Investigation of the threonine metabolism of Echinococcus multilocularis: EmTDH as a potential drug target in alveolar echinococcosis

ABSTRACT: Alveolar echinococcosis is a severe zoonotic disease caused by the fox tapeworm E. multilocularis. New treatment options are urgently needed. Whole-organism-based screening allows the discovery of potential new anti-echinococcal compounds, but suffers from low- to medium-throughput. An alternative approach to identify novel compounds is target-based screening, which requires in-depth knowledge of the basic biology of the pathogen of interest. We recently showed that E. multilocularis metacestode vesicles scavenge large amounts of L-threonine from the culture medium that was neither stored nor overused for protein synthesis. This motivated us to study the effect of L-threonine on the parasite and how it is metabolized. We showed that L-threonine significantly increased growth of E. multilocularis metacestode vesicles and the formation of novel metacestode vesicles from germinal layer cell cultures. The non-proteinogenic threonine analog 3-hydroxynorvaline had the contrary effect. We traced [U-13C]-L-threonine and metabolites in metacestode vesicles and culture medium and detected [13C]-labeling in aminoacetone and glycine, indicating that L-threonine was metabolized by threonine dehydrogenase (TDH) and 2-amino-3-ketobutyrate coenzyme A ligase (KBL), but not by threonine deaminase. We further detected [13C2]-glutathione, which suggested that E. multilocularis metacestode vesicles synthesize glutathione via L-threonine-derived glycine in vitro. EmTDH mediated threonine metabolism in E. multilocularis metacestode vesicles was further confirmed by quantitative real-time PCR which showed high expression of emtdh. EmTDH was enzymatically active in metacestode vesicle extracts and thus we established an enzymatic assay to evaluate the activity of compounds against recombinantly expressed EmTDH. We tested 11 compounds (disulfiram, myricetin, seven quinazoline carboxamides, quercetin and sanguinarine) and of these, disulfiram, myricetin and sanguinarine were highly active against recEmTDH. Upon testing of these three compounds against E. multilocularis metacestode vesicles and GL cells, sanguinarine showed high in vitro activity and was thus further characterized by calculating IC50 values against metacestode vesicles, GL cells, as well as mammalian cells. Our results suggested that sanguinarine should be further followed for its potential activity against E. multilocularis in mouse models of AE. Furthermore, the here-established TDH assay could serve as high-throughput target-based discovery platform for novel anti-echinococcal compounds in the future.

INSTRUMENT(S): Liquid Chromatography MS - positive - hilic, Liquid Chromatography MS - negative - hilic

PROVIDER: MTBLS10738 | MetaboLights | 2026-09-14

REPOSITORIES: MetaboLights