Regulation of epithelial-mesenchymal transition and stem cell activity by PTEN in breast cancer

PTEN 对乳腺癌上皮间质转化和干细胞活性的调节

• 批准号: 9250104
• 负责人: Shaohua Li
• 金额: $ 16.45万
• 依托单位: RBHS-ROBERT WOOD JOHNSON MEDICAL SCHOOL
• 依托单位国家: 美国
• 财政年份: 2016
• 资助国家: 美国
• 起止时间: 2016-04-01 至 2019-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9250104
• 关键词: Actins; Address; Alleles; Animals; Area; BRCA1 gene; Binding Sites; Breast Cancer Cell; Breast Epithelial Cells; Breast cancer metastasis; Cancer Prognosis; Cellular biology; Chromatin Remodeling Factor; Clinical Trials; Complex; Epithelial; Event; FRAP1 gene; Fatty acid glycerol esters; Foundations; Gene Expression Regulation; Generations; Genes; Goals; Gray unit of radiation dose; Growth; Human; Immunoprecipitation; Knockout Mice; Light; Link; Lipids; Malignant Neoplasms; Mammary Neoplasms; Mammary gland; Maps; Mass Spectrum Analysis; Mediating; Mesenchymal; Modeling; Molecular; Morphogenesis; Mus; Mutation; Neoplasm Metastasis; PTEN gene; Pathway interactions; Penetrance; Phenotype; Phosphatidylinositol 4,5-Diphosphate; Phosphatidylinositols; Phosphoric Monoester Hydrolases; Phosphotransferases; Phosphotyrosine; Predictive Value; Process; Protein Dephosphorylation; Protein p53; Protein phosphatase; Proteins; Regulation; Relapse; Reporting; Resistance; Role; SWI/SNF Family Complex; Signal Pathway; Signal Transduction; Site-Directed Mutagenesis; Specificity; Stem cells; Susceptibility Gene; TP53 gene; Testing; Therapeutic; Time; Transitional Cell; Tumor Suppression; Tumor Suppressor Proteins; Tumorigenicity; Xenograft procedure; base; cancer recurrence; cancer risk; cancer stem cell; cell growth; chromatin remodeling; clinically relevant; conventional therapy; drug discovery; early onset; epigenetic regulation; genetic analysis; inhibitor/antagonist; insight; knock-down; loss of function; mTOR Inhibitor; malignant breast neoplasm; mammary gland development; migration; mutant; new therapeutic target; novel; overexpression; promoter; public health relevance; reconstitution; transcription factor; tumor; tumor progression; tumorigenesis

 DESCRIPTION (provided by applicant): Epithelial-mesenchymal transition (EMT) has been implicated in promoting breast cancer invasion and metastasis. Recent studies have shown that this important process is also responsible for the generation of breast cancer stem cells (CSCs), which are resistant to conventional therapy and contribute to tumor metastasis and relapse. Independent of these findings, mutations in BRCA1/2, TP53 and PTEN have emerged as high- penetrance susceptibility genes and are clinically relevant for the determination of breast cancer risk and prognosis. Among these genes, PTEN inactivation promotes both EMT and CSC enrichment. However, the underlying molecular mechanisms behind these activities are largely unknown. In our genetic analysis of PTEN functions in epithelial morphogenesis, we have identified Abi1, a key component of the WAVE regulatory complex (WRC), as a new substrate for PTEN. Protein interaction studies have further demonstrated a novel interaction between Abi1 and Smarcc1 of the SWI/SNF chromatin remodeling complex, which links PTEN to SWI/SNF-mediated epigenetic regulation of cancer. PTEN dephosphorylates Abi1 and consequently downregulates Abi1 and Smarcc1. Overexpression of Abi1 or Smarcc1 in human mammary epithelial cells promotes EMT and enhances stem cell activity. Based on these findings, we hypothesize that PTEN loss induces EMT and CSC enrichment through Abi1 and Smarcc1 in breast cancer. To test this hypothesis, we propose two specific aims. In Aim 1, the role of Abi1 in PTEN loss-induced EMT, CSC enrichment, tumorigenicity and metastasis will be analyzed by gain- and loss-of-function approaches in breast cancer cells and a mouse xenograft mammary fat pad model. In addition, we will delete one or both Abi1 alleles in PTEN- null mouse mammary glands to determine if Abi1 elevation mediates PTEN loss-induced breast tumor formation. To elucidate the signaling events downstream of PTEN-Abi1, we will focus on the interaction of Abi1 with WRC components and Abl kinase and their role in the EMT and CSC enrichment. In Aim 2, we will map the binding sites that mediate the Abi1-Smarcc1 interaction by site-directed mutagenesis. We will then analyze the functional consequences of the Abi1-Smarcc1 interaction in SWI/SNF complex assembly and chromatin remodeling activity at the promoters of the EMT-inducing transcription factors. The impact on EMT and CSC activity will be analyzed both in breast cancer cells and mouse xenografts. Furthermore, we will test the hypothesis that elevated Smarcc1 competes with Smarcc2 in the SWI/SNF complex and thus induces EMT and CSC activity by overexpressing Smarcc2 and knocking down Smarcc1 in PTEN-deficient breast cancer cells. Successful completion of the proposed studies will lay the foundation for developing therapeutic strategies targeting EMT and CSCs in PTEN-deficient breast cancer. It will also shed light on the new PTEN- Abi1-Smarcc1 pathway.

 描述(由申请人提供)：上皮-间充质转化(EMT)与促进乳腺癌的侵袭和转移有关。最近的研究表明，这一重要过程也是乳腺癌干细胞(CSCs)产生的原因，CSCs对传统疗法具有抵抗力，并有助于肿瘤的转移和复发。与这些发现无关的是，BRCA1/2、TP53和PTEN突变已成为高外显性易感基因，并在临床上与乳腺癌风险和预后的确定相关。在这些基因中，PTEN的失活同时促进了EMT和CSC的丰富。然而，这些活动背后的潜在分子机制在很大程度上是未知的。在我们对PTEN在上皮形态发生中作用的遗传分析中，我们已经确定Abi1是波形调节复合体(WRC)的关键成分，是PTEN的新底物。蛋白质相互作用研究进一步证明了SWI/SNF染色质重塑复合体中Abi1和Smarcc1之间的一种新的相互作用，它将PTEN与SWI/SNF介导的癌症表观遗传调控联系起来。PTEN使ABI1去磷酸化，从而下调ABI1和Smarcc1的表达。在人乳腺上皮细胞中过表达Abi1或Smarcc1可促进EMT并增强干细胞活性。基于这些发现，我们假设PTEN缺失通过Abi1和Smarcc1在乳腺癌中诱导EMT和CSC的丰富。为了检验这一假设，我们提出了两个具体目标。在目标1中，将通过在乳腺癌细胞和小鼠异种移植乳腺脂肪垫模型中获得和丧失功能的方法来分析ABI1在PTEN缺失诱导的EMT、CSC富集性、致瘤性和转移中的作用。此外，我们将在PTEN缺失的小鼠乳腺中删除一个或两个Abi1等位基因，以确定Abi1的升高是否介导了PTEN缺失诱导的乳腺肿瘤的形成。为了阐明PTEN-Abi1下游的信号事件，我们将重点研究Abi1与WRC组分和Abl激酶的相互作用以及它们在EMT和CSC浓缩中的作用。在目标2中，我们将通过定点突变来定位介导ABI1-Smarcc1相互作用的结合位点。然后，我们将分析ABI1-Smarcc1相互作用在SWI/SNF复合体组装中的功能后果以及EMT诱导转录因子启动子的染色质重塑活性。对EMT和CSC活性的影响将在乳腺癌细胞和小鼠异种移植中进行分析。此外，我们将检验这一假说，即上调的Smarcc1与Smarcc2在SWI/SNF复合体中竞争，从而通过在PTEN缺陷的乳腺癌细胞中过表达Smarcc2并击倒Smarcc1来诱导EMT和CSC活性。建议研究的成功完成将为开发针对PTEN缺陷乳腺癌的EMT和CSCs的治疗策略奠定基础。这也将揭示新的PTEN-Abi1-Smarcc1途径。