Antagonistic interactions between azoles and amphotericin B with yeasts depend on azole lipophilia for special test conditions in vitro

Abstract

The interactions of the azole antifungal agents fluconazole, itraconazole, ketoconazole, or miconazole with amphotericin B (AmB) in their effect on Candida albicans were investigated. These four azoles antagonized the fungistatic activity of AmB at sub-MICs if both substances acted simultaneously. This coincubation test was primarily developed to observe the azole-mediated demethylase inhibition quantitatively by bioassay. Interestingly, the occurrence of azole-AmB antagonism depended on azole lipophilia if specially selected test conditions were applied. By a consecutive incubation regimen, preincubation at high azole concentrations (1 to 50 g/ml) and then subsequent incubation with AmB (1 g/ml), only preincubation with the three lipophilic azoles decreased the fungicidal activity of AmB but not that of FCZ. It was shown that yeasts absorb only lipophilic azoles to a remarkable extent. This fact might be responsible for the absence of antagonism of FCZ to AmB when yeasts were incubated consecutively. It can be concluded with caution that consecutive treatment of candidiasis with FCZ and AmB does not necessarily result in a clinically relevant antagonism.Alterations of the effects of amphotericin B (AmB) on pathogenic fungi by azoles were observed repeatedly in vivo as well as in vitro. The majority of reports mention antagonisms (2-4, 6, 7, 11, 12, 16, 17), but indifference (1, 10, 18) and synergism (10, 14, 18) have also been described. The results were influenced by a variety of factors, especially experimental conditions as well as the particular fungal strains or species. The fact that the results depend on the properties of antifungal agents has been described sporadically (12). At first, a postexposure antagonism of azoles to AmB was assumed. In experiments dealing with this question, it was found that lipophilic azoles are able to influence the fungicidal activity of subsequently applied AmB on yeasts but not that of fluconazole (FCZ). To detect this behavior of lipophilic azoles, special test conditions (unusually high drug concentrations as well as a high density of organisms) had to be selected. Decreasing both inoculum size and antifungal concentration resulted in a failure to detect an influence of azoles on the fungicidal activity of AmB. For this aim of the investigation, the test conditions had to be very different from those that work with drug concentrations similar to those found in vivo, that is, sub-MICs, to observe postantifungal effects (9) with possible clinical significance.Additional tests that were performed in order to explain the dependence of the long-lasting effects of azoles on their lipophilicities also required large quantities of test organisms.The results presented in this report focus on a further aspect of the azole-AmB antagonism. The antagonism between azoles and AmB has been explained by the inhibition of ergosterol synthesis and then a disappearance of ergosterol from the cell membrane (12,13,17,19). The affinity of AmB for ergosterol precursors is not as high as that fo...

Yeasts as a model for assessing the toxicity of the fungicides Penconazol, Cymoxanil and Dichlofluanid

“…These results are in agreement with the described activity of penconazol as an inhibitor of ergosterol biosynthesis in fungi (K oller, 1992). In fact, cells may accumulate ergosterol reserves in the membranes, and the azole-induced depletion of these reserves may take some hours (Scheven and Schwegler, 1995). This would allow growth to proceed normally for some time, after which it would arrest.…”

Combination Antifungal Therapy

“…Antagonism among antifungal agents might occur in one of several ways. (i) Direct antifungal action at the same site results in decreased ability of other agents to exert their competitive effects on that site or at an altered target, as proposed for the azoles and amphotericin B (31,49,182,183,185,186,203). Azoles block the synthesis of ergosterol in the fungal cell membrane and may thus render amphotericin B inactive, since this agent exerts its activity by binding to ergosterol in the cell membrane.…”

“…(ii) Adsorption to the cell surface by one agent inhibits binding of another antifungal agent to its target site of activity. This mechanism is proposed for lipophilic azoles (such as itraconazole and ketoconazole) which may adsorb to the fungal cell surface and inhibit binding of amphotericin B to fungal cell membrane sterols (183,185). (iii) Modification of a target upon exposure to an antifungal agent occurs that renders the pathogen less susceptible to the effects of other antifungals.…”

“…In contrast, antagonism was reported when low concentrations of each agent were used against C. parapsilosis and indifference was observed when higher concentrations of both ketoconazole and flucytosine were employed (19). (107,122,154,185,186,198,214). Most of these studies have been performed using C. albicans isolates, but results with the non-albicans Candida spp.…”

Stable Phenotypic Resistance of Candida Species to Amphotericin B Conferred by Preexposure to Subinhibitory Levels of Azoles.

“…Moreover, several investigators have found synergistic interactions by simultaneous exposure of C. albicans to azoles and AmB (13,30). One group, on the other hand, described antagonisms with preexposures of Candida to the more lipophilic azoles, such as itraconazole, but not to fluconazole (31,32).…”