Xikui Sun, Junhao Fan, Xiaolu Xie, Yuting Shi, Xiaoqing Liu, Tiefeng Xu, Sidi Yang, Keda Shi, Chengshu Xie, Shixiong Li, Siwen Chen, Ruoxi Cai, Ping He, Yuzhen Ye, Tao Chen, Xiancai Ma, Chunmei Li, Jingyou Yu, Deyin Guo
The rapid spread of immune-evasive viral variants, exemplified by SARS-CoV-2, highlights the urgent need for broad-spectrum coronavirus vaccines. Although multimeric display of the receptor-binding domain (RBD) using exogenous scaffolds improve immune responses, such approaches may elicit off-target immune responses against the scaffolds themselves and remain limited by rapid RBD antigenic drift. Here, we report the rational design of a self-assembling trimeric subunit vaccine, termed RBD-heptad repeat 1 (HR1)-stem helix (SH)-heptad repeat 2 (HR2) (RHS), which integrates the JN.1 RBD with a highly conserved SH epitope from SARS-CoV-2 within a native HR1-HR2 trimeric scaffold via optimized linkers. RHS exhibits high structural stability, efficient trimerization, and enhanced antigen presentation. In mice, RHS elicits robust humoral and cellular immune responses, including potent cross- neutralizing antibodies against diverse SARS-CoV-2 variants and pan-betacoronavirus SH-specific antibodies. In K18-hACE2 mice, RHS confers strong protection against both antigen-matched and antigen-mismatched Omicron variants, while intranasal immunization induces potent mucosal IgA and IgG responses. Furthermore, the modular RHS platform is readily adaptable to antigens from SARS-CoV and MERS-CoV, underscoring its versatility for pan-coronavirus vaccine development. Together, these findings establish RHS as a generalizable strategy for overcoming viral immune evasion and advancing next-generation vaccine design.