The current SARS-CoV-2 vaccines have successfully prevented severe disease, however the immune-protection they induce has not provided sufficient durability or breadth to prevent transmission of highly diverged new variants. Consequently, booster-vaccine updates targeting new spike variants aim for longer-lasting immunity. Our study introduces an mRNA vaccine candidate, termed 3RBD, which focuses immunity on the receptor binding domain (RBD) of spike, the primary target of neutralising antibodies, by presenting a polyvalent antigen linking three RBDs via a flexible linker. This design allows each RBD segment to maintain its unique strain-specific antigenicity, fostering antibody maturation toward common epitopes presented across each segment of the 3RBD polyvalent antigen.
Our proof-of-concept 3RBD antigen, incorporating Beta, Delta, and BA.1 variant RBDs with a C-terminal transmembrane domain, was expressed using optimised in vitro transcribed mRNA and delivered via lipid nanoparticle (LNP) formulation. C7BL/6 mice were immunised intramuscularly with three doses of 1 µg, 3 µg, or 10 µg, and subsequently challenged with a mouse-adapted strain of SARS-CoV-2 not represented in the vaccine.
The optimal antibody response was obtained after three doses of 10 µg of the vaccine, as assessed by ELISA binding and neutralising antibody assays. Mice elicited broad polyspecific RBD-antibodies against each variant expressed in the 3RBD antigen, indicating that the linkers do not interfere with antibody binding to individual RBDs. Mouse sera equally reacted with divergent strains, including a mouse-infectious ancestral strain not present in the vaccine. Importantly, vaccinated mice were protected from infection upon challenge with the non-homologous mouse-infectious strain.
Our study highlights that an mRNA vaccine expressing a membrane-tethered 3RBD antigen can induce robust polyspecific antibody responses against multiple variants of SARS-CoV-2, including those absent in the vaccine. Thus, the 3RBD mRNA vaccine emerges as a promising candidate for broad-spectrum protection against evolving SARS-CoV-2 variants, warranting further clinical investigation.