Three molecular hybrids containing 1,4-naphthoquinones, 1,3,5-triazines, morpholine and 7-chloroquinoline, which have recognized contributions to the biological activity of many drugs, were synthesized in yields ranging from 43-84%. All hybrids were obtained in three steps starting from readily available reactants: lawsone, cyanuric chloride, morpholine and 4,7-dichloroquinoline. A previous docking study was carried out to identify the binding energy and pharmacophore conformation of the promising anticancer compounds with PI3Kγ (phosphoinositide 3-kinase) and AMPK (5’ AMP-activated protein kinase). The cancer activity in human metastatic melanoma cells (SKMEL-103) were performed, and the synthetized compounds presented half maximal inhibitory concentration (IC50) values around 25 µM. The expressions of PI3K and AMPK were also determined using western blotting technique, and all molecular hybrids negatively modulated both targets.

Dengue virus (DENV) has been characterized as having great clinical importance in the world, as there is no specific treatment against this virus. The NS2B-NS3pro complex is essential for the replication and maturation of DENV and is a potential pharmacological target. The present study aims to evaluate and understand the interactions and affinities (via molecular docking/AutoDock Vina) of 16 peptidomimetic derivatives applied to a NS2B-NS3pro DENV-2 complex constructed by homology modeling (via SWISS-MODEL). Two compounds were selected as potential inhibitors of this protein complex. In addition, these compounds possess important interactions involving Ser135, Gly169 and Tyr161, which have been described previously to be fundamental to the recognition of inhibitors directed to this receptor. Thus, the involvement of these residues is significant pharmacologically because they may contribute to the inhibitory action of this molecular target against DENV.

Hydroxynaphthoquinones such as lawsone (2-hydroxy-1,4-naphthoquinone) have proven to be effective antifungal agents. These compounds were tested for antifungal activity against yeast standard and clinical strains by the broth microdilution method. Among the synthetic lawsone derivatives, 2-hydroxy-3-((2-hydroxyphenyl)(pyrrolidin-1-yl)methyl)naphthalene-1,4-dione, 2-hydroxy-3-(((4-nitrophenyl)amino)(phenyl)methyl)naphthalene-1,4-dione and 2-hydroxy-3-((2-hydroxyphenyl)((4-nitrophenyl)amino)methyl)naphthalene-1,4-dione showed high activity against Candida albicans ATCC 10231, with minimal inhibitory concentrations (MICs) and minimal fungicidal concentrations (MFCs) ranging from 20 to 330 and from 80 to 330 µg mL-1, respectively. Moreover, they also showed a mechanism of action on exogenous ergosterol. Therapeutic concentrations (CC50) of 2-hydroxy-3-((2-hydroxyphenyl)(pyrrolidin-1-yl)methyl)naphthalene-1,4-dione, 2-hydroxy-3-(((4-nitrophenyl)amino)(phenyl)methyl)naphthalene-1,4-dione and 2-hydroxy-3-((2-hydroxyphenyl)((4-nitrophenyl)amino)methyl)naphthalene-1,4-dione were 52.81, 52.58 and 85.94 µg mL-1, respectively, which can be considered moderate or low. In addition, docking studies showed that these compounds had similar binding energy to standard ketoconazole, which are recognized as the molecular target by van der Waals interactions. Furthermore, they are under Lipinski's rule of 5 with a druglikeness score better than ketoconazole and nystatin. These findings suggest that 2-hydroxy-3-((2-hydroxyphenyl)(pyrrolidin-1-yl)methyl)naphthalene-1,4-dione, 2-hydroxy-3-(((4-nitrophenyl)amino)(phenyl)methyl)naphthalene-1,4-dione and 2-hydroxy-3-((2-hydroxyphenyl)((4-nitrophenyl)amino)methyl)naphthalene-1,4-dione have potential as leading compounds against human fungal infections.

We herein report the synthesis of hybrid kauranoid molecules of type 1,2,3-triazole-1,4-disubstituted aiming to improve the antimalarial activity of kaurenoic and xylopic acids. The CuI-catalyzed cycloaddition of azides and kauranoid terminal alkynes was explored as a hybridization strategy. Kauranoid terminal alkynes were prepared from kaurenoic and xylopic acids that were isolated from Wedelia paludosa D. C. (Asteraceae) and Xylopia frutescens Aubl. (Annonaceae). A total of 15 kauranoid derivatives, including nine new triazoles, were obtained and five out of these were more active than the original diterpenes. Interestingly, an increased activity was observed for a kauranoid propargyl ether. Interaction between ent-kaurane diterpene derivatives and Ca2+-ATPase (PfATP6) was investigated. Synthesis of diterpene derivatives emerges as a possible route to be explored in the quest of potentially new inhibitors of PfATP6.

Eleven 2,3-(substituted)-1,4-naphthoquinone derivatives were synthesized in yields ranging from 52-89%. These derivatives were evaluated for their cytotoxic effects on human lungs (H460), triple-negative breast (MDA-MB-231) and ovarian (A2780) cancer cell lines. Compounds 5f and 8 showed IC50values of 3.048 × 10-5 mol L-1 and 4.24 × 10-6 mol L-1 for H460; 5c and 8showed IC50 values of 2.16 × 10-5 mol L-1 and 1.60 × 10-5 mol L-1 for MDA-MB-231, and 5gand 8 showed IC50 values of 2.68 × 10-6 mol L-1 and 3.89 × 10-6 mol L-1 for A2780. Additionally, we conducted a docking study with the four most active compounds and the therapeutic targets PI3K and topoisomerase II showing the pharmacophoric conformation of these compounds.

A enzima quitinase de Moniliophthora perniciosa, o fungo causador da doença vassoura de bruxa em Theobroma cacao, foi parcialmente purificada por precipitação com sulfato de amônio e filtração em coluna de Sephacryl S-200 usando fosfato de sódio como tampão de extração. A metodologia de Superfície de resposta (MSR) foi usada para determinar o pH e temperatura ótimos. Foram isoladas quatro quitinases diferentes: ChitMp I, ChitMp II, ChitMp III e ChitMp IV. A ChitMp I mostrou temperatura ótima de 44-73ºC e pH ótimo de 7,0-8,4. A ChitMp II apresentou temperatura ótima entre 45-73ºC e pH ótimo de 7,0-8,4. ChitMp III apresentou temperatura ótima de 54-67ºC e pH ótimo de 7,3-8,8. ChitMp IV apresentou temperatura ótima de 60ºC e pH ótimo em 7,0. Em relação à biologia computacional a sequência primária foi feita in silico usando banco de dados do Projeto Genoma/Proteoma do M. perniciosa, obtendo-se uma sequência com 564 pb e 188 aminoácidos, que foi utilizada para a obtenção da estrutura tridimensional por metodologia de modelagem comparativa. Os modelos gerados foram submetidos à validação usando Procheck 3.0 and ANOLEA. O modelo proposto para a quitinase foi submetido à análise dinâmica por 1 ns, resultando em um modelo com 91,7% de seus resíduos em locais regiões favoráveis no gráfico de Ramachandran e RMS de 2,68.

The enzyme chitinase from Moniliophthora perniciosa the causative agent of the witches' broom disease in Theobroma cacao, was partially purified with ammonium sulfate and filtration by Sephacryl S-200 using sodium phosphate as an extraction buffer. Response surface methodology (RSM) was used to determine the optimum pH and temperature conditions. Four different isoenzymes were obtained: ChitMp I, ChitMp II, ChitMp III and ChitMp IV. ChitMp I had an optimum temperature at 44-73ºC and an optimum pH at 7.0-8.4. ChitMp II had an optimum temperature at 45-73ºC and an optimum pH at 7.0-8.4. ChitMp III had an optimum temperature at 54-67ºC and an optimum pH at 7.3-8.8. ChitMp IV had an optimum temperature at 60ºC and an optimum pH at 7.0. For the computational biology, the primary sequence was determined in silico from the database of the Genome/Proteome Project of M. perniciosa, yielding a sequence with 564 bp and 188 amino acids that was used for the three-dimensional design in a comparative modeling methodology. The generated models were submitted to validation using Procheck 3.0 and ANOLEA. The model proposed for the chitinase was subjected to a dynamic analysis over a 1 ns interval, resulting in a model with 91.7% of the residues occupying favorable places on the Ramachandran plot and an RMS of 2.68.

A enzima UDP-N-acetilglicosamina pirofosforilase de Moniliophthora perniciosa (CCMB 0257), o fungo patogênico causador da doença vassoura-de-bruxa do Theobroma cacao, foi parcialmente purificada por precipitação com sulfato de amônio e cromatografia de gel filtração em Sephacryl S-200. O tampão de extração da enzima foi o fosfato de sódio, 0,050 mol L-1, pH 7,0, contendo 1,0 mol L-1 de NaCl. A metodologia de superfície de resposta (MSR) foi usada para a obtenção do pH e temperatura ótima. Os resultados mostraram quatro diferentes isoenzimas (PyroMp I, PyroMp II, PyroMp III e PyroMp IV) que apresentaram pH ótimo na faixa de 6,9-8,4 e temperatura ótima variando entre 28 a 68 °C. A estrutura 3D de pirofosforilase de M. perniciosa foi obtida por modelagem comparativa. O modelo obtido mostrou uma boa qualidade, possuindo 78,6% de aminoácidos nas regiões energeticamente favoráveis. O modelo foi então submetido a simulações de dinâmica molecular (DM). O modelo apresentou uma boa qualidade geométrica após as simulações de DM (91,1% -gráfico de Ramachandran). A procura pelo sítio ativo da enzima mostrou que este é mantido extremamente conservado. Este modelo pode ser útil para desenvolvimento de inibidores contra a doença vassoura de bruxa.

The enzyme UDP-N-acetylglucosamine pyrophosphorylase (PyroMp) from Moniliophthora perniciosa (CCMB 0257), a pathogenic fungal strain and the causative agent of the witches' broom disease in Theobroma cacao, was partially purified by precipitation with ammonium sulfate and gel filtration on Sephacryl S-200. The buffer for enzyme extraction was sodium phosphate, 0.050 mol L-1, pH 7.0, containing 1.0 mol L-1 NaCl. Response surface methodology (RSM) was used to determine the optimum pH and temperature conditions. Four different isoenzymes (PyroMp I, PyroMp II, PyroMp III and PyroMp IV) were obtained with optimal pH ranging from 6.9-8.4 and optimum temperature ranging from 28 to 68 °C. The 3D structure of pyrophosphorylase of M. perniciosa was determined by comparative modeling. The model obtained showed a good quality, possessing 78.6% of amino acids in energetically allowed regions. The model was then submitted for DM simulation and showed a good geometric quality (91.1% Ramachandran plot). The active site of the enzyme was found to be extremely well conserved. This model will be useful for developing new inhibitors against witches' broom disease.

A enzima glucanase de Moniliophthora perniciosa foi produzida em meio líquido e purificada a partir do sobrenadante da cultura. A metodologia de superfície de resposta (MSR) foi usada para avaliar os efeitos das variáveis, incluindo indutor (extrato de levedura) e tempo de fermentação, na atividade da glucanase de M. perniciosa detectada no meio de cultura. A enzima presente no sobrenadante foi purificada em duas etapas: precipitação com sulfato de amônio (70%) e cromatografia de filtração em gel em Sephacryl S-200. A produção da enzima glucanase foi maior na concentração de 5,9 g L-1 de extrato de levedura e 13 dias de fermentação. Os resultados mostraram três diferentes isoformas (GLUI, GLUII e GLUIII) com fatores de purificação de 4,33, 1,86 e 3,03, respectivamente. O extrato enzimático parcialmente purificado mostrou um pH ótimo de 5,0 e uma temperatura ótima de 40°C. A atividade enzimática aumentou na presença de KCl em todas as concentrações estudadas. A atividade da glucanase foi maior na presença de NaCl 0,2 M. A enzima apresentou alta estabilidade térmica, perdendo apenas 10,20% de sua atividade específica após 40 minutos de incubação a 90°C. Os resultados de termoestabilidade e a atividade em baixo pH mostraram que a enzima glucanase de M. perniciosa tem características promissoras para futuras aplicações industriais.

The enzyme glucanase from Moniliophthora perniciosa was produced in liquid medium and purified from the culture supernatant. A multivariate statistical approach (Response Surface Methodology - RSM) was employed to evaluate the effect of variables, including inducer (yeast extract) and fermentation time, on secreted glucanase activities M. perniciosa detected in the culture medium. The crude enzyme present in the supernatant was purified in two steps: precipitation with ammonium sulfate (70%) and gel filtration chromatography on Sephacryl S-200. The best inducer and fermentation time for glucanase activities were 5.9 g L-1 and 13 days, respectively. The results revealed three different isoforms (GLUI, GLUII and GLUIII) with purification factors of 4.33, 1.86 and 3.03, respectively. The partially purified enzymatic extract showed an optimum pH of 5.0 and an optimum temperature of 40°C. The enzymatic activity increased in the presence of KCl at all concentrations studied. The glucanase activity was highest in the presence of 0.2 M NaCl. The enzyme showed high thermal stability, losing only 10.20% of its specific activity after 40 minutes of incubation at 90°C. A purified enzyme with relatively good thermostability that is stable at low pH might be used in future industrial applications.