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Epimastigote and trypomastigote forms of Trypanosoma cruzi attach to the macrophage surface and are internalized with the formation of a membrane bounded vacuole, known as the parasitophorous vacuole (PV). In order to determine if components of the host cell membrane are internalized during formation of the PV we labeled the macrophage surface with fluorescent probes for proteins, lipids and sialic acid residues and then allowed the labeled cells to interact with the parasites. The interaction process was interrupted after 1 hr at 37ºC and the distribution of the probes analyzed by confocal laser scanning microscopy. During attachment of the parasites to the macrophage surface an intense labeling of the attachment regions was observed. Subsequently labeling of the membrane lining the parasitophorous vacuole containing epimastigote and trypomastigote forms was seen. Labeling was not uniform, with regions of intense and light or no labeling. The results obtained show that host cell membrane lipids, proteins and sialoglycoconjugates contribute to the formation of the membrane lining the PV containing epimastigote and trypomastigote T. cruzi forms. Lysosomes of the host cell may participate in the process of PV membrane formation.

Ethnopharmacological Relevance: Acmella oleracea is traditionally used by Amazonian folks to treat skin and mucous wounds, influenza, cough, toothache, bacterial and fungal infections. Its phytoconstituents, such as alkylamides, phenolic compounds, and terpenes, are reported to produce therapeutic effects, which justify the medicinal use of A. oleracea extracts. However, the scientific evidence supporting the application A. oleracea bioactive products for wound treatment of remains unexplored so far., Objective: This work aimed to characterize the phytochemical composition of methanolic extract of A. oleracea leaves (AOM) and to investigate their antioxidant, anti-inflammatory, antimicrobial and healing potential focusing on its application for wound healing., Material and Methods: The dried leaves from A. oleracea submitted to static maceration in methanol for 40 days. The phytochemical constitution of AOM was analyzed based on the total phenolic dosage method and by UFLC-QTOF-MS analysis. Antioxidant activity was assessed by DPPH and NO scavenging activities, as well as MDA formation, evaluation of ROS levels, and phosphomolybdenum assays. In vitro anti-inflammatory activities were assessed by reduction of NO, IL-6, and TNF-α production and accumulation of LDs in peritoneal macrophages cells. Antimicrobial activity was evaluated by determining MIC and MBC/MFC values against P. aeruginosa, E. coli, S. epidermidis, S. aureus and C. albicans, bacterial killing assay, and biofilm adhesion assessment. In vitro wound healing activity was determined by means of the scratch assay with L929 fibroblasts., Results: Vanillic acid, quercetin, and seven other alkamides, including spilanthol, were detected in the UFLC-QTOF-MS spectrum of AOM. Regarding the biocompatibility, AOM did not induce cytotoxicity in L929 fibroblasts and murine macrophages. The strong anti-inflammatory activity was evidenced by the fact that AOM reduced the cellular production of inflammatory mediators IL-6, TNF-α, NO, and LDs in macrophages by 100%, 96.66 ± 1.95%, 99.21 ± 3.82%, and 67.51 ± 0.72%, respectively. The antioxidant effects were confirmed, since AOM showed IC 50 values of 44.50 ± 4.46 and 127.60 ± 14.42 μg/mL in the DPPH and NO radical inhibition assays, respectively. Additionally, AOM phosphomolybdenium reducing power was 63.56 ± 13.01 (RAA% of quercetin) and 104.01 ± 21.29 (RAA% of rutin). Finally, in the MDA quantification assay, AOM showed 63,69 ± 3.47% of lipid peroxidation inhibition. It was also observed that the production of ROS decreased by 69.03 ± 3.85%. The MIC values of AOM ranged from 1000 to 125 μg/mL. Adhesion of S. aureus, P. Aeruginosa, and mixed biofilms was significantly reduced by 44.71 ± 4.44%, 95.50 ± 6.37 %, and 51.83 ± 1.50%, respectively. AOM also significantly inhibited the growth of S. aureus (77.17 ± 1.50 %) and P. aeruginosa (62.36 ± 1.01%). Furthermore, AOM significantly enhanced the in vitro migration of L929 fibroblasts by 97.86 ± 0.82% compared to the control (P < 0.05)., Conclusions: This study is the first to report total antioxidant capacity and intracellular LD reduction by AOM. The results clearly demonstrated that AOM exerts potent anti-inflammatory, antioxidant, antimicrobial, and wound healing effects, encouraging its further investigation and promising application in wound treatment., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Etnopharmacological Relevance: Lantana camara L. is a species known for its broad spectrum of bioactivities and is commonly used in folk therapy to address inflammatory, dermatological, gastrointestinal, intestinal worms and protozoan diseases. It boasts a diverse array of secondary metabolites such as terpenes, flavonoids, and saponins. However, despite its rich chemical profile, there remains a scarcity of studies investigating its antileishmanial properties., Aim of the Study: This research aims to explore the antileishmanial potential of L. camara, focusing also on its mechanism of action against Leishmania amazonensis., Material and Methods: The ethanolic extract of L. camara leaves (LCE) was obtained through static maceration, and its phytoconstituents were identified using UFLC-QTOF-MS. The colorimetric MTT method was conducted to determine the effect of LCE on promastigotes of L. amazonensis and murine macrophages. The anti-amastigote activity was evaluated by counting intracellular parasites in macrophages after Giemsa staining. Additionally, investigations into the mechanisms underlying its action were conducted using cellular and biochemical approaches., Results: LCE exhibited significant activity against both promastigotes and intracellular amastigotes of L. amazonensis, with IC 50 values of 12.20 μg/mL ± 0.12 and 7.09 μg/mL ± 1.24, respectively. These IC 50 values indicate very promising antileishmanial activity, comparable to those found for the positive control miltefosine (5.10 μg/mL ± 1.79 and 8.96 μg/mL ± 0.50, respectively). Notably, LCE exhibited negligible cytotoxicity on macrophages (IC 50  = 223.40 μg/mL ± 47.02), demonstrating selectivity towards host cells (SI = 31.50). The antileishmanial activity of LCE involved a multi-targeted cell death process, characterized by morphological and ultrastructural alterations observed through SEM and TEM analyses, as well as oxidative effects evidenced by the inhibition of trypanothione reductase, elevation of ROS and lipid levels, and mitochondrial dysfunction evaluated using DTNB, H 2 DCFDA, Nile red, and JC-1 assays. Additionally, extraction of ergosterol and double labeling with annexin V and PI revealed modifications to the organization and permeability of the treated parasite's plasma membrane. LCE was found to consist predominantly of terpenes, with lantadenes A, B, and C being among the eleven compounds identified through UFLC-QTOF-MS analysis., Conclusions: The extract of L. camara presents a diverse array of chemical constituents, prominently featuring high terpene content, which may underlie its antileishmanial properties through a combination of apoptotic and non-apoptotic mechanisms of cell death induced by LCE. This study underscores the therapeutic potential of L. camara as a candidate for antileishmanial treatment, pending further validation., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Leishmaniasis is a disease caused by protozoa Leishmania spp., considered as a significant and urgent public health problem mainly in developing countries. In the absence of an effective vaccine, the treatment of infected people is one of the most commonly prophylactic measures used to control this disease. However, the therapeutic arsenal is reduced to a few drugs, with serious side effects and variability in efficacy. Attempting to this problem, in this work, a series of benzothiazole derivatives was synthetized and assayed against promastigotes and intracellular amastigotes of L. amazonensis, as well as the toxicity on macrophages. In addition, studies about the mechanism of action were also performed. Among the synthesized molecules, the substitution at position 4 of the aromatic ring appears to be critical for activity. The best compound exhibited IC 50 values of 28.86 and 7.70 μM, against promastigotes and amastigotes of L. amazonensis, respectively, being more active than miltefosine, used as reference drug. The in silico analysis of physicochemical and pharmacokinetic (ADMET) properties of this compound suggested a good profile of oral bioavailability and safety. In conclusion, the strategy of using benzothiazole nucleous in the search for new antileishmanial agents was advantageous and preliminar data provide information about the mechanism of action as well as in silico parameters suggest a good profile for preclinical studies., (© 2024 John Wiley & Sons Ltd.)

Despite efforts, available alternatives for the treatment of leishmaniasis are still scarce. In this work we tested a class of 15 quinolinylhydrazone analogues and presented data that support the use of the most active compound in cutaneous leishmaniasis caused by Leishmania amazonensis. In general, the compounds showed activity at low concentrations for both parasitic forms (5.33-37.04 μM to promastigotes, and 14.31-61.98 μM to amastigotes). In addition, the best compound (MHZ15) is highly selective for the parasite. Biochemical studies indicate that the treatment of promastigotes with MHZ15 leads the loss of mitochondrial potential and increase in ROS levels as the primary effects, which triggers accumulation of lipid droplets, loss of plasma membrane integrity and apoptosis hallmarks, including DNA fragmentation and phosphatidylserine exposure. These effects were similar in the intracellular form of the parasite. However, in this parasitic form there is no change in plasma membrane integrity in the observed treatment time, which can be attributed to metabolic differences and the resilience of the amastigote. Also, ultrastructural changes such as vacuolization suggesting autophagy were observed. The in vivo effectiveness of MHZ15 in the experimental model of cutaneous leishmaniasis was carried out in mice of the BALB/c strain infected with L. amazonensis. The treatment by intralesional route showed that MHZ15 acted with great efficiency with significantly reduction in the parasite load in the injured paws and draining lymph nodes, without clinical signs of distress or compromise of animal welfare. In vivo toxicity was also evaluated and null alterations in the levels of hepatic enzymes aspartate aminotransferase, and alanine aminotransferase was observed. The data presented herein demonstrates that MHZ15 exhibits a range of favorable characteristics conducive to the development of an antileishmanial agent., (© 2024 John Wiley & Sons Ltd.)

Leishmania poses a substantial threat to the human population all over the globe because of its visceral and cutaneous spread engendered by all 20 species. Unfortunately, the available drugs against leishmania are already hobbled with toxicity, prolonged treatment, and increasing instances of acquirement of resistance. Under these grave circumstances, the development of new drugs has become imperative to keep these harmful microbes at bay. To this end, a Groebke-Blackburn-Bienaymé multicomponent reaction-based library of different imidazo-fused heterocycles has been synthesized and screened against Leishmania amazonensis promastigotes and amastigotes. Among the library compounds, the imidazo-pyrimidine 24 has been found to be the most effective (inhibitory concentration of 50% (IC 50 ) < 10 μM), with selective antileishmanial activity on amastigote forms, a stage of the parasite related to human disease. The compound 24 has exhibited an IC 50 value of 6.63 μM, being ∼two times more active than miltefosine, a reference drug. Furthermore, this compound is >10 times more destructive to the intracellular parasites than host cells. The observed in vitro antileishmanial activity along with suitable in silico physicochemical and absorption, distribution, metabolism, excretion, and toxicity (ADMET) properties of compound 24 reinforce the imidazo-pyrimidine scaffold as a new antileishmanial pharmacophore and encourage further murine experimental leishmaniasis studies., Competing Interests: The authors declare no competing financial interest., (© 2023 The Authors. Published by American Chemical Society.)

Objectives: To evaluate the ability of the aqueous extract of Mitracarpus frigidus (MFAq) to inhibit lipid body formation and inflammatory mediator production in macrophages stimulated with lipopolysaccharide (LPS) and interferon gamma (IFN-γ)., Methods: MFAq was chemically characterized by ultrafast liquid chromatography/quadruple time-of-flight tandem mass spectrometry. The macrophages obtained from mice were incubated with MFAq. Cell viability and membrane integrity were evaluated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and propidium iodide assays, respectively. Moreover, NO, reactive oxygen species (ROS), transforming growth factor beta (TGF-β), prostaglandin E2 (PGE2) levels and lipid bodies (LBs) were examined in macrophages that were stimulated with LPS and IFN-γ and treated with MFAq. Finally, molecular docking analysis was conducted to investigate the interaction of MFAq with the cyclooxygenase 2 (COX-2) enzyme., Key Findings: Chlorogenic acid, clarinoside, harounoside, rutin, kaempferol-3O-rutinoside and 2-azaanthraquinone were identified in MFAq. MFAq significantly inhibited NO, ROS and LBs, and did not affect the membrane integrity of macrophages. MFAq-treated cells showed significantly lower levels of TGF-β and PGE2. Molecular docking demonstrated that the compounds found in MFAq are able to inhibit COX-2 by binding to important residues in the catalytic site., Conclusions: MFAq interferes with lipid metabolism in stimulated macrophages, leading to the reduction of important inflammatory mediators. Furthermore, MFAq can directly inhibit the COX-2 enzyme or inhibit its expression owing to its ability to reduce NO production., (© The Author(s) 2023. Published by Oxford University Press on behalf of the Royal Pharmaceutical Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Leishmaniasis is a group of infectious diseases caused by protozoan parasites that belong to the genus Leishmania . Currently, there is no human vaccine, and the available treatments are associated with toxicity, high cost, and the emergence of resistant strains. These factors highlight the need to identify new antileishmanial candidates. In this study, we synthesized twenty-four methoxylated cinnamides containing 1,2,3-triazole fragments and evaluated their antileishmanial activity against the Leishmania braziliensis species, which is the main etiological agent responsible for American Tegumentary Leishmaniasis (ATL). The cinnamides were synthetically prepared using nucleophilic acyl substitution and copper(I)-catalyzed azide-alkyne cycloaddition (CuAAC) reactions. The compounds were characterized using infrared, nuclear magnetic resonance, and high-resolution mass spectrometry techniques. We performed preliminary studies to evaluate the biological activity of these compounds against L. braziliensis promastigotes and axenic amastigotes. Compound 28 , N -((1-(7-(diethylamino)-2-oxo-2 H -chromen-3-yl)-1 H -1,2,3-triazole-4-yl) methyl)-3,4-dimethoxy cinnamide, demonstrated relevant antileishmanial activity with low toxicity in murine cells. The selectivity index values for this compound were superior compared with data obtained using amphotericin B. Furthermore, this cinnamide derivative reduced the infection percentage and number of recovered amastigotes in L. braziliensis -infected macrophages. It also induced an increase in reactive oxygen species production, depolarization of the mitochondrial potential, and disruption of the parasite membrane. Taken together, these findings suggest that this synthetic compound holds potential as an antileishmanial candidate and should be considered for future studies in the treatment of ATL.

The treatment against leishmaniasis presents problems, mainly due to their toxicity of the drugs, high cost and/or by the emergence of parasite resistant strains. In this context, new therapeutics should be searched. In this study, two novel synthetic derivatives from vanillin: [4-(2-hydroxy-3-(4-octyl-1H-1,2,3-triazol-1-yl)propoxy)-3-methoxybenzaldehyde] or 3s and [4-(3-(4-decyl-1H-1,2,3-triazol-1-yl)-2-hydroxypropoxy)-3-methoxybenzaldehyde] or 3t, were evaluated regarding their antileishmanial activity against distinct parasite species able to cause cutaneous and visceral leishmaniasis. Results showed that compounds 3s and 3t were effective against Leishmania infantum, L. amazonensis and L. braziliensis promastigote and amastigote-like forms, showing selectivity index (SI) of 25.1, 18.2 and 22.9, respectively, when 3s was used against promastigotes, and of 45.2, 7.5 and 15.0, respectively, against amastigote-like stage. Using the compound 3t, SI values were 45.2, 53.0 and 80.0, respectively, against promastigotes, and of 35.9, 46.0 and 58.4, respectively, against amastigote-like forms. Amphotericin B (AmpB) showed SI values of 5.0, 7.5 and 15.0, respectively, against promastigotes, and of 3.8, 5.0 and 7.5, respectively, against amastigote-like stage. The treatment of infected macrophages and inhibition of the infection upon pre-incubation with the molecules showed that they were effective in reducing the infection degree and inhibiting the infection in pre-incubated parasites, respectively, as compared to data obtained using AmpB. The mechanism of action of 3s and 3t was evaluated in L. infantum, revealing that both 3s and 3t altered the parasite mitochondrial membrane potential leading to reactive oxygen species production, increase in lipid corps and changes in the cell cycle, causing the parasite' death. A preliminary assay using the cell culture supernatant from treated and infected macrophages showed that 3s and 3t induced higher IL-12 and lower IL-10 values; suggesting the development of an in vitro Th1-type response in the treated cells. In this context, data indicated that 3s and 3t could be considered therapeutic agents to be tested in future studies against leishmaniasis., Competing Interests: Declaration of competing interest The authors declare no commercial or financial conflict of interest., (Copyright © 2023 Elsevier Inc. All rights reserved.)

Visceral leishmaniasis (VL) is a progressive, debilitating, and potentially fatal disease if left untreated. As a neglected tropical disease (NTD), the available treatment is restricted to a few drugs, which typically must be administered over a long period but are associated with serious adverse effects and have variability in efficacy. In this sense, drug repositioning has been considered an excellent strategy in the search for alternative treatments, especially in reducing the time and cost of the research. In this work, the repositioning potential of amodiaquine (AQ), a well-known antimalarial drug, was investigated for the treatment of VL. AQ showed significant and selective activity against promastigotes (IC 50  = 11.6 μg/mL) and intracellular amastigotes (IC 50  = 2.4 μg/mL) of L. infantum, being 10 times more destructive to the intracellular parasites than the host cell. In addition, pre-treatment of macrophages with AQ caused a significant reduction in the infection index, indicating a prophylactic effect of this drug. SEM images showed that AQ induces strong shape alterations of the promastigotes with an increase in cell volume with rounding and ribbing (vertical ridges), as well as a shortened flagellum. In addition, AQ induced depolarization of the ΔΨm, an increase in ROS and neutral lipids levels, and changes in the cell cycle in promastigotes, without alterations to the permeability of the parasite plasma membrane. L. infantum-infected macrophages treated with AQ induced the activation of oxidative mechanisms by infected host cells, with an increase in ROS and NO levels. Finally, in vitro interactions between AQ and miltefosine were found to have an additive effect in both biological stages of the parasite, with the ∑FIC 50 values ranging from 0.74 to 1.16 μg/mL and 0.54-1.11 μg/mL for promastigotes and intracellular amastigotes, respectively. Overall, these data highlight the utility of drug repurposing and indicate future preclinical testing for AQ itself or in combination as a potential VL treatment., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2022 Elsevier B.V. All rights reserved.)

In recent years, natural products with biological activities have been increasingly researched. The elucidation of phytoconstituents is necessary for the development of drugs as a natural alternative for the treatment of various diseases. The work aimed to evaluate in vitro and in silico bioactivities of hexane (CCHE) and methanol (CCME) fractions of ethanolic extract from Centrosema coriaceum Benth (Fabaceae) leaves and elucidate their phytoconstituents. CCHE and CCME showed antifungal activity for Candida glabrata (MIC of 1000 μg/mL) with fungistatic effect and action in cell envelope by sorbitol and ergosterol assays. CCHE and CCME presented promising antioxidant activity against the DPPH radical with IC 50 of 13.61±0.50 and 6.31±0.40 μg/mL, respectively, and relative antioxidant activity (RAA%) of 45.77±3.61/ 28.53±2.25 % for CCHE and 82.18±2.25/51.99±3.23 % for CCME when compared to rutin and quercetin, respectively. Moreover, these fractions demonstrated promising results for the inhibition of lipid peroxidation by β-carotene/linoleic acid assay. For anti-inflammatory and cytotoxicity activities, CCHE and CCME significantly inhibited the production of nitric oxide and TNF-α, without toxicity on murine intraperitoneal macrophages, respectively. Esters, alkanes, steroids, tocopherols, and terpenes were identified in CCHE by GC/MS. Flavonoids, phenolic acids, and disaccharides were detected in CCME by UFLC-QTOF-MS and FACE. Furthermore, rutin was purified from CCME. In silico predictions evidenced that compounds present in both fractions have high affinity to the fungal membrane besides antioxidant and anti-inflammatory activities. Based on these observations, CCHE and CCME have a noteworthy potential for the design of novel antifungal and anti-inflammatory agents that should be explored in future studies., (© 2022 Wiley-VHCA AG, Zurich, Switzerland.)

Malaria is an infectious disease widespread in underdeveloped tropical regions. The most severe form of infection is caused by Plasmodium falciparum, which can lead to development of cerebral malaria (CM) and is responsible for deaths and significant neurocognitive sequelae throughout life. In this context and considering the emergence and spread of drug-resistant P. falciparum isolates, the search for new antimalarial candidates becomes urgent. β-carbolines alkaloids are good candidates since a wide range of biological activity for these compounds has been reported. Herein, we designed 20 chemical entities and performed an in silico virtual screening against a pool of P. falciparum molecular targets, the Brazilian Malaria Molecular Targets (BRAMMT). Seven structures showed potential to interact with PfFNR, PfPK7, PfGrx1, and PfATP6, being synthesized and evaluated for in vitro antiplasmodial activity. Among them, compounds 3−6 and 10 inhibited the growth of the W2 strain at µM concentrations, with low cytotoxicity against the human cell line. In silico physicochemical and pharmacokinetic properties were found to be favorable for oral administration. The compound 10 provided the best results against CM, with important values of parasite growth inhibition on the 5th day post-infection for both curative (67.9%) and suppressive (82%) assays. Furthermore, this compound was able to elongate mice survival and protect them against the development of the experimental model of CM (>65%). Compound 10 also induced reduction of the NO level, possibly by interaction with iNOS. Therefore, this alkaloid showed promising activity for the treatment of malaria and was able to prevent the development of experimental cerebral malaria (ECM), probably by reducing NO synthesis.

Studies have previously demonstrated the importance of serine proteases in Leishmania . A well-known serine protease inhibitor, TPCK, was used in the present study to evaluate its in vitro and in vivo antileishmanial effects and determine its mechanism of action. Despite slight toxicity against mammalian cells (CC 50 = 138.8 µM), TPCK was selective for the parasite due to significant activity against L. amazonensis and L. infantum promastigote forms (IC 50 = 14.6 and 31.7 µM for L. amazonensis PH8 and Josefa strains, respectively, and 11.3 µM for L. infantum ) and intracellular amastigotes (IC 50 values = 14.2 and 16.6 µM for PH8 and Josefa strains, respectively, and 21.7 µM for L. infantum ). Leishmania parasites treated with TPCK presented mitochondrial alterations, oxidative stress, modifications in lipid content, flagellar alterations, and cytoplasmic vacuoles, all of which are factors that could be considered as contributing to the death of the parasites. Furthermore, BALB/c mice infected with L. amazonensis and treated with TPCK had a reduction in lesion size and parasite loads in the footpad and spleen. In BALB/c mice infected with L. infantum , TPCK also caused a reduction in the parasite loads in the liver and spleen. Therefore, we highlight the antileishmanial effect of the assessed serine protease inhibitor, proposing a potential therapeutic target in Leishmania as well as a possible new alternative treatment for leishmaniasis.

Objectives: This study aimed to evaluate the potential of aqueous extract from Mitracarpus frigidus aerial parts (MFAq) in the treatment of inflammation and oxidative stress, as well as to characterize its chemical constituents., Methods: Total phenolic and flavonoid contents were determined, and phytoconstituents were detected by ultra-fast liquid chromatography/quadrupole-time-of-flight tandem mass spectrometry (UFLC-QTOF-MS). The antioxidant activity was evaluated by DPPH, TAC and β-carotene/linoleic acid assays. In-vitro anti-inflammatory activity, cell viability and cell cycle were performed in J774A.1 cell line. In-vivo anti-inflammatory activity was investigated by two ear oedema assays (croton oil and phenol)., Key Findings: Chlorogenic acid, clarinoside, quercetin-hexosylpentoside, rutin, kaempferol-3-O-rutinoside, kaempferol-rhamnosylhexoside, quercetin-pentosylrhamnosylhexoside, harounoside, 2-azaanthraquinone and sucrose were identified by UFLC-QTOF-MS. MFAq showed antioxidant activity, which was positively correlated to the content of phenolic compounds. MFAq significantly inhibited the production of nitric oxide, did not decrease viability in MTT assay (all concentrations) and showed no changes in membrane permeability and cell cycle of J774A.1 cell line. Furthermore, MFAq showed a reduction in ear oedema in all tested doses., Conclusion: MFAq was effective in some antioxidant and inflammatory parameters, in the experimental conditions that were used in the study. This is the first report of chemical composition and bioactivities from this extract., (© The Author(s) 2022. Published by Oxford University Press on behalf of the Royal Pharmaceutical Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Visceral leishmaniasis (VL) is the most severe clinical form of leishmaniasis, being fatal if untreated. In search of a more effective treatment for VL, one of the main strategies is the development and screening of new antileishmanial compounds. Here, we reported the synthesis of seven new acetyl functionalized 1,2,3-triazolium salts, together with four 1,2,3-triazole precursors, and investigated their effect against different strains of L. infantum from dogs and humans. The 1,2,3-triazolium salts exhibited better activity than the 1,2,3-triazole derivatives with IC 50 range from 0.12 to 8.66 μM and, among them, compound 5 showed significant activity against promastigotes (IC 50 from 4.55 to 5.28 μM) and intracellular amastigotes (IC 50 from 5.36 to 7.92 μM), with the best selective index (SI ~ 6-9) and reduced toxicity. Our findings, using biochemical and ultrastructural approaches, demonstrated that compound 5 targets the mitochondrion of L. infantum promastigotes, leading to the formation of reactive oxygen species (ROS), increase of the mitochondrial membrane potential, and mitochondrial alteration. Moreover, quantitative transmission electron microscopy (TEM) revealed that compound 5 induces the reduction of promastigote size and cytoplasmic vacuolization. Interestingly, the effect of compound 5 was not associated with apoptosis or necrosis of the parasites but, instead, seems to be mediated through a pathway involving autophagy, with a clear detection of autophagic vacuoles in the cytoplasm by using both a fluorescent marker and TEM. As for the in vivo studies, compound 5 showed activity in a mouse model of VL at 20 mg/kg, reducing the parasite load in both spleen and liver (59.80% and 26.88%, respectively). Finally, this compound did not induce hepatoxicity or nephrotoxicity and was able to normalize the altered biochemical parameters in the infected mice. Thus, our findings support the use of 1,2,3-triazolium salts as potential agents against visceral leishmaniasis., (© 2022. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Chloroquine (CQ) was the most effective and widely used drug for the prophylaxis and treatment of severe and non-severe malaria. Although its prophylactic use has led to resistance to P. falciparum in all endemic countries, CQ still remains the drug of choice for the treatment of vivax malaria. Otherwise, the speed in which parasite resistance to available antimalarials rises and spreads in endemic regions points to the urgent need for the development of new antimalarials. Quinoline derivatives have been used as a tool in the search for new drugs and were investigated in the present study in an attempt to produce a HIT compound to avoid the cerebral malarial (CM). Seven compounds were synthesized, including three quinoline derivate salts. The cytotoxicity and antiplasmodial activity were assayed in vitro, highlighting compound 3 as a HIT, which also showed interaction with ferriprotoporphyrin IX similarly to CQ. Physicochemical and pharmacokinetic properties of absorption were found to be favorable when analyzed in silico. The in vivo assays, using the experimental cerebral malaria (ECM) model, showed important values of parasite growth inhibition on the 7th day-post infection (Q15 15 mg/kg: 76.9%, Q30 30 mg/kg: 90,1% and Q50 50 mg/kg: 92,9%). Compound 3 also showed significant protection against the development of CM, besides hepatic and renal parameters better than CQ. In conclusion, this quinoline derivative demonstrated promising activity for the treatment of malaria and was able to avoid the development of severe malaria in mice., (Copyright © 2022 Elsevier B.V. All rights reserved.)

Quinoline and 1,2,3-triazoles are well-known nitrogen-based heterocycles presenting diverse pharmacological properties, although their antileishmanial activity is still poorly exploited. As an effort to contribute with studies involving these interesting chemical groups, in the present study, a series of compounds derived from 4-aminoquinoline and 1,2,3-triazole were synthetized and biological studies using L. amazonensis species were performed. The results pointed that the derivative 4, a hybrid of 4-aminoquinoline/1,2,3-triazole exhibited the best antileishmanial action, with inhibitory concentration (IC 50 ) values of ~1 µM against intramacrophage amastigotes of L. amazonensis , and being 16-fold more active to parasites than to the host cell. The mechanism of action of derivative 4 suggest a multi-target action on Leishmania parasites, since the treatment of L. amazonensis promastigotes caused mitochondrial membrane depolarization, accumulation of ROS products, plasma membrane permeabilization, increase in neutral lipids, exposure of phosphatidylserine to the cell surface, changes in the cell cycle and DNA fragmentation. The results suggest that the antileishmanial effect of this compound is primarily altering critical biochemical processes for the correct functioning of organelles and macromolecules of parasites, with consequent cell death by processes related to apoptosis-like and necrosis. No up-regulation of reactive oxygen and nitrogen intermediates was promoted by derivative 4 on L. amazonensis -infected macrophages, suggesting a mechanism of action independent from the activation of the host cell. In conclusion, data suggest that derivative 4 presents selective antileishmanial effect, which is associated with multi-target action, and can be considered for future studies for the treatment against disease., (Copyright © 2021 The Authors. Published by Elsevier Masson SAS.. All rights reserved.)

Objectives: Evaluation of the anti-Leishmanial activity of imidazoquinoline-based TLR7/8 agonists., Methods: TLR7/8-active imidazoquinolines (2 and 3) were synthesized and assessed for activity against Leishmania amazonensis-intracellular amastigotes using mouse peritoneal macrophages. The production of reactive oxygen species (ROS), nitric oxide (NO) and cytokines was determined in infected and non-infected macrophages., Key Findings: The imidazoquinolines, 2 and 3, were primarily agonists of TLR7 with compound 3 also showing modest TLR8 activity. Docking studies showed them to occupy the same binding pocket on TLR7 and 8 as the known agonists, imiquimod and resiquimod. Compounds 2 and 3 inhibited the growth of L. amazonensis-intracellular amastigotes with the most potent compound (3, IC50 = 5.93 µM) having an IC50 value close to miltefosine (IC50 = 4.05 µM), a known anti-Leishmanial drug. Compound 3 induced macrophages to produce ROS, NO and inflammatory cytokines that likely explain the anti-Leishmanial effects., Conclusions: This study shows that activating TLR7 using compounds 2 or 3 induces anti-Leishmanial activity associated with induction of free radicals and inflammatory cytokines able to kill the parasites. While 2 and 3 had a very narrow cytotoxicity window for macrophages, this identifies the possibility to further develop this chemical scaffold to less cytotoxic TLR7/8 agonist for potential use as anti-Leishmanial drug., (© The Author(s) 2021. Published by Oxford University Press on behalf of the Royal Pharmaceutical Society. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.)

Background: Despite high seroprevalence of asymptomatic infection in humans, toxoplasmosis can manifest as a severe systemic disease, as occurs in the congenital infection. Here we evaluate the seroprevalence of Toxoplasma infection among pregnant women in a highly urbanized area of Brazil., Methods: A robust seroepidemiological study was conducted using laboratory databases of anti-Toxoplasma gondii serological results together with information on age, month/year of diagnosis and place of residence of pregnant women in the public health system of the city of Juiz de Fora, Brazil., Results: Of 5895 pregnant women analysed, 54.7% showed seronegativity and 44.4% showed seropositivity for immunoglobulin G (IgG) antibodies against Toxoplasma gondii. This seropositivity rate increased to 68.3% when only considering participants from rural areas. Multivariate analysis revealed higher odds of being seropositive associated with age (odds ratio [OR] 1.06 [confidence interval {CI} 1.05 to 1.07]) and with living in rural areas (OR 2.96 [CI 1.64 to 5.36]). The spatial distribution of IgG seropositivity indicated a higher prevalence concentrated in rural and peripheral neighbourhoods., Conclusions: This is the first report to use spatial analysis to show a cluster of Toxoplasma infection in rural and peripheral neighbourhoods of a highly urbanized municipality, which highlights the need for adequate healthcare actions to be implemented for women living in these areas., (© The Author(s) 2020. Published by Oxford University Press on behalf of Royal Society of Tropical Medicine and Hygiene.)

Treatment against visceral leishmaniasis (VL) is mainly hampered by drug toxicity, long treatment regimens and/or high costs. Thus, the identification of novel and low-cost antileishmanial agents is urgent. Acarbose (ACA) is a specific inhibitor of glucosidase-like proteins, which has been used for treating diabetes. In the present study, we show that this molecule also presents in vitro and in vivo specific antileishmanial activity against Leishmania infantum. Results showed an in vitro direct action against L. infantum promastigotes and amastigotes, and low toxicity to mammalian cells. In addition, in vivo experiments performed using free ACA or incorporated in a Pluronic ® F127-based polymeric micelle system called ACA/Mic proved effective for the treatment of L. infantum-infected BALB/c mice. Treated animals presented significant reductions in the parasite load in their spleens, livers, bone marrows and draining lymph nodes when compared to the controls, as well as the development of antileishmanial Th1-type humoral and cellular responses based on high levels of IFN-γ, IL-12, TNF-α, GM-CSF, nitrite and IgG2a isotype antibodies. In addition, ACA or ACA-treated animals suffered from low organ toxicity. Treatment with ACA/Mic outperformed treatments using either Miltefosine or free ACA based on parasitological and immunological evaluations performed one and 15 days post-therapy. In conclusion, data suggest that the ACA/Mic is a potential therapeutic agent against L. infantum and merits further consideration for VL treatment.

Schistosomiasis, caused by a blood fluke of the genus Schistosoma, afflicts over 230 million people worldwide. Treatment of the disease relies on just one drug, praziquantel. Cnicin (Cn) is the sesquiterpene lactone found in blessed thistle (Centaurea benedicta) that showed antiparasitic activities but has not been evaluated against Schistosoma. However, cnicin has poor water solubility, which may limit its antiparasitic activities. To overcome these restrictions, inclusion complexes with cyclodextrins may be used. In this work, we evaluated the in vitro and in vivo antischistosomal activities of cnicin and its complexes with β-cyclodextrin (βCD) and 2-hydroxypropyl-β-cyclodextrin (HPβCD) against Schistosoma mansoni. Cnicin were isolated from C. benedicta by chromatographic fractionation. Complexes formed by cnicin and βCD (Cn/βCD), as well as by cnicin and HPβCD (Cn/HPβCD), were prepared by coprecipitation and characterized. In vitro schistosomicidal assays were used to evaluate the effects of cnicin and its complexes on adult schistosomes, while the in vivo antischistosomal assays were evaluated by oral and intraperitoneal routes. Results showed that cnicin caused mortality and tegumental alterations in adult schistosomes in vitro, also showing in vivo efficacy after intraperitoneal administration. The oral treatment with cnicin or Cn/βCD showed no significant worm reductions in a mouse model of schistosomiasis. In contrast, Cn/HPβCD complex, when orally or intraperitoneally administered to S. mansoni-infected mice, decreased the total worm load, and markedly reduced the number of eggs, showing high in vivo antischistosomal effectiveness. Permeability studies, using Nile red, indicated that HPβCD complex may reach the tegument of adult schistosomes in vivo. These results demonstrated the antischistosomal potential of cnicin in preparations with HPβCD.

Treatment for visceral leishmaniasis (VL) is hindered mainly by the toxicity and/or high cost of therapeutic drugs. In addition, parasite resistance has been registered. Thus, there is an urgent need for the identification of novel, effective and low-cost antileishmanial agents. Since drug discovery is a long and expensive process, drug repositioning for treatment of leishmaniasis should be considered. In the present study, Ivermectin (IVE), a broad-spectrum drug used for treatment of parasitic diseases, was evaluated in vitro and in vivo against Leishmania infantum species. Results in vitro showed that IVE presented 50% Leishmania and macrophage inhibitory concentrations (IC 50 and CC 50 , respectively) of 3.64 ± 0.48 μM and 427.50 ± 17.60 μM, respectively, with a selectivity index (SI) of 117.45; whereas Amphotericin B (AmpB), which was used as control, showed IC 50 and CC 50 values of 0.12 ± 0.05 μM and 1.06 ± 0.23 μM, respectively, with a corresponding SI of 8.90. Treatment with IVE effectively reduced the infection percentage and parasite burden in infected and treated macrophages and displayed a prophylactic activity by inhibiting macrophage infection with pre-treated parasites. Furthermore, preliminary studies suggested that IVE targets the parasite's mitochondria. Activity of IVE in its free format or incorporated into Pluronic® F127-based polymeric micelles (IVE/Mic) was also evaluated in vivo as a treating drug for L. infantum-infected BALB/c mice. Miltefosine was used as a control. Results showed that Miltefosine, IVE and IVE/Mic-treated animals presented significant reductions in the parasite load in their spleens, livers, bone marrows and draining lymph nodes, as well as development of an antileishmanial Th1-type immune response one and 15 days after treatment. Notably, IVE/Mic showed a better parasitological and immunological response in comparison to other alternative treatments. In conclusion, results suggest that IVE/Mic could be considered in future studies as a therapeutic alternative to treat VL., (Copyright © 2020 Elsevier Inc. All rights reserved.)

PF-429242 is an inhibitor of subtilisin, an important protease found in Leishmania . However, studies regarding the effect of PF-429242 on Leishmania are scarce. In this work we evaluated the antileishmanial effect of PF-429242 against Leishmania infantum and the mechanism involved in the death of the parasite. PF-429242 had low toxicity against mammalian cells (peritoneal macrophages) (CC 50 = 189.07 μM) and presented activity against L. infantum promastigotes (IC 50 = 2.78 μM) and intracellular amastigotes (IC 50 = 14.07 μM), indicating selectivity toward the parasite. Transmission electron microscopy (TEM), as well as staining of L. infantum promastigotes with MitoTracker ® Red, rhodamine 123 and MitoSOX, revealed that the mitochondria was a potential target of PF-429242. In addition, PF-429242 caused an accumulation of neutral lipids in promastigotes, which was demonstrated by Nile Red staining and TEM, and induced oxidative stress (H 2 DCFDA staining). Furthermore the formation of autophagic vacuoles in L. infantum promastigotes was observed by MDC staining and TEM. However, the killing induced by PF-429242 in L. infantum promastigotes appeared to be unrelated to apoptosis and/or necrosis as there was no phosphatidylserine externalization, DNA fragmentation or alterations in the permeability of the parasite plasma membrane, as assessed by annexin V-FITC, TUNEL and propidium iodide staining, respectively. The morphological and ultrastructural evaluation of the promastigotes by optical microscopy, scanning electron microscopy (SEM) and TEM, revealed the presence of parasites with flagellar defects. TEM analysis of the intracellular amastigotes indicated that mitochondrial damage and autophagy could also be involved in the death of these forms after treatment with PF-429242. In addition, PF-429242 treatment stimulated NO production from infected macrophage, but only at a high concentration (100 μM), as well as an increase of TNF levels after treatment with 10 μM of PF-429242. The compound did not stimulate ROS or IL-10 production. Together, these data highlight the antileishmanial potential of PF-429242, inducing several cellular alterations in the parasite, such as mitochondrial damage, neutral lipids accumulation, oxidative stress and autophagy which culminate in the death of L. infantum , as well as modulating host cellular responses that favor the development of an immune response against the parasite., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2021 Machado, Gomes, Midlej, Coimbra and de Matos Guedes.)

Current treatments of visceral leishmaniasis face limitations due to drug side effects and/or high cost, along with the emergence of parasite resistance. Novel and low-cost antileishmanial agents are therefore required. We report herein the antileishmanial activity of β-acetyl-digitoxin (b-AD), a cardenolide isolated from Digitalis lanata leaves, assayed in vitro and in vivo against Leishmania infantum. Results showed direct action of b-AD against parasites, as well as efficacy for the treatment of Leishmania-infected macrophages. In vivo experiments using b-AD-containing Pluronic ® F127 polymeric micelles (b-AD/Mic) to treat L. infantum-infected mice showed that this composition reduced the parasite load in distinct organs in more significant levels. It also induced the development of anti-parasite Th1-type immunity, attested by high levels of IFN-γ, IL-12, TNF-α, GM-CSF, nitrite and specific IgG2a antibodies, in addition to low IL-4 and IL-10 contents, along with higher IFN-γ-producing CD4 + and CD8 + T-cell frequency. Furthermore, low toxicity was found in the organs of the treated animals. Comparing the therapeutic effect between the treatments, b-AD/Mic was the most effective in protecting animals against infection, when compared to the other groups including miltefosine used as a drug control. Data found 15 days after treatment were similar to those obtained one day post-therapy. In conclusion, the results obtained suggest that b-AD/Mic is a promising antileishmanial agent and deserves further studies to investigate its potential to treat visceral leishmaniasis., (© C.S. Freitas et al., published by EDP Sciences, 2021.)

Schistosomiasis is a neglected disease caused by helminth flatworms of the genus Schistosoma, affecting over 240 million people in more than 70 countries. The treatment relies on a single drug, praziquantel, making urgent the discovery of new compounds. Aurones are a natural type of flavonoids that display interesting pharmacological activities, particularly as chemotherapeutic agents against parasites. In pursuit of treatment alternatives, the present work conducted an in vitro and in vivo antischistosomal investigation with aurone derivatives against Schistosoma mansoni. After preparation of aurone derivatives and their in vitro evaluation on adult schistosomes, the three most active aurones were evaluated in cytotoxicity and haemolytic assays, as well as in confocal laser-scanning microscope studies, showing tegumental damage in parasites in a concentration-dependent manner with no haemolytic or cytotoxic potential toward mammalian cells. In a mouse model of schistosomiasis, at a single oral dose of 400 mg/kg, the selected aurones showed worm burden reductions of 35% to 65.0% and egg reductions of 25% to 70.0%. The most active thiophenyl aurone derivative 18, unlike PZQ, had efficacy in mice harboring juvenile S. mansoni, also showing significant inhibition of oviposition by parasites, giving support for the antiparasitic potential of aurones as lead compounds for novel antischistosomal drugs., (Copyright © 2020. Published by Elsevier B.V.)

Treatment for visceral leishmaniasis (VL) is hampered mainly by drug toxicity, their high cost, and parasite resistance. Drug development is a long and pricey process, and therefore, drug repositioning may be an alternative worth pursuing. Cardenolides are used to treat cardiac diseases, especially those obtained from Digitalis species. In the present study, cardenolide digitoxigenin (DIGI) obtained from a methanolic extract of Digitalis lanata leaves was tested for its antileishmanial activity against Leishmania infantum species. Results showed that 50% Leishmania and murine macrophage inhibitory concentrations (IC 50 and CC 50 , respectively) were of 6.9 ± 1.5 and 295.3 ± 14.5 μg/mL, respectively. With amphotericin B (AmpB) deoxycholate, used as a control drug, values of 0.13 ± 0.02 and 0.79 ± 0.12 μg/mL, respectively, were observed. Selectivity index (SI) values were of 42.8 and 6.1 for DIGI and AmpB, respectively. Preliminary studies suggested that the mechanism of action for DIGI is to cause alterations in the mitochondrial membrane potential, to increase the levels of reactive oxygen species and induce accumulation of lipid bodies in the parasites. DIGI was incorporated into Pluronic® F127-based polymeric micelles, and the formula (DIGI/Mic) was used to treat L. infantum-infected mice. Miltefosine was used as a control drug. Results showed that animals treated with either miltefosine, DIGI, or DIGI/Mic presented significant reductions in the parasite load in their spleens, livers, bone marrows, and draining lymph nodes, as well as the development of a specific Th1-type response, when compared with the controls. Results obtained 1 day after treatment were corroborated with data corresponding to 15 days after therapy. Importantly, treatment with DIGI/Mic induced better parasitological and immunological responses when compared with miltefosine- and DIGI-treated mice. In conclusion, DIGI/Mic has the potential to be used as a therapeutic agent to protect against L. infantum infection, and it is therefore worth of consideration in future studies addressing VL treatment.

Vitamin D has been reported to activate macrophage microbicidal mechanisms by inducing the production of antimicrobial peptides and nitric oxide (NO), but conversely has been shown to contribute to a greater susceptibility to Leishmania amazonensis infection in mice. Thus, this study aimed to evaluate the role of vitamin D during intracellular infection with L. amazonensis by examining its effect on macrophage oxidative mechanisms and parasite survival in vitro. Vitamins D2 and D3 significantly inhibited promastigote and amastigote growth in vitro. Vitamin D3 was not able to induce NO and reactive oxygen species (ROS) production in uninfected macrophages or macrophages infected with L. amazonensis. In addition, vitamin D3 in combination with interferon (IFN)-γ did not enhance amastigote killing and in fact, significantly reduced NO and ROS production when compared with the effect of IFN-γ alone. In this study, we demonstrated that vitamin D directly reduces parasite growth in infected macrophages (approximately 50-60% at 50 μm) but this effect is independent of the activation of macrophage oxidative mechanisms. These findings will contribute to a better understanding of the role of vitamin D in cutaneous leishmaniasis.

Aims: Treatment for visceral leishmaniasis (VL) is hampered by the toxicity and/or high cost of drugs, as well as by emergence of parasite resistance. Therefore, there is an urgent need for new antileishmanial agents., Methods and Results: In this study, the antileishmanial activity of a diprenylated flavonoid called 5,7,3,4'-tetrahydroxy-6,8-diprenylisoflavone (CMt) was tested against Leishmania infantum and L amazonensis species. Results showed that CMt presented selectivity index (SI) of 70.0 and 165.0 against L infantum and L amazonensis promastigotes, respectively, and of 181.9 and 397.8 against respective axenic amastigotes. Amphotericin B (AmpB) showed lower SI values of 9.1 and 11.1 against L infantum and L amazonensis promastigotes, respectively, and of 12.5 and 14.3 against amastigotes, respectively. CMt was effective in the treatment of infected macrophages and caused alterations in the parasite mitochondria. L infantum-infected mice treated with miltefosine, CMt alone or incorporated in polymeric micelles (CMt/Mic) presented significant reductions in the parasite load in distinct organs, when compared to the control groups. An antileishmanial Th1-type cellular and humoral immune response were developed one and 15 days after treatment, with CMt/Mic-treated mice presenting a better protective response., Conclusion: Our data suggest that CMt/Mic could be evaluated as a chemotherapeutic agent against VL., (© 2020 John Wiley & Sons Ltd.)

Leishmaniasis is a disease impacting public health worldwide due to its high incidence, morbidity and mortality. Available treatments are costly, lengthy and toxic, not to mention the problem of parasite resistance. The development of alternative treatments is warranted and natural products demonstrate promising activity. This study investigated the activity of Connarus suberosus extracts and compounds against Leishmania species. Several C. suberosus extracts were tested against L. amazonensis promastigotes. Active and inactive extracts were analyzed by UHPLC-MS and data evaluated using a metabolomics platform, revealing an unknown neoflavonoid (connarin, 3), isolated together with the pterocarpans: hemileiocarpin (1) and leiocarpin (2). The aforementioned compounds (1-3), together with the benzoquinones: rapanone (4), embelin (5) and suberonone (6) previously isolated by our group from the same species, were tested against: (i) L. amazonensis and L. infantum promastigotes, and (ii) L. amazonensis intracellular amastigotes, with the most active compound (3) also tested against L. infantum amastigotes. Cytotoxicity against murine peritoneal macrophages was also investigated. Compounds 2 and 3 presented an IC50 33.8 μM and 11.4 μM for L. amazonensis promastigotes; and 44.3 μM and 13.3 μM for L. infantum promastigotes, respectively. For L. amazonensis amastigotes, the IC50 of 2 was 20.4 μM with a selectivity index (SI) of 5.7, while the IC50 of 3 was 2.9 μM with an SI of 6.3. For L. infantum amastigotes, the IC50 of 3 was 7.7 μM. Compounds 2 and 3 presented activity comparable with the miltefosine positive control, with compound 3 found to be 2-4 times more active than the positive control, depending on the Leishmania species and form. The extracts and isolated compounds showed moderate toxicity against macrophages. Compounds 2 and 3 altered the mitochondrial membrane potential (ΔΨm) and neutral lipid body accumulation, while 2 also impacted plasma membrane permeabilization, culminating in cellular disorder and parasite death. Transmission electron microscopy of L. amazonensis promastigotes treated with compound 3 confirmed the presence of lipid bodies. Leiocarpin (2) and connarin (3) demonstrated antileishmanial activity. This study provides knowledge of natural products with antileishmanial activity, paving the way for prototype development to fight this neglected tropical disease., Competing Interests: The authors have declared that no competing interests exist.

Introduction: The drugs currently available for leishmaniasis treatment have major limitations., Methods: In vitro and in vivo studies were performed to evaluate the effect of a quinoline derivative, Hydraqui (7-chloro-4-(3-hydroxy-benzilidenehydrazo)quinoline, against Leishmania amazonensis. In silico analyses of absorption, distribution, metabolism, excretion, and toxicity (ADMET) parameters were performed., Results: Hydraqui showed significant in vitro anti-amastigote activity. Also, Hydraqui-treated mice exhibited high efficacy in lesion size (48.3%) and parasitic load (93.8%) reduction, did not cause hepatic and renal toxicity, and showed appropriate ADMET properties., Conclusions: Hydraqui presents a set of satisfactory criteria for its application as an antileishmanial agent.

Purpose: Due to serious problems with the treatment of leishmaniasis all around the world, here is an urgent need in the search for new drugs that are more effective and safer for the treatment of the various forms of leishmaniasis. Actual therapy is limited and lacks sufficient efficacy due to incomplete elimination of the parasites form of patients. In this sense, we decided to evaluate, by first-time, a series of seventeen camphor hydrazone derivatives (2a-2p) against Leishmania amazonensis., Methods: The compounds previously synthesized from camphor, an abundant natural compound, were evaluated in vitro against the extra and intracellular forms of Leishmania amazonensis, and murine macrophages., Results: The majority of compounds, fourteen, displayed activity against the intracellular form of the parasite (amastigote) with IC 50 values ranging from 21.78 to 58.23 µM, being six compounds active for both forms of the parasite. The compound 2i exhibited higher activity against the amastigote form with the value of IC 50 (21.78 µM) close to standard utilized miltefosine (12.74 µM) and selectivity index of at least 6.9. Six compounds displayed activity against promastigote form of Leishmania amazonensis 2g, 2j-2n (41.17-69.59 µM), with the compound 2m being the more active with IC 50  = 41.17 µM, 1.9 times less active than the reference drug (IC 50  = 21.39 µM). The compound 2m was the more selective to this form, with a selectivity index of at least 3.6. All the compounds were non-cytotoxic to macrophages., Conclusions: Most compounds showed activity against amastigote form of Leishmania amazonensis, being that they were not cytotoxic to macrophage at the maximum tested concentration, showing the selective property of these compounds. Since amastigotes are the parasite stages that cause the disease in humans, these results highlight the antileishmanial effect of the compounds. This study indicates the possible development of candidates to leishmanicidal drugs from an abundant natural compound of easy access.

1,2,3-triazolium salts are poorly understood regarding their antileishmanial activity. Hence, as an effort to identify novel chemical scaffolds as antileishmanial agents, a series of 1,2,3-triazolium salts (TS) and corresponding 1,2,3-triazole (T) precursors including new epoxide derivatives were synthesized and assayed against Leishmania amazonensis promastigote and intracellular amastigote forms. Among them, the compound TS-6 exhibited promising activity on promastigotes (IC 50  = 3.61 μM) and intracellular amastigotes (IC 50  = 7.61 μM) of L. amazonensis, superior to miltefosine (IC 50  > 10.0 μM), used as reference drug. In addition, TS-6 showed negligible cytotoxicity on murine peritoneal macrophages with a SI of about 10. Studies on the mode of action of TS-6 indicate mitochondrial dysfunction through an increase in 'total' and mitochondrial-ROS as well as depolarization of mitochondrial membrane potential of L. amazonensis promastigotes. In silico physicochemical studies indicate that the TS-6 could potentially be used as an oral drug., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2019 Elsevier B.V. All rights reserved.)

In this work, we report the antileishmanial activity of 15 compounds based on 2-pyrimidinyl hydrazone and N-acylhydrazone derivatives, being 13 new compounds. All compounds were tested against promastigotes and Leishmania amazonensis-GFP amastigotes, as well as murine macrophages. Besides, studies about the mechanism of action of the best antileishmanial compounds and in silico physicochemical and pharmacokinetic properties were performed. Studies about the mechanism of action of representative compounds of each class showed slight differences in mode of action and both are able to cause mitochondrial depolarization and increase of intracellular ROS levels. Through computational tool and further analysis of the physicochemical and pharmacokinetic parameters, the results indicating good oral bioavailability. These results confirm the potential of 2-pyrimidinyl derivatives as lead compounds in antileishmanial drug discovery., (Copyright © 2019. Published by Elsevier Masson SAS.)

The alkylaminoalkanethiosulfuric acids (AAATs) are amphipathic compounds effective against experimental schistosomiasis, of low toxicity, elevated bioavailability after a single oral dose and prompt tissue absorption., Objectives: To explore the in-vitro antileishmanial potential of AAATs using five compounds of this series against Leishmania (Viannia) braziliensis., Methods: Their effects on promastigotes and axenic amastigotes, and cytotoxicity to macrophages were tested by the MTT method, and on Leishmania-infected macrophages by Giemsa stain. Effects on the mitochondrial membrane potential of promastigotes and axenic amastigotes and DNA of intracellular amastigotes were tested using JC-1 and TUNEL assays, respectively., Key Findings: The 2-(isopropylamino)-1-octanethiosulfuric acid (I) and 2-(sec-butylamino)-1-octanethiosulfuric acid (II) exhibit activity against both promastigotes and intracellular amastigotes (IC 50 25-35 µm), being more toxic to intracellular parasites than to the host cell. Compound I induced a loss of viability of axenic amastigotes, significantly reduced (30%) the mitochondrial membrane potential of both promastigotes and axenic amastigotes and promoted selective DNA fragmentation of the nucleus and kinetoplast of intracellular amastigotes., Conclusions: In this previously unpublished study of trypanosomatids, it is shown that AAATs could also exhibit selective antileishmanial activity, a new possibility to be investigated in oral treatment of leishmaniasis., (© 2019 Royal Pharmaceutical Society.)

Objectives: The in vitro antileishmanial effect of analogues of resveratrol (AR) present in the N-aryl imines and N-aryl hydrazones series was investigated. In addition, possible parasite targets were evaluated., Methods: Antipromastigote activity of Leishmania amazonensis, L. braziliensis and L. infantum, as well as the cytotoxicity on macrophages was determined by MTT assay and L. braziliensis-infected macrophages effect by Giemsa stain. After staining, effects on the parasite targets were analysed by flow cytometry or by fluorescence microscopy., Key-Findings: Among the tested compounds, the derivative AR26 showed the best effect against promastigotes of all Leishmania species (IC 50  < 3.0 µg/ml), being more active than miltefosine, the control drug. AR26 was also effective against amastigotes of L. braziliensis (IC 50  = 15.9 µg/ml), with low toxicity to mammalian cells. The evaluation of mechanism of action of AR26 on L. braziliensis promastigotes indicates mitochondrial potential depolarization, plasma membrane permeabilization, interference in the progression of the cell cycle and accumulation of autophagic vacuoles. In addition, any increase of the reactive oxygen species levels was detected in the treated L. braziliensis-macrophages., Conclusions: Data indicate that the antileishmanial activity of AR26 is related to multitarget action, and the resveratrol analogues could be used in future studies as antileishmanial agent., (© 2019 Royal Pharmaceutical Society.)

The identification of new therapeutics to treat leishmaniasis is desirable, since available drugs are toxic and present high cost and/or poor availability. Therefore, the discovery of safer, more effective and selective pharmaceutical options is of utmost importance. Efforts towards the development of new candidates based on molecule analogs with known biological functions have been an interesting and cost-effective strategy. In this context, quinoline derivatives have proven to be effective biological activities against distinct diseases. In the present study, a new chloroquinoline derivate, AM1009, was in vitro tested against two Leishmania species that cause leishmaniasis. The present study analyzed the necessary inhibitory concentration to preclude 50% of the Leishmania promastigotes and axenic amastigotes (EC 50 value), as well as the inhibitory concentrations to preclude 50% of the murine macrophages and human red blood cells (CC 50 and RBC 50 values, respectively). In addition, the treatment of infected macrophages and the inhibition of infection using pre-treated parasites were also investigated, as was the mechanism of action of the molecule in L. amazonensis. To investigate the in vivo therapeutic effect, BALB/c mice were infected with L. amazonensis and later treated with AM1009. Parasitological and immunological parameters were also evaluated. Clioquinol, a known antileishmanial quinoline derivate, and amphotericin B (AmpB), were used as molecule and drug controls, respectively. Results in both in vitro and in vivo experiments showed a better and more selective action of AM1009 to kill the in vitro parasites, as well as in treating infected mice, when compared to results obtained using clioquinol or AmpB. AM1009-treated animals presented significantly lower average lesion diameter and parasite burden in the infected tissue and organs evaluated in this study, as well as a more polarized antileishmanial Th1 immune response and low renal and hepatic toxicity. This result suggests that AM1009 should be considered a possible therapeutic target to be evaluated in future studies for treatment against leishmaniasis., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Schistosomiasis is a neglected tropical disease that affects million people worldwide, mostly in developing countries. Ruta graveolens (Rutaceae) is a plant used in folk medicine to treat several diseases, including parasitic infections. In this study, we reported the in vitro schistosomicidal activity of the R. graveolens extract ( Rg ) and its active fraction ( Rg-FAE ). Also, the characterization of Rg-FAE by UPLC-ESI-QTOF-MS analysis and its in vitro antileishmanial activity against Leishmania braziliensis were also performed. In vitro schistosomicidal assays were assessed against adult worms of S. mansoni , while cell viability against peritoneal macrophages was measured by MTT assay. Rg (100  μ g/mL) exhibited noticeable schistosomicidal activity, causing 100% mortality and decreasing motor activity of all adult male and female schistosomes, but with low activity against L. braziliensis . After chromatographic fractionation of Rg , fraction Rg-FAE was obtained, showing high activity against adult schistosomes. UPLC-ESI-QTOF-MS analysis of Rg-FAE revealed the presence of eleven alkaloids and one furanocoumarin. No significant antileishmanial activity was found for Rg , while Rg-FAE exhibited activity against L. braziliensis promastigotes. We demonstrated, for the first time, that the R. graveolens extract ( Rg ) and its alkaloid-rich fraction ( Rg-FAE ) are active against adult worms of S. mansoni , with no significant cytotoxicity on macrophages. Our findings open the route to further antiparasitic studies with the active fraction of R. graveolens and its identified compounds, especially alkaloids., Competing Interests: The authors declare that there are no conflicts of interest., (Copyright © 2019 Lara Soares Aleixo de Carvalho et al.)

In this work, we combined molecular modeling, computational docking and in vitro analysis to explore the antileishmanial effect of some resveratrol analogs (ResAn), focusing on their pro-oxidant effect. The molecular target was the trypanothione reductase of Leishmania braziliensis (LbTryR), an essential component of the antioxidant defenses in trypanosomatid parasites. Three-dimensional structures of LbTryR were modeled and molecular docking studies of ResAn1-5 compounds showed the following affinity: ResAn1 > ResAn2 > ResAn4 > ResAn5 > ResAn3. Positive correlation was observed between these compounds' affinity to the LbTryR and the IC 50 values against Leishmania sp (ResAn1 < ResAn2 < ResAn4), which allows for TryR being considered an important target for them. As the compound ResAn1 showed the best antileishmanial activity, and docking studies showed its high affinity for NADP binding site (NS) of TryR, plus having been able to induce ROS production in L. braziliensis promastigotes treated, ResAn1 probably occupies NS interfering in the electron transfer processes responsible for the catalytic reaction. The in silico prediction of ADMET properties suggests that ResAn1 may be a promising drug candidate with properties to cross biological membranes and high gastrointestinal absorption, not violating Lipinski's rules. Ultimately, the antileishmanial effect of ResAn can be associated with a pro-oxidant effect which, in turn, can be exploited as an antimicrobial agent. Communicated by Ramaswamy H. Sarma.

A nucleoside triphosphate diphosphohydrolase-1 (NTPDase 1) was identified on the surface, flagellum and kinetoplast from L. infantum promastigotes by immunocytochemistry and confocal laser scanning microscopy, using immune sera that recognized specifically the B domain of NTPDase 1 and produced against synthetic peptides (LbB1LJ and LbB2LJ) derived from this domain. The polyclonal antibodies had effective antileishmanial effect, reducing significantly in vitro promastigotes growth (21-25%), an antiproliferative effect also demonstrated by immune sera produced against recombinant r-pot B domain, and two other synthetic peptides (potB1LJ and potB2LJ). In addition, using these biomolecules in ELISA technique, IgG1 and IgG2 subclasses reactivities of either healthy dogs or infected by L. infantum and classified clinically as asymptomatic, oligosymptomatic and symptomatic were tested. Analysis of distinct IgG1 and IgG2 seropositivities patterns suggested antibody subclasses binding epitopes along B domain for protection against infection, indicating this domain as a new tool for prophylactic and immunotherapeutic investigations., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Nucleoside triphosphate diphosphohydrolase (NTPDase) 1 from intracellular amastigotes of Leishmania infantum-infected macrophage was identified by immunocytochemistry and confocal laser scanning microscopy using antibodies that specifically recognize its B-domain. This enzyme was previously characterized in Leishmania promastigote form, and here it is shown to be susceptible to pentamidine isethionate (PEN). In initial assays, this antileishmanial compound (100 μM) reduced 60% phosphohydrolytic activity of promastigotes preparation. An active NTPDase 1 was then isolated by non-denaturing gel electrophoresis, and PEN (10 μM) inhibited 74% and 35% of the ATPase and ADPase activities, respectively, of this pure protein. In addition, PEN 0.1-1 μM inhibited 56% potato apyrase activity, a plant protein that shares high identity with Leishmania NTPDase 1. In contrast, amphotericin B, fluconazole, ketoconazole or allopurinol did not significantly affect phosphohydrolytic activity of either promastigotes preparation or potato apyrase. This work suggests amastigote NTPDase 1 as a new molecular target, and inhibition of its catalytic activity by pentamidine can be part of the mode of action of this drug contributing with the knowledge of its antileishmanial effect., (Copyright © 2019 Elsevier Inc. All rights reserved.)

The treatment against leishmaniasis presents problems, since the currently used drugs are toxic and/or have high costs. In addition, parasite resistance has increased. As a consequence, in this study, a chloroquinolin derivative, namely 7-chloro-N,N-dimethylquinolin-4-amine or GF1059, was in vitro and in vivo tested against Leishmania parasites. Experiments were performed to evaluate in vitro antileishmanial activity and cytotoxicity, as well as the treatment of infected macrophages and the inhibition of infection using pre-treated parasites. This study also investigated the GF1059 mechanism of action in L. amazonensis. Results showed that the compound was highly effective against L. infantum and L. amazonensis, presenting a selectivity index of 154.6 and 86.4, respectively, against promastigotes and of 137.6 and 74.3, respectively, against amastigotes. GF1059 was also effective in the treatment of infected macrophages and inhibited the infection of these cells when parasites were pre-incubated with it. The molecule also induced changes in the parasites' mitochondrial membrane potential and cell integrity, and caused an increase in the reactive oxygen species production in L. amazonensis. Experiments performed in BALB/c mice, which had been previously infected with L. amazonensis promastigotes, and thus treated with GF1059, showed that these animals presented significant reductions in the parasite load when the infected tissue, spleen, liver, and draining lymph node were evaluated. GF1059-treated mice presented both lower parasitism and low levels of enzymatic markers, as compared to those receiving amphotericin B, which was used as control. In conclusion, data suggested that GF1059 can be considered a possible therapeutic target to be tested against leishmaniasis., (Copyright © 2019 Elsevier Inc. All rights reserved.)

New therapeutics against leishmaniasis are desirable, since the current drugs applied against this disease complex presents problems, such as the toxicity, high cost and/or parasite resistance. In the present study, a new fluoroquinoline derivate, namely 7-chloro-N-(4-fluorophenethyl)quinolin-4-amine or GF1061, was evaluated regarding to its in vitro antileishmanial action against Leishmania infantum and L. amazonensis species, as well as by its toxicity in mammalian cells and efficacy in the treatment of infected macrophages. The mechanism of action of this molecule in L. amazonensis and the therapeutic efficacy in infected BALB/c mice were also evaluated. Results showed that GF1061 was effective against both parasite species, showing selectivity index (SI) of 38.7 and 42.7 against L. infantum and L. amazonensis promastigotes, respectively, and of 45.0 and 48.9 against the amastigotes, respectively. Amphotericin B (AmpB), used as control, showed SI values of 6.6 and 8.8 against L. infantum and L. amazonensis promastigotes, respectively, and of 2.2 and 2.7 against the amastigotes, respectively. The molecule was effective in treat infected macrophages, as well as it induced alterations in the mitochondrial membrane potential, increase in the reactive oxygen species production, and in the cell integrity of the parasites. Regarding to the in vivo experiments, BALB/c mice (n = 8 per group) were subcutaneously infected with 10 6 L. amazonensis stationary promastigotes and, 60 days post-infection, they received saline or were treated during 10 days, once a day, with AmpB (1 mg/kg body weight) or GF1061 (5 mg/kg body weight). One day after the treatment, the infected tissue, spleen, liver, and draining lymph node (dLN) of the animals were collected, and the parasite load was evaluated. GF1061-treated mice, as compared to the saline and AmpB groups, showed significant reductions in the parasitism in the infected tissue (66% and 62%, respectively), liver (69% and 44%, respectively), spleen (71% and 38%, respectively), and dLN (72% and 48%, respectively). In conclusion, results suggested that GF1061 may be considered as a possible therapeutic target to be evaluated against leishmaniasis in other mammalian hosts., (Copyright © 2018 Elsevier B.V. All rights reserved.)

Clioquinol (5-chloro-7-iodoquinolin-8-ol or ICHQ) was recently showed to presents an in vitro effective antileishmanial action, causing changes in membrane permeability, mitochondrial functionality, and parasite morphology. In the present study, ICHQ was incorporated into a Poloxamer 407-based polymeric micelles system (ICHQ/M), and its antileishmanial activity was in vivo evaluated in L. amazonensis-infected BALB/c mice. Amphotericin B (AmpB) and its liposomal formulation (Ambisome®) were used as controls. Parasitological and immunological evaluations were performed 30 days after the treatment. Results indicated more significant reductions in the average lesion diameter and parasite burden in ICHQ or ICHQ/M-treated mice, which were associated with the development of a polarized Th1 immune response, based on production of high levels of IFN-γ, IL-12, TNF-α, GM-CSF, and antileishmanial IgG2a antibody. Control groups´ mice produced high levels of IL-4, IL-10, and IgG1 isotype antibody. No organic toxicity was found by using ICHQ or ICHQ/M to treat the animals, although those receiving AmpB and Ambisome® have presented higher levels of renal and hepatic damage markers. In conclusion, results suggested that the ICHQ/M composition can be considered as an antileishmanial candidate to be tested against human leishmaniasis., (Copyright © 2018 Elsevier B.V. All rights reserved.)

New therapeutic strategies against leishmaniasis are desirable, since the treatment against disease presents problems, such as the toxicity, high cost and/or parasite resistance. As consequence, new antileishmanial compounds are necessary to be identified, as presenting high activity against Leishmania parasites, but low toxicity in mammalian hosts. Flau-A is a naphthoquinone derivative recently showed to presents an in vitro effective action against Leishmania amazonensis and L. infantum species. In the present work, the in vivo efficacy of Flau-A, which was incorporated into a Poloxamer 407-based micelle system, was evaluated in a murine model against L. amazonensis infection. Amphotericin B (AmB) and Ambisome ® were used as controls. The animals were infected and later treated with the compounds. Thirty days after the treatment, parasitological and immunological parameters were evaluated. Results showed that AmB, Ambisome ® , Flau-A or Flau-A/M-treated animals presented significantly lower average lesion diameter and parasite burden in tissue and organs evaluated, when compared to the control (saline and micelle) groups. Flau-A or Flau-A/M-treated mice were those presenting the most significant reductions in the parasite burden, when compared to the others. These animals developed also a more polarized antileishmanial Th1 immune response, which was based on significantly higher levels of IFN-γ, IL-12, TNF-α, GM-CSF, and parasite-specific IgG2a isotype; associated with low levels of IL-4, IL-10, and IgG1 antibody. The absence of toxicity was found in these animals, although mice receiving AmB have showed high levels of renal and hepatic damage markers. In conclusion, results suggested that the Flau-A/M compound may be considered as a possible therapeutic target to be evaluated against human leishmaniasis., (Copyright © 2018 Elsevier Masson SAS. All rights reserved.)

Currently, available treatment options for leishmaniasis are limited and unsatisfactory. In a previous study, a quinoline derivative (AMQ-j), exhibited a strong effect against Leishmania amazonensis and its antileishmanial activity was preliminarily associated with mitochondrial dysfunction. The present study further explores the antileishmanial effect of this compound against L. amazonensis, as well as determines the main cellular processes involved in the death of the parasite. Moreover, this study evaluated the in vivo effect of the AMQ-j in BALB/c mice experimentally infected by L. amazonensis. The results showed that the compound AMQ-j induces a set of morphological and biochemical features that could correlate with both autophagy-related and apoptosis-like processes, indicating intense mitochondrial swelling, a collapse of the mitochondrial membrane potential, an abnormal chromatin condensation, an externalization of phosphatidylserine, an accumulation of lipid bodies, a disorganization of cell cycle, a formation of autophagic vacuoles, and an increase of acidic compartments. Treatment with AMQ-j through an intralesional route was effective in reducing the parasite burden and size of the lesion. No significant increase in the serum levels of hepatic or renal damage toxicity markers was observed. These findings contribute to the understanding of the mode of action of quinoline derivatives involved in the death of Leishmania parasites and encourage new studies in other experimental models of Leishmania infection., (Copyright © 2018 Elsevier Inc. All rights reserved.)

In this study, we evaluated the in vitro and in vivo schistosomicidal activities of chalcones against Schistosoma mansoni worms. In vitro assays revealed that chalcones 1 and 3 were the most active compounds, without affecting significantly mammalian cells. Confocal laser scanning microscopy and scanning electron microscopy studies revealed reduction on the numbers of tubercles and morphological alterations in the tegument of S. mansoni worms after in vitro incubation with chalcones 1 and 3. In a mouse model of schistosomiasis, the oral treatment (400 mg/kg) with chalcone 1 or 3 significantly caused a total worm burden reduction in mice. Chalcone 1 showed significant inhibition of the S. mansoni ATP diphosphohydrolase activity, which was corroborated by molecular docking studies. The results suggested that chalcones could be explored as lead compounds with antischistosomal properties., (© 2018 Wiley-VHCA AG, Zurich, Switzerland.)

The search for new drugs for the treatment of leishmaniasis is an important strategy for improving the current therapeutic arsenal for the disease. There are several limitations to the available drugs including high toxicity, low efficacy, prolonged parenteral administration, and high costs. Steroids are a diverse group of compounds with various applications in pharmacology. However, the antileishmanial activity of this class of molecules has not yet been explored. Therefore, in the present study, we investigated the antileishmanial activity and cytotoxicity of novel steroids against murine macrophages with a focus on the derivatives of cholesterol (CD), cholic acid (CA), and deoxycholic acid (DA). Furthermore, the mechanism of action of the best compound was assessed, and in silico studies to evaluate the physicochemical and pharmacokinetic properties were also conducted. Among the sixteen derivatives, schiffbase2, CD2 and deoxycholic acid derivatives (DOCADs) were effective against promastigotes of Leishmania species. Despite their low toxicity to macrophages, the majority of DOCADs were active against intracellular amastigotes of L. amazonensis, and DOCAD5 exhibited the best biological effect against these parasitic stages (IC 50  = 15.34 μM). Neither the CA derivatives (CAD) nor DA alone inhibited the intracellular parasites. Thus, the absence of hydroxyl in the C-7 position of the steroid nucleus, as well as the modification of the acid group in DOCADs were considered important for antileishmanial activity. The treatment of L. amazonensis promastigote forms with DOCAD5 induced biochemical changes such as depolarization of the mitochondrial membrane potential, increased ROS production and cell cycle arrest. No alterations in parasite plasma membrane integrity were observed. In silico physicochemical and pharmacokinetic studies suggest that DOCAD5 could be a good candidate for an oral drug. The data demonstrate the potential antileishmanial effect of certain steroid derivatives and encourage new in vivo studies., (Copyright © 2018 Elsevier Masson SAS. All rights reserved.)

Leishmaniases are infectious diseases, caused by protozoa of the Leishmania genus. These drugs present high toxicity, long-term administration, many adverse effects and are expensive, besides the identification of resistant parasites. In this work, the antileishmanial activity of quinoline derivative salts (QDS) was evaluated, as well as the toxicity on mammalian cells and the mechanism of action of the most promising compound. Among the compound tested, only the compound QDS3 showed activity against promastigotes and amastigotes of Leishmania spp., being more active against the intracellular amastigotes of L. amazonensis-GFP (IC 50 of 5.48 μM). This value is very close to the one observed for miltefosine (IC 50 of 4.05 μM), used as control drug. Furthermore, the compound QDS3 exhibited a selective effect, being 40.35 times more toxic to the amastigote form than to the host cell. Additionally, promastigotes of L. amazonensis treated with this compound exhibited characteristics of cells in the process of apoptosis such as mitochondrial membrane depolarization, mitochondrial swelling, increase of ROS production, phosphatidylserine externalization, reduced and rounded shape, and cell cycle alteration. The integrity of the plasma membrane remained unaltered, excluding necrosis in treated promastigotes. The compound QDS3 inhibited the formation of autophagic vacuoles, which may have contributed to parasite death by preventing autophagic mechanisms in the removal of damaged organelles, intensifying the damage caused by the treatment, highlighting the antileishmanial effect of this compound. In addition, treatment with QDS3 induced increased ROS levels in L. amazonensis-infected macrophages, but not in uninfected host cell. These data reinforce that the induction of oxidative stress is one of the main toxic effects caused by the treatment with the compound QDS3 in L. amazonensis, causing irreversible damage and triggering a selective death of intracellular parasites. Data shown here confirm the biological activity of quinoline derivatives and encourage future in vivo studies with this compound in the murine model., (Copyright © 2018 Elsevier B.V. All rights reserved.)

In this study, a quinoline derivate, clioquinol (5-chloro-7-iodoquinolin-8-ol), was evaluated against Leishmania amazonensis and Leishmania infantum promastigotes and amastigotes. The cytotoxicity in murine macrophages and human red blood cells, as well as the efficacy in treating infected macrophages and the inhibition of infection using pre-treated parasites were also evaluated. Results showed that clioquinol inhibited L. amazonensis and L. infantum promastigotes with effective concentration 50% (EC 50 ) values of 2.55 ± 0.25 and 1.44 ± 0.35 μg/mL, respectively, and of 1.88 ± 0.13 and 0.98 ± 0.17 μg/mL against axenic amastigotes, respectively. The cytotoxic EC 50 concentrations of clioquinol in murine macrophages and human red blood cells were, respectively, 255 ± 23 and 489 ± 20 μg/mL. With these results, the selectivity index was calculated, showing values of 99.9 and 177.1 against promastigotes, respectively, and of 135.6 and 260.1 against axenic amastigotes, respectively. Significant reductions in the percentage of infected macrophages after treatment using clioquinol were also observed, as well as when parasites were pre-treated with clioquinol and used to infect murine macrophages. The mechanism of action of clioquinol was investigated in L. amazonensis, and results revealed morphological and biochemical alterations in the clioquinol-treated parasites, including reduction in cell volume, loss of mitochondrial membrane potential, increase in the ROS production and rupture of the plasma membrane. The externalization of phosphatidylserine (PS) at the cell surface was evaluated in treated parasites that had been doubly labelled with annexin and propidium iodide (PI). The results showed no significant difference for PS exposure when compared to the untreated control, although a significant increase in the PI/annexin V-labelled cell population was found in the treated parasites. Results suggest that clioquinol induces a discontinuity of the parasite membrane, possibly related to a characteristic event of cell death caused by necrosis. This study demonstrates, for the first time, the antileishmanial activity of clioquinol against two relevant Leishmania species and suggests that the mitochondria of the parasites may be a possible biological target leading to parasite necrosis. Our findings suggest that clioquinol may have a potential application in treatment of leishmaniasis and further studies should be performed in infected mammalian hosts., (© 2018 Nordic Association for the Publication of BCPT (former Nordic Pharmacological Society).)

In this work, we report the antileishmanial activity of 23 compounds based on 2-pyrazyl and 2-pyridylhydrazone derivatives. The compounds were tested against the promastigotes of Leishmania amazonensis and L. braziliensis, murine macrophages, and intracellular L. amazonensis amastigotes. The most potent antileishmanial compound was selected for investigation into its mechanism of action. Among the evaluated compounds, five derivatives [(E)-3-((2-(pyridin-2-yl)hydrazono)methyl)benzene-1,2-diol (2 b), (E)-4-((2-(pyridin-2-yl)hydrazono)methyl)benzene-1,3-diol (2 c), (E)-4-nitro-2-((2-(pyrazin-2-yl)hydrazono)methyl)phenol (2 s), (E)-2-(2-(pyridin-2-ylmethylene)hydrazinyl)pyrazine (2 u), and (E)-2-(2-((5-nitrofuran-2-yl)methylene)hydrazinyl)pyrazine (2 v)] exhibited significant activity against L. amazonensis amastigote forms, with IC 50 values below 20 μm. The majority of the compounds did not show any toxic effect on murine macrophages. Preliminary studies on the mode of action of members of this hydrazine-derived series indicate that the accumulation of reactive oxygen species (ROS) and disruption of parasite mitochondrial function are important for the pharmacological effect on L. amazonensis promastigotes., (© 2018 Wiley-VCH Verlag GmbH & Co. KGaA, Weinheim.)