Prevention of Tumor Progression by a Novel Approach

描述(申请人提供)：新的证据表明，癌症的进展往往与获得上皮间充质转化(EMT)表型有关，这使人想起“癌症干细胞(CSCs)”，这在一定程度上导致胰腺癌(PC)细胞获得侵袭性，从而导致肿瘤转移。我们发表的数据显示，在吉西他滨耐药的EMT细胞中，miR-200的表达降低，导致ZEB1的表达上调，从而下调E-钙粘蛋白的表达，同时miR-21的表达增加，导致PTEN的表达失活。我们还发现，与正常志愿者相比，PC患者血浆中miR-21水平显著升高，这与生存恶化有关。基于这些结果，我们推测寻找一种新的无毒途径，通过这种途径可以上调miR-200，下调miR-21，这将是一种防止肿瘤进展的新途径。为此，我们首次发表了一种无毒天然药物姜黄素的合成类似物，名为CDF，其靶向组织(胰腺)的生物利用度更高，没有任何副作用，CDF在消除EMT表型细胞方面非常有效，部分原因是miR-200上调，miR-21下调，导致ZEB1下调，E-钙粘素和PTEN表达增加。因此，我们似乎找到了一种新的方法，通过对耐药的EMT型细胞的生物学环境进行条件调节，从而有效地消除EMT型细胞或CSCs，这可能有助于通过消除肿瘤复发的根本原因来防止肿瘤进展。根据我们的初步结果，我们假设miR-21的激活在获得吉西他滨耐药(GR)PC细胞的EMT表型过程中起关键作用，这些细胞可以被CDF单独或与传统疗法联合消除。我们将验证我们的假设，从机制上了解miR-21和PTEN在GR细胞获得EMT表型过程中的作用，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和上调其靶标的表达。这可能导致PC细胞的生物学环境的“条件作用”，导致细胞对常规药物的敏感性(AIM-2；体外研究)。最后，我们将确定与单独使用CDF(AIM-3)相比，CDF与使用异种移植和K-ras转基因动物模型的体内常规治疗药物联合使用时，是否可以导致抗肿瘤活性增加。我们的研究结果将为深入了解miR-21在耐药EMT细胞(CSCs)中的作用提供机制方面的见解，也将为CDF在预防肿瘤进展和/或治疗PC中的作用提供临床前数据支持。因此，我们的研究结果将对预防肿瘤复发具有重要意义，这将导致PC患者获得更好的生存结果。

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.