hplc检测白血病细胞内药物代谢酶cyp3a5活性.docVIP

HPLC检测白血病细胞内药物代谢酶CYP3A5活性 摘 要 迄今为止，化疗仍是急性白血病的主要治疗手段，但每一个体接受标准剂量药物治疗时产生的反应，往往存在着极大的个体差异，有些会出现耐药，有些会出现难以耐受的毒副反应。这在临床上是一个非常重要的问题，它可以导致治疗失败，甚至产生严重的药物毒副作用。其中涉及的因素很多，药物代谢酶是最关键的一环，它是介导机体药物代谢活化与代谢解毒之间平衡的关键物质。作为机体最重要的药物代谢酶，细胞色素P450酶系（CYPs）3A亚家族，参与代谢约60％的处方药。本文应用特异性检测CYP3A亚家族酶活性的氢化考的松6β-羟化活性的检测方法，结合高压液相色谱仪(HPLC)检测不同化疗药干预后对K562白血病细胞株CYP3A5活性的影响，以及白血病患者细胞CYP3A5的活性。 第一部分：不同化疗药对K562白血病细胞株CYP3A5酶活性的影响 选取具有CYP3A5高活性的细胞株（K562白血病细胞株）经化疗药长春新碱( 0.08 μg/ml )、高三尖杉酯碱( 0.12 μg/ml )、卡铂( 20.0 μg/ml )、阿糖胞苷( 6.0 μg/ml )、甲氨蝶呤( 120.0 μg/ml )、环磷酰胺( 48.0 μg/ml )、米托蒽醌( 0.40 μg/ml )作用24h，与未加药K562细胞比较，CYP3A5酶活性均增加，差异具有显著统计学意义。 结论：不同化疗药均可诱导K562细胞CYP3A5酶活性的增加。 第二部分：急性白血病患者骨髓原代细胞CYP3A5酶活性检测 检测88例急性白血病患者原代细胞CYP3A5酶活性，均值为0.142±0.083，原发耐药组、继发耐药组、敏感组分别为：0.108±0.0310、0.046±0.0214、0.023±0.0051，敏感组与原发耐药组比较，差异具有统计学意义。 结论：急性白血病患者骨髓原代细胞具有CYP3A5代谢活性，且与耐药有相关性。 关键词：白血病，CYP3A5基因，药物代谢酶，耐药，高压液相色谱 DETECT CYP3A5 ACTIVITIES IN LEUKEMIA CELL WITH HPLC ASSAY ABSTRACT Up to now, chemotherapy is still the principal means for curing the acute leukemia, however there is great heterogeneity in the way individuals respond to the standard dose chemotherapy. Some patients resistant to chemotherapy or someone get serious side effects or toxicity. This is an important problem. It may lead to a failure therapy or bad clinical outcome. Among those potential causes for such variability, drug metabolism is a key link. Drug-metabolizing enzymes (DMEs) play central roles in the detoxification of xenobiotics or foreign compounds. As one of the most important DMEs, human cytochrome P450 (CYP) 3A gene subfamily is responsible for the metabolism of more than 60% of clinically used drugs (including anticancer agents). The enzymatic activities of CYP3A5 gene induced by different drugs in K562 cell line were obtained by hydrocortisone 6β-hydroxylation with HPLC assay，and this method can also detect CYP3A5 activities of AL bone marrow leukemia cell. Part one: Enzymatic activity of CYP3A5 gene induced by drugs in K562 cell line We selected K562