Chagas disease is a neglected tropical disease endemic to Latin America, though migratory movements have recently spread it to other regions. Here, we have applied a cascade virtual screening campaign combining ligand- and structure-based methods. In order to find novel inhibitors of putrescine uptake in Trypanosoma cruzi, an ensemble of linear ligand-based classifiers obtained by has been applied as initial screening filter, followed by docking into a homology model of the putrescine permease TcPAT12. 1,000 individual linear classifiers were inferred from a balanced dataset. Subsequently, different schemes were tested to combine the individual classifiers: MIN operator, average ranking, average score, average voting, with MIN operator leading to the best performance. The homology model was based on the arginine/agmatine antiporter (AdiC) from Escherichia coli as template. It showed 64% coverage of the entire query sequence and it was selected based on the normalized Discrete Optimized Protein Energy parameter and the GA341 score. The modeled structure had 96% in the allowed area of Ramachandran's plot, and none of the residues located in non-allowed regions were involved in the active site of the transporter. Positivity Predictive Value surfaces were applied to optimize the score thresholds to be used in the ligand-based virtual screening step: for that purpose Positivity Predictive Value was charted as a function of putative yields of active in the range 0.001–0.010 and the Se/Sp ratio. With a focus on drug repositioning opportunities, DrugBank and Sweetlead databases were subjected to screening. Among 8 hits, cinnarizine, a drug frequently prescribed for motion sickness and balance disorder, was tested against T. cruzi epimastigotes and amastigotes, confirming its trypanocidal effects and its inhibitory effects on putrescine uptake. Furthermore, clofazimine, an antibiotic with already proven trypanocidal effects, also displayed inhibitory effects on putrescine uptake. Two other hits, meclizine and butoconazole, also displayed trypanocidal effects (in the case of meclizine, against both epimastigotes and amastigotes), without inhibiting putrescine uptake.

No abstract

IL-10 is a pleiotropic cytokine with immunoregulatory functions affecting various cell types.In a model of experimental infection with the protozoan Trypanosoma cruzi (T. cruzi), we found increased morbidity and lower parasite control in IL-10 deficient mice (IL-10 KO) compared to wild-type (WT) mice. Despite enhanced M function and dendritic cell activation, IL-10 KO mice were more susceptible to infection. The kinetics of T cells in spleen and peripheral blood revealed that infected IL-10 KO mice failed to increase the number of spleen and circulating total CD8 + T cells, a phenomenon observed from the second week of infection in WT mice. Total CD8 + T cells from IL-10 KO mice exhibited diminished proliferation, cytotoxic potential and IFN-production than their WT counterparts and T. cruzi-specific CD8 + T cells displayed reduced in vivo cytotoxicity. The absence of IL-10 selectively affected expansion, survival, and increased PD-1 expression of CD8 + T cells without altering these same parameters on CD4 + T cells. Increased inhibitory receptors expression and down-modulation of T-bet by CD8 + T cells from IL-10 KO infected mice were compatible with a T cell exhaustion phenotype. Collectively, these findings reveal that during acute infection, IL-10 plays a previously unrecognized stimulatory role on CD8 + T cells, the most relevant lymphocyte population for the control of intracellular T. cruzi stages. A clear knowledge of the underlying mechanisms that drive effector functions of cytotoxic T cells is critical to understand pathogen persistence and rational design of prophylactic strategies against T. cruzi.