Trypanothione Reductase High-Throughput Screening Campaign Identifies Novel Classes of Inhibitors with Antiparasitic Activity

Abstract

High-throughput screening of 100,000 lead-like compounds led to the identification of nine novel chemical classes of trypanothione reductase (TR) inhibitors worthy of further investigation. Hits from five of these chemical classes have been developed further through different combinations of preliminary structure-activity relationship rate probing and assessment of antiparasitic activity, cytotoxicity, and chemical and in vitro metabolic properties. This has led to the identification of novel TR inhibitor chemotypes that are drug-like and display antiparasitic activity. For one class, a series of analogues have displayed a correlation between TR inhibition and antiparasitic activity. This paper explores the process of identifying, investigating, and evaluating a series of hits from a high-throughput screening campaign.

Discovery of novel and potent benzhydryl-tropane trypanocides highly selective for Trypanosoma cruzi

“…We have previously reported on a high-throughput screening campaign conducted to identify inhibitors of trypanothione reductase (TR) that might consequently be toxic to trypanosomatids. 4,5 Amongst the identified hits was a benzhydryl tropinone oxime that was found to be potently toxic towards T. cruzi with an EC 50 of 110 nM and highly selective for this organism relative to other trypanosomatids such as Leishmania donovani and Trypanosoma brucei. 5 These data are summarized in Figure 2.…”

“…4,5 Amongst the identified hits was a benzhydryl tropinone oxime that was found to be potently toxic towards T. cruzi with an EC 50 of 110 nM and highly selective for this organism relative to other trypanosomatids such as Leishmania donovani and Trypanosoma brucei. 5 These data are summarized in Figure 2. For several reasons, significantly amongst these being the very weak trypanothione reductase inhibitory activity of compound 1 (EC 50 35uM, Figure 2), we have previously argued that the observed potent T.cruzi-selective trypanocidal activity could likely be due to off-target activity.…”

“…For several reasons, significantly amongst these being the very weak trypanothione reductase inhibitory activity of compound 1 (EC 50 35uM, Figure 2), we have previously argued that the observed potent T.cruzi-selective trypanocidal activity could likely be due to off-target activity. 5 Subsequently, we gained access to a higher throughput parasite assay and it was principally for this reason that we initiated a hit optimisation campaign around compound 1, using a whole parasite primary assay. This communication will disclose the results of our SAR investigation into this compound.…”

High Throughput Screening against the Peroxidase Cascade of African Trypanosomes Identifies Antiparasitic Compounds That Inactivate Tryparedoxin

“…Different chemotypes were identified that showed selectivity for TR over human glutathione reductase (15). Another approach revealed compounds with potent antiparasitic activity, but only moderate correlation with TR inhibition (6). All enzymes establishing the parasite peroxidase system, namely TR, Tpx, and both types of tryparedoxin peroxidases, have been shown to be essential for T. brucei (16 -19) and thus fulfill a crucial prerequisite of a putative drug target molecule.…”

“…The dithiol is kept reduced by NADPH and the flavoenzyme trypanothione reductase (TR), the system replacing the glutathione/glutathione reductase couple of the mammalian host (4). TR is the most thoroughly studied enzyme of the trypanothione metabolism, and a large number of inhibitors has been identified (5)(6)(7)(8)(9). Tpx is a distant member of the thioredoxin (Trx) protein family.…”