Systematic dissection of function and mechanism of long non-coding RNAs in glioblastoma

系统解析长非编码RNA在胶质母细胞瘤中的功能和机制

基本信息

批准号：
10390363
负责人：
Yiwen Chen
金额：
$ 40.5万
依托单位：
UNIVERSITY OF TX MD ANDERSON CAN CTR
依托单位国家：
美国
项目类别：
财政年份：
2021
资助国家：
美国
起止时间：
2021-04-15 至 2026-02-28
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10390363
关键词：
Adult Binding Biological Assay Biology Brain CCND1 gene CCNE2 gene CRISPR interference Cell Cycle Cell Cycle Progression Cell Proliferation Cells Clinical Sciences Code DNA Double Strand Break DNA Repair Data Development Diagnostic Diffuse Disease Dissection Double Strand Break Repair E2F transcription factors FOXM1 gene G1/S Transition Genes Genetic Transcription Genomics Glioblastoma Glioma Gliomagenesis Goals Human In Vitro Infiltration Ionizing radiation Maintenance Malignant - descriptor Mass Spectrum Analysis Mediating Messenger RNA Molecular Neuroglia Nucleotides Pathogenesis Patients Post-Transcriptional Regulation Primary Brain Neoplasms Prognostic Marker Proteins Proteomics RNA RNA-Binding Proteins Radiation Tolerance Radiation therapy Regulation Research Resistance Role Spinal Cord Neoplasms Stress Therapeutic Untranslated RNA Xenograft Model base brain tissue cell growth clinical prognosis crosslinking and immunoprecipitation sequencing effective therapy gain of function homologous recombination in vivo insight loss of function multiple omics novel novel therapeutic intervention protein complex public health relevance ribosome profiling self-renewal standard care stem cell proliferation stem cell self renewal stem cells therapeutically effective transcription factor transcriptome sequencing tumor

项目摘要

PROJECT SUMMARY/ABSTRACT Glioblastoma (GBM), a high-grade glioma (grade IV), is the most prevalent and malignant primary brain tumor in adults. There is no effective treatment of GBM. After standard treatment, the median survival of GBM patients is around 15 months. GBM is featured by enhanced cell proliferation, high propensity of invasion/diffuse infiltration throughout the brain, and resistance to chemo-/radiation therapy. Emerging evidence supports that long non-coding RNAs (lncRNAs, ~18,000 human lncRNA genes) can mediate tumor- promoting/suppressing effects and serve as independent diagnostic/prognostic biomarkers. Our previous integrative genomic study revealed that many lncRNAs show dysregulated expression, are associated with clinical prognosis, or harbor frequent somatic copy number alterations in GBM, suggesting an important role of lncRNAs in GBM pathogenesis. However, the function and mechanism of most lncRNAs in GBM are unknown. To fill this gap, our long-term goal is to leverage systematic multi-omic approaches to characterize the function and mechanism of lncRNAs in GBM, and to help develop new therapeutic strategies based on novel insight into lncRNA regulation in GBM. Our large-scale loss-of-function screen using CRISPR interference (CRISPRi) identified an antisense lncRNA, lnc-YINC (YBX1-interacting lncRNA) that was critical for GBM cell growth and was significantly up-regulated in GBM compared with normal brain tissue and low-grade glioma (LGG). The higher expression of lnc-YINC was associated with shorter overall survival of GBM patients. Functionally, lnc- YINC acted in-trans and was a key regulator of cell cycle progression of GBM cells/glioma stem cells (GSCs) and self-renewal of GSCs. It also protected GBM cells/GSCs from DNA double-strand breaks (DSBs) caused by endogenous stress or exogenous ionizing radiation. Mechanistically, lnc-YINC interacted with YBX1, and post-transcriptionally regulated the expression of key regulators of cell cycle or DSB repair, at least partially by stabilizing their mRNAs. Based on these new findings, we hypothesize that lnc-YINC is a GBM-promoting lncRNA that promotes proliferation of and modulates radio-sensitivity of GBM cells/GSCs, by enhancing YBX1 binding to key regulators of cell cycle/DSB repair and co-regulating their expression at post-transcriptional level. The objectives of this proposal are (a) to determine the role of lnc-YINC in gliomagenesis, (b) to integrate enhanced CLIP-seq (eCLIP-seq), ribosome profiling, RNA-seq and quantitative mass spectrometry data, with functional assays to identify the downstream targets of lnc-YINC/YBX1 that are key to regulating cell cycle or DSB repair, (c) to dissect the molecular mechanisms whereby lnc-YINC/YBX1 axis post-transcriptionally regulates the expression of key regulators of cell cycle or DSB repair, (d) to determine the role of lnc-YINC in modulating radio-sensitivity of GBM cells/GSCs and to investigate the therapeutic impact of combining inhibition of lnc-YINC with radiation therapy on GBM maintenance in vivo. This application is strengthened by a team of experts of basic/clinical science of GBM, DNA damage repair, RNA biology and proteomics.

项目概要/摘要胶质母细胞瘤 (GBM) 是一种高级胶质瘤（IV 级），是最常见和恶性的原发性脑肿瘤在成人中。 GBM 尚无有效治疗方法。标准治疗后，GBM 的中位生存期患者病程约15个月。 GBM 的特点是细胞增殖增强，整个大脑的侵袭/弥漫性浸润，以及对化疗/放疗的抵抗。新兴有证据表明长非编码 RNA（lncRNA，约 18,000 个人类 lncRNA 基因）可以介导肿瘤促进/抑制作用并作为独立的诊断/预后生物标志物。我们之前的综合基因组研究表明，许多 lncRNA 表现出表达失调，与临床预后，或 GBM 中存在频繁的体细胞拷贝数改变，表明 GBM 发病机制中的 lncRNA。然而，大多数lncRNA在GBM中的功能和机制尚不清楚。为了填补这一空白，我们的长期目标是利用系统的多组学方法来表征该功能 GBM 中 lncRNA 的研究和机制，并根据新的见解帮助开发新的治疗策略 GBM 中的 lncRNA 调控。我们使用 CRISPR 干扰 (CRISPRi) 进行大规模功能丧失筛查鉴定出一种对 GBM 细胞生长至关重要的反义 lncRNA lnc-YINC（YBX1 相互作用 lncRNA）与正常脑组织和低级别胶质瘤（LGG）相比，GBM 中的表达显着上调。这 lnc-YINC 的较高表达与 GBM 患者的总生存期较短相关。从功能上讲，lnc- YINC 发挥反式作用，是 GBM 细胞/神经胶质瘤干细胞 (GSC) 细胞周期进展的关键调节因子和 GSC 的自我更新。它还可以保护 GBM 细胞/GSC 免受 DNA 双链断裂 (DSB) 的影响通过内源性应力或外源性电离辐射。从机制上讲，lnc-YINC 与 YBX1 相互作用，并且转录后调节细胞周期或 DSB 修复关键调节因子的表达，至少部分是通过稳定它们的 mRNA。基于这些新发现，我们假设 lnc-YINC 是一种促进 GBM 的药物 lncRNA 通过增强 YBX1 促进 GBM 细胞/GSC 增殖并调节放射敏感性与细胞周期/DSB 修复的关键调节因子结合，并在转录后水平共同调节它们的表达。该提案的目标是 (a) 确定 lnc-YINC 在神经胶质瘤发生中的作用，(b) 整合增强的 CLIP-seq (eCLIP-seq)、核糖体分析、RNA-seq 和定量质谱数据，功能测定，以确定 lnc-YINC/YBX1 的下游靶标，这些靶标对于调节细胞周期或 DSB 修复，(c) 剖析 lnc-YINC/YBX1 轴转录后的分子机制调节细胞周期或 DSB 修复关键调节因子的表达，(d) 确定 lnc-YINC 在调节 GBM 细胞/GSC 的放射敏感性并研究组合的治疗影响放疗抑制lnc-YINC对体内GBM维持的影响。该应用程序通过以下方式得到加强： GBM基础/临床科学、DNA损伤修复、RNA生物学和蛋白质组学专家团队。