Identifying the Role of Tumor Cell Intrinsic DNMT1 in Anti-Tumor Immunity in Pancreatic Ductal Adenocarcinoma

确定肿瘤细胞内在 DNMT1 在胰管腺癌抗肿瘤免疫中的作用

• 批准号: 10577787
• 负责人: Erin Elizabeth Hollander
• 金额: $ 3.54万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2021
• 资助国家: 美国
• 起止时间: 2021-03-01 至 2025-02-28
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10577787
• 关键词: Azacitidine; Biological Assay; CD8-Positive T-Lymphocytes; CRISPR screen; Cancer Etiology; Cell Survival; Cell physiology; Cells; Cessation of life; Clone Cells; Coculture Techniques; Combination immunotherapy; DNA Methylation; DNA Modification Methylases; DNA biosynthesis; DNA methyltransferase inhibition; Data; Double-Stranded RNA; Early Diagnosis; Elements; Endogenous Retroviruses; Epigenetic Process; Exposure to; Flow Cytometry; Genes; Genetic Transcription; Genetically Engineered Mouse; Genome; Goals; Growth; Immune; Immune system; Immunocompetent; Immunosuppression; Immunotherapy; Infiltration; Injections; KPC model; Knock-out; Libraries; Maintenance; Malignant Neoplasms; Malignant neoplasm of pancreas; Measures; Methylation; Mus; Pancreatic Ductal Adenocarcinoma; Pathway interactions; Patients; Phenotype; Primary Neoplasm; Refractory; Regimen; Reporting; Resistance; Role; Survival Rate; T cell infiltration; T-Cell Activation; T-Cell Depletion; T-Lymphocyte; T-cell inflamed; Therapeutic; Tumor Immunity; Tumor Suppression; Tumor-infiltrating immune cells; United States; Viral; Virus Activation; Western Blotting; bisulfite sequencing; cell motility; comparative efficacy; derepression; effective therapy; effector T cell; experimental study; gene repression; immune cell infiltrate; immune checkpoint blockade; immunoregulation; immunotherapy trials; in vivo; inhibitor; insight; methylation pattern; mimicry; mouse model; neoplastic cell; new therapeutic target; novel; overexpression; pancreatic cancer cells; pancreatic ductal adenocarcinoma cell; pancreatic ductal adenocarcinoma model; pharmacologic; release factor; response; sensor; subcutaneous; therapeutic target; tool; transcriptome sequencing; translational potential; tumor; tumor growth; tumor microenvironment; tumor-immune system interactions

Project Summary Pancreatic ductal adenocarcinoma (PDAC) is currently the third leading cause of cancer-related death in the United States. The five-year survival rate of less than nine percent is attributed mainly to a difficulty in early detection and a lack of effective treatments for PDAC. Novel immunotherapies such as immune checkpoint blockade which have revolutionized treatment of other cancers have failed to achieve efficacy in PDAC. This is thought to be due to the immunosuppressive microenvironment of PDAC which limits the effector T cell infiltration and activation necessary for effective immunotherapy. Understanding how to increase T cell activation and infiltration despite an immunosuppressive microenvironment is essential to increasing the efficacy of immunotherapies in PDAC. Using a genetically engineered mouse model (“KPCY”) of pancreatic cancer, our lab has demonstrated that clones derived from primary KPCY tumors can be divided into “T-cell-inflamed” or “non-T-cell-inflamed” phenotypes. While T-cell-inflamed tumors are responsive to combination immunotherapy, non-T-cell-inflamed tumors are resistant. A CRISPR screen of non-T-cell-inflamed cells found that DNMT1, a DNA methyltransferase involved in maintaining methylation marks through DNA replication, is important for tumor growth in PDAC. Knockout of DNMT1 in a non-T-cell-inflamed tumor line was found to significantly increase T cell infiltration and decrease tumor growth in vivo. T cell depletion, however, was found to rescue the wild type phenotype. Based on recent studies of the non-specific DNMT inhibitor azacytidine, loss of DNMT1 is hypothesized to de-repress endogenous retroviruses in the genome, leading to dsRNA induction which activates a “viral mimicry” immune mechanism. The goals of this proposal are: (1) to determine how loss of DNMT1 promotes T-cell dependent anti-tumor immunity, and (2) to assess the therapeutic potential of targeting DNMT1 in conjunction with immunotherapy. To examine the mechanism of tumor cell intrinsic DNMT1 in inhibition of T-cell dependent suppression of tumor growth, I will first assess the changes in DNA methylation at endogenous retroviruses and genes related to the viral mimicry pathway provoked by loss of DNMT1. I will then determine if loss of DNMT1 leads to activation of the viral mimicry pathway, and whether inhibition of this pathway rescues the wild type phenotype (Aim 1). To determine the translational potential of DNMT1 as a target, I will first identify anti-tumor changes in T cells and tumor cells provoked by loss of DNMT1. I will trial immunotherapy in mice with DNMT1 knockout tumors to determine changes in sensitivity to immunotherapy (Aim 2). Ultimately, the insights gained from this study will provide us with a better understanding of the role of DNMT1 in regulating the PDAC immune microenvironment and its potential as a therapeutic target.

项目摘要 胰腺导管腺癌(PDAC)是目前癌症相关死亡的第三大原因。 在美国。不到9%的五年存活率主要归因于 早期发现，缺乏有效的治疗方法。新的免疫疗法，如免疫 检查站封锁使其他癌症的治疗发生了革命性的变化，但在 PDAC。这被认为是由于PDAC的免疫抑制微环境限制了 有效的免疫治疗所必需的效应T细胞的渗透和激活。了解如何提高 尽管处于免疫抑制的微环境中，T细胞的激活和渗透对于增加T细胞的 免疫治疗在PDAC中的疗效。 使用胰腺癌的基因工程小鼠模型(KPCY)，我们的实验室已经证明 来源于原发KPCY肿瘤的克隆可分为“T细胞炎症”和“非T细胞炎症”。 表型。虽然T细胞炎症的肿瘤对联合免疫治疗有反应，但非T细胞炎症的肿瘤 肿瘤是耐药的。CRISPR对非T细胞炎症细胞的筛查发现，DNMT1，一种DNA 甲基转移酶通过DNA复制参与维持甲基化标记，对肿瘤非常重要 PDAC的增长。在非T细胞炎症的肿瘤系中，DNMT1的敲除被发现显著增加T 体内细胞浸润，减少肿瘤生长。然而，T细胞的耗尽被发现挽救了野生型 表型。根据最近对非特异性DNMT抑制剂氮胞苷的研究，DNMT1的丢失是 假设去抑制基因组中的内源性逆转录病毒，导致dsRNA诱导， 激活一种“病毒模仿”免疫机制。这项建议的目标是：(1)确定损失如何 DNMT1促进T细胞依赖的抗肿瘤免疫，以及(2)评估DNMT1的治疗潜力 靶向DNMT1结合免疫治疗。 肿瘤细胞内源性DNMT1抑制T细胞依赖抑制机制的研究 关于肿瘤的生长，我将首先评估内源性逆转录病毒和基因的DNA甲基化的变化 与DNMT1缺失引起的病毒拟态途径有关。然后我将确定DNMT1导联是否丢失 病毒模拟途径的激活，以及抑制这一途径是否拯救了野生型 表型(目标1)。为了确定DNMT1作为靶点的翻译潜力，我将首先鉴定抗肿瘤 DNMT1缺失引起的T细胞和肿瘤细胞的变化。我将在携带DNMT1的小鼠身上试验免疫疗法 剔除肿瘤以确定免疫治疗敏感性的变化(目标2)。最终，这些洞察获得了 这项研究将使我们更好地了解DNMT1在调节PDAC中的作用 免疫微环境及其作为治疗靶点的潜力。