Effect of adjuvanting RBD-dimer-based subunit COVID-19 vaccines with Sepivac SWE™.

Protein subunit vaccines have been widely used to combat infectious diseases, including the current COVID-19 pandemic. Adjuvants play the key role in shaping the quality and magnitude of the immune response to protein and inactivated vaccines. We previously developed a protein subunit COVID-19 vaccine, termed ZF2001, based on an aluminium hydroxide-adjuvanted tandem-repeat dimeric receptor-binding domain (RBD) of the viral spike (S) protein. Here, we described the use of a squalene-based oil-in-water adjuvant, Sepivac SWE™ (abbreviated to SWE), to further improve the immunogenicity of this RBD-dimer-based subunit vaccines. Compared with ZF2001, SWE adjuvant enhanced the antibody and CD4+ T-cell responses in mice with at least 10 fold of dose sparing compared with ZF2001 adjuvanted with aluminium hydroxide. SWE-adjuvanted vaccine protected mice against SARS-CoV-2 challenge. To ensure adequate protection against the currently circulating Omicron variant, we evaluated this adjuvant in combination with Delta-Omicron chimeric RBD-dimer. SWE significantly increased antibody responses compared with aluminium hydroxide adjuvant and afforded greater neutralization breadth. These data highlight the advantage of emulsion-based adjuvants to elevate the protective immune response of protein subunit COVID-19 vaccines.

Structural characterization of SARS-CoV-2 dimeric ORF9b reveals potential fold-switching trigger mechanism.

The constant emergence of severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) variants indicates the evolution and adaptation of the virus. Enhanced innate immune evasion through increased expression of viral antagonist proteins, including ORF9b, contributes to the improved transmission of the Alpha variant; hence, more attention should be paid to these viral proteins. ORF9b is an accessory protein that suppresses innate immunity via a monomer conformation by binding to Tom70. Here, we solved the dimeric structure of SARS-CoV-2 ORF9b with a long hydrophobic tunnel containing a lipid molecule that is crucial for the dimeric conformation and determined the specific lipid ligands as monoglycerides by conducting a liquid chromatography with tandem mass spectrometry analysis, suggesting an important role in the viral life cycle. Notably, a long intertwined loop accessible for host factor binding was observed in the structure. Eight phosphorylated residues in ORF9b were identified, and residues S50 and S53 were found to contribute to the stabilization of dimeric ORF9b. Additionally, we proposed a model of multifunctional ORF9b with a distinct conformation, suggesting that ORF9b is a fold-switching protein, while both lipids and phosphorylation contribute to the switching. Specifically, the ORF9b monomer interacts with Tom70 to suppress the innate immune response, whereas the ORF9b dimer binds to the membrane involving mature virion assembly. Our results provide a better understanding of the multiple functions of ORF9b.

A tandem-repeat dimeric RBD protein-based covid-19 vaccine zf2001 protects mice and nonhuman primates.

Safe, efficacious, and deployable vaccines are urgently needed to control COVID-19 in the large-scale vaccination campaigns. We report here the preclinical studies of an approved protein subunit vaccine against COVID-19, ZF2001, which contains tandem-repeat dimeric receptor-binding domain (RBD) protein with alum-based adjuvant. We assessed vaccine immunogenicity and efficacy in both mice and non-human primates (NHPs). ZF2001 induced high levels of RBD-binding and SARS-CoV-2 neutralizing antibody in both mice and non-human primates, and elicited balanced TH1/TH2 cellular responses in NHPs. Two doses of ZF2001 protected Ad-hACE2-transduced mice against SARS-CoV-2 infection, as detected by reduced viral RNA and relieved lung injuries. In NHPs, vaccination of either 25 µg or 50 µg ZF2001 prevented infection with SARS-CoV-2 in lung, trachea, and bronchi, with milder lung lesions. No evidence of disease enhancement was observed in both animal models. ZF2001 has been approved for emergency use in China, Uzbekistan, Indonesia, and Columbia. The high safety, immunogenicity, and protection efficacy in both mice and NHPs found in this preclinical study was consistent with the results in human clinical trials.