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A serum-free Vero production platform for a chimeric virus vaccine candidate

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Abstract

MedImmune Vaccines has engineered a live, attenuated chimeric virus that could prevent infections caused by parainfluenza virus type 3 (PIV3) and respiratory syncytial virus (RSV), causative agents of acute respiratory diseases in infants and young children. The work here details the development of a serum-free Vero cell culture production platform for this virus vaccine candidate. Efforts to identify critical process parameters and optimize culture conditions increased infectious virus titers by approximately 2 log10 TCID50/ml over the original serum-free process. In particular, the addition of a chemically defined lipid concentrate to the pre-infection medium along with the shift to a lower post-infection cultivation temperature increased virus titers by almost 100-fold. This improved serum-free process achieved comparable virus titers to the serum-supplemented process, and demonstrated consistent results upon scale-up: Vero cultures in roller bottles, spinner flasks and bioreactors reproducibly generated maximum infectious virus titers of 8 log10 TCID50/ml.

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Abbreviations

CDLC:

Chemically defined lipid concentrate

F:

Fusion

FBS:

Fetal bovine serum

HN:

Hemagglutinin–neuraminidase

hPIV3:

Human parainfluenza virus type 3

MEDI-534:

Chimeric human parainfluenza virus type 3/respiratory syncytial virus

MOI:

Multiplicity of infection

PIV3:

Parainfluenza virus type 3

RB:

Roller bottle

RSV:

Respiratory syncytial virus

SFM:

Serum-free medium

TCID50 :

50% Tissue culture infective dose

Vero:

African green monkey kidney

VP-SFM:

Virus production serum-free medium

WME:

Williams’ Medium E

References

  • Asher DM (1999) The transmissible spongiform encephalopathy agents: concerns and responses of United States regulatory agencies in maintaining the safety of biologics. Dev Biol Stand 100:103–118

    CAS  Google Scholar 

  • Bender FC, Whitbeck JC, de Leon MP, Lou H, Eisenbery RJ, Cohen GH (2003) Specific association of glycoprotein B with lipid rafts during Herpes Simplex Virus entry. J Virol 77:9542–9552

    Article  CAS  Google Scholar 

  • Berry JM, Barnabe N, Coombe KM, Butler M (1999) Production of reovirus type-1 and type-3 from Vero cells grown on solid and macroporous microcarriers. Biotechnol Bioeng 62:12–19

    Article  CAS  Google Scholar 

  • Brown G, Rixon HWM, Steel J, McDonald TP, Pitt AR, Graham S, Sugrue RJ (2005) Evidence for an association between heat shock protein 70 and the respiratory syncytial virus polymerase complex within lipid-raft membranes during virus infection. Virology 338:69–80

    Article  CAS  Google Scholar 

  • Castle P, Robertson JS (1999) Animal sera, animal sera derivatives and substitutes used in the manufacture of pharmaceuticals: viral safety and regulatory aspects. Dev Biol Stand 99:191–196

    CAS  Google Scholar 

  • Erickson GA, Landgraf JG, Wessman SJ, Koski TA, Moss LM (1989) Detection and elimination of adventitious agents in continuous cell lines. Dev Biol Stand 70:59–66

    CAS  Google Scholar 

  • Feller JA, Smallwood S, Skiadopoulos MH, Murphy BR, Moyer SA (2000) Comparison of identical temperature-sensitive mutations in the l polymerase proteins of Sendai and Parainfluenza 3 viruses. Virology 275:190–201

    Article  Google Scholar 

  • Galbraith DN (2002) Transmissible spongiform encephalopathies and tissue cell culture. Cytotechnology 39:117–124

    Article  Google Scholar 

  • Hall CD (2001) Respiratory syncytial virus and parainfluenza virus. N Engl J Med 344:1917–1927

    Article  CAS  Google Scholar 

  • Haller AA, Miller T, Mitiku M, Coelingh K (2000) Expression of the surface glycoproteins of human parainfluenza virus type 3 by bovine parainfluenza virus type 3, a novel attenuated virus vaccine vector. J Virol 74:11626–11635

    Article  CAS  Google Scholar 

  • Haller AA, Mitiku M, MacPhail M (2003) Bovine parainfluenza virus type 3 (PIV3) expressing the respiratory syncytial virus (RSV) attachment and fusion proteins protects hamsters from challenge with human PIV3 and PSV. J Gen Virol 84:2153–2162

    Article  CAS  Google Scholar 

  • Henry O, Dormond E, Perrier M, Kamen A (2004) Insights into adenoviral vector production kinetics in acoustic filter-based perfusion cultures. Biotechnol Bioeng 86:765–774

    Article  CAS  Google Scholar 

  • Hu WS, Wang DIC (1987) Selection of microcarrier diameter for the cultivation of mammalian cells on microcarriers. Biotechnol Bioeng 30:548–557

    Article  Google Scholar 

  • Kaptein LCM, Greijer AE, Valerio D, van Beusechem VW (1997) Optimized conditions for the production of recombinant amphotropic retroviral vector preparations. Gene Ther 4:172–176

    Article  CAS  Google Scholar 

  • Kamen A, Henry O (2004) Development and optimization of an adenovirus production process. J Gene Med 6:S184–S192

    Article  CAS  Google Scholar 

  • Kotani H, Newton PB, Zhang S, Chiang YL, Otto E, Weaver L, Blaese RM, Anderson WF, McGarrity GJ (1994) Improved methods of retroviral vector transduction and production for gene therapy. Hum Gene Ther 5:19–28

    CAS  Google Scholar 

  • Lee SG, Kim S, Robbins PD, Kim BG (1996) Optimization of environmental factors for the production and handling of recombinant retrovirus. Appl Microbiol Biotechnol 45:477–483

    CAS  Google Scholar 

  • Litwin J (1992) The growth of Vero cells in suspension as cell-aggregates in serum-free media. Cytotechnology 10:169–174

    Article  CAS  Google Scholar 

  • Montagnon BJ (1989) Polio and rabies vaccines produced in continuous cell lines: a reality for Vero cell line. Dev Biol Stand 70:27–47

    CAS  Google Scholar 

  • Montagnon BJ, Vincent-Falquet JC (1998) Experience with the Vero cell line. Dev Biol Stand 93:119–123

    CAS  Google Scholar 

  • Nadeau I, Gilbert PA, Jacob D, Perrier M, Kamen A (2002) Low-protein medium affects the 293SF central metabolism during growth and infection with adenovirus. Biotechnol Bioeng 77:91–104

    Article  CAS  Google Scholar 

  • Nadeau I, Kamen A (2003) Production of adenovirus vector for gene therapy. Biotechnol Adv 20:475–489

    Article  CAS  Google Scholar 

  • Skiadopoulos MH, Surman S, Tatem JM, Paschalis M, Wu SL, Udem SA, Durbin P, Collins PL, Murphy BR (1999) Identification of mutations contributing to the temperature-sensitive, cold-adapted, and attenuation phenotypes of the live-attenuated, cold-passaged 45 (cp45) human parainfluenza virus 3 candidate vaccine. J Virol 73:1374–1381

    CAS  Google Scholar 

  • Tang RS, Schickli JH, MacPhail M, Fernandes F, Bicha L, Spaete J, Fouchier RAM, Osterhaus ADME, Spaete R, Haller AA (2003) Effects of human metapneumovirus and respiratory syncytial virus antigen insertion in two 3′ proximal genome positions of bovine/human parainfluenza virus type 3 on virus replication and immunogenicity. J Virol 77:10819–10828

    Article  CAS  Google Scholar 

  • Tang RS, MacPhail M, Schickli JH, Kaur J, Robinson CL, Lawlor HA, Guzzetta JM, Spaete RR, Haller AA (2004) Parainfluenza virus type 3 expressing the native or soluble fusion (F) protein of respiratory syncytial virus (RSV) confers protection from RSV infection in African green monkeys. J Virol 78:11198–11207

    Article  CAS  Google Scholar 

  • Vincent-Falquet JC, Peyron L, Souvras M, Moulin JC, Tektoff J, Patet J (1989) Qualification of working cell banks for the Vero cell line to produce licensed human vaccines. Dev Biol Stand 70:153–156

    CAS  Google Scholar 

  • Williams GM, Gunn JM (1974) Long-term cell culture of adult rat liver epithelial cells. Exp Cell Res 89:139–142

    Article  CAS  Google Scholar 

  • World Health Organization (1987a) Requirements for continuous cell lines used for biological substances. WHO Tech Rep Ser 745:99–115

    Google Scholar 

  • World Health Organization (1987b) Requirements for rabies vaccine (inactivated) for human use produced in continuous cell lines. WHO Tech Rep Ser 760:167–189

    Google Scholar 

  • Wu SC, Liu CC, Lian WC (2004) Optimization of microcarrier cell culture process for the inactivated enterovirus type 71 vaccine development. Vaccine 22:3858–3864

    Article  CAS  Google Scholar 

  • Yokomizo AY, Antoniazzi MM, Galdino PL, Azambunja N Jr, Jorge SAC, Pereira CA (2004) Rabies virus production in high Vero cell density cultures on macroporous microcarriers. Biotechnol Bioeng 85:506–515

    Article  CAS  Google Scholar 

  • Yuk IH, Olsen MM, Geyer S, Forestell SP (2004) Perfusion cultures of human tumor cells: a scalable production platform for oncolytic adenoviral vectors. Biotechnol Bioeng 86:637–642

    Article  CAS  Google Scholar 

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Acknowledgements

We thank Richard R. Spaete, Roderick S. Tang, Mia MacPhail, Jeanne H. Schickli, Jeanne M. Guzzetta, Jasmine Kaur, Elizabeth Stillman, and Aurelia Haller for generating the original MEDI-534 vectors by plasmid rescue.

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Correspondence to Inn H. Yuk.

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Yuk, I.H., Lin, G.B., Ju, H. et al. A serum-free Vero production platform for a chimeric virus vaccine candidate. Cytotechnology 51, 183–192 (2006). https://doi.org/10.1007/s10616-006-9030-7

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  • DOI: https://doi.org/10.1007/s10616-006-9030-7

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