The School of Basic Medical Sciences, Fujian Medical University, Fuzhou 350122, China.
Laboratory of Immuno-Oncology, Fujian Cancer Hospital & Fujian Medical University Cancer Hospital, Fuzhou 350014, China.
Cancer Biol Med. 2021 Nov 24;18(4):1053-65. doi: 10.20892/j.issn.2095-3941.2021.0022.
Neoantigens arising from gene mutations in tumors can induce specific immune responses, and neoantigen-based immunotherapies have been tested in clinical trials. Here, we characterized the efficacy of altered neoepitopes in improving immunogenicity against gastric cancer.
Raw data of whole-exome sequencing derived from a patient with gastric cancer were analyzed using bioinformatics methods to identify neoepitopes. Neoepitopes were modified by P1Y (the first amino acid was replaced by tyrosine) and P2L (the second amino acid was replaced by leucine). T2 binding and stability assays were used to detect the affinities between the neoepitopes and the HLA molecules, as well as the stabilities of complexes. Dendritic cells (DCs) presented with neoepitopes stimulated naïve CD8 T cells to induce specific cytotoxic T lymphocytes. ELISA and carboxyfluorescein succinimidyl ester were used to detect IFN-γ and TNF-α levels, and T cell proliferation. Perforin was detected by flow cytometry. The cytotoxicity of T cells was determined using the lactate dehydrogenase assay.
Bioinformatics analysis, T2 binding, and stability assays indicated that residue substitution increased the affinity between neoepitopes and HLA molecules, as well as the stabilities of complexes. DCs presented with altered neoepitopes stimulated CD8T cells to release more IFN-γ and had a greater effect on promoting proliferation than wild-type neoepitopes. CD8T cells stimulated with altered neoepitopes killed more wild-type neoepitope-pulsed T2 cells than those stimulated with wild-type neoepitopes, by secreting more IFN-γ, TNF-α, and perforin.
Altered neoepitopes exhibited greater immunogenicity than wild-type neoepitopes. Residue substitution could be used as a new strategy for immunotherapy to target neoantigens.
肿瘤基因突变产生的新抗原可诱导特异性免疫反应，新抗原为基础的免疫疗法已在临床试验中进行了测试。在这里，我们对改变的新抗原改善对胃癌的免疫原性的功效进行了表征。
使用生物信息学方法分析来自胃癌患者的全外显子组测序的原始数据，以鉴定新抗原。通过 P1Y（第一个氨基酸被酪氨酸取代）和 P2L（第二个氨基酸被亮氨酸取代）对新抗原进行修饰。T2 结合和稳定性测定用于检测新抗原与 HLA 分子之间的亲和力以及复合物的稳定性。树突状细胞（DC）呈递新抗原刺激幼稚 CD8 T 细胞诱导特异性细胞毒性 T 淋巴细胞。ELISA 和羧基荧光素琥珀酰亚胺酯用于检测 IFN-γ和 TNF-α水平以及 T 细胞增殖。通过流式细胞术检测穿孔素。用乳酸脱氢酶测定法测定 T 细胞的细胞毒性。
生物信息学分析、T2 结合和稳定性测定表明，残基取代增加了新抗原与 HLA 分子之间的亲和力以及复合物的稳定性。DC 呈递改变的新抗原刺激 CD8T 细胞释放更多的 IFN-γ，对促进增殖的作用大于野生型新抗原。与野生型新抗原刺激的 CD8T 细胞相比，用改变的新抗原刺激的 CD8T 细胞通过分泌更多的 IFN-γ、TNF-α和穿孔素杀死更多的野生型新抗原脉冲 T2 细胞。
改变的新抗原比野生型新抗原具有更强的免疫原性。残基取代可以作为针对新抗原的免疫疗法的新策略。
