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1.
Inorg Chem ; 59(8): 5662-5673, 2020 Apr 20.
Artigo em Inglês | MEDLINE | ID: mdl-32255617

RESUMO

A family of stable anticancer gold(III)-based therapeutic complexes containing cyclometalated triphenylphosphine sulfide ligands have been prepared. The anticancer properties of the newly developed complexes [AuCl2{κ2-2-C6H4P(S)Ph2}] (1), [Au(κ2-S2CNEt2){κ2-2-C6H4P(S)Ph2}]PF6 (2), [AuCl(dppe){κC-2-C6H4P(S)Ph2}]Cl (3), and [Au(dppe){κ2-2-C6H4P(S)Ph2}][PF6]2 (4) were investigated toward five human cancer cell lines [cervical (HeLa), lung (A549), prostate (PC3), fibrosarcoma (HT1080), and breast (MDA-MB-231)]. In vitro cytotoxicity studies revealed that compounds 2-4 displayed potent cell growth inhibition (IC50 values in the range of 0.17-2.50 µM), comparable to, or better than, clinically used cisplatin (0.63-6.35 µM). Preliminary mechanistic studies using HeLa cells indicate that the cytotoxic effects of the compounds involve apoptosis induction through ROS accumulation. Compound 2 also demonstrated significant inhibition of endothelial cell migration and tube formation in the angiogenesis process. Evaluation of the in vivo antitumor activity of compound 2 in nude mice bearing cervical cancer cell (HeLa) xenografts indicated significant tumor growth inhibition (55%) with 1 mg/kg dose (every 3 days) compared with the same dose of cisplatin (28%). These results demonstrate the potential of gold(III) complexes containing cyclometalated triphenylphosphine sulfide ligands as novel metal-based anticancer agents.


Assuntos
Inibidores da Angiogênese/uso terapêutico , Complexos de Coordenação/uso terapêutico , Neoplasias/tratamento farmacológico , Fosfinas/uso terapêutico , Sulfetos/uso terapêutico , Inibidores da Angiogênese/síntese química , Animais , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Movimento Celular/efeitos dos fármacos , Complexos de Coordenação/síntese química , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/uso terapêutico , Feminino , Ouro/química , Humanos , Ligantes , Camundongos Endogâmicos BALB C , Camundongos Nus , Fosfinas/síntese química , Espécies Reativas de Oxigênio/metabolismo , Sulfetos/síntese química , Tiorredoxina Dissulfeto Redutase/antagonistas & inibidores , Ensaios Antitumorais Modelo de Xenoenxerto
3.
Chemistry ; 26(31): 7092-7108, 2020 Jun 02.
Artigo em Inglês | MEDLINE | ID: mdl-32037581

RESUMO

Many cancer cells critically rely on antioxidant systems for cell survival and are vulnerable to further oxidative impairment triggered by agents generating reactive oxygen species (ROS). Therefore, the classical design and development of inhibitors that target antioxidant defense enzymes such as thioredoxin reductase (TrxR) can be a promising anticancer strategy. Herein, it is shown that a gold(I) complex containing an oleanolic acid derivative (4 b) induces apoptosis of ovarian cancer A2780 cells by activating endoplasmic reticulum stress (ERS). It can inhibit TrxR enzyme activity to elevate ROS, mediate ERS and mitochondrial dysfunction, and finally leads to cell cycle arrest and apoptosis of A2780 cells. Notably, this complex inhibits A2780 xenograft tumor growth accompanied by increased ERS level and decreased TrxR activity in tumor tissues.


Assuntos
Antineoplásicos/farmacologia , Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Complexos de Coordenação/farmacologia , Ouro/química , Ácido Oleanólico/química , Neoplasias Ovarianas/tratamento farmacológico , Espécies Reativas de Oxigênio/metabolismo , Tiorredoxina Dissulfeto Redutase/antagonistas & inibidores , Antineoplásicos/química , Pontos de Checagem do Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Complexos de Coordenação/química , Estresse do Retículo Endoplasmático , Feminino , Ouro/farmacologia , Humanos , Oxirredução , Tiorredoxina Dissulfeto Redutase/química
4.
Chemistry ; 26(45): 10175-10184, 2020 Aug 12.
Artigo em Inglês | MEDLINE | ID: mdl-32097513

RESUMO

The thioredoxin system is highly conserved system found in all living cells and comprises NADPH, thioredoxin, and thioredoxin reductase. This system plays a critical role in preserving a reduced intracellular environment, and its involvement in regulating a wide range of cellular functions makes it especially vital to cellular homeostasis. Its critical role is not limited to healthy cells, it is also involved in cancer development, and is overexpressed in many cancers. This makes the thioredoxin system a promising target for cancer drug development. As such, over the last decade, many inhibitors have been developed that target the thioredoxin system, most of which are small molecules targeting the thioredoxin reductase C-terminal redox center. A few inhibitors of thioredoxin have also been developed. We believe that more efforts should be invested in developing protein/peptide-based inhibitors against both thioredoxin reductase and/or thioredoxin.


Assuntos
Antineoplásicos/farmacologia , NADP/química , Neoplasias/tratamento farmacológico , Tiorredoxina Dissulfeto Redutase/metabolismo , Tiorredoxinas/uso terapêutico , Antineoplásicos/química , Desenvolvimento de Medicamentos , Humanos , Oxirredução , Tiorredoxina Dissulfeto Redutase/química , Tiorredoxinas/metabolismo
5.
J Enzyme Inhib Med Chem ; 35(1): 506-510, 2020 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31928252

RESUMO

The hypothesis that sulfocoumarin acting as inhibitors of human carbonic anhydrase (CA, EC 4.2.1.1) cancer-associated isoforms hCA IX and - hCA XII is being able to also inhibit thioredoxin reductase was verified and confirmed. The dual targeting of two cancer cell defence mechanisms, i.e. hypoxia and oxidative stress, may both contribute to the observed antiproliferative profile of these compounds against many cancer cell lines. This unprecedented dual anticancer mechanism may lead to a new approach for designing innovative therapeutic agents.


Assuntos
Antineoplásicos/farmacologia , Anidrase Carbônica IX/antagonistas & inibidores , Anidrases Carbônicas/metabolismo , Cumarínicos/farmacologia , Inibidores Enzimáticos/farmacologia , Tiorredoxina Dissulfeto Redutase/antagonistas & inibidores , Antígenos de Neoplasias/metabolismo , Antineoplásicos/síntese química , Antineoplásicos/química , Anidrase Carbônica IX/metabolismo , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Cumarínicos/síntese química , Cumarínicos/química , Relação Dose-Resposta a Droga , Ensaios de Seleção de Medicamentos Antitumorais , Inibidores Enzimáticos/síntese química , Inibidores Enzimáticos/química , Humanos , Isoenzimas/antagonistas & inibidores , Isoenzimas/metabolismo , Células MCF-7 , Estrutura Molecular , Relação Estrutura-Atividade , Tiorredoxina Dissulfeto Redutase/metabolismo , Células Tumorais Cultivadas
6.
Parasitol Res ; 119(2): 695-711, 2020 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-31907668

RESUMO

Amoebiasis is caused by the protozoan Entamoeba histolytica that affects millions of people throughout the world. The standard treatment is metronidazole, however, this drug causes several side effects, and is also mutagenic and carcinogenic. Therefore, the search for therapeutic alternatives is necessary. Quinoxaline 1,4-di-N-oxides (QdNOs) derivatives have been shown to exhibit activity against different protozoan. In the present study, the effects of esters of quinoxaline-7-carboxylate 1,4-di-N-oxide (7-carboxylate QdNOs) derivatives on E. histolytica proliferation, morphology, ultrastructure, and oxidative stress were evaluated, also their potential as E. histolytica thioredoxin reductase (EhTrxR) inhibitors was analyzed. In vitro tests showed that 12 compounds from n-propyl and isopropyl series, were more active (IC50 = 0.331 to 3.56 µM) than metronidazole (IC50 = 4.5 µM). The compounds with better biological activity have a bulky, trifluoromethyl and isopropyl group at R1-, R2-, and R3-position, respectively. The main alterations found in trophozoites treated with some of these compounds included changes in chromatin, cell granularity, redistribution of vacuoles with cellular debris, and an increase in reactive oxygen species. Interestingly, docking studies suggested that 7-carboxylate QdNOs derivatives could interact with amino acid residues of the NADPH-binding domain and/or the redox-active site of EhTrxR. Enzymatic assays demonstrated that selected 7-carboxylate QdNOs inhibits EhTrxR disulfide reductase activity, and diaphorase activity shows that these compounds could act as electron acceptor substrates for the enzyme. Taken together, these data indicate that among the mechanisms involved in the antiamoebic effect of the 7-carboxylate QdNOs derivatives studied, is the induction of oxidative stress and the inhibition of EhTrxR activity.


Assuntos
Entamoeba histolytica/efeitos dos fármacos , Quinoxalinas/farmacologia , Tiorredoxina Dissulfeto Redutase/antagonistas & inibidores , Óxidos N-Cíclicos , Entamoeba histolytica/enzimologia , Ésteres , Humanos , Metronidazol/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Quinolinas , Espécies Reativas de Oxigênio/metabolismo
7.
Psychopharmacology (Berl) ; 237(1): 127-136, 2020 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-31473777

RESUMO

RATIONALE: Selective serotonin reuptake inhibitors (SSRIs) and serotonin-norepinephrine reuptake inhibitors (SNRIs) are the most commonly used drugs for the treatment of depression. Studies have shown that chronic treatment with SSRIs and SNRIs produces a protective effect against oxidative stress. Thioredoxin (Trx) is an antioxidant protein that reverses protein cysteine oxidation and facilitates scavenging reactive oxygen species. OBJECTIVES: The current study is to determine whether the SSRI fluoxetine and the SNRI venlafaxine regulate Trx and protect neuronal cells against protein cysteine oxidation. METHODS: HT22 mouse hippocampal cells were incubated with fluoxetine or venlafaxine for 5 days. Protein levels of Trx, Trx reductase (TrxR), and Trx-interacting protein (Txnip) were measured by immunoblotting analysis. Trx and TrxR activities were analyzed by spectrophotometric method. Protein cysteine sulfenylation was measured by dimedone-conjugation assay, while nitrosylation was measured by biotin-switch assay. RESULTS: We found that treatment with fluoxetine or venlafaxine for 5 days increased Trx and TrxR protein levels but produced no effect on Txnip protein levels. These treatments also increased Trx and TrxR activities. Although treatment with fluoxetine or venlafaxine alone had no effect on sulfenylated and nitrosylated protein levels, both drugs inhibited H2O2-increased sulfenylated protein levels and nitric oxide donor nitrosoglutathione-increased nitrosylated protein levels. Stress increases risk of depression. We also found that treatment with fluoxetine or venlafaxine for 5 days inhibited stress hormone corticosterone-increased total sulfenylated and nitrosylated protein levels. CONCLUSIONS: Our findings suggest that chronic treatment with antidepressants may upregulate Trx, subsequently inhibiting protein sulfenylation and nitrosylation, which may contribute to the protective effect of antidepressants against oxidative stress. Our findings also indicate that thioredoxin is a potential therapeutic target for the treatment of depression.


Assuntos
Fluoxetina/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Tiorredoxinas/metabolismo , Regulação para Cima/efeitos dos fármacos , Cloridrato de Venlafaxina/farmacologia , Animais , Antidepressivos/farmacologia , Proteínas de Transporte/metabolismo , Linhagem Celular , Hipocampo/efeitos dos fármacos , Hipocampo/metabolismo , Camundongos , Neurônios/efeitos dos fármacos , Neurônios/metabolismo , Espécies Reativas de Oxigênio/metabolismo , Inibidores de Captação de Serotonina/farmacologia , Tiorredoxina Dissulfeto Redutase/metabolismo
8.
Int J Cancer ; 146(1): 123-136, 2020 01 01.
Artigo em Inglês | MEDLINE | ID: mdl-31090219

RESUMO

Triple-negative breast cancer (TNBCs) is a very aggressive and lethal form of breast cancer with no effective targeted therapy. Neoadjuvant chemotherapies and radiotherapy remains a mainstay of treatment with only 25-30% of TNBC patients responding. Thus, there is an unmet clinical need to develop novel therapeutic strategies for TNBCs. TNBC cells have increased intracellular oxidative stress and suppressed glutathione, a major antioxidant system, but still, are protected against higher oxidative stress. We screened a panel of antioxidant genes using the TCGA and METABRIC databases and found that expression of the thioredoxin pathway genes is significantly upregulated in TNBC patients compared to non-TNBC patients and is correlated with adverse survival outcomes. Treatment with auranofin (AF), an FDA-approved thioredoxin reductase inhibitor caused specific cell death and impaired the growth of TNBC cells grown as spheroids. Furthermore, AF treatment exerted a significant in vivo antitumor activity in multiple TNBC models including the syngeneic 4T1.2 model, MDA-MB-231 xenograft and patient-derived tumor xenograft by inhibiting thioredoxin redox activity. We, for the first time, showed that AF increased CD8+Ve T-cell tumor infiltration in vivo and upregulated immune checkpoint PD-L1 expression in an ERK1/2-MYC-dependent manner. Moreover, combination of AF with anti-PD-L1 antibody synergistically impaired the growth of 4T1.2 primary tumors. Our data provide a novel therapeutic strategy using AF in combination with anti-PD-L1 antibody that warrants further clinical investigation for TNBC patients.


Assuntos
Anticorpos/uso terapêutico , Auranofina/uso terapêutico , Antígeno B7-H1/imunologia , Inibidores Enzimáticos/uso terapêutico , Tiorredoxina Dissulfeto Redutase/antagonistas & inibidores , Neoplasias de Mama Triplo Negativas/tratamento farmacológico , Animais , Auranofina/farmacologia , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Inibidores Enzimáticos/farmacologia , Feminino , Humanos , Camundongos , Camundongos Endogâmicos BALB C , Espécies Reativas de Oxigênio/metabolismo , Neoplasias de Mama Triplo Negativas/metabolismo , Neoplasias de Mama Triplo Negativas/patologia , Ensaios Antitumorais Modelo de Xenoenxerto
9.
Proc Natl Acad Sci U S A ; 117(1): 741-751, 2020 01 07.
Artigo em Inglês | MEDLINE | ID: mdl-31871212

RESUMO

Seeds preserve a far developed plant embryo in a quiescent state. Seed metabolism relies on stored resources and is reactivated to drive germination when the external conditions are favorable. Since the switchover from quiescence to reactivation provides a remarkable case of a cell physiological transition we investigated the earliest events in energy and redox metabolism of Arabidopsis seeds at imbibition. By developing fluorescent protein biosensing in intact seeds, we observed ATP accumulation and oxygen uptake within minutes, indicating rapid activation of mitochondrial respiration, which coincided with a sharp transition from an oxidizing to a more reducing thiol redox environment in the mitochondrial matrix. To identify individual operational protein thiol switches, we captured the fast release of metabolic quiescence in organello and devised quantitative iodoacetyl tandem mass tag (iodoTMT)-based thiol redox proteomics. The redox state across all Cys peptides was shifted toward reduction from 27.1% down to 13.0% oxidized thiol. A large number of Cys peptides (412) were redox switched, representing central pathways of mitochondrial energy metabolism, including the respiratory chain and each enzymatic step of the tricarboxylic acid (TCA) cycle. Active site Cys peptides of glutathione reductase 2, NADPH-thioredoxin reductase a/b, and thioredoxin-o1 showed the strongest responses. Germination of seeds lacking those redox proteins was associated with markedly enhanced respiration and deregulated TCA cycle dynamics suggesting decreased resource efficiency of energy metabolism. Germination in aged seeds was strongly impaired. We identify a global operation of thiol redox switches that is required for optimal usage of energy stores by the mitochondria to drive efficient germination.


Assuntos
Arabidopsis/fisiologia , Ciclo do Ácido Cítrico/fisiologia , Germinação/fisiologia , Mitocôndrias/metabolismo , Sementes/metabolismo , Trifosfato de Adenosina/metabolismo , Proteínas de Arabidopsis/genética , Proteínas de Arabidopsis/metabolismo , Glutationa Redutase/genética , Glutationa Redutase/metabolismo , Oxirredução , Oxigênio/metabolismo , Plantas Geneticamente Modificadas , Proteômica/métodos , Sementes/citologia , Sementes/crescimento & desenvolvimento , Tiorredoxina h/genética , Tiorredoxina h/metabolismo , Tiorredoxina Dissulfeto Redutase/genética , Tiorredoxina Dissulfeto Redutase/metabolismo
10.
Chem Commun (Camb) ; 56(2): 297-300, 2019 Dec 19.
Artigo em Inglês | MEDLINE | ID: mdl-31808475

RESUMO

Here we report the design and synthesis of a chlorambucil-alkynyl (CHL-CCH) ligand, mononuclear gold(i) complex K[(CHL-CC)AuCl], 1, and heteronuclear complex (CHL-CC)Au(µ2-η2-CS3)Ti(η5-Cp)2, 2 for renal cancer. Complex 2 is significantly more cytotoxic than complex 1 and cisplatin against renal cancer cells with a high selectivity index value. The mechanism of action of these complexes against renal cancer cells was studied in detail by experimental and computational methods.


Assuntos
Antineoplásicos/farmacologia , Clorambucila/análogos & derivados , Clorambucila/farmacologia , Complexos de Coordenação/farmacologia , Inibidores Enzimáticos/farmacologia , Tiorredoxina Dissulfeto Redutase/antagonistas & inibidores , Antineoplásicos/síntese química , Apoptose/efeitos dos fármacos , Linhagem Celular Tumoral , Clorambucila/síntese química , Complexos de Coordenação/síntese química , Teoria da Densidade Funcional , Desenho de Fármacos , Inibidores Enzimáticos/síntese química , Ouro/química , Humanos , Neoplasias Renais/tratamento farmacológico , Modelos Químicos , Simulação de Acoplamento Molecular , Titânio/química
11.
Artigo em Inglês | MEDLINE | ID: mdl-31801208

RESUMO

Methylmercury (MeHg) is a highly neurotoxic compound to which human populations are exposed via fish consumption. Once in cells, MeHg actively binds thiols and selenols, interfering with the activity of redox enzymes such as thioredoxin (Trx) and the selenoenzyme thioredoxin reductase (TrxR) which integrate the thioredoxin system. In fact, it has been shown that inhibition of this system by MeHg is a critical step in the unfolding of cell death. Current clinical approaches to mitigate the toxicity of MeHg rely on the use of chelators, such as meso-2,3-dimercaptosuccinic acid (DMSA) which largely replaced British anti-Lewisite or 2,3-dimercapto-1-propanol (BAL) as the prime choice. However, therapeutic efficacy is limited and therefore new therapeutic options are necessary. In this work, we evaluated the efficacy of a macrocyclic chelator, 1-thia-4,7,10,13-tetraazacyclopentadecane ([15]aneN4S), in preventing MeHg toxicity, namely by looking at the effects over relevant molecular targets, i.e., the thioredoxin system, using both purified enzyme solutions and cell experiments with human neuroblastoma cells (SH-SY5Y). Results showed that [15]aneN4S had a similar efficacy to DMSA and BAL in reversing the inhibition of MeHg over purified TrxR and Trx by looking at both the 5,5'-dithiobis(2-nitrobenzoic acid) (DTNB) reduction assay and insulin reduction capability. In experiments with cells, none of the chelating agents could reverse the inhibition of TrxR by MeHg, which corroborates the high affinity of MeHg to the selenol in TrxR active site. [15]aneN4S and BAL, unlike DMSA, could prevent inhibition of Trx, which allows the maintenance of downstream functions, although BAL showed higher toxicity to cells. Overall these findings highlight the potential of using [15]aneN4S in the treatment of MeHg poisoning and encourage further studies, namely in vivo.


Assuntos
Compostos Aza/farmacologia , Quelantes/farmacologia , Compostos Macrocíclicos/farmacologia , Compostos de Metilmercúrio/toxicidade , Linhagem Celular Tumoral , Humanos , Tiorredoxina Dissulfeto Redutase/metabolismo , Tiorredoxinas/metabolismo
12.
Biomed Res Int ; 2019: 6502793, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31828114

RESUMO

Nuclear factor erythroid 2-related factor 2 (Nrf2) is involved in tumor drug resistance, but its role in imatinib resistance of chronic myeloid leukemia (CML) remains elusive. We aimed to investigate the effects of Nrf2 on drug sensitivity, thioredoxin reductase (TrxR) expression, reactive oxygen species (ROS) production, and apoptosis induction in imatinib-resistant CML K562/G01 cells and explored their potential mechanisms. Stable K562/G01 cells with knockdown of Nrf2 were established by infection of siRNA-expressing lentivirus. The mRNA and protein expression levels of Nrf2 and TrxR were determined by real-time quantitative polymerase chain reaction and western blot, respectively. ROS generation and apoptosis were assayed by flow cytometry, while drug sensitivity was measured by the Cell Counting Kit-8 assay. Imatinib-resistant K562/G01 cells had higher levels of Nrf2 expression than the parental K562 cells at both mRNA and protein levels. Expression levels of Nrf2 and TrxR were positively correlated in K562/G01 cells. Knockdown of Nrf2 in K562/G01 cells enhanced the intracellular ROS level, suppressed cell proliferation, and increased apoptosis in response to imatinib treatments. Nrf2 expression contributes to the imatinib resistance of K562/G01 cells and is positively correlated with TrxR expression. Targeted inhibition of the Nrf2-TrxR axis represents a potential therapeutic approach for imatinib-resistant CML.


Assuntos
Antineoplásicos/farmacologia , Mesilato de Imatinib/farmacologia , Células K562 , Fator 2 Relacionado a NF-E2/antagonistas & inibidores , Tiorredoxina Dissulfeto Redutase/antagonistas & inibidores , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos/efeitos dos fármacos , Técnicas de Silenciamento de Genes , Humanos , Células K562/efeitos dos fármacos , Células K562/metabolismo , Fator 2 Relacionado a NF-E2/análise , Fator 2 Relacionado a NF-E2/genética , Fator 2 Relacionado a NF-E2/metabolismo , Tiorredoxina Dissulfeto Redutase/análise , Tiorredoxina Dissulfeto Redutase/metabolismo
13.
Mol Med Rep ; 20(5): 4540-4550, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31702035

RESUMO

Rheumatoid arthritis (RA) is characterized by chronic inflammatory synovitis resulting in progressive joint destruction. Persistent synovial inflammation is induced by activation of various inflammatory cells. G­protein­coupled bile acid receptor 1 (TGR5) is a G­protein­coupled receptor activated by various bile acids, which has been reported to act as a key adaptor in regulating various signaling pathways involved in inflammatory responses and a diverse array of physiological processes, including bile acid synthesis, lipid and carbohydrate metabolism, carcinogenesis, immunity and inflammation. In the present study, TGR5 expression was detected in RA peripheral blood mononuclear cells (PBMCs), and its association with clinical disease activity, histological synovitis severity and radiological joint destruction was analyzed. Subsequently, the role and potential underlying mechanisms of TGR5 in the PBMCs of patients with RA and mice with collagen II­induced arthritis (CIA) were investigated. PBMCs were obtained from 50 patients with RA and 40 healthy controls (HCs). The mRNA and protein expression levels of TGR5 were detected in PBMCs via reverse transcription­quantitative polymerase chain reaction (RT­qPCR) and immunofluorescence staining, respectively. Additionally, the levels of proinflammatory cytokines were analyzed by RT­qPCR and enzyme­linked immunosorbent assay (ELISA). The activation of nuclear factor­κB (NF­κB) and IκB kinase a was determined via western blot analysis. The anti­arthritic and anti­inflammatory effects of LCA on mice with CIA were then investigated. The arthritis score was assessed, and the protein levels of proinflammatory cytokines in the plasma of mice were detected via ELISA. TGR5 mRNA expression was significantly downregulated in the PBMCs of patients with RA compared with in those of the HCs (0.53±0.58 for patients vs. 1.49±0.83 for HCs; P<0.001); similar findings were observed at the protein level. The mRNA expression levels of TGR5 in the PBMCs of patients with RA with a high 28­Joint Disease Activity Score (DAS28) were significantly decreased compared with in patients with a low DAS28 (0.81±0.65 for low score vs. 0.35±0.46 for high score; P=0.002). Furthermore, TGR5 expression was significantly correlated with the levels of C­reactive protein (r=­0.429; P=0.002) and the DAS28 (r=­0.383; P=0.006). RT­qPCR and ELISA analyses indicated that lithocholic acid (LCA, 10 mg/kg/day) attenuated lipopolysaccharide­induced proinflammatory cytokine production via inhibition of NF­κB activity in the PBMCs of patients with RA. In addition, the arthritis score was significantly decreased in LCA­treated CIA mice compared with in non­treated CIA mice. The increased production of tumor necrosis factor­α, interleukin (IL)­1ß, IL­6 and IL­8 was significantly reduced in the plasma of LCA­treated CIA mice compared with the control. In conclusion, TGR5 may contribute to the inflammation of PBMCs in patients with RA and mice with CIA.


Assuntos
Artrite Experimental/metabolismo , Artrite Reumatoide/metabolismo , Regulação da Expressão Gênica , Leucócitos Mononucleares/metabolismo , Receptores Acoplados a Proteínas-G/biossíntese , Tiorredoxina Dissulfeto Redutase/biossíntese , Animais , Artrite Experimental/patologia , Artrite Reumatoide/patologia , Citocinas/metabolismo , Feminino , Humanos , Inflamação/metabolismo , Inflamação/patologia , Leucócitos Mononucleares/patologia , Masculino , Camundongos , Pessoa de Meia-Idade
14.
Eur J Med Chem ; 183: 111721, 2019 Dec 01.
Artigo em Inglês | MEDLINE | ID: mdl-31577978

RESUMO

Thioredoxin reductase (TrxR) is often overexpressed in different types of cancer cells including hepatocellular carcinoma (HCC) cells and regarded as a target with great promise for anticancer drug research and development. Here, we have synthesized and characterized nine new designed rhodium(I) N-heterocyclic carbene (NHC) complexes. All of them were effective towards cancer cells, especially complex 1e was more active than cisplatin and manifested strong antiproliferative activity against HCC cells. In vivo anticancer studies showed that 1e significantly repressed tumor growth in an HCC nude mouse model and ameliorated liver lesions in a chronic HCC model caused by CCl4. Notably, a mechanistic study revealed that 1e can strongly inhibit TrxR system both in vitro and in vivo. Furthermore, 1e promoted intracellular ROS accumulation, damaged mitochondrial membrane potential, promoted cancer cell apoptosis and blocked the cells in the G1 phase.


Assuntos
Antineoplásicos/síntese química , Carcinoma Hepatocelular/tratamento farmacológico , Complexos de Coordenação/síntese química , Ciclo-Octanos/química , Imidazóis/química , Neoplasias Hepáticas/tratamento farmacológico , Ródio/química , Tiorredoxina Dissulfeto Redutase/antagonistas & inibidores , Animais , Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Permeabilidade da Membrana Celular , Sobrevivência Celular/efeitos dos fármacos , Complexos de Coordenação/farmacologia , Desenho de Fármacos , Células Hep G2 , Humanos , Masculino , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Camundongos Nus , Estrutura Molecular , Espécies Reativas de Oxigênio/metabolismo , Relação Estrutura-Atividade
15.
Plant Physiol ; 181(3): 976-992, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31527089

RESUMO

NADPH-thioredoxin reductase C (NTRC) forms a separate thiol-reduction cascade in plastids, combining both NADPH-thioredoxin reductase and thioredoxin activities on a single polypeptide. While NTRC is an important regulator of photosynthetic processes in leaves, its function in heterotrophic tissues remains unclear. Here, we focus on the role of NTRC in developing tomato (Solanum lycopersicum) fruits representing heterotrophic storage organs important for agriculture and human diet. We used a fruit-specific promoter to decrease NTRC expression by RNA interference in developing tomato fruits by 60% to 80% compared to the wild type. This led to a decrease in fruit growth, resulting in smaller and lighter fully ripe fruits containing less dry matter and more water. In immature fruits, NTRC downregulation decreased transient starch accumulation, which led to a subsequent decrease in soluble sugars in ripe fruits. The inhibition of starch synthesis was associated with a decrease in the redox-activation state of ADP-Glc pyrophosphorylase and soluble starch synthase, which catalyze the first committed and final polymerizing steps, respectively, of starch biosynthesis. This was accompanied by a decrease in the level of ADP-Glc. NTRC downregulation also led to a strong increase in the reductive states of NAD(H) and NADP(H) redox systems. Metabolite profiling of NTRC-RNA interference lines revealed increased organic and amino acid levels, but reduced sugar levels, implying that NTRC regulates the osmotic balance of developing fruits. These results indicate that NTRC acts as a central hub in regulating carbon metabolism and redox balance in heterotrophic tomato fruits, affecting fruit development as well as final fruit size and quality.


Assuntos
Frutas/enzimologia , Lycopersicon esculentum/enzimologia , Amido/metabolismo , Tiorredoxina Dissulfeto Redutase/metabolismo , Metabolismo dos Carboidratos , Frutas/genética , Frutas/crescimento & desenvolvimento , Frutas/fisiologia , Lycopersicon esculentum/genética , Lycopersicon esculentum/crescimento & desenvolvimento , Lycopersicon esculentum/fisiologia , Metabolômica , Oxirredução , Fotossíntese , Folhas de Planta/enzimologia , Folhas de Planta/genética , Folhas de Planta/crescimento & desenvolvimento , Folhas de Planta/fisiologia , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Interferência de RNA , Tiorredoxina Dissulfeto Redutase/genética
16.
Chem Commun (Camb) ; 55(71): 10627-10630, 2019 Aug 29.
Artigo em Inglês | MEDLINE | ID: mdl-31429450

RESUMO

We report the synthesis of a novel hydroxyl-functionalised heteroleptic bis-NHC gold(i) complex that permits conjugation to various amines via carbamate bond formation. The resulting derivatives were studied in vitro using cell proliferation assays and fluorescent microscopic imaging of human cancer cell lines.


Assuntos
Aminas/química , Antineoplásicos/síntese química , Carbamatos/química , Complexos de Coordenação/síntese química , Corantes Fluorescentes/síntese química , Ouro/química , Células A549 , Antineoplásicos/farmacologia , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Complexos de Coordenação/farmacologia , Doxorrubicina/química , Humanos , Imagem Óptica , Tiorredoxina Dissulfeto Redutase/antagonistas & inibidores , Triazóis/química
17.
J Biol Chem ; 294(38): 14105-14118, 2019 09 20.
Artigo em Inglês | MEDLINE | ID: mdl-31366732

RESUMO

Thioredoxin (Trx) is a conserved, cytosolic reductase in all known organisms. The enzyme receives two electrons from NADPH via thioredoxin reductase (TrxR) and passes them on to multiple cellular reductases via disulfide exchange. Despite the ubiquity of thioredoxins in all taxa, little is known about the functions of resurrected ancestral thioredoxins in the context of a modern mesophilic organism. Here, we report on functional in vitro and in vivo analyses of seven resurrected Precambrian thioredoxins, dating back 1-4 billion years, in the Escherichia coli cytoplasm. Using synthetic gene constructs for recombinant expression of the ancestral enzymes, along with thermodynamic and kinetic assays, we show that all ancestral thioredoxins, as today's thioredoxins, exhibit strongly reducing redox potentials, suggesting that thioredoxins served as catalysts of cellular reduction reactions from the beginning of evolution, even before the oxygen catastrophe. A detailed, quantitative characterization of their interactions with the electron donor TrxR from Escherichia coli and the electron acceptor methionine sulfoxide reductase, also from E. coli, strongly hinted that thioredoxins and thioredoxin reductases co-evolved and that the promiscuity of thioredoxins toward downstream electron acceptors was maintained during evolution. In summary, our findings suggest that thioredoxins evolved high specificity for their sole electron donor TrxR while maintaining promiscuity to their multiple electron acceptors.


Assuntos
Tiorredoxina Dissulfeto Redutase/metabolismo , Tiorredoxinas/metabolismo , Citoplasma/metabolismo , Citosol/metabolismo , Dissulfetos/metabolismo , Escherichia coli/metabolismo , Evolução Molecular , História Antiga , Cinética , NADP/metabolismo , Oxidantes/metabolismo , Oxirredução , Relação Estrutura-Atividade
18.
Int J Nanomedicine ; 14: 4991-5015, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31371943

RESUMO

Purpose: This study evaluates the cytotoxicity of AuNPs coated with polyallylamine (AuNPs-PAA) and conjugated or not to the epidermal growth factor receptor (EGFR)-targeting antibody Cetuximab (AuNPs-PAA-Ctxb) in normal human kidney (HK-2), liver (THLE-2) and microvascular endothelial (TIME) cells, and compares it with two cancer cell lines that are EGFR-overexpressing (A431) or EGFR-negative (MDA-MB-453). Results: Conjugation of Cetuximab to AuNPs-PAA increased the AuNPs-PAA-Ctxb interactions with cells, but reduced their cytotoxicity. TIME cells exhibited the strongest reduction in viability after exposure to AuNPs-PAA(±Ctxb), followed by THLE-2, MDA-MB-453, HK-2 and A431 cells. This cell type-dependent sensitivity was strongly correlated to the inhibition of thioredoxin reductase (TrxR) and glutathione reductase (GR), and to the depolarization of the mitochondrial membrane potential. Both are suggested to initiate apoptosis, which was indeed detected in a concentration- and time-dependent manner. The role of oxidative stress in AuNPs-PAA(±Ctxb)-induced cytotoxicity was demonstrated by co-incubation of the cells with N-acetyl L-cysteine (NAC), which significantly decreased apoptosis and mitochondrial membrane depolarization. Conclusion: This study helps to identify the cells and tissues that could be sensitive to AuNPs and deepens the understanding of the risks associated with the use of AuNPs in vivo.


Assuntos
Antioxidantes/metabolismo , Ouro/química , Nanopartículas Metálicas/química , Acetilcisteína/farmacologia , Apoptose/efeitos dos fármacos , Caspase 3/metabolismo , Caspase 7/metabolismo , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Cetuximab/farmacologia , Endocitose/efeitos dos fármacos , Glutationa Redutase/metabolismo , Humanos , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Nanopartículas Metálicas/ultraestrutura , Mitocôndrias/efeitos dos fármacos , Mitocôndrias/metabolismo , Tamanho da Partícula , Poliaminas/química , Substâncias Protetoras/farmacologia , Eletricidade Estática , Tiorredoxina Dissulfeto Redutase/metabolismo
19.
Toxicol In Vitro ; 61: 104590, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31279089

RESUMO

The thioredoxin (Trx) system controls cellular redox in vascular smooth muscle cells. The present study investigated the roles of Trx1 and Trx reductase1 (TrxR1) proteins in regulation of cell growth, death, reactive oxygen species (ROS) and glutathione (GSH) levels in hydrogen peroxide (H2O2)-treated human pulmonary artery smooth muscle (HPASM) cells. H2O2 induced growth inhibition and cell death in HPASM cells over 24 h. Overexpression of Trx1 and TrxR1 using adenoviruses significantly weakened cell growth inhibition and cell death caused by H2O2. Increases in ROS levels including mitochondrial superoxide anion (O2•-) were observed as early as 5-30 min after H2O2 addition. Administration of adTrxR1 attenuated H2O2-induced increases in ROS levels at 30-180 min. adTrx1 and adTrxR1 significantly reduced the increases in O2•- level in H2O2-treated HPASM cells at 24 h. Furthermore, HPASM cells transfected with Trx1 or TrxR1 siRNA showed increases in ROS levels with or without H2O2 at 5 min. While H2O2 transiently decreased GSH level at 5 min, Trx1 and TrxR1 siRNA intensified the decrease in GSH level. In conclusion, upregulation of Trx1 and TrxR1 significantly attenuated cell growth inhibition and death in H2O2-treated HPASM cells. As a whole, Trx-related adenoviruses diminished H2O2-induced ROS level in HPASM cells whereas Trx-related siRNAs increased ROS levels and decreased GSH level in these cells.


Assuntos
Peróxido de Hidrogênio/toxicidade , Miócitos de Músculo Liso/efeitos dos fármacos , Tiorredoxina Dissulfeto Redutase/metabolismo , Tiorredoxinas/metabolismo , Morte Celular/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Glutationa/metabolismo , Humanos , Miócitos de Músculo Liso/metabolismo , Artéria Pulmonar/citologia , RNA Interferente Pequeno/administração & dosagem , Espécies Reativas de Oxigênio/metabolismo , Tiorredoxina Dissulfeto Redutase/genética , Tiorredoxinas/genética , Transfecção , Regulação para Cima
20.
J Biol Chem ; 294(36): 13336-13343, 2019 09 06.
Artigo em Inglês | MEDLINE | ID: mdl-31320475

RESUMO

Dynamic control of thioredoxin (Trx) oxidoreductase activity is essential for balancing the need of cells to rapidly respond to oxidative/nitrosative stress and to temporally regulate thiol-based redox signaling. We have previously shown that cytokine stimulation of the respiratory epithelium induces a precipitous decline in cell S-nitrosothiol, which depends upon enhanced Trx activity and proteasome-mediated degradation of Txnip (thioredoxin-interacting protein). We now show that tumor necrosis factor-α-induced Txnip degradation in A549 respiratory epithelial cells is regulated by the extracellular signal-regulated protein kinase (ERK) mitogen-activated protein kinase pathway and that ERK inhibition augments both intracellular reactive oxygen species and S-nitrosothiol. ERK-dependent Txnip ubiquitination and proteasome degradation depended upon phosphorylation of a PXTP motif threonine (Thr349) located within the C-terminal α-arrestin domain and proximal to a previously characterized E3 ubiquitin ligase-binding site. Collectively, these findings demonstrate the ERK mitogen-activated protein kinase pathway to be integrally involved in regulating Trx oxidoreductase activity and that the regulation of Txnip lifetime via ERK-dependent phosphorylation is an important mediator of this effect.


Assuntos
Proteínas de Transporte/metabolismo , MAP Quinases Reguladas por Sinal Extracelular/metabolismo , Complexo de Endopeptidases do Proteassoma/metabolismo , Tiorredoxina Dissulfeto Redutase/metabolismo , Células A549 , Humanos , Espectrometria de Massas , Células Tumorais Cultivadas
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