title: Decoration of outer membrane vesicles with multiple antigens by using an autotransporter approach.
abstract: Outer membrane vesicles (OMVs) are spherical nanoparticles that naturally shed from Gram-negative bacteria. They are rich in immunostimulatory proteins and lipopolysaccharide but do not replicate, which increases their safety profile and renders them attractive vaccine vectors. By packaging foreign polypeptides in OMVs, specific immune responses can be raised toward heterologous antigens in the context of an intrinsic adjuvant. Antigens exposed at the vesicle surface have been suggested to elicit protection superior to that from antigens concealed inside OMVs, but hitherto robust methods for targeting heterologous proteins to the OMV surface have been lacking. We have exploited our previously developed hemoglobin protease (Hbp) autotransporter platform for display of heterologous polypeptides at the OMV surface. One, two, or three of the Mycobacterium tuberculosis antigens ESAT6, Ag85B, and Rv2660c were targeted to the surface of Escherichia coli OMVs upon fusion to Hbp. Furthermore, a hypervesiculating ΔtolR ΔtolA derivative of attenuated Salmonella enterica serovar Typhimurium SL3261 was generated, enabling efficient release and purification of OMVs decorated with multiple heterologous antigens, exemplified by the M. tuberculosis antigens and epitopes from Chlamydia trachomatis major outer membrane protein (MOMP). Also, we showed that delivery of Salmonella OMVs displaying Ag85B to antigen-presenting cells in vitro results in processing and presentation of an epitope that is functionally recognized by Ag85B-specific T cell hybridomas. In conclusion, the Hbp platform mediates efficient display of (multiple) heterologous antigens, individually or combined within one molecule, at the surface of OMVs. Detection of antigen-specific immune responses upon vesicle-mediated delivery demonstrated the potential of our system for vaccine development.
authors.affiliation: Section Molecular Microbiology, Department of Molecular Cell Biology, Faculty of Earth and Life Sciences, VU University, Amsterdam, The Netherlands Abera Bioscience AB, Stockholm, Sweden maria.daleke@aberabio.com s.luirink@vu.nl.
authors.forename: Maria H
authors.initials: MH
authors.lastname: Daleke-Schermerhorn
authors.affiliation: Institut Pasteur, Unité de Régulation Immunitaire et Vaccinologie, Paris, France INSERM, U1041, Paris, France.
authors.forename: Tristan
authors.initials: T
authors.lastname: Felix
authors.affiliation: Section Molecular Microbiology, Department of Molecular Cell Biology, Faculty of Earth and Life Sciences, VU University, Amsterdam, The Netherlands.
authors.forename: Zora
authors.initials: Z
authors.lastname: Soprova
authors.affiliation: Section Molecular Microbiology, Department of Molecular Cell Biology, Faculty of Earth and Life Sciences, VU University, Amsterdam, The Netherlands Abera Bioscience AB, Stockholm, Sweden.
authors.forename: Corinne M
authors.initials: CM
authors.lastname: Ten Hagen-Jongman
authors.affiliation: Center for Biomembrane Research, Department of Biochemistry and Biophysics, Stockholm University, Stockholm, Sweden Xbrane Bioscience AB, Stockholm, Sweden.
authors.forename: David
authors.initials: D
authors.lastname: Vikström
authors.affiliation: Institut Pasteur, Unité de Régulation Immunitaire et Vaccinologie, Paris, France INSERM, U1041, Paris, France.
authors.forename: Laleh
authors.initials: L
authors.lastname: Majlessi
authors.affiliation: Section Molecular Microbiology, Department of Molecular Cell Biology, Faculty of Earth and Life Sciences, VU University, Amsterdam, The Netherlands.
authors.forename: Joep
authors.initials: J
authors.lastname: Beskers
authors.affiliation: Chlamydia Vaccine Research, Department of Infectious Disease Immunology, Statens Serum Institut, Copenhagen, Denmark.
authors.forename: Frank
authors.initials: F
authors.lastname: Follmann
authors.affiliation: The Netherlands Cancer Institute, Antonie van Leeuwenhoek Hospital, Amsterdam, The Netherlands.
authors.forename: Karin
authors.initials: K
authors.lastname: de Punder
authors.affiliation: The Netherlands Cancer Institute, Antonie van Leeuwenhoek Hospital, Amsterdam, The Netherlands.
authors.forename: Nicole N
authors.initials: NN
authors.lastname: van der Wel
authors.affiliation: Center for Biomembrane Research, Department of Biochemistry and Biophysics, Stockholm University, Stockholm, Sweden.
authors.forename: Thomas
authors.initials: T
authors.lastname: Baumgarten
authors.affiliation: Department of Medical Oncology, OncoProteomics Laboratory, VU University Medical Center, Amsterdam, The Netherlands.
authors.forename: Thang V
authors.initials: TV
authors.lastname: Pham
authors.affiliation: Department of Medical Oncology, OncoProteomics Laboratory, VU University Medical Center, Amsterdam, The Netherlands.
authors.forename: Sander R
authors.initials: SR
authors.lastname: Piersma
authors.affiliation: Department of Medical Oncology, OncoProteomics Laboratory, VU University Medical Center, Amsterdam, The Netherlands.
authors.forename: Connie R
authors.initials: CR
authors.lastname: Jiménez
authors.affiliation: Section Molecular Microbiology, Department of Molecular Cell Biology, Faculty of Earth and Life Sciences, VU University, Amsterdam, The Netherlands.
authors.forename: Peter
authors.initials: P
authors.lastname: van Ulsen
authors.affiliation: Center for Biomembrane Research, Department of Biochemistry and Biophysics, Stockholm University, Stockholm, Sweden.
authors.forename: Jan-Willem
authors.initials: JW
authors.lastname: de Gier
authors.affiliation: Institut Pasteur, Unité de Régulation Immunitaire et Vaccinologie, Paris, France INSERM, U1041, Paris, France.
authors.forename: Claude
authors.initials: C
authors.lastname: Leclerc
authors.affiliation: Section Molecular Microbiology, Department of Molecular Cell Biology, Faculty of Earth and Life Sciences, VU University, Amsterdam, The Netherlands Abera Bioscience AB, Stockholm, Sweden.
authors.forename: Wouter S P
authors.initials: WS
authors.lastname: Jong
authors.affiliation: Section Molecular Microbiology, Department of Molecular Cell Biology, Faculty of Earth and Life Sciences, VU University, Amsterdam, The Netherlands Abera Bioscience AB, Stockholm, Sweden maria.daleke@aberabio.com s.luirink@vu.nl.
authors.forename: Joen
authors.initials: J
authors.lastname: Luirink
mesh_terms: D000217:Acyltransferases / Q000235:genetics / Q000378:metabolism; D000942:Antigens, Bacterial / Q000235:genetics / Q000378:metabolism*; D001426:Bacterial Proteins / Q000235:genetics / Q000378:metabolism; D002692:Chlamydia trachomatis / Q000235:genetics; D010450:Endopeptidases / Q000378:metabolism*; D004926:Escherichia coli / Q000235:genetics / Q000378:metabolism*; D026901:Membrane Transport Proteins / Q000378:metabolism*; D009169:Mycobacterium tuberculosis / Q000235:genetics; D021381:Protein Transport; D012486:Salmonella typhimurium / Q000235:genetics / Q000378:metabolism*; D022142:Secretory Vesicles / Q000378:metabolism*
chemical_list: D000942:Antigens, Bacterial; D001426:Bacterial Proteins; C093276:ESAT-6 protein, Mycobacterium tuberculosis; D026901:Membrane Transport Proteins; C570849:Rv2660c protein, Mycobacterium tuberculosis; D000217:Acyltransferases; C121433:antigen 85B, Mycobacterium tuberculosis; D010450:Endopeptidases; C114676:hemoglobin protease Hbp