RESUMO
A series of 3-aryl-naphtho[2,3-d]isoxazole-4,9-diones and some of their 6-aza analogues were synthesized and found to inhibit the heat shock protein 90 (Hsp90). The compounds were tested for their binding to Hsp90 and for their effects on Hsp90 client proteins expression in a series of human tumour cell lines. Representative compounds (7f, 10c) downregulated the Hsp90 client proteins EGFR, Akt, Cdk4, Raf-1, and survivin, and upregulated Hsp70. Most of the compounds, in particular the alkylated 3-pyridyl derivatives, exhibited potent antiproliferative activity, down to two-digit nanomolar range. Preliminary results indicated in vivo activity of 7f against human epithelial carcinoma A431 model growing as tumour xenograft in nude mice, thus supporting the therapeutic potential of this novel series of Hsp90 inhibitors.
Assuntos
Antineoplásicos/química , Antineoplásicos/farmacologia , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Isoxazóis/química , Naftoquinonas/química , Naftoquinonas/farmacologia , Animais , Antineoplásicos/síntese química , Linhagem Celular Tumoral , Feminino , Proteínas de Choque Térmico HSP90/química , Humanos , Concentração Inibidora 50 , Camundongos , Modelos Moleculares , Naftoquinonas/síntese química , Conformação Proteica , Relação Quantitativa Estrutura-Atividade , Ensaios Antitumorais Modelo de XenoenxertoRESUMO
A series of mold metabolites of Ascomycetes, structurally belonging to the class of azaphilones, were found to inhibit the heat shock protein Hsp90. In particular, bulgarialactone B was tested for its binding to Hsp90 using surface plasmon resonance and limited proteolysis assays and for its effects on Hsp90 client proteins expression in a series of human tumor cell lines. This compound showed high affinity for Hsp90, interacting with the 90-280 region of the N-terminal domain and down-regulated the Hsp90 client proteins Raf-1, survivin, Cdk4, Akt, and EGFR. Bulgarialactone B and other natural azaphilones showed antiproliferative activity in a panel of human tumor cell lines; their conversion into semisynthetic derivatives by reaction with primary amines increased the antiproliferative activity. Preliminary results indicated in vivo activity of bulgarialactone B against an ascitic ovarian carcinoma xenograft, thus supporting the therapeutic potential of this novel series of Hsp90 inhibitors.
Assuntos
Antineoplásicos/química , Antineoplásicos/uso terapêutico , Ascomicetos/química , Benzopiranos/química , Benzopiranos/uso terapêutico , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Pigmentos Biológicos/química , Pigmentos Biológicos/uso terapêutico , Animais , Antineoplásicos/isolamento & purificação , Antineoplásicos/farmacologia , Benzopiranos/isolamento & purificação , Benzopiranos/farmacologia , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Feminino , Proteínas de Choque Térmico HSP90/metabolismo , Humanos , Camundongos , Camundongos Nus , Neoplasias/tratamento farmacológico , Pigmentos Biológicos/isolamento & purificação , Pigmentos Biológicos/farmacologiaRESUMO
The naturally occurring coumarins and resveratrol, attract great attention due to their wide range of biological properties, including anticancer, antileukemic, antibacterial and anti-inflammatory activities; moreover, their cancer chemopreventive property have been recently emphasized. A novel class of hybrid compounds, obtained by introducing a substituted trans-vinylbenzene moiety on a coumarin backbone, was synthesized and evaluated for the antitumor profile. A number of derivatives showed a good antiproliferative activity, in some cases higher to that of the reference compound resveratrol. The most promising compounds in this series were 14 and 17, endowed with excellent antiproliferative and proapoptotic activities. The present study suggests that the 7-methoxycoumarin nucleus, together with the 3,5-disubstitution pattern of the trans-vinylbenzene moiety, are likely promising structural features to obtain excellent antitumor compounds endowed with a apoptosis-inducing capability.
Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Estilbenos/farmacologia , Anticarcinógenos , Antineoplásicos/química , Antineoplásicos Fitogênicos/farmacologia , Antioxidantes , Linhagem Celular Tumoral , Quimera , Cumarínicos/química , Cumarínicos/farmacologia , Desenho de Fármacos , Avaliação Pré-Clínica de Medicamentos , Resistencia a Medicamentos Antineoplásicos , Ensaios de Seleção de Medicamentos Antitumorais , Humanos , Estrutura Molecular , Óxidos/farmacologia , Resveratrol , Estilbenos/química , Relação Estrutura-AtividadeRESUMO
Curcumin, a natural polyphenol, has been described to exhibit effects on signaling pathways, leading to induction of apoptosis. In this study, we observed that curcumin inhibited Hsp90 activity causing depletion of client proteins implicated in survival pathways. Based on this observation, this study was designed to investigate the cellular effects of curcumin combination with the pan-HDAC inhibitors, vorinostat and panobinostat, which induce hyperacetylation of Hsp90, resulting in inhibition of its chaperone function. The results showed that, at subtoxic concentrations, curcumin markedly sensitized tumor cells to vorinostat- and panobinostat-induced growth inhibition and apoptosis. The sensitization was associated with persistent depletion of Hsp90 client proteins (EGFR, Raf-1, Akt, and survivin). In conclusion, our findings document a novel mechanism of action of curcumin and support the therapeutic potential of curcumin/HDAC inhibitors combination, because the synergistic interaction was observed at pharmacologically achievable concentrations, which were ineffective when each drug was used alone.
Assuntos
Apoptose/efeitos dos fármacos , Curcumina/farmacologia , Proteínas de Choque Térmico HSP90/antagonistas & inibidores , Inibidores de Histona Desacetilases/farmacologia , Ácidos Hidroxâmicos/farmacologia , Ligação Competitiva , Proliferação de Células/efeitos dos fármacos , Regulação para Baixo/efeitos dos fármacos , Proteínas de Choque Térmico HSP90/metabolismo , Desacetilase 6 de Histona , Histona Desacetilases/metabolismo , Humanos , Indóis , Panobinostat , Relação Estrutura-Atividade , Células Tumorais Cultivadas , VorinostatRESUMO
A number of recent observations have suggested a potential role for membrane-bound gamma-glutamyltransferase (GGT) in tumor progression and appearance of more aggressive and resistant phenotypes, through redox interactions leading to production of reactive oxygen species. The present study was aimed to evaluate whether such pro-oxidant activity of GGT can promote oxidative DNA damage, thus contributing to cancer genomic instability. Human GGT-transfected melanoma cells were studied, and DNA damage was measured by using the alkaline comet assay. Our results indicate that higher levels of GGT activity are associated with higher levels of background DNA damage and oxidized bases. This association cannot be explained by differences in cell cycle distribution or apoptotic rates. GGT-over-expressing cells also presented with a markedly higher glucose uptake, a phenomenon potentially leading to higher metabolic rate and oxidative DNA damage. Anyway, when GGT-over-expressing cells were incubated in the presence of GGT substrates and a source of catalytic iron, increased levels of DNA damage and oxidized bases were observed, an effect completely prevented in the presence of GGT inhibitors or various antioxidants.The findings reported indicate that GGT activity is able to promote iron-dependent DNA oxidative damage, thus potentially representing an important mechanism in initiation/progression of neoplastic transformation.
Assuntos
Membrana Celular/enzimologia , Dano ao DNA , DNA de Neoplasias/efeitos dos fármacos , Compostos Férricos/farmacologia , Melanoma/enzimologia , Melanoma/genética , gama-Glutamiltransferase/metabolismo , Antioxidantes/farmacologia , Apoptose/efeitos dos fármacos , Ciclo Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Cloretos , Ensaio Cometa , Fibrinolíticos/farmacologia , Humanos , Melanoma/patologia , Oxirredução , Estresse Oxidativo , Espécies Reativas de Oxigênio/metabolismoRESUMO
The cell ability of tumor cells to tolerate stress conditions is a typical feature of solid tumors. In particular, the resistance to oxidative stress of melanoma cells likely contributes to their intrinsic drug resistance. In an attempt to develop novel strategies for overcoming the mechanisms of cellular protection against oxidative stress, in this study we have explored the efficacy of the combination of two prooxidant agents in two human melanoma cell clones. The selected clones are characterized by a marked difference in expression of gamma-glutamyltransferase, which is known to produce a persistent low level of oxidative stress resulting in the stimulation of protective systems. The gamma-glutamyltransferase-overexpressing clone exhibited a low susceptibility to arsenic trioxide-induced apoptosis, associated with low reactive oxygen species induction and increased catalase activity. The combination of arsenic trioxide with subtoxic concentrations of ascorbic acid resulted in a sensitization to apoptotic cell death. The expression of protective mechanisms, in particular catalase activity, accounted for the behavior of the resistant clone. The sensitization achieved by the combination was associated with a cellular response involving the ASK1/p38 axis, which is implicated in the regulation of catalase expression and the activation of apoptotic signals. In conclusion, the results of our study provide evidence that a rational combination of prooxidant agents may be effective in overcoming cellular tolerance to oxidative stress.
Assuntos
Arsenicais/uso terapêutico , Ácido Ascórbico/metabolismo , Inibidores do Crescimento/uso terapêutico , Melanoma/tratamento farmacológico , Óxidos/uso terapêutico , gama-Glutamiltransferase/metabolismo , Apoptose , Trióxido de Arsênio , Catalase/metabolismo , Linhagem Celular Tumoral , Citoproteção , Resistencia a Medicamentos Antineoplásicos , Ativação Enzimática , Regulação Neoplásica da Expressão Gênica , Humanos , MAP Quinase Quinase Quinase 5/genética , MAP Quinase Quinase Quinase 5/metabolismo , Melanoma/genética , Melanoma/metabolismo , Melanoma/patologia , Estresse Oxidativo , gama-Glutamiltransferase/genéticaRESUMO
The extracellular gamma-glutamyltransferase-mediated metabolism of glutathione has been implicated in prooxidant events which may have impact on cellular functions including drug resistance. This study was performed in two GGT-transfected melanoma clones to explore the hypothesis that GGT expression in tumour cells is implicated in modulation of cell behaviour under stress conditions. Our results show that GGT-overexpression in melanoma cells was associated with resistance to oxidative stress produced by prooxidant agents such as hydrogen peroxide and ascorbic acid. In GGT-overexpressing cells, ability to tolerate oxidative stress was evidenced by the presence of a moderate level of ROS and lack of DNA damage response following treatment with H(2)O(2). Cellular response to oxidative stress induced by ascorbic acid was detectable only in the clone with low GGT activity which also exhibited an increased susceptibility to apoptosis. The increased resistance of the GGT-overexpressing clone was not related to intracellular GSH content but rather to the increased expression of catalase and to a reduced efficiency of iron-mediated formation of toxic free radicals. Taken together, these findings are consistent with a contribution of GGT in the mechanisms of drug resistance, because induction of oxidative stress is a relevant event in the apoptotic response to cytotoxic agents.
Assuntos
Antioxidantes/farmacologia , Ácido Ascórbico/farmacologia , Melanoma/enzimologia , Estresse Oxidativo/fisiologia , gama-Glutamiltransferase/metabolismo , Apoptose/fisiologia , Western Blotting , Catalase/farmacologia , Ciclo Celular , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Citocromos c/metabolismo , Resistencia a Medicamentos Antineoplásicos , Humanos , Peróxido de Hidrogênio/farmacologia , Mitocôndrias/metabolismo , Espécies Reativas de Oxigênio/metabolismoRESUMO
Gamma-glutamyltransferase, a key enzyme of GSH metabolism, can modulate crucial redox-sensitive functions, such as antioxidant/antitoxic defences and cellular proliferative/apoptotic balance, with potential implications in tumour progression and drug resistance. Recent studies have elucidated the mechanisms of GGT involvement in various pathological processes suggesting its potential role as therapeutic target and diagnostic/prognostic marker.