The mucosal immune response plays a crucial role in the prevention of respiratory viruses. Given the risk of recurrent SARS-CoV-2 infections in the population, the rapid development of next-generation intranasal COVID-19 vaccines with high safety and efficacy is paramount. In the current study, we developed a protein-based intranasal vaccine comprising the XBB.1.5 receptor binding domain (RBD)-derived trimeric recombinant protein (RBD_XBB.1.5-HR) and an MF59-like oil-in-water adjuvant. Intranasal administration of RBD_XBB.1.5-HR vaccine elicited robust and sustained humoral immune responses in mice and rats, resulting in high levels of neutralizing antibodies against XBB-lineage subvariants, with protection lasting for at least six months. The intranasal RBD_XBB.1.5-HR vaccine generated potent mucosal immune responses, characterized by the inductions of tissue-resident T (T_RM) cells, local cellular immunity, germinal center, and memory B cell responses in the respiratory tract. The combination of intramuscular and intranasal delivery of the RBD_XBB.1.5-HR vaccine demonstrated exceptional systemic and mucosal protective immunity. Furthermore, intranasal delivery of RBD_XBB.1.5-HR vaccine as a heterologous booster shot showed more effective boosting effects after mRNA administration compared to homologous vaccination, as evidenced by the induction of superior systemic and extra mucosal immune response. Importantly, the intranasal RBD_XBB.1.5-HR vaccine conferred efficient protection against the challenge with authentic EG.5.1 viruses in vivo. These findings identify the intranasal RBD_XBB.1.5-HR vaccine as a potential mucosal vaccine candidate for the prevention of SARS-CoV-2 infection.

粘膜免疫反应在预防呼吸道病毒中起着至关重要的作用。鉴于人群中反复感染 SARS-CoV-2 的风险，快速开发具有高安全性和有效性的下一代鼻内 COVID-19 疫苗至关重要。在目前的研究中，我们开发了一种基于蛋白质的鼻内疫苗，包括 XBB.1.5 受体结合域 （RBD） 衍生的三聚体重组蛋白 （RBD_XBB.1.5-HR） 和一种 MF59 样水包油佐剂。RBD_XBB.1.5-HR 疫苗的鼻内给药在小鼠和大鼠中引起了强烈而持续的体液免疫反应，导致针对 XBB 谱系亚变体的高水平中和抗体，保护持续至少六个月。鼻内 RBD_XBB.1.5-HR 疫苗产生有效的粘膜免疫反应，其特征是组织驻留 T （T_RM） 细胞的诱导、局部细胞免疫、生发中心和呼吸道记忆 B 细胞反应。RBD_XBB.1.5-HR 疫苗的肌内和鼻内给药相结合表现出卓越的全身和粘膜保护性免疫。此外，与同源疫苗接种相比，作为异源加强针的鼻内递送 RBD_XBB.1.5-HR 疫苗在 mRNA 给药后显示出更有效的增强效果，诱导优越的全身和粘膜外免疫反应证明了这一点。重要的是，鼻内 RBD_XBB.1.5-HR 疫苗在体内有效保护了真正的 EG.5.1 病毒的攻击。这些发现确定鼻内 RBD_XBB.1.5-HR 疫苗是预防 SARS-CoV-2 感染的潜在粘膜疫苗候选者。