The Determinants of CD8 T cell Dysfunction in Oral Cavity Squamous Cell Carcinoma

项目摘要 包括口腔鳞状细胞癌（OCSCC）在内的头颈部癌症是导致口腔癌的第六大原因。 癌症和发病率和死亡率的主要原因。不幸的是，OCSCC的五年生存率 在过去的30年里只略有改善。最近，出现了免疫检查点抑制（ICI）疗法， 作为改善OCSCC结局的有希望的治疗，但只有14-22%的OCSCC患者对ICI有反应。 因此，描绘ICI抗性的机制代表了一个机会，以确定新的目标， 增加ICI功效并改善OCSCC患者存活率。CD 8 T细胞是主要的免疫细胞类型， 在ICI期间杀死恶性细胞，并且它们的功能障碍可能有助于ICI抗性。在慢性 在对肿瘤的应答中，CD 8 T细胞进入耗竭细胞状态-一种功能失调的表型，其特征在于 降低裂解靶细胞的能力。胸腺细胞选择相关转录因子HMG的表达 框（TOX）对于诱导CD 8 T细胞耗竭是关键的，并且在CD 8 T细胞亚群中富集。 OCSCC患者肿瘤样品的单细胞RNA-seq数据集。直接转录的基因 然而，由TOX调节的促进CD 8 T细胞功能障碍并且可能是治疗靶向的， 定义不好。除TOX外，恶性细胞信号可能导致CD 8 T细胞功能障碍和ICI。 阻力我们在口腔鳞状细胞癌中发现了一种上皮/间质混合型（HEM）恶性细胞状态， 与邻近CD 8 T细胞的肿瘤边缘相关，并与这些邻近CD 8 T细胞的耗竭相关。的 HEM细胞在这种关联下的机制还没有很好地理解。这个问题的主要假设是 推测OCSCC中的CD 8 T细胞功能障碍是由CD 8 T细胞中TOX的直接基因靶点驱动的， 通过免疫抑制接触和HEM细胞的旁分泌信号。该提案的目的是定义 通过以下目的可以靶向改善ICI的CD 8 T细胞功能障碍的机制：在目的1中， TOX的直接基因靶点及其在CD 8 T细胞功能障碍中的作用将使用CD 8 T细胞在体外确定 分离自口腔鳞状细胞癌样本。在目标2中，将患者来源的CD 8 T细胞与患者匹配的CD 8 T细胞共培养。 OCSCC肿瘤衍生的类器官或表达模型抗原的癌细胞系。在这些共同培养中， 特异性膜和分泌的蛋白质将在恶性细胞中被敲低，以确定它们在恶性细胞中的作用。 抑制CD 8 T细胞。拟议的研究将提供机制洞察CD 8 T细胞功能障碍， OCSCC，从而支持开发治疗性活化CD 8 T细胞并改善 ICI和OCSCC结果。重要的是，在这个奖学金，我将追求科学和临床活动下， 由医生-科学家组成的综合研究金咨询委员会的直接指导， 癌症和免疫学研究专家。我所建议的训练对我实现 研究癌症与免疫的相互作用，并成为一名独立的学术医生和科学家。

PROJECT SUMMARY Head and neck cancer, including oral cavity squamous cell carcinoma (OCSCC), is the sixth leading cause of cancer and a major cause of morbidity and mortality. Unfortunately, the five-year survival rate of OCSCC has only slightly improved in the past 30 years. Recently, immune checkpoint inhibition (ICI) therapies have emerged as a promising treatment to improve OCSCC outcomes, yet only 14-22% of OCSCC patients respond to ICI. Thus, delineating the mechanisms of ICI resistance represents an opportunity to identify novel targets that may increase ICI efficacy and improve OCSCC patient survival. CD8 T cells are the primary immune cell type that kills malignant cells during ICI, and their dysfunction may contribute to ICI resistance. During the chronic response to a tumor, CD8 T cells enter an exhausted cell state – a dysfunctional phenotype characterized by decreased ability to lyse target cells. Expression of the transcription factor thymocyte selection-associated HMG box (TOX) is critical for inducing CD8 T cell exhaustion and is enriched in the exhausted CD8 T cell subset within single cell RNA-seq datasets of OCSCC patient tumor samples. The genes that are directly transcriptionally regulated by TOX that promote CD8 T cell dysfunction and may be therapeutically targeted, however, remain poorly defined. In addition to TOX, malignant cell signals may contribute to CD8 T cell dysfunction and ICI resistance. We discovered a hybrid epithelial/mesenchymal (HEM) malignant cell state in OCSCC that localizes to the tumor edge adjacent to CD8 T cells and is associated with exhaustion of these adjacent CD8 T cells. The mechanisms of HEM cells underlying this association are not well understood. The primary hypothesis of this proposal is that CD8 T cell dysfunction in OCSCC is driven by the direct gene targets of TOX in CD8 T cells and by immunosuppressive contact and paracrine signaling from HEM cells. The goal of this proposal is to define mechanisms of CD8 T cell dysfunction that may be targeted to improve ICI through the following Aims: In Aim 1, the direct gene targets of TOX and their role in CD8 T cell dysfunction will be defined in vitro using CD8 T cells isolated from OCSCC samples. In Aim 2, patient-derived CD8 T cells will be co-cultured with patient-matched OCSCC tumor-derived organoids or cancer cell lines expressing a model antigen. In these co-cultures, HEM- specific membrane and secreted proteins will be knocked-down in the malignant cells to determine their role in suppressing CD8 T cells. The proposed research will provide mechanistic insight into CD8 T cell dysfunction in OCSCC, thereby supporting development of new strategies to therapeutically activate CD8 T cells and improve ICI and OCSCC outcomes. Importantly, during this fellowship, I will pursue scientific and clinical activities under the direct mentorship of a comprehensive fellowship advisory committee composed of physician-scientists and experts in cancer and immunology research. My proposed training will be vital to accomplishing my goal of investigating cancer-immune interactions and maturing as an independent, academic physician-scientist.