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Physiologically based pharmacokinetic evaluation of cefuroxime in perioperative antibiotic prophylaxis

Aims Adequate plasma concentrations of antibiotics during surgery are essential for the prevention of surgical site infections. We examined the pharmacokinetics of 1.5 g cefuroxime administered during induction of anaesthesia with follow‐up doses every 2.5 hours until the end of surgery. We built a physiologically based pharmacokinetic model with the aim to ensure adequate antibiotic plasma concentrations in a heterogeneous population. Methods A physiologically based pharmacokinetic model (PK‐Sim®/MoBi®) was developed to investigate unbound plasma concentrations of cefuroxime. Blood samples from 25 thoracic surgical patients were analysed with high‐performance liquid chromatography. To evaluate optimized dosing regimens, physiologically based pharmacokinetic model simulations were conducted. Results Dosing simulations revealed that a standard dosing regimen of 1.5 g every 2.5 hours reached the pharmacokinetic/pharmacodynamic target for Staphylococcus aureus. However, for Escherichia coli, >50% of the study participants did not reach predefined targets. Effectiveness of cefuroxime against E. coli can be improved by administering a 1.5 g bolus immediately followed by a continuous infusion of 3 g cefuroxime over 3 hours. Conclusion The use of cefuroxime for perioperative antibiotic prophylaxis to prevent staphylococcal surgical site infections appears to be effective with standard dosing of 1.5 g preoperatively and follow‐up doses every 2.5 hours. In contrast, if E. coli is relevant in surgeries, this dosing regimen appears insufficient. With our derived dose recommendations, we provide a solution for this issue.


| INTRODUCTION
Surgical site infections (SSI) are among the most common hospitalacquired infections, causing increased patient morbidity and mortality, as well as increased cost for healthcare systems. 1 Therefore, the prevention of SSIs is of paramount importance in the management of surgical patients. Contemporary strategies for the reduction of SSI-rates typically combine several different approaches into prevention bundles, e.g. aspects of skin preparation, disinfection, perioperative homeostasis and anti-infectives. [2][3][4][5] The perioperative application of parenteral antibiotics is a cornerstone in the reduction of postoperative infections and the efficacy of perioperative antibiotic prophylaxis (PAP) is well established in a wide range of surgical procedures. Various guidelines provide recommendations on the application of PAP. 6 Besides the choice of antibiotic to cover skin flora and the flora of the operative field, several pharmacological aspects must be considered to ensure adequacy of PAP. Typically, the antibiotic should be infused within approximately 60 minutes before skin incision, with repeated doses given after approximately 2 half-lives of the respective drug, if the surgery continues. [6][7][8] This practice recommendation reflects the tenet that concentrations of antibiotic in the operative field must be high enough to ensure eradication of pathogens that enter tissues because of the incision. As β-lactam antibiotics are the mainstay of PAP, the relevant pharmacokinetic (PK)/pharmacodynamic (PD) parameter is the fraction of time of the dosing interval (ƒT) that the free unbound antibiotic concentration remains higher than the minimal inhibitory concentration (MIC) of bacteria (ƒT > MIC) at the focus of infection. 9 In the absence of a widespread availability of therapeutic drug monitoring of antibiotic concentrations in plasma or tissues, dosing strategies of antibiotic therapies and prophylaxis are empirical, assuming efficacy for a wide range of different patients with standard doses.
Cefuroxime is a second-generation cephalosporin with widespread use in PAP. The half-life of cefuroxime is approximately 1.17 hours (70 min) in patients with normal renal function (glomerular filtration rate [GFR] 90-130 mL/min). 10 After initial doses of 1.5 g preoperatively, guidelines recommend a follow-up dose after 3-4 hours if the surgery continues beyond this period. 6,9 This recommendation is somewhat peculiar, as 2 half-lives of cefuroxime would rather mean a repetition after 2.5 hours and the span of 3-4 hours appears arbitrary. In addition, a considerable interindividual variation in the disposition of cefuroxime has been described. 10 In unfavourable circumstances (e.g. augmented renal clearance, increased blood loss), a reduction in cefuroxime concentration below the MIC of relevant pathogens might result in insufficient protection against SSI.
For cefuroxime, no relevant active distribution (e.g. P-glycoprotein transporter) or metabolism (e.g. cytochrome P-system) is known. A single dose of cefuroxime is excreted unchanged in the urine within 24 hours. 11,12 Superposition of literature concentration-time (CT) curves and the observed proportionality between area under the CT curve (AUC) and dosage 10 suggest linearity in the renal elimination.
Application of probenecid as co-medication causes a significant decrease of cefuroxime clearance. [13][14][15] Cephalosporins have similar affinities for OAT-1 and -3 while the role of OAT-4, P-glycoprotein, MRP2/4 and NPT1 for renal absorption and excretion is at least questionable. 16,17 To gain insight into PK properties of cefuroxime used as PAP, we collected perioperative blood samples in patients undergoing thoracic surgical procedures. Cefuroxime concentrations were measured and used for building a physiologically based PK (PBPK) model with the aim to ensure adequate antibiotic protection in a heterogeneous population.

| Patient population
This study was conducted prospectively at the University Hospital Muenster after approval of the study protocol by the local ethics committee (Study Code 02-AnIT-12) and was conducted in accordance with the principles of the Declaration of Helsinki and Good Clinical Practice. Written informed consent was obtained from 25 competent adult patients scheduled for routine thoracic surgery with intercostal thoracotomy. Exclusion criteria were an allergy to cephalosporine, end-stage renal failure and preoperative application of any antibiotic within 7 days. Patients were recruited on the day before surgery.
Age, height, weight and sex were recorded during the preoperative visit. Laboratory measurements of creatinine, urea, albumin and protein were performed on the day of surgery from blood samples drawn at incision.

What is already known about this subject
• Cefuroxime is 1 of the standard drugs used in the field of perioperative antibiotic prophylaxis.
• Current guidelines recommend to administer cefuroxime 1.5 g intravenously before the start of surgery. Followup doses are recommended after approximately 2 halflives.

What this study adds
• During surgery, a timely follow-up dose of 1.5 g cefuroxime after 2.5 hours is warranted to reach minimal inhibitory concentration targets of common pathogens of surgical site infection (e.g. Staphylococcus aureus).
• In case of pathogens with higher minimal inhibitory concentrations than S. aureus (e.g. Escherichia coli), a change in the dosing strategy with a prolonged infusion of cefuroxime should be considered.
• We provide simulated dosing regimens which should enable physicians to individualize perioperative antibiotic prophylaxis.

| Administration of cefuroxime
During intravenous induction of anaesthesia (followed by inhalational maintenance), a dose of 1.5 g cefuroxime (Fresenius Kabi, Germany) was administered intravenously over 10-15 minutes. A follow-up dose of 1.5 g cefuroxime was given every 2.5 hours (150 min) until wound closure (standard dosing regimen at the University Hospital Muenster). No postoperative applications of antibiotic were prescribed in accordance with local standards.

| Blood sampling for cefuroxime measurement
Blood samples were exclusively drawn from an arterial line into ethylenediaminetetraacetic acid-containing tubes (S-Monovette 2.7 mL K3E, Sarstedt, Nuembrecht, Germany) at the following time points: 10 minutes after the initial dose, at incision, shortly before each successive dose of cefuroxime, 10 minutes after every repetition dose and at skin closure. These acquisition points represent peak and trough concentrations of cefuroxime with samples at incision and wound closure providing additional information between these instances. In several patients, supplementary blood samples were collected during the operation. Cefuroxime concentrations of these extra blood samples were included into the PK analyses.

| Cefuroxime assay
For sample preparation the blood was centrifuged (10 min, 2500 g) and 200 μL plasma were transferred; 4 μL internal standard solution (cefazolin 1 mg/mL in water) and 500 μL acetonitrile were added and vortex-mixed (15 s) followed by centrifugation at 12 000 g for  www.mathworks.de/products/matlab/). A more detailed insight into software PK-Sim ® is given by Eissing et al. 18 and Willmann et al. 19,20 The software R (version 3.4.0, R Foundation for Statistical Computing, Vienna, Austria; http://www.r-project.org) was used for PK-analyses, statistical analyses and graphics creation. The software SigmaPlot ® version 12.5 (© 2011 Systat Software, Inc) for 2 ANOVA testing.

| Model building-evaluation
Each step in the model building process was evaluated by comparing simulation results with observed in vivo PK data taken from the literature or measured in the study, considering the European Medicines Agency guideline on the qualification and reporting of PBPK modelling and simulation. 21 For each step in the workflow, the model predictions were visually inspected against the observed data using visual predictive checks, goodness of fit (GOF) and relative-residuals-vs-observedconcentrations plots. 22 Percentage error (PE) and absolute PE (APE) were calculated for every concentration point according to Equations (1 and 2): where C pred is the PBPK-simulated plasma concentration of cefuroxime and C obs is the actual concentration of cefuroxime. Additionally, mean PE (MPE) to quantify the bias and mean APE (MAPE) were calculated to quantify the precision. 23 PK parameters such as AUC and maximum plasma concentration (C max ) were also used for model evaluation by comparison with published AUC values extrapolated to infinity (AUC inf ) or calculated to the last simulated time point in the respective study. The relative changes in AUC and C max were calculated on the basis of Equation 1.

| Model building-healthy adults
For model building we used a generally accepted procedure. 24 (Table S1). As GFR, either GFR from the literature or estimated GFR (eGFR) values calculated by CKD-EPI 25 were used. If no GFR or serum creatinine values were available, we assumed 120 mL/min/1.73m 2 for healthy adults. In both cases we considered body surface area, using the equation described by Dubois and Dubois, 26 to correct the standardized eGFR.
The standard GFR of the virtual twin (sGFR v-twin ) can be calculated using PK-Sim ® specific values for the GFR (GFR specific ) and the kidney volume of the virtual twin (V k-v-twin ) Equation (3): The sGFR v-twin calculated by PK-Sim ® was corrected using the fraction of the eGFR (GFR fraction ) and sGFR v-twin to achieve the same passive renal elimination in silico, as in the literature or later in our study patients Equation (4): The remaining tubular secretion via OAT-transporter (integrated as tubular secretion) was fitted via parameter identification using the integrated Monte-Carlo algorithm within PK-Sim ® for all studies. The obtained mean value for healthy adults was used for simulations.

| Model building-renally impaired adults
The model was then scaled to renally impaired individuals. Drug-specific parameters (e.g. molecular weight and lipophilicity) were not changed.
With a deteriorating kidney function of 50 mL/min or less, cefuroxime is completely eliminated by glomerular filtration, 27 indicating that tubular secretion does not contribute to the renal elimination in case of renal impairment. To describe the decreasing function of the OAT-systems in renal impairment, the tubular excretion was implemented according to available literature values. 28,29 In Figure   , the ratio of the standard kidney volume (V ks = 0.44 L; a PK-Sim ® specific value) and the V k-v-twin was considered as follows Equation (6): The correlation of renal clearance (Equation 5) obtained through regression analyses is lacking, especially at lower kidney functions.
We used our study population to confirm this equation using the above described procedure. The open symbols in Figure 2b present the optimal clearance values derived via parameter identification using the integrated Monte-Carlo algorithm within PK-Sim ® .
In the ESM (electronic supplementary material), an example calculation for a theoretical 30-year-old person with a kidney function of 115 mL/min is given.

| Model building-study patients' individual simulation
In a next step, the evaluated model for healthy and renal impaired adults was used to simulate the study patients. The low albumin concentrations in our study population leads to a possible change in protein binding compared to a healthy adult population.
Serum protein binding of cefuroxime is described as a fraction unbound (fu) of 0.67 12,14 with reported values ranging from 0.5 to 0.91. 30 35 were used Equation (7): The plasma unbound concentration of cefuroxime was calculated with the aid of the generated fu i and the measured total plasma concentrations.

FIGURE 2
Calculation of the total renal clearance as a function of the estimated glomerular filtration. Left side: fitted tubular secretion plotted against the described estimated glomerular filtration (eGFR). Right side: percentage of the glomerular filtration and tubular secretion in relation to the total renal clearance plotted against their estimated glomerular filtration values. The blue regression line was used for calculation of the renal clearance (equations 5, 6). Filled circles represent eGFR taken from the literature, filled triangles represent the remaining renal clearance described as tubular secretion fitted against literature concentration-time profiles (literature values are described in Table 3), open triangles symbolize the estimated glomerular filtration obtained from the study population, open circles visualize the fitted tubular secretion obtained from the study population, blue and black line: ratio between glomerular filtration and tubular secretion as a function of the eGFR

| Model building-study patients' population simulation
In a last step, a population with 5000 individuals was built corresponding to the study patients' characteristics. For final dose recommendations, this population was split into three groups with defined ranges of GFR: mild renal impairment (40-<80 mL/min) = low, no renal impairment (80-<120 mL/min) = normal and increased clearance (120- ƒT > 2 mg/L, 100% ƒT > 8 mg/L and 100% ƒT > 32 mg/L).

| Dosing strategies
We tested different dosing regimens (bolus vs prolonged infusion), for three different kidney populations matching the anthropometric characteristics of our study population (see Model building):

Prolonged infusion
Bolus + prolonged infusion 1 g every 1 h 3 g over 3 h 0.5-g bolus + 3 g over 3 h 1.5 g every 2.5 h 4.5 g over 3 h 1-g bolus + 3 g over 3 h 4.5 g at start of surgery 1.5-g bolus + 3 g over 3 h Furthermore, we set an upper limit of 3 × 1.5 g cefuroxime over 7.5 hours, as this was the maximum amount of antibiotic used and the longest duration of surgery observed in this study population.

| Sensitivity analyses
Sensitivity analyses were performed on the cefuroxime model to investigate the impact of single model parameters on the predicted AUC inf and C max at the first dose interval given a 1.5-g dose.
Parameters were included into the analysis if they have been optimized, if they could have a strong influence on the PK due to their use in calculation of permeabilities or partition coefficients (e.g. fraction unbound). Sensitivity is calculated as the ratio of the relative change of the simulated AUC inf according to Equation 8: where S is the sensitivity of the AUC to the tested model parameter, ΔAUC is the change of the AUC, and AUC is calculated with the original model parameter value. Sensitivity analyses were performed using a relative perturbation of ±100%. For body weight and height, a relative perturbation of ±10% and for logP and pKa the corresponding values were changed with ±1. This means a 10-fold change due to the octanol:water ratio or the concentration of the oxonium ion! 4 | RESULTS

| Patient demographics
In total, 25 surgical patients treated with cefuroxime were enrolled yielding a total of 135 blood samples. No measured concentration fell below the lower limit of quantification. Twenty-two patients received a second follow-up dose. Detailed demographic data and patient characteristics are summarized in Table 1.

| Evaluation of the model
The relevant input parameters for the PBPK-model are given in   Table 3.

| Simulated dosing regimes
A dosing regimen of 1.5-g bolus immediately followed by a 3 hours extended infusion of 3 g cefuroxime and the standard application scheme ( Figure 5) showed the longest protection times.   When taking PK targets for E. coli, the standard dosing regimen appears to be insufficient (Table 4)

| DISCUSSION
The prevention of SSI is an important component of perioperative medicine. The application of antibiotics as PAP is part of a multifactorial prevention bundle and should be performed in an optimal fashion.
To better characterize the PK properties of cefuroxime in PAP, we used literature data on various aspects of cefuroxime disposition and excretion to build a PBPK model of cefuroxime. The PBPK model was able to accurately describe and predict the PK of cefuroxime. In  In a next step, the model was used to examine the PK adequacy of Besides a timely repetition of PAP, an application scheme considering PK properties of cefuroxime as a β-lactam might be helpful to reduce the number of patients at risk of underdosing. As a time-dependent antibiotic, a bolus directly followed by continuous application during surgery might offer advantages. Using our model, we could show an improvement of E. coli coverage by using a 1.5-g bolus of cefuroxime directly followed by a continuous infusion of 3 g cefuroxime over 3 hours. In long-lasting surgeries, an extension of the continuous infusion appears pragmatic and feasible. The beneficial effects of prolonged or continuous infusion of β-lactam antibiotics have recently been demonstrated. 52 In any inoculum, pathogens occur with elevated MICs beyond clinical breakpoints for sensitive. In this case, a prolonged infusion might be beneficial in order to provide extended coverage above necessary thresholds. 53 Clones with higher MICs carry the risk of further mutations (mutation selection window) and a β-lactam concentration of >6 × MIC is assumed to suppress resistance. 48   As a common approach, missing anthropometric data like age, weight and height were extrapolated with the ICRP data. It has to be considered, that the described healthy volunteers are often a small group of individuals. The possibility that the anthropometric characteristics differ from the average in the ICRP is high. To take into account mean changes of height and weight related to the different decades of the studies, we used the best matching ICRP report for each study (ESM Table S1). The possible bias was tested with a sensitivity analyses (ESM).
The sensitivity analyses showed no relevant impact for cefuroxime Range of the population For simulations of the dose recommendations, the populations were built on basis of the anthropometric values taken from the study collective (age: 18-77 y; weight: 53-120 kg; height 160-193 cm) Predictions for extreme small or obese person should be made carefully. This also needs to be considered for certain age groups (≥78 and ≤17 y) Model is validated with venous data Although a dedicated arterial compartment is not validated, an extrapolation is a strength of PBPK. Both venous and arterial compartment simulations were evaluated, showing minimal differences (MPE arterial = − 1.1% and venous = 5.4%).

No significant effect
Calculation of fraction unbound Measured f u concentrations of cefuroxime were not available in this study. To consider intra-and interindividual differences, we chose an approach by taking the individual albumin concentrations into account. This approach was evaluated and validated before. 22,34 Small possible effect, due to the large effect of f u in the sensitivity analysis eGFR range validated from 34.1-169.3 mL/ min The correlation between eGFR and TS is built and validated on sparse data. For severe renal injury, extrarenal clearance pathways described by Walstad et al. 54  aureus SSI, seems to be reasonable and recommendable. However, the protection against E. coli using the actual standard dosing regimen appears to be insufficient. With our dose recommendations, we provide a potential solution for this issue.