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）目前是与癌症相关死亡的第三大主要原因 在美国。五年的生存率少于9％，主要归因于困难 PDAC的早期检测和缺乏有效的治疗方法。新颖的免疫疗法，例如免疫 彻底改变其他癌症治疗的检查点封锁未能达到效率 PDAC。这被认为是由于PDAC的免疫抑制微环境限制了 有效免疫疗法所需的效应T细胞浸润和激活。了解如何增加 T细胞激活和渗透任务免疫抑制微环境对于增加 PDAC免疫疗法的功效。 使用胰腺癌的一般工程鼠标模型（“ KPCY”），我们的实验室已证明 源自原发性kpcy肿瘤的克隆可以分为“ T细胞爆发”或“非T细胞爆发” 表型。虽然T细胞增添的肿瘤对联合免疫疗法有反应，但非T细胞增长 肿瘤具有抗性。非T-Cell Inflymed细胞的CRISPR屏幕发现DNMT1，DNA 通过DNA复制维持甲基化标记的甲基转移酶对肿瘤很重要 PDAC的增长。发现在非T细胞增长的肿瘤系中DNMT1的敲除可显着增加T 细胞浸润并减少体内肿瘤的生长。但是，发现T细胞部署可以挽救野生类型 表型。基于对非特异性DNMT抑制剂偶氮丁胺的最新研究，DNMT1的损失为 假设在基因组中被抑制内源性逆转录病毒，导致dsRNA诱导， 激活“病毒模仿”免疫力学。该提案的目标是：（1）确定如何损失 DNMT1促进T细胞依赖性抗肿瘤免疫，（2）评估治疗潜力 与免疫疗法结合使用DNMT1。 检查肿瘤细胞固有的DNMT1抑制T细胞依赖抑制的机制 在肿瘤生长中，我将首先评估内源性逆转录病毒和基因DNA甲基化的变化 与DNMT1丢失引起的病毒模仿途径有关。然后，我将确定dnmt1导线的损失是否 激活病毒模仿途径，以及该途径的抑制是否反应野生型 表型（目标1）。为了确定DNMT1作为目标的翻译潜力，我将首先确定抗肿瘤 DNMT1损失引起的T细胞和肿瘤细胞的变化。我将在使用DNMT1的小鼠中试用免疫疗法 敲除肿瘤以确定对免疫疗法敏感性的变化（AIM 2）。最终，洞察力获得了 从这项研究中，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标的潜力。