Fluorescence and analytical ultracentrifugation analyses of the interaction of the tyrosine kinase inhibitor, tyrphostin AG1478-mesylate, with albumin
- 作者：Clayton ; Andrew H.A. ; Perugini ; Matthew A. ; Weinstock ; Janet ; Rothacker ; Julie ; Watson ; Keith G. ; et. al.
- 关键词：Analytical ultracentrifugation ; Fluorescence ; Drug binding ; Tyrphostin ; AG1478 ; Tyrosine kinase inhibitor
- 刊名：Analytical Biochemistry
- 出版年：2005
- 期刊代码：93_00032697
- 类别：imm
- 出版时间：July 15, 2005
- 卷：342
- 期：2
- 页码：292-299
- 文件大小：340 K
摘要
Quantifying the interaction of drugs with carrier proteins in plasma is of importance for understanding effective drug delivery to disease-affected tissues. In this study, we employed analytical ultracentrifugation and steady-state fluorescence spectroscopy to characterize the interaction of a potential new anticancer drug, AG1478-mesylate, with plasma proteins in a suspension of normal serum albumin (NSA). We found that mesylate salt of AG1478, an epidermal growth factor receptor kinase inhibitor, sediments in 0.1%(w/v) NSA as a complex with a sedimentation coefficient of 3.8 S. This is consistent with the size of human serum albumin. This interaction was quantitated by meniscus depletion sedimentation and fluorescence titration analyses. AG1478-mesylate binds to albumin with an apparent single-site affinity (Kd) of 120 μM. In this article, we show that the cyclodextrin carrier molecule, Captisol, increases the apparent affinity of the hydrophobic AG1478-mesylate for albumin (Kd = 4–6 μM), and we propose that the AG1478-mesylate–Captisol (1:1) complex binds to albumin with at least 10-fold higher affinity than does AG1478-mesylate ligand alone. A fluorenylmethoxycarbonyl–sulfonic acid (FMS) derivative of the 6-aminoquinazoline analog of AG1478, which was designed to have improved serum-binding properties, was shown by fluorescence analysis to bind with approximately 100-fold greater affinity than the parent compound. This has significant implications in the effective delivery of therapeutic agents in vivo.