表达CRT/E7融合蛋白的人乳头瘤病毒DNA疫苗对肿瘤生长和血管生成抑制作用的研究

HPV DNA Vaccines Expressing Recombinant CRT/ HPV6bE7 Fusion Protein Inhibit Tumor Growth and Angiogenic Activity

研究HPV6bE7／CRTDNA疫苗免疫保护，清除已有感染和相关肿瘤细胞及其血管生成抑制作用，分析CRT抗血管生成的功能片段，为筛选高效的HPV疫苗提供实验依据。用重组质粒pcDNA3．1（＋）GFP—CRT120／HPV6bE7、pcDNA3．1（＋）-GFP—HPV6bE7、pcDNA3．1（＋）GFP—CRT120、pcDNA3．1（＋）-GFP—CRT180／HPV6bE7、pcDNA3．1（＋）-GFPCRT180通过肌内注射途径免疫C57BL／6小鼠。Matrigel法进行抗血管活性检测；B16／HPV6bE7细胞接种于C57BL／6雌性小鼠建立荷瘤模型，观察各组DNA疫苗对HPV6bE7基因的荷瘤组织的出瘤时间和肿瘤大小的影响。结果显示：重组DNA疫苗pcDNA3．1-CRT180／HPV6bE7和pcDNA3．1CRT180在动物体内能对bFGF诱导的新生血管的生成有明显的抑制作用；CRT180／HPV6bE7和CRT180能显著抑制荷瘤的大小且CRT180／HPV6bE7免疫组较其他组能明显延缓荷瘤的形成时间、生长速率以及肿瘤重量。CRT180／HPV6bE7免疫组较其他组能诱导更强的血管抑制作用和部分抑制肿瘤生长，推测抑制血管的功能片段存在于CRT120～180aa片段上。

This paper was to study the angiogenic inhibitory effect and the potential antitumor effect of the constructed recombinant DNA vaccine CRT/HPV6bE7 in vivo . The C57BL/6 mice were vaccinated respectively with recombinant CRT/HPV6bE7 DNA plamids. The inhibitory effects on angiogenesis of generated vaccines in vivo were evaluated by a bFGF-induced angiogenesis assay using the Matrigel kit. To investigate the potential antitumor effect, the mean tumor weights, sizes and tumor appearing times were measured in C57BL/6 mice treated with HPV6bE7-expressing B16 cells. The results indicated that the recombinants CRT180/HPV6bE7 and CRT180 showed strong anti-angiogenic effects in bFGF-induced angiogenesis in vivo. Moreover, CRT180/HPV6bE7 and CRT180 DNA vaccines could significantly inhibit the tumor growth in tumor challenge experiment, and CRT180/HPV6bE7 was superior to other vaccines in delaying tumor formation time, limiting tumor size and weight in tumor protection experiment. In conclusion,recombinant CRT180/HPV6bE7 DNA could elicit a most efficient anti-angiogenic effect and inhibit tumor growth in mice inoculated with DNA vaccines. The antiangiogenic activity of CRT were suggested residing in a domain between CRT 120 -180 aa.