Zentralinstitut For Klinische Chemie Und Laboratoriumsmedizin

Stuttgart, Germany

Zentralinstitut For Klinische Chemie Und Laboratoriumsmedizin

Stuttgart, Germany
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van Gelder T.,Erasmus University Rotterdam | Fischer L.,University of Hamburg | Shihab F.,University of Utah | Shipkova M.,Zentralinstitut For Klinische Chemie Und Laboratoriumsmedizin
Transplantation Reviews | Year: 2017

The mammalian target of rapamycin (mTOR) inhibitor everolimus is a narrow therapeutic index drug for which optimal exposure levels are essential. The consistent pharmacokinetic profile of everolimus allows trough concentration (C0) measurement to be an appropriate and reliable index for therapeutic drug monitoring (TDM). Exposure-response analyses of data from early fixed-dose trials demonstrated that rates of biopsy-proven acute rejection (BPAR) are significantly higher if everolimus C0 declines below 3 ng/mL, an observation confirmed in subsequent concentration-controlled trials. Evidence for the most favorable upper limit is less clear but with reduced-exposure calcineurin inhibitor (CNI) therapy, an upper limit of 8 ng/mL appears to balance efficacy and safety outcomes. The recommended C0 range is 3-8 ng/mL in kidney, liver and heart transplantation patients, based on LC-MS/MS monitoring in whole blood. Randomized clinical trials based on this target range have demonstrated rates of BPAR comparable to a regimen of mycophenolic acid with standard-exposure CNI. Everolimus exhibits moderate intrapatient pharmacokinetic variability, and it can be challenging to maintain stable concentrations within target range in some individuals. Many factors can influence everolimus exposure for a given dose, including hepatic function, activity of the drug efflux pump P-glycoprotein, the rate of everolimus metabolism, drug-drug interactions (predominantly with CYP3A4 and P-glycoprotein inhibitors, including cyclosporine), intake of fatty food, and patient adherence to the prescribed regimen. Trough concentration levels should be monitored 4-5days after the first dose and after any change in everolimus dose, with additional monitoring in response to any change in concomitant medication or other clinical circumstances which could alter everolimus exposure. Although LC-MS/MS is the gold standard for everolimus monitoring, various immunoassays are widely used due to their relative simplicity and lower cost, and results can show considerable discrepancies with reference methods due to issues such as interassay variability and cross-reactivity. Method standardization will be important in the future to improve the consistency and reproducibility of results between centers. In conclusion, based on an extensive program of clinical trials, the optimal exposure range for everolimus in combination with reduced-exposure CNI therapy has been established and can be achieved in most transplant recipients through careful, planned TDM. © 2017 The Authors.


Kirchherr H.,Medizinisches Labor Bremen | Shipkova M.,Zentralinstitut For Klinische Chemie Und Laboratoriumsmedizin | Ahsen N.V.,Medizinisches Labor Bremen
Therapeutic Drug Monitoring | Year: 2013

Objectives: Thiopurine drugs (azathioprine, 6-mercaptopurine) show wide interindividual variability and a narrow therapeutic range thus making therapeutic monitoring of their active metabolite 6-thioguanine nucleotides (6-TGN) desirable. We improved the currently available laborious and complex methodology of therapeutic drug monitoring of 6-TGN and the metabolite 6-methylmercaptopurine (6-MMP) in washed erythrocytes (ery) based on a whole-blood method. Methods: The analytes were hydrolyzed and extracted from 25-mL ethylenediaminetetraacetic acid-Anticoagulated whole-blood spiked with isotope labeled 6-TG-13C2 15N and 6-MMP-d3 internal standards. Chromatography was performed in 5.1 minutes on a C18 reverse phase column followed by detection via electrospray interface- coupled API 4000 mass spectrometer set up in the positive multiple reaction monitoring mode. The hemoglobin concentration was measured in 20 mL of the original sample (AHD575 method), and the results were standardized to 120 g/L of hemoglobin. Results: Calibration curves were linear with r2 < 0.999 (6-TGN and 6-MMP up to 10,000 pmol/0.2 mL). The limit of quantification was 30 pmol/0.2 mL for 6-TGN and 6-MMP. Intraassay and interassay imprecision was >7.5% at 3 tested levels for 6-TGN and 6-MMP, respectively. Method comparisons were as follows: Ery 6-TGN: y = 1.3x 2 11 and ery 6-MMP y = 1.1x 2 124. Conclusions: The new method compares favorably with established ones, allowing for rapid single run determination of 6-TGN and 6-MMP from >50 mL of fresh or frozen whole blood. Linearity and limits of quantification cover the clinically relevant range. Variability during sample preparation and matrix effects are compensated by the use of isotope-labeled internal standards. The whole-blood method is hemoglobin standardized to avoid falsely low results in the case of anemia. The method correlates well with 6-TGN measured in washed erythrocytes, but it requires significantly less hands-on time. Preliminary therapeutic ranges for the most common indications of azathioprine and 6-MP are provided. © 2013 by Lippincott Williams and Wilkins.


Wieland E.,Zentralinstitut For Klinische Chemie Und Laboratoriumsmedizin | Olbricht C.J.,Klinik fur Nieren und Hochdruckkrankheiten und Transplantationszentrum | Susal C.,University of Heidelberg | Gurragchaa P.,Asklepios Klinikum Uckermark | And 8 more authors.
Therapeutic Drug Monitoring | Year: 2010

Therapeutic drug monitoring is a well-established approach in transplantation medicine to guide immunosuppressive therapy. However, it cannot always predict the effects of immunosuppressive drugs on immune cells, because it does not reflect any aspect of an individual patient's immune system. Pharmacodynamic monitoring is a more recent strategy to provide information about the biologic effect of a specific drug or drug combination on the individual transplant patient. Currently, there is a large number of different biomarkers that either directly (specific markers) or indirectly (global markers) relate to the pharmacodynamic effects of immunosuppressive drugs and are under investigation as potential candidates to be introduced in clinical practice. Such biomarkers may be useful to identify patients at risk of developing acute rejection, infection, or cancer as well as patients who are suitable for minimization of immunosuppressant therapy and may be helpful to manage the timing and rate of immunosuppressant weaning. Serial longitudinal monitoring may allow maintenance of an individualized immunosuppressive regimen. Thus, biomarker monitoring is a potential complementary tool to therapeutic drug monitoring. This review summarizes the current state of knowledge about the use of a number of global or drug-specific pharmacodynamic biomarkers. It is not a comprehensive overview of the literature available, but rather an evidence-based reflection by experts who are intensively involved in scientific work in this field. Copyright © 2010 by Lippincott Williams & Wilkins.


Schettler V.,Nephrologisches Zentrum Gottingen GbR | Neumann C.L.,Nephrologisches Zentrum Gottingen GbR | Hagenah G.C.,Nephrologisches Zentrum Gottingen GbR | Schulz E.G.,Nephrologisches Zentrum Gottingen GbR | Wieland E.,Zentralinstitut For Klinische Chemie Und Laboratoriumsmedizin
Atherosclerosis Supplements | Year: 2013

Background: Lipoprotein apheresis (LA) is used in hypercholesterolemic patients suffering from cardiovascular disease (CHD) if a modified diet and lipid-lowering drug regimens had failed. During the first LA treatments LDL-cholesterol (LDL-C) and lipoprotein (a) (Lp(a)) can be decreased very effectively when using generally accepted formulas for calculating plasma (PV) (e.g. Pearson) or blood volumes (BV) as a basis for calculating treatment volume (e.g. Nadler). With respect to LDL-C and Lp(a) levels after LA treatment not all treated patients on steady state with apheresis treatment procedures may achieve the desired target concentrations for LDL-C (<70 mg/dl) and Lp(a) (<30 mg/dl). Are there further ways to increase the effectiveness of LA? Methods: Over months or years of LA the treated volumes were stepwise increased in patients to achieve target cholesterol concentrations but not sufficiently in all cases. Therefore the patients' actual LA treatment volumes were compared to the calculated PV or BV. To possibly optimize the treatment capacity of LA procedures independent of calculated PV or BV the capacity threshold was determined in addition. During LA procedures every 20 min cholesterol, triglycerides, LDL-C, HDL-C and Lp(a) concentrations were determined and related to the hematocrit to exclude dilution effects. Results: In patients undergoing regular LA treated volumes vs. calculated volumes were different: for PV 28 ± 18% (n = 7); for BV 28 ± 20% (n = 6). The mean treated volumes were 1.3 fold larger than the calculated volumes to achieve cholesterol target levels in most LA treatments. With respect to the capacity threshold we observed in only 1 of 13 patients an ineffective long treatment time. No LA procedure failed due to exhausted capacity. Conclusions: Lipoprotein apheresis treatment is a very effective treatment procedure in lowering LDL-C and Lp(a). However, not in all procedures the optimal treatment volume for LA patients may be calculated. However calculations of PV and BV are more or less error-prone. An increase of 1.3 fold in the calculated volumes may be the first step in optimizing individual LA treatment options. In addition, to exclude an exhaustion of LA procedures the determination of the individual capacity threshold in every LA patient may be further helpful to adjust treatment volumes. To substantiate our demand on changed treatment volumes further data are necessary. © 2012 Elsevier Ireland Ltd.


Huttemann M.,Zentralinstitut For Klinische Chemie Und Laboratoriumsmedizin | Shipkova M.,Zentralinstitut For Klinische Chemie Und Laboratoriumsmedizin | Klett C.,Zentralinstitut For Klinische Chemie Und Laboratoriumsmedizin | Hasche G.,PHV Dialysezentrum | And 4 more authors.
Transplantation Proceedings | Year: 2013

Plasma concentrations of A771726, the active moiety of leflunomide, have been suggested to be associated with antiviral efficacy and/or an increased risk of toxicity. A771726 is >99% bound to serum albumin, which can be relevant in kidney transplant recipients (KTRs) displaying impaired function, which leads to increased pharmacologically active free drug concentrations. This study investigated the relationship of total (t-) and free (f-) A771726 concentrations with clinical outcomes. The 20 KTRs displayed a median daily dose and time on leflunomide of 20 mg (range, 10-50) and 16.5 months (range, 2-28), respectively. A median of 6 (range, 1-15) trough concentrations were measured in each patient. All patients received steroids and a calcineurin inhibitor (CNI) as well as 4 of them, cidofovir. To evaluate therapeutic efficacy, we monitored viral loads in the urine and blood, serum creatinine, and kidney histology. To detect toxicity, we recorded blood and platelet counts, hematocrit, hemoglobin concentrations, liver enzymes (alanine aminotransferase [ALT], and aspartate aminotransferase [AST]), and skin diseases. The median t-A771726 concentration was 31.5 mg/L (interindividual range, 11.0-56.4); the median f-A771726 concentration and fraction were 55.8 μg/L and 0.19% (interindividual ranges, 27.9-148.4 μg/L and 0.12%-0.50%), respectively. A weak but significant inverse correlation was observed between the free drug fraction and both the glomerular filtration rate estimated by the Modification of Diet in Renal Disease formula (MDRD-GFR) (r = -0.202) and serum albumin (r = -0.358). Higher MDRD-GFRs were associated with greater t-A771726 concentrations. There were no significant associations between efficacy parameters and either the t- or f-A771726 concentration or between the t-A771726 concentration and toxicity parameters. In contrast, the f-A771726 concentration was significantly associated with leukopenia. These results indicated that f-A771726 concentrations may be more reliable than t-A771726 content to estimate the risk of leukopenia. Intensified elimination due to a higher free drug fraction and compromised absorption associated with a low GFR may have been responsible for the positive correlation between MDRD-GFR and t-A771726. © 2013 Elsevier Inc. All rights reserved.


Feichtiger H.,Zentralinstitut For Klinische Chemie Und Laboratoriumsmedizin | Wieland E.,Zentralinstitut For Klinische Chemie Und Laboratoriumsmedizin | Armstrong V.W.,University of Gottingen | Shipkova M.,Zentralinstitut For Klinische Chemie Und Laboratoriumsmedizin
Clinical Biochemistry | Year: 2010

Objectives: The acyl glucuronide (AcMPAG) of mycophenolic acid (MPA) forms covalent protein adducts and possesses antiproliferative properties independent of IMPDH inhibition. The underlying mechanism is unknown. Disorganized tubulin polymerization prevents cell cycle progression. We investigated whether AcMPAG interacts with tubulin polymerization. Design and methods: AcMPAG (1.0-100 μM) was incubated with bovine tubulin in the presence of GTP. Polymerization was followed at 340 nm. The time until onset and the extent of polymerization were determined. MPA (100 μM), phenolic glucuronide MPAG (100 μM), and paclitaxel (10 μM) served as controls. Results: MPAG was without effect. The AcMPAG effect on tubulin polymerization was dose dependent and significantly stronger (about 2.5-fold) than that of MPA (n = 4; p < 0.05), but weaker than paclitaxel. Conclusions: MPA and AcMPAG can induce tubulin polymerization in the presence of GTP with AcMPAG being significantly stronger. This property of AcMPAG may contribute to its IMPDH independent antiproliferative effect. © 2009.


Leicht S.,Zentralinstitut For Transfusionsmedizin | Shipkova M.,Zentralinstitut For Klinische Chemie Und Laboratoriumsmedizin | Klett C.,Zentralinstitut For Klinische Chemie Und Laboratoriumsmedizin | Gert H.,Zentralinstitut For Klinische Chemie Und Laboratoriumsmedizin | And 8 more authors.
Clinical Biochemistry | Year: 2013

Objectives: Human CD26 is co-stimulatory for lymphocytes, circulates in a soluble form in blood (sCD26), and has intrinsic dipeptidyl peptidase IV (DPPIV) activity. Associations between CD26 expression on the surface of T cells (CD26. +/CD3. +) and acute rejection and between (CD26. +/CD3. +)/DPPIV and clinical immunosuppression have been reported. These results encouraged the investigation of CD26 as a potential biomarker to optimize immunosuppressive therapy. To better understand the significance of CD26, a comparative study of CD26 expression on CD3. + cells, sCD26 concentration, and DPPIV activity in healthy persons (HP) and kidney transplant recipients (KTR) was performed. Design and methods: Thirty-one HP and 34 KTR were included in the study. CD26. +/CD3. + was determined by FACS, sCD26 concentration was determined by ELISA, and DPP activity was determined by spectrophotometry. For KTR, these parameters were studied on the day before transplantation (preTx) and 7 ± 1. days after transplantation (postTx). Results: There was no significant difference in the CD26. +/CD3. +, sCD26, and DPPIV data regarding gender, donor type (16 living donors), delayed graft function (n = 8), or presence of ≥. 4HLA mismatches (n = 16). Compared to the HP data, preTx CD26. +/CD3. + was 4.5-fold higher, sCD26 was 1.2-fold higher, and DPPIV showed no significant difference. PostTx, CD26. +/CD3. + was 3.8-fold higher, and sCD26 and DPPIV decreased significantly, reaching lower values than that observed in HP. Re-transplanted patients (n = 5) showed significantly lower preTx CD26. +/CD3. + expression than patients receiving their first transplant. Patients with preemptive transplantation (n = 7) showed higher postTx CD26. +/CD3. + expression than patients on dialysis. Conclusions: CD26 expression on CD3. + cells was strongly increased in patients with end stage kidney disease compared to HP and remained high early postTx. The differences in sCD26 and DPPIV behavior compared to that of CD26. +/CD3. + postTx may reflect a regulatory response to the new immunological situation and the effects of therapy. © 2013 The Canadian Society of Clinical Chemists.


Aufenanger J.,Klinikum Ingolstadt GmbH | Schernikau E.,Zentralinstitut For Klinische Chemie Und Laboratoriumsmedizin | Wieland E.,Zentralinstitut For Klinische Chemie Und Laboratoriumsmedizin
LaboratoriumsMedizin | Year: 2010

Although the costs for laboratory medication are only very minor compared to the total expenditures in the healthcare system, cost-cutting is frequently focused on hospital laboratories. Only 7.3% of German hospitals have a laboratory headed by a laboratory physician. To discontinue this trend it is mandatory that the hospital is not considered as a cost centre any longer but should be transformed into a profit and competence centre. To achieve this goal, the laboratory needs to show that it adds to the value of patient care and that it is an essential link in the value chain of the total hospital enterprise. The hospital laboratory should be a workflow management system. In particular, microbiology can serve as an example to demonstrate the economic advantage of a local hospital laboratory. The competence of the medical laboratory and its staff has to be offered to the hospital and its medical disciplines around the clock. However, in the future it will also be necessary to provide laboratory competence and excellence to outpatients. To achieve this, hospital laboratories have to consolidate and cooperate in regional networks. © 2010 by Walter de Gruyter Berlin New York 2010.


Shipkova M.,Zentralinstitut For Klinische Chemie Und Laboratoriumsmedizin
Therapeutic Drug Monitoring | Year: 2015

ABSTRACT:: In response to the urgent need for new, reliable biomarkers to complement the guidance of the immunosuppressive therapy a huge number of biomarker candidates to be implemented in clinical practice have been introduced to the transplant community. This includes a diverse range of molecules with very different molecular weights, chemical and physical properties, ex vivo stabilities, in vivo kinetic behaviors and levels of similarity to other molecules, etc. In addition a large body of different analytical techniques and assay protocols can be used to measure biomarkers. Sometimes a complex software-based data evaluation is a prerequisite for appropriate interpretation of the results and for their reporting. Although some analytical procedures are of great value for research purposes, they may be too complex for implementation in a clinical setting. Whereas the proof of “fitness for purpose” is appropriate for validation of biomarker assays used in exploratory drug development studies, a higher level of analytical validation must be achieved and eventually advanced analytical performance might be necessary before diagnostic application in transplantation medicine. A high level of consistency of results between-laboratories as well as between-methods (if applicable) should be obtained and maintained to make biomarkers effective instruments in support of therapeutic decisions.This overview focuses on pre-analytical and analytical aspects to be considered for the implementation of new biomarkers for adjusting immunosuppression in a clinical setting and highlights critical points to be addressed on the way to make them suitable as diagnostic tools. These include but are not limited to appropriate method validation, standardization, education, automation and commercialization. © 2015 Wolters Kluwer Health, Inc. All rights reserved.


PubMed | Zentralinstitut For Klinische Chemie Und Laboratoriumsmedizin
Type: Journal Article | Journal: Transplantation proceedings | Year: 2013

Plasma concentrations of A771726, the active moiety of leflunomide, have been suggested to be associated with antiviral efficacy and/or an increased risk of toxicity. A771726 is >99% bound to serum albumin, which can be relevant in kidney transplant recipients (KTRs) displaying impaired function, which leads to increased pharmacologically active free drug concentrations. This study investigated the relationship of total (t-) and free (f-) A771726 concentrations with clinical outcomes. The 20 KTRs displayed a median daily dose and time on leflunomide of 20 mg (range, 10-50) and 16.5 months (range, 2-28), respectively. A median of 6 (range, 1-15) trough concentrations were measured in each patient. All patients received steroids and a calcineurin inhibitor (CNI) as well as 4 of them, cidofovir. To evaluate therapeutic efficacy, we monitored viral loads in the urine and blood, serum creatinine, and kidney histology. To detect toxicity, we recorded blood and platelet counts, hematocrit, hemoglobin concentrations, liver enzymes (alanine aminotransferase [ALT], and aspartate aminotransferase [AST]), and skin diseases. The median t-A771726 concentration was 31.5 mg/L (interindividual range, 11.0-56.4); the median f-A771726 concentration and fraction were 55.8 g/L and 0.19% (interindividual ranges, 27.9-148.4 g/L and 0.12%-0.50%), respectively. A weak but significant inverse correlation was observed between the free drug fraction and both the glomerular filtration rate estimated by the Modification of Diet in Renal Disease formula (MDRD-GFR) (r = -0.202) and serum albumin (r = -0.358). Higher MDRD-GFRs were associated with greater t-A771726 concentrations. There were no significant associations between efficacy parameters and either the t- or f-A771726 concentration or between the t-A771726 concentration and toxicity parameters. In contrast, the f-A771726 concentration was significantly associated with leukopenia. These results indicated that f-A771726 concentrations may be more reliable than t-A771726 content to estimate the risk of leukopenia. Intensified elimination due to a higher free drug fraction and compromised absorption associated with a low GFR may have been responsible for the positive correlation between MDRD-GFR and t-A771726.

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