Engineered receptor binding domain immunogens elicit pan-sarbecovirus neutralizing antibodies outside the receptor binding motif

Abstract

Effective countermeasures are needed against emerging coronaviruses of pandemic potential, similar to severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). Designing immunogens that elicit broadly neutralizing antibodies to conserved viral epitopes on the major surface glycoprotein, spike, such as the receptor binding domain (RBD) is one potential approach. Here, we report the generation of homotrimeric RBD immunogens from different sarbecoviruses using a stabilized, immune-silent trimerization tag. We find that that a cocktail of homotrimeric sarbecovirus RBDs can elicit a neutralizing response to all components even in context of prior SARS-CoV-2 imprinting. Importantly, the cross-neutralizing antibody responses are focused towards conserved RBD epitopes outside of the ACE-2 receptor-binding motif. This may be an effective strategy for eliciting broadly neutralizing responses leading to a pan-sarbecovirus vaccine.

Rationally designed immunogens enable immune focusing to the SARS-CoV-2 receptor binding motif

“…S10 ). Notably, these results are in contrast to our previous work showing that a cocktail of sarbecovirus that included SARS-1 and WIV1 RBDs could predominantly focus the antibody response towards the conserved CR3022 and S309 epitopic regions ( 30 ).…”

“…Importantly, these epitopic regions were shown to be a significant portion of the SARS-2 RBD-directed response in murine immunizations and thus any RBM focusing would require masking of these regions ( Fig. S5 ) ( 27, 28, 30 ). While SARS-2 hg effectively abrogated S309 and CR3022 binding, the engineered PNGs at the antibody:antigen interface on rsSARS-1 hg and rsWIV1 hg did not completely abrogate S309 and CR3022 binding.…”

“…We then tested the immunogenicity and antigenicity of our optimized constructs and assessed their RBM immune-focusing properties, in the murine model. In order to increase avidity and to minimize any off-target tag-specific responses, we generated trimeric versions of each immunogen using our previously characterized hyperglycosylated, cysteine-stabilized GCN4 tag ( hg GCN4 cys ) ( 30, 31 ). We first primed all cohorts with SARS-2 spike to reflect pre-existing SARS-2 immunity and to imprint an initial RBM response that may be recalled and selectively expanded by our immunogens.…”

Naive human B cells engage the receptor binding domain of SARS-CoV-2, variants of concern, and related sarbecoviruses

“…We first designed a SARS-CoV-2 RBD construct that positions two glycans at residues 475 and 501 to selectively block binding to ACE2 and the receptor-binding motif (RBM)-directed antibody, B38 (fig. S1) (45). Using this "∆RBM" probe, in addition to wildtype SARS-CoV-2 spike, and RBD probes, we isolated naive (CD19 + /IgD + /IgG -) B cells specific to the RBD and, more finely, the RBM from the peripheral blood of 8 SARS-CoV-2 seronegative human donors (Fig.…”

Stabilization of the SARS-CoV-2 Spike receptor-binding domain using deep mutational scanning and structure-based design

“…The SARS-CoV-2 RBD is more structurally homogeneous than metastable S ectodomains, with far higher expression yields (25,26). RBD-based immunogens in various oligomeric states have been investigated as genetic or protein-based vaccines for SARS-CoV-2, including monomers (27)(28)(29)(30)(31), dimers (32), trimers (33)(34)(35) and highly multivalent nanoparticles (25,(36)(37)(38)(39), several of which are now being evaluated in clinical trials. Although the RBD comprises only a minority of the total mass and antigenic surface of S and could in principle be more susceptible to escape mutations, this theoretical disadvantage may be mitigated by both functional constraints and the elicitation of antibodies targeting multiple distinct neutralizing epitopes in the RBD by infection or vaccination (25,(40)(41)(42)(43).…”