Roles for Rho Proteins During Specific Stages of Breast Cancer Metastasis.

Rho 蛋白在乳腺癌转移特定阶段的作用。

• 批准号: 8242866
• 负责人: John Lamar
• 金额: $ 5.39万
• 依托单位: MASSACHUSETTS INSTITUTE OF TECHNOLOGY
• 依托单位国家: 美国
• 财政年份: 2010
• 资助国家: 美国
• 起止时间: 2010-04-01 至 2013-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8242866
• 关键词: American; Biological Assay; Biological Models; Blood; Breast Carcinoma; Cell Count; Cell Line; Cells; Citofur; Data; Detection; Development; Disease; Extravasation; Fatty acid glycerol esters; Fellowship; Goals; Health; Implant; Individual; Liver; Lung; Malignant Epithelial Cell; Mammary Neoplasms; Mammary gland; Measures; Mediating; Metastatic Lesion; Molecular; Mouse Mammary Tumor Virus; Mus; National Research Service Awards; Neoplasm Metastasis; Organ; Pathway interactions; Polyomavirus; Process; Proteins; Research Design; Role; Screening procedure; Series; Spinal Cord; Staging; Testing; Woman; base; cellular transduction; combat; implantation; in vitro Assay; killings; lymph nodes; malignant breast neoplasm; metastatic process; retroviral transduction; rho; small hairpin RNA; therapy design; tumor; tumor growth

DESCRIPTION (provided by applicant): Although RhoA and RhoC have been implicated in invasion and metastasis, the molecular mechanisms behind Rho-mediated metastasis are unclear, and it is unknown which metastatic processes involve Rho proteins. Our long-term goals for this project are to understand the importance of these two proteins during key stages of the metastatic cascade, and to identify which Rho effector pathways are involved. We hypothesize that both RhoA and RhoC will promote the spread of breast cancer by enhancing multiple stages of metastasis. We further predict that Rho-mediated metastasis will require specific Rho effectors, and that different Rho effectors may mediate each Rho-mediated process during metastasis. Specific Aims: (1) Determine if breast cancer metastasis requires RhoA or RhoC, and identify which stages of the metastatic cascade are dependent upon these proteins. (2) Identify which stages of breast cancer metastasis are promoted by increased expression of RhoA or RhoC. (3) Identify Rho effectors that are required for Rho-mediated metastasis. (4) Test the involvement of RhoA, RhoC, and Rho effectors during mammary carcinoma development, progression, and metastasis. Study design: We will use retroviral expression constructs and miR30-based shRNAs to alter the expression of RhoA or RhoC in a series of mammary carcinoma (MC) cell lines with varying metastatic potentials. Cells will then be implanted into the mammary fat pads of syngeneic mice to assay differences in several metastatic processes (invasion, intravasation, extravasation and metastatic tumor formation). We will also screen for Rho effectors involved in metastasis by orthotopically implanting pools of MC cells expressing barcoded miR30-based shRNAs that each target a known Rho effector, and then screening for enrichment or dramatic reduction of each barcode in metastatic lesions that form in the lungs. As an additional model system, we will also isolate primary MC cells from tumor-bearing MMTV Polyoma middle T mice, manipulate Rho protein and effector expression, and then orthotopically implant the cells into donor mice and assay the effects on metastatic processes. PUBLIC HEALTH RELEVANCE: Breast cancer metastasis kills 40,000 American women each year, and until we fully understand how specific proteins influence the metastatic processes, we will be unable to design therapies that combat this disease. Our studies aim to identify which metastatic processes involve Rho proteins, as well as determine which molecular pathways downstream of Rho proteins are required for each of these processes.

描述(申请人提供)：尽管RhoA和RhoC与侵袭和转移有关，但Rho介导的转移背后的分子机制尚不清楚，也不清楚哪些转移过程涉及Rho蛋白。我们这个项目的长期目标是了解这两个蛋白在转移级联的关键阶段的重要性，并确定哪些Rho效应通路参与其中。我们假设RhoA和RhoC都会通过促进多个阶段的转移来促进乳腺癌的扩散。我们进一步预测，Rho介导的转移将需要特定的Rho效应器，并且不同的Rho效应器可能在转移过程中介导每个Rho介导的过程。具体目标：(1)确定乳腺癌转移是否需要RhoA或RhoC，并确定转移级联反应的哪些阶段依赖于这些蛋白。(2)明确RhoA或RhoC表达增加促进乳腺癌转移的阶段。(3)确定Rho介导的肿瘤转移所需的Rho效应子。(4)检测RhoA、RhoC和Rho效应分子在乳腺癌发生、发展和转移过程中的作用。研究设计：我们将使用逆转录病毒表达载体和基于miR30的shRNAs来改变RhoA或RhoC在一系列具有不同转移潜能的乳腺癌(MC)细胞系中的表达。然后将细胞移植到同基因小鼠的乳房脂肪垫中，以分析几种转移过程(侵袭、渗入、渗出和转移肿瘤形成)的差异。我们还将通过以下方式筛选参与转移的Rho效应器：将表达条形码miR30的shRNA的MC细胞池原位移植，每个条形码都针对一个已知的Rho效应器，然后在肺部形成的转移灶中筛选每个条形码的丰富或显著减少。作为一个额外的模型系统，我们还将从荷瘤的MMTV多瘤中间T鼠中分离出原代MC细胞，操纵Rho蛋白和效应器的表达，然后将这些细胞原位移植到供体小鼠体内，并检测其对转移过程的影响。公共卫生相关性：每年有40,000名美国女性死于乳腺癌转移，在我们完全了解特定蛋白质如何影响转移过程之前，我们将无法设计出对抗这种疾病的疗法。我们的研究旨在确定哪些转移过程涉及Rho蛋白，以及确定这些过程中每个过程都需要Rho蛋白下游的哪些分子通路。