Cisplatin resistance mediated by cooper export pathways

描述（由申请人提供）：铜（Cu）出口物ATP7A和ATP7B通过改变其细胞内隔离和外排来调节肿瘤细胞对含铂药物的敏感性的能力现已在多个实验系统中得到证实。本项目的总体目标是确定ATP7A和ATP7B介导顺铂、卡铂和奥沙利铂从卵巢癌细胞外排的机制。我们的假设是，在临床相关浓度下，铂药物进入细胞，分布到不同的亚细胞区室，并通过转运蛋白和伴侣蛋白从细胞输出，这些转运蛋白和伴侣蛋白已经进化到控制Cu稳态。这一假设的一个推论是，对于铜，这些pi型atp酶通过将Pt药物隔离到分泌途径的囊泡中或附着在囊泡上来解毒，最终从细胞中输出。具体目的是：1)确定ATP7A和ATP7B是否与顺铂、卡铂和奥沙利铂结合并转运到分泌囊泡中；2)研究铂类药物与ATP7A、ATP7B和ATOX1蛋白金属结合序列中cu结合基序的结合能力；3)确定Cu伴侣ATOX1是否对ATP7A和ATP7B在活细胞中介导Pt耐药、隔离和输出的能力至关重要。仔细分析铜外排转运体调节铂药物输出的机制，有望深入了解顺铂、卡铂和奥沙利铂在疗效和毒性方面存在差异的原因，并确定提高这些药物治疗指数的策略。这些研究还将进一步阐明介导对这类重要化疗药物耐药的机制，并提出逆转耐药的临床相关策略。

DESCRIPTION (provided by applicant): The ability of the copper (Cu) exporters ATP7A and ATP7B to regulate tumor cell sensitivity to the platinum-containing drugs by altering their intracellular sequestration and efflux has now been demonstrated in multiple experimental systems. The overall goal of this project is to determine the mechanism by which ATP7A and ATP7B mediate the efflux of cisplatin, carboplatin and oxaliplatin from ovarian carcinoma cells. Our hypothesis is that at clinically relevant concentrations, the platinum drugs enter the cell, are distributed to various subcellular compartments and are exported from the cell using transporters and chaperones that have evolved to control Cu homeostasis. A corollary to this hypothesis is that, as for Cu, these PI-type ATPases function to detoxify the Pt drugs by sequestering them into or onto vesicles of the secretory pathway that are eventually exported from the cell. The specific aims are to: 1) determine whether ATP7A and ATP7B bind and transport cisplatin, carboplatin and oxaliplatin into secretory vesicles; 2) investigate the ability of the Pt drugs to bind to the Cu-binding motif in the metal binding sequences of ATP7A, ATP7B and ATOX1 proteins; and, 3) determine whether the Cu chaperone ATOX1 is essential to the ability of ATP7A and ATP7B to mediate Pt drug resistance, sequestration, and export in living cells. Careful dissection of the mechanism by which the Cu efflux transporters modulate the export of the Pt drugs is expected to offer insight into why cisplatin, carboplatin and oxaliplatin differ in their efficacy and toxicity, and to identify strategies for improving the therapeutic index of these agents. These studies will also further elucidate the mechanisms that mediate resistance to this important class of chemotherapeutic agents and suggest clinically relevant strategies for reversing resistance.