Characterisation of Recombinant Cell Lines

A couple of quality concerns for cell-derived biological products have originated from the possible presence of adventitious contaminants or from the properties of the cells used to prepare the product. Recombinant DNA (rDNA) - derived products also carry quality concerns regarding the expression construct contained in the cell substrate. Thus, it is well established that the properties of the cell substrate and events linked to generation of the cell substrate can affect resultant product quality and safety and, further, that effective quality control of these products requires appropriate controls on all aspects of handling the cell substrate.

Especially for biopharmaceutical products the characterisation and safety testing of cell substrates are the initial steps in warranting the safety. The intention of characterisation is to confirm the identity and purity of the cell substrate. The cell line is additionally tested for safety to make certain that it is free of adventitious agents which could potentially compromise the biopharmaceutical product. All the tests required to prove the suitability of the cell line for biopharmaceutical production must be performed to GLP/GMP standards using validated test procedures and according to the international guidelines, as listed:

  • ICH harmonised guideline Q5A: Viral Safety Evaluation of Biotechnology Products Derived from Cell Lines of Human or Animal Origin (1997) (Download document from ICH site)
  • ICH harmonised guideline Q5B: Quality of Biotechnological Products: Analysis of the Expression Construct in Cells Used for Production of r-DNA Derived Protein Products (1996) (Download document from ICH site)
  • ICH harmonised guideline Q5D: Derivation and Characterisation of Cell Substrates Used for Production of Biotechnological/Biological Products (1997) (Download document from ICH site)
  • ICH harmonised guideline Q6B: Specifications: Test Procedures and Acceptance Criteria for Biotechnological/Biological Products (1999) (Download document from ICH site)
  • ICH harmonised guideline S6: Preclinical Safety Evaluation of Biotechnology-Derived Pharmaceuticals (1997) (Download document from ICH site)
  • EMEA:CHMP: Note for Guidance on Minimising the Risk of Transmitting Animal Spongiform Encephalopathy Agents via Human and Veterinary Medicinal Products (2004, DRAFT, EMEA/410/01 Rev. 3) (Download document from EMEA site)
  • EMEA/CHMP: Note for Guidance on the Use of Bovine Serum in the Manufacture of Human Biological Medicinal Products (CPMP/BWP/1793/02 - August 2003) (Download document from EMEA site)

The test routine used for the cell line characterisation and safety testing depends upon the specific production system, the intended use of the product, the history of the expression system and the used culture media and supplementary reagents (e.g., FCS, trypsin). The following table provides an tests suitable for the characterisation of eukaryotic cell lines and the product.


Eukaryotic Cells

Aspect

Method

Identity

Isoenzyme pattern
Karyotyping
Banding cytogenetics
FACS analysis
Southern / Northern blot analysis
Plasmid copy number
RT-PCR
DNA sequencing

Stabilty during Production

Productivity
Karyotyping
Banding cytogenetics
FACS analysis
Gene copy number
Expression rate (Northern blot analysis)

Purity / Safety

Sterility
Mycoplasma
test for tumorigenicity
In vitro assays for adventitious viruses
In vivo assays for adventitious viruses
Assays for retroviruses(e.g. S+L- focus or XC plaque)
Assays for specific viruses
Transmission electron microscopy
Reverse transcriptase activity
test on host cell derived DNA in the final product
test on host cell derived protein (HCP) in the final product.

  • Normally two cell banks are produced for production of biologicals: the Master Cell Bank (MCB) and the Working Cell Bank (WCB). Each cell bank type requires an individual testing regime.
  • Additionally to the testing of the cell banks, the Post-Production Cells (PPC) or End-of-Production Cells (EPC) as well as bulk harvests may require specific tests to be performed.