Inhibition of sterol 4-demethylation in Candida albicans by 6-amino-2-n-pentylthiobenzothiazole, a novel mechanism of action for an antifungal agent

Abstract

The effects of 6-amino-2-n-pentylthiobenzothiazole (APB), a new antifungal agent, on ergosterol biosynthesis in Candida albicans and Saccharomyces cerevisiae were studied, using [ 14 C]acetate incorporation. In C. albicans, the inhibition of growth was accompanied by a marked inhibition of acetate incorporation in 4-desmethylsterols, with a significant portion of the radiolabel being incorporated in 4,4-dimethylsterols, lanosterol, and 4,4-dimethylzymosterol and minor amounts being incorporated in 4-methylsterols and squalene. The data are interpreted as evidence of a block of the ergosterol biosynthesis pathway at the level of 4-demethylation of 4,4-dimethylzymosterol, with partial inhibition of lanosterol 14-demethylation and squalene epoxidation also being possible. In 6-amino-2-n-pentylthiobenzothiazole-treated S. cerevisiae, a significant amount of the radiolabel was incorporated also in 4-methylsterols, 4-methylzymosterol, and 4-methylfecosterol, indicating that in this microorganism there are different sensitivities of the two 4-demethylations and that the pathway is blocked at the level of 4-demethylation of 4-methylsterols.Growth of the population with altered immunity, which has taken place during the last 15 years, has led to an increase in the incidence of invasive and systemic mycoses. In spite of recent advances in the development of antimycotic agents, antifungal chemotherapy remains in many cases problematic and the search for new antifungal agents continues (16).A promising new antifungal agent is 6-amino-2-n-pentylthiobenzothiazole (APB) (Fig. 1). It has been shown to be active against various Candida species in vitro, to inhibit the yeastmycelium conversion in Candida albicans, and to be active in vivo, curing systemic candidosis in mice (4,5,12).The mechanism of action of APB in Saccharomyces cerevisiae was studied (11): there was a reduction in the cellular content of ergosterol, which was accompanied by an accumulation of squalene and 4-methylated sterols, following APB treatment. These effects differed from those caused by other antifungal drugs, such as azoles, which are established ergosterol biosynthesis inhibitors. It was previously suggested (11) that APB was an inhibitor of a new target site in the ergosterol biosynthesis pathway, namely, sterol 4-demethylation.In this paper, we report the effects of APB on growth and on ergosterol biosynthesis in a clinically important yeast-like microorganism, C. albicans, and present evidence that the antifungal agent inhibits ergosterol biosynthesis at the level of 4-demethylation. So as to better understand the mechanism of action of the compound, the effects on C. albicans are compared with those observed with S. cerevisiae. MATERIALS AND METHODSAntifungal agent. APB (Fig. 1) was provided by E. Sidóová, Chemical Institute, Faculty of Natural Sciences, Comenius University, Bratislava, Slovakia. Solutions of APB were freshly prepared in dimethyl sulfoxide when required.The amount of the solvent in the culture media did not exceed 0.1% (vol/vol) a...

Steroid-transforming enzymes in fungi

“…It has been confirmed that 6-amino-2-n-pentylthiobenzothiazole inhibits ergosterol biosynthesis at the level of 4-demethylation in S. cerevisiae and C. albicans [46]. Over the last years, two compounds have been isolated from Penicillium sp.…”

Inhibition of Sterol Biosynthesis Reduces Tombusvirus Replication in Yeast and Plants

“…To further test the role of ERG25 in TBSV replication, we took advantage of a potent chemical inhibitor, namely, 6-amino-2-n-pentylthiobenzothiazole (APB). APB has been shown to bind to Erg25p and inhibits its catalytic function in a competitive manner (9,21). The treatment of yeast with APB inhibited the biosynthesis of ergosterol and led to the accumulation of the methylated sterol precursors, such as ergosta-5,7-dienol and squalene, but had no significant effect on the composition and the rate of biosynthesis of fatty acids (22).…”

New Anticandidous 2-Alkylthio-6-aminobenzothiazoles

“…The best antibacterial activity against Staphylococcus aureus, both against strain ATEC 25923 and a clinical isolate, was manifested by compund 6 with alkenyl substituent of right chain C 6 ( Table 2). The mechanism of candida inhibition by 2-alkenylthio-6-aminobenzothiazoles 1 -6 is probably the same as in the case of APB [11,12]. For this compound the mechanism of inhibition has been studied for Saccharomyces cerevisiae as well.…”