DISCOVERY OF SMALL MOLECULAR ACTIVATORS OF NR4A ORPHAN NUCLEAR RECEPTORS

描述（由申请人提供）：本申请的总体目标是通过激活孤儿核受体NR4A亚类来鉴定用于治疗急性髓性白血病（AMLs）的新型抗白血病药物。aml主要是骨髓细胞发育正常转录控制被破坏的疾病，导致未成熟细胞的积累和转化白血病起始细胞（LIC）群体的出现，这些细胞具有长期自我更新特性，能够维持白血病的扩展。NR4A1和NR4A3是AML的有效肿瘤抑制因子。无论患者细胞遗传学如何，它们在所有人类AML LICs中都是沉默的。由于造血干细胞（HSC）稳态的破坏和转化的抗辐射LIC的出现，小鼠中的缺失导致极快的出生后AML。此外，NR4A1或NR4A3在人类AML细胞中的急性抢救抑制了它们的增殖，并重新编程了一组基因特征，这些基因特征将所有原代人LICs与正常hsc区分开来，而不管细胞遗传学如何。我们假设NR4A沉默是AML发展的必要步骤，针对其再激活的策略可能在AML治疗中具有普遍的治疗益处。通过我们基于NR4A靶点的基因组学数据与硅化学基因组学筛选的交叉，我们采用了一种新的整合策略，成功地鉴定了AML细胞中重新激活NR4A的小分子，并可用于揭示NR4A沉默的机制。我们这项建议的具体目标是：1)通过分析NR4A在人AML细胞中再激活的细胞和分子后果，进一步验证NR4As作为治疗人AML的新治疗靶点；2)通过分析NR4As小分子激活剂在AML细胞系中消除LICs的功效，测试NR4As在人AML细胞中的抗白血病特性。来自AML患者和人类AML动物模型的原代细胞，并通过比较它们对正常人类造血干细胞的影响来询问它们的AML选择性，以及3)揭示AML细胞中NR4As的沉默和再激活机制。为此，我们将结合小分子激活因子的信息学途径分析和NR4A基因染色质景观的分子分析，以鉴定AML中NR4A沉默的标记和介质，并揭示参与其再激活的信号通路。考虑到NR4As在AML患者中的广泛沉默以及它们在引起AML时的充分失活，我们相信这些研究在新药发现方面的成功结果将很快导致新的临床试验，这将对AML患者的治疗产生重大影响。

DESCRIPTION (provided by applicant): The overall objective of this proposal is to identify novel antileukemic drugs for treatment of acute myeloid leukemias (AMLs) via activation of the NR4A subclass of orphan nuclear receptors. AMLs are primarily diseases of corruption of normal transcriptional control of myeloid cell development leading to accumulation of immature blasts and emergence of a transformed leukemic initiating cell (LIC) population with long term self renewal properties capable of sustaining leukemic expansion. NR4A1 and NR4A3 are potent tumor suppressors of AML. They are silenced in all human AML LICs irrespective of patient cytogenetics. Deletions in mice lead to extremely rapid postnatal AML due to disruption of hematopoietic stem cell (HSC) homeostasis and the emergence of a transformed radio-resistant LIC. Further, acute rescue of NR4A1 or NR4A3 in human AML cells inhibits their proliferation and reprograms a subset of gene signatures that distinguish all primary human LICs from normal HSCs regardless of cytogenetics. We hypothesize that NR4A silencing is an obligate step in AML development and that strategies directed toward their reactivation may be of general therapeutic benefit in treatment of AMLs. By intersection of our NR4A target based genomics data with in silico chemical genomics screening, we have employed a novel integrative strategy to successfully identify small molecules that reactivate NR4As in AML cells and that can be used to disclose mechanisms of NR4A silencing. Our specific aims in this proposal are: 1) to further validate NR4As as novel therapeutic targets for treatment of human AMLs by analysis of the cellular and molecular consequences of NR4A reactivation in human AML cells, 2) to test the anti-leukemic properties of small molecule activators of NR4As in human AML cells by analysis of their efficacy in elimination of LICs using AML cell lines, primary cells from AML patients and animal models of human AML and by interrogation of their AML selectivity by comparison of their effects on normal human HSCs, and 3) to disclose mechanisms of silencing and reactivation of NR4As in AML cells. In this aim, we will combine informatics pathway analysis of small molecule activators with molecular analysis of the chromatin landscape of the NR4A genes to identify marks and mediators of NR4A silencing in AML and to disclose signaling pathways involved in their reactivation. Given the widespread silencing of NR4As in AML patients and the sufficiency of their inactivation in causing AML, we believe that a successful outcome from these studies in terms of new drug discovery will quickly result in new clinical trials that will have major impact on treatment of AML patients.