Stable overexpression of mutated PTEN in Chinese hamster ovary cells enhances their performance and therapeutic antibody production

Abstract

Chinese hamster ovary (CHO) cells with a high viable cell density (VCD), resilience to culture stress, and the capacity to continuously express recombinant proteins are highly desirable. Phosphatase and tension homology deleted on chromosome ten (PTEN) functions as a key negative regulator of the PI3K/Akt signaling pathway, mediating cell growth and survival. Its oncogenic mutant endows cells with an enhanced proliferation rate and resistance to death. In this study, the role of oncogenic PTEN C124S or G129E on the performance of CHO‐K1 and CHO‐IgG cells was investigated. Our results showed that CHO‐K1 cells stably expressing PTEN C124S or G129E exhibited enhanced proliferation, reduced apoptosis rate, and increased transient expression of therapeutic antibodies compared to the control cells. Moreover, the stable overexpression of PTEN C124S or G129E endowed CHO‐IgG cells with higher cell viability, VCD, and antibody titers (yield increased by approximately 0.77‐fold) in the fed‐batch culture process and enhanced their performance in response to the addition of sodium lactate. Moreover, the engineering of mutated PTEN in CHO‐IgG cells did not alter antibody quality. Collectively, our data suggest that mutated PTEN is a potential target for improving the manufacture of therapeutic antibodies.

Long term culture promotes changes to growth, gene expression, and metabolism in CHO cells that are independent of production stability

“…The PI3K‐Akt signaling with its association with the mechanistic target of rapamycin (mTOR) pathway mediates multiple aspects of cell function, such as anabolic reactions, nutrient consumption, cell growth, and survival (Saxton & Sabatini, 2017). Several studies have engineered CHO cells to overexpress components of this signaling pathway through different targets (e.g., Akt [Hwang & Lee, 2009], PTEN [Zhou et al, 2021], mTOR [Dreesen & Fussenegger, 2011]), leading to increased cell proliferation and modulated apoptosis. The MAPK/ERK signaling pathway also positively regulates cell proliferation, differentiation, and apoptosis (Sun et al, 2015).…”

“…Several studies have engineered CHO cells to overexpress components of this signaling pathway through different targets (e.g., Akt [Hwang & Lee, 2009], PTEN [Zhou et al, 2021], mTOR [Dreesen & Fussenegger, 2011]), leading to increased cell proliferation and modulated apoptosis. The MAPK/ERK signaling pathway also positively regulates cell proliferation, differentiation, and apoptosis (Sun et al, 2015).…”

Life at the periphery: what makes CHO cells survival talents

“…Previous efforts in CHO cell engineering have already focused on this pathway by overexpression of active AKT, which induced bioprocess relevant and beneficial effects as reduced apoptosis and decreased level of autophagosome accumulation (Hwang and Lee 2009 ). Furthermore, overexpression of oncogenic mutant phosphatase and tension homology deleted on chromosome ten (PTEN), a negative regulator of AKT, resulted in enhanced proliferation, reduced apoptosis rate, and increased transient recombinant protein expression (Zhou et al 2021 ) and the overexpression of EGF receptor in CHO cells resulted in higher sensitivity to EGF supplementation and higher growth rates in the presence of EGF (Chen et al 2001 ). On the other hand, medium supplementation may be a relevant strategy based on expressed or missing ligands of identified surface receptors.…”

Strategies and Considerations for Improving Recombinant Antibody Production and Quality in Chinese Hamster Ovary Cells

“…Owing to the difference between the growth and metabolic characteristics of animal cells, the expression level and modification ability of recombinant antibody are also different ( Dhara et al, 2018 ). Therefore, we can further transform the host cells, improve the expression level by engineering cell line, and meet the production capacity and quality requirement of antibody drugs as much as possible ( Lu et al, 2018 ; Liu et al, 2021 ; Zhou et al, 2021 ).…”