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Optimized Selection Marker and CHO Host Cell Combinations for Generating High Monoclonal Antibody Producing Cell
Jessna H M Yeo1, Steven C L Ho1, Mariati Mariati1
1Bioprocessing Technology Institute, Agency for Science, Technology and Research (A*STAR), Singapore, Singapore.
Biotechnology Journal
|November 2, 2017
Summary
Optimizing selection markers in Chinese hamster ovary (CHO) cells enhances antibody production. Different markers and stringencies improve cell line productivity and stability for biopharmaceutical manufacturing.
Area of Science:
- Biotechnology
- Cell Line Development
- Protein Expression
Background:
- Chinese hamster ovary (CHO) cell lines are crucial for biopharmaceutical production.
- Selection markers are essential for generating stable, high-producing cell lines.
- Optimizing selection stringency is key to maximizing productivity.
Purpose of the Study:
- To optimize expression of antibiotics resistance genes (ARGs) and dihydrofolate reductase (DHFR) as selection markers in CHO-K1 and CHO DG44 cells, respectively.
- To investigate the impact of selection marker expression levels on cell line productivity and stability.
- To compare the production stability and characteristics of CHO K1 and CHO DG44 cell lines generated using different selection strategies.
Main Methods:
- Utilized an internal ribosome entry site (IRES)-mediated tricistronic vector with varying IRES strengths.
- Optimized expression of five ARGs in CHO-K1 cells and DHFR in CHO DG44 cells.
- Assessed cell line productivity, gene copy number, expression stability, and monoclonal antibody (mAb) glycan profiles.
Main Results:
- Identified optimal expression levels for selection markers, with significant productivity drops outside this range.
- Enhanced productivity in ARG-selected CHO K1 cells resulted from selective active site integration; in DHFR-amplified CHO DG44 cells, it was due to increased gene copies.
- CHO K1 cell lines exhibited superior production stability compared to CHO DG44 cell lines, with distinct mechanisms of expression loss (gene copy loss vs. transcriptional silencing).
- Significant differences in mAb glycan profiles were observed between CHO K1 and CHO DG44 cell lines.
Conclusions:
- Optimized selection marker expression and stringency are critical for developing high-producing and stable CHO cell lines.
- ARG selection in CHO K1 cells offers better production stability than DHFR amplification in CHO DG44 cells.
- Understanding cell line-specific mechanisms of productivity enhancement and expression loss is vital for bioprocess development.
- Differences in mAb glycosylation highlight the importance of cell line choice for specific product quality attributes.
