Multivariate analysis of the effect of operating conditions on hybridoma cell metabolism and glycosylation of produced antibody

Green, Amy; Glassey, Jarka

doi:10.1002/jctb.4481

Cited by 19 publications

(12 citation statements)

References 26 publications

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“…One reason for this lies in the natural fear to lose a culture if the cells should be exposed to substrate limitations and starve, for instance because a stoichiometric feeding regime did not work as expected. Another reason for running at a high target glucose concentration range may have to do with glycosylation patterns of the final product which may be influenced by GLC levels [ 109 ]. However, the intracellular flux and concentrations might surpass the extracellular concentration of substrates in importance for the final structure of a complex protein [ 110 ].…”

Section: Figure A1mentioning

confidence: 99%

Metabolic Control in Mammalian Fed-Batch Cell Cultures for Reduced Lactic Acid Accumulation and Improved Process Robustness

et al. 2016

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Biomass and cell-specific metabolic rates usually change dynamically over time, making the “feed according to need” strategy difficult to realize in a commercial fed-batch process. We here demonstrate a novel feeding strategy which is designed to hold a particular metabolic state in a fed-batch process by adaptive feeding in real time. The feed rate is calculated with a transferable biomass model based on capacitance, which changes the nutrient flow stoichiometrically in real time. A limited glucose environment was used to confine the cell in a particular metabolic state. In order to cope with uncertainty, two strategies were tested to change the adaptive feed rate and prevent starvation while in limitation: (i) inline pH and online glucose concentration measurement or (ii) inline pH alone, which was shown to be sufficient for the problem statement. In this contribution, we achieved metabolic control within a defined target range. The direct benefit was two-fold: the lactic acid profile was improved and pH could be kept stable. Multivariate Data Analysis (MVDA) has shown that pH influenced lactic acid production or consumption in historical data sets. We demonstrate that a low pH (around 6.8) is not required for our strategy, as glucose availability is already limiting the flux. On the contrary, we boosted glycolytic flux in glucose limitation by setting the pH to 7.4. This new approach led to a yield of lactic acid/glucose (Y L/G) around zero for the whole process time and high titers in our labs. We hypothesize that a higher carbon flux, resulting from a higher pH, may lead to more cells which produce more product. The relevance of this work aims at feeding mammalian cell cultures safely in limitation with a desired metabolic flux range. This resulted in extremely stable, low glucose levels, very robust pH profiles without acid/base interventions and a metabolic state in which lactic acid was consumed instead of being produced from day 1. With this contribution, we wish to extend the basic repertoire of available process control strategies, which will open up new avenues in automation technology and radically improve process robustness in both process development and manufacturing.

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Section: Figure A1mentioning

confidence: 99%

Metabolic Control in Mammalian Fed-Batch Cell Cultures for Reduced Lactic Acid Accumulation and Improved Process Robustness

et al. 2016

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“…A significant amount of work has been done in cell culture technology over the last few decades for improving mAb productivity with maintaining quality attributes while reducing manufacturing cost and time to market . To improve the mAb titer, cell line engineering, cell culture operation conditions, and media optimization have been studied. Another idea of improving mAb productivity can be obtained by increasing understanding of cell organisms by applying metabolic flux analysis.…”

Section: Introductionmentioning

confidence: 99%

Metabolic responses and pathway changes of mammalian cells under different culture conditions with media supplementations

Park

Reimonn

Agarabi

et al. 2018

Biotechnology Progress

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Amino acids and glucose consumption, cell growth and monoclonal antibody (mAb) production in mammalian cell culture are key considerations during upstream process and particularly media optimization. Understanding the interrelations and the relevant cellular physiology will provide insight for setting strategy of robust and effective mAb production. The aim of this study was to further our understanding of nutrient consumption metabolism, since this could have significant impact on enhancing mAb titer, cell proliferation, designing feeding strategies, and development of feed media. The nutrient consumption pattern, mAb concentration, and cell growth were analyzed in three sets of cell cultures with media supplementation of glucose, methionine, threonine, tryptophan, and tyrosine. The amino acids metabolism and its impact on cell growth and mAb production during the batch and fed-batch culture were closely analyzed. It was shown that the phenylalanine, tyrosine and tryptophan biosynthesis pathways were significantly altered under different culture conditions with different media. These changes were more apparent in the fed-batch process in which higher mAb titer was observed due to the metabolic changes than mAb titer in the batch process. The pathway analysis approach was well utilized for evaluating the impact on the relevant pathways involved under different cell culture conditions to improve cell growth and mAb titer. © 2018 American Institute of Chemical Engineers Biotechnol. Prog., 34:793-805, 2018.

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“…Glycosylation is another aspect that has to be taken into consideration as change in glycosylation pattern could affect functionality as well as impact PK/PD characteristics of mAbs (Liu 2015). Successful attempts have been made from a bioengineering point of view to investigate the effects of the production process on glycosylation profiles of monoclonal antibodies by using multivariate techniques, such as principal component analysis, partial least squares and parallel factor analysis (Green and Glassey 2015; Glassey 2012). Glycoengineered antibodies were produced by CHO cells with higher glycosyltransferase which enabled the production of engineered antibodies with the N-acetylglucosamine profiles required to achieve higher neutrophil-mediated phagocytosis activity and thus greater efficacy in killing tumour cells (Umaña et al 1999; Golay et al 2013).…”

Section: Discussion: Status Quo and Scope For Mab-based Applicationmentioning

confidence: 99%

Applicability of predictive toxicology methods for monoclonal antibody therapeutics: status Quo and scope

2016

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Biopharmaceuticals, monoclonal antibody (mAb)-based therapeutics in particular, have positively impacted millions of lives. MAbs and related therapeutics are highly desirable from a biopharmaceutical perspective as they are highly target specific and well tolerated within the human system. Nevertheless, several mAbs have been discontinued or withdrawn based either on their inability to demonstrate efficacy and/or due to adverse effects. Approved monoclonal antibodies and derived therapeutics have been associated with adverse effects such as immunogenicity, cytokine release syndrome, progressive multifocal leukoencephalopathy, intravascular haemolysis, cardiac arrhythmias, abnormal liver function, gastrointestinal perforation, bronchospasm, intraocular inflammation, urticaria, nephritis, neuropathy, birth defects, fever and cough to name a few. The advances made in this field are also impeded by a lack of progress in bioprocess development strategies as well as increasing costs owing to attrition, wherein the lack of efficacy and safety accounts for nearly 60 % of all factors contributing to attrition. This reiterates the need for smarter preclinical development using quality by design-based approaches encompassing carefully designed predictive models during early stages of drug development. Different in vitro and in silico methods are extensively used for predicting biological activity as well as toxicity during small molecule drug development; however, their full potential has not been utilized for biological drug development. The scope of in vitro and in silico tools in early developmental stages of monoclonal antibody-based therapeutics production and how it contributes to lower attrition rates leading to faster development of potential drug candidates has been evaluated. The applicability of computational toxicology approaches in this context as well as the pitfalls and promises of extending such techniques to biopharmaceutical development has been highlighted.Electronic supplementary materialThe online version of this article (doi:10.1007/s00204-016-1876-7) contains supplementary material, which is available to authorized users.

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Multivariate analysis of the effect of operating conditions on hybridoma cell metabolism and glycosylation of produced antibody

Cited by 19 publications

References 26 publications

Metabolic Control in Mammalian Fed-Batch Cell Cultures for Reduced Lactic Acid Accumulation and Improved Process Robustness

Metabolic Control in Mammalian Fed-Batch Cell Cultures for Reduced Lactic Acid Accumulation and Improved Process Robustness

Metabolic responses and pathway changes of mammalian cells under different culture conditions with media supplementations

Applicability of predictive toxicology methods for monoclonal antibody therapeutics: status Quo and scope

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