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1.
J Enzyme Inhib Med Chem ; 34(1): 1158-1163, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31179772

RESUMO

Even though phenazines have been extensively reported as anticancer molecules, the molecular target of these compounds is severely lagging behind. Our study consequently focuses on the anticancer target of a phenazine analogue (CPUL1) for its potently antitumor activities in initial stage. Along with redox status courses of Hep G2 cells, thioredoxin reductase I (TrxR1) was speculated as anticancer target of CPUL1. By virtue of zymologic, immunological and molecular biological experiments, we demonstrated that TrxR1 could be the anticancer target of CPUL1. The knowledge on phenazine targeting to TrxR1 have not been reported previously. Thus, it can provide valuable information for further development of the TrxR1 inhibitors.


Assuntos
Antineoplásicos/farmacologia , Inibidores Enzimáticos/farmacologia , Fenazinas/química , Fenazinas/farmacologia , Tiorredoxina Redutase 1/antagonistas & inibidores , Antineoplásicos/síntese química , Antineoplásicos/química , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Células Hep G2 , Humanos , Modelos Moleculares , Estrutura Molecular , Fenazinas/síntese química , Espécies Reativas de Oxigênio/metabolismo , Relação Estrutura-Atividade , Tiorredoxina Redutase 1/metabolismo
2.
Nat Commun ; 10(1): 2745, 2019 06 21.
Artigo em Inglês | MEDLINE | ID: mdl-31227705

RESUMO

Small molecule probes are indispensable tools to explore diverse cellular events. However, finding a specific probe of a target remains a high challenge. Here we report the discovery of Fast-TRFS, a specific and superfast fluorogenic probe of mammalian thioredoxin reductase, a ubiquitous enzyme involved in regulation of diverse cellular redox signaling pathways. By systematically examining the processes of fluorophore release and reduction of cyclic disulfides/diselenides by the enzyme, structural factors that determine the response rate and specificity of the probe are disclosed. Mechanistic studies reveal that the fluorescence signal is switched on by a simple reduction of the disulfide bond within the probe, which is in stark contrast to the sensing mechanism of published probes. The favorable properties of Fast-TRFS enable development of a high-throughput screening assay to discover inhibitors of thioredoxin reductase by using crude tissue extracts as a source of the enzyme.


Assuntos
Descoberta de Drogas/métodos , Corantes Fluorescentes/química , Imagem Molecular/métodos , Sondas Moleculares/química , Tiorredoxina Redutase 1/metabolismo , Animais , Produtos Biológicos/farmacologia , Misturas Complexas , Dissulfetos/química , Corantes Fluorescentes/metabolismo , Células HeLa , Ensaios de Triagem em Larga Escala/métodos , Humanos , Microscopia Intravital/métodos , Microscopia de Fluorescência/métodos , Sondas Moleculares/metabolismo , Oxirredução , RNA Interferente Pequeno/metabolismo , Proteínas Recombinantes/metabolismo , Especificidade por Substrato , Tiorredoxina Redutase 1/antagonistas & inibidores , Tiorredoxina Redutase 1/genética
3.
Oxid Med Cell Longev ; 2019: 1503478, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31049125

RESUMO

When dairy cows are exposed to high-temperature environment, their antioxidant capacity and productive performance decrease, leading to economic losses. Emerging evidence has shown that selenium (Se) can effectively alleviate heat stress in dairy cows; however, the cellular mechanism underlying this protection is not clear. The purpose of this study was to investigate and compare the protective effects of inorganic Se (sodium selenite, SS) and organic Se (selenite methionine, SM) in MAC-T (mammary alveolar cells-large T antigen, a bovine mammary epithelial cell (BMEC) line) cells during heat stress. MAC-T cells were treated in 4 ways unless otherwise described: (i) cells in the heat treatment (HT) group were cultured at 42.5°C for 1 h and then recovered in 37°C for another 12 h; (ii) the SM group was pretreated with organic Se for 2 h, cultured at 42.5°C for 1 h, and then recovered in 37°C for 12 h; (iii) the SS group was treated similarly to the SM group except that the cells were pretreated with inorganic Se instead of organic Se; and (iv) the control group was continuously cultured in 37°C and received no Se treatment. The results showed that heat shock at 42.5°C for 1 h triggered heat shock response, sabotaged the redox balance, and reduced cell viability in MAC-T cells; and pretreatment of cells with SM or SS effectively alleviated the negative effects of heat shock on the cells. However, the cells were much more sensitive to SS treatment but more tolerant to SM. In addition, two forms of Se appeared to affect the expression of different genes, including nuclear factor erythroid 2-related factor 2 (Nrf2) and inducible nitric oxide synthase (iNOS) in the SM group and thioredoxin reductase 1 (TXNRD1) in the SS group in Nrf2-ARE (antioxidant response element) antioxidant pathway and inflammation response. In summary, results showed the mechanistic differences in the protective effects of organic and inorganic Se on heat stress in BMECs.


Assuntos
Antioxidantes/farmacologia , Células Epiteliais/metabolismo , Resposta ao Choque Térmico/efeitos dos fármacos , Glândulas Mamárias Animais/metabolismo , Selênio/farmacologia , Selenito de Sódio/farmacologia , Animais , Bovinos , Linhagem Celular , Células Epiteliais/patologia , Feminino , Glândulas Mamárias Animais/patologia , Fator 2 Relacionado a NF-E2/metabolismo , Óxido Nítrico Sintase Tipo II/metabolismo , Tiorredoxina Redutase 1/metabolismo
4.
Nutrients ; 11(4)2019 Mar 28.
Artigo em Inglês | MEDLINE | ID: mdl-30925798

RESUMO

The application of plant extracts for therapeutic purposes has been used in traditional medicine since the plants are a source of a great variety of chemical compounds that possess biological activity. Actually, the effect of these extracts on diseases such as cancer is being widely studied. Colorectal adenocarcinoma is one of the main causes of cancer related to death and the second most prevalent carcinoma in Western countries. The aim of this work is to study the possible effect of two fenugreek (Trigonella foenum graecum) protein hydrolysates on treatment and progression of colorectal cancer. Fenugreek proteins from seeds were hydrolysed by using two enzymes separately, which are named Purafect and Esperase, and were then tested on differentiated and undifferentiated human colonic adenocarcinoma Caco2/TC7 cells. Both hydrolysates did not affect the growth of differentiated cells, while they caused a decrease in undifferentiated cell proliferation by early apoptosis and cell cycle arrest in phase G1. This was triggered by a mitochondrial membrane permeabilization, cytochrome C release to cytoplasm, and caspase-3 activation. In addition, the hydrolysates of fenugreek proteins displayed antioxidant activity since they reduce the intracellular levels of ROS. These findings suggest that fenugreek protein hydrolysates could be used as nutraceutical molecules in colorectal cancer treatment.


Assuntos
Proliferação de Células/efeitos dos fármacos , Neoplasias do Colo/tratamento farmacológico , Suplementos Nutricionais , Proteínas de Plantas/química , Hidrolisados de Proteína/farmacologia , Trigonella/química , Apoptose/efeitos dos fármacos , Células CACO-2 , Caspase 3 , Neoplasias do Colo/metabolismo , Citocromos c , Regulação Neoplásica da Expressão Gênica/efeitos dos fármacos , Humanos , Hidrolisados de Proteína/química , Espécies Reativas de Oxigênio , Tiorredoxina Redutase 1/genética , Tiorredoxina Redutase 1/metabolismo
5.
J Enzyme Inhib Med Chem ; 34(1): 665-671, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-30746961

RESUMO

Human thioredoxin reductase 1 (TrxR1) is a selenocysteine-containing enzyme which plays a crucial role in regulating numerous redox signalling pathways within the cell. While its functioning is important in all cells, levels of TrxR1 expression are higher in cancer cells, possibly as an adaptation to much higher levels of reactive oxygen species and the need for more extensive DNA synthesis. This makes TrxR1 an attractive target for cancer therapy development. Inspired by the structure of disulphide compounds which have advanced through various stages of clinical development, we designed a series of dithiodiglycolic acid derivatives. These were prepared from respective thiol synthons using an iodine- or benzotriazolyl chloride-promoted oxidative disulphide bond formation. Inhibition of TrxR present in cell lysates from human neuroblastoma cells (SH-SY5Y) and rat liver cells indicated several compounds with a potential for TrxR inhibition. Some of these compounds were also tested for growth inhibition against two human cancer cell lines and normal human keratinocytes.


Assuntos
Antineoplásicos/farmacologia , Desenho de Drogas , Inibidores Enzimáticos/farmacologia , Glicolatos/farmacologia , Compostos de Sulfidrila/farmacologia , Tiorredoxina Redutase 1/antagonistas & inibidores , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Glicolatos/síntese química , Glicolatos/química , Humanos , Queratinócitos/efeitos dos fármacos , Estrutura Molecular , Oxirredução , Ratos , Relação Estrutura-Atividade , Compostos de Sulfidrila/química , Tiorredoxina Redutase 1/metabolismo , Células Tumorais Cultivadas
6.
Oxid Med Cell Longev ; 2019: 7945983, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30805084

RESUMO

Background: Aurothioglucose- (ATG-) mediated inhibition of thioredoxin reductase-1 (TXNRD1) improves alveolarization in experimental murine bronchopulmonary dysplasia (BPD). Glutathione (GSH) mediates susceptibility to neonatal and adult oxidative lung injury. We have previously shown that ATG attenuates hyperoxic lung injury and enhances glutathione- (GSH-) dependent antioxidant defenses in adult mice. Hypothesis: The present studies evaluated the effects of TXNRD1 inhibition on GSH-dependent antioxidant defenses in newborn mice in vivo and lung epithelia in vitro. Methods: Newborn mice received intraperitoneal ATG or saline prior to room air or 85% hyperoxia exposure. Glutamate-cysteine ligase (GCL) catalytic (Gclc) and modifier (Gclm) mRNA levels, total GSH levels, total GSH peroxidase (GPx) activity, and Gpx2 expression were determined in lung homogenates. In vitro, murine transformed club cells (mtCCs) were treated with the TXNRD1 inhibitor auranofin (AFN) or vehicle in the presence or absence of the GCL inhibitor buthionine sulfoximine (BSO). Results: In vivo, ATG enhanced hyperoxia-induced increases in Gclc mRNA levels, total GSH contents, and GPx activity. In vitro, AFN increased Gclm mRNA levels, intracellular and extracellular GSH levels, and GPx activity. BSO prevented AFN-induced increases in GSH levels. Conclusions: Our data are consistent with a model in which TXNRD1 inhibition augments hyperoxia-induced GSH-dependent antioxidant responses in neonatal mice. Discrepancies between in vivo and in vitro results highlight the need for methodologies that permit accurate assessments of the GSH system at the single-cell level.


Assuntos
Antioxidantes/metabolismo , Displasia Broncopulmonar/enzimologia , Displasia Broncopulmonar/patologia , Glutationa/metabolismo , Tiorredoxina Redutase 1/antagonistas & inibidores , Animais , Animais Recém-Nascidos , Aurotioglucose , Displasia Broncopulmonar/genética , Células Epiteliais/metabolismo , Glutamato-Cisteína Ligase/genética , Glutamato-Cisteína Ligase/metabolismo , Glutationa Peroxidase/metabolismo , Hiperóxia/genética , Hiperóxia/patologia , Pulmão/metabolismo , Pulmão/patologia , Camundongos , RNA Mensageiro/genética , RNA Mensageiro/metabolismo , Tiorredoxina Redutase 1/metabolismo
7.
Free Radic Res ; 53(1): 104-114, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30668191

RESUMO

Thioredoxin reductase 1 (TrxR1) has emerged as a potential target for cancer therapy, because it is overexpressed in several types of cancers and associated with increased tumour growth and poor patient prognosis. Alantolactone (ALT), a natural sesquiterpene lactone originated from traditional folk medicine Inula helenium L., has been reported to exert antitumor activity in various tumours. However, the effect of ALT on human gastric cancer cells and its underlying mechanism remains unknown. In this study, we showed that ALT inhibited cell proliferation and induced cell apoptosis in gastric cancer cells. Mechanistically, our data found that ALT induced reactive oxygen species (ROS) production by inhibiting TrxR1 activity, resulting in the activation of p38 mitogen-activated protein kinase (MAPK) pathway and eventually cell apoptosis in gastric cancer cells. And the effects of ALT were reversed by pre-treatment with NAC (a scavenger of ROS). Further investigation revealed that ALT displayed synergistic lethality with erastin against gastric cancer cells, which demonstrating combined inhibition of TrxR1 and glutathione (GSH) leads to a synergistic effect in gastric cancer cells. More importantly, ALT treatment markedly reduced the activity of TrxR1 in vivo and inhibited the growth of gastric cancer xenografts without exhibiting significant toxicity. Taken together, these findings suggest that ALT may be used as a novel therapeutic agent against human gastric cancer.


Assuntos
Antineoplásicos Fitogênicos/farmacologia , Lactonas/farmacologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Espécies Reativas de Oxigênio/metabolismo , Sesquiterpenos de Eudesmano/farmacologia , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/patologia , Tiorredoxina Redutase 1/antagonistas & inibidores , Proteínas Quinases p38 Ativadas por Mitógeno/metabolismo , Animais , Antineoplásicos Fitogênicos/química , Antineoplásicos Fitogênicos/isolamento & purificação , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Feminino , Humanos , Inula/química , Lactonas/química , Lactonas/isolamento & purificação , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Nus , Neoplasias Experimentais/tratamento farmacológico , Neoplasias Experimentais/metabolismo , Neoplasias Experimentais/patologia , Sesquiterpenos de Eudesmano/química , Sesquiterpenos de Eudesmano/isolamento & purificação , Neoplasias Gástricas/metabolismo , Relação Estrutura-Atividade , Tiorredoxina Redutase 1/metabolismo , Células Tumorais Cultivadas
8.
Redox Biol ; 21: 101061, 2019 02.
Artigo em Inglês | MEDLINE | ID: mdl-30590310

RESUMO

Gastric cancer is one of the leading causes of cancer-related deaths. Chemotherapy has improved long-term survival of patients with gastric cancer. Unfortunately, cancer readily develops resistance to apoptosis-inducing agents. New mechanisms, inducing caspase-independent paraptosis-like cell death in cancer cells is presently emerging as a potential direction. We previously developed a curcumin analog B63 as an anti-cancer agent in pre-clinical evaluation. In the present study, we evaluated the effect and mechanism of B63 on gastric cancer cells. Our studies show that B63 targets TrxR1 protein and increases cellular reactive oxygen species (ROS) level, which results in halting gastric cancer cells and inducing caspase-independent paraptotic modes of death. The paraptosis induced by B63 was mediated by ROS-mediated ER stress and MAPK activation. Either overexpression of TrxR1 or suppression of ROS normalized B63-induced paraptosis in gastric cancer cells. Furthermore, B63 caused paraptosis in 5-fluorouracil-resistant gastric cancer cells, and B63 treatment reduced the growth of gastric cancer xenografts, which was associated with increased ROS and paraptosis. Collectively, our findings provide a novel strategy for the treatment of gastric cancer by utilizing TrxR1-mediated oxidative stress generation and subsequent cell paraptosis.


Assuntos
Apoptose/efeitos dos fármacos , Curcumina/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Neoplasias Gástricas/metabolismo , Tiorredoxina Redutase 1/antagonistas & inibidores , Animais , Antineoplásicos/química , Antineoplásicos/farmacologia , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Curcumina/análogos & derivados , Curcumina/química , Modelos Animais de Doenças , Estresse do Retículo Endoplasmático/efeitos dos fármacos , Feminino , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Camundongos , Modelos Biológicos , Modelos Moleculares , Terapia de Alvo Molecular , Estresse Oxidativo/efeitos dos fármacos , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/patologia , Relação Estrutura-Atividade , Tiorredoxina Redutase 1/química , Tiorredoxina Redutase 1/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
9.
J Immunol Res ; 2018: 6140320, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30225271

RESUMO

Thioredoxin reductase (TrxR), an antioxidant enzyme dependent on nicotinamide adenine dinucleotide phosphate, plays a vital role in defense against oxidative stress. However, the role of microRNAs targeting TrxR under oxidative stress has not yet been determined. In this study, we tested the involvement of miRNA-mediated posttranscriptional regulation in H2O2-induced TrxR1 expression in endothelial cells. Dual luciferase assay combined with expression analysis confirmed that miR-125a suppressed TrxR1 expression by targeting its 3'-UTR. Furthermore, H2O2 induced TrxR1 expression partly through downregulation of miR-125a. These findings indicate that miRNA-mediated posttranscriptional mechanism is involved in H2O2-induced TrxR1 expression in endothelial cells, suggesting an important role of miRNAs in the response to oxidative stress.


Assuntos
Endotélio Vascular/fisiologia , MicroRNAs/genética , Tiorredoxina Redutase 1/genética , Regiões 3' não Traduzidas/genética , Regulação da Expressão Gênica , Células HEK293 , Células Endoteliais da Veia Umbilical Humana , Humanos , Peróxido de Hidrogênio/metabolismo , NADP/metabolismo , Estresse Oxidativo , Processamento Pós-Transcricional do RNA , Tiorredoxina Redutase 1/metabolismo
10.
Eur J Med Chem ; 156: 493-509, 2018 Aug 05.
Artigo em Inglês | MEDLINE | ID: mdl-30025345

RESUMO

In this study, twenty curcumin analogue hybrids as potential anticancer agents through regulation protein of TrxR were designed and synthesized. Results of anticancer activity showed that 5,7-dimethoxy-3-(3-(2-((1E, 4E)-3-oxo-5-(pyridin-2-yl)penta-1,4-dien-1- yl)phenoxy)propoxy)-2-(3,4,5-trimethoxyphenyl)-4H-chromen-4-one (compound 7d) could induce gastric cancer cells apoptosis by arresting cell cycle, break mitochondria function and inhibit TrxR activity. Meanwhile, western blot revealed that this compound could dramatically up expression of Bax/Bcl-2 ratio and high expression of TrxR oxidation. These results preliminarily show that the important role of ROS mediated activation of ASK1/MAPK signaling pathways by this title compound.


Assuntos
Antineoplásicos/química , Antineoplásicos/farmacologia , Curcumina/análogos & derivados , Curcumina/farmacologia , Neoplasias/tratamento farmacológico , Antineoplásicos/síntese química , Apoptose/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Curcumina/síntese química , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Neoplasias/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Neoplasias Gástricas/tratamento farmacológico , Neoplasias Gástricas/metabolismo , Tiorredoxina Redutase 1/antagonistas & inibidores , Tiorredoxina Redutase 1/metabolismo
11.
J Trace Elem Med Biol ; 49: 43-50, 2018 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-29895371

RESUMO

The present observation based research was designed to evaluate the influence of occupational human exposure to metallic mercury (Hg°) vapor on the biomarkers of selenium status involved in the antioxidant defense system. For this purpose we determined Hg and selenium (Se) concentrations in body fluids, the markers of antioxidant effect measured as an activity of Se-dependent enzymes (red blood cell and plasma glutathione peroxidase: GPx1-RBC and GPx3-P), concentration of selenoprotein P in the plasma (SeP-P) and total antioxidant activity in the plasma (TAA-P) in 131 male workers from a chloralkali plant exposed to Hg° and 67 non-exposed males (control group). The mRNA expression levels of glutathione peroxidases (GPX1, GPX3), selenoprotein P (SEPP1), thioredoxin reductase 1 (TRXR1), thioredoxin 1 (TRX1), peroxiredoxins (PRDX1, PRDX2) were also examined in the leukocytes of peripheral blood. Hg concentration in the blood (Hg-B) and urine (Hg-U) samples was determined using the thermal decomposition amalgamation/atomic absorption spectrometry (TDA-AAS) method and Se concentrations in plasma (Se-P) and urine (Se-U) using the inductively coupled plasma mass spectrometry (ICP-MS) method. Activities of GPx1-RBC, GPx3-P and TAA-P were determined using the kinetic and spectrophotometric method, respectively. Gene expression analysis was performed using the quantitative Real-Time PCR. The results showed significant higher Hg levels among the Hg°-exposed workers in comparison to control group (12-times higher median for Hg-B and almost 74-times higher median for Hg-U concentration in chloralkali workers). Se-P was also significantly higher (Me (median): 82.85 µg/L (IQR (interquartile range) 72.03-90.28 µg/L) for chloralkali workers vs. Me: 72.74 µg/L (IQR 66.25-80.14 µg/L) for control group; p = 0.0001) but interestingly correlated inversely with Hg-U in chloralkali workers suggesting depletion of the Se protection among the workers with the highest Hg-U concentration. The mRNA level for GPX1, PRXD1 were markedly but significantly higher in the workers compared to the control group. Moreover, concentrations of Hg-B and Hg-U among the workers were significantly positively correlated with the levels of selenoprotein P at both the mRNA and selenoprotein levels. In the multivariate model, after adjusting to cofounders (dental amalgam fillings, age, BMI, job seniority time, smoking), we confirmed that Hg-U concentration was inversely correlated with genes expression of TRXR1. This is the first comprehensive assessment of the impact of occupational exposure of workers to Hg° at both the mRNA and selenoprotein levels, with investigation of fish intake obtained by means of a questionnaire. These findings suggest that exposure to Hg° alters gene expression of the antioxidant enzymes and the level of Se-containing selenoproteins.


Assuntos
Antioxidantes/metabolismo , Mercúrio/sangue , Mercúrio/urina , Selênio/sangue , Selênio/urina , Adulto , Glutationa Peroxidase/metabolismo , Humanos , Masculino , Pessoa de Meia-Idade , Peroxirredoxinas/metabolismo , Reação em Cadeia da Polimerase em Tempo Real , Selenoproteína P/metabolismo , Selenoproteínas/metabolismo , Tiorredoxina Redutase 1/metabolismo , Tiorredoxinas/metabolismo
12.
Redox Biol ; 15: 297-315, 2018 05.
Artigo em Inglês | MEDLINE | ID: mdl-29304480

RESUMO

The system (PTTRS) formed by typical 2-Cys peroxiredoxins (Prx), thioredoxin (Trx), Trx reductase (TrxR), and sulfiredoxin (Srx) is central in antioxidant protection and redox signaling in the cytoplasm of eukaryotic cells. Understanding how the PTTRS integrates these functions requires tracing phenotypes to molecular properties, which is non-trivial. Here we analyze this problem based on a model that captures the PTTRS' conserved features. We have mapped the conditions that generate each distinct response to H2O2 supply rates (vsup), and estimated the parameters for thirteen human cell types and for Saccharomyces cerevisiae. The resulting composition-to-phenotype map yielded the following experimentally testable predictions. The PTTRS permits many distinct responses including ultra-sensitivity and hysteresis. However, nearly all tumor cell lines showed a similar response characterized by limited Trx-S- depletion and a substantial but self-limited gradual accumulation of hyperoxidized Prx at high vsup. This similarity ensues from strong correlations between the TrxR, Srx and Prx activities over cell lines, which contribute to maintain the Prx-SS reduction capacity in slight excess over the maximal steady state Prx-SS production. In turn, in erythrocytes, hepatocytes and HepG2 cells high vsup depletes Trx-S- and oxidizes Prx mainly to Prx-SS. In all nucleated human cells the Prx-SS reduction capacity defined a threshold separating two different regimes. At sub-threshold vsup the cytoplasmic H2O2 concentration is determined by Prx, nM-range and spatially localized, whereas at supra-threshold vsup it is determined by much less active alternative sinks and µM-range throughout the cytoplasm. The yeast shows a distinct response where the Prx Tsa1 accumulates in sulfenate form at high vsup. This is mainly due to an exceptional stability of Tsa1's sulfenate. The implications of these findings for thiol redox regulation and cell physiology are discussed. All estimates were thoroughly documented and provided, together with analytical approximations for system properties, as a resource for quantitative redox biology.


Assuntos
Antioxidantes/metabolismo , Oxirredução , Peroxirredoxinas/metabolismo , Tiorredoxinas/metabolismo , Linhagem Celular Tumoral , Glutationa/genética , Glutationa/metabolismo , Células Hep G2 , Humanos , Estresse Oxidativo/genética , Oxirredutases atuantes sobre Doadores de Grupo Enxofre/metabolismo , Peroxidases/genética , Peroxidases/metabolismo , Peroxirredoxinas/genética , Saccharomyces cerevisiae/genética , Saccharomyces cerevisiae/metabolismo , Proteínas de Saccharomyces cerevisiae/genética , Proteínas de Saccharomyces cerevisiae/metabolismo , Transdução de Sinais , Tiorredoxina Redutase 1/metabolismo
13.
Redox Biol ; 14: 237-249, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-28965082

RESUMO

Hepatocellular carcinoma (HCC) accounts for most of primary liver cancer, of which five-year survival rate remains low and chemoprevention has become a strategy to reduce disease burden of HCC. We aim to explore the in vivo chemopreventive effect of an organoselenium-containing compound butaselen (BS) against hepatocarcinogenesis and its underlying mechanisms. Pre- and sustained BS treatment (9, 18 and 36mg/Kg BS) could dose-dependently inhibit chronic hepatic inflammation, fibrosis, cirrhosis and HCC on murine models with 24 weeks treatment scheme. The thioredoxin reductase (TrxR), NF-κB pathway and pro-inflammatory factors were activated during hepatocarcinogenesis, while their expression were decreased by BS treatment. BS treatment could also significantly reduce tumor volume in H22-bearing models and remarkably slow tumor growth. HCC cell lines HepG2, Bel7402 and Huh7 were time- and dose-dependently inhibited by BS treatment. G2/M arrest and apoptosis were observed in HepG2 cells after BS treatment, which were mediated by TrxR/Ref-1 and NF-κB pathways inhibition. BS generated reactive oxygen species (ROS), which could be reduced by antioxidant N-acetyl-L-cysteine (NAC) and NADPH oxidase inhibitor DPI. NAC could markedly increase HepG2 cells viability. TrxR activity of HepG2 cells treated with BS were significantly decreased in parallel with proliferative inhibition. The TrxR1-knockdown HepG2 cells also exhibited low TrxR1 activity, high ROS level, relatively low proliferation rate and increased resistance to BS treatment. In conclusion, BS can prevent hepatocarcinogenesis through inhibiting chronic inflammation, cirrhosis and tumor progression. The underlying mechanisms may include TrxR activity inhibition, leading to ROS elevation, G2/M arrest and apoptosis.


Assuntos
Antineoplásicos/uso terapêutico , Carcinoma Hepatocelular/tratamento farmacológico , Neoplasias Hepáticas/tratamento farmacológico , Compostos Organosselênicos/uso terapêutico , Tiorredoxina Redutase 1/antagonistas & inibidores , Animais , Antineoplásicos/química , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Carcinoma Hepatocelular/metabolismo , Carcinoma Hepatocelular/patologia , Linhagem Celular Tumoral , Células Hep G2 , Humanos , Neoplasias Hepáticas/metabolismo , Neoplasias Hepáticas/patologia , Masculino , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , NF-kappa B/metabolismo , Compostos Organosselênicos/química , Compostos Organosselênicos/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Tiorredoxina Redutase 1/metabolismo , Carga Tumoral/efeitos dos fármacos
14.
Adv Cancer Res ; 136: 139-151, 2017.
Artigo em Inglês | MEDLINE | ID: mdl-29054416

RESUMO

The cytosolic selenoprotein thioredoxin reductase 1 (TrxR1, encoded in human by TXNRD1) is implied to have several different roles in relation to cancer. Its physiologic functions may protect normal cells from carcinogenesis, but may also promote cancer progression if carcinogenesis nonetheless occurs. With distinct links to Nrf2 signaling, ribonucleotide reductase-dependent production of deoxyribonucleotides and its support of several antioxidant systems counteracting oxidative stress, the metabolic pathways regulated, and affected by TrxR1, are altogether of crucial importance in cancer. These pathways and causal relationships are at the same time highly intricate. In spite of the complexity in the cellular redox networks, several observations discussed in this chapter suggest that specific targeting of TrxR1 may be promising as a mechanistic principle for anticancer therapy.


Assuntos
Antineoplásicos/farmacologia , Antineoplásicos/uso terapêutico , Neoplasias/tratamento farmacológico , Neoplasias/metabolismo , Selenoproteínas/metabolismo , Tiorredoxina Redutase 1/metabolismo , Animais , Antioxidantes/metabolismo , Humanos , Oxirredução/efeitos dos fármacos , Estresse Oxidativo/efeitos dos fármacos , Transdução de Sinais/efeitos dos fármacos
15.
J Infect Dis ; 216(4): 477-488, 2017 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-28931237

RESUMO

NQO1 and TRXR1 are important host reductases implicated in the regulation of inflammation and apoptosis. Although the transcriptional machinery governing these processes have been extensively investigated, the associated epigenetic regulatory events remain unclear. Here, we report that SET8, a histone H4 lysine 20 monomethylase (H4K20me1), is highly induced during Mycobacterium tuberculosis infection that orchestrates immune evasion strategies through the induction of NQO1 and TRXR1 in vivo. SET8, along with FoxO3a, mediates an active NQO1-PGC1-α complex, which promotes the anti-inflammatory M2 macrophage phenotype, and assists TRXR1-regulated arrest of tumor necrosis factor-related apoptosis-inducing ligand-induced apoptosis. Strikingly, the loss-of-function analysis in an in vivo mouse tuberculosis model further corroborated the pivotal role of SET8-responsive NQO1 and TRXR1 in mycobacterial survival. Thus, augmenting host immune responses against Mycobacterium tuberculosis by harnessing the SET8-NQO1/TRXR1 axis with its specific and potent inhibitors could lead to promising host-directed therapeutic adjuvants for tuberculosis treatment.


Assuntos
Epigênese Genética , Histona-Lisina N-Metiltransferase/metabolismo , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/imunologia , Mycobacterium tuberculosis/crescimento & desenvolvimento , Tuberculose/imunologia , Animais , Apoptose/efeitos dos fármacos , Modelos Animais de Doenças , Proteína Forkhead Box O3/genética , Proteína Forkhead Box O3/metabolismo , Regulação da Expressão Gênica , Histona-Lisina N-Metiltransferase/genética , Humanos , Evasão da Resposta Imune , Leucócitos Mononucleares/microbiologia , Camundongos , NAD(P)H Desidrogenase (Quinona)/genética , NAD(P)H Desidrogenase (Quinona)/metabolismo , Células RAW 264.7 , Reprodutibilidade dos Testes , Transdução de Sinais , Tiorredoxina Redutase 1/genética , Tiorredoxina Redutase 1/metabolismo , Tuberculose/microbiologia
16.
Free Radic Biol Med ; 112: 277-286, 2017 11.
Artigo em Inglês | MEDLINE | ID: mdl-28774816

RESUMO

Intracellular redox status influences the oxidation and enzyme activity of the tumor suppressor phosphatase and tensin homolog on chromosome 10 (PTEN). Cumene hydroperoxide (CuHP), an organic hydroperoxide, is a known tumor promoter. However, molecular targets and action mechanism of CuHP in tumor promotion have not been well characterized. In this study, we investigated the effect of CuHP on the redox state of PTEN in HeLa cells. In addition, the intracellular reducing system of oxidized PTEN was analyzed using a biochemical approach and the effect of CuHP on this reducing system was also analyzed. While PTEN oxidized by hydrogen peroxide is progressively converted to its reduced form, PTEN was irreversibly oxidized by exposure to CuHP in HeLa cells. A combination of protein fractionation and mass analysis showed that the reducing system of PTEN was comprised of NADPH, thioredoxin reductase (TrxR), and thioredoxin (Trx). Although CuHP-mediated PTEN oxidation was not reversible in cells, CuHP-oxidized PTEN was reactivated by the exogenous Trx system, indicating that the cellular Trx redox system for PTEN is inactivated by CuHP. We present evidence that PTEN oxidation and the concomitant inhibition of thioredoxin by CuHP results in irreversible oxidation of PTEN in HeLa cells. In addition, ablation of peroxiredoxin (Prdx) enhanced CuHP-induced PTEN oxidation in cells. These results provide a new line of evidence that PTEN might be a crucial determinant of cell fate in response to cellular oxidative stress induced by organic hydroperoxides.


Assuntos
Derivados de Benzeno/farmacologia , Carcinógenos/farmacologia , Fibroblastos/efeitos dos fármacos , PTEN Fosfo-Hidrolase/química , Tiorredoxina Redutase 1/metabolismo , Tiorredoxinas/metabolismo , Animais , Linhagem Celular , Embrião de Mamíferos , Fibroblastos/citologia , Fibroblastos/metabolismo , Expressão Gênica , Células HeLa , Humanos , Peróxido de Hidrogênio/farmacologia , Camundongos , NADP/metabolismo , Oxirredução , Estresse Oxidativo , PTEN Fosfo-Hidrolase/genética , PTEN Fosfo-Hidrolase/metabolismo , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Tiorredoxina Redutase 1/genética , Tiorredoxinas/genética
17.
J Biol Chem ; 292(35): 14371-14380, 2017 09 01.
Artigo em Inglês | MEDLINE | ID: mdl-28684416

RESUMO

Regulation of growth factor signaling involves reversible inactivation of protein tyrosine phosphatases (PTPs) through the oxidation and reduction of their active site cysteine. However, there is limited mechanistic understanding of these redox events and their co-ordination in the presence of cellular antioxidant networks. Here we investigated interactions between PTP1B and the peroxiredoxin 2 (Prx2)/thioredoxin 1 (Trx1)/thioredoxin reductase 1 (TrxR1) network. We found that Prx2 becomes oxidized in PDGF-treated fibroblasts, but only when TrxR1 has first been inhibited. Using purified proteins, we also found that PTP1B is relatively insensitive to inactivation by H2O2 but found no evidence for a relay mechanism in which Prx2 or Trx1 facilitates PTP1B oxidation. Instead, these proteins prevented PTP1B inactivation by H2O2 Intriguingly, we discovered that TrxR1/NADPH directly protects PTP1B from inactivation when present during the H2O2 exposure. This protection was dependent on the concentration of TrxR1 and independent of Trx1 and Prx2. The protection was blocked by auranofin and required an intact selenocysteine residue in TrxR1. This activity likely involves reduction of the sulfenic acid intermediate form of PTP1B by TrxR1 and is therefore distinct from the previously described reactivation of end-point oxidized PTP1B, which requires both Trx1 and TrxR1. The ability of TrxR1 to directly reduce an oxidized phosphatase is a novel activity that can help explain previously observed increases in PTP1B oxidation and PDGF receptor phosphorylation in TrxR1 knockout cells. The activity of TrxR1 is therefore of potential relevance for understanding the mechanisms of redox regulation of growth factor signaling pathways.


Assuntos
NADP/metabolismo , Proteína Tirosina Fosfatase não Receptora Tipo 1/metabolismo , Proteínas Tirosina Fosfatases Classe 3 Semelhantes a Receptores/metabolismo , Tiorredoxina Redutase 1/metabolismo , Animais , Auranofina/farmacologia , Domínio Catalítico , Células Cultivadas , Dimerização , Embrião de Mamíferos/citologia , Proteínas de Homeodomínio/química , Proteínas de Homeodomínio/genética , Proteínas de Homeodomínio/metabolismo , Humanos , Peróxido de Hidrogênio/farmacologia , Camundongos , Oxidantes/farmacologia , Oxirredução , Fragmentos de Peptídeos/química , Fragmentos de Peptídeos/genética , Fragmentos de Peptídeos/metabolismo , Domínios e Motivos de Interação entre Proteínas , Proteína Tirosina Fosfatase não Receptora Tipo 1/antagonistas & inibidores , Proteína Tirosina Fosfatase não Receptora Tipo 1/genética , Ratos , Proteínas Tirosina Fosfatases Classe 3 Semelhantes a Receptores/química , Proteínas Tirosina Fosfatases Classe 3 Semelhantes a Receptores/genética , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/metabolismo , Selenocisteína/química , Selenocisteína/metabolismo , Tiorredoxina Redutase 1/antagonistas & inibidores , Tiorredoxina Redutase 1/química , Tiorredoxina Redutase 1/genética , Tiorredoxinas/química , Tiorredoxinas/genética , Tiorredoxinas/metabolismo
18.
J Proteomics ; 166: 27-38, 2017 08 23.
Artigo em Inglês | MEDLINE | ID: mdl-28688915

RESUMO

Cytoprotective effects by upregulating cellular expression levels of antioxidant proteins are attributed to a significant number of food ingredients. Evaluation of those cytoprotective effects and identification of the most active components requires reliable and comprehensive proteomic strategies. Thus, promising potential bioactive ingredients could be determined for the prevention of various diseases, including colon cancer formation. We established a multiplexed nanoLC-MS/MS targeted proteomic method, operated in scheduled selected reaction monitoring mode (sSRM), to record modulation of the expression levels of six major antioxidant proteins induced by dietary phytochemicals. Relative quantification was achieved by isotope-coded protein labels (ICPLs) and based on two to three proteotypic peptides per target protein. The assay provided accurate (mean relative error 6.4%) and precise (mean RSD 7.4%) quantification. Incubation experiments were carried out in primary human colon epithelial cells (HCoEpiCs) and revealed significant upregulation of NAD(P)H dehydrogenase [quinone] 1 (up to threefold) and thioredoxin reductase 1 (up to twofold) by 10µM sulforaphane (from broccoli), 5µM carnosol (rosemary), and 20µM cinnamaldehyde (cinnamon). The latter two substances additionally upregulated heme oxygenase-1 and were identified as the most active components in the test set. The results provide additional evidence for nutritive cytoprotection in human colon cells. SIGNIFICANCE: Targeted proteome analysis using LC coupled to scheduled selected reaction monitoring (sSRM)-MS is a highly flexible and reliable method to monitor protein expression profiles. The present study screened modulators occurring in food, which may be protective against colon cancer by inducing cytoprotective enzymes. Primary human colonic epithelial cells were used because they model the conditions in healthy gut tissue better than immortalized cells. Thus, an LC-MS/MS-sSRM protocol was established and validated including relative quantification of cytoprotective protein expression by isotope-coded protein labels, because metabolic labelling cannot be applied for primary cells. The present study demonstrated that the major components of cinnamon and rosemary, respectively, i.e. cinnamaldehyde and carnosol, are potent alimentary candidates to increase the expression of cytoprotective enzymes in the human colon. Among the investigated enzymes, NAD(P)H dehydrogenase [quinone] 1 (NQO1) was most susceptible towards modulation by phytochemicals. NQO1 exerts its cytoprotective activity by detoxifying electrophilic and oxidative xenobiotics with quinone structure.


Assuntos
Colo/efeitos dos fármacos , Citoproteção/efeitos dos fármacos , Células Epiteliais/enzimologia , Compostos Fitoquímicos/farmacologia , Acroleína/análogos & derivados , Acroleína/farmacologia , Células Cultivadas , Colo/citologia , Colo/enzimologia , Neoplasias do Colo/prevenção & controle , Diterpenos de Abietano/farmacologia , Células Epiteliais/citologia , Células Epiteliais/efeitos dos fármacos , Humanos , Marcação por Isótopo , NAD(P)H Desidrogenase (Quinona)/metabolismo , Proteômica/métodos , Espectrometria de Massas em Tandem , Tiorredoxina Redutase 1/metabolismo
19.
J Cancer Res Clin Oncol ; 143(11): 2257-2265, 2017 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-28653098

RESUMO

PURPOSE: Intracellular antioxidant enzymes are commonly upregulated in various cancer types and are associated with treatment outcomes. Because the relationship has rarely been examined in oral squamous cell carcinoma (OSCC), we aimed to evaluate the association between the levels of glutathione peroxidase (GPX)1, GPX4, and thioredoxin reductase (TrxR)1 expression and prognosis in patients with OSCC who underwent curative surgical resection. METHODS: This study included 233 patients who underwent curative surgery for previously untreated OSCC between 2000 and 2012. Tumour GPX1, GPX4, and TrxR1 expression was evaluated by immunohistochemistry and was dichotomised to low and high values according to defined expression levels. The association between GPX1, GPX4, and TrxR1 expression and clinicopathological results was analysed. Univariate and multivariate analyses using the Cox proportional hazards model were conducted to assess the significance of differences in recurrence or survival outcomes between variables. RESULTS: High GPX1, GPX4, and TrxR1 expression was observed in 99 (42.5%), 133 (57.1%), and 46 (19.7%) patients, respectively. GPX1 overexpression was significantly correlated with nodal metastasis, advanced overall stage, depth of invasion of >10 mm, high grade and perineural invasion (P < 0.05). High GPX4 expression was also related to nodal metastasis, overall advanced stage and high grade (P < 0.05). Univariate and multivariate analyses showed that increased GPX1 expression was significantly associated with poor disease-free, cancer-specific and overall survival (all P < 0.05), while increased GPX4 or TrxR1 expression was not (all P > 0.1). CONCLUSIONS: Tumour GPX1 expression is a useful biomarker predictive of recurrence and survival in OSCC patients.


Assuntos
Biomarcadores Tumorais/metabolismo , Carcinoma de Células Escamosas/secundário , Glutationa Peroxidase/metabolismo , Neoplasias Bucais/patologia , Tiorredoxina Redutase 1/metabolismo , Adulto , Idoso , Idoso de 80 Anos ou mais , Carcinoma de Células Escamosas/metabolismo , Feminino , Seguimentos , Humanos , Metástase Linfática , Masculino , Pessoa de Meia-Idade , Neoplasias Bucais/metabolismo , Invasividade Neoplásica , Estadiamento de Neoplasias , Prognóstico , Taxa de Sobrevida , Adulto Jovem
20.
Cancer Lett ; 403: 98-107, 2017 09 10.
Artigo em Inglês | MEDLINE | ID: mdl-28624622

RESUMO

Organometallic complexes offer the prospect of targeting multiple pathways that are important in cancer biology. Here, the preclinical activity and mechanism(s) of action of a silver-bis(N-heterocyclic carbine) complex (Ag8) were evaluated. Ag8 induced DNA damage via several mechanisms including topoisomerase I/II and thioredoxin reductase inhibition and induction of reactive oxygen species. DNA damage induction was consistent with cytotoxicity observed against proliferating cells and Ag8 induced cell death by apoptosis. Ag8 also inhibited DNA repair enzyme PARP1, showed preferential activity against cisplatin resistant A2780 cells and potentiated the activity of temozolomide. Ag8 was substantially less active against non-proliferating non-cancer cells and selectively inhibited glycolysis in cancer cells. Ag8 also induced significant anti-tumour effects against cells implanted intraperitoneally in hollow fibres but lacked activity against hollow fibres implanted subcutaneously. Thus, Ag8 targets multiple pathways of importance in cancer biology, is less active against non-cancer cells and shows activity in vivo in a loco-regional setting.


Assuntos
Antineoplásicos/farmacologia , Imidazóis/farmacologia , Neoplasias/tratamento farmacológico , Compostos Organometálicos/farmacologia , Antígenos de Neoplasias/metabolismo , Antineoplásicos/química , Antineoplásicos/toxicidade , Protocolos de Quimioterapia Combinada Antineoplásica/farmacologia , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Cisplatino/farmacologia , Dano ao DNA , DNA Topoisomerases Tipo I/metabolismo , DNA Topoisomerases Tipo II/metabolismo , Proteínas de Ligação a DNA/metabolismo , Dacarbazina/análogos & derivados , Dacarbazina/farmacologia , Relação Dose-Resposta a Droga , Resistencia a Medicamentos Antineoplásicos , Sinergismo Farmacológico , Glicólise/efeitos dos fármacos , Humanos , Imidazóis/química , Imidazóis/toxicidade , Concentração Inibidora 50 , Neoplasias/metabolismo , Neoplasias/patologia , Compostos Organometálicos/química , Compostos Organometálicos/toxicidade , Estresse Oxidativo/efeitos dos fármacos , Poli(ADP-Ribose) Polimerase-1/antagonistas & inibidores , Poli(ADP-Ribose) Polimerase-1/metabolismo , Inibidores de Poli(ADP-Ribose) Polimerases/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Transdução de Sinais/efeitos dos fármacos , Temozolomida , Tiorredoxina Redutase 1/antagonistas & inibidores , Tiorredoxina Redutase 1/metabolismo , Inibidores da Topoisomerase I/farmacologia , Inibidores da Topoisomerase II/farmacologia
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