Thread by @veeslerlab

Curious about #COVID19 #SARSCoV2 therapeutic antibodies, bivalent vaccines & immune imprinting?

Here is our latest work on these topics in collaboration with @DavideCorti6 @msdiamondlab

Led by @AminAddetia @LucaPiccoli9 Brett Case @YoungjunPark11

1/
biorxiv.org/content/10.110…

Currently circulating #SARSCoV2 variants are evolving from the Omicron BA.2 and BA.5 lineages, through independent accumulation of amino acid mutations at shared receptor-binding domain (RBD) hot spots, underscoring the importance of antibody-mediated selective pressure.

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We first studied how mutations in the #SARSCoV2 BQ.1.1, XBB.1(.5) and BA.2.75.2 variants affect function.
@AminAddetia @johnbowenbio in our lab previously showed that BA.5 affinity for ACE2 was >> Wu but fusogenicity was worse

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science.org/doi/10.1126/sc…

We found that the (dominant) BQ.1.1 RBD and the BA.2.75.2 RBD have similar ACE2 binding affinity to the BA.5 RBD whereas XBB.1 RBD has lower ACE2 affinity (similar to Wu RBD).
Modulation results from off-rate differences, as we have seen throughout the pandemic

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We also observed that spike-mediated membrane fusion (leading to viral entry) is more efficient with BQ.1.1, XBB.1(.5) and BA.2.75.2 than previous Omicron variants, almost reaching Wu levels

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BQ.1.1 and BA.2.75.2 viral fitness is therefore not limited by ACE2 binding whereas the lower ACE2-binding affinity of XBB.1 might have limited its spread.
Recent data from @yunlong_cao showing enhanced ACE2 binding of XBB.1.5 (S486P) support this!

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biorxiv.org/content/10.110…

BA.1, BA.2 and BA.5 had altered cell entry pathway vs previous #SARSCoV2 strains, with Omicron variants entering preferentially through the endosomal entry route (cathepsin-mediated) as opposed to plasma membrane fusion (TMPRSS2-mediated)
@GuptaR_lab

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nature.com/articles/s4158…

To assess the preferred cell entry route of currently circulating variants, we investigated the impact of protease inhibitors on spike-mediated entry into VeroE6-TMPRSS2 cells (enabling both plasma membrane and endosomal entry routes)

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Camostat and nafamostat (TMPRSS2 inhibitors) had limited impact on Omicron variants, including BQ.1.1, BA.2.75.2 and XBB.1, whereas E64d (cathepsin B/L inhibitor) reduced BA.1, BA.2 and BA.5 entry more than BA.2.75.2, BQ.1.1 or XBB.1, relative to Wu-G614

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Inefficient use of TMPRSS2 concurs with the identical BQ.1.1, BA.2.75.2 and XBB.1 S2 subunit sequences, including spike N969K which @PeacockFlu previously showed to account for preferred endosomal cell entry of prior Omicron variants (BA.2)

10/

biorxiv.org/content/10.110…

To reveal how amino acid substitutions in the BQ.1.1 and XBB.1 RBDs alter receptor recognition and key antigenic sites, we determined #cryoEM structures of the RBDs bound to ACE2 & S309 Fab (parent of the sotrovimab monoclonal antibody therapeutic)

11/

The R493Q reversion likely relieves repulsion with ACE2 residue K31 and restores a network of local interactions similar to that made with the Wu RBD

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These data provide a blueprint to contextualize deep-mutational scanning and mutagenesis data we recently reported in the context of the BA.1 and BA.2 RBD backgrounds with @tylernstarr @jbloom_lab

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journals.plos.org/plospathogens/…

Our #cryoEM structures reveal how the BQ.1.1 K444T and the BQ.1.1/XBB.1 R346T would abrogate key electrostatic interactions with the LY-CoV1404 (bebtelovimab parent) and COV2-2130 (cilgavimab parent, Evushield cocktail), explaining dampened binding and neutralization

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We show how S309 binds both the BQ.1.1 and XBB.1 RBDs and accommodates the XBB.1 H339 residue in the epitope! S309 binding pose is indistinguishable from that observed when bound to the Wu/BA.1 RBDs

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The S371F mutation (BA.2, BA.5, BQ.1.1, XBB.1 and BA.2.75.2) changes conformation of the RBD helix 364-372 which becomes sterically incompatible with the glycan N343 conformation observed in S309-bound spike structures

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science.org/doi/10.1126/sc…

The BQ.1.1 helix has a conformation similar to that in the S309-bound BA.1 structure, but distinct from apo BA.2 /BA.5 structures, and is disordered for XBB.1, indicative of conformational frustration upon S309 binding which could explain reduced neutralizing activity

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We found that S309 retains detectable neutralizing activity of currently circulating variants, including the rising XBB.1.5 and the dominant BQ.1.1, with potencies correlating with 1:1 Fab binding affinity to variant RBDs

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Sotrovimab (S309 derivative) IgG binds avidly to cell surface-expressed currently dominant Omicron #SARSCoV2 spike variants & promoted antibody-dependent cell cytotoxicity (ADCC) using primary natural killer effector cells (independently of 1:1 Fab affinity)!

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Prophylactic administration of S309 protected K18-hACE2 transgenic mice from weight loss and reduced both viral RNA and infectious virus titers in the lungs.
Despite marked reduction of in vitro neutralizing activity, S309 can protect mice from BQ.1.1 challenge!

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To assess the impact of the BQ1.1, XBB.1 and BA.2.75.2 S mutations on vaccine-elicited antibody responses, we quantified plasma neutralizing activity in various human cohorts infected and/or vaccinated with monovalent and bivalent mRNA vaccines

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Wu/BA.5 or Wu/BA.1 bivalent mRNA vaccinated subjects had detectable neutralizing activity vs vaccine-mismatched XBB.1, BA.2.75.2 and BQ.1.1 (w/wo prior infection) whereas little to no neutralization of these variants was detected for the Wu-only vaccinated subjects

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These data suggest that bivalent (Wu/BA.1 or Wu/BA.5) mRNA vaccination elicits more potent and broader antibody responses against vaccine-matched and mismatched Omicron variants than monovalent Wu mRNA vaccination in healthy subjects

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We further observed than vaccine-elicited polyclonal plasma antibodies retain comparable binding titers across all Omicron variants and that in some subjects they are triggering Fc effector functions against circulating Omicron variants.

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Analysis of memory B cell responses in these human cohorts showed that bivalent Wu/BA.5 mRNA vaccination enriches for MBCs that are cross-reactive with the vaccine-matched and mismatched RBD variants, relative to monovalent Wu mRNA vaccination, in line with neutralization.

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Surprisingly, few de novo elicited Omicron-specific memory B cells were detected even after two exposures to an Omicron spike via breakthrough infection followed by Wu/BA.5 bivalent mRNA vaccination.

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We did not detect any de novo elicited Omicron-specific memory B cells after Wu/BA.1 bivalent mRNA vaccination (w/wo Omicron infection) and most RBD-directed IgGs secreted by stimulated memory B cells cross-react with BA.1 and (to a lesser extent) with BQ.1.1 and XBB.1 RBDs

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Collectively, these data suggest that two Omicron spike exposures are not sufficient to overcome the immunological imprinting induced by repeated Wu spike exposures but does continue to enrich for memory B cells cross-reacting with multiple RBD variants

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Thank you so much to @sprouse_kaitlin Cameron Stewart, Courtney Yoshiyama & all our wonderful collaborators including @LanzavecchiaB @DrLisaPurcell @HelenChuMD @HaVanDang2712 @SkiClimbSciNick