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The role and mechanism of METTL3/METTL14-mediated RNA modification in the pathogenesis and drug-resistance of AML

METTL3/METTL14介导的RNA修饰在AML发病及耐药中的作用及机制

基本信息

批准号：
10329928
负责人：
Jianjun Chen
金额：
$ 45.07万
依托单位：
BECKMAN RESEARCH INSTITUTE/CITY OF HOPE
依托单位国家：
美国
项目类别：
财政年份：
2019
资助国家：
美国
起止时间：
2019-02-01 至 2024-01-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10329928
关键词：
11q23 Acute Myelocytic Leukemia Address Affect Anthracycline Ara-C Basic Science Biological Process Biology Bone Marrow Transplantation Cell Differentiation process Cells Chemoresistance Complex Coupled Cytarabine Cytogenetics Data Data Set Daunorubicin Deposition Development Disease Disease remission Drug resistance FLT3 gene Frequencies Genes Goals Hematopoietic Hematopoietic Neoplasms Hematopoietic stem cells Human In Vitro Knock-in Mouse Knockout Mice Lead Leukemic Cell Life Light MLL-AF9 MLL-rearranged leukemia Maintenance Malignant Neoplasms Mediating Messenger RNA Methyltransferase Modeling Modification Molecular Mus Mutation NPM1 gene Neoadjuvant Therapy Oncogenic Pathogenesis Pathologic Pathway interactions Patients Plant Roots Play Post-Transcriptional Regulation Prognosis Proteins RNA RNA Stability Reader Refractory Regulation Relapse Reporting Research Design Resistance Role Stress Therapeutic Time Tissues Transgenic Organisms Translational Research Translations Transplant Recipients Treatment Failure Validation Xenograft procedure acute myeloid leukemia cell bioprocess cancer type chemotherapy crosslinking and immunoprecipitation sequencing drug relapse in vivo insight leukemia loss of function mouse model mutant novel novel therapeutic intervention novel therapeutics overexpression response ribosome profiling self-renewal side effect stem stem cell self renewal stem-like cell success transcriptome transcriptome sequencing transplant model validation studies virtual

项目摘要

PROJECT SUMMARY (ABSTRACT): Background: N6-methyladenosine (m6A) modification is the most abundant internal modification in eukaryotic messenger RNAs (mRNAs) and plays critical roles in various normal bioprocesses. Evidence is emerging that m6A and the associated machinery also play roles in cancers. Acute myeloid leukemia (AML) is one of the most common and fatal forms of hematopoietic malignancies with distinct cytogenetic (e.g., MLL-rearranged) and molecular (e.g., mutations in FLT3 and NPM1) abnormalities. Over 70% of AML patients cannot be cured by contemporary therapies due to drug resistance and relapse. The leukemia stem/initiating cells (LSCs/LICs) are considered to be the root cause for the treatment failure and relapse of AML. Thus, it is critical to better understand the molecular mechanisms underlying AML pathogenesis, LSC/LIC self-renewal and drug resistance, which may lead to the development of effective novel therapeutic strategies to treat AML. RNA m6A modification is deposited by the METTL3-METTL14 methyltransferase complex (MTC; i.e., writer), and may affect the stability and/or translation efficiency of target mRNAs. We and others have reported that both METTL3 and METTL14 play oncogenic roles in MLL-rearranged AML, but their definitive roles in the development and maintenance of other (i.e., non-MLL- rearranged) subtypes of AMLs and especially in LSC/LIC self-renewal remain elusive. Our unpublished preliminary data imply that METTL3 and METTL14 may also play roles in drug resistance in AML, which warrants further systematic studies. Furthermore, their critical targets in AMLs have yet to be well investigated. Hypothesis: METTL3 and METTL14 play essential oncogenic roles in AML pathogenesis, LSC/LIC maintenance and drug resistance through m6A-dependent regulation of expression of critical target genes. Specific Aims: 1) To determine the roles of METTL3 and METTL14 in the development and maintenance of different subtypes of AMLs, as well as in the self-renewal of relevant LSCs/LICs and normal HSCs; 2) To determine the roles of METTL3 and METTL14 in drug resistance of AMLs; and 3) To decipher the molecular mechanisms underlying the pathological roles of METTL3 and METTL14 in AML development/maintenance, LSC/LIC self-renewal and drug resistance. Study Design: 1) We will use inducible Mettl3 and Mettl14 knockout mouse models coupled with mouse AML models, as well as patient-derived xeno-transplantation (PDX) models, to investigate the roles of METTL3 and METTL14 in development/maintenance of different subtypes of AMLs and self-renewal of LSCs/ LICs and normal hematopoietic stem cells (HSCs). 2) We will conduct both gain- and loss-of-function in vivo studies to determine whether METTL3 and METTL14 play critical roles in drug resistance of AMLs. 3) We will perform PAR- CLIP-seq, m6A-seq, RNA-seq, RNA life-time profiling-seq, and ribosome profiling-seq to identify all direct targets of METTL3 and METTL14 in AMLs and to understand how METTL3 and METTL14 regulate expression of their target RNAs, followed by validation and functional studies of a set of top candidate targets.

项目总结（摘要）：背景：N6-甲基腺苷（m6 A）修饰是细胞内最丰富的内部修饰，真核生物的信使RNA（mRNA），并在各种正常的生物过程中发挥关键作用。证据 m6 A和相关机制也在癌症中发挥作用。急性髓细胞白血病（AML）是一种最常见和致命的具有不同细胞遗传学的造血系统恶性肿瘤（例如，MLL重排）和分子（例如，FLT 3和NPM 1突变）异常。超过70%的AML患者无法通过由于耐药性和复发而导致的当代疗法。白血病干细胞/起始细胞（LSC/LIC）被认为是AML治疗失败和复发的根本原因。因此，更好地了解 AML发病机制的分子机制，LSC/LIC自我更新和耐药性，这可能导致开发有效的新型治疗策略来治疗AML。RNA m6 A修饰由胃L3-L14甲基转移酶复合物（MTC;即，作者），并可能影响稳定性和/或翻译靶mRNA的效率。我们和其他人已经报道了胃L3和胃L14都具有致癌作用在MLL重排的AML中，但它们在其他（即，非MLL- 重排的）AML亚型，特别是LSC/LIC自我更新中的AML亚型仍然难以捉摸。我们未发表初步数据表明，胃L3和胃L14也可能在AML的耐药性中发挥作用，进一步系统研究。此外，它们在AML中的关键靶点尚未得到很好的研究。假设：CXL 3和CXL 14在AML发病机制中发挥重要的致癌作用，LSC/LIC 通过m6 A依赖性调节关键靶基因的表达来维持和耐药性。具体目的：1）确定胃L3和胃L14在发育和维持中的作用不同AML亚型的自我更新，以及相关LSC/LIC和正常HSC的自我更新; 2）明确胃L3和胃L14在AML耐药中的作用; 3）破译AML耐药的分子机制，在AML发展/维持中，胃L3和胃L14的病理作用的潜在机制， LSC/LIC自我更新和耐药性。研究设计：1）我们将使用诱导型Mettl 3和Mettl 14敲除小鼠模型， AML模型以及患者来源的异种移植（PDX）模型，以研究胃L3的作用和胃L14在AML的不同亚型的发展/维持和LSC/LIC的自我更新中的作用，正常造血干细胞（HSC）。2)我们将进行功能获得和丧失的体内研究，确定胃L3和胃L14是否在AML耐药中起关键作用。3)我们将执行PAR- CLIP-seq、m6 A-seq、RNA-seq、RNA寿命分析-seq和核糖体分析-seq，用于鉴定所有直接靶标了解AML中胃L3和胃L14的表达，并了解胃L3和胃L14如何调节其表达，目标RNA，随后是一组顶级候选靶标的验证和功能研究。