Centre for Cancer Immunology, Faculty of Medicine, University of Southampton, Southampton, UK.
Centre for Immuno-Oncology, Nuffield Department of Medicine, University of Oxford, Oxford, UK
J Immunother Cancer. 2023 Aug;11(8). doi: 10.1136/jitc-2023-007114.
CD8 T cells are a highly diverse population of cells with distinct phenotypic functions that can influence immunotherapy outcomes. Further insights on the roles of CD8 specificities and TCR avidity of naturally arising tumor-specific T cells, where both high and low avidity T cells recognizing the same peptide-major histocompatibility complex (pMHC) coexist in the same tumor, are crucial for understanding T cell exhaustion and resistance to PD-1 immunotherapy.
CT26 models were treated with anti-PD-1 on days 3, 6 and 9 following subcutaneous tumor implantation generating variable responses during early tumor development. Tetramer staining was performed to determine the frequency and avidity of CD8 T cells targeting the tumor-specific epitope GSW11 and confirmed with tetramer competition assays. Functional characterization of high and low avidity GSW11-specific CD8 T cells was conducted using flow cytometry and bulk RNA-seq. In vitro cytotoxicity assays and in vivo adoptive transfer experiments were performed to determine the cytotoxicity of high and low avidity populations.
Treatment success with anti-PD-1 was associated with the preferential expansion of low avidity (Tet) GSW11-specific CD8 T cells with Vβ TCR expressing clonotypes. High avidity T cells (Tet), if present, were only found in progressing PD-1 refractory tumors. Tet demonstrated precursor exhausted or progenitor T cell phenotypes marked by higher expression of Tcf-1 and T-bet, and lower expression of the exhaustion markers CD39, PD-1 and Eomes compared with Tet, whereas Tet cells were terminally exhausted. Transcriptomics analyses showed pathways related to TCR signaling, cytotoxicity and oxidative phosphorylation were significantly enriched in Tet found in both regressing and progressing tumors compared with Tet, whereas genes related to DNA damage, apoptosis and autophagy were downregulated. In vitro studies showed that Tet exhibits higher cytotoxicity than Tet. Adoptive transfer of Tet showed more effective tumor control than Tet, and curative responses were achieved when Tet was combined with two doses of anti-PD-1.
Targeting subdominant T cell responses with lower avidity against pMHC affinity neoepitopes showed potential for improving PD-1 immunotherapy. Future interventions may consider expanding low avidity populations via vaccination or adoptive transfer.
CD8 T 细胞是一个具有不同表型功能的高度多样化细胞群体，可影响免疫疗法的结果。进一步了解自然产生的肿瘤特异性 T 细胞的 CD8 特异性和 TCR 亲合力的作用，其中在同一肿瘤中同时存在高亲和性和低亲和性识别相同肽-MHC（pMHC）的 T 细胞，对于理解 T 细胞衰竭和对 PD-1 免疫疗法的抵抗至关重要。
在皮下肿瘤植入后第 3、6 和 9 天，用抗 PD-1 治疗 CT26 模型，在早期肿瘤发展过程中产生不同的反应。进行四聚体染色以确定靶向肿瘤特异性表位 GSW11 的 CD8 T 细胞的频率和亲和性，并通过四聚体竞争试验进行验证。使用流式细胞术和批量 RNA-seq 对高亲和性和低亲和性 GSW11 特异性 CD8 T 细胞进行功能表征。进行体外细胞毒性测定和体内过继转移实验以确定高亲和性和低亲和性群体的细胞毒性。
抗 PD-1 治疗的成功与低亲和性（Tet）GSW11 特异性 CD8 T 细胞的优先扩增有关，这些细胞具有表达 Vβ TCR 的克隆型。如果存在高亲和性 T 细胞（Tet），则仅在进展的 PD-1 难治性肿瘤中发现。与 Tet 相比，Tet 表现出前体衰竭或祖细胞 T 细胞表型，其特征是 Tcf-1 和 T-bet 的表达更高，而衰竭标志物 CD39、PD-1 和 Eomes 的表达更低，而 Tet 细胞则处于终末衰竭状态。转录组学分析表明，与 TCR 信号转导、细胞毒性和氧化磷酸化相关的途径在回归和进展肿瘤中均明显富集于 Tet，而与 DNA 损伤、细胞凋亡和自噬相关的基因则下调。体外研究表明，Tet 比 Tet 具有更高的细胞毒性。Tet 的过继转移显示出比 Tet 更有效的肿瘤控制效果，并且当 Tet 与两剂抗 PD-1 联合使用时，可实现治愈反应。
针对针对 pMHC 亲和力新表位的低亲和力亚优势 T 细胞反应进行靶向治疗，可能有助于改善 PD-1 免疫疗法。未来的干预措施可能需要通过疫苗接种或过继转移来扩大低亲和力群体。
