Understanding the mechanism of virus removal by Q sepharose fast flow chromatography during the purification of CHO-cell derived biotherapeutics

Authors

  • Daniel M. Strauss,

    1. Process Research and Development, Genentech, Inc., One DNA Way, South San Francisco, California 94080; telephone: 650-225-1000; fax: 650-225-7592
    Current affiliation:
    1. Bioproduct Research and Development, Lilly Research Laboratories, Eli Lilly and Company, Lilly Technology Center, Indianapolis, IN 46221.
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  • Scott Lute,

    1. Division of Monoclonal Antibodies, Center for Drug Evaluation and Research, Food and Drug Administration, Silver Spring, Maryland
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  • Zinaida Tebaykina,

    1. Process Research and Development, Genentech, Inc., One DNA Way, South San Francisco, California 94080; telephone: 650-225-1000; fax: 650-225-7592
    Current affiliation:
    1. Department of Microbiology & Immunology, University of British Columbia, Vancouver, BC, Canada.
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  • Douglas D. Frey,

    1. Department of Chemical and Biochemical Engineering, University of Maryland Baltimore County, Baltimore, Maryland
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  • Cintia Ho,

    1. Process Research and Development, Genentech, Inc., One DNA Way, South San Francisco, California 94080; telephone: 650-225-1000; fax: 650-225-7592
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  • Gregory S. Blank,

    1. Process Research and Development, Genentech, Inc., One DNA Way, South San Francisco, California 94080; telephone: 650-225-1000; fax: 650-225-7592
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  • Kurt Brorson,

    1. Division of Monoclonal Antibodies, Center for Drug Evaluation and Research, Food and Drug Administration, Silver Spring, Maryland
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  • Qi Chen,

    1. Process Research and Development, Genentech, Inc., One DNA Way, South San Francisco, California 94080; telephone: 650-225-1000; fax: 650-225-7592
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  • Bin Yang

    Corresponding author
    1. Process Research and Development, Genentech, Inc., One DNA Way, South San Francisco, California 94080; telephone: 650-225-1000; fax: 650-225-7592
    • Process Research and Development, Genentech, Inc., One DNA Way, South San Francisco, California 94080; telephone: 650-225-1000; fax: 650-225-7592.
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  • Statements in this article represent the views of the authors and do not constitute official positions or policies of the Food and Drug Administration or the U.S. Government.

Abstract

During production of therapeutic monoclonal antibodies (mAbs) in mammalian cell culture, it is important to ensure that viral impurities and potential viral contaminants will be removed during downstream purification. Anion exchange chromatography provides a high degree of virus removal from mAb feedstocks, but the mechanism by which this is achieved has not been characterized. In this work, we have investigated the binding of three viruses to Q sepharose fast flow (QSFF) resin to determine the degree to which electrostatic interactions are responsible for viral clearance by this process. We first used a chromatofocusing technique to determine the isoelectric points of the viruses and established that they are negatively charged under standard QSFF conditions. We then determined that virus removal by this chromatography resin is strongly disrupted by the presence of high salt concentrations or by the absence of the positively charged Q ligand, indicating that binding of the virus to the resin is primarily due to electrostatic forces, and that any non-electrostatic interactions which may be present are not sufficient to provide virus removal. Finally, we determined the binding profile of a virus in a QSFF column after a viral clearance process. These data indicate that virus particles generally behave similarly to proteins, but they also illustrate the high degree of performance necessary to achieve several logs of virus reduction. Overall, this mechanistic understanding of an important viral clearance process provides the foundation for the development of science-based process validation strategies to ensure viral safety of biotechnology products. Biotechnol. Bioeng. 2009; 104: 371–380 © 2009 Wiley Periodicals, Inc.

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