Experimental and Clinical Pharmacology Unit-CRO Aviano, National Cancer Institute, IRCCS, Aviano, Italy.
Clinical Trial Office, Scientific Direction-CRO Aviano, National Cancer Institute, IRCCS, Aviano, Italy.
Ther Drug Monit. 2024 Aug 1;46(4):485-493. doi: 10.1097/FTD.0000000000001174. Epub 2024 Feb 6.
Therapeutic drug monitoring (TDM) using cyclin-dependent kinase inhibitors (CDK4/6is) is a novel approach for optimizing treatment outcomes. Currently, palbociclib, ribociclib, and abemaciclib are the available CDK4/6is and are primarily coadministered with letrozole. This study aimed to develop and validate an LC-MS/MS method for the simultaneous analysis of CDK4/6is, 2 active metabolites of abemaciclib (M2 and M20), and letrozole in human plasma for use in TDM studies.
Sample pretreatment comprised protein precipitation with methanol and dilution of the supernatant with an aqueous mobile phase. Chromatographic separation was achieved using a reversed-phase XBridge BEH C18 column (2.5 μm, 3.0 × 75 mm XP), with methanol serving as the organic mobile phase and pyrrolidine-pyrrolidinium formate (0.005:0.005 mol/L) buffer (pH 11.3) as the aqueous mobile phase. A triple quadrupole mass spectrometer was used for the detection, with the ESI source switched from negative to positive ionization mode and the acquisition performed in multiple reaction monitoring mode.
The complete validation procedure was successfully performed in accordance with the latest regulatory guidelines. The following analytical ranges (ng/mL) were established for the tested compounds: 6-300, palbociclib and letrozole; 120-6000, ribociclib; 40-800, abemaciclib; and 20-400, M2 and M20. All results met the acceptance criteria for linearity, accuracy, precision, selectivity, sensitivity, matrix effects, and carryover. A total of 85 patient samples were analyzed, and all measured concentrations were within the validated ranges. The percent difference for the reanalyzed samples ranged from -11.2% to 7.0%.
A simple and robust LC-MS/MS method was successfully validated for the simultaneous quantification of CDK4/6is, M2, M20, and letrozole in human plasma. The assay was found to be suitable for measuring steady-state trough concentrations of the analytes in patient samples.
使用细胞周期蛋白依赖性激酶抑制剂（CDK4/6is）进行治疗药物监测（TDM）是优化治疗效果的一种新方法。目前，帕博西尼、瑞博西尼和阿贝西利是可用的 CDK4/6is，主要与来曲唑联合使用。本研究旨在开发和验证一种 LC-MS/MS 方法，用于同时分析人血浆中的 CDK4/6is、阿贝西利的 2 种活性代谢物（M2 和 M20）和来曲唑，用于 TDM 研究。
样品预处理包括用甲醇沉淀蛋白质，然后用含有水性流动相的上清液稀释。使用反相 XBridge BEH C18 柱（2.5μm，3.0×75mm XP）进行色谱分离，甲醇作为有机流动相，吡咯烷-吡咯烷甲酸（0.005：0.005mol/L）缓冲液（pH 11.3）作为水性流动相。采用电喷雾源从负离子切换到正离子模式的三重四极杆质谱仪进行检测，以多反应监测模式进行采集。
根据最新的监管指南，成功完成了完整的验证程序。为测试化合物建立了以下分析范围（ng/mL）：6-300，帕博西尼和来曲唑；120-6000，瑞博西尼；40-800，阿贝西利；20-400，M2 和 M20。所有结果均符合线性、准确性、精密度、选择性、灵敏度、基质效应和携带污染的验收标准。共分析了 85 例患者样本，所有测量浓度均在验证范围内。重新分析样本的百分比差异在-11.2%至 7.0%之间。
成功验证了一种简单而稳健的 LC-MS/MS 方法，用于同时定量测定人血浆中的 CDK4/6is、M2、M20 和来曲唑。该测定法适用于测量患者样本中分析物的稳态谷浓度。
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