Abstract
We have developed an efficient, reproducible, and scaleable cell culture process for a recombinant adenoviral vector expressing therapeutic transgenes for clinical trials. HEK 293 cells — which support the propagation of E1 deficient adenovirus — were first adapted to serum free media and suspension growth. Subsequent studies focused on the infection, virus production and harvest from suspension culture bioreactors. Future studies are planned to address the kinetics of adenovirus production in HEK 293 as well as in other cell lines.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Preview
Unable to display preview. Download preview PDF.
Abbreviations
- Ad5:
-
adenovirus serotype 5
- cGMP:
-
current good manufacturing practice
- CPE:
-
cytopathic effect
- DMEM:
-
Dulbecco’s modified Eagle’s medium
- EPD:
-
end point dilution assay
- FBS:
-
fetal bovine serum
- HEK 293:
-
human embryonic kidney cells transformed with Ad5 E1 DNA
- HPLC:
-
high pressure liquid chromatography
- ip:
-
number of infectious virus particles
- MOI:
-
multiplicity of infection
- p:
-
total number pf virus particles (infectious and non-infectious)
- PBS:
-
phosphate buffered saline
- SFM:
-
serum free medium
References
Crystal, RG (1995) Transfer of genes to humans: early lessons and obstacles to success. Science 270: 404–410.
Gamier A, Cote J, Nadeau I, Kamen A and Massie B (1994) Scaleup of the adenovirus expression system for the production of recombinant protein in human 293S cells. Cytotechnology 15: 145–155.
Graham FL and Prevec L (1991) Manipulation of adenovirus vectors. In: Murray EJ (ed.) Methods in Molecular Biology. Vol. 7, The Humana Press, Clifton, NJ, pp. 109–128.
Green M and Wold WSM (1979) Human adenoviruses: Growth, purification, and transfection assay. Meth Enzym 58: 425–435.
Hehir K, Keegan J, Martin J, Pratt D, Auger C, Davis M, Everton M, Narayana R and Karey K (1998) Production issues for adenoviral gene therapy vectors. Presented at Cell Culture Engineering VI, February 7-12, 1998, San Diego, CA.
Marcel T and Grausz JD (1997) The TMC worldwide gene therapy enrollment report, end 1996. Human Gene Therapy 8: 775–800.
Nadeau I, Garnier A, Cote J, Massie B, Chavarie C and Kamen A (1996) Improvement of recombinant protein production with the human adenovirus/293S expression system using fed-batch strategies. Biotech Bioeng 51: 613–623.
Nielsen LK, Smyth GK and Greenfield PF (1992) Accuracy of the endpoint assay for virus titration. Cytotechnology 8: 231–236.
Roth G, Smith C, Schoofs GM, Montgomery TJ, Ayala JL and Horwitz JI (1997) Using an external vortex flow filtration device for perfusion cell culture. BioPharm 10: 30–35.
Shabram PW, Giroux DD, Goudreau AM, Gregory RJ, Horn MT, Huyghe BG, Liu X, Nunnally MH, Sugarman BJ and Sutjipto S (1997) Analytical anion-exchange HPLC of recombinant type-5 adenoviral particles. Human Gene Therapy 8: 453–465.
Author information
Authors and Affiliations
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 1998 Springer Science+Business Media Dordrecht
About this chapter
Cite this chapter
Schoofs, G. et al. (1998). A high-yielding serum-free, suspension cell culture process to manufacture recombinant adenoviral vectors for gene therapy. In: Betenbaugh, M.J., Chalmers, J.J., Arathoon, R., Chaplen, F.W.R., Mastrangelo, A.J. (eds) Cell Culture Engineering VI. Current Applications of Cell Culture Engineering, vol 3. Springer, Dordrecht. https://doi.org/10.1007/978-94-011-4786-6_10
Download citation
DOI: https://doi.org/10.1007/978-94-011-4786-6_10
Publisher Name: Springer, Dordrecht
Print ISBN: 978-94-010-6011-0
Online ISBN: 978-94-011-4786-6
eBook Packages: Springer Book Archive