Development of a Manufacturing Process for the Production of a Canine Adenovirus Type 2 (CAV-2) Vector Using MDCK Cells
Adenovirus vectors are one of the most efficient vehicles for delivering nucleic acids into mammalian cells. However, human adenoviruses are ubiquitous in all population, posing memory immunity responses obstacles for their use during clinical gene transfer. To circumvent this drawback, nonhuman adenovirus vectors like CAV-2 are being developed. It was demonstrated that HD CAV-2 vectors have numerous advantages for clinical gene transfer leading to long-term expression in vivo gene transfer in the rodent CNS. Since the final goal of this work is the establishment of a complete manufacturing process for first generation (ΔE1) and helper dependent (HD) CAV-2 vectors, allowing the reduction of production costs and higher safety, first it is necessary the development of a robust producer cell line that enables high production in serum-free media. In order to evaluate the cell growth and ΔE1 CAV-2 vectors production, MDCK E1, a cell line based on Madin-Darby Canine Kidney (MDCK), a cell line commonly used for the production of biopharmaceuticals and vaccines, was adapted to two commercial serum-free media: Ex-cell and Optipro. When comparing with serum supplemented medium, Optipro medium allows almost 4 times higher cell densities while maintaining the vector production yields. These results show Optipro as the best serum-free medium for the growth and production of CAV-2 vectors.
KeywordsMDCK Cell Adenovirus Vector Maximum Cell Density Maximum Cell Concentration Helper Dependent
The authors acknowledge the financial support received from Fundação para a Ciência e Tecnologia – Portugal (PTDC/BIO/69452/2006: Production of Canine Adenoviral Vectors for Gene Therapy: development of MDCK CAV-2 E1 complementing cell lines) and European Commission (BRAINCAV HEALTH – HS_2008_222992).