Sensitive bioassay for ketoconazole in serum and cerebrospinal fluid

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...

Section: Methodsmentioning

confidence: 99%

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

Louie

Kaw

Banerjee

et al. 2001

“…A bioassay was used to determine azole levels in the serum, cerebrospinal fluid (CSF), aqueous humor, and vitreous body of each animal. The assay used the agar-well diffusion method of Bennett et al (4), modified by the method of Jorgensen et al for assaying azole compounds (25). The standard concentrations were dissolved in serum or normal saline.…”

Section: Methodsmentioning

confidence: 99%

Penetration of new azole compounds into the eye and efficacy in experimental Candida endophthalmitis

Savani¹,

Perfect²,

Cobo³

et al. 1987

198

We studied the penetration of three azole compounds, ketoconazole, itraconazole, and fluconazole, into the ocular tissues and fluids of rabbits in the presence and absence of ocular inflammation. Drug concentrations were compared with those found in serum and cerebrospinal fluid. The rank order of penetration into eye tissue was fluconazole > ketoconazole > itraconazole. Fluconazole penetrated freely into both inflamed and uninflamed eyes. The presence of inflammation improved penetration of all three compounds into ocular fluids and tissues. Penetration of these azoles into the anterior chamber of uninflamed eyes and into the cerebrospinal fluid was similar. All three azole compounds reduced the number of yeasts found in the eye in hematogenous Candida albicans endophthalmitis in rabbits when therapy was initiated within 24 h of inoculation. However, only ketoconazole significantly reduced yeast counts in the eye when therapy was postponed for 7 days.The incidence of ocular fungal infections has increased (10, 11, 26) due to a number of factors: increased prevalence of immunosuppression associated with organ transplantation and malignancies; prolonged recovery from complex surgical procedures; and increasing use of antibiotics, immunosuppressive agents, intravenous catheters, and hyperalimentation fluids. Endogenous Candida endophthalmitis (ECE) also occurs in heroin addicts

“…Blood samples were decanted, and serum and CSF were stored at -70°C. Drug concentrations in the serum and CSF were measured by a modification of the well diffusion microbiological assay described by Jorgensen et al (9). Several modifications aimed at improving the sensitivity and reproducibility of this assay were examined: varying the inoculum, the agar well depth, and the temperature and duration of incubation.…”

mentioning

confidence: 99%

Pharmacokinetics of fluconazole in cerebrospinal fluid and serum of rabbits: validation of an animal model used to measure drug concentrations in cerebrospinal fluid

Madu

Cioffe

Mian

et al. 1994