Use this URL to cite or link to this record in EThOS:
Title: Cascading effects in bioprocessing : the impact of cell culture environment on CHO cell behaviour and host cell protein species
Author: Goey, Cher Hui
ISNI:       0000 0004 6422 7714
Awarding Body: Imperial College London
Current Institution: Imperial College London
Date of Award: 2017
Availability of Full Text:
Access from EThOS:
Access from Institution:
One of the reasons for the rejection of new drugs during clinical trials is the presence of host cell proteins (HCPs) in the drug formulation. HCPs are immunogenic impurities that can compromise patient safety. Moreover, proteolytic and binding HCPs compromise the integrity, and, hence, the stability and efficacy of a recombinant protein. Therefore, HCPs should be removed from the bioprocess train as soon as possible. Current downstream purification platforms are challenged by HCP-mAb co-elution. A Quality by Design strategy to overcome this problem is to reduce the number of HCPs in the downstream feedstock by tracing their source back to upstream culture and eliminating it. Previous studies have shown that upstream cell culture parameters, e.g. harvest time and culture temperature, significantly affect HCP profiles. However, little is known about how host cells coordinate and regulate their molecular machinery under different cell culture environment that results in different HCP profiles. This study presents experimental results linking cell culture temperature and key process indicators of CHO cell cultures and post-Protein A purification (mAb titre, HCP level and HCP species) by considering the cellular behaviour in terms of cell growth, cell cycle distribution and cell health). This study involved the application of single-use fed-batch bioreactors to culture IgG4 producing GS-CHO cell line, cell health and cell cycle analysis with NucleoCounter, Protein A purification and proteomic analysis with HCP ELISA kits and LC-MS/MS. Cells were more robust under mild hypothermia: over 90% of cells were maintained in a healthy state until the decline phase, and the onset of apoptosis was less evident compared to the results for physiological temperature. IgG4 titre and HCP level at harvest were comparable between the two cases. However, mild hypothermia reduced the HCP variety in HCCF by 36%, including 44% and 27% lower proteases and chaperones, respectively. The differences in HCP variety at harvest resulted in a significantly different HCP profile post-Protein A purification between the two cases. Half of the critically immunogenic HCPs species as determined by the CHOPPI tool were different between the two cases. This study shows that cell culture conditions significantly affect the HCP profile at harvest and that of purified samples.
Supervisor: Kontoravdi, Cleo Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral