Process‐induced cell cycle oscillations in CHO cultures: Online monitoring and model‐based investigation

Möller, J.; Bhat, Krathika; Riecken, Kristoffer; Pörtner, Ralf; Zeng, An‐Ping; Jandt, Uwe

doi:10.1002/bit.27124

Cited by 16 publications

(18 citation statements)

References 68 publications

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“…The transition of the cultures into the stationary growth phase occurs along with apoptosis and necrosis [49,50]. Furthermore, growth inhibitory media components can accumulate (e.g., ammonium and putative autocrine factors), which can lead to changes in the metabolism [16,17,51]. In this study, a different bulk cell growth was observed for mixed shake flask cultures comprised of populations from different growth phases.…”

Section: Discussionmentioning

confidence: 87%

“…Automated and online flow cytometry An automated and online flow cytometry (AFC) set-up was previously introduced [17] and also applied in this study. In brief, a flow cytometer (Beckman Coulter, Germany) was connected to the bioreactor, and samples were drawn automatically every hour.…”

Section: Case Study IImentioning

confidence: 99%

“…The total cell number (X t ) was determined with gating, and the viable cell density (X v ) was calculated from offline samples and re-iterated on the online data. More information about the AFC set-up can be found in ESM and in [17].…”

Section: Case Study IImentioning

confidence: 99%

“…However, population heterogeneities can significantly affect the bioprocess performance, e.g., with differences in the intraclonal protein expression or metabolically different cell populations [4][5][6][7][8]. It was shown that even in monoclonal CHO cell lines ( [9], recently discussed in [10]), population heterogeneities with different phenotypes can spontaneously evolve [4,[11][12][13], and that cell-cycle-dependent population dynamics with variable productivities and metabolic regulations can be present in cell culture processes [14][15][16][17]. The formation and dynamics of physically-derived process heterogeneities are studied intensively in bioprocesses, e.g., evaluating spatial gradients of gas bubbles [18], concentration gradients of substrates [19] and oxygen [20], and pH gradients [21].…”

Section: Introductionmentioning

confidence: 99%

“…These individually cultivated cells were pooled together and used for inoculation of mixed shake flask cultures, which were tracked using offline flow cytometry. Automated (online) flow cytometry [17] was used in addition to assess population changes in a bioreactor experiment with the same mixed population.…”

Section: Introductionmentioning

confidence: 99%

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Quantification of the dynamics of population heterogeneities in CHO cultures with stably integrated fluorescent markers

et al. 2020

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Cell population heterogeneities and their changes in mammalian cell culture processes are still not well characterized. In this study, the formation and dynamics of cell population heterogeneities were investigated with flow cytometry and stably integrated fluorescent markers based on the lentiviral gene ontology (LeGO) vector system. To achieve this, antibodyproducing CHO cells were transduced with different LeGO vectors to stably express single or multiple fluorescent proteins. This enables the tracking of the transduced populations and is discussed in two case studies from the field of bioprocess engineering: In case study I, cells were co-transduced to express red, green, and blue fluorescent proteins and the development of sub-populations and expression heterogeneities were investigated in high passage cultivations (total 130 days). The formation of a fast-growing and more productive population was observed with a simultaneous increase in cell density and product titer. In case study II, different preculture growth phases and their influence on the population dynamics were investigated in mixed batch cultures with flow cytometry (offline and automated). Four cell line derivatives, each expressing a different fluorescent protein, were generated and cultivated for different time intervals, corresponding to different growth phases. Mixed cultures were inoculated from them, and changes in the composition of the cell populations were observed during the first 48 h of cultivation with reduced process productivity. In summary, we showed how the dynamics of population heterogeneities can be characterized. This represents a novel approach to investigate the dynamics of cell population heterogeneities under near-physiological conditions with changing productivity in mammalian cell culture processes.

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Section: Discussionmentioning

confidence: 87%

Section: Case Study IImentioning

confidence: 99%

Section: Case Study IImentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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Quantification of the dynamics of population heterogeneities in CHO cultures with stably integrated fluorescent markers

et al. 2020

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Analysis of Transformed Upstream Bioprocess Data Provides Insights into Biological System Variation

Richelle¹,

Lee²,

Portela³

et al. 2020

Biotechnology Journal

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In recent years, multivariate data analysis (MVDA) and modeling approaches have found increasing applications for upstream bioprocess studies (e.g., monitoring, development, optimization, scale-up, etc.). Many of these studies look at variations in the concentrations of metabolites and cell-based measurements. However, these measures are subject to system inherent variations (e.g., changes in metabolic activity) but also intentional operational changes. It is proposed to perform MVDA and modeling on data representative of the underlying biological system operation, that is, the specific rates, which are per se independent of the scale, operational strategy (e.g., batch, fed-batch), and biomass content. Two industrial case studies are highlighted to showcase the approach: one is HEK medium performance comparison study and the other is CHO scale-up/-down study. It is shown that analyzing processes in this way reveals insights into behavior of the underlying biological system, which cannot to the same degree be deducted from the analysis of concentrations.

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Comparison of mechanistic and hybrid modeling approaches for characterization of a CHO cultivation process: Requirements, pitfalls and solution paths

Bayer¹,

Duerkop²,

Pörtner

et al. 2022

Biotechnology Journal

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Despite the advantages of mathematical bioprocess modeling, successful model implementation already starts with experimental planning and accordingly can fail at this early stage. For this study, two different modeling approaches (mechanistic and hybrid) based on a four-dimensional antibody-producing CHO fed-batch process are compared. Overall, 33 experiments are performed in the fractional factorial fourdimensional design space and separated into four different complex data partitions subsequently used for model comparison and evaluation. The mechanistic model demonstrates the advantage of prior knowledge (i.e., known equations) to get informative value relatively independently of the utilized data partition. The hybrid approach displayes a higher data dependency but simultaneously yielded a higher accuracy on all data partitions. Furthermore, our results demonstrate that independent of the chosen modeling framework, a smart selection of only four initial experiments can already yield a very good representation of a full design space independent of the chosen modeling structure. Academic and industry researchers are recommended to pay more attention to experimental planning to maximize the process understanding obtained from mathematical modeling.

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Process‐induced cell cycle oscillations in CHO cultures: Online monitoring and model‐based investigation

Cited by 16 publications

References 68 publications

Quantification of the dynamics of population heterogeneities in CHO cultures with stably integrated fluorescent markers

Quantification of the dynamics of population heterogeneities in CHO cultures with stably integrated fluorescent markers

Analysis of Transformed Upstream Bioprocess Data Provides Insights into Biological System Variation

Comparison of mechanistic and hybrid modeling approaches for characterization of a CHO cultivation process: Requirements, pitfalls and solution paths

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