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.

项目概要/摘要 项目 1：中危 AML 进展和复发的基因组学。本次活动的长远目标 该项目是为了更好地了解有助于进展的遗传和表观遗传事件 中危 AML 患者的复发和复发，并对其进行治疗。中等风险 AML（最常由 DNMT3A 突变引发）可能会产生截然不同的结果，例如 其原因尚不十分清楚。在本提案中，我们将探讨遗传和表观遗传因素 影响白血病前期的祖先克隆如何进展为 AML（即“进展”），然后演化为 缓解后复发（即“复发”）。事实上，所有 AML 样本在呈现时都是克隆异质的， 通常包含源自创始克隆（或其他亚克隆）的一个或多个亚克隆。亚克隆 通常对治疗表现出不同的敏感性，并且抗治疗亚克隆通常会随着新的 不被认为是复发驱动因素的突变。确定表观遗传因素是否也可能是 与亚克隆进化相关，我们使用单细胞 RNA 测序（scRNA- seq），并定义了复发时的转录进化，并检测了复发特异性的差异表达 大量 RNA 测序无法检测到的基因。在本提案中，我们将利用单细胞方法来 更好地了解复发时的亚克隆进化，并评估 DNMT3A 突变对疾病模式的作用 就诊和复发时的基因表达。我们还将确定 DNMT3A 是否引发突变 仅需要创建白血病前“状态”，或者它们是否也与维持完全相关 转化的 AML 细胞。这两个目标的研究可能会改善 AML 的风险评估和治疗： 具体目标 1：我们将定义导致克隆进化和复发的事件 中危 AML 患者。我们将进行增强型全基因组测序 (eWGS) 和 scRNA- 对来自中等风险 AML 样本的匹配呈现样本和复发样本进行 seq，这将使我们能够 估算亚克隆的表达特征、它们在复发时如何进展，并识别基因和/或 在显性复发亚克隆中通常失调的途径。 具体目标 2：我们将定义 DNMT3A 突变在 AML 启动和维持中的作用。我们 已经产生了具有可诱导的 WT DNMT3A 转基因的 Dnmt3a 缺陷小鼠（“Dnmt3a null-3A addback” 小鼠），可以准确地对移植的骨髓细胞的基因组进行再甲基化。我们将生成类似的 具有条件性 Dnmt3aR878H 突变的 addback 小鼠，定义其 DNA 甲基化表型，以及 通过 DNMT3A 修复表征其再甲基化动力学和准确性。我们将创造各种 两种模型中的协同突变导致 AML，然后确定这些 AML 是否可以发挥作用 通过恢复 DNMT3A 表达来改变生长和/或分化。这些研究将为临床前研究提供信息 试验将研究针对人类 AML 细胞中 DNMT3AR882H 突变的药物的效果。