Parallel evolution of the elite neutralizer phenotype in divergent HIV-1 clades

The development of an effective vaccine against HIV-1 requires understanding how broadly neutralizing antibodies (bNAbs) evolve in natural viral infections. Here, we recovered 152 envelope sequences from two elite neutralizers (ENs) and five viral controllers and determined the neutralization sensitivity (IC₅₀) of each envelope glycoprotein (Env) to broadly neutralizing monoclonal antibodies (bN-mAbs). For the combined EN/controller data set, we observed that the median IC₅₀ value for a CD4-binding site (CD4bs) bN-mAb (VRC01) was significantly lower for viruses lacking an N465 glycan. For a clade AE EN viral population, Env mutations implicating a VRC01-like antibody occurred in concert with positive selection at N465. Using rabbit immunizations, N465 was found to shield immunogenic VRC01 contacts in the V5/β24 domain. In the clade B EN viral population, positive selection was observed at N332, a CD4bs epitope, and a 2G12-like glycan-dependent epitope (N295-N339-N392). In rabbit immunizations, a recombinant antigen derived from the basal virus showed a greater median positivity response to cladal consensus peptides overlapping N332 than from a later virus. However, the latter virus, which displayed an N49 glycan, elicited a greater median response to consensus peptides overlapping a VRC01 contact in the C1 domain. Rarely observed C1 glycans (N49, N97) evolved in both EN viral populations. Signals of positive selection, while largely absent for glycan polymorphism present in the controllers, were detected at glycans shielding bNAb epitope contacts in both EN viral populations. Positive selection of shielding glycans suggested constraints on the escape pathways of glycan-dependent antibodies imposed by bNAb responses.IMPORTANCEIn a small subset of HIV-1-infected individuals, natural viral evolution leads to the appearance of antibodies capable of neutralizing a broad assemblage of viruses from divergent clades (bNAbs). In this study, we determined that two viral populations with divergent infections exhibited commonalities in glycan evolution that accompanied the acquisition of exceptional serum neutralization breadth. Positive selection of glycans shielding immunogenic bNAb epitope contacts suggested conflicts for escape from glycan-dependent antibodies, and rare C1 glycan introductions highlighted the immunogenicity of a region overlapping a VRC01 epitope contact. Conflicts for glycan loss could lead to their persistence in a viral population. However, a heightening of a regional cross-reactive immune response concomitant with extraordinary glycosylation pointed to evolution of specific sequence for expanding antibody neutralization breadth. These results suggest that antigens displaying immunogenic bNAb epitopes in combination with rare glycosylation might help realize the production of an effective vaccine.