Ral in human bladder cancer progression

Ral 在人膀胱癌进展中的作用

基本信息

批准号：
7600785
负责人：
DAN THEODORESCU
金额：
$ 3.81万
依托单位：
UNIVERSITY OF VIRGINIA
依托单位国家：
美国
项目类别：
财政年份：
1997
资助国家：
美国
起止时间：
1997-09-01 至 2008-06-30
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7600785
关键词：
1-Phosphatidylinositol 3-Kinase Actins Address Adhesions Affect Anus Back Basement membrane Biochemistry Biological Biological Assay Biology Bladder Bladder Neoplasm Bladder Urothelium Cancer cell line Cell Line Cell membrane Cells Cellular Morphology Clinical Oncology Clinical Research Complex Connective Tissue Cytoskeleton Data Development Diagnostic Disease EGF gene EGFR Protein Overexpression Endopeptidases Epidermal Growth Factor Receptor Epithelial Family Fibroblasts Foundations Freezing Funding Future GTP-Binding Proteins Genes Genetic Growth Factor Guanine Nucleotide Dissociation Inhibitors Guanosine Triphosphate Phosphohydrolases HRAS gene Human In Vitro Incidence Invaded Invasive Knockout Mice Laboratories Lead Life Malignant Neoplasms Malignant neoplasm of urinary bladder Mediating Membrane Modeling Molecular Molecular Weight Mucous Membrane Mus Muscle Mutate Mutation PTEN gene Pathway interactions Patients Peptide Hydrolases Phenotype Phosphotransferases Process Production Protein Overexpression Proteins RRAS2 gene Range Rattus Receptor Activation Receptor Signaling Regulation Research Personnel Risk Role Role playing therapy Running Second Messenger Systems Sequence Homology Signal Pathway Signal Transduction Specimen Stimulus Stress Fibers Technology Testing Time Tissue Banks Tissues Transgenic Animals Transgenic Organisms Transport Vesicles Tumor Cell Invasion Tumor Suppressor Genes United States base bladder Carcinoma cancer cell cell growth cell motility follow-up human RRAS2 protein human tissue in vivo mRNA Expression member migration novel p120 GTPase Activating Protein phospholipase C gamma polymerization prognostic programs ras GTPase-Activating Proteins receptor reconstitution response rho second messenger tool tumor tumor progression

项目摘要

Background and Significance: 40% of patients presenting with "superficial" (non-muscle-invasive) bladder cancer develop the "invasive" life-threatening form of the disease during follow up. In clinical studies, overexpression of Epidermal Growth Factor Receptor (EGFR), Ha-Ras mutation and loss of tumor suppressor gene PTEN have been associated with this phenotypic tumor transition. However, the exact molecular pathway by which these genes effectively trigger or facilitate the invasive process is incompletely understood. Our original R29 hypothesized that EGFR signaling enhances bladder tumor motility in vitro and invasion in vivo and intended to determine the signaling pathways used by EGFR in this process. Since funding of the R29 in 9/97, we have made the following important observations which support the original hypothesis and address the aims of the original application: 1) EGFR and Ras inhibition diminished the motility of invasive bladder cancer cells; 2) EGF stimulates motility in non-invasive cells via PI3K and this requires activity of Rho and Ras effector Rat; 3) In non-invasive cells, baseline RalA activity is low while invasive cells have constitutively higher activation; 4) Invasive cells have low levels of RhoGDI2 expression. Reconstitution of this gene leads to diminished motility and activity of RalA but not RhoA suggesting this gene may be the first RalGDI identified to function as an invasion suppressor; 5) Inhibition of PI3K activity via PTEN reconstitution in invasive cells with inactive PTEN, results in an inhibition of orthotopic invasion in vivo and a decrease in RhoA activity. Since the overall biology of both Ral and RhoGDI2 is poorly understood, but might be critical for regulating tumor invasion in patients with bladder cancer, we propose the Guiding Hypothesis that EGF mediates bladder tumor invasion via Ral activation. We will test this hypothesis with a matrix of technologies ranging from basic biochemistry to clinical oncology to address Ral biology in human bladder cancer. These include: 1) unique paired human bladder cancer cell lines with different invasive abilities; 2) a novel organotypic bladder model allowing in vitro study of tumor invasion; 3) an orthotopic assay evaluating the effects of candidate molecules on in vivo bladder cancer invasion; 4) transgenic and knockout mice with appropriate genetic and phenotypic profiles; 5) a human tissue bank with pathologically and clinically well characterized frozen specimens. Specific Aims: 1) Determine the role and pathobiology of Ral in bladder cancer invasion in organotypic, murine orthotopie and human tumor studies; 2) Determine the regulators of Ral activation (RhoGDI2, etc..) and their effect on intracellular Ral localization and bladder cancer nfigration and invasion; 3) Determine the protein complexes associated with Ral in vitro and in vivo, including those found in human cancer. Conclusion: Completion of these specific aims will provide biologically relevant molecular information on the signaling pathways regulating bladder cancer invasion in vivo and lead to the rational development of diagnostic and prognostic tools predicting the development of invasive disease and therapies to interfere with this process in patients with superficial bladder cancer.

背景和意义：40% 的“浅表”（非肌肉浸润性）膀胱癌患者会发展为在随访期间出现“侵袭性”危及生命的疾病。在临床研究中，表皮生长因子的过度表达受体 (EGFR)、Ha-Ras 突变和抑癌基因 PTEN 缺失与这种表型肿瘤相关过渡。然而，这些基因有效触发或促进侵袭过程的确切分子途径是不完全理解。我们最初的 R29 假设 EGFR 信号传导在体外增强膀胱肿瘤的运动性，并且侵入体内并旨在确定EGFR在此过程中使用的信号通路。自 9/97 为 R29 提供资金以来，我们做出了以下重要的观察，这些观察支持了最初的假设并解决了最初的目标应用： 1）抑制EGFR和Ras，降低侵袭性膀胱癌细胞的运动能力； 2) EGF 刺激运动通过 PI3K 非侵入性细胞，这需要 Rho 和 Ras 效应大鼠的活性； 3) 在非侵袭性细胞中，基线 RalA 活性为低，而侵袭细胞具有较高的活化； 4)侵袭细胞RhoGDI2表达水平较低。该基因的重建导致 RalA 的运动性和活性减弱，但 RhoA 则不然，表明该基因可能是第一个 RalGDI 被鉴定为入侵抑制因子； 5) 通过侵袭细胞中 PTEN 重建抑制 PI3K 活性具有失活的 PTEN，导致体内原位侵袭的抑制和 RhoA 活性的降低。由于整体 Ral 和 RhoGDI2 的生物学特性尚不清楚，但可能对于调节膀胱患者的肿瘤侵袭至关重要癌症，我们提出指导假设：EGF 通过 Ral 激活介导膀胱肿瘤侵袭。我们将测试这个假设采用从基础生物化学到临床肿瘤学的一系列技术来解决人类 Ral 生物学问题膀胱癌。其中包括：1）具有不同侵袭能力的独特配对人类膀胱癌细胞系； 2）一本小说器官型膀胱模型允许体外研究肿瘤侵袭； 3) 评估候选物效果的原位测定体内膀胱癌侵袭的分子； 4) 具有适当遗传和表型特征的转基因和基因敲除小鼠； 5）具有病理学和临床特征的冷冻标本的人体组织库。具体目标：1）确定角色以及器官型、小鼠原位和人类肿瘤研究中 Ral 在膀胱癌侵袭中的病理学； 2）确定 Ral 激活调节因子（RhoGDI2 等）及其对细胞内 Ral 定位和膀胱癌浸润的影响入侵； 3) 在体外和体内测定与 Ral 相关的蛋白质复合物，包括在人类癌症中发现的蛋白质复合物。结论：完成这些特定目标将提供信号通路的生物学相关分子信息调节膀胱癌体内侵袭并导致合理开发预测膀胱癌的诊断和预后工具开发侵袭性疾病和疗法来干扰浅表性膀胱癌患者的这一过程。