Development of Inhibitors of the Leukemia Fusion Protein CBFbeta-SMMHC

描述(申请人提供)：编码CBF2(Cbfb)的基因被16号染色体倒位[inv(16)(P13q22)]破坏，与人类约12%的急性髓系白血病有关，导致融合蛋白包含大部分CBF2融合到平滑肌肌球蛋白重链(SMMHC)的卷曲尾部区域。CBF2-SMMHC融合蛋白作为CBF功能的显性抑制因子，与RUNX1结合，并失调正常造血所需的多个基因的表达。目前使用细胞毒性化疗的治疗结果是45-65%的五年总存活率，但对于60岁以上的患者只有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启动细胞群体，这些细胞具有长期自我更新的潜力，并可以在复发时完全概括肿瘤表型。我们的假设是，抑制CBF2-SMMHC与RUNX1 Run结构域相互作用的小分子可能是有效的治疗药物，诱导细胞凋亡和/或分化，消除白血病起始细胞群，从而获得更好的长期生存。我们已经确定了一种初步的铅，它抑制CBF2-SMMHC和RUNX1之间的蛋白质-蛋白质相互作用。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铅的效力。将测试化合物是否有能力解除CBF2-SMMHC介导的抑制，并影响白血病细胞系的生长、凋亡和分化。选定的化合物将在具有良好特征的inv(16)白血病小鼠模型中进行体内测试。正如目标1中建议的抑制剂所做的那样，同时抑制CBF2和CBF2-SMMHC功能可能会导致毒性或缺乏疗效，因此我们建议开发对CBF2-SMMHC具有选择性的小分子抑制剂。将测试化合物是否有能力解除CBF2-SMMHC介导的抑制，并影响白血病细胞系的生长、凋亡和分化。选定的化合物将在具有良好特征的inv(16)白血病小鼠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的白血病前期祖细胞的分化和细胞死亡，而对骨髓中的正常干细胞和早期祖细胞的影响最小。我们建议研究选定的化合物在体内对正常造血的影响。对正常造血影响最小或没有影响的化合物将被优先研究它们在表达CBF2-SMMHC的敲入小鼠中减少/根除白血病前期祖细胞的效果。 公共卫生意义：我们建议开发CBF2-SMMHC融合蛋白的小分子抑制剂，该融合蛋白是一种急性髓系白血病(AML)的病原体。这种靶向抑制物有可能通过消除导致复发的白血病细胞群来提高这种白血病患者的长期存活率。这些抑制剂也有可能比标准化疗毒性更小，从而减少治疗的副作用。

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.