Epigenetic control of the metabolic tumor suppressor FBP1 in liver cancer

肝癌代谢肿瘤抑制因子 FBP1 的表观遗传控制

• 批准号: 9758641
• 负责人: Jason Godfrey
• 金额: $ 4.5万
• 依托单位: UNIVERSITY OF PENNSYLVANIA
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-07-01 至 2022-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9758641
• 关键词: Address; Area; Binding; Binding Proteins; Biological; Biological Markers; Cells; Cellular Metabolic Process; ChIP-seq; Chromatin; Clinical; Clustered Regularly Interspaced Short Palindromic Repeats; Complex; Data; Diagnosis; Disease; Enhancers; Enzymes; Epigenetic Process; Fructose; Fructose-1,6-Bisphosphatase; Gene Silencing; Genes; Genetic Heterogeneity; Genetic Transcription; Gluconeogenesis; Glucose; Glycolysis; Goals; Growth; Hepatic; Hepatocyte; Heterochromatin; Histones; Homologous Gene; Laboratories; Liver; Lysine; Malignant Epithelial Cell; Malignant Neoplasms; Malignant neoplasm of liver; Mass Spectrum Analysis; Measures; Mediating; Messenger RNA; Metabolic; Metabolic Control; Metabolism; Methylation; Methyltransferase; Modeling; Molecular; Mus; Mutation; Nature; Oncogenic; Ontology; Pathologic; Pathway Analysis; Pathway interactions; Phenotype; Phosphotransferases; Physiological; Primary carcinoma of the liver cells; Process; Proteins; Regulation; Repression; Research; Resistance; Role; Signal Transduction; Site; TP53 gene; Telomerase; Testing; Therapeutic; Tumor Suppressor Genes; Tumor Suppressor Proteins; Tumorigenicity; Work; Xenograft procedure; advanced disease; aerobic glycolysis; base; beta catenin; cancer type; candidate validation; cellular targeting; combinatorial; experimental study; glucose metabolism; hepatocellular carcinoma cell line; histone methylation; histone methyltransferase; human embryonic stem cell; improved; in vivo; inhibitor/antagonist; mRNA Expression; mass flux; metabolic phenotype; mortality; neoplastic; novel therapeutic intervention; novel therapeutics; overexpression; patient population; promoter; recruit; resistance mechanism; response; small hairpin RNA; targeted treatment; tumor; tumor growth; validation studies

Abstract Hepatocellular carcinoma (HCC) is the primary form of liver cancer and afflicts over half a million people worldwide. Alarmingly, mortality rates have steadily increased in recent years, highlighting the importance of identifying new therapies. One promising area of treating HCC is in targeting cellular metabolism. Specifically, aerobic glycolysis is a common feature of liver cancer, which is an otherwise genetically and pathologically heterogeneous disease. Neoplastic hepatocyte cells attain this glycolytic phenotype in part by decreasing physiological gluconeogenesis. Indeed, the mRNA and protein levels of a rate-limiting gluconeogenic enzyme, fructose-1,6-bisphosphatase 1 (FBP1), are significantly depleted in HCC. Our lab has found that ectopically expressing FBP1 in HCC xenografts reduces tumor growth, whereas deleting Fbp1 accelerates autochthonous models of liver cancer. These results suggest a strong tumor suppressive role for FBP1, and substantiate the therapeutic potential of FBP1 activity. Therefore, my overall goal is to elucidate the mechanism of FBP1 repression to discover new therapeutic strategies for HCC. Three key pieces of preliminary data indicate that the histone 3 lysine 27 (H3K27) methyltransferase, Enhancer of Zeste Homolog 2 (EZH2), contributes to FBP1 suppression: (1) publicly available ChIP-seq data reveals EZH2 enrichment sites at the FBP1 gene locus in human embryonic stem cells. (2) EZH2 mRNA levels progressively increase with HCC stage and negatively correlate with FBP1 expression. (3) Functionally, depleting EZH2 in HCC cells increases FBP1 mRNA, whereas elevating EZH2 expression in normal hepatocytes diminishes FBP1 protein abundance. I hypothesize that EZH2 directly catalyzes H3K27 trimethylation at the FBP1 promoter to suppress its transcription, and inhibiting EZH2 will reduce HCC growth, in part, through metabolic consequences of FBP1 activation. To address these ideas, I will first characterize the physical interaction of EZH2 with the FBP1 locus in HCC by ChIP. Additionally, I will determine how EZH2 is recruited to the FBP1 promoter by performing co-IP and mass spectrometry, followed by validation of candidate binding partners. Second, to determine biological effects relevant to clinical response, I will perform comprehensive metabolite analyses of HCC cells under conditions of EZH2 loss. Importantly, I expect to see both FBP1-dependent and independent alterations in metabolism upon EZH2 reduction, which will have implications for metabolite biomarkers, synthetic lethality, and resistance mechanisms. This work will decipher the complex nature of FBP1 epigenetic silencing in HCC, and outline new treatment paradigms that reactivate gluconeogenesis in HCC.

摘要 肝细胞癌是肝癌的主要形式，有50多万人患有此病。 全世界。令人担忧的是，近年来死亡率稳步上升，突显了 确定新的治疗方法。治疗肝细胞癌的一个有希望的领域是靶向细胞代谢。具体来说， 有氧糖酵解是肝癌的共同特征，它是一种遗传和病理上不同的疾病。 异质性疾病。肿瘤肝细胞获得这种糖酵解表型的部分原因是 生理性的糖异生。事实上，一种限速糖异生酶的mRNA和蛋白质水平， 果糖-1，6-二磷酸酶1(FBP1)在肝细胞癌中显著缺失。我们的实验室发现了这种异位现象 在肝细胞癌移植瘤中表达FBP1可减少肿瘤生长，而缺失FBP1则可加速原位生长 肝癌的模型。这些结果表明FBP1具有很强的肿瘤抑制作用，并证实了 FBP1活性的治疗潜力。因此，我的总体目标是阐明FBP1的作用机制 抑制，以发现新的肝癌治疗策略。三个关键的初步数据表明 组蛋白3赖氨酸27(H3K27)甲基转移酶，Zust Homolog 2(EZH2)的增强子，有助于 FBP1抑制：(1)公开的CHIP-SEQ数据揭示了FBP1基因座位上的EZH2富集点 在人类胚胎干细胞中。(2)EZH2基因表达水平随肝细胞癌分期的增加而逐渐升高，呈负相关。 与FBP1的表达呈正相关。(3)在功能上，在肝癌细胞中耗尽EZH2可增加FBP1基因的表达。 而EZH2在正常肝细胞中的表达增加则会降低FBP1蛋白的丰度。我 假设EZH2直接催化FBP1启动子上的H3K27三甲基化抑制其 转录，并抑制EZH2将减少肝癌的生长，部分通过代谢后果 Fbp1激活。为了解决这些想法，我将首先描述EZH2与 肝细胞癌Fbp1基因座芯片检测此外，我将通过以下方式确定EZH2如何被招募到FBP1启动子 进行共IP和质谱分析，然后验证候选结合伙伴。第二，到 确定与临床反应相关的生物效应，我将进行全面的代谢产物分析 EZH2缺失条件下的肝癌细胞。重要的是，我希望看到依赖于FBP1的和独立的 EZH2还原时代谢的变化，这将对代谢物生物标志物产生影响， 合成杀伤力和抵抗机制。这项工作将破译FBP1表观遗传学的复杂性质 肝细胞癌中的沉默，并概述了重新激活肝细胞癌中糖异生的新治疗范例。