Molecular understanding of leukemic bone marrow cytokine-Ras signals and metabolic dependence

项目摘要/摘要 儿童急性淋巴细胞性白血病(ALL)是一种幼稚骨髓扩张性疾病 细胞，并用非特异性化疗治疗。ALL仍然是#年癌症死亡的主要原因之一 年满20岁的人。T细胞急性淋巴细胞白血病(T-ALL)的发育起源对临床有着深远的影响。 基因突变表明RAS信号异常在50%的T-ALL中起因果作用，但这种异常是如何发生的 信号影响T-ALL的发育轨迹尚不清楚。同样，大体上还不知道如何 有效抑制异常RAS信号。 在1R01CA187318期间，我们在RAS信号中发现了一个新的分子范式；致癌RAS 突变(KrasG12D)和RAS激活物RASGRP1的过度表达导致两种非常独特的白血病 RAS信号。在我们新的RoLoRiG/Mx1CRE鼠标模型的进度报告中，我们展示了这两个 RAS信号的异常均可诱导异常的造血，但具有相反的干细胞特征。大写字母 在我们的定量、多重流式细胞仪平台上，我们确定RAS和RASGRP1经常 连接到T-ALL中的PI3K(肌醇磷脂3-激酶)-AKT/mTOR信号。接下来，我们表演了四个 使用PI3K抑制剂进行广泛的合成致死性筛选，以确定T-ALL的有效联合治疗。我们 证实了十种预测的小分子抑制剂和PI3K抑制剂GDC0941的联合治疗 与微管蛋白抑制剂长春新碱在五种癌症类型中产生广泛的协同作用，并在我们的临床前小鼠试验中 活着。这些筛查预测了许多代谢目标，但这些目标仍未被探索。 我们的更新侧重于了解异常的RAS-PI3K信号与细胞之间的联系 新陈代谢的目标是解决T-ALL的发育起源并测试新的组合 治疗。我们将在我们的10人小组中确定RAS-PI3K信号和细胞代谢之间的联系 通过研究我们筛查中确定的六种营养转运蛋白(SLCs)，对所有细胞系进行T-all检测(目标1)。我们会 获得对信号、代谢特征和造血成分的机械性理解 作为白血病RAS-PI3K信号的函数的轨迹，具有前所未有的分辨率。我们将继续努力 RoLoRiG/Mx1CRE和KrasG12D/Mx1CRE小鼠模型的表型完全相反 骨髓干细胞(目标2)。最后，我们将获得发展的一个全面的表征-- 单细胞分辨率的儿科T-ALL中的生化和代谢程序。我们将把这些结合起来 通过临床前试验测试PI3Kγ和δ抑制、SLC13A2和SLC13A2抑制的体内疗效 SLC25A44，以及我们T-ALL/NGS平台中的组合(目标3)。协同目标、新陈代谢洞察力、 新的鼠标模型、患者样本-NSG管道以及创新的单细胞分辨率技术平台 (CyTOF，SCENITH，PORTO-FLOW)将使我们能够对分子 对T-ALL和精确医学在癌症中的认识。

PROJECT SUMMARY/ABSTRACT Childhood Acute Lymphoblastic Leukemia (ALL) is a disease of the bone marrow with expansion of immature cells and treated with non-specific chemotherapy. ALL remains one of the leading causes of cancer death in persons <20 years old. The developmental origin of T cell ALL (T-ALL) is and profoundly impacts the clinic. Genetic mutations pointed aberrant Ras signals with causal role in 50% of T-ALL, but how such aberrant signals impact developmental trajectories in T-ALL is not known. Similarly, it is by and large unknown how to effectively inhibit aberrant Ras signals. During 1R01CA187318 we uncovered a new molecular paradigm in Ras signaling; Oncogenic Ras mutations (KRasG12D) and overexpression of the Ras activator RasGRP1 drive two very distinctive leukemic Ras signals. In the progress report with our new RoLoRiG/Mx1CRE mouse model, we show that these two aberrant Ras signals both induce abnormal hematopoiesis, but with opposing stem cell features. Capitalizing on our quantitative, multiplex flow cytometry platform we determined that Ras and RasGRP1 frequently connect to PI3K (phosphoinositide 3-kinase)-AKT/mTOR signaling in T-ALL. Next, we performed four extensive synthetic lethality screens with PI3K inhibitors to identify effective combination therapy in T-ALL. We confirmed ten predicted combination therapies with small molecule inhibitors and the PI3K inhibitor GDC0941 with tubulin inhibitor Vincristine yields broad synergy in five cancer types and in our preclinical mouse trials in vivo. The screens predict many metabolic targets, which remained unexplored. Our renewal focuses on understanding the connection between aberrant Ras-PI3K signals and cell metabolism with the goal of resolving the developmental origin of T-ALL and testing new combination therapies. We will determine the connections between Ras-PI3K signals and cell metabolism in our panel of 10 T-ALL cell lines by investigating six nutrient transporters (SLCs), identified in our screens (Aim 1). We will obtain mechanistic understanding of the signals, metabolic features, and hematopoietic composition and trajectories, as a function of leukemic Ras-PI3K signals with unprecedented resolution. We will continue efforts on RoLoRiG/Mx1CRE and KRasG12D/Mx1CRE mouse models that display completely opposite phenotypes in bone marrow stem cells (Aim 2). Lastly, we will obtain a comprehensive characterization of developmental-, biochemical- and metabolic- programs in pediatric T-ALL with single cell resolution. We will combine these efforts with preclinical trial to test the in vivo efficacy of PI3Kγ- and δ- inhibition, inhibition of SLC13A2 and SLC25A44, and combinations in our T-ALL/NGS platform (Aim 3). The synergistic aims, metabolic insights, new mouse models, patient sample-NSG pipeline, and innovative, single cell-resolution technology platforms (CyTOF, SCENITH, phospho-flow) will allow us to make significant contributions towards molecular understanding of T-ALL and precision medicine in cancer.