Optimizing Rabies mRNA Vaccine Efficacy: Role of RABV-G Structural Domains and Heterologous Prime-Boost Immunization Strategies
Authors: Li Dongdong , Wang Xuan , Li Gaotian , Zhou Jingying , Bian Lijun , Zhao Xiaoyan , Xing Liao , Zeng Juanmei , Cui Jiaxing , Cui Lili , Zhang Yong , Chen Yan 
Published: November 2024

Background: Rabies remains a critical global public health issue, highlighting the need for effective and affordable vaccines. mRNA technology offers a promising solution due to its advantages in safety, immunogenicity, and production efficiency. This study investigates the impact of different structural domains of rabies virus glycoprotein (RABV-G) on mRNA vaccine efficacy and explores the potential of heterologous prime-boost strategies combining RABV mRNA vaccines and inactivated rabies virus (IRV) for enhanced immune responses and protection.
Methods: mRNA vaccines encoding full-length RABV-G (RABV-Full), ectodomain/transmembrane domain (RABV-TE), ectodomain (RABV-E), and a mutant variant (RABV-R333Q) were encapsulated in lipid nanoparticles (LNPs) and administered to BALB/c mice, respectively. Immunogenicity and protective efficacy were evaluated in terms of binding antibody titers, virus-neutralizing titers (VNTs), cell-mediated responses, and protection against lethal rabies virus challenge. Additionally, heterologous prime-boost strategies were assessed to determine optimal immunization regimens. Transcriptome analysis was conducted on lymph nodes to evaluate the impact of mRNA vaccines and IRV on the immune microenvironment.
Results: mRNA vaccines encoding full-length RABV-G, particularly with the R333Q mutation, induced strong humoral and cellular immune responses, achieving full protection against lethal challenge. RABV-TE showed reduced immune responses and protection compared to RABV-Full, while RABV-E induced binding antibodies but lacked neutralizing and cellular responses. Furthermore, we found that although mRNA vaccine produced high levels of VNTs after the second immunization, the short-term VNTs were lower than that of inactivated rabies vaccine (IRV). Therefore, a heterologous prime-boost immunization that combined mRNA vaccine and IRV was performed. The results showed that mRNA-primed strategies (RABV-Full>IRV and 2´RABV-Full) yielded higher long-lasting VNTs, but lower early VNTs than IRV-primed strategies (IRV>RABV-Full and 2´IRV) or a mixed formulation (1:1). It was worth noting that IRV>RABV-Full or mixed formulation yielded higher long-lasting VNTs and better protection than 2×IRV, although heterologous immunizations resulted in reduced cellular immune response. Transcriptome analysis showed that IRV primarily induced MHCII antigen presentation and B cell activation, while rabies mRNA vaccine induced both MHCI and MHCII antigen presentation, as well as B/T cell activation, highlighting its stronger immune-activating properties at the lymph node.
Conclusion: Full-length RABV-G mRNA vaccines, especially with the R333Q mutation, hold significant potential for rabies prevention. While the cytoplasmic domain of RABV-G may not be essential for neutralizing antibody production, it enhances vaccine immunogenicity and protection efficacy. The 'IRV prime and RABV-Full boost' regimen shows promise as an effective immunization strategy. These findings underscore the promise of mRNA vaccines as a cost-effective and potent strategy for rabies control.