Developing an eVLP mRNA vaccine for respiratory syncytial virus with enhanced pre-fusion targeting humoral responses.

Respiratory syncytial virus (RSV) is a leading cause of lower respiratory tract disease (LRTD) in infants, the elderly, and immunocompromised individuals. In this study, we leveraged the innovative EABR (ESCRT- and ALIX-binding region) vaccine technology to develop a pre-fusion (Pre-F)-based RSV mRNA vaccine (Pre-F-EABR) that encodes self-assembling enveloped virus-like particles (eVLPs). In vitro expression of Pre-F-EABR mRNA manifested pronounced Pre-F protein expression, particularly on cell membranes and in cell culture supernatant, compared to classical Pre-F and Pre-F-Ferritin nanoparticle (Pre-F-Fe) mRNAs, suggesting efficient eVLP self-assembly. Immunization of mice with lipid nanoparticle-encapsulated, nucleoside-modified mRNAs (mRNA-LNPs) demonstrated that the Pre-F-EABR vaccine elicited enhanced neutralizing activities and more robust cellular immunity when compared to Pre-F or Pre-F-Fe mRNA vaccines. Notably, the Pre-F-EABR mRNA vaccine induced a significantly higher level of protective pre-fusion-specific antibodies in contrast to post-fusion-targeting antibodies, dramatically reducing pulmonary viral load following RSV challenge in vaccinated mice. Furthermore, the Pre-F-EABR mRNA vaccine demonstrated significantly improved efficacy against the contemporary clinical ON1 and BA9 isolates. Overall, these findings demonstrate that the Pre-F-EABR mRNA vaccine induces more robust immune responses, highlighting its potential in preventing RSV infection in vulnerable populations. IMPORTANCE: Respiratory syncytial virus (RSV) is a major cause of lower respiratory tract disease in infants, the elderly, and immunocompromised individuals. It is well-established that enhancing the neutralizing antibody levels and Th1-biased cellular immune responses can potentially improve the efficacy and safety of RSV vaccines. In this study, we developed an RSV pre-fusion protein-based mRNA vaccine that encodes self-assembling enveloped virus-like particles. Mice immunized with this vaccine showed significantly enhanced pre-fusion protein-targeted humoral responses and improved protection against RSV infection compared to the conventional RSV mRNA vaccine. Additionally, this vaccine demonstrated a considerably stronger neutralizing ability against contemporary clinical RSV isolates and induced more robust Th1-biased cellular immune responses, suggesting its potential as a promising RSV vaccine candidate.