Development of Inhibitors of the Leukemia Fusion Protein CBFbeta-SMMHC

描述（由申请人提供）：编码CBF2 （CBFB）的基因被16号染色体倒置破坏[inv(16)(p13q22)]，与人类约12%的急性髓性白血病相关，导致含有大部分CBF2的融合蛋白融合到平滑肌肌球蛋白重链（SMMHC）的卷曲尾区。CBF2- SMMHC融合蛋白作为CBF功能的主要抑制因子，结合RUNX1并失调正常造血所需的多种基因的表达。目前使用细胞毒性化疗的5年总生存率为45-65%，但对于60岁以上的患者仅为20%。这些数据清楚地表明，能够改善inv(16) AML患者治疗反应的靶向治疗是必不可少的。新出现的文献表明，目前的治疗方法无法治愈癌症可能归因于所谓的癌症干细胞或癌症起始细胞群，它们具有长期的自我更新潜力，并且在复发时可以完全再现肿瘤表型。我们的假设是，抑制CBF2-SMMHC与RUNX1的Runt结构域相互作用的小分子可能是诱导细胞凋亡和/或分化并根除白血病起始细胞群的有效治疗药物，从而获得更好的长期生存。我们已经确定了一种抑制CBF2-SMMHC和RUNX1之间蛋白质相互作用的初始先导。我们建议使用基于结构的药物设计和药物化学方法进一步优化这种先导物的效力。化合物将被测试其缓解CBF2-SMMHC介导的抑制和影响白血病细胞系生长、凋亡和分化的能力。选定的化合物将在具有良好特征的小鼠白血病模型中进行体内测试。正如Aim 1中提出的抑制剂所做的那样，抑制CBF2和CBF2- smmhc功能可能会导致毒性或缺乏疗效，因此我们建议开发选择性CBF2- smmhc而不是CBF2的小分子抑制剂。化合物将被测试其缓解CBF2-SMMHC介导的抑制和影响白血病细胞系生长、凋亡和分化的能力。选定的化合物将在具有良好特征的小鼠白血病模型中进行体内测试。我们假设有效的抑制剂应该特异性地影响表达CBF2-SMMHC的白血病前祖细胞的分化和细胞死亡，而骨髓中正常干细胞和早期祖细胞的改变最小。我们建议研究选定的化合物对体内正常造血的影响。在表达CBF2-SMMHC的敲入小鼠中，将优先研究对正常造血影响很小或没有影响的化合物对减少/根除白血病前祖细胞的影响。

DESCRIPTION (provided by applicant): The gene encoding CBF2 (CBFB) is disrupted by the chromosome 16 inversion [inv(16)(p13q22)], associated with ~12% of acute myeloid leukemia in humans, resulting in a fusion protein containing most of the CBF2 fused to the coiled-coil tail region of smooth muscle myosin heavy chain (SMMHC). The CBF2- SMMHC fusion protein acts as a dominant repressor of CBF function, binding RUNX1 and dysregulating the expression of multiple genes required for normal hematopoiesis. Current treatment utilizing cytotoxic chemotherapy results in 45-65% five year overall survival but only 20% for patients older than 60. These data clearly indicate that targeted therapies that can improve the therapeutic response for inv(16) AML patients is essential. Emerging literature suggests that inability to cure cancers with current therapies may be attributed to a population of so-called cancer stem cells or cancer initiating cells that have long term self-renewal potential and can fully recapitulate tumor phenotype at time of relapse. Our hypothesis is that small molecules that inhibit the interaction of CBF2-SMMHC with the Runt domain of RUNX1 could be effective therapeutic drugs that induce apoptosis and/or differentiation and eradicate the leukemia initiating cell population, thereby achieving better long term survival. We have identified an initial lead that inhibits the protein-protein interaction between CBF2-SMMHC and RUNX1. We are proposing to further optimize the potency of this lead using structure-based drug design and medicinal chemistry approaches. Compounds will be tested for their ability to relieve CBF2-SMMHC mediated repression and affect the growth, apoptosis, and differentiation of leukemia cell lines. Selected compounds will be tested in vivo in a well-characterized mouse model of inv(16) leukemia. It is possible that inhibiting both CBF2 and CBF2-SMMHC function, as the inhibitors proposed in Aim 1 will do, could result in toxicity or a lack of efficacy, therefore we are proposing to develop small molecule inhibitors which are selective for CBF2-SMMHC over CBF2. Compounds will be tested for their ability to relieve CBF2-SMMHC mediated repression and affect the growth, apoptosis, and differentiation of leukemia cell lines. Selected compounds will be tested in vivo in a well-characterized mouse model of inv(16) leukemia. We hypothesize that effective inhibitors should specifically affect differentiation and cell death of pre- leukemic progenitor cells expressing CBF2-SMMHC with minimal alteration of the normal stem and early progenitor cells in the bone marrow. We propose to study the effect of selected compounds in vivo in normal hematopoiesis. Compounds with minimal or no effect on normal hematopoiesis will be prioritized to study their effect on the reduction/eradication of pre-leukemic progenitors in knock-in mice expressing CBF2-SMMHC. PUBLIC HEALTH RELEVANCE: We are proposing to develop small molecule inhibitors of the fusion protein CBF2- SMMHC which is a causative agent in one type of acute myeloid leukemia (AML). Such targeted inhibitors have the potential to improve the long term survival of patients with this leukemia by eliminating the population of leukemia cells which cause relapse to occur. These inhibitors also have the potential to be less toxic than standard chemotherapy, resulting in reduced side effects from treatment.