ALTERED TOPOISOMERASE IN ATYPICAL MULTIDRUG RESISTANCE

多药耐药(MDR)是指肿瘤细胞 可对多种“天然”表现出广泛的交叉抗性 “产品”化合物的结构和机理大相径庭。 行动。一种类型的MDR(PGP-MDR)的特征是 减少药物积聚和编码基因的过度表达 对于一种大的膜糖蛋白，称为Pgp。我们已经开发出 对VM-26渐进性耐药的人白血病细胞系(CEM/VM-1 和CEM/Vm-1-5)具有交叉耐药的“非典型”模式 细胞药理学。这些细胞对长春新碱保持敏感。 生物碱，在药物的运输过程中不会受到损害，也不会 过表达Pgp基因。现在有大量证据表明 “非典型MDR”(at-MDR)是由于 关键的核酶，拓扑异构酶II，一种独特的 是许多抗癌药物的重要靶标。这是不是 改变是编码基因突变的结果 拓扑异构酶II或在调节它的基因中尚不清楚。这个 拟议工作的长期目标是确定 AT-MDR的分子基础。 为了实现这一目标，将解决五个具体目标。这个 第一个目标是纯化和鉴定拓扑异构酶II 来自at-mdr细胞的蛋白质。第二个目标是描述 拓扑异构酶II在AT-MDR细胞中的表达及其调控 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基因的特定突变 使用核酸酶作图技术和cDNA序列测定。 第四个目标是确认在 AT-MDR拓扑异构酶II确实是该基因表达的原因 At-mdr表型。最后，如果拓扑异构酶II的mRNA和 酶在耐药细胞中是不变的，第五个目标是 以生物化学和分子程序为特征的性质 拓扑异构酶的调节因子II。 AT-MDR细胞中突变的特征 负责拓扑异构酶II活性变化的Will 提供有关临床重要疾病相互作用的信息 含有关键酶的药物，可能会提供相关信息 拓扑异构酶II的作用机制，并可能提供 对新药和治疗模式设计的见解 以克服这种形式的抵抗。

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.