Immunopurification and Mass Spectrometric Quantification of the Active Form of a Chimeric Therapeutic Antibody in Human Serum

详细信息    查看全文

• 作者：Mathieu Dubois ; Fran&ccedil ; ois Fenaille ; Gilles Clement ; Martin Lechmann ; Jean-Claude Tabet ; Eric Ezan ; Fran&ccedil ; ois Becher
• 刊名：Analytical Chemistry
• 出版年：2008
• 出版时间：March 1, 2008
• 年：2008
• 卷：80
• 期：5
• 页码：1737 - 1745
• 全文大小：161K
• 年卷期：v.80,no.5(March 1, 2008)
• ISSN：1520-6882

文摘

In this study, we show that liquid chromatography coupledwith tandem mass spectrometry provides a sensitive,specific, and accurate absolute quantification of Erbitux,a human:murine chimeric mAb used for the treatment ofcolorectal cancer. Micrometric magnetized beads, functionalized with soluble epidermal growth factor receptor(sEGFR), the pharmacological target of Erbitux, were usedfor specific immunocapture of Erbitux allowing assessment of the antibody's biological potency and samplepurification. Following digestion with trypsin, specificpeptides from light and heavy chains were monitored inthe selected reaction monitoring (SRM) mode. Assayvariability below 20% was provided through optimizationof the digestion step and rigorous monitoring of the wholeanalytical process using an appropriate internal standard.The 20 ng/mL lower limit of quantification was similar tothat of ELISA methods. These results show that this massspectrometric approach is a potential alternative forpharmacokinetic evaluation of mAbs during clinical development.