title: In vitro reconstitution of B cell receptor-antigen interactions to evaluate potential vaccine candidates.
abstract: Predicting immune responses before vaccination is challenging because of the complexity of the governing parameters. Nevertheless, recent work has shown that B cell receptor (BCR)-antigen engagement in vitro can prove a powerful means of informing the design of antibody-based vaccines. We have developed this principle into a two-phased immunogen evaluation pipeline to rank-order vaccine candidates. In phase 1, recombinant antigens are screened for reactivity to the germline precursors that produce the antibody responses of interest. To both mimic the architecture of initial antigen engagement and facilitate rapid immunogen screening, these antibodies are expressed as membrane-anchored IgM (mIgM) in 293F indicator cells. In phase 2, the binding hits are multimerized by nanoparticle or proteoliposome display, and they are evaluated for BCR triggering in an engineered B cell line displaying the IgM sequences of interest. Key developments that complement existing methodology in this area include the following: (i) introduction of a high-throughput screening step before evaluation of more time-intensive BCR-triggering analyses; (ii) generalizable multivalent antigen-display platforms needed for BCR activation; and (iii) engineered use of a human B cell line that does not display endogenous antibody, but only ectopically expressed BCR sequences of interest. Through this pipeline, the capacity to initiate favorable antibody responses is evaluated. The entire protocol can be completed within 2.5 months.
authors.affiliation: The Ragon Institute of Massachusetts General Hospital, The Massachusetts Institute for Technology and Harvard University, Cambridge, Massachusetts, USA.
authors.forename: Grant C
authors.initials: GC
authors.lastname: Weaver
authors.affiliation: The Ragon Institute of Massachusetts General Hospital, The Massachusetts Institute for Technology and Harvard University, Cambridge, Massachusetts, USA.
authors.forename: Rina F
authors.initials: RF
authors.lastname: Villar
authors.affiliation: Vaccine Research Center, National Institute for Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: Masaru
authors.identifier: http://orcid.org/0000-0001-5767-1532
authors.initials: M
authors.lastname: Kanekiyo
authors.affiliation: Sanofi, Cambridge, Massachusetts, USA.
authors.forename: Gary J
authors.initials: GJ
authors.lastname: Nabel
authors.affiliation: Vaccine Research Center, National Institute for Allergy and Infectious Diseases, National Institutes of Health, Bethesda, Maryland, USA.
authors.forename: John R
authors.initials: JR
authors.lastname: Mascola
authors.affiliation: The Ragon Institute of Massachusetts General Hospital, The Massachusetts Institute for Technology and Harvard University, Cambridge, Massachusetts, USA.
authors.forename: Daniel
authors.initials: D
authors.lastname: Lingwood
mesh_terms: D000941:Antigens / Q000378:metabolism*; D001402:B-Lymphocytes / Q000276:immunology*; D002460:Cell Line; D006801:Humans; D011485:Protein Binding; D011947:Receptors, Antigen, B-Cell / Q000378:metabolism*; D014612:Vaccines / Q000276:immunology*
chemical_list: D000941:Antigens; D011947:Receptors, Antigen, B-Cell; D014612:Vaccines