Division of Dermatology, Department of Medicine, University of Washington, Seattle, WA, USA.
Clinical Research Division, Fred Hutchinson Cancer Research Center, Seattle, Washington, USA.
J Immunother Cancer. 2022 Sep;10(9). doi: 10.1136/jitc-2022-005328.
Merkel cell carcinoma (MCC) often responds to PD-1 pathway blockade, regardless of tumor-viral status (~80% of cases driven by the Merkel cell polyomavirus (MCPyV)). Prior studies have characterized tumor-specific T cell responses to MCPyV, which have typically been CD8, but little is known about the T cell response to UV-induced neoantigens.
A patient in her mid-50s with virus-negative (VN) MCC developed large liver metastases after a brief initial response to chemotherapy. She received anti-PD-L1 (avelumab) and had a partial response within 4 weeks. Whole exome sequencing (WES) was performed to determine potential neoantigen peptides. Characterization of peripheral blood neoantigen T cell responses was evaluated via interferon-gamma (IFNγ) ELISpot, flow cytometry and single-cell RNA sequencing. Tumor-resident T cells were characterized by multiplexed immunohistochemistry.
WES identified 1027 tumor-specific somatic mutations, similar to the published average of 1121 for VN-MCCs. Peptide prediction with a binding cut-off of ≤100 nM resulted in 77 peptides that were synthesized for T cell assays. Although peptides were predicted based on class I HLAs, we identified circulating CD4 T cells targeting 5 of 77 neoantigens. In contrast, no neoantigen-specific CD8 T cell responses were detected. Neoantigen-specific CD4 T cells were undetectable in blood before anti-PD-L1 therapy but became readily detectible shortly after starting therapy. T cells produced robust IFNγ when stimulated by neoantigen (mutant) peptides but not by the normal (wild-type) peptides. Single cell RNAseq showed neoantigen-reactive T cells expressed the Th1-associated transcription factor (T-bet) and associated cytokines. These CD4 T cells did not significantly exhibit cytotoxicity or non-Th1 markers. Within the pretreatment tumor, resident CD4 T cells were also Th1-skewed and expressed T-bet.
We identified and characterized tumor-specific Th1-skewed CD4 T cells targeting multiple neoantigens in a patient who experienced a profound and durable partial response to anti-PD-L1 therapy. To our knowledge, this is the first report of neoantigen-specific T cell responses in MCC. Although CD4 and CD8 T cells recognizing viral tumor antigens are often detectible in virus-positive MCC, only CD4 T cells recognizing neoantigens were detected in this patient. These findings suggest that CD4 T cells can play an important role in the response to anti-PD-(L)1 therapy.
默克尔细胞癌（MCC）通常对 PD-1 通路阻断有反应，无论肿瘤病毒状态如何（~80%的病例由默克尔细胞多瘤病毒（MCPyV）驱动）。先前的研究已经描述了针对 MCPyV 的肿瘤特异性 T 细胞反应，这些反应通常是 CD8，但对于紫外线诱导的新抗原的 T 细胞反应知之甚少。
一位 50 多岁的女性患者患有病毒阴性（VN）MCC，在最初接受化疗后短暂缓解后出现大肝转移。她接受了抗 PD-L1（avelumab）治疗，并在 4 周内出现部分缓解。进行全外显子组测序（WES）以确定潜在的新抗原肽。通过干扰素-γ（IFNγ）ELISpot、流式细胞术和单细胞 RNA 测序评估外周血新抗原 T 细胞反应。通过多重免疫组织化学检测肿瘤驻留 T 细胞。
WES 鉴定出 1027 个肿瘤特异性体细胞突变，与 VN-MCC 发表的平均 1121 个相似。用结合截止值≤100 nM 的肽预测导致了 77 个用于 T 细胞测定的合成肽。尽管肽是基于 I 类 HLA 预测的，但我们鉴定出了针对 77 个新抗原中的 5 个的循环 CD4 T 细胞。相比之下，没有检测到新抗原特异性 CD8 T 细胞反应。在接受抗 PD-L1 治疗之前，新抗原特异性 CD4 T 细胞在血液中无法检测到，但在开始治疗后不久即可轻易检测到。T 细胞在受到新抗原（突变）肽刺激时会产生大量 IFNγ，但不会受到正常（野生型）肽的刺激。单细胞 RNAseq 显示新抗原反应性 T 细胞表达 Th1 相关转录因子（T-bet）和相关细胞因子。这些 CD4 T 细胞没有明显的细胞毒性或非 Th1 标志物。在预处理肿瘤内，常驻 CD4 T 细胞也呈 Th1 偏向性并表达 T-bet。
我们在一名接受抗 PD-L1 治疗后获得深刻和持久部分缓解的患者中鉴定并描述了针对多种新抗原的肿瘤特异性 Th1 偏向性 CD4 T 细胞。据我们所知，这是 MCC 中首例新抗原特异性 T 细胞反应的报告。尽管在病毒阳性 MCC 中经常检测到识别病毒肿瘤抗原的 CD4 和 CD8 T 细胞，但在该患者中仅检测到识别新抗原的 CD4 T 细胞。这些发现表明 CD4 T 细胞在抗 PD-（L）1 治疗反应中可能发挥重要作用。
