UC San Diego

Proteases are fundamental to successful parasitism, including for the schistosome flatworm parasite, which causes the disease schistosomiasis in 200 million people worldwide. One protease complex, the proteasome, is receiving much attention as a potential drug target for treatment of a variety of infectious parasitic diseases but has been relatively understudied in the schistosome. Adult Schistosoma mansoni was incubated in the presence of 1 µM of the commercial proteasome inhibitors bortezomib, carfilzomib and MG132. After 24 h, bortezomib and carfilzomib decreased worm motility by more than 85% and inhibited endogenous proteasome activity by >75% relative to DMSO controls. The clear association between the engagement of the proteasome target, and the phenotypic and biochemical effects recorded, encouraged the two-step chromatographic enrichment of the S. mansoni proteasome (Sm20S). Proteomic analysis of Sm20S identified 14 subunits each with an ortholog in the constitutive human proteasome. Activity assays with standard fluorescent substrates for the β1, β2 and β5 proteasome subunits revealed that Sm20S contains caspase-type (β1), trypsin-type (β2) and chymotrypsin-type (β5) activities. Sm20S was screened with 11 peptide epoxyketone inhibitors derived from the marine natural product, carmaphycin B. Analog 17 exhibited preferential inhibition of the Sm20S β2 and β5 subunits over that of c20S and i20S. Furthermore, 1 µM of 17 decreased both worm motility and endogenous Sm20S activity by more than 90% after 24 h. We provide direct evidence of the proteasome’s importance to schistosome viability and identify a lead for which future studies will aim to improve the potency, selectivity and safety.