Amoeboid Membrane Dynamics in Prostate Cancer

描述（由申请人提供）：前列腺腺癌（PCa）是癌症死亡的主要原因，目前尚无有效治疗“去势抵抗”疾病的方法。这个项目是基于我们最近的发现，一种肌动蛋白成核蛋白，透明相关的形成蛋白-3 (DRF3)，是人类PCa细胞中“变形虫”表型的抑制剂，并且DRF3编码位点的染色体丢失在转移性疾病中发生频率很高，但不是原发疾病。功能研究表明，DRF3是EGF受体（EGFR）网络的成员，位于控制间充质癌细胞表型向变形虫癌细胞表型转变的信号节点。间充质-变形虫转变（MAT）最近被确定为一种动态表型转变，可以影响肿瘤细胞通过基质的迁移并降低其对蛋白酶抑制剂的敏感性。DRF3还抑制生物活性微泡的分泌，这些微泡能够改变肿瘤微环境，并与富含胆固醇的脂质筏膜微域相关。这些数据表明，DRF3网络可能对胆固醇或脂肪酸靶向治疗和/或化学预防敏感，这在人类研究和动物模型中显示出前景。我们将验证由DRF3缺失引起的变形虫特性导致更具侵袭性表型的假设。我们进一步假设变形虫行为与脂质合成代谢上调相一致，变形虫肿瘤细胞可能对脂质依赖途径上瘾。第三，我们假设存在一个独特的变形虫信号网络，并且该网络的组成部分可能代表新的肿瘤生物标志物。具体目标是：目标1。确定前列腺癌中DRF3缺失的生物学后果。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是否积极介导向间质表型的转变。将评估DRF3沉默对肿瘤生长、转移、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。目标2。确定在前列腺癌中介导间质-变形虫转变的信号网络。在DRF3稳定沉默和EGFR激活的条件下，一系列定向和无偏实验，评估RNA、蛋白质和棕榈酰蛋白网络的扰动，将被用来揭示前列腺癌细胞中控制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MAT的一小部分信息指标。这些标记物将用于确定是否可以使用与预后评估相关的最先进平台在体内检测到变形虫表型。这些研究将为研究膜动力学、脂质代谢与前列腺癌发展为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.