Role of therapeutic drug monitoring in pulmonary infections : use and potential for expanded use of dried blood spot samples

Respiratory tract infections are among the most common infections in men. We reviewed literature to document their pharmacological treatments, and the extent to which therapeutic drug monitoring (TDM) is needed during treatment. We subsequently examined potential use of dried blood spots as sample procedure for TDM. TDM was found to be an important component of clinical care for many (but not all) pulmonary infections. For gentamicin, linezolid, voriconazole and posaconazole dried blood spot methods and their use in TDM were already evident in literature. For glycopeptides, beta-lactam antibiotics and fluoroquinolones it was determined that development of a dried blood spot (DBS) method could be useful. This review identifies specific antibiotics for which development of DBS methods could support the optimization of treatment of pulmonary infections

Analytical and pharmacological aspects of therapeutic drug monitoring of mTOR inhibitors

Mammalian Target Of Rapamycin (mTOR) inhibitors represent a new class of immunosuppressant drugs extensively used for the prevention and the treatment of graft rejection in organ transplant recipients. Their current use is due to referred low nephrotoxic effects, particularly important in kidney transplanted and/or patients with renal failure. The most representative drugs of such class are Sirolimus (Siro) and Everolimus (Rad). Both drugs show a narrow therapeutic window, therefore, monitoring of whole-blood drug levels is recommended in order to optimize the therapy. Among the available assays, Liquid Chromatography coupled with UltraViolet or Electrospray Tandem Mass Spectrometry methods (LC/UV or LC/ESI-MSMS) are the most accurate and specific ones. A reliable alternative is represented by immunoassays, which offer the opportunity to minimize sample pre-treatment, thus reducing the time between drawing blood sample and measuring the drug concentration, an important aspect in high-throughput analyses. Despite this, a limitation in the use of immunoassays for therapeutic drug monitoring is the lower specifity compared with the chromatographic methods when analysing structurally-related drugs. New insights to optimize mTOR inhibitors regimens seem to be offered by the evaluation of CYP450 3A activity by using the probe drug approach. To such purpose, there are a number of major probe drugs used for in vivo studies including: midazolam, cortisol, lidocaine, nifedipine, dextromethorphan, erythromycin, dapsone and alfentanil. The aim of the present paper is to report the most recent knowledge concerning this issue, supplying a critical and comprehensive review for whom are involved both in the clinical and analytical areas

Optimizing immunosuppression with mTOR inhibitors in renal transplant recipients

The aim of this thesis was to optimize immunosuppressive therapy, especially everolimus therapy in renal transplantation recipients by identifying pharmacological and pharmacogenetic risk factors influencing pharmacokinetics, and dynamics such as side effects and patient outcome. Therapeutic Drug Monitoring (TDM) of oral immunosuppressive agents is essential to prevent toxicity and or rejection; therefore it is very important to use a reliable and accurate bioanalytical assay. Differences between the most used analytical assays of measuring everolimus in whole blood and its effect on dosing advice were investigated. TDM is performed based on either trough or AUC monitoring and pharmacogenetics might be a valuable addition to TDM. Therefore the population pharmacokinetics of everolimus in a calcineurin free regimen was investigated and predictive factors such as pharmacogenetics were evaluated. In addition a limited sampling model was developed. MTOR inhibitors are known for a variety of side effects and high dropout rates. In this thesis a comprehensive analysis was performed aimed at identifying risk factors for discontinuation and a number of serious side effects. This thesis also describes an analysis aimed at identifying risk factor associated with delayed graft function, acute rejection and subclinical rejection in patients on a cyclosporine based immunosuppressive regimen.Nederlandse Transplantatie Vereniging de Nierstichting AZL ontwikkelingskrediet (OOK) Apotheek Afdeling Nierziekten LUMCUBL - phd migration 201

Comparison of blood sirolimus, tacrolimus and everolimus concentrations measured by LC-MS/MS, HPLC-UV and immunoassay methods

ObjectivesAn LC-MS/MS method was developed for simultaneous quantitation of tacrolimus, sirolimus and everolimus in whole blood, and compared to HPLC-UV and immunoassay methods.Design and methodsBlood (0.1mL) was analysed following solid-phase extraction and chromatographic resolution using a C18 column (45°C) and mobile phase of methanol/40mM ammonium acetate/glacial acetic acid (83/17/0.1) at 200μL/min, with positive electrospray ionisation and multiple reaction monitoring.ResultsIntra- and inter-day imprecision and inaccuracy were ≤12.2% over a 1.5-40μg/L calibration range. An external quality assurance programme confirmed acceptable inaccuracy and imprecision of the LC-MS/MS method, but highlighted problems with immunoassay quantitation, particularly for everolimus, showing a >30% bias in FPIA everolimus concentrations measured in pooled patient samples versus spiked drug-free whole blood.ConclusionsLC-MS/MS provides significant accuracy and precision advantages compared to HPLC and immunoassays. Discrepancies in everolimus concentrations measured by the Seradyn FPIA immunoassay require further investigation.Benedetta C. Sallustio, Benjamin D. Noll, Raymond G. Morri

Therapeutic and toxic blood concentrations of nearly 1,000 drugs and other xenobiotics

Introduction: In order to assess the significance of drug levels measured in intensive care medicine, clinical and forensic toxicology, as well as for therapeutic drug monitoring, it is essential that a comprehensive collection of data is readily available. Therefore, it makes sense to offer a carefully referenced compilation of therapeutic and toxic plasma concentration ranges, as well as half-lives, of a large number of drugs and other xenobiotics for quick and comprehensive information. Methods: Data have been abstracted from original papers and text books, as well as from previous compilations, and have been completed with data collected in our own forensic and clinical toxicology laboratory. The data presented in the table and corresponding annotations have been developed over the past 20 years and longer. A previous compilation has been completely revised and updated. In addition, more than 170 substances, especially drugs that have been introduced to the market since 2003 as well as illegal drugs, which became known to cause intoxications, were added. All data were carefully referenced and more than 200 new references were included. Moreover, the annotations providing details were completely revised and more than 100 annotations were added. Results: For nearly 1,000 drugs and other xenobiotics, therapeutic ("normal") and, if data were available, toxic and comatose-fatal blood-plasma concentrations and elimination half-lives were compiled in a table. Conclusions: In case of intoxications, the concentration of the ingested substances and/or metabolites in blood plasma better predicts the clinical severity of the case when compared to the assumed amount and time of ingestion. Comparing and contrasting the clinical case against the data provided, including the half-life, may support the decision for or against further intensive care. In addition, the data provided are useful for the therapeutic monitoring of pharmacotherapies, to facilitate the diagnostic assessment and monitoring of acute and chronic intoxications, and to support forensic and clinical expert opinions

Trends in Precision Medicine and Pharmacogenetics as an Adjuvant in Establishing a Correct Immunosuppressive Therapy for Kidney Transplant: An Up-to-Date Historical Overview

Kidney transplantation is currently the treatment of choice for patients with end-stage kidney diseases. Although significant advancements in kidney transplantation have been achieved over the past decades, the host’s immune response remains the primary challenge, often leading to potential graft rejection. Effective management of the immune response is essential to ensure the long-term success of kidney transplantation. To address this issue, immunosuppressives have been developed and are now fully integrated into the clinical management of transplant recipients. However, the considerable inter- and intra-patient variability in pharmacokinetics (PK) and pharmacodynamics (PD) of these drugs represents the primary cause of graft rejection. This variability is primarily attributed to the polymorphic nature (genetic heterogeneity) of genes encoding xenobiotic-metabolizing enzymes, transport proteins, and, in some cases, drug targets. These genetic differences can influence drug metabolism and distribution, leading to either toxicity or reduced efficacy. The main objective of the present review is to report an historical overview of the pharmacogenetics of immunosuppressants, shedding light on the most recent findings and also suggesting how relevant is the research and investment in developing validated NGS-based commercial panels for pharmacogenetic profiling in kidney transplant recipients. These advancements will enable the implementation of precision medicine, optimizing immunosuppressive therapies to improve graft survival and kidney transplanted patient outcomes

CD4 T-HELPER CELL COUNT IS AN ALTERNATIVE PROMISING MARKER FOR DOSING CYCLOSPORINE IN KIDNEY TRANSPLANT PATIENT

Objective: The study was aimed to find out the correlation between cyclosporine blood concentrations and (Clusters of Differentiation 4) CD4 T-helper cell count (percentage) in order to use the latter parameter as an alternative marker for cyclosporine dosing. Besides, the study was also aimed to find out the optimum dosing strategy for Iraqi patients requiring cyclosporine therapy in Iraqi hospitals using TDM approach.Methods: One hundred and twenty subjects participated in the study. The subjects are involved two groups; group A was 80 patients (53 males and 27 females) using cyclosporine twice daily (Sandimmune® oral solution containing cyclosporine 100 mg/ml) and they had kidney transplantation for more than one year. The ages of the patients were 15-45 y (mean±SD =31.962±8.8207); and. Group B included 40 healthy control subjects (24 males and 16 females) with ages of 15-45 y (mean±SD =31.666±8.1606). According to the condition and the need of the patients, they were administered cyclosporine dose range of 1-10 mg/kg/d. Ten ml blood samples were withdrawn from each patient after fasting for about 12 h for monitoring trough/minimum blood concentration (C0) of cyclosporine and for determination of (Clusters of Differentiation 4) CD4 T-helper cells count at C0. Other 10 ml of blood was then withdrawn after 2 h of cyclosporine administration to be used for monitoring maximum/peak cyclosporine blood cyclosporine (Cmax) after 2 h of drug intake (C2) and for determination of (Clusters of Differentiation 4) CD4 T-helper cells count at C2. Five ml of blood samples were withdrawn from each control subject for determination of (Clusters of Differentiation 4) CD4 T-helper cells count.Results: Good correlations were found between cyclosporine dose administered to each patient and the resulted C0 and C2. The majority of patients (66 patients=82.5%) had C0 of 150-200 ng/ml and C2 of 700-900 ng/ml, which are within the therapeutic range. The range of cyclosporine doses that produce therapeutic C0 and C2 was 4.1-9 mg/kg/d. The mean total lymphocyte count and percentage decreased significantly in all patients compared to the control subjects (1.26±0.60 vs.1.98±0.66 e3/uL) and (19.92±13.77 vs. 28.88±10.22), respectively. A similar trend was found for the total lymphocyte count and percentage of patients with cyclosporine C0, and C2 within the therapeutic range (66 patients) compared to the control subjects (1.34±0.57 vs. 1.98±0.66) and (18.98±10.93 vs. 28.88±10.22), respectively. Good negative correlations were found between lymphocyte count and percentage versus C0 for all patients and for patients with C0 within the therapeutic range. Similarly, good negative correlations were found between lymphocyte count and percentage versus C2 for all patients and for patients with C2 within the therapeutic range. The (Clusters of Differentiation 4) CD4 T-helper cell percentage at C0 decreased significantly in all patients and patients with cyclosporine blood concentrations within the therapeutic range (66 patients) compared to the control subjects (24.33±10.31 vs. 35.83±9.11) and (25.50±2.44 vs. 35.83±9.11), respectively. Similarly, (Clusters of Differentiation 4) CD4 T-helper cell percentage at C2 decreased significantly in all patients and patients with cyclosporine blood concentrations within the therapeutic range compared to the control subjects (22.60±9.28 vs. 35.83±9.11) and (21.50±2.16 vs. 35.83±9.11), respectively. The range of (Clusters of Differentiation 4) CD4 T-helper cell percentages at C0 for patients with cyclosporine blood levels above the therapeutic concentrations was 21.65-23.43; for patients with cyclosporine blood levels within the therapeutic concentrations, the range was 23.70-29.00; and for patients with cyclosporine blood levels below the therapeutic concentrations, the range was 29.80-34.60. Good negative correlations were found between (Clusters of Differentiation 4) CD4 T-helper cell percentage and C0 for all patients and for patients with blood concentrations of cyclosporine within the therapeutic range. The (Clusters of Differentiation 4) CD4 T-helper cell percentage range at C2 for patients with cyclosporine blood levels above the therapeutic concentrations was 13.40-18.20; for patients with cyclosporine blood levels within the therapeutic concentrations, the range was 18.50-22.23; and for patients with cyclosporine blood levels below the therapeutic concentrations, the range was 22.76-24.42. Identically, good negative correlations were found between (Clusters of Differentiation 4) CD4 T-helper cell percentage and C2 for all patients and for patients with blood concentrations of cyclosporine within the therapeutic range. For patients with cyclosporine blood levels above therapeutic concentrations; the minimum percentage of (Clusters of Differentiation 4) CD4 T-helper cell at C2 was 13.40, whereas, the maximum percentage of (Clusters of Differentiation 4) CD4 T-helper cell at C0 was 20.23. For patients with cyclosporine blood levels within therapeutic concentrations; the minimum percentage of (Clusters of Differentiation 4) CD4 T-helper cell at C2 was 18.50, whereas, the maximum percentage of (Clusters of Differentiation 4) CD4 T-helper cell at C0 was 29.00. For patients with cyclosporine blood levels below therapeutic concentrations; the minimum percentage of (Clusters of Differentiation 4) CD4 T-helper cell at C2 was 23.40, whereas, the maximum percentage of (Clusters of Differentiation 4) CD4 T-helper cell at C0 was 34.60.Conclusion: Good negative (reciprocal) correlations were demonstrated between cyclosporine blood concentrations at C0 and C2 versus The percentage of (Clusters of Differentiation 4) CD4 T-helper cell. Therefore, the percentage of (Clusters of Differentiation 4) CD4 T-helper cell may be used as an alternative or surrogate marker for optimum cyclosporine dosing than the traditional dosing strategy using TDM, since the former approach is direct for reflecting drug safety and efficacy, beside, it is the affordable, fast and simple approach. The range of cyclosporine doses that produce therapeutic C0 and C2 in Iraqi kidney transplant patients was 4.1-9 mg/kg/d

Assessment Of The Pharmacodynamic Effects Of Cyclosporine In Dogs

Cyclosporine is a commonly used immunosuppressive drug in dogs, but dosing is often empirical and based primarily on clinical response. Pharmacokinetic monitoring of blood drug concentrations can be performed, but target blood concentrations for various disease states in dogs are not well described. Pharmacodynamic assays measuring the effects of cyclosporine on target cells are being used to evaluate immunosuppressive effectiveness in humans, but have been minimally explored in veterinary medicine. This dissertation describes the development of pharmacodynamic assays for measuring the effects of cyclosporine on canine T cell cytokine production and surface antigen expression. Incubation with cyclosporine in vitro caused significant suppression of activated T cell production of interleukin-2 (IL-2), IL-4, interferon-gamma (IFN-gamma), CD25, and CD95 measured in peripheral blood mononuclear cells using flow cytometry. IL-2 and IFN-gamma were then evaluated using flow cytometry and quantitative reverse transcription polymerase chain reaction (qRT-PCR) in whole blood incubated with cyclosporine and dexamethasone in vitro. Cyclosporine caused concentration-dependent inhibition of both cytokines, and a greater degree of suppression was noted with qRT-PCR than flow cytometry. Dexamethasone caused concentration-dependent inhibition of IFN-gamma with both methods, but IL-2 reduction was only significant for qRT-PCR. Both methods were then used to evaluate IL-2 and IFN-gamma after administration of high dose oral cyclosporine to dogs. Both qRT-PCR and flow cytometry identified marked cytokine suppression after cyclosporine dosing, but qRT-PCR was uniformly suppressed across the 12-hour dosing interval, while flow cytometry results were significantly higher at trough blood drug concentrations than at peak blood concentrations and subsequent post-dosing time points. Both flow cytometry and qRT-PCR are valid methods for evaluation of T cell cytokine expression in dogs. Further study at lower drug doses is needed to correlate pharmacodynamic results with pharmacokinetic drug concentrations, and to confirm the best method for cytokine monitoring. Studies in clinic patients are also needed to determine the level of cytokine suppression associated with clinical effectiveness in different disease states. Pharmacodynamic evaluation of cyclosporine’s effects shows promise, and may allow for more individualized dosing of cyclosporine in dogs