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Development of a Manufacturing Process for the Production of a Canine Adenovirus Type 2 (CAV-2) Vector Using MDCK Cells

  • Paulo Fernandes
  • Virgínia Santiago
  • Núria Viana
  • Eric J. Kremer
  • Ana S. Coroadinha
  • Paula M. Alves
Conference paper
Part of the ESACT Proceedings book series (ESACT, volume 5)

Abstract

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.

Keywords

MDCK Cell Adenovirus Vector Maximum Cell Density Maximum Cell Concentration Helper Dependent 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Notes

Acknowledgments

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).

References

  1. Keriel, A. et al (2006) Canine Adenovirus Vectors for Lung-Directed Gene Transfer: Efficacy, Immune Response, and Duration of Transgene Expression Using Helper- Dependent Vectors. J Virol 80 (3), 1487–1496PubMedCrossRefGoogle Scholar
  2. Kremer, EJ. et al (2000) Canine Adenovirus Vectors: An Alternative for Adenovirus- Mediated Gene Transfer. J Virol 74 (1), 505–512PubMedCrossRefGoogle Scholar
  3. Soudais, C. et al (2004) Long-Term In Vivo Transduction of Neurons Throughout the Rat Central Nervous System Using Novel Helper-Dependent CAV-2 Vectors. FASEB J 18 (2), 391–393PubMedGoogle Scholar

Copyright information

© Springer Science+Business Media B.V. 2012

Authors and Affiliations

  • Paulo Fernandes
    • 1
    • 2
  • Virgínia Santiago
    • 1
    • 2
  • Núria Viana
    • 1
    • 2
  • Eric J. Kremer
    • 3
  • Ana S. Coroadinha
    • 1
    • 4
  • Paula M. Alves
    • 5
    • 6
  1. 1.Instituto de Biologia Experimental e Tecnológica (IBET)OeirasPortugal
  2. 2.Instituto de Tecnologia Química e Biológica (ITQB)Universidade Nova de LisboaOeirasPortugal
  3. 3.Institut de Génétique Moléculaire de Montpellier, CNRSMontpellierFrance
  4. 4.Instituto de Tecnologia Química e Biológica (ITQB)Universidade Nova de Lisboa (UNL), Av. da República, Estação Agronómica NacionalOeirasPortugal
  5. 5.Instituto de Tecnologia Química e Biológica (ITQB)Universidade Nova de Lisboa (UNL)OeirasPortugal
  6. 6.Animal Cell Technology UnitInstituto de Biologia Experimental e Tecnológica (IBET)OeirasPortugal

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