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Aims: Tacrolimus is a critical dose drug and to avoid under‐ and overexposure, therapeutic drug monitoring is standard practice. However, rejection and drug‐related toxicity occur despite whole‐blood tacrolimus pre‐dose concentrations ([Tac]blood) being on target. Monitoring tacrolimus concentrations at the target site (within peripheral blood mononuclear cells; [Tac]cells) may better correlate with drug‐efficacy. The aim of this study was to (1) investigate the relationship between [Tac]blood and [Tac]cells, (2) identify factors affecting the tacrolimus distribution in cells and whole‐blood, and (3) study the relationship between [Tac]cells and clinical outcomes after kidney transplantation. Methods: A total of 175 renal transplant recipients were prospectively followed. [Tac]blood and [Tac]cells were determined at Months 3, 6 and 12 post‐transplantation. Patients were genotyped for ABCB1 1199G>A and 3435C>T, CYP3A4 15389C>T, and CYP3A5 6986G>A. Data on rejection and tacrolimus‐related nephrotoxicity and post‐transplant diabetes mellitus were collected. Results: Correlations between [Tac]blood and [Tac]cells were moderate to poor (Spearman's r = 0.31; r = 0.41; r = 0.61 at Months 3, 6 and 12, respectively). The [Tac]cells/[Tac]blood ratio was stable over time in most patients (median intra‐patient variability 39.0%; range 3.5%–173.2%). Age, albumin and haematocrit correlated with the [Tac]cells/[Tac]blood ratio. CYP3A5 and CYP3A4 genotype combined affected both dose‐corrected [Tac]blood and [Tac]cells. ABCB1 was not significantly related to tacrolimus distribution. Neither [Tac]blood nor [Tac]cells correlated with clinical outcomes. Conclusions: The correlation between [Tac]blood and [Tac]cells is poor. Age, albumin and haematocrit correlate with the [Tac]cells/[Tac]blood ratio, whereas genetic variation in ABCB1, CYP3A4 and CYP3A5 do not. Neither [Tac]blood nor [Tac]cells correlated with clinical outcomes. [ABSTRACT FROM AUTHOR]

After solid organ transplantation, tacrolimus is given to prevent rejection. Therapeutic drug monitoring is used to reach target concentrations of tacrolimus in whole blood. Because the site of action of tacrolimus is the lymphocyte, and tacrolimus binds ~80% to erythrocytes, the intracellular tacrolimus concentration in lymphocytes is possibly more relevant. For this purpose, we aimed to develop, improve and validate a UPLC–MS/MS method to measure tacrolimus concentrations in isolated peripheral blood mononuclear cells (PBMCs). PBMCs were isolated using a Ficoll separation technique, followed by a washing step using red blood cell lysis. A cell suspension of 50 μL containing 1 million PBMCs was used in combination with MagSiMUS‐TDMPREP. To each sample we added 30 μL lysis buffer, 20 μL reconstitution buffer containing 13C2H4‐tacrolimus as internal standard, 40 μL MagSiMUS‐TDMPREP Type I Particle Mix and 175 μL Organic Precipitation Reagent VI for methanol‐based protein precipitation. A 10 μL aliquot of the supernatant was injected into the UPLC–MS/MS system. The method was validated, resulting in high sensitivity and specificity. The method was linear (r2 = 0.997) over the range 5.0–1250 pg/1 × 106 PBMCs. The inaccuracy was <5% and the imprecision was <15%. The washing steps following Ficoll isolation could be performed at either room temperature or on ice, with no effect of the temperature on the results. A method for the analysis of tacrolimus concentrations in PBMCs was developed and successfully validated. Further research will be performed to investigate the correlation between concentrations in PBMCs and clinical outcome. [ABSTRACT FROM AUTHOR]

Tacrolimus (Tac) is well established as main immunosuppressant in most immunosuppressive regimens in solid organ transplantation. Due to the narrow therapeutic window, pre dose Tac levels (C0) are monitored in all patients receiving Tac to reach optimal therapeutic levels. Tac is metabolized in the liver and intestine by the cytochrome P450 3A (CYP3A) isoforms CYP3A4 and CYP3A5. We present a case of an African American woman who underwent a liver transplantation in which adequate Tac levels were difficult to accomplish due to differences in cytochrome P450 3A4/5 (CYP3A4/5) polymorphisms of the transplant recipient and the donor liver graft. This case report highlights that genotyping the liver transplant recipient and the donor liver graft might provide data which could be used to predict the tacrolimus metabolism post transplantation. [ABSTRACT FROM AUTHOR]

• A multicomponent dried blood spot method was clinically validated in transplant recipients. • A dried blood spot method was validated for cyclosporine A, tacrolimus, creatinine and hematocrit. • A correction for hematocrit is needed for clinical use of dried blood spot for cyclosporine A and tacrolimus. [ABSTRACT FROM AUTHOR]

• The correlation between [Tac] cells and [Tac] blood, is poor. • Tacrolimus exposure and distribution appeared stable in the early phase after transplantation. • The inter-occasion variability in [Tac] cells is lower than the inter-patient variability. • [Tac] cells was not significantly associated with the occurrence of rejection. After kidney transplantation, rejection and drug-related toxicity occur despite tacrolimus whole-blood pre-dose concentrations ([Tac] blood) being within the target range. The tacrolimus concentration within peripheral blood mononuclear cells ([Tac] cells) might correlate better with clinical outcomes. The aim of this study was to investigate the correlation between [Tac] blood and [Tac] cells, the evolution of [Tac] cells and the [Tac] cells /[Tac] blood ratio, and to assess the relationship between tacrolimus concentrations and the occurrence of rejection. In this prospective study, samples for the measurement of [Tac] blood and [Tac] cells were collected on days 3 and 10 after kidney transplantation, and on the morning of a for-cause kidney transplant biopsy. Biopsies were reviewed according to the Banff 2019 update. Eighty-three [Tac] cells samples were measured of 44 kidney transplant recipients. The correlation between [Tac] cells and [Tac] blood was poor (Pearson's r = 0.56 (day 3); r = 0.20 (day 10)). Both the dose-corrected [Tac] cells and the [Tac] cells /[Tac] blood ratio were not significantly different between days 3 and 10, and the median inter-occasion variability of the dose-corrected [Tac] cells and the [Tac] cells /[Tac] blood ratio were 19.4% and 23.4%, respectively (n = 24). Neither [Tac] cells, [Tac] blood, nor the [Tac] cells /[Tac] blood ratio were significantly different between patients with biopsy-proven acute rejection (n = 4) and patients with acute tubular necrosis (n = 4) or a cancelled biopsy (n = 9; p > 0.05). Tacrolimus exposure and distribution appeared stable in the early phase after transplantation. [Tac] cells was not significantly associated with the occurrence of rejection. A possible explanation for these results might be related to the low number of patients included in this study and also due to the fact that PBMCs are not a specific enough matrix to monitor tacrolimus concentrations. [ABSTRACT FROM AUTHOR]