Phyla-specific sterol pathway in the trypanosomes and its chemotherapeutic implications
The sterol composition and growth of Trypanosoma cruzi epimastigotes, the causative agent for Chagas' Disease and Trypanosoma brucei procyclic cells, the causative agent for African sleeping sickness, grown in the absence and presence of a series of sterol analogs with C24/C25- nitrogen- or methylenecyclopropane-containing derivatives were determined, as well as the incorporation of 2H- and 13C-labeled intermediates into 24-alkyl sterols by T. brucei. The concentrations of inhibitors in the culture medium required for 50% inhibition of growth (IC50) was associated with the mechanism of action of the test drug as a postulated transition state analog or mechanism-based inactivator of the C24-methylation reaction catalyzed by sterol C24-methyltransferase (24-SMT). The most potent growth inhibitors had IC50 values in the low nanomolar range. As expected, the main sterols to accumulate in the treated cultures were C24-desalkyl sterols accompanied by a loss of 24-alkyl sterols essential for the membrane architecture and normal growth. The de novo sterol pathway in the two parasites was found to be different and distinct from the human host that lacks the gene for the 24-SMT. According to the literature the T. cruzi pathway can operate a leucine-based fluxome to the final sterols ergosterol (24?-methylcholesta-5,7,22-trienol) 1 and 24-ethylcholesta-5,7,22-trienol 2. In contradistinction, we found that the T. brucei pathway synthesizes a novel set of 24-alkyl sterol end products, including a new functional sterol trypanosterol (24?-methylcholesta-5,7,25(27)-trienol) 3 and its companion ergosta-5,7,24(25)-trienol 4 by the acetate-mevalonate pathway, as evidenced by the sterol identities determined by GC-MS and 1H and 13C-NMR and the pattern of isotope labeling in the 24-alkyl sterols isolated from the cells incubated with [methyl-2H3]AdoMet and [1-13C]glucose. The results are interpreted to imply that T. brucei operates a phyla-distinct biosynthetic pathway from lanosterol to trypanosterol, affording the possibility for new chemotherapeutic leads.