Mechanistic Basis for ERK in driving KRAS-dependent pancreatic cancer

项目摘要/摘要 胰腺导管腺癌(PDAC)是美国第三大癌症死亡原因，是 以95%的KRAS癌基因突变激活率为特征。在经历了近40年的失败后， 最近临床批准针对一个KRAS突变(G12C)的直接KRAS抑制剂用于肺癌标记物 这是KRAS突变癌症治疗方法发展的一个重要里程碑。KRASG12C特异性抑制剂 在一组KRASG12C突变患者中显示出肿瘤显著缩小，但基本上全部复发 由于治疗引起的获得性耐药性。对复发患者的基因分析确定了发病机制 其中大部分涉及驱动重新激活的信号组件的突变激活 KRAS的关键效应通路是三级RAF-MEK-ERK丝裂原活化蛋白激酶级联通路。 因此，ERK的重新激活将限制直接KRAS抑制剂的长期疗效。尽管取得了巨大的成功 ERK MAPK级联每个节点的有效和选择性抑制剂的开发 作为单一疗法，它们对RAS突变的癌症几乎没有临床疗效。有两个关键问题 导致这一结果的原因是，对正常组织的毒性和从头开始或治疗诱导的获得性耐药性 癌细胞。我建议进一步阐明ERK驱动KRAS依赖性癌症的机制 增长将引导更有效的抗ERK疗法的发展。然而，通过这些机制 ERK推动PDAC增长的机制仍鲜为人知。一个尚未解决的主要问题是ERK活动如何不同 亚细胞隔间支持癌症的生长。目标1学习包括我的K99培训阶段，我将 采取两种互补的方法来更好地理解细胞质和核ERK的作用 支持依赖KRAS的PDAC增长的活动。首先，我将确定细胞质对 核ERK活性在支持KRAS突变型PDAC生长中的作用第二，我会用一种药理 核出口蛋白Exportin-1(Selinexor)的抑制剂，以确定它是否破坏ERK细胞质- 核动力学，并使PDAC模型对KRAS抑制敏感。我的目标2研究包括我的R00- 支持独立研究，并基于我们全面的依赖ERK的磷酸蛋白质组/ KRAS突变体PDAC的转录组研究。利用这些数据，我设计了CRISPR-Cas9基因缺失- 功能筛选文库，靶向ERK调节的磷酸蛋白和/或转录本。我现在将执行一个系统- 广泛确定ERK在PDAC肿瘤发生中的作用以及识别新的ERK依赖性 与KRAS抑制剂结合的靶点。帮助我实现这些研究目标并成功过渡到 独立的第一阶段有一个特殊的指导委员会，由研究中的主要研究人员组成 KRAS信号和治疗学(Chning Der)、ERK时空信号(张进)和ERK 底物利用(John Blenis)。在他们的指导下，我相信我将成功过渡到建立 一个独立的研究项目，在那里我将提高我们对ERK信号转导和治疗的知识。

Project Summary/Abstract Pancreatic ductal adenocarcinoma (PDAC), the third leading cause of cancer deaths in the United States, is characterized by a 95% rate of mutational activation of the KRAS oncogene. After nearly four decades of failure, the recent clinical approval of a direct KRAS inhibitor targeting one KRAS mutation (G12C) for lung cancer marks a significant milestone in the development of therapies for KRAS-mutant cancers. KRASG12C-specific inhibitors have demonstrated dramatic tumor shrinkage in a subset of KRASG12C-mutant patients but essentially all relapse due to treatment-induced acquired resistance. Genetic analyses of relapsed patients have identified mechanisms of resistance, with a majority involving mutational activation of signaling components that drive reactivation of the key KRAS effector pathway, the three-tiered RAF-MEK-ERK mitogen-activated protein kinase cascade. Thus, ERK reactivation will limit the long-term efficacy of direct KRAS inhibitors. Despite the highly successful development of potent and selective inhibitors of each node of the ERK MAPK cascade, when used as monotherapy, they have shown little to no clinical efficacy against RAS-mutant cancers. Two key issues have contributed to this outcome, toxicity for normal tissues and de novo or treatment-induced acquired resistance in cancer cells. I propose that further delineation of the mechanisms by which ERK drives KRAS-dependent cancer growth will guide the development of more effective anti-ERK therapies. However, the mechanisms by which ERK drives PDAC growth remain poorly understood. One major unresolved issue is how ERK activity in different subcellular compartments supports cancer growth. Aim 1 studies comprise my K99 phase of training where I will take two complementary approaches to gain a better understanding of the role of cytoplasmic and nuclear ERK activity in supporting KRAS-dependent PDAC growth. First, I will determine the capacity of cytoplasmic versus nuclear ERK activity in supporting the growth of KRAS-mutant PDAC. Second, I will use a pharmacological inhibitor of the nuclear export protein exportin-1 (Selinexor) to determine whether it disrupts ERK cytoplasmic- nuclear dynamics and sensitizes PDAC models to KRAS inhibition. My Aim 2 studies comprise my R00- supported independent research and are based on our comprehensive ERK-dependent phosphoproteome/ transcriptome studies in KRAS-mutant PDAC. Using these data, I designed a CRISPR-Cas9 genetic loss-of- function screen library, targeting ERK regulated phosphoproteins and/or transcripts. I will now perform a system- wide determination of how ERK contributes to PDAC tumorigenesis as well as identify new ERK dependent targets to combine with KRAS inhibitors. To help me achieve these research goals and successfully transition to the independent phase I have an exceptional mentoring committee comprised of leading researchers in the study of KRAS signaling and therapeutics (Channing Der), in ERK spatiotemporal signaling (Jin Zhang), and ERK substrate utilization (John Blenis). With their guidance I am confident that I will successfully transition to establish an independent research program where I will advance our knowledge on ERK signaling and therapeutics.