A comparison of orbitally‐shaken and stirred‐tank bioreactors: pH modulation and bioreactor type affect CHO cell growth and protein glycosylation

Orbitally shaken bioreactors (OSRs) support the suspension cultivation of animal cells at volumetric scales up to 200 L and are a potential alternative to stirred‐tank bioreactors (STRs) due to their rapid and homogeneous mixing and high oxygen transfer rate. In this study, a Chinese hamster ovary cell line producing a recombinant antibody was cultivated in a 5 L OSR and a 3 L STR, both operated with or without pH control. Effects of bioreactor type and pH control on cell growth and metabolism and on recombinant protein production and glycosylation were determined. In pH‐controlled bioreactors, the glucose consumption and lactate production rates were higher relative to cultures grown in bioreactors without pH control. The cell density and viability were higher in the OSRs than in the STRs, either with or without pH control. Volumetric recombinant antibody yields were not affected by the process conditions, and a glycan analysis of the antibody by mass spectrometry did not reveal major process‐dependent differences in the galactosylation index. The results demonstrated that OSRs are suitable for recombinant protein production from suspension‐adapted animal cells. © 2016 American Institute of Chemical Engineers Biotechnol. Prog., 32:1174–1180, 2016

[1]  Alfio Quarteroni,et al.  Efficient oxygen transfer by surface aeration in shaken cylindrical containers for mammalian cell cultivation at volumetric scales up to 1000 L , 2009 .

[2]  J. Büchs,et al.  Advances in shaking technologies. , 2012, Trends in biotechnology.

[3]  J. Chalmers,et al.  Physiological responses of CHO cells to repetitive hydrodynamic stress , 2009, Biotechnology and bioengineering.

[4]  R. Jordan,et al.  Galactosylation variations in marketed therapeutic antibodies , 2012, mAbs.

[5]  R. Senger,et al.  Effect of Shear Stress on Intrinsic CHO Culture State and Glycosylation of Recombinant Tissue‐Type Plasminogen Activator Protein , 2008, Biotechnology progress.

[6]  Tongtong Wang,et al.  pH measurement and a rational and practical pH control strategy for high throughput cell culture system , 2009, Biotechnology progress.

[7]  F. Wurm,et al.  New disposable tubes for rapid and precise biomass assessment for suspension cultures of mammalian cells. , 2006, Biotechnology and bioengineering.

[8]  A. Nienow.  Reactor Engineering in Large Scale Animal Cell Culture , 2006, Cytotechnology.

[9]  Tilman Schlothauer,et al.  In Vitro Glycoengineering of IgG1 and Its Effect on Fc Receptor Binding and ADCC Activity , 2015, PloS one.

[10]  W M Miller,et al.  Glycosylation of CHO‐Derived Recombinant tPA Produced under Elevated pCO2 , 1997, Biotechnology progress.

[11]  Z. Li,et al.  Optimal and consistent protein glycosylation in mammalian cell culture. , 2009, Glycobiology.

[12]  G. Kresse.  Biosimilars--science, status, and strategic perspective. , 2009, European journal of pharmaceutics and biopharmaceutics : official journal of Arbeitsgemeinschaft fur Pharmazeutische Verfahrenstechnik e.V.

[13]  M. Butler,et al.  Glycosylation of an immunoglobulin produced from a murine hybridoma cell line: The effect of culture mode and the anti‐apoptotic gene, bcl‐2 , 2007, Biotechnology and bioengineering.

[14]  Roy Jefferis,et al.  Glycosylation as a strategy to improve antibody-based therapeutics , 2009, Nature Reviews Drug Discovery.

[15]  Alfio Quarteroni,et al.  Numerical simulation of orbitally shaken viscous fluids with free surface , 2013 .

[16]  F. Wurm,et al.  k(L)a as a predictor for successful probe-independent mammalian cell bioprocesses in orbitally shaken bioreactors. , 2012, New biotechnology.

[17]  C -M. Liu,et al.  Development of a shaking bioreactor system for animal cell cultures. , 2001, Biochemical engineering journal.

[18]  Stefan Schillberg,et al.  Correlation between mass transfer coefficient kLa and relevant operating parameters in cylindrical disposable shaken bioreactors on a bench-to-pilot scale , 2013, Journal of Biological Engineering.

[19]  Massimo Morbidelli,et al.  Adaptation for survival: phenotype and transcriptome response of CHO cells to elevated stress induced by agitation and sparging. , 2014, Journal of biotechnology.

[20]  David L Hacker,et al.  Generation of stable, high‐producing cho cell lines by lentiviral vector‐mediated gene transfer in serum‐free suspension culture , 2011, Biotechnology and bioengineering.

[21]  J. Büchs,et al.  Scaled‐up manufacturing of recombinant antibodies produced by plant cells in a 200‐L orbitally‐shaken disposable bioreactor , 2015, Biotechnology and bioengineering.

[22]  Gyun Min Lee,et al.  Effect of culture pH on erythropoietin production by Chinese hamster ovary cells grown in suspension at 32.5 and 37.0 degrees C. , 2005, Biotechnology and bioengineering.

[23]  Uwe Kärst,et al.  Effects of buffering conditions and culture pH on production rates and glycosylation of clinical phase I anti-melanoma mouse IgG3 monoclonal antibody R24. , 2003, Biotechnology and bioengineering.

[24]  F. Wurm,et al.  Efficient and reproducible mammalian cell bioprocesses without probes and controllers? , 2011, New biotechnology.

[25]  Alvin W. Nienow,et al.  On impeller circulation and mixing effectiveness in the turbulent flow regime , 1997 .

[26]  J. Büchs,et al.  Time efficient way to calculate oxygen transfer areas and power input in cylindrical disposable shaken bioreactors , 2014, Biotechnology progress.

[27]  B O Palsson,et al.  Growth, Metabolic, and Antibody Production Kinetics of Hybridoma Cell Culture: 2. Effects of Serum Concentration, Dissolved Oxygen Concentration, and Medium pH in a Batch Reactor , 1991, Biotechnology progress.

[28]  Stephan Kaiser,et al.  Disposable bioreactors: the current state-of-the-art and recommended applications in biotechnology , 2010, Applied Microbiology and Biotechnology.

[29]  A. Quarteroni,et al.  Use of orbital shaken disposable bioreactors for mammalian cell cultures from the milliliter-scale to the 1,000-liter scale. , 2009, Advances in biochemical engineering/biotechnology.

[30]  P. Aebischer,et al.  Genetic engineering of cell lines using lentiviral vectors to achieve antibody secretion following encapsulated implantation. , 2014, Biomaterials.

[31]  P. Chatellard,et al.  Transient gene expression: recombinant protein production with suspension-adapted HEK293-EBNA cells. , 2001, Biotechnology and bioengineering.

[32]  Jochen Büchs,et al.  Orbitally shaken single-use bioreactors. , 2014, Advances in biochemical engineering/biotechnology.

[33]  A. Nienow,et al.  Control of pH in large-scale, free suspension animal cell bioreactors: alkali addition and pH excursions. , 1999, Biotechnology and bioengineering.

[35]  T. Raju,et al.  Terminal sugars of Fc glycans influence antibody effector functions of IgGs. , 2008, Current opinion in immunology.

[36]  F. Wurm,et al.  Disposable 600-mL orbitally shaken bioreactor for mammalian cell cultivation in suspension , 2013 .

[37]  F. Wurm,et al.  Novel Orbital Shake Bioreactors for Transient Production of CHO Derived IgGs , 2007, Biotechnology progress.

[38]  Michael C. Borys,et al.  Ammonia affects the glycosylation patterns of recombinant mouse placental lactogen‐I by chinese hamster ovary cells in a pH‐dependent manner , 1994, Biotechnology and bioengineering.

[39]  Natalie Muller,et al.  Orbital shaker technology for the cultivation of mammalian cells in suspension. , 2005, Biotechnology and bioengineering.

[40]  F. Wurm,et al.  Glycan variability on a recombinant IgG antibody transiently produced in HEK-293E cells. , 2012, New biotechnology.