EGFR-VDR signals in diet-promoted inflammation and cancer

描述（由申请人提供）：饮食被认为在散发性结肠癌的发展中起重要作用。实验性结肠癌的氮氧甲烷（AOM）模型模拟了人类结肠癌的许多特征，包括西式高脂肪饮食对肿瘤的促进作用。炎症被认为有助于饮食增强的肿瘤发生。我们最近发现，膳食脂肪诱导的肿瘤促进需要表皮生长因子受体（EGFR）信号，并涉及促炎环氧合酶-2 （Cox-2）的增加。相反，我们证明膳食补充维生素D类似物F6-D3抑制炎症和肿瘤发生。维生素D效应是由维生素D受体（VDR）介导的。我们发现VDR可以拮抗来自关键炎症和致癌转录因子的信号，包括NFkB和b-catenin。在最近的初步研究中，我们发现在肿瘤发生过程中上调的EGFR信号抑制了VDR的表达。此外，VDR缺失小鼠似乎更容易发生肿瘤。在Aim 1中，我们将使用AOM模型来描述饮食和EGFR信号以及VDR下调之间的关系，并确定抑制饮食促进的炎症和肿瘤发生的VDR途径。我们将利用aom处理的VDR野生型和VDR零突变小鼠来解剖VDR通路。我们将调节膳食脂肪来评估饮食对炎症和肿瘤发生的影响。我们还将包括单独补充F6-D3或F6-D3加EGFR抑制剂吉非替尼的饮食，以分析饮食促进炎症和肿瘤发生中的VDR和EGFR信号。我们假设膳食脂肪会增加EGFR信号，降低VDR表达，而F6-D3会增加VDR水平，减少炎症和致癌信号，抑制肿瘤发生。在Aim 2中，我们将剖析结肠癌细胞和结肠成纤维细胞中下调VDR的EGFR通路，并揭示VDR对抗EGFR和促炎细胞因子信号的机制。Caco-2结肠癌细胞和结肠成纤维细胞表达功能性EGFR和VDR。在初步研究中，EGF下调了这些细胞的VDR，而EGFR抑制或活性维生素D上调了VDR。因此，在这些转化和未转化的结肠细胞中，EGFR的激活在体外模拟了EGFR信号对体内结肠肿瘤发生中VDR表达的影响。我们将揭示EGFR介导VDR下调的转录和转录后机制。潜在的机制包括抑制启动子激活，减少转录物或蛋白质半衰期。使用可诱导的显性负EGFR表达的转染物来控制EGFR，使用siRNA和cDNA策略来调节VDR水平，我们还将剖析控制炎症和肿瘤发生的EGFR和VDR途径。利用途径特异性蛋白质组学和基因组学方法，我们还将寻找新的EGFR和vdr调节的细胞因子和生长因子信号，这些信号可以在体外控制炎症，并在我们的体内模型中测试它们的相关性。这些研究将确定潜在的新目标，以开发化学预防策略。

DESCRIPTION (provided by applicant): Diet is believed to play a major role in the development of sporadic colon cancer. The azoxymethane (AOM) model of experimental colon cancer mimics many features of human colon cancer, including tumor promotion by Western style high fat diets. Inflammation is thought to contribute to diet-enhanced tumorigenesis. We recently showed that tumor promotion induced by dietary fat required epidermal growth factor receptor (EGFR) signals and involved increased pro-inflammatory cyclooxygenase-2 (Cox-2). In contrast, we demonstrated that dietary supplementation with a vitamin D analogue F6-D3 inhibited inflammation and tumorigenesis. Vitamin D effects are mediated by the vitamin D receptor (VDR). We showed that the VDR could antagonize signals from key inflammatory and oncogenic transcription factors, including NFkB and b-catenin. In recent preliminary studies, we demonstrated that EGFR signals, which are up-regulated in tumorigenesis, suppressed VDR expression. Furthermore, VDR null mice appear to be more susceptible to tumorigenesis. In Aim 1 using the AOM model we will characterize the relationships between diet and EGFR signals and VDR down- regulation and identify VDR pathways that inhibit diet-promoted inflammation and tumorigenesis. We will utilize AOM-treated VDR wild type and VDR null mutant mice to dissect VDR pathways. We will modulate dietary fat to assess the effects of diet on inflammation and tumorigenesis. We will also include a diet supplemented with F6-D3 alone, or F6-D3 plus EGFR inhibitor Gefitinib to dissect VDR and EGFR signals in diet-promoted inflammation and tumorigenesis. We postulate that dietary fat will increase EGFR signals and decrease VDR expression, whereas F6-D3 will increase VDR levels and reduce inflammatory and oncogenic signals and inhibit tumorigenesis. In Aim 2 we will dissect EGFR pathways in colon cancer cells and colonic fibroblasts that down-regulate VDR and uncover VDR mechanisms that oppose EGFR and pro- inflammatory cytokine signals. Caco-2 colon cancer cells and colonic fibroblasts express functional EGFR and VDR. In preliminary studies, EGF down-regulated the VDR in these cells, whereas EGFR inhibition or active vitamin D up-regulated VDR. Thus, EGFR activation in these transformed and non-transformed colonic cells mimics in vitro the effects of EGFR signals on VDR expression in colonic tumorigenesis in vivo. We will uncover the transcriptional and post-transcriptional mechanisms that mediate this VDR down-regulation by EGFR. Potential mechanisms include inhibited promoter activation, and reduced transcript or protein half-life. Using inducible dominant negative EGFR expressing transfectants to control EGFR, and siRNA and cDNA strategies to modulate VDR levels, we will also dissect EGFR and VDR pathways that control inflammation and oncogenesis. Using pathway-specific proteomic and genomic approaches, we will also search for new EGFR and VDR-regulated cytokine and growth factor signals that control inflammation in vitro and test their relevance in our in vivo models. These studies will identify potential new targets to exploit for chemopreventive strategies. PUBLIC HEALTH RELEVANCE: Colon cancer is the 3rd leading cause of cancer-related deaths in the US. More than half of these malignancies are believed to be preventable by dietary modifications. We have shown that tumor promotion by Western style high fat diets requires epidermal growth factor receptor (EGFR) signals that inhibit anti-inflammatory vitamin D signals. We will uncover EGFR and vitamin D cellular pathways that interact to modulate dietary inflammation and colonic tumor risk.