PDK-1 as an attractive cancer therapeutic

描述(申请人提供)：3-磷酸肌醇依赖的激酶-1(PDK-1)是通过磷酸化和激活蛋白激酶B(PKB)的能力来鉴定的，PKB是介导PI3依赖的信号转导的关键蛋白激酶。PDK-1随后被证明在激活环中的同源激活磷酸化位点上，对AGC激酶亚家族中的一些额外的蛋白激酶进行磷酸化。其中一些也被PI3依赖的信号激活(如p70S6和SGK)，而另一些被认为是不依赖PI3的信号(如p90rsk)。在PDK-1缺失的ES细胞中，PDK-1的活性被取消，这证明了PDK-1在介导这些蛋白激酶的激活中的关键作用。肿瘤细胞被认为依赖于其中几种蛋白激酶的活性来实现其增殖和生存。此外，PDK-1的过度表达已被证明是致癌的。因此，抑制PDK-1有望抑制肿瘤生长。然而，目前尚不清楚哪些肿瘤对PDK-1抑制最敏感，也不知道PDK-1抑制对完整生物体的毒性有多大。缺乏PDK-1的小鼠在胚胎发育早期死亡。为了绕过这些障碍，分析在哺乳动物生物体中肿瘤发生过程中PDK-1抑制的影响，我们建议创建一种敲入小鼠，借此扩大PDK-1中的ATP结合口袋，允许小分子对其他蛋白激酶具有惰性的特异性抑制。我们将使用来自这些动物的细胞来展示急性PDK-1抑制的生化和生物学后果。此外，我们将在这些肿瘤中启动肿瘤形成，以检查急性和特异性PDK-1抑制的后果。我们提出了两个明确定义的小鼠肿瘤模型来定义对PDK-1活性的需求。第一种是化学致癌物诱导的皮肤肿瘤模型，DMBA启动，TPA导致表皮乳头状瘤的进展。第二种是髓系白血病模型，在造血细胞中，激活的K-RAS在其内源性启动子下表达，导致一种致命的骨髓增生性疾病。这些实验将确定抑制PDK-1是否代表了一种有效和无毒的抑制肿瘤形成的方法。

DESCRIPTION (provided by applicant): 3-phosphoinositide-dependent kinase-1 (PDK-1) was identified by its ability to phosphorylate and activate protein kinase B (PKB), a key protein kinase in mediating PI3-kinase-dependent signal transduction. PDK-1 has subsequently been demonstrated to phosphorylate a number of additional protein kinases in the AGC kinase sub-family, on a homologous activating phosphorylation site in the activation loop. Some of these are also activated by PI3-kinase-dependent signals (eg p70S6 and SGK), whereas others are thought to be PI3- kinase-independent (eg p90rsk). The critical role for PDK-1 in mediating the activation of these protein kinases is demonstrated by the fact that their activity is abolished in PDK-1 null ES cells. Tumor cells are thought to rely on the activity of several of these protein kinases for their proliferation and survival. In addition, over-expression of PDK-1 has been shown to be oncogenic. Therefore, inhibition of PDK-1 would be predicted to inhibit tumor growth. However, it is not known which tumors would be most sensitive to PDK-1 inhibition, or how toxic PDK-1 inhibition would be to an intact organism. Mice lacking PDK-1 die early in embryogenesis. To circumvent these obstacles analyzing the effects of PDK-1 inhibition during tumorigenesis in a mammalian organism, we propose to create a knock-in mouse whereby the ATP binding pocket in PDK-1 is enlarged, allowing specific inhibition by small molecules that should be inert to other protein kinases. We will use cells derived from these animals to demonstrate the biochemical and biological consequences of acute PDK-1 inhibition. In addition, we will initiate tumor formation in these to examine the consequences of acute and specific PDK-1 inhibition. We propose two well-defined mouse tumor models to define the requirement for PDK-1 activity. The first is a chemical carcinogen induced skin tumor model, whereby DMBA initiates and TPA causes progression of epidermal papillomas. The second is a myeloid leukemia model whereby activated K-Ras expressed under its endogenous promoter in hematopoietic cells causes a fatal myeloproliferative disorder. These experiments will determine whether PDK-1 inhibition represents an effective and non-toxic approach to curb tumorigenesis.