Yeung Priscilla S-W, Miller Paige, Lai-Nyugen Tran Bao, Cheng Phil, Ibrahim Amira, Shi Run-Zhang, Bowen Raffick A R, Luo Ruben Yiqi
Department of Pathology, Stanford University, Stanford, CA, USA.
Clinical Laboratories, Stanford Health Care, Palo Alto, CA, USA.
J Mass Spectrom Adv Clin Lab. 2023 Mar 7;28:99-104. doi: 10.1016/j.jmsacl.2023.03.002. eCollection 2023 Apr.
Therapeutic drug monitoring (TDM) of immunosuppressants is essential for optimal care of transplant patients. Immunoassays and liquid chromatography-mass spectrometry (LC-MS) are the most commonly used methods for TDM. However, immunoassays can suffer from interference from heterophile antibodies and structurally similar drugs and metabolites. Additionally, nominal-mass LC-MS assays can be difficult to optimize and are limited in the number of detectable compounds.
The aim of this study was to implement a mass spectrometry-based test for immunosuppressant TDM using online solid-phase extraction (SPE) and accurate-mass full scan-single ion monitoring (FS-SIM) data acquisition mode.
LC-MS analysis was performed on a TLX-2 multi-channel HPLC with a Q-Exactive Plus mass spectrometer. TurboFlow online SPE was used for sample clean up. The accurate-mass MS was set to positive electrospray ionization mode with FS-SIM for quantitation of tacrolimus, sirolimus, everolimus, and cyclosporine A. MS fragmentation pattern was used for compound confirmation.
The method was validated in terms of precision, analytical bias, limit of quantitation, linearity, carryover, sample stability, and interference. Quantitation of tacrolimus, sirolimus, everolimus, and cyclosporine A correlated well with results from an independent reference laboratory (r = 0.926-0.984).
Accurate-mass FS-SIM can be successfully utilized for immunosuppressant TDM with good correlation with results generated by standard methods. TurboFlow online SPE allows for a simple "protein crash and shoot" sample preparation protocol. Compared to traditional MRM, analyte quantitation by FS-SIM facilitates a streamlined assay optimization process.
免疫抑制剂的治疗药物监测（TDM）对于移植患者的最佳护理至关重要。免疫测定法和液相色谱 - 质谱联用（LC - MS）是TDM最常用的方法。然而，免疫测定法可能会受到嗜异性抗体以及结构相似的药物和代谢物的干扰。此外，标称质量的LC - MS测定法可能难以优化，并且可检测化合物的数量有限。
本研究的目的是使用在线固相萃取（SPE）和精确质量全扫描 - 单离子监测（FS - SIM）数据采集模式实施基于质谱的免疫抑制剂TDM检测。
在配备Q - Exactive Plus质谱仪的TLX - 2多通道高效液相色谱仪上进行LC - MS分析。TurboFlow在线SPE用于样品净化。精确质量质谱仪设置为正电喷雾电离模式，采用FS - SIM对他克莫司、西罗莫司、依维莫司和环孢素A进行定量。MS碎片模式用于化合物确认。
该方法在精密度、分析偏差、定量限、线性、残留、样品稳定性和干扰方面得到了验证。他克莫司、西罗莫司、依维莫司和环孢素A的定量结果与独立参考实验室的结果相关性良好（r = 0.926 - 0.984）。
精确质量FS - SIM可成功用于免疫抑制剂TDM，与标准方法产生的结果具有良好的相关性。TurboFlow在线SPE允许采用简单的“蛋白质沉淀和进样”样品制备方案。与传统的多反应监测（MRM）相比，通过FS - SIM进行分析物定量有助于简化分析方法的优化过程。
