RESUMO
Benznidazole is the first-line drug used in treating Chagas disease, which is caused by the parasite Trypanosoma cruzi (T. cruzi). However, benznidazole has limited efficacy and several adverse reactions. Pentamidine is an antiprotozoal drug used in the treatment of leishmaniasis and African trypanosomiasis. In T. cruzi, pentamidine blocks the transport of putrescine, a precursor of trypanothione, which constitutes an essential molecule in the resistance of T. cruzi to benznidazole. In the present study, we describe the effect of the combination of benznidazole and pentamidine on isolated parasites, mammalian cells and in mice infected with T. cruzi. In isolated trypomastigotes, we performed a dose-matrix scheme of combinations, where pentamidine antagonized the effect of benznidazole, mainly at concentrations below the EC50 of pentamidine. In T. cruzi-infected mammalian cells, pentamidine reversed the effect of benznidazole (measured by qPCR). In comparison, in infected BALB/c mice, pentamidine failed to get synergy with benznidazole, measured on mice survival, parasitemia and amastigote nest quantification. To further explain the in vitro antagonism, we explored whether pentamidine affects intracellular trypanothione levels, however, pentamidine produced no change in trypanothione concentrations. Finally, the T. cruzi polyamine permease (TcPAT12) was overexpressed in epimastigotes, showing that pentamidine has the same trypanocidal effect, independently of transporter expression levels. These results suggest that, in spite of the high potency in the putrescine transport blockade, TcPAT12 permease is not the main target of pentamidine, and could explain the lack of synergism between pentamidine and benznidazole.
Assuntos
Doença de Chagas/tratamento farmacológico , Nitroimidazóis/antagonistas & inibidores , Pentamidina/farmacologia , Tripanossomicidas/farmacologia , Trypanosoma cruzi/efeitos dos fármacos , Animais , Doença de Chagas/patologia , Chlorocebus aethiops , DNA de Protozoário/análise , DNA de Protozoário/isolamento & purificação , Relação Dose-Resposta a Droga , Glutationa/análogos & derivados , Glutationa/efeitos dos fármacos , Glutationa/metabolismo , Macrófagos/parasitologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Miocárdio/patologia , Parasitemia/tratamento farmacológico , Parasitemia/parasitologia , Putrescina/metabolismo , Distribuição Aleatória , Espermidina/análogos & derivados , Espermidina/metabolismo , Timidina/metabolismo , Tripanossomicidas/antagonistas & inibidores , Células VeroRESUMO
Tumor cells principally exhibit increased mitochondrial transmembrane potential (ΔΨ(m)) and altered metabolic pathways. The therapeutic targeting and delivery of anticancer drugs to the mitochondria might improve treatment efficacy. Gallic acid exhibits a variety of biological activities, and its ester derivatives can induce mitochondrial dysfunction. Four alkyl gallate triphenylphosphonium lipophilic cations were synthesized, each differing in the size of the linker chain at the cationic moiety. These derivatives were selectively cytotoxic toward tumor cells. The better compound (TPP(+)C10) contained 10 carbon atoms within the linker chain and exhibited an IC50 value of approximately 0.4-1.6 µM for tumor cells and a selectivity index of approximately 17-fold for tumor compared with normal cells. Consequently, its antiproliferative effect was also assessed in vivo. The oxygen consumption rate and NAD(P)H oxidation levels increased in the tumor cell lines (uncoupling effect), resulting in a ΔΨ(m) decrease and a consequent decrease in intracellular ATP levels. Moreover, TPP(+)C10 significantly inhibited the growth of TA3/Ha tumors in mice. According to these results, the antineoplastic activity and safety of TPP(+)C10 warrant further comprehensive evaluation.
Assuntos
Antineoplásicos/síntese química , Antineoplásicos/farmacologia , Ácido Gálico/análogos & derivados , Ácido Gálico/síntese química , Trifosfato de Adenosina/metabolismo , Análise de Variância , Animais , Apoptose/efeitos dos fármacos , Caspase 3/efeitos dos fármacos , Inibidores de Caspase/síntese química , Inibidores de Caspase/farmacologia , Cátions/química , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Sobrevivência Celular/efeitos dos fármacos , Ensaios de Seleção de Medicamentos Antitumorais , Ácido Gálico/farmacologia , Humanos , L-Lactato Desidrogenase/metabolismo , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Camundongos , Dilatação Mitocondrial/efeitos dos fármacos , NADP/metabolismo , Oxirredução , Consumo de Oxigênio/efeitos dos fármacos , Reprodutibilidade dos Testes , Desacopladores/síntese química , Desacopladores/farmacologiaRESUMO
The extracts of several plants of Central Chile exhibited anti-Trypanosoma cruzi trypomastigotes activity. Most active extracts were those obtained from Podanthus ovatifolius, Berberis microphylla, Kageneckia oblonga, and Drimys winteri. The active extract of Drimys winteri (IC50 51.2 microg/mL) was purified and three drimane sesquiterpenes were obtained: polygodial, drimenol, and isodrimenin. Isodrimenin and drimenol were found to be active against the trypomastigote form of T. cruzi with IC50 values of 27.9 and 25.1 microM, respectively.
Assuntos
Antiprotozoários/farmacologia , Extratos Vegetais/farmacologia , Plantas Medicinais/química , Trypanosoma cruzi/efeitos dos fármacos , Animais , Chile , Chlorocebus aethiops , Cromatografia em Camada Fina , Espectroscopia de Ressonância Magnética , Células VeroRESUMO
Changes in mitochondrial ATP synthesis can affect the function of tumor cells due to the dependence of the first step of glycolysis on mitochondrial ATP. The oxidative phosphorylation (OXPHOS) system is responsible for the synthesis of approximately 90% of the ATP in normal cells and up to 50% in most glycolytic cancers; therefore, inhibition of the electron transport chain (ETC) emerges as an attractive therapeutic target. We studied the effect of a lipophilic isoprenylated catechol, 3-hydroxybakuchiol (3-OHbk), a putative ETC inhibitor isolated from Psoralea glandulosa. 3-OHbk exerted cytotoxic and anti-proliferative effects on the TA3/Ha mouse mammary adenocarcinoma cell line and induced a decrease in the mitochondrial transmembrane potential, the activation of caspase-3, the opening of the mitochondrial permeability transport pore (MPTP) and nuclear DNA fragmentation. Additionally, 3-OHbk inhibited oxygen consumption, an effect that was completely reversed by succinate (an electron donor for Complex II) and duroquinol (electron donor for Complex III), suggesting that 3-OHbk disrupted the electron flow at the level of Complex I. The inhibition of OXPHOS did not increase the level of reactive oxygen species (ROS) but caused a large decrease in the intracellular ATP level. ETC inhibitors have been shown to induce cell death through necrosis and apoptosis by increasing ROS generation. Nevertheless, we demonstrated that 3-OHbk inhibited the ETC and induced apoptosis through an interaction with Complex I. By delivering electrons directly to Complex III with duroquinol, cell death was almost completely abrogated. These results suggest that 3-OHbk has antitumor activity resulting from interactions with the ETC, a system that is already deficient in cancer cells.
Assuntos
Antineoplásicos Fitogênicos/farmacologia , Apoptose/efeitos dos fármacos , Catecóis/farmacologia , Transporte de Elétrons/efeitos dos fármacos , Mitocôndrias/efeitos dos fármacos , Fenóis/farmacologia , Animais , Antineoplásicos Fitogênicos/química , Catecóis/química , Técnicas de Cultura de Células , Ciclo Celular/efeitos dos fármacos , Morte Celular/efeitos dos fármacos , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Relação Dose-Resposta a Droga , Potencial da Membrana Mitocondrial/efeitos dos fármacos , Camundongos , Mitocôndrias/metabolismo , Mitocôndrias/patologia , Dilatação Mitocondrial/efeitos dos fármacos , Estrutura Molecular , Necrose , Fenóis/química , Espécies Reativas de Oxigênio/metabolismoRESUMO
Chronic Chagas' disease affects 10-30 % of patients infected with Trypanosoma cruzi, and it mainly manifests as cardiomyopathy. Important pathophysiological mechanisms involved in the cardiac lesions include activation of the endothelium and induced microvascular alterations. These processes involve the production of endothelial adhesion molecules and thromboxane A2, which are involved in inflammatory cell recruitment and platelet aggregation, respectively. Cyclooxygenase inhibitors such as aspirin decrease thromboxane production and alter the course of Chagas' disease, both in the acute and chronic phases. We studied the effects of the administration of low and high doses of aspirin during the early phase of T. cruzi infection, following microvascular damage in the context of a chronic murine model of Chagas' disease. The effects of both schedules were assessed at 24 and 90 days postinfection by evaluating parasitemia, mortality, and cardiac histopathological changes as well as the expression of ICAM, VCAM, and E-selectin in cardiac tissue. Thromboxane A2, soluble ICAM, and E-selectin blood levels were also measured. While aspirin did not affect parasitemia or mortality in the infected mice, it decreased both cardiac inflammatory infiltrates and thromboxane levels. Additionally, at 90 days postinfection, aspirin normalized sICAM and sE-selectin levels. Considering the improved endothelial function induced by aspirin, we propose the possibility of including this drug in clinical therapy to treat chronic Chagas' disease.
Assuntos
Anti-Inflamatórios/uso terapêutico , Aspirina/uso terapêutico , Cardiomiopatia Chagásica/tratamento farmacológico , Cardiomiopatia Chagásica/prevenção & controle , Endotélio Vascular/patologia , Animais , Cardiomiopatia Chagásica/patologia , Doença Crônica , Modelos Animais de Doenças , Inibidores Enzimáticos/uso terapêutico , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Parasitemia/tratamento farmacológico , Análise de Sobrevida , Tromboxano A2/sangueRESUMO
Chagas' disease, produced by Trypanosoma cruzi, affects more than 8 million people, producing approximately 10,000 deaths each year in Latin America. Migration of people from endemic regions to developed countries has expanded the risk of infection, transforming this disease into a globally emerging problem. PGE2 and other eicosanoids contribute to cardiac functional deficits after infection with T. cruzi. Thus, the inhibition of host cyclooxygenase (COX) enzyme emerges as a potential therapeutic target. In vivo studies about the effect of acetylsalicylic acid (ASA) upon T. cruzi infection are controversial, and always report the effect of ASA at a single dose. Therefore, we aimed to analyze the effect of ASA at different doses in an in vivo model of infection and correlate it with the production of arachidonic acid metabolites. ASA decreased mortality, parasitemia, and heart damage in T. cruzi (Dm28c) infected mice, at the low doses of 25 and 50 mg/Kg. However, this effect disappeared when the high ASA doses of 75 and 100 mg/Kg were used. We explored whether this observation was related to the metabolic shift toward the production of 5-lipoxygenase derivatives, and although we did not observe an increase in LTB4 production in infected RAW cells and mice infected, we did find an increase in 15-epi-LXA4 (an ASA-triggered lipoxin). We also found high levels of 15-epi-LXA4 in T. cruzi infected mice treated with the low doses of ASA, while the high ASA doses decreased 15-epi-LXA4 levels. Importantly, 15-epi-LXA4 prevented parasitemia, mortality, and cardiac changes in vivo and restored the protective role in the treatment with a high dose of ASA. This is the first report showing the production of ASA-triggered lipoxins in T. cruzi infected mice, which demonstrates the role of this lipid as an anti-inflammatory molecule in the acute phase of the disease.
Assuntos
Aspirina/uso terapêutico , Doença de Chagas/prevenção & controle , Lipoxinas/metabolismo , Trypanosoma cruzi/efeitos dos fármacos , Trypanosoma cruzi/patogenicidade , Animais , Linhagem Celular , Modelos Animais de Doenças , Masculino , Camundongos , Camundongos Endogâmicos BALB CRESUMO
In Latin America, there are about 10-12 million people infected with Trypanosoma cruzi, the agent of Chagas' disease, one of the most important neglected tropical parasitism. Identification of molecular targets, specific for the aggressor or host cells or both, may be useful in the development of pharmacological and/or immunological therapeutic tools. Classic efforts in Chagas' disease explore those strategies. Although the immune system frequently controls parasite aggressions, sterile immunity is seldom achieved and chronic interactions are thus established. However, laboratory-modified immunologic probes aimed at selected parasite targets, may be more effective than their unmodified counterparts. Calreticulin (CRT) from vertebrates is a calcium binding protein, present mainly in the endoplasmic reticulum (ER), where it directs the conformation of proteins and controls calcium levels. We have isolated, gene-cloned, expressed and characterized T. cruzi calreticulin (TcCRT). Upon infection, the parasite can translocate this molecule from the ER to the surface, where it inhibits both the classical and lectin complement pathways. Moreover, by virtue of its capacity to bind and inactivate first complement component C1, it promotes parasite infectivity. These two related properties reside in the central domain of this molecule. A different domain, amino terminal, binds to endothelial cells, thus inhibiting their angiogenic capacity. Since tumor growth depends, to a large extent on angiogenesis, their growth is also inhibited.
Assuntos
Calreticulina/metabolismo , Doença de Chagas/imunologia , Doença de Chagas/parasitologia , Interações Hospedeiro-Parasita , Neoplasias/patologia , Trypanosoma cruzi/fisiologia , Inibidores da Angiogênese , Animais , Proliferação de Células , Doença de Chagas/patologia , Humanos , Neoplasias/metabolismo , Trypanosoma cruzi/imunologia , Trypanosoma cruzi/patogenicidadeRESUMO
Chagas disease is one of the most important endemic diseases in Latin America, caused by Trypanosoma cruzi. The drugs used for the treatment of this disease, nifurtimox and benznidazole, are toxic and present severe side effects. The need of effective drugs, without adverse effects, has stimulated the search for new compounds with potential clinical utility. An overview of a number of natural naphthoquinones tested against T. cruzi parasites is provided. Among natural naphthoquinones, lapachol, ß-lapachone and its α-isomer have demonstrated useful trypanocidal activities. In the search for new trypanocidal agents, this review outlines different structural modifications of natural quinones, as well as synthetic quinones, which have been subjected to trypanocidal studies. This review summarizes the mechanism of action and structure-activity relationships of the quinone derivatives, including some theoretical calculations that discuss the correlation of stereo electronic properties with the trypanocidal activity. In this context, this review will be useful for the development of new antichagasic drugs based mainly on structural modification of natural quinones.
Assuntos
Doença de Chagas/tratamento farmacológico , Naftoquinonas/química , Tripanossomicidas/química , Trypanosoma cruzi/efeitos dos fármacos , Humanos , Naftoquinonas/síntese química , Naftoquinonas/uso terapêutico , Relação Estrutura-Atividade , Tripanossomicidas/síntese química , Tripanossomicidas/uso terapêuticoRESUMO
Nifurtimox and benznidazole are the only active drugs against Trypanosoma cruzi; however, they have limited efficacy and severe side effects. During primoinfection, T. cruzi infected macrophages mount an antiparasitic response, which the parasite evades through an increase of tumor growth factor beta and PGE(2) activation as well as decreased iNOS activity. Thus, prostaglandin synthesis inhibition with aspirin might increase macrophage antiparasitic activity and increase nifurtimox and benznidazole effect. Aspirin alone demonstrated a low effect upon macrophage antiparasitic activity. However, isobolographic analysis of the combined effects of aspirin, nifurtimox and benznidazole indicated a synergistic effect on T. cruzi infection of RAW cells, with combinatory indexes of 0.71 and 0.61, respectively. The observed effect of aspirin upon T. cruzi infection was not related with the PGE(2) synthesis inhibition. Nevertheless, NO() levels were restored by aspirin in T. cruzi-infected RAW cells, contributing to macrophage antiparasitic activity improvement. Thus, the synergy of aspirin with nifurtimox and benznidazole is due to the capability of aspirin to increase antiparasitic activity of macrophages.
Assuntos
Aspirina/farmacologia , Inibidores de Ciclo-Oxigenase/farmacologia , Nifurtimox/farmacologia , Nitroimidazóis/farmacologia , Tripanossomicidas/farmacologia , Trypanosoma cruzi/efeitos dos fármacos , Animais , Linhagem Celular , Sobrevivência Celular/efeitos dos fármacos , Sinergismo Farmacológico , Concentração Inibidora 50 , Macrófagos/efeitos dos fármacos , Macrófagos/imunologia , Macrófagos/parasitologia , Camundongos , Óxido Nítrico/metabolismoRESUMO
There are approximately 7.8 million people in Latin America, including Chile, who suffer from Chagas disease and another 28 million who are at risk of contracting it. Chagas is caused by the flagellate protozoan Trypanosoma cruzi. It is a chronic disease, where 20%-30% of infected individuals develop severe cardiopathy, with heart failure and potentially fatal arrhythmias. Currently, Chagas disease treatment is more effective in the acute phase, but does not always produce complete parasite eradication during indeterminate and chronic phases. At present, only nifurtimox or benznidazole have been proven to be superior to new drugs being tested. Therefore, it is necessary to find alternative approaches to treatment of chronic Chagas. The current treatment may be rendered more effective by increasing the activity of anti-Chagasic drugs or by modifying the host's immune response. We have previously shown that glutathione synthesis inhibition increases nifurtimox and benznidazole activity. In addition, there is increasing evidence that cyclooxygenase inhibitors present an important effect on T. cruzi infection. Therefore, we found that aspirin reduced the intracellular infection in RAW 264.7 cells and, decreased myocarditis extension and mortality rates in mice. However, the long-term benefit of prostaglandin inhibition for Chagasic patients is still unknown.