title: Engineering new mycobacterial vaccine design for HIV-TB pediatric vaccine vectored by lysine auxotroph of BCG.
abstract: In this study, we have engineered a new mycobacterial vaccine design by using an antibiotic-free plasmid selection system. We assembled a novel Escherichia coli (E. coli)-mycobacterial shuttle plasmid p2auxo.HIVA, expressing the HIV-1 clade A immunogen HIVA. This shuttle vector employs an antibiotic resistance-free mechanism for plasmid selection and maintenance based on glycine complementation in E. coli and lysine complementation in mycobacteria. This plasmid was first transformed into glycine auxotroph of E. coli strain and subsequently transformed into lysine auxotroph of Mycobacterium bovis BCG strain to generate vaccine BCG.HIVA(2auxo). We demonstrated that the episomal plasmid p2auxo.HIVA was stable in vivo over a 7-week period and genetically and phenotypically characterized the BCG.HIVA(2auxo) vaccine strain. The BCG.HIVA(2auxo) vaccine in combination with modified vaccinia virus Ankara (MVA). HIVA was safe and induced HIV-1 and Mycobacterium tuberculosis-specific interferon-γ-producing T-cell responses in adult BALB/c mice. Polyfunctional HIV-1-specific CD8+ T cells, which produce interferon-γ and tumor necrosis factor-α and express the degranulation marker CD107a, were induced. Thus, we engineered a novel, safer, good laboratory practice-compatible BCG-vectored vaccine using prototype immunogen HIVA. This antibiotic-free plasmid selection system based on "double" auxotrophic complementation might be a new mycobacterial vaccine platform to develop not only recombinant BCG-based vaccines expressing second generation of HIV-1 immunogens but also other major pediatric pathogens to prime protective response soon after birth.
authors.affiliation: AIDS Research Group, Hospital Clinic/HIVACAT, School of Medicine, University of Barcelona , Barcelona, Catalonia, Spain.
authors.forename: Narcís
authors.initials: N
authors.lastname: Saubi
authors.affiliation: AIDS Research Group, Hospital Clinic/HIVACAT, School of Medicine, University of Barcelona , Barcelona, Catalonia, Spain.
authors.forename: Ester
authors.initials: E
authors.lastname: Gea-Mallorquí
authors.affiliation: Department of Chemical Engineering, Group of Bioprocess Engineering and Applied Biocatalysis, School of Engineering, Autonomous University of Barcelona , Barcelona, Catalonia, Spain.
authors.forename: Pau
authors.initials: P
authors.lastname: Ferrer
authors.affiliation: AIDS Research Group, Hospital Clinic/HIVACAT, School of Medicine, University of Barcelona , Barcelona, Catalonia, Spain.
authors.forename: Carmen
authors.initials: C
authors.lastname: Hurtado
authors.affiliation: AIDS Research Group, Hospital Clinic/HIVACAT, School of Medicine, University of Barcelona , Barcelona, Catalonia, Spain.
authors.forename: Sara
authors.initials: S
authors.lastname: Sánchez-Úbeda
authors.affiliation: AIDS Research Group, Hospital Clinic/HIVACAT, School of Medicine, University of Barcelona , Barcelona, Catalonia, Spain.
authors.forename: Yoshiki
authors.initials: Y
authors.lastname: Eto
authors.affiliation: AIDS Research Group, Hospital Clinic/HIVACAT, School of Medicine, University of Barcelona , Barcelona, Catalonia, Spain.
authors.forename: Josep M
authors.initials: JM
authors.lastname: Gatell
authors.affiliation: The Jenner Institute, University of Oxford , Oxford, UK ; MRC Human Immunology Unit, Weatherall Institute of Molecular Medicine, University of Oxford , Oxford, UK.
authors.forename: Tomáš
authors.initials: T
authors.lastname: Hanke
authors.affiliation: AIDS Research Group, Hospital Clinic/HIVACAT, School of Medicine, University of Barcelona , Barcelona, Catalonia, Spain.
authors.forename: Joan
authors.initials: J
authors.lastname: Joseph