PDK-1 as an attractive cancer therapeutic

描述（由申请方提供）：通过磷酸化和激活蛋白激酶B（PK B）（介导PI 3-激酶依赖性信号转导的关键蛋白激酶）的能力鉴定了3-磷酸肌醇依赖性激酶-1（PDK-1）。随后已证明PDK-1在活化环中的同源活化磷酸化位点上磷酸化AGC激酶亚家族中的许多另外的蛋白激酶。其中一些也被PI 3激酶依赖性信号（如p70 S6和SGK）激活，而其他被认为是PI 3激酶独立的（如p90 rsk）。PDK-1在介导这些蛋白激酶的活化中的关键作用通过它们的活性在PDK-1缺失ES细胞中被消除的事实证明。肿瘤细胞被认为依赖于这些蛋白激酶中的几种蛋白激酶的活性来增殖和存活。此外，PDK-1的过度表达已被证明是致癌的。因此，预测抑制PDK-1将抑制肿瘤生长。然而，目前尚不清楚哪些肿瘤对PDK-1抑制最敏感，或者PDK-1抑制对完整生物体的毒性如何。缺乏PDK-1的小鼠在胚胎发育早期死亡。为了规避这些障碍，分析PDK-1抑制在哺乳动物生物体中肿瘤发生期间的作用，我们提出创建敲入小鼠，由此PDK-1中的ATP结合口袋被扩大，允许对其他蛋白激酶惰性的小分子的特异性抑制。我们将使用来自这些动物的细胞来证明急性PDK-1抑制的生物化学和生物学后果。此外，我们将在这些中启动肿瘤形成，以检查急性和特异性PDK-1抑制的后果。我们提出了两个定义明确的小鼠肿瘤模型来定义PDK-1活性的要求。第一种是化学致癌物诱导的皮肤肿瘤模型，其中DMBA启动和TPA导致表皮乳头状瘤的进展。第二种是骨髓性白血病模型，其中在造血细胞中在其内源性启动子下表达的活化的K-Ras引起致命的骨髓增殖性疾病。这些实验将确定PDK-1抑制是否代表抑制肿瘤发生的有效且无毒的方法。

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.