The oncogenic role of EPIC1MYC axis in breast cancer

EPIC1MYC轴在乳腺癌中的致癌作用

基本信息

批准号：
10214561
负责人：
Da Yang
金额：
$ 35.8万
依托单位：
UNIVERSITY OF PITTSBURGH AT PITTSBURGH
依托单位国家：
美国
项目类别：
财政年份：
2018
资助国家：
美国
起止时间：
2018-08-02 至 2023-07-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10214561
关键词：
Attenuated BT 474 Binding Binding Sites Bioinformatics Biological Assay Breast Cancer Cell Breast Cancer Patient Breast Cancer therapy CCNA2 gene CDC45L gene Cancer Cell Growth Cancer cell line Cell Culture Techniques Cell Cycle Arrest Cell model Cells Characteristics Clinical Code DNA Methylation Data Dependence Development Disease Epigenetic Process Event Future Gene Expression Gene Expression Regulation Genes Genetic Transcription Goals High-Throughput Nucleotide Sequencing Immunoprecipitation In Vitro Light Luciferases MCF7 cell MYC Family Protein MYC gene Mediating Messenger RNA Meta-Analysis Molecular Mus Nucleotides Oncogenic Outcome Patients Pattern Play Promoter Regions Proteins Proto-Oncogene Proteins c-myc RNA RNA Binding Reporter Reporter Genes Research Resistance Role Sampling Sequence Deletion Series Small Interfering RNA Testing The Cancer Genome Atlas Transcript Untranslated RNA Xenograft Model Xenograft procedure base breast cancer survival c-myc Genes cancer subtypes chromatin immunoprecipitation cohort crosslink crosslinking and immunoprecipitation sequencing deletion analysis effective therapy in vivo infancy knock-down locked nucleic acid malignant breast neoplasm mouse model mutant novel overexpression prognostic promoter targeted cancer therapy transcriptome sequencing tumor tumor growth tumorigenesis

项目摘要

A better understanding of the molecular events that contribute to the aggressiveness and poor outcome of breast cancer is essential to develop more effective therapies for this deadly disease. Emerging evidence suggests that lncRNAs play important roles in breast cancer tumorigenesis. However, research of lncRNAs in breast cancer is still in its infancy. Through an integrated bioinformatic analysis of the lncRNA epigenetic landscape in 6475 tumor samples and 781 cancer cell lines, we discovered a novel intergenic lncRNA, EPIC1 (EPigenetically Induced lnCRNA1, or ENSG00000224271). EPIC1 is significantly overexpressed in the luminal B breast cancer subtype due to the loss of DNA methylation at the EPIC1 gene promoter. We went on to show that EPIC1 promotes luminal B breast cancer tumorigenesis in cell cultures and in mouse xenograft models by directly interacting with the oncogenic transcription factor MYC. In this project, we hypothesize that EPIC1 is an oncogenic lncRNA in breast cancer, particularly in the luminal B subtype. Mechanistically, EPIC1 enhances the transcriptional activity of MYC by directly interacting with the MYC protein. We propose three specific aims to test this hypothesis. In Aim 1, we will characterize the EPIC1-MYC interaction using HITS-CLIP assay and investigate if the identified MYC binding sequences in EPIC1 are necessary and sufficient for EPIC1-MYC interaction. We will also determine the functional domain of MYC protein responsible for the EPIC1 binding. In Aim 2, we will determine the oncogenic role of the EPIC1-MYC axis in breast cancer by overexpressing wt-EPIC1 or MYC binding sequence deletion mutant EPIC1 (ΔMYC-EPIC1) in ZR-75-1 and BT-474 cells. We will then determine the dependency of EPIC1’s oncogenic role on MYC protein in xenograft mouse models. Finally, we will investigate EPIC1’s association with breast cancer clinical characteristics and outcomes in eight independent patient cohorts including 1882 breast cancer tumors. In Aim 3, we will first determine the pattern of MYC binding to its target gene promoters in wt- or ΔMYC-EPIC1 overexpressing ZR-75-1 and BT-474 cells by performing RNA-seq and MYC ChIP-seq. We will then determine whether EPIC1 regulates the MYC targets through EPIC1-MYC binding sites using ChIP-PCR and luciferase reporter analyses. Finally, we will construct the EPIC1-cMYC regulatory network in breast cancer by integrating ChIP-seq, RNA-seq, and TCGA breast cancer data. Our study will uncover the role of a novel oncogenic lncRNA, EPIC1, in breast cancer through its functional crosstalk with the well-established oncogene MYC.

更好地了解导致乳房侵袭性和不良结果的分子事件新的证据表明，癌症对于开发针对这种致命疾病的更有效疗法至关重要。 lncRNAs在乳腺癌肿瘤发生中发挥重要作用，然而，lncRNAs在乳腺癌中的研究尚不深入。仍处于起步阶段，通过对 6475 肿瘤的 lncRNA 表观遗传景观进行综合生物信息学分析。在样本和 781 个癌细胞系中，我们发现了一种新的基因间 lncRNA，EPIC1（EPigenically Induced lnCRNA1 或 ENSG00000224271) 在管腔 B 乳腺癌亚型中显着过度表达。由于 EPIC1 基因启动子处 DNA 甲基化的丢失，我们继续证明 EPIC1 促进。细胞培养物和小鼠异种移植模型中的管腔 B 乳腺癌肿瘤发生通过直接与致癌转录因子 MYC 在这个项目中，我们发现 EPIC1 是一种致癌 lncRNA。乳腺癌，特别是管腔 B 亚型中，EPIC1 增强转录活性。我们提出了三个具体目标来检验这一假设。目标 1，我们将使用 HITS-CLIP 测定来表征 EPIC1-MYC 相互作用，并研究是否已识别 EPIC1 中的 MYC 结合序列对于 EPIC1-MYC 相互作用是必要且充分的。确定负责 EPIC1 结合的 MYC 蛋白的功能域在目标 2 中，我们将确定。通过过度表达 wt-EPIC1 或 MYC 结合来研究 EPIC1-MYC 轴在乳腺癌中的致癌作用然后我们将确定 ZR-75-1 和 BT-474 细胞中的序列缺失突变体 EPIC1 (ΔMYC-EPIC1)。最后，我们将研究异种移植小鼠模型中 EPIC1 致癌作用对 MYC 蛋白的依赖性。 EPIC1 与八个独立患者队列中乳腺癌临床特征和结果的关联包括 1882 个乳腺癌肿瘤在目标 3 中，我们将首先确定 MYC 与其靶标结合的模式。通过执行 RNA-seq 和 wt- 或 ΔMYC-EPIC1 过表达 ZR-75-1 和 BT-474 细胞中的基因启动子然后，我们将确定 EPIC1 是否通过 EPIC1-MYC 结合来调节 MYC 靶标。最后，我们将构建 EPIC1-cMYC 监管。我们的研究将通过整合 ChIP-seq、RNA-seq 和 TCGA 乳腺癌数据来建立乳腺癌网络。揭示一种新型致癌 lncRNA EPIC1 通过其与成熟的癌基因 MYC。