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Multiplex genome editing eliminates the Warburg Effect without impacting growth rate in mammalian cells

Hefzi, Hooman ; Martínez-Monge, Iván ; Marin de Mas, Igor ; Cowie, Nicholas Luke ; Toledo, Alejandro Gomez LU ; Noh, Soo Min ; Karottki, Karen Julie la Cour ; Decker, Marianne ; Arnsdorf, Johnny and Camacho-Zaragoza, Jose Manuel , et al. (2024) In bioRxiv : the preprint server for biology
Abstract

The Warburg effect is ubiquitous in proliferative mammalian cells, including cancer cells, but poses challenges for biopharmaceutical production, as lactate accumulation inhibits cell growth and protein production. Previous efforts to eliminate lactate production via knockout have failed in mammalian bioprocessing since lactate dehydrogenase has proven essential. However, here we eliminated the Warburg effect in Chinese hamster ovary (CHO) and HEK293 cells by simultaneously knocking out lactate dehydrogenase and regulators involved in a negative feedback loop that typically inhibits pyruvate conversion to acetyl-CoA. In contrast to long-standing assumptions about the role of aerobic glycolysis, Warburg-null cells maintain wildtype... (More)

The Warburg effect is ubiquitous in proliferative mammalian cells, including cancer cells, but poses challenges for biopharmaceutical production, as lactate accumulation inhibits cell growth and protein production. Previous efforts to eliminate lactate production via knockout have failed in mammalian bioprocessing since lactate dehydrogenase has proven essential. However, here we eliminated the Warburg effect in Chinese hamster ovary (CHO) and HEK293 cells by simultaneously knocking out lactate dehydrogenase and regulators involved in a negative feedback loop that typically inhibits pyruvate conversion to acetyl-CoA. In contrast to long-standing assumptions about the role of aerobic glycolysis, Warburg-null cells maintain wildtype growth rate while producing negligible lactate. Further characterization of Warburg-null CHO cells showed a compensatory increase in oxygen consumption, a near total reliance on oxidative metabolism, and higher cell densities in fed-batch cell culture. These cells remained amenable for production of diverse biotherapeutic proteins, reaching industrially relevant titers and maintaining product glycosylation. Thus, the ability to eliminate the Warburg effect is an important development for biotherapeutic production and provides a tool for investigating a near-universal metabolic phenomenon.

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publishing date
type
Working paper/Preprint
publication status
published
subject
in
bioRxiv : the preprint server for biology
publisher
bioRxiv
external identifiers
  • pmid:39211256
ISSN
2692-8205
DOI
10.1101/2024.08.02.606284
language
English
LU publication?
yes
id
99a0cba8-e91e-4a78-875d-80446a90e6ea
date added to LUP
2024-09-05 07:32:08
date last changed
2024-09-05 08:03:37
@misc{99a0cba8-e91e-4a78-875d-80446a90e6ea,
  abstract     = {{<p>The Warburg effect is ubiquitous in proliferative mammalian cells, including cancer cells, but poses challenges for biopharmaceutical production, as lactate accumulation inhibits cell growth and protein production. Previous efforts to eliminate lactate production via knockout have failed in mammalian bioprocessing since lactate dehydrogenase has proven essential. However, here we eliminated the Warburg effect in Chinese hamster ovary (CHO) and HEK293 cells by simultaneously knocking out lactate dehydrogenase and regulators involved in a negative feedback loop that typically inhibits pyruvate conversion to acetyl-CoA. In contrast to long-standing assumptions about the role of aerobic glycolysis, Warburg-null cells maintain wildtype growth rate while producing negligible lactate. Further characterization of Warburg-null CHO cells showed a compensatory increase in oxygen consumption, a near total reliance on oxidative metabolism, and higher cell densities in fed-batch cell culture. These cells remained amenable for production of diverse biotherapeutic proteins, reaching industrially relevant titers and maintaining product glycosylation. Thus, the ability to eliminate the Warburg effect is an important development for biotherapeutic production and provides a tool for investigating a near-universal metabolic phenomenon.</p>}},
  author       = {{Hefzi, Hooman and Martínez-Monge, Iván and Marin de Mas, Igor and Cowie, Nicholas Luke and Toledo, Alejandro Gomez and Noh, Soo Min and Karottki, Karen Julie la Cour and Decker, Marianne and Arnsdorf, Johnny and Camacho-Zaragoza, Jose Manuel and Kol, Stefan and Schoffelen, Sanne and Pristovšek, Nuša and Hansen, Anders Holmgaard and Miguez, Antonio A and Bjorn, Sara Petersen and Brøndum, Karen Kathrine and Javidi, Elham Maria and Jensen, Kristian Lund and Stangl, Laura and Kreidl, Emanuel and Kallehauge, Thomas Beuchert and Ley, Daniel and Ménard, Patrice and Petersen, Helle Munck and Sukhova, Zulfiya and Bauer, Anton and Casanova, Emilio and Barron, Niall and Malmström, Johan and Nielsen, Lars K and Lee, Gyun Min and Kildegaard, Helene Faustrup and Voldborg, Bjørn G and Lewis, Nathan E}},
  issn         = {{2692-8205}},
  language     = {{eng}},
  month        = {{08}},
  note         = {{Preprint}},
  publisher    = {{bioRxiv}},
  series       = {{bioRxiv : the preprint server for biology}},
  title        = {{Multiplex genome editing eliminates the Warburg Effect without impacting growth rate in mammalian cells}},
  url          = {{http://dx.doi.org/10.1101/2024.08.02.606284}},
  doi          = {{10.1101/2024.08.02.606284}},
  year         = {{2024}},
}