Proteomics Core Facility, Department of Biochemistry and Molecular Biology, Faculty of Medicine, University of Debrecen, Egyetem tér 1, 4032 Debrecen, Hungary.
Metabolomics Research Group, Department of Biochemistry and Molecular Biology, Faculty of Medicine, University of Debrecen, Egyetem tér 1, 4032 Debrecen, Hungary.
Int J Mol Sci. 2024 Oct 18;25(20):11233. doi: 10.3390/ijms252011233.
Autoimmune hepatitis (AIH) is a chronic inflammatory liver disease treated by steroids and immunomodulator thiopurine drugs such as azathioprine (AZA). AZA is metabolized in the human body into bioactive forms such as 6-thioguanine (6-TG) and 6-methyl-mercaptopurine (6-MMP). Monitoring the levels of bioactive AZA metabolites is very important for proper treatment of patients. In this study, our aim was to develop and validate a fast and sensitive ultra-high performance liquid chromatography-mass spectrometry (UHPLC-MS) method for the analysis of 6-TG and 6-MMP from blood samples of patients with AIH to monitor the level of these bioactive metabolites. The detection and quantification of the analytes was carried out by Selected Reaction Monitoring (SRM)-based targeted mass spectrometry. The method was validated according to the EMA guidelines. Blood samples from patients with AIH treated with AZA were analysed with the developed method. The method was successfully validated with appropriate accuracy and precision for the target biomolecules and their concentration in the samples from patients with AIH was determined. The developed and validated UHPLC-MS method enables the fast and precise analysis of AZA metabolites.
自身免疫性肝炎（AIH）是一种慢性炎症性肝病，采用类固醇和免疫调节剂硫嘌呤类药物（如硫唑嘌呤，AZA）治疗。AZA 在人体内代谢为 6-硫鸟嘌呤（6-TG）和 6-甲基巯基嘌呤（6-MMP）等生物活性形式。监测生物活性 AZA 代谢物的水平对于患者的正确治疗非常重要。在这项研究中，我们的目的是开发和验证一种快速灵敏的超高效液相色谱-质谱联用（UHPLC-MS）方法，用于分析 AIH 患者血液样本中的 6-TG 和 6-MMP，以监测这些生物活性代谢物的水平。通过基于选择反应监测（SRM）的靶向质谱法进行分析物的检测和定量。该方法按照 EMA 指南进行了验证。采用所开发的方法分析了接受 AZA 治疗的 AIH 患者的血液样本。该方法对目标生物分子具有适当的准确性和精密度，成功地进行了验证，并确定了 AIH 患者样本中的浓度。所开发和验证的 UHPLC-MS 方法能够快速准确地分析 AZA 代谢物。
