Background
Several classes of HIV-1 broadly neutralizing antibodies (bnAbs) exhibit distinct immunoglobulin heavy chain variable (IGHV) gene usage, and many HIV vaccine strategies aim to elicit public bnAb clonotypes. Here, we performed a systematic analysis of HIV-1 envelope (Env) trimer reactive B cell receptor (BCR) repertoires in bnAb inducers to define public features of IGHV and IGLV/KV gene segment usage, enabling the discovery of novel bnAbs.
Methods
bnAb inducers (N=36) included in the XbnAb cohort (Pasin et al. biorXiv 2024) were analyzed by single cell BCR cloning (10X genomics with Libra-Seq) to identify Env-trimer reactive BCRs. Neutralization activity was measured against a 41-virus multi-clade Env pseudotype panel. bnAb epitopes were defined by Env mutant screens (mutational scanning, binding, neutralization) and Cryo-EM structure analysis.
Results
BCR repertoire analysis of 36 bnAb inducers identified a total of 1,218 individual IgG and IgA trimer-reactive BCRs assigned to 869 BCR lineages. IG gene segment usage was generally comparable between trimer-reactive and non-trimer BCRs. IGHV4-34/IGKV2-24 was rare in non-trimer (0.06% total Ig, 0.07%, IgA) but remarkably enriched in trimer-reactive BCR families (4.26% total Ig, 9.84% IgA). We identified IGHV4-34 bnAbs SHCS-0209 (IGKV2-24), SHCS-3L6 (IGKV2-24) and SHCS-3D14 (IGKV3-15) with 63%, 78%, and 88% breadth, respectively. IGHV4-34 usage is known to be associated with autoreactivity, and the autoreactive IGHV4-34 codon 26 was retained in SHCS-3L6 and SHCS-3D14 but mutated in SHCS-0209. Trimer-reactive IGHV4-34 BCRs were found in 30/36 bnAb inducers, and those with similarity to SHCS-3L6 (short CDR-H3/ IGKV2-24 usage) in 12/36 donors, suggesting the existence of IGHV4-34 public clonotypes. Cryo-EM structure analysis showed that SHCS-0209 has a clear silent face specificity. However, SHCS-3L6 targets V3-glycans (N301, N332) as well as glycans in the silent face (N262, N295). SHCS-3D14 targets the V3 (N301), V2-loop and CD4bs of the adjacent protomer. Neutralization correlation analyses with reference bnAbs and bnAb inducer plasmas (N=304) showed that SHCS-3L6 and SHCS-3D14 cluster distinctly from V3-glycan and silent face bnAbs, forming a novel subclass we termed V3-glycan type II.
Conclusions
The high frequency of trimer-reactive IGHV4-34 BCRs, and the identification of novel IGHV4-34 bnAbs suggests that, provided the autoreactivity associated with IGHV4-34 could be mitigated, targeting IGHV4-34 for bnAb vaccines could be valuable.