sb-415286 and Stomach-Neoplasms

Resistance to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis has been reported in some cancer cells, including AGS human gastric adenocarcinoma cells. Reducing this resistance might shed light on the treatment of human gastric adenocarcinoma. In this study, we examined whether glycogen synthase kinase-3 (GSK-3) inhibitors can restore TRAIL responsiveness in gastric adenocarcinoma cells. The effect of two GSK-3 inhibitors, SB-415286, and LiCl, on apoptosis signaling of TRAIL in human gastric adenocarcinoma cell lines and primary gastric epithelial cells was analyzed. Both inhibitors can sensitize gastric adenocarcinoma cells, but not primary gastric epithelial cells, to TRAIL-induced apoptosis by increasing caspase-8 activity and its downstream signal transmission. Adding p53 siRNA can downregulate GSK-3 inhibitor-related sensitization to TRAIL-induced apoptosis and caspase-3 activity. GSK-3 inhibitors strongly activate the phosphorylation of JNK. Inhibition of JNK leads to earlier and more intense apoptosis, showing that the activation of JNK may provide anti-apoptotic equilibrium of pro-apoptotic cells. Our observations indicate that GSK-3 inhibitors can sentize AGS gastric adenocarcinoma cells to TRAIL-induced apoptosis. Therefore, in certain types of gastric adenocarcinoma, GSK-3 inhibitor might enhance the antitumor activity of TRAIL and mightbe a promising candidate for the treatment of certain types of gastric adenocarcinoma.

Topics: Adenocarcinoma; Aminophenols; Apoptosis; Cell Line, Tumor; Drug Screening Assays, Antitumor; Glycogen Synthase Kinase 3; Humans; Lithium Chloride; Maleimides; Protein Kinase Inhibitors; RNA, Small Interfering; Stomach Neoplasms; TNF-Related Apoptosis-Inducing Ligand; Tumor Suppressor Protein p53

Gastric cancer is the second most common cause of global cancer-related mortality. Although dedifferentiation predicts poor prognosis in gastric cancer, the molecular mechanism underlying dedifferentiation, which could provide fundamental insights into tumor development and progression, has yet to be elucidated. Furthermore, the molecular mechanism underlying the effects of hexamethylene bisacetamide (HMBA), a recently discovered differentiation inducer, requires investigation and there are no reported studies concerning the effect of HMBA on gastric cancer.. Based on the results of FACS analysis, the levels of proteins involved in the cell cycle or apoptosis were determined using western blotting after single treatments and sequential combinations of HMBA and LiCl. GSK-3β and proton pump were investigated by western blotting after up-regulating Akt expression by Ad-Akt infection. To investigate the effects of HMBA on protein localization and the activities of GSK-3β, CDK2 and CDK4, kinase assays, immunoprecipitation and western blotting were performed. In addition, northern blotting and RNase protection assays were carried out to determine the functional concentration of HMBA.. HMBA increased p27(Kip1) expression and induced cell cycle arrest associated with gastric epithelial cell differentiation. In addition, treating gastric-derived cells with HMBA induced G0/G1 arrest and up-regulation of the proton pump, a marker of gastric cancer differentiation. Moreover, treatment with HMBA increased the expression and activity of GSK-3β in the nucleus but not the cytosol. HMBA decreased CDK2 activity and induced p27(Kip1) expression, which could be rescued by inhibition of GSK-3β. Furthermore, HMBA increased p27(Kip1) binding to CDK2, and this was abolished by GSK-3β inhibition.. The results presented herein suggest that GSK-3β functions by regulating p27(Kip1) assembly with CDK2, thereby playing a critical role in G0/G1 arrest associated with HMBA-induced gastric epithelial cell differentiation.

Topics: Acetamides; Aminophenols; Antineoplastic Agents; Biomarkers, Tumor; Cell Cycle; Cell Differentiation; Cell Line, Tumor; Cyclin-Dependent Kinase 2; Cyclin-Dependent Kinase Inhibitor p27; Gene Expression Regulation, Neoplastic; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; Humans; Intestinal Mucosa; Maleimides; Proton Pumps; Stomach Neoplasms

Cyclooxygenase-2 (COX-2) expression is a marker of poor prognosis in gastric cancer patients, and its inhibition suppresses gastric tumorigenesis in experimental animal models. The mechanism that leads to COX-2 overexpression in this tumor type is unknown. We have now shown that inhibition of phosphatidylinositol 3-kinase by LY294002 suppresses both basal and phorbol myristate acetate-induced COX-2 expression in TMK-1 and MKN-28 gastric cancer cells. Furthermore, inhibition of glycogen synthase kinase-3beta (GSK-3beta) by SB415286 induced expression of COX-2 mRNA and protein as well as the enzyme activity in the gastric cancer cells. The effect of SB415286 was confirmed by the use of two additional GSK-3beta inhibitors, lithium chloride and SB216763. SB415286 had a modest 1.6-fold stimulatory effect on a 2-kb COX-2 promoter reporter construct, but more importantly, it was shown to block the decay of COX-2 mRNA. In contrast to modulation of phosphatidylinositol 3-kinase/Akt/GSK-3beta pathway, inhibitors of mitogen-activated protein kinases (MEK 1/2, p38, JNK) or the mammalian target of rapamycin did not alter COX-2 expression in gastric cancer cells. Our data show that inhibition of GSK-3beta stimulates COX-2 expression in gastric cancer cells, which seems to be primarily facilitated via an increase in mRNA stability and to a lesser extent through enhanced transcription.

Topics: Aminophenols; Blotting, Northern; Blotting, Western; Cell Line, Tumor; Chromones; Cyclooxygenase 2; Dinoprostone; Enzyme Inhibitors; Gene Expression Regulation, Enzymologic; Gene Expression Regulation, Neoplastic; Genes, Reporter; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; Humans; Indoles; Lithium Chloride; Maleimides; Models, Biological; Models, Statistical; Morpholines; Phosphatidylinositol 3-Kinases; Phosphoinositide-3 Kinase Inhibitors; Proto-Oncogene Proteins c-akt; Reverse Transcriptase Polymerase Chain Reaction; RNA; RNA, Messenger; Signal Transduction; Stomach Neoplasms; Tetradecanoylphorbol Acetate; Time Factors; Transfection