CHO cells knocked out for TSC2 display an improved productivity of antibodies under fed batch conditions. Issue 9 (6th March 2016)

• Record Type: Journal Article
• Authors: McVey, Duncan; Aronov, Michael; Rizzi, Giovanni; Cowan, Alexis; Scott, Charo; Megill, John; Russell, Reb; Tirosh, Boaz
• Is Part Of: Biotechnology and bioengineering. Volume 113:Issue 9(2016)
• Page Start: 1942
• Page End: 1952
• Publication Date: 2016-03-06
• Subjects: mTOR -- CRISPR/Cas9 -- CHO cells -- productivity; Biotechnology -- Periodicals; Bioengineering -- Periodicals; 660.6
• DOI: 10.1002/bit.25951
• Languages: English
• ISSNs: 0006-3592

Abstract

ABSTRACT: The kinase mTOR operates in two cellular complexes, mTORC1 and mTORC2. mTORC1 adjusts metabolic activity according to external growth conditions and nutrients availability. When conditions are prosperous, mTOR facilitates protein and lipid biosyntheses and inhibits autophagy, while under metabolic constraints, however, its attenuation induces a catabolic program, energy preservation and autophagy. CHO is a key cell line for manufacturing of biologics owing to its remarkable ability to grow to high densities and maintain protein production and secretion for extended times. While high mTOR activity has been associated with high productivity in CHO cells, its inhibition by rapamycin has also been documented to augment productivity via promotion of viability. Here using CRISPR/Cas9 editing we engineered CHO cells to enforce high mTORC1 activity by knocking‐out TSC2, a major mTOR inhibitory protein, or PTEN, a phosphatase that attenuates the PI3K/AKT/mTOR pathway. Only TSC2‐deleted cells exhibited a constitutive activation of mTORC1 under fed batch conditions. Cells grew larger in size, synthesized more proteins and displayed an over twofold elevation in their specific productivity. While peak viable cell density was compromised, overall titers increased to an extent dependent upon the parental clone. Our data underscore manipulation of TSC as a strategy to improve performance of CHO cell in bioreactors. Biotechnol. Bioeng. 2016;113: 1942–1952. © 2016 Wiley Periodicals, Inc. Abstract : Tirosh and co‐workers show that under fed batch conditions, mTORC1 output in normal CHO cells undergoes progressive attenuation at the stationary phase of growth. Deletion of TSC2 completely aborts this attenuation, allowing mTORC1 to be constitutively active. This results in cells that are bigger, synthesize more proteins and ultimately producing more antibodies.