Department of Microbiology and Immunology, Medical School of Southeast University, Nanjing, 210009, Jiangsu, China.
Department of Hepatobiliary Oncology, The Second Hospital of Nanjing Affiliated To Southeast University, Nanjing, 210003, Jiangsu, China.
Cancer Immunol Immunother. 2022 Jul;71(7):1569-1582. doi: 10.1007/s00262-021-03096-y. Epub 2021 Nov 1.
Hepatocellular carcinoma (HCC) is a malignant tumor with high mortality, but lacks effective treatments. Carcinoembryonic antigen glypican-3 (GPC3) is a tumor-associated antigen overexpressed in HCC but rarely expressed in healthy individuals and thus is one of the most promising therapeutic targets. T cell epitope-based vaccines may bring light to HCC patients, especially to the patients at a late stage. However, few epitopes from GPC3 were identified to date, which limited the application of GPC3-derived epitopes in immunotherapy and T cell function detection. In this study, a total of 25 HLA-A0201 restricted GPC3 epitopes were in silico predicted and selected as candidate epitopes. Then, HLA-A0201/GPC3 HCC patients' PBMCs were collected and co-stimulated with the candidate epitope peptides in ex vivo IFN-γ Elispot assay, by which five epitopes were identified as real-world epitopes. Their capacity to elicit specific CD8 T cells activation and proliferation was further confirmed by in vitro co-cultures of patients' PBMCs with peptide, in vitro co-cultures of healthy donors' PBLs with DCs and peptide, T2 cell binding assay as well as HLA-A2 molecule stability assay. Moreover, the in vivo immunogenicity of the five validated epitopes was confirmed by peptides cocktail/poly(I:C) vaccination in HLA-A0201/DR1 transgenic mice. Robust epitope-specific CD8 T cell responses and cytotoxicity targeting HepG2 cells were observed as detected by IFN-γ Elispot, intracellular IFN-γ staining and cytolysis assay. This study provided novel GPC3 CTL epitopes for the development of T cell epitope vaccines and evaluation of GPC3 specific T cell responses.
肝细胞癌 (HCC) 是一种死亡率很高的恶性肿瘤，但缺乏有效的治疗方法。癌胚抗原糖蛋白 3 (GPC3) 是 HCC 中过表达的肿瘤相关抗原，但在健康个体中很少表达，因此是最有前途的治疗靶点之一。基于 T 细胞表位的疫苗可能为 HCC 患者带来希望，尤其是晚期患者。然而，迄今为止，从 GPC3 中鉴定出的表位很少，这限制了 GPC3 衍生表位在免疫治疗和 T 细胞功能检测中的应用。在这项研究中，共预测和选择了 25 个 HLA-A0201 限制的 GPC3 表位作为候选表位。然后，从 HLA-A0201/GPC3 HCC 患者的 PBMC 中收集，并在体外 IFN-γ Elispot 检测中与候选表位肽共刺激，鉴定出五个表位为真实世界的表位。通过体外共培养患者 PBMC 与肽、体外共培养健康供体 PBL 与 DC 和肽、T2 细胞结合试验以及 HLA-A2 分子稳定性试验进一步证实了它们诱导特异性 CD8 T 细胞激活和增殖的能力。此外，通过肽鸡尾酒/聚 I:C 在 HLA-A0201/DR1 转基因小鼠中进行疫苗接种，证实了这五个验证表位的体内免疫原性。通过 IFN-γ Elispot、细胞内 IFN-γ 染色和细胞溶解试验检测到针对 HepG2 细胞的强大的表位特异性 CD8 T 细胞反应和细胞毒性。这项研究为开发 T 细胞表位疫苗和评估 GPC3 特异性 T 细胞反应提供了新的 GPC3 CTL 表位。
