Influenza vaccine manufacturers traditionally use egg-derived candidate vaccine viruses (CVVs) to produce high-yield influenza viruses for seasonal or pandemic vaccines; however, these egg-derived CVVs need an adaptation process for the virus to grow in mammalian cells. The low yields of cell-based manufacturing systems using egg-derived CVVs remain an unsolved issue. This study aimed to develop high-growth cell-derived CVVs for MDCK cell-based vaccine manufacturing platforms. Four H7N9 CVVs were generated in characterized Vero and adherent MDCK (aMDCK) cells. Furthermore, reassortant viruses were amplified in adherent MDCK (aMDCK) cells with certification, and their growth characteristics were detected in aMDCK cells and new suspension MDCK (sMDCK) cells. Finally, the plaque-forming ability, biosafety, and immunogenicity of H7N9 reassortant viruses were evaluated. The HA titers of these CVVs produced in proprietary suspension MDCK (sMDCK) cells and chicken embryos were 2- to 8-fold higher than those in aMDCK cells. All H7N9 CVVs showed attenuated characteristics by trypsin-dependent plaque assay and chicken embryo lethality test. The alum-adjuvanted NHRI-RG5 (derived from the fifth wave H7N9 virus A/Guangdong/SP440/2017) vaccine had the highest immunogenicity and cross-reactivity among the four H7N9 CVVs. Finally, we found that AddaVax adjuvant improved the cross-reactivity of low pathogenic H7N9 virus against highly pathogenic H7N9 viruses. Our study indicates that cell-derived H7N9 CVVs possessed high growth rate in new sMDCK cells and low pathogenicity in chicken embryo, and that CVVs generated by this platform are also suitable for both cell- and egg-based prepandemic vaccine production.

中文翻译：

---

悬浮MDCK细胞中高生长性流感H7N9大流行前候选疫苗病毒的开发。

传统上，流感疫苗制造商使用鸡蛋衍生的候选疫苗病毒（CVV）生产季节性或大流行疫苗的高产流感病毒；但是，这些鸡蛋衍生的CVV需要适应过程才能使病毒在哺乳动物细胞中生长。使用鸡蛋衍生的CVV的基于细胞的生产系统的低产量仍然是一个尚未解决的问题。这项研究旨在为基于MDCK细胞的疫苗生产平台开发高生长细胞来源的CVV。在特征化的Vero和贴壁MDCK（aMDCK）细胞中产生了四个H7N9 CVV。此外，经认证的重配病毒在贴壁MDCK（aMDCK）细胞中扩增，并在aMDCK细胞和新的悬浮MDCK（sMDCK）细胞中检测到它们的生长特征。最后，噬菌斑形成能力，生物安全性，评估了H7N9重配病毒的免疫原性。在专有的悬浮MDCK（sMDCK）细胞和鸡胚中产生的这些CVV的HA滴度比aMDCK细胞中的HA滴度高2至8倍。通过胰蛋白酶依赖性噬斑测定和鸡胚致死率测试，所有H7N9 CVVs均显示出减弱的特征。佐剂明矾的NHRI-RG5（源自第五波H7N9病毒A / Guangdong / SP440 / 2017）疫苗在四个H7N9 CVV中具有最高的免疫原性和交叉反应性。最后，我们发现AddaVax佐剂改善了低致病性H7N9病毒与高致病性H7N9病毒的交叉反应性。我们的研究表明，源自细胞的H7N9 CVV在新的sMDCK细胞中具有高生长速率，在鸡胚中具有低致病性，