Ki-Ras Signaling in Pancreatic Cancer: Role of a Novel Akt Regulator

描述（由申请人提供）：胰腺导管腺癌（PDAC）患者的5年生存率约为5%，在过去25年中基本保持不变。当务之急是迅速确定这种癌症的潜在机制 并进行研究，以便开发新的分子疗法。在大多数情况下，响应于Ki-Ras基因中的遗传改变以及胰腺癌发生的其他公认标志而失调的信号传导机制仍不完全清楚。长期目标是阐明磷酸肌醇3-激酶（PI 3 K）信号通路及其蛋白激酶介导剂Akt在PDAC中的作用和调节，并开发比目前可用的更有效的针对它们的拮抗剂用于癌症预防和治疗。为了实现这一目标，以Akt为诱饵的改良酵母双杂交筛选有助于将BSTA鉴定为先前未知的PI 3 K/Akt信号转导介体。本提案的目的是进一步了解BSTA的调节和功能以及阻断其活性对PDAC的影响。中心假设是BSTA是致癌Ki-Ras诱导的Akt信号传导的关键调节因子，并促进胰腺肿瘤发生。基于强有力的初步数据，这一假设将通过追求以下两个具体目标进行检验：1）确定 Ki-Ras激活Akt的机制;和2）研究BSTA在体外和体内PDAC对mTOR抑制剂的抗性中的作用。将使用基于RNAi的方法和蛋白质磷酸化测定来检验工作假设（目的1），即致癌Ki-Ras通过以下方式激活PDAC细胞中的Akt：a）通过涉及Ral小GTP酶和microRNA miR-21的机制增加BSTA表达，和B）在BSTA依赖性机制中刺激Akt激酶（mTORC 2）和/或抑制Akt磷酸酶（PHLPP）。为了测试BSTA是否促进胰腺肿瘤发生和耐药性（目标2），将描绘BSTA上的最小Akt结合序列，并设计细胞可穿透的肽以在PDAC细胞上测试。BSTA也将在PDAC细胞中下调，以研究原位小鼠模型中的肿瘤发生和对mTOR抑制剂的抗性。拟议项目的基本原理是，了解BSTA的作用和调节可能有助于确定靶向PI 3 K通路和致癌Ki-Ras信号传导的新方法，以有效治疗PDAC。这一贡献是重要的，因为它有望帮助揭示K-RasG 12 D额外控制Akt的新机制的细节，并导致开发靶向胰腺导管癌中PI 3 K/Akt通路的替代药理学策略。这项研究提案是创新的，因为它描述了致癌Ki-Ras和Akt之间的一种新的机制联系，一旦正确划定，将提供一种新的和实质上不同的方法来靶向PI 3 K/Akt通路。总之，这些研究有望通过从根本上推进对Ki-Ras和PI 3 K/Akt信号传导的理解并发现治疗胰腺导管癌的新分子靶点而产生积极影响。 公共卫生相关性：胰腺癌是一种高度侵袭性和快速致命的疾病。这项资助申请中提出的研究有望从根本上推进我们对Ki-Ras和PI 3 K/Akt信号传导的理解，并发现治疗胰腺导管癌的新分子靶点和替代治疗策略。因此，拟议的研究与公共卫生和NIH使命相关，如“NCI胰腺癌治疗临床试验计划会议共识报告”所定义。

DESCRIPTION (provided by applicant): The 5-year survival rate for Pancreatic ductal adenocarcinoma (PDAC) patients is about 5% and has remained largely unchanged over the past 25 years. It is imperative that the mechanisms that underlie this cancer be rapidly identified and investigated so that novel molecular therapies can be developed. For the most part, the signaling mechanisms deregulated in response to genetic alterations in the Ki-Ras gene, and by other well- established hallmarks of pancreatic carcinogenesis, are still incompletely understood. The long-term goal is to elucidate the role and regulation of the phosphoinositide 3-kinase (PI3K) signaling pathway and its protein ki- nase mediator Akt in PDAC and develop more effective antagonists against them for cancer prevention and therapy than are currently available. In pursuit of this goal, a modified yeast two-hybrid screen with Akt as bait helped identify BSTA as a previously unknown mediator of PI3K/Akt signaling. The objective of this proposal is to further understand BSTA regulation and function and the effects of blocking its activity on PDAC. The central hypothesis is that BSTA is a key regulator of oncogenic Ki-Ras-induced Akt signaling and promotes pancreatic tumorigenesis. Based on strong preliminary data, this hypothesis will be tested by pursuing the fol- lowing two specific aims: 1) Identify the mechanism by which Ki-Ras activates Akt; and 2) Investigate the role of BSTA in the resistance of PDAC to mTOR inhibitors in vitro and in vivo. An RNAi-based approach, and pro- tein phosphorylation assays, will be used to test the working hypothesis (Aim 1) that oncogenic Ki-Ras acti- vates Akt in PDAC cells by a) increasing BSTA expression via a mechanism involving the Ral small GTPases and microRNA miR-21, and b) stimulating Akt kinase (mTORC2) and/or suppressing Akt phosphatase (PHLPP) in a BSTA-dependent mechanism. To test if BSTA promotes pancreatic tumorigenesis and drug re- sistance (Aim 2), the minimal Akt-binding sequence on BSTA will be delineated and cell-penetrable peptides designed to test on PDAC cells. BSTA will be also be downregulated in PDAC cells to investigate tumorigene- sis and resistance to mTOR inhibitors in an orthotopic mouse model. The rationale for the proposed project is that understanding the role and regulation of BSTA may help identify novel ways to target the PI3K pathway and oncogenic Ki-Ras signaling for the effective treatment of PDAC. This contribution is significant because it is expected to help reveal details of a novel mechanism by which K-RasG12D additionally controls Akt and lead to the development of alternative pharmacologic strategies for targeting the PI3K/Akt pathway in pancrea- tic ductal cancer. The research proposal is innovative because it describes a novel mechanistic link between oncogenic Ki-Ras and Akt that will, once properly delineated, offer a new and substantially different approach for targeting the PI3K/Akt pathway than currently available. Together, these studies are expected to have a positive impact by fundamentally advancing the understanding of Ki-Ras and PI3K/Akt signaling and uncover new molecular targets for the treatment of pancreatic ductal cancer. PUBLIC HEALTH RELEVANCE: Pancreatic cancer is a highly aggressive and rapidly fatal disease. The studies proposed in this grant application may be expected to fundamentally advance our understanding of Ki-Ras and PI3K/Akt signaling and uncover new molecular targets and alternative therapeutic strategies for the treatment of pancreatic ductal cancer. The proposed research is therefore relevant to public health and to the NIH mission as defined in the "Consensus Report of the NCI Clinical Trials Planning Meeting on Pancreas Cancer Treatment".