RESUMO
PURPOSE: Fasciola hepatica is a globally distributed trematode that causes significant economic losses. Triclabendazole is the primary pharmacological treatment for this parasite. However, the increasing resistance to triclabendazole limits its efficacy. Previous pharmacodynamics studies suggested that triclabendazole acts by interacting mainly with the ß monomer of tubulin. METHODS: We used a high-quality method to model the six isotypes of F. hepatica ß-tubulin in the absence of three-dimensional structures. Molecular dockings were conducted to evaluate the destabilization regions in the molecule against the ligands triclabendazole, triclabendazole sulphoxide and triclabendazole sulphone. RESULTS: The nucleotide binding site demonstrates higher affinity than the binding sites of colchicine, albendazole, the T7 loop and pßVII (p < 0.05). We suggest that the binding of the ligands to the polymerization site of ß-tubulin can lead a microtubule disruption. Furthermore, we found that triclabendazole sulphone exhibited significantly higher binding affinity than other ligands (p < 0.05) across all isotypes of ß-tubulin. CONCLUSIONS: Our investigation has yielded new insight on the mechanism of action of triclabendazole and its sulphometabolites on F. hepatica ß-tubulin through computational tools. These findings have significant implications for ongoing scientific research ongoing towards the discovery of novel therapeutics to treat F. hepatica infections.
Assuntos
Anti-Helmínticos , Fasciola hepatica , Fasciolíase , Animais , Triclabendazol/farmacologia , Triclabendazol/metabolismo , Triclabendazol/uso terapêutico , Tubulina (Proteína)/genética , Simulação de Acoplamento Molecular , Benzimidazóis/farmacologia , Benzimidazóis/química , Benzimidazóis/metabolismo , Ligantes , Sulfonas/metabolismo , Sulfonas/uso terapêutico , Anti-Helmínticos/farmacologia , Anti-Helmínticos/uso terapêutico , Fasciolíase/parasitologiaRESUMO
Focal adhesion kinase (FAK), a cytoplasmic protein tyrosine kinase (PTK), is implicated in diverse cellular processes, including the regulation of F-actin dynamics. Host cell F-actin rearrangement is critical for invasion of Trypanosoma cruzi, the protozoan parasite that causes Chagas disease. It is unknown whether FAK is involved in the internalization process of metacyclic trypomastigote (MT), the parasite form that is important for vectorial transmission. MT can enter the mammalian host through the ocular mucosa, lesion in the skin, or by the oral route. Oral infection by MT is currently a mode of transmission responsible for outbreaks of acute Chagas disease. Here we addressed the question by generating HeLa cell lines deficient in FAK. Host cell invasion assays showed that, as compared to control wild type (WT) cells, FAK-deficient cells were significantly more susceptible to parasite invasion. Lysosome spreading and a disarranged actin cytoskeleton, two features associated with susceptibility to MT invasion, were detected in FAK-deficient cells, as opposed to WT cells that exhibited a more organized F-actin arrangement, and lysosomes concentrated in the perinuclear area. As compared to WT cells, the capacity of FAK-deficient cells to bind a recombinant protein based on gp82, the MT surface molecule that mediates invasion, was higher. On the other hand, when treated with FAK-specific inhibitor PF573228, WT cells exhibited a dense meshwork of actin filaments, lysosome accumulation around the nucleus, and had increased resistance to MT invasion. In cells treated with PF573228, the phosphorylation levels of FAK were reduced and, as a consequence of FAK inactivation, diminished phosphorylation of extracellular signal-regulated protein kinases (ERK1/2) was observed. Fibronectin, known to impair MT invasion, induced the formation of thick bundles of F-actin and ERK1/2 dephosphorylation.
Assuntos
Suscetibilidade a Doenças/metabolismo , Proteína-Tirosina Quinases de Adesão Focal/metabolismo , Proteínas de Protozoários/metabolismo , Trypanosoma cruzi/metabolismo , Glicoproteínas Variantes de Superfície de Trypanosoma/metabolismo , Actinas/metabolismo , Doença de Chagas/metabolismo , Doença de Chagas/parasitologia , Suscetibilidade a Doenças/parasitologia , Quinase 1 de Adesão Focal/metabolismo , Proteína-Tirosina Quinases de Adesão Focal/genética , Células HeLa , Interações Hospedeiro-Patógeno/genética , Interações Hospedeiro-Patógeno/fisiologia , Humanos , Lisossomos/metabolismo , Sistema de Sinalização das MAP Quinases , Fosforilação , Proteínas de Protozoários/genética , Quinolonas/metabolismo , Proteínas Recombinantes/metabolismo , Sulfonas/metabolismo , Trypanosoma cruzi/genética , Trypanosoma cruzi/patogenicidade , Glicoproteínas Variantes de Superfície de Trypanosoma/genéticaRESUMO
Acute kidney injury (AKI) and metabolic dysfunction are critical complications in sepsis syndrome; however, their pathophysiological mechanisms remain poorly understood. Therefore, we evaluated whether the pharmacological properties of 6-gingerol (6G) and 10-gingerol (10G) could modulate AKI and metabolic disruption in a rat model of sepsis (faecal peritonitis). Animals from the sham and AKI groups were intraperitoneally injected with 6G or 10G (25 mg/kg). Septic AKI decreased creatinine clearance and renal antioxidant activity, but enhanced oxidative stress and the renal mRNA levels of tumour necrosis factor-α, interleukin-1ß, and transforming growth factor-ß. Both phenol compounds repaired kidney function through antioxidant activity related to decreased oxidative/nitrosative stress and proinflammatory cytokines. Metabolomics analysis indicated different metabolic profiles for the sham surgery group, caecal ligation and puncture model alone group, and sepsis groups treated with gingerols. 1H nuclear magnetic resonance analysis detected important increases in urinary creatine, allantoin, and dimethylglycine levels in septic rats. However, dimethylamine and methylsulfonylmethane metabolites were more frequently detected in septic animals treated with 6G or 10G, and were associated with increased survival of septic animals. Gingerols attenuated septic AKI by decreasing renal disturbances, oxidative stress, and inflammatory response through a mechanism possibly correlated with increased production of dimethylamine and methylsulfonylmethane.
Assuntos
Injúria Renal Aguda/prevenção & controle , Catecóis/administração & dosagem , Álcoois Graxos/administração & dosagem , Peritonite/complicações , Sepse/complicações , Injúria Renal Aguda/etiologia , Injúria Renal Aguda/metabolismo , Injúria Renal Aguda/mortalidade , Animais , Dimetil Sulfóxido/metabolismo , Dimetilaminas/metabolismo , Modelos Animais de Doenças , Fezes/microbiologia , Humanos , Injeções Intraperitoneais , Masculino , Metaboloma/efeitos dos fármacos , Metabolômica , Estresse Oxidativo/efeitos dos fármacos , Peritonite/metabolismo , Peritonite/microbiologia , Peritonite/mortalidade , Ratos , Ratos Wistar , Sepse/metabolismo , Sepse/microbiologia , Sepse/mortalidade , Sulfonas/metabolismo , Análise de Sobrevida , Resultado do TratamentoRESUMO
This study was designed to verify the in vivo efficacy of sulfoxide and sulfone fexinidazole metabolites following oral administration in a murine model of Chagas disease. Female Swiss mice infected with the Y strain of Trypanosoma cruzi were treated orally once per day with each metabolite at doses of 10 to 100 mg/kg of body weight for a period of 20 days. Parasitemia was monitored throughout, and cures were detected by parasitological and PCR assays. The results were compared with those achieved with benznidazole treatment at the same doses. Fexinidazole metabolites were effective in reducing the numbers of circulating parasites and protecting mice against death, compared with untreated mice, but without providing cures at daily doses of 10 and 25 mg/kg. Both metabolites were effective in curing mice at 50 mg/kg/day (30% to 40%) and 100 mg/kg/day (100%). In the benznidazole-treated group, parasitological cure was detected only in animals treated with the higher dose of 100 mg/kg/day (80%). Single-dose pharmacokinetic parameters for each metabolite were obtained from a parallel group of uninfected mice and were used to estimate the profiles following repeated doses. Pharmacokinetic data suggested that biological efficacy most likely resides with the sulfone metabolite (or subsequent reactive metabolites formed following reduction of the nitro group) following administration of either the sulfoxide or the sulfone and that prolonged plasma exposure over the 24-h dosing window is required to achieve high cure rates. Fexinidazole metabolites were effective in treating T. cruzi in a mouse model of acute infection, with cure rates superior to those achieved with either fexinidazole itself or benznidazole.
Assuntos
Doença de Chagas/tratamento farmacológico , Nitroimidazóis/farmacologia , Sulfonas/farmacologia , Sulfóxidos/farmacologia , Tripanossomicidas/farmacologia , Trypanosoma cruzi/efeitos dos fármacos , Administração Oral , Animais , Biotransformação , Doença de Chagas/mortalidade , Doença de Chagas/parasitologia , Modelos Animais de Doenças , Relação Dose-Resposta a Droga , Esquema de Medicação , Feminino , Camundongos , Nitroimidazóis/farmacocinética , Sulfonas/metabolismo , Sulfóxidos/metabolismo , Análise de Sobrevida , Tripanossomicidas/farmacocinética , Trypanosoma cruzi/crescimento & desenvolvimentoRESUMO
The flukicidal compound triclabendazole (TCBZ) has a complex metabolic pattern that includes the systemic presence of its sulphoxide (TCBZ.SO) and sulphone (TCBZ.SO2) metabolites, usually recovered from the bile of treated animals. The aim of the current work was to evaluate the time-course and pattern of in vivo accumulation of TCBZ/metabolites into adult Fasciola hepatica specimens recovered from infected sheep. Twelve (12) healthy Corriedale sheep were orally infected with one hundred (100) metacercariae of the TCBZ-susceptible Cullomptom isolate of F. hepatica. Sixteen weeks after infection, animals were intraruminally treated with TCBZ (10mg/kg). At 3, 24, 48 and 60h post-treatment (pt), animals were sacrificed (n=3/time period) and samples of blood, bile, liver tissue and adult F. hepatica specimens were collected. The concentrations of TCBZ/metabolites were measured by HPLC. TCBZ.SO and TCBZ.SO2 were the only molecules recovered in the bloodstream, with peak plasma concentrations of 10.8µg/mL (TCBZ.SO) and 12.6µg/mL (TCBZ.SO2). The same metabolites were also the main analytes accumulated within the adult flukes, reaching peak concentrations between 6.35µg/g (TCBZ.SO) and 13.9µg/g (TCBZ.SO2) at 24h pt, which was coincident with the time when the maximum plasma concentration was attained. Low levels of TCBZ parent drug (0.14µg/g at 24h pt) were measured within collected flukes. TCBZ parent drug and its sulpho- and hydroxy-derivatives were recovered in bile collected from treated sheep between 3 and 60h pt. Although relatively high concentrations of hydroxy-TCBZ (ranging from 0.86 to 10.1µg/mL) were measured in bile, this metabolite was not recovered within the flukes at any time pt. Finally, TCBZ parent drug was the main compound accumulated in liver tissue over the 60h pt period. The time-course and drug concentration patterns within the adult liver fluke after TCBZ treatment followed a similar trend to those observed in plasma. Overall, the data reported here confirm that oral ingestion is a main route of drug entry into the trematode in vivo exposed to TCBZ/metabolites. However, the presence of TCBZ within the adult fluke (despite being absent in the systemic circulation) may be related to some degree of trans-tegumental diffusion from bile or by a direct oral ingestion from portal blood.
Assuntos
Anti-Helmínticos/farmacocinética , Benzimidazóis/farmacocinética , Bile/metabolismo , Fasciolíase/veterinária , Fígado/metabolismo , Doenças dos Ovinos/tratamento farmacológico , Animais , Anti-Helmínticos/sangue , Anti-Helmínticos/uso terapêutico , Área Sob a Curva , Benzimidazóis/sangue , Benzimidazóis/metabolismo , Benzimidazóis/uso terapêutico , Fasciola hepatica/efeitos dos fármacos , Fasciola hepatica/metabolismo , Fasciolíase/tratamento farmacológico , Fasciolíase/metabolismo , Masculino , Ovinos , Doenças dos Ovinos/metabolismo , Doenças dos Ovinos/parasitologia , Sulfonas/metabolismo , Sulfóxidos/metabolismo , TriclabendazolRESUMO
Benzimidazole 2-carbamates, such as albendazole (ABZ) and mebendazole (MBZ), used for the treatment of helmintic infections, have low aqueous solubility and poor bioavailability, both of which lead to high interindividual variability when used for human systemic helmintiosis; therefore, it is necessary to search for new anthelmintics with better biopharmaceutical properties. In the present study the solubility, pKa, logP and apparent permeability in the Caco-2 cells system of four novel anthelmintic (1H-benzimidazol-5(6)-yl)carboxamide derivatives (compounds 1-4) with a 2-methylthyo group were evaluated. Also the pharmacokinetic parameters of compound 1 which in previous studies showed activity similar to ABZ against T. spirallis, was evaluated in BALB/c mice, as a representative molecule of the series. The novel anthelmintics, showed better solubility than ABZ in aqueous acid pH and in organic solvents. The logP, P(app) and Caco-2 data indicate that the 4 derivatives are highly permeable drugs, but it is possible that an efflux system could be involved in the transport of these compounds. Plasma levels of compound 1 and its sulfoxide (compound 5) were high after the first 5 min. This fact strongly suggests that compound 1 is rapidly metabolized in the small intestine. On the other hand, the sulfone metabolite (compound 6) levels were lower than those of compound 5. The half life and mean residence time (MRT) of compound 1 and its main metabolites indicate that their elimination is very rapid. More studies in mammalian species are necessary in order to understand the pharmacokinetic behavior of these novel compounds.
Assuntos
Anti-Helmínticos/química , Anti-Helmínticos/farmacocinética , Benzimidazóis/química , Benzimidazóis/farmacocinética , 1-Octanol/química , Animais , Células CACO-2 , Humanos , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Solubilidade , Sulfonas/metabolismo , Sulfóxidos/metabolismo , Água/químicaRESUMO
The nitric oxide/cyclic-guanosine 3',5'-monophosphate signaling cascade plays an essential role in cardiovascular homeostasis but its involvement in the pathophysiology of refractory hypertension is unclear. The acute vasodilatory effect of a single oral dose of a phosphodiesterase-5 inhibitor (sildenafil citrate) on the brachial artery dilatation was evaluated in 25 normal healthy volunteers (NL) and in 25 refractory hypertensive patients (RH). Endothelial and vascular smooth muscle functions were assessed two times. First, the brachial artery response to endothelium-dependent (flow-mediated dilatation [FMD]) and independent (glyceryl trinitrate [GTN]) stimuli was examined. The FMD in NL was 14.2+/-3.2% compared to 10.3+/-3.5% in RH (P<0.001) and the GTN-induced responses were 23.5+/-6.3 in NL compared to 18.4+/-5.7% in RH (P<0.001). Two weeks later, the brachial artery responses to FMD were determined before and after the administration of sildenafil citrate. Sildenafil caused a significant, slow and progressive dilatation of the brachial artery until 45 min after administration (4.7+/-3.0%, 6.7+/-3.0% and 9.4+/-3.9% after 15', 30' and 45', respectively, in RH and 3.7+/-1.9%, 7.4+/-2.7% and 10.1+/-3.0%, respectively, in NL). A second FMD stimulus, applied 45 min after ingesting 50mg of sildenafil resulted in an additional significant increase in the vasodilatory response (from 9.4+/-3.9% to 13.0+/-4.0% in RH; P<0.001 and from 10.1+/-3.0 to 14.6+/-4.1 in NL; P<0.001), but this was still significantly less than the response to GTN. Sildenafil citrate caused brachial artery vasodilatation similar to that caused by NO released during FMD in patients with refractory hypertension.
Assuntos
GMP Cíclico/metabolismo , Endotélio/metabolismo , Hipertensão/tratamento farmacológico , Óxido Nítrico/metabolismo , Inibidores de Fosfodiesterase , Piperazinas , Sulfonas , Vasodilatação/fisiologia , 3',5'-GMP Cíclico Fosfodiesterases/metabolismo , Adulto , Nucleotídeo Cíclico Fosfodiesterase do Tipo 5 , Humanos , Masculino , Pessoa de Meia-Idade , Nitroglicerina/uso terapêutico , Inibidores de Fosfodiesterase/metabolismo , Inibidores de Fosfodiesterase/uso terapêutico , Piperazinas/metabolismo , Piperazinas/uso terapêutico , Purinas/metabolismo , Purinas/uso terapêutico , Transdução de Sinais/fisiologia , Citrato de Sildenafila , Sulfonas/metabolismo , Sulfonas/uso terapêutico , Vasodilatadores/metabolismo , Vasodilatadores/uso terapêuticoRESUMO
Estudou-se o efeito do DDS, sobre a transformaçao linfocitaria estimulada pela PHA, in vitro, utilisando-se concentraçoes variaveis de DDS. Os linfocitos foram doados por voluntarios, divididos em tres gupos, e o estudo foi desenvolvido em tres fases. Foram empregadas 4 concentraçoes diferentes de DDS para 10 a sexta de linfocitos em cultura de tecidos alem de 0,02 ml de PHA. Foi observada uma diminuiçao estatisticamente significativa (P<0,05) no percentual de formaçao de celulas blasticas induzida pela PHA com todas as concentraçoes de DDS, exceto com a concentraçao mais baixa (0,01 ug de DDS). Enquanto que a diminuiçao observada na primeira e na terceira fases mostrou-se dependente de dose de DDS, na segunda fase nao se encontrou correlaçao significativa entre a concentraçao de DDS e a depressao da blastogenese induzida pela PHA. Discute-se a significancia destas observaçoes.