硫嘌呤药物是免疫调节性抗代谢药，其特点是剂量依赖性不良反应，例如骨髓抑制和肝毒性，通常与个体间差异有关，涉及生物转化基础上重要酶的活性，例如硫嘌呤S-甲基转移酶（TPMT） ）。表面增强拉曼散射 (SERS) 光谱学正在成为一种新兴的生物分析工具，与最常用的药物药代动力学分析方法相比，它在成本可承受、分析时间更短和样品制备更容易方面代表了一种有效的替代方法。本研究的目的是通过 SERS 研究 B 类淋巴母细胞系 (NALM-6) 的细胞裂解物中的巯基嘌呤和硫鸟嘌呤药代动力学，该细胞系 (TPMT*1) 或未 (MOCK) 过度表达野生型TPMT 作为体外细胞淋巴细胞模型，可根据形成的硫鸟苷核苷酸 (TGN) 代谢物的量区分具有不同 TPMT 活性水平的细胞。使用由沉积在纸上的银纳米粒子构成的SERS基底对细胞裂解物进行SERS分析，并使用平行样品通过液相色谱-串联质谱（LC-MS/MS）对硫嘌呤核苷酸进行定量。已经建立了一种直接的 SERS 检测方法，该方法可以作为在体外细胞模型中研究硫嘌呤药物药代动力学的工具，以定性区分过度表达 TPMT 酶和不过度表达 TPMT 酶的细胞，作为其他更费力的技术的替代方法。结果强调，在巯嘌呤和硫鸟嘌呤处理后，当 TPMT 过度表达时，TGN 水平下降，甲基化代谢物水平增加。通过 LC-MS/MS 获得的 TGMP 绝对定量 (pmol/1x106cells) 与 SERS 信号（915 cm-1 处的 TGN 强度）之间存在强正相关性（Spearman 等级相关系数 rho=0.96）。在未来的研究中，我们的目标是应用这种方法来研究患者白细胞中的 TPMT 活性。版权所有 © 2023。由 Elsevier B.V. 出版。

The thiopurine drugs are immunomodulatory antimetabolites that are characterized by dose-dependent adverse effects such as myelosuppression and hepatotoxicity, often related to inter-individual differences, involving the activity of important enzymes at the basis of their biotransformation, such as thiopurine S-methyltransferase (TPMT). Surface Enhanced Raman Scattering (SERS) spectroscopy is emerging as a bioanalytical tool and represents a valid alternative in terms of affordable costs, shorter analysis time and easier sample preparation in comparison to the most employed methods for pharmacokinetic analysis of drugs. The aim of this study is to investigate mercaptopurine and thioguanine pharmacokinetics by SERS in cell lysates of a B-lymphoblastoid cell line (NALM-6), that did (TPMT*1) or did not (MOCK) overexpress the wild-type form of TPMT as an in vitro cellular lymphocyte model to discriminate between cells with different levels of TPMT activity on the base of the amount of thioguanosine nucleotides (TGN) metabolites formed. SERS analysis of the cell lysates was carried out using SERS substrates constituted by Ag nanoparticles deposited on paper and parallel samples were used for quantification of thiopurine nucleotides with liquid chromatography-tandem mass-spectrometry (LC-MS/MS). A direct SERS detection method has been set up that could be a tool to study thiopurine drug pharmacokinetics in in vitro cellular models to qualitatively discriminate between cells that do and do not overexpress the TPMT enzyme, as an alternative to other more laborious techniques. Results underlined decreased levels of TGN and increased levels of methylated metabolites when TPMT was over expressed, both after mercaptopurine and thioguanine treatments. A strong positive correlation (Spearman's rank correlation coefficient rho = 0.96) exists between absolute quantification of TGMP (pmol/1x106 cells), obtained by LC-MS/MS, and SERS signal (intensity of TGN at 915 cm-1). In future studies, we aim to apply this method to investigate TPMT activity in patients' leukocytes.Copyright © 2023. Published by Elsevier B.V.