ALTERED TOPOISOMERASE IN ATYPICAL MULTIDRUG RESISTANCE

多药耐药（MDR）是指肿瘤细胞 可以显示出对多种"天然"的广泛的交叉抗性 产品"化合物的广泛不同的结构和机制， 行动上 一种类型的MDR（Pgp-MDR）的特征在于： 减少药物蓄积和基因编码的过度表达 一个大的膜糖蛋白，称为Pgp。 我们已经开发 VM-26耐药白血病细胞系CEM/Vm-1 和CEM/VM-1 - 5），具有"非典型"交叉耐药模式， 细胞药理学 这些细胞保持对神经细胞的敏感性。 生物碱，在药物的运输中不受损害， 过度表达Pgp基因。 现在有大量证据表明， "药物-MDR"（at-MDR）是由于药物活性的改变， 必需的核酶，拓扑异构酶II，一种独特的， 是许多抗癌剂重要靶点。 这是否 改变是基因编码突变的结果， 拓扑异构酶II或调节它的基因尚不清楚。 的 拟议工作的长期目标是确定 at-MDR的分子基础。 为实现这一目标，将实现五个具体目标。 的 第一个目的是纯化和表征拓扑异构酶II 来自at-MDR细胞的蛋白质。 第二个目的是描述 拓扑异构酶Ⅱ mRNA及其在at-MDR细胞中的调控 拓扑异构酶II特异性核苷酸探针。 第三个目标是 鉴定at-MDR拓扑异构酶II mRNA中的特定突变 使用核酸酶作图技术和cDNA序列测定。 第四个目的是确认， at-MDR拓扑异构酶II确实负责表达 耐多药表型。 最后，如果拓扑异构酶II mRNA和 酶在抗性细胞中不变，第五个目的是 通过生物化学和分子程序表征 改变的拓扑异构酶II调节剂 表征at-MDR细胞中的突变， 负责拓扑异构酶II活性改变的 产生关于临床重要的 关键酶的药物，可以提供相关信息 拓扑异构酶II的作用机制，并可能提供 对新药和治疗方式设计的见解 克服这种形式的阻力。

Multiple drug resistance (MDR) refers to the fact that tumor cells can display a broad cross-resistance to a variety of "natural product" compounds of widely differing structure and mechanism of action. One type of MDR (Pgp-MDR) has been characterized by decreased drug accumulation and overexpression of the gene coding for a large membrane glycoprotein, termed, Pgp. We have developed progressively-VM-26-resistant human leukemic cell lines (CEM/Vm-1 and CEM/VM-1-5) with an "atypical" pattern of cross-resistance and cellular pharmacology. These cells retain sensitivity to the Vinca alkaloids, are not impaired in their transport of drugs, and do not overexpress the pgp gene. There is now considerable evidence that "atypical-MDR" (at-MDR) is due to an alteration in the activity of the essential nuclear enzyme, topoisomerase II, an unique and important target for many anticancer agents. Whether this alteration is a consequence of a mutation in the gene coding for topoisomerase II or in a gene that modulates it is not clear. The long-term objective of the proposed work is to determine the molecular basis of at-MDR. To accomplish this goal, five specific aims will be addressed. The first aim is to purify and characterize the topoisomerase II protein from at-MDR cells. A second aim is to characterize the topoisomerase II mRNA and its regulation in the at-MDR cells using a topoisomerase II-specific nucleotide probe. The third aim is to identify specific mutations in the at-MDR topoisomerase II mRNA using nuclease mapping techniques and cDNA sequence determinations. The fourth aim is to confirm that any identified alterations in the at-MDR topoisomerase II are indeed responsible for the expression of the at-MDR phenotype. Finally, if the topoisomerase II mRNA and enzyme is unaltered in the resistant cells, the fifth aim is to characterize by biochemical and molecular procedures the nature of the altered modulator of topoisomerase II. Characterization of mutations in the at-MDR cells that are responsible for the alterations in topoisomerase II activity will yield information about the interaction of clinically important drugs with the critical enzyme, may provide information pertinent to the mechanism of action of topoisomerase II, and may provide insights into the design of new drugs and therapeutic modalities to overcome this form of resistance.