Project 1 - Genomics of Intermediate-Risk AML Progression and Relapse.

项目 1 - 中危 AML 进展和复发的基因组学。

• 批准号: 10541166
• 负责人: TIMOTHY J. LEY
• 金额: $ 43.14万
• 依托单位: WASHINGTON UNIVERSITY
• 依托单位国家: 美国
• 财政年份: 2003
• 资助国家: 美国
• 起止时间: 2003-09-19 至 2024-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10541166
• 关键词: Acute Myelocytic Leukemia; Address; Back; Bone Marrow Cell Transplantation; Bone Marrow Cells; CRISPR/Cas technology; Cells; Cessation of life; Clonal Evolution; Cryopreservation; DNA; DNA Methylation; DNMT3a; DNMT3a mutation; Data; Disease remission; Dominant-Negative Mutation; Drug Design; Engineering; Engraftment; Epigenetic Process; Event; Evolution; FLT3 gene; Gene Expression; Gene Expression Profile; Genes; Genetic; Genetic Transcription; Genome; Genomics; Goals; Growth; Hematopoiesis; Hematopoietic; Hematopoietic Neoplasms; Human; Immunodeficient Mouse; Individual; KRASG12D; Kinetics; Maintenance; Mediating; Methods; Methylation; Modeling; Molecular; Mus; Mutation; NPM1 gene; Normal tissue morphology; Outcome; Pathogenesis; Pathway interactions; Patients; Pattern; Phenotype; Pilot Projects; Predisposition; Preleukemia; Proteins; Recurrence; Relapse; Risk; Risk Assessment; Role; Sampling; Skin Tissue; TP53 gene; Techniques; Therapeutic; Transgenes; WT1 gene; biomarker identification; cell transformation; differential expression; drug relapse; drug sensitivity; gain of function mutation; genome editing; genome sequencing; high risk; improved; insertion/deletion mutation; mouse model; novel; novel strategies; null mutation; preclinical trial; relapse patients; relapse prediction; relapse risk; restoration; single-cell RNA sequencing; therapy resistant; transcriptome sequencing; transcriptomics; vector; virtual; whole genome

Project Summary/Abstract Project 1: Genomics of intermediate-risk AML progression and relapse. The long-term goal of this project is to better understand the genetic and epigenetic events that contribute to the progression and relapse of patients with intermediate-risk AML, and exploit them therapeutically. Intermediate-risk AML (which is most commonly initiated by DNMT3A mutations) can have vastly different outcomes, for reasons that are still not well understood. In this proposal, we will explore the genetic and epigenetic factors that influence how a pre-leukemic, ancestral clone progresses to AML (i.e. “progression”), and then evolves to recur after a remission (i.e. “relapse”). Virtually all AML samples are clonally heterogeneous at presentation, generally containing one or more subclones derived from a founding clone (or other subclones). Subclones often display different susceptibilities to therapies, and therapy-resistant subclones often evolve with new mutations that are not recognized as drivers at relapse. To determine whether epigenetic factors may also be relevant for subclonal evolution, we have performed pilot studies using single-cell RNA-sequencing (scRNA- seq), and defined transcriptional evolution at relapse, and detected relapse-specific, differentially expressed genes that were not detectable by bulk RNA-sequencing. In this proposal, we will exploit single cell methods to better understand subclonal evolution at relapse, and evaluate the role of DNMT3A mutations for patterns of gene expression at presentation and relapse. We will also determine whether initiating mutations in DNMT3A are required only for creating the preleukemic “state”, or whether they are also relevant for maintaining fully transformed AML cells. The studies of both aims may improve risk assessment and therapeutics for AML: Specific Aim 1: We will define the events that contribute to clonal evolution and relapse in intermediate-risk AML patients. We will perform enhanced whole genome sequencing (eWGS) and scRNA- seq on matched presentation and relapse samples from intermediate-risk AML samples, which will allow us to impute the expression signatures of subclones, how they progress at relapse, and identify genes and/or pathways that are commonly dysregulated in dominant relapse subclones. Specific Aim 2: We will define the role of DNMT3A mutations for AML initiation and maintenance. We have generated Dnmt3a deficient mice with an inducible WT DNMT3A transgene (“Dnmt3a null-3A addback” mice) that can accurately remethylate the genomes of transplanted bone marrow cells. We will generate similar addback mice with a conditional Dnmt3aR878H mutation, define their DNA methylation phenotype, and characterize their remethylation kinetics and accuracy with DNMT3A restoration. We will create a variety of cooperating mutations in both models to cause AML, and then determine whether these AMLs can have their growth and/or differentiation altered by restoring DNMT3A expression. These studies will inform preclinical trials that will study the effects of drugs designed to target the DNMT3AR882H mutation in human AML cells.

项目摘要/摘要 项目1：中等风险AML进展和缓解的基因组学。这个长期目标 项目将更好地了解有助于进展的遗传和表观遗传事件 和中等风险AML患者的继电器，并热探索它们。 AML（最常见的是由DNMT3A突变引发的）可能具有截然不同的结果 原因仍然不太了解。在此提案中，我们将探索遗传和表观遗传因素 这影响了祖先克隆的前美核血症如何发展为AML（即“进展”），然后发展为 缓解后复发（即“复发”）。实际上，所有AML样品在演示时都是克隆的异质性， 通常包含一个或多个源自创始克隆（或其他子克隆）的子克隆。子克隆 通常对治疗表现出不同的敏感性，耐药子克隆通常会随着新的方式而发展 不被公认为驱动因素的突变。确定表观遗传因素是否也可能是 与亚克隆进化相关，我们使用了单细胞RNA-seques（SCRNA- seq），并在浮雕时定义了转录演变，并检测到了继电器特异性，表达不同 无法通过大量RNA测序检测的基因。在此提案中，我们将利用单细胞方法 更好地了解退休时克子的进化，并评估DNMT3A突变的作用 介绍和浮雕中的基因表达。我们还将确定是否启动DNMT3A突变 仅需要创建统计学的“状态”，或者需要与完全保持 转化的AML细胞。两种目标的研究都可以改善AML的风险评估和治疗： 特定目的1：我们将定义有助于克隆进化和缓解的事件 中等风险的AML患者。我们将执行增强的整个基因组测序（EWGS）和SCRNA- SEQ在匹配的演示文稿和中型风险AML样本中的继电器样本中，这将使我们能够 估算亚克隆的表达特征，它们在继电器方面的进展以及识别基因和/或 通常在主要继电器亚克隆中失调的途径。 具体目标2：我们将定义DNMT3A突变在AML启动和维护中的作用。我们 已经用诱导WT DNMT3A变换生成了DNMT3A缺乏的小鼠（“ DNMT3A NULL-3A倒数倒数” 小鼠）可以准确地将移植的骨髓细胞的基因组重新甲基化。我们将生成类似的 具有条件DNMT3AR878H突变的倒数小鼠，定义其DNA甲基化表型，并且 通过DNMT3A恢复来表征它们的脱甲基化动力学和准确性。我们将创建各种各样的 在两个模型中合作突变以引起AML，然后确定这些AML是否可以具有 通过恢复DNMT3A表达改变了生长和/或分化。这些研究将为临床前提供信息 试验将研究旨在针对人类AML细胞中DNMT3AR882H突变的药物的作用。