Anti-pancreatic tumorigenesis by inactivation of SAG/RBX2 E3 ubiquitin ligase

描述(申请人提供)：凋亡敏感基因(SAG，人；SAG，小鼠)，也被称为RBX2/ROC2，是SCF(Skp1，Cullins，F-box蛋白)E3泛素连接酶的重要环成分，通过促进各种关键调控蛋白的泛素化和降解，控制几个重要的生物学过程，包括细胞周期进程、信号转导、转录和DNA复制。我们已发表和未发表的资料显示SAG与胰腺癌的直接关联如下：a)SAG在胰腺癌组织中过表达，SAG核表达与胰腺癌患者的预后不良有关；b)SAG siRNA敲除导致肿瘤抑制蛋白的积聚，抑制胰腺癌细胞的存活和贴壁非依赖性生长，以及c)SAG siRNA敲除显著抑制胰腺癌细胞的体内原位生长和基质侵袭。这些研究表明，SAG的过度表达可能在胰腺肿瘤的发生过程中起致癌作用。本应用的目的是利用胰腺特异性SAG转基因或KO小鼠模型，从机制上研究SAG在Kras激活(KrasG12D)和p53突变(P53R172H)引发的胰腺肿瘤发生中的作用。中心假说是SAG通过促进肿瘤抑制底物如DEPTOR、I？B和p21/p27的降解，导致mTOR和NF？B通路的激活，从而促进肿瘤的发生。因此，SAG-胰腺转基因表达将促进KrasG12D启动的胰腺肿瘤的发生，而SAG胰腺的缺失将通过肿瘤抑制蛋白的积聚来阻断肿瘤信号，从而抑制KrasG12D和p53R172H引发的胰腺肿瘤的发生。有两个特定的目的：1)确定SAG在KrasG12D/p53R172H诱导的胰腺肿瘤发生中的作用；2)阐明SAG调控胰腺肿瘤发生的机制(S)。影响：本研究采用小鼠胰腺导管腺癌(PDAC)模型，探讨SAG E3连接酶在PDAC发生发展中的作用，并阐明其作用机制。我们的研究将从机制上验证SAG E3泛素连接酶作为一种新的抗胰腺癌靶点，并为未来发现SAG E3连接酶的特异性抑制剂作为一类新的抗胰腺癌药物提供概念验证证据。因此，我们的工作具有很高的创新性和重大影响，具有翻译价值。

DESCRIPTION (provided by applicant): Sensitive to Apoptosis Gene (SAG, human; Sag, mouse) also known as RBX2/ROC2, is an essential RING component of SCF (SKP1, Cullins, F-box proteins) E3 ubiquitin ligase which, by promoting ubiquitination and degradation of various key regulatory proteins, controls several important biological processes including cell cycle progression, signal transduction, transcription, and DNA replication. Our published and unpublished data showed a direct association of SAG with pancreatic cancer as follows: a) SAG is overexpressed in pancreatic cancer tissues, and SAG nuclear expression is associated with poor prognosis of pancreatic cancer patients; b) SAG siRNA knockdown caused accumulation of tumor suppressive proteins and inhibited survival and anchorage-independent growth of pancreatic cancer cells, and c) orthotopic in vivo growth and matrigel invasion of pancreatic cancer cells were significantly inhibited, respectively, by SAG siRNA knockdown. All these studies suggested that SAG overexpression could be oncogenic during pancreatic tumorigenesis. The objectives of this application are to use pancreatic specific Sag transgenic or KO mouse models to mechanistically study the role of Sag in pancreatic tumorigenesis triggered by Kras activation (KrasG12D) and p53 mutation (p53R172H). The central hypothesis is that SAG promotes tumorigenesis by promoting the degradation of tumor suppressive substrates such as DEPTOR, I¿B, and p21/p27, leading to activation of the mTOR and NF¿B pathways. Thus, SAG-pancreatic transgenic expression would promote pancreatic tumorigenesis, initiated by KrasG12D, whereas Sag pancreatic deletion would abrogate oncogenic signals by accumulation of tumor suppressive proteins, thus suppressing pancreatic tumorigenesis triggered by KrasG12D and p53R172H. Two specific aims are proposed to 1) determine the role of Sag in KrasG12D/p53R172H-induced pancreatic tumorigenesis and 2) elucidate mechanism(s) by which Sag regulates pancreatic tumorigenesis. IMPACT: Our study uses mouse models that recapitulate the development of pancreatic ductal adenocarcinoma (PDAC) to investigate the role of Sag E3 ligase in the initiation and progression of PDAC and to elucidate its mechanism of action. Our study will mechanistically validate SAG E3 ubiquitin ligase as a novel anti-pancreatic cancer target and provide proof-of-concept evidence for future discovery of specific inhibitor of SAG E3 ligase as a novel class of anti-pancreatic cancer drugs. Our work is, therefore, highly innovative and of significant impact with translational value.