Targeting m6A RNA methylation for glioma therapy

靶向 m6A RNA 甲基化治疗神经胶质瘤

• 批准号: 10264814
• 负责人: Sean T Pianka
• 金额: $ 3.79万
• 依托单位: UNIVERSITY OF CALIFORNIA LOS ANGELES
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-10 至 2022-09-09
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10264814
• 关键词: Acute; Address; Adenosine; Adult; Affect; American; Attenuated; Biological Markers; Biology; Cell Proliferation; Cell physiology; Cells; Characteristics; Clinical; CpG Islands; DNA; Data; Dioxygenases; Disease; Elements; Epigenetic Process; Evaluation; Exhibits; Family; Gene Expression; Gene Expression Regulation; Genetic; Glioma; Goals; Hypermethylation; In Vitro; Isocitrate Dehydrogenase; Knowledge; Link; MGMT gene; Malignant Neoplasms; Malignant neoplasm of brain; Mediating; Messenger RNA; Methodology; Methylation; Modeling; Modification; Molecular; Mus; Mutation; Nuclear Export; Nucleotides; Oncogenic; Outcome; Pathogenesis; Pathway interactions; Patients; Pharmacology; Phenotype; Play; Production; Prognosis; Prognostic Marker; Progression-Free Survivals; Proto-Oncogenes; RNA; RNA Splicing; RNA immunoprecipitation sequencing; RNA methylation; Reproducibility; Resolution; Role; Route; Sampling; Techniques; Testing; Therapeutic Intervention; Transcript; Transcription Process; Translating; Translations; Work; Xenograft procedure; alpha ketoglutarate; base; clinical efficacy; demethylation; efficacious treatment; epitranscriptome; epitranscriptomics; field study; improved; improved outcome; in vivo; insight; jun Oncogene; mRNA Stability; molecular phenotype; novel therapeutic intervention; novel therapeutics; personalized medicine; personalized therapeutic; profiles in patients; prognostic; promoter; stem cells; targeted treatment; therapeutic development; transcriptome sequencing; transcriptomics; treatment response; tumor; tumor growth

PROJECT SUMMARY/ABSTRACT Glioma is the most common primary adult brain cancer, affecting over 20,000 Americans each year. Though past molecular characterizations defined glioma subtypes exhibiting different disease trajectories and treatment responses, there remains a paucity of personalized therapeutic approaches. The goal of this study is to delineate and exploit a putative mechanism of decreased malignancy governed by RNA hypermethylation. Gliomas exhibiting characteristic mutations in isocitrate dehydrogenase 1 (IDHMUT) grow significantly slower than IDHWT tumors. This confers a clear clinical advantage. IDHMUT disrupts normal IDH1 enzymatic activity, leading to the production of an oncometabolite called D-2-hydroxyglutarate (D2HG). D2HG directly inhibits α- ketoglutarate-dependent dioxygenases involved in a range of cellular functions, including demethylation of DNA and RNA. D2HG-mediated inhibition of TET-family DNA demethylases leads to hypermethylated DNA profiles in IDHMUT gliomas, with the degree of methylation being positively associated with improved survival. FTO and AlkBH5 are the only two known RNA demethylases, and both are sensitive to D2HG-mediated inhibition. However, RNA hypermethylation has not been previously investigated in glioma. This proposal aims to perform the first `epitranscriptomic' profiling of IDHMUT and IDHWT gliomas, evaluating mRNA methylation in patient tumor samples and patient-derived glioma stem cells. RNA methylation occurs most frequently in the form of N6-methyladenosine (m6A), and is linked to a number of post-transcriptional processes including mRNA stability, intracellular localization, and translation. I will employ a recently developed methodology known as m6A RNA immunoprecipitation sequencing (MeRIP-Seq) to study m6A abundance and localization with transcript-level resolution. MeRIP-Seq is a modified version of RNA-Seq, and I will utilize both techniques to identify m6A-enriched transcripts and quantify overall transcript abundances using these respective approaches. This will allow me to investigate whether IDHMUT gliomas exhibit a reproducible set of m6A- enriched transcripts, and evaluate the effect of m6A enrichment on transcript stability. I will also assess m6A profiles in patient tumors as a biomarker of improved outcomes. To elucidate whether m6A hypermethylation is a key factor in reducing cell proliferation as seen in the clinically favored IDHMUT subtype, I will inhibit FTO in IDHWT glioma cells and determine if inhibition recapitulates the IDHMUT m6A hypermethylation phenotype, decreases tumor proliferation, and increases overall survival. Taken together, this work will provide insight into the mechanism of m6A-mediated gene regulation and malignancy in glioma, while evaluating the clinical efficacy of FTO inhibition as a novel therapeutic approach.

项目总结/摘要 胶质瘤是最常见的原发性成人脑癌，每年影响超过20，000名美国人。虽然 过去的分子表征定义了表现出不同疾病轨迹的胶质瘤亚型， 尽管治疗反应不佳，但仍然缺乏个性化的治疗方法。本研究的目的是 描绘和开发一个假定的机制，减少恶性肿瘤的RNA超甲基化。 异柠檬酸脱氢酶1（IDHMUT）特征性突变的胶质瘤生长明显减慢 IDHWT肿瘤。这具有明显的临床优势。IDHMUT破坏正常的IDH 1酶活性， 导致产生称为D-2-羟基戊二酸（D2 HG）的肿瘤代谢物。D2 HG直接抑制α- 酮戊二酸依赖性双加氧酶参与一系列细胞功能，包括 DNA和RNA。D2 HG介导的TET家族DNA去甲基化酶抑制导致DNA高甲基化 在IDHMUT胶质瘤中，甲基化程度与生存率的提高呈正相关。 FTO和AlkBH 5是仅有的两种已知的RNA去甲基化酶，并且两者都对D2 HG介导的RNA去甲基化酶敏感。 抑制作用然而，RNA高甲基化以前没有在胶质瘤中进行过研究。这项建议旨在 进行IDHMUT和IDHWT胶质瘤的首次“表观转录组学”分析，评估IDHMUT和IDHWT胶质瘤中mRNA的甲基化。 患者肿瘤样品和患者来源的神经胶质瘤干细胞。RNA甲基化最常发生在 N6-甲基腺苷（m6 A）的形式，并与许多转录后过程，包括 mRNA稳定性、细胞内定位和翻译。我将采用一种新的方法 称为m6 A RNA免疫沉淀测序（MeRIP-Seq），以研究m6 A丰度和定位 具有转录水平的分辨率。MeRIP-Seq是RNA-Seq的改良版，我将同时使用这两种技术 为了鉴定富含m6 A的转录物并使用这些相应的转录物定量总体转录物丰度， 接近。这将使我能够研究IDHMUT胶质瘤是否表现出一组可重复的m6 A- 富集的转录物，并评估m6 A富集对转录物稳定性的影响。我还将评估m6 a 作为改善结果的生物标志物。为了阐明m6 A高甲基化是否是 如在临床上有利的IDHMUT亚型中所见，FTO是减少细胞增殖的关键因素， IDHWT神经胶质瘤细胞，并确定抑制是否重演IDHMUT m6 A超甲基化表型， 降低肿瘤增殖并增加总存活率。总之，这项工作将提供深入了解 m6 A介导的基因调控机制与胶质瘤恶性程度的关系，同时评价其临床应用价值。 FTO抑制作为一种新的治疗方法的功效。