| 摘要: | 季節性流感病毒對全球健康構成重大威脅，迫切需要有效的預防策略。疫苗接種仍然是預
防的基石，但當前的方法主要依賴於使用胚胎雞蛋（ECE）生產的滅活裂解病毒疫苗，此
方法存在各種缺點，包括過敏反應、血凝素（HA）基因的適應性突變、蛋供應中斷以及
生產放大問題。為了克服對 ECE的依賴，極需利用高效和穩定生產平台來開發流感疫苗。
已經有幾個疫苗生產系統顯示出作為替代品的潛力，包括類病毒顆粒（VLP）平台。在這
項研究中，我們使用桿狀病毒表達系統（BES），選殖流感 A/H1N1、A/H3N2、
B/Yamagata-like 和 B/Victoria-like 病 毒 株 （ A/Hawaii/70/2019, A/Minnesota/41/2019,
B/Brisbane/09/2014 & B/Brisbane/63/2014）的 HA、NA 和 M1 基因，以生產 VLP 疫苗抗原
H1N1-VLP、H3N2-VLP、Yamagata-VLP 和 Victoria-VLP，因此四種 homologus VLP 的 HA
抗原產量差距四倍以上。然後，我們開發了嵌合 VLP，以提高 HA 蛋白的產量。我們評估
了它們的功能和抗原特性，包括凝血試驗、蛋白組成、形態學、穩定性和免疫原性。我們
發現，同源和嵌合 VLP 的功能活性`形態和大小上類似於流感病毒顆粒，同時保持結構完
整性。小鼠的免疫原性比較評估顯示，我們的四價 VLP在誘導凝血抑制抗體、中和抗體與
上市的雞胚蛋及重組蛋白疫苗相比具有優越性，並對同源病毒提供 100%的保護。這些發
現表明，昆蟲細胞產出的 VLP疫苗作為四價季節性流感疫苗的候選者具有良好的商業潛力
，值得進一步開發。
;Seasonal influenza viruses pose a significant threat to global health, necessitating effective
prevention strategies. While vaccination remains the cornerstone, current methods primarily rely
on inactivated split virus vaccines generated using embryonated chicken eggs (ECE), present
various drawbacks including allergic reactions, adaptive mutation in hemagglutinin (HA) gene,
disruptions in egg supply, and scalability issues. To overcome the dependency on ECE,
developments of influenza vaccines based on efficient and robust production platforms are urgently
needed. Several potential vaccine production systems have shown potential as replacers, including
a virus-like particle (VLP) platform. In this study, using a Baculovirus Expression System (BES),
we engineered HA, NA, and M1 genes of influenza A/H1N1, A/H3N2, B/Yamagata-like, and
B/Victoria-like virus strains (A/Hawaii/70/2019, A/Minnesota/41/2019, B/Brisbane/09/2014 &
B/Brisbane/63/2014) to produce VLP vaccine antigens H1N1-VLP, H3N2-VLP, Yamagata-VLP,
Victoria-VLP, respectively. We then assessed their functional and antigenic characteristics,
including hemagglutination assay, protein composition, HA protein productivity, VLPs
morphology, stability and immunogenicity. We found that recombinant VLPs displayed functional
activity, resembling influenza virions in morphology and size, while maintaining structural
integrity. Comparative immunogenicity assessments in mice showed that our quadrivalent VLPs
were consistent in inducing hemagglutination inhibition (HAI) and neutralizing (NT) antibody
titers against homologous viruses compared to both commercial recombinant HA (Flublok) and
egg-based vaccines (Vaxigrip). Since the HA antigen yields of the homologous VLPs varied more
than four folds, we further developed chimeric VLPs to improve HA protein production and
address the limitations of the homologous VLPs. We assessed their antigenic characteristics,
comparative HA protein productivity, immunogenicity and vaccine efficacy. We found that similar
to homologous VLPs, chimeric VLPs displayed functional activity, resembling influenza virions
in morphology and size. Comparative immunogenicity assessments in mice showed that the VLP
vaccine consistent in inducing higher hemagglutination inhibition and neutralizing antibody titers
and provided 100% protection against homologous viruses compared to the commercial vaccine.
The findings highlight insect cell-based VLP vaccines as promising candidates for quadrivalent
seasonal influenza vaccines. |
