Studies on Epstein-Barr virus vector for human gene therapy

eb病毒（Epstein-Barr virus， EBV）感染后，还没有像winch那样产生病毒的感染系统，因此不可能大量生产重组EBV用于基因治疗。在本研究中，我们旨在建立EBV扩增子系统。在病毒复制过程中，包含病毒复制起始点的DNA序列和包装信号被有效地扩增并包装到病毒颗粒中。我们获得了基因组大小超过200kbp的重组EBV感染细胞。如此大的EBV基因组超过了被包装成病毒颗粒的大小限制。因此，这种细胞应成为扩增子病毒繁殖的理想包装细胞。我们证实，携带大量EBV基因组的Akata细胞产生抒情抗原，但在抗lg抗体治疗后不产生子代病毒。在此基础上，我们制备了含有EBV和GFP基因复制起始和包装信号的质粒，并将其转染到携带大量EBV基因组的Akata细胞中。我们证实转染的质粒经抗lg处理后包装并生产为感染性病毒。高田博士和伊迈博士于1998年6月访问了汉默施密特博士的实验室。在访问期间，我们对彼此的实验结果进行了讨论，并相互获得了许多重要的信息。

There has been no infection system such as that winch produces viruses following Epstein-Barr virus (EBV infection, Therefore, it has not been possible to produce a large amount of recombinant EBV for gene therapy. In this study, we aimed to generate an amplicon system of EBV.DNA sequences containing the replication origin in a virus replicative phase and packaging signals are efficiently amplified and packaged into virus particles during virus replication. We generated cells infected with recombinant EBV with more than 200kbp genome size. Such large EBV genomes exceed the size limitation for being packaged into virus particles. Therefore, such cells should become an ideal packaging cells for propagating amplicon virus. We confirmed that Akata cells carrying huge EBV genome produce lyric antigen but do not produce progeny viruses upon treatment with anti-lg antibodies. Base on these results, we genierated a plasmid containing the replication origin and packaging signal of EBV and GFP gene, and transfected it into the Akata cells carrying huge EBV genome. We confirmed that the transfected plasmid is packaged and produced as infectious viruses after anti-lg treatment.Dr.Takada and Dr.lmai visited Dr.Hammerschmidt's laboratory in June, 1998. During its visit, we discussed about mutual experimental results, and obtained many important informations each other.