Prevention of Tumor Progression by a Novel Approach

描述（由申请人提供）：新出现的证据表明，癌症进展通常与上皮细胞向间充质细胞转化（EMT）表型的获得相关，EMT表型使人联想到“癌症干细胞样细胞（CSC）"，其部分负责胰腺癌（PC）细胞获得侵袭性的能力，从而促进肿瘤转移。我们发表的数据显示，在吉西他滨抗性EMT型细胞中，miR-200的表达降低，导致ZEB 1的上调，从而导致E-钙粘蛋白的下调，并且还显示miR-21的表达增加，导致PTEN表达失活。我们还发现，与正常志愿者相比，PC患者血浆中的miR-21水平显著较高，这与生存率恶化相关。基于这些结果，我们推断，找到一种新的无毒途径，通过这种途径可以上调miR- 200，下调miR-21，这将是一种预防肿瘤进展的新方法。为此，我们首次发表了一种名为CDF的无毒天然药物姜黄素的合成类似物在其靶组织中具有上级效果CDF在消除EMT表型细胞方面非常有效，这部分是由于miR-200的上调和miR-21的下调，导致ZEB 1表达下调，E-cadherin和PTEN表达增加。因此，我们似乎发现了一种新的方法，通过该方法可以实现耐药EMT型细胞的生物环境的“调节”，以有效消除EMT型细胞或CSC，这可以通过消除肿瘤复发的“根本”原因来预防肿瘤进展。基于我们的初步结果，我们假设miR-21的激活在吉西他滨耐药（GR）PC细胞中获得EMT表型过程中至关重要，并且这些细胞可以通过CDF单独或与常规治疗剂组合来消除。我们将测试我们的假设，以获得对miR-21和PTEN在GR细胞中获得EMT表型过程中的作用的机制见解，并通过操纵miR-21（Aim-1）的表达来研究这些细胞的生物学后果。我们还将研究CDF如何下调miR- 21并上调其靶点的表达。这可能导致PC细胞的生物环境的“调节”，导致细胞对常规试剂的致敏（Aim-2;体外研究）。最后，我们将使用异种移植物和K-ras转基因动物模型确定与单独的CDF相比，CDF与体内常规治疗剂组合时是否可以引起增加的抗肿瘤活性（Aim-3）。我们的研究结果将提供关于miR-21在耐药EMT型细胞（CSC）中的作用的机制见解，并且还将提供支持CDF在预防肿瘤进展和/或治疗PC中的作用的临床前数据。因此，我们的研究结果将对预防肿瘤复发产生显著的高度影响，这将导致诊断为PC的患者获得更好的生存结局。

DESCRIPTION (provided by applicant): Emerging evidence suggests that cancer progression is often associated with the acquisition of Epithelial-to- Mesenchymal Transition (EMT) phenotype that is reminiscent of "Cancer Stem-like Cells (CSCs)", which is partly responsible for the ability of pancreatic cancer (PC) cells to acquire aggressiveness, contributing to tumor metastasis. Our published data showed decreased expression of miR-200 in gemcitabine resistant EMT- type cells, resulting in the up-regulation of ZEB1 and thereby down-regulation of E-cadherin, and also showed increased expression of miR-21, resulting in the inactivation of PTEN expression. We also found significantly higher levels of miR-21 in the plasma of PC patients compared to normal volunteers, which was correlated with worsened survival. Based on these results, we reasoned that finding a novel non-toxic avenue by which miR- 200 could be up-regulated and miR-21 could be down-regulated would be a novel approach for the prevention of tumor progression. To that end, we have published, for the first time, that a synthetic analog of a non-toxic natural agent curcumin, named CDF is superior in its target tissue (pancreas) bioavailability without any adverse side effects, and the CDF was very effective in the elimination of EMT phenotypic cells, which was in part due to up-regulation of miR-200 and down-regulation of miR-21, resulting in the down-regulation of ZEB1, and increased expression of E-cadherin and PTEN. Therefore, it appears that we found a novel approach by which "conditioning" of the biological milieu of drug-resistant EMT-type cells could be achieved toward effective elimination of EMT-type cells or CSCs, which could be useful for the prevention of tumor progression by eliminating the "root" cause of tumor recurrence. Based on our preliminary results, we hypothesize that the activation of miR-21 is critical during the acquisition of EMT phenotype in gemcitabine-resistant (GR) PC cells, and these cells could be eliminated by CDF alone or in combination with conventional therapeutics. We will test our hypothesis to gain mechanistic insight on the role of miR-21 and PTEN during the acquisition of EMT phenotype in GR cells, and investigate the biological consequence of these cells by manipulating the expression of miR-21 (Aim-1). We will also investigate how CDF could down-regulate miR- 21 and up-regulate the expression of its targets. This may lead to the "conditioning" of the biological milieu of PC cells, resulting in sensitization of cells to conventional agents (Aim-2; in vitro studies). Finally, we will determine whether CDF could cause increased anti-tumor activity when combined with conventional therapeutics in vivo using xenograft and K-ras transgenic animal models compared to CDF alone (Aim-3). The results of our studies will provide mechanistic insight as to the role of miR-21 in drug-resistant EMT-type cells (CSCs), and will also provide pre-clinical data in support of the role of CDF for the prevention of tumor progression and/or treatment of PC. Therefore, our results will have significantly high impact toward preventing tumor recurrence, which will lead to achieve better survival outcome of patients diagnosed with PC.