Bivalent SARS-CoV-2 mRNA vaccines increase breadth of neutralization and protect against the BA.5 Omicron variant

Abstract

The emergence of SARS-CoV-2 variants in the Omicron lineage with large numbers of substitutions in the spike protein that can evade antibody neutralization has resulted in diminished vaccine efficacy and persistent transmission. One strategy to broaden vaccine-induced immunity is to administer bivalent vaccines that encode for spike proteins from both historical and newly-emerged variant strains. Here, we evaluated the immunogenicity and protective efficacy of two bivalent vaccines that recently were authorized for use in Europe and the United States and contain two mRNAs encoding Wuhan-1 and either BA.1 (mRNA-1273.214) or BA.4/5 (mRNA-1273.222) spike proteins. As a primary immunization series in BALB/c mice, both bivalent vaccines induced broader neutralizing antibody responses than the constituent monovalent vaccines (mRNA-1273 [Wuhan-1], mRNA-1273.529 [BA.1], and mRNA-1273-045 [BA.4/5]). When administered to K18-hACE2 transgenic mice as a booster at 7 months after the primary vaccination series with mRNA-1273, the bivalent vaccines induced greater breadth and magnitude of neutralizing antibodies compared to an mRNA-1273 booster. Moreover, the response in bivalent vaccine-boosted mice was associated with increased protection against BA.5 infection and inflammation in the lung. Thus, boosting with bivalent Omicron-based mRNA-1273.214 or mRNA-1273.222 vaccines enhances immunogenicity and protection against currently circulating SARS-CoV-2 strains.

Figure 1.. Robust antibody responses in BALB/c mice after a primary immunization series with preclinical versions of monovalent and bivalent mRNA vaccines.

Six-to-eight-week-old female BALB/c mice were immunized twice over a three-week interval with PBS or 1 μg total dose of preclinical versions of mRNA-1273 [Wuhan-1 spike], mRNA-1273.529 [BA.1 spike], mRNA-1273.045 [BA.4/5 spike], mRNA-1273.214 [benchside 1:1 mixture of mRNA-1273 + mRNA-1273.529], or mRNA-1273.222 [benchside 1:1 mixture of mRNA-1273 + mRNA-1273.045]. Immediately before (day 21) or two weeks after (day 35) the second vaccine dose, serum was collected. A. Scheme of immunization and blood draws. B. Serum antibody binding to Wuhan-1 (S2P), BA.1 (S2P.529), or BA.4/5 (S2P.045) spike proteins by ELISA at Day 21 and Day 35 (n = 8, boxes illustrate mean values, dotted lines show the limit of detection [LOD]). C. Neutralizing activity of serum at day 35 against VSV pseudoviruses displaying the spike proteins of Wuhan-1 D614G, BA.1, BA.2.75, or BA.4/5 (n = 8, boxes illustrate geometric mean values, dotted lines show the LOD). GMT values are indicated above the columns. D. Neutralizing activity of serum at day 35 against pseudotyped lentiviruses displaying the spike proteins of Wuhan-1, BA.1, or BA.4/5 (n = 8, boxes illustrate geometric mean values, dotted lines show the LOD). GMT values are indicated above the columns. Statistical analysis. B. One-way ANOVA with Dunnett’s post-test. C-D. Kruskal-Wallis with Dunn’s post-test (ns, not significant; * P < 0.05; ** P < 0.01; *** P < 0.001; **** P < 0.0001).

Figure 2.. Robust neutralizing antibody responses in BALB/c mice after primary series immunization with clinically representative versions of mRNA-1273, mRNA-1273.214, and mRNA-1273.222.

Six-to-eight-week-old female BALB/c mice were immunized twice over a three-week interval with PBS or 1 μg total dose of clinically representative versions of mRNA-1273, mRNA-1273.214 [1::1 mixture in the vial of separately formulated mRNA-1273 and mRNA-1273.529], or mRNA-1273.222 [1:1 mixture in the vial of separately formulated mRNA-1273 and mRNA-1273.045]. Immediately before (day 21) or two weeks after (day 35) the second vaccine dose, serum was collected. A. Scheme of immunization and blood draws. B. Neutralizing activity of serum at day 35 against VSV pseudoviruses displaying the spike proteins of Wuhan-1 D614G, BA.1, BA.2.75, or BA.4/5 (n = 16, boxes illustrate geometric mean values, dotted lines show the LOD). GMT values are indicated above the columns. Statistical analysis. Kruskal-Wallis with Dunn’s post-test (ns, not significant; ** P < 0.01; **** P < 0.0001).

Figure 3.. Neutralizing antibody responses in K18-hACE2 mice after boosting with clinically representative versions of mRNA-1273, mRNA-1273.214, and mRNA-1273.222.

Seven-week-old female K18-hACE2 mice were immunized with 0.25 μg of control mRNA or mRNA-1273 vaccine and then boosted 31 weeks later with PBS, 0.25 μg of control mRNA, or 0.25 μg of clinically representative versions of mRNA-1273, mRNA-1273.214, or mRNA-1273.222 vaccines. A. Scheme of immunizations, blood draws and virus challenge. B-C. Serum neutralizing antibody responses immediately before (B, pre-boost) and four weeks after (C, post-boost) receiving the indicated mRNA boosters or PBS as judged by focus reduction neutralization test (FRNT) with authentic WA1/2020 D614G, B.1.617.2, BA.1, and BA.5 viruses (n = 9–10, two experiments, boxes illustrate geometric mean values, GMT values are indicated at the top of the graphs, dotted lines show the LOD). Statistical analysis. C: Kruskal-Wallis with Dunn’s post-test, ns, not significant; * P < 0.05; ** P < 0.01; *** P < 0.001; **** P < 0.0001).

Figure 4.. Protection of K18-hACE2 mice from BA.5 challenge after boosting with clinically representative versions of mRNA-1273, mRNA-1273.214, and mRNA-1273.222.

Seven-week-old female K18-hACE2 mice were immunized with 0.25 μg of control mRNA or mRNA-1273, boosted 31 weeks later with PBS, 0.25 μg of control mRNA, or 0.25 μg of clinically representative versions of mRNA-1273, mRNA-1273.214, or mRNA-1273.222 vaccines, and then one month later challenged via intranasal route with 10⁴ focus-forming units (FFU) of BA.5. A. Viral RNA levels at 4 dpi in the nasal washes, nasal turbinates, and lungs (n = 8–10 per group, two experiments, boxes illustrate mean values, dotted lines show LOD; One-way ANOVA with Dunnett’s post-test: * P < 0.05; ** P < 0.01; *** P < 0.001; **** P < 0.0001). B. Infectious viral load at 4 dpi in the lungs after BA.5 challenge of vaccinated and boosted mice as determined by plaque assay (n = 8–10 per group, two experiments, boxes illustrate mean values, dotted lines show LOD; Kruskal Wallis with Dunn’s post-test: ns, P > 0.05; **** P < 0.0001). C. Cytokine and chemokine levels in lung homogenates at 4 dpi. Data are expressed as fold-change relative to naive mice, and log₂ values are plotted (n = 8–10 per group except naïve where n = 4, two experiments, lines illustrates median values, dotted lines indicate LOD for each respective analyte based on standard curves; one-way Kruskal Wallis ANOVA with Dunn’s post-test: ns, P > 0.05; * P < 0.05; ** P < 0.01; *** P < 0.001; **** P < 0.0001). The absolute values are shown in Supplementary Table S1.