Defective sterol C5-6 desaturation and azole resistance: A new hypothesis for the mode of action of azole antifungals

Resistant P45051A1 activity in azole antifungal tolerant Cryptococcus neoformans from AIDS patients

“…The molecular basis of azole resistance in fungi is a subject of current interest due to its emergence as a clinical problem and also in agriculture [10]. Sterol A 5,6 desaturase has been shown to play a critical role in azole mode of action and resistance in S. cerevisiae, being required for the formation of 14a-methylergosta-24(28)-dien-3fl,6a-diol under azole antifungal treatment, which is incompatible with sterol function in membranes [15]. Mutants blocked in this enzyme do not synthesise the diol and accumulate the precursor sterol, 14a-methylfecosterol, which is capable of supporting growth.…”

“…For microsomal fractions the cell-free extract was subject to further centrifugation, first at 10000 x g to pellet mitochondria and subsequently at 100000 × g to produce the microsomal pellet. This was resuspended to approximately 10 mg protein/ml in 20% glycerol, 100 mM phosphate pH 7.4 and P450 estimated using the method of Omura and Sato [15]. the cell-free biosynthesis of ergosterol was assayed.…”

Altered P450 activity associated with direct selection for fungal azole resistance

“…However, one 14~-methylated sterol does satisfy the growth requirements of S. cerevisiae. 14~-Methylfecosterol accumulates in sterol A 5'6 desaturase mutants under azole treatment instead of 14-methyl-3,6-diol [10] and results in azole resistance. This mechanism of resistance was not apparent in the U. maydis studies reported here in which mutants were challenged directly with azole antifungal, but a possible leaky sterol/[5(6) desaturase mutant which is azole resistant is the subject of further investigation.…”

“…The predominant mechanism of azole resistance for S. eerevisiae is through a block in sterol A 5'6 desaturase activity [10]. This results instead in the accumulation of 14~-methylfecosterol which can fulfill requirements for growth.…”

Microbial cytochromes P450: biodiversity and biotechnology. Where do cytochromes P450 come from, what do they do and what can they do for us?

“…This is not yet clear for resistance in filamentous fungi. The other mechanisms clearly identified in C. albicans are increased expression of drug transporters, particularly CDR1, through mutation in a transcription factor [160] and altering the pathway of 14a-demethylated sterols accumulating under treatment away from a toxic 14a-methyl-3,6-diol end product to 14a-methylfecosterol that C. albicans can grow on [161,162]. This latter mechanism involves suppression of the effect of azoles through mutation in the sterol C5-desaturase (Erg3).…”