MECHANISM OF ATP DEPENDENT TRANSPORT BY MRP1 PROTEIN

MRP1 蛋白依赖 ATP 的转运机制

• 批准号: 6833474
• 负责人: Xiu-Bao Chang
• 金额: $ 24.1万
• 依托单位: MAYO CLINIC ARIZONA
• 依托单位国家: 美国
• 财政年份: 2001
• 资助国家: 美国
• 起止时间: 2001-01-01 至 2006-04-28
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6833474
• 关键词: P glycoprotein; active sites; adenosine triphosphate; adenosinetriphosphatase; binding sites; cell line; chemical binding; enzyme activity; enzyme inhibitors; monoclonal antibody; multidrug resistance; phosphoproteins; protein kinase; site directed mutagenesis

Overexpression of P-glycoprotein (P-gp) and/or multidrug resistance-associated protein (MRP1) is associated with resistance of tumors to a wide range of chemotherapeutic drugs. Many cancers initially respond well to chemotherapeutic treatment, but these tumors eventually become resistant to those cytotoxic drugs. Some other cancers are resistant to anticancer drugs, even at the beginning of treatment. Therefore, no matter whether the resistance is "acquired" or "intrinsic," multidrug-resistance is one of the major obstacles to the successful treatment of many types of cancers. The mechanism of multidrug resistance conferred by either P-gp or MRP1 involves extrusion of the drugs out of the tumor cells. Both P-gp and MRP1 are members of the ABC superfamily of transport proteins, typically containing two membrane-spanning domains and two nucleotide binding domains. However, MRP1 differs from P-gp in that it contains an extra membrane-spanning domain at the N-terminus. ATP (binding and hydrolysis) is required in both cases. However, the transport steps differ in that P-gp extrudes the unmodified drugs directly, while the drugs transported by MRP1 protein require the presence of a hydrophilic compound, e.g., glutathione- or glucuronide-conjugates. Since the mechanism of multidrug resistance conferred by P-gp or MRP1 is to reduce the level of drug accumulation inside of the cell, inhibiting of the function of P-gp or MRP1 may reverse the drug resistance. Therefore, much effort is being devoted to discover specific inhibitors of these pumps. Present candidates as modulators of the process include verapamil, cyclosporin A, Cremophor, ardeemins, PSC833, rifamycins, RU486, MS-209, non-steroidal inflammatory drugs, acrolein, pyridine analogues, ONO-1078, chloroacetaldehyde, imidazothiazole derivatives, and even some protein kinase inhibitors. However, in order to develop well-designed specific modulators, it is important to understand the mechanisms of drug transport carried out by each individual protein. The first specific aim is to further characterize the ATP finding/hydrolysis and substrate transport by MRP1 protein. The second aim is to determine whether MRP1 protein phosphorylates itself or is a substrate for other protein kinases. This is based on our hypotheses that the phosphorylated MRP1 protein is an intermediate (i.e., there is an autophosphorylation) during ATP hydrolysis and substrate transport. The third aim is to define the substrate binding site(s). New insights gained from these aims should provide the basis for novel means of combating multidrug resistance.

P-糖蛋白（P-GP）和/或多药的过表达 抗性相关蛋白（MRP1）与肿瘤的抗性有关 多种化学治疗药物。许多癌症最初对 化学治疗方法，但这些肿瘤最终对 那些细胞毒性药物。其他一些癌症对抗癌药有抵抗力， 即使在治疗开始时。因此，无论抵抗是否 是“获得的”或“固有的”，多药耐药是主要的之一 成功治疗多种癌症的障碍。机制 由P-gp或MRP1赋予的多药电阻涉及挤出 这些药物从肿瘤细胞中出来。 P-GP和MRP1都是ABC的成员 运输蛋白的超家族，通常包含两个膜跨度 域和两个核苷酸结合结构域。但是，MRP1与p-gp不同 它在N末端包含一个额外的跨膜域。 ATP 在两种情况下都需要（结合和水解）。但是，运输 步骤不同，P-gp直接挤出了未修饰的药物，而 MRP1蛋白运输的药物需要存在亲水性的药物 化合物，例如谷胱甘肽或葡萄糖醛酸 - 偶联物。由于机制 P-gp或MRP1赋予的多药电阻是降低药物的水平 细胞内部的积累，抑制P-gp或MRP1功能的功能可能 逆转耐药性。因此，正在努力 发现这些泵的特定抑制剂。将候选人作为调节器 该过程包括Verapamil，Cyclosporin A，Cremophor，Ardeemins，PSC833， 利福米霉素，RU486，MS-209，非甾体炎症药物，丙烯醛，吡啶 类似物，Ono-1078，氯乙醛醛，咪唑硫氰酸衍生物，甚至 一些蛋白激酶抑制剂。但是，为了开发精心设计的 特定的调节剂，重要的是要了解药物的机制 每个蛋白质进行的运输。第一个具体目的是 进一步表征MRP1的ATP发现/水解和底物运输 蛋白质。第二个目的是确定MRP1蛋白是否磷酸化是否磷酸化 本身或是其他蛋白激酶的底物。这是基于我们的 假设磷酸化的MRP1蛋白是中间体（即 是ATP水解和底物运输过程中的自磷酸化）。这 第三目的是定义底物结合位点。从中获得的新见解 这些目的应该为打击多药的新颖手段提供基础 反抗。