Alternative splicing and epithelial-mesenchymal plasticity in prostate tumors

上皮细胞中成纤维细胞生长因子受体2（FGFR 2）转录本的选择性剪接 导致FGFR 2（IIIb）同种型的表达，而在间充质细胞中， 导致包括外显子IIIc的FGFR 2（IIIc）的合成。这种受调控的剪接被破坏 在前列腺癌的进展过程中导致FGFR 2的不适当表达 同种型。为了观察FGFR 2外显子IIIc在同基因大鼠Dunning AT 3肿瘤中的应用，我们 构建了报告可变剪接的小基因结构。想象这些选择性剪接 结果显示，在这些原发性肿瘤中存在意想不到的间充质-上皮转化（MET）。 这种上皮间质可塑性的理解，揭示了剪接的差异，是 这个应用程序的首要目标。 为此，我们建议实现以下具体目标：具体目标1。到 表征FGFR 2剪接和上皮-间充质可塑性之间的联系 前列腺肿瘤我们已经开发出一种新的双色报告，可用于检测两者 MET和上皮-间充质转化（EMT）。我们还将我们的观察扩展到 在培养物中表征人前列腺肿瘤细胞。在这方面完成的工作将检验 假设上皮可塑性是侵袭性前列腺癌的一般特性， 人类具体目标2。为了表征原发性骨髓瘤中上皮-间充质可塑性， 通过剪接的改变揭示的前列腺肿瘤。在这个目标中，我们将表征分子 原发性肿瘤中MET簇的特征和行为。具体目标3。表征 上皮-间质可塑性和转移行为之间的联系。我们将 研究上皮-间质可塑性和转移行为之间的联系。 最后，我们将从AT 3大鼠肿瘤和患者中分离循环前列腺肿瘤细胞， 我们将确定他们的FGFR 2剪接状态和他们的上皮状态。目标完成的工作 两个，并在这一目标将测试假设，上皮间质可塑性导致增加 肿瘤适应性 这项工作的意义是双重的：首先，我们将揭示基因的基本信息， 在前列腺肿瘤进展过程中由选择性剪接控制的表达程序。二是 前列腺癌作为一种健康威胁的严重性强调了其对人类健康的潜在重要性 问题.识别将发展为转移性疾病的肿瘤/细胞是一个挑战， 要务这里提出的工作应该确定有用的标志物，甚至可能是治疗性的 目标的

In epithelial cells alternative splicing of fibroblast growth factor receptor-2 (FGFR2) transcripts leads to the expression of the FGFR2(IIIb) isoform, whereas in mesenchymal cells the same process results in the synthesis of FGFR2(IIIc), which include exon IIIc. This regulated splicing is disrupted during progression of prostate carcinomas leading to the inappropriate expression of FGFR2 isoforms. To visualize the use of FGFR2 exon IIIc in Dunning AT3 tumors in syngeneic rats we constructed minigene constructs that report on alternative splicing. Imaging these alternative splicing decisions revealed unexpected mesenchymal-epithelial transitions (MET) in these primary tumors. The understanding of this epithelial-mesenchymal plasticity, revealed by differences in splicing, is the overarching goal of this application. To this end we propose to accomplish the following specific aims: Specific Aim 1. To characterize the connections between FGFR2 splicing and epithelial-mesenchymal plasticity in prostate tumors. We have developed a new bichromatic reporter that can be used to detect both MET and epithelial-mesenchymal transitions (EMT). We have also extended our observations to well characterized human prostate tumor cells in culture. Work accomplished in this aim will test the hypothesis that epithelial plasticity is a general property of aggressive prostate carcinomas in humans. Specific Aim 2. To characterize the epithelial-mesenchymal plasticity in primary prostate tumors revealed by alterations in splicing. In this aim we will characterize the molecular signatures and behavior of the MET clusters in primary tumors. Specific aim 3. To characterize the connections between epithelial-mesenchymal plasticity and metastatic behavior. We will investigate the connections between epithelial-mesenchymal plasticity and metastatic behavior. Finally, we will isolate circulating prostate tumor cells from both the AT3 rat tumors and patients, and we will determine their FGFR2 splicing status and their epithelial state. Work accomplished in aim two and in this aim will test the hypothesis that epithelial-mesenchymal plasticity leads to increased tumor fitness. The significance of this work is twofold: First we will unravel basic information about gene expression programs controlled by alternative splicing during prostate tumor progression. Second, the potential significance to human health is underscored by the magnitude of prostate cancer as a health problem. Identifying the tumors/cells that will progress to metastatic disease is a challenge and a priority. The work proposed here should identify useful markers and perhaps even therapeutic targets.