title: Synergistic cytotoxicity and apoptosis of JTE-522, a selective cyclooxygenase-2 inhibitor, and 5-fluorouracil against bladder cancer.
abstract: OBJECTIVE
  Cyclooxygenase-2 (COX-2) is a key inducible enzyme involved in the production of prostaglandins that has been shown to induce apoptosis in various cancer cells. Several anticancer agents also mediate apoptosis and may share the common intracellular pathways leading to apoptosis. Since over expression of COX-2 has been demonstrated in bladder cancer, we reasoned that combination treatment with a COX-2 inhibitor and anticancer agents in bladder cancer cells may result in synergistic apoptosis. We examined whether the selective COX-2 inhibitor JTE-522 induces apoptosis in bladder cancer cells and whether JTE-522 may act synergistically with anticancer agents to achieve cytotoxicity and apoptosis in these cells.
  
  
  METHODS
  Cytotoxicity was determined by microculture tetrazolium dye assay. Synergy was assessed by isobolographic analysis.
  
  
  RESULTS
  COX-2 mRNA expression was observed in the HT1197 bladder cancer cell line. JTE-522 was cytotoxic in HT1197 cells. Treating HT1197 cells with JTE-522 combined with doxorubicin or mitomycin C did not show synergistic cytotoxicity. However, combination treatment of HT1197 cells with JTE-522 and 5-fluorouracil (5-FU) resulted in a synergistic cytotoxic effect. Synergy was also achieved in the T24 bladder cancer line. Synergistic cytotoxicity was noted irrespective of treatment sequence but the highest percent cytotoxicity was obtained when HT1197 cells were treated with JTE-522 and 5-FU simultaneously. The synergy achieved in cytotoxicity with JTE-522 and 5-FU was shown to be due to apoptosis. The mechanisms responsible for synergistic cytotoxicity and apoptosis was examined. Treating HT1197 cells with 5-FU enhanced expression of the pro-apoptotic molecule Bax, while JTE-522 treatment reduced expression of the anti-apoptotic molecule Bcl-XL, resulting in a significantly higher ratio of Bax-to-Bcl-XL.
  
  
  CONCLUSIONS
  This study shows that combination treatment of bladder cancer cells with the selective COX-2 inhibitor JTE-522 and 5-FU results in synergistic cytotoxicity and apoptosis due to the enhanced Bax-to-Bcl-XL expression ratio. These findings support the in vivo potential application of a combination of JTE-522 and 5-FU for bladder cancer.
authors.affiliation: Department of Urology, Kyoto Prefectural University of Medicine, Japan.
authors.forename: Yoichi
authors.initials: Y
authors.lastname: Mizutani
authors.forename: Kazumi
authors.initials: K
authors.lastname: Kamoi
authors.forename: Osamu
authors.initials: O
authors.lastname: Ukimura
authors.forename: Akihiro
authors.initials: A
authors.lastname: Kawauchi
authors.forename: Tsuneharu
authors.initials: T
authors.lastname: Miki
mesh_terms: D000964:Antimetabolites, Antineoplastic / Q000008:administration & dosage / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects*; D001557:Benzenesulfonates / Q000008:administration & dosage / Q000494:pharmacology*; D015153:Blotting, Western; D051546:Cyclooxygenase 2; D052246:Cyclooxygenase 2 Inhibitors; D016861:Cyclooxygenase Inhibitors / Q000008:administration & dosage / Q000494:pharmacology*; D004357:Drug Synergism; D005472:Fluorouracil / Q000008:administration & dosage / Q000494:pharmacology*; D006801:Humans; D007527:Isoenzymes / Q000037:antagonists & inhibitors* / Q000235:genetics / Q000378:metabolism; D008297:Male; D008565:Membrane Proteins; D010080:Oxazoles / Q000008:administration & dosage / Q000494:pharmacology*; D011451:Prostaglandin-Endoperoxide Synthases / Q000235:genetics / Q000378:metabolism; D011471:Prostatic Neoplasms / Q000378:metabolism / Q000473:pathology; D011518:Proto-Oncogene Proteins / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D012333:RNA, Messenger / Q000378:metabolism; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014407:Tumor Cells, Cultured / Q000187:drug effects; D001749:Urinary Bladder Neoplasms / Q000378:metabolism / Q000473:pathology*; D051028:bcl-2-Associated X Protein; D051020:bcl-X Protein; D000096926:Benzenesulfonamides
chemical_list: D000964:Antimetabolites, Antineoplastic; D001557:Benzenesulfonates; D051546:Cyclooxygenase 2; D052246:Cyclooxygenase 2 Inhibitors; D016861:Cyclooxygenase Inhibitors; D005472:Fluorouracil; D007527:Isoenzymes; D008565:Membrane Proteins; D010080:Oxazoles; D011451:Prostaglandin-Endoperoxide Synthases; D011518:Proto-Oncogene Proteins; D019253:Proto-Oncogene Proteins c-bcl-2; D012333:RNA, Messenger; D051028:bcl-2-Associated X Protein; D051020:bcl-X Protein; C110453:4-(4-cyclohexyl-2-methyloxazol-5-yl)-2-fluorobenzenesulfonamide; D000096926:Benzenesulfonamides; C494826:BAX protein, human; C494811:BCL2L1 protein, human; C496540:PTGS2 protein, human