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High-throughput screening and selection of yeast cell lines expressing monoclonal antibodies

  • Original Paper
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Journal of Industrial Microbiology & Biotechnology

Abstract

The methylotrophic yeast Pichia pastoris has recently been engineered to express therapeutic glycoproteins with uniform human N-glycans at high titers. In contrast to the current art where producing therapeutic proteins in mammalian cell lines yields a final product with heterogeneous N-glycans, proteins expressed in glycoengineered P. pastoris can be designed to carry a specific, preselected glycoform. However, significant variability exists in fermentation performance between genotypically similar clones with respect to cell fitness, secreted protein titer, and glycan homogeneity. Here, we describe a novel, multidimensional screening process that combines high and medium throughput tools to identify cell lines producing monoclonal antibodies (mAbs). These cell lines must satisfy multiple selection criteria (high titer, uniform N-glycans and cell robustness) and be compatible with our large-scale production platform process. Using this selection process, we were able to isolate a mAb-expressing strain yielding a titer (after protein A purification) in excess of 1 g/l in 0.5-l bioreactors.

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Authors and Affiliations

  1. Adimab Inc., 16 Cavendish Court, Lebanon, NH, 03766, USA

    Gavin C. Barnard & Piotr Bobrowicz

  2. GlycoFi Inc. (wholly owned subsidiary of Merck & Co, Inc.), 21 Lafayette Street, Lebanon, NH, 03766, USA

    Angela R. Kull, Nathan S. Sharkey, Seemab S. Shaikh, Alissa M. Rittenhour, Irina Burnina, Youwei Jiang, Fang Li, Heather Lynaugh, Teresa Mitchell, Juergen H. Nett, Adam Nylen, Thomas I. Potgieter, Bianka Prinz, Sandra E. Rios, Dongxing Zha, Natarajan Sethuraman & Terrance A. Stadheim

Authors
  1. Gavin C. Barnard
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  2. Angela R. Kull
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  3. Nathan S. Sharkey
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  4. Seemab S. Shaikh
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  5. Alissa M. Rittenhour
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  6. Irina Burnina
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  7. Youwei Jiang
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  8. Fang Li
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  9. Heather Lynaugh
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  10. Teresa Mitchell
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  11. Juergen H. Nett
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  12. Adam Nylen
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  13. Thomas I. Potgieter
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  14. Bianka Prinz
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  15. Sandra E. Rios
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  16. Dongxing Zha
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  17. Natarajan Sethuraman
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  18. Terrance A. Stadheim
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  19. Piotr Bobrowicz
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Corresponding author

Correspondence to Terrance A. Stadheim.

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Barnard, G.C., Kull, A.R., Sharkey, N.S. et al. High-throughput screening and selection of yeast cell lines expressing monoclonal antibodies. J Ind Microbiol Biotechnol 37, 961–971 (2010). https://doi.org/10.1007/s10295-010-0746-1

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  • DOI: https://doi.org/10.1007/s10295-010-0746-1

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