MULTIDRUG RESISTANCE IN MAMMALIAN CELLS

观察到对化疗剂的耐药性的发展， 临床上，并已在组织培养细胞中进行了广泛的研究。一 不寻常的表型是多药耐药，其中细胞， 受到任何一种细胞毒性药物的挑战 不仅对选择剂有抗性，而且对 其他看似无关的化合物这种多药耐药（MDR） 在许多情况下，表型与小的 称为P-糖蛋白（Pgp）的膜糖蛋白家族，其 被认为是ATP激活的外排泵。三大家族中 仓鼠中的成员，它是人类mdr 1的同源物（已知是 在人类肿瘤中过度表达），pgp 1，其最常涉及 随着组织中多药耐药性的建立和维持， 培养细胞系，并且是本申请的主要主题。 关于pgp 1支持的外排机制知之甚少， 建立交叉耐药模式的方式，或如何 通过诸如维拉帕米和环孢菌素A的药物逆转mdr， 办妥了一批也不知道如果有的话， 所有哺乳动物细胞中存在的小pgp 1相关RNA转录物。而 这些问题中的每一个对于pgp 1介导的MDR都是重要的， 多药耐药支持的非PGP相关形式的MDR 相关蛋白（MRP）和拓扑异构酶，也存在，必须 如果我们要扩展我们对肿瘤细胞药物的理解， 阻力利用各种重组DNA技术，基因 转染试验、定点诱变和短期药物治疗 我们提出的选择方案是：1）继续分子遗传学研究 pgp 1中跨膜结构域6的一个区域，该区域已知介导 交叉耐药模式，如果不是直接参与， 与环孢素A逆转机制，2）分析额外的 进一步描述其药物结合的天然存在的Pgp 1突变体 3）确定MDR机制出现的顺序 短期选择使用不同的药物和4）尝试 为了阐明2.3kb poly（A）+ RNA转录物的功能， 是PGP 1的剪接产物。

The development of resistance to chemotherapeutic agents is observed in the clinic, and has been studied extensively in tissue culture cells. One unusual phenotype is that of multidrug resistance, in which cells that are challenged with any one of a variety of cytotoxic drugs develop resistance not only to the selective agent but also cross-resistance to other, seemingly unrelated, compounds. This multidrug-resistant (mdr) phenotype is associated in many cases with the overproduction of a small family of membrane glycoproteins called p-glycoproteins (pgp's), which are thought to act as ATP-activated efflux pumps. Of the three family members in hamster, it is the homolog of human mdr1 (known to be overexpressed in human tumors), pgp1, that is most frequently involved with the establishment and maintenance of multidrug-resistance in tissue culture cell lines, and is the primary subject of this application. Little is known concerning the efflux mechanism supported by pgp1, the manner in which cross-resistance patterns are established, or how reversal of mdr by agents such as Verapamil and Cyclosporin A is achieved. Nor is it understood what function if any is played by the small pgp1 related RNA transcripts present in all mammalian cells. While each of these issues is of importance to pgp1 mediated mdr, it is clear that non-pgp related forms of mdr, supported by multidrug-resistance associated protein (MRP), and topoisomerase, also exist and must be accounted for if we are to extend our understanding of tumor cell drug resistance. Using a variety of recombinant DNA techniques, gene transfection assays, site directed mutagenesis, and short-term drug selection protocols we propose to 1) continue the molecular genetic study of transmembrane domain 6 in pgp1, a region that is known to mediate cross-resistance patterns and to be closely, if not directly, involved with the mechanism of Cyclosproin A reversal, 2) analyze additional naturally occurring mutants of pgp1 to further delineate its drug binding site(s) 3) determine the order of emergence of mdr mechanisms during short term selections using different antineoplastic drugs and 4) attempt to clarify the function of the 2.3kb poly (A)+ RNA transcript thought to be a splicing product of pgp1.