title: A Chimeric HIV-1 gp120 Fused with Vaccinia Virus 14K (A27) Protein as an HIV Immunogen.
abstract: In the HIV vaccine field, there is a need to produce highly immunogenic forms of the Env protein with the capacity to trigger broad B and T-cell responses. Here, we report the generation and characterization of a chimeric HIV-1 gp120 protein (termed gp120-14K) by fusing gp120 from clade B with the vaccinia virus (VACV) 14K oligomeric protein (derived from A27L gene). Stable CHO cell lines expressing HIV-1 gp120-14K protein were generated and the protein purified was characterized by size exclusion chromatography, electron microscopy and binding to anti-Env antibodies. These approaches indicate that gp120-14K protein is oligomeric and reacts with a wide spectrum of HIV-1 neutralizing antibodies. Furthermore, in human monocyte-derived dendritic cells (moDCs), gp120-14K protein upregulates the levels of several proinflammatory cytokines and chemokines associated with Th1 innate immune responses (IL-1β, IFN-γ, IL-6, IL-8, IL-12, RANTES). Moreover, we showed in a murine model, that a heterologous prime/boost immunization protocol consisting of a DNA prime with a plasmid expressing gp120-14K protein followed by a boost with MVA-B [a recombinant modified vaccinia virus Ankara (MVA) expressing HIV-1 gp120, Gag, Pol and Nef antigens from clade B], generates stronger, more polyfunctional, and greater effector memory HIV-1-specific CD4+ and CD8+ T-cell immune responses, than immunization with DNA-gp120/MVA-B. The DNA/MVA protocol was superior to immunization with the combination of protein/MVA and the latter was superior to a prime/boost of MVA/MVA or protein/protein. In addition, these immunization protocols enhanced antibody responses against gp120 of the class IgG2a and IgG3, together favoring a Th1 humoral immune response. These results demonstrate that fusing HIV-1 gp120 with VACV 14K forms an oligomeric protein which is highly antigenic as it activates a Th1 innate immune response in human moDCs, and in vaccinated mice triggers polyfunctional HIV-1-specific adaptive and memory T-cell immune responses, as well as humoral responses. This novel HIV-1 gp120-14K immunogen might be considered as an HIV vaccine candidate for broad T and B-cell immune responses.
authors.affiliation: Department of Molecular and Cellular Biology, Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas (CNB-CSIC), Madrid, Spain.
authors.forename: Aneesh
authors.initials: A
authors.lastname: Vijayan
authors.affiliation: Department of Molecular and Cellular Biology, Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas (CNB-CSIC), Madrid, Spain.
authors.forename: Juan
authors.initials: J
authors.lastname: García-Arriaza
authors.affiliation: Department of Molecular and Cellular Biology, Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas (CNB-CSIC), Madrid, Spain.
authors.forename: Suresh C
authors.initials: SC
authors.lastname: Raman
authors.affiliation: Department of Structure of Macromolecules, Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas (CNB-CSIC), Madrid, Spain.
authors.forename: José Javier
authors.initials: JJ
authors.lastname: Conesa
authors.affiliation: Department of Structure of Macromolecules, Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas (CNB-CSIC), Madrid, Spain.
authors.forename: Francisco Javier
authors.initials: FJ
authors.lastname: Chichón
authors.affiliation: X-ray Crystallization Unit, Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas (CNB-CSIC), Madrid, Spain.
authors.forename: César
authors.initials: C
authors.lastname: Santiago
authors.affiliation: Biocomputing Unit, Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas (CNB-CSIC), Madrid, Spain.
authors.forename: Carlos Óscar S
authors.initials: CÓ
authors.lastname: Sorzano
authors.affiliation: Department of Structure of Macromolecules, Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas (CNB-CSIC), Madrid, Spain.
authors.forename: José L
authors.initials: JL
authors.lastname: Carrascosa
authors.affiliation: Department of Molecular and Cellular Biology, Centro Nacional de Biotecnología, Consejo Superior de Investigaciones Científicas (CNB-CSIC), Madrid, Spain.
authors.forename: Mariano
authors.initials: M
authors.lastname: Esteban
mesh_terms: D016915:AIDS Vaccines / Q000235:genetics / Q000276:immunology; D056704:Adaptive Immunity; D000818:Animals; D057134:Antibodies, Neutralizing / Q000276:immunology; D015704:CD4 Antigens / Q000378:metabolism; D002352:Carrier Proteins / Q000235:genetics / Q000276:immunology*; D002460:Cell Line; D003713:Dendritic Cells / Q000276:immunology / Q000378:metabolism; D015483:HIV Antibodies / Q000276:immunology; D015488:HIV Antigens / Q000235:genetics / Q000276:immunology*; D015699:HIV Envelope Protein gp120 / Q000235:genetics / Q000276:immunology*; D015497:HIV-1 / Q000235:genetics / Q000276:immunology*; D006801:Humans; D007113:Immunity, Innate; D007074:Immunoglobulin G / Q000276:immunology; D007156:Immunologic Memory; D008565:Membrane Proteins; D011134:Polysaccharides / Q000378:metabolism; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology* / Q000302:isolation & purification; D016176:T-Lymphocyte Subsets / Q000276:immunology / Q000378:metabolism; D018417:Th1 Cells / Q000276:immunology / Q000378:metabolism; D014760:Viral Fusion Proteins / Q000235:genetics / Q000276:immunology*
chemical_list: C526377:A27 protein, vaccinia virus; D016915:AIDS Vaccines; D057134:Antibodies, Neutralizing; D015704:CD4 Antigens; D002352:Carrier Proteins; D015483:HIV Antibodies; D015488:HIV Antigens; D015699:HIV Envelope Protein gp120; D007074:Immunoglobulin G; D008565:Membrane Proteins; D011134:Polysaccharides; D011993:Recombinant Fusion Proteins; D014760:Viral Fusion Proteins