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Enhancing CHO cell productivity through a dual selection system using Aspg and Gs in glutamine free medium
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  • Tae Ha,
  • Andreu Òdena,
  • Karen Julie la Cour Karottki,
  • Che Lin Kim,
  • Hooman Hefzi,
  • Gyun Min Lee,
  • Helene Faustrup Kildegaard,
  • Lars K. Nielsen,
  • Lise Marie Grav,
  • Nathan Lewis
Tae Ha
The Novo Nordisk Foundation Center For Biosustainability Technical University Of Denmark Kgs Lyngby Denmark
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Andreu Òdena
The Novo Nordisk Foundation Center For Biosustainability Technical University Of Denmark Kgs Lyngby Denmark
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Karen Julie la Cour Karottki
The Novo Nordisk Foundation Center For Biosustainability Technical University Of Denmark Kgs Lyngby Denmark
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Che Lin Kim
The Novo Nordisk Foundation Center For Biosustainability Technical University Of Denmark Kgs Lyngby Denmark
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Hooman Hefzi
Departments of Pediatrics and Bioengineering University of California San Diego USA
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Gyun Min Lee
Department Of Biological Sciences Kaist 291 Daehak-Ro Yuseong-Gu Daejeon 305-701 Republic Of Korea
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Helene Faustrup Kildegaard
The Novo Nordisk Foundation Center For Biosustainability Technical University Of Denmark Kgs Lyngby Denmark
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Lars K. Nielsen
The Novo Nordisk Foundation Center For Biosustainability Technical University Of Denmark Kgs Lyngby Denmark
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Lise Marie Grav
The Novo Nordisk Foundation Center For Biosustainability Technical University Of Denmark Kgs Lyngby Denmark
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Nathan Lewis
Departments of Pediatrics and Bioengineering University of California San Diego USA
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Abstract

The dominant method for generating Chinese hamster ovary (CHO) cell lines that produce high titers of biotherapeutic proteins utilizes selectable markers such as dihydrofolate reductase (Dhfr) or glutamine synthetase (Gs), alongside inhibitory compounds like methotrexate (MTX) or methionine sulfoximine (MSX), respectively. Recent work has shown the importance of asparaginase (Aspg) for growth in media lacking glutamine–the selection medium for Gs-based selection systems. We generated a Gs/Aspg double knockout CHO cell line and evaluated its utility as a novel dual selectable system via co-transfection of Gs-Enbrel and Aspg-Enbrel plasmids. Using the same selection conditions as the standard Gs system, the resulting cells from the Gs/Aspg dual selection showed substantially improved specific productivity and titer compared to the standard Gs selection method, however, with reduced growth rate and viability. Following adaptation in selection medium, the cells improved viability and growth while still achieving ~5-fold higher specific productivity and ~3-fold higher titer than Gs selection alone. We anticipate that with further optimization of culture medium and selection conditions this approach would serve as an effective addition to workflows for the industrial production of recombinant biotherapeutics.
24 Jun 2022Submitted to Biotechnology and Bioengineering
24 Jun 2022Assigned to Editor
24 Jun 2022Submission Checks Completed
27 Jun 2022Reviewer(s) Assigned
27 Jul 2022Review(s) Completed, Editorial Evaluation Pending
27 Jul 2022Editorial Decision: Revise Major