Visceral leishmaniasis (VL), caused by the protozoan parasites Leishmania donovani and L. infantum, is responsible for ∼30 000 deaths annually. Available treatments are inadequate, and there is a pressing need for new therapeutics. N-Myristoyltransferase (NMT) remains one of the few genetically validated drug targets in these parasites. Here, we sought to pharmacologically validate this enzyme in Leishmania. A focused set of 1600 pyrazolyl sulfonamide compounds was screened against L. major NMT in a robust high-throughput biochemical assay. Several potent inhibitors were identified with marginal selectivity over the human enzyme. There was little correlation between the enzyme potency of these inhibitors and their cellular activity against L. donovani axenic amastigotes, and this discrepancy could be due to poor cellular uptake due to the basicity of these compounds. Thus, a series of analogues were synthesized with less basic centers. Although most of these compounds continued to suffer from relatively poor antileishmanial activity, our most potent inhibitor of LmNMT (DDD100097, Ki of 0.34 nM) showed modest activity against L. donovani intracellular amastigotes (EC50 of 2.4 μM) and maintained a modest therapeutic window over the human enzyme. Two unbiased approaches, namely, screening against our cosmid-based overexpression library and thermal proteome profiling (TPP), confirm that DDD100097 (compound 2) acts on-target within parasites. Oral dosing with compound 2 resulted in a 52% reduction in parasite burden in our mouse model of VL. Thus, NMT is now a pharmacologically validated target in Leishmania. The challenge in finding drug candidates remains to identify alternative strategies to address the drop-off in activity between enzyme inhibition and in vitro activity while maintaining sufficient selectivity over the human enzyme, both issues that continue to plague studies in this area.
Wild rodents constitute a very large biomass of potential reservoirs for Leishmania spp. Therefore, an epidemiological study was carried out in a well-known focus of canine leishmaniasis from southern Spain, with the objective of detecting and characterizing Leishmania infantum infection in wild rodents. Blood, liver, spleen, bone marrow, and skin from 37 rodents (24 Apodemus sylvaticus, 9 Rattus rattus, and 4 Mus musculus) were analyzed by optical microscopy, culture, and two different polymerase chain reactions. L. infantum DNA was found in 27% (10 out of 37) of the trapped rodents, in a variety of tissues: bone marrow, spleen, or healthy skin (ear lobe). High prevalences of L. infantum infection were found in the three investigated rodent species. The presence of other trypanosomatids was also evidenced. These rodent species are abundant, widely distributed in Europe, and have a long enough lifespan to overcome the low sandfly activity season. They live in a suitable habitat for sandflies and serve as blood sources for these insects, which can become infected when induced to feed on Leishmania-infected animals. Whether they are reservoirs or just irrelevant incidental hosts, it is clear that the epidemiology of L. infantum is more complex than previously thought, and so is its control. The classic epidemiological cycle dog-sandfly-human is turning into a network of animal species that collaborate with the dog in the maintenance of the parasite under natural conditions and probably showing local differences.
Visceral leishmaniasis (VL) is a parasitic disease endemic across multiple regions of the world and is fatal if untreated. Current therapies are unsuitable, and there is an urgent need for safe, short-course, and low-cost oral treatments to combat this neglected disease. The benzoxaborole chemotype has previously delivered clinical candidates for the treatment of other parasitic diseases. Here, we describe the development and optimization of this series, leading to the identification of compounds with potent in vitro and in vivo antileishmanial activity. The lead compound (DNDI-6148) combines impressive in vivo efficacy (>98% reduction in parasite burden) with pharmaceutical properties suitable for onward development and an acceptable safety profile. Detailed mode of action studies confirm that DNDI-6148 acts principally through the inhibition of Leishmania cleavage and polyadenylation specificity factor (CPSF3) endonuclease. As a result of these studies and its promising profile, DNDI-6148 has been declared a preclinical candidate for the treatment of VL.
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