Zhang Lumin, Daikoku Tohru, Ohtake Koichiro, Ohtsuka Junpei, Nawa Akihiro, Kudoh Ayumi, Iwahori Satoko, Isomura Hiroki, Nishiyama Yukihiro, Tsurumi Tatsuya
Division of Virology, Aichi Cancer Center Research Institute, 1-1, Kanokoden, Chikusa-ku, Nagoya 464-8681, Japan.
J Virol Methods. 2006 Nov;137(2):177-83. doi: 10.1016/j.jviromet.2006.06.017. Epub 2006 Jul 18.
Herpes simplex virus type 1 (HSV-1)-based amplicon vectors have been used widely in genetic engineering with many advantages for gene delivery, being easily constructed. An attenuated and replication-competent HSV-1 HF10 clone demonstrating an oncolytic effect on cancer cells in vitro and in vivo has been applied recently for clinical virotherapy of breast cancers and the present studies were conducted to test its efficacy in combination with an HSV-1 amplicon. For this purpose, a new system was developed to produce high titers of the HSV-1 amplicon vector and the results showed that its package efficiency and the titer ratio to HF10 were improved by passage through two cell lines. A high ratio of amplicon/helper virus HF10 (A/H) (>1) was required to express the foreign gene efficiently. Furthermore, in order to express the foreign gene conditionally, an HSV-1 ICP8 promoter was introduced in place of the human cytomegalovirus MIE promoter, this driving expression of the transgene when replication of HF10 progressed. The methodology for simple preparation of mixtures of viruses containing the amplicon with the oncolytic virus is documented. This system should find application for studies of cancer therapy.
基于1型单纯疱疹病毒（HSV-1）的扩增子载体已在基因工程中广泛应用，具有许多基因递送优势，且易于构建。一种减毒且具有复制能力的HSV-1 HF10克隆在体外和体内对癌细胞均显示出溶瘤作用，最近已应用于乳腺癌的临床病毒治疗，本研究旨在测试其与HSV-1扩增子联合使用的疗效。为此，开发了一种新系统来生产高滴度的HSV-1扩增子载体，结果表明，通过两种细胞系传代后，其包装效率和与HF10的滴度比均有所提高。高效表达外源基因需要高比例的扩增子/辅助病毒HF10（A/H）（>1）。此外，为了条件性表达外源基因，引入了HSV-1 ICP8启动子以取代人巨细胞病毒MIE启动子，当HF10复制进行时，该启动子驱动转基因表达。本文记录了简单制备含有扩增子与溶瘤病毒的病毒混合物的方法。该系统应可用于癌症治疗研究。
