Survivin表位肽诱导CTL免疫学效应及杀瘤效应研究

Study on the immune and anti-tumor responses of CTL cells generated from Survivin-derived epitopes

目的:探讨Survivin HLA-A2限制性细胞毒性T淋巴细胞(CTL)表位SV20-28、SV23-31、SV88-96、负载DC诱导CTL免疫学效应和杀瘤活性。方法:从外周血中分离PBMC诱导DC,用Survivin高亲和性SV20-28、中等亲和性SV23-31及低亲和性表位肽SV88-96负载DC并诱导CTL;用酶联免疫斑点法(ELISPOT),检测CTL细胞IFN-γ的分泌情况;用乳酸脱氢酶(LDH)释放法检测其对靶细胞的杀伤作用。结果:Survivin高亲和性表位SV20-28和中等亲和力表位SV23-31负载DC诱导的CTL能产生分泌IFN-γ细胞,在ELISPOT试验中效靶比为1∶1时产生的斑点数分别为183.42±16.07、76.08±8.42,显著高于阴性对照组斑点数,P<0.05;而低亲和力表位SV88-96诱导的CTL未能产生明显的IFN-γ分泌细胞。高亲和力及中等亲和力表位肽诱导的CTL能以MHCⅠ限制方式杀伤Survivin阳性细胞,在效靶比为50∶1、25∶1、12.5∶1时的杀伤率分别为(62.40±5.16)%、(44.0±2.32)%、(26.53±1.07)%和(33.42±4.76)%、(25.16±2.64)%、(15.83±1.57)%,显著高于阴性对照组:(5.59±0.16)%、(4.76.0±0.32)%、(2.93±0.07)%;而低亲和力表位肽SV88-96诱导的免疫细胞对靶细胞没有明显的杀伤作用。结论:Survivin表位肽负载DC可有效诱导出CTL,Survivin表位肽SV20-28、SV23-31可有效诱导CTL产生免疫反应,并能以MHCⅠ限制方式杀伤Survivin阳性细胞。

Objective：To investigate immune and anti-tumor responses of CTL cells generated from DC loaded with HLA-restricted Survivin-derived peptide.Methods：The ability of producing IFN-γ and killing tumor cell of CTL induced by using high-affinity peptide SV20-28,intermediate-affinity peptide SV23-31 and low-affinity peptide SV88-96 respectively were measured by using ELISPOT method and LDH release assay.Results：Survivin-specific CTLs could be generated from the high-affinity survivin peptide of SV20-28-and the intermediate-affinity survivin pepticd SV23-31-loaded dendritic cells.The CTLs secreted IFN-γ.The number of the spots for CTL/SV20-28 cells and CTL/SV23-31 cells was 183.42±16.07 and 76.08±8.42 respectively,markedly higher than that of negative control with P〈0.05,while no obvious immuno-responses were observed for CTL/SV88-96cells.Survivin-specific CTL induced from high-affinity and intermediate-affinity survivin peptide SV20-28 and SV23-31 could kill survivin-positive cancerous cells in MHC class I-restricted fashion.The specific cytolytic rates of the CTLs were （62.40±5.16）%,（44.0±2.32）% and （26.53±1.07）% at effector to target ratio of 50 to 1,25 to 1 and 12.5 to 1 when induced by DCs loading SV20-28,and the rates for SV23-31 were （33.42±4.76）% and （25.16±2.64）% and （15.83±1.57）% at the same effector to target ratio.Conclusion：Survivin-derived peptides SV20-28 and SV23-31 can efficiently induce survivin-specific CTLs which can recognize and kill survivin positive cancerous cells in MHC class I-restricted fashion.