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: 8039275
• 负责人: QIHONG HUANG
• 金额: $ 34.19万
• 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/8039275
• 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)：我们最近在小鼠全基因组RNAi筛选中发现Kr|pple-like factor7(KLF17)是上皮-间充质转化(EMT)的负调控因子。人类KLF17基因位于染色体1p34，等位基因杂合性缺失(LOH)与乳腺癌预后不良相关。本研究的总体目标是确定转录因子KLF17在肿瘤维持和EMT中的分子机制和基因调控，并确定KLF17在乳腺癌中的治疗价值和预后价值。我们将使用分子图谱、细胞分析和动物模型来阐明KLF17的功能和调控机制。我们将从以下目标出发：目的1.确定KLF17在肿瘤生长和EMT中的调控和作用的分子机制。我们将确定由KLF17单独调控的Id2和ARHGAP29的作用，以及这两个基因在体外和小鼠模型中对肿瘤细胞生长和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。上位性分析将被用来确定这些基因是否是KLF17在肿瘤生长和EMT中的主要中介。我们还将研究已知的EMT调节因子SNAI1(Snail)在转录和转录后通过microRNA miR-129调控KLF17的分子机制。我们将确定KLF17是否是SNAI1的直接靶点，以及miR-129是否直接针对KLF17的3‘非编码区。SNAI1和miR-129在EMT和肿瘤侵袭中的作用将通过体外实验和小鼠移植瘤模型来确定。上位性分析将确定KLF17是否是SNAI1和miR-129在EMT和肿瘤侵袭中的主要介导者。目的2.研究KLF17在肿瘤维持中的作用。我们将使用异种移植模型和在诱导性载体中表达KLF17短发夹状RNA(ShRNA)和RAS的细胞来测试KLF17表达的调节是否单独或联合RAS抑制可以导致乳腺癌的肿瘤消退。我们将利用异种移植动物模型研究KLF17基因敲除是否拮抗RAS诱导的肿瘤衰老，并确定KLF17在RAS诱导的衰老中的干预途径。这些实验将确定KLF17是否可以单独或与RAS联合作为乳腺癌治疗的潜在治疗靶点。目的：探讨KLF17、Id1、SNAI1和miR-129在乳腺癌临床标本中的预后价值。我们将检测KLF17和Id1以及Snail和miR-129在临床上已知临床结果的乳腺癌标本中的表达，以评估它们是否可以作为乳腺癌复发的预后标志。 公共卫生相关性：这项建议研究转录因子KLF17在肿瘤维持、上皮-间充质转化(EMT)和转移中的作用。肿瘤从原发部位扩散到远处是大多数癌症死亡的原因。EMT在肿瘤扩散中起着关键作用。目前，对转移性疾病的预后标记物和治疗方法很少。最近，我的实验室通过全基因组RNAi筛查发现KLF17参与了EMT。阐明KLF17在EMT和转移中的分子机制将为这一致命疾病的治疗提供潜在的预后标志物和治疗方法。