JAK/STAT signaling in the pathogenesis of DNMT3A mutant T-ALL

摘要 与其他癌症一样，T细胞急性淋巴细胞性白血病(T-ALL)是由基因积聚引起的 损害未成熟T细胞祖细胞功能的异常。DNMT3A，编码从头DNA 催化在基因组上建立新的DNA甲基化标记的甲基转移酶，是 在10-18%的成人T-ALL病例中反复突变，临床预后较差。我们最近 使用遗传小鼠模型显示，DNMT3A作为T细胞肿瘤抑制因子发挥作用。AN简介 将Notch1突变激活为DNMT3A功能丧失遗传背景(一种常见的遗传组合 患者)产生了致命性T-ALL，与野生型DNMT3A的T-ALL相比，潜伏期缩短了一半。 无论是在体内还是在不同的应激条件下，DNMT3A突变的T-ALL母细胞都能抵抗细胞死亡。这 提示T-ALL细胞中DNMT3A突变的一个主要生物学功能是使它们更难被杀死，以及 这些患者的临床结果较差可能是由于对标准化疗方案耐药所致。 我们现在已经发现了一种潜在的机制来解释这一点，这种机制在DNMT3A-突变体中都是保守的 小鼠T-ALL模型和原始人类DNMT3A突变T-ALL患者样本。DNMT3A-突变体T-ALL 细胞对IL-6和IL-7等细胞因子高度敏感，导致JAK/STAT信号增强 触发支持生存的基因表达程序。具体地说，我们假设DNMT3A突变T- 由于依赖pSTAT5的bclxl表达上调，所有细胞都对凋亡具有抵抗作用。的目标 这项提议是为了了解驱动这些表型的分子机制，并利用它们来开发新的 治疗性干预。我们将通过以下具体目标来检验这一假设； 确定STAT5和bclxl在DNMT3A突变型T-ALL发病机制中的作用 决定了JAK/STAT信号在DNMT3A突变的T-ALL耐药中的作用。 鉴定了支持DNMT3A突变T-ALL功能表型的分子机制。 我们将利用初步的发现，使用互补的组合来验证这一假设 遗传小鼠模型，人类患者来源的异种移植，以及CRSIPR/Cas9基因组工程。在目标1中， 我们将评估STAT5和BCL-XL对DNMT3A的开发和维护的重要性- 突变的T-ALL(包括小鼠和人类)使用最先进的遗传工具。在目标2中，我们将使用基因组 检测以了解DNMT3A突变的T-ALL细胞如何对化疗耐药，以及JAK/STAT 在体内，抑制可以使这些细胞对化疗药物重新敏感。在目标3中，我们将确定 致病基因表达程序中增强子DNA甲基化的重要性 这一T-ALL亚型，并评价SHP-1在赋予DNMT3A-突变体T- 所有牢房。我们将利用这个项目的结果来指导设计合理靶向的精确医学。 基于潜在遗传学的T-ALL患者的治疗策略。

ABSTRACT Like other cancers, T-cell acute lymphoblastic leukemia (T-ALL) arises from the accumulation of genetic abnormalities that impair function of immature T-cell progenitors. DNMT3A, which encodes a de novo DNA methyltransferase enzyme that catalyzes the establishment of new DNA methylation marks on the genome, is recurrently mutated in 10-18% of adult T-ALL cases and confers a poor clinical prognosis. We recently showed using genetic mouse models that Dnmt3a acts as a T-cell tumor suppressor. Introduction of an activating Notch1 mutation into a Dnmt3a loss-of-function genetic background (a common genetic combination in patients) generated a lethal T-ALL with half the latency period compared to T-ALL with wild-type Dnmt3a. Dnmt3a-mutant T-ALL blasts are resistant to cell death, both in vivo and under different stress conditions. This suggests a major biological function of DNMT3A mutations in T-ALL cells is to make them “harder to kill”, and the inferior clinical outcomes of these patients may be due to resistance to standard chemotherapy regimens. We have now uncovered a potential mechanism to explain this which is conserved in both Dnmt3a-mutant mouse T-ALL models and primary human DNMT3A-mutant T-ALL patient samples. DNMT3A-mutant T-ALL cells are hypersensitive to cytokines such as IL-6 and IL-7, which results in elevated JAK/STAT signaling triggering a pro-survival gene expression program. Specifically, we hypothesize that DNMT3A-mutant T- ALL cells are resistant to apoptosis due to pSTAT5-dependent upregulation of BCL-xL. The goals of this proposal are to understand the molecular mechanisms driving these phenotypes and exploit them for novel therapeutic interventions. We will test this hypothesis with the following Specific Aims;  Determine the role of STAT5 and BCL-xL in the pathogenesis of DNMT3A-mutant T-ALL  Define the role of JAK/STAT signaling in chemoresistance of DNMT3A-mutant T-ALL.  Identify molecular mechanisms underpinning the functional phenotypes of Dnmt3a-mutant T-ALL. We will leverage preliminary findings to interrogate this hypothesis using a complementary combination of genetic mouse models, human patient-derived xenografts, and CRSIPR/Cas9 genome engineering. In Aim 1, we will evaluate the importance of STAT5 and BCL-xL for the development and maintenance of DNMT3A- mutant T-ALL (both mouse and human) using state-of-the-art genetic tools. In Aim 2, we will use genomic assays to understand how DNMT3A-mutant T-ALL cells are resistant to chemotherapy, and if JAK/STAT inhibition can resensitize these cells to chemotherapeutic agents in vivo. In Aim 3, we will determine the importance of DNA methylation at enhancers in the generation of pathogenic gene expression programs for this T-ALL subtype, and evaluate the role of SHP-1 in conferring cytokine hypersentivity to DNMT3A-mutant T- ALL cells. We will use the results of this project to inform design of rationally-targeted precision medicine strategies for the treatment of T-ALL patients based on their underlying genetics.