A number of biological medicinal products are derived from cells that either: naturally produce the substance desired, are genetically modified to do so, or as is the case with viral vaccines, are inoculated with vaccine virus which then replicates to produce large quantities. Traditionally animal cells have been used for the production of viral vaccines, although one of the commercially available recombinant human papillomavirus virus-like particle vaccines is produced in insect cells.
Cell substrate refers to the cells that are used to produce the desired biotechnological/biological products. Cell substrates may be microbial cells (e.g. yeast) or cells derived from various animal sources. Within the animal cell group, there are a number of cell types use for production:
- primary cells or tissues (used without passage in tissue culture)
- diploid cells (cells with a finite lifespan and passaged in tissue culture), or
- continous cell lines (cells with an apparently unlimited capacity to replicate).
Cell substrates also differ in their host range (ability to be infected or transfected by microorganisms) and their degree of differentiation.
Primary cells are obtained by dissociating cells from specific tissues, and many vaccines continue to be produced on such highly differentiated cell substrates (e.g. primary monkey kidney cells).
Diploid cell lines contain the full complement of genetic material, often retain many characteristics of the cell types from which they were derived, and normally cease to replicate in cell culture conditions after a limited number of generations. On the other hand, continuous cell lines replicate in cell culture for indefinite periods, but often lose the characteristics of their parent cells.
To reduce variability and avoid potential contaminants, various diploid and continuous cell lines have been established and characterized. Because these well characterized cells can be stored, they have improved the consistency of production of biological products including vaccines.
Cell substrates differ in the degree to which they can be adapted to grow in defined culture media such as serum-free media. This is desirable for production in order to avoid potential contaminants from serum. Cell substrates also have different characteristics regarding their adhesiveness to solid supports. For example, some cell substrates can grow on microcarrier beads suspended in a large fermenter, a technique capable of generating high yields of consistent products.
The challenge in standardizing cell substrates is to balance the desire for a very efficient production system with the goal of minimizing risks. The current approach to manufacturing a safe product includes the thorough characterization of the cell substrate, validation of the manufacturing process for removal and/or inactivation of potential adventious agents; and testing of the bulk and final product.
The objectives of cell line characterization are to confirm the identity and purity of the cell substrate and to provide a high level of confidence in its safety as a component in the manufacture of biologicals. To facilitate this, WHO regularly reviews current practices in the use of cells as production substrates, evaluates the risks inherent in specific cell types, and makes recommendations to manufacturers and regulatory authorities on their correct handling and use.
WHO has generated stocks of the widely used Vero cell line (Vero 10-87) for use as a cell seed from which master cell banks may be established through additional qualification. This is distributed to qualified vaccine manufacturers upon request.
Cell Substrates standardization
The first WHO requirements for cell cultures used in the production of biologicals were formulated in 1959 as part of the production of inactivated poliomyelitis vaccine. The 1996 revision describes the characterization and testing of continuous-cell line and diploid cell substrates, with the general manufacturing requirements applicable to primary cell substrates as well.
In 2010, the ECBS adopted the WHO Recommendations for the evaluation of animal cell cultures as substrates for the manufacture of biological medicinal products and for the characterization of cell banks.
Recommendations for the Evaluation of Animal Cell Cultures as Substrates for the Manufacture of Biological Medicinal Products and for the Characterization of Cell banks. Proposed replacement of TRS, 878, Annex 1 - ECBS 18 to 22 October 2010