Cutaneous and visceral leishmaniasis are amongst the most devastating infectious diseases of our time, affecting millions of people worldwide. The treatment of these serious diseases rely on a few chemotherapeutic agents, most of which are of parenteral use and induce severe side-effects. Furthermore, rates of treatment failure are high and have been linked to drug resistance in some areas. Here, we reviewed data on current chemotherapy practice in leishmaniasis. Drug resistance and mechanisms of resistance are described as well as the prospects for applying drug combinations for leishmaniasis chemotherapy. It is clear that efforts for discovering new drugs applicable to leishmaniasis chemotherapy are essential. The main aspects on the various steps of drug discovery in the field are discussed.

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Miltefosine was the first oral compound approved for visceral leishmaniasis chemotherapy, and its efficacy against Leishmania donovani has been well documented. Leishmania amazonensis is the second most prevalent species causing cutaneous leishmaniasis and the main etiological agent of diffuse cutaneous leishmaniasis in Brazil. Driven by the necessity of finding alternative therapeutic strategies for a chronic diffuse cutaneous leishmaniasis patient, we evaluated the susceptibility to miltefosine of the Leishmania amazonensis line isolated from this patient, who had not been previously treated with miltefosine. In vitro tests against promastigotes and intracellular amastigotes showed that this parasite isolate was less susceptible to miltefosine than L. amazonensis type strains. Due to this difference in susceptibility, we evaluated whether genes previously associated with miltefosine resistance were involved. No mutations were found in the miltefosine transporter gene or in the Ros3 or pyridoxal kinase genes. These analyses were conducted in parallel with the characterization of L. amazonensis mutant lines selected for miltefosine resistance using a conventional protocol to select resistance in vitro, i.e., exposure of promastigotes to increasing drug concentrations. In these mutant lines, a single nucleotide mutation G852E was found in the miltefosine transporter gene. In vivo studies were also performed to evaluate the correlation between in vitro susceptibility and in vivo efficacy. Miltefosine was effective in the treatment of BALB/c mice infected with the L. amazonensis type strain and with the diffuse cutaneous leishmaniasis isolate. On the other hand, animals infected with the resistant line bearing the mutated miltefosine transporter gene were completely refractory to miltefosine chemotherapy. These data highlight the difficulties in establishing correlations between in vitro susceptibility determinations and response to chemotherapy in vivo. This study contributed to establish that the miltefosine transporter is essential for drug activity in L. amazonensis and a potential molecular marker of miltefosine unresponsiveness in leishmaniasis patients.

Leishmaniasis chemotherapy remains very challenging. The high cost of active drugs, along with the severity of their side effects and the increasing failure rate of the current therapeutic schemes, calls for the discovery of new active drugs and schemes of treatment. The use of combination therapy has gained much attention in recent years as a possible strategy for overcoming the various shortcomings in the present arsenal. We recently described the effectiveness of tamoxifen in murine models of leishmaniasis, and here, we investigated the interactions between tamoxifen and amphotericin B, one of the most potent drugs used in leishmaniasis treatment. The in vitro interactions were indifferent for the association of tamoxifen and amphotericin B. The association was also assayed in vivo in Leishmania amazonensis-infected BALB/c mice and was found to yield at least additive effects at low doses of both drugs.

Leishmaniasis is a disease found throughout the (sub)tropical parts of the world caused by protozoan parasites of the Leishmania genus. Despite the numerous problems associated with existing treatments, pharmaceutical companies continue to neglect the development of better ones. The high toxicity of current drugs combined with emerging resistance makes the discovery of new therapeutic alternatives urgent. We report here the evaluation of a binuclear cyclopalladated complex containing Pd(II) and N,N=-dimethylbenzylamine (Hdmba) against Leishmania amazonensis. The compound [Pd(dmba)(-N 3)] 2 (CP2) inhibits promastigote growth (50% inhibitory concentration [IC 50 ] 13.2 0.7 M) and decreases the proliferation of intra-cellular amastigotes in in vitro incubated macrophages (IC 50 10.2 2.2 M) without a cytotoxic effect when tested against peritoneal macrophages (50% cy-totoxic concentration 506.0 10.7 M). In addition, CP2 was also active against T. cruzi intracellular amastigotes (IC 50 2.3 0.5 M, selective index 225), an indication of its potential for use in Chagas disease therapy. In vivo assays using L. amazonensis-infected BALB/c showed an 80% reduction in parasite load compared to infected and nontreated animals. Also, compared to ampho-tericin B treatment, CP2 did not show any side effects, which was corroborated by the analysis of plasma levels of different hepatic and renal biomarkers. Furthermore , CP2 was able to inhibit Leishmania donovani topoisomerase 1B (Ldtopo1B), a potentially important target in this parasite. (This study has been registered at ClinicalTrials.gov under identifier NCT02169141.)