sulindac and Ovarian-Neoplasms

Epithelial ovarian cancer (EOC) remains the most lethal gynecologic malignancy in developed countries. Chronic endogenous sterile pro-inflammatory responses are strongly linked to EOC progression and chemoresistance to anti-cancer therapeutics. In the present study, the activity of epithelial NF-κB, a key pro-inflammatory transcription factor, was enhanced with the progress of EOC. This result was mechanistically linked with an increased expression of NSAID-Activated Gene 1 (NAG-1) in MyD88-positive type I EOC stem-like cells, compared with that in MyD88-negative type II EOC cells. Elevated NAG-1 as a potent biomarker of poor prognosis in the ovarian cancer was positively associated with the levels of NF-κB activation, chemokines and stemness markers in type I EOC cells. In terms of signal transduction, NAG-1-activated SMAD-linked and non-canonical TGFβ-activated kinase 1 (TAK-1)-activated pathways contributed to NF-κB activation and the subsequent induction of some chemokines and cancer stemness markers. In addition to effects on NF-κB-dependent gene regulation, NAG-1 was involved in expression of EGF receptor and subsequent activation of EGF receptor-linked signaling. The present study also provided evidences for links between NAG-1-linked signaling and chemoresistance in ovarian cancer cells. NAG-1 and pro-inflammatory NF-κB were positively associated with resistance to paclitaxel in MyD88-positive type I EOC cells. Mechanistically, this chemoresistance occurred due to enhanced activation of the SMAD-4- and non-SMAD-TAK-1-linked pathways. All of the present data suggested NAG-1 protein as a crucial mediator of EOC progression and resistance to the standard first-line chemotherapy against EOC, particularly in MyD88-positive ovarian cancer stem-like cells.

Topics: Anti-Inflammatory Agents, Non-Steroidal; Antineoplastic Agents, Phytogenic; Biomarkers, Tumor; Carcinogenesis; Carcinoma, Ovarian Epithelial; Cell Line, Tumor; Chemokines; Disease Progression; Disease-Free Survival; Drug Resistance, Neoplasm; ErbB Receptors; Female; Fluorescent Antibody Technique; Growth Differentiation Factor 15; Humans; Immunohistochemistry; Inflammation; Kaplan-Meier Estimate; MAP Kinase Kinase Kinases; Microscopy, Confocal; Myeloid Differentiation Factor 88; Neoplasm Staging; Neoplasms, Glandular and Epithelial; Neoplastic Stem Cells; NF-kappa B; Ovarian Neoplasms; Ovary; Paclitaxel; Prognosis; Signal Transduction; Smad4 Protein; Sulindac; Up-Regulation

Sulindac, a non-steroidal anti-inflammatory drug, suppresses carcinogenesis and inhibits growth of tumor cells. Pyrrolidine dithiocarbamate (PDTC), a potent NF-κB inhibitor, has been also identified as a potential anti-neoplastic agent. We hypothesized that combination of sulindac and PDTC could result in augmentation of cytotoxicity against ovarian cancer cells. The effect of sulindac and PDTC was examined on several ovarian cancer lines. Tumor cell viability was assessed using the MTT assay. Annexin-V/PI staining was used to detect apoptosis, cell cycle distribution was analyzed in FACS, and expression of cellular proteins was detected by western blotting. Incubation of OVA-14, OVP-10 and CAOV-1 ovarian cancer cells with sulindac and PDTC resulted in significantly greater inhibition of cell viability compared to either compound alone. In a model of OVA-14 cells it was evident that this effect was not related to the expression of COX enzymes since both active (sulindac sulfide) and inactive (sulindac) in vitro compounds affected the growth of tumor cells to a similar extent and synergized in cytotoxicity with PDTC. Combination of sulindac and PDTC lead to G0 arrest and massive apoptosis in co-treated cultures. Western blotting analysis argued for induction of the mitochondrial apoptotic pathway. These data demonstrate the synergistic cytotoxic effect of sulindac and PDTC on ovarian cancer cells through apoptosis and cell cycle arrest and prompt to test the efficacy of this combination in animal models.

Topics: Anti-Inflammatory Agents, Non-Steroidal; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Blotting, Western; Cell Cycle Checkpoints; Cell Line, Tumor; Cell Proliferation; Cell Survival; Dose-Response Relationship, Drug; Drug Synergism; Female; Flow Cytometry; Humans; NF-kappa B; Ovarian Neoplasms; Prostaglandin-Endoperoxide Synthases; Pyrrolidines; Resting Phase, Cell Cycle; Sulindac; Thiocarbamates; Time Factors

The purpose of our study was to assess susceptibility of cells of various ovarian cell lines on different nonsteroidal anti-inflammatory drugs (NSAIDs).. Cytotoxic effect of NSAIDs was tested using MTT colorimetric assay.. Amongst 6 NSAIDs tested: sulindac, sulindac sulfide, sulindac sulfone, acetylsalicylic acid, nimesulide, and rofecoxib, viability of ovarian carcinoma cells was compromised most strongly by sulindac and sulindac sulfide and concerned all the cell lines tested: SKOV-3, MDAH 2774, OVCA-1, and OVP-10. Sulindac sulfone and rofecoxib also displayed some cytotoxic effect during prolonged 72-hour incubation. Other NSAIDs tested: nimesulide and acetylsalicylic acid were devoid of cytotoxic effect on ovarian cancer cells.. Our results are encourage enough to conduct clinical trials that could allow to draw conclusions regarding potential application of sulindac in the adjuvant treatment of a standard chemotherapy of ovarian cancer.

Topics: Anti-Inflammatory Agents, Non-Steroidal; Antineoplastic Agents; Aspirin; Cell Cycle; Cell Size; Cell Survival; Colorimetry; Female; Humans; Lactones; Ovarian Neoplasms; Sulfonamides; Sulfones; Sulindac; Tumor Cells, Cultured

Several epidemiological studies have correlated the use of non-steroidal anti-inflammatory drugs (NSAID) with reduced risk of ovarian cancer, the most lethal gynecological cancer, diagnosed usually in late stages of the disease. We have previously established that the pro-apoptotic cytokine melanoma differentiation associated gene-7/Interleukin-24 (mda-7/IL-24) is a crucial mediator of NSAID-induced apoptosis in prostate, breast, renal and stomach cancer cells. In this report we evaluated various structurally different NSAIDs for their efficacies to induce apoptosis and mda-7/IL-24 expression in ovarian cancer cells. While several NSAIDs induced apoptosis, Sulindac Sulfide and Diclofenac most potently induced apoptosis and reduced tumor growth. A combination of these agents results in a synergistic effect. Furthermore, mda-7/IL-24 induction by NSAIDs is essential for programmed cell death, since inhibition of mda-7/IL-24 by small interfering RNA abrogates apoptosis. mda-7/IL-24 activation leads to upregulation of growth arrest and DNA damage inducible (GADD) 45 α and γ and JNK activation. The NF-κB family of transcription factors has been implicated in ovarian cancer development. We previously established NF-κB/IκB signaling as an essential step for cell survival in cancer cells and hypothesized that targeting NF-κB could potentiate NSAID-mediated apoptosis induction in ovarian cancer cells. Indeed, combining NSAID treatment with NF-κB inhibitors led to enhanced apoptosis induction. Our results indicate that inhibition of NF-κB in combination with activation of mda-7/IL-24 expression may lead to a new combinatorial therapy for ovarian cancer.

Topics: Animals; Anti-Inflammatory Agents, Non-Steroidal; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Blotting, Western; Cell Cycle Proteins; Cell Line, Tumor; Diclofenac; Drug Synergism; Enzyme Activation; Female; Gene Expression Regulation, Neoplastic; Humans; Interleukins; Intracellular Signaling Peptides and Proteins; JNK Mitogen-Activated Protein Kinases; Mice; Mice, SCID; NF-kappa B; Nuclear Proteins; Ovarian Neoplasms; Reverse Transcriptase Polymerase Chain Reaction; RNA Interference; Sesquiterpenes; Sulindac; Tumor Burden; Xenograft Model Antitumor Assays

Although the chemopreventive and antitumorigenic activities of nonsteroidal anti-inflammatory drug (NSAID) against colorectal cancer are well established, the molecular mechanisms responsible for these properties in ovarian cancer have not been elucidated. Therefore, there is an urgent need to develop mechanism-based approaches for the management of ovarian cancer. To this end, the effect of several NSAIDs on ovarian cancer cells was investigated as assessed by the induction of NAG-1/MIC-1/GDF-15, a proapoptotic gene belonging to the transforming growth factor-beta superfamily. Sulindac sulfide was the most significant NSAID activated gene 1 (NAG-1) inducer and its expression was inversely associated with cell viability as determined by 3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium assay. This growth suppression by sulindac sulfide was recovered by transfection of NAG-1 small interfering RNA. These results indicate that NAG-1 is one of the genes responsible for growth suppression by sulindac sulfide. Furthermore, we observed down-regulation of p21 WAF1/CIP1 by introduction of NAG-1 small interfering RNA into sulindac sulfide-treated cells. In addition, to elucidate other potential molecular mechanisms involved in sulindac sulfide treatment of ovarian cancer cells, we did a membrane-based microarray experiment. We found that cyclin D1, MMP-1, PI3KR1, and uPA were down-regulated by sulindac sulfide. In conclusion, a novel molecular mechanism is proposed to explain the experimental results and provide a rationale for the chemopreventive activity of NSAIDs in ovarian cancer.

Topics: Anti-Inflammatory Agents, Non-Steroidal; Apoptosis; Blotting, Western; Cell Cycle Proteins; Cell Line, Tumor; Cell Survival; Coloring Agents; Cyclin D1; Cyclin-Dependent Kinase Inhibitor p21; Cytokines; Down-Regulation; Female; Growth Differentiation Factor 15; Humans; Luciferases; Oligonucleotide Array Sequence Analysis; Ovarian Neoplasms; Promoter Regions, Genetic; Reverse Transcriptase Polymerase Chain Reaction; RNA, Small Interfering; Sulindac; Tetrazolium Salts; Thiazoles; Time Factors; Up-Regulation