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42 p.-8 fig. Drug repurposing affords the implementation of new treatments at a moderate cost and under a faster time-scale. Most of the clinical drugs against Leishmania share this origin. The antidepressant sertraline has been successfully assayed in a murine model of visceral leishmaniasis. Nevertheless, sertraline targets in Leishmania were poorly defined. In order to get a detailed insight into the leishmanicidal mechanism of sertraline on Leishmania infantum, unbiased multiplatform metabolomics and transmission electron microscopy were combined with a focused insight into the sertraline effects on bioenergetics metabolism of the parasite. Sertraline induced respiration uncoupling, a significant decrease of intracellular ATP level, and oxidative stress in L. infantum promastigotes. Metabolomics evidenced an extended metabolic disarray caused by sertraline. This encompasses a remarkable variation of the levels of thiol-redox and polyamine biosynthetic intermediates, as well as shortage of intracellular amino acids used as metabolic fuel by Leishmania Sertraline killed Leishmania through a multitarget mechanism of action, tackling essential metabolic pathways of the parasite. As such, sertraline is a valuable candidate for visceral leishmaniasis treatment under a drug repurposing strategy. This work was supported by grants of the Fondo de Investigaciones Sanitarias-ISCIII-FEDER (PI12-02706), and (RD16/0027/0010, Red de Enfermedades Tropicales, subprogram RETICS del Plan Estatal de I+D+i (2013-2016) co-financed with FEDER funds, SAF2015-65740-R, and CSIC PIE 201620E038 to L. R., São Paulo State Research Foundation (FAPESP 2015/23403-9 to A. G.T.); EADS-CASA/ Brazilian Air Force (FAB) mobility program to M.L.L., and Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) to M.L.L.

A lack of viable hits, increasing resistance, and limited knowledge on mode of action is hindering drug discovery for many diseases. To optimize prioritization and accelerate the discovery process, a strategy to cluster compounds based on more than chemical structure is required. We show the power of metabolomics in comparing effects on metabolism of 28 different candidate treatments for Leishmaniasis (25 from the GSK Leishmania box, two analogues of Leishmania box series, and amphotericin B as a gold standard treatment), tested in the axenic amastigote form of Leishmania donovani. Capillary electrophoresis–mass spectrometry was applied to identify the metabolic profile of Leishmania donovani, and principal components analysis was used to cluster compounds on potential mode of action, offering a medium throughput screening approach in drug selection/prioritization. The comprehensive and sensitive nature of the data has also made detailed effects of each compound obtainable, providing a resource to assist in further mechanistic studies and prioritization of these compounds for the development of new antileishmanial drugs.

Amphotericin B (AmB) is a drug of choice against life-threatening systemic fungal infections and an alternative therapy for the treatment of all forms of leishmaniasis. It is known that AmB and its conventional formulation cause renal damage; however, the lipid formulations can reduce these effects. The aim of the present study was to identify metabolic changes in mice treated with two different AmB formulations, a nanoemulsion (NE) (lipid system carrier) loaded with AmB and the conventional formulation (C-AmB). For this purpose, metabolic fingerprinting represents a valuable strategy to monitor, in a non-targeted manner, the changes that are at the base of the toxicity mechanism of AmB. Plasma samples of BALB-c mice were collected after treatment with 3 alternate doses of AmB at 1 mg kg

Leishmaniases have a broad spectrum of clinical manifestations, ranging from a cutaneous to a progressive and fatal visceral disease. Chemotherapy is nowadays the almost exclusive way to fight the disease but limited by its scarce therapeutic arsenal, on its own compromised by adverse side effects and clinical resistance. Cyclobenzaprine (CBP), an FDA-approved oral muscle relaxant drug has previously demonstrated in vitro and in vivo activity against Leishmania sp., but its targets were not fully unveiled. This study aimed to define the role of energy metabolism as a target for the leishmanicidal mechanisms of CBP. Methodology to assess CBP leishmanicidal mechanism variation of intracellular ATP levels using living Leishmania transfected with a cytoplasmic luciferase. Induction of plasma membrane permeability by assessing depolarization with DiSBAC(2)3 and entrance of the vital dye SYTOX® Green. Mitochondrial depolarization by rhodamine 123 accumulation. Mapping target site within the respiratory chain by oxygen consumption rate. Reactive oxygen species (ROS) production using MitoSOX. Morphological changes by transmission electron microscopy. CBP caused on L. infantum promastigotes a decrease of intracellular ATP levels, with irreversible depolarization of plasma membrane, the collapse of the mitochondrial electrochemical potential, mild uncoupling of the respiratory chain, and ROS production, with ensuing intracellular Ca2+ imbalance and DNA fragmentation. Electron microscopy supported autophagic features but not a massive plasma membrane disruption. The severe and irreversible mitochondrial damage induced by CBP endorsed the bioenergetics metabolism as a relevant target within the lethal programme induced by CBP in Leishmania. This, together with the mild-side effects of this oral drug, endorses CBP as an appealing novel candidate as a leishmanicidal drug under a drug repurposing strategy., This work was supported by grants of Sao Paulo State Research Foundation (FAPESP 2019/10434-4 to S.E.T.B. and 2021/04464-8 and 2017/50333-7 to A.G.T.); Subdirección General de Redes y Centros de Investigación Cooperativa - FEDER (RICET RD16/0027/0010, and Ministerio de Ciencia e Innovación España. Plan Estatal de Investigación Científica y Técnica y de Innovación 2017-2020-FEDER PID2019- 108166GB-100 /AEI/10.13039/501100011033 to L. R.), EADS-CASA/Brazilian Air Force (FAB) mobility program to M.L.L., and Coordenaçao de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) to M. L.L.

42 p.-8 fig., Drug repurposing affords the implementation of new treatments at a moderate cost and under a faster time-scale. Most of the clinical drugs against Leishmania share this origin. The antidepressant sertraline has been successfully assayed in a murine model of visceral leishmaniasis. Nevertheless, sertraline targets in Leishmania were poorly defined. In order to get a detailed insight into the leishmanicidal mechanism of sertraline on Leishmania infantum, unbiased multiplatform metabolomics and transmission electron microscopy were combined with a focused insight into the sertraline effects on bioenergetics metabolism of the parasite. Sertraline induced respiration uncoupling, a significant decrease of intracellular ATP level, and oxidative stress in L. infantum promastigotes. Metabolomics evidenced an extended metabolic disarray caused by sertraline. This encompasses a remarkable variation of the levels of thiol-redox and polyamine biosynthetic intermediates, as well as shortage of intracellular amino acids used as metabolic fuel by Leishmania Sertraline killed Leishmania through a multitarget mechanism of action, tackling essential metabolic pathways of the parasite. As such, sertraline is a valuable candidate for visceral leishmaniasis treatment under a drug repurposing strategy., This work was supported by grants of the Fondo de Investigaciones Sanitarias-ISCIII-FEDER (PI12-02706), and (RD16/0027/0010, Red de Enfermedades Tropicales, subprogram RETICS del Plan Estatal de I+D+i (2013-2016) co-financed with FEDER funds, SAF2015-65740-R, and CSIC PIE 201620E038 to L. R., São Paulo State Research Foundation (FAPESP 2015/23403-9 to A. G.T.); EADS-CASA/ Brazilian Air Force (FAB) mobility program to M.L.L., and Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) to M.L.L.