Regulation of cell cycle and productivity in NS0 cells by the over‐expression of p21<sup>CIP1</sup>

Watanabe, Shikiko; Shuttleworth, James; Al‐Rubeai, Mohamed

doi:10.1002/bit.10112

Cited by 70 publications

(22 citation statements)

References 31 publications

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“…p21 CIP1 -arrested CHO are approximately 4-fold bigger than proliferating CHO, are metabolically more active and demonstrate a corresponding increase in protein synthesis (Bi et al 2004). Similarly specific productivity increased 4-fold in NS0 cells expressing a chimeric IgG4 antibody following p21 CIP1 arrested NS0 cells (Watanabe et al 2002;Ibarra et al 2003). p27 KIP1 exerts G1-cell cycle arrest by binding to cyclin/CDK complexes, i.e.…”

Section: (Ii) Cell Engineering Based Approachesmentioning

confidence: 99%

Proliferation control strategies to improve productivity and survival during CHO based production culture

2007

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Chinese Hamster Ovary cells are the primary system for the production of recombinant proteins for therapeutic use. Protein productivity is directly proportional to viable biomass, viability and culture longevity of the producer cells and a number of approaches have been taken to optimise these parameters. Cell cycle arrest, particularly in G1 phase, typically using reduced temperature cultivation and nutritional control have been used to enhance productivity in production cultures by prolonging the production phase, but the mechanism by which these approaches work is still not fully understood. In this article, we analyse the public literature on proliferation control approaches as they apply to production cell lines with particular reference to what is known about the mechanisms behind each approach.

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Section: (Ii) Cell Engineering Based Approachesmentioning

confidence: 99%

Proliferation control strategies to improve productivity and survival during CHO based production culture

2007

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“…To increase productivity, many cellular pathways including energy metabolism, cell cycle, apoptosis and protein secretion have been extensively studied (Chen et al 2001;Kim and Lee 2002;Watanabe et al 2002;Ifandi and Al-Rubeai 2005;Seth et al 2006;Jayapal et al 2007;Kim et al 2012). Increasing cell density by either reducing apoptosis (Meents et al 2002;Figueroa et al 2007) or altering cell metabolism (Chen et al 2001) can lead to increase in product titers.…”

Section: Introductionmentioning

confidence: 99%

Dynamics of unfolded protein response in recombinant CHO cells

Prashad

Mehra

2014

Cytotechnology

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Genes in the protein secretion pathway have been targeted to increase productivity of monoclonal antibodies in Chinese hamster ovary cells. The results have been highly variable depending on the cell type and the relative amount of recombinant and target proteins. This paper presents a comprehensive study encompassing major components of the protein processing pathway in the endoplasmic reticulum (ER) to elucidate its role in recombinant cells. mRNA profiles of all major ER chaperones and unfolded protein response (UPR) pathway genes are measured at a series of time points in a high-producing cell line under the dynamic environment of a batch culture. An initial increase in IgG heavy chain mRNA levels correlates with an increase in productivity. We observe a parallel increase in the expression levels of majority of chaperones. The chaperone levels continue to increase until the end of the batch culture. In contrast, calreticulin and ERO1-L alpha, two of the lowest expressed genes exhibit transient time profiles, with peak induction on day 3. In response to increased ER stress, both the GCN2/PKR-like ER kinase and inositol-requiring enzyme-1alpha (Ire1a) signalling branch of the UPR are upregulated. Interestingly, spliced X-Box binding protein 1 (XBP1s) transcription factor from Ire1a pathway is detected from the beginning of the batch culture. Comparison with the expression levels in a low producer, show much lower induction at the end of the exponential growth phase. Thus, the unfolded protein response strongly correlates with the magnitude and timing of stress in the course of the batch culture.

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“…Strategies for controlling cell growth include the expression or addition of cell cycle inhibitors like p27 (Kaufmann et al 2001), p21cip1 (Watanabe et al 2002), and rapamycin (Balcarcel and Stephanopoulos 2001); or exposure to hyperosmotic pressure (Ryu et al 2000) or low culture temperature. Low temperature cultivation is a simple and very effective method of controlling cell proliferation (Reuveny et al 1993).…”

Section: Introductionmentioning

confidence: 99%

pH Condition in temperature shift cultivation enhances cell longevity and specific hMab productivity in CHO culture

et al. 2006

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Controlling cell proliferation during cell culturing is an effective way to improve the production yield in mammalian cell culture. We examined the effect of temperature shifts (TS) under pH control conditions in Chinese hamster ovary cells. When we shifted the culture temperature from 37°C to 31°C before a stationary phase at pH 6.8 (TS/pH 6.8), cell viability remained high, and the final human monoclonal antibody (hMab) concentration increased to 2.3 times that in the culture remaining at 37°C. However, there were no significant effects on the cell viability or production yield with the same TS at pH 7.0 (TS/pH 7.0). The average specific hMab productivity and mRNA level of TS/pH 7.0 were the same as that of TS/pH 6.8. The control of cell growth by the TS or the addition of rapamycin was effective in the maintenance of cell viability, but there was no significant increase of the average specific hMab productivity in the culture where cell proliferation was controlled with rapamycin. The hMab mRNA concentration decreased to 55%-65% at a 37°C culture with the addition of actinomycin D. In contrast, actinomycin D did not affect the mRNA level in the TS culture. This result suggested that the increase in the mRNA level in the TS condition was caused by an increase in mRNA stability. In this study, we show that TS can produce two unrelated effects: a prolongation of cell longevity and an improvement in mRNA stability.

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Regulation of cell cycle and productivity in NS0 cells by the over‐expression of p21^CIP1

Cited by 70 publications

References 31 publications

Proliferation control strategies to improve productivity and survival during CHO based production culture

Proliferation control strategies to improve productivity and survival during CHO based production culture

Dynamics of unfolded protein response in recombinant CHO cells

pH Condition in temperature shift cultivation enhances cell longevity and specific hMab productivity in CHO culture

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Regulation of cell cycle and productivity in NS0 cells by the over‐expression of p21CIP1

Cited by 70 publications

References 31 publications

Proliferation control strategies to improve productivity and survival during CHO based production culture

Proliferation control strategies to improve productivity and survival during CHO based production culture

Dynamics of unfolded protein response in recombinant CHO cells

pH Condition in temperature shift cultivation enhances cell longevity and specific hMab productivity in CHO culture

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Regulation of cell cycle and productivity in NS0 cells by the over‐expression of p21^CIP1