Whole Blood Tacrolimus Concentrations Research Articles

A Phase I Trial of the Pharmacokinetic Interaction Between Cannabidiol and Tacrolimus.

One in six Americans uses cannabidiol-based or cannabis-derived products. Cannabidiol is a substrate of CYP3A, but its role as a potential CYP3A inhibitor remains unclear. We hypothesized that cannabidiol would inhibit CYP3A-mediated metabolism of tacrolimus. This report is an interim analysis of an open-label, three-period, fixed-sequence, crossover study in healthy participants. Participants first received a single dose of tacrolimus 5 mg orally. After washout, participants later received cannabidiol titrated to 5 mg/kg twice daily for 14 days to reach a steady state, followed by a second single dose of tacrolimus 5 mg orally. Tacrolimus concentrations in whole blood were measured by UHPLC-MS/MS method. Pharmacokinetic parameters were calculated by noncompartmental analysis. Twelve participants completed all periods of the study. The maximum concentration (Cmax) of tacrolimus increased 4.2-fold (P < 0.0001) with cannabidiol (40.2 ± 13.5 ng/mL) compared with without cannabidiol (9.85 ± 4.63 ng/mL). The area under the concentration-vs.-time curve (AUC0-∞) increased 3.1-fold (P < 0.0001). No change in half-life (t1/2) was observed. This study demonstrates that cannabidiol increases tacrolimus exposure. Our data suggest the need for dose reduction in tacrolimus and frequent therapeutic dose monitoring in transplant patients taking cannabidiol concomitantly. Whether this observed interaction occurred due to the inhibition of CYP3A4 and/or CYP3A5 in the liver, intestine, or both, or intestinal drug transporters (e.g., p-glycoprotein) during the first-pass elimination remains to be elucidated.

Examining Whole Blood, Total and Free Plasma Tacrolimus in Elderly Kidney Transplant Recipients.

Therapeutic monitoring is routinely performed to ensure tacrolimus whole-blood concentrations fall within a predefined target. Despite this, patients still experience inefficacy and toxicity that could be related to variability in free (unbound) tacrolimus exposure. Therefore, the aim of this study was to compare tacrolimus-free plasma (Cu), total plasma (Cp), and whole-blood (Cwb) concentrations in adult kidney transplant recipients and to characterize tacrolimus disposition across different matrices. Twelve-hour concentration-time profiling was performed in 15 recipients, allowing simultaneous measurement of Cu, Cp, and Cwb. Pharmacokinetic parameters were estimated using noncompartmental analysis. The relationship between Cwb and Cp were examined using a capacity-limited binding model, incorporating the hematocrit fraction (fHCT) to estimate maximum binding concentration (Bmax) and dissociation constant (Kd). The relationship between Cp and Cu was evaluated using a linear binding model to estimate the nonspecific binding parameter (Nplasma). Nonlinear regression analysis was used to obtain estimates of Bmax, Kd, and Nplasma. A total of 195 paired Cwb, Cp, and Cu values were collected. The median ratios of Cwb:Cp, Cp:Cu, and Cwb:Cu were 9:1, 20:1, and 138:1, respectively. Variability in free plasma exposure was large; free trough values ranged from 8 to 51 ng/L and free area-under-the-concentration-time-curve values ranged from 424 to 7160 ng·h/L. Median (range) estimates of Bmax, Kd, and Nplasma were 90.4 µg/L (22.4-752.5 µg/L), 2.36 µg/L (0-69.2 µg/L), and 0.05 (0.035-0.085), respectively. The interindividual variability (CV%) in binding parameters was considerable (Bmax 117.2%; Nplasma 32.5%). Large variability was observed in tacrolimus-free plasma exposure and binding parameters. Future research to characterize the relationship between tacrolimus Cu and patient outcomes may be of benefit.

Guiding the starting dose of the once-daily formulation of tacrolimus in "de novo" adult renal transplant patients: a population approach.

The once-daily extended-release tacrolimus formulation (ER-Tac) has demonstrated similar efficacy and safety to the twice-daily immediate-release formulation (IR-Tac), but few population-based pharmacokinetic models have been developed in de novo kidney transplant patients to optimize doses. Therefore, this study aimed i) at developing a population pharmacokinetic model for ER-Tac in de novo adult kidney transplant patients ii) and identifying genetic factors and time-varying covariates predictive of pharmacokinetic variability to guide tacrolimus dosage during the early post-transplant period. A total of 1,067 blood tacrolimus concentrations from 138 kidney transplant patients were analyzed. A total of 29 out of 138 patients were intensively sampled for 24h on the day 5 post-transplantation; meanwhile, for the remaining patients, concentrations were collected on days 5, 10, and 15 after transplantation. Tacrolimus daily doses and genetic and demographic characteristics were retrieved from the medical files. Biochemistry time-varying covariates were obtained on different days over the pharmacokinetic (PK) study. A simultaneous PK analysis of all concentrations was carried out using the non-linear mixed-effects approach with NONMEM 7.5. A two-compartment model with linear elimination and delayed absorption best described the tacrolimus pharmacokinetics. Between-patient variability was associated with oral blood clearance (CL/F) and the central compartment distribution volume (Vc/F). Tacrolimus concentrations standardized to a hematocrit value of 45% significantly improved the model (p < 0.001). This method outperformed the standard covariate modeling of the hematocrit-blood clearance relationship. The effect of the CYP3A5 genotype was statistically (p < 0.001) and clinically significant on CL/F. The CL/F of patients who were CYP3A5*1 carriers was 51% higher than that of CYP3A5*1 non-carriers. Age also influenced CL/F variability (p < 0.001). Specifically, CL/F declined by 0.0562 units per each increased year from the value estimated in patients who were 60years and younger. The 36% between-patient variability in CL/F was explained by CYP3A5 genotype, age, and hematocrit. Hematocrit standardization to 45% explained the variability of tacrolimus whole-blood concentrations, and this was of utmost importance in order to better interpret whole-blood tacrolimus concentrations during therapeutic drug monitoring. The dose requirements of CYP3A5*/1 carriers in patients aged 60years or younger would be highest, while CYP3A5*/1 non-carriers older than 60years would require the lowest doses.

Clinical validation of two volumetric absorptive microsampling devices to support home-based therapeutic drug monitoring of immunosuppression.

Dried blood volumetric absorptive microsamples (VAMS) may facilitate home-based sampling to enhance therapeutic drug monitoring after transplantation. This study aimed to clinically validate a liquid chromatography-tandem mass spectrometry assay using 2 VAMS devices with different sampling locations (Tasso-M20 for the upper arm and Mitra for the finger). Patient preferences were also evaluated. Clinical validation was performed for tacrolimus and mycophenolic acid by comparison of paired VAMS and venipuncture samples using Passing-Bablok regression and Bland-Altman analysis. Conversion of mycophenolic acid VAMS to serum concentrations was evaluated using haematocrit-dependent formulas and fixed correction factors defined a priori. Patients' perspectives, including useability, acceptability and feasibility, were also investigated using established questionnaires. Paired samples (n =50) were collected from 25 kidney transplant recipients. Differences for tacrolimus whole-blood concentration were within ±20% for 86 and 88% of samples from the upper arm and fingerstick, respectively. Using correction factors of 1.3 for the upper-arm and 1.47 for finger-prick samples, 84 and 76% of the paired samples, respectively, were within ±20% for mycophenolic acid serum concentration. Patient experience surveys demonstrated limited pain and acceptable useability of the upper-arm device. Tacrolimus and mycophenolic acid can be measured using 2 common VAMS devices with similar analytical performance. Patients are supportive of home-based monitoring with a preference for the Tasso-M20 device.

Tacrolimus trough level and oxidative stress in Tunisian kidney transplanted patients

Background The effect of tacrolimus (TAC) on oxidative stress after kidney transplantation (KT) is unclear. This study aimed to evaluate the influence of TAC trough levels of oxidative stress status in Tunisian KT patients during the post-transplantation period (PTP). Methods A prospective study including 90 KT patients was performed. TAC whole-blood concentrations were measured by the microparticle enzyme immunoassay method and adjusted according to the target range. Plasma levels of oxidants (malondialdehyde (MDA) and advanced oxidation protein products (AOPP)) and antioxidants (ascorbic acid, glutathione (GSH), glutathione peroxidase (GPx), and superoxide dismutase (SOD)) were measured using spectrophotometry. The subjects were subdivided according to PTP into three groups: patients with early, intermediate, and late PT. According to the TAC level, they were subdivided into LL-TAC, NL-TAC, and HL-TAC groups. Results A decrease in MDA levels, SOD activity, and an increase in GSH levels and GPx activity were observed in patients with late PT compared to those with early and intermediate PT (p < 0.05). Patients with LL-TAC had lower MDA levels and higher GSH levels and GPx activity compared with the NL-TAC and HL-TAC groups (p < 0.05). Conclusion Our results have shown that in KT patients, despite the recovery of kidney function, the TAC reduced but did not normalize oxidative stress levels in long-term therapy, and the TAC effect significantly depends on the concentration used.

Effects of Wuzhi Capsule on Whole-Blood Tacrolimus Concentration Levels: A Systematic Review and Meta-Analysis.

Wuzhi Capsule (WZC) is a traditional Chinese medicinal herb widely used to treat drug-induced hepatitis or liver dysfunction and is usually prescribed in China to increase tacrolimus concentration. Several studies with small sample sizes have shown that WZC can increase tacrolimus concentration levels in clinical practice. This study aimed to evaluate the effect of WZC on whole-blood tacrolimus concentration levels and safety. We searched 7 databases for randomized clinical trials (RCTs) and observational studies (OSs) comparing whole-blood tacrolimus concentration levels between WZC and non-WZC treatments. Data analysis was performed using Review Manager version 5.3. This study was conducted according to the Preferred Reporting Items for Systematic Reviews and Meta-Analyses reporting guidelines. Eleven studies involving 6 RCTs and 5 OSs were included. The meta-analysis indicated that whole-blood tacrolimus concentration levels in the WZC group was significantly higher than that of the non-WZC group [weighted mean difference = 1.38, 95% CI (confidence interval), 1.21-1.56, P < 0.001], and similar results were shown in all the subgroups of follow-up time, different primary disease, and different WZC doses. In the self-control OSs, the whole-blood tacrolimus concentration levels in the WZC group was significantly higher than the non-WZC group (weighted mean difference = 1.17, 95% CI, 0.71-1.64, P < 0.001). WZC was generally well tolerated and there was no significant difference in the incidence of adverse reactions between the 2 groups. WZC can increase whole-blood tacrolimus concentration levels. This may be an economical and practical treatment choice for patients, especially those with poor oral tacrolimus absorption capabilities. Nevertheless, RCTs and OSs with large sample sizes and high quality are needed in the future to confirm these positive results.

Influencing factors of post-transplantation diabetes mellitus in kidney transplant recipients and establishment of a risk prediction model.

The aim was to explore the influencing factors of post-trans-plantation diabetes mellitus (PTDM) in kidney transplant recipients and to es-tablish a risk prediction model. A retrospective analysis was performed on the clinical data of 408 patients subjected to kidney transplantation from May 2015 to March 2022. With the simple random sampling method, they were divided into a training set (n=306) and a test set (n=102) at a ratio of 3:1. According to the occurrence of PTDM, the training set was further classified into PTDM and non-PTDM groups. The influencing factors of PTDM were identified by least absolute shrinkage and selection operator and multivariate logistic regression analysis. A nomogram prediction model was constructed and validated. Non-PT-DM and PTDM groups had significantly different preoperative body mass index (BMI), family history of diabetes mellitus, 2-h preoperative and postprandial blood glucose, 2-hpreoperative and postprandial peptide index, postoperative hypomagnesemia, whole blood concentration of tacrolimus, triacylglycerol, glycated albumin and fasting blood glucose (P&lt;0.05). BMI, family history of diabetes mellitus, 2-h preoperative and postprandial blood glucose, and post-operative whole blood tacrolimus concentration were independent risk factors for PTDM. In contrast, the 2-h preoperative and postprandial peptide index was an independent protective factor (P&lt;0.05). The incidence of PTDM in patients receiving kidney transplantation correlates with the family history of diabetes mellitus, preoperative BMI, 2-h postprandial blood glucose, 2-h postprandial peptide index, and postoperative whole blood tacrolimus concentration.

A highly sensitive method for determination of tacrolimus in peripheral blood mononuclear cells by nano liquid chromatography–high resolution accurate mass spectrometry

The determination of intracellular tacrolimus concentration in peripheral blood mononuclear cells (PBMCs) is crucial for assessing the effect-site concentration of tacrolimus. Analytical methods previously reported required a minimum of 3 mL of whole blood sample for measuring the tacrolimus concentration. In this study, we developed a highly sensitive method using EASY nLC 1200 combined with Q Exactive orbitrap mass spectrometer for detecting tacrolimus in PBMCs, requiring only 0.5–2 mL of sample. Furthermore, we compared two primary normalization methods for PBMCs tacrolimus concentration using Passing–Bablok regression, Bland–Altman analysis, Spearman's rank correlation, and Mountain plot. The newly established method was employed to compare tacrolimus concentrations in whole blood and PBMCs among 194 lung transplant recipients. The developed method exhibited high sensitivity with a lower limit of quantitation at 5 pg/mL, and excellent intra- and inter-days accuracy and precision. The comparison between different normalization methods for PBMCs tacrolimus concentration revealed a strong correlation between PBMCs count and intracellular protein amount within these cells. This finding suggests that both PBMCs count and intracellular protein amount can be used for normalizing intracellular tacrolimus levels and can be mutually converted. However, a weaker correlation was observed between PBMCs and whole-blood tacrolimus concentrations in lung transplant recipients, warranting further investigation. The method reported herein enables the quantification of PBMCs tacrolimus concentration using smaller volumes of whole blood samples, which has significant implications for both patients and laboratory personnel.

A Population Pharmacokinetic Model of Whole-Blood and Intracellular Tacrolimus in Kidney Transplant Recipients.

Background and ObjectiveThe tacrolimus concentration within peripheral blood mononuclear cells may correlate better with clinical outcomes after transplantation compared to concentrations measured in whole blood. However, intracellular tacrolimus measurements are not easily implemented in clinical practice. The prediction of intracellular concentrations based on whole-blood concentrations would be a solution for this. Therefore, the aim of this study was to describe the relationship between intracellular and whole-blood tacrolimus concentrations in a population pharmacokinetic (popPK) model.MethodsPharmacokinetic analysis was performed using non-linear mixed effects modelling software (NONMEM). The final model was evaluated using goodness-of-fit plots, visual predictive checks, and a bootstrap analysis.ResultsA total of 590 tacrolimus concentrations from 184 kidney transplant recipients were included in the study. All tacrolimus concentrations were measured in the first three months after transplantation. The intracellular tacrolimus concentrations (n = 184) were best described with an effect compartment. The distribution into the effect compartment was described by the steady-state whole-blood to intracellular ratio (RWB:IC) and the intracellular distribution rate constant between the whole-blood and intracellular compartments. Lean body weight was negatively correlated [delta objective function value (ΔOFV) −8.395] and haematocrit was positively correlated (ΔOFV = − 6.752) with RWB:IC, and both lean body weight and haematocrit were included in the final model.ConclusionWe were able to accurately describe intracellular tacrolimus concentrations using whole-blood concentrations, lean body weight, and haematocrit values in a popPK model. This model may be used in the future to more accurately predict clinical outcomes after transplantation and to identify patients at risk for under- and overexposure.Dutch National Trial Registry number NTR2226Supplementary InformationThe online version contains supplementary material available at 10.1007/s13318-022-00767-8.

The use of freeze-dried blood samples affects the results of a dried blood spot analysis

Dried blood spot (DBS) microsampling has several advantages over venous blood sampling. In a clinical validation study of tacrolimus microsampling it was noted that tacrolimus DBS concentrations ([Tac]DBS) were systematically higher than tacrolimus whole-blood concentrations ([Tac]WB). This observation was explored by investigating the effect of using freeze-dried standards (STFD) for [Tac]DBS measurement.For all experiments, both non-frozen whole-blood samples and whole-blood samples that were frozen and thawed (to simulate freeze-drying) of 10 patients were analyzed. Multiple tacrolimus concentrations were measured: 1) [Tac]WB, 2) [Tac]DBS, where 15 μL was volumetrically applied to a pre-punched DBS disk, and 3) [Tac]DBS, where 50 μL was applied before a 6 mm DBS disk was punched from the card. All tacrolimus concentrations were determined independently using STFD and standards made of non-frozen blood spiked with tacrolimus (STSP).In both non-frozen and frozen and thawed whole-blood samples, [Tac]WB measured with STFD appeared similar to [Tac]WB measured with STSP (Ratios 1.061 and 1.077, respectively). In non-frozen samples, the median ratio between the [Tac]DBS measured with STFD, and [Tac]WB measured with STFD (the reference method), was 1.396. When blood was volumetrically applied to the DBS card (to eliminate the effect of the spreading over the filter paper), this ratio was 1.009.In conclusion, when using DBS microsampling to quantify concentrations of analytes, one should be aware that using the commercially available freeze-dried blood samples for the preparation of standards may affect the spreading of blood on the filter-paper, leading to a systematic error in the results.

Monitoring Tacrolimus Concentrations in Whole Blood and Peripheral Blood Mononuclear Cells: Inter- and Intra-Patient Variability in a Cohort of Pediatric Patients.

Tacrolimus (TAC) is a first-choice immunosuppressant for solid organ transplantation, characterized by high potential for drug-drug interactions, significant inter- and intra-patient variability, and narrow therapeutic index. Therapeutic drug monitoring (TDM) of TAC concentrations in whole blood (WB) is capable of reducing the incidence of adverse events. Since TAC acts within lymphocytes, its monitoring in peripheral blood mononuclear cells (PBMC) may represent a valid future alternative for TDM. Nevertheless, TAC intracellular concentrations and their variability are poorly described, particularly in the pediatric context. Therefore, our aim was describing TAC concentrations in WB and PBMC and their variability in a cohort of pediatric patients undergoing constant immunosuppressive maintenance therapy, after liver transplantation. TAC intra-PBMCs quantification was performed through a validated UHPLC–MS/MS assay over a period of 2–3 months. There were 27 patients included in this study. No significant TAC changes in intracellular concentrations were observed (p = 0.710), with a median percent change of −0.1% (IQR −22.4%–+46.9%) between timings: this intra-individual variability was similar to the one in WB, −2.9% (IQR −29.4–+42.1; p = 0.902). Among different patients, TAC weight-adjusted dose and age appeared to be significant predictors of TAC concentrations in WB and PBMC. Intra-individual seasonal variation of TAC concentrations in WB, but not in PBMC, have been observed. These data show that the intra-individual variability in TAC intracellular exposure is comparable to the one observed in WB. This opens the way for further studies aiming at the identification of therapeutic ranges for TAC intra-PBMC concentrations.

Distribution evaluation of tacrolimus in the ascitic fluid of liver transplant recipients with liver cirrhosis by a sensitive ultra-performance liquid chromatography-tandem mass spectrometry method.

Tacrolimus has a narrow therapeutic index and large individual differences in pharmacokinetics. The distribution of tacrolimus in ascitic fluid and its influence on whole-blood tacrolimus were unclear. In this study, a sensitive ultra-performance liquid chromatography-tandem mass spectrometry method was established and validated for the quantification of tacrolimus in the ascitic fluid of liver transplant recipients. Chromatographic separation was achieved on an Agilent ZORBAX Eclipse Plus Phenyl-Hexyl column (2.1 × 100mm, 3.5 μm). Mass spectrometry was performed in multiple reaction monitoring conditions of transitions m/z 821.4→768.5 for tacrolimus. The concentrations of tacrolimus in the ascitic fluid range from 0.2 to 3.0ng/mL, accounting for 1.19-31.87% of whole-blood tacrolimus concentrations. A linear mixed model showed a statistically significant positive correlation between the steady-state trough blood concentration of tacrolimus and the corresponding amount of tacrolimus excreted in the ascitic fluid for 24 consecutive hours, especially after normalization by daily dose per unit body weight. These data suggested that the distribution of tacrolimus in the ascitic fluid has great individual differences. The whole-blood tacrolimus concentration, dose per unit body weight, and other confounding factors may contribute to the excretion of tacrolimus in ascitic fluid, but the influence of tacrolimus excretion in drained ascitic fluid on the whole-blood tacrolimus concentration is negligible.

Volumetric absorptive microsampling and dried blood spot microsampling vs. conventional venous sampling for tacrolimus trough concentration monitoring

Objectives Monitoring tacrolimus blood concentrations is important for preventing allograft rejection in transplant patients. Our hospital offers dried blood spot (DBS) sampling, giving patients the opportunity to sample a drop of blood from a fingerprick at home, which can be sent to the laboratory by mail. In this study, both a volumetric absorptive microsampling (VAMS) device and DBS sampling were compared to venous whole blood (WB) sampling. Methods A total of 130 matched fingerprick VAMS, fingerprick DBS and venous WB samples were obtained from 107 different kidney transplant patients by trained phlebotomists for method comparison using Passing-Bablok regression. Bias was assessed using Bland-Altman. A multidisciplinary team pre-defined an acceptance limit requiring >80% of all matched samples within 15% of the mean of both samples. Sampling quality was evaluated for both VAMS and DBS samples. Results 32.3% of the VAMS samples and 6.2% of the DBS samples were of insufficient quality, leading to 88 matched samples fit for analysis. Passing-Bablok regression showed a significant difference between VAMS and WB, with a slope of 0.88 (95% CI 0.81-0.97) but not for DBS (slope 1.00; 95% CI 0.95-1.04). Both VAMS (after correction for the slope) and DBS showed no significant bias in Bland-Altman analysis. For VAMS and DBS, the acceptance limit was met for 83.0% and 96.6% of the samples, respectively. Conclusions VAMS sampling can replace WB sampling for tacrolimus trough concentration monitoring, but VAMS sampling is currently inferior to DBS sampling, both regarding sample quality and agreement with WB tacrolimus concentrations.

Pharmacokinetics and Pharmacodynamics of Once-daily Prolonged-release Tacrolimus in Liver Transplant Recipients

PurposeLimited published data are available regarding the pharmacokinetic (PK) and pharmacodynamic (PD) variables of prolonged-release tacrolimus (PRT) after liver transplantations. The goal of this study was to compare the PK and PD profiles of PRT in early and stable liver transplant recipients by developing a population PK model of PRT and investigating the profile of calcineurin activity (CNA) in the peripheral blood mononuclear cells. MethodsA conversion from BID immediate-release tacrolimus (IRT) to once-daily PRT based on a one-to-one daily dose was performed at day 7 (D7) and D90 posttransplantation in groups A (n = 12) and B (n = 12), respectively. Extensive PK samplings, including whole-blood tacrolimus (TAC) concentration, and CNA assessments were performed at D14 and D104 in groups A and B, respectively. TAC concentration–time data (N = 221) were analyzed by using nonlinear mixed effects modeling. FindingsA 2-compartment model with linear elimination and a delayed first-order absorption characterized by 2 transit compartments best described the PK data. Model-predicted dose-normalized (6.0 mg/d) area under the TAC concentration–time curve over the dosing interval in groups A and B was similar (geometric mean, 235.6 ng/mL · h [95% CI, 139.6–598.7] vs 224.6 ng/mL · h [95% CI, 117.6–421.5], respectively; P = 0.94). Area under the CNA versus time curve over the dosing interval did not differ between groups (4897 [3437] and 4079 [1008] pmol/min/106 cells; P = 0.50). In group A, trough CNA at D14 posttransplantation was statistically higher than that measured just before the switch to PRT (ie, D7 posttransplantation) (198 [92] vs 124 [72] pmol/min/106cells, n = 8; P = 0.048); no statistical difference in TAC concentration was observed (P = 0.11). In group B, no statistical difference between D90 and D104 was observed in either trough CNA (149 [78] vs 172 [82] pmol/min/106 cells, n = 6; P = 0.18) or TAC (P = 0.17) concentration. No graft rejection was observed in either of the groups. ImplicationsThis study suggests that one-to-one dosage conversion to once-daily PRT during the early posttransplantation period could result in significant CNA variations but without causing graft rejection. Further investigations in larger cohorts are warranted to confirm these results. ClinicalTrials.gov identifier: NCT02105155.

The level of intracellular tacrolimus in T cell is affected by CD44+ABCB1+activities triggered by inflammation

Rejection is an important issue in kidney transplant. Although with adequate trough level of tacrolimus, acute rejection occurs, and we are focused on these cases. We hypothesized that the lower concentration of tacrolimus in the peripheral blood mononuclear cell would be a cause of rejection; in this regard, we describe ABCB1, which regulates intracellular concentration of tacrolimus. The effect of inflammation on the intracellular concentration of tacrolimus was evaluated, as was the association between that concentration and ABCB1 and CD44 activities. Seven kidney recipients experiencing acute rejection were prospectively enrolled. Both the whole blood concentration of tacrolimus and intracellular concentration of tacrolimus were measured at the time of enrollment and after stabilization. A human T lymphoblastoid cell line (Jurkat T cell) was treated with various concentrations of tacrolimus for 21 h and then stimulated for 3 h. The levels of mRNA interleukin-2, interleukin-8, and interferon-γ decreased dose dependently by tacrolimus. Furthermore, a fluorescence-activated cell sorter was used to count cells expressing CD44 and ABCB1; changes in intracellular concentration of tacrolimus were explored after tacrolimus treatment and stimulation. Also, B6 splenocytes were tested in the same manner as previous Jurkat T cell experiments. The tacrolimus ratio of three patients was lower at the time of acute rejection than when patients were stabilized. In vitro, the intracellular concentration of tacrolimus decreased after stimulation. Under the same conditions, CD44+ABCB1+cells increased in proportion on tacrolimus treatment and stimulation. This work supports the hypothesis that inflammation reduces the intracellular tacrolimus level, possibly via drug efflux mediated by CD44+ABCB1+and inflammation could lead to acute rejection.

Population Pharmacokinetic Analysis of Immediate-Release Oral Tacrolimus Co-administered with Mycophenolate Mofetil in Corticosteroid-Free Adult Kidney Transplant Recipients.

Tacrolimus is the mainstay calcineurin inhibitor frequently administered with mycophenolic acid with or without corticosteroids to prevent graft rejection in adult kidney transplant recipients. The primary objective of this study was to develop and evaluate a population pharmacokinetic model characterizing immediate-release oral tacrolimus co-administered with mycophenolate mofetil (a pro-drug of mycophenolic acid) in adult kidney transplant recipients on corticosteroid-free regimens. The secondary objective was to investigate the effects of clinical covariates on the pharmacokinetics of tacrolimus, emphasizing the interacting effects of mycophenolic acid. Population modeling and evaluation were conducted with Monolix (Suite-2018R1) using the stochastic approximation expectation-maximization algorithm in 49 adult subjects (a total of 320 tacrolimus whole-blood concentrations). Effects of clinical variables on tacrolimus pharmacokinetics were determined by population covariate modeling, regression modeling, and categorical analyses. A two-compartment, first-order absorption with a lag-time, linear elimination, and constant error model best represented the population pharmacokinetics of tacrolimus. The apparent clearance value for tacrolimus was 17.9l/h (6.95% relative standard error) in our model, which is lower compared with similar subjects on corticosteroid-based therapy. The glomerular filtration rate had significant effects on the apparent clearance and central compartment volume of distribution. Conversely, mycophenolic acid did not affect the apparent clearance of tacrolimus. We have developed and internally evaluated a novel population pharmacokinetic model for tacrolimus co-administered with mycophenolate mofetil in corticosteroid-free adult kidney transplant patients. These findings are clinically important and provide further reasons for conducting therapeutic drug monitoring in this specific population.

Dose optimization of tacrolimus with therapeutic drug monitoring and CYP3A5 polymorphism in patients with myasthenia gravis.

Tacrolimus is beneficial for treatment of myasthenia gravis (MG) and has a narrow therapeutic range. Therefore, therapeutic drug monitoring is essential for tacrolimus to optimize dosage and prevent adverse reactions. However, no studies have explored the factors influencing tacrolimus blood concentration in patients with MG. Thus, we aimed to analyze these factors and discuss how to optimize tacrolimus dosage for MG treatment. Data regarding clinical characteristics, therapeutic drugs and adverse reactions of patients with MG who received tacrolimus were collected from 2013 to 2015 at Beijing Hospital. Tacrolimus whole-blood concentrations were measured by chemiluminescent microparticle immunoassay and CYP3A5*3 gene polymorphism was detected by digital fluorescence molecule hybridization fluorescence. Regression analysis was applied to analyze the factors influencing blood concentrations and adverse reactions. It was shown that there was a correlation between concentration and dosage. Furthermore, co-administration of proton pump inhibitor and clarithromycin and CYP3A5*3 gene polymorphism could significantly increase the tacrolimus whole-blood levels. Adverse reactions were related to blood concentration, CYP3A5 genetic polymorphisms and combined medication though logistic regression analysis. The concentration of tacrolimus is affected by many factors. Therapeutic drug monitoring and detection of CYP3A5 gene polymorphism was essential for dosage optimization in patients with MG.

New tablet formulation of tacrolimus with smaller interindividual variability may become a better treatment option than the conventional capsule formulation in organ transplant patients.

To evaluate the pharmacokinetic (PK) and tolerability profiles of a new tablet formulation of tacrolimus and its interindividual variability (IIV) in the systemic exposure, and to compare them with those of the conventional capsule formulation, a randomized, open-label, two-treatment, two-period, two-sequence, crossover study was performed in 47 healthy males. The capsule or tablet formulation of tacrolimus was orally administered, and serial blood samples were collected up to 96 hours after dosing. Whole-blood tacrolimus concentration was determined using liquid chromatography–tandem mass spectrometry. The maximum whole-blood tacrolimus concentration (Cmax) and the area under the whole-blood tacrolimus concentration–time curve from 0 hour to the last quantifiable concentration (AUClast) were compared between the two formulations. The similarity factor (f2) of the in vitro dissolution profiles was calculated. The geometric mean ratio (90% confidence interval) of tablet to capsule was 0.9680 (0.8873–1.0560) and 1.0322 (0.9359–1.1385) for Cmax and AUClast, respectively. The IIV of Cmax and AUClast of the tablet was smaller than the capsule. The f2 values were >50 in all media. Both formulations were well tolerated. Thus, the tablet formulation of tacrolimus has smaller IIV in the systemic exposure than capsule, while having comparable PK and tolerability profiles, which may render it as a better treatment option for organ transplant patients.

A New CYP3A5*3 and CYP3A4*22 Cluster Influencing Tacrolimus Target Concentrations: A Population Approach.

Single nucleotide polymorphisms (SNPs) in the CYP3A5 and CYP3A4 genes have been reported to be an important cause of variability in the pharmacokinetics of tacrolimus in renal transplant patients. The aim of this study was to merge all of the new genetic information available with tacrolimus pharmacokinetics to generate a more robust population model with data from renal transplant recipients. Tacrolimus exposure data from 304 renal transplant recipients were collected throughout the first year after transplantation and were simultaneously analyzed with a population pharmacokinetic approach using NONMEM® version 7.2. The tacrolimus whole-blood concentration versus time data were best described by a two-open-compartment model with inter-occasion variability assigned to plasma clearance. The following factors led to the final model, which significantly decreased the minimum objective function value (p<0.001): a new genotype cluster variable combining the CYP3A5*3 and CYP3A4*22 SNPs defined as extensive, intermediate, and poor metabolizers; the standardization of tacrolimus whole blood concentrations to a hematocrit value of 45%; and age included as patients <63years versus patients ≥63years. External validation confirmed the prediction ability of the model with median bias and precision values of 1.17ng/mL (95% confidence interval [CI] -3.68 to 4.50) and 1.64ng/mL (95% CI 0.11-5.50), respectively. Simulations showed that, for a given age and hematocrit at the same fixed dose, extensive metabolizers required the highest doses followed by intermediate metabolizers and then poor metabolizers. Tacrolimus disposition in renal transplant recipients was described using a new population pharmacokinetic model that included the CYP3A5*3 and CYP3A4*22 genotype, age, and hematocrit.

五酯胶囊对肾移植术后早期他克莫司药代动力学和肝肾功能的影响

目的：五味子，木兰科的多年生草本植物北五味子属的成熟果实，其在许多国家广泛用作滋补类营养品和恢复性加强肝脏及其他器官的功能。五酯胶囊，一种五味子醇提制剂，是一个著名的中药，在中国广泛使用。五酯胶囊及其活性成分木酚素/木脂素显著保护肝损伤，在中国通常与TAC联合应用来防止药物引起的肝炎。本研究旨在探讨五酯胶囊对肾移植术后早期他克莫司血药浓度和肝肾功能的影响。方法：回顾性研究113名肾移植受者，未用五酯胶囊者55人，匹配年龄和性别后应用五酯胶囊者58人，所有患者接受Tac + MMF + Pred三联免疫抑制治疗。检测3天、7天和30天时Tac的血药浓度的变化，使用微粒酶免疫测定(MEIA)法。与此同时，测试所有患者的肝脏、肾脏功能。肝功能、肾功能、全血他克莫司浓度进行了两组之间的分析比较。结果：我们发现五酯胶囊可以显著提高3天、7天和30天时TAC血药浓度。3天、7天和30天时，五酯胶囊组全血Tac浓度平均增加了40.1%、23.3%、15.0%。更重要的是，直接胆红素也显著降低。两组之间其他的肝肾功能指标没有显著的差异(P > 0.05)。结论：五酯胶囊可以显著提高肾移植受者的Tac血药浓度，且具有保护肝脏的作用，没有明显的不良反应。 Objective: Fructus Schisandrae, the ripe fruits of perennial plant of Schisandra chinensis (Turcz.) Baill, perennial herb of family Magnoliaceae, is widely used as a restorative, tonic nutrition to enhance the function of liver and other organs in many countries. Wuzhi capsule, an ethanol extract preparation of Fructus Schisandrae, is a well-known herbal medicine widely used in China. Wuzhi capsule and its active lignans significantly protect liver injury, and are often prescribed with TAC to prevent drug-induced hepatitis in China. This study aimed to investigate the effects of Wuzhi capsule on blood concentration of tacrolimus and liver function, renal function after renal transplantation. Methods: One hundred and thirteen renal transplant recipients were randomly assigned to two groups, i.e., 58 in the WC group and 55 in the control group. All patients received Tac + MMF + Pred triple immunosuppressive therapy. One month was taken as one therapeutic course. Changes of the blood concentration of Tac were detected 3, 7, 30 d after medication in the two groups using microparticle enzyme immune assay (MEIA). Meanwhile, the liver and kidney functions were tested. Liver function, renal function, and whole-blood concentrations of tacrolimus were analyzed to compare between the two groups. Results: We found that WZ capsule could significantly increase TAC blood concentration at 3, 7, 30 d. At 3, 7, 30 d after the application of WC capsule, the blood Tac concentration increased 40.1%, 23.3%, 15.0% averagely. More importantly, serum DBIL was also remarkably reduced. There was no significant difference in the liver and renal functions between the two groups (P > 0.05). Conclusion: WC capsule could significantly increase the blood Tac concentration of renal transplant recipients with liver protection, and has no obvious adverse reaction.