Targeting AML with PI3K/AKT inhibitors and BH3-mimetics

描述(由申请人提供)：迫切需要治疗难治性急性髓细胞白血病(AML)的新策略。Bcl2家族的凋亡调控蛋白家族和PI3K/AKT/mTOR和MEK/ERK通路在AML中经常发生异常，越来越多的证据表明，除了它们的相对丰度外，Bcl2家族成员之间的相互作用在决定细胞命运方面起着关键作用。我们现在提出了一种基于机制的抗白血病策略，将PI3K/mTOR抑制剂与模拟BH3的ABT-737/263结合起来，其中这三条重要的生存途径被协同扰乱。这种方法利用了我们最近的发现，与其他类型的肿瘤不同，在AML中，PI3K/mTOR抑制剂不同于其他AKT途径拮抗剂，既抑制AKT又抑制ERK。在BH3-模拟物的存在下，这导致促凋亡蛋白(如Bim，Bak)和抗凋亡蛋白(如Mcl-1，Bcl2/Xl)之间的关联发生关键变化，以及它们表达的扰动(例如，Mcl-1下调和Bim上调)。在特定的目标#1中，我们将使用遗传策略(例如，Bim AKT或ERK磷酸化突变体，结构性活性或激酶死亡的AKT或MEK突变体)来测试我们的假设，即除了Mcl-1下调外，协同作用还源于Mcl-1下调Bim(以及Bak)从Mcl-1/Bcl2/xl的抑制影响中释放。在具体目标2中，我们将测试假设，即协同作用也将发生在原始的、遗传定义的AML样本中，并且类似的机制将起作用。我们还将测试PI3K/AKT途径突变，特别是基础AKT激活状态可以预测其易感性的假设 策略。在具体目标#3中，我们将测试PI3K/mTOR抑制剂和BH3-模拟物的假设，它们分别针对原始白血病启动细胞(例如，CD34/CD38-/CD123)，将协同作用来根除这一经典耐药群体。我们会 还首次表征了AKT和ERK在这些细胞中的激活情况。在具体目标#4中，将使用体内多个全身性AML模型系统来推断完整动物的体外研究结果，并确定抗白血病协同作用是否存在类似的机制。这些目标的实现将为开发和实施PI3K途径抑制剂/BH3模拟抗白血病策略提供必要的理论基础，这是第一个协调中断三条相互作用的白血病相关途径的策略。它还可能有助于验证药效反应决定因素，并确定最有可能从这一策略中受益的AML患者个体。

DESCRIPTION (provided by applicant): New treatment strategies for refractory acute myelogenous leukemia (AML) are urgently needed. Dysregulation of the Bcl-2 family of apoptotic regulatory proteins and the PI3K/AKT/mTOR and MEK/ERK pathways occurs frequently in AML, and accumulating evidence indicates that interactions between Bcl-2 family members, in addition to their relative abundance, play key roles in determining cell fate. We now propose a mechanism-based anti-leukemic strategy combining PI3K/mTOR inhibitors with the BH3-mimetic ABT-737/263 in which these three important survival pathways are coordinately disrupted. This approach exploits our recent discovery that in AML, in contrast to other tumor types, PI3K/mTOR inhibitors, unlike other AKT pathway antagonists, inhibit both AKT and ERK. In the presence of BH3-mimetics, this leads to critical alterations in the association between pro-(e.g., Bim, Bak) and anti-(e.g., Mcl-1,Bcl-2/xL) apoptotic proteins, as well as perturbations in their expression (e.g., Mcl-1 down-regulation and Bim up-regulation). In Specific Aim #1, we will employ genetic strategies (e.g., Bim AKT or ERK phosphorylation mutants, constitutively active or kinase-dead AKT or MEK mutants) to test our hypothesis that synergism stems from, in addition to Mcl-1 down-regulation, unleashing of up-regulated Bim (as well as Bak) from the inhibitory influence of Mcl-1/Bcl- 2/xL. In Specific Aim 2, we will test the hypotheses that synergistic interactions will also occur in primary, genetically defined AML specimens, and that analogous mechanisms will be responsible. We will also test the hypothesis that PI3K/AKT pathway mutations, and particularly basal AKT activation status, can predict susceptibility to this strategy. In Specific Aim #3, we will test the hypothesis that PI3K/mTOR inhibitors and BH3-mimetics, which individually target primitive leukemia-initiating cells (e.g., CD34+/CD38-/CD123+), will interact synergistically to eradicate this classically resistant population. We will also characterize, for the first time, AKT and ERK activation in these cells. In Specific Aim #4, multiple in vivo systemic AML model systems will be employed to extrapolate in vitro findings to intact animals, and to determine whether similar mechanisms underlie anti-leukemic synergism. Fulfillment of these aims will provide the theoretical foundation needed to develop and implement a PI3K pathway inhibitor/BH3-mimetic anti-leukemic strategy, which represents the first to interrupt coordinately three mutually interactive leukemia-related pathways. It may also help to validate pharmacodynamic response determinants, and identify individual AML patients most likely to benefit from this strategy.