Wan Bing, Qin Lu, Ma Weihong, Wang He
Gynecologist Tumor Department, Guangxi Medical University Cancer Hospital, 71 Hedi Road, Zhong Shan Street, Nanning, 530021, Guangxi Zhuang Autonomous Region, China.
Infect Agent Cancer. 2022 Feb 23;17(1):5. doi: 10.1186/s13027-022-00417-3.
This study aims to prepare candidate vaccines for cervical cancer immunotherapy by inserting the fused genes of human papillomavirus (HPV)16/18/58 mE6E7 lacking transforming activity into an adenovirus vector and to verify its efficiency in model mice with tumor expressing the associated HPV genes.
The E6/E7 genes of HPV16/18/58 were point-mutated to abolish their transforming activity, and adenovirus (AD)-HPV16/18/58 mE6E7 adenovirus vaccine was constructed. The immune effect of the adenovirus vaccine against HPV16/18/58-type tumors was analyzed by tumor morphology, enzyme linked immunosorbent assay, enzyme-linked immunospot and specific cytotoxic T lymphocyte (CTL) and T lymphocyte subsets.
The HPV16/18/58 mE6E7 plasmid containing point mutations was verified by quantitative real-time polymerase chain reaction (qRT-PCR), enzyme digestion and electrophoresis, and gene sequencing. qRT-PCR and Western blots verified that AD-HPV16/18/58 mE6E7 could express the HPV16 mE6E7, HPV18 mE6E7 and HPV58 mE6E7 fusion genes and proteins in cells. The results of animal experiments were as follows: In the vaccine group, the tumors formed later, the incubation period was longer, the growth was slower, growth was inhibited, and the survival period was significantly prolonged. The immunological results all showed that the vaccine could induce effective humoral and cellular immunity in mice with three types of tumors, compared with the phosphate buffered saline (PBS) group and the adenovirus-negative control (AD-NC) group, the differences were statistically significant (P < 0.05).
We successfully constructed the HPV16/18/58 trivalent therapeutic adenovirus vaccine AD-HPV16/18/58 mE6E7. The AD-HPV16/18/58 mE6E7 adenovirus vaccine can protect immunized mice to a certain extent from TC-1, U14/LV-HPV18 E6E7 and U14/LV-HPV58 E6E7 cells, which contain HPV16, 18 and 58 E6 and/or E7 genes, respectively.
本研究旨在通过将缺乏转化活性的人乳头瘤病毒（HPV）16/18/58 mE6E7融合基因插入腺病毒载体来制备用于宫颈癌免疫治疗的候选疫苗，并在表达相关HPV基因的肿瘤模型小鼠中验证其有效性。
对HPV16/18/58的E6/E7基因进行点突变以消除其转化活性，并构建腺病毒（AD）-HPV16/18/58 mE6E7腺病毒疫苗。通过肿瘤形态学、酶联免疫吸附测定、酶联免疫斑点法以及特异性细胞毒性T淋巴细胞（CTL）和T淋巴细胞亚群分析腺病毒疫苗对HPV16/18/58型肿瘤的免疫效果。
通过定量实时聚合酶链反应（qRT-PCR）、酶切、电泳和基因测序验证了含有点突变的HPV16/18/58 mE6E7质粒。qRT-PCR和蛋白质免疫印迹法验证了AD-HPV16/18/58 mE6E7可在细胞中表达HPV16 mE6E7、HPV18 mE6E7和HPV58 mE6E7融合基因及蛋白。动物实验结果如下：在疫苗组中，肿瘤形成较晚，潜伏期较长，生长较慢，生长受到抑制，生存期显著延长。免疫学结果均显示该疫苗可在三种肿瘤类型的小鼠中诱导有效的体液免疫和细胞免疫，与磷酸盐缓冲液（PBS）组和腺病毒阴性对照（AD-NC）组相比，差异具有统计学意义（P < 0.05）。
我们成功构建了HPV16/18/58三价治疗性腺病毒疫苗AD-HPV16/18/58 mE6E7。AD-HPV16/18/58 mE6E7腺病毒疫苗可在一定程度上保护免疫小鼠免受分别含有HPV16、18和58 E6和/或E7基因的TC-1、U14/LV-HPV18 E6E7和U14/LV-HPV58 E6E7细胞的侵害。
