RESUMO
Pristimerin (PM) is a quinonemethide triterpenoid with cytotoxic activity against a wide range of cancer cell lines. However, the effect of PM on IL-2 induced activation of T lymphocytes, which play a major role in antitumor immunity has not been studied. The objective of the present study was to evaluate the effect of PM on IL-2 induced proliferation of T cells, generation of lymphokine activated killer cells (LAK cells) and the signaling pathways involved in activation of T cells by IL-2. PM inhibited the IL-2 induced proliferation of mouse splenic T cells and the generation LAK cells at very low concentrations. The suppression of T cell proliferation by PM was associated with the inhibition of IL-2 induced Janus kinase/signal transducers and activators of transcription (Jak/STAT) and extracellular signal-regulated kinase 1 and 2 (Erk1/2) signaling pathways. PM also inhibited the proliferation and differentiation-related immediate early gene products such as p-c-fos, p-c-jun, c-myc and cyclin D1. In addition, antiapoptotic (prosurvival) NF-кB, p-Akt and p-mTOR were also inhibited by PM. These data demonstrated that PM inhibits IL-2 induced T cell activation and generation of LAK cells by disrupting multiple cell signaling pathways induced by IL-2.
Assuntos
Antineoplásicos/farmacologia , Interleucina-2/farmacologia , Ativação Linfocitária/efeitos dos fármacos , Baço/efeitos dos fármacos , Linfócitos T/efeitos dos fármacos , Triterpenos/farmacologia , Animais , Proteínas Reguladoras de Apoptose/metabolismo , Proteínas de Ciclo Celular/metabolismo , Diferenciação Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Relação Dose-Resposta a Droga , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Janus Quinases/metabolismo , Células Matadoras Ativadas por Linfocina/efeitos dos fármacos , Células Matadoras Ativadas por Linfocina/imunologia , Células Matadoras Ativadas por Linfocina/metabolismo , Camundongos , Triterpenos Pentacíclicos , Fosforilação , Fatores de Transcrição STAT/metabolismo , Transdução de Sinais/efeitos dos fármacos , Baço/imunologia , Baço/metabolismo , Linfócitos T/imunologia , Linfócitos T/metabolismoRESUMO
Trichothecenes are powerful mycotoxins that inhibit protein synthesis and induce ribotoxic stress response in mammalian cells. Verrucarin A (VC-A) is a Type D macrocyclic mycotoxin which inhibits cell proliferation and induces apoptosis in cancer cells. However, the antitumor activity of VC-A for prostate cancer cells has not been investigated. The objective of the present study was to determine the anticancer activity and its mechanism of action in hormone-responsive (LNCaP) and hormone-refractory (PC-3) carcinoma of the prostate (CaP) cell lines. VC-A strongly inhibited the proliferation and induced cell cycle arrest in G2/M phase associated with the inhibition of cell cycle regulatory proteins cyclin D, cyclin E, cyclin-dependent kinases (cdks) cdk2, cdk4, cdk6 and cdk inhibitors WAF1/21 and KIP1/27. VC-A also induced apoptosis in CaP cells as characterized by the cleavage of poly (ADP-ribose) polymerase (PARP-1), procaspases-3, -8 and -9 and the inhibition of Bcl-2 family proteins that regulate apoptosis (Bcl-2, Bcl-xL, Bax, Bak and Bad). In addition, VC-A also down-regulated the expression of prosurvival phospho-AKT (p-AKT), nuclear factor kappa B (NF-kB) (p65) and phospho-mammalian target of rapamycin (p-mTOR) signaling proteins. Taken together, these results demonstrated strong antiproliferative and apoptosis-inducing activity of verrucarin A against CaP cells through cell cycle arrest and inhibition of the prosurvival (antiapoptotic) AKT/NF-kB/mTOR signaling pathway.
Assuntos
Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , NF-kappa B/metabolismo , Neoplasias da Próstata/tratamento farmacológico , Proteínas Proto-Oncogênicas c-akt/metabolismo , Serina-Treonina Quinases TOR/metabolismo , Tricotecenos/farmacologia , Proteínas Reguladoras de Apoptose/metabolismo , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Regulação para Baixo/efeitos dos fármacos , Pontos de Checagem da Fase G2 do Ciclo Celular/efeitos dos fármacos , Humanos , Masculino , Micotoxinas/farmacologia , Próstata/efeitos dos fármacos , Próstata/metabolismo , Neoplasias da Próstata/metabolismo , Transdução de Sinais/efeitos dos fármacosRESUMO
Pristimerin (PM) is a quinonemethide triterpenoid present in various plant species with strong antiprolifertive and proapoptotic activities in cancer cells. The effect of PM on telomerase which is reactivated in most cancers including carcinoma of the prostate (CaP) has not been studied. We investigated the effect of PM on the expression of human telomerase reverse transcriptase (hTERT) gene that codes for the catalytic subunit of the telomerase holoenzyme complex in prostate cancer cell lines LNCaP and PC-3 cells. The inhibition of cell proliferation and induction of apoptosis by PM in both cell lines was associated with the inhibition of hTERT mRNA expression, suppression of native and phosphorylated hTERT protein and hTERT telomerase activity. The ablation of hTERT expression increased the sensitivity of cancer cells to PM. In addition, results also revealed that the inhibition of hTERT expression is attributed to the inhibition of transcription factors SP1, c-Myc and STAT3 and protein kinase B/Akt which regulate hTERT transcriptionally and post-translationally, respectively. These data provide evidence that telomerase is a potential target of PM in prostate cancer.
Assuntos
Antineoplásicos/farmacologia , Inibidores Enzimáticos/farmacologia , Neoplasias da Próstata/enzimologia , Telomerase/antagonistas & inibidores , Triterpenos/farmacologia , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Masculino , Triterpenos Pentacíclicos , Fosforilação , Neoplasias da Próstata/genética , Neoplasias da Próstata/patologia , Interferência de RNA , RNA Mensageiro/metabolismo , Transdução de Sinais/efeitos dos fármacos , Telomerase/genética , Telomerase/metabolismo , Transcrição Gênica/efeitos dos fármacos , TransfecçãoRESUMO
Pristimerin isaquinonemethidetriterpenoidthathasshown anticancer activity against some cancer types. However, the antitumor effects of pristimerin (PM) in ovarian cancer cells have not been adequately studied. The objective of the present study was to determine the anticancer activity and its mechanism of action in human ovarian carcinoma cell lines. PM strongly inhibited the proliferation of ovarian cancer cells by inducing apoptosis characterized by increased annexin V-binding, cleavage of poly (ADP-ribose) polymerase (PARP-1) and procaspases-3, -8 and -9. Furthermore, PM caused mitochondrial depolarization. Western blot analysis showed inhibition of prosurvival phospho-AKT (p-AKT), nuclear factor kappa B (NF-κB) (p65) and phospho-mammalian target of rapamycin (p-mTOR) signaling proteins in cells treated with PM. Treatment with PM also inhibited the expression of NF-κB-regulated antiapoptotic Bcl-2, Bcl-xL, c-IAP1 and survivin. Thus, our data showing potent antiproliferative and apoptosis-inducing activity of PM in ovarian carcinoma cells through the inhibition of AKT/ NF-κB/ mTOR signaling pathway warrant further investigation of PM for the management of ovarian cancer.
Assuntos
Antineoplásicos/farmacologia , NF-kappa B/antagonistas & inibidores , Neoplasias Ovarianas/tratamento farmacológico , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Transdução de Sinais/efeitos dos fármacos , Serina-Treonina Quinases TOR/antagonistas & inibidores , Triterpenos/farmacologia , Apoptose/efeitos dos fármacos , Carcinoma/tratamento farmacológico , Carcinoma/metabolismo , Carcinoma/patologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Feminino , Humanos , Mitocôndrias/efeitos dos fármacos , NF-kappa B/metabolismo , Neoplasias Ovarianas/metabolismo , Neoplasias Ovarianas/patologia , Triterpenos Pentacíclicos , Proteínas Proto-Oncogênicas c-akt/metabolismo , Serina-Treonina Quinases TOR/metabolismoRESUMO
Methyl-2-cyano-3,12-dioxooleana-1,9(11)-dien-28-oate (CDDO-Me) is a synthetic derivative of oleanolic acid, a triterpene, with apoptosis-inducing activity in a wide range of cancer cells. Induction of apoptosis by CDDO-Me is associated with the generation of reactive oxygen species (ROS) and inhibition of telomerase activity. In the present study, we investigated the role of ROS in inhibition of telomerase by CDDO-me. Treatment of MiaPaCa-2 and Panc-1 pancreatic cancer cell lines with CDDO-Me induced the production of hydrogen peroxide and superoxide anions and inhibited the telomerase activity. Pretreatment of cells with N-acetylcycsteine, a general purpose antioxidant or overexpression of glutathione peroxidase (GPx) or superoxide dismutase-1 (SOD-1) blocked the telomerase inhibitory activity of CDDO-Me. Furthermore, blocking ROS generation also prevented the inhibition of hTERT gene expression, hTERT protein production and expression of a number of hTERT-regulatory proteins by CDDO-Me (e.g., c-Myc, Sp1, NF-κB and p-Akt). Data also showed that Akt plays an important role in the activation of telomerase activity. Together, these data suggest that inhibition of telomerase activity by CDDO-Me is mediated through a ROS-dependent mechanism; however, more work is needed to fully understand the role of ROS in down-regulation of hTERT gene and hTERT-regulatory proteins by CDDO-Me.
Assuntos
Ácido Oleanólico/análogos & derivados , Neoplasias Pancreáticas/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Telomerase/antagonistas & inibidores , Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Ácido Oleanólico/farmacologia , Neoplasias Pancreáticas/genética , Proteínas Proto-Oncogênicas c-akt/metabolismo , Telomerase/genética , Telomerase/metabolismoRESUMO
Our previous studies have shown that methyl-2-cyano-3,12-dioxooleana-1,9(11)-dien-28-oate (CDDO-Me), a oleanane synthetic triterpenoid induces apoptosis in prostate cancer cells by inhibiting the Akt/NF-κB/mTOR signaling cascade; however, the mechanism by which CDDO-Me inhibits Akt/NF-κB/mTOR signaling has remained undetermined. Present studies show that Akt plays a critical role in the response of prostate cancer cells to CDDO-Me. Silencing of Akt sensitized PC-3 cells to CDDO-Me, whereas its overexpression rendered them resistant to CDDO-Me. Evaluation of the effect of CDDO-Me on Akt which lies upstream of NF-κB and mTOR showed that CDDO-Me directly inhibits the Akt kinase activity in cell-free kinase activity assay and in vivo without modulating the activity of PDK1, the upstream kinase that phosphorylates and activates Akt. The inhibition of Akt activity resulted in inhibition of phosphorylation/inactivation of proapoptotic procaspase-9, Bad and Foxo3a. Further, inhibition of p-Akt by CDDO-Me was not attributable to an increase in the activity of protein phosphatase 2A (PP2A) or PH domain/leucine-rich repeat protein phosphatase1 (PHLPP1) both of which dephosphorylate p-Akt. These findings show that Akt is a direct target of CDDO-Me in the Akt/NF-κB/mTOR prosurvival signaling axis.
Assuntos
Antineoplásicos/farmacologia , Ácido Oleanólico/análogos & derivados , Neoplasias da Próstata/enzimologia , Proteína Fosfatase 2/metabolismo , Proteínas Serina-Treonina Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Linhagem Celular Tumoral , Humanos , Masculino , NF-kappa B/metabolismo , Ácido Oleanólico/farmacologia , Piruvato Desidrogenase Quinase de Transferência de Acetil , Serina-Treonina Quinases TOR/metabolismoRESUMO
Methyl-2-cyano-3,12-dioxooleana-1,9(11)-dien-28-oate (CDDO-Me) is a multifunctional oleanane synthetic triterpenoid with potent anti-inflammatory and antitumorigenic properties. The mechanisms of the antisurvival and apoptosis-inducing activities of CDDO-Me and related derivatives of oleanolic acid have been defined; however, to date, no study has been carried out on the effect of CDDOs on human telomerase reverse transcriptase (hTERT) gene or telomerase activity. Here we report for the first time that inhibition of cell proliferation and induction of apoptosis by CDDO-Me in pancreatic cancer cell lines is associated with the inhibition of hTERT gene expression, hTERT telomerase activity and a number of proteins that regulate hTERT expression and activity. Furthermore, abrogation or overexpression of hTERT protein altered the susceptibility of tumor cells to CDDO-Me. These findings suggest that telomerase (hTERT) is a relevant target of CDDO-Me in pancreatic cancer cells.
Assuntos
Antineoplásicos/farmacologia , Proliferação de Células/efeitos dos fármacos , Regulação Enzimológica da Expressão Gênica/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Ácido Oleanólico/análogos & derivados , Neoplasias Pancreáticas/enzimologia , Supressão Genética , Telomerase/antagonistas & inibidores , Linhagem Celular Tumoral , Humanos , Ácido Oleanólico/farmacologia , Neoplasias Pancreáticas/genética , Telomerase/genéticaRESUMO
Oleanolic acid-derived synthetic triterpenoids are broad spectrum antiproliferative and antitumorigenic agents. In this study, we investigated the role of reactive oxygen species (ROS) in induction of apoptosis and inhibition of prosurvival Akt, NF-kappaB and mTOR signaling pro-teins by methyl-2-cyano-3,12-dioxooleana-1,9(11)-dien-28-oate (CDDO-Me) in pancreatic cancer cells. Micromolar concentrations of CDDO-Me inhibited proliferation and induced apoptosis in MiaPaCa-2 and Panc-1 pancreatic cancer cells. Treatment with CDDO-Me caused the generation of hydrogen peroxide and superoxide anion and pretreatment of cells with NADPH oxidase inhibitor diphylene iodonium (DPI) or respiratory chain complex 1 inhibitor rotenone prevented ROS generation. Pretreatment with N-acetylcysteine (NAC) or overexpression of glutathione peroxidase (GPx) or superoxide dismutase-1 (SOD-1) blocked the antiproliferative effects of CDDO-Me. Likewise, NAC prevented the induction of apoptosis (annexin V-FITC binding and cleavage of PARP-1 and procaspases-3,-8 and -9) and reversed the loss of mitochondrial membrane potential and release of cytochrome c from mitochondria by CDDO-Me. CDDO-Me down-regulated p-Akt, p-mTOR and NF-kappaB (p65) but increased the activation of Erk1/2 and NAC blocked the modulation of these cell signaling proteins by CDDO-Me. Thus, the results of this study indicate that the antiproliferative and apoptosis inducing effects of CDDO-Me are mediated through a ROS-dependent mechanism and the role of ROS in modulation of signaling proteins by CDDO-Me warrants further investigation.
Assuntos
Acetilcisteína/farmacologia , Ácido Oleanólico/análogos & derivados , Neoplasias Pancreáticas/metabolismo , Espécies Reativas de Oxigênio , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Glutationa Peroxidase/metabolismo , Humanos , Peróxido de Hidrogênio/metabolismo , Potencial da Membrana Mitocondrial , Ácido Oleanólico/administração & dosagem , Espécies Reativas de Oxigênio/antagonistas & inibidores , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Superóxido Dismutase/metabolismo , Superóxido Dismutase-1 , Superóxidos/metabolismoRESUMO
Xanthohumol (XN), a prenylated chalcone present in hops exhibits anti-inflammatory, antioxidant and anticancer activity. In the present study we show that XN inhibits the proliferation of mouse lymphoma cells and IL-2 induced proliferation and cell cycle progression in mouse splenic T cells. The suppression of T cell proliferation by XN was due to the inhibition of IL-2 induced Janus kinase/signal transducers and activators of transcription (Jak/STAT) and extracellular signal-regulated kinase 1 and 2 (Erk1/2) signaling pathways. XN also inhibited proliferation-related cellular proteins such as c-Myc, c-Fos and NF-kappaB and cyclin D1. Thus, understanding of IL-2 induced cell signaling pathways in normal T cells, which are constitutively turned on in T cell lymphomas may facilitate development of XN for the treatment of hematologic cancers.
Assuntos
Proliferação de Células , Flavonoides/farmacologia , Interleucina-2/metabolismo , Propiofenonas/farmacologia , Linfócitos T , Animais , Proteínas de Ciclo Celular/efeitos dos fármacos , Proteínas de Ciclo Celular/metabolismo , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Janus Quinases/metabolismo , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Camundongos , Camundongos Endogâmicos C57BL , Baço/citologia , Baço/metabolismo , Linfócitos T/efeitos dos fármacos , Linfócitos T/metabolismoRESUMO
Methyl-2-cyano-3,12-dioxooleana-1,9(11)-dien-28-oate (CDDO-Me) is an synthetic oleanane triterpenoid with strong antiprolifertive and proapoptotic activities in cancer cells. However, the effect of CDDO-Me on human telomerase reverse transcriptase (hTERT) and its telomerase activity in prostate cancer cells has not been studied. We investigated the role of hTERT in mediating the anticancer activity of CDDO-Me in prostate cancer cells in vitro and in vivo. The inhibition of cell proliferation and induction of apoptosis by CDDO-Me in LNCaP and PC-3 prostate cancer cell lines was associated with the inhibition of hTERT gene expression, hTERT telomerase activity and a number of proteins that regulate hTERT transcriptionally and post-translationally. Furthermore, ablation of hTERT protein increased the sensitivity of cancer cells to CDDO-Me, whereas its overexpression rendered them resistant to CDDO-Me. In addition, inhibition of progression of preneoplastic lesions (i.e., low and high-grade prostate intraepithelial neoplasms, PINs) to adenocarcinoma of the prostate by CDDO-Me in TRAMP mice was associated with significant decrease in TERT and its regulatory proteins in the prostate gland. These data provide evidence that telomerase is a potential target of CDDO-Me for the prevention and treatment of prostate cancer.
Assuntos
Ácido Oleanólico/análogos & derivados , Telomerase/antagonistas & inibidores , Telomerase/metabolismo , Animais , Apoptose/efeitos dos fármacos , Western Blotting , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Regulação da Expressão Gênica , Humanos , Masculino , Camundongos , NF-kappa B/antagonistas & inibidores , NF-kappa B/metabolismo , Ácido Oleanólico/farmacologia , Neoplasias da Próstata/tratamento farmacológico , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-akt/metabolismo , Fator de Transcrição STAT3/antagonistas & inibidores , Fator de Transcrição STAT3/metabolismoRESUMO
In an extension of our previous studies showing potent antitumorigenic activity of synthetic triterpenoids of oleanolic acid against prostate cancer cell lines, we examined the efficacy of 2-cyano-3,12-dioxooleana-1,9(11)-dien-28-oic acid (CDDO) in preventing the development and/or progression of prostate cancer in the transgenic adenocarcinoma of the mouse prostate (TRAMP) model. Data show that oral gavage with CDDO (10 µmol/kg) for 20 weeks resulted in inhibition of the progression of preneoplastic lesions in the dorsolateral prostate and ventral prostate to adenocarcinoma without toxicity. CDDO also inhibited metastasis of tumor to the distant organs. Treatment with CDDO significantly inhibited cell proliferation, reduced the density of blood vessels and promoted apoptosis in the prostatic tissue. Further, Akt, NF-κB and NF-κB regulated Bcl-2, Bcl-xL, survivin and cIAP1 appear to be the molecular targets of CDDO for inhibiting the progression of prostate cancer in TRAMP mice. Thus, these studies show for the first time the potential of CDDO for chemoprevention of human prostate cancer.
Assuntos
Apoptose/efeitos dos fármacos , Neoplasias Renais/tratamento farmacológico , Neoplasias Hepáticas/tratamento farmacológico , Neoplasias Pulmonares/tratamento farmacológico , Ácido Oleanólico/análogos & derivados , Neoplasias da Próstata/tratamento farmacológico , Transdução de Sinais/efeitos dos fármacos , Adenocarcinoma/tratamento farmacológico , Adenocarcinoma/patologia , Animais , Animais Geneticamente Modificados , Western Blotting , Proliferação de Células/efeitos dos fármacos , Progressão da Doença , Feminino , Humanos , Técnicas Imunoenzimáticas , Neoplasias Renais/secundário , Neoplasias Hepáticas/secundário , Neoplasias Pulmonares/secundário , Metástase Linfática , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Óxido Nítrico/antagonistas & inibidores , Ácido Oleanólico/uso terapêutico , Neoplasia Prostática Intraepitelial/tratamento farmacológico , Neoplasia Prostática Intraepitelial/patologia , Neoplasias da Próstata/patologia , RNA Mensageiro/genética , Reação em Cadeia da Polimerase Via Transcriptase ReversaRESUMO
CDDO-Me, an oleanane synthetic triterpenoid has shown strong antitumorigeic activity towards diverse cancer cell types including colorectal cancer cells. In the present study, we investigated the role of free radicals in the growth inhibitory and apoptosis-inducing activity of CDDO-Me in colorectal cancer cells lines. Results demonstrated that CDDO-Me potently inhibited the growth of colorectal cancer cells and pretreatment of cancer cells with small-molecule antioxidant N-acetylcysteine (NAC) completely blocked the growth inhibitory activity of CDDO-Me. CDDO-Me caused the generation of reactive oxygen species, which was inhibited by NAC and mitochondrial chain 1 complex inhibitors DPI and rotenone. CDDO-Me induced apoptosis as demonstrated by the cleavage of PARP-1, activation of procaspases -3, -8, and -9 and mitochondrial depolarization and NAC blocked the activation of these apoptosis related processes. Furthermore, induction of apoptosis by CDDO-Me was associated with the inhibition of antiapoptotic/ prosurvival Akt, mTOR and NF-kappaB signaling proteins and the inhibition of these signaling molecules was blocked by NAG. Together these studies provided evidence that CDDO-Me is a potent anticancer agent, which imparts growth inhibition and apoptosis in colorectal cancer cells through the generation of free radicals.
Assuntos
Antineoplásicos/farmacologia , Neoplasias Colorretais/tratamento farmacológico , Ácido Oleanólico/análogos & derivados , Espécies Reativas de Oxigênio/metabolismo , Acetilcisteína/metabolismo , Antioxidantes/metabolismo , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Neoplasias Colorretais/patologia , Radicais Livres/metabolismo , Humanos , Ácido Oleanólico/farmacologiaRESUMO
BACKGROUND: The current method to determine the efficacy of chemoprevention in TRAMP mouse model of carcinoma of prostate (CaP) is by extracting and weighing the prostate at different time points or by immunohistochemistry analysis. Non-invasive determination of volumes of prostate glands and seminal vesicles before, during and after treatment would be valuable in investigating the efficacy of newer chemopreventive agents in CaP. The purpose of this study was to determine whether in vivo magnetic resonance imaging (MRI) using a 3 tesla clinical MRI system can be used to follow the effect of chemoprevention in TRAMP model of mouse CaP. METHODS: Mice were randomized into control and treated groups. The animals in treated group received 10 µmol/kg of CDDO, 5 days a week for 20 weeks. Animals underwent in vivo MRI of prostate gland and seminal vesicles by a clinical 3 Tesla MRI system just before (at 5 weeks), during and at the end of treatment, at 25 weeks. T1-weighted and fat saturation (FATSAT) multiecho fast spin echo T2-weighted images (T2WI) were acquired. Volume of the prostate glands and seminal vesicles was determined from MR images. T2 signal intensity changes in the seminal vesicles were determined by subtracting higher echo time (TE) from lower TE T2WI. Following treatments all animals were sacrificed, prostate and seminal vesicles collected, and the tissues prepared for histological staining. All data were expressed as mean ± 1 standard deviation. Two-way or multivariate analysis of variance followed by post-hoc test was applied to determine the significant differences. A p-value of <0.05 was considered significant. RESULTS: Histological analysis indicated tumor in 100% of control mice, whereas 10% of the treated mice showed tumor in prostate gland. Both MRI and measured prostate weights showed higher volume/weight in control mouse group. MRI showed significantly higher volume of seminal vesicles in control animals and T2 signal intensity changes in seminal vesicles of control mice indicating higher number of tumor foci, which was also proven by histology. CONCLUSIONS: In vivo MRI is helpful in determining the efficacy of chemoprevention of prostate cancer in TRAMP mice.
Assuntos
Quimioprevenção , Imageamento por Ressonância Magnética , Ácido Oleanólico/análogos & derivados , Neoplasias da Próstata/prevenção & controle , Animais , Progressão da Doença , Masculino , Camundongos , Camundongos Transgênicos , Ácido Oleanólico/uso terapêutico , Próstata/patologia , Neoplasias da Próstata/patologia , Glândulas Seminais/patologiaRESUMO
BACKGROUND: Synthetic triterpenoids are potent anticancer agents, but their therapeutic efficacy or mechanism of action for prostate cancer has not been investigated. The goal of this study was to determine the antitumor activity and the mechanism of action of methyl-2-cyano-3,12-dioxooleana-1,9(11)-dien-28-oate (CDDO-Me), a oleanane-derived synthetic triterpenoid for human prostate cancer cells. METHODS: The antitumor activity of CDDO-Me for hormone-refractory PC-3 (AR(-)) and C4-2 (AR(+)) prostate cancer cell lines was determined by effects on cell growth and induction of apoptosis, identification of molecular targets, and therapeutic efficacy in vivo in PC-3 xenograft model. RESULTS: CDDO-Me inhibited the growth and induced apoptosis in PC-3 and C4-2 cells at extremely low concentrations. The antitumor activity of CDDO-Me was associated with the inhibition of p-Akt, mammalian target of rapamycin (mTOR), and nuclear factor kappa B (NF-kappaB) signaling proteins and their downstream targets such as p-Bad and p-Foxo3a (Akt); p-S6K1, p-eIF-4E and p-4E-BP1 (mTOR); and COX-2, VEGF and cyclin D1(NF-kappaB). Silencing of Akt sensitized the PC-3 cells to CDDO-Me, whereas overexpression of Akt induced resistance to CDDO-Me. Targeted silencing of Akt showed that Akt does not regulate mTOR activation in PC-3 cells, but targeted silencing of mTOR sensitized PC-3 cells to CDDO-Me mediated growth inhibition. Further, treatment with CDDO-Me inhibited the growth of PC-3 xenografts in nude mice. CONCLUSIONS: This study demonstrated potent antitumor activity of CDDO-Me against prostate cancer cells both in vitro and in vivo. Data also identified Akt and mTOR as molecular targets of CDDO-Me in prostate cancer cells.
Assuntos
Apoptose/efeitos dos fármacos , Divisão Celular/efeitos dos fármacos , Ácido Oleanólico/análogos & derivados , Neoplasias da Próstata/patologia , Proteínas Quinases/genética , Proteínas Proto-Oncogênicas c-akt/genética , Triterpenos/farmacologia , Animais , Western Blotting , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Masculino , Camundongos , Camundongos Nus , NF-kappa B/efeitos dos fármacos , NF-kappa B/metabolismo , Proteínas Nucleares/efeitos dos fármacos , Proteínas Nucleares/isolamento & purificação , Ácido Oleanólico/farmacologia , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/genética , Proteínas Quinases/efeitos dos fármacos , Proteínas Proto-Oncogênicas c-akt/efeitos dos fármacos , Serina-Treonina Quinases TOR , Transplante HeterólogoRESUMO
Xanthohumol (XN), a prenylated chalcone present in hops (Humulus lupus L.) and beer, exhibits anti-inflammatory, antioxidant and antiproliferative activity, but has not been studied for effects on T cell-mediated immune responses. Here we demonstrate that XN has profound immunosuppressive effects on T cell proliferation, development of IL-2 activated killer (LAK) cells, cytotoxic T lymphocytes (CTLs), and production of Th1 cytokines (IL-2, IFN-gamma and TNF-alpha). The suppression of these cell-mediated immune responses by XN was at, least in part, due to the inhibition of nuclear factor kappa B (NF-kappaB) transcription factor through suppression of phosphorylation of IkappaBalpha, an inhibitor of NF-kappaB.
Assuntos
Proliferação de Células/efeitos dos fármacos , Flavonoides/farmacologia , Imunossupressores/farmacologia , Linfócitos/imunologia , NF-kappa B/antagonistas & inibidores , Propiofenonas/farmacologia , Animais , Citocinas/biossíntese , Citocinas/imunologia , Flavonoides/química , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/imunologia , Humulus/química , Proteínas I-kappa B/imunologia , Proteínas I-kappa B/metabolismo , Imunidade Celular/efeitos dos fármacos , Imunidade Celular/imunologia , Imunossupressores/química , Linfócitos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C3H , Inibidor de NF-kappaB alfa , NF-kappa B/imunologia , NF-kappa B/metabolismo , Fosforilação/efeitos dos fármacos , Fosforilação/imunologia , Propiofenonas/químicaRESUMO
Chemoprevention represents a promising strategy to reducing the incidence of prostate cancer which afflicts more than 240,000 males annually in the U.S. 2-cyano-3,12-dioxooleana-1,9(11)-dien-28-oic acid (CDDO) and its C-28 methyl ester (CCDO-Me) and C-28 imidazole (CDDO-Im) derivatives are synthetic oleanane triterpenoids that exhibit several-fold more potent antiinflammatory activity than naturally occurring oleanolic acid, but have not been investigated for prevention of the prostate. In order to evaluate the anticancer activity of CDDOs for prostate cancer, we have investigated the effect of synthetic oleanane triterpenoids on molecular targets relevant to the chemoprevention and treatment of prostate cancer in vitro in TRAMPC-1 cells derived from the primary tumor in the prostate of a transgenic adenocarcinoma of the mouse prostate (TRAMP) mouse. Data demonstrate that CDDOs strongly inhibit the proliferation of TRAMPC-1 cells with a potency order of CDDO-Me>CDDO-Im>CDDO. Because CDDO-Me showed the most growth inhibitory activity it was further analyzed for the anticancer activity. CDDO-Me induced apoptosis in TRAMPC-1 cells as shown by the increased binding of annexin V-FITC and cleavage of procaspases 3, -8, and -9. It effectively inhibited the molecular targets such as p-Akt, NF-kappaB, and p-mTOR and downstream effectors of mTOR (p-S6K1, cyclin-D1, and cdk4). Further, CDDO-Me inhibited NF-kappaB-regulated antiapoptotic Bcl-2, Bcl-xL, and XIAP and proangiogenic VEGF. Taken together, these data demonstrate that CDDO-Me is potentially a potent chemopreventive agent that inhibits several molecular targets that are known to play critical roles in the development and progression of prostate cancer.
Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Ácido Oleanólico/análogos & derivados , Neoplasias da Próstata/metabolismo , Animais , Animais Geneticamente Modificados , Antineoplásicos/uso terapêutico , Linhagem Celular Tumoral/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Inibidores Enzimáticos/uso terapêutico , Humanos , Masculino , Camundongos , NF-kappa B/antagonistas & inibidores , NF-kappa B/metabolismo , Óxido Nítrico/antagonistas & inibidores , Óxido Nítrico/metabolismo , Ácido Oleanólico/farmacologia , Ácido Oleanólico/uso terapêutico , Neoplasias da Próstata/tratamento farmacológico , Neoplasias da Próstata/patologia , Proteínas Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-akt/metabolismo , Serina-Treonina Quinases TORRESUMO
Because of lack of effective treatment options for hormone-refractory prostate cancer at the present time, the need for developing novel therapeutic strategies and targets to treat and prevent the progression of hormone-sensitive prostate cancer to the hormone-refractory stage is paramount. Our previous in vitro studies have shown that curcumin sensitizes both hormone-sensitive and hormone-resistant prostate cancer cells to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) and that combined curcumin/TRAIL treatment induces apoptosis in cancer cells by inhibiting antiapoptotic p-Akt and nuclear factor-kappaB (NF-kappaB). In the present study, we demonstrate that curcumin and TRAIL combination regimen is also the most effective treatment for inhibiting the growth of PC3 xenografts compared to curcumin or TRAIL monotherpy. The inhibition of PC3 tumors by combined treatment correlated with significant reduction in expression of p-Akt and NF-kappaB in tumor tissue. Furthermore, tumor growth inhibition by curcumin/TRAIL combination regimen was associated with significant decrease in cell proliferation and an increase in terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL)-positive cells in the tumors without significant change in microvessel density. Based on the significant efficacy in this preclinical model, combined curcumin/TRAIL regimen may be an effective adjuvant therapy for hormone-refractory prostate cancer.
Assuntos
Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Neoplasias da Próstata/tratamento farmacológico , Animais , Antineoplásicos Hormonais/uso terapêutico , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Curcumina/administração & dosagem , Resistencia a Medicamentos Antineoplásicos , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , NF-kappa B/metabolismo , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Ligante Indutor de Apoptose Relacionado a TNF/administração & dosagem , Resultado do Tratamento , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
Synthetic oleanane triterpenoids (CDDO, CDDO-Im and CDDO-Me) are potent anti-inflammatory agents, but have not been investigated for effects on T cell-mediated immune responses. Here we demonstrate that CDDOs have profound immunosuppressive effects on T cell proliferation, development of IL-2 activated LAK cells and cytotoxic T lymphocytes (CTLs), and expression of cytokines at concentrations of 1.25 microM to 0.078 microM. Treatment with CDDO-Me also inhibited the generation of allo-reactive T cell responses in vivo. The suppression of these cell-mediated immune responses by CDDO-Me was associated with the inhibition of NF-kappaB transcription factor.
Assuntos
Proliferação de Células/efeitos dos fármacos , Citocinas/metabolismo , Citotoxicidade Imunológica/efeitos dos fármacos , Imunidade Celular/efeitos dos fármacos , Imunossupressores/farmacologia , Ativação Linfocitária/efeitos dos fármacos , NF-kappa B/antagonistas & inibidores , Ácido Oleanólico/farmacologia , Linfócitos T/efeitos dos fármacos , Animais , Células Cultivadas , Citocinas/genética , Relação Dose-Resposta a Droga , Regulação da Expressão Gênica/efeitos dos fármacos , Proteínas I-kappa B/metabolismo , Imidazóis/farmacologia , Isoantígenos/imunologia , Células Matadoras Ativadas por Linfocina/efeitos dos fármacos , Células Matadoras Ativadas por Linfocina/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Inibidor de NF-kappaB alfa , NF-kappa B/metabolismo , Ácido Oleanólico/análogos & derivados , Fosforilação , Linfócitos T/imunologia , Linfócitos T Citotóxicos/efeitos dos fármacos , Linfócitos T Citotóxicos/imunologiaRESUMO
BACKGROUND: Synthetic oleanolic acid triterpenoid 2-cyano-3,12-dioxooleana-, 9(11)-dien-28-oic acid (CDDO) and its methyl ester (CDDO-Me) and imidazole (CDDO-Im) derivatives exhibit potent antitumor activity against diverse types of tumor cell lines. However, the anticancer activity of these triterpenoids against prostate cancer cells has not been reported. MATERIALS AND METHODS: The apoptosis-inducing activity of CDDO-Me in human prostate cancer cell lines was investigated using flow cytometry and immunoblotting. RESULTS: Prostate cancer cells are highly sensitive to CDDO-Me at concentrations of 1.25 to 10 microM. The primary mode of tumor cell destruction was apoptosis as demonstrated by increase in annexin V-FITC binding, activation of procaspases, release of cytochrome c from mitochondria, and inhibition of anti-apoptotic proteins. Furthermore, CDDO-Me inhibited the levels of anti-apoptotic Akt, mTOR and NF-kappaB (p65) signaling molecules. CONCLUSION: These studies provide a rationale for clinical evaluation of CDDO-Me as adjuvant therapy for treatment of advanced and fatal form of prostate cancer.
Assuntos
Apoptose/efeitos dos fármacos , NF-kappa B/antagonistas & inibidores , Ácido Oleanólico/análogos & derivados , Neoplasias da Próstata/tratamento farmacológico , Proteínas Quinases/metabolismo , Proteínas Proto-Oncogênicas c-akt/antagonistas & inibidores , Anexina A5/metabolismo , Caspases/metabolismo , Processos de Crescimento Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Citocromos c/metabolismo , Precursores Enzimáticos/metabolismo , Fluoresceína-5-Isotiocianato/metabolismo , Humanos , Isoenzimas/metabolismo , Masculino , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , NF-kappa B/metabolismo , Ácido Oleanólico/farmacologia , Neoplasias da Próstata/metabolismo , Neoplasias da Próstata/patologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Proteínas Proto-Oncogênicas c-bcl-2/antagonistas & inibidores , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Transdução de Sinais/efeitos dos fármacos , Serina-Treonina Quinases TOR , Proteínas Inibidoras de Apoptose Ligadas ao Cromossomo X/antagonistas & inibidores , Proteínas Inibidoras de Apoptose Ligadas ao Cromossomo X/metabolismo , Proteína bcl-X/antagonistas & inibidores , Proteína bcl-X/metabolismoRESUMO
We have previously demonstrated that TNF-alpha gene therapy with myeloid progenitor cells inhibits the progression of 32Dp210 myeloid leukemia in mice. Because TNF-alpha has been shown to induce the activation and maturation of dendritic cells (DCs), we investigated the efficacy of DC-based leukemia vaccine for eradication of residual disease when administered following cytoreductive therapy. Immunization with DC cells loaded with 32Dp210 myeloid leukemia cells (32Dp210 vaccine) was far more effective in preventing the development of leukemia compared to immunization with irradiated leukemia cells alone. The resistance to leukemia could be adoptively transferred to naïve mice with the spleen cells of mice immunized with DC-32Dp210 vaccine, and splenic cells responsible for adoptive transfer of resistance were identified as CD90 + T lymphocytes. Development of immunity in vaccinated mice was associated with the generation of leukemia specific cytotoxic T lymphocytes (CTLs) and secretion of cytokines TNF-alpha and IFN-gamma. Further, immunization with DC-32Dp210 vaccine following cytoreductive therapy with Cytoxan was effective in eradicating residual disease in approximately 50 percent of the animals. However, eradication of residual disease was significantly improved (approximately 74%) when animals were treated with DC-32Dp210 vaccine in which DCs were activated with TNF-alpha prior to loading of 32Dp210 leukemia cells. Cured mice were in molecular remission since Bcr/Abl oncogene could not be amplified from the DNA isolated from the marrow, spleen, or liver of cured mice. Taken together, these data demonstrate the efficacy of DC-based leukemia vaccine for eradication of residual disease and prevention of relapse.