钱其军　车小燕　岑信棠　崔贞福　薛惠斌　朱斌　徐建国　Billy 　吴孟超

【摘要】　
目的　
研究一种结合肿瘤基因治疗与病毒治疗优势的新型肿瘤治疗载体系统,即基因2 病毒载体系统。方法　利用病毒重组技术将抗癌基因插入肿瘤细胞特异性的增殖病毒的病毒基因组中,通过细胞病理作用、荧光显微镜、免疫酶链技术及电镜等技术分别观察病毒的杀伤效应、报告基因绿色荧光蛋白、抗癌基因小鼠白细胞介素12 表达量及病毒复制情况。
结果　
构建了一种新型基因2病毒载体系统,该载体系统腺病毒E1b 55 000 蛋白缺失, 保留了腺病毒E1a 蛋白。该载体系统具有肿瘤增殖病毒ONYX2015 的相似功能, 即它可在肿瘤细胞内复制及增殖,而在正常细胞内不能复制及增殖, 从而特异性杀灭肿瘤细胞。该载体系统还可携带各种抗癌基因以进一步提高抗肿瘤的疗效。应用该载体系统携带该报告基因绿色荧光蛋白可使绿色荧光蛋白在肿瘤细胞内高效表达, 其表达量明显高于传统基因治疗的腺病毒载体系,而在正常细胞内低表达, 表达量与传统腺病毒载体系统相似或更低。应用该载体系统携带抗癌基因小鼠白细胞介素12 ,也产生类似结果。电镜也证实该载体系统携带抗癌基因小鼠白细胞介素12 可在肿瘤细胞株中复制及增殖。
结论　
基因2病毒载体将抗癌基因插入肿瘤增殖病毒基因组,可数百倍乃至上万倍提高抗癌基因的表达量,进一步提高抗肿瘤的疗效。

【关键词】　基因,病毒; 　载体蛋白质类; 　肿瘤; 　基因疗法

Development of gene2viral vector and its anti2tumor effect , a primary study

【Abstract 】　
Objective 　To develop a new kind of vector system , named as gene2viral vector , which combines the advantages of the gene therapy and virus therapy. Method 　An anti2tumor gene was inserted into the genome of the replicative virus specific for the tumor cells by virus recombination technology. The killing effect , report gene expression of the green fluorescence protein、expression of the anti2tumor gene of mouse IL12 , and the replication of the virus were observed respectively by cell pathology , fluorescence microscopy , ELISA and electron microscopy. Results 　A new kind of gene2viral vector system , in which the E1b255 000 gene is deleted but the E1a gene of adenovirus is preserved , was constructed. The vector system possessed the same characteristics as the replicative virus ONYX2015 , replication and proliferation in the tumor cells but not in the normal cells , thus specifically killing the tumor cells. Besides , it carried many kinds of anti2tumor genes. When carrying the report gene of the green fluorescence protein it made the expression of this gene in tumor cells far more effectively than the adenovirus vector employed in the traditional gene therapy did. However in the normal cells the expression of green fluorescence protein caused by this vector system was as little as or even less than that by the traditional adenovirus system. The similar result was also observed in the experiments of this vector system carrying the anti2 tumor gene , gene of mouse IL12. The replication and proliferation of the virus carrying the gene of mouse IL12 in the tumor cells were confirmed by electron microscopy. Conclusion 　Gene2viral vector is a new kind of vector in which the anti2tumor gene is inserted into the genome of the replicative virus specific for the tumor cells. It increases the expression of the anti2tumoe gene by hundreds even tens of thousand times. It posseses all the advantages of gene therapy and virus therapy , thus further enhancing the curative effect and it overcomes such disadvantages as low transfer rate , low expression , lack of target tropism and low anti2tumor activity. It will become one of the most promising means in tumor treatment.
【Key words】　Genes ,viral ; 　Carrier proteins ; 　Neoplasms ; 　Gene therapy