Rosato A, Zambon A, Milan G, Ciminale V, D'Agostino D M, Macino B, Zanovello P, Collavo D
Division of Immunology, Department of Oncology and Surgical Sciences, University of Padova, Italy.
Hum Gene Ther. 1997 Aug 10;8(12):1451-8. doi: 10.1089/hum.1997.8.12-1451.
A DNA immunization approach was used to induce an immune response against the tumor-specific antigen P815A in DBA/2 mice. The P1A gene, which encodes the P815A antigen, was modified by the addition of a short sequence coding for a tag epitope recognized by the monoclonal antibody AU1, and cloned into the eukaryotic expression vector pBKCMV, resulting in plasmid pBKCMV-P1A. L1210 cells stably transfected with pBKCMV-P1A expressed P1A mRNA and were lysed by the syngeneic P815A-specific cytotoxic clone CTL-P1:5, thus confirming that the tag-modified P1A protein underwent correct processing and presentation. A single intramuscular injection of 100 microg of pBKCMV-P1A induced the expression of P1A mRNA for at least 4 months. Eighty percent of DBA/2 mice injected three times with 100 microg of pBKCMV-P1A generated cytotoxic T lymphocytes (CTL) that lysed P815 tumor cells, whereas mock-inoculated animals failed to show any cytotoxicity. Moreover, experiments designed to evaluate the protection of pBKCMV-P1A-immunized mice against a lethal challenge with P815 tumor cells showed that 6 of 10 immunized mice rejected the tumor, and 2 mice showed prolonged survival compared to control animals.
采用DNA免疫方法在DBA/2小鼠中诱导针对肿瘤特异性抗原P815A的免疫反应。编码P815A抗原的P1A基因通过添加一段编码单克隆抗体AU1识别的标签表位的短序列进行修饰，并克隆到真核表达载体pBKCMV中，得到质粒pBKCMV-P1A。用pBKCMV-P1A稳定转染的L1210细胞表达P1A mRNA，并被同基因的P815A特异性细胞毒性克隆CTL-P1:5裂解，从而证实标签修饰的P1A蛋白经过了正确的加工和呈递。单次肌内注射100μg pBKCMV-P1A可诱导P1A mRNA表达至少4个月。80%接受三次100μg pBKCMV-P1A注射的DBA/2小鼠产生了可裂解P815肿瘤细胞的细胞毒性T淋巴细胞（CTL），而模拟接种的动物未表现出任何细胞毒性。此外，旨在评估用pBKCMV-P1A免疫的小鼠对P815肿瘤细胞致死性攻击的保护作用的实验表明，10只免疫小鼠中有6只排斥了肿瘤，2只小鼠与对照动物相比存活时间延长。
