中文摘要：

目的用SYFPEITHI数据库筛选针对HIA-A2的人乳头瘤病毒（HPV）18E7抗原表位多肽联合CpG制备HPV18疫苗。方法（1）利用生物信息学平台预测和筛选HPV18E7蛋白的HLA-A2抗原结合表位。（2）收集HLA-A2和HPV18阳性／阴性的病人／志愿者外周血和相应的肿瘤组织标本，并将这些标本随机分为7组：ETPA＋CpG、E7PB＋CpG、E7PC＋CpG、E7PD＋CpG、CpG、IRT＋CpG及对照组。利用MTT比色法检测各组不同时间段的T细胞增殖效应；利用乳酸脱氢酶（LDH）释放法检测人外周血淋巴细胞（PBMC）在不同效靶比下对肿瘤细胞的细胞毒活性；利用酶联免疫斑点测定法（ELISPOT）检测外周血中分泌IFN-γ的T细胞水平。结果（1）筛选出一致性较高，特异性及亲和力较强，综合评分前4位的4段多肽作为研究对象，分别命名为E7PA，E7PB，ETPC，E7PD。（2）各实验组中T细胞较对照组增殖明显（P〈0．05），在接种疫苗后不同时间点连续观察中以E7PA＋CpG组增殖效应最为突出；LDH释放实验示：效靶比为100：1时，E7PA＋CpG组与对照组相比，差异有统计学意义（P〈0．01），且效靶比呈明显的浓度依赖性。而除IR-T＋CpG组外其余各组较对照组差异有统计学意义（P〈0．05），但组间差异无统计学意义；外周血中分泌IFN-γ的T细胞在E7PA组激活最明显，与对照组比差异有统计学意义（P〈0．01）；E7PA＋CpG组在HPV18阳性组中可引起大量的T细胞分泌IFN-γ，与其他组相比差异有统计学意义（P〈0．05）。结论HPV18E7PA联合CpG可以在体外激活特异性的细胞免疫，是治疗HPV18相关疾病的有效靶点。

英文摘要：

Objective To screen and prepare the vaccine of human papillomavirus （HPV）18 E7 peptide target at human leukocyte antigen （HLA） -A2 plus CpG through SYFPEITHI. Methods （ 1 ） The SYFPEITHI database was employed for predicting and screening of HPV18 E7 HLA-A2 restricted T cell epitopes. （2） The peripheral blood and tumor tissue sample of HLA-A2 positive and HPV18 positive/ negative patients were collected and randomly divided into 7 groups, i. e. E7PA ＋ CpG, E7PB ＋ CpG, E7PC ＋ CpG, E7PD ＋ CpG, CpG, IR-T ＋ CpG and control groups respectively. T cell proliferation was detected by thiazolyl blue tetrazolium bromide （MTT） assay at different timepoints. Lactate dehydrogenase delivery method （LDH） was used to test the cytolytic t lymphocyte （CTL） activity of peripheral blood mononuclear cell （PBMC） in different ratios of effect and target （E： T）. And the level of activity T cells was evaluated by interferon gamma （IFN-γ）-related enzyme-linked immuno-spot assay （ELISPOT）. Results （1） Four peptides named E7PA, E7PB, E7PC and E7PD were obtained separately with high levels of affinity and specificity. （2） During continuous observations after vaccination, the E7PA ＋ CpG group had the most pronounced proliferation rate. When E： T = 100： 1, the E7PA ＋ CpG group had more powerful CTL effect than the control group with statistic significance （P 〈 0.00）. E： T was concentration-dependent. Except for IR-T ＋ CpG, all other groups had great difference than control group with statistic significance （P 〈 0. 05 ） but no significant difference between the groups. The levels of IFN-γ/spot-forming T cells were higher in the E7PA ＋ CpG group than the control group with statistic significance （ P 〈 0. 01 ）. In terms of specificity, E7PA ＋ CpG in the HPVI8 positive group could induce the proliferation of IFN-γ-secreting T cells. And there was statistical difference with the control group （ P 〈 0. 05 ）. Conclusion Screening the HPV18 E7 pep