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Metabolism of PER.C6TM cells cultivated under fed-batch conditions at low glucose and glutamine levels

Authors

  • Luis Maranga,

    Corresponding author
    1. Fermentation and Cell Culture, Bioprocess R&D, Merck Research Laboratories, Merck & Co., Inc., 770 Sumneytown Pike, WP17-201 P.O. Box 4, West Point, Pennsylvania 19486; telephone: ++1-215-652-8636; fax: ++1-215-993-4884
    • Fermentation and Cell Culture, Bioprocess R&D, Merck Research Laboratories, Merck & Co., Inc., 770 Sumneytown Pike, WP17-201 P.O. Box 4, West Point, Pennsylvania 19486; telephone: ++1-215-652-8636; fax: ++1-215-993-4884
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  • Charles F. Goochee

    1. Fermentation and Cell Culture, Bioprocess R&D, Merck Research Laboratories, Merck & Co., Inc., 770 Sumneytown Pike, WP17-201 P.O. Box 4, West Point, Pennsylvania 19486; telephone: ++1-215-652-8636; fax: ++1-215-993-4884
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Abstract

This is the first study to examine PER.C6TM cell glucose/energy and glutamine metabolism with fed-batch cultures at controlled low glutamine, low glucose, and simultaneous low glucose and low glutamine levels. PER.C6TM cell metabolism was investigated in serum-free suspension bioreactors at two-liter scale. Control of glucose and/or glutamine concentrations had a significant effect on cellular metabolism leading to an increased efficiency of nutrient utilization, altered byproduct synthesis, while having no effect on cell growth rate. Cultivating cells at a controlled glutamine concentration of 0.25 mM reduced qGln and qmath image by approximately 30%, qAla 85%, and qNEAA 50%. The fed-batch control of glutamine also reduced the overall accumulation of ammonium ion by approximately 50% by minimizing the spontaneous chemical degradation of glutamine. No major impact upon glucose/energy metabolism was observed. Cultivating cells at a glucose concentration of 0.5 mM reduced qGlc about 50% and eliminated lactate accumulation. Cells exhibited a fully oxidative metabolism with Ymath image of approximately 6 mol/mol. However, despite no increase in qGln, an increased ammonium ion accumulation and Ymath image were also observed. Effective control of lactate and ammonium ion accumulation by PER.C6TM cells was achieved using fed-batch with simultaneously controlled glucose and glutamine. A fully oxidative glucose metabolism and a complete elimination of lactate production were obtained. The qGln value was again reduced and, despite an increased qmath image compared with batch culture, ammonium ion levels were typically lower than corresponding ones in batch cultures, and the accumulation of non-essential amino acids (NEAA) was reduced about 50%. In conclusion, this study shows that PER.C6TM cell metabolism can be confined to a state with improved efficiencies of nutrient utilization by cultivating cells in fed-batch at millimolar controlled levels of glucose and glutamine. In addition, PER.C6TM cells fall into a minority category of mammalian cell lines for which glutamine plays a minor role in energy metabolism. © 2006 Wiley Periodicals, Inc.

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