Mechanisms of resistance to azole antifungal agents in Candida albicans isolates from AIDS patients involve specific multidrug transporters

Abstract

Azole antifungal agents, and especially fluconazole, have been used widely to treat oropharyngeal candidiasis in patients with AIDS. An increasing number of cases of clinical resistance against fluconazole, often correlating with in vitro resistance, have been reported. To investigate the mechanisms of resistance toward azole antifungal agents at the molecular level in clinical C. albicans isolates, we focused on resistance mechanisms related to the cellular target of azoles, i.e., cytochrome P450 14DM (14DM) and those regulating the transport or accumulation of fluconazole. The analysis of sequential isogenic C. albicans isolates with increasing levels of resistance to fluconazole from five AIDS patients showed that overexpression of the gene encoding 14DM either by gene amplification or by gene deregulation was not the major cause of resistance among these clinical isolates. We found, however, that fluconazole-resistant C. albicans isolates failed to accumulate 3 H-labelled fluconazole. This phenomenon was reversed in resistant cells by inhibiting the cellular energy supply with azide, suggesting that resistance could be mediated by energy-requiring efflux pumps such as those described as ATP-binding cassette (ABC) multidrug transporters. In fact, some but not all fluconazole-resistant clinical C. albicans isolates exhibited up to a 10-fold relative increase in mRNA levels for a recently cloned ABC transporter gene called CDR1. In an azole-resistant C. albicans isolate not overexpressing CDR1, the gene for another efflux pump named BEN r was massively overexpressed. This gene was cloned from C. albicans for conferring benomyl resistance in Saccharomyces cerevisiae. Therefore, at least the overexpression or the deregulation of these two genes potentially mediates resistance to azoles in C. albicans clinical isolates from AIDS patients with oropharyngeal candidiasis. Involvement of ABC transporters in azole resistance was further evidenced with S. cerevisiae mutants lacking specific multidrug transporters which were rendered hypersusceptible to azole derivatives including fluconazole, itraconazole, and ketoconazole.

CaIPF14030 negatively modulates intracellular ATP levels during the development of azole resistance in Candida albicans

“…As shown in Figure 1B, the CaIPF14030 gene was overexpressed in the DSF7, DSF14, DSF21, and DSF28 strains. These results are similar to those found for other known resistance-related genes such as ERG11, CDR1, and CaMDR1 [4,24] . These data suggest that CaIPF14030 is involved in the development of azole resistance in C albicans.…”

“…However, there is a rapid acquisition of resistance to azoles in C albicans. This is due to a limited number of multidrug resistance related genes, which include ERG11, CDR1, CDR2, and CaMDR1 [3][4][5][6] . Recently, DNA microarray and proteomic analyses have revealed many differentially regulated genes in either laboratory [7,8] or clinical [9][10][11][12] azole-resistant C albicans isolates.…”

A novel polyamide SL-A92 as a potential fungal resistance blocker: synthesis and bioactivities in Candida albicans

“…The reduced intracellular accumulation of drugs is a common mechanism of resistance, which is correlated with the increased expression of the genes CDR1 and CDR2 (members of the ABC efflux pump family) and of the gene CaMDR1 (a member of the MFS efflux pump family) [37][38][39] . Four kinds of CDR genes in C albicans have been cloned [13,[40][41][42] , but only over-expression of CDR1 and CDR2 resulted in azole resistance [43] .…”

Sphingolipids and membrane biology as determined from genetic models

“…In these mutant backgrounds, the authors also analyzed the surface localization of Cdr1p, a major ABC drug efflux protein that functions in the drug resistance mechanism [102,103]. In the mutant C. albicans, Cdr1p shows impaired surface localization and defective efflux function.…”