Stereoselective interaction of the azole antifungal agent SCH39304 with the cytochrome P-450 monooxygenase system isolated from Cryptococcus neoformans.
We investigated the stereoselective inhibition of growth and ergosterol biosynthesis by SCH39304 in the pathogenic fungus Cryptococcus neoformans obtained from four AIDS patients who failed fluconazole therapy and compared the results to those obtained with a wild-type strain. For all strains, the MICs of the RR isomer were approximately half those of the racemate, with the SS enantiomer showing no inhibitory activity. The 50% inhibitory concentrations for in vitro ergosterol biosynthesis correlated with the MIC data, indicating stereoselective inhibition of their target P-450 enzyme, sterol 14alpha-demethylase, as the cause of this difference. The RR enantiomer produced classical type II spectra on addition to microsomal extracts of the strains, whereas the SS enantiomer showed an absence of binding. Stereo- and regio-specific localization of N-1 substituent groups of SCH39304 within the active site of the enzyme determined the unique discrimination between its two enantiomers, and the inability to bind to sterol 14alpha-demethylase is also true of other P-450 enzymes contained in the microsomal fraction. As previously observed for other antifungal azoles, isolates obtained following failure of fluconazole therapy showed resistance to SCH39304 and its RR enantiomer. This resistance could be associated with an alteration in the sensitivity of ergosterol biosynthesis in vitro. These alterations did not cause any changes allowing the SS enantiomer to bind to the P-450 mediating sterol 14alpha-demethylation.
Lamb
D C
DC
Department of Molecular Biology and Biotechnology, Krebs Institute for Biomolecular Research, Sheffield University, United Kingdom.
Baldwin
B C
BC
Kwon-Chung
K J
KJ
Kelly
S L
SL
D000935:Antifungal Agents / Q000627:therapeutic use*; D003455:Cryptococcus neoformans / Q000187:drug effects* / Q000201:enzymology; D003577:Cytochrome P-450 Enzyme System / Q000187:drug effects* / Q000302:isolation & purification; D004875:Ergosterol / Q000096:biosynthesis; D006801:Humans; D008826:Microbial Sensitivity Tests; D008861:Microsomes / Q000187:drug effects; D013237:Stereoisomerism; D014230:Triazoles / Q000627:therapeutic use*
D000935:Antifungal Agents; D014230:Triazoles; C056513:Sch 39304; D003577:Cytochrome P-450 Enzyme System; D004875:Ergosterol