sulindac and Carcinoma--Hepatocellular

Sulindac, a widely used non-steroidal anti-inflammatory drug (NSAID), has been shown to inhibit chemically induced carcinogenesis in animal models. In the present study, we have investigated the molecular mechanism by which sulindac affects the activity and expression of the enzymes that mediate the initial detoxification steps of many environmental carcinogens, the cytochromes P450 1A1, 1A2 and 1B1. Sulindac treatment of Sprague-Dawley rats resulted in a dose-dependent increase in hepatic cytochrome P450 (CYP) enzyme activity and in the expression of hepatic CYPs 1A1 and 1B1 mRNA. In the HepG2 human liver cancer cell line, sulindac caused a sustained, dose-dependent increase in CYP enzyme activity. Sulindac treatment resulted in a profound, dose-dependent increase in CYP 1A1 mRNA and a modest increase in 1A2 mRNA. The increase in CYP 1A1 mRNA induced by sulindac was, like enzyme activity, sustained for several days after the initial treatment. Sulindac induced the transcription of the CYP1A1 gene, as measured by the level of heterogeneous nuclear 1A1 RNA and by actinomycin D chase experiment. Since the transcription of CYP1A1 is under the control of the aryl hydrocarbon receptor (AhR), we examined the ability of sulindac to activate the receptor. Sulindac bound to the AhR, as measured by ligand-binding assay, and induced the binding of the AhR with the xenobiotic-responsive element present in the promoter region of the CYP1A1 gene. These results are the first demonstration that NSAIDs modulate carcinogen metabolic enzymes and provide a novel mechanism to explain the established chemopreventive activity of sulindac.

Topics: Animals; Antineoplastic Agents; Breast Neoplasms; Carcinoma, Hepatocellular; Cytochrome P-450 CYP1A1; Dactinomycin; Dose-Response Relationship, Drug; Gene Expression Regulation, Enzymologic; Humans; In Vitro Techniques; Liver; Liver Neoplasms; Male; Promoter Regions, Genetic; Protein Synthesis Inhibitors; Rats; Rats, Sprague-Dawley; Receptors, Aryl Hydrocarbon; RNA, Messenger; Sulindac; Transcription, Genetic; Tumor Cells, Cultured; Xenobiotics

Hepatoma is one of the most frequently occurring cancers worldwide. However, effective chemotherapeutic agents for this disease have not been developed. Acyclic retinoid, a novel synthetic retinoid, can reduce the incidence of postsurgical recurrence of hepatoma and improve the survival rate. OSI-461, a potent derivative of exisulind, can increase intracellular levels of cyclic GMP, which leads to activation of protein kinase G and induction of apoptosis in cancer cells. In the present study, we examined the combined effects of acyclic retinoid plus OSI-461 in the HepG2 human hepatoma cell line. We found that the combination of as little as 1.0 micromol/L acyclic retinoid and 0.01 micromol/L OSI-461 exerted synergistic inhibition of the growth of HepG2 cells. Combined treatment with low concentrations of these two agents also acted synergistically to induce apoptosis in HepG2 cells through induction of Bax and Apaf-1, reduction of Bcl-2 and Bcl-xL, and activation of caspase-3, -8, and -9. OSI-461 enhanced the G0-G1 arrest caused by acyclic retinoid, and the combination of these agents caused a synergistic decrease in the levels of expression of cyclin D1 protein and mRNA, inhibited cyclin D1 promoter activity, decreased the level of hyperphosphorylated forms of the Rb protein, induced increased cellular levels of the p21(CIP1) protein and mRNA, and stimulated p21(CIP1) promoter activity. Moreover, OSI-461 enhanced the ability of acyclic retinoid to induce increased cellular levels of retinoic acid receptor beta and to stimulate retinoic acid response element-chloramphenicol acetyltransferase activity. A hypothetical model involving concerted effects on p21(CIP1) and retinoic acid receptor beta expression is proposed to explain these synergistic effects. Our results suggest that the combination of acyclic retinoid plus OSI-461 might be an effective regimen for the chemoprevention and chemotherapy of human hepatoma and possibly other malignancies.

Topics: Antineoplastic Agents; Apoptosis; beta Catenin; Blotting, Western; Carcinoma, Hepatocellular; Cell Adhesion Molecules; Cell Cycle Proteins; Cell Line, Tumor; Cell Proliferation; Chloramphenicol O-Acetyltransferase; Cyclic GMP; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p21; Cytoskeletal Proteins; Dose-Response Relationship, Drug; Drug Synergism; G1 Phase; Gene Expression Regulation, Neoplastic; Humans; Intracellular Space; Liver Neoplasms; Microfilament Proteins; Models, Biological; Phosphoproteins; Phosphorylation; Promoter Regions, Genetic; Receptors, Retinoic Acid; Recombinant Fusion Proteins; Response Elements; Resting Phase, Cell Cycle; Retinoblastoma Protein; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger; Sulindac; Trans-Activators; Transfection; Tretinoin; Tumor Suppressor Protein p53

To evaluate the effects of sulindac in inducing growth inhibition and apoptosis of human gastric cancer cells in comparison with human hepatocellular carcinoma (HCC) cells.. The human gastric cancer cell lines MKN45 and MKN28 and human hepatocellular carcinoma cell lines HepG(2) and SMMC7721 were used for the study. Anti-proliferative effect was measured by MTT assay, and apoptosis was determined by Hoechst-33258 staining, electronography and DNA fragmentation. The protein of cyclooxygenase-2 (COX-2) and Bcl-2 were detected by Western dot blotting.. Sulindac could initiate growth inhibition and apoptosis of MKN45, MKN28, HepG(2) and SMMC7721 cells in a dose-and time-dependent manner. Growth inhibitory activity and apoptosis were more sensitive in HepG(2) cells than in SMMC7721 cells, MKN45 and MKN28 cells. After 24 hours incubation with sulindac at 2mmol x L(-1) and 4mmol x L(-1), the level of COX-2 and Bcl-2 protein were lowered in MKN45, SMMC7721 and HepG(2) cells but not in MKN28 cells.. Sulindac could inhibit the growth of gastric cancer cells and HCC cells effectively in vitro by apoptosis induction, which was associated with regression of COX-2 and Bcl-2 expression. The growth inhibition and apoptosis of HCC cells were greater than that of human gastric cancer cells. The different effects of apoptosis in gastric cancer cells may be related to the differentiation of the cells.

Topics: Anti-Inflammatory Agents, Non-Steroidal; Apoptosis; Carcinoma, Hepatocellular; Cell Division; Cells, Cultured; Humans; Liver Neoplasms; Stomach Neoplasms; Sulindac

This is the first report enumerating a superb antiproliferative effect of both sulindac and exisulind on hepatocellular cancer cell lines. The growth inhibition and cytotoxicity of sulindac in human hepatocellular carcinoma cell lines HepG2, Huh-7, and KYN-2 were investigated by studying cell growth, cell cycle distribution, and induction of apoptosis. In the presence of sulindac, there was a marked time- and dose-dependent decrease in cell proliferation and viability. Also, exisulind exhibited a similar growth-inhibitory effect on the KYN-2 cell line. The findings of this study suggest that sulindac exhibits a growth-inhibitory effect on human hepatocellular carcinoma cell lines; therefore, these drugs might serve as an effective tool for hepatocellular carcinoma chemoprevention.

Topics: Anti-Inflammatory Agents, Non-Steroidal; Antineoplastic Agents; Apoptosis; Carcinoma, Hepatocellular; Cell Division; Cyclooxygenase 2; DNA, Neoplasm; Dose-Response Relationship, Drug; Flow Cytometry; Humans; Interphase; Isoenzymes; Liver Neoplasms; Membrane Proteins; Necrosis; Prostaglandin-Endoperoxide Synthases; RNA, Messenger; Sulindac; Time Factors; Tumor Cells, Cultured