Puy Jean-Yves, Jordheim Lars Petter, Cros-Perrial Emeline, Dumontet Charles, Peyrottes Suzanne, Lefebvre-Tournier Isabelle
IBMM, UMR 5247, CNRS - UM - ENSCM, Université de Montpellier, Place Eugène Bataillon, 34095 Montpellier Cedex 5, France.
Univ Lyon, Université Claude Bernard Lyon 1, INSERM 1052, CNRS 5286, Centre Léon Bérard, Centre de Recherche en Cancérologie de Lyon, Lyon, 69008, France.
J Chromatogr B Analyt Technol Biomed Life Sci. 2017 May 15;1053:101-110. doi: 10.1016/j.jchromb.2017.03.024. Epub 2017 Mar 24.
Purine nucleoside analogues are widely used in the treatment of haematological malignancies, and their biological activity is dependent on the intracellular accumulation of their triphosphorylated metabolites. In this context, we developed and validated a liquid chromatography tandem mass spectrometry (LC-MS/MS) method to study the formation of 5'-triphosphorylated derivatives of cladribine, fludarabine, clofarabine and 2'-deoxyadenosine in human cancer cells. Br-ATP was used as internal standard. Separation was achieved on a hypercarb column. Analytes were eluted with a mixture of hexylamine (5 mM), DEA (0.4%, v/v, pH 10.5) and acetonitrile, in a gradient mode at a flow rate of 0.3mLmin. Multiple reactions monitoring (MRM) and electrospray ionization in negative mode (ESI-) were used for detection. The application of this method to the quantification of these phosphorylated cytotoxic compounds in a human follicular lymphoma cell line, showed that it was suitable for the study of relevant biological samples.
嘌呤核苷类似物广泛用于血液系统恶性肿瘤的治疗，其生物学活性取决于其三磷酸化代谢产物在细胞内的积累。在此背景下，我们开发并验证了一种液相色谱串联质谱（LC-MS/MS）方法，用于研究克拉屈滨、氟达拉滨、氯法拉滨和2'-脱氧腺苷在人癌细胞中5'-三磷酸化衍生物的形成。溴代三磷酸腺苷（Br-ATP）用作内标。在高碳柱上实现分离。分析物用己胺（5 mM）、二乙胺（0.4%，v/v，pH 10.5）和乙腈的混合物以梯度模式洗脱，流速为0.3mL/min。采用多反应监测（MRM）和负离子模式电喷雾电离（ESI-）进行检测。将该方法应用于定量人滤泡性淋巴瘤细胞系中这些磷酸化细胞毒性化合物，结果表明它适用于相关生物样品的研究。
