The cell line integrity
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The cell lines used to manufacture biopharmaceuticals must meet strict quality criteria to ensure high-quality, stable products. Master cell banks are critical starting materials, and should be sterile, mycoplasma-free, and tested for identity and purity. Cell lines must stably express the drug with high productivity, appropriate quality and structure, and be robust enough for large-scale production. Comprehensive testing of cell lines and banks is required by guidelines to confirm identity, purity, freedom from contamination, and genetic stability.
The cell line integrity
- 2. JensMartensson The quality of the cell lines used to manufacture biopharmaceuticals are crucial for the production of high-quality, stable biopharmaceuticals. 2 Aug 01, 2018 By Feliza Mirasol BioPharm International Volume 31, Issue 8, pg 10–15
- 3. JensMartensson 3 Cell banks are critical starting materials for the production of biological products and, as such, the quality of these cell banks directly affects the characteristics and safety of the products. Master cell banks (MCBs) should be prepared from seed cells that, at a minimum, have tested negative in compendial sterility and mycoplasma tests. Tingfeng Lai, et al. Advances in Mammalian Cell Line Development Technologies for Recombinant Protein Production. Pharmaceuticals (Basel). 2013 Apr 26;6(5):579-603 SOA/Bio-MCteam
- 5. JensMartensson Cell lines used for manufacturing processes need to fulfill the following criteria: 5 1- Cell lines need to be capable of producing the drug substance with high productivity. For example, CHO cell lines are the most frequently applied mammalian manufacturing cell lines in the biopharmaceutical industry. They usually express classical monoclonal therapeutic antibodies with titers of more than 5 g/L.
- 6. JensMartensson Cell lines used for manufacturing processes need to fulfill the following criteria: 6 2- Cell lines need to produce the drug molecules with the appropriate quality, meaning that the structure of the molecule needs to be ‘as expected’. For example, it is known that CHO cell lines express human antibodies with a suitable glycan pattern.
- 7. JensMartensson Cell lines used for manufacturing processes need to fulfill the following criteria: 7 3- Cell lines need to accept foreign DNA, which provides the coding instruction for the drug molecule together with elements directing the gene expressions (e.g., promoters, enhancers, terminator structures). The cell lines need to stably keep this genetic information within the cells, and they need to be able to stably express this genetic information with a reproducibly high productivity over a long time (i.e., in the case of mammalian cell lines over many generation times for months, and after freezing and thawing cycles)..
- 8. JensMartensson Cell lines used for manufacturing processes need to fulfill the following criteria: 8 4- Cell lines need to be robust enough to grow to high density in large bioreactors (i.e., thousands of liters) 5- The cell lines need to be free of any virus or other harmful contamination and should not have been in contact with serum or undefined media components.
- 9. JensMartensson 9 Culture mode • Most mammalian upstream processes, in which cells grow to high cell density and produce the drug molecules, run as fed-batch processes. This means that the cells grow over a period of approximately two weeks with optimal feeding and then are harvested. Perfusion • Some upstream processes run as continuous processes, in which cell density is kept constant, while media is continuously added and removed. • Usually a cell line that is well suited for fed-batch processes is not necessarily good in continuous culturing processes and vice versa. Fed-batch
- 11. JensMartensson The analytical processes required to test cell-line quality 11
- 12. JensMartensson The details in the testing program can vary depending on the source of the cell line as well as on the final product. However, there are standard analytical procedures to be followed as described in national and international guidelines and as summarized in the International Conference on Harmonization (ICH) Q5D 12
- 13. JensMartensson 13 test of cell´s identity (e.g., short tandem repeat analysis) test of purity (absence of adventitious cellular or microbial contaminants and potential cross-contaminations with other cell lines) the complete package of in vitro, in vivo, and polymerase chain reaction (PCR)-based assays for adventitious agents. Depending on the source of the cell and the final product, tests for tumorigenicity of the cells and oncogenicity of the cell´s genomic DNA might be required.
- 14. JensMartensson During bioprocessing and manufacturing of the product, cell integrity should be further monitored in realtime to evaluate the process and the cell´s performance. Critical parameters, such as stability of cell doubling time, productivity, and a cell’s metabolite profile are subject to continuous monitoring. 14
- 15. JensMartensson Cells should be purchased only from qualified vendors that have been audited by the purchasing laboratory. At minimum, the vendor should provide a full history of the cells being provided along with a CoA demonstrating the cells to be free of bacterial and mycoplasma contamination. Clear details as to the sterility and mycoplasma testing methodologies should be provided and/or the purchaser should audit these testing methods during vendor qualification. A simple negative result on the CoA should not be considered sufficient. 15
- 16. JensMartensson With regard to the MCB testing should include: 1- identity testing. 2- sterility and mycoplasma testing 3- viral testing : using a variety of suitable tests designed to be both broad ranging and specific for particular viruses. 4- Genetic stability testing should : performed using methods such as good manufacturing practices (GMP) sequencing, Northern and Southern blotting, and copy number by quantitative polymerase chain reaction (qPCR). 16
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