MOLECULAR MECHANISMS OF ARA C THERAPY IN MAN

我们先前已经证明了Ara-C整合到白血病细胞中 DNA和这种结合的程度与失去 克隆生存。我们还证明了公司的 阿拉伯糖基残基是一种较差的引物末端，因此导致 抑制DNA合成。(Ara-C)DNA结构异常， 暴露在碱中时会发生链断裂。此外，抑制 通过掺入Ara-C而导致DNA合成的改变导致 DNA在修复和复制合成过程中的链断裂。最后， (Ara-C)DNA的形成导致末端的诱导 分化和致命的细胞事件。 这些研究结果为进一步深入了解银杏叶提取物的作用机制提供了新的思路。 治疗急性髓系白血病最有效的药物。 拟议的研究将延长我们关于纳入Ara-C的工作 通过监测DNA修复过程中Ara-C的掺入来进行复制合成。 (Ara-C)DNA在复制和复制过程中的生物化学效应 然后，修复合成将作为暴露于 3-氨基苯甲酰胺和咖啡因。分子参数将会相互关联 其生物学效应由诱变和克隆丧失决定 生死存亡。最后，我们将使用一种新的体外后标记试验来 监测临床标本中(Ara-C)DNA的形成。这种方法将是 适用于从接受低剂量连续注射的患者获得的标本 输液C区与我们的生物和生化的临床相关性 学习。 这些研究将在HL-60早幼粒母细胞和 密度受阻的平台期二倍体成纤维细胞。成立为法团 在复制和修复DNA合成过程中的Ara-C将通过 平衡离心和DNA损伤将使用 碱性洗脱技术。这种方法应该提供分子 了解Ara-C在致死诱导中作用的基础 细胞事件。这项工作对于提供实验数据具有重要意义 阿糖胞苷设计生化合理临床试验的基础 以及了解人类白血病的分子控制。

We have previously demonstrated that ara-C incorporates in leukemic cell DNA and that the extent of this incorporation correlates with loss of clonogenic survival. We have also demonstrated that the incorporated arabinosyl residue serves as a poor primer terminus and thereby results in inhibition of DNA synthesis. The (ara-C)DNA is structurally abnormal and undergoes strand scission upon exposure to alkali. Furthermore, inhibition of DNA synthesis by the incorporation of ara-C causes the accumulation of strand breaks in DNA undergoing repair and replicative synthesis. Finally, the formation of (ara-C)DNA results in both the induction of terminal differentiation and lethal cellular events. These findings provide major new insights into the mechanism of action of the most effective agent in the treatment of acute myelogenous leukemia. The proposed studies will extend our work on incorporation of ara-C during replicative synthesis by monitoring ara-C incorporation during DNA repair. The biochemical effects of (ara-C)DNA formation during both replicative and repair synthesis will then be monitored as a result of exposue to 3-aminobenzamide and caffeine. The molecular parameters will be correlated with biologic effect as determined by mutagenesis and loss of clonogenic survival. Finally, we will employ a new in vitro post-labeling assay to monitor formation of (ara-C)DNA in clinical samples. This approach will be applied to speciments obtained from patients receiving low dose continuous infusion area-C as a clinical correlate of our biologic and biochemical studies. These studies will be performed on HL-60 promyeloblasts and density-arrested, plateau phase diploid fibroblasts. The incorporation of ara-C during replicative and repair DNA synthesis will be monitored by equilibrium centrifugation and DNA damage will be monitored using the alkaline elution technique. This approach should provide the molecular basis for understanding the effects of ara-C on the induction of lethal cellular events. This work is important in providing the experimental basis for the design of biochemically rational clinical trials with ara-C and for gaining an understanding of the molecular control of human leukemia.