Targeting the kinome in K-ras driven colorectal cancers

项目概要/摘要 RAS蛋白的突变，特别是K-ras，在人类癌症中很常见， 已经详细检查了这种癌蛋白的来源。尽管我们努力翻译这本书 尽管K-ras基因突变体肿瘤的研究已经被用于改善癌症护理，但K-ras基因突变体肿瘤仍然是一个巨大的科学和临床挑战。 在这项提案中，我们专注于结直肠癌（CRC）中常见的单个KRAS突变， 因为这些突变与对靶向抑制剂的不同敏感性相关， 不同的信号程序，需要个性化的治疗策略。为了获取和利用这些独特的 程序，我们采用了一种创新的质谱技术，使大多数人 同时分析激酶组，提供了在无偏倚的情况下探索K-ras诱导的激酶组的方法。 时尚.我们独特的方法结合了质谱和蛋白激酶捕获珠， 从肿瘤细胞中提取活化的蛋白激酶，提供肿瘤激酶活性的“全貌”。使用此 在目的1中，我们将测量（i）模型等基因CRC系统中激酶组的基础活性， 代表常见的CRC相关KRAS突变等位基因，和（ii）一组充分表征的人KRAS突变等位基因， 突变CRC细胞系，在KRAS敲低之前和之后，以鉴定其活性依赖于 该致癌基因的各种突变形式的表达。在目标2中，我们将探讨CRC的弹性 KRAS激酶组对MEK抑制的作用，以鉴定和靶向促进抗性的补偿激酶。在Aim中 3、我们将利用这些信息在体内进行靶向的合成致死筛选，测试其有效性。 在KRAS依赖性CRC异种移植物和CRC敲入小鼠中抑制这些活化的蛋白激酶 由常见的KRAS外显子2（G12 D）或外显子4（A146 T）突变驱动的模型。这样，我们希望 鉴定有效治疗这些常见类型K-ras的个体化激酶抑制剂组合， 驱动CRC。

PROJECT SUMMARY/ABSTRACT Mutations in RAS proteins, in particular K-ras, are common in human cancer, and the signaling pathways emanating from this oncoprotein have been examined in detail. Despite intensive efforts to translate this knowledge into improved cancer care, K-ras mutant tumors remain a formidable scientific and clinical challenge. In this proposal, we focus on individual KRAS mutations that are commonly found in colorectal cancer (CRC), because these mutations are associated with different sensitivities to targeted inhibitors and thus may engage distinct signaling programs and require individualized therapeutic strategies. To access and exploit these distinct programs, we employ an innovative mass spectrometry-based technique that enables the majority of the human kinome to be analyzed simultaneously, providing the means to explore the K-ras induced kinome in an unbiased fashion. Our unique approach combines mass spectrometry and protein kinase-capture beads to monitor activated protein kinases from tumor cells, providing the ‘big picture’ of tumor kinase activity. Using this technology, in Aim 1 we will measure the basal activity of the kinome in (i) a model isogenic CRC system that represents common CRC associated KRAS mutant alleles and (ii) a panel of well characterized human KRAS mutant CRC cell lines, before and after KRAS knockdown, to identify kinases whose activity is dependent on expression of the various mutant forms of this oncogene. In Aim 2, we will explore the resiliency of the CRC KRAS kinome(s) to MEK inhibition to identify and target compensatory kinases that promote resistance. In Aim 3, we will use this information to carry out a targeted synthetic lethal screen in vivo, testing the effectiveness of inhibiting these activated protein kinases in KRAS dependent CRC xenografts and in CRC knock-in mouse models driven by a common KRAS exon 2 (G12D) or an exon 4 (A146T) mutation. In this way, we hope to identify individualized kinase inhibitor combinations that are effective in treating these common types of K-ras- driven CRC.