Prediction of anticancer effect of 1-p-D-arabinofuranosylcytostee by sensitive monitoring of its intracellular active metabolite in leukemic cells

1.阿糖胞苷胞内活性代谢产物ara-CTP的药代动力学研究。用新建立的灵敏方法测定了25例白血病患者接受低剂量或常规剂量阿糖胞苷或对数作用阿糖胞苷、BHAC治疗后细胞内ara-CTP的浓度。ara-CTP浓度因给药方法、剂量和患者而异，并且不能从血浆ara-C浓度预测。由于血浆ara-C的维持对于ara-CTP在细胞中的保留是重要的，因此连续输注ara-C和BHAC有效地产生ara-CTP。在BHAC治疗中，完全缓解的患者比未缓解的患者获得更大的ara-CTP量，这表明ara-CTP将是BHAC治疗效果的关键参数。骨髓抑制与血浆阿糖胞苷水平相关，提示正常造血干细胞对阿糖胞苷的敏感性在个体间相似.对DNA中掺入的阿糖胞苷的检测方法进行了探讨， 关于我们 r ara-C掺入DNA的方法。用阿糖胞苷处理白血病细胞后，从其酸不溶性部分中分离出DNA。将DNA酶消化为包括ara-C的核苷。通过高效液相色谱法分离ara-C，然后冻干。各步回收率均在90%以上。用抗阿糖胞苷血清对分离的样品进行放射免疫分析，以确定其阿糖胞苷浓度.用计算机控制的体外药代动力学模拟系统评价细胞毒作用，比较药代动力学模拟条件与传统培养系统条件下的细胞毒作用。K562细胞与模拟阿糖胞苷孵育2、4、8和16 h后的存活率表明，阿糖胞苷的细胞毒性具有时间依赖性。相反，在常规培养系统下，未观察到时间依赖性抑制。类似地，输注柔红霉素0.5、2、4和8小时的模拟显示柔红霉素的细胞毒性作用是浓度依赖性的。

1. Pharmacokinetic study of ara-CTP, an intracellular active metabolite of ara-C.ara-CTP was measured in leukemic cells by the newly established sensitive method in 25 leukemic patients receiving ara-C or log-acting ara-C, BHAC at low or conventional doses. The ara-CTP concentrations differed by the administration methods, doses, and patients, and were not predicted from the plasma ara-C concentrations. As the maintenance of the plasma ara-C was important for the retention of ara-CTP in the cell, continuous infusion of ara-C and BHAC produced ara-CTP efficiently. In BHAC therapy, patients with complete remission achieved greater ara-CTP amounts than those without remission, suggesting that ara-CTP would 6e a crucial parameter for the therapeutic efficacy of BHAC. Myelosuppression was correlated to the plasma ara-C level, suggesting that normal hematopoietic stem cells have similar sensitivity to ara-C among individuals.2. Detection of ara-C incorporated into DNA.The detection method fo … More r ara-C incorporated into DNA was established. DNA was separated from acid insoluble fraction of leukemic cells after treatment with ara-C. The DNA was digested enzymatically to nucleosides that included ara-C. ara-C was isolated by high performance liquid chromatography, followed by liophilization. The recovery of each step was over 90 %. Radioimmunoassay will be applied to the isolated sample using anti-ara-C serum to confirm its ara-C concentration.3. Cytotpxic effects evaluated by a new computer-controlled in vitro pharmacokinetic simulation system.Cytotoxicity was compared between a pharmacokinetically simulated condition and a conventional culture system condition. The survival rates of the cell line K562 incubated with the simulated ara-C infusions for 2, 4, 8, and 16 h demonstrated that the cytotoxicity of ara-C was time-dependent. In contrast, under a conventional culture system, no time-dependent inhibition was observed. Similarly, the simulations of the infusion of daunorubicin for 0.5, 2, 4, and 8 h revealed that the cytotoxic effect of daunorubicin was concentration-dependent Less