Human serum from SARS-CoV-2-vaccinated and COVID-19 patients shows reduced binding to the RBD of SARS-CoV-2 Omicron variant

Abstract

Background: The COVID-19 pandemic is caused by the betacoronavirus SARS-CoV-2. In November 2021, the Omicron variant was discovered and immediately classified as a variant of concern (VOC), since it shows substantially more mutations in the spike protein than any previous variant, especially in the receptor-binding domain (RBD). We analyzed the binding of the Omicron RBD to the human angiotensin-converting enzyme-2 receptor (ACE2) and the ability of human sera from COVID-19 patients or vaccinees in comparison to Wuhan, Beta, or Delta RBD variants.

Methods: All RBDs were produced in insect cells. RBD binding to ACE2 was analyzed by ELISA and microscale thermophoresis (MST). Similarly, sera from 27 COVID-19 patients, 81 vaccinated individuals, and 34 booster recipients were titrated by ELISA on RBDs from the original Wuhan strain, Beta, Delta, and Omicron VOCs. In addition, the neutralization efficacy of authentic SARS-CoV-2 wild type (D614G), Delta, and Omicron by sera from 2× or 3× BNT162b2-vaccinated persons was analyzed.

Results: Surprisingly, the Omicron RBD showed a somewhat weaker binding to ACE2 compared to Beta and Delta, arguing that improved ACE2 binding is not a likely driver of Omicron evolution. Serum antibody titers were significantly lower against Omicron RBD compared to the original Wuhan strain. A 2.6× reduction in Omicron RBD binding was observed for serum of 2× BNT162b2-vaccinated persons. Neutralization of Omicron SARS-CoV-2 was completely diminished in our setup.

Conclusion: These results indicate an immune escape focused on neutralizing antibodies. Nevertheless, a boost vaccination increased the level of anti-RBD antibodies against Omicron, and neutralization of authentic Omicron SARS-CoV-2 was at least partially restored. This study adds evidence that current vaccination protocols may be less efficient against the Omicron variant.

Keywords: Antibody titer; Beta variant (B.1.351); COVID-19; Delta variant (B.1.617.2); Human angiotensin-converting enzyme-2 receptor (ACE2); Omicron variant (B.1.1.529); Receptor-binding domain (RBD); SARS-CoV-2; Vaccination; Virus neutralization.

Fig. 1

RBD variants binding to human ACE2. 300 ng/well immobilized Wuhan wt, Beta, Delta, or Omicron RBD were detected with human ACE2 (fusion protein with human Fc part) in titration ELISA. BSA was used as a negative control. Experiments were performed in triplicates and mean values are given. EC₅₀ were calculated with OriginPro Version 9.1, fitting to a five-parameter logistic curve

Fig. 2

Human serum binding to SARS-CoV-2 Wuhan original strain, Beta, Delta, and Omicron RBD. A ELISA using sera from hospitalized COVID-19 patients. B ELISA using sera from 2×BNT162b2-vaccinated persons (7–52 days after 2nd immunization). C ELISA using sera from 1×Ad26.COV2.S-vaccinated (14–33 days 1st immunization). D ELISA using sera from 2×mRNA-1273 (5–55 days after 2nd immunization). E ELISA using sera from 2×BNT162b2 or 1× Ad26.COV2.S vaccinated + boosted with BNT162b2 or mRNA-1273 (5–49 days after 3rd or in case of Ad26.COV2.S 2nd immunization) binding to the Omicron variant. F Rearranged representation of the data presented in A–E. The ELISAs were performed as single-point titrations. The software Gen5 version 3.03 was used to calculate EC₅₀ values, further expressed as relative potency in respect to an internal calibrant, for which the Binding Antibody Unit (BAU) was calculated using the WHO International Standard 20/136 titrated on Wuhan wt as reference. The geometrical mean values and the 95% CI are given in the graphs. The graphics and statistical analysis were performed with Graphpad Prism 9.1. For A–E: Friedman test with Dunn’s multiple comparisons test was performed on the four conditions per graph (WT, Beta, Delta, Omicron). For F: Kruskal-Wallis test with Dunn’s multiple comparisons test was performed. Geometric mean and 95% confidence interval are represented by error bars. Multiplicity adjusted P values are shown as follows: ns: P > 0.05, *P ≤ 0.05, **P ≤ 0.01, ***P ≤ 0.001, ****P ≤ 0.0001

Fig. 3

Neutralization of SARS-CoV-2 wild type (D614G), Delta, and Omicron. Neutralization of authentic SARS-CoV-2 wt (including D614G mutation), Delta, and Omicron using sera of 2× BNT162b-vaccinated (A) and BNT162b2 boost-vaccinated (B) individuals. Delta, WT, and Omicron 90% neutralization titers (NT90) and median of values are shown from healthcare workers that underwent two-dose vaccination series (a) and three-dose vaccination series (b). Samples were collected 2 weeks and 3 weeks after the last dose, respectively. Reciprocal titers were log10 converted (1 was added to all titers to allow undetectable neutralization to be plotted). Upper and lower dotted lines represent the upper and lower limit of detection (the equivalent of 1:640 and 1:10 titers, respectively). Gray lines represent matched samples from the same donor. Friedman test with Dunn’s multiple comparison was calculated and P values are shown in asterisks