针对癌睾抗原PLAC1、PL2L60、MAGE-3的长肽疫苗设计及其免疫活性检测

摘要

目的:设计针对癌睾抗原PLAC1、PL2L60、MAGE-3的长肽,并检测其免疫活性.方法:利用在线数据库预测PLAC1、PL2L60、MAGE-3的CTL表位,在表位集中区域选取适当长度的长肽,化学合成并纯化后,通过体外和体内活性实验验证长肽是否具有免疫活性.结果:针对PLAC1、PL2L60、MAGE-3预测出HLA-A2限制性CTL表位并设计出7条长肽.体外自提呈、DC荷肽的ELISPOT结果和体外LDH结果显示,PLAC1 P42-64、PL2L60 P169-191、MAGE-3 P101-122、MAGE3-P152-175等4条长肽具有较好的免疫活性;体内ELISA结果显示MAGE-3 P152-175在HLA-A2.1/Kb转基因小鼠中诱导了较多的IFN-γ释放,体内LDH结果显示PLAC1 P42-64、PL2L60 P169-191、MAGE-3 P101-122、MAGE-3 P152-175等4条长肽所诱导的HLA-A2.1/Kb转基因小鼠脾淋巴细胞对靶细胞具有较高的杀伤率.结论:成功筛选鉴定出针对癌睾抗原PLAC1、PL2L60、MAGE-3的4条长肽,可用作免疫治疗疫苗.%Aim:To design long peptides based on CTL epitope prediction for cancer testis antigen PLAC 1, PL2L60 and MAGE-3, and to study the immune activity of the long peptides .Methods:The CTL epitope of PLAC1, PL2L60 and MAGE-3 were predicted using online databases , the long peptides with appropriate length in the epitope region was chosen , and then they were synthesized chemically and purified .Finally in vitro and in vivo activity experiments were used to verify if the long peptides had immune activity .Results: HLA-A2 restricted CTL epitopes for PLAC1, PL2L60 and MAGE-3 were predicted, and 7 long peptides were designed .The autopresentation, long peptide-pulsed DC ELISPOT assay and LDH results showed that long peptides PLAC1 P42-64, PL2L60 P169-191, MAGE-3 P101-122 and MAGE-3 P152-175 had better immune activity; the ELISA results in vivo showed that MAGE-3 P152-175 induced more IFN-γrelease in HLA-A2.1/Kb transgenic mice, in vivo LDH results showed that the spleen lymphocytes from HLA-A2.1/Kb transgenic mice induced by the long peptides PLAC1 P42-64, PL2L60 P169-191, MAGE-3 P101-122 and MAGE-3 P152-175 had higher killing rate on target cells .Conclusion:Four long peptides pointing to cancer testis antigen PLAC 1, PL2L60 and MAGE-3 have been successfully identified , which could be used as immunotherapy vaccine in future .