MECHANISMS OF RESISTANCE TO AZOLE ANTIFUNGAL AGENTS IN CANDIDA-ALBICANS ISOLATES FROM AIDS PATIENTS INVOLVE SPECIFIC MULTIDRUG TRANSPORTERS

Azole antifungal agents, and especially fluconazole, have been used wi dely to treat oropharyngeal candidiasis in patients with AIDS, An incr easing number of cases of clinical resistance against fluconazole, oft en correlating with in vitro resistance, have been reported. To invest igate the mechanisms of resistance toward azole antifungal agents at t he molecular level in clinical C. albicans isolates, we focused on res istance mechanisms related to the cellular target of azoles, i.e., cyt ochrome P450(14DM) (14DM) and those regulating the transport or accumu lation of fluconazole. The analysis of sequential isogenic C. albicans isolates with increasing levels of resistance to fluconazole from fiv e AIDS patients showed that overexpression of the gene encoding 14DM I either by gene amplification or by gene deregulation was not the majo r cause of resistance among these clinical isolates. We found, however , that fluconazole-resistant C. albicans isolates failed to accumulate H-3-labelled fluconazole. This phenomenon was reversed in resistant c ells by inhibiting the cellular energy supply with azide, suggesting t hat resistance could be mediated by energy-requiring efflux pumps such as those described as ATP-binding cassette (ABC) multidrug transporte rs, In fact, some but not all fluconazole-resistant clinical C. albica ns isolates exhibited up to a 10-fold relative increase in mRNA levels for a recently cloned ABC transporter gene called CDR1. In an azole-r esistant C. albicans isolate not overexpressing CDR1, the gene for ano ther efflux pump named BEN(r) was massively overexpressed. This gene w as cloned from C. albicans for conferring benomyl resistance in Saccha romyces cerevisiae. Therefore, at least the overexpression or the dere gulation of these two genes potentially mediates resistance to azoles in C. albicans,ls clinical isolates from AIDS patients with oropharyng eal candidiasis. Involvement of ABC transporters in azole resistance w as further evidenced with S. cerevisiae mutants lacking specific multi drug transporters which were rendered hypersnsceptible to azole deriva tives including fluconazole, itraconazole, and ketoconazole.