The p110 alpha and delta isoforms of PI3 kinase in hematopoiesis and leukemia

描述（由申请人提供）：急性髓性白血病（AML）每年影响25000人中的1人，只有40%的病例是可治愈的。主要的治疗方法是多药化疗和骨髓移植，这两种方法都有许多并发症和高死亡率。这在很大程度上归因于大多数抗白血病药物对造血干细胞（HSC）的毒性作用，经常导致细胞减少。因此，确定哪些信号通路在白血病细胞中特别重要，而在造血干细胞中不重要，这对于设计更特异性和毒性更小的AML治疗方法至关重要。PI3K激酶（PI3K）通路在包括AML在内的许多人类癌症中被病理激活。PI3K在造血过程中也可能很重要，因为它被造血生长因子受体激活，如促红细胞生成素受体、c-kit受体和fms样酪氨酸激酶3 （FLT3）。在造血细胞中，受体酪氨酸激酶通过PI3K的催化p110亚基发出信号，PI3K有3个亚型（,,），激活靶激酶AKT的磷酸化。然而，PI3K及其特异性亚基在正常和白血病干细胞功能中的作用尚不清楚。我们假设，靶向PI3K的单个亚基可能实现最具选择性的抗白血病作用，并且潜在的不良影响最小。本研究将利用两种新的遗传工具来研究PI3K亚基p110和p110单独或共同在白血病细胞和正常造血干细胞中的作用：(1)造血细胞中p110条件缺失的小鼠，(2)种系p110和p110条件缺失的双突变小鼠。初步数据显示，造血干细胞中p110的条件缺失会导致造血功能受损，并略微降低造血干细胞的再生能力。由于AKT信号在p110缺失的骨髓中维持，我们假设p110，一个只在造血细胞中表达的亚基，可以在造血干细胞中补充p110。在Specific Aim 1中，我们将分析p110和p110复合缺失对造血细胞HSC功能和AKT信号传导的影响。初步数据表明，至少在一些小鼠白血病模型中，p110的缺失不足以阻止疾病进展。基于p110切除不能消除造血细胞中的AKT信号，我们假设可能需要同时消除p110和p110来抑制白血病细胞的生长。Specific Aim 2将使用骨髓移植系统，通过逆转录病毒引入突变酪氨酸激酶，包括BCR-ABL、FLT3- ITD和JAK2V617F，检测p110、p110或两者缺失后小鼠白血病的发生和进展。特异性Aim 3将研究p110和p110在mll - af9诱导的白血病中白血病干细胞功能中的作用，这是一种特别难以靶向治疗的白血病。总之，这些研究将为了解PI3K信号转导在造血干细胞中的作用和机制，以及在白血病中靶向治疗这一途径的最佳策略提供重要见解。

DESCRIPTION (provided by applicant): Acute myeloid leukemia (AML) affects 1 in 25,000 people per year, and is curable in only about 40% of cases. The mainstay of treatment is multi-agent chemotherapy and bone marrow transplantation, both of which have many complications and high mortality. This is largely attributed to the toxic effects that most antileukemic drugs have on hematopoietic stem cells (HSC), frequently leading to cytopenias. Therefore, it is crucial to determine which signaling pathways are important specifically in leukemic cells but not in HSCs, to enable the design of more specific and less toxic treatments for AML. The PI3 kinase (PI3K) pathway is pathologically activated in many human cancers, including AML. PI3K may also be important during hematopoiesis, as it is activated by hematopoietic growth factor receptors, such as erythropoietin receptor, c-kit receptor, and fms-like tyrosine kinase 3 (FLT3). In hematopoietic cells, receptor tyrosine kinases signal through the catalytic p110 subunit of PI3K, which has 3 isoforms (, , ), to activate the phosphorylation of the target kinase AKT. However, the roles of PI3K and its specific subunits in normal and leukemic stem cell function are poorly understood. We hypothesize that targeting individual subunits of PI3K may achieve the most selective anti-leukemic effect with minimal potential adverse affects. This proposal will utilize two novel genetic tools to investigate the roles of the PI3K subunits p110 and p110, individually and together, in leukemic cells and in normal HSCs: (1) mice with conditional deletion of p110 in hematopoietic cells and (2) double mutant mice with germline deletion of p110 and conditional deletion of p110. Preliminary data reveal that conditional deletion of p110 in HSCs results in impaired erythropoiesis, and slightly reduced HSC repopulating ability. As AKT signaling is maintained in p110-deleted bone marrow, we hypothesize that p110, a subunit expressed exclusively in hematopoietic cells, can complement for p110 in HSCs. In Specific Aim 1, we will analyze the effects of compound loss of p110 and p110 on HSC function and on AKT signaling in hematopoietic cells. Preliminary data suggests that deletion of p110 is insufficient to halt disease progression in at least some murine models of leukemia. Based on the failure of p110 excision to abrogate AKT signaling in hematopoietic cells, we hypothesize that it may be necessary to eliminate both p110 and p110 to inhibit the growth of leukemic cells. Specific Aim 2 will examine leukemia initiation and progression in mice after loss of p110, p110, or both using the bone marrow transplant system with retrovirally introduced mutated tyrosine kinases, including BCR-ABL, FLT3- ITD, and JAK2V617F. Specific Aim 3 will examine the roles of p110 and p110 in leukemic stem cell function in MLL-AF9-induced leukemia, which has been particularly difficult to target therapeutically. Together, these studies will provide important insights into the role and mechanism of PI3K signal transduction in HSCs, and the best strategies for therapeutic targeting of this pathway in leukemia. PUBLIC HEALTH RELEVANCE: PI3 kinase signaling is of central importance in solid tumors, but its roles in normal and leukemic stem cell function are poorly understood. This proposal will use mouse knockout models to examine whether the p110 and p110 subunits of PI3 kinase are important for normal HSC function and are feasible targets for leukemia therapy.