3-Alkoxy-1-Benzyl-5-Nitroindazole Derivatives Are Potent Antileishmanial Compounds.

Indazoles have previously been identified as molecules with antiprotozoal activity. In this study, we evaluate the in vitro activity of thirteen 3-alkoxy-1-benzyl-5-nitroindazole derivatives (series D) against L. amazonensis, L. infantum, and L. mexicana. In vitro, cytotoxicity against mouse peritoneal macrophages and growth inhibitory activity in promastigotes were evaluated for all compounds, and those showing adequate activity and selectivity were tested against intracellular amastigotes. Transmission and scanning electron microscopy were employed to study the effects of 3-alkoxy-1-benzyl-5-nitroindazole and 2-benzyl-5-nitroindazolin-3-one derivatives on promastigotes of L. amazonensis. Compounds NV6 and NV8 were active in the two life stages of the three species, with the latter showing the best indicators of activity and selectivity. 3-alkoxy-1-benzyl-5-nitroindazole derivatives (series D) showed in vitro activity comparable to that of amphotericin B against the promastigote stage of Leishmania spp. Two compounds were also found to be active the amastigote stage. Electron microscopy studies confirmed the antileishmanial activity of the indazole derivatives studied and support future research on this family of compounds as antileishmanial agents.

Detection and counting of Leishmania intracellular parasites in microscopy images.

Problem: Leishmaniasis is a disease caused by protozoan parasites of the genus Leishmania and has a high prevalence and impact on global health. Currently, the available drugs for its treatment have drawbacks, such as high toxicity, resistance of the parasite, and high cost. Therefore, the search for new, more effective, and safe drugs is a priority. The effectiveness of an anti-leishmanial drug is analyzed through in vitro studies in which a technician manually counts the intracellular form of the parasite (amastigote) within macrophages, which is slow, laborious, and prone to errors. Objectives: To develop a computational system that facilitates the detection and counting of amastigotes in microscopy images obtained from in vitro studies using image processing techniques. Methodology: Segmentation of objects in the microscope image that might be Leishmania amastigotes was performed using the multilevel Otsu method on the saturation component of the hue, saturation, and intensity color model. In addition, morphological operations and the watershed transform combined with the weighted external distance transform were used to separate clustered objects. Then positive (amastigote) objects were detected (and consequently counted) using a classifier algorithm, the selection of which as well as the definition of the features to be used were also part of this research. MATLAB was used for the development of the system. Results and discussion: The results were evaluated in terms of sensitivity, precision, and the F-measure and suggested a favorable effectiveness of the proposed method. Conclusions: This system can help researchers by allowing large volumes of images of amastigotes to be counted using an automatic image analysis technique.

In Vitro and In Vivo Antileishmanial Activity of Thioridazine.

INTRODUCTION: Leishmaniasis is a neglected disease with high prevalence and incidence in tropical and subtropical areas. Existing drugs are limited due to cost, toxicity, declining efficacy and unavailability in endemic places. Drug repurposing has established as an efficient way for the discovery of drugs for a variety of diseases. PURPOSE: The objective of the present work was testing the antileishmanial activity of thioridazine, an antipsychotic agent with demonstrated effect against other intracellular pathogens. METHODS: The cytotoxicity for mouse peritoneal macrophages as well as the activity against Leishmania amazonensis, Leishmania mexicana and Leishmania major promastigotes and intracellular amastigotes, as well as in a mouse model of cutaneous leishmaniasis, were assessed. RESULTS: Thioridazine inhibited the in vitro proliferation of promastigotes (50% inhibitory concentration-IC50-values in the range of 0.73 µM to 3.8 µM against L. amazonensis, L. mexicana and L. major) and intracellular amastigotes (IC50 values of 1.27 µM to 4.4 µM for the same species). In contrast, in mouse peritoneal macrophages, the 50% cytotoxic concentration was 24.0 ± 1.89 µM. Thioridazine inhibited the growth of cutaneous lesions and reduced the number of parasites in the infected tissue of mice. The dose of thioridazine that inhibited lesion development by 50% compared to controls was 23.3 ± 3.1 mg/kg and in terms of parasite load, it was 11.1 ± 0.97 mg/kg. CONCLUSIONS: Thioridazine was effective against the promastigote and intracellular amastigote stages of three Leishmania species and in a mouse model of cutaneous leishmaniasis, supporting the potential repurposing of this drug as an antileishmanial agent.

Antileishmanial activity of 5-nitroindazole derivatives.

Background: Currently, there is no safe and effective vaccine against leishmaniasis and existing therapies are inadequate due to high toxicity, cost and decreased efficacy caused by the emergence of resistant parasite strains. Some indazole derivatives have shown in vitro and in vivo activity against Trichomonas vaginalis and Trypanosoma cruzi. On that basis, 20 indazole derivatives were tested in vitro against Leishmania amazonensis. Objective: To evaluate the in vitro activity of twenty 2-benzyl-5-nitroindazolin-3-one derivatives against L. amazonensis. Design: For the selection of promising compounds, it is necessary to evaluate the indicators for in vitro activity. For this aim, a battery of studies for antileishmanial activity and cytotoxicity were implemented. These results enabled the determination of the substituents in the indazole derivatives responsible for activity and selectivity, through the analysis of the structure-activity relationship (SAR). Methods: In vitro cytotoxicity against mouse peritoneal macrophages and growth inhibitory activity in promastigotes were evaluated for 20 compounds. Compounds that showed adequate selectivity were tested against intracellular amastigotes. The SAR from the results in promastigotes was represented using the SARANEA software. Results: Eight compounds showed selectivity index >10% and 50% inhibitory concentration <1 µM against the promastigote stage. Against intracellular amastigotes, four were as active as Amphotericin B. The best results were obtained for 2-(benzyl-2,3-dihydro-5-nitro-3-oxoindazol-1-yl) ethyl acetate, with 50% inhibitory concentration of 0.46 ± 0.01 µM against amastigotes and a selectivity index of 875. The SAR study showed the positive effect on the selectivity of the hydrophilic fragments substituted in position 1 of 2-benzyl-5- nitroindazolin-3-one, which played a key role in improving the selectivity profile of this series of compounds. Conclusion: 2-bencyl-5-nitroindazolin-3-one derivatives showed selective and potent in vitro activity, supporting further investigations on this family of compounds as potential antileishmanial hits.

Evaluación de la toxicidad del desodorante RLV en ojos / Evaluation of RLV deodorant toxicity in eyes

Se realizó la evaluación del posible efecto irritante de un formulado (RLV) que se empleará como desodorante y que contiene como principio activo la hexamina, la cual es empleada como antiséptico urinario. Este formulado se aplicó por vía oftálmica en 6 conejos de la raza Nueva Zelandia, durante 7 días. Las valoraciones se basan en las observaciones macroscópicas de los posibles efectos adversos que se presentan en las estructuras oculares. Para esta evaluación nos basamos en el método propuesto por Draize, así como las guías de la OECD, de acuerdo con los resultados obtenidos la forma farmacéutica elaborada a una concentración del 3 (por ciento) para el desodorante, no ocasiona irritación en las estructuras oculares(AU)

Evaluación de la toxicidad del desodorante RLV en ojos / Evaluation of RLV deodorant toxicity in eyes

Se realizó la evaluación del posible efecto irritante de un formulado (RLV) que se empleará como desodorante y que contiene como principio activo la hexamina, la cual es empleada como antiséptico urinario. Este formulado se aplicó por vía oftálmica en 6 conejos de la raza Nueva Zelandia, durante 7 días. Las valoraciones se basan en las observaciones macroscópicas de los posibles efectos adversos que se presentan en las estructuras oculares. Para esta evaluación nos basamos en el método propuesto por Draize, así como las guías de la OECD, de acuerdo con los resultados obtenidos la forma farmacéutica elaborada a una concentración del 3 (por ciento) para el desodorante, no ocasiona irritación en las estructuras oculares