A nucleoside-modified mRNA vaccine forming rabies virus-like particle elicits strong cellular and humoral immune responses against rabies virus infection in mice

Rabies is a lethal zoonotic disease that threatens human health. As the only viral surface protein, the rabies virus (RABV) glycoprotein (G) induces main neutralizing antibody (Nab) responses; however, Nab titre is closely correlated with the conformation of G. Virus-like particles (VLP) formed by the co-expression of RABV G and matrix protein (M) improve retention and antigen presentation, inducing broad, durable immune responses. RABV nucleoprotein (N) can elicit humoral and cellular immune responses. Hence, we developed a series of nucleoside-modified RABV mRNA vaccines encoding wild-type G, soluble trimeric RABV G formed by an artificial trimer motif (tG-MTQ), membrane-anchored prefusion-stabilized G (preG). Furthermore, we also developed RABV VLP mRNA vaccine co-expressing preG and M to generate VLPs, and VLP/N mRNA vaccine co-expressing preG, M, and N. The RABV mRNA vaccines induced higher humoral and cellular responses than inactivated rabies vaccine, and completely protected mice against intracerebral challenge. Additionally, the IgG and Nab titres in RABV preG, VLP and VLP/N mRNA groups were significantly higher than those in G and tG-MTQ groups. A single administration of VLP or VLP/N mRNA vaccines elicited protective Nab responses, the Nab titres were significantly higher than that in inactivated rabies vaccine group at day 7. Moreover, RABV VLP and VLP/N mRNA vaccines showed superior capacities to elicit potent germinal centre, long-lived plasma cell and memory B cell responses, which linked to high titre and durable Nab responses. In summary, our data demonstrated that RABV VLP and VLP/N mRNA vaccines could be promising candidates against rabies.

狂犬病是一种威胁人类健康的致死性人畜共患病。狂犬病病毒（Rabies Virus, RABV）糖蛋白（G蛋白）作为唯一的病毒表面蛋白，可诱导主要的中和抗体（Neutralizing Antibody, Nab）应答；然而中和抗体滴度与G蛋白的构象密切相关。由RABV G蛋白与基质蛋白（M蛋白）共表达形成的病毒样颗粒（Virus-like Particle, VLP）可提升抗原保留与抗原呈递效率，诱导广谱且持久的免疫应答。RABV核蛋白（N蛋白）可激发体液免疫与细胞免疫应答。据此，我们开发了一系列核苷修饰的RABV mRNA疫苗，分别编码野生型G蛋白、经人工三聚体基序构建的可溶性三聚体RABV G蛋白（tG-MTQ），以及膜锚定的预融合稳定型G蛋白（preG）。此外，我们还开发了共表达preG与M蛋白以生成VLP的RABV VLP mRNA疫苗，以及共表达preG、M与N蛋白的VLP/N mRNA疫苗。相较于灭活狂犬病疫苗，本研究开发的RABV mRNA疫苗可诱导更强的体液与细胞免疫应答，并可完全保护小鼠抵御脑内攻毒。此外，preG、VLP及VLP/N mRNA疫苗组的IgG与中和抗体滴度均显著高于G蛋白组与tG-MTQ组。单次接种VLP或VLP/N mRNA疫苗即可诱导保护性中和抗体应答，在接种后第7天，其抗体滴度显著高于灭活狂犬病疫苗组。进一步研究发现，RABV VLP与VLP/N mRNA疫苗可更高效地诱导强效的生发中心、长寿浆细胞与记忆B细胞应答，这与高滴度且持久的中和抗体应答密切相关。综上，本研究数据表明，RABV VLP与VLP/N mRNA疫苗有望成为抗狂犬病的候选疫苗。