We investigated the in-vitro antiproliferative effects and mechanism of action of both enantiomers of the bis-
triazole derivative
ICI 195,739 against epimastigotes and amastigotes of Trypanosoma cruzi, the aetiological agent of
Chagas' disease. It has recently been shown that the R(+) enantiomer,
D0870, can induce radical parasitological cure in murine models of the acute and chronic forms of the disease.
D0870 dose-dependently affected the growth rate of the epimastigote form (IC50 = 0.1 microM; MIC = 1-3 microM). The S(-) enantiomer was much less active (IC50 = 3 microM). Growth arrest and cell lysis induced by
D0870 coincided with the complete depletion of endogenous 4,14-desmethyl
sterols and their replacement by 4,14-trimethyl and dimethyl
sterols. The S(-) enantiomer produced qualitatively similar changes but to a lesser extent.
D0870 inhibited the incorporation of radioactivity from [2-14C]
acetate into the epimastigote's 4,14-desmethyl
sterols with an IC50 of 50 nM while the corresponding concentration for the S(-) enantiomer was 3 microM.
D0870 eradicated the intracellular (amastigote) form of the parasite from cultured Vero cells
at 10 nM; a 100-fold higher concentration of the S(-) enantiomer was required to produce a similar effect, and deleterious effects of the host cells were observed at > 100 nM. At the MIC of
D0870 the endogenous amastigote
sterols (ergosta-7-en-3beta-ol, 24-ethyl-cholesta-7-en-3beta-ol and ergosta-7, 24(24[1])-dien-3beta-ol) were also largely replaced by
lanosterol and 24-methyl-dihydrolanosterol. Combinations of
D0870 and inhibitors of
sterol delta24(25)
sterol methyltransferase (such as 22,26-azasterol and 24(R,S),25-epiminolanosterol) acted synergically against the intracellular forms. Taken together these results indicate that, although both enantiomers have
anti-T. cruzi activity, the specific activity of the R(+) enantiomer (
D0870) is nearly two orders of magnitude higher than that of its S(-) analogue. However, as the in-vitro activity of
D0870 is comparable to that of standard
azoles, such as
ketoconazole, its remarkable in-vivo
antiparasitic activity may only be explained by its particular pharmacokinetic properties.