Radioligand-Binding Assay Employing P-Glycoprotein-Overexpressing Cells: Testing Drug Affinities to the Secretory Intestinal Multidrug Transporter
Radioligand-Binding Assay Employing P-Glycoprotein-Overexpressing Cells: Testing Drug Affinities to the Secretory Intestinal Multidrug Transporter
复制标题
采用 P-糖蛋白过表达细胞的放射性配体结合测定：测试对分泌性肠道多药物转运蛋白的药物亲和力
DOI：
10.1023/a:1011965707998
复制
发表时间：
1998
影响因子：
3.7
通讯作者：
P. Langguth
中科院分区：
文献类型：
--
作者：
Stephan Döppenschmitt;H. Spahn‐Langguth;C. Regårdh;P. Langguth
目的。建立一种快速、可靠的测定传统及新合成化合物与P-gp亲和力的方法。
方法.将放射性配体结合分析法的原理应用于人小肠P-gp的测定。从人癌细胞系Caco-2中获得受体蛋白，其中在细胞生长抑制药物长春碱存在下通过生长细胞诱导P-gp的过表达。选择3H-维拉帕米作为放射性配体。
结果证明了~3H-维拉帕米作为放射性配体与P-gp结合的饱和性和特异性。从浓度依赖性的放射性配体的各种非标记的P-gp配体的位移，亲和常数P-gp结合位点进行了计算。所获得的结合结果与先前发表的结果一致，其中已进行了细胞单层的流入和流出实验，以在功能上表征P-gp-药物相互作用。
结论.建立了一种基于P-gp过表达的Caco-2细胞的放射性配体结合试验。该方法适用于药物与人P-gp相互作用的高通量筛选。它将允许药物与人多药转运蛋白的相互作用的建模，也有可能作为一个高通量筛选工具，以检测化合物倾向于P-gp介导的肠道分泌和潜在的P-gp相关的药物/药物相互作用的药物发现和早期开发。
AbstractPurpose. To develop a rapid and reliable system for affinity determination of conventional as well as newly synthesized compounds to P-gp.
Methods. The principles of radioligand-binding assay were adapted to the human intestinal P-gp. Acceptor protein was obtained from the human carcinoma cell line Caco-2, where overexpression of P-gp was induced by growing cells in the presence of the cytostatic drug vinblastine. 3H-Verapamil was chosen as radioligand.
Results. The saturability and specificity of 3H-verapamil as the radioligand for the binding to P-gp was demonstrated. From concentration dependence of displacement of the radioligand by various non-labeled ligands for P-gp, affinity constants to P-gp binding sites were calculated. The binding results obtained were in agreement with those published earlier where influx and efflux experiments with cell monolayers had been conducted in order to functionally characterize the P-gp -drug interaction.
Conclusions. A radioligand-binding assay on the basis of P-gp overexpressing Caco-2 cells has been developed. The method might be suitable for high-throughput screening of drug interaction with human P-gp. It will allow modeling of the interaction of drugs with the human multi-drug transporter and has also the potential to serve as a high-throughput screening tool to detect compounds prone to P-gp mediated intestinal secretion and potential P-gp related drug/drug interactions in drug discovery and early development.
脂质对纯化的中国仓鼠 P-糖蛋白 ATP 酶活性的影响。
影响因子：
3.9
作者：
Urbatsch,IL;Senior,AE
通讯作者：
Senior,AE