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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.
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
4.
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
5.
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
6.
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
7.
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
8.
Cancer Lett ; 412: 46-58, 2018 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-29037867

RESUMO

Colon cancer is one of the leading causes of cancer-related deaths. A natural sesquiterpene lactone, costunolide (CTD), showed inhibition of cancer development. However, the underlying mechanisms are not known. Here, we have examined the therapeutic activity and novel mechanisms of the anti-cancer activities of CTD in colon cancer cells. Using SPR analysis and enzyme activity assay on recombinant TrxR1 protein, our results show that CTD directly binds and inhibits the activity of TrxR1, which caused enhanced generation of ROS and led to ROS-dependent endoplasmic reticulum stress and cell apoptosis in colon cancer cells. Overexpression of TrxR1 in HCT116 cells reversed CTD-induced cell apoptosis and ROS increase. CTD treatment of mice implanted with colon cancer cells showed tumor growth inhibition and reduced TrxR1 activity and ROS level. In addition, it was observed that TrxR1 was significantly up-regulated in existing colon cancer gene database and clinically obtained colon cancer tissues. Our studies have uncovered the mechanism underlying the biological activity of CTD in colon cancer and suggest that targeting TrxR1 may prove to be beneficial as a treatment option.


Assuntos
Apoptose/efeitos dos fármacos , Neoplasias do Colo/tratamento farmacológico , Sesquiterpenos/farmacologia , Tiorredoxina Redutase 1/antagonistas & inibidores , Fator 4 Ativador da Transcrição/fisiologia , Animais , Antineoplásicos Fitogênicos/farmacologia , Neoplasias do Colo/patologia , Estresse do Retículo Endoplasmático , Feminino , Células HCT116 , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Espécies Reativas de Oxigênio/metabolismo , Sesquiterpenos/metabolismo , Ensaios Antitumorais Modelo de Xenoenxerto
9.
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
10.
Inorg Chem ; 56(22): 14237-14250, 2017 Nov 20.
Artigo em Inglês | MEDLINE | ID: mdl-29095609

RESUMO

We report here on the synthesis of a series of mono- and dinuclear gold(I) complexes exhibiting sulfonated bis(NHC) ligands and novel hydroxylated mono(NHC) Au(I) compounds, which were also examined for their biological activities. Initial cell viability assays show strong antiproliferative activities of the hydroxylated mono(NHC) gold compounds (8 > 9 > 10) against 2008 human ovarian cancer cells even after 1 h incubation. In order to gain insight into the mechanism of biological action of the gold compounds, their effect on the pivotal cellular target seleno-enzyme thioredoxin reductase (TrxR), involved in the maintenance of intracellular redox balance, was investigated in depth. The compounds' inhibitory effects on TrxR and glutathione reductase (GR) were studied comparatively, using either the pure proteins or cancer cell extracts. The results show a strong and selective inhibitory effect of TrxR, specifically for the hydroxyl-functionalized NHC gold(I) complexes (8-10). Valuable information on the gold compounds' molecular reactivity with TrxR was gained using the BIAM (biotin-conjugated iodoacetamide) assay and performing competition experiments by mass spectrometry (MS). In good agreement, both techniques suggest the binding affinity of the mono(NHC) Au(I) complexes toward selenols and thiols. Notably, for the first time, bis-carbene formation from mono-carbenes in buffered solution could be observed by MS, which may provide new insights into the speciation mechanisms of bioactive Au(I) NHC complexes. Furthermore, the compounds' interactions with another relevant in cellulo target, namely telomeric G-quadruplex DNA-a higher-order DNA structure playing key roles in telomere function-was investigated by means of FRET melting assays. The lack of interactions with this type of nucleic acid secondary structure support the idea of selective targeting of the hydrophilic Au(I) NHC compounds toward proteins such as TrxR.


Assuntos
Complexos de Coordenação/farmacologia , Ouro/química , Tiorredoxina Redutase 1/antagonistas & inibidores , Tiorredoxina Redutase 2/antagonistas & inibidores , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Antineoplásicos/farmacologia , Linhagem Celular Tumoral , Complexos de Coordenação/síntese química , Complexos de Coordenação/química , DNA/metabolismo , Estabilidade de Medicamentos , Quadruplex G , Glutationa Redutase/antagonistas & inibidores , Humanos , Interações Hidrofóbicas e Hidrofílicas , Ligantes , Compostos Organoáuricos/química , Ratos , Espécies Reativas de Oxigênio/metabolismo , Prata/química , Solubilidade
11.
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
12.
Inorg Chem ; 56(14): 8562-8579, 2017 Jul 17.
Artigo em Inglês | MEDLINE | ID: mdl-28682069

RESUMO

New gold(I) thiolate complexes have been synthesized and characterized, and their physicochemical properties and anticancer activity have been tested. The coordination of PTA derivatives provides optimal hydrophilicity/lipophilicity properties to the complexes, which present high solution stability. Moreover, the complexes show a high anticancer activity against Caco-2 cells, comparable to that of auranofin, and a very low cytotoxic activity against enterocyte-like differentiated cells. Their activity has been shown to produce cell death by apoptosis and arrest of the cell cycle because of interaction with the reductase enzymes and consequent reactive oxygen species production. Some of these new complexes are also able to decrease the necessary dose of 5-fluorouracil, a drug used for the treatment of colon cancer, by a synergistic mechanism.


Assuntos
Antineoplásicos/farmacologia , Neoplasias do Colo/tratamento farmacológico , Fluoruracila/farmacologia , Compostos Organoáuricos/farmacologia , Animais , Antineoplásicos/síntese química , Antineoplásicos/química , Apoptose/efeitos dos fármacos , Auranofina/farmacologia , Células CACO-2 , Bovinos , Ciclo Celular/efeitos dos fármacos , Ácido Ditionitrobenzoico/química , Estabilidade de Medicamentos , Sinergismo Farmacológico , Humanos , Compostos Organoáuricos/síntese química , Compostos Organoáuricos/química , Espécies Reativas de Oxigênio/metabolismo , Soroalbumina Bovina/metabolismo , Tiorredoxina Redutase 1/antagonistas & inibidores
13.
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
14.
Toxicol Appl Pharmacol ; 329: 58-66, 2017 08 15.
Artigo em Inglês | MEDLINE | ID: mdl-28551108

RESUMO

Karenia brevis, the Florida red tide dinoflagellate produces a suite of neurotoxins known as the brevetoxins. The most abundant of the brevetoxins PbTx-2, was found to inhibit the thioredoxin-thioredoxin reductase system, whereas the PbTx-3 has no effect on this system. On the other hand, PbTx-2 activates the reduction of small disulfides such as 5,5'-dithio-bis-(2-nitrobenzoic acid) by thioredoxin reductase. PbTx-2 has an α, ß-unsaturated aldehyde moiety which functions as an efficient electrophile and selenocysteine conjugates are readily formed. PbTx-2 blocks the inhibition of TrxR by the inhibitor curcumin, whereas curcumin blocks PbTx-2 activation of TrxR. It is proposed that the mechanism of inhibition of thioredoxin reduction is via the formation of a Michael adduct between selenocysteine and the α, ß-unsaturated aldehyde moiety of PbTx-2. PbTx-2 had no effect on the rates of reactions catalyzed by related enzymes such as glutathione reductase, glutathione peroxidase or glutaredoxin.


Assuntos
Inibidores Enzimáticos/toxicidade , Toxinas Marinhas/toxicidade , Oxocinas/toxicidade , Tiorredoxina Redutase 1/antagonistas & inibidores , Animais , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Curcumina/farmacologia , Relação Dose-Resposta a Droga , Humanos , Peroxidação de Lipídeos/efeitos dos fármacos , Oxirredução , Domínios Proteicos , Ratos , Selenocisteína , Tiorredoxina Redutase 1/química , Tiorredoxina Redutase 1/metabolismo , Fatores de Tempo
15.
J Zhejiang Univ Sci B ; 18(5): 373-382, 2017 May.
Artigo em Inglês | MEDLINE | ID: mdl-28471109

RESUMO

It has been reported that Ethaselen shows inhibitory effects on thioredoxin reductase (TrxR) activity and human tumor cell growth. In order to find an efficient way to reverse cisplatin resistance, we investigated the reversal effects of Ethaselen on cisplatin resistance in K562/cisplatin (CDDP) cells that were established by pulse-inducing human erythrocyte leukemic cell line K562, which are fivefold more resistant to cisplatin compared to K562 cells. The morphology and growth showed that the adhesion of K562/CDDP further decreased while the cell volume increased. The proliferation of K562/CDDP is strengthened. The antitumor activities in vitro were assessed by MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay and combination index (CI), showing the significant synergic effects of cisplatin and Ethaselen. Focusing on apoptosis, a series of comparisons was made between K562 and K562/CDDP. Cisplatin induced higher reactive oxygen species (ROS) generation in K562 and subsequently induced the formation of mitochondrial permeability transition pores (PTPs). In addition, cisplatin increased the ratio of Bax to Bcl-2 in K562, which can influence the mitochondrial membrane permeability. PTP formation and mitochondrial membrane permeabilization eventually resulted in the release of cytochrome c and activation of the Caspase pathway. However, these effects were not clearly seen in K562/CDDP, which may be the reason for the acquired CDDP resistance. However, Ethaselen can induce a high level of ROS in K562/CDDP by TrxR activity inhibition and increased ratio of Bax to Bcl-2 in K562/CDDP by nuclear factor κB (NF-κB) suppression, which subsequently induces the release of cytochrome c in K562/CDDP. This response is partly responsible for the reversal of the cisplatin resistance in K562/CDDP cells.


Assuntos
Apoptose/efeitos dos fármacos , Compostos Bicíclicos Heterocíclicos com Pontes/administração & dosagem , Cisplatino/administração & dosagem , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Compostos Organosselênicos/administração & dosagem , Tiorredoxina Redutase 1/antagonistas & inibidores , Tiorredoxina Redutase 1/metabolismo , Antineoplásicos/administração & dosagem , Proliferação de Células/efeitos dos fármacos , Tamanho Celular/efeitos dos fármacos , Relação Dose-Resposta a Droga , Ativação Enzimática/efeitos dos fármacos , Humanos , Células K562 , Terapia de Alvo Molecular/métodos , Resultado do Tratamento
16.
Sci Rep ; 6: 36860, 2016 11 15.
Artigo em Inglês | MEDLINE | ID: mdl-27845427

RESUMO

Thioredoxin reductase 1 (TrxR1) is a pivotal intracellular redox sensor and antioxidant enzyme. On the other hand, overexpression of TrxR1 is closely correlated with the initiation of various tumors including breast cancer, though the detailed mechanism remains unclear. Here we investigated the role of TrxR1 in dysplastic transformation of human breast epithelial cell line MCF-10A induced by chronic oxidative stress. Not surprisingly, sustained exposure to H2O2 significantly augmented the expression and activity of TrxR1 in MCF-10A cells. The dysplastically transformed MCF-10A (MCF-10AT) cells undergoing 8-week H2O2 treatment exhibited a certain degree of malignancy in tumorigenicity evaluation. Moreover, TrxR1 inhibitor ethaselen (BBSKE) could partially reverse some malignant phenotypes including epithelial to mesenchymal transition (EMT) of MCF-10AT as well as MCF-7 cells. Collectively, our results supported the considerable involvement of TrxR1 in the onset of breast cancer and BBSKE may be a promising agent against breast cancer.


Assuntos
Peróxido de Hidrogênio/toxicidade , Estresse Oxidativo/efeitos dos fármacos , Tiorredoxina Redutase 1/metabolismo , Adulto , Animais , Neoplasias da Mama/metabolismo , Neoplasias da Mama/patologia , Compostos Bicíclicos Heterocíclicos com Pontes/farmacologia , Caderinas/metabolismo , Linhagem Celular , Movimento Celular/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Epiteliais/citologia , Células Epiteliais/metabolismo , Transição Epitelial-Mesenquimal/efeitos dos fármacos , Feminino , Doença da Mama Fibrocística/patologia , Humanos , Queratina-7/metabolismo , Células MCF-7 , Camundongos , Camundongos Nus , Microscopia Confocal , Pessoa de Meia-Idade , Modelos Biológicos , Compostos Organosselênicos/farmacologia , Espécies Reativas de Oxigênio/metabolismo , Tiorredoxina Redutase 1/antagonistas & inibidores , Tiorredoxina Redutase 1/sangue , Transplante Heterólogo
17.
Free Radic Biol Med ; 101: 53-70, 2016 12.
Artigo em Inglês | MEDLINE | ID: mdl-27693380

RESUMO

Oxidative damage and aggregation of cellular proteins is a hallmark of neuronal cell death after neurotrauma and chronic neurodegenerative conditions. Autophagy and ubiquitin protease system are involved in degradation of protein aggregates, and interruption of their function is linked to apoptotic cell death in these diseases. Oxidative modification of cysteine groups in key molecular proteins has been linked to modification of cellular systems and cell death in these conditions. Glutathione and thioredoxin systems provide reducing protons that can effectively reverse protein modifications and promote cell survival. The central role of Thioredoxin in inhibition of apoptosis is well identified. Additionally, its involvement in initiation of autophagy has been suggested recently. We therefore aimed to investigate the involvement of Thioredoxin system in autophagy-apoptosis processes. A model of serum deprivation in SH-SY5Y was used that is associated with autophagy and apoptosis. Using pharmacological and RNA-editing technology we show that Thioredoxin reductase deficiency in this model enhances oxidative stress and interrupts the early protective autophagy and promotes apoptosis. This was associated with decreased protein-degradation in lysosomes due to altered lysosomal acidification and accumulation of autophagosomes as well as impairment in proteasome pathway. We further confirmed that the extent of oxidative stress is a determining factor in autophagy- apoptosis interplay, as upregulation of cellular reducing capacity by N-acetylcysteine prevented impairment in autophagy and proteasome systems thus promoted cell viability. Our study provides evidence that excessive oxidative stress inhibits protein degradation systems and affects the final stages of autophagy by inhibiting autolysosome maturation: a novel mechanistic link between protein aggregation and conversion of autophagy to apoptosis that can be applicable to neurodegenerative diseases.


Assuntos
Proteína 7 Relacionada à Autofagia/genética , Autofagia/efeitos dos fármacos , Lisossomos/efeitos dos fármacos , Neurônios/metabolismo , Complexo de Endopeptidases do Proteassoma/efeitos dos fármacos , Tiorredoxina Redutase 1/genética , Acetilcisteína/farmacologia , Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Proteína 7 Relacionada à Autofagia/antagonistas & inibidores , Proteína 7 Relacionada à Autofagia/metabolismo , Linhagem Celular Tumoral , Meios de Cultura Livres de Soro/farmacologia , Regulação da Expressão Gênica , Vetores Genéticos/química , Vetores Genéticos/metabolismo , Humanos , Lentivirus/genética , Lentivirus/metabolismo , Lisossomos/metabolismo , Neurônios/efeitos dos fármacos , Neurônios/patologia , Oxirredução , Fagossomos/efeitos dos fármacos , Fagossomos/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Proteólise/efeitos dos fármacos , RNA Interferente Pequeno/genética , RNA Interferente Pequeno/metabolismo , Espécies Reativas de Oxigênio , Tiorredoxina Redutase 1/antagonistas & inibidores , Tiorredoxina Redutase 1/metabolismo
18.
Free Radic Biol Med ; 97: 375-385, 2016 08.
Artigo em Inglês | MEDLINE | ID: mdl-27377780

RESUMO

Mammalian thioredoxin 1 (Trx1) and the selenoprotein Trx reductase 1 (TrxR1) are key cellular enzymes that function coordinately in thiol-based redox regulation and signaling. Recent studies have revealed that the Trx1/TrxR1 system has an S-nitrosothiol reductase (denitrosylase) activity through which it can regulate nitric oxide-related cellular processes. In this study we revealed that TrxR1 is itself susceptible to nitrosylation, characterized the underlying mechanism, and explored its functional significance. We found that nitrosothiol or nitric oxide donating agents rapidly and effectively inhibited the activity of recombinant or endogenous TrxR1. In particular, the NADPH-reduced TrxR1 was partially and reversibly inhibited upon exposure to low concentrations (<10µM) of S-nitrosocysteine (CysNO) and markedly and continuously inhibited at higher doses. Concurrently, TrxR1 very efficiently reduced low, but not high, levels of CysNO. Biochemical and mass spectrometric analyses indicated that its active site selenocysteine residue renders TrxR1 highly susceptible to nitrosylation-mediated inhibition, and revealed both thiol and selenol modifications at the two redox active centers of the enzyme. Studies in HeLa cancer cells demonstrated that endogenous TrxR1 is sensitive to nitrosylation-dependent inactivation and pointed to an important role for glutathione in reversing or preventing this process. Notably, depletion of cellular glutathione with l-buthionine-sulfoximine synergized with nitrosating agents in promoting sustained nitrosylation and inactivation of TrxR1, events that were accompanied by significant oxidation of Trx1 and extensive cell death. Collectively, these findings expand our knowledge of the role and regulation of the mammalian Trx system in relation to cellular nitroso-redox imbalance. The observations raise the possibility of exploiting the nitrosylation susceptibility of TrxR1 for killing tumor cells.


Assuntos
Tiorredoxina Redutase 1/metabolismo , Sequência de Aminoácidos , Animais , Domínio Catalítico , Cisteína/análogos & derivados , Cisteína/química , Cisteína/farmacologia , Glutationa/metabolismo , Células HeLa , Humanos , NADP/química , Doadores de Óxido Nítrico/química , Doadores de Óxido Nítrico/farmacologia , Oxirredução , Processamento de Proteína Pós-Traducional , Ratos , S-Nitrosotióis/química , S-Nitrosotióis/farmacologia , Selenocisteína/química , Tiorredoxina Redutase 1/antagonistas & inibidores , Tiorredoxina Redutase 1/química
19.
J Inorg Biochem ; 160: 296-304, 2016 07.
Artigo em Inglês | MEDLINE | ID: mdl-27130146

RESUMO

The synthesis and the biological effects of two ferrocifen analogs in the osmium series, namely the monophenolic complex 1, the tamoxifen-like complex 2 and their oxidized quinone methide (QM) derivatives, 1-QM and 2-QM, are reported. Inhibition of purified thioredoxin reductase (TrxR) is observed with 1 and 2 only after their enzymatic oxidation by the hydrogen peroxide/horseradish peroxidase (H2O2/HRP) system with IC50 of 2.4 and 1.2µM respectively. However, this inhibition is larger than that obtained with the corresponding quinone methides (IC50=5.4µM for 1-QM and 3.6µM for 2-QM). The UV-Vis spectra of 1 or 2 incubated in the presence of H2O2/HRP show that the species generated is not a quinone methide, but probably the corresponding cation. In Jurkat cells, 2 shows high toxicity (IC50=7.4µM), while 1 is less effective (IC50=42µM). Interestingly, a significant inhibition of TrxR activity is observed in cells incubated with 2 (about 70% inhibition with 15µM) while the inhibition induced by 1 is much weaker (about 30% inhibition with 50µM). This strong inhibition of TrxR by 2 leads to accumulation of thioredoxin and peroxiredoxin 3 in oxidized form and to a decrease of the mitochondrial membrane potential (MMP). These results show that cytotoxicity of the osmocifens depends on their oxidation within the cell and that inhibition of thioredoxin reductase by oxidized species is a key factor in rationalizing the cytotoxicity of these complexes on Jurkat cells.


Assuntos
Antineoplásicos/farmacologia , Complexos de Coordenação/farmacologia , Compostos Ferrosos/farmacologia , Compostos Organometálicos/farmacologia , Tamoxifeno/química , Tiorredoxina Redutase 1/antagonistas & inibidores , Antineoplásicos/síntese química , Morte Celular/efeitos dos fármacos , Complexos de Coordenação/síntese química , Compostos Ferrosos/síntese química , Expressão Gênica , Glutationa/metabolismo , Humanos , Indolquinonas/química , Células Jurkat , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Compostos Organometálicos/síntese química , Osmio/química , Oxirredução , Estresse Oxidativo , Peroxirredoxina III/química , Peroxirredoxina III/metabolismo , Espécies Reativas de Oxigênio/agonistas , Espécies Reativas de Oxigênio/metabolismo , Tiorredoxina Redutase 1/genética , Tiorredoxina Redutase 1/metabolismo , Tiorredoxinas/química , Tiorredoxinas/metabolismo
20.
Cancer Lett ; 375(1): 114-126, 2016 May 28.
Artigo em Inglês | MEDLINE | ID: mdl-26963494

RESUMO

Piperlongumine (PL), a natural alkaloid isolated from the fruit of long pepper, is known to selectively kill tumor cells while sparing their normal counterparts. However, the cellular target and potent anticancer efficacy of PL in numerous types of human cancer cells have not been fully defined. We report here that PL may interact with the thioredoxin reductase 1 (TrxR1), an important selenocysteine (Sec)-containing antioxidant enzyme, to induce reactive oxygen species (ROS)-mediated apoptosis in human gastric cancer cells. By inhibiting TrxR1 activity and increasing intracellular ROS levels, PL induces a lethal endoplasmic reticulum stress and mitochondrial dysfunction in human gastric cancer cells. Importantly, knockdown of TrxR1 sensitizes cells to PL treatment, and PL displays synergistic lethality with GSH inhibitors (BSO and Erastin) against gastric cancer cells. In vivo, PL treatment markedly reduces the TrxR1 activity and tumor cell burden. Remarkably, TrxR1 was significantly overexpressed in gastric cancer cell lines and human gastric cancer tissues. Targeting TrxR1 with PL thus discloses a previously unrecognized mechanism underlying the biological activity of PL and provides an in-depth insight into the action of PL in the treatment of gastric cancer.


Assuntos
Antineoplásicos/farmacologia , Dioxolanos/farmacologia , Neoplasias Gástricas/tratamento farmacológico , Tiorredoxina Redutase 1/antagonistas & inibidores , Animais , Linhagem Celular Tumoral , Estresse do Retículo Endoplasmático , Feminino , Humanos , Potencial da Membrana Mitocondrial , Camundongos Endogâmicos BALB C , Camundongos Nus , Proteínas Proto-Oncogênicas c-bcl-2/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Neoplasias Gástricas/patologia , Carga Tumoral , Ensaios Antitumorais Modelo de Xenoenxerto
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