Mechanism of Long Non-coding RNAs Action in leiomyoma

长链非编码RNA在平滑肌瘤中的作用机制

• 批准号: 10436359
• 负责人: OMID A. KHORRAM
• 金额: $ 40.27万
• 依托单位: LUNDQUIST INSTITUTE FOR BIOMEDICAL INNOVATION AT HARBOR-UCLA MEDICAL CENTER
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-08 至 2024-06-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10436359
• 关键词: Achievement; Address; African American population; Age; Animal Model; Area; Benign; Bioinformatics; Cell Cycle; Cell Cycle Progression; Cell Proliferation; Cells; Characteristics; Code; Complex; Data; Development; Disease; Down-Regulation; Enantone; Epigenetic Process; Epithelial; Estrogens; Ethnic group; Extracellular Matrix; Female Genital Neoplasms; Fibroid Tumor; Fibrosis; Genes; Genetic Transcription; Gonadal Steroid Hormones; Growth; High Prevalence; Human Genome; Immunoprecipitation; Inflammation; Inflammatory; Investigation; Kidney; Knock-in; Knowledge; Laboratories; Leiomyoma; Lentivirus; Luteal Phase; Mediator of activation protein; Mesenchymal; Messenger RNA; Modeling; Modification; Mus; Mutation; Ovarian; Pain; Pathogenesis; Pathway interactions; Patients; Phase; Porifera; Post-Transcriptional Regulation; Preclinical Testing; Progesterone; Proteins; Publishing; RNA; Race; Regulation; Rest; Role; Severities; Small Interfering RNA; Small RNA; Smooth Muscle Myocytes; Sp1 Transcription Factor; Steroids; Symptoms; Testing; Therapeutic; Tissues; Transplantation; United States National Institutes of Health; Untranslated RNA; Up-Regulation; Uterine Fibroids; Uterine Neoplasms; Uterine hemorrhage; Woman; Work; base; capsule; cdc Genes; chromosomal location; clinically relevant; experimental study; fibrogenesis; gene network; in vivo imaging; in vivo monitoring; knock-down; mRNA Expression; myometrium; new therapeutic target; next generation; next generation sequencing; overexpression; pressure; reproductive; therapeutic target; transcriptome; transcriptome sequencing; tumor; tumor growth; tumorigenesis

Uterine leiomyoma (fibroids) are benign tumors afflicting a significant number of reproductive age women and without a known cause. Our laboratory has focused on understanding how miRNAs impact pro-inflammatory and pro-fibrotic pathways in leiomyoma, and identified two miRNAsmiR-200c and miR-29c which primarily target cell cycle, inflammation and extracellular matrix (ECM) component genes respectively as key in the development of leiomyoma. Our recent findings using next generation sequencing has demonstrated a whole host of non-coding RNAs including long non-coding RNAs (lncRNAs) are dysregulated in fibroids. LncRNAs can act as sponge for miRNAs and therefore may be a driver of gene dysregulation in leiomyomas. Based on the level of expression and functional association with tumorigenesis and tissue fibrosis we selected a group of lncRNAs to further characterize. Using QRT-PCR we detected marked overexpression of XIST and MIAT in leiomyomas. The sequences of these lncRNAs and preliminary data suggest they can act as miRNA sponges for miR-29c and miR-200c. Therefore, based on our published and preliminary data we hypothesize that leiomyoma as compared to myometrium display an altered expression of a number of lncRNAs that are transcriptionally regulated and independently or through miRNA-guided mechanisms regulate the expression of specific target genes functionally associated with cell cycle progression, inflammation, fibrosis and epigenetic modification all of which are central to leiomyoma pathogenesis. To test our core hypothesis we propose 3 specific aims. In Aim 1 we will build on our preliminary RNA sequencing study and provide a comprehensive profile of lncRNAs, miRNAs, with concurrent mRNAs expression in large sets of paired leiomyoma and myometrium from different race/ethnic groups derived from proliferative and secretory phase of the menstrual cycle. Through bioinformatic analysis we will identify lncRNAs subtypes, their chromosomal locations at loci often rearranged in leiomyoma and identify lncRNAs which could potentially act as miRNA sponges, or competing endogenous RNAs (ceRNAs). We will also determine if presence of MED12 mutations has any impact on the profiles of these ncRNAs. Aim 2 will address the regulation and function of MIAT and XIST in leiomyoma. Using RNA immunoprecipitation studies we will determine if these lncRNAs will interact with miR-29c and miR-200c known to be important in fibrogenesis. At the cellular level we will assess the role of ovarian steroids on their expression and through knockdown and knock-in strategies identify their effects on expression of ECM and cell cycle genes. To establish the clinical relevance and therapeutic potential of lncRNAs in Aim 3 we will determine the effect of altering the expression of XIST and MIAT which are highly overexpressed in fibroids on fibroid progression and/or establishment in a leiomyoma animal model. This translational proposal addresses a significant gap in knowledge in the pathogenesis of fibroids which is a priority area of investigation for NIH, and could have potential therapeutic applications for treatment of fibroids.

子宫肌瘤（肌瘤）是良性肿瘤，困扰着大量育龄妇女和 没有已知原因。我们的实验室致力于了解 miRNA 如何影响促炎症 和平滑肌瘤中的促纤维化途径，并鉴定了两个 miRNA——miR-200c 和 miR-29c，它们主要 靶细胞周期、炎症和细胞外基质 (ECM) 组成基因分别作为关键 平滑肌瘤的发展。我们最近使用下一代测序的发现证明了整个 包括长非编码 RNA (lncRNA) 在内的非编码 RNA 宿主在肌瘤中失调。长链非编码RNA 可以充当 miRNA 的海绵，因此可能是平滑肌瘤基因失调的驱动因素。基于 为了研究表达水平以及与肿瘤发生和组织纤维化的功能相关性，我们选择了一组 lncRNA 需要进一步表征。使用 QRT-PCR，我们检测到 XIST 和 MIAT 在 平滑肌瘤。这些 lncRNA 的序列和初步数据表明它们可以充当 miRNA 海绵 对于 miR-29c 和 miR-200c。因此，根据我们发布的初步数据，我们假设 与子宫肌瘤相比，平滑肌瘤显示许多 lncRNA 的表达发生改变，这些 lncRNA 转录调节并独立或通过 miRNA 引导的机制调节 与细胞周期进程、炎症、功能相关的特定靶基因的表达 纤维化和表观遗传修饰都是平滑肌瘤发病机制的核心。来测试我们的 核心假设我们提出了 3 个具体目标。在目标 1 中，我们将基于我们的初步 RNA 测序研究 并提供 lncRNA、miRNA 的全面概况，以及大量的并发 mRNA 表达 来自不同种族/民族的成对平滑肌瘤和子宫肌瘤源自增殖和分泌 月经周期的阶段。通过生物信息学分析，我们将识别lncRNA亚型，它们的亚型 平滑肌瘤中经常重排的基因座的染色体位置并鉴定出可能发挥作用的lncRNA 例如 miRNA 海绵或竞争性内源 RNA (ceRNA)。我们还将确定是否存在 MED12 突变对这些 ncRNA 的谱有影响。目标 2 将解决以下问题的监管和职能： 平滑肌瘤中的 MIAT 和 XIST。通过 RNA 免疫沉淀研究，我们将确定这些 lncRNA 是否会 与已知在纤维发生中重要的 miR-29c 和 miR-200c 相互作用。在细胞水平上，我们将评估 卵巢类固醇对其表达的作用，并通过敲低和敲入策略识别其表达 对 ECM 和细胞周期基因表达的影响。确定临床相关性和治疗潜力 在目标 3 中，我们将确定改变 XIST 和 MIAT 表达的效果，这两个因子高度依赖于 lncRNA。 在肌瘤动物模型中肌瘤进展和/或建立时过度表达。这 转化提案解决了肌瘤发病机制方面的巨大知识空白，这是一个 NIH 的优先研究领域，并且可能具有治疗肌瘤的潜在治疗应用。