sulindac and Uterine-Cervical-Neoplasms

To observe whether cisplatin-induced apoptosis were increased when SiHa cells were preincubated with nuclear factor-kappa B (NF-kappaB) inhibitors [aspirin, sulindac, curcumin or pyrrolidine dithiocarbamate (PDTC)].. SiHa cells were preincubated 2 hours with aspirin, sulindac, curcumin and PDTC respectively, then a further incubation were done with cisplatin, and Western blot analysis were applied to detect P65 level of nuclear extraction. MTT assay was done to detect relative cell viability. TUNEL was applied to detect apoptosis rates. Flow cytometryies with PI staining were also used to detect apoptosis as well as cell cycle.. When SiHa cells were pretreated with aspirin, sulindac, curcumin or PDTC, Western blot showed that the expression of P65 was inhibited upon cisplatin stimulus (P < 0.05). MTT assay demonstrated that a preincubation with NF-kappaB inhibitor could signifianctly increase cisplatin-induced chemosensitivity (P < 0.05). When cells pretreated with aspirin, sulindac, curcumin, or PDTC, TUNEL and flow cytometries assay showed that the apoptotic rates were all increased after 24 hours cisplatin stimulus (P < 0.05). Results of flow cytometries were also showed that a pretreation with aspirin, sulindac, curcumin, or PDTC could significantly increase cisplatin-induced apoptosis.. Aspirin, sulindac, curcumin and PDTC could all inhibit cisplatin induced NF-kappaB activiation, which could increase cispaltin-induced chemosensativity by augments of apoptosis.

Topics: Apoptosis; Aspirin; Cell Cycle; Cell Line, Tumor; Cell Survival; Cisplatin; Curcumin; Female; Humans; NF-kappa B; Pyrrolidines; Sulindac; Thiocarbamates; Uterine Cervical Neoplasms

Epidemiologic and experimental studies have shown that cyclooxygenase-2 (COX-2) inhibitors as non-steroidal anti-inflammatory drugs (NSAIDs) are effective chemopreventive agents. The mechanisms underlying the antitumor activity of COX-2 inhibitors are thought to involve inhibition of COX-2 enzyme activity and induction of apoptosis. The aim of the current work was to study the mechanisms of synergistic action noted in HeLa cervical carcinoma cells under doxorubicin (DOX) and sulindac (SUL) co-treatment.. Cytotoxic activity of the drugs was defined with MTT test, apoptosis was detected with TUNEL test, DOX transmembrane efflux was measured fluorometrically, expression of MDR-1 and MRP-1 was determined with quantitative real time - PCR (QRT-PCR).. It was shown that SUL at non-toxic concentrations, 10 and 50 microM, is an effective enhancer of cytotoxic action for DOX in 0.5 and 1 microM, respectively; however, only for SUL concentration equal to 50 microM potentiated apoptosis induced by 1 microM of DOX. Moreover, blocking DOX efflux outside the cells was observed. The QRT - PCR analysis has shown that, when used simultaneously, DOX 1 microM and SUL 50 microM results in decreased mRNA level for MDR-1 and MRP-1.. It is concluded that cytotoxic action of DOX against HeLa cells is enhanced by non-toxic concentrations of SUL. The observed effect is due to quenching of MDR-1 and MRP-1 genes expression, which results in blocking of efflux of DOX outside the cells, which in turn correlates with enhanced apoptotic effects. According to obtained results the mechanisms of potentiating of DOX action by SUL are dose specific.

Topics: Apoptosis; ATP Binding Cassette Transporter, Subfamily B; ATP Binding Cassette Transporter, Subfamily B, Member 1; Cyclooxygenase Inhibitors; DNA Primers; Dose-Response Relationship, Drug; Doxorubicin; Drug Synergism; Female; Gene Expression Regulation, Neoplastic; HeLa Cells; Humans; In Situ Nick-End Labeling; Multidrug Resistance-Associated Proteins; Reverse Transcriptase Polymerase Chain Reaction; Sulindac; Tetrazolium Salts; Thiazoles; Uterine Cervical Neoplasms

Sulindac, a nonsteroidal anti-inflammatory drug (NSAID), induces growth arrest in HeLa cells and causes strong inhibition of the G1 to S transition of the cell cycle in a concentration-dependent manner. The G1 arrest is preceded by suppression of cyclin E and A, inactivation of cdk2, and the complete loss of the viral oncoprotein E7, despite ongoing HPV transcription. As shown by inhibitors specific for cyclooxygenase (COX) 1 and 2 loss of E7 is COX-independent. Moreover, inhibition of the proteasome activity with MG132 partially blocked the ability of sulindac to suppress E7 suggesting that sulindac induces degradation of E7 by the proteasomal pathway. In addition to inhibiting growth, sulindac strongly induces apoptosis, which can be abrogated by using the general caspase inhibitor zVAD-fmk. Unchanged expression of the pro-apoptotic protein Bax and suppression of the anti-apoptotic molecules Bcl-2 and Bcl-x(L) argues for the engagement of the mitochondrial apoptotic pathway. These results support the notion that sulindac is a potent growth inhibitor and inducer of apoptosis on cervical cancer cells in vitro and may offer new perspectives as a chemopreventive or supplementary anti-cervical cancer drug.

Topics: Anti-Inflammatory Agents, Non-Steroidal; Apoptosis; Blotting, Northern; Blotting, Western; Cell Cycle; Cyclins; Cyclooxygenase 2; Cyclooxygenase Inhibitors; Cysteine Proteinase Inhibitors; DNA-Binding Proteins; Dose-Response Relationship, Drug; Female; G1 Phase; Gene Expression Regulation, Viral; HeLa Cells; Humans; Leupeptins; Meloxicam; Oncogene Proteins, Viral; Proteasome Endopeptidase Complex; Proteasome Inhibitors; Proto-Oncogene Proteins c-bcl-2; Pyrazoles; S Phase; Sulfonamides; Sulindac; Thiazines; Thiazoles; Uterine Cervical Neoplasms

Transcription factor NF-kappaB plays a pivotal role in cancer cells in the resistance to apoptosis, since NF-kappaB is frequently activated in many primary carcinoma cells. Indeed, several NF-kappaB inhibitors are found to be promising anti-cancer agents. However, some anti-cancer agents activate NF-kappaB signals and may reduce their potential, including tumor necrosis factor (TNF)-alpha. Recently, the nonsteroidal anti-inflammatory drug (NSAID) sulindac and its metabolites have been shown to inhibit the NF-kappaB-mediated survival signals through inhibition of IKK-beta by their direct interaction. We thus investigate whether sulindac and its metabolite can augment TNF-alpha-mediated apoptosis in human carcinoma cells and be applicable for in vivo clinical usage. We here demonstrate that sulindac inhibited TNF-alpha-mediated NF-kappaB activation and greatly enhanced TNF-alpha-induced apoptosis in human gastric MKN45 and cervical HeLa carcinoma cell lines. The in vivo tumor growth of MKN45 cells was most strongly inhibited by a combination of TNF-alpha with sulindac compared with TNF-alpha or sulindac alone. Moreover, we demonstrate that sulindac sulfide further augmented TNF-alpha-mediated apoptosis. Our data strongly suggest that combination therapy of TNF-alpha with sulindac and its metabolites may sensitize cancer cells to TNF-alpha and augment its pro-apoptotic potential. Therefore, in combination with sulindac or its metabolites, TNF-alpha may become a potentially useful anti-cancer agent to suppress tumor.

Topics: Apoptosis; Cell Line, Tumor; Female; Humans; NF-kappa B; Sulindac; Tumor Necrosis Factor-alpha; Uterine Cervical Neoplasms