Summary:Voriconazole, a new antifungal agent, is increasingly being used after HSCT. The hepatic cytochrome P450 isoenzyme 2C19 plays a significant role in voriconazole metabolism. As CYP2C19 exhibits significant genetic polymorphism, some patients metabolize voriconazole poorly resulting in increased plasma drug levels. The clinical significance of this is unknown, and the utility of monitoring voriconazole levels is unclear. Steady-state trough plasma voriconazole levels were obtained in 25 allogeneic HSCT recipients using an HPLC assay. Patients had drug levels checked once (n ¼ 13), twice (n ¼ 10), or X3 times (n ¼ 2) 5-18 days (median 10) after starting voriconazole or dose modification. The 41 voriconazole levels were 0.2-6.8 lg/ml (median 1.6); 6 (15%) were o0.5 (possibly below the in vitro MIC 90 for Aspergillus spp.). Voriconazole concentrations correlated with aspartate aminotranferase (AST) (r ¼ 0.5; P ¼ 0.0009) and alkaline phosphatase (r ¼ 0.34; P ¼ 0.03), but not with creatinine, bilirubin and alanine aminotransferase (ALT). Since liver dysfunction is common after HSCT, it was not possible to determine if elevated AST and alkaline phosphatase levels were the cause or the consequence of higher voriconazole levels. We conclude that trough voriconazole levels vary considerably between patients, and suggest monitoring levels in patients receiving voriconazole for confirmed fungal infections, and in those with elevated AST or alkaline phosphatase levels. Bone Marrow Transplantation (2005) 35, 509-513.

BACKGROUND. Low voriconazole levels have been associated with a higher failure rate in patients with confirmed fungal infections. METHODS. Steady‐state plasma trough voriconazole levels were measured after at least 5 days of therapy in 87 patients with hematologic malignancies on 201 separate occasions (1–5 levels per patient; median, 2). Most patients (90%) had undergone allogeneic hematopoietic stem cell transplantation. The daily voriconazole dose, administered in 2 divided doses, was 200 mg (n = 4), 400 mg (n = 151), 500 mg (n = 20), 600 mg (n = 18), and 800 mg (n = 8); corresponding to 2.0–16.3 (median, 5.4) mg/kg. Plasma voriconazole levels were 0–12.5 μg/mL (median, 1.2). Voriconazole was undetectable (<0.2 μg/mL) in 15%. RESULTS. The correlation between dose and levels was weak (r = 0.14; P = .045). The median absolute daily drug dose (400 mg) was identical in groups of patients with levels of 0, 0.2 to 0.5, >0.5 to 2.0, >2.0 to 5.0, and >5.0. Whereas the daily drug dose in mg/kg was significantly higher when the levels were >5.0 μg/mL, there was no consistent relation between dose and level below that threshold. In adult patients getting standard doses of voriconazole orally, the drug levels are highly variable. Based on limited available data, between a quarter and two‐thirds of these levels could potentially be associated with a lower likelihood of response or a higher likelihood of failure. CONCLUSIONS. Future voriconazole studies should incorporate prospective therapeutic drug monitoring and consideration should be given to checking levels in patients receiving the drug for confirmed, life‐threatening fungal infections. Cancer 2007. © 2007 American Cancer Society.

An analytical method for the determination of voriconazole (UK-109,496; Pfizer) in plasma was developed and validated. The method utilizes solid-phase extraction technology and high-performance liquid chromatography. The lower limit of quantitation is 0.2 g/ml, and the range of linearity tested was 0.2 to 10 g/ml.Voriconazole (VRC; 496 [C 16 H 14 N 5 OF 3 ]; Pfizer Pharmaceuticals) is a novel broad-spectrum triazole antifungal that is used in the treatment of a wide range of opportunistic fungal infections, including aspergillosis (4). VRC is marketed in formulations for administration both orally (tablet) and intravenously. Previously described assays for VRC include a bioassay procedure and two different high-performance liquid chromatography (HPLC) methods. The former lacks the required sensitivity, and the latter HPLC methods either lacked the necessary sensitivity (4) or were lengthy and technically difficult (6). This assay includes the use of an internal standard (UK-115,794) and sample preparation by solid-phase extraction (SPE). Validation guidelines published by Shah et al. (5) were used to determine the method's accuracy, precision, reproducibility, and specificity.Pfizer Research and Development, Sandwich, United Kingdom, provided VRC and internal-standard powders. The HPLC system (Beckman Coulter, Fullerton, Calif.) consisted of a 168 diode array detector, a 126 solvent pump, a 508 autosampler, an IBM NT-based computer work station, and 32 Karat software.Stock and working VRC standards (1,000, 100, and 10 g/ ml, respectively) and stock and working internal standards (1,000 and 100 g/ml, respectively) were all made in methanol (MeOH) and stored at Ϫ20°C. Calibration standards (0.2, 0.5, 1, 2, 4, 6, 8, and 10 g/ml) were prepared in pooled plasma from the VRC working standards on the day of the analysis.Plasma controls (0.2, 0.5, 4, and 8 g/ml) were made in batches and frozen for analyses over a period of time from an independent weighing of VRC powder. Five hundred microliters of each blank, standard, or control was pipetted into an appropriately labeled tube. Ten microliters of a 100-g/ml internal standard was added to each tube except the "blankblank" tube. All samples were buffered with 700 l of 0.2 M borate buffer (pH 9.0). Samples were extracted by SPE with C 18 , 100 mg, 1-ml Bond Elut columns (Varian, Inc., Harbor City, Calif.). The columns were conditioned with separate washings in the following order: 1 ml of MeOH, 1 ml of H 2 O, and 1 ml of 0.2 M borate buffer (pH 9.0). The buffered plasma samples were added to each respective column. After the columns completely drained, they were washed with separate and independent washings of the following reagents: 1 ml of 0.2 M borate buffer, followed by 1 ml of MeOH-H 2 O (50:50, vol/vol). Inside the vacuum manifold glass chamber, microcentrifuge tubes were positioned for collection of each eluted sample from its respective SPE column. One milliliter of the eluent, MeOH-glacial acetic acid (99:1, vol/vol), was added to each column. The collected elua...

A new selective high-performance liquid chromatography (HPLC) method with UV detection for the determination of the investigational triazole voriconazole in human plasma by using acetonitrile precipitation followed by reverse-phase HPLC on a C 18 column was compared with a simple agar well diffusion bioassay method with Candida kefyr ATCC 46764 as the assay organism. Pooled plasma was used to prepare standard and control samples for both methods. The results of analyses with spiked serum samples (run as unknowns) were concordant by the bioassay and HPLC methods, with expected values being obtained. HPLC demonstrated an improved precision (3.47 versus 12.12%) and accuracy (0.81 versus 1.28%) compared to those of the bioassay method. The range of linearity obtained by both methods (from 0.2 to 10 g/ml for HPLC and from 0.25 to 20 g/ml for the bioassay) includes the range of concentrations of voriconazole (from 1.2 to 4.7 g/ml) which are considered clinically relevant. Although either methodology could be used for the monitoring of patient therapy, the smaller variability observed with HPLC compared to that observed with the bioassay favors the use of HPLC for pharmacokinetic studies.

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