人肝肿瘤细胞株及临床肝肿瘤组织S9中药物的Ⅰ相代谢酶动力学研究

Enzyme kinetics study of phaseⅠmetabolic profile in the human liver tumor cells and clinical tumor tissue S9

目的研究人肝肿瘤细胞株及临床人肝肿瘤组织S9中的Ⅰ相代谢功能。方法采用体外培养人肝肿瘤细胞株与人正常肝细胞株,提取临床来源肝肿瘤组织与肿瘤旁组织S9,考察5种P450亚型(CYP1A2、CYP2C9、CYP2C19、CYP2D6和CYP3A4)的代谢水平,依次选用非那西丁、双氯芬酸、奥美拉唑、右美沙芬、咪达唑仑和睾酮等特异性探针药物,借助LC-MS/MS、HPLC等分析手段测定生成的特定代谢产物量,来对比和表征细胞株和组织中相应P450同工酶的活性大小。结果人肝肿瘤癌旁组织S9相代谢水平远高于人肝癌组织S9。在体外细胞系中,除去双氯芬酸、奥美拉唑的代谢产物低于定量限外,其余底物咪达唑仑、睾酮、非那西丁、右美沙芬在肝肿瘤细胞株中的代谢水平远高于在人正常肝细胞中(P<0.01)。肝肿瘤细胞株与临床组织S9中酶的表达和功能均一致。结论 2种体外代谢模型,即组织S9与细胞S9的Ⅰ相代谢规律并不一致。癌旁组织更能反映肿瘤在体状态对药物Ⅰ相代谢的规律。

AIM To assess the differences in phase I metabolic profile in the human liver tumor cells and clinical tumor tissue S9. METHODS Tumor ceils were treated with a series of concentration of phenacetin, dex- tromethorphan, diclofenac, testosterone, midazolam and omeprazole for I-phase enzyme CYP1A2, CYP2D6, CYP2C9, CYP3A4 and CYP2C19 study with normal cells of the corresponding organ as the control group. HPLC and LC-MS/MS were used to determine the levels of drugs and their metabolites intracellularly or extracellularly to qualitatively analyze the differences of metabolic capacities between the two cells. In vitro and in vivo consistency was compared by using clinical hepatocellular carcinoma（HCC） and adjacent tissue microsomal incubation. RE- SULTS Metabolism of the in vitro model was inconsistent with in vivo one that adjacent tissue had more level. However enzyme activity of CYP1A2, CYP3A4 and CYP2D6 was higher in HepG2 than that in L-02, vitality of CYP2C9 and CYP2C19 was under detection. CONCLUSION The phase I metabolism in tissueS9 does not co- incide with the cell. The adjacent tissue may more reflect the metabolism of tumor in vivo to some extent.