutics and Pharmacodynamics, Center for Pasrmaceutical Biotechnology, College of Pharmacy, Department of Pharmaceutics, College of Pharmacy, Rutgers University, Environmental and Occupational Health Science Institute)
  • Published : 2001.04.01
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Abstract

Quinacrine (QU), a phospholipase-A2 (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 cytochrome c-dependent apoptotic signaling. The release of pro-apoptotic cytochrome c was QU concentration- and time-dependent, and preceded activation of caspase-9 and -3. Flow cytometric FACScan analysis using fluorescence intensities of $DiOC_6$/ demonstrated that QU-induced cytochrome c release was independent of mitochondrial permeability transition (MPT), since the concentrations of QU that induced cytochrome c release did not alter mitochondrial membrane potential (${\blacktriangle}{\Psi}_m$). 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 cellular arachidonate metabolism of lipooxygenase (Nordihydroxyguaiaretic Acid, NDGA) and cyclooxygenase (5,8,11,14-Eicosatetraynoic Acid, ETYA) demonstrated that QU-induced apoptotic signaling may be dependent on its role as a PLA-2 inhibitor. Interestingly, NDCA attenuated QU-induced cytochrome c release, caspase activity as well as apoptotic cell death. The blockade of cytochrome c release by NDCA 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 event is caspase dependent, and caspase-8 activation may be upstream of the mitochondrial 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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