Correspondence of In Vitro and In Vivo Fluconazole Dose-Response Curves for Cryptococcus neoformans

Abstract

We conducted in vitro experiments to evaluate the susceptibility of a clinical isolate of Cryptococcus neoformans to a wide range of concentrations of fluconazole. In vitro susceptibility was tested using broth macrodilution methods modified to provide a numeric count of viable organisms. The association between the quantitative in vitro response and fluconazole drug concentrations was estimated using local nonparametric regression. Regression analysis was used to assess the correspondence between the in vitro fluconazole concentration-response curve and the murine dose-response curve observed in our previously reported murine model. The regression model was then used to predict the murine response. There was a strong correspondence between in vitro measures of response to fluconazole alone and the previously reported biologic effects seen in the mouse. In vitro antifungal drug susceptibility testing can reliably predict the murine response to fluconazole.The goal of in vitro susceptibility testing is to predict the clinical outcome. Having an in vitro measure that can reliably predict the response following 2 weeks of treatment would permit physicians to select the drug(s) with the greatest activity, extend intensive treatments past 2 weeks, and/or use combination drug therapy. During the last 20 years, considerable effort has been expended to establish a reproducible standard for in vitro testing of yeast susceptibility to antifungal drugs. This standard has been most useful for testing Candida species causing oral thrush (14,18,20). Limited data suggest this method might also be useful for Cryptococcus neoformans; however, no interpretive guidelines or breakpoints have been identified (16,19). The usual approach to identifying breakpoints has been to measure outcome as a categorical response (e.g., sterile cerebrospinal fluid [CSF] versus nonsterile CSF) (1,11,24). This approach forces the search for a breakpoint into a search for a drug concentration that predicts an outcome measure that takes on just two values, "sterile CSF" versus "nonsterile CSF." This search is greatly confounded by the wide range in the severity of meningitis that occurs in clinical practice (21, 24).In our previously published murine model, we used a clinical isolate of C. neoformans to evaluate the effects of fluconazole (12). The response was the number of CFU per gram of brain recovered at day 16. By using this quantitative response and by testing fluconazole over a wide range of doses, we could use regression methods to estimate the dose-response curve for fluconazole. In the present paper, we report the in vitro susceptibility of this same isolate to fluconazole tested over a wide range of concentrations. MATERIALS AND METHODS Test isolate. The Cryptococcus neoformans var. neoformans isolate (USC 1597)was obtained from a patient with AIDS-associated cryptococcal meningitis that responded promptly to treatment with fluconazole plus flucytosine. The isolate was stored in 20% skim milk at Ϫ70°C and subcultured on Sabouraud's dextros...

Pharmacokinetics and Pharmacodynamics of Fluconazole for Cryptococcal Meningoencephalitis: Implications for Antifungal Therapy andIn VitroSusceptibility Breakpoints

“…Experimental studies in mice and retrospective clinical studies suggest that the MIC (or other in vitro measures of antifungal potency-see, for example, Bauer et al [24]) is an important prognostic factor for determining the ultimate clinical outcome (22,(25)(26)(27)(28). For example, the probability of clinical failure in pa- mg/kg/day of fluconazole orally.…”

Results Obtained with Various Antifungal Susceptibility Testing Methods Do Not Predict Early Clinical Outcome in Patients with Cryptococcosis

“…Nevertheless, no relationship between in vitro data and early outcome was found for any of the three drugs. For fluconazole, a recent experimental study showed a correspondence of in vitro and in vivo dose-response curves for one isolate of C. neoformans (13). Nevertheless, it has also been shown in an experimental model of disseminated cryptococcosis that factors other than the MIC, such as the size of the fungal burden in the brain, influenced the in vivo efficacy of fluconazole (14).…”

In Vitro Susceptibility of the Yeast Pathogen Cryptococcus to Fluconazole and Other Azoles Varies with Molecular Genotype

“…Consistent with this, in the current study, Canadian isolates had significantly lower MICs to FLC than isolates from Australia. Most studies evaluating the correlation between in vitro susceptibility and the in vivo response to antifungal drugs have demonstrated that susceptibility testing is a reliable predictor of clinical response (22,47) and that the clinical outcome may be poorer when higher concentrations of FLC are required (MIC Ն 16 mg/liter) (1).…”