Development of a fluorescence liposomal ABCG2 Multidrug Transporter assay

荧光脂质体 ABCG2 多药物转运蛋白测定的开发

• 批准号: 8644055
• 负责人: DONALD Lee MELCHIOR
• 金额: $ 36.36万
• 依托单位: GLSYNTHESIS, INC.
• 依托单位国家: 美国
• 财政年份: 2014
• 资助国家: 美国
• 起止时间: 2014-05-07 至 2016-05-06
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8644055
• 关键词: ABCG2 gene; ATP phosphohydrolase; Adverse effects; Antineoplastic Agents; Biological Assay; Carrier Proteins; Clinical; Detergents; Development; Drug Efflux; Drug Industry; Drug Transport; Drug or chemical Tissue Distribution; Drug resistance; Excision; Fluorescence; Future; Goals; Housing; Human; In Vitro; Industry; International; Legal patent; Letters; Lipid Bilayers; Lipids; Liposomes; Malignant Neoplasms; Marketing; Materials Testing; Measurement; Mediating; Oral; P-Glycoprotein; Paper; Pharmaceutical Preparations; Predisposition; Prevalence; Production; Proteins; Protocols documentation; Pump; Reader; Reagent; Resistance; Resistance development; Robotics; Series; Services; Staging; Substrate Specificity; System; Techniques; Testing; Time; Tissues; United States; absorption; aqueous; base; chemotherapeutic agent; chemotherapy; drug candidate; drug development; drug discovery; expectation; high throughput screening; human ABCG2 protein; in vitro Assay; inhibitor/antagonist; instrumentation; multi drug transporter; multidrug transport; new technology; novel; oncology; programs; public health relevance; reconstitution; research study; scale up; sensor; tool; tumor; uptake

ABSTRACT In this project we propose to exploit the recent successful isolation and reconstitution of the important ATP- dependent drug efflux transporter, ABCG2 (BCRP, Breast Cancer Resistance Protein), and combine it with a new transport assay system, the Fluorosome platform, to characterize the interaction of ABCG2 with drugs and drug candidates. The result will provide a useful, novel, highly specific and rapid in vitro transport assay for future general use. We shall employ this assay together with ATPase studies to characterize a wide variety of ABCG2 substrates and modulators. Additionally, by means of this unique construct we shall screen the 89 anticancer drugs in the NCI Oncology Drug Set Plate series for inhibition of ABCG2-mediated transport. The ABCG2 protein is a ubiquitous high capacity drug transporter with wide substrate specificity. The "White Paper" recently issued by the International Transporter Consortium formed to propose industry and FDA standards for studies of drug:transporter interactions lists ABCG2 as second in importance to P-glycoprotein (Pgp) among those drug transporters involved in clinical absorption and disposition of drugs. Drug effects are often modified by the highly variable oral uptake of drugs and by limited tissue distribution. Tumors frequently develop resistance against treatment with multiple chemotherapeutic agents. The consequence of this is the expectation that systemic chemotherapy of nearly one-half of the one million new cancer cases annually in the United States will fail due to the resistance of tumors to drugs. A major factor in this resistance is the prevalence of energy-driven efflux transporter proteins which actively pump drugs out of their target tissues. In addition, many side effects and incompatibilities accompanying multidrug use result from the interference of one drug with another's susceptibility to active efflux. The recent successful production, isolation, and reconstitution of the ABCG2 transporter combined with new technology employing the Fluorosome platform will provide an effective in vitro assay to determine the susceptibility of drug candidates to extrusion from their target tissue by the ABCG2 transporter and to determine if compounds interact with this transporter. These studies will employ a novel reagent "Fluorosome-trans-abcg2" that is unambiguously specific for the ABCG2 transporter, applicable to a wide range of compounds, uses very small amounts of test material, and, with respect to instrumentation, requires only a standard injecting multiwell fluorescence plate reader. The assay will be amenable to moderate and high throughput screening.

抽象的 在这个项目中，我们建议利用最近成功分离和重建重要的 ATP- 依赖性药物外排转运蛋白 ABCG2（BCRP，乳腺癌耐药蛋白），并将其与新的药物相结合 转运分析系统，Fluorosome 平台，用于表征 ABCG2 与药物以及药物的相互作用 候选人。该结果将为未来提供一种有用、新颖、高度特异性和快速的体外转运测定方法。 一般用途。我们将采用该测定法与 ATP 酶研究一起来表征各种 ABCG2 底物和调制器。此外，通过这种独特的构建，我们将筛选 89 种抗癌药物 NCI 肿瘤药物套装板系列中用于抑制 ABCG2 介导的转运的药物。 ABCG2 蛋白是一种普遍存在的高容量药物转运蛋白，具有广泛的底物特异性。的“白 国际运输联盟最近发布的论文”，旨在向行业和 FDA 提出建议 药物与转运蛋白相互作用研究标准将 ABCG2 列为仅次于 P-糖蛋白的第二位 (Pgp) 参与临床药物吸收和处置的药物转运蛋白。 药物的作用常常因药物的口服摄取的高度可变性和有限的组织分布而改变。 肿瘤经常对多种化疗药物产生耐药性。这 这样做的结果是，预计一百万新患者中的近二分之一将接受全身化疗 美国每年的癌症病例都会因肿瘤对药物的耐药性而失败。这其中的一个主要因素 耐药性是能量驱动的外排转运蛋白的普遍存在，这些蛋白主动将药物从其细胞中泵出 靶组织。此外，伴随多种药物使用的许多副作用和不相容性是由 一种药物干扰另一种药物对主动外排的敏感性。 最近成功生产、分离和重构了 ABCG2 转运蛋白，并与新的 采用 Fluorosome 平台的技术将提供有效的体外测定来确定 候选药物对 ABCG2 转运蛋白从其靶组织中挤出的敏感性，并确定是否 化合物与该转运蛋白相互作用。这些研究将采用一种新型试剂“Fluorosome-trans-abcg2” 明确针对 ABCG2 转运蛋白，适用于多种化合物，使用非常广泛 少量的测试材料，并且就仪器而言，仅需要标准的注射多孔 荧光读板机。该测定将适合中度和高通量筛选。