Construction of an rAAV Producer Cell Line through Synthetic Biology,ACS Synthetic Biology

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Construction of an rAAV Producer Cell Line through Synthetic Biology
ACS Synthetic Biology ( IF 3.7 ) Pub Date : 2022-10-11 , DOI: 10.1021/acssynbio.2c00207
Zion Lee ₁ , Min Lu ₁ , Eesha Irfanullah ₁ , Morgan Soukup ₁ , Wei-Shou Hu ₁

Affiliation

Recombinant adeno-associated viruses (rAAV) are important gene delivery vehicles for gene therapy applications. Their production relies on plasmid transfection or virus infection of producer cells, which pose a challenge in process scale-up. Here, we describe a template for a transfection-free, helper virus-free rAAV producer cell line using a synthetic biology approach. Three modules were integrated into HEK293 cells including an rAAV genome and multiple inducible promoters controlling the expression of AAV Rep, Cap, and helper coding sequences. The synthetic cell line generated infectious rAAV vectors upon induction. Independent control over replication and packaging activities allowed for manipulation of the fraction of capsid particles containing viral genomes, affirming the feasibility of tuning gene expression profiles in a synthetic cell line for enhancing the quality of the viral vector produced. The synthetic biology approach for rAAV production presented in this study can be exploited for scalable biomanufacturing.

中文翻译：

通过合成生物学构建 rAAV 生产细胞系

重组腺相关病毒（rAAV）是基因治疗应用的重要基因递送载体。它们的生产依赖于生产细胞的质粒转染或病毒感染，这对工艺放大提出了挑战。在这里，我们描述了使用合成生物学方法的无转染、无辅助病毒的 rAAV 生产细胞系的模板。三个模块被整合到 HEK293 细胞中，包括 rAAV 基因组和控制 AAV Rep、Cap 和辅助编码序列表达的多个诱导型启动子。合成细胞系在诱导后产生感染性 rAAV 载体。对复制和包装活动的独立控制允许操纵含有病毒基因组的衣壳颗粒部分，证实了调整合成细胞系中的基因表达谱以提高所产生的病毒载体的质量的可行性。本研究中提出的 rAAV 生产的合成生物学方法可用于可扩展的生物制造。

更新日期：2022-10-11

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