Quantification of farnesylmethylcysteine in lysates of peripheral blood mononuclear cells using liquid chromatography coupled with electrospray tandem mass spectrometry: Pharmacodynamic assay for farnesyl transferase inhibitors
Quantification of farnesylmethylcysteine in lysates of peripheral blood mononuclear cells using liquid chromatography coupled with electrospray tandem mass spectrometry: Pharmacodynamic assay for farnesyl transferase inhibitors
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DOI：
10.1021/ac051786s
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发表时间：
2006-04-15
影响因子：
7.4
通讯作者：
Beijnen, JH
中科院分区：
文献类型：
--
作者：
Appels, NMGM;Rosing, H;Beijnen, JH
生物有效性是确定法尼基转移酶抑制剂等分子靶向药物最佳剂量的重要参数。为了确定浓度-效应关系，可靠和定量的生物检测是先决条件。在这里，我们提出了一种基于生物标记物法尼基甲基半胱氨酸(FMC)产生的蛋白质法尼化的新方法。采用高效液相色谱与串联质谱仪联用进行定量。该分析已经根据FDA最新的生物分析验证指南进行了验证，所有结果都在要求的范围内。FMC是在细胞裂解时被激活的内源性蛋白酶的作用下形成的。该生物标记物可以在A549人肺癌细胞和人外周血单核细胞中检测到。A549细胞与新型异丙烯基转移酶抑制剂AZD3409孵育后，裂解物中FMC浓度显著降低。这些发现为在FTI的临床前和临床剂量发现研究中使用该分析提供了一个非常好的起点。
Biological effectiveness is an important parameter in determining optimal dosages of molecular targeted drugs, such as farnesyl transferase inhibitors. To determine concentration-effect relationships, robust and quantitative biological assays are a prerequisite. Here, we present a novel assay for protein farnesylation that is based on generation of the biomarker farnesylimethylicysteine (FmC). Quantification was performed with liquid chromatography coupled to tandem mass spectrometry. The assay has been validated based on the most recent FDA guidelines for bioanalytical validation, and all results were within requirements. FmC is formed under the action of an endogenous protease that is activated upon cell lysis. The biomarker could be detected in A549 human lung cancer cells as well as in human peripheral blood mononuclear cells. Incubation of A549 cells with AZD3409, a novel prenyl transferase inhibitor, resulted in a significant decrease of the FmC concentration in the lysates. These findings provide a very good starting point for use of this assay in preclinical and clinical dose finding studies with FTIs.