Stereochemical insights into ß-amino-N-acylhydrazones and their impact on DPP-4 inhibition.

Dipeptidyl peptidase IV (DPP-4) is a key enzyme that regulates several important biological processes and it is better known to be targeted by gliptins as a modern validated approach for the management of type 2 diabetes mellitus (T2DM). However, new generations of DPP-4 inhibitors capable of controlling inflammatory processes associated with chronic complications of T2DM are still needed. In this scenario, we report here the design by molecular modelling of new ß-amino-N-acylhydrazones, their racemic synthesis, chiral resolution, determination of physicochemical properties and their DPP4 inhibitory potency. Theoretical and experimental approaches allowed us to propose a preliminary SAR, as well as to identify LASSBio-2124 (6) as a new lead for DPP-4 inhibition, with good physicochemical properties, favourable eudismic ratio, scalable synthesis and anti-diabetes effect in a proof-of-concept model. These findings represent an interesting starting point for the development of a new generation of DPP-4 inhibitors, useful in the treatment of T2DM and comorbidities.

Pharmacokinetic Profile Evaluation of Novel Combretastatin Derivative, LASSBio-1920, as a Promising Colorectal Anticancer Agent.

LASSBio-1920 was synthesized due to the poor solubility of its natural precursor, combretastatin A4 (CA4). The cytotoxic potential of the compound against human colorectal cancer cells (HCT-116) and non-small cell lung cancer cells (PC-9) was evaluated, yielding IC50 values of 0.06 and 0.07 µM, respectively. Its mechanism of action was analyzed by microscopy and flow cytometry, where LASSBio-1920 was found to induce apoptosis. Molecular docking simulations and the enzymatic inhibition study with wild-type (wt) EGFR indicated enzyme-substrate interactions similar to other tyrosine kinase inhibitors. We suggest that LASSBio-1920 is metabolized by O-demethylation and NADPH generation. LASSBio-1920 demonstrated excellent absorption in the gastrointestinal tract and high central nervous system (CNS) permeability. The pharmacokinetic parameters obtained by predictions indicated that the compound presents zero-order kinetics and, in a human module simulation, accumulates in the liver, heart, gut, and spleen. The pharmacokinetic parameters obtained will serve as the basis to initiate in vivo studies regarding LASSBio-1920's antitumor potential.

Methyl Effect on the Metabolism, Chemical Stability, and Permeability Profile of Bioactive N-Sulfonylhydrazones.

Sulfonylhydrazones are privileged structures with multifaceted pharmacological activity. Exploring the hypoglycemic properties of these organic compounds, we previously revealed a new series of N-sulfonylhydrazones (NSH) as antidiabetic drug candidates. Here, we evaluated the microsomal metabolism, chemical stability, and permeability profile of these NSH prototypes, focusing on the pharmacokinetic differences in N-methylated and non-N-methylated analogs. Our results demonstrated that the N-methylated analogs (LASSBio-1772 and LASSBio-1774) were metabolized by CYP, forming three and one metabolites, respectively. These prototypes exhibited chemical stability at pH 2.0 and 7.4 and brain penetration ability. On the other hand, non-N-methylated analogs (LASSBio-1771 and LASSBio-1773) were hydrolyzed in acid pH and could not cross the artificial blood-brain barrier. The cyano group in LASSBio-1771 was postulated as a possible site of interaction with the heme group, potentially inhibiting CYP enzymes. Moreover, prototypes with the methyl ester group were metabolized by carboxylesterase, and non-N-methylated analogs did not show oxidative metabolism. The prototypes (except LASSBio-1774) showed excellent gastrointestinal absorption. Altogether, our data support the idea that the methyl effect on NSH strongly alters their pharmacokinetic profile, enhances the recognition by CYP enzymes, promotes brain penetration, and plays a protective effect upon acid hydrolysis.

Natural products as new antimitotic compounds for anticancer drug development.

Cell cycle control genes are frequently mutated in cancer cells, which usually display higher rates of proliferation than normal cells. Dysregulated mitosis leads to genomic instability, which contributes to tumor progression and aggressiveness. Many drugs that disrupt mitosis have been studied because they induce cell cycle arrest and tumor cell death. These antitumor compounds are referred to as antimitotics. Vinca alkaloids and taxanes are natural products that target microtubules and inhibit mitosis, and their derivatives are among the most commonly used drugs in cancer therapy worldwide. However, severe adverse effects such as neuropathies are frequently observed during treatment with microtubule-targeting agents. Many efforts have been directed at developing improved antimitotics with increased specificity and decreased likelihood of inducing side effects. These new drugs generally target specific components of mitotic regulation that are mainly or exclusively expressed during cell division, such as kinases, motor proteins and multiprotein complexes. Such small molecules are now in preclinical studies and clinical trials, and many are products or derivatives from natural sources. In this review, we focused on the most promising targets for the development of antimitotics and discussed the advantages and disadvantages of these targets. We also highlighted the novel natural antimitotic agents under investigation by our research group, including combretastatins, withanolides and pterocarpans, which show the potential to circumvent the main issues in antimitotic therapy.

Natural products as new antimitotic compounds for anticancer drug development

Cell cycle control genes are frequently mutated in cancer cells, which usually display higher rates of proliferation than normal cells. Dysregulated mitosis leads to genomic instability, which contributes to tumor progression and aggressiveness. Many drugs that disrupt mitosis have been studied because they induce cell cycle arrest and tumor cell death. These antitumor compounds are referred to as antimitotics. Vinca alkaloids and taxanes are natural products that target microtubules and inhibit mitosis, and their derivatives are among the most commonly used drugs in cancer therapy worldwide. However, severe adverse effects such as neuropathies are frequently observed during treatment with microtubule-targeting agents. Many efforts have been directed at developing improved antimitotics with increased specificity and decreased likelihood of inducing side effects. These new drugs generally target specific components of mitotic regulation that are mainly or exclusively expressed during cell division, such as kinases, motor proteins and multiprotein complexes. Such small molecules are now in preclinical studies and clinical trials, and many are products or derivatives from natural sources. In this review, we focused on the most promising targets for the development of antimitotics and discussed the advantages and disadvantages of these targets. We also highlighted the novel natural antimitotic agents under investigation by our research group, including combretastatins, withanolides and pterocarpans, which show the potential to circumvent the main issues in antimitotic therapy.

Association between allergic responses and Schistosoma mansoni infection in residents in a low-endemic setting in Brazil

Introduction Schistosomiasis is endemic in 76 countries and territories. Several studies have found an inverse correlation between parasitic disease and the development of allergies. The purpose of the present study was to determine whether infection with Schistosoma mansoni in subjects with a low parasite load is protective against allergy. The final sample consisted of 39 S. mansoni-positive and 52 S. mansoni-negative residents of a small community in northeastern Brazil. Methods All subjects were submitted to the Kato-Katz test, anti-S. mansoni IgG measurement, the prick test for aeroallergens, eosinophil counts and serum IgE measurement. Results Subjects who reacted to one or more antigens in the prick test were considered allergic. Only 7 S. mansoni-positive subjects (17.9%) reacted to one or more antigens, whereas 20 S. mansoni-negative subjects (38.5%) tested positive for allergy. Conclusions Our findings suggest that, in areas of low endemicity, infection with S. mansoni significantly reduces the risk of the development of allergy in subjects with a low parasite load. .

Association between allergic responses and Schistosoma mansoni infection in residents in a low-endemic setting in Brazil.

INTRODUCTION: Schistosomiasis is endemic in 76 countries and territories. Several studies have found an inverse correlation between parasitic disease and the development of allergies. The purpose of the present study was to determine whether infection with Schistosoma mansoni in subjects with a low parasite load is protective against allergy. The final sample consisted of 39 S. mansoni-positive and 52 S. mansoni-negative residents of a small community in northeastern Brazil. METHODS: All subjects were submitted to the Kato-Katz test, anti-S. mansoni IgG measurement, the prick test for aeroallergens, eosinophil counts and serum IgE measurement. RESULTS: Subjects who reacted to one or more antigens in the prick test were considered allergic. Only 7 S. mansoni-positive subjects (17.9%) reacted to one or more antigens, whereas 20 S. mansoni-negative subjects (38.5%) tested positive for allergy. CONCLUSIONS: Our findings suggest that, in areas of low endemicity, infection with S. mansoni significantly reduces the risk of the development of allergy in subjects with a low parasite load.

A conventional polymerase chain reaction-based method for the diagnosis of human schistosomiasis in stool samples from individuals in a low-endemicity area

The aim of this study was to evaluate the efficacy of a polymerase chain reaction (PCR)-based method to detect Schistosoma mansoni DNA in stool samples from individuals living in a low-endemicity area in Brazil. Of the 125 initial stool samples, 80 were ELISA reactive and eggs were identified in 19 of the samples by parasitological examination. For the PCR evaluations, 56 stool samples were selected and divided into five groups. Groups I-IV were scored negative for S. mansoni eggs by parasitological examination. Groups I and II were ELISA reactive, whereas Groups III and IV were ELISA nonreactive. Groups II and III were positive for other intestinal parasites. PCR testing scored eight samples as positive from these four groups. Group V represented the S. mansoni -positive group and it included ELISA-reactive samples that were scored positive for S. mansoni by one or more parasitological examinations (6/19 were positive by Kato-Katz method, 9/17 by saline gradient and 10/13 by Helmintex®). PCR scored 13 of these 19 samples as positive for S. mansoni . We conclude that while none of these methods yielded 100% sensitivity, a combination of techniques should be effective for improving the detection of S. mansoni infection in low-endemicity areas.

A conventional polymerase chain reaction-based method for the diagnosis of human schistosomiasis in stool samples from individuals in a low-endemicity area.

The aim of this study was to evaluate the efficacy of a polymerase chain reaction (PCR)-based method to detect Schistosoma mansoni DNA in stool samples from individuals living in a low-endemicity area in Brazil. Of the 125 initial stool samples, 80 were ELISA reactive and eggs were identified in 19 of the samples by parasitological examination. For the PCR evaluations, 56 stool samples were selected and divided into five groups. Groups I-IV were scored negative for S. mansoni eggs by parasitological examination. Groups I and II were ELISA reactive, whereas Groups III and IV were ELISA nonreactive. Groups II and III were positive for other intestinal parasites. PCR testing scored eight samples as positive from these four groups. Group V represented the S. mansoni -positive group and it included ELISA-reactive samples that were scored positive for S. mansoni by one or more parasitological examinations (6/19 were positive by Kato-Katz method, 9/17 by saline gradient and 10/13 by Helmintex®). PCR scored 13 of these 19 samples as positive for S. mansoni . We conclude that while none of these methods yielded 100% sensitivity, a combination of techniques should be effective for improving the detection of S. mansoni infection in low-endemicity areas.

The combination of three faecal parasitological methods to improve the diagnosis of schistosomiasis mansoni in a low endemic setting in the state of Ceará, Brazil.

Laboratory diagnosis of intestinal schistosomiasis mansoni can be accomplished through various methods of stool examination to detect parasites, ranging from the most classic tests (Kato-Katz) to several methods that are still undergoing validation. This study was conducted to assess two new parasite identification methods for diagnosing schistosomiasis mansoni in residents of a low endemic area in the municipality of Maranguape, in the state of Ceará, Brazil using the Kato-Katz method as a reference and serology (enzyme-linked immunosorbent assay) for the screening of patients. The Kato-Katz, the saline gradient method and the Helmintex(®) method parasite identification methods were employed only in subjects who exhibited positive serologic tests. The test results were then analysed and treatment of positive individuals was subsequently performed. After comparing the test results, we observed that the saline gradient method and the Helmintex(®) method were more effective in diagnosing schistosomiasis mansoni in the study area compared with the Kato-Katz method.

The combination of three faecal parasitological methods to improve the diagnosis of schistosomiasis mansoni in a low endemic setting in the state of Ceará, Brazil

Laboratory diagnosis of intestinal schistosomiasis mansoni can be accomplished through various methods of stool examination to detect parasites, ranging from the most classic tests (Kato-Katz) to several methods that are still undergoing validation. This study was conducted to assess two new parasite identification methods for diagnosing schistosomiasis mansoni in residents of a low endemic area in the municipality of Maranguape, in the state of Ceará, Brazil using the Kato-Katz method as a reference and serology (enzyme-linked immunosorbent assay) for the screening of patients. The Kato-Katz, the saline gradient method and the Helmintex® method parasite identification methods were employed only in subjects who exhibited positive serologic tests. The test results were then analysed and treatment of positive individuals was subsequently performed. After comparing the test results, we observed that the saline gradient method and the Helmintex® method were more effective in diagnosing schistosomiasis mansoni in the study area compared with the Kato-Katz method.

Increased detection of schistosomiasis with Kato-Katz and SWAP-IgG-ELISA in a Northeastern Brazil low-intensity transmission area.

INTRODUCTION: The laboratory diagnosis of schistosomiasis is based mainly on the detection of parasite eggs in stool samples through the Kato-Katz (KK) technique, reading one slide by test. However, a widely known limitation of parasitological methods is reduced sensitivity, particularly in low endemic areas. METHODS: To increase sensitivity, we conducted further slide readings from the same stool sample using the parasitological method associated with a serological test. We used the KK method (three slides) and the IgG anti-Schistosoma mansoni-enzyme-linked immunosorbent assay (ELISA) technique to diagnose schistosomiasis in low endemic areas in the Brazilian State of Ceará. Fecal samples and sera from 250 individuals were analyzed. RESULTS: Sixteen percent and 47.2% of samples were positive in parasitological tests and serological tests, respectively. Parasitological methods showed that 32 (80%) individuals tested positive on the first slide, 6 (15%) on the second slide, and 2 (5%) on the third. The performance of the ELISA test in the diagnosis, using the KK method as diagnostic reference, showed a negative predictive value of 100%, with specificity and positive predictive values of 62.8% and 33.9%, respectively. CONCLUSIONS: In this study, the increase from one to three slides analyzed per sample using the KK technique was shown to be a useful procedure for increasing the diagnostic sensitivity of this technique.

Increased detection of schistosomiasis with Kato-Katz and SWAP-IgG-ELISA in a Northeastern Brazil low-intensity transmission area / Aumento da detecção da esquistossomose com Kato-Katz e SWAP-IgG-ELISA, em área de baixa endemicidade, no nordeste do Brasil

INTRODUCTION: The laboratory diagnosis of schistosomiasis is based mainly on the detection of parasite eggs in stool samples through the Kato-Katz (KK) technique, reading one slide by test. However, a widely known limitation of parasitological methods is reduced sensitivity, particularly in low endemic areas. METHODS: To increase sensitivity, we conducted further slide readings from the same stool sample using the parasitological method associated with a serological test. We used the KK method (three slides) and the IgG anti-Schistosoma mansoni-enzyme-linked immunosorbent assay (ELISA) technique to diagnose schistosomiasis in low endemic areas in the Brazilian State of Ceará. Fecal samples and sera from 250 individuals were analyzed. RESULTS: Sixteen percent and 47.2% of samples were positive in parasitological tests and serological tests, respectively. Parasitological methods showed that 32 (80%) individuals tested positive on the first slide, 6 (15%) on the second slide, and 2 (5%) on the third. The performance of the ELISA test in the diagnosis, using the KK method as diagnostic reference, showed a negative predictive value of 100%, with specificity and positive predictive values of 62.8% and 33.9%, respectively. CONCLUSIONS: In this study, the increase from one to three slides analyzed per sample using the KK technique was shown to be a useful procedure for increasing the diagnostic sensitivity of this technique.

INTRODUÇÃO: O diagnóstico laboratorial da esquistossomose é baseada principalmente na detecção de ovos do parasito nas fezes, realizada pela técnica de Kato-Katz (KK), com a leitura de uma lâmina por teste. No entanto, uma limitação conhecida dos métodos coproscópicos é a reduzida sensibilidade, especialmente nas áreas de baixa endemicidade. MÉTODOS: A fim de reduzir essa limitação, realizamos mais leituras da mesma amostra de fezes pelo método coproscópico e associamos a um teste sorológico.Utilizamos o método de KK (três lâminas) e a técnica de IgG-ELISA, buscando aumentar a sensibilidade do diagnóstico da esquistossomose em área de baixa endemicidade, no Estado Brasileiro do Ceará. Amostras de fezes e soro de 250 indivíduos foram analisadas. RESULTADOS: Destas, 40 e 118 foram positivas nos testes coproscópico e sorológico, respectivamente. Na coproscopia, 32 (80%) indivíduos tiveram testes positivos na primeira lâmina, 6 (15%) na segunda e 2 (5%) apenas na terceira lâmina. O desempenho do teste de ELISA no diagnóstico utilizando-se o método KK como referência de diagnóstico, demonstrou valor preditivo negativo de 100% mas a especificidade e o valor preditivo positivo foram de 62,8% e 33,9%, respectivamente. CONCLUSÕES: Neste estudo, o aumento de uma para três lâminas analisadas por amostra pelo KK, mostrou ser um procedimento útil para o aumento da sensibilidade diagnóstica desta técnica.

Combination of Kato-Katz faecal examinations and ELISA to improve accuracy of diagnosis of intestinal schistosomiasis in a low-endemic setting in Brazil.

Considering the decrease of disease burden caused by intestinal schistosomiasis in many endemic settings, more sensitive diagnostic methods are needed to plan and monitor control measures. We conducted a cross-sectional survey in a rural community in northeast Brazil (317 inhabitants). A combined approach including repeated faecal examinations and ELISA testing was applied. In a first round, single stool samples were collected from 305 (96.2%) participants. Three Kato-Katz (KK) smears were prepared from each sample, and IgG ELISA was performed from serum samples. In the 85 cases of negative KK smears, but positive ELISA results, three additional faecal samples were collected in a second round, and another five KK smears prepared. In the first round of KK analysis, 11/287 (3.8%; 95% confidence interval; 1.92-6.75) were positive. After examining up to eight smears per individual (second round), prevalence of schistosomiasis increased to 8.7% (95% confidence interval: 5.9-12.5). In total, 96/287 (33.4%, 95% confidence interval: 28.0-39.2) samples were positive by ELISA testing. There were no false negative ELISA results. Specificity, positive and negative predictive values of ELISA as compared to up to eight KK smears from three stool samples (reference diagnosis) were 72.9%, 26.0% and 100%, respectively. A single KK smear detected only 12% of the 25 infections; this increased to 44% (three smears, one stool sample), 84% (five smears, three stool samples) and 96% (six smears, four stool samples). We conclude that in low-endemic areas in Brazil the use of KK continues being an important tool. The additional benefit of preparing more than six KK smears from repeated stool samples is negligible. ELISA may be useful for screening populations, with subsequent confirmation of diagnosis by KK or other more sensitive, but highly specific methods.