Anti-trypanosomal activity and target validation of the M17 leucyl-aminopeptidase inhibitor DDD00057570 in Trypanosoma cruzi

Abstract

Background: Chagas disease, caused by the parasite Trypanosoma cruzi, affects six to seven million people, and its treatment still relies on two drugs introduced more than fifty years ago, both of limited efficacy and poorly tolerated. The major acidic M17 leucyl-aminopeptidase of T. cruzi is expressed in all developmental stages of the parasite and has been proposed as a drug target, but it has not been pharmacologically validated. Methodology/Principal Findings: We assessed DDD00057570, a compound originally identified as an inhibitor of the orthologue enzyme of Leishmania major, against the recombinant T. cruzi enzyme and against the parasite. The compound inhibited the recombinant enzyme with an IC50 of 2.7 {+/-} 0.9 {micro}M and reduced the proliferation of insect-stage epimastigotes, with a half-maximal effective concentration of 3.7 {+/-} 1.1 {micro}M. Parasites engineered to overexpress the enzyme were twice as resistant to the compound as wild-type parasites, indicating that the enzyme is a primary target in the living parasite. Treatment increased the proportion of epimastigotes with fragmented DNA. In infected mammalian cells, the compound reduced the release of trypomastigotes, the stage responsible for dissemination in the mammalian host, with a half-maximal effective concentration of 1.3 {+/-} 0.6 {micro}M, while its half-maximal cytotoxic concentration on CHO-K1 mammalian host cells was 41.5 {+/-} 3.9 {micro}M, resulting in a selectivity index of approximately 32. Conclusions/Significance: These results validate the major M17 leucyl-aminopeptidase as a drug target in T. cruzi through target overexpression, and identify DDD00057570 as a selective lead structure warranting mechanistic, pharmacokinetic and in vivo studies toward a new treatment for Chagas disease.