Targeting the Pim 1 Protein Kinase to Overcome Resistance to AKT Inhibitors

描述(由申请人提供)：靶向信号转导通路用于开发新的前列腺癌治疗方法的成功迄今受到随后耐药机制发展的限制。在近70%的转移性前列腺癌患者中发现了高活性的AKT蛋白激酶，这是该疾病治疗的重要靶点。这项提案中的初步数据表明，在前列腺癌细胞系中加入AKT抑制剂可诱导细胞表面受体酪氨酸激酶(RTK)显着增加，该受体在一定程度上通过提高ERK活性来限制这些抑制剂的活性。重要的是，它还证明了AKT抑制剂诱导Pim-1蛋白激酶，该酶与前列腺癌的发生和发展有关。该研究小组发现，无论是通过siRNA还是在基因工程小鼠成纤维细胞中敲除Pim-1，都会抑制AKT抑制剂诱导RTK的反馈。使用卡夫实验室团队开发的一种小分子Pim-1抑制剂，他们已经证明，AKT和Pim-1抑制剂的组合在组织培养中协同阻止前列腺癌细胞的生长，并显著抑制免疫抑制动物的肿瘤生长。数据表明，AKT和Pim抑制剂调节RTK的翻译。这些令人兴奋的发现导致了一个独特的假说，即AKT抑制剂治疗引起Pim-1导向的反馈环，该反馈环诱导RTK，进而刺激ERK活性的增加。因此，AKT和Pim抑制剂的结合将阻断Pim-1的诱导，并协同作用杀死前列腺癌。这个建议的具体目的是探索和验证这一假说，方法是：1)在复杂的细胞培养和前列腺癌动物模型中证明下调Pim-1活性可增强AKT抑制剂的肿瘤杀伤作用；2)破译AKT抑制剂如何增加Pim-1蛋白激酶并调节翻译以增加RTK水平；3)探讨这些药物如何最好地联合用于肿瘤杀伤和抑制转移癌，并检查AKT和Pim抑制剂联合使用是否会导致前列腺癌中活性氧物种(ROS)的显著增加。这些研究将确定RTKs、磷酸化ERK和Pim-1水平可能是AKT抑制剂作用的潜在临床重要中间标志物。拟议的研究设计将利用独特的基因工程小鼠模型，以及核糖体图谱和脚印来探索这些问题。当这些研究完成后，这些研究将把注意力集中在开发与Pim和AKT抑制剂联合治疗以针对反馈抵抗机制的潜力上。这一组合将显著提高对目前正在研究的前列腺癌治疗单剂疗法的反应。

DESCRIPTION (provided by applicant): The success of targeting signal transduction pathways for the development of new prostate cancer therapies has been limited to date by the subsequent development of drug resistance mechanisms. Highly activated AKT protein kinase found in almost 70% of cases of metastatic prostate cancer are an important target for therapies in this disease. Preliminary data in this proposal demonstrate that the addition of AKT inhibitors to prostate cancer cell lines induces a marked increase in cell surface receptor tyrosine kinases (RTKs) that function to limit the activity of these inhibitors in part by elevating ERK activity. Importantly, it is also demonstrated that AKT inhibitors induce the Pim-1 protein kinase, an enzyme that has been implicated in prostate cancer initiation and progression. This research team has discovered that knocking down Pim-1 either by siRNA or in genetically engineering mouse fibroblasts will inhibit the feedback in which AKT inhibitors induce RTKs. Using a small molecule Pim-1 inhibitor developed by the Kraft laboratory team, they have demonstrated that the combination of an AKT and Pim-1 inhibitor synergistically blocks prostate cancer cell growth in tissue culture, and markedly inhibits the growth of tumors in immunosuppressed animals. Data obtained suggests that AKT and Pim inhibitors regulate the translation of RTKs. These exciting findings lead to the unique hypothesis that AKT inhibitor treatment causes a Pim-1-directed feedback loop that induces RTKs that in turn stimulates increases in ERK activity. Thus, the combination of an AKT and Pim inhibitor will interrupt the induction of Pim-1 and synergize to kill prostate cancer. The specific aims in this proposal are to explore and validate this hypothesis by: 1) demonstrating in complex cell culture and animal models of prostate cancer that knocking down Pim-1 activity enhances AKT inhibitor tumor killing; 2) deciphering how AKT inhibitors increase the Pim-1 protein kinase and modulate translation to increase RTK levels; and 3) exploring how these agents can be best combined for tumor killing and to inhibit metastatic cancer, and examining whether the combination of AKT and Pim inhibitors induces a marked increase in reactive oxygen species (ROS) in prostate tumors. These studies will identify RTKs, phosphorylated ERK, and Pim-1 levels as potentially clinically important intermediate markers of AKT inhibitor action. The proposed study designs will make use of unique genetically engineered mouse models, and ribosome profiling and foot printing to explore these questions. When completed, these studies will focus attention on the potential for the development of combination therapies with Pim and AKT inhibitors to target feedback resistance mechanisms. This combination would markedly enhance responses to single agent therapies currently under investigation for the treatment of prostate cancer.