Discovering small molecules that overcome differentiation arrest in acute myeloid

描述（由申请人提供）：成人急性髓性白血病（AML）是一种破坏性疾病，5年生存率仅为25%。AML缺乏新的治疗方法，目前的化疗护理标准在过去三十年中没有改变。治疗AML的一个成功案例是开发了促进白血病细胞成熟或分化的疗法。在一小部分（约10%）急性早幼粒细胞白血病（APL）AML患者中，全反式维甲酸（ATRA）和砷形式的分化治疗耐受性良好且非常有效，5年生存率接近80%。不幸的是，分化治疗不能用于其余90%的急性髓性白血病患者。同源异型盒蛋白HoxA9在早期造血中表达，并且对沿着骨髓谱系的细胞的正常发育至关重要。HoxA9的不适当表达已经在大约70%的AML中被证实。此外，表达涉及MLL（混合谱系白血病）基因的融合癌蛋白的白血病亚群依赖于HoxA9的表达。这些观察结果使得HoxA9及其下游靶标成为小分子探针抑制的有吸引力的候选者。研究一直受到白血病模型系统不足和原始患者样本有限的阻碍。已经开发了一种新的AML体外模型，其中原代鼠骨髓细胞被癌蛋白HoxA9阻滞在未成熟状态。这些细胞允许鉴定可以克服骨髓分化停滞的生物学相关化合物。可以鉴定两种类型的分子：直接干扰由HoxA9建立的分化停滞机制的分子，以及能够以不依赖于HoxA9的方式促进分化的分子。用内置的分化标志物对测定细胞系进行工程改造，因为其从仅在成熟细胞中有活性的启动子表达绿色荧光蛋白（GFP）。因此，通过测定细胞的绿色荧光，可以以高通量方式容易地鉴定促进分化的化合物。二次和反筛选试验将消除潜在的自体荧光化合物，并通过测定基因表达和细胞表面标志物表达的变化来确认对骨髓分化的影响。开发一种能够促进急性髓细胞白血病分化的小分子药物将是目前白血病化疗的一个重要进展。页1的1 公共卫生相关性：急性骨髓性白血病（AML）是一种毁灭性疾病，迫切需要新的AML治疗方法。传统的化疗通常会使快速分裂的白血病细胞中毒，在75%的病例中最终无效。已经设计了一种新的AML模型系统，以鉴定触发白血病细胞恢复正常成熟过程，从而失去其增殖和白血病潜能的新化合物。页1的1

DESCRIPTION (provided by applicant): Acute myeloid leukemia (AML) in adults is a devastating disease with a 5-year survival rate of only 25%. New treatments for AML are lacking, and the current standard of care for chemotherapy has not changed in the last thirty years. One success story in the treatment of AML has been the development of therapies which promote the maturation, or differentiation of the leukemic cells. In the small subset (~10%) of AML patients with acute promyelocytic leukemia (APL), differentiation therapy in the form of all-trans retinoic acid (ATRA) and arsenic are both well-tolerated and extremely effective, leading to 5-year survival rates approaching 80%. Unfortunately, differentiation therapy is not available for the remaining 90% of acute myeloid leukemia patients. The homeobox protein HoxA9 is expressed in early hematopoiesis and is critical to the normal development of cells along the myeloid lineage. The inappropriate expression of HoxA9 has been demonstrated in approximately 70% of AML. Furthermore, the subset of leukemias which express a fusion oncoprotein involving the MLL (mixed lineage leukemia) gene are dependent upon the expression of HoxA9. These observations make HoxA9 and its downstream targets attractive candidates for inhibition by small molecule probes. Research has been hindered by inadequate model systems of leukemia and the limited availability of primary patient samples. A novel in vitro model of AML has been developed whereby primary murine myeloid cells are arrested in an immature state by the oncoprotein HoxA9. These cells allow for the identification of biologically relevant compounds that can overcome myeloid differentiation arrest. Two types of molecules can be identified: those that directly interfere with the mechanism of differentiation arrest established by HoxA9, and those that are capable of promoting differentiation in a HoxA9-independent manner. The assay cell line is engineered with a built-in marker of differentiation as it expresses the green fluorescent protein (GFP) from a promoter which is active only in mature cells. Thus, compounds that promote differentiation can be readily identified in a high-throughput fashion by assaying cells for green fluorescence. Secondary and counterscreen assays will eliminate potentially autofluorescent compounds and will confirm the effect on myeloid differentiation by assaying for changes in gene expression and cell surface marker expression. The development of a small molecule capable of promoting differentiation in acute myeloid leukemia will be an important advancement in the current state of leukemia chemotherapy. Page 1 of 1 PUBLIC HEALTH RELEVANCE: Acute myeloid leukemia (AML) is a devastating disease and new treatments for AML are in desperate need. Traditional chemotherapy, which typically poisons rapidly dividing leukemia cells, is ultimately ineffective in 75% of cases. A novel model system of AML has been devised to identify novel compounds which trigger leukemic cells to resume the normal process of maturation, thereby losing their proliferative and leukemic potential. Page 1 of 1