Function and targeting of a stable transcription factor complex in leukemia

描述（由申请人提供）：转录因子在许多类型的癌症中发挥着关键作用，但长期以来，由于缺乏可被大多数当前药物典型的小分子类型靶向的特定活性位点，转录因子被认为是“不可药物的”。然而，由于不同的相互作用模式可能赋予肽高度的选择性，短肽的使用最近成为一种有希望的策略，以靶向通过特定蛋白质复合物起作用的转录因子。在本提案中，我们寻求使用这种新策略来靶向急性髓系白血病中最常见的白血病融合蛋白/转录因子AML1-ETO。我们发现，在白血病细胞中，AML1- ETO存在于包含多种转录因子和辅助因子的稳定蛋白复合体中。在这个复合体中，二聚化的AML1-ETO直接与一系列传统的转录激活因子E蛋白相互作用，这些转录激活因子与造血谱系发育事件有关。我们还发现，AML1-ETO二聚化结构域（NHR2）利用二聚化α -螺旋的独特表面介导AML1-ETO与E蛋白中保守基序的相互作用，这在之前被证明对白血病发生至关重要。这种特殊的相互作用模式理想地允许设计抑制剂特异性地破坏相互作用并操纵AML1-ETO的活性，从而为白血病治疗提供潜在的靶点。在这方面，进一步支持NHR2- e蛋白相互作用作为治疗靶点的研究表明，一种破坏NHR2- e蛋白相互作用但不破坏NHR2二聚化的特定突变，已被证明会损害AML1-ETO增强人类造血干细胞自我更新的能力。基于对含有aml1 - eto的转录因子/辅因子（AETFC）复合物的生化和功能研究，我们计划(i)通过详细的机制研究进一步鉴定和表征AETFC复合物（es）；（ii）通过全基因组ChIP分析鉴定直接AML1- ETO靶基因；（iii）通过基于细胞和无细胞的体外转录系统，阐明AML1-ETO和其他成分合作（去）调节转录的详细机制；（iv）研究（并验证）单个成分的生物学功能及其在白血病细胞和小鼠模型中的相互作用；(v)设计特异性的拟肽抑制剂来操纵AML1-ETO在转录和白血病发生中的作用。

DESCRIPTION (provided by applicant): Transcription factors play critical roles in many types of cancer but, for a long time, were considered "undruggable" because of the lack of specific active sites that can be targeted by the types of small molecules that typify most current drugs. However, the use of short peptides has recently emerged as a promising strategy to target transcription factors that function through specific protein complexes, since distinct interaction patterns may confer a high degree of selectivity on the peptide. In this proposal, we seek to use this new strategy to target the leukemogenic fusion protein/transcription factor, AML1-ETO, that is most frequently involved in acute myeloid leukemia. We have found that, in leukemic cells, AML1- ETO resides in a stable protein complex containing multiple transcription factors and cofactors. Within this complex, the dimerized AML1-ETO directly interacts with a family of conventional transcriptional activators, E proteins, that are implicated in hematopoietic lineage developmental events. We also have found that the AML1-ETO dimerization domain (NHR2), which previously was shown to be critical for leukemogenesis, utilizes a distinct surface of the dimerized alpha-helixes to mediate the AML1-ETO interaction with a conserved motif in E proteins. This particular interaction pattern ideally allows the design of inhibitors to specifically disrupt the interaction and to manipulate the activities of AML1-ETO, thus providing a potential target for leukemia treatment. In this regard, and in further support of this NHR2-E protein interaction as a therapeutic target, a specific mutation that disrupts the NHR2-E protein interaction, but not NHR2 dimerization, has been shown to impair the capacity of AML1-ETO to enhance human hematopoietic stem cell self-renewal. Based on these biochemical and functional studies of the AML1-ETO-containing transcription factor/cofactor (AETFC) complex(es), we plan (i) to further identify and characterize the AETFC complex(es) by detailed mechanistic studies; (ii) to identify direct AML1- ETO target genes by genome-wide ChIP analyses; (iii) to clarify, through cell-based and cell-free in vitro transcription systems, the detailed mechanisms by which AML1-ETO and other components cooperate to (de)regulate transcription; (iv) to study (and validate) the biological functions of individual components and their interactions in leukemic cellular and mouse models; and (v) to design specific peptidomimetic inhibitors to manipulate the action of AML1-ETO in transcription and leukemogenesis.