The emergence of XBB- and JN.1-lineages with remarkable immune evasion characteristics have led to rises in breakthrough infections within populations. In addition, the unfavorable impacts of immune imprinting, stemming from continuous exposure to antigens from circulated viruses, have been observed to incline immune response against earlier lineages, thereby declining the neutralization to newly emerged Omicron subvariants. In response to this, the advancement of next-generation vaccines against COVID-19 targeting components from new subvariants such as XBB-lineage is imperative. In the current study, a self-assembled trimeric recombinant protein (RBDXBB.1.5-HR) was generated by concatenating the sequences of the receptor binding domain (RBD) derived from XBB.1.5 with heptad-repeat 1 (HR1) and HR2 sequences from the spike S2 subunit. Adjuvanted-RBDXBB.1.5-HR induced robust humoral and cellular immune responses, characterized by elevated neutralization against JN.1-inculuded subvariants and a substantial population of antigen-specific T memory cells. Protective immunity conferred by RBDXBB.1.5-HR vaccine was preserved post-immunization, as evidenced by germinal center B (GC B) and T follicular helper (Tfh) responses, sustained neutralization potency, and an increase in memory B cells (MBCs) and long-lived plasma cells (LLPCs). The RBDXBB.1.5-HR vaccine showed a favorable boosting effect when administered heterologously after three doses of inactivated virus (IV) and mRNA vaccines. Significantly, it provided protection against live Omicron EG.5.1 viruses in vivo. The monovalent RBDXBB.1.5-HR vaccine showed favorable safety and immunogenicity, boosting neutralizing antibodies against JN.1- and XBB-lineage subvariants in individuals with prior COVID-19 vaccinations. These findings highlight its clinical potential in safeguarding against circulating Omicron subvariants.© 2025. The Author(s).