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.

A high-throughput screening campaign of a library of 100,000 lead-like compounds identified 2-iminobenzimidazoles as a novel class of trypanothione reductase inhibitors. These 2-iminobenzimidazoles display potent trypanocidal activity against Trypanosoma brucei rhodesiense, do not inhibit closely related human glutathione reductase and have low cytotoxicity against mammalian cells. KeywordsTropical diseases; Trypanosomiasis therapeutics; Trypanothione reductase inhibitors; Highthroughput screening; Medicinal chemistry; Imino benzimidazoles Parasitic protozoa of the family Trypanosomatidae are the causative agent of many significant tropical diseases including African trypanosomiasis, Chagas disease and Leishmaniasis. In the 2004 world health report 1 African trypanosomiasis was reported to cause 48 thousand deaths and a disease burden of 1525 thousand DALYs (disability adjusted life years) annually, Chagas disease 14 thousand deaths and a disease burden of 667 thousand DALYs and Leishmaniasis 51 thousand deaths and a disease burden of 2090 DALYs. There are currently nine key drugs in use for the treatment of these disease states ( Fig. 1): suramin and pentamidine against early stage African trypanosomiasis, and eflornithine and melarsoprol against late stage disease; nifurtimox and benznidazole against early stage Chagas disease; meglumine antimonite and sodium stibogluconate against Leishmaniasis, and amphotericin B against anti-mony-resistant strains. All of these drugs have severe limitations including administration diffculties, long treatment regimes, lifethreatening side effects, varied parasitological cure rates for different strains, lack of effcacy against late stage diseases, and increasing incidence of drug resistance. [2][3][4][5] The intracellular reducing environment of trypanosomatids is maintained by a unique thiol redox system where the glutathione/glutathione reductase (GR) couple found in mammalian cells is replaced by the (bis-glutathionyl)spermidine trypanothione/trypanothione reductase (TR) couple. 6 TR is a key enzyme of the parasite antioxidant defence, 7,8 does not occur in the mammalian host and has been found to be essential for all trypanosomatids currently studied. 9 Europe PMC Funders Author ManuscriptsEurope PMC Funders Author Manuscripts TR and human GR have similar catalytic mechanisms with 14 of the 19 amino acid residues close to the substrate binding site being conserved. However, they are specific to their respective disulfide substrates (Fig. 2). 11 GR has a hydrophilic, positively charged region in its active site that interacts with the glycine carboxylates of glutathione disulfide, while TR has a larger binding site with a negatively charged region with which the spermidine moiety of trypanothione disulfide binds. 12 The absence of TR from the mammalian host and the sensitivity of trypanosomatids to oxidative stress makes TR an attractive target for trypanosomiasis therapeutics. 13 The objective of this work, therefore, was to identify novel classes of TR inhibit...

We report that TBK-1 inhibition using WEHI-112 abrogated antibody-dependent CIA. As WEHI-112 failed to inhibit non-antibody-driven joint inflammation, we conclude that the major effect of this compound was most likely the targeting of TBK-1-mediated mechanisms in the GC reaction. This approach may have therapeutic potential in RA and in other GC-associated autoantibody-driven inflammatory diseases.

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