Impact of the Order of Initiation of Fluconazole and Amphotericin B in Sequential or Combination Therapy on Killing of Candida albicans In Vitro and in a Rabbit Model of Endocarditis and Pyelonephritis

Abstract

In vitro time-kill studies and a rabbit model of endocarditis and pyelonephritis were used to define the impact that the order of exposure of Candida albicans to fluconazole (FLC) and amphotericin B (AMB), as sequential and combination therapies, had on the susceptibility of C. albicans to AMB and on the outcome. The contribution of FLC-induced resistance to AMB for C. albicans also was assessed. In vitro, AMB monotherapy rapidly killed each of four C. albicans strains; FLC alone was fungistatic. Preincubation of these fungi with FLC for 18 h prior to exposure to AMB decreased their susceptibilities to AMB for 8 to >40 h. Induced resistance to AMB was transient, but the duration of resistance increased with the length of FLC preincubation. Yeast sequentially incubated with FLC followed by AMB plus FLC (FLC3AMB؉FLC) showed fungistatic growth kinetics similar to that of fungi that were exposed to FLC alone. This antagonistic effect persisted for at least 24 h. Simultaneous exposure of C. albicans to AMB and FLC [AMB؉FLC(simult)] demonstrated activity similar to that with AMB alone for AMB concentrations of >1 g/ml; antagonism was seen using an AMB concentration of 0.5 g/ml. The in vitro findings accurately predicted outcomes in our rabbit infection model. In vivo, AMB monotherapy and treatment with AMB for 24 h followed by AMB plus FLC (AMB3AMB؉FLC) rapidly sterilized kidneys and cardiac vegetations. AMB؉FLC(simult) and FLC3AMB treatments were slower in clearing fungi from infected tissues. FLC monotherapy and FLC3AMB؉FLC were both fungistatic and were the least active regimens. No adverse interaction was observed between AMB and FLC for the AMB3FLC regimen. However, FLC3AMB treatment was slower than AMB alone in clearing fungi from tissues. Thus, our in vitro and in vivo studies both demonstrate that preexposure of C. albicans to FLC reduces fungal susceptibility to AMB. The length of FLC preexposure and whether AMB is subsequently used alone or in combination with FLC determine the duration of induced resistance to AMB.With the increased capacity of medical science to prolong the lives of the immunocompromised host, the incidence of systemic Candida albicans infections is rising (5). Yet despite treatment with fluconazole (FLC) or amphotericin B (AMB) monotherapy, the mortality associated with deep-seated candidal infections remains substantial (2,22,23,25,30).In an attempt to improve survival rates, there is much interest in using FLC and AMB in combination. However, the interaction between FLC and AMB remains poorly characterized when these drugs are used concurrently or sequentially. In vitro studies in which FLC and AMB were simultaneously or sequentially introduced to cultures of C. albicans frequently showed this drug combination to be antagonistic (13,19,21,28; P. Banerjee, Q.-F. Liu, A. Louie, M. Shayegani, H. Taber, G. Drusano, and M. Miller, Abstr. 97th Gen. Meet. Am. Soc. Microbiol., abstr. C-252a, p. 164, 1997). Further, using time-kill studies, Vazquez et al. (28) demonstrated that sequential expo...

Combination Treatment of Invasive Fungal Infections

“…However, removal of FLU from the culture medium reversed the AmB inhibition within a very short period (Ͻ6 h). Similar studies were performed recently by Louie et al (135), who used in vitro timekill studies to show that preincubation of C. albicans with FLU for 18 h prior to the addition of AmB resulted in transient AmB antagonism. The duration of this induced AmB resistance was directly related to the duration of FLU preincubation.…”

“…The duration of this induced AmB resistance was directly related to the duration of FLU preincubation. These in vitro findings were also reflected in an in vivo model used by these investigators (described below) (135).…”

“…Consequently, Vazquez et al (229) investigated the interaction between AmB and FLU against Candida cells and demonstrated that preexposure of C. albicans to the azole leads to transient protection against subsequent exposure to AmB (229). In a subsequent study of FLU and AmB interactions, the order of drug initiation in this combination was examined in a two-part study using four Candida isolates (135). The in vivo implications of in vitro antagonism were examined in the first part by using the time-kill method and macrobroth dilutions (incubation for up to 48 h) and FLU preincubation (up to 40 hs).…”

Combination Antifungal Therapy

“…When studied sequentially, the order of administration and duration of exposure appear to be important factors affecting the activity of the polyene-azole combinations. Pretreatment with fluconazole has generally resulted in the reduction of subsequent amphotericin B activity (34,67,105,107,114,153,175,216), but in some series high azole concentrations, long duration of pretreatment, or high levels of inocula were required to produce these effects (67,107,185). Other azoles have also been reported to exert antagonistic effects on amphotericin B activity (31,185,216,217), and preexposure with more-lipophilic azoles such as itraconazole has produced more profoundly negative effects in some experimental systems (184,185,217) but not in others (216).…”

Competitive Binding Inhibition Enzyme-Linked Immunosorbent Assay That Uses the Secreted Aspartyl Proteinase ofCandida albicansas an Antigenic Marker for Diagnosis of Disseminated Candidiasis

“…In contrast, the tissue burden for the spleen and liver was somewhat reduced over time, suggesting that the rabbit model described in this report better reflects acute disseminated candidiasis (found in both oncologic and nononcologic patients [59]) rather than the chronic, hepatosplenic infection most often observed in patients treated for acute leukemia and in bone marrow transplant recipients who have recovered from neutropenia (31,59,62,77). Most animal models of acute disseminated infection, particularly when animals are infected via the intravenous route, demonstrate an increased organ burden for the kidney with disease progression and a reduction in tissue burden for other organs (5,21,37,38,86). This result occurs because the osmotic pressure within the kidney tubules reduces the effectiveness of phagocytic cells to kill and remove C. albicans cells, and fungus balls can grow and obstruct kidney function (86).…”