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目的构建胰腺癌黏液蛋白核芯肽连续重复序列(MUC1-VNTR)核酸疫苗,研究疫苗所诱生的免疫应答。方法将重组人VNTR基因定向克隆入真核表达载体pcDNA3．1／Myc-his(+) A质粒的多克隆位点中,构建胰腺癌MUC1-VNTR核酸疫苗。C57BL／6(H-2b)小鼠2次免疫2周后取血检测抗VNTR抗体,取脾细胞体外以VNTR合成肽特异刺激培养6 d后进行细胞毒性T淋巴细胞 (CTL)杀伤试验。结果疫苗组小鼠脾细胞CTL的特异性杀伤率(34．8±3．1)％显著高于载体对照组[(9．2±0．8)％,P<0．01]和空白对照组[(6．1±0．6)％,P<0．01]。CTL对未经VNTR合成肽孵育的靶细胞EL4-VNTR杀伤较低(P<0．01)。VU 3C6可抑制CTL对经VNTR合成肽孵育的靶细胞E14-VNTR+的杀伤活性(P<0．01)。疫苗组小鼠血清抗VNTR抗体等效浓度(2324±238)μg/ml 高于载体对照组(1896±533)μg／ml和空白对照组(1736±142)μg／ml,P<0．01。结论本实验构建的胰腺癌MUC1—VNTR核酸疫苗免疫C57BL／6小鼠后可诱生VNTR特异的CTL应答和抗体应答。
Objective To construct the Muc1-VNTR DNA vaccine of pancreatic cancer mucosal protein core peptide to study the immune response induced by the vaccine. Methods The recombinant human VNTR gene was cloned into the multi-cloning site of the eukaryotic expression vector pcDNA3.1 / Myc-his (+) A to construct the MUC1-VNTR DNA vaccine. Anti-VNTR antibody was detected in blood of C57BL / 6 (H-2b) mice two weeks after two immunizations. The cytotoxic T lymphocyte (CTL) cytotoxicity test was performed after the spleen cells were cultured for 6 days with VNTR synthetic peptides. Results The specific killing rate of CTL in the vaccine group was significantly higher than that in the control group [(9.2 ± 0.8)%, P <0.01] and the blank control Group [(6.1 ± 0.6)%, P <0.01]. CTL killed EL4-VNTR target cells that had not been incubated with the VNTR synthetic peptide (P <0.01). VU 3C6 inhibited CTL killing activity (P <0.01) on target cells E14-VNTR + incubated with VNTR synthetic peptide. The serum concentration of anti-VNTR antibody in vaccine group mice was 2324 ± 238 μg / ml higher than that in vehicle control group (1896 ± 533 μg / ml) and blank control group (1736 ± 142 μg / ml, P <0.01) . Conclusions The VNTR-specific CTL response and antibody response of C57BL / 6 mice immunized with the MUC1-VNTR DNA vaccine of pancreatic cancer were induced by this experiment.