Diethylcarbamazine-related antimicrobial activity in Mycobacterium tuberculosis-infected blood

Abstract

The cytochrome P-450 lanosterol 14alpha-demethylase (CYP51A1) of yeasts is involved in an important step in the biosynthesis of ergosterol. Since CYP51A1 is the target of azole antifungal agents, this enzyme is potentially prone to alterations leading to resistance to these agents. Among them, a decrease in the affinity of CYP51A1 for these agents is possible. We showed in a group of Candida albicans isolates from AIDS patients that multidrug efflux transporters were playing an important role in the resistance of C. albicans to azole antifungal agents, but without excluding the involvement of other factors (D. Sanglard, K. Kuchler, F. Ischer, J.-L. Pagani, M. Monod, and J. Bille, Antimicrob. Agents Chemother. 39:2378-2386, 1995). We therefore analyzed in closer detail changes in the affinity of CYP51A1 for azole antifungal agents. A strategy consisting of functional expression in Saccharomyces cerevisiae of the C. albicans CYP51A1 genes of sequential clinical isolates from patients was designed. This selection, which was coupled with a test of susceptibility to the azole derivatives fluconazole, ketoconazole, and itraconazole, enabled the detection of mutations in different cloned CYP51A1 genes, whose products are potentially affected in their affinity for azole derivatives. This selection enabled the detection of five different mutations in the cloned CYP51A1 genes which correlated with the occurrence of azole resistance in clinical C. albicans isolates. These mutations were as follows: replacement of the glycine at position 129 with alanine (G129A), Y132H, S405F, G464S, and R467K. While the S405F mutation was found as a single amino acid substitution in a CYP51A1 gene from an azole-resistant yeast, other mutations were found simultaneously in individual CYP51A1 genes, i.e., R467K with G464S, S405F with Y132H, G129A with G464S, and R467K with G464S and Y132H. Site-directed mutagenesis of a wild-type CYP51A1 gene was performed to estimate the effect of each of these mutations on resistance to azole derivatives. Each single mutation, with the exception of G129A, had a measurable effect on the affinity of the target enzyme for specific azole derivatives. We speculate that these specific mutations could combine with the effect of multidrug efflux transporters in the clinical isolates and contribute to different patterns and stepwise increases in resistance to azole derivatives.

Structural Insights into Binding of the Antifungal Drug Fluconazole to Saccharomyces cerevisiae Lanosterol 14α-Demethylase

“…Mutations which reduce the effectiveness of azole drugs are frequently found in resistant pathogenic fungi. Of the residues within 4 Å of FLC in ScErg11p6ϫHis that have residues equivalent to those found to be mutated in C. albicans Erg11p, only Y132F/H (Y140F/H in ScErg11p) is known to confer resistance to FLC as a single mutation (13,45). The others confer resistance in combination with additional mutations or confer susceptible phenotypes alone (46).…”

“…Fungi have evolved several mechanisms to circumvent the action of azoles, including mutations in the drug-binding site of Erg11p that reduce the binding affinity of triazoles and thus confer resistance (12)(13)(14). The innate resistance of Aspergillus fumigatus to FLC is thought to be due to the presence of two homologues of the drug target, with CYP51B appearing sensitive to fluconazole whereas CYP51A confers significantly reduced susceptibility (15).…”

Distinct Roles of Candida albicans Drug Resistance Transcription Factors TAC1 , MRR1 , and UPC2 in Virulence

“…As for TAC1, GOF mutations in MRR1 are associated with MDR1 upregulation (16). Upregulation of ERG11 can mediate azole resistance, especially when this gene contains mutations modifying azole affinity (11,17,18). ERG11 expression is mainly controlled by the TF UPC2, and GOF mutations in this gene result in ERG11 upregulation (19)(20)(21).…”

Functional characterization of Candida albicans Hos2 histone deacetylase

“…is mediated by up regulation of genes encoding ERG11, a lanosterol demethylase 4– 6 , MDR1 5, 7 and by CDR (Candida drug resistance) efflux pumps 5, 7– 9 . A combination of existing anti-fungals with new classes of drugs that act by different mechanisms will be viable alternatives to the current monotherapy regimen, which contributes to the emergence of drug resistance.…”