Inhibition of pancreatic carcinogenesis via targeting c-Raf and sEH

描述(申请人提供)：该项目的意义是开发一种通过靶向c-Raf和sEH来预防高度致命的胰腺癌的有效药物。早期发现和预防对降低胰腺癌死亡率具有重要意义。突变的Kras是参与人类胰腺癌发生的最常见和最早的分子事件；利用突变的Kras启动的小鼠肿瘤发生，已经证明c-Raf在Kras启动的肿瘤发生中是必需的，而敲除c-Raf阻止了Kras启动的肿瘤的发展。慢性胰腺炎是公认的胰腺癌危险因素。在突变的Kras启动的Pdx1-Cre/LSL-KrasG12D小鼠(称为PanKras)的胰腺肿瘤发生中，雨蛙素诱导的胰腺炎促进了恶性进展。除了胰腺炎，突变的Kras与突变的p53(第二种最常见的基因改变)合作，主要通过协同调节ERK-MAPK和NF-kB，导致小鼠胰腺癌的发生。抗炎EET可被促炎症的可溶性环氧化物水解酶(SEH)迅速灭活。我们的初步数据显示：1)96%(27/28)的人胰腺癌标本和90%(27/30)的人胰腺癌标本分别显示出明显的sEH和p-c-Raf的过度表达；2)我们的新型双重c-Raf/sEH抑制剂t-CUPM对PanKras小鼠的胰腺炎和mPanIN的形成有很强的抑制作用。我们的假设是，c-Raf和sEH在突变的kras启动的胰腺癌发生中起关键作用，并被胰腺炎或突变的p53增强；同时靶向这两种酶是一种新的和相关的胰腺癌预防策略。现提出三个具体目标来验证这一假说：1)确定c-Raf/sEH双重抑制剂t-CUPM、sEH抑制剂t-AUCB和Raf抑制剂Raf265/sorafenib对突变的Kras启动和胰腺炎促进的PanKras小鼠癌变的影响；2)确定使用抑制剂或基因敲除来阻断突变的Kras激活的c-Raf是否能够抑制突变的Kras-p53的相互作用和Pdx1-KrasG12D-p53R172小鼠的恶性进展；以及3)利用sEH基因敲除的方法确定sEH在PanKras小鼠的胰腺炎和癌症发生中的作用。这项研究的成功不仅将引导我们阐明c-Raf和sEH在胰腺癌发生中的作用，而且还将确立t-CUPM作为一种新的抑制胰腺癌发展的药物的意义。

DESCRIPTION (provided by applicant): The significance of this project is to develop an efficient agent for the prevention of highly lethal pancreatic cancer through targeting both c-Raf and sEH. Early detection and prevention is of great importance to reduce the mortality of pancreatic cancer. Mutant Kras is the most common and earliest molecular event involved in human pancreatic carcinogenesis; and using mutant Kras-initiated tumorigenesis in mice, it has been demonstrated that c-Raf is required for Kras-initiated tumorigenesis and knockout c-Raf blocks Kras- initiated cancer development. Chronic pancreatitis is a well-recognized risk factor of pancreatic cancer. In mutant Kras-initiated pancreatic tumorigenesis in Pdx1-Cre/LSL-KrasG12D mice (called PanKras), caerulein- induced pancreatitis enhances malignant progression. In addition to pancreatitis, mutant Kras cooperates with mutant p53 (the second most common genetic alteration) leading to pancreatic cancer development in mice, mainly via synergistically regulating ERK-MAPK and NF-kB. Anti-inflammatory EETs are quickly inactivated by pro-inflammatory soluble epoxide hydrolase (sEH). Our preliminary data showed that 1) 96% (27/28) and 90% (27/30) of human pancreatic adenocarcinoma samples displayed a distinct over-expression of sEH and p-c-Raf, respectively, and 2) our novel dual c-Raf/sEH inhibitor t-CUPM exhibited strong inhibitory effect on pancreatitis and mPanIN formation in PanKras mice. Our hypothesis is that c-Raf and sEH play key roles in pancreatic carcinogenesis initiated by mutant Kras and enhanced by pancreatitis or mutant p53; and targeting these two enzymes together is a novel and relevant preventive strategy for pancreatic cancer. Three specific aims are proposed to test this hypothesis: 1) to determine the effects of c-Raf/sEH dual inhibitor t-CUPM, sEH inhibitor t- AUCB and Raf inhibitor Raf265/sorafenib on mutant Kras-initiated and pancreatitis-enhanced carcinogenesis in PanKras mice; 2) to determine whether blocking mutant Kras-activated c-Raf using inhibitors or gene knockout suppresses mutant Kras-p53 interaction and malignant progression in Pdx1-KrasG12D-p53R172 mice; and 3) to determine the role of sEH in pancreatitis and carcinogenesis using the approach of sEH gene knockout in PanKras mice. Successfulness of the proposed study will not only lead us to demonstrate the role of c-Raf and sEH in pancreatic carcinogenesis, but also to establish a significance of t-CUPM as a novel agent for inhibiting the pancreatic cancer development.