携带mIL-12基因的增殖腺病毒在鼻咽癌细胞中增殖及mIL-12高效表达  

High-level mIL-12 expression and replication of replicating adenovirus carrying mIL12gene in naso-pharyngeal carcinoma cells

作　　者：聂飚[1] 钱其军[2] 车小燕[1] 薛惠斌[2] 岑信棠[3] 

机构地区：[1]第一军医大学珠江医院中心实验室,广东广州510282 [2]第二军医大学东方肝胆外科医院研究所,上海200438 [3]香港大学玛丽医院临床肿瘤系

出　　处：《第一军医大学学报》2002年第4期296-298,共3页Journal of First Military Medical University

基　　金：国家自然科学基金重大国际合作研究项目(30120160823);全军十五面上项目(01MA135);广东省自然科学基金(01063) ;国家十五"863"计划(2001AA217171)

摘　　要：目的联合选择性增殖腺病毒溶肿瘤细胞和小鼠白细胞介素12(mouse interleukin-12, mIL12)免疫治疗的作用,提高对肿瘤细胞的杀伤作用。方法用携带mIL12的选择性增殖腺病毒CNHK200-mIL12转染鼻咽癌细胞株CNE3和915,测定其在鼻咽癌细胞株中的增殖能力和mIL12的表达水平。结果CNHK200-mIL12转染鼻咽癌细胞后,免疫组化染色显示大部分细胞腺病毒六联体蛋白(hexon)阳性,用流式细胞仪检测,转染24 h 后CNE3和915细胞的阳性率分别比0 h增高39%和4%,转染72 h后,病毒增殖超过1 000倍,增殖能力同dl1520相似,并能高水平表达mIL12,在1×104个CNE3和915细胞中分别达到(84.5±4.6)和(75.6±3.4)ng/105空斑形成单位(plaque forming unit, PFU)。结论携带mIL12 的CNHK200在p53缺陷的鼻咽癌细胞中增殖,并且高效表达mIL12,为进一步研究体内外对肿瘤杀伤作用奠定基础。Objective To enhance the therapeutic effects on nasopharyngeal carcinoma by combining treatment with selectively replicating adenovirus and IL12. Methods Replicating adenovirus with mouse IL12 gene insert (CNHK200-mIL12) was constructed to transfect nasopharyngeal carcinoma cell lines CNE3 and 915. Adenovirus hexon was detected by immunohistochemical staining and flow cytometry (FCM), and mIL12 expression examined by enzyme-linked immunosorbent assay (ELISA). The replication rates of CNHK200-mIL12 and dl1520 was determined by 50% tissue culture infectious dose (TCID 50 ). Results Twenty-four hours after transfection with CNHK200-mIL12, most of cells were positive for adenovirus hexon and FCM demonstrated increased positivity rates of 39% and 4% among CNE3 and 915 cells respectively. It was observed that CNHK200-mIL12 replication increased by 1 000 folds with mIL12 expression level reaching as high as 84.5±4.6 ng in CNE3 cells and 75.6±3.4 ng in 915 cells as determined 72 h after transfection with 1×10 5 PFU CNHK200-mIL12 into 1×10 4 cells. Conclusion CNHK200-mIL12 can replicate in vitro in nasopharyngeal carcinoma cells (dl1520 cells, for instance) with high mIL12 expression, which suggests that CNHK200-mIL12 may potentially be used to treat nasopharyngeal carcinoma.

关 键 词：增殖腺病毒 白细胞介素12 鼻咽癌 基因治疗 

分 类 号：R739.63[医药卫生—肿瘤]