Kocur Arkadiusz, Olkowski Bartosz, Moczulski Mateusz, Miszewska-Szyszkowska Dorota, Rostkowska Olga Maria, Polak Katarzyna, Korniluk Katarzyna, Bączkowska Teresa, Durlik Magdalena, Pawiński Tomasz
Department of Drug Chemistry, Pharmaceutical and Biomedical Analysis, Medical University of Warsaw, Banacha 1, 02-097 Warsaw, Poland.
Students' Scientific Club by the Department of Transplantation, Immunology, Nephrology and Internal Medicine, Medical University of Warsaw, 02-006 Warsaw, Poland.
Molecules. 2025 Jul 26;30(15):3139. doi: 10.3390/molecules30153139.
Everolimus (EVE), an mTOR inhibitor, is widely used in solid organ transplantation (SOT) because of its immunosuppressive properties. Due to its narrow therapeutic window and significant pharmacokinetic variability, therapeutic drug monitoring (TDM) is essential for achieving optimal outcomes. We developed and thoroughly validated a robust LC-MS/MS method to measure EVE levels in venous whole blood (WB) and capillary blood collected using two microsampling devices: Mitra™ (volumetric absorptive microsampling, VAMS) and Capitainer (quantitative dried blood spot, qDBS). The validation followed EMA and IATDMCT guidelines, assessing linearity (1.27-64.80 ng/mL for WB and 0.50-60 ng/mL for VAMS/qDBS), as well as selectivity, accuracy, precision, matrix effects, recovery, stability, and incurred sample reanalysis. Clinical validation involved 66 matched samples from 33 adult SOT recipients. The method demonstrated high accuracy and precision across all matrices, with no significant carryover or matrix interference. Statistical analysis using Passing-Bablok regression and Bland-Altman plots showed excellent agreement between the microsampling methods and the venous reference. Hematocrit effects were tested both in laboratory conditions and on clinical samples and were found to be negligible. This study provides the first comprehensive analytical and clinical validation of the Mitra and Capitainer devices for EVE monitoring. The validated LC-MS/MS microsampling method supports decentralized, patient-centred TDM, offering a reliable alternative to conventional blood sampling in transplant care.
依维莫司（EVE）是一种mTOR抑制剂，因其免疫抑制特性而广泛应用于实体器官移植（SOT）。由于其治疗窗狭窄且药代动力学变异性大，治疗药物监测（TDM）对于实现最佳疗效至关重要。我们开发并全面验证了一种稳健的液相色谱-串联质谱（LC-MS/MS）方法，用于测量静脉全血（WB）以及使用两种微量采样装置采集的毛细血管血中的EVE水平：Mitra™（体积吸收性微量采样，VAMS）和Capitainer（定量干血斑，qDBS）。该验证遵循欧洲药品管理局（EMA）和国际治疗药物监测与临床毒理学协会（IATDMCT）的指南，评估线性（WB为1.27 - 64.80 ng/mL，VAMS/qDBS为0.50 - 60 ng/mL）以及选择性、准确性、精密度、基质效应、回收率、稳定性和测定样本再分析。临床验证涉及来自33名成年SOT受者的66对匹配样本。该方法在所有基质中均显示出高准确性和精密度，无明显残留或基质干扰。使用Passing-Bablok回归和Bland-Altman图进行的统计分析表明，微量采样方法与静脉参考之间具有极好的一致性。在实验室条件和临床样本中均测试了血细胞比容的影响，发现其可忽略不计。本研究首次对用于EVE监测的Mitra和Capitainer装置进行了全面的分析和临床验证。经过验证的LC-MS/MS微量采样方法支持以患者为中心的分散式TDM，为移植护理中的传统血液采样提供了可靠的替代方法。
