gw9662 and Lung-Neoplasms

Peroxisome proliferator-activated receptor γ (PPARγ) agonists frequently induce cell death in human non-small-cell lung cancer (NSCLC) cells. However, majority of NSCLC patients acquire resistance after cancer therapy, and it is still unclear.. In this study we investigated the apoptotic mechanism and the anti-cancer effects of a novel purine-based PPARγ agonist, CB11 (8-(2-aminophenyl)-3-butyl-1,6,7-trimethyl-1H-imidazo[2,1-f]purine-2,4(3H,8H)-dione), on human NSCLC cells. CB11 mediates PPARγ-dependent cell death, reactive oxygen species (ROS) production, mitochondrial membrane potential (MMP) collapse, cell cycle arrest, lactate dehydrogenase (LDH) cytotoxicity, and caspase-3 activity in human NSCLC cells.. CB11 causes cell death via ROS-mediated ATM-p53-GADD45α signalling in human NSCLC cells, and diphenyleneiodonium (DPI), an NADPH oxidase inhibitor, decreases cell death by inhibiting CB11-mediated ATM signalling. In a xenograft experiment, CB11 dramatically reduced tumour volume when compared to a control group. Furthermore, CB11 induced cell death by inhibiting epithelial-to-mesenchymal transition (EMT) under radiation exposure in radiation-resistant human NSCLC cells. However, PPARγ deficiency inhibited cell death by blocking the ATM-p53 axis in radiation/CB11-induced radiation-resistant human NSCLC cells.. Taken together, our results suggest that CB11, a novel PPARγ agonist, may be a novel anti-cancer agent, and it could be useful in a therapeutic strategy to overcome radio-resistance in radiation-exposed NSCLC.

Topics: 3T3 Cells; Adipocytes; Anilides; Animals; Apoptosis; Ataxia Telangiectasia Mutated Proteins; Azo Compounds; Carcinoma, Non-Small-Cell Lung; Caspase 3; Caspase 9; Cell Cycle Checkpoints; Cell Cycle Proteins; Cell Death; Cell Differentiation; Cell Line, Tumor; DNA Damage; Epithelial-Mesenchymal Transition; Female; Humans; Imidazoles; L-Lactate Dehydrogenase; Ligands; Luciferases; Lung Neoplasms; Membrane Potential, Mitochondrial; Mice; Mice, Inbred BALB C; Mice, Nude; Onium Compounds; PPAR gamma; Purines; Radiation Tolerance; Reactive Oxygen Species; RNA, Small Interfering; Signal Transduction; Thiazolidinediones; Tumor Burden; Tumor Suppressor Protein p53

Statins (3-hydroxy-3-methylglutaryl coenzyme A [HMG-CoA] reductase inhibitors) are well-established agents to treat hyperlipidemic states. Experimental and epidemiological evidence further implies an anticancer effect of these substances. This study investigates the mechanism underlying human lung cancer cell death by lovastatin and the role of the prostaglandin (PG)-synthesizing enzyme cyclooxygenase-2 (COX-2) in this process. In A549 and H358 lung carcinoma cells the lipophilic prodrug lovastatin lactone led to a concentration-dependent decrease of viability and induction of DNA fragmentation, whereas its HMG-CoA-inhibitory, ring-open acid form was inactive in this respect. Apoptotic cell death by lovastatin was accompanied by high intracellular levels of the lactone form, by upregulation of COX-2 mRNA and protein, as well as by increased formation of peroxisome proliferator-activated receptor γ (PPARγ)-activating PGD2 and 15-deoxy-Δ12,14-PGJ2. Cells were significantly less sensitive to lovastatin-induced apoptotic cell death, when the expression or activity of COX-2 was suppressed by siRNA or by the COX-2 inhibitor NS-398. Apoptosis by lovastatin was likewise reversed by the PPARγ antagonist GW9662. Fluorescence microscopy analyses revealed a lovastatin-induced cytosol-to-nucleus translocation of PPARγ that was inhibited by NS-398. Collectively, this study demonstrates COX-2 induction and subsequent COX-2-dependent activation of PPARγ as a hitherto unknown mechanism by which lovastatin lactone induces human lung cancer cell death.

Topics: A549 Cells; Anilides; Apoptosis; Cell Line, Tumor; Cell Survival; Cyclooxygenase 2; Cyclooxygenase 2 Inhibitors; DNA Fragmentation; Humans; Hydroxymethylglutaryl-CoA Reductase Inhibitors; Lovastatin; Lung Neoplasms; Nitrobenzenes; PPAR gamma; Prostaglandin D2; RNA Interference; RNA, Messenger; RNA, Small Interfering; Sulfonamides

Our previous study demonstrated that quercetin-metabolite-enriched plasma (QP) but not quercetin itself upregulates peroxisome proliferator-activated receptor gamma (PPAR-γ) expression to induce G2/M arrest in A549 cells. In the present study, we incubated A549 cells with QP as well as quercetin-3-glucuronide (Q3G) and quercetin-3'-sulfate (Q3'S), two major metabolites of quercetin, to investigate the effects of quercetin metabolites on cell invasion and migration, the possible mechanisms and the role of PPAR-γ. We also compared the effects of QP with those of quercetin and troglitazone (TGZ), a PPAR-γ ligand. The results showed that QP significantly suppressed cell invasion and migration, as well as matrix metalloproteinases (MMPs)-2 activity and expression in a dose-dependent manner. The effects of 10% QP on those parameters were similar to those of 10μM quercetin and 20μM TGZ. However, QP and TGZ rather than quercetin itself increased the expressions of nm23-H1 and tissue inhibitor of metalloproteinase (TIMP-2). Furthermore, we demonstrated that Q3G and Q3'S also inhibited the protein expression of MMP-2. GW9662, a PPAR-γ antagonist, significantly diminished such an effect of Q3G and Q3'S. Silencing PPAR-γ expression in A549 cells also significantly diminished the suppression effect of Q3G and Q3'S on MMP-2 expression. Taken together, our study demonstrated that QP inhibited cell invasion and migration through nm23-H1/TIMP-2/MMP-2 associated mechanisms. The upregulation of PPAR-γ by quercetin metabolites such as Q3G and Q3'S could play an important role in the effects of QP.

Topics: A549 Cells; Anilides; Animals; Anticarcinogenic Agents; Cell Movement; Chromans; Dietary Supplements; Enzyme Repression; G2 Phase; Gerbillinae; Glucuronides; Humans; Ligands; Lung Neoplasms; Male; Matrix Metalloproteinase 2; Neoplasm Invasiveness; Neoplasm Proteins; PPAR gamma; Quercetin; RNA Interference; Thiazolidinediones; Troglitazone; Up-Regulation

The antitumorigenic mechanism of the selective cyclooxygenase-2 (COX-2) inhibitor celecoxib is still a matter of debate. Among different structurally related COX-2 inhibitors, only celecoxib was found to cause apoptosis and cell death of human lung cancer cells (IC₅₀ values of 19.96 µM [A549], 12.48 µM [H460], and 41.39 µM [H358]) that was paralleled by a time- and concentration-dependent upregulation of COX-2 and peroxisome proliferator-activated receptor γ (PPARγ) at mRNA and protein levels. Apoptotic death of celecoxib-treated cancer cells was suppressed by the PPARγ antagonist GW9662 and by siRNA targeting PPARγ and, surprisingly, also by the selective COX-2 inhibitor NS-398 and siRNA targeting COX-2. NS-398 (1 µM) was shown to suppress celecoxib-induced COX-2 activity. Among the COX-2-dependent prostaglandins (PG) induced upon celecoxib treatment, PGD₂ and 15-deoxy-Δ¹²,¹⁴-PGJ₂ were found to induce a cytosol-to-nucleus translocation of PPARγ as well as a PPARγ-dependent apoptosis. Celecoxib-elicited PPARγ translocation was inhibited by NS-398. Finally, a COX-2- and PPARγ-dependent cytotoxic action of celecoxib was proven for primary human lung tumor cells. Together, our data demonstrate a proapoptotic mechanism of celecoxib involving initial upregulation of COX-2 and PPARγ and a subsequent nuclear translocation of PPARγ by COX-2-dependent PGs.

Topics: Anilides; Antineoplastic Agents; Apoptosis; Carcinoma, Non-Small-Cell Lung; Celecoxib; Cell Line, Tumor; Cell Nucleus; Cyclooxygenase 2; Cyclooxygenase 2 Inhibitors; Enzyme Induction; Gene Expression Regulation, Neoplastic; Humans; Intracellular Space; Lung Neoplasms; Nitrobenzenes; PPAR gamma; Prostaglandins; Protein Transport; Pyrazoles; Sulfonamides

The antitumorigenic mechanism of cannabidiol is still controversial. This study investigates the role of COX-2 and PPAR-γ in cannabidiol's proapoptotic and tumor-regressive action. In lung cancer cell lines (A549, H460) and primary cells from a patient with lung cancer, cannabidiol elicited decreased viability associated with apoptosis. Apoptotic cell death by cannabidiol was suppressed by NS-398 (COX-2 inhibitor), GW9662 (PPAR-γ antagonist), and siRNA targeting COX-2 and PPAR-γ. Cannabidiol-induced apoptosis was paralleled by upregulation of COX-2 and PPAR-γ mRNA and protein expression with a maximum induction of COX-2 mRNA after 8 hours and continuous increases of PPAR-γ mRNA when compared with vehicle. In response to cannabidiol, tumor cell lines exhibited increased levels of COX-2-dependent prostaglandins (PG) among which PGD(2) and 15-deoxy-Δ(12,14)-PGJ(2) (15d-PGJ(2)) caused a translocation of PPAR-γ to the nucleus and induced a PPAR-γ-dependent apoptotic cell death. Moreover, in A549-xenografted nude mice, cannabidiol caused upregulation of COX-2 and PPAR-γ in tumor tissue and tumor regression that was reversible by GW9662. Together, our data show a novel proapoptotic mechanism of cannabidiol involving initial upregulation of COX-2 and PPAR-γ and a subsequent nuclear translocation of PPAR-γ by COX-2-dependent PGs.

Topics: Anilides; Animals; Antineoplastic Agents; Apoptosis; Cannabidiol; Cell Line, Tumor; Cell Nucleus; Cell Survival; Cyclooxygenase 2; Cyclooxygenase 2 Inhibitors; Female; Gene Expression; Gene Knockdown Techniques; Humans; Lung Neoplasms; Mice; Mice, Nude; Nitrobenzenes; PPAR gamma; Prostaglandins; RNA, Small Interfering; Sulfonamides; Tumor Burden; Xenograft Model Antitumor Assays

In this study, we incubated human A549 lung cancer cells with quercetin-metabolite-enriched plasma (QMP) obtained from Mongolian gerbils 2 h after quercetin feeding (100 mg/kg body wt/week). We investigated the effects of QMP on the growth of A549 cells and the possible mechanisms for these effects. We found that QMP but not control plasma (CP) reduced the cell growth in A549 cells. QMP led to cell cycle arrest at the G (2)/M phase by downregulating the expression of cdk1 and cyclin B. QMP but not CP or quercetin itself significantly increased PPAR- γ expression (p < 0.05), which was accompanied by an increase of phosphatase and tensin homologue deleted on the chromosome ten level and a decrease of phosphorylation of Akt. Furthermore, quercetin-3-glucuronide and quercetin-3'-sulfate also significantly increased PPAR- γ expression in A549 cells. GW9662, a PPAR- γ antagonist, significantly suppressed the effects of 10 % QMP on cell proliferation and on the expression of cyclin B and cdk1. Taken together, these data suggest that the activation of PPAR- γ plays an important role, at least in part, in the antiproliferative effects of quercetin metabolites.

Topics: Administration, Oral; Anilides; Animals; CDC2 Protein Kinase; Cell Division; Cell Line, Tumor; Cell Proliferation; Cyclin B; G2 Phase; Gerbillinae; Humans; Lung Neoplasms; Male; Oncogene Protein v-akt; Phosphorylation; Plasma; PPAR gamma; PTEN Phosphohydrolase; Quercetin; Up-Regulation

Emerging preclinical data suggests that tea possess anticarcinogenic and antimutagenic properties. We therefore hypothesize that white tea extract (WTE) is capable of favorably modulating apoptosis, a mechanism associated with lung tumorigenesis. We examined the effects of physiologically relevant doses of WTE on the induction of apoptosis in non-small cell lung cancer cell lines A549 (adenocarcinoma) and H520 (squamous cell carcinoma) cells. We further characterized the molecular mechanisms responsible for WTE-induced apoptosis, including the induction of peroxisome proliferator-activated receptor-gamma (PPAR-gamma) and the 15-lipoxygenase (15-LOX) signaling pathways. We found that WTE was effective in inducing apoptosis in both A549 and H520 cells, and inhibition of PPAR-gamma with GW9662 partially reversed WTE-induced apoptosis. We further show that WTE increased PPAR-gamma activation and mRNA expression, concomitantly increased 15(S)-hydroxy-eicosatetraenoic acid release, and upregulated 15-LOX-1 and 15-LOX-2 mRNA expression by A549 cells. Inhibition of 15-LOX with nordihydroguaiaretic acid (NGDA), as well as caffeic acid, abrogated WTE-induced PPAR-gamma activation and upregulation of PPAR-gamma mRNA expression in A549 cells. WTE also induced cyclin-dependent kinase inhibitor 1A mRNA expression and activated caspase-3. Inhibition of caspase-3 abrogated WTE-induced apoptosis. Our findings indicate that WTE is capable of inducing apoptosis in non-small cell lung cancer cell lines. The induction of apoptosis seems to be mediated, in part, through the upregulation of the PPAR-gamma and 15-LOX signaling pathways, with enhanced activation of caspase-3. Our findings support the future investigation of WTE as an antineoplastic and chemopreventive agent for lung cancer.

Topics: Anilides; Apoptosis; Arachidonate 15-Lipoxygenase; Carcinoma, Non-Small-Cell Lung; Catechin; Drug Evaluation, Preclinical; Gene Expression Regulation, Enzymologic; Gene Expression Regulation, Neoplastic; Hormone Antagonists; Humans; Hydroxyeicosatetraenoic Acids; Lung Neoplasms; Plant Extracts; PPAR gamma; Tea; Tumor Cells, Cultured

Thiazolidinediones (TZDs) are synthetic peroxisome proliferator-activated receptor-gamma (PPARgamma) ligands that are widely used in type II diabetes treatment. In addition to their ability to improve glucose homeostasis, TZDs possess anti-inflammatory properties and inhibit growth of many cells, particularly cancerous airway epithelial cells. However, the functional effects of PPARgamma ligands on nonmalignant human bronchial epithelial cells have never been investigated. In the present study, we questioned whether PPARgamma ligands may regulate proliferation of human bronchial epithelial cells, and we studied their potential molecular mechanisms. We found that synthetic PPARgamma agonists, rosiglitazone (RGZ) and troglitazone (TGZ), induced proliferation of human bronchial epithelial cells, whereas the endogenous PPARgamma ligand, 15-deoxy-Delta(12,14)-prostaglandin J(2) (15d-PGJ(2)), inhibited cell growth. RGZ and TGZ (10 microM) induced a rapid and transient intracellular Ca(2+) mobilization from thapsigargin-sensitive intracellular stores, whereas 15d-PGJ(2) (5 microM) did not induce any Ca(2+) signal. The PPARgamma antagonist GW-9662 did not inhibit any biological responses, but it reversed the effect of 15d-PGJ(2) on cell growth. Using RT-PCR, we detected mRNA expression of the GPR40 receptor, a G protein-coupled receptor recently identified as a receptor for free fatty acids and TZDs, in human bronchial epithelial cells. Downregulation of GPR40 by small-interfering RNA led to a significant inhibition of TZD-induced Ca(2+) mobilization and proliferation. This study provides evidence for the proliferative effect of anti-diabetic drug TZDs in nonmalignant human bronchial epithelial cells through GPR40 receptor activation, involving an intracellular Ca(2+) signaling pathway.

Topics: Adenocarcinoma; Anilides; Bronchi; Calcium Signaling; Cell Division; Cell Line, Transformed; Cell Line, Tumor; Chromans; Humans; Hypoglycemic Agents; Lung Neoplasms; PPAR gamma; Receptors, G-Protein-Coupled; Respiratory Mucosa; RNA, Small Interfering; Rosiglitazone; Thiazolidinediones; Troglitazone

Activation of peroxisome proliferator-activated receptors gamma (PPARgamma) induces diverse effects on cancer cells. The thiazolidinediones (TZDs), such as troglitazone and ciglitazone, are PPARgamma agonists exhibiting antitumor activities; however, the underlying mechanism remains inconclusive. Rosiglitazone (RGZ), a synthetic ligand of PPARgamma used in the treatment of Type 2 diabetes, inhibits growth of some tumor cells and is involved in other processes related to cancer progression. Opposing results have also been reported with different ligands on tumor cells. The purpose of this study was to determine if RGZ and 15d-PGJ2 induce antitumor effects in vivo and in vitro on the murine mammary tumor cell line LMM3.. The effect on LMM3 cell viability and nitric oxide (NO) production of different doses of RGZ, 15-dPGJ2, BADGE and GW9662 were determined using the MTS colorimetric assay and the Griess reaction respectively. In vivo effect of orally administration of RGZ on tumor progression was evaluated either on s.c. primary tumors as well as on experimental metastasis. Cell adhesion, migration (wound assay) and invasion in Transwells were performed. Metalloproteinase activity (MMP) was determined by zymography in conditioned media from RGZ treated tumor cells. PPARgamma expression was detected by inmunohistochemistry in formalin fixed tumors and by western blot in tumor cell lysates.. RGZ orally administered to tumor-bearing mice decreased the number of experimental lung metastases without affecting primary s.c. tumor growth. Tumor cell adhesion and migration, as well as metalloproteinase MMP-9 activity, decreased in the presence of 1 muM RGZ (non-cytotoxic dose). RGZ induced PPARgamma protein expression in LMM3 tumors. Although metabolic activity -measured by MTS assay- diminished with 1-100 microM RGZ, 1 microM-treated cells recovered their proliferating capacity while 100 microM treated cells died. The PPARgamma antagonist Biphenol A diglicydyl ether (BADGE) did not affect RGZ activity. On the contrary, the specific antagonist GW9662 completely abrogated RGZ-induced decrease in cell viability. A decrease in NO levels was detected in the presence of either 1 or 100 microM RGZ. The natural ligand 15d-PGJ2 did not affect metabolic activity although it induced a significant decrease in NO production.. A significant decrease in the number of experimental LMM3 lung metastasis, but not on primary tumor growth, after oral RGZ administration was observed. In vitro, 100 microMRGZ also reduced cell viability and NO production, while no changes were observed in the presence of 15d-PGJ2. BADGE did not reverse RGZ effect while the antagonist GW9662 completely abrogated it, suggesting a PPARgamma- dependent mechanism. Inhibition of lung metastatic nodules by RGZ administered in vivo, might be associated with the observed decrease in MMP-9 expression, in cell adhesion, migration and invasion. RGZ augmented its expression. PPARgamma was detected in cell lysates by western blot and by immunohistochemistry in tumors from RGZ-treated mice. In summary we can suggest that RGZ or any other TZDs might be possible future approaches in the treatment of metastasis of PPARgamma-expressing cells.

Topics: Administration, Oral; Anilides; Animals; Cell Adhesion; Cell Movement; Dose-Response Relationship, Drug; Lung Neoplasms; Mammary Neoplasms, Experimental; Matrix Metalloproteinases; Mice; Mice, Inbred BALB C; Neoplasm Invasiveness; Nitric Oxide; PPAR gamma; Rosiglitazone; Thiazolidinediones; Tumor Cells, Cultured

Peroxisome proliferator-activated receptor gamma (PPARgamma) plays a critical albeit poorly defined role in the development and progression of several cancer types including those of the breast, colon, and lung. A PPAR response element (PPRE) reporter assay was utilized to evaluate the selective transactivation of PPARgamma in 10 different cell lines including normal mammary epithelial, breast, lung, and colon cancer cells. Cells were treated with one of four compounds including rosglitizone (Ros), ciglitizone (Cig), 15-deoxy-Delta(12,14)-prostaglandin J2 (PGJ2), or GW 9662 (GW). We observed differences in transactivation between cell lines from different tissue origin, across cell lines from a single tissue type, and selective modulation of PPARgamma within a single cell line by different ligands. Interestingly, GW, a PPARgamma antagonist in adipocytes, enhanced PPRE reporter activation in normal mammary epithelial cells while it had virtually no effect in any of the cancer cell lines tested. Within each cancer type, individual cell lines were found to respond differently to distinct PPARgamma ligands. For instance, Ros, Cig, and PGJ2 were all potent agonist of PPARgamma transactivation in lung adenocarcinoma cell lines while these same ligands had no effect in squamous cell or large cell carcinomas of the lung. Message levels of PPARgamma and retinoid X receptor alpha (RXRalpha) in the individual cell lines were quantitated by real time-polymerase chain reaction (RT-PCR). The ratio of PPARgamma to RXRalpha was predictive of how cells responded to co-treatment of Ros and 9-cis-retinoic acid, an RXRalpha agonist, in two out of three cell lines tested. These data indicate that PPARgamma can be selectively modulated and suggests that it may be used as a therapeutic target for individual tumors.

Topics: Alitretinoin; Anilides; Breast Neoplasms; Caco-2 Cells; Cell Line, Tumor; Colonic Neoplasms; Female; Gene Expression Regulation, Neoplastic; Genes, Reporter; HT29 Cells; Humans; Ligands; Lung Neoplasms; PPAR gamma; Prostaglandin D2; Retinoid X Receptor alpha; RNA, Messenger; Rosiglitazone; Thiazolidinediones; Transfection; Tretinoin