Mapping the Pro-Peptide of the <i>Schistosoma mansoni</i> Cathepsin B1 Drug Target: Modulation of Inhibition by Heparin and Design of Mimetic Inhibitors

Blood flukes of the genus <i>Schistosoma</i> cause the disease schistosomiasis that infects over 200 million people worldwide. Treatment relies on just one drug, and new therapies are needed should drug resistance emerge. <i>Schistosoma mansoni</i> cathepsin B1 (SmCB1) is a gut-associated protease that digests host blood proteins as source of nutrients. It is under evaluation as a therapeutic target. Enzymatic activity of the SmCB1 zymogen is prevented by the pro-peptide that sterically blocks the active site until activation of the zymogen to the mature enzyme. We investigated the structure−inhibition relationships of how the SmCB1 pro-peptide interacts with the enzyme core using a SmCB1 zymogen model and pro-peptide-derived synthetic fragments. Two regions were identified within the pro-peptide that govern its inhibitory interaction with the enzyme core: an “active site region” and a unique “heparin-binding region” that requires heparin. The latter region is apparently only found in the pro-peptides of cathepsins B associated with the gut of trematode parasites. Finally, using the active site region as a template and a docking model of SmCB1, we designed a series of inhibitors mimicking the pro-peptide structure, the best of which yielded low micromolar inhibition constants. Overall, we identify a novel glycosaminoglycan-mediated mechanism of inhibition by the pro-peptide that potentially regulates zymogen activation and describe a promising design strategy to develop antischistosomal drugs.