Efficacy, metabolism and pharmacokinetics of Ro 15-5458, a forgotten schistosomicidal 9-acridanone hydrazone.

BACKGROUND: Treatment of schistosomiasis, a neglected disease, relies on just one partially effective drug, praziquantel. We revisited the 9-acridanone hydrazone, Ro 15-5458, a largely forgotten antischistosomal lead compound. METHODS: Ro 15-5458 was evaluated in juvenile and adult Schistosoma mansoni-infected mice. We studied dose-response, hepatic shift and stage specificity. The metabolic stability of Ro 15-5458 was measured in the presence of human and mouse liver microsomes, and human hepatocytes; the latter also served to identify metabolites. Pharmacokinetic parameters were measured in naive mice. The efficacy of Ro 15-5458 was also assessed in S. haematobium-infected hamsters and S. japonicum-infected mice. RESULTS: Ro 15-5458 had single-dose ED50 values of 15 and 5.3 mg/kg in mice harbouring juvenile and adult S. mansoni infections, respectively. An ED50 value of 17 mg/kg was measured in S. haematobium-infected hamsters; however, the compound was inactive at up to 100 mg/kg in S. japonicum-infected mice. The drug-induced hepatic shift occurred between 48 and 66 h post treatment. A single oral dose of 50 mg/kg of Ro 15-5458 had high activity against all tested S. mansoni stages (1-, 7-, 14-, 21- and 49-day-old). In vitro, human hepatocytes produced N-desethyl and glucuronide metabolites; otherwise Ro 15-5458 was metabolically stable in the presence of microsomes or whole hepatocytes. The maximum plasma concentration was approximately 8.13 µg/mL 3 h after a 50 mg/kg oral dose and the half-life was approximately 4.9 h. CONCLUSIONS: Ro 15-5458 has high activity against S. mansoni and S. haematobium, yet lacks activity against S. japonicum, which is striking. This will require further investigation, as a broad-spectrum antischistosomal drug is desirable.

The Proteasome as a Drug Target in the Metazoan Pathogen, Schistosoma mansoni.

Proteases are fundamental to successful parasitism, including that of the schistosome flatworm parasite, which causes the disease schistosomiasis in 200 million people worldwide. The proteasome is receiving attention as a potential drug target for treatment of a variety of infectious parasitic diseases, but it has been understudied in the schistosome. Adult Schistosoma mansoni were incubated with 1 µM concentrations of the proteasome inhibitors bortezomib, carfilzomib, and MG132. After 24 h, bortezomib and carfilzomib decreased worm motility by more than 85% and endogenous proteasome activity by >75%, and after 72 h, they increased caspase activity by >4.5-fold. The association between the engagement of the proteasome target and the phenotypic and biochemical effects recorded encouraged the chromatographic enrichment of the S. mansoni proteasome (Sm20S). Activity assays with fluorogenic proteasome substrates revealed that Sm20S contains caspase-type (ß1), trypsin-type (ß2), and chymotrypsin-type (ß5) activities. Sm20S was screened with 11 peptide epoxyketone inhibitors derived from the marine natural product carmaphycin B. Analogue 17 was 27.4-fold less cytotoxic to HepG2 cells than carmaphycin B and showed equal potency for the ß5 subunits of Sm20S, human constitutive proteasome, and human immunoproteasome. However, this analogue was 13.2-fold more potent at targeting Sm20S ß2 than it was at targeting the equivalent subunits of the human enzymes. Furthermore, 1 µM 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.