Transcriptomic analysis of clonal growth rate variation during CHO cell line development

Doolan, Padraig; Clarke, Colin; Kinsella, Paula; Breen, Laura; Leonard, Mark; Zhang, Lin; Clynes, Martin; Aherne, Sinéad; Barron, Niall

doi:10.1016/j.jbiotec.2013.04.014

Cited by 25 publications

(24 citation statements)

References 30 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

Section: Introductionmentioning

confidence: 99%

“…In order to address this issue, a number of proteomic, transcriptomic and metabolic based studies have now been undertaken and the subsequent data used to develop models to predict the phenotype of a given cell line (e.g. Clarke et al, 2011;Clarke et al, 2012;Doolan et al, 2013;Jacob et al, 2010;Mead et al, 2009;Mead et al, 2012;Meleady et al, 2011;Sanchez et al, 2014;Selvarasu et al, 2012), however these models are usually developed from cell line data at the end of the cell line development process.The goal of this work was to develop a screening system that would allow the selection of highly productive cell lines for monoclonal antibody (mAb) production early in the cell line development process that would use substantially less resource to achieve the same or better success rate as current methods. The vision was to be able to select a small number of cell lines based upon the analysis of data generated in multi-well plates, and take these straight to a lab-scale bioreactorevaluation stage (10 L) with a high probability that the selected cell lines were highly productive.…”

mentioning

confidence: 99%

See 1 more Smart Citation

Rapid high-throughput characterisation, classification and selection of recombinant mammalian cell line phenotypes using intact cell MALDI-ToF mass spectrometry fingerprinting and PLS-DA modelling

Povey

O'Malley

Root

et al. 2014

Journal of Biotechnology

View full text Add to dashboard Cite

Despite many advances in the generation of high producing recombinant mammalian cell lines over the last few decades, cell line selection and development is often slowed by the inability to predict a cell line's phenotypic characteristics (e.g. growth or recombinant protein productivity) at larger scale (large volume bioreactors) using data from early cell line construction at small culture scale. Here we describe the development of an intact cell MALDI-ToF mass spectrometry fingerprinting method for mammalian cells early in the cell line construction process whereby the resulting mass spectrometry data is used to predict the phenotype of mammalian cell lines at larger culture scale using a Partial Least Squares Discriminant Analysis (PLS-DA) model. Using MALDI-ToF mass spectrometry, a library of mass spectrometry fingerprints was generated for individual cell lines at the 96 deep well plate stage of cell line development. The growth and productivity of these cell lines were evaluated in a 10 L bioreactor model of Lonza's large-scale (up to 20,000 L) fed-batch cell culture processes.Using the mass spectrometry information at the 96 deep well plate stage and phenotype information at the 10 L bioreactor scale a PLS-DA model was developed to predict the productivity of unknown cell lines at the 10 L scale based upon their MALDI-ToF fingerprint at the 96 deep well plate scale. This approach provides the basis for the very early prediction of cell lines' performance in cGMP manufacturing-scale bioreactors and the foundation for methods and models for predicting other mammalian cell phenotypes from rapid, intact-cell mass spectrometry based measurements.Keywords: Cell line development, Chinese hamster ovary cells, whole cell MALDI-ToF mass spectrometry, PLS-DA modelling, cell line prediction 3 IntroductionThe majority of recombinant protein biopharmaceuticals are produced from cultured mammalian cells (Walsh, 2010), with the most commonly used industrial mammalian cell host being the Chinese hamster ovary (CHO) cell (Kim et al., 2012). Despite the development of high throughput methods that allow the screening of many recombinant cell lines to isolate those with desirable phenotypes (e.g. high growth and productivity), the ability of such methods to select or predict the performance of a given cell line at manufacturing scale remains limited with the best cell lines at a manufacturing scale often distributed across the phenotypic performance of cell lines at smaller-scale (Porter et al., 2010a;Porter et al., 2010b). As a result, early use of a simple productivity based approach does not necessarily allow the identification of high producers in the population of cell lines, and some potentially high producers are discarded early in the process (Porter et al., 2010b). In order to address this issue, a number of proteomic, transcriptomic and metabolic based studies have now been undertaken and the subsequent data used to develop models to predict the phenotype of a given cell line (e.g. Clarke et al., 2011;Clarke...

show abstract

Section: Introductionmentioning

confidence: 99%

mentioning

confidence: 99%

Rapid high-throughput characterisation, classification and selection of recombinant mammalian cell line phenotypes using intact cell MALDI-ToF mass spectrometry fingerprinting and PLS-DA modelling

Povey

O'Malley

Root

et al. 2014

Journal of Biotechnology

View full text Add to dashboard Cite

show abstract

“…Jing and colleagues observed that angiotensinogen (serpin peptidase inhibitor, clade A, member 8) was upregulated 2.15 fold in dexamethasone treated CHO cultures, and they associated the upregulation to the anti-apoptotic effects of dexamethasone (Jing et al, 2012). Most recently, Doolan et al concluded that Serpinf1 negatively correlated to growth rate in their transcriptomic studies on biomarker discovery for higher growth in CHO cells (Doolan et al, 2013). Although these Serpins may all have different substrates of inhibition and functions in their native tissue and cell types, they may play similar roles as Serpinb1 in CHO cells.…”

Section: Discussionmentioning

confidence: 99%

Overexpression of Serpinb1 in Chinese hamster ovary cells increases recombinant IgG productivity

Lin

Brooks

Sealover

et al. 2015

Journal of Biotechnology

View full text Add to dashboard Cite

“…Specifically, two sets of values for μ g and q p were extracted from each reference, the first corresponding to a low productivity to growth rate ratio (Dool-L24, Chus-L25, and Kant-L26), and the second corresponding to a high productivity to growth ratio (Dool-H24, Chus-H25, and Kant-H26) (Table 1). All but the μ g and q p values obtained from Kant-H correspond to industrial CHO cell lines continuously expressing a recombinant mAb while growing exponentially.…”

Section: Methodsmentioning

confidence: 99%

“…By combining the assumed values for M cell = 271 pg / cell 34 and Z prot = 74.2%35 with the μ g values obtained from literature242526, cellular protein productivity can be estimated (Eq. 22).…”

Section: Methodsmentioning

confidence: 99%

A theoretical estimate for nucleotide sugar demand towards Chinese Hamster Ovary cellular glycosylation

Val

Polizzi

Kontoravdi

2016

Sci Rep

View full text Add to dashboard Cite

Glycosylation greatly influences the safety and efficacy of many of the highest-selling recombinant therapeutic proteins (rTPs). In order to define optimal cell culture feeding strategies that control rTP glycosylation, it is necessary to know how nucleotide sugars (NSs) are consumed towards host cell and rTP glycosylation. Here, we present a theoretical framework that integrates the reported glycoproteome of CHO cells, the number of N-linked and O-GalNAc glycosylation sites on individual host cell proteins (HCPs), and the carbohydrate content of CHO glycosphingolipids to estimate the demand of NSs towards CHO cell glycosylation. We have identified the most abundant N-linked and O-GalNAc CHO glycoproteins, obtained the weighted frequency of N-linked and O-GalNAc glycosites across the CHO cell proteome, and have derived stoichiometric coefficients for NS consumption towards CHO cell glycosylation. By combining the obtained stoichiometric coefficients with previously reported data for specific growth and productivity of CHO cells, we observe that the demand of NSs towards glycosylation is significant and, thus, is required to better understand the burden of glycosylation on cellular metabolism. The estimated demand of NSs towards CHO cell glycosylation can be used to rationally design feeding strategies that ensure optimal and consistent rTP glycosylation.

show abstract

Transcriptomic analysis of clonal growth rate variation during CHO cell line development

Cited by 25 publications

References 30 publications

Rapid high-throughput characterisation, classification and selection of recombinant mammalian cell line phenotypes using intact cell MALDI-ToF mass spectrometry fingerprinting and PLS-DA modelling

Rapid high-throughput characterisation, classification and selection of recombinant mammalian cell line phenotypes using intact cell MALDI-ToF mass spectrometry fingerprinting and PLS-DA modelling

Overexpression of Serpinb1 in Chinese hamster ovary cells increases recombinant IgG productivity

A theoretical estimate for nucleotide sugar demand towards Chinese Hamster Ovary cellular glycosylation

Contact Info

Product

Resources

About