title: Comparison of lanosterol-14 alpha-demethylase (CYP51) of human and Candida albicans for inhibition by different antifungal azoles.
abstract: Inhibition of fungal lanosterol-14 alpha-demethylase (CYP51) is the working principle of the antifungal activity of azoles used in agriculture and medicine. Inhibition of human CYP51 may result in endocrine disruption since follicular fluid-meiosis activating steroid (FF-MAS), the direct product of lanosterol demethylation, is involved in the control of meiosis. To investigate the specificity of antifungal agents for the fungal enzyme, assays to determine inhibitory potencies of 13 agricultural fungicides and 6 antimycotic drugs were established. FF-MAS product formation was measured by LC-MS/MS analysis in the incubations using lanosterol as substrate. Recombinant human enzyme (hCYP51) was available from BD Gentest. CYP51 of Candida albicans (cCYP51) was co-expressed with Candida tropicalis oxidoreductase in the baculovirus system. IC(50) values of 13 fungicides for cCYP51 ranged about six-fold (0.059-0.35 microM); for hCYP51 the range was about 30-fold (1.3-37.2 microM). The most favourable IC(50) ratio human to Candida was observed for imazalil (440-fold), while the specificity of epoxiconazole and tebuconazole for cCYP51 was only by a factor of 10. For the antimycotic drugs, the range of IC(50) values for cCYP51 was similar to those of fungicides (0.039-0.30 microM). For the inhibition of hCYP51, IC(50) values split into two classes: the newer drugs fluconazole and itraconazole showed little inhibition (> or = 30 microM) while the older drugs were even more potent than the agricultural fungicides, with miconazole being the most potent (0.057 microM). No correlation was seen between the IC(50) values determined for the two enzymes, indicating that a housekeeping gene can show significant diversity if inhibition is concerned. Our data indicate that fungicide residues in food are unlikely to exert a relevant inhibition of CYP51 in humans whereas systemic use of some antimycotic drugs, e.g. ketoconazole or miconazole, should be carefully considered regarding disturbance of human steroid biosynthesis.
authors.affiliation: Department of Toxicology, University of Würzburg, Versbacher Str 9, Würzburg, Germany. lutz@toxi.uni-wuerzburg.de
authors.forename: Eva R
authors.initials: ER
authors.lastname: Trösken
authors.forename: Magdalena
authors.initials: M
authors.lastname: Adamska
authors.forename: Michael
authors.initials: M
authors.lastname: Arand
authors.forename: Jürg A
authors.initials: JA
authors.lastname: Zarn
authors.forename: Christopher
authors.initials: C
authors.lastname: Patten
authors.forename: Wolfgang
authors.initials: W
authors.lastname: Völkel
authors.forename: Werner K
authors.initials: WK
authors.lastname: Lutz
mesh_terms: D000935:Antifungal Agents / Q000494:pharmacology*; D001393:Azoles / Q000737:chemistry / Q000145:classification / Q000494:pharmacology*; D002176:Candida albicans / Q000201:enzymology*; D065607:Cytochrome P-450 Enzyme Inhibitors; D004305:Dose-Response Relationship, Drug; D004791:Enzyme Inhibitors / Q000145:classification / Q000494:pharmacology*; D005506:Food Contamination; D005656:Fungal Proteins / Q000037:antagonists & inhibitors*; D006801:Humans; D020128:Inhibitory Concentration 50; D010088:Oxidoreductases / Q000037:antagonists & inhibitors*; D010573:Pesticide Residues; D018570:Risk Assessment; D013045:Species Specificity; D058886:Sterol 14-Demethylase; D013329:Structure-Activity Relationship
chemical_list: D000935:Antifungal Agents; D001393:Azoles; C548418:CYP51A1 protein, human; D065607:Cytochrome P-450 Enzyme Inhibitors; D004791:Enzyme Inhibitors; D005656:Fungal Proteins; D010573:Pesticide Residues; D010088:Oxidoreductases; D058886:Sterol 14-Demethylase