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

描述(由申请人提供)：急性髓系白血病(AML)每年影响25000人中有1人，只有大约40%的病例是可治愈的。目前主要的治疗方法是多药联合化疗和骨髓移植，两者并发症多，死亡率高。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，从而能够设计出更特异、毒性更低的AML治疗方法，这是至关重要的。PI3K通路在包括急性髓系白血病在内的许多人类癌症中被病理激活。PI3K在造血过程中可能也很重要，因为它是由造血生长因子受体激活的，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亚单位(有3种异构体(、、))来激活靶蛋白激酶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还知之甚少。我们假设，靶向PI3K的单个亚基可能以最小的潜在不良影响达到最具选择性的抗白血病效果。这项建议将利用两种新的遗传工具来单独和共同研究PI3K亚基p110和p110在白血病细胞和正常HSC中的作用：(1)有条件地删除造血细胞中的p110的小鼠；(2)具有种系缺失和有条件缺失的p110的双突变小鼠。初步数据显示，HSC中p110的条件性缺失会导致红细胞生成受损，并略微降低HSC的再生能力。由于AKT信号在p110缺失的骨髓中保持不变，我们假设p110是一个仅在造血细胞中表达的亚单位，可以在造血干细胞中补充p110。在特定的目标1中，我们将分析p110和p110的复合缺失对HSC功能和造血细胞AKT信号的影响。初步数据表明，至少在某些白血病小鼠模型中，p110的缺失不足以阻止疾病的进展。基于p110切除未能阻断造血细胞中的AKT信号，我们推测可能有必要同时消除p110和p110以抑制白血病细胞的生长。特殊目的2将使用带有逆转录病毒引入的突变酪氨酸激酶的骨髓移植系统，检测p110和/或p110缺失后小鼠白血病的发生和发展，包括bcr-abl、flt3-itd和JAK2V617F。具体目标3将研究p110和p110在MLL-AF9诱导的白血病中的白血病干细胞功能中的作用，这是特别难在治疗中靶向的。总之，这些研究将为深入了解PI3K信号转导在造血干细胞中的作用和机制，以及针对该途径在白血病中的治疗靶向的最佳策略提供重要的见解。 公共卫生相关性：PI3激酶信号在实体瘤中具有中心重要性，但其在正常和白血病干细胞功能中的作用尚不清楚。这项建议将使用小鼠基因敲除模型来检查PI3激酶的p110和p110亚单位是否对正常的HSC功能重要，是否为白血病治疗的可行靶点。

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.