Human B-cell Ontogeny in Humanized NOD/SCID γc\(^{null}\) Mice Generates a Diverse Yet Auto/Poly- and HIV-1 Reactive Antibody Repertoire

Abstract

Characterization of the human antibody (Ab) repertoire in mouse models of the human immune system is essential to establish their relevance in translational studies. Single human B-cells were sorted from bone marrow and periphery of humanized NOD/SCID γc\(^{null}\) mice at 8–10 months post-engraftment with human cord blood-derived CD34\(^+\) stem cells. Human immunoglobulin variable heavy (V\(_H\)) and kappa (V\(_κ\)) genes were amplified, cognate V\(_H\)-V\(_κ\) gene-pairs assembled as single-chain variable fragment-Fc antibodies (scFvFcs) and functional studies performed. Although overall distribution of V\(_H\) genes approximated the normal human Ab repertoire, analysis of the V\(_H\)-third complementarity determining regions (H-CDR3) in the mature B-cell subset demonstrated an increase in length and positive charges suggesting autoimmune characteristics. Additionally, >70% of Vκ sequences utilized V\(_κ\)4-1, a germline gene associated with autoimmunity. The mature B-cell subset-derived scFvFcs displayed the highest frequency of autoreactivity and polyspecificity, suggesting defects in checkpoint control mechanisms. Furthermore, these scFvFcs demonstrated binding to recombinant HIV envelope corroborating previous observations of poly/autoreactivity in anti-HIVgp140 antibodies. These data lend support to the hypothesis that anti-HIV BnAbs may be derived from auto/polyspecific Abs that escaped immune elimination and that the hNSG mouse could provide a new experimental platform for studying the origin of anti-HIV neutralizing Ab responses.