Methods

Patients and Data collection

The prospective and observational study was conducted on liver dysfunction patients who received voriconazole between February 28, 2018 and December 11, 2018. The inclusion criteria were (1) Age≥15 years; (2) Patients were diagnosed with liver dysfunction, such as liver failure or liver cirrhosis according to the Child-Pugh classifications; (3) Treatment or prevention of invasive fungal infections with voriconazole; (4) patients contributed at least one blood sample. The exclusion criteria were: (1) Patients who were allergic or intolerant to voriconazole; (2) Pregnant or lactating patients; (3) Using potent CYP450 inducer or inhibitor such as rifampicin, isoniazid, phenytoin, carbamazepine during voriconazole treatment, but did not include proton pump inhibitors (PPIs). (4) Patients who lacked the necessary data such as genotype of CYP2C19, renal and liver function index. This study was approved by Ethics Committee of The Second XiangYa Hospital of Central South University (Changsha, China). All of the patients were provided written informed consent before participating in the study.
Information of the following potential covariates was collected and analyzed: age, gender (Gen), body weight (WT), platelet counts (PLT), alanine aminotransferase (ALT), aspartate aminotransferase (AST), total bilirubin (TBIL), direct bilirubin (DBIL), albumin (ALB), creatinine clearance rate (CLcr) which is calculated using the Cockcroft and Gault equation [19], international normalized ratio (INR), CYP2C19 genotype and concomitant medication (PPIs). Liver dysfunction was classified using Child-Pugh scores [20], and Model for End-stage Liver Disease (MELD) scores. The MELD score according to the following formula [21]:\(MELD\ score=0.957\times\log_{e}\left(creatinine,mg/dl\right)+0.378\times\log_{e}\left(bilirubin,mg/dl\right)+1.12\times\log_{e}\left(\text{INR}\right)+6.43\)\(MELD\ score\ =0.957\times\log_{e}\left(creatinine,mg/dl\right)+\log_{e}(bilirubin,\ mg/dl)+1.12\times\log_{e}(INR)\ +6.43\)

Dosing regimen and Specimen collection

Voriconazole dosing was according to the product information, where patients with mild to moderate liver dysfunction (Child-Pugh A and B) received standard loading doses (400 mg twice daily PO or 6 mg/kg twice daily IV) on the first day, but half the standard maintenance dose (100 mg twice daily PO or 2 mg/kg twice daily IV). Due to the limited data on the dosing of voriconazole in patients with severe liver dysfunction (Child-Pugh C), dosing of these patients was based on the clinician’s experience. The subsequent doses for all patients were adjusted according to the measured voriconazole trough concentration (Ctrough) and the patient’s clinical response to voriconazole (effective or ineffective, with or without adverse effects).
Venous blood samples (2 mL) were collected into anticoagulant tubes. Patients were randomly collected 2-3 blood samples without intervention in treatment at 0.5 h, 1 h, 1.5 h, 2 h, 4 h, 6 h, 8 h, 12 h, 24 h after intravenous or oral administration. In addition, blood samples such as Ctrough from TDM were collected from all patients after 24 hours. All voriconazole plasma concentrations were analysed by automatic two-dimensional liquid chromatography (2D-HPLC, Demeter Instrument Co., Ltd., Hunan, China) as previously described [17].

DNA sequencing and CYP2C19 genetic polymorphism

Genomic DNA was extracted using commercially available EZNA® SQ Blood DNA Kit II. Sanger dideoxy DNA sequencing method with ABI3730xl‐full automatic sequencing instrument (ABI Co.) from Boshang Biotechnology Co. Ltd. (Shanghai, China) was used for CYP2C19 genotyping. CYP2C19 phenotypes were classified into five categories: ultrarapid metabolizer (UM, CYP2C19*17/*17), rapid metabolizer (RM, CYP2C19*1/*17), extensive metabolizer (EM, CYP2C19*1/*1), intermediate metabolizer (IM, CYP2C19*1/*2, CYP2C19*1/*3, CYP2C19*2/*17) and poor metabolizer (PM, CYP2C19*2/*2, CYP2C19*2/*3, CYP2C19*3/*3) [22].

Statistical analysis

The Wilcoxon two-sample test or Kruskal–Wallis test was used to compare voriconazole Ctrough. Proportions were compared with the Chi-square test or Fisher’s exact test. Univariate analysis was performed to assess the association between voriconazole Ctrough and adverse events. Receiver operating characteristic (ROC) curves were used to explore the relationship between voriconazole Ctrough and adverse events. Statistical analysis was performed with SPSS version 22.0 (IBM Corporation, Armonk, New York).

Population Pharmacokinetics analysis

The concentration–time data of voriconazole was developed using Phoenix NLME (version 8.0, Pharsight Corporation, USA). The first-order conditional estimation method with the η-ε interaction option (FOCE ELS) was used throughout the model development.
One- and two-compartment structural kinetic models with first-order and Michaelis–Menten elimination were evaluated to describe the pharmacokinetics of voriconazole. Finally, we comprehensively compared the objective function value (OFV), graphical goodness of fit, the evaluation of parameter estimates (including precision) and scientific and physiological plausibility to choose the best base model. The oral absorption rate constant (ka) was fixed to a value of 1.1 h−1 based on the results from a previous study [23].
The inter-individual variability in voriconazole pharmacokinetic parameters was described with an exponential error model. Residual error models for voriconazole were tested as follows: the proportional error model, the additive error model and combined error model, including proportional plus additive error model.
Potential demographic and biochemical covariates were evaluated by visual inspection of covariates possible relationships with pharmacokinetic parameters included in the model. For continuous covariates, a linear, piece-wise, exponential, and power parameter-covariate relations were tested. Categorical covariates were linearly included. Then, a covariate model in a stepwise forward-inclusion and backward-elimination procedure were carried out. A covariate was considered to be significant when inclusion of the covariate resulted in a decrease in the objective function value (OFV) of greater than 6.64 (p< 0.01) and elimination of the covariate resulted in an increase in the OFV of greater than 10.83 (p< 0.001).
Goodness-of-fit (GOF) plots were used to evaluate the adequacy of fitting. The bootstrap method was used to assess the robustness and stability of the final model. 1000 resamples from the original data were performed. All of the model parameters were estimated, and their median and 2.5 and 97.5 percentiles were calculated. That was stable if the 95% CI for the parameter estimates derived from the 1000 bootstrap runs encompassed the original final parameter estimate.

Monte Carlo simulation

1000 individuals receiving the dosing regimens including loading doses of 200, 300 and 400 mg every 12 hours (q12h), and maintenance doses of 50, 100, 150 and 200 mg once daily (qd) or q12h orally or intravenously were simulated by the final model. The dosing regimens were simulated for 30-days and stratified by TBIL (TBIL-1: TBIL < 51 μmol/L; TBIL-2: 51 μmol/L ≤ TBIL < 171 μmol/L; TBIL-3: TBIL ≥ 171 μmol/L) were performed. The voriconazole Ctrough range of 0.5-5.0 mg/L [24] was used as the target range. The probability of target attainment (PTA) for the Ctrough range was examined for each of the different dosing regimens.