Defining the Molecular Mechanisms of KLF17 Regulation and Function in EMT

定义 KLF17 在 EMT 中调节和功能的分子机制

• 批准号: 8215813
• 负责人: QIHONG HUANG
• 金额: $ 33.96万
• 依托单位: WISTAR INSTITUTE
• 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 2015-01-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/8215813
• 关键词: 1p34; 3' Untranslated Regions; Affect; Animal Model; Breast Cancer Cell; Breast Cancer Treatment; Cancer cell line; Cells; Cellular Assay; Cessation of life; Chromosomes; Clinical; Development; Disease; Distant; Embryonic Development; Epithelial; Epithelial Cells; Event; Exhibits; Gametogenesis; Gene Expression; Genes; Genetic; Genetic Epistasis; Genetic Screening; Goals; Human; In Vitro; Laboratories; Loss of Heterozygosity; Maintenance; Malignant Neoplasms; Mammary gland; Mediating; Mediator of activation protein; Mesenchymal; MicroRNAs; Molecular; Molecular Profiling; Mus; Neoplasm Metastasis; Outcome; Pathway interactions; Play; Primary Carcinoma; Primary Neoplasm; Prognostic Marker; RNA Interference; Regulation; Relapse; Role; Sampling; Signal Pathway; Site; Snails; Testing; Therapeutic; Transplantation; Tumor Angiogenesis; Tumor Cell Invasion; Tumor Suppressor Proteins; Xenograft Model; Xenograft procedure; base; cell growth; genome-wide; in vitro Assay; in vivo; insight; lymph nodes; malignant breast neoplasm; mouse model; mutant; neoplastic cell; outcome forecast; prognostic; programs; promoter; public health relevance; research study; senescence; small hairpin RNA; therapeutic target; transcription factor; tumor; tumor growth; tumor xenograft; tumorigenesis; vector

DESCRIPTION (provided by applicant): We recently identified Kr|pple-like factor 7 (KLF17) as a negative regulator of epithelial-mesenchymal transition (EMT) using a genome-wide RNAi screen in mice. The human KLF17 gene is located at chromosome 1p34 where allelic loss of heterozygosity (LOH) correlates with poor prognosis in breast cancer. The overall goal of this proposal is to determine the molecular mechanisms and genetic regulation of transcription factor KLF17 in tumor maintenance and EMT, and determine the therapeutic value of KLF17 modulation and prognostic value of KLF17 in breast cancer. We will use molecular profiling, cellular assays and animal models to elucidate the mechanisms of KLF17 functions and regulation. We will proceed from the following aims: Aim 1. Determine the molecular mechanisms of KLF17 regulation and function in tumor growth and EMT. We will determine the effect of Id2 and ARHGAP29, regulated by KLF17, individually and the combined effect of these two genes on tumor cell growth and EMT both in vitro and in mouse models. We will use in vitro assays and in vivo orthotopic transplantation mouse models to test their functions in tumor invasion and metastasis. Epistasis analysis will be performed to determine whether these genes are major mediators of KLF17 in tumor growth and EMT. We will also investigate the molecular mechanisms of KLF17 regulation by SNAI1 (Snail), a known EMT regulator, transcriptionally and by microRNA miR-129 post-transcriptionally. We will determine whether KLF17 is a direct target of SNAI1 and whether miR-129 directly targets KLF17 at its 3'UTR. The functions of SNAI1 and miR-129 in EMT and tumor invasion will be determined by in vitro assays and tumor xenograft mouse models. Epistasis analysis will be performed to determine whether KLF17 is a major mediator of SNAI1 and miR-129 in EMT and tumor invasion. Aim 2. Characterize the function of KLF17 in tumor maintenance. We will use xenograft models and cells expressing KLF17 short hairpin RNA (shRNA) and Ras in an inducible vector to test whether modulation of KLF17 expression alone or in combination with Ras inhibition can cause tumor regression in breast cancer. We will investigate whether KLF17 knockdown antagonizes Ras-induced senescence in tumors using xenograft animal models and determine the pathways that KLF17 interferes with in Ras-induced senescence. These experiments will determine whether KLF17 alone or in combination with Ras can serve as a potential therapeutic target in the treatment of breast cancer. Aim 3. Determine the prognostic value of KLF17, Id1, SNAI1 and miR-129 in clinical breast cancer samples. We will determine the expression of KLF17 and Id1, along with Snail and miR-129 in clinical primary breast cancer samples with known clinical outcomes to assess whether they can be used as prognostic markers for relapse in breast cancer. PUBLIC HEALTH RELEVANCE: This proposal studies transcription factor KLF17 in tumor maintenance, epithelial- mesenchymal transition (EMT) and metastasis. Tumor spread from a primary site to distant sites is responsible for most cancer death. EMT plays a critical role in tumor spread. Currently there are few prognostic markers and therapeutics for metastatic disease. KLF17 was recently discovered in my laboratory to be involved in EMT by a genome-wide RNAi screen. Elucidation of the molecular mechanisms of KLF17 in EMT and metastasis will provide potential prognostic marker and therapeutics for the treatment of this deadly disease.

描述（由申请人提供）：我们最近发现Kr|使用小鼠全基因组RNAi筛选，发现pple样因子7（KLF 17）是上皮-间充质转化（EMT）的负调节因子。人KLF 17基因位于染色体1 p34，其中等位基因杂合性丢失（洛）与乳腺癌的不良预后相关。本研究的总体目标是明确转录因子KLF 17在肿瘤维持和EMT中的分子机制和遗传调控，并确定KLF 17调节乳腺癌的治疗价值和预后价值。我们将使用分子分析，细胞分析和动物模型来阐明KLF 17的功能和调节机制。我们将从以下目标出发：目标1。确定KLF 17在肿瘤生长和EMT中的调节和功能的分子机制。我们将在体外和小鼠模型中分别确定由KLF 17调节的Id 2和ARHGAP 29的作用，以及这两个基因对肿瘤细胞生长和EMT的联合作用。我们将使用体外实验和体内原位移植小鼠模型来测试它们在肿瘤侵袭和转移中的功能。将进行上位性分析，以确定这些基因是否是KLF 17在肿瘤生长和EMT中的主要介质。我们还将研究SNAI 1（Snail）（一种已知的EMT调节因子）在转录和转录后通过microRNA miR-129调节KLF 17的分子机制。我们将确定KLF 17是否是SNAI 1的直接靶点，以及miR-129是否直接靶向KLF 17的3 'UTR。SNAI 1和miR-129在EMT和肿瘤侵袭中的功能将通过体外测定和肿瘤异种移植小鼠模型来确定。将进行上位性分析，以确定KLF 17是否是EMT和肿瘤侵袭中SNAI 1和miR-129的主要介导因子。目标2.描述KLF 17在肿瘤维持中的功能。我们将使用异种移植模型和细胞表达KLF 17短发夹RNA（shRNA）和Ras在诱导型载体，以测试是否KLF 17表达单独或与Ras抑制相结合的调制可以导致乳腺癌肿瘤消退。我们将使用异种移植动物模型研究KLF 17敲低是否拮抗Ras诱导的肿瘤衰老，并确定KLF 17干扰Ras诱导的衰老的途径。这些实验将确定KLF 17单独或与Ras组合是否可以作为治疗乳腺癌的潜在治疗靶点。目标3.确定KLF 17、Id 1、SNAI 1和miR-129在临床乳腺癌样本中的预后价值。我们将在具有已知临床结果的临床原发性乳腺癌样本中确定KLF 17和Id 1的表达，沿着Snail和miR-129，以评估它们是否可用作乳腺癌复发的预后标志物。 公共卫生相关性：本研究旨在探讨转录因子KLF 17在肿瘤维持、上皮-间质转化（EMT）和转移中的作用.肿瘤从原发部位向远处扩散是大多数癌症死亡的原因。EMT在肿瘤扩散中起关键作用。目前，转移性疾病的预后标志物和治疗方法很少。最近，我的实验室通过全基因组RNAi筛选发现KLF 17参与EMT。阐明KLF 17在EMT和转移中的分子机制将为EMT的治疗提供潜在的预后指标和治疗方法。