The role of AKT signaling in NOTCH1 induced leukemias

描述(申请人提供)：NOTCH1受体的激活突变导致的Notch信号的异常激活在超过50%的人类T淋巴细胞白血病(T-ALL)的发病机制中发挥了关键作用。T-ALL病例中NOTCH1基因突变的发现促使临床试验的启动，以测试用β-分泌酶抑制剂(GSI)阻断NOTCH1信号通路在本病中的有效性。然而，GSI只在一小部分NOTCH1突变的T-ALL中活跃，这表明导致GSI耐药的机制可能限制了GSI在T-ALL中的有效性。我们的初步数据表明：(I)NOTCH1上调T-ALL中PI3K/AKT通路；(Ii)NOTCH1信号下调PTEN抑癌基因的表达；(Iii)PTEN缺失与GSI抵抗NOTCH1抑制有关。因此，我们的中心假设是，PI3K-AKT通路的活性至少部分地介导了NOTCH1信号异常的致癌效应。我们进一步认为，PTEN突变缺失导致的PI3K-AKT的结构性激活促进了细胞的生长、增殖和存活，使T-ALL细胞对NOTCH1的抑制不敏感。本研究的短期目标是确定NOTCH1和PI3K/AKT信号通路之间的相互作用在T细胞转化和治疗反应中的意义。为了实现这些目标，我们提出了以下具体目标：目的1.分析NOTCH1和Pten-PI3K-AKT在T细胞转化过程中的相互作用。为此，我们将利用逆转录病毒癌基因转移在野生型和Pten基因敲除小鼠的造血祖细胞中表达激活的NOTCH1，以评估致癌NOTCH1和PI3K-Akt信号在T细胞淋巴母细胞肿瘤发病机制中的相互作用。目的2.分析白血病干细胞中NOTCH1和Pten的功能关系。在这一目标中，我们将分析NOTCH1和PI3K-Akt信号通路在Pten野生型、Pten单倍体不足和Pten缺失型肿瘤中表达激活的NOTCH1的白血病干细胞的动态平衡中的作用。目的3.分析T-ALL对分子靶向药物反应中Noch和Akt信号的相互作用。Pten-Lost是否会引起对NOTCH1抑制剂的耐药性？抑制AKT信号通路是否逆转了对NOTCH1抑制剂的耐药性？Pten杂合子和/或Pten缺失T-ALL肿瘤对Akt抑制剂或针对Akt信号下游效应因子的药物(如mTOR、NFKB和FOXO因子)是否显示出更高的敏感性？公共卫生相关性：该项目旨在分析NOTCH1和PI3K-AKT通路在T细胞淋巴母细胞性白血病(T-ALL)发病机制中的遗传相互作用，以及它们在针对NOTCH1和AKT癌蛋白的新兴疗法中所起的作用。这里概述的实验将为合理开发针对T-ALL的新疗法奠定基础。

DESCRIPTION (provided by applicant): Aberrant activation of NOTCH signaling due to the presence of activating mutations in the NOTCH1 receptor play a critical role in the pathogenesis in more than 50% human T-cell lymphoblastic leukemias (T-ALL). The identification of NOTCH1 mutations of T-ALL cases has prompted the initiation of clinical trials to test the effectiveness of blocking NOTCH1 signaling with ?-secretase inhibitors (GSIs) in this disease. However, GSIs are active only in a small fraction of T-ALLs with NOTCH1 mutations, suggesting that mechanisms responsible for GSI resistance may limit the effectiveness of GSIs in T-ALL. Our preliminary data shows that: (i) NOTCH1 upregulates the PI3K/AKT pathway in T-ALL; (ii) NOTCH1 signaling downregulates the expression of the PTEN tumor suppressor gene; and (iii) loss of PTEN is associated with resistance to NOTCH1 inhibition with GSIs. Thus, our central hypothesis is that the activity of the PI3K-AKT pathway mediates, at least in part, the oncogenic effects of aberrant NOTCH1 signaling. We further propose that constitutive activation of PI3K-AKT due to mutational loss of PTEN promotes cell growth proliferation and survival rendering T-ALL cells insensitive to NOTCH1 inhibition. The short-term goals of this research are to establish the significance of the interaction between NOTCH1 and PI3K/AKT signaling pathways in T-cell transformation and therapy response. To achieve these objectives, we propose the following specific aims: Aim 1. To analyze the interaction between NOTCH1 and Pten-PI3K-AKT in T-cell transformation. In this aim we will use retroviral oncogene transfer to express activated NOTCH1 in hematopoietic progenitors from wild type and Pten knockout mice to assess the relevance of the interaction between oncogenic NOTCH1 and PI3K-Akt signaling in the pathogenesis of T-cell lymphoblastic tumors. Aim 2. To analyze the functional relationship between NOTCH1 and Pten in leukemia stem cells. In this aim we will analyze the role of the NOTCH1 and PI3K-Akt signaling pathways in the homeostasis of the leukemia stem cells from Pten-wild type, Pten-haploinsuficient and Pten-null tumors expressing activated NOTCH1. Aim 3. To analyze the interaction between NOTCH and Akt signaling in the response of T-ALL to molecularly targeted drugs. Does Pten-loss induce resistance to NOTCH1 inhibitors? Does inhibition of the AKT signaling pathway reverse resistance to NOTCH1 inhibitors? Do Pten-heterozygous and/or Pten-null T-ALL tumors show increased sensitivity to Akt inhibitors or to drugs targeting downstream effectors of Akt signaling such as mTOR, NFKB and FOXO factors? PUBLIC HEALTH RELEVANCE: This project aims to analyze the genetic interaction between NOTCH1 and PI3K-AKT pathways in the pathogenesis of T-cell lymphoblastic leukemias (T-ALL) and their role in tumor resistance to emerging therapies targeting the NOTCH1 and AKT oncoproteins. The experiments outlined here will set the basis for the rational development of new therapies against T-ALL.