Low Caspofungin Exposure in Patients in Intensive Care Units.

In critically ill patients, drug exposure may be influenced by altered drug distribution and clearance. Earlier studies showed that the variability in caspofungin exposure was high in intensive care unit (ICU) patients. The primary objective of this study was to determine if the standard dose of caspofungin resulted in adequate exposure in critically ill patients. A multicenter prospective study in ICU patients with (suspected) invasive candidiasis was conducted in the Netherlands from November 2013 to October 2015. Patients received standard caspofungin treatment, and the exposure was determined on day 3 of treatment. An area under the concentration-time curve from 0 to 24 h (AUC0-24) of 98 mg · h/liter was considered adequate exposure. In case of low exposure (i.e., <79 mg · h/liter, a ≥20% lower AUC0-24), the caspofungin dose was increased and the exposure reevaluated. Twenty patients were included in the study, of whom 5 had a positive blood culture. The median caspofungin AUC0-24 at day 3 was 78 mg · h/liter (interquartile range [IQR], 69 to 97 mg · h/liter). A low AUC0-24 (<79 mg · h/liter) was seen in 10 patients. The AUC0-24 was significantly and positively correlated with the caspofungin dose in mg/kg/day (P = 0.011). The median AUC0-24 with a caspofungin dose of 1 mg/kg was estimated using a pharmacokinetic model and was 114.9 mg · h/liter (IQR, 103.2 to 143.5 mg · h/liter). In conclusion, the caspofungin exposure in ICU patients in this study was low compared with that in healthy volunteers and other (non)critically ill patients, most likely due to a larger volume of distribution. A weight-based dose regimen is probably more suitable for patients with substantially altered drug distribution. (This study has been registered at ClinicalTrials.gov under registration no. NCT01994096.).

Voriconazole Therapeutic Drug Monitoring Practices in Intensive Care.

BACKGROUND: Routine therapeutic drug monitoring of voriconazole seems to be beneficial. This study investigated the therapeutic drug monitoring practices in intensive care to derive possible recommendations for improvement. METHODS: A retrospective chart review was performed for patients aged ≥18 years who started treatment with voriconazole, which lasted for at least 3 days while being admitted to an intensive care unit to assess possible differences between the patients with and without voriconazole trough concentrations measured. RESULTS: In 64 (76%) of the 84 patients, voriconazole trough concentrations were measured. The groups differed significantly with respect to the duration of voriconazole treatment and intensive care unit admission. Time of sampling was very early and therefore inappropriate for 49% of the first measured voriconazole trough concentrations and in 48% of the subsequent measured concentrations. Of the 349 trough concentrations measured, 129 (37%) were outside the therapeutic window. In 11% of these cases, no recommendation was provided without identifiable reason. In addition, 27% of recommended dose adjustments were not implemented, probably because the advice was not suited for the specific clinical situation. CONCLUSIONS: The performance of voriconazole therapeutic drug monitoring can still be improved although voriconazole concentrations were monitored in most patients. A multidisciplinary approach-for instance by means of antifungal stewardship-will probably be able to overcome problems encountered such as timing of sampling, incompleteness of data in clinical context, and lack of implementation of recommendations.

Limited-sampling strategies for anidulafungin in critically ill patients.

Efficacy of anidulafungin is driven by the area under the concentration-time curve (AUC)/MIC ratio. Determination of the anidulafungin AUC along with MIC values can therefore be useful. Since obtaining a full concentration-time curve to determine an AUC is not always feasible or appropriate, limited-sampling strategies may be useful in adequately estimating exposure. The objective of this study was to develop a model to predict the individual anidulafungin exposure in critically ill patients using limited-sampling strategies. Pharmacokinetic data were derived from 20 critically ill patients with invasive candidiasis treated with anidulafungin. These data were used to develop a two-compartment model in MW\Pharm using an iterative 2-stage Bayesian procedure. Limited-sampling strategies were subsequently investigated using two methods, a Bayesian analysis and a linear regression analysis. The best possible strategies for these two methods were evaluated by a Bland-Altman analysis for correlation of the predicted and observed AUC from 0 to 24 h (AUC0-24) values. Anidulafungin exposure can be adequately estimated with the concentration from a single sample drawn 12 h after the start of the infusion either by linear regression (R2=0.99; bias, 0.05%; root mean square error [RMSE], 3%) or using a population pharmacokinetic model (R2=0.89; bias, -0.1%; RMSE, 9%) in critically ill patients and also in less severely ill patients, as reflected by healthy volunteers. Limited sampling can be advantageous for future studies evaluating the pharmacokinetics and pharmacodynamics of anidulafungin and for therapeutic drug monitoring in selected patients. (This study has been registered at ClinicalTrials.gov under registration no. NCT01047267.).

Subtherapeutic Posaconazole Exposure and Treatment Outcome in Patients With Invasive Fungal Disease.

BACKGROUND: Posaconazole exposure seems to be subtherapeutic in some patients with invasive fungal disease. Due to the pharmacokinetic variability of posaconazole, therapeutic drug monitoring may help to optimize the efficacy of this antifungal drug. METHODS: A retrospective study of patients treated with posaconazole from January 2008 to April 2014 and for whom posaconazole serum concentrations were available was conducted. Risk factors for underexposure of posaconazole were detected, and the relationship between posaconazole exposure and treatment outcome according to the European Organization for Research and Treatment of Cancer (EORTC) criteria was assessed. RESULTS: Seventy patients met the inclusion criteria, 45 patients received posaconazole as treatment, and 25 patients received posaconazole as a prophylactic. Posaconazole serum trough concentrations were <1.25 mg/L in 44.4% of patients receiving treatment and <0.7 mg/L in 40.0% of patients receiving prophylactic posaconazole. Multiple linear regression analysis showed a significant, independent, and negative association of the posaconazole serum trough concentration with a lack of enteral nutrition (P < 0.001), vomiting (P = 0.035), the use of a proton pump inhibitor or H2-receptor antagonist (P < 0.001), a liquid diet (P = 0.002), concomitant chemotherapy (P = 0.004), and a posaconazole dose frequency of 2 times daily (P = 0.015). A higher posaconazole concentration was associated with a better treatment outcome [odds ratio = 22.22 (95% confidence interval, 3.40-145.33); P = 0.001]. CONCLUSIONS: Posaconazole exposure is insufficient in more than 40% of patients at risk of or with invasive fungal disease, and posaconazole exposure is positively correlated with a successful treatment outcome. Therapeutic drug monitoring of posaconazole can detect underexposure and can be helpful in treatment optimization.

Low but sufficient anidulafungin exposure in critically ill patients.

The efficacy of anidulafungin is driven by the area under the concentration-time curve (AUC)/MIC ratio. Patients in intensive care may be at risk for underexposure. In critically ill patients with an invasive Candida infection, the anidulafungin exposure and a possible correlation with disease severity or plasma protein levels were explored. Concentration-time curves were therefore obtained at steady state. Anidulafungin concentrations were measured with a validated liquid chromatography-tandem mass spectrometry (LC-MS/MS) method. The MIC values of the Candida species were determined with the Etest. The target AUC/MIC ratio was based on European Committee on Antimicrobial Susceptibility Testing (EUCAST) data. Twenty patients were included. The patients received a maintenance dose of 100 mg once daily after a loading dose of 200 mg on the first day. The mean (±standard deviation) AUC, maximum concentration of drug in plasma (Cmax), and minimum concentration of drug in plasma (Cmin) were 69.8 ± 24.1 mg · h/liter, 4.7 ± 1.4 mg/liter, and 2.2 ± 0.8 mg/liter, respectively. The MIC values of all cultured Candida species were below the EUCAST MIC breakpoints. The exposure to anidulafungin in relation to the MIC that was determined appeared sufficient in all patients. The anidulafungin exposure was low in our critically ill patients. However, combined with the low MICs of the isolated Candida strains, the lower exposure observed in comparison to the exposure in the general patient population resulted in favorable AUC/MIC ratios, based on EUCAST data. No correlation was observed between anidulafungin exposure and disease severity or plasma protein concentrations. In patients with less-susceptible Candida albicans or glabrata strains, we recommend considering determining the anidulafungin exposure to ensure adequate exposure. (This trial has been registered at ClinicalTrials.gov under registration no. NCT01047267.).

Inflammation is associated with voriconazole trough concentrations.

Voriconazole concentrations display a large variability, which cannot completely be explained by known factors. Inflammation may be a contributing factor, as inflammatory stimuli can change the activities and expression levels of cytochrome P450 isoenzymes. We explored the correlation between inflammation, reflected by C-reactive protein (CRP) concentrations, and voriconazole trough concentrations. A retrospective chart review of patients with at least one steady-state voriconazole trough concentration and a CRP concentration measured on the same day was performed. A total of 128 patients were included. A significantly (P < 0.001) higher voriconazole trough concentration was observed in patients with severe inflammation (6.2 mg/liter; interquartile range [IQR], 3.4 to 8.7 mg/liter; n = 20) than in patients with moderate inflammation (3.4 mg/liter; IQR, 1.6 to 5.4 mg/liter; n = 60) and in patients with no to mild inflammation (1.6 mg/liter; IQR, 0.8 to 3.0 mg/liter; n = 48). The patients in all three groups received similar voriconazole doses based on mg/kg body weight (P = 0.368). Linear regression analyses, both unadjusted and adjusted for covariates of gender, age, dose, route of administration, liver enzymes, and interacting coadministered medications, showed a significant association between voriconazole and CRP concentration (P < 0.001). For every 1-mg/liter increase in the CRP concentration, the voriconazole trough concentration increased by 0.015 mg/liter (unadjusted 95% confidence interval [CI], 0.011 to 0.020 mg/liter; adjusted 95% CI, 0.011 to 0.019 mg/liter). Inflammation, reflected by the C-reactive protein concentration, is associated with voriconazole trough concentrations. Further research is necessary to assess if taking the inflammatory status of a patient into account is helpful in therapeutic drug monitoring of voriconazole to maintain concentrations in the therapeutic window, thereby possibly preventing suboptimal treatment or adverse events.

Application of sweat patch screening for 16 drugs and metabolites using a fast and highly selective LC-MS/MS method.

BACKGROUND: To facilitate the monitoring of drug abuse by patients, a method was developed and validated for fast and highly selective screening for amphetamine, methamphetamine, 3,4-methylenedioxymethamphetamine, methylenedioxyamphetamine, methylenedioxyethylamphetamine, methylphenidate, cocaine, benzoylecgonine, morphine, codeine, heroin, 6-monoacteylmorphine, methadone, 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine, nicotine, and cotinine in PharmCheck sweat patches. The analysis of sweat patches would provide a noninvasive alternative matrix to urine or blood samples. METHODS: The sweat patches were extracted during vigorous shaking for 10 minutes with 1.5 mL of 20 mmol/L ammonium formate, pH 7, and methanol (50%:50% vol/vol). The extracts were cleaned up by filtering through Whatman mini-Uniprep syringeless filter vials before injection. The method uses a single injection to detect and confirm all 16 drugs and metabolites within 9.6 minutes. RESULTS: The validated substances have a linear range of 3.0-300 nanograms per patch, except for nicotine which has a linear range of 30-3750 nanograms per patch. Stabilities of all substances in worn sweat patches were validated at room temperature for 7 days and as a processed sample in the autosampler at 10°C for 5 days. Only heroin was unstable, with high individual variability and reported bias and coefficient of variation of, respectively, -30.6% and 22.1% in worn sweat patches at room temperature. The monitoring of ion ratios was added to the validation criteria. This resulted in analytical cutoff concentrations of 3.0 and 60 nanograms per patch for nicotine with validated qualifier/quantifier ratios. All analytical cutoff concentrations were lower than the cutoff concentrations proposed by the Substance Abuse and Mental Health Services Administration. CONCLUSIONS: The method uses validated cutoff concentrations, qualifier/quantifier ratios, and a simple extraction without extensive sample treatment for the analysis of 16 drugs and metabolites with a runtime of 9.6 minutes. This method was successfully applied for the analysis of 96 worn sweat patches to monitor patients for drug abuse. The results provided the physician or health-care professional with information about drug abuse and could be used to improve patient care with patient-specific therapy.

Fast and highly selective LC-MS/MS screening for THC and 16 other abused drugs and metabolites in human hair to monitor patients for drug abuse.

BACKGROUND: To facilitate the monitoring of drug abuse by patients, a method was developed and validated for the analysis of amphetamine, methamphetamine, 3,4-methylenedioxymethamphetamine, methylenedioxyamphetamine, methylenedioxyethylamphetamine, methylphenidate, cocaine, benzoylecgonine, morphine, codeine, heroin, 6-monoacteylmorphine, methadone, 2-ethylidene-1,5-dimethyl-3,3-diphenylpyrrolidine (EDDP), delta-9-tetrahydrocannabinol (THC), nicotine, and cotinine in human hair. METHODS: The hair preparation method contains a 3-step wash procedure with dichloromethane followed by a simultaneous hair pulverization and extraction procedure with disposable metal balls. The developed liquid chromatography tandem mass spectrometry method uses a single injection to detect and confirm all 17 abused drugs, including THC, within 4.8 minutes. RESULTS: Nicotine was validated with a linear range of 800-25,000 pg/mg hair, and all other substances were validated with a linear range of 30.0-2500 pg/mg hair. For inaccuracy and imprecision, the overall bias did not exceed -8.2% and the overall coefficient of variation did not exceed 17.7%. Autosampler stability was proven for 48 hours at 10°C for all substances. Analytical cutoff concentrations were defined for each substance at the lowest validated inaccuracy and imprecision concentration with a bias and coefficient of variation within 15% and qualifier/quantifier ratios within 20% of the set ratio. The analytical cutoff concentrations were 200 pg/mg for codeine and 80.0 pg/mg for 6-MAM, heroin, EDDP, and THC. The analytical cutoff concentration for nicotine was 800 pg/mg and for all other validated substances 30.0 pg/mg. This method was successfully applied to analyze hair samples from patients who were monitored for drug abuse. Hair samples of 47 subjects (segmented into 129 samples) showed 3,4-methylenedioxymethamphetamine, methylphenidate, cocaine, benzoylecgonine, codeine, methadone, EDDP, THC, nicotine, and cotinine above the analytical cutoff. CONCLUSIONS: The method was fully validated, including the validation of the qualifier/quantifier ratios. The analysis of real hair samples proved the efficacy of the developed method for monitoring drug abuse. The results obtained by this method provide the physician or health-care professional with extensive information about actual drug abuse or relapse and can be used for patient-specific therapy.

Assessing the availability of the teratogenic drug isotretinoin outside the pregnancy prevention programme: a survey of e-pharmacies.

PURPOSE: The increase in online purchasing of medications raises safety concerns regarding teratogenic drugs. The use of the teratogenic drug 'isotretinoin' for women of childbearing age requires strict adherence to the Pregnancy Prevention Programme (PPP), a risk minimisation measure imposed on prescribers and users. We sought to determine how readily consumers can purchase isotretinoin online and the associated safety procedures and information. METHODS: A descriptive cross-sectional survey was conducted of 50 e-pharmacies identified from commonly used search engines. E-pharmacy characteristics and isotretinoin PPP specific criteria were evaluated. Purchases of isotretinoin from seven e-pharmacies not bearing authentication logos and not requiring a prescription were assessed for PPP policy adherence, purchasing procedures and compound quality. RESULTS: Forty-three (86%) of the e-pharmacies did not have an authentication seal/logo. Isotretinoin could be purchased from 42 sites without a valid prescription. Information on isotretinoin causing birth defects was lacking in 25 of the 50 sites, on not taking isotretinoin in pregnancy in 24 sites and not taking isotretinoin if planning or at risk of a pregnancy in 33 sites. Of the eight attempted purchases, seven arrived, all without any patient information leaflet. All were verified as isotretinoin. CONCLUSION: The Internet provides a loophole for purchasing of medications known to cause congenital abnormalities, which needs to be addressed by medicines regulatory agencies worldwide. The current PPP for isotretinoin may be failing to protect mothers and babies from preventable harm-clinicians need to be aware of this, and the public needs to be educated about the potential risks.

Dried blood spot analysis suitable for therapeutic drug monitoring of voriconazole, fluconazole, and posaconazole.

Invasive aspergillosis and candidemia are important causes of morbidity and mortality in immunocompromised and critically ill patients. The triazoles voriconazole, fluconazole, and posaconazole are widely used for the treatment and prophylaxis of these fungal infections. Due to the variability of the pharmacokinetics of the triazoles among and within individual patients, therapeutic drug monitoring is important for optimizing the efficacy and safety of antifungal treatment. A dried blood spot (DBS) analysis was developed and was clinically validated for voriconazole, fluconazole, and posaconazole in 28 patients. Furthermore, a questionnaire was administered to evaluate the patients' opinions of the sampling method. The DBS analytical method showed linearity over the concentration range measured for all triazoles. Results for accuracy and precision were within accepted ranges; samples were stable at room temperature for at least 12 days; and different hematocrit values and blood spot volumes had no significant influence. The ratio of the drug concentration in DBS samples to that in plasma was 1.0 for voriconazole and fluconazole and 0.9 for posaconazole. Sixty percent of the patients preferred DBS analysis as a sampling method; 15% preferred venous blood sampling; and 25% had no preferred method. There was significantly less perception of pain with the DBS sampling method (P = 0.021). In conclusion, DBS analysis is a reliable alternative to venous blood sampling and can be used for therapeutic drug monitoring of voriconazole, fluconazole, and posaconazole. Patients were satisfied with DBS sampling and had less pain than with venous sampling. Most patients preferred DBS sampling to venous blood sampling.

Clarithromycin increases linezolid exposure in multidrug-resistant tuberculosis patients.

The use of linezolid for the treatment of multidrug-resistant tuberculosis is limited by dose- and time-dependent toxicity. Recently, we reported a case of pharmacokinetic drug-drug interaction between linezolid and clarithromycin that resulted in increased linezolid exposure. The aim of this prospective pharmacokinetic study is to quantify the effect of clarithromycin on the exposure of linezolid. Subjects were included in an open-label, single-centre, single-arm, fixed-order pharmacokinetic interaction study. All subjects received 300 mg linezolid twice daily during the entire study, consecutively co-administered with 250 mg and 500 mg clarithromycin once daily. Steady-state serum curves of linezolid and clarithromycin were analysed using validated methods, and differences between pharmacokinetic parameters were calculated. Linezolid exposure increased by a median (interquartile range) of 44% (23-102%, p=0.043) after co-administration of 500 mg clarithromycin (n=5) compared to baseline, whereas 250 mg clarithromycin had no statistically significant effect. Co-administration was well tolerated by most patients; none experienced severe adverse effects. One patient reported common toxicity criteria grade 2 gastrointestinal adverse events. In this study, we showed that clarithromycin significantly increased linezolid serum exposure after combining clarithromycin with linezolid in multidrug-resistant tuberculosis patients. The drug-drug interaction is possibly P-glycoprotein-mediated. Due to large interpatient variability, therapeutic drug monitoring is advisable to determine individual effect size.

Simultaneous quantification of anidulafungin and caspofungin in plasma by an accurate and simple liquid chromatography tandem mass-spectrometric method.

INTRODUCTION: Echinocandins are a valuable addition for the treatment of invasive fungal infections, as they are efficacious, demonstrate low toxicity, and have limited drug-drug interactions. In specific clinical situations when altered pharmacokinetics can be expected or dosing guidelines are conflicting, it may be useful to measure concentrations. For this purpose, a liquid chromatography tandem mass-spectrometric method to measure anidulafungin and caspofungin in ethylenediaminetetraacetic acid plasma was developed. METHODS: The method was developed on a Thermo Fisher TSQ Quantum LC-MS/MS. For separation, a BetaBasic C4 (100 mm × 3.0 mm; 5 µm) analytical column was used. Sample preparation consisted of protein precipitation directly in the autosampler vial. The internal standard aculeacin A is structurally related, not used in humans, and commercially available. The method was validated according to the guidelines for bioanalytical method validation of the Food and Drug Administration. RESULTS: The method was accurate (bias ranging from -3.0% to 1.9%) and precise (within-run and between-run coefficients of variation of 2.2% to 7.7% and 1.6% to 9.0%, respectively). All calibration curves were linear over a range of 0.5-10.0 mg/L for anidulafungin and 0.1-20.0 mg/L for caspofungin, and if necessary, samples can be diluted 10-fold. The samples were stable for 3 freeze-thaw cycles, with a bias ranging from 0.6% to 11%. The maximum bias from the worst storage condition, 72 hours at room temperature, was -14.7%. In patient samples, anidulafungin peak concentrations ranged from 2.8 to 8.6 mg/L (n = 20) and trough concentrations ranged from 1.0 to 4.7 mg/L (n = 79). The measured caspofungin concentrations ranged from 1.9 to 7.3 mg/L (n = 20). CONCLUSIONS: The method developed has a straightforward sample preparation and uses a structural analog as the internal standard. This method has been applied successfully for the measurement of anidulafungin and caspofungin concentrations in patient samples, both for clinical practice and for research.

Effect of supplementation with zinc and other micronutrients on malaria in Tanzanian children: a randomised trial.

BACKGROUND: It is uncertain to what extent oral supplementation with zinc can reduce episodes of malaria in endemic areas. Protection may depend on other nutrients. We measured the effect of supplementation with zinc and other nutrients on malaria rates. METHODS AND FINDINGS: In a 2×2 factorial trial, 612 rural Tanzanian children aged 6-60 months in an area with intense malaria transmission and with height-for-age z-score≤-1.5 SD were randomized to receive daily oral supplementation with either zinc alone (10 mg), multi-nutrients without zinc, multi-nutrients with zinc, or placebo. Intervention group was indicated by colour code, but neither participants, researchers, nor field staff knew who received what intervention. Those with Plasmodium infection at baseline were treated with artemether-lumefantrine. The primary outcome, an episode of malaria, was assessed among children reported sick at a primary care clinic, and pre-defined as current Plasmodium infection with an inflammatory response, shown by axillary temperature ≥37.5°C or whole blood C-reactive protein concentration ≥ 8 mg/L. Nutritional indicators were assessed at baseline and at 251 days (median; 95% reference range: 191-296 days). In the primary intention-to-treat analysis, we adjusted for pre-specified baseline factors, using Cox regression models that accounted for multiple episodes per child. 592 children completed the study. The primary analysis included 1,572 malaria episodes during 526 child-years of observation (median follow-up: 331 days). Malaria incidence in groups receiving zinc, multi-nutrients without zinc, multi-nutrients with zinc and placebo was 2.89/child-year, 2.95/child-year, 3.26/child-year, and 2.87/child-year, respectively. There was no evidence that multi-nutrients influenced the effect of zinc (or vice versa). Neither zinc nor multi-nutrients influenced malaria rates (marginal analysis; adjusted HR, 95% CI: 1.04, 0.93-1.18 and 1.10, 0.97-1.24 respectively). The prevalence of zinc deficiency (plasma zinc concentration <9.9 µmol/L) was high at baseline (67% overall; 60% in those without inflammation) and strongly reduced by zinc supplementation. CONCLUSIONS: We found no evidence from this trial that zinc supplementation protected against malaria. TRIAL REGISTRATION: ClinicalTrials.gov NCT00623857

Limited-sampling strategies for therapeutic drug monitoring of moxifloxacin in patients with tuberculosis.

BACKGROUND: Moxifloxacin (MFX) is a potent drug for multidrug resistant tuberculosis(TB) treatment and is also useful if first-line agents are not tolerated. Therapeutic drug monitoring may help to prevent treatment failure. Obtaining a full concentration-time curve of MFX for therapeutic drug monitoring is not feasible in most settings. Developing a limited-sampling strategy based on population pharmacokinetics (PK) may help to overcome this problem. METHODS: Steady-state plasma concentrations after the administration of 400 mg of MFX once daily were determined in 21 patients with TB, using a validated liquid chromatography-tandem mass spectrometry method. A one-compartment population model was generated and crossvalidated. Monte Carlo data simulation (n=1000) was used to calculate limited-sampling strategies. The correlation between predicted MFX AUC0-24h (area under the concentration-time curve 0 to 24 hours) and observed AUC0-24h was investigated by Bland-Altman analysis. Finally, the predictive performance of the final model was tested prospectively using MFX profiles from patients with TB receiving 400, 600, or 800 mg once daily. RESULTS: Median minimum inhibitory concentration of Mycobacterium tuberculosis isolates was 0.25 mg/L (interquartile range: 0.25-0.5 mg/L). The geometric mean AUC0-24h was 24.5 mg·h/L (range: 8.5-72.2 mg·h/L), which resulted in a geometric mean AUC0-24h/minimum inhibitory concentration ratio of 72 (range: 21-321). PK analysis, based on PK profiles of 400 mg of MFX once daily, resulted in a crossvalidated population PK model with the following parameters: apparent clearance (Cl) 18.5±8.6 L/h per 1.85 m, Vd 3.0±0.7 L/kg corrected lean body mass, Ka 1.15±1.16 h, and F was fixed at 1. After the Monte Carlo simulation, the best predicting strategy for MFX AUC0-24h for practical use was based on MFX concentrations 4 and 14 hours postdosing (r=0.90, prediction bias=-1.5%, and root mean square error=15%). CONCLUSIONS: MFX AUC0-24h in patients with TB can be predicted with acceptable accuracy for clinical management, using limited sampling. AUC0-24h prediction based on 2 samples, 4 and 14 hours postdose, can be used to individualize treatment.

Gemcitabine and epirubicin plasma concentration-related excretion in saliva in patients with non-small cell lung cancer.

AIM: The excretion in saliva of gemcitabine and its metabolite 2',2'-difluorodeoxyuridine (dFdU) as well as epirubicin (Epi) and its metabolite epirubicinol (Epi-ol) was studied in patients with non-small cell lung cancer, treated with gemcitabine plus epirubicin. METHODS: Patients (n = 12) were treated with gemcitabine 1125 mg/m, followed by Epi 100 mg/m. Blood, saliva, and oral mucosa cells were collected during 22 hours for analysis of gemcitabine, Epi, and their metabolites. Gemcitabine, dFdU, Epi, and Epi-ol were quantified by high-performance liquid chromatography. RESULTS: Gemcitabine was cleared rapidly from plasma and undetectable after 3 hours in all patients. Gemcitabine was detectable in saliva during only the first hour after infusion. The Cmax in saliva was 0.66 +/- 0.61 mg/L, and the saliva to plasma ratio (S/P ratio) was 0.038 +/- 0.037. The Cmax of dFdU was reached 1.5-2 hours after gemcitabine infusion and was 1.03 +/- 0.63 mg/L. The dFdU S/P ratios gradually increased from 0.021 +/- 0.013 at t = 1 hour to 0.050 +/- 0.027 at t = 6 hours after infusion. Epi displayed a triexponential plasma concentration-time profile. The Epi and Epi-ol concentrations in saliva at t = 6 hours after administration were 55 +/- 27 and 9 +/- 9 microg/L, respectively, and decreased to 28 +/- 14 and 7 +/- 4 microg/L, respectively, at t = 22 hours. The corresponding S/P ratios were 1.28 +/- 0.73 and 0.36 +/- 0.31 at t = 6 hours and 1.72 +/- 1.00 and 0.62 +/- 0.34 at t = 22 hours, respectively. The amount of Epi in mucosal cells ranged from 135-598 ng per 10 cells at t = 3 hours and decreased to 33-196 ng per 10 cells at t = 22 hours. CONCLUSION: Gemcitabine and Epi, as well as their main metabolites dFdU and Epi-ol, are excreted in detectable amounts in saliva, although their absolute concentrations remain relatively low.

Limited sampling strategies for therapeutic drug monitoring of linezolid in patients with multidrug-resistant tuberculosis.

INTRODUCTION: Linezolid is a potential drug for the treatment of multidrug-resistant tuberculosis but its use is limited because of severe adverse effects such as anemia, thrombocytopenia, and peripheral neuropathy. This study aimed to develop a model for the prediction of linezolid area under the plasma concentration-time curve from 0 to 12 hours (AUC0-12h) by limited sampling strategy to enable individualized dosing. PATIENTS AND METHODS: Fourteen patients with multidrug-resistant tuberculosis received linezolid twice daily as part of their antituberculosis treatment. Linezolid concentrations were determined at steady state by high-performance liquid chromatography tandem mass spectrometry before and at 1, 2, 4, 8, and 12 hours after dosing. Linezolid AUC0-12h population model and limited sampling models were calculated with MWPharm software. The correlation between predicted linezolid AUC0-12h and observed linezolid AUC0-12h was investigated by Bland-Altman analysis. RESULTS: A total of 26 pharmacokinetic profiles were obtained. The median AUC0-12h was 51.8 (interquartile range, 41.8-65.9) mg*h/L at 300 mg and 123.8 (interquartile range, 100.9-152.5) mg*h/L at 600 mg, both twice daily. The most relevant model clinically for prediction of linezolid AUC0-12h used a linezolid trough concentration (r = 0.91, prediction bias = -2.9% and root mean square error = 15%). DISCUSSION: The difference between choosing a trough concentration and two to three samples increased the correlation from 0.90 to 0.95 but appeared not clinically relevant because it did not result in different dosing advice. CONCLUSION: This study showed that linezolid AUC0-12h in patients with multidrug-resistant tuberculosis could be predicted accurately by a minimal sampling strategy and could be used to individualize the dose.

Pharmacokinetics of gemcitabine in non-small-cell lung cancer patients: impact of the 79A>C cytidine deaminase polymorphism.

OBJECTIVE: To study the impact of the 79A>C polymorphism in the cytidine deaminase (CDA) gene on the pharmacokinetics of gemcitabine and its metabolite 2',2'-difluorodeoxyuridine (dFdU) in non-small-cell lung cancer (NSCLC) patients. PATIENTS AND METHODS: Patients (n = 20) received gemcitabine 1,125 mg/m(2) as a 30 min i.v. infusion as part of treatment for NSCLC. Plasma samples were collected during 0-6 h after gemcitabine administration. Gemcitabine and dFdU were quantified by high performance liquid chromatography with ultraviolet detection. The CDA 79A>C genotype was determined with PCR and DNA sequencing. RESULTS: Gemcitabine was rapidly cleared from plasma and undetectable after 3 h. The allele frequency of the 79A>C polymorphism was 0.40. Diplotypes were distributed as A/A n = 8, A/C n = 8 ,and C/C n = 4. No significant differences were found between the different CDA genotypes and gemcitabine or dFdU AUC, clearance, or half-life. CONCLUSION: The 79A>C polymorphism in the CDA gene does not have a major consistent and signficant impact on gemcitabine pharmacokinetics.

Ciclosporin does not influence bone marrow-derived cell differentiation to myofibroblasts early after renal ischemia/reperfusion.

BACKGROUND: Ischemia/reperfusion injury (IRI) is a risk factor for the development of interstitial fibrosis. Previously we had shown that after renal IRI, bone marrow-derived cells (BMDC) can differentiate to interstitial myofibroblasts. Here we hypothesized that the immunosuppressant ciclosporin A (CsA), known for its profibrotic side effect, promotes myofibroblast differentiation of BMDC in the postischemic kidney. METHODS: Using a model of unilateral renal IRI in rats reconstituted with R26-human placental alkaline phosphatase transgenic bone marrow, CsA was administered in a previously defined critical window for differentiation of BMDC to myofibroblasts. We evaluated fibrotic changes in the kidney and myofibroblast differentiation of BMDC on day 14 after CsA treatment. RESULTS: CsA treatment for 14 days led to increased transforming growth factor-beta transcript levels and collagen III deposition in the postischemic kidney. However, neither the total number of alpha-smooth-muscle-actin-positive interstitial myofibroblasts, nor the bone marrow-derived fraction thereof was affected by CsA administration, irrespective of dosage and duration of treatment. CONCLUSIONS: In the critical postischemic window of BMDC differentiation to myofibroblasts, CsA did not promote BMDC differentiation to myofibroblasts, suggesting that, in the clinical setting, CsA is not involved in myofibroblastic differentiation of BMDC.

Robust, high-throughput LC-MS/MS method for therapeutic drug monitoring of cyclosporine, tacrolimus, everolimus, and sirolimus in whole blood.

The authors describe a fast, robust, and straightforward liquid chromatography and tandem mass spectrometry (LC-MS/MS) method with the use of a single LC-MS/MS system for cyclosporine A, tacrolimus, sirolimus, and everolimus in whole blood. The purpose of this method was to replace the immunoassay (IA) methods used in the laboratory of a hospital performing most organ transplantations (including heart, lung, liver, kidneys, bone marrow, and intestinal tract). Several LC-MS/MS methods have been described so far; however, most of them require complicated online extraction procedures. The described LC-MS/MS method uses a chromatographic gradient in combination with protein precipitation as sample preparation. The chromatographic method is capable of separating otherwise interfering peaks, with an analysis time of 2.6 minutes. Analyses were performed on a triple quadrupole LC-MS/MS system, with a C18 column held at 60 degrees C. Sample preparation required only 1 precipitation/dilution step. Sirolimus and everolimus are prepared and measured separately from tacrolimus and cyclosporine. During method development, it was found that the use of zinc sulfate provides process efficiency results of about 100% for tacrolimus and cyclosporine A, but only 81% and 87% for sirolimus and everolimus, respectively. With the developed sample preparation without zinc sulfate for sirolimus and everolimus, process efficiencies were 99% and 108%, respectively. The methods have been fully validated, and in a comparative study, patient samples were analyzed with IA and our developed LC-MS/MS methods. In the comparative studies, correlations (R2 values) of more than 0.85 were found between the IA and the new LC-MS/MS patient blood levels. There was a systematic deviation in blood levels measured by LC-MS/MS compared with IAs for cyclosporine A (17% lower than with immunoassay) and everolimus (30% lower than with IA). There seemed to be little or no systematic deviation for sirolimus and tacrolimus. The controls determined by the LC-MS/MS method over the past 10 months showed coefficient of variations of no more than 8.0% for each of the 4 immunosuppressants. In conclusion, the authors found the developed methods to be cost saving, more flexible, and more sensitive and that these methods have larger linear ranges than the previously used IA methods. The methods are already used for more than 20,000 patient samples in the daily routine, analyzing approximately 70 patient samples per day.