Itraconazole preexposure attenuates the efficacy of subsequent amphotericin B therapy in a murine model of acute invasive pulmonary aspergillosis.

Antagonism has been described in vitro and in vivo for azole-polyene combinations against Aspergillus species. Using an established murine model of invasive pulmonary aspergillosis, we evaluated the efficacy of several amphotericin B (AMB) dosages given alone or following preexposure to itraconazole (ITC). Mice were immunosuppressed with cortisone acetate and cyclophosphamide. During immunosuppression, animals were administered either ITC solution (50 mg/kg of body weight) or saline by oral gavage twice daily for 3 days prior to infection. Infection was induced by intranasally inoculating mice with a standardized conidial suspension (1 x 10(8) CFU/ml) of Aspergillus fumigatus strain AF 293. AMB was then administered by daily intraperitoneal injections (0.25, 0.5, 1.0, and 3.0 mg/kg) starting 24 h after inoculation and continuing for a total of 72 h. Drug pharmacokinetics of AMB and ITC in plasma were determined by high-performance liquid chromatography. Four different endpoints were used to examine the efficacy of antifungal therapy: (i) viable counts from harvested lung tissue (in CFU per milliliter), (ii) the whole-lung chitin assay, (iii) mortality at 96 h, and (iv) histopathology of representative lung sections. At AMB doses of >0.5 mg/kg/day, fewer ITC-preexposed mice versus non-ITC-preexposed mice were alive at 96 h (0 to 20 versus 60%, respectively). At all time points, the fungal lung burden was consistently and significantly higher in animals preexposed to ITC, as measured by the CFU counts (P = 0.001) and the chitin assay (P = 0.03). Higher doses of AMB did not overcome this antagonism. ITC preexposure was associated with poorer mycological efficacy and survival in mice treated subsequently with AMB for invasive pulmonary aspergillosis.

Evaluation of polyene-azole antagonism in liquid cultures of Candida albicans using an automated turbidometric method.

BACKGROUND: A computerized machine, SPECTRAmax 340, was used to evaluate the recently reported phenomenon of antagonism of the polyene amphotericin B (AMB) in Candida albicans pre-exposed to the triazole fluconazole (FLZ). METHODS: We investigated growth inhibition by varying concentrations of AMB in seven isolates of C. albicans pre-exposed to FLZ (50 microg/ml) for 18 h. All isolates were obtained on sequential visits from human immunodeficiency virus-infected patients not treated with FLZ. RESULTS: Antagonism of AMB activity was observed in 5, 4, 2 and a single isolate for 0.5, 1, 2 and 3 microg/ml of the antifungal, respectively. In the majority of Candida isolates, antagonism was seen within a concentration range of 0.5-1.0 microg/ml AMB; 1 Candida strain (HK1-Sa) was resistant to 3 microg/ml AMB. Higher concentrations of AMB (>3 microg/ml) killed both the controls and FLZ-pre-exposed Candida cells. No significant differences were observed between the periods of antagonism observed for any of the sequential isolates or for any of the AMB concentrations or in the maxima of the growth curves obtained for all Candida isolates. CONCLUSION: We conclude that the SPECTRAmax system is a useful tool to evaluate in vitro pharmacodynamic interactions between antifungal regimens within a fluid culture system, and provides information that cannot be obtained using traditional plate assay systems.

Effect of salt supplementation on amphotericin B nephrotoxicity.

It has been suggested that salt loading protects against amphotericin B-induced nephrotoxicity. The influence of saline loading on the nephrotoxic response to amphotericin B (50 mg/dose given i.v. over 4 hr 3 X/week for 10 weeks) was assessed in two groups of ten patients each who were diagnosed with mucocutaneous leishmaniasis. Patients were randomized to receive either 1 liter of 0.9% saline or 1 liter of 5% dextrose in water, administered i.v. over one hour in a double-blinded manner, directly prior to amphotericin B administration. Renal function was monitored on a weekly basis two days after the last dose of amphotericin B. Baseline characteristics were similar in both groups except for a slightly higher serum creatinine concentration (Cr) in the saline group (0.8 +/- 0.05 vs. 0.6 +/- 0.04 mg/dl). Baseline sodium (Na) excretion was relatively high (262 +/- 23 mmol/day in the dextrose group and 224 +/- 17 mmol/day in the saline group). None of the patients sustained an increase in Cr to values greater than 1.7 mg/dl. Although mean Cr remained within normal, there was a significant difference between the two groups over the ten week period, with the dextrose group sustaining a significant increase in Cr and the saline group remaining unchanged. Serum potassium (K) levels fell in both groups necessitating oral K supplementation. The saline group required significantly greater amounts of K supplementation to maintain a normal serum K. Amphotericin B caused a rapid reduction in the acidification ability of the kidney in response to an ammonium chloride load. Under these conditions, the saline group had a poorer ability to acidify the urine.(ABSTRACT TRUNCATED AT 250 WORDS)

Limited protection by small unilamellar liposomes against the renal tubular toxicity induced by repeated amphotericin B infusions in rats.

Amphotericin B (AMB), either alone or incorporated into small unilamellar vesicles of pure dipalmitoylphosphatidyl choline (DPPC SUV-AMB), was administered intravenously to male Sprague-Dawley rats once daily for 5 days. Either 1.5 or 3.5 mg of AMB or DPPC SUV-AMB per kg was given, since these concentrations corresponded, respectively, to the lowest nephrotoxic dose and the sublethal dose of AMB in our model. Tubular functions were evaluated daily, and AMB concentrations in plasma, urine, and tissues were measured by high-performance liquid chromatography. AMB at both doses induced tubular toxicity, hyposthenuria being the earliest symptom. DPPC SUV-AMB at 1.5 mg/kg/day was atoxic, but the tubular alterations induced by 3.5 mg of DPPC SUV-AMB per kg were similar to those observed with 3.5 mg of AMB per kg, except that the ability to concentrate urine was partly restored 72 h after the last infusion. Incorporating AMB into DPPC SUV did not influence the pharmacokinetics of the drug. Using this lipidic AMB formulation, we thus observed a beneficial effect toward limiting the renal tubular toxicity of repeated low doses of AMB but, unexpectedly, not that of high doses. These results indicate that tubular renal functions and electrolyte serum values should be closely monitored in patients treated with AMB liposomal formulations, especially high-dose regimens.

Activity of amphotericin B and intraconazole against intraphagocytic Candida albicans.

The activity of amphotericin B and intraconazole against intracellular Candida albicans was determined in vitro using murine resident peritoneal macrophages. Amphotericin B at concentrations of 0.5 and 2 micrograms/ml produced significantly less rapid killing of intracellular than of extracellular Candida albicans as measured in macrophage-free medium. Amphotericin B at concentrations of 0.1 micrograms/ml or itraconazole concentrations of up to 3 micrograms/ml produced only fungistatic or limited fungicidal activity against both intracellular and extracellular organisms. Against intracellular Candida albicans amphotericin B acted by direct antifungal action rather than through stimulation of macrophage function, as demonstrated by the fact that (i) activity persisted when macrophages were successively exposed to amphotericin B, washed and disrupted by sonication, and (ii) no activity was seen when amphotericin B was tested against intracellular amphotericin B-resistant Candida tropicalis or Salmonella typhimurium. Pre-exposure of macrophages to itraconazole (0.4 micrograms/ml) inhibited subsequent killing activity of amphotericin B (2 micrograms/ml) against intracellular susceptible Candida albicans. These experiments validate the conventional methods of susceptibility testing for determining the fungistatic activity of antifungal agents.