Immunopurification and mass spectrometric quantification of the active form of a chimeric therapeutic antibody in human serum

被引:104
作者
Dubois, Mathieu [1 ]
Fenaille, Francois [1 ]
Clement, Gilles [2 ]
Lechmann, Martin [3 ]
Tabet, Jean-Claude [4 ]
Ezan, Eric [1 ]
Becher, Francois [1 ]
机构
[1] CEA Saclay, Serv Pharmacol & Immunoanalyse, F-91191 Gif Sur Yvette, France
[2] INRA, Lab Immunoallergie Alimentaire, F-91191 Gif Sur Yvette, France
[3] Merck KGaA, D-85567 Merck Serona, Grafing, Germany
[4] Univ Paris 06, CNRS, UMR 7613, Lab Synth Struct & Fonct Mol Bioact, F-75252 Paris, France
关键词
D O I
10.1021/ac7021234
中图分类号
O65 [分析化学];
学科分类号
070302 ; 081704 ;
摘要
In this study, we show that liquid chromatography coupled with tandem mass spectrometry provides a sensitive, specific, and accurate absolute quantification of Erbitux, a human:murine chimeric mAb used for the treatment of colorectal cancer. Micrometric magnetized beads, functionalized with soluble epidermal growth factor receptor (sEGFR), the pharmacological target of Erbitux, were used for specific immunocapture of Erbitux allowing assessment of the antibody's biological potency and sample purification. Following digestion with trypsin, specific peptides from light and heavy chains were monitored in the selected reaction monitoring (SRM) mode. Assay variability below 20% was provided through optimization of the digestion step and rigorous monitoring of the whole analytical process using an appropriate internal standard. The 20 ng/mL lower limit of quantification was similar to that of ELISA methods. These results show that this mass spectrometric approach is a potential alternative for pharmacokinetic evaluation of mAbs during clinical development.
引用
收藏
页码:1737 / 1745
页数:9
相关论文
共 35 条
  • [11] Kinetic characterization of sequencing grade modified trypsin
    Finehout, EJ
    Cantor, JR
    Lee, KH
    [J]. PROTEOMICS, 2005, 5 (09) : 2319 - 2321
  • [12] Improving mass accuracy of high performance liquid chromatography/electrospray ionization time-of-flight mass spectrometry of intact antibodies
    Gadgil, Himanshu S.
    Pipes, Gary D.
    Dillon, Thomas M.
    Treuheit, Michael J.
    Bondarenko, Pavel V.
    [J]. JOURNAL OF THE AMERICAN SOCIETY FOR MASS SPECTROMETRY, 2006, 17 (06) : 867 - 872
  • [13] Effect of the mobile phase composition on the separation and detection of intact proteins by reversed-phase liquid chromatography -: electro spray mass spectrometry
    García, MC
    Hogenboom, AC
    Zappey, H
    Irth, H
    [J]. JOURNAL OF CHROMATOGRAPHY A, 2002, 957 (02) : 187 - 199
  • [14] Gerber Scott A, 2007, Methods Mol Biol, V359, P71
  • [15] LC-MS/MS method for confirmation of recombinant human erythropoietin and darbepoetin α in equine plasma
    Guan, Fuyu
    Uboh, Cornelius E.
    Soma, Lawrence R.
    Birks, Eric
    Chen, Jinwen
    Mitchell, Janis
    You, Youwen
    Rudy, Jeffrey
    Xu, Fran
    Li, Xiaoqing
    Mbuy, Gustave
    [J]. ANALYTICAL CHEMISTRY, 2007, 79 (12) : 4627 - 4635
  • [16] Identification of multiple sources of charge heterogeneity in a recombinant antibody
    Harris, RJ
    Kabakoff, B
    Macchi, FD
    Shen, FJ
    Kwong, M
    Andya, JD
    Shire, SJ
    Bjork, N
    Totpal, K
    Chen, AB
    [J]. JOURNAL OF CHROMATOGRAPHY B, 2001, 752 (02): : 233 - 245
  • [17] Selective quantitative bioanalysis of proteins in biological fluids by on-line immunoaffinity chromatography-protein digestion-liquid chromatography-mass spectrometry
    Hoos, JS
    Sudergat, H
    Hoelck, JP
    Stahl, M
    de Vlieger, JSB
    Niessen, WMA
    Lingeman, H
    Irth, H
    [J]. JOURNAL OF CHROMATOGRAPHY B-ANALYTICAL TECHNOLOGIES IN THE BIOMEDICAL AND LIFE SCIENCES, 2006, 830 (02): : 262 - 269
  • [18] A multiple reaction monitoring method for absolute quantification of the human liver alcohol dehydrogenase ADH1C1 isoenzyme
    Janecki, Dariusz J.
    Bemis, Kerry G.
    Tegeler, Tony J.
    Sanghani, Paresh C.
    Zhai, Lanmin
    Hurley, Thomas D.
    Bosron, William F.
    Wang, Mu
    [J]. ANALYTICAL BIOCHEMISTRY, 2007, 369 (01) : 18 - 26
  • [19] LC-MS development strategies for quantitative bioanalysis
    Jemal, Mohammed
    Xia, Yuan-Qing
    [J]. CURRENT DRUG METABOLISM, 2006, 7 (05) : 491 - 502
  • [20] A strategy of plasma protein quantitation by selective reaction monitoring of an intact protein
    Ji, QC
    Rodila, R
    Gage, EM
    El-Shourbagy, TA
    [J]. ANALYTICAL CHEMISTRY, 2003, 75 (24) : 7008 - 7014