Hefzi Hooman, Martínez-Monge Iván, Marin de Mas Igor, Cowie Nicholas Luke, Toledo Alejandro Gomez, Noh Soo Min, Karottki Karen Julie la Cour, Decker Marianne, Arnsdorf Johnny, Camacho-Zaragoza Jose Manuel, Kol Stefan, Schoffelen Sanne, Pristovšek Nuša, Hansen Anders Holmgaard, Miguez Antonio A, Bjørn Sara Petersen, Brøndum Karen Kathrine, Javidi Elham Maria, Jensen Kristian Lund, Stangl Laura, Kreidl Emanuel, Kallehauge Thomas Beuchert, Ley Daniel, Ménard Patrice, Petersen Helle Munck, Sukhova Zulfiya, Bauer Anton, Casanova Emilio, Barron Niall, Malmström Johan, Nielsen Lars K, Lee Gyun Min, Kildegaard Helene Faustrup, Voldborg Bjørn G, Lewis Nathan E
Department of Bioengineering, University of California, University of California, San Diego, La Jolla, CA, USA.
Novo Nordisk Foundation Center for Biosustainability, University of California, San Diego, School of Medicine, La Jolla, CA, USA.
Nat Metab. 2025 Jan;7(1):212-227. doi: 10.1038/s42255-024-01193-7. Epub 2025 Jan 14.
The Warburg effect, which describes the fermentation of glucose to lactate even in the presence of oxygen, 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 in cells for bioprocessing have failed as lactate dehydrogenase is essential for cell growth. Here, we effectively eliminate lactate production in Chinese hamster ovary and in the human embryonic kidney cell line HEK293 by simultaneous knockout of lactate dehydrogenases and pyruvate dehydrogenase kinases, thereby removing a negative feedback loop that typically inhibits pyruvate conversion to acetyl-CoA. These cells, which we refer to as Warburg-null cells, maintain wild-type growth rates while producing negligible lactate, show a compensatory increase in oxygen consumption, near total reliance on oxidative metabolism, and higher cell densities in fed-batch cell culture. Warburg-null cells remain amenable for production of diverse biotherapeutic proteins, reaching industrially relevant titres and maintaining product glycosylation. The ability to eliminate lactate production may be useful for biotherapeutic production and provides a tool for investigating a common metabolic phenomenon.
瓦伯格效应描述了即使在有氧存在的情况下葡萄糖仍发酵生成乳酸的现象，这种效应在包括癌细胞在内的增殖性哺乳动物细胞中普遍存在，但对生物制药生产构成挑战，因为乳酸积累会抑制细胞生长和蛋白质生产。以往为生物加工而消除细胞中乳酸生成的努力均告失败，因为乳酸脱氢酶对细胞生长至关重要。在此，我们通过同时敲除乳酸脱氢酶和丙酮酸脱氢酶激酶，有效消除了中国仓鼠卵巢细胞和人胚肾细胞系HEK293中的乳酸生成，从而消除了通常抑制丙酮酸转化为乙酰辅酶A的负反馈回路。这些细胞，我们称之为无瓦伯格效应细胞，在产生可忽略不计的乳酸的同时保持野生型生长速率，显示出耗氧量的代偿性增加，几乎完全依赖氧化代谢，并且在分批补料细胞培养中具有更高的细胞密度。无瓦伯格效应细胞仍然适合生产多种生物治疗蛋白，达到工业相关滴度并维持产物糖基化。消除乳酸生成的能力可能对生物治疗生产有用，并为研究一种常见的代谢现象提供了一种工具。
