携带IL-24的溶瘤腺病毒对裸鼠乳腺癌移植瘤的抑制作用

An in vivo study on the effect of oncolytic adenovirus CNHK600-IL24 on breast cancer

目的构建携带IL-24基因的溶瘤腺病毒CNHK600-IL24，评估该病毒在裸鼠体内对乳腺癌的抑瘤能力。方法将IL-24基因插入腺病毒穿梭载体SG502-△CR2，与5型腺病毒骨架载体pPE3共转染至293细胞，获得溶瘤腺病毒CNHK600-IL24。建立乳腺癌原位成瘤模型、尾静脉注射及左心室注射模拟转移瘤模型，通过尾静脉注射CNHK600-IL24，应用“活体内光学成像系统”动态地观察病毒的疗效。结果CNHK600-IL24经测序及PCR鉴定正确，滴度为1．9×10＾10pfu／ml。乳腺癌原位成瘤模型：对照组的光子数以及肿瘤体积均明显高于CNHK600-IL24各治疗组（P〈0．05）。CNHK600-IL24治疗后肿瘤组织出现明显的坏死，肿瘤细胞发生显著的凋亡，免疫组化检测可见肿瘤细胞内Hexon和IL-24的表达。尾静脉注射模拟转移瘤模型：对照组的裸鼠大部分于38d前死亡，而CNHK600-IL24组的生存天数明显延长（P〈0．05）。左心室注射模拟转移瘤模型：活体内光学成像可见对照组与治疗组之间具有明显差别。结论高滴度的溶瘤腺病毒CNHK600．1124对于乳腺癌具有明显的抑瘤效果。

Objective To construct an oncolytic adenovirus CNHK600-IL24, and to observe the in vivo effects of CNHK600-IL24 in treating breast cancer. Methods The IL-24 gene was cloned into adenovirus shuttle vector SGS02-ACR2, and CNHK600-IL24 was obtained by cotransfection of SGS02-INS- IL24 and pPE3 plasmids and subsequent recombination in 293 ceils. Based on the establishment of the athymic mice model of breast cancer in situ and imitated metastatic breast cancer by injection in the vena caudalis and the left artfium, we administered the virus by the tail vein. We used the optical imaging in vivo system to monitor the effects. Results The oneolytic adenovirus CNHK600-IL24 was correctly constructed and confirmed by restriction DNA sequence analysis and PCR. The titer of CNHK600-IL24 reached 1.9 × 10^10 pfu/ml. Establishing athymic mice model of breast cancer in situ, the volume and photon number of the tumors in the control group was significantly larger than those of the CNHK600-IL24 group（ P 〈 0. 05 ）. The tumor had conspicuous necrosis after the treatment of CNHK600-IL24. There was noticeable apoptosis of the tumor cells. Immunohistochemistry showed the expression of IL-24 and the Hexon protein in the tumor cells. In athymic mice model of imitated metastatic breast cancer by infusion into the vena caudalis, most of the mice in the control group died before 38 days, the mice of the CNHK600-IL24 group survived significantly longer（ P 〈 0. 05 ）. Using athymic mice model of imitated metastatic breast cancer by infusion in the left artrium, the optical imaging in vivo system showed obvious difference between the control group and the CNHK600-IL24 group. Conclusions The high-titer oncolytic adenovirus CNHK600-IL24 was successfully constructed and purified. The oncolytic adenovirus had obvious antitumor effect on breast cancer.