Wang Liqun, Kaneko Shuichi, Honda Masao, Kobayashi Kenichi
Cancer Gene Regulation, Graduate School of Medical Sciences, Kanazawa University, Takara Machi 13-1 Kanazawa 920-8641, Japan.
Virus Res. 2002 May 10;85(2):187-97. doi: 10.1016/s0168-1702(02)00043-6.
The development of liver-directed virus vector may play a crucial role in hepatic gene therapy. Hepatitis B virus (HBV) is the only known DNA virus that has hepatocyte specificity. In order to construct an efficient HBV-based vector for targeting the liver, we studied the potential use of naturally occurring defective HBVs obtained from hepatitis patients. The enhanced green fluorescent protein (EGFP) gene or small tag sequences (Flag) were introduced in frame into the deleted sites of the defective HBVs. One HBV defective in site for putative T cell epitope and a part of the polymerase gene tolerated EGFP insertion and was successfully packaged. This defective recombinant HBV harboring 48 bp Flag tag sequence instead of EGFP (rHBV-7-Flag) replicated well. Human primary hepatocytes could uptake rHBV-7-Flag virions, though in a low frequency, when exposed to the virions at a high density in the culture medium, and also express Flag tag sequences. This defective HBV-based vector may have a potential application in liver targeting gene therapy.
肝靶向病毒载体的开发可能在肝脏基因治疗中发挥关键作用。乙型肝炎病毒（HBV）是唯一已知的具有肝细胞特异性的DNA病毒。为了构建一种高效的靶向肝脏的基于HBV的载体，我们研究了从肝炎患者中获得的天然存在的缺陷型HBV的潜在用途。将增强型绿色荧光蛋白（EGFP）基因或小标签序列（Flag）读框插入缺陷型HBV的缺失位点。一个在假定的T细胞表位和部分聚合酶基因位点有缺陷的HBV能够耐受EGFP插入并成功包装。这种携带48bp Flag标签序列而非EGFP的缺陷型重组HBV（rHBV-7-Flag）复制良好。当在培养基中以高密度暴露于人原代肝细胞时，人原代肝细胞能够摄取rHBV-7-Flag病毒粒子，尽管频率较低，并且还能表达Flag标签序列。这种基于缺陷型HBV的载体可能在肝靶向基因治疗中有潜在应用。
