title: APC targeting enhances immunogenicity of a novel multistage Fc-fusion tuberculosis vaccine in mice.
abstract: Numerous studies have demonstrated that targeting immunogens to FcγR on antigen-presenting cells (APCs) can selectively uptake and increase cellular immunity in vitro and in vivo. Therefore, the present study was conducted to evaluate immunogenicity of a novel multistage tuberculosis vaccine, a combination of an early and a dormant immunogenic protein, ESAT6 and HspX, fused to Fcγ2a fragment of mouse IgG2a to target all forms of tuberculosis. Codon-optimized genes consisting of ESAT6, a linker, and HspX fused either to mouse Fcγ2a (ESAT6:HspX:mFcγ2a) or 6× His-tag (ESAT6:HspX:His) were synthesized. The resulting proteins were then produced in Pichia pastoris. The fusion proteins were separately emulsified in dimethyldioctadecylammonium bromide(DDA)-trehalose-6,6-dibehenate(TDB) adjuvant, and their immunogenicity with and without bacille Calmette-Guérin (BCG) was assessed in C57BL/6 mice. Th1, Th2, Th17, and T-reg cytokine patterns were evaluated using the ELISA method. Both multistage vaccines induced very strong IL-12 and IFN-γ secretion from splenic cells; the Fc-tagged subunit vaccine induced a more effective Th1 immune response (IFN-γ, 910 pg/mL, and IL-12, 854 pg/mL) with a very low increase in IL-17 (∼0.1 pg/mL) and IL-4 (37 pg/mL) and a mild increase in TGF-β (543 pg/mL) compared to the BCG or ESAT6:HspX:His primed and boosted groups. The production of IFN-γ to ESAT6:HspX:Fcγ2a was very consistent and showed an increasing trend for IL-12 compared to the BCG or ESAT6:HspX:His primed and boosted groups. Fcγ2a used as a delivery vehicle supported the idea of selective uptake, inducing cross-presentation and forming a proper anti-tuberculosis response in context of Th1/Th2 and Th17/T-reg balances, which is important for protection and prevention of damage.
authors.affiliation: Microbiology & Virology Research Center, Bu-Ali Research Institute, Mashhad University of Medical Sciences, Mashhad, Iran.
authors.forename: Saman
authors.initials: S
authors.lastname: Soleimanpour
authors.affiliation: Biotechnology Research Center, Semnan University of Medical Sciences, Semnan, Iran.
authors.forename: Hadi
authors.initials: H
authors.lastname: Farsiani
authors.affiliation: Microbiology & Virology Research Center, Bu-Ali Research Institute, Mashhad University of Medical Sciences, Mashhad, Iran.
authors.forename: Arman
authors.initials: A
authors.lastname: Mosavat
authors.affiliation: Microbiology & Virology Research Center, Bu-Ali Research Institute, Mashhad University of Medical Sciences, Mashhad, Iran.
authors.forename: Kiarash
authors.initials: K
authors.lastname: Ghazvini
authors.affiliation: Biotechnology Research Center, Semnan University of Medical Sciences, Semnan, Iran.
authors.forename: Mohammad Reza Akbari
authors.initials: MR
authors.lastname: Eydgahi
authors.affiliation: Immunobiochemistry Lab, Immunology Research Center, Bu-Ali Research Institute, Mashhad University of Medical Sciences, Mashhad, Iran.
authors.forename: Mojtaba
authors.initials: M
authors.lastname: Sankian
authors.affiliation: Organic Chemistry, Department of Laboratory Sciences, Mashhad University of Medical Sciences, Mashhad, Iran.
authors.forename: Hamid
authors.initials: H
authors.lastname: Sadeghian
authors.affiliation: Microbiology & Virology Research Center, Bu-Ali Research Institute, Mashhad University of Medical Sciences, Mashhad, Iran.
authors.forename: Zahra
authors.initials: Z
authors.lastname: Meshkat
authors.affiliation: Microbiology & Virology Research Center, Bu-Ali Research Institute, Mashhad University of Medical Sciences, Mashhad, Iran. RezaeeR@mums.ac.ir.|Inflammation and Inflammatory Diseases Research Center, Medical School, Mashhad University of Medical Sciences, Mashhad, Iran. RezaeeR@mums.ac.ir.
authors.forename: Seyed Abdolrahim
authors.initials: SA
authors.lastname: Rezaee
mesh_terms: D000276:Adjuvants, Immunologic / Q000008:administration & dosage; D000818:Animals; D000938:Antigen-Presenting Cells / Q000276:immunology* / Q000378:metabolism*; D000942:Antigens, Bacterial / Q000235:genetics / Q000276:immunology*; D001426:Bacterial Proteins / Q000235:genetics / Q000276:immunology; D016207:Cytokines / Q000378:metabolism; D004797:Enzyme-Linked Immunosorbent Assay; D015870:Gene Expression; D007141:Immunoglobulin Fc Fragments / Q000235:genetics / Q000378:metabolism*; D051379:Mice; D008810:Mice, Inbred C57BL; D010843:Pichia / Q000235:genetics / Q000378:metabolism; D017452:Receptors, IgG / Q000378:metabolism*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism; D016176:T-Lymphocyte Subsets / Q000276:immunology; D032581:Tuberculosis Vaccines / Q000008:administration & dosage / Q000235:genetics / Q000276:immunology*; D014614:Vaccines, Synthetic / Q000008:administration & dosage / Q000235:genetics / Q000276:immunology
chemical_list: D000276:Adjuvants, Immunologic; D000942:Antigens, Bacterial; D001426:Bacterial Proteins; D016207:Cytokines; C093276:ESAT-6 protein, Mycobacterium tuberculosis; C118127:HspX protein, Mycobacterium tuberculosis; D007141:Immunoglobulin Fc Fragments; D017452:Receptors, IgG; D011993:Recombinant Fusion Proteins; D032581:Tuberculosis Vaccines; D014614:Vaccines, Synthetic
keywords: APC-targeting; ESAT-6; Fc-fusion vaccine; HspX; Tuberculosis