Understanding Cisplatin Resistance Mediated by a Copper Efflux Protein

描述（由申请人提供）：顺铂是一种广泛使用的抗癌药物，但由于其内在和获得性细胞耐药，其疗效有限。最近的研究表明，铜稳态蛋白，特别是铜转运泵和伴侣蛋白，可能介导顺铂耐药。这种关系的细节尚不完全清楚。拟议的研究将检查顺铂与铜泵ATP7B（威尔逊病蛋白）分泌途径中的调节铜结合域（MBDs）和人类铜伴侣Atox1之间的相互作用。初步数据表明，顺铂可以直接结合这些蛋白，它们具有共同的折叠，并通过保守的半胱氨酸基序结合Cu(I)。随后的实验将确定产生稳定和均匀的顺铂-蛋白质加合物的条件，确定顺铂结合的化学计量学和铂结合位点的性质，以及它对这些蛋白质整体结构的影响。对一种不太复杂的ATP7B同源物——fulgidus Archaeoglobus CopA的研究将阐明顺铂与MBDs相互作用对铜结合、ATP水解和离子运输的功能影响。在这项工作中获得的信息将提供顺铂和铜转运体之间相互作用的分子水平表征，并可能影响新治疗方法的发展。

DESCRIPTION (provided by applicant): Cisplatin is a widely used anticancer drug but its efficacy is limited due to intrinsic and acquired cellular resistance. Recent studies indicate that copper homeostatic proteins, particularly copper transport pumps and chaperones, may mediate cisplatin resistance. The details of this relationship are not fully understood. The proposed study will examine the interaction between cisplatin and regulatory copper binding domains (MBDs) from the secretory pathway copper pump ATP7B (the Wilson disease protein) and the human copper chaperone Atox1. Preliminary data indicate that cisplatin can directly bind these proteins, all of which share a common fold and bind Cu(I) via conserved cysteine motifs. Subsequent experiments will identify conditions that produce stable and homogenous cisplatin-protein adducts, determine the stoichiometry of cisplatin binding and the nature of the platinum binding site as well as its effect on the overall structure of these proteins. Studies with a less complex ATP7B homolog, Archaeoglobus fulgidus CopA, will elucidate the functional consequences of cisplatin interaction with MBDs for copper binding, ATP hydrolysis, and ion transport. The information gained in this work will provide molecular level characterization of the interaction between cisplatin and copper transporters and could impact the development of new therapeutic approaches. PUBLIC HEALTH RELEVANCE: This project will result in molecular level characterization of the interaction between cisplatin, a widely prescribed anticancer drug, and copper transporters. The information gained will provide insight into the mechanism of drug resistance mediated by copper homeostasis machinery and could impact the development of new therapeutic approaches.