Synthesis, molecular modeling and preliminary biological evaluation of a set of 3-acetyl-2,5-disubstituted-2,3-dihydro-1,3,4-oxadiazole as potential antibacterial, anti-Trypanosoma cruzi and antifungal agents

AbstractA series of 3-acetyl-2,5-disubstituted-2,3-dihydro-1,3,4-oxadiazole derivatives was synthesized and their activity screened in vitro against Staphylococcus aureus, Trypanosoma cruzi, and Candida albicans. The bioactivity was expressed as minimum inhibitory concentration (MIC) for S. aureus strains, and as fifty-percent inhibitory concentration (IC50) of parasite population growth for T. cruzi. A molecular modeling approach was performed to establish qualitative relationships regarding the biological data and the compounds’ physicochemical properties. The 5-(4-OC4H9Ph, 5l), and 5-(4-CO2CH3Ph, 5o) derivatives were the most active compounds for S. aureus ATCC 25923 (MIC=1.95–1.25μg/mL) and T. cruzi (IC50=7.91μM), respectively. Also, a preliminary evaluation against C. albicans involving some compounds was performed and the 5-(4-CH3Ph, 5e) derivative was the most active compound (MIC=3.28–2.95μg/mL). In this preliminary study, all synthesized 3-acetyl-2,5-disubstituted-2,3-dihydro-1,3,4-oxadiazole derivatives were active against all microorganisms tested

Preliminary in vitro evaluation of N '-(benzofuroxan-5-yl)methylene benzohydrazide derivatives as potential anti-Trypanosoma cruzi agents

A set of benzofuroxan derivatives was tested in vitro against Trypanosoma cruzi epimastigote forms. The influence of physicochemical properties on these benzofuroxan derivatives' activity was observed, and the presence of electron-withdrawing and hydrophobic groups attached to the benzene ring seems to make a favorable contribution at lower concentrations.CNPqCNPqCAPESCAPESFAPESPFAPES

Novel nutraceutical supplements with yeast β-glucan, prebiotics, minerals, and Silybum marianum (silymarin) ameliorate obesity-related metabolic and clinical parameters: A double-blind randomized trial

PurposeIt is known that obesity has a multifactorial etiology that involves genetic and environmental factors. The WHO estimates the worldwide prevalence of 1.9 billion overweight adults and more than 650 million people with obesity. These alarming data highlight the high and growing prevalence of obesity and represent a risk factor for the development and aggravation of other chronic diseases, such as nonalcoholic fatty liver disease (NAFLD) that is frequently considered the hepatic outcome of type 2 diabetes. The use of non-pharmacological therapies such as food supplements, nutraceuticals, and natural integrative therapies has grown as an alternative tool for obesity-related diseases compared to conventional medications. However, it is a still little explored research field and lacks scientific evidence of therapeutic effectiveness. Considering this, the aim is to evaluate whether a new nutraceutical supplement composition can improve and supply essential mineral nutrients, providing an improvement of obesity-related metabolic and endocrine parameters.MethodsSedentary volunteers (women and men) with body mass index (BMI) ≤34.9 kg/m2 were divided into two groups: Novel Nutraceutical Supplement_(S) (n = 30) and Novel Nutraceutical Supplement (n = 29), differing in the absence (S) or presence of silymarin, respectively. Volunteers were instructed to take two capsules in the morning and two capsules in the evening. No nutritional intervention was performed during the study period. The data (anthropometrics and anamneses) and harvest blood (biochemistry and hormonal exams) were collected at three different time points: baseline time [day 0 (T0)], day 90 (T90), and day 180 (T180) post-supplementation.ResultsIn the anthropometric analysis, the waist circumference in middle abdomen (WC-mid) and waist circumference in iliac crest (WC-IC) were reduced. Also, the waist-to-height ratio (WHt R) and waist-to-hip ratio (WHR) seem to slightly decrease alongside the supplementation period with both nutraceutical supplements tested as well as transaminase enzyme ratio [aspartate aminotransferase (AST)/alanine aminotransferase (ALT) ratio (AAR)], a known as a biomarker of NAFLD, and endocrine hormones cortisol and thyroid-stimulating hormone (TSH) at 90 and 180 days post-supplementation.ConclusionsIn a condition associated with sedentary and no nutritional intervention, the new nutraceutical supplement composition demonstrated the ability to be a strong and newfangled tool to improve important biomarkers associated with obesity and its comorbidities

Design of anti-T. cruzi compounds: Synthesis of 5-nitrofuran derivatives, in vitro biological activity and study of structure-activity relationships

A doença de Chagas afeta cerca de 6 a 7 millhões de pessoas no mundo, principalmente América Latina. A busca de alternativas terapêuticas para esta enfermidade tem grande relevância para a sociedade, já que as opções atuais são limitadas, sendo disponível apenas o benznidazol (BZD) e nifurtimox. Os derivados nitroheterocíclicos são considerados compostos bioativos com número crescente de estudos na comunidade científica contra seu agente etiológico, o Trypanosoma cruzi. Neste sentido, o presente trabalho tem por objetivo a identificação de derivados do 5-nitrofurano com atividade frente a diferentes cepas do T. cruzi, assim como estudar possíveis modo de ação desta classe de compostos. Esta investigação envolve estudos computacionais com o propósito de construir modelos quantitativos de relações estrutura-atividade (QSAR multivariado) que possam auxiliar na previsão de novas estruturas com perfil farmacológico otimizado. No presente trabalho foram realizadas as etapas de planejamento, síntese e identificação de 36 compostos com resultados satisfatórios quanto à identificação estrutural, pureza e rendimento, que foi da ordem de 70%. A determinação da atividade anti-T. cruzi in vitro dos compostos obtidos foi realizada frente às cepas Silvio X10 cl1, Y, Bug 2149 cl10 e Colombiana na forma epimastigota do parasito. A maioria dos compostos analisados apresentou maior capacidade de inibição de crescimento do parasito, comparado ao BZD: Silvio X10 cl1 - IC50 = 29,16 ±2,90 µM, Y - IC50 = 40,40 ±3,37µM, Bug 2149 cl10 - IC50 = 30,63 ±3,21 µM, Colombiana - IC50 = 47,91 ±4,96 µM. O composto mais ativo (BSF-35) apresentou os seguintes valores: Silvio X10 cl1 - IC50 = 3,17 ±0,32 µM, Y - IC50 = 1,17 ±0,12 µM, Bug 2149 cl10 - IC50 = 1,81 ±0,18 µM e Colombiana - IC50 = 3,06 ±0,23 µM. Foram realizados cálculos de propriedades moleculares das estruturas tridimensionais dos compostos, seguido pela análise exploratória de dados por análise de agrupamentos hierárquicos (HCA) e análise de componentes principais (PCA), possibilitando o reconhecimento de padrões do conjunto. Considerando esta análise prévia, foram obtidos modelos QSAR com abordagem multivariada, aplicando algorítmo OPS e método de regressão por quadrados mínimos parciais, PLS. Os melhores modelos gerados foram obtidos considerando os compostos benzenos substituídos para as quatro cepas estudadas. Os descritores que mais influenciaram na análise foram o ClogP (coeficiente de partição) e cargas CHELPG. Considerando as informações obtidas, foram planejados e sintetizados quatro novos compostos com objetivo de obter compostos mais ativos e validar os modelos QSAR. Estes compostos apresentaram alta atividade frente a forma epimastigota das quatro cepas estudadas. Os compostos mais ativos foram avaliados quanto a citotoxicidade frente células LLC-MK2 e apresentaram seletividade até 25 vezes superior ao BZD. Estudos in vitro frente a forma amastigota da cepa Y em células U2OS foram realizados com metodologia fenotípica de análise de alto conteúdo (HCA\') e os compostos apresentaram atividade até 64 vezes superior ao BZD e com seletividade de até 50 vezes superior a este fármaco. Quanto à determinação da atividade dos compostos frente às enzimas tripanotiona redutase (TcTR) e glutationa redutase (GR), os compostos analisados não apresentaram atividade relevante, indicando não ser este o mecanismo desta classe de compostos. Com finalidade de explorar outro possível mecanismo de ação dos compostos 5-nitrofurânicos, foi realizada a análise de potencial de redução da membrana mitocondrial, porém a morte parasitária não foi atribuída à despolarização da membrana em estudos simultâneos com iodeto de propídio.Chagas disease affects approximately 6-7 millions people worldwide, especially Latin America. The search for therapeutic alternatives for this disease is of great relevance to society, as current options are limited and there are only two available drugs: benznidazole (BZD) and nifurtimox. The nitroheterocyclic derivatives are considered bioactive compounds with increasing number of studies in the scientific community against its etiologic agent, Trypanosoma cruzi. In this sense, this work aims to identify derivatives of 5-nitrofuran with activity against different strains of T. cruzi, and to study possible mode of action of this compounds. This research involves computational studies to obtain models of quantitative structure-activity relationships (QSAR multivariate) that can help predict new structures with optimized pharmacological profile. In this work were carried out the design, synthesis and identification of 36 compounds with satisfactory results regarding the structural identification, purity and yield (approximately 70%). The determination of anti-T. cruzi activity in vitro of the compounds obtained was carried out with Silvio X10 cl1, Y, Bug 2149 CL10 and Colombiana strains of epimastigote form of the parasite. Most of the compounds examined showed greater capacity of growth inhibition of the parasite compared to the BZD (Silvio X10 CL1 - IC 50 = 29.16 ± 2.90 µM, Y - IC50 = 40.40 ± 3,37µM, 2149 CL10 Bug - IC 50 = 30.63 ± 3.21 µM, Colombiana - IC 50 = 47.91 ± 4.96 µM). The most active compound (BSF-35) showed the following values: Silvio X10 cl1 - IC 50 = 3.17 ± 0.32 uM, Y - IC 50 = 1.17 ± 0.12 µM, Bug 2149 CL10 - IC50 = 1, 81 ± 0.18 µM and Colombiana - IC 50 = 3.06 ± 0.23 µM. Calculations were performed for the molecular properties of three-dimensional structures of the compounds, followed by exploratory data analysis by hierarchical cluster analysis (HCA) and principal component analysis (PCA), allowing the recognition of the set. Considering this preliminary analysis were obtained QSAR models with multivariate approach, using OPS algorithm and regression method of partial least squares, PLS. The best generated models were obtained considering the benzyl substituted compounds for the four strains. The descriptors that most influenced the analysis were ClogP (partition coefficient) and CHELPG charges. Considering the information obtained, four new compounds were designed and synthesized to obtain more active compounds and validate QSAR models. These compounds showed high activity against epimastigote form of the four strains studied. The most active compounds were evaluated for cytotoxicity against LLC-MK2 cells and the compounds selectivity values were up to 25 times higher than BZD. In vitro studies against amastigote form of the Y strain in U2OS cells were performed with phenotypic method of high content analysis (HCA\') and the compounds showed activity to 64 times higher than BZD and selectivity of up to 50 times. The activity of the compounds against trypanothione reductase enzymes (TcTR) and glutathione reductase (GR) showed no significant activity, indicating that this is not the mechanism of this class of compounds. In order to exploit another possible mechanism of action of 5-nitrofuran derivatives, analysis reduction of mitochondrial membrane potential was held, however the cell death was not attributed to membrane depolarization in simultaneous studies with propidium iodide

Design, synthesis, identification and anti-Trypanosoma cruzi activity evaluation of furfurylidene azomethine and oxadiazole derivatives

A busca de alternativas terapêuticas para o tratamento da doença de Chagas é de grande importância, visto que atualmente existem apenas dois fármacos disponíveis, o benznidazol e o nifurtimox. Ambos apresentam efeitos adversos consideráveis, sendo utilizado, no Brasil, apenas o benznidazol. Compostos nitro-heterocíclicos com atividade frente ao Trypanosoma cruzi, agente causal da doença de Chagas, tem apresentado resultados promissores. Assim, este trabalho abrange o planejamento, síntese, identificação, avaliação da atividade anti-T. cruzi de 5-nitro furfurilidênicos (IC50 T. cruzi) e a citotoxicidade destes compostos frente a macrófagos de linhagem J774 (IC50 J774). A nifuroxazida, composto-protótipo, inspirou as modificações moleculares originando duas séries de compostos furfurilidênicos, uma com estrutura azometínica, série I, e outra com estrutura oxadiazolínica, série II. A escolha de substituintes foi baseada no diagrama de Craig, sendo selecionados dez substituintes para cada série. Foi avaliada a atividade dos vinte compostos planejados frente ao T. cruzi, dos quais os mais ativos foram: 4-butil-[N\'- (5-nitrofuran-2-il)metileno]benzidrazida (4g - IC50 T. cruzi = 1,05 µM, DP = 0,07) e 3-Acetil-5-(4-butilfenil)-2-(5-nitrofuran-2-il)-2,3-di-idro-1,3,4-oxadiazol (5g - IC50 T. cruzi = 8,27 µM, DP = 0,42). Comparados com os fármacos de referência, benznidazol (IC50 T. cruzi = 22,69 µM, DP = 1,96) e nifurtimox (IC50 T. cruzi = 3,78 µM, DP = 0,10), o composto 4g demonstrou atividade anti-T. cruzi superior a ambos. Todos os compostos apresentaram atividade maior do que a nifuroxazida (IC50 T. cruzi = 120,46 µM, DP = 4,06). Para os ensaios de citotoxicidade, obteve-se para o composto mais ativo frente ao T. cruzi, 4g, IC50 J774 = 28,05 µM, DP = 1,05, e para o composto 5g, obteve-se IC50 J774 = > 400 µM, o que representa que a concentração máxima avaliada deste composto não afetou as células. Ambos apresentaram boa seletividade ao efetuar a relação entre o IC50 J774 e o IC50 T. cruzi. Adicionalmente, foram realizados cálculos de propriedades físico-químicas das estruturas tridimensionais dos compostos, seguido pela análise exploratória de dados por análise de agrupamentos hierárquicos (hierarchical clusters analysis - HCA) e análise de componentes principais (principal component analysis, PCA), possibilitando a identificação das propriedades que mais influenciam na atividade anti-T. cruzi nas séries dos compostos estudados. Os resultados indicaram uma forte influência das propriedades ClogP e momento dipolo, evidenciando a necessidade de um equilíbrio lipofílico/hidrofílico no planejamento de novas moléculas com ação anti-T. cruzi.The search for alternative therapies for the treatment of Chagas disease presents great importance, since there are only two currently available drugs, nifurtimox and benznidazole. Both have considerable adverse effects and, in Brazil, is used only benznidazole. Nitro-heterocyclic compounds with activity against Trypanosoma cruzi, the causative agent of Chagas disease, has shown promising results. Thus, this work includes the design, synthesis, identification, evaluation of anti-T. cruzi activity of 5-nitro-2- furfuriliden (IC50 T. cruzi) and cytotoxicity of these compounds against J774 macrophages cell line (IC50 J774). The nifuroxazide, as a lead compound, inspired the molecular modification leading to two series of furfuriliden compounds, a azometinic structure, series I, and other with oxadiazolinic structure, series II. The choice of substituents was based on the Craig\'s diagram, and ten substituents were selected for each series. We evaluated the activity of twenty compounds designed against T. cruzi, and the most active compounds were: 4-butyl-[N\'-(5-nitrofuran-2-yl) methylene] benzidrazide (4g - IC50 T. cruzi = 1.05 µM, SD = 0.07) and 3-acetyl-5-(4-butylphenyl)-2 -(5-nitrofuran-2-yl)- 2,3-dihydro, 1,3,4-oxadiazole (5g - IC50 T. cruzi = 8.27 µM, SD = 0.42). Compared to the reference drugs, benznidazole (IC50 T. cruzi = 22.69 µM, SD = 1.96) and nifurtimox (IC50 T. cruzi = 3.78 µM, SD = 0.10), the compound 4g demonstrated anti-T. cruzi activity superior to both drugs. All compounds showed better activity than nifuroxazide (IC50 T. cruzi = 120.46 µM, SD = 4.06). For cytotoxicity assays, was found for the most active compound against T. cruzi, 4g, IC50 J774 = 28.05 µM, SD = 1.05, and for compound 5g was obtained IC50 J774 = >400 µM, that represents the maximum concentration of the compound evaluated which did not affect the cells. Both showed good selectivity in the calculation of the ratio between the IC50 T. cruzi and IC50 J774. Additionally, we performed calculations of the physicochemical properties of three-dimensional structures of the compounds, followed by exploratory data analysis including hierarchical cluster analysis (HCA) and principal component analysis (PCA), which contributed to the identification of properties that influence the activity anti-T. cruzi in the series of compounds studied. The findings indicated a significant influence of ClogP and dipole moment properties, pointing out the need of a lipophilic/hydrophilic balance in the designing of novel anti-T. cruzi molecules

Nitro-Heterocyclic compounds induce apoptosis-like effects in Leishmania (L). amazonensis promastigotes

Abstract Background: Three drugs - pentavalent antimonials, amphotericin B and pentamidine - are currently used for leishmaniasis treatment. They are administered for long periods, only parenterally, and have high cardiac, renal and hepatic toxicities. Therefore, the investigation of new compounds is required. Nitro-heterocyclic derivatives have been used as possible drug candidates to treat diseases caused by trypanosomatids. Methods: Leishmania (L.) amazonensis promastigotes (MHO/BR/73/M2269), maintained in the Laboratório de Soroepidemiologia - Instituto de Medicina Tropical- USP, were exposed to five nitroheterocyclic derivatives, with differences at phenyl-ring position 4: BSF-C4H9, BSF-H, BSF-NO2, BSF-CH3 and BSF-Cl, for 48 hours. After analyzing viability (MTT assay), we evaluated cellular-morphology activity of compounds by transmission electron microscopy (TEM) and measurement of apoptosis (phosphatidylserine expression) by flow cytometry. Results: EC50 of amphotericin B and BSF-CH3 were 0.50 (M and 0.39 (M respective. Other nitro-heterocyclic compounds presented EC50 higher than amphotericin B. All compounds showed greater AV- and PI-positive expression than amphotericin B at 100 (M, except BSF-NO2. TEM showed complete nuclear disfigurement with 100 (M of BSF-NO2, 25 and 6.25 (M of BSF-H, and 6.25 (M BSF-Cl; presence of vesicles within the flagellar pocket with 25 (M BSF-H; alteration of the kinetoplast with 25 (M BSF-C4H9, 25 (M of BSF-H, 6.25 (M BSF-CH3 and 6.25 (M of BSF-Cl. Conclusions: Nitro-heterocyclic compounds have shown activity against promastigotes of L. amazonensis, at lower concentrations. However, improvement of compound scaffolds are needed to assist the elucidation of the mechanism of action and to achieve greater activity

BFD-22 a new potential inhibitor of BRAF inhibits the metastasis of B16F10 melanoma cells and simultaneously increased the tumor immunogenicity

Benzofuroxan is an interesting ring system, which has shown a wide spectrum of biological responses against tumor cell lines. We investigated, herein, the antitumor effects of benzofuroxan derivatives (BFDs) in vitro and in a melanoma mouse model. Cytotoxic effects of twenty-two BFDs were determined by MIT assay. Effects of BFD-22 in apoptosis and cell proliferation were evaluated using Annexin V-FITC/PI and CFSE staining. In addition, the effects in the cell cycle were assessed. Flow cytometry, western blot, and fluorescence microscopy analysis were employed to investigate the apoptosis-related proteins and the BRAF signaling. Cell motility was also exploited through cell invasion and migration assays. Molecular docking approach was performed in order to verify the BFD-22 binding mode into the ATP catalytic site of BRAF kinase. Moreover, the BFD-22 antitumor effects were evaluated in a melanoma murine model using B16F10. BFD-22 was identified as a potential hit against melanoma cells. BFD-22 induced apoptosis and inhibited cell proliferation of B16F10 cells. BFD-22 has suppressed, indeed, the migratory and invasive behavior of B16F10 cells. Cyclin D1 and CDK4 expression were reduced leading to cell cycle arrest at G0/G1 phase. Of note, phosphorylation of BRAF at Ser338 was strongly down-regulated by BFD-22 in B16F10 cells. The accommodation/orientation into the binding site of BRAF was similar of BAY43-9006 (co-crystallized inhibitor of BRAF, sorafenib). Importantly, BFD-22 presented in vivo antimetastatic effects and showed better therapeutic efficacy than sorafenib and taxol. BFD-22 can be considered as a new lead compound and, then, can be helpful for the designing of novel drug candidates to treat melanoma. (C) 2016 Elsevier Inc. All rights reserved.Sao Paulo Research Foundation (FAPESP)Brazilian Governmental Agency CNPqBrazilian Governmental Agency CAPESUniv Sao Paulo, Lab Tumor Immunol, Sao Paulo, SP, BrazilButantan Inst, Biochem & Biophys Lab, Sao Paulo, SP, BrazilUniv Groningen, Univ Med Ctr Groningen, Dept Med Oncol, Hanzepl 1, Groningen, NetherlandsUniv Sao Paulo, Lab Drug Design & Dev, Sao Paulo, SP, BrazilUniv Sao Paulo, Lab Cytopathol, Dept Clin Chem & Toxicol, Fac Pharmaceut Sci, Sao Paulo, SP, BrazilButantan Inst, Genet Lab, Sao Paulo, SP, BrazilUniv Fed Sao Paulo, Expt Oncol Sect, Sao Paulo, SP, BrazilUniv Sao Paulo, Cell & Mol Therapy Ctr NUCEL NETCEM, Sao Paulo, SP, BrazilUniv Fed Sao Paulo, Expt Oncol Sect, Sao Paulo, SP, BrazilFAPESP: 2009/54599-5FAPESP: 2013/07273-2FAPESP: 2013/05396-0FAPESP: 2014/07341-0FAPESP: 2014/14267-1Web of Scienc