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Brunner, Matthias

Loeffler, Michael

Schaub, Jochen

Unsoeld, Andreas

Wucherpfennig, Thomas

English

Engineering Conferences International

Poster Number 96 ADVANCED CELL CULTURE PERFORMANCE BY RATIONAL MEDIA DESIGN BASED ON IN DEPTH PROCESS UNDERSTANDING   Michael Loeffler, Bioprocess Development Biologicals, Boehringer Ingelheim Michael.loeffler@boehringer-ingelheim.com Andreas Unsoeld, Bioprocess Development Biologicals, Boehringer Ingelheim Matthias Brunner, HP BioP Mammalian, Boehringer Ingelheim Thomas Wucherpfennig, Bioprocess Development Biologicals, Boehringer Ingelheim Jochen Schaub, Bioprocess Development Biologicals, Boehringer Ingelheim   Key Words: Cysteine, titer optimum, media optimization, feed, intracellular metabolism  Monoclonal antibodies are the leading class of biopharmaceuticals. These products are mainly produced in processes with CHO cells. A robust cell culture media platform becomes key to develop an efficient and robust production process. A comprehensive screening was performed to detect substances with the potential to improve the cell culture performance without impacting the media stability. Nineteen substances with potential effects on cysteine metabolism, cell cycle, reactive oxygen scavenger, histone deacetylase inhibitor or intracellular cAMP were spiked to the standard fed-batch process in different concentrations. Especially substances impacting the cysteine metabolism showed the ability to prolong specific productivity and viability during the last days of the fed-batch process. The intracellular data revealed a metabolic understanding of the connection between cysteine concentration and detox pathways (Figure 1).  Additional optimization experiments focusing on the cysteine metabolism showed a sweet spot with an increased process performance (viability and titer) without negatively affecting PQ and media stability (Figure 2).   Among the high number of media components, cysteine is of particular interest as it plays a pivotal role in achieving prolonged cell viability and high product concentrations. Furthermore, it also has a high impact on PQ, media reactivity and stability.          Figure 2 - Cys sweet spot to ensure titer optimum without affecting PQ. 0 5 10 1502468Process time [day]Homocysteine [µM]0 5 10 1505101520Process time [day]Cysteine [µM]0 5 10 150123  y-GluCys [µM]0 5 10 150200400600800  GSH [µM]0 5 10 15020406080  GSSG [µM]0 5 10 15050100150Process time [day]5-Oxoproline [µM]0 5 10 15012345Process time [day]Cystathione [µM]0 5 10 15050010001500Process time [day]Serine [µM]0 5 10 150,00,10,20,3Process time [day]Cysteine sulfinic acid [µM]0 5 10 15050100150200250  Taurine [µM]Cys conc. 1Cys conc.2Basal Cys conc.0 5 10 150200400600800Process time [day]Glutamate [µM]0 5 10 150200040006000Process time [day]Glycine [µM]MethionineHomocysteineSerineCystathionineCysteineCysteine Sulfinic AcidHypotaurineTauriney-Glutamyl-CysGlutathioneDisulfide (GSSG)Cys-GlyGlycineGlutamateFree Amino Acidy-Glutamyl-Amino AcidFree Amino AcidProteinSynthesisAntioxidant RoleDetox Pathway5-OxoprolineGlutathione (GSH)Figure 1 - Exemplary overview of intracellular metabolites associated with Cys metabolism when feeding different amounts of Cys. 

Advanced cell culture performance by rational media design based on in depth process understanding

https://dc.engconfintl.org/cgi/viewcontent.cgi?filename=0&article=1143&context=ccexviii&type=additional

Advanced cell culture performance by rational media design based on in depth process understanding

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Engineering Conferences International