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

项目总结 头颈部癌症，包括口腔鳞状细胞癌(OCSCC)，是导致 癌症是发病率和死亡率的主要原因。不幸的是，OCSCC的五年存活率 在过去的30年里，只有轻微的改善。最近，出现了免疫检查点抑制(Ici)疗法。 作为一种有希望改善OCSCC预后的治疗方法，然而只有14-22%的OCSCC患者对ICI有反应。 因此，描绘ICI抵抗的机制代表着识别新的靶点的机会 提高ICI疗效，改善OCSCC患者生存。CD8 T细胞是主要的免疫细胞类型 在ICI过程中杀死恶性细胞，它们的功能障碍可能导致ICI抵抗。在慢性病期间 对肿瘤的反应，CD8T细胞进入耗尽的细胞状态--一种功能失调的表型，其特征是 裂解靶细胞的能力下降。胸腺细胞选择相关转录因子HMG的表达 BOX(TOX)是诱导CD8 T细胞耗竭的关键因子，在体内耗尽的CD8 T细胞亚群中富含 口腔鳞状细胞癌患者肿瘤样本的单细胞RNA-SEQ数据集。直接转录的基因 然而，由TOX调节的促进CD8 T细胞功能障碍并可能被作为治疗靶点的药物仍然存在 定义不明确。除TOX外，恶性细胞信号可能导致CD8 T细胞功能障碍和ICI 抵抗。我们在口腔鳞癌中发现了一种混合上皮/间充质(HEM)恶性细胞状态，定位于 与CD8 T细胞相邻的肿瘤边缘，并与这些邻近的CD8 T细胞的耗竭有关。这个 这种联系背后的HEM细胞机制还不是很清楚。这一点的主要假设是 认为口腔鳞癌中CD8T细胞功能障碍是由CD8T细胞中TOX的直接基因靶点驱动的。 通过免疫抑制接触和来自HEM细胞的旁分泌信号。这项提案的目标是界定 CD8 T细胞功能障碍的机制可能通过以下目的改善ICI：在目标1中， 将利用CD8T细胞在体外确定TOX的直接基因靶点及其在CD8T细胞功能障碍中的作用 从OCSCC样本中分离出来。在目标2中，患者来源的CD8 T细胞将与患者匹配的患者共同培养 表达模型抗原的OCSCC肿瘤衍生器官或癌细胞系。在这些共同的文化中，嗯- 将特定的细胞膜和分泌的蛋白质击倒在恶性细胞中，以确定它们在其中的作用 抑制CD8T细胞。这项拟议的研究将为CD8 T细胞功能障碍提供机制方面的见解 OCSCC，从而支持开发新的策略来治疗性地激活CD8T细胞并改善 ICI和OCSCC结果。重要的是，在这次奖学金期间，我将继续从事科学和临床活动 直接指导一个由内科科学家和 癌症和免疫学研究方面的专家。我提议的培训对实现我的目标至关重要 研究癌症和免疫的相互作用，并作为一名独立的、学术的内科医生兼科学家而成熟。

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.