摘要
目的:利用TCGA数据库筛选和分析膀胱癌肿瘤抑制基因CDKN1A的突变特征,筛选具有肿瘤新抗原特征的CDKN1A抗原突变肽并分析其在人群中的免疫反应性。方法:利用TCGA公共数据库中413例膀胱癌全外显子测序数据,根据亲和力预测和基因表达量,筛选出人类白细胞抗原(HLA)-A2限制的CDKN1A突变型肽段与野生型肽段。利用T2细胞实验检测配对肽段的结合力。采用酶联免疫斑点试验检测HLA-A2基因型膀胱癌患者外周血单个核细胞对配对抗原肽的免疫反应。结果:根据预测的突变型肽段高亲和力而野生型肽段低亲和力的原则,筛选获得HLA-A2限制性野生型和突变型抗原肽段。体外T2亲和力结合实验结果显示,来自CDKN1A基因编码的突变型肽段亲和力(Kd=3.724 mg/ml)显著高于野生型肽段。40例HLA-A2阳性膀胱癌患者突变型肽段刺激产生的抗原肽特异性IFN-γ平均点数明显高于野生型(P=0.045),同时突变型肽段在40例患者中的阳性反应率达到25%(10/40)。结论:利用TCGA数据筛选获得膀胱癌抑癌基因CDKN1A的突变型肽段能够在膀胱癌患者中诱导高于野生型肽段的特异性免疫反应性,具有膀胱癌肿瘤新抗原的免疫学特征,为其应用于膀胱癌的免疫治疗提供了理论依据。
Objective:To use the mutation characteristics of the bladder cancer suppressor gene CDKN1 A screened and analyzed by the TCGA database,CDKN1 A coded mutant peptides with neoantigen property were screened and their immune reactivity in the population was analyzed.Methods:By using bioinformatics,HLA-A2 restricted mutant type(MT)peptides and wild type(WT)counterparts were selected based on whole exosome sequencing data of bladder cancer from the TCGA database mainly based on predictive affinity as well as expressing frequency. The binding affinity of predicted peptide pairs were determined by using T2 cell-based assay. Peptides with high affinity to HLA-A2 molecules were subjected to the determination of immunoreactivity in bladder cancer patients by using enzyme-linked immunospot(ELISpot)assay upon in vitro stimulation.Results:CDKN1 A coded paired peptides were synthesized with the high affinity of mutant type and low affinity of wild type subjected to T2 cells based validation. MT peptides exhibited higher and stronger affinity to HLA-A2 molecules when compared to WT counterparts. Their immunoreactivity was determined in PBMCs from 40 HLA-A2 positive bladder cancer patients. It was shown that bladder cancer patients exhibited higher responsiveness to MT peptides than to WT peptides. The average number of peptide-specific IFN-γ spots upon MT peptide stimulation was dramatically higher than that upon WT stimulation(P=0.045). Meanwhile,the positive rate of mutant peptide was 25%(10/40).Conclusion:CDKN1 A coded MT peptides screened from the TCGA database can induce stronger immune reactivity than WT peptides in bladder cancer,which possess the immunological characteristics of neoantigens in bladder cancer,provides the theoretical basis for its application in immunotherapy of bladder cancer.
作者
王晨
刘源涌
夏维敏
季萍
余奇文
曹志伟
王颖
沈海波
WANG Chen;LIU Yuan-Yong;XIA Wei-Min;JI Ping;YU Qi-Wen;CAO Zhi-Wei;WANG Ying;SHEN Hai-Bo(Department of Urology,Xinhua Hospital,Shanghai Jiaotong University School of Medicine,Shanghai 200082,China)
出处
《中国免疫学杂志》
CAS
CSCD
北大核心
2021年第10期1246-1251,共6页
Chinese Journal of Immunology
作者简介
王晨,男,硕士,主要从事膀胱癌免疫治疗方面研究,E-mail:845258355@qq.com;通信作者:沈海波,男,博士,副主任医师,硕士生导师,主要从事膀胱癌免疫治疗与免疫环境研究,E-mail:shenhaibo@xinhuamed.com.cn。
