Manasyan, L. | bioRxiv Science

bioRxiv2025

mRNA delivery of genetically encoded mosaic-8 pan-sarbecovirus RBD vaccines

Effective pan-sarbecovirus vaccines could prevent future zoonotic spillovers of SARS-like betacoronaviruses. We previously developed protein-based mosaic-8 nanoparticles displaying eight diverse sarbecovirus RBDs, either individually (mosaic-8 RBD-NPs) or as two "quartets" of four tandemly-arranged RBDs (dual-quartet RBD-NPs), which elicited broadly cross-reactive antibodies but require multi-component manufacturing. Here, we address scalability challenges by extending the mosaic-8 concept to mRNA by encoding membrane-bound RBD quartets as dual-quartet RBD-mRNA and dual-quartet RBD-EABR-mRNA, the latter leveraging ESCRT- and ALIX-binding region (EABR) technology for immunogen display on cell surfaces and secreted virus-like particles. Compared with protein-based mosaic-8 immunogens, mRNA-encoded mosaic-8 vaccines induced equivalent or enhanced antibody breadth, neutralization potencies, T-cell responses, and targeting of conserved RBD epitopes. In addition, mRNA-encoded mosaic-8 vaccines elicited more balanced IgG subclass profiles and increased Fc{gamma} receptor-binding IgGs, consistent with potentially superior Fc effector functions. These findings demonstrate successful translation of mosaic-8 RBD-NPs into mRNA/EABR-mRNA vaccines, enabling scalable manufacturing and improving protection against future sarbecovirus outbreaks. Finally, our newly developed technique, Systems Serology-Polyclonal Epitope Mapping (SySPEM), revealed distinct IgG-subclass-specific epitope signatures across mRNA, EABR-mRNA, and protein vaccines, demonstrating that the mode of antigen display can shape epitope recognition. SummaryWe translated a pan-sarbecovirus RBD vaccine from protein nanoparticles to scalable mRNA and EABR-mRNA platforms encoding RBD quartets. Compared with protein-based immunogens, mRNA-based vaccines matched or improved antibody breadth, T-cell responses, Fc functionality, and conserved epitope targeting. A newly-developed Systems Serology-Polyclonal Epitope Mapping (SySPEM) technique revealed that antigen presentation modality shapes IgG subclass-specific epitope recognition.

immunology↗

bioRxiv2025

Single administration of mosaic-8b RBD-nanoparticle vaccine prepared with atomic layer deposition technology elicits broadly neutralizing anti-sarbecovirus responses

Atomic layer deposition (ALD), a new vaccine technology, permits multiple dosing with a single administration by pulsatile release of one or more immunogens. We evaluated ALD delivery of mosaic-8b [60-mer nanoparticles presenting 8 different SARS-like betacoronavirus (sarbecovirus) receptor-binding domains (RBDs)] that elicits broadly cross-reactive antibodies and protects against mismatched sarbecoviruses not represented by RBDs on mosaic-8b. Compared with conventional prime-boost immunizations, ALD-delivered mosaic-8b RBD-nanoparticles elicited antibodies in both naive and pre-vaccinated mice with improved mismatched binding and neutralization. Results of RBD epitope mapping of serum antibodies from ALD-delivered mosaic-8b were consistent with broader coverage of RBD epitopes compared to conventional immunizations, and systems serology revealed distinct IgG subclass and Fc{gamma}R-binding IgG distributions. These results suggest that ALD is a promising technology for use with mosaic-8b RBD-nanoparticle vaccines to protect against future sarbecovirus spillovers and support applications for ALD vaccine delivery to elicit cross-reactive antibodies against rapidly mutating or diverse pathogens.

immunology↗