Centre for Heart Research, The Westmead Institute for Medical Research, Westmead 2145, Australia.
Sydney Medical School, Faculty of Medicine and Health, The University of Sydney, Sydney 2006, Australia.
Viruses. 2022 Jul 26;14(8):1623. doi: 10.3390/v14081623.
Gene therapy is making significant impact on a modest, yet growing, number of human diseases. Justifiably, the preferred viral vector for clinical use is that based on recombinant adeno-associated virus (rAAV). There is a need to scale up rAAV vector production with the transition from pre-clinical models to human application. Standard production methods based on the adherent cell type (HEK293) are limited in scalability and other methods, such as those based on the baculovirus and non-adherent insect cell (Sf9) system, have been pursued as an alternative to increase rAAV production. In this study, we compare these two production methods for cardiotropic rAAVs. Transduction efficiency for both production methods was assessed in primary cardiomyocytes, human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs), and in mice following systemic delivery. We found that the rAAV produced by the traditional HEK293 method out-performed vector produced using the baculovirus/Sf9 system in vitro and in vivo. This finding provides a potential caveat for vector function when using the baculovirus/Sf9 production system and underscores the need for thorough assessment of vector performance when using diverse rAAV production methods.
基因治疗正在为数以千计的人类疾病带来重大影响。有理由认为，用于临床应用的首选病毒载体是基于重组腺相关病毒（rAAV）的。随着从临床前模型向人类应用的转变，需要扩大 rAAV 载体的生产规模。基于贴壁细胞类型（HEK293）的标准生产方法在可扩展性方面存在限制，因此已经探索了其他方法，如基于杆状病毒和非贴壁昆虫细胞（Sf9）系统的方法，以增加 rAAV 的产量。在这项研究中，我们比较了这两种用于心脏靶向 rAAV 的生产方法。在原代心肌细胞、人诱导多能干细胞衍生的心肌细胞（hiPSC-CMs）以及系统给药后的小鼠中评估了这两种生产方法的转导效率。我们发现，传统的 HEK293 方法生产的 rAAV 在体外和体内均优于杆状病毒/Sf9 系统生产的载体。这一发现为使用杆状病毒/Sf9 生产系统时的载体功能提供了一个潜在的警示，并强调了在使用不同的 rAAV 生产方法时需要对载体性能进行彻底评估。
