Omicron BQ.1.1 and XBB.1 unprecedentedly escape broadly neutralizing antibodies elicited by prototype vaccination

Abstract

Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) Omicron subvariants have seriously attacked the antibody barrier established by natural infection and/or vaccination, especially the recently emerged BQ.1.1 and XBB.1. However, crucial mechanisms underlying the virus escape and the broad neutralization remain elusive. Here, we present a panoramic analysis of broadly neutralizing activity and binding epitopes of 75 monoclonal antibodies isolated from prototype inactivated vaccinees. Nearly all neutralizing antibodies (nAbs) partly or totally lose their neutralization against BQ.1.1 and XBB.1. We report a broad nAb, VacBB-551, that effectively neutralizes all tested subvariants including BA.2.75, BQ.1.1, and XBB.1. We determine the cryoelectron microscopy (cryo-EM) structure of VacBB-551 complexed with the BA.2 spike and perform detailed functional verification to reveal the molecular basis of N460K and F486V/S mutations mediating the partial escape of BA.2.75, BQ.1.1, and XBB.1 from the neutralization of VacBB-551. Overall, BQ.1.1 and XBB.1 raised the alarm over SARS-CoV-2 evolution with unprecedented antibody evasion from broad nAbs elicited by prototype vaccination.

Keywords: BQ.1.1; CP: Immunology; CP: Microbiology; Omicron subvariant; SARS-CoV-2; XBB.1; broadly neutralizing activity; structure analysis.

Graphical abstract

Figure 1

Neutralizing activity and binding epitope of 75 monoclonal nAbs isolated from prototype inactivated vaccinees (A and B) The neutralizing activity (IC₅₀) was measured based on the SARS-CoV-2 pseudovirus-neutralization assay, whose cutoff value was set as 50 μg/mL (summarized in A and detailed in B). The data are means of at least two independent experiments. The positive rate, geometric mean IC₅₀, fold change, and significance of difference are labeled on the top. “-” represents decreased neutralization. The statistical significance was performed using two-tailed paired Wilcoxon test. ^∗∗∗∗p < 0.0001; ^∗∗p < 0.01. The neutralizing potency is represented by a heatmap. Red: high, yellow: moderate, and green: weak. (B–D) Competition ELISA was performed to predict the binding epitope of 75 nAbs (detailed in B and summarized in C and D). Human ACE2 and four representative mAbs of four classes (class 1: P2C-1F11, class 2: BD-368-2, class 3: S309, and class 4: EY6A) were used as the competitor for binding to the WT RBD. The data are means of at least two independent experiments. Dark blue: high competition (>90%), light blue: moderate competition (45%–90%), and none: weak or no competition (<45%). “Undefined” means that mAbs do not compete with any tested references (P2C-1F11, BD-368-2, S309, and EY6A). The classifications of binding epitopes of mAbs are indicated in different colors, which are identical in (A), (B), and (D). See also Figures S1 and S2 and Table S1.

Figure 2

Cryo-EM structure basis of VacBB-551 binding to Omicron BA.2 RBD (A) The cryo-EM density map of VacBB-551 in complex with Omicron BA.2 spike trimer. Spike is shown in gray and RBDs in orange. Heavy chain of VacBB-551 is shown in dark cyan and light chain in deep pink. Atomic model of RBD-Fab interacting region is shown as a cartoon, fitted in the corresponding transparency density map. (B) Structure and binding footprint of VacBB-551 on BA.2 RBD. ACE2 is shown in medium violet red and VacBB-551 in orange. Epitopes recognized by VacBB-551 are listed, and mutant positions that appeared in tested Omicron subvariants are highlighted in red. The interface area of the ACE2-BA.2 RBD was calculated based on a published structure (PDB: 7XB0). (C–F) Interactions of heavy chain (C–E) and light chain (F) of VacBB-551 with BA.2 RBD. Heavy chain is shown in cyan and light chain in magenta. Interface area (Ų) and potential hydrogen bond (black dotted line) or salt bridge (black solid line) were calculated by PISA v.1.52 (https://www.ebi.ac.uk/pdbe/pisa). See also Figures S3 and S4 and Tables S2 and S3.

Figure 3

Sequence alignment and structural analysis of Omicron BA.2.75, BQ.1.1, and XBB.1 subvariants (A) Key mutations in the spike protein of Omicron BA.2, BA.2.75, BQ.1.1, and XBB.1. “^∗” represents mutations appeared in the binding epitopes of VacBB-551. (B–E) Local region of N460 and F486 of BA.2 RBD (B), K460 and F486 of BA.2.75 RBD (C), K460 and V486 of BQ.1.1 RBD (D), and K460 and S486 of XBB.1 RBD (E) with G54 and R97 of VacBB-551 heavy chain, respectively. The structures of BA.2.75, BQ.1.1, and XBB.1 RBDs were predicted by AlphaFold2 (https://colab.research.google.com/github/sokrypton/ColabFold/blob/main/AlphaFold2.ipynb). The clash discs were displayed using the “Show bumps” plugin (https://pymolwiki.org/index.php/Show_bumps) in Pymol (C–E). The pseudoatom of F486 is shown as nb_sphere and the cation-π interaction is shown in a yellow dashed line (B and C). N460, K460, F486, V486, S486, G54, and R97 residues were shown as sticks. The VacBB-551 heavy chain was shown as cartoon/surface colored by cyan. RBDs of BA.2, BA.2.75, BQ.1.1, and XBB.1 are shown in gray, orange, smudge, and yellow, respectively. See also Figure S5.

Figure 4

Binding affinity of Fab-form and IgG-form VacBB-551 to WT and mutated RBD proteins SPR analysis of Fab-form VacBB-551 (A) and IgG-form VacBB-551 (B) binding to WT, N460K, F486V, F486S, N460K+F486V, or N460K+F486S RBD proteins, respectively. The dissociation constant (K_D), association rate constant (K_on), and dissociation rate constant (K_off) are calculated from three independent experiments and represented in mean values ±standard deviation (SD). One representative curve is presented here.

Figure 5

Functional verification of N460 and F486 mutations influencing the neutralization susceptibility of SARS-CoV-2 variants to VacBB-551 The neutralization of VacBB-551 against the WT-related (A), BA.2-related (B), BA.2.75-related (C), BQ.1.1-related (D), and XBB.1-related (E) mutated pseudoviruses. Fold changes of neutralization (IC₅₀) are calculated on mean values of two independent experiments. One representative curve is presented here. “-” represents decreased neutralization, and “+” represents increased neutralization.