A recent study revealed the dynamic evolutionary pathway of the SARS-CoV-2 spike (S) protein for the first time (Zhao et al., 2023). By tracking the evolution and mutation of SARS-CoV-2 S proteins, the research team proposed a novel strategy to optimize vaccine immunogen design to fight Delta, Omicron, and potential future variants.
Under selective pressure from human and vaccine immunity, SARS-CoV-2 has evolved from the original strain into a multi-lineage variant. The World Health Organization named SARS-CoV-2 variants as variants of concern (VOC) and variants of interest (VOI) based on their transmissibility, virulence, disease severity, and impact on existing drugs and vaccines. The leading global spread of Omicron and its sublineage is affiliated with VOC.
However, most of the vaccines approved for marketing are still designed with the original strain of SARS-CoV-2. The neutralization ability for Omicron and its sublineage strains has decreased to varying degrees in the real world. The breakthrough infection still occurs after multiple vaccinations, which significantly challenges preventing and controlling the pandemic. Therefore, the development of a broad-spectrum COVID-19 vaccine is crucial.
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The S protein of SARS-CoV-2 contains two fragments, the amino-terminal S1 subunit that recognizes and interacts with the host ACE2 receptor and the carboxy-terminal S2 subunit that catalyzes the fusion of the virus and the cell membrane. The S2 subunit drives the release of viral RNA into the cell, where it replicates within the infected cell. Compared with the original strain of SARS-CoV-2, most of the mutation sites of the main VOC strains are located in the S protein, so predicting the mutation pattern of the S protein may become a candidate strategy for vaccine development.
To explore SARS-CoV-2 variants and variant S proteins, the researchers constructed 54 pseudoviruses expressing the original strain S protein (Swt) and variant S proteins. Susceptible cell lines such as BHK21-hACE2, Vero E6, Caco- 2, and A549-hACE2 were evaluated for the infectivity of the pseudoviruses mentioned above. The results showed that the infectivity of the Alpha, Beta, Kappa, Delta, and Lambda variants was increased by 1.77, 3.38, 2.90, 4.24, and 4.05 fold, respectively, compared with the D614G control group.
The emerging Omicron variant showed high infectivity (4.74-fold increase) in the Vero E6 cell line. Still, its infectivity decreased by an average of 1.58 fold in the other three cell lines, indicating that Omicron prefers the endocytic pathway.
The above results suggest that the SARS-CoV-2 variants have significant in vitro infectious activity.
Immune Escape Ability
To study the immune escape ability of the variant S protein, the researchers tested the neutralizing effect of 14 commercially available monoclonal neutralizing antibodies against the original strain S1 or the original strain RBD on 54 pseudoviruses. The average sensitivity of the alpha variant to mAbs was similar to that of the control D614G variant, whereas the beta variant showed higher immune evasion. Due to the E484K mutation, the Gamma and Eta variants exhibit a broader escape spectrum against partially neutralizing antibodies. It is worth noting that the Delta and Lambda variants showed only slight immune escape, and the Omicron variant showed resistance to neutralizing antibodies.
The above results indicate a decrease in the neutralizing activity of monoclonal neutralizing antibodies against SARS-CoV-2 variants.
S Protein Evolution
The different mutation paths of the S protein lead to changes in the antigenicity of the mutant strains, and the vaccine antigen components of a single strain cannot effectively protect the population against other variants on different evolutionary paths. To further understand the evolution trend and characteristics of the SARS-CoV-2 S protein, the researchers drew an evolutionary dynamics map based on the immune escape and infection capabilities of the S protein of different mutant SARS-CoV-2 strains to guide the design of the immunogen Span antigen sequence.
Subsequently, by reconstructing the phylogenetic tree, the researchers confirmed that the Span antigen sequence is located in the center of the phylogenetic tree, showing high consistency characteristics with other evolutionary lines. The Omicron variant and the Span antigen sequence are clustered in the same clade, suggesting that the Span antigen sequence has universality across evolutionary branches. The Span antigen sequence is the most representative and universal sequence among the SARS-CoV-2 strains.
Broad-Spectrum COVID-19 Vaccine
To verify whether the Span antigen can be used as a universal S protein immunogen against SARS-CoV-2 variants, the researchers expressed the S protein trimer of Swt and SSwt through the CHO-K1 eukaryotic expression system (Swt and Span reagents related to protein expression and purification were provided by GenScript). Swt and Span protein vaccines were prepared and vaccinated in keratin 18 (K18)-hACE2 mice.
Sera from mice vaccinated with Swt protein showed reduced neutralizing activity against all VOIs or VOCs (especially Omicron and Beta). In contrast, those from mice immunized with Span showed neutralizing activity against all ten variants, suggesting that antibodies induced by Span antigen vaccines can broadly neutralize SARS-CoV-2 variants.
To evaluate the protective effect of the Span vaccine against the actual SARS-CoV-2 virus, the researchers immunized K18-hACE2 mice with human doses of Span and Swt vaccines (25 μg per dose) to achieve appropriate titers of neutralizing antibodies.
Mice in the placebo group died between day 5 and 7 after infection. The Swt vaccine protected mice against WT and Delta virus challenge but completely lost protection when challenged with the Beta variant. In contrast, the Span vaccine was protective against WT and Delta viruses (80% and 100%, respectively). It remained protective against the Beta variant (60%), suggesting that the Span vaccination protects K18-hACE2 mice from WT, Beta, and Delta SARS-CoV-2 infection.
Most primary vaccine immunizations have been based on the original strain SARS-CoV-2 S protein, so the researchers further evaluated the protective effect of Span as a heterologous booster vaccine against the circulating strain BA.1.
The study showed that a third dose of Span or Swt booster vaccination produced higher titers of neutralizing antibodies against both Beta and Delta pseudoviruses. But only the serum of mice inoculated with the Span booster vaccine can produce effective neutralizing antibodies against Omicron and its subvariants.
This work is the first to report the evolutionary dynamics of the SARS-CoV-2 S protein. It is worth noting that the universal S protein immunogen Span was designed before the emergence of the Delta variant. Still, it can efficiently induce broad-spectrum neutralizing antibodies against the Delta and Omicron variants, indicating that the design based on the common evolutionary mutations of Span is expected to create a broad-spectrum vaccine candidate for the prevention of current and potential future strains of SARS-CoV-2.
Yongliang ZHAO, et al. Vaccination with Span, an antigen guided by SARS-CoV-2 S protein evolution, protects against challenge with viral variants in mice. Sci Transl Med. 2023,15（677）:eabo3332. doi: 10.1126/scitranslmed.abo3332.