应用色质联用法测定人白血病细胞亚细胞组分内的阿霉素及其代谢物水平.docVIP

  Subcellular Quantification of Doxorubicin and its Metabolite in Cultured Human Leukemia Cells using Liquid Chromatography-Tandem Mass Spectrometry 5 10 15 20 25 30 35 40 XU Jinhui, CHEN Yun (School of Pharmacy,Nanjing Medical University,Nanjing, 210029) Abstract: Doxorubicin (DOX) is widely used in the world as an anti-cancer agent for the treatment of leukemia and solid tumors. However, its clinic use is largely limited by the emergence of cardiotoxicity. One of the most frequently proposed mechanisms for DOX induced cardiotoxicity is the formation of metabolites. However, the enzymatic pathways involved in DOX intracellular metabolism have not been fully elucidated thus far. To provide a more detailed description of DOX metabolism, an assay using liquid chromatography-tandem mass spectrometry (LC/MS/MS) was developed in our lab to simultaneously determine DOX and its primary metabolite doxorubicinol (DOXol) in subcellular compartments. Good accuracy and precision were achieved. Using this assay, the accumulation of DOX and DOXol in whole cells, nuclear enriched fraction (NEF) and organelle-enriched fraction (OEF) were compared between two human T leukemia cell lines (i.e., Jurkat and CCRF-CEM). A time-course analysis was also carried out. The resulting varieties of DOX and DOXol subcellular distributions and concentration-time profiles might be attributed to the differential expression, activities and localization of reductive enzymes within these cell lines. More importantly, this work demonstrated that simultaneous determination of drug and its metabolite in subcellular compartments could be achieved using LC/MS/MS. Key words: Doxorubicin; Metabolite; Subcellular Fractions; Liquid Chromatography-Tandem Mass Spectrometry; Time-course Analysis 0 Introduction Doxorubicin (DOX) is a widely used anthracycline that has been proven highly effective against a variety of human malignancies such as breast cancer, leukemia, Hodgkin’s and Non-Hodgkin’s lymphomas, etc