Model uncertainty-based evaluation of process strategies during scale-up of biopharmaceutical processes

Möller, J.; Rodríguez, Tanja Hernández; Müller, Jan; Arndt, Lukas; Kuchemüller, Kim B.; Frahm, Björn; Eibl, Regine; Eibl, Dieter; Pörtner, Ralf

doi:10.1016/j.compchemeng.2019.106693

Cited by 28 publications

(59 citation statements)

References 52 publications

(53 reference statements)

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“…The peak antibody titer was

c_{normalAb} = 85 mg {normalL}^{- 1}

at the end of the cultivation, as seen in Figure 2H. The product concentrations were comparable to previous studies using the same medium and cell line [43, 48–50].…”

Section: Resultssupporting

confidence: 85%

“…Hence, any effects of pH on cell metabolism were absent. Second, many studies report lactate consumption to occur close to or during the stationary phase, which is usually influenced by cell death mechanisms [42–45]. Therefore, the initial glucose concentration in the medium was reduced to

20 mmol {normalL}^{- 1}

(typically

42 mmol {normalL}^{- 1}

) to maintain steady cell growth and lactate uptake.…”

Section: Resultsmentioning

confidence: 99%

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Regulation of pyruvate dehydrogenase complex related to lactate switch in CHO cells

Möller

Bhat

Guhl

et al. 2020

Engineering in Life Sciences

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The metabolism of Chinese hamster ovary (CHO) cell lines is typically characterized by high rates of aerobic glycolysis with increased lactate formation, known as the ”Warburg” effect. Although this metabolic state can switch to lactate consumption, the involved regulations of the central metabolism have only been partially studied so far. An important reaction transferring the lactate precursor, pyruvate, into the tricarboxylic acid cycle is the decarboxylation reaction catalyzed by the pyruvate dehydrogenase enzyme complex (PDC). Among other mechanisms, PDC is mainly regulated by phosphorylation–dephosphorylation at the three sites Ser232, Ser293, and Ser300. In this work, the PDC phosphorylation in antibody‐producing CHO DP‐12 cell culture is investigated during the lactate switch. Batch cultivations were carried out with frequent sampling (every 6 h) during the transition from lactate formation to lactate uptake, and the PDC phosphorylation levels were quantified using a novel indirect flow cytometry protocol. Contrary to the expected activation of PDC (i.e., reduced PDC phosphorylation) during lactate consumption, Ser293 and Ser300 phosphorylation levels were 33% higher compared to the phase of glucose excess. At the same time, the relative phosphorylation level of Ser232 increased steadily throughout the cultivation (66% increase overall). The intracellular pyruvate was found to accumulate only during the period of high lactate production, while acetyl‐CoA showed nearly no accumulation. These results indicate a deactivation of PDC and reduced oxidative metabolism during lactate switch even though the cells undergo a metabolic transition to lactate‐based cell growth and metabolism. Overall, this study provides a unique view on the regulation of PDC during the lactate switch, which contributes to an improved understanding of PDC and its interaction with the bioprocess.

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“…The peak antibody titer was

c_{normalAb} = 85 mg {normalL}^{- 1}

at the end of the cultivation, as seen in Figure 2H. The product concentrations were comparable to previous studies using the same medium and cell line [43, 48–50].…”

Section: Resultssupporting

confidence: 85%

20 mmol {normalL}^{- 1}

(typically

42 mmol {normalL}^{- 1}

) to maintain steady cell growth and lactate uptake.…”

Section: Resultsmentioning

confidence: 99%

Regulation of pyruvate dehydrogenase complex related to lactate switch in CHO cells

Möller

Bhat

Guhl

et al. 2020

Engineering in Life Sciences

Self Cite

View full text Add to dashboard Cite

show abstract

“…The identified population changes can affect process variability with different productivities in large scale, e.g., if cells are pooled from different bioreactors for inoculation [6,22]. Moreover, this study demonstrates the strong impact of preculture treatment on the bulk process performance and the need for an efficient seed train design [54] and scale-up strategy [55]. Regarding the application of the cell labeling concept, the changes in the bulk composition behavior would be not observable if the different populations were not individually marked.…”

Section: Discussionmentioning

confidence: 80%

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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“…To overcome the previously mentioned limitations of DoE, a new model-assisted Design of Experiments (mDoE) concept was recently introduced for knowledge-driven bioprocess development and optimization [ 1 , 14 , 19 ]. In the mDoE approach, the recommended experiments in statistical DoE designs are simulated using mathematical process models instead of being performed experimentally.…”

Section: Introductionmentioning

confidence: 99%

Model-assisted DoE software: optimization of growth and biocatalysis in Saccharomyces cerevisiae bioprocesses

Moser

Kuchemüller

Deppe

et al. 2021

Bioprocess Biosyst Eng

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Bioprocess development and optimization are still cost- and time-intensive due to the enormous number of experiments involved. In this study, the recently introduced model-assisted Design of Experiments (mDoE) concept (Möller et al. in Bioproc Biosyst Eng 42(5):867, 10.1007/s00449-019-02089-7, 2019) was extended and implemented into a software (“mDoE-toolbox”) to significantly reduce the number of required cultivations. The application of the toolbox is exemplary shown in two case studies with Saccharomyces cerevisiae. In the first case study, a fed-batch process was optimized with respect to the pH value and linearly rising feeding rates of glucose and nitrogen source. Using the mDoE-toolbox, the biomass concentration was increased by 30% compared to previously performed experiments. The second case study was the whole-cell biocatalysis of ethyl acetoacetate (EAA) to (S)-ethyl-3-hydroxybutyrate (E3HB), for which the feeding rates of glucose, nitrogen source, and EAA were optimized. An increase of 80% compared to a previously performed experiment with similar initial conditions was achieved for the E3HB concentration.

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Model uncertainty-based evaluation of process strategies during scale-up of biopharmaceutical processes

Cited by 28 publications

References 52 publications

Regulation of pyruvate dehydrogenase complex related to lactate switch in CHO cells

Regulation of pyruvate dehydrogenase complex related to lactate switch in CHO cells

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

Model-assisted DoE software: optimization of growth and biocatalysis in Saccharomyces cerevisiae bioprocesses

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