Omicron-specific mRNA vaccination alone and as a heterologous booster against SARS-CoV-2

Abstract

The Omicron variant of SARS-CoV-2 recently swept the globe and showed high level of immune evasion. Here, we generate an Omicron-specific lipid nanoparticle (LNP) mRNA vaccine candidate, and test its activity in animals, both alone and as a heterologous booster to WT mRNA vaccine. Our Omicron-specific LNP-mRNA vaccine elicits strong antibody response in vaccination-naïve mice. Mice that received two-dose WT LNP-mRNA show a > 40-fold reduction in neutralization potency against Omicron than WT two weeks post boost, which further reduce to background level after 3 months. The WT or Omicron LNP-mRNA booster increases the waning antibody response of WT LNP-mRNA vaccinated mice against Omicron by 40 fold at two weeks post injection. Interestingly, the heterologous Omicron booster elicits neutralizing titers 10-20 fold higher than the homologous WT booster against Omicron variant, with comparable titers against Delta variant. All three types of vaccination, including Omicron alone, WT booster and Omicron booster, elicit broad binding antibody responses against SARS-CoV-2 WA-1, Beta, Delta variants and SARS-CoV. These data provide direct assessments of an Omicron-specific mRNA vaccination in vivo, both alone and as a heterologous booster to WT mRNA vaccine.

Fig. 1. Design and biophysical characterization of Omicron-specific LNP-mRNA vaccine.

a Illustration of mRNA vaccine construct expressing SARS-CoV-2 WT and Omicron spike genes. The spike open reading frame were flanked by 5′ untranslated region (UTR), 3′ UTR, and polyA tail. The Omicron mutations (red) and HexaPro mutations (black) were numbered based on WA-1 spike residue number. b Distribution of Omicron spike mutations (magenta) were displayed in one protomer of spike trimer of which N-terminal domain (NTD), receptor-binding domain (RBD), hinge region and S2 were colored in purple, blue, green, and orange respectively (PDB: 7SBL). The HexaPro mutations in S2 were colored in cyan. c Schematics illustrating the formulation and biophysical characterization of lipid nanoparticle (LNP)-mRNA. Created with BioRender.com. d Dynamic light scattering derived histogram depicting the particle radius distribution of Omicron spike LNP-mRNA. e Omicron LNP-mRNA image collected on transmission electron microscope. f Human ACE2 receptor binding of LNP-mRNA encoding Omicron spike expressed in 293T cells as detected by human ACE2-Fc fusion protein and PE-anti-human Fc antibody on Flow cytometry.

Fig. 2. Omicron-specific LNP-mRNA vaccine-elicited neutralizing antibodies against SARS-CoV-2 Omicron variant.

a Immunization and sample collection schedule. Retro-orbital blood was collected prior Omicron LNP-mRNA vaccination on day 0, day 13, and day 21. Ten mice (n = 10) were intramuscularly injected with 10 µg Omicron LNP-mRNA on day 0 (prime, Omicron × 1) and day 14 (boost, Omicron × 2). The plasma and peripheral blood mononuclear cells (PBMCs) were separated from blood for downstream assays. The slight offset of the labels reflects the fact that each of the blood collections were performed prior to the vaccination injections. Data were collected from two independent experiments and each experiment has five mice. Created with BioRender.com. b Binding antibody titers of plasma from mice vaccinated with Omicron LNP-mRNA against Omicron spike RBD as quantified by area under curve of log₁₀-transformed titration curve (Log₁₀ AUC) in Supplementary Fig. 1. Each dot in bar graphs represents the value of one mouse (n = 10 mice). c Neutralization of Omicron pseudovirus by plasma from Omicron LNP-mRNA vaccinated mice. d Omicron live virus titration curves over serial dilution points of plasma from mice before and after immunization with Omicron LNP-mRNA at defined time points. Data of each sample were collected from three replicates (n = 10 mice). e Neutralization of Omicron infectious virus by plasma from Omicron LNP-mRNA vaccinated mice (n = 10 mice). Data on dot-bar plots are shown as mean ± s.e.m. with individual data points in plots. One-way ANOVA with Dunnett’s multiple comparisons test was used to assess statistical significance. Statistical significance labels: *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001.

Fig. 3. Heterologous booster with Omicron LNP-mRNA as compared to homologous booster with WT LNP-mRNA in mice that previously received a two-dose WT LNP-mRNA vaccination.

a Schematics showing the immunization and blood sampling schedule of mice administered with 1 µg WT LNP-mRNA prime (WT × 1) and boost (WT × 2) as well as 10 µg WT or Omicron-specific LNP-mRNA booster shots. The data was collected and combined from two independent experiments shown in Supplementary Fig. 2 and 3. Created with BioRender.com. b Bar graph comparing binding antibody titers of mice administered with PBS or WT and Omicron LNP-mRNA against Omicron, Delta, and WA-1 RBD (ELISA antigens). The antibody titers were quantified as Log₁₀ AUC based on titration curves in Supplementary Fig. S1a. PBS sub-groups (n = 6 each) collected from different matched time points showed no statistical differences between each other, and were combined as one group (n = 18). c Pseudovirus neutralizing antibody titers in the form of log₁₀-transformed reciprocal IC50 calculated from fitting the titration curve with a logistic regression model (n = 12 mice before booster, n = 5 in WT × 3, n = 7 in WT × 2 + Omicron). d Infectious virus neutralization titer comparisons between mice before and after vaccination with WT or Omicron boosters (n = 9 mice before booster, n = 5 in WT × 3, n = 4 in WT × 2 + Omicron). Titer ratios were indicated in each graph and fold change described in manuscript is calculated from (ratio − 1). Data on dot-bar plots are shown as mean ± s.e.m. with individual data points in plots. Two-way ANOVA with Tukey’s multiple comparisons test was used to assess statistical significance. Statistical significance labels: *p < 0.05; **p < 0.01; ***p < 0.001; ****p < 0.0001. Non-significant comparisons are not shown, unless otherwise noted as n.s., not significant. Sample number is designated as n from biologically independent samples.

Fig. 4. Cross reactivity and targeting sites characterization of plasma antibodies elicited by Omicron and WT LNP-mRNAs against SARS-CoV-2 VoCs and Betacoronavirus species.

a Cross reactivity of plasma antibody from mice immunized with Omicron LNP mRNA (prime and boost) to SARS-CoV-2 VoCs and pathogenic coronavirus species (n = 10 mice). b cross reactivity of plasma antibody from mice immunized with WT (WT × 3) or Omicron (WT × 2 + Omicron) boosters to SARS-CoV-2 beta variant and pathogenic coronavirus species (n = 6 mice in PBS, n = 5 in WT × 3, n = 7 in WT × 2 + Omicron). c Representative antibodies from major classes of RBD epitopes were shown by aligning spike RBDs in each of complex structures. The Omicron RBD surface was set to semi-transparent to visualize 15 RBD mutations and their relative positions to antibody epitopes. d baseline titers of plasma from mice of different vaccination status (WT × 3, WT × 2 + Omicron, Omicron × 2) were shown as log₁₀ AUC determined in hACE2 and antibody competition ELISA. Each group sample number is denoted with n (n = 10 in Omicron × 2, n = 5 in WT × 3, n = 7 in WT × 2 + Omicron) in two independent assays (hACE2 and antibody competition ELISA). e Significant portion of plasma antibody from mice receiving Omicron (Omicron × 2, left panel) or WT + Omicron (WT × 3 middle, or WT × 2 + Omicron, right panel) LNP-mRNA competed with hACE2 for Omicron RBD binding in ELISA (n = 10 in Omicron × 2, n = 5 in WT × 3, n = 7 in WT × 2 + Omicron). f Plasma antibody from mice receiving Omicron (Omicron × 2, n = 10, left panel) or WT + Omicron (WT × 3, n = 5, middle or WT × 2 + Omicron, n = 7, right panel) LNP-mRNA showed various extent of binding reduction in the presence of blocking antibodies with known epitopes on RBD. The error bar and statistical information are identical with Fig. 3 and described in Method section.