HIV newly infects 1.8 million people each year, making development of an HIV vaccine a global health priority. Nearly all licensed vaccines protect by inducing antibodies, but highly variable pathogens such as HIV and influenza virus have eluded traditional vaccine strategies. The discoveries of broadly neutralizing antibodies (bnAbs) that bind to conserved epitopes on the surface proteins of these viruses have inspired vaccine design strategies to induce bnAbs. Antibodies are produced by B cells, and highly effective antibodies like bnAbs acquire affinity-enhancing mutations when a bnAb-precursor B cell mutates and matures from the original naïve B cell (or germline) state. Among several new vaccine strategies, germline-targeting vaccine design aims to induce bnAbs by first stimulating bnAb-precursor B cells and then shepherding B cell affinity maturation with a series of rationally designed boosting immunogens. A key rationale for this strategy is that germline-reverted forms of bnAbs—precursors with all recognizable amino acid mutations reverted to germline—typically have no detectable affinity for HIV envelope (Env). Thus, for a vaccine to initiate bnAb induction, a germline-targeting priming immunogen with appreciable affinity for bnAb precursors must be engineered.

Most HIV bnAbs (and most antibodies to any pathogen) bind to their target by using their heavy chain complementarity-determining region 3 (HCDR3) as a major binding determinant. Hence, an optimal HIV vaccine that induces multiple bnAbs, and a general solution to germline-targeting vaccine design that could be applied broadly to other pathogens, will need to work with HCDR3-dependent antibodies. However, the need to design germline-targeting immunogens to initiate HCDR3-dependent bnAb responses faces major technical challenges. Although each B cell expresses a single unique antibody, different B cells produce diverse antibodies encoded by different combinations of antibody genes, with the greatest antibody genetic diversity encoded in the HCDR3 portion of the molecule. The exceptional diversity in the human B cell repertoire makes any single HCDR3 sequence an impractical vaccine target. Rather, a pool of precursors sharing a set of bnAb-associated genetic features must be identified and targeted. Thus, owing to the enormous diversity of human antibodies, a germline-targeting immunogen should have affinity for diverse bnAb precursors in order to succeed in diverse vaccine recipients.

Herein we report a solution to the above challenges. Using the strongly HCDR3-dependent bnAb BG18 that binds a conserved site on HIV Env as a high-value target and a proof of principle, we demonstrate a method to identify pools of potential bnAb precursors in an ultradeep human antibody sequence database, guided by key genetic features that enable bnAb structural recognition of the antigen. We then use a representative set of those potential bnAb precursors as design targets to guide our engineering of HIV Env immunogens that bind to diverse potential bnAb precursors. Lastly, we provide critical preclinical validation of immunogen design by assessing these immunogens for (i) their ability to select rare potential bnAb-precursor naïve B cells from the blood of healthy human donors, (ii) their modes of binding to bnAb precursors, and (iii) their capacity to prime rare bnAb-precursor B cells with physiologically relevant affinities in a mouse model.

Overall, we demonstrate a new approach to defining diverse precursors for a target antibody and designing vaccine immunogens that take advantage of that …