Wang Lizheng, Wang Zixuan, Zhang Fangfang, Zhu Rui, Bi Jinpeng, Wu Jiaxin, Zhang Haihong, Wu Hui, Kong Wei, Yu Bin, Yu Xianghui
1. National Engineering Laboratory for AIDS Vaccine, School of Life Sciences, Jilin University, Changchun 130012, China.
1. National Engineering Laboratory for AIDS Vaccine, School of Life Sciences, Jilin University, Changchun 130012, China; 2. Key Laboratory for Molecular Enzymology and Engineering, the Ministry of Education, School of Life Sciences, Jilin University, Changchun 130012, China.
Int J Med Sci. 2016 Apr 1;13(4):286-91. doi: 10.7150/ijms.14152. eCollection 2016.
Adeno-associated virus (AAV) vectors have been utilized extensively in gene therapy and gene function studies, as strong transgene expression is a prerequisite for positive outcomes. AAV8 was reported as the most efficient AAV serotype for transduction of the liver, brain and muscle compared with other serotypes. However, AAV8-mediated transduction of human hepatocytes is rather poor with approximately 20-fold lower efficiency compared with that of mouse hepatocytes. Therefore, we applied the woodchuck hepatitis virus post-transcriptional regulatory element (WPRE) to enhance AAV8-mediated transgene expression driven by a combination promoter (CAG promoter) with a CMV-IE enhancer and chicken beta-actin promoter for a more efficient viral vector. Transgene expression from recombinant AAV8 (rAAV8) vectors harboring a red fluorescent protein (RFP) reporter gene with or without WPRE were evaluated in vitro and in vivo. The results demonstrated that WPRE improved AAV8-mediated RFP expression in different cell lines with clear increases of transgene expression in the liver, brain or muscle of animals. The findings of this study will help to substantially reduce the quantity of viral particles that must be injected in order to reach a therapeutic level of transgene expression in gene therapy. Consequently, such dose reductions may lessen the potential risks associated with high doses of viral vectors.
腺相关病毒（AAV）载体已广泛应用于基因治疗和基因功能研究，因为强大的转基因表达是取得积极成果的先决条件。与其他血清型相比，AAV8被报道为用于肝脏、大脑和肌肉转导的最有效的AAV血清型。然而，AAV8介导的人类肝细胞转导相当差，与小鼠肝细胞相比效率低约20倍。因此，我们应用土拨鼠肝炎病毒转录后调控元件（WPRE）来增强由具有CMV-IE增强子和鸡β-肌动蛋白启动子的组合启动子（CAG启动子）驱动的AAV8介导的转基因表达，以构建更有效的病毒载体。在体外和体内评估了携带红色荧光蛋白（RFP）报告基因且有或无WPRE的重组AAV8（rAAV8）载体的转基因表达。结果表明，WPRE改善了AAV8介导的不同细胞系中的RFP表达，动物肝脏、大脑或肌肉中的转基因表达明显增加。本研究结果将有助于大幅减少为达到基因治疗中转基因表达的治疗水平而必须注射的病毒颗粒数量。因此，这种剂量的减少可能会降低与高剂量病毒载体相关的潜在风险。
