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Klf4 in tumor initiation and maintenance of squamous cell carcinoma

Klf4在鳞状细胞癌肿瘤发生和维持中的作用

基本信息

批准号：
7795119
负责人：
John Michael Ruppert
金额：
$ 27.84万
依托单位：
WEST VIRGINIA UNIVERSITY
依托单位国家：
美国
项目类别：
财政年份：
2007
资助国家：
美国
起止时间：
2007-03-01 至 2011-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/7795119
关键词：
Acetylation Acetylglucosamine Affect Alleles Animal Model Anthracenes Antibodies Binding Binding Sites Breast Carcinogens Cell Cycle Cell Nucleus Cells Chimeric Proteins Chromatin Codon Nucleotides Complementary DNA Cultured Cells Cyclin D1 Cytokeratin DNA Binding Development Dominant-Negative Mutation Dysplasia EP300 gene ERBB2 gene Elements Enzymes Epithelial Epithelial Cells Event Frequencies GKLF protein Genetic Transcription Genetically Engineered Mouse Genetically Modified Animals Goals Growth HRAS gene Half-Life Histologic Human Hyperplasia In Situ In Vitro Incidence Indium Keratin Ligands Link MCF10A cells Maintenance Malignant Neoplasms Mammary gland Maps Mediating Messenger RNA Modeling Modification Molecular Mus Mutagenesis Mutation Neoplasms Nuclear Oncogenes Open Reading Frames Oral Pathogenesis Pathway interactions Physiologic pulse Post-Translational Protein Processing Prevention Proteins RNA RNA analysis Regulation Research Personnel Role Run-On Assays Sequence Analysis Skin Skin Neoplasms Small Interfering RNA Squamous cell carcinoma Staining method Stains Structure TP53 gene Testing Tetanus Helper Peptide Tetracyclines Tissues Transactivation Transcript Transgenes Transgenic Mice Tumor Cell Line Untranslated Regions Up-Regulation Wild Type Mouse cell fate specification cell growth cell type chromatin remodeling gain of function in vivo loss of function malignant breast neoplasm mouse model mutant neoplastic cell notch protein novel programs promoter protein function protein protein interaction skin squamous cell carcinoma transcription factor tumor tumor growth tumor initiation tumor progression tumorigenesis

项目摘要

The KLF4 transcription factor is important in cell fate and differentiation of epithelial cells. KLF4 rapidly initiates cutaneous squamous cell carcinoma (SCC) in mice, and is deregulated at an early step in human skin tumor progression. Using a conditional KLF4-ER fusion protein, we identified the cell fate determinant Notch 1 as a transcriptional target of KLF4. In cultured cells, in transgenic mice and in human tumors, KLF4 expression is associated with expression of Notchl, Notchl ligands, and Notchl effectors such as Cyclin D1. Transformation of RK3E epithelial cells requires Notchl. These studies identify a KLF4-Notch1 pathway that appears important in cell fate specification of normal epithelial cells and tumor cells. Mutagenesis of KLF4 identified dominant negative (DN) alleles that block transformation in vitro by KLF4. KLF4-DNs identify a role of KLF4 in growth of specific human cell types in vitro, and in transformation by specific transforming oncogenes such as ErbB2 and c-MYC. To study the role of Klf4 in cutaneous SCC we developed a novel highly penetrant model of SCC by administering carcinogens to genetically-engineered mice. Tumors were histologically similar to human SCC, and expressed SCC markers such as p53, keratin 14, and Notch! SCCs that developed in this model included KLF4-positive tumors and some tumors that were KLF4-negative. To understand the mechanisms that normally regulate KLF4, we will study the role of posttranslational modifications in the DN activity of mutant KLF4 proteins (Aim 1). The role of endogenous Klf4 in tumor initiation and maintenance will be studied in the new SCC mouse model using conditional, Cre-mediated deletion of Klf4/LoxP alleles, and by tetracycline-inducible expression of siRNA or KLF4-DN in transgenic mice (Aim 2). Using similar loss- and gain-of-function strategies, the role of Klf4 in mammary epithelial development and in transformation by ErbB2 will be determined (Aim 3). In Aim 4, a posttranslational control that appears responsible for upregulation of Klf4 transcripts in mouse and human SCC will be analyzed by identifying RNA destabilizing elements in the KLF4 cDNA. These studies may provide support of a role for KLF4 and Notchl in pathogenesis of common human cancers such as SCC and breast cancer. Thus, completion of the studies would identify this pathway as a target for prevention and treatment, and provide new animal models in which to assess such therapies.

KLF 4转录因子在上皮细胞的细胞命运和分化中是重要的。KLF 4快速在小鼠中引发皮肤鳞状细胞癌（SCC），在人类中在早期阶段失调皮肤肿瘤进展。使用条件性KLF 4-ER融合蛋白，我们鉴定了细胞命运决定因子 Notch 1作为KLF 4的转录靶点。在培养细胞、转基因小鼠和人类肿瘤中，KLF 4 表达与刻缺蛋白1、刻缺蛋白1配体和刻缺蛋白1效应物如细胞周期蛋白D1的表达相关。 RK 3E上皮细胞的转化需要Notchl。这些研究确定了KLF 4-Notch 1通路，在正常上皮细胞和肿瘤细胞的细胞命运特化中显得重要。 KLF 4的诱变鉴定了在体外阻断KLF 4转化的显性阴性（DN）等位基因。 KLF 4-DNs鉴定了KLF 4在体外特定人类细胞类型的生长中的作用，以及通过特异性转化癌基因，如ErbB 2和c-MYC。为了研究Klf 4在皮肤SCC中的作用，我们通过以下方法建立了一种新的高度渗透的SCC模型：给基因工程小鼠注射致癌物质。肿瘤在组织学上与人类相似 SCC，并表达SCC标志物，如p53，角蛋白14，和Notch！在这个模型中产生的SCC 包括KLF 4阳性肿瘤和一些KLF 4阴性肿瘤。为了了解正常情况下调节KLF 4的机制，我们将研究翻译后蛋白的作用。突变KLF 4蛋白的DN活性的修饰（Aim 1）。内源性Klf 4在肿瘤中的作用启动和维持将在新的SCC小鼠模型中使用条件性，Cre介导的 Klf 4/LoxP等位基因的缺失，以及通过在转基因小鼠中四环素诱导的siRNA或KLF 4-DN表达，小鼠（Aim 2）。使用类似的功能丧失和获得策略，Klf 4在乳腺上皮细胞中的作用将确定ErbB 2的发育和转化（目标3）。在目标4中，翻译后控制在小鼠和人SCC中似乎负责Klf 4转录物上调的基因将通过以下方法进行分析：鉴定KLF 4 cDNA中的RNA去稳定化元件。这些研究为KLF 4和Notchl在人类白血病发病机制中的作用提供了支持。癌症如SCC和乳腺癌。因此，研究的完成将确定这一途径为用于预防和治疗靶点，并提供新的动物模型来评估这种疗法。