Abstract

BackgroundCruzain, the major cysteine protease of Trypanosoma cruzi, is an essential enzyme for the parasite life cycle and has been validated as a viable target to treat Chagas' disease. As a proof-of-concept, K11777, a potent inhibitor of cruzain, was found to effectively eliminate T. cruzi infection and is currently a clinical candidate for treatment of Chagas' disease.Methodology/Principal FindingsWRR-483, an analog of K11777, was synthesized and evaluated as an inhibitor of cruzain and against T. cruzi proliferation in cell culture. This compound demonstrates good potency against cruzain with sensitivity to pH conditions and high efficacy in the cell culture assay. Furthermore, WRR-483 also eradicates parasite infection in a mouse model of acute Chagas' disease. To determine the atomic-level details of the inhibitor interacting with cruzain, a 1.5 Å crystal structure of the protease in complex with WRR-483 was solved. The structure illustrates that WRR-483 binds covalently to the active site cysteine of the protease in a similar manner as other vinyl sulfone-based inhibitors. Details of the critical interactions within the specificity binding pocket are also reported.ConclusionsWe demonstrate that WRR-483 is an effective cysteine protease inhibitor with trypanocidal activity in cell culture and animal model with comparable efficacy to K11777. Crystallographic evidence confirms that the mode of action is by targeting the active site of cruzain. Taken together, these results suggest that WRR-483 has potential to be developed as a treatment for Chagas' disease.Author SummaryCurrent drugs for Chagas' disease, caused by Trypanosoma cruzi infection, are limited in efficacy and are severely toxic. Hence the development of novel chemotherapeutic agents targeting T. cruzi infections is an important undertaking. In recent years, there has been considerable interest in cruzain, the major protease in T. cruzi, as a target to treat Chagas' disease. Herein, we present the synthesis of WRR-483, a small molecule designed as an irreversible cysteine protease inhibitor, and an assessment of its biological activity against cruzain and T. cruzi infection. This compound displays pH-dependent affinity for cruzain and highly effective trypanocidal activity in both cell cuture and a mouse model of acute Chagas' disease. The crystal structure of WRR-483 bound to cruzain elucidates the details of inhibitor binding to the enzyme. Based on these results, this inhibitor is a promising compound for the development of therapeutics for Chagas' disease.