Stable expression of lentiviral antigens by quality-controlled recombinant mycobacterium bovis BCG vectors

Bryan E. Hart, Rose Asrican, So Yon Lim, Jaimie D. Sixsmith, Regy Lukose, Sommer J.R. Souther, Swati D.G. Rayasam, Joseph W. Saelens, Ching Ju Chen, Sarah A. Seay, Linda Berney-Meyer, Leslie Magtanong, Kim Vermeul, Priyadharshini Pajanirassa, Amanda E. Jimenez, Tony W. Ng, David M. Tobin, Steven A. Porcelli, Michelle H. Larsen, Joern E. SchmitzBarton F. Haynes, William R. Jacobs, Sunhee Lee, Richard Frothingham

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11 Scopus citations

Abstract

The well-established safety profile of the tuberculosis vaccine strain, Mycobacterium bovis bacille Calmette-Guérin (BCG), makes it an attractive vehicle for heterologous expression of antigens from clinically relevant pathogens. However, successful generation of recombinant BCG strains possessing consistent insert expression has encountered challenges in stability. Here, we describe a method for the development of large recombinant BCG accession lots which stably express the lentiviral antigens, human immunodeficiency virus (HIV) gp120 and simian immunodeficiency virus (SIV) Gag, using selectable leucine auxotrophic complementation. Successful establishment of vaccine stability stems from stringent quality control criteria which not only screen for highly stable complemented BCG ΔleuCD transformants but also thoroughly characterize postproduction quality. These parameters include consistent production of correctly sized antigen, retention of sequence-pure plasmid DNA, freezethaw recovery, enumeration of CFU, and assessment of cellular aggregates. Importantly, these quality assurance procedures were indicative of overall vaccine stability, were predictive for successful antigen expression in subsequent passaging both in vitro and in vivo, and correlated with induction of immune responses in murine models. This study has yielded a quality-controlled BCG ΔleuCD vaccine expressing HIV gp120 that retained stable full-length expression after 1024-fold amplification in vitro and following 60 days of growth in mice. A second vaccine lot expressed full-length SIV Gag for>1068-fold amplification in vitro and induced potent antigen-specific T cell populations in vaccinated mice. Production of large, well-defined recombinant BCG ΔleuCD lots can allow confidence that vaccine materials for immunogenicity and protection studies are not negatively affected by instability or differences between freshly grown production batches.

Original languageEnglish (US)
Pages (from-to)726-741
Number of pages16
JournalClinical and Vaccine Immunology
Volume22
Issue number7
DOIs
StatePublished - Jul 1 2015

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ASJC Scopus subject areas

  • Immunology and Allergy
  • Immunology
  • Clinical Biochemistry
  • Microbiology (medical)

Cite this

Hart, B. E., Asrican, R., Lim, S. Y., Sixsmith, J. D., Lukose, R., Souther, S. J. R., Rayasam, S. D. G., Saelens, J. W., Chen, C. J., Seay, S. A., Berney-Meyer, L., Magtanong, L., Vermeul, K., Pajanirassa, P., Jimenez, A. E., Ng, T. W., Tobin, D. M., Porcelli, S. A., Larsen, M. H., ... Frothingham, R. (2015). Stable expression of lentiviral antigens by quality-controlled recombinant mycobacterium bovis BCG vectors. Clinical and Vaccine Immunology, 22(7), 726-741. https://doi.org/10.1128/CVI.00075-15