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

Cohen, A. A.; Keeffe, J. R.; Manasyan, L.; Madireddy, I.; Fils, A.-C. I. P.; Dam, K.-M. A.; Stober, H. E.; Hills, R. A.; Moon, W. J.; Lin, P. J. C.; Howarth, M. R.; Hoffmann, M. A. G.; Bjorkman, P. J.

doi:10.1101/2025.10.21.683747

bioRxiv · 10.1101/2025.10.21.683747

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

Cohen, A. A.·Keeffe, J. R.·Manasyan, L.·Madireddy, I.·Fils, A.-C. I. P.·Dam, K.-M. A.·Stober, H. E.·Hills, R. A.·Moon, W. J.·Lin, P. J. C.·Howarth, M. R.·Hoffmann, M. A. G.·Bjorkman, P. J.

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Abstract

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.

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Cohen, A. A., Keeffe, J. R., Manasyan, L., Madireddy, I., Fils, A.-C. I. P., Dam, K.-M. A., Stober, H. E., Hills, R. A., Moon, W. J., Lin, P. J. C., Howarth, M. R., Hoffmann, M. A. G., Bjorkman, P. J.. 2025-10-22. mRNA delivery of genetically encoded mosaic-8 pan-sarbecovirus RBD vaccines. https://doi.org/10.1101/2025.10.21.683747

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