Amoeboid Membrane Dynamics in Prostate Cancer

描述(申请人提供)：前列腺癌(PCA)是癌症死亡的主要原因，目前还没有有效的治疗方法来治疗“去势抵抗”疾病。这个项目是基于我们最近的发现，一种肌动蛋白核蛋白，透明相关的Forin-3(DRF3)，是人前列腺癌细胞中“阿米巴”表型的抑制因子，并且DRF3编码区的染色体丢失在转移性疾病中发生的频率很高，但在原发病中不是。功能研究表明，DRF3是表皮生长因子受体(EGFR)网络的一员，位于控制间充质细胞向阿米巴癌细胞表型转化的信号节点。间充质-阿米巴样转变(MAT)是一种动态的表型转变，可影响肿瘤细胞在基质中的迁移，降低其对蛋白水解酶抑制剂的敏感性。DRF3还抑制能够改变肿瘤微环境的生物活性微囊的分泌，并与富含胆固醇的脂筏膜微域相关。这些数据表明，DRF3网络可能对胆固醇或脂肪酸靶向治疗和/或化学预防很敏感，这在人类研究和动物模型中显示出了希望。我们将检验这一假设，即DRF3缺失引起的变形体特性会导致更具侵略性的表型。我们进一步假设，变形体的行为与脂质合成代谢的上调相一致，并且变形体肿瘤细胞可能会对脂质依赖的途径上瘾。第三，我们假设存在一个独特的阿米巴信号网络，该网络的组成部分可能代表新的肿瘤生物标记物。具体目标是：目的1.确定DRF3缺失在前列腺癌中的生物学后果。雄激素、雄激素受体和ERBB受体酪氨酸激酶信号在阿米巴表型中的作用将在DRF3沉默的背景下确定。我们将评估DRF3是否积极地介导了向间充质表型的转变。我们将评估DRF3沉默对肿瘤生长、转移以及对ERBB和胆固醇靶向治疗以及雄激素消融的敏感性的影响。DRF3在人类前列腺癌中的表达模式将被评估并与临床参数相关联。目的2.确定前列腺癌间充质-阿米巴转化的信号网络。在稳定的DRF3沉默和EGFR激活的条件下，评估RNA、蛋白质和棕榈酰蛋白质网络的扰动，将采用一系列定向和无偏倚的实验，以揭示控制前列腺癌细胞MAT的信号网络。将根据这些数据构建模型，并进行验证，以确定垫子的一小系列信息性指标。这些标记物将被用来确定是否可以使用与预后评估相关的最先进的平台在体内检测到阿米巴表型。这些研究将为膜动力学、脂代谢和前列腺癌进展到致死性疾病之间的关系提供新的见解。

DESCRIPTION (provided by applicant): Prostate adenocarcinoma (PCa) is a leading cause of death from cancer and there is no effective treatment for "castration-resistant" disease. This project is based on our recent findings that an actin-nucleating protein, Diaphanous related formin-3 (DRF3), is an inhibitor of the "amoeboid" phenotype in human PCa cells, and that chromosomal loss at the DRF3 coding locus occurs at high frequency in metastatic but not primary disease. Functional studies have demonstrated that DRF3 is a member of the EGF receptor (EGFR) network and is positioned at a signaling node that controls the transition from mesenchymal to amoeboid cancer cell phenotypes. The mesenchymal-amoeboid transition (MAT) has recently been identified as a dynamic phenotypic shift that can affect migration of tumor cells through matrices and reduce their sensitivity to protease inhibitors. DRF3 also inhibits the secretion of bioactive microvesicles capable of altering the tumor microenvironment and associates with cholesterol-rich lipid raft membrane microdomains. These data suggest that the DRF3 network may be sensitive to cholesterol- or fatty acid-targeting therapies and/or chemoprevention, which have shown promise in human studies and animal models. We will test the hypothesis that amoeboid properties arising from DRF3 loss results in a more aggressive phenotype. We further hypothesize that amoeboid behavior coincides with upregulation of lipid anabolism and that amoeboid tumor cells may become addicted to lipid-dependent pathways. Thirdly, we hypothesize that a distinct amoeboid signaling network exists and that components of this network may represent novel tumor biomarkers. The specific aims are: Aim 1. Determine the biological consequences of DRF3 loss in prostate cancer. The role of androgen, the androgen receptor, and ERBB receptor tyrosine kinase signaling in the amoeboid phenotype will be determined in the context of DRF3 silencing. We will assess whether DRF3 actively mediates the transition to the mesenchymal phenotype. The effect of DRF3 silencing on tumor growth, metastasis, and sensitivity to ERBB- and cholesterol-targeting therapy, and androgen ablation will be assessed. Patterns of DRF3 expression in human prostate cancers will be evaluated and correlated with clinical parameters. Aim 2. Identify the signaling network that mediates the mesenchymal-amoeboid transition in prostate cancer. A series of directed as well as unbiased experiments, evaluating perturbations in RNA, protein, and palmitoyl-protein networks under conditions of stable DRF3 silencing and EGFR activation will be employed to uncover the signaling network controlling the MAT in prostate cancer cells. Models will be constructed from these data and validated to identify a small series of informative indicators of the MAT. These markers will be used to determine whether the amoeboid phenotype can be detected in vivo using a state-of-the-art platform relevant to prognostic evaluation. These studies will provide new insight into the relationship between membrane dynamics, lipid metabolism and PCa progression to lethal disease.