Puente-Massaguer Eduard, Andrade Thales Galdino, Scherm Michael J, Vasilev Kirill, Abozeid Hassanein, Rodriguez Alesandra J, Yueh Josh, Bhavsar Disha, Campbell John D, Yu Dong, Webby Richard J, Kawaoka Yoshihiro, Neumann Gabriele, Han Julianna, Ward Andrew B, Krammer Florian
Department of Microbiology, Icahn School of Medicine at Mount Sinai, New York, NY, USA.
Department of Integrative Structural and Computational Biology, The Scripps Research Institute, La Jolla, CA, USA.
bioRxiv. 2025 Aug 14:2025.08.14.670375. doi: 10.1101/2025.08.14.670375.
The continuous evolution and widespread dissemination of highly pathogenic avian influenza (HPAI) H5N1 viruses, particularly clade 2.3.4.4b, pose critical challenges to global pandemic preparedness. In this study, we assessed a low-dose inactivated split virus vaccine derived from clade 2.3.4.4b H5N1, formulated with an Alum/CpG adjuvant, using a preclinical mouse model. This vaccine induced potent humoral and cellular immune responses, generating high titers of cross-reactive antibodies targeting both hemagglutinin (HA) and neuraminidase (NA) glycoproteins across homologous and heterologous H5 clades. The Alum/CpG adjuvant enabled significant antigen dose-sparing while promoting a balanced Th1/Th2 immune profile. Functional analyses demonstrated strong virus neutralization, neuraminidase inhibition, and potent antibody-dependent cellular cytotoxicity activity. Additionally, the vaccine elicited robust antigen-specific CD4 and CD8 T cell responses and effectively controlled viral replication in the lungs, accompanied by reduced lung inflammation. Importantly, vaccinated mice were fully protected against lethal challenges with both the homologous clade 2.3.4.4b and heterologous clade 1 H5N1 viruses, despite low hemagglutination inhibition titers. Electron microscopy polyclonal epitope mapping revealed serum antibodies targeting multiple epitopes on homologous HA and NA, with some cross-reacting to conserved epitopes on heterologous proteins, underscoring broad immune recognition. Collectively, these results highlight the potential of this vaccine candidate to provide broad, multifunctional, and durable immunity against both current and emerging H5N1 threats, supporting its further development for pandemic preparedness.
高致病性禽流感（HPAI）H5N1病毒，尤其是2.3.4.4b分支病毒的持续进化和广泛传播，对全球大流行防范构成了严峻挑战。在本研究中，我们使用临床前小鼠模型评估了一种源自2.3.4.4b分支H5N1病毒的低剂量灭活裂解病毒疫苗，该疫苗采用铝盐/甲基化CpG寡脱氧核苷酸（Alum/CpG）佐剂配制。这种疫苗诱导了强效的体液免疫和细胞免疫反应，产生了高滴度的针对同源和异源H5分支血凝素（HA）和神经氨酸酶（NA）糖蛋白的交叉反应抗体。Alum/CpG佐剂在促进Th1/Th2免疫平衡的同时，实现了显著的抗原剂量节省。功能分析表明该疫苗具有强大的病毒中和、神经氨酸酶抑制以及有效的抗体依赖性细胞毒性活性。此外，该疫苗引发了强烈的抗原特异性CD4和CD8 T细胞反应，并有效控制了肺部的病毒复制，同时减轻了肺部炎症。重要的是，尽管血凝抑制滴度较低，但接种疫苗的小鼠对同源的2.3.4.4b分支和异源的1分支H5N1病毒的致死性攻击均具有完全保护作用。电子显微镜多克隆表位图谱显示，血清抗体靶向同源HA和NA上的多个表位，其中一些与异源蛋白上的保守表位发生交叉反应，突出了广泛的免疫识别。总体而言，这些结果凸显了这种候选疫苗针对当前和新出现的H5N1威胁提供广泛、多功能和持久免疫力的潜力，支持其为大流行防范进行进一步开发。
