title: Induction of G0/G1 cell cycle arrest in ovarian carcinoma cells by the anti-inflammatory drug NS-398, but not by COX-2-specific RNA interference.
abstract: Cyclooxygenases, particularly COX-2, play an important role in tumor development and progression. We have previously shown that COX-2 expression is an independent prognostic factor in human ovarian carcinoma. In this study, we investigated the effects of the inhibition of COX isoforms by the NSAID NS-398 as well as by COX-isoform-specific RNA interference (RNAi) in the human ovarian carcinoma cell lines OVCAR-3 and SKOV-3. OVCAR-3 cells showed a constitutive expression of COX-1 and an induction of high levels of COX-2 and PGE(2) after stimulation with interleukin-1beta. In contrast, SKOV-3 cells were negative for both COX isoforms. In OVCAR-3 cells, PGE(2) production was inhibited by NS-398 in concentrations of 1 microM and by a COX-2-specific silencing RNA (siRNA), while a COX-1-specific siRNA did not have an effect. This suggests that COX-2 is the major source of PGE(2) in this cell line. To dissociate COX-2-specific and non-COX-2-specific effects on cell proliferation, a proliferation assay was performed after incubation of cells with NS-398 and COX siRNAs. NS-398 induced an inhibition of cell proliferation at concentrations of 50-500 microM, which are above the concentrations needed for the inhibition of PGE(2) production. This inhibitory effect was present in the COX-positive cell line OVCAR-3 as well as in the COX-negative cell line SKOV-3 and could not be reverted by addition of exogenous PGE(2). Neither COX-1- nor COX-2-specific siRNAs had an effect on cell proliferation of OVCAR-3 cells. Cell cycle analysis showed an increased accumulation of cells in the G0/G1 phase after treatment with NS-398, but not with COX siRNAs. These experiments suggest that NS-398 reduced cell proliferation in ovarian carcinoma cells by induction of G0/G1 cell cycle arrest independent of COX-2 inhibition. Our study shows that specific inhibition of COX isoforms by RNAi could be used to dissociate effects of NSAIDs. Furthermore, our results suggest that cell cycle arrest is one of the primary mechanisms responsible for the antiproliferative effects of NS-398 on ovarian carcinoma cells.
authors.affiliation: 1Institute of Pathology, Charité Hospital, Humboldt University, Campus Mitte, Schumannstr. 20/21, D-10117 Berlin, Germany. carsten.denkert@charite.de
authors.forename: Carsten
authors.initials: C
authors.lastname: Denkert
authors.forename: Antje
authors.initials: A
authors.lastname: Fürstenberg
authors.forename: Peter Ted
authors.initials: PT
authors.lastname: Daniel
authors.forename: Ines
authors.initials: I
authors.lastname: Koch
authors.forename: Martin
authors.initials: M
authors.lastname: Köbel
authors.forename: Wilko
authors.initials: W
authors.lastname: Weichert
authors.forename: Antje
authors.initials: A
authors.lastname: Siegert
authors.forename: Steffen
authors.initials: S
authors.lastname: Hauptmann
mesh_terms: D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000494:pharmacology*; D002455:Cell Division / Q000187:drug effects; D045744:Cell Line, Tumor; D002470:Cell Survival / Q000187:drug effects; D051545:Cyclooxygenase 1; D051546:Cyclooxygenase 2; D015232:Dinoprostone / Q000096:biosynthesis; D005260:Female; D016193:G1 Phase / Q000187:drug effects*; D015971:Gene Expression Regulation, Enzymologic; D006801:Humans; D007527:Isoenzymes / Q000037:antagonists & inhibitors* / Q000235:genetics; D008565:Membrane Proteins; D009578:Nitrobenzenes / Q000494:pharmacology*; D010051:Ovarian Neoplasms / Q000188:drug therapy* / Q000201:enzymology / Q000473:pathology; D011451:Prostaglandin-Endoperoxide Synthases / Q000235:genetics; D034622:RNA Interference; D016192:Resting Phase, Cell Cycle / Q000187:drug effects*; D013449:Sulfonamides / Q000494:pharmacology*
chemical_list: D000894:Anti-Inflammatory Agents, Non-Steroidal; D007527:Isoenzymes; D008565:Membrane Proteins; D009578:Nitrobenzenes; D013449:Sulfonamides; C080955:N-(2-cyclohexyloxy-4-nitrophenyl)methanesulfonamide; D051545:Cyclooxygenase 1; D051546:Cyclooxygenase 2; C496511:PTGS1 protein, human; C496540:PTGS2 protein, human; D011451:Prostaglandin-Endoperoxide Synthases; D015232:Dinoprostone