TOR kinase inhibitors for leukemia therapy: mechanisms of action and resistance

描述（由申请人提供）：大多数癌细胞表现出磷酸肌苷3激酶（PI3K）和下游酶AKT和TOR的激活。靶向PI3K/AKT/TOR网络是一种很有前景的癌症治疗策略，但目前尚不清楚哪种靶点能提供疗效和耐受性的最佳平衡。部分抑制TOR的化合物（如雷帕霉素）或同时抑制PI3K和TOR的化合物（如PI-103）在疗效和/或耐受性方面有明显的局限性。该领域的一个重大突破是发现了高效、选择性、小分子竞争性TOR抑制剂的新化合物。这些化合物被称为“活性位点TOR抑制剂”，当TOR酶存在于TORC1或TORC2（两种不同的多蛋白复合物）中时，它们可以完全抑制TOR酶。与变构抑制剂雷帕霉素相比，活性位点TOR抑制剂对关键信号通路的影响更广泛，在抑制小鼠和人类白血病细胞存活方面更有效。值得注意的是，这些化合物的免疫抑制作用比雷帕霉素小，尽管它们具有更大的抗白血病功效。活性位点TOR抑制剂在白血病模型中与选择性较低的PI3K/TOR抑制剂同样有效，但在小鼠中耐受性明显更好。几种TOR活性位点抑制剂正处于早期临床试验阶段。本提案的总体目标是完善和扩展我们对活性位点TOR抑制剂的理解，最终目标是改善癌症患者的健康。我们将重点关注B细胞恶性肿瘤，因为活性位点TOR抑制剂在B细胞白血病模型中具有显著作用，但其作用机制尚不清楚。这项提议有两个具体目的。首先，我们将建立激活位点TOR抑制剂触发白血病细胞死亡的机制。在这个目的中，我们将使用遗传方法来验证TORC1和TORC2介导白血病细胞存活信号的假设。这将通过诱导cre介导的敲除系统和shrna介导的敲除来完成。然后，我们将测试TORC1和TORC2底物在维持白血病细胞存活中的作用。其次，我们将定义细胞对活性位点TOR抑制剂的抗性机制。与任何靶向分子方法一样，不同类型的癌细胞对活性位点TOR抑制剂表现出不同的敏感性。药物开发中的一个新主题是需要确定靶向药物的有效组合。我们将使用不发生凋亡的细胞系来响应TOR抑制，我们将使用候选和全局方法来确定耐药机制。然后，我们将在体外和体内测试联合策略。确定耐药机制并应用适当的药物组合将扩大活性位点TOR抑制剂的潜在应用。

DESCRIPTION (provided by applicant): Most cancer cells display activation of phosphoinositide 3-kinase (PI3K) and the downstream enzymes AKT and TOR. Targeting the PI3K/AKT/TOR network is a promising strategy for cancer therapeutics, yet it is not clear which target profile will provide the best balance of efficacy and tolerability. Compounds that partially inhibit TOR (such as rapamycin) or compounds that inhibit both PI3K and TOR (such as PI-103) have significant limitations in efficacy and/or tolerability. A major breakthrough in this field has been the identification of novel compounds that are highly potent, selective, small molecule competitive inhibitors of TOR. Termed "active-site TOR inhibitors", these compounds fully inhibit the TOR enzyme when it is present in TORC1 or TORC2, two distinct multiprotein complexes. Compared to rapamycin, an allosteric inhibitor, active-site TOR inhibitors have a broader effect on key signaling pathways and are more effective at suppressing survival of murine and human leukemia cells. Remarkably, these compounds are less immunosuppressive than rapamycin despite their greater anti-leukemic efficacy. Active-site TOR inhibitors appear equally effective in leukemia models as the less selective PI3K/TOR inhibitors, yet are considerably better tolerated in mice. Several active-site TOR inhibitors are in early stage clinical trials. The overall objective of this proposal is to refine and extend our understanding of active-site TOR inhibitors, with the ultimate goal of improving the health of cancer patients. We will focus on B cell malignancies, since active-site TOR inhibitors have dramatic effects in B cell leukemia models yet the mechanism of action remains poorly understood. The proposal has two specific aims. First, we will establish the mechanisms by which active-site TOR inhibitors trigger leukemia cell death. In this aim we will use genetic approaches to test the hypothesis that both TORC1 and TORC2 mediate survival signaling in leukemia cells. This will be accomplished using inducible Cre-mediated knockout systems and shRNA-mediated knockdown. We will then test the roles of TORC1 and TORC2 substrates in maintaining survival in leukemia cells. Second, we will define mechanisms of cellular resistance to active-site TOR inhibitors. As with any targeted molecular approach, subtypes of cancer cells display differing sensitivity to active-site TOR inhibitors. An emerging theme in drug development is the need to identify effective combinations of targeted agents. Using cell lines that do not undergo apoptosis in response to TOR inhibition, we will use candidate and global approaches to identify druggable mechanisms of resistance. We will then test combination strategies in vitro and in vivo. Identifying mechanisms of resistance and applying appropriate drug combinations will broaden the potential application of active-site TOR inhibitors.