Medium development beyond production media: Chemically defined media for transfection and single cell cultivation

Beckmann, Tim; Klausing, Sandra; Püngel, Sebastian; Coronel, Juliana; Welsink, Tim; Heinrich, Christoph A.

doi:10.1186/1753-6561-9-s9-p27

Cited by 1 publication

(5 citation statements)

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“…24 Cells were routinely cultivated in a customized chemically-defined ACF medium (TC-LECC, Xell AG), supplemented with 0.1 mg/L LONG R 3 IGF-I, in baffled shake flasks at 180 rpm, 37 C, and 5% CO 2. Wells were monitored over time using Cellavista (SynenTec) to determine which wells contained colonies originating from a single cell.…”

Section: Cho Cell Clone and Subclonementioning

confidence: 99%

“…Wells were monitored over time using Cellavista (SynenTec) to determine which wells contained colonies originating from a single cell. 24 Cells were routinely cultivated in a customized chemically-defined ACF medium (TC-LECC, Xell AG), supplemented with 0.1 mg/L LONG R 3 IGF-I, in baffled shake flasks at 180 rpm, 37 C, and 5% CO 2. The same medium was used for all cultivations, including as feed of the perfusion bioreactor.…”

Section: Cho Cell Clone and Subclonementioning

confidence: 99%

“…As an approach to further increase perfusion process productivity, cultivations with a high-producer subclone isolated from the parental clone 24 were carried out. Prior to testing it in perfusion cultures, we subcultured it in parallel to the parental clone for 4 weeks in spin tubes.…”

Section: Perfusion Cultivations At 31 C With Va With a High-producementioning

confidence: 99%

“…15 Depending on the product characteristics (e.g., protein structure), the short half-life can vary between~11 and 22 hr. 24 This subclone was then used for the production of FVIII in perfusion mode under the conditions of temperature and fattyacid supplementation optimized for the parental clone, enabling further enhancement of FVIII productivity. 17 For improving recombinant FVIII production, a screening for cultivation conditions in batch was done by our group and significant increases in q p were obtained at low temperatures with short-chain fatty acids supplementation.…”

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confidence: 99%

“…Based on the knowledge of CHO genome instability and heterogeneity of cell populations gathered over the recent years, [19][20][21][22][23] we additionally proceeded with subclone isolation from the parental clone in order to screen for a higher producer subclone. 24 This subclone was then used for the production of FVIII in perfusion mode under the conditions of temperature and fattyacid supplementation optimized for the parental clone, enabling further enhancement of FVIII productivity.…”

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confidence: 99%

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Perfusion process combining low temperature and valeric acid for enhanced recombinant factor VIII production

Coronel

Heinrich²,

Klausing³

et al. 2019

Biotechnology Progress

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Perfusion operation mode remains the preferred platform for production of labile biopharmaceuticals (e.g., blood factors) and is also being increasingly adopted for production of stable products (e.g., monoclonal antibodies). Regardless of the product, process development typically aims at maximizing production capacity. In this work, we investigated the impact of perfusion cultivation conditions on process productivity for production of human factor VIII (FVIII). Recombinant CHO cells were cultivated in bioreactors coupled to inclined settlers and the effects of reducing the temperature to 31 C with or without valeric acid (VA) supplementation were evaluated. Increases in cell specific productivity (q p ) up to 2.4-fold (FVIII concentration) and up to 3.0-fold (FVIII biological activity) were obtained at 31 C with VA compared to the control at 37 C. Biological activity is the most important quality attribute for FVIII and was positively affected by mild hypothermia in combination with the chemical inducer. The low temperature conditions resulted in enhanced product transcript levels, suggesting that the higher q p is related to the increased mRNA levels. Furthermore, a high-producer subclone was evaluated under the perfusion conditions optimized for the parental clone (31 C with VA), yielding increases in q p of 6-fold and 15-fold compared to the parental clone cultivated under the same condition and at 37 C, respectively. The proposed perfusion strategy enables increased product formation without increasing production costs, being potentially applicable to perfusion production of other CHO-derived biopharmaceuticals. To the best of our knowledge, this is the first report showing the benefits of perfusion combining mild hypothermia with VA supplementation. K E Y W O R D SCHO perfusion cultivation, mild hypothermia, productivity, recombinant factor VIII, valeric acid

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Section: Cho Cell Clone and Subclonementioning

confidence: 99%

Section: Cho Cell Clone and Subclonementioning

confidence: 99%

Section: Perfusion Cultivations At 31 C With Va With a High-producementioning

confidence: 99%

mentioning

confidence: 99%