Engineering Cells for Cell Culture Bioprocessing – Physiological Fundamentals

Seth, Gargi; Hossler, Patrick; Yee, Joon Chong; Hu, Wei‐Shou

doi:10.1007/10_017

Cited by 59 publications

(62 citation statements)

References 187 publications

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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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Section: Introductionmentioning

confidence: 99%

Dynamics of unfolded protein response in recombinant CHO cells

Prashad

Mehra

2014

Cytotechnology

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“…Mammalian cells have the unique ability to produce therapeutic proteins with oligosaccharides attached to their serine/threonine (O-linked glycosylation) or asparagines (N-linked glycosylation) residues (Butler 2005;Seth et al 2006a). The functional biological activity of the glycoprotein is directly affected by glycosylation, including its trafficking and folding within the host cell (Scallon et al 2006;Walsh and Jefferis 2006;Willey 1999).…”

Section: Optimisation Of Glycosylation Patternsmentioning

confidence: 99%

“…Once secreted, solubility, aggregation, stability and immunogenicity of the protein may be affected by its glycosylation pattern (Kobata 1992;Sinclair and Elliott 2005;Willey 1999;Wyss and Wagner 1996). N-glycosylation is initiated in the ER, and O-glycosylation can be initiated in either the ER or Golgi apparatus, but due to processing inconstancies glycans can have frequent structural heterogeneities (Patel et al 1992;Seth et al 2006a;Varki 1998). The most common therapeutic monoclonal antibody approved biopharmaceutical is of the immunoglobulin G (IgG) class (Walsh and Jefferis 2006).…”

Section: Optimisation Of Glycosylation Patternsmentioning

confidence: 99%

“…For instance, by enhancing pyruvate carboxylase expression, which converts pyruvate to oxaloacetate (OAA). The increase in oxalacetate conversion to malate via the malate-aspartate shuttle for entry into the tricarboxylic acid cycle decreased pyruvate concentration which in turn reduced lactate formation (Irani et al 2002, Seth et al 2006a); however, no data have been presented on the effect of oxygen uptake rate which should give a more conclusive picture. In another study, antisense technology has been employed to knockdown the enzyme LDH in CHO cells.…”

Section: Engineering Of Metabolic Pathwaysmentioning

confidence: 99%

“…In another study, antisense technology has been employed to knockdown the enzyme LDH in CHO cells. LDH is involved in the reversible reaction of lactate acid from pyruvate and interestingly, this strategy reduced the overall glycolysis rate, likely due to reduced NADH to NAD + formation, which is coupled to the pyruvate-lactate reaction (Seth et al 2006a). This work illustrates the complexity of metabolic pathway engineering and how multiple gene engineering in a single host might be a possible future strategy to overcome some of these problems.…”

Section: Engineering Of Metabolic Pathwaysmentioning

confidence: 99%

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Using cell engineering and omic tools for the improvement of cell culture processes

et al. 2007

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Significant strides have been made in mammalian cell based biopharmaceutical process and cell line development over the past years. With several established mammalian host cell lines and expression systems, optimization of selection systems to reduce development times and improvement of glycosylation patterns are only some of the advances being made to improve cell culture processes. In this article, the advances pertaining to cell line development and cell engineering strategies are discussed. An overview of the cell engineering strategies to enhance cellular characteristics by genetic manipulation are illustrated, focusing on the use of genomics and proteomics tools and their application in such endeavors. Included in this review are some of the early studies using the 'omic' technique to understand cellular mechanisms of product synthesis and secretion, apoptosis, cell proliferation and the influence of the physicochemical environment. The article highlights the significance of integrating genomics and proteomics data with the vast amounts of bioprocess data for improved analysis of the biological pathways involved. Further improvements of the techniques and methodologies used are needed but ultimately, the new cell engineering strategies should provide great insight into the regulatory networks within the cell in a bioprocess environment and how to manipulate them to increase overall productivity.

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Process Development of Protein Therapeutics

Thomas

Guhan

Pettit

2010

Burger's Medicinal Chemistry and Drug Discovery

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The application of recombinant DNA technology to the production of protein therapeutics has undergone considerable progress since first introduced 30 years ago. Considerable scientific effort has been devoted to developing robust processes that produce large amounts of complex proteins with the desired product quality attributes. An overview of the contributions from the four major disciplines working in concert to deliver these processes and methods will be covered. This chapter will discuss some of the tools, methods, and approaches used to produce, purify, formulate, and analyze the drugs of modern biotechnology. Future trends in each of the disciplines will also be presented.

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Engineering Cells for Cell Culture Bioprocessing – Physiological Fundamentals

Cited by 59 publications

References 187 publications

Dynamics of unfolded protein response in recombinant CHO cells

Dynamics of unfolded protein response in recombinant CHO cells

Using cell engineering and omic tools for the improvement of cell culture processes

Process Development of Protein Therapeutics

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