Inhibition of GSK-3β reverses the pro-apoptotic effect of proadifen (SKF-525A) in HT-29 colon adenocarcinoma cells.
Proadifen (SKF-525A) is a well-known inhibitor of cytochrome P450 monooxygenases. Besides the prevention of drug metabolism it affects the proliferation of cancer cells, although the mechanisms of possible anti-cancer activity of proadifen have not been fully understood yet. The aim of this study therefore was to evaluate the potential anti-proliferative effect of proadifen on HT-29 colon cancer cells. Our results show that proadifen inhibited the growth of HT-29 cells by the accumulation of cells in the G1 phase of the cell cycle, reduction of metabolic activity and colony formation and by the induction of apoptosis. Analyses of Western blots and flow cytometry revealed time- and dose-dependent phosphatidylserine externalization, caspase-3 activation and PARP cleavage. Intense upregulation of NAG-1 and ATF3 and downregulation of Mcl-1 and Egr-1 were also observed. Further investigation showed that NAG-1 gene silencing by siRNA had no effect on the pro-apoptotic action of proadifen. In contrast, we found that AR-A014418, the specific inhibitor of glycogen synthase kinase-3 β (GSK-3β), significantly decreased proadifen-induced apoptosis. Inactivation of GSK-3β (phosphorylation at serine 9) resulted in changes in phosphatidylserine externalization and caspase-3 activation. These data suggest that GSK-3β is an important factor in the induction of apoptosis in HT-29 colon cancer cells treated with proadifen.
Institute of Biology and Ecology, Department of Cellular Biology, Pavol Jozef Šafárik University in Košice, Slovakia.
Rastislav
R
Jendželovský
Ján
J
Koval
Jaromír
J
Mikeš
Zuzana
Z
Papčová
Jana
J
Plšíková
Peter
P
Fedoročko
D000230:Adenocarcinoma; D000970:Antineoplastic Agents / Q000494:pharmacology*; D017209:Apoptosis / Q000187:drug effects; D053148:Caspase 3 / Q000378:metabolism; D002453:Cell Cycle / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D003110:Colonic Neoplasms; D038362:Glycogen Synthase Kinase 3 / Q000037:antagonists & inhibitors*; D000071679:Glycogen Synthase Kinase 3 beta; D019073:HT29 Cells; D006801:Humans; D011335:Proadifen / Q000494:pharmacology*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology*; D013844:Thiazoles / Q000494:pharmacology*; D014508:Urea / Q000031:analogs & derivatives* / Q000494:pharmacology
D000970:Antineoplastic Agents; D047428:Protein Kinase Inhibitors; D013844:Thiazoles; C479831:N-(4-methoxybenzyl)-N'-(5-nitro-1,3-thiazol-2-yl)urea; D014508:Urea; D011335:Proadifen; C000605506:GSK3B protein, human; D000071679:Glycogen Synthase Kinase 3 beta; D038362:Glycogen Synthase Kinase 3; D053148:Caspase 3