PARP1-Chromatin and NAD-Metabolism in EBV Epithelial Cancers

EBV 上皮癌中的 PARP1-染色质和 NAD-代谢

• 批准号: 10627691
• 负责人: Italo Tempera
• 金额: $ 47.45万
• 依托单位: WISTAR INSTITUTE
• 依托单位国家: 美国
• 财政年份: 2023
• 资助国家: 美国
• 起止时间: 2023-05-11 至 2028-04-30
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10627691
• 关键词: 3-Dimensional; Benign; Carcinoma; Cells; Cessation of life; Chromatin; Chromatin Loop; Chromatin Structure; Chromosomes; Collaborations; Complex; CpG Island Methylator Phenotype; DNA; DNA Damage; DNA Methylation; DNA Repair; Data; Dependence; Development; Drug Targeting; Enzymes; Epigenetic Process; Episome; Epithelial Cells; Epithelium; Epstein-Barr Virus Infections; Epstein-Barr Virus latency; Epstein-Barr Virus-Related Malignant Neoplasm; Gene Expression; Gene Expression Regulation; Genes; Genome; Goals; Human Herpesvirus 4; In Vitro; Link; Location; MEKs; Maintenance; Malignant Neoplasms; Mediating; Metabolic; Metabolic Control; Metabolism; Molecular Conformation; Mus; Mutation; NADH; Nasopharynx Carcinoma; Oncogenic; Organoids; Outcome; PIK3CG gene; Pathway interactions; Pharmaceutical Preparations; Play; Poly(ADP-ribose) Polymerase Inhibitor; Poly(ADP-ribose) Polymerases; Pre-Clinical Model; Production; Proteins; Reader; Recycling; Regulation; Role; Signal Transduction; Site; Stomach Carcinoma; Testing; Viral; Virus Diseases; Virus Latency; base; cancer cell; cytotoxicity; enzyme activity; epigenome; gastric cancer cell; genome integrity; in vivo; inhibitor; latent gene expression; malignant phenotype; malignant stomach neoplasm; methylation pattern; novel; programs; response; small molecule; synergism; therapeutically effective; three dimensional structure; tumor; tumorigenesis

PROJECT 2 – PROJECT SUMMARY EBV(+) epithelial cancers represent 75% of all the EBV(+) malignancies. Despite the presence of virus infection, these EBV(+) tumors receive the same treatment as EBV(-) cancers. The long-term goal of this Program Project is to identify specific targetable mechanisms of EBV-mediated oncogenesis in epithelial cells. We previously showed that EBV latency and oncogenicity are regulated by factors that link epigenetics with metabolism, such Poly-ADP-ribose Polymerases (PARPs). Drugs that target PARPs have the potential to be effective therapeutic options for EBV(+) tumors. However, our incomplete understanding of the epigenetic and metabolic mechanisms regulating EBV latency in epithelial cells limits the application of such drug options to treat EBV(+) epithelial malignancies. In this project (Project 2), we investigated the role of PARP1 in regulating EBV genome maintenance, gene expression, and metabolic sensing in epithelial cells. We found that PARP1 forms a complex with CTCF on the EBV genome. We now show that this complex includes UHRF1, an epigenetic reader protein involved in DNA methylation pattern propagation. We found that treatment of EBV(+) cells with PARP1 inhibitors disrupts this complex altering both viral and cellular gene expression and causing global DNA hypomethylation. Consistently, PARP inhibitors synergized with DNA hypomethylating agents to elicit cytotoxicity in EBV+ gastric cancers both in vitro and in vivo. Based on these data, we hypothesize that in EBV+ epithelial cells, PARP1, by interacting with CTCF and UHRF1 and modulating their functions, is a key epigenetic factor that connects 3D chromatin conformation and DNA methylation to enable EBV latency and EBV-driven epithelial cell oncogenesis .We base our hypothesis on our preliminary data showing that EBV+ epithelial cells hyper-activate PARP1 and that this drives changes in the 3D conformation of the EBV genome supporting viral latent gene expression. We observed that multiple mechanisms for hyper-activation of PARP1 exist in EBV+ cells, including regulation of the ERK/MEK pathway. We observed that differences exist in NAD+/NADH ration between EBV+ and EBV- epithelial cells, indicating that EBV+ cells are metabolically equipped to sustain PARP1 activation. This observation is consistent with our observation that EBV+ cells are sensitive to inhibitors of NAD salvage pathways, indicating that NAD metabolism plays a key and underappreciated role in EBV -driven oncogenesis in epithelial cancer cells. Programmatic Interactions. This project is highly integrated with other projects and cores. Collaborations with Project 1 on the effect of PARP1 and NAD metabolism on EBNA1 functions; and with Project 3 on the relevance of the PARP1/CTCF/UHRF1 complex for CIMP reversal and PI3K blockade. We have engaged all three cores: Core B for the development of novel small molecule PARP1 degraders, Core D for analysis of chromosome conformations and gene regulation, and Core A and C for analysis of EBV epithelial tumor response in mouse and organoids, and production of site-directed mutations in EBV.

项目 2 – 项目摘要 EBV(+) 上皮癌占所有 EBV(+) 恶性肿瘤的 75%。尽管存在病毒感染， 这些 EBV(+) 肿瘤接受与 EBV(-) 癌症相同的治疗。本计划项目的长期目标 目的是确定上皮细胞中 EBV 介导的肿瘤发生的特定靶向机制。我们之前 研究表明，EBV 潜伏期和致癌性受到表观遗传学与代谢相关因素的调节，例如 聚 ADP 核糖聚合酶 (PARP)。靶向 PARP 的药物有可能成为有效的治疗药物 EBV(+) 肿瘤的选择。然而，我们对表观遗传和代谢机制的了解还不完全 调节上皮细胞中的 EBV 潜伏期限制了此类药物在治疗 EBV(+) 上皮细胞中的应用 恶性肿瘤。 在这个项目（项目2）中，我们研究了PARP1在调节EBV基因组维护、基因 上皮细胞中的表达和代谢传感。我们发现 PARP1 与 CTCF 在 EB 病毒基因组。我们现在证明该复合物包含 UHRF1，一种参与 DNA 的表观遗传读取蛋白 甲基化模式传播。我们发现用 PARP1 抑制剂治疗 EBV(+) 细胞会破坏这种现象 复合物改变病毒和细胞基因表达并导致整体 DNA 低甲基化。一贯地， PARP 抑制剂与 DNA 低甲基化剂协同作用，对 EBV+ 胃癌产生细胞毒性 体外和体内。基于这些数据，我们假设在 EBV+ 上皮细胞中，PARP1 通过相互作用 与 CTCF 和 UHRF1 结合并调节其功能，是连接 3D 染色质的关键表观遗传因子 构象和 DNA 甲基化使 EBV 潜伏期和 EBV 驱动的上皮细胞肿瘤发生成为可能。我们基于 我们对初步数据的假设表明 EBV+ 上皮细胞过度激活 PARP1，并且这 驱动 EBV 基因组 3D 构象的变化，支持病毒潜伏基因的表达。我们观察到 EBV+ 细胞中存在多种 PARP1 过度激活机制，包括 ERK/MEK 的调节 途径。我们观察到 EBV+ 和 EBV- 上皮细胞之间的 NAD+/NADH 比率存在差异， 表明 EBV+ 细胞在代谢上具备维持 PARP1 激活的能力。这个观察结果是一致的 根据我们的观察，EBV+ 细胞对 NAD 挽救途径的抑制剂敏感，表明 NAD 代谢在 EBV 驱动的上皮癌细胞肿瘤发生中发挥着关键但未被充分认识的作用。 程序化交互。该项目与其他项目和核心高度集成。合作伙伴 项目1 PARP1和NAD代谢对EBNA1功能的影响；以及与项目 3 的相关性 PARP1/CTCF/UHRF1 复合物用于 CIMP 逆转和 PI3K 阻断。我们已经调动了所有三个核心： Core B 用于开发新型小分子 PARP1 降解剂，Core D 用于染色体分析 构象和基因调控，以及用于分析小鼠 EBV 上皮肿瘤反应的核心 A 和 C 和类器官，以及 EBV 定点突变的产生。