title: Rescue of HIV-1 broad neutralizing antibody-expressing B cells in 2F5 VH x VL knockin mice reveals multiple tolerance controls.
abstract: The HIV-1 broadly neutralizing Ab (bnAb) 2F5 has been shown to be poly-/self-reactive in vitro, and we previously demonstrated that targeted expression of its VDJ rearrangement alone was sufficient to trigger a profound B cell developmental blockade in 2F5 V(H) knockin (KI) mice, consistent with central deletion of 2F5 H chain-expressing B cells. In this study, we generate a strain expressing the entire 2F5 bnAb specificity, 2F5 V(H) × V(L) KI mice, and find an even higher degree of tolerance control than observed in the 2F5 V(H) KI strain. Although B cell development was severely impaired in 2F5 V(H) × V(L) KI animals, we demonstrate rescue of their B cells when cultured in IL-7/BAFF. Intriguingly, even under these conditions, most rescued B cell hybridomas produced mAbs that lacked HIV-1 Envelope (Env) reactivity due to editing of the 2F5 L chain, and the majority of rescued B cells retained an anergic phenotype. Thus, when clonal deletion is circumvented, κ editing and anergy are additional safeguards preventing 2F5 V(H)/V(L) expression by immature/transitional B cells. Importantly, 7% of rescued B cells retained 2F5 V(H)/V(L) expression and secreted Env-specific mAbs with HIV-1-neutralizing activity. This partial rescue was further corroborated in vivo, as reflected by the anergic phenotype of most rescued B cells in 2F5 V(H) × V(L) KI × Eμ-Bcl-2 transgenic mice and significant (yet modest) enrichment of Env-specific B cells and serum Igs. The rescued 2F5 mAb-producing B cell clones in this study are the first examples, to our knowledge, of in vivo-derived bone marrow precursors specifying HIV-1 bnAbs and provide a starting point for design of strategies aimed at rescuing such B cells.
authors: Department of Medicine, Duke Human Vaccine Institute, Duke University Medical Center, Durham, NC 27710, USA. laurent.verkoczy@duke.edu
authors: Laurent
authors: L
authors: Verkoczy
authors: Yao
authors: Y
authors: Chen
authors: Hilary
authors: H
authors: Bouton-Verville
authors: Jinsong
authors: J
authors: Zhang
authors: Marilyn
authors: M
authors: Diaz
authors: Jennifer
authors: J
authors: Hutchinson
authors: Ying-Bin
authors: YB
authors: Ouyang
authors: S Munir
authors: SM
authors: Alam
authors: T Matt
authors: TM
authors: Holl
authors: Kwan-Ki
authors: KK
authors: Hwang
authors: Garnett
authors: G
authors: Kelsoe
authors: Barton F
authors: BF
authors: Haynes
mesh_terms: D000818:Animals; D057134:Antibodies, Neutralizing / Q000276:immunology*; D001402:B-Lymphocytes / Q000276:immunology*; D002469:Cell Separation; D004797:Enzyme-Linked Immunosorbent Assay; D005434:Flow Cytometry; D055879:Gene Knock-In Techniques; D005803:Genes, Immunoglobulin / Q000235:genetics / Q000276:immunology; D015658:HIV Infections / Q000276:immunology*; D015497:HIV-1 / Q000276:immunology; D007108:Immune Tolerance / Q000276:immunology*; D007135:Immunoglobulin Variable Region / Q000235:genetics / Q000276:immunology; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic
chemical_list: D057134:Antibodies, Neutralizing; D007135:Immunoglobulin Variable Region