PDF(Pubmed)
Abstract：
Hotspot driver mutations presented by human leukocyte antigens might be recognized by anti-tumor T cells. Based on their advantages of tumor-specificity and immunogenicity, neoantigens derived from hotspot mutations, such as PIK3CAH1047L, may serve as emerging targets for cancer immunotherapies. NetMHCpan V4.1 was utilized for predicting neoepitopes of PIK3CA hotspot mutation. Using in vitro stimulation, antigen-specific T cells targeting the HLA-A*11:01-restricted PIK3CA mutation were isolated from healthy donor-derived peripheral blood mononuclear cells. T cell receptors (TCRs) were cloned using single-cell PCR and sequencing. Their functionality was assessed through T cell activation markers, cytokine production and cytotoxic response to cancer cell lines pulsed with peptides or transduced genes of mutant PIK3CA. Immunogenic mutant antigens from PIK3CA and their corresponding CD8+ T cells were identified. These PIK3CA mutation-specific CD8+ T cells were subsequently enriched, and their TCRs were isolated. The TCR clones exhibited mutation-specific and HLA-restricted reactivity, demonstrating varying degrees of functional avidity. Identified TCR genes were transferred into CD8+ Jurkat cells and primary T cells deficient of endogenous TCRs. TCR-expressing cells demonstrated specific recognition and reactivity against the PIK3CAH1047L peptide presented by HLA-A*11:01-expressing K562 cells. Furthermore, mutation-specific TCR-T cells demonstrated an elevation in cytokine production and profound cytotoxic effects against HLA-A*11:01+ malignant cell lines harboring PIK3CAH1047L. Our data demonstrate the immunogenicity of an HLA-A*11:01-restricted PIK3CA hotspot mutation and its targeting therapeutic potential, together with promising candidates of TCR-T cell therapy.
摘要：
由人白细胞抗原呈递的热点驱动突变可能被抗肿瘤T细胞识别。基于其肿瘤特异性和免疫原性的优势，来自热点突变的新抗原，如PIK3CAH1047L，可能作为癌症免疫疗法的新兴靶标。NetMHCPanV4.1用于预测PIK3CA热点突变的新表位。使用体外刺激，从健康供体来源的外周血单核细胞中分离出靶向HLA-A*11:01限制性PIK3CA突变的抗原特异性T细胞.使用单细胞PCR和测序克隆T细胞受体(TCR)。它们的功能是通过T细胞活化标记来评估的，用突变型PIK3CA的肽或转导基因脉冲的癌细胞系的细胞因子产生和细胞毒性反应。鉴定了来自PIK3CA的免疫原性突变抗原及其相应的CD8+T细胞。这些PIK3CA突变特异性CD8+T细胞随后被富集，和他们的TCR被分离。TCR克隆表现出突变特异性和HLA限制性反应性，显示不同程度的功能亲合力。将鉴定的TCR基因转移到缺乏内源性TCR的CD8+Jurkat细胞和原代T细胞中。表达TCR的细胞表现出对表达HLA-A*11:01的K562细胞呈递的PIK3CAH1047L肽的特异性识别和反应性。此外，突变特异性TCR-T细胞对携带PIK3CAH1047L的HLA-A*11:01+恶性细胞系表现出细胞因子产生的升高和深远的细胞毒性作用。我们的数据证明了HLA-A*11:01限制性PIK3CA热点突变的免疫原性及其靶向治疗潜力，以及TCR-T细胞疗法的有希望的候选者。
