Screening approaches towards trypanothione reductase

Trypanosomes and Leishmania, the causative agents of African sleeping sickness, Chagas disease, and Leishmaniasis, are responsible for over half a million human deaths per year in subtropical and tropical regions around the world. Only a handful of chemotherapeutics are available, and their efficacy suffers from drug resistance and serious side-effects. Therefore, new inhibiting compounds of essential targets are urgently needed. Trypanosomes lack both glutathione reductase and thioredoxin reductase, but possess the phylogenetically related trypanothione reductase (TR), a validated drug target and key enzyme of the unique trypanothione-based thiol metabolism. By inhibiting this essential flavoenzyme the parasite becomes vulnerable against oxidative stress induced by the host defense system and drug treatment. Here, we provide an overview of published TR screenings, and present an alternative approach combining in silico and in vitro experiments. Starting from an in vitro screen of 2816 highly diverse compounds, 21 novel TR actives could be identified. These inhibitors were used as the starting point to identify similar substances within a compound library of 200 000 available chemicals by a ligand-based in silico screening. In vitro screening of the resulted in silico-enriched dataset led to the identification of 61 additional TR actives. All 82 TR inhibitors showed IC50 values down to the nanomolar range. Subsequently, the dataset of known actives was used to implement a novel structure-based in silico screening approach that might identify further potential novel TR inhibitors