Targeting Quiescence Pathways to Overcome Chemoresistance in EVI1-Overexpressing Leukemia

项目摘要/摘要 急性髓系白血病(AML)是一种以异常过度增殖为特征的侵袭性白血病 骨髓(BM)中的髓系祖细胞，5年存活率不到25%。 癌基因EVI1在AML中的过度表达与较短的生存期相关 持续时间和较高的复发率。与AML相关的复发是多因素的，先前的研究表明 细胞周期停滞的激活在化疗期间保护AML亚克隆，导致其 生死存亡。鉴于EVI1过度表达AML的严重性，缺乏对EVI1过度表达AML的作用的深入了解 EVI1在AML患者生存期较短和复发率较高的情况下，目前治疗的不足 治疗策略，我们的目标是更好地了解EVI1相关的化疗耐药与长期- 为这种严重的急性髓细胞白血病开发新疗法的长期目标。我们的初步研究使用了EVI1- 过表达AML小鼠模型和患者样本表明EVI1过表达激活静止 小路。我们的CHIP-SEQ和ATAC-SEQ数据揭示了EVI1诱导的静止的机制包括 两条途径：1)细胞周期蛋白依赖性激酶抑制因子1C(CDKN1C/p57Kip2)的上调 造血干细胞静止，其表达与AML复发有关；以及2) 激活富含嘌呤的盒结合蛋白1(PU1)，它是骨髓生成的主要调节因子，足以 触发造血干细胞(HSCs)的细胞周期停滞。因此，我们假设EVI1过度表达 通过CDKN1C和PU.1通路激活静止期，从而保护AML细胞免受化疗的影响。 为了验证我们的假设，我们将阐明EVI1诱导CDKN1C表达的机制及其在 静默(Aim 1)，并研究PU1激活在EVI1相关静默(Aim 2)中的作用。 过度表达急性髓系白血病。这将通过集成来自RNA-SEQ、CHIP-SEQ、ATAC-SEQ和 从我们的体外和体内模型以及从 主要的人类急性髓系白血病样本。将拟议的机械论见解转化为临床环境，并 治疗策略，我们将在临床前研究中使用EVI1过表达来测试新的治疗方案 AML患者来源的异种移植(PDX)模型(AIM 3)。综上所述，拟议的工作将集中于调查 EVI1诱导的静止机制，其发现不仅有助于解释较短的存活时间 与EVI1过表达白血病相关的较高复发率，但也揭示了新的治疗方法 重新激活细胞周期并改善EVI1过表达AML患者生存的策略。

PROJECT SUMMARY/ABSTRACT Acute myeloid leukemia (AML), an aggressive leukemia characterized by the excessive proliferation of abnormal myeloid progenitor cells in the bone marrow (BM), and carries a 5-year survival rate less than 25%. Overexpression of the oncogene ecotropic viral integration site 1 (EVI1) in AML is associated with shorter survival durations and higher relapse rates. AML-associated relapse is multifactorial, and previous studies have shown that the activation of cell cycle quiescence protects AML subclones during chemotherapy, resulting in their survival. Given the severity of EVI1-overexpressing AML, the lack of an in-depth understanding of the role of EVI1 in AML patients’ shorter survival durations and higher relapse rates, and the inadequacy of current therapeutic strategies, we aim to gain a better understanding of EVI1-associated chemoresistance with the long- term goal of developing novel therapies for this sever form of AML. Our preliminary studies using an EVI1- overexpressing AML mouse model and patient samples indicate that EVI1 overexpression activates quiescence pathways. Our ChIP-seq and ATAC-seq data revealed that the mechanism of EVI1-induced quiescence involves two pathways: 1) the upregulation of cyclin-dependent kinase inhibitor 1C (CDKN1C/P57kip2), a critical activator of hematopoietic stem cell quiescence, whose expression has been linked with AML relapse; and 2) the activation of purine-rich box binding protein 1 (PU.1), a master regulator of myelopoiesis, which is sufficient to trigger cell cycle quiescence in hematopoietic stem cells (HSCs). Thus, we hypothesize that EVI1 overexpression protects AML cells from chemotherapy by activating quiescence through CDKN1C and PU.1 pathways. To test our hypothesis, we will elucidate the mechanism of EVI1-induced CDKN1C expression and its role in quiescence (Aim 1), and investigate the role of PU.1 activation in EVI1-associated quiescence (Aim 2) in EVI1- overexpressing AML. This will be accomplished by integrating data from RNA-seq, ChIP-seq, ATAC-seq, and other techniques to analyze EVI1-overexpressing leukemia cells from our in vitro and in vivo models and from primary human AML samples. To translate the proposed mechanistic insights into clinical settings and therapeutic strategies, we will test new treatment regiments in preclinical studies using EVI1-overexpressing AML patient-derived xenograft (PDX) models (Aim 3). In summary, the proposed work will focus on investigating EVI1-induced quiescence mechanisms, and its findings will not only help explain the shorter survival durations and higher relapse rates associated with EVI1-overexpressing leukemia but also unveil new therapeutic strategies that reactivate the cell cycle and improve the survival of patients with EVI1-overexpressing AML.