Effects of carbonyl cyanide p-(trifluoromethoxy) phenylhydrazone on the growth inhibition in human pulmonary adenocarcinoma Calu-6 cells.
Carbonyl cyanide p-(trifluoromethoxy) phenylhydrazone (FCCP) is an uncoupler of mitochondrial oxidative phosphorylation in eukaryotic cells. Here, we evaluated the in vitro effects of FCCP on the growth of Calu-6 lung cancer cells. FCCP inhibited the growth of Calu-6 cells with an IC(50) of approximately 6.64+/-1.84 microM at 72 h, as shown by MTT. DNA flow cytometric analysis indicated that FCCP induced G1 phase arrest below 20 microM of FCCP. Treatment with FCCP decreased the level of CDKs and cyclines in relation to G1 phase. In addition, FCCP not only increased the p27 level but also enhanced its binding with CDK4, which was associated with hypophosphorylation of Rb protein. While transfection of p27 siRNA inhibited G1 phase arrest in FCCP-treated cells, it did not enhance Rb phosphorylation. FCCP also efficiently induced apoptosis. The apoptotic process was accompanied with an increase in sub-G1 cells, annexin V staining cells, mitochondria membrane potential (MMP) loss and cleavage of PARP protein. All of the caspase inhibitors (caspase-3, -8, -9 and pan-caspase inhibitor) markedly rescued the Calu-6 cells from FCCP-induced cell death. However, knock down of p27 protein intensified FCCP-induced cell death. Moreover, FCCP induced the depletion of GSH content in Calu-6 cells, which was prevented by all of the caspase inhibitors. In summary, our results demonstrated that FCCP inhibits the growth of Calu-6 cells in vitro. The growth inhibitory effect of FCCP might be mediated by cell cycle arrest and apoptosis via decrease of CDKs and caspase activation, respectively. These findings now provide a better elucidation of the mechanisms involved in FCCP-induced growth inhibition in lung cancer.
Han
Yong Hwan
YH
Department of Physiology, Medical School, Institute for Medical Sciences, Chonbuk National University, Jeonju 561-180, Republic of Korea.
Moon
Hwa Jin
HJ
You
Bo Ra
BR
Kim
Sung Zoo
SZ
Kim
Suhn Hee
SH
Park
Woo Hyun
WH
D000230:Adenocarcinoma / Q000188:drug therapy* / Q000473:pathology; D017304:Annexin A5 / Q000378:metabolism; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone / Q000494:pharmacology*; D002453:Cell Cycle / Q000187:drug effects; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D018844:Cyclin-Dependent Kinases / Q000378:metabolism; D004273:DNA, Neoplasm / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D004353:Drug Evaluation, Preclinical; D005562:Formazans / Q000378:metabolism; D016193:G1 Phase / Q000187:drug effects; D006801:Humans; D020128:Inhibitory Concentration 50; D008175:Lung Neoplasms / Q000188:drug therapy* / Q000473:pathology; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D008928:Mitochondria / Q000187:drug effects; D051336:Mitochondrial Membranes / Q000187:drug effects; D013778:Tetrazolium Salts / Q000378:metabolism; D013997:Time Factors; D014475:Uncoupling Agents / Q000494:pharmacology*
D017304:Annexin A5; D000970:Antineoplastic Agents; D004273:DNA, Neoplasm; D005562:Formazans; D013778:Tetrazolium Salts; D014475:Uncoupling Agents; C079782:MTT formazan; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; D018844:Cyclin-Dependent Kinases