Roles of tGLI1 and microRNA Network in Breast Cancer Brain Metastasis

tGLI1 和 microRNA 网络在乳腺癌脑转移中的作用

基本信息

批准号：
10702139
负责人：
HUI-WEN LO
金额：
$ 41.4万
依托单位：
UNIVERSITY OF TEXAS HLTH SCI CTR HOUSTON
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-04-01 至 2025-03-31
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10702139
关键词：
Alternative Splicing Astrocytes Brain Breast Cancer Cell Breast Cancer Model Breast cancer metastasis Cancer Biology Cell Line Cessation of life Clinical Data Distant ERBB2 gene Exons Family Fatty acid glycerol esters GLI gene Genes Goals Hand Human In Vitro Injections Laboratories Malignant Neoplasms Mediating Mediator of activation protein Metastatic Neoplasm to Lymph Nodes Metastatic malignant neoplasm to brain MicroRNAs Modeling Molecular Mouse Mammary Tumor Virus Mus Neoplasm Circulating Cells Neoplasm Metastasis Organ Pathway interactions Patients Pattern Play Prognosis Protein Isoforms Proteins Role Route SHH gene Sampling Sampling Studies Signal Transduction Stains TP53 gene Testing Transcription Repressor Transgenic Mice Transgenic Organisms Up-Regulation Validation Xenograft procedure Zinc Fingers angiogenesis autocrine base brain cell cancer cell cytokine exosome gain of function implantation in vivo insight knock-down malignant breast neoplasm malignant phenotype mammary migration monomer mouse model neoplastic cell neurotropic novel nucleocytoplasmic transport overexpression paracrine patient derived xenograft model transcription factor tumor tumor growth

项目摘要

The goal of this proposal is to determine the role of tGLI1 (truncated glioma-associated oncogene homolog 1) in breast cancer brain metastasis (BCBM). Our laboratory discovered tGLI1 as an alternatively spliced GLI1 that lacks entire exon 3 and part of exon 4, but retains the ability to undergo nuclear transport and respond to sonic hedgehog-smoothened signaling. In addition to activating known GLI1 target genes, tGLI1 gains the ability to activate genes not regulated by GLI1, leading to increased migration, invasion and angiogenesis. Whether tGLI1 plays any role in metastasis of any tumor type is unknown. Our mouse studies indicated that tGLI1 promotes breast cancer preferential metastasis to the brain and conversely, tGLI1 knockdown selectively suppressed BCBM. tGLI1 protein is overexpressed in lymph node metastases and BCBM samples. In elucidating how tGLI1 promotes BCBM, we found that exosomes secreted from tGLI1-high cancer cells strongly activated astrocytes, the most abundant brain cells known to promote tumor growth when activated, and that tGLI1-high cancer cells had an increased ability to interact with and activate astrocytes in vitro and in vivo. Since microRNAs can be loaded into exosomes and circulating tumor exosomal miRNAs can prime distant organs for organ-specific metastasis, we conducted an exosomal miRNA microarray followed by validations and found that tGLI1-high cancer cells secreted high levels of exosomal miR-1290 and miR-1246 in vitro and in vivo. Whether miR-1290 and miR-1246 play any role in brain metastasis of any cancer or astrocytes is unknown. We observed that miR-1290/1246 inhibited expression of two transcription repressors (FOXA2 and SOX9), which leads to secretion of ciliary neurotropic factor (CNTF) to activate astrocytes and stimulate cancer cells. We hypothesize that tumoral tGLI1 promotes brain metastasis by priming astrocytes in the metastatic niche, and activated astrocytes in turn facilitate BCBM through secreting CNTF. We further hypothesize that tGLI1-high breast cancer cells prime astrocytes via the novel signaling axis: tumoral tGLI1→exosomal miR-1290/1246┤astrocyte FOXA2/ SOX9┤astrocyte CNTF→astrocyte activation. In Aim 1, we will determine the extent to which tGLI1 promotes BCBM using three mouse models (cell line-derived xenograft, PDX, and transgenic mice), overexpression and knockdown approaches, two inoculation routes (intracardiac and mammary fat pad injections), and human patient samples. In Aim 2, we will examine whether and how tumoral tGLI1 upregulates exosomal miR- 1290/1246 to activate astrocytes in the metastatic niche, and clarify how miR-1290/1246 suppress FOXA2 and SOX9 expression within astrocytes, how FOXA2 and SOX9 repress CNTF expression, and the role of CNTF in astrocyte activation and BCBM progression. In Aim 3, we will examine if miR-1290 and miR-1246 play essential roles in tGLI1-mediated BCBM, identify their downstream targets, and elucidate the roles of intracellular miR- 1290/1246 in BCBM. The project could define tGLI1, miR-1290/1246, and CNTF as novel mediators of BCBM and regulators of astrocytes in the metastatic niche, thus providing novel mechanistic insights into BCBM.

该提案的目标是确定 tGLI1（截短的神经胶质瘤相关癌基因同源物 1）的作用在乳腺癌脑转移 (BCBM) 中，我们的实验室发现了 tGLI1 作为选择性剪接的 GLI1。缺乏整个外显子 3 和部分外显子 4，但保留进行核运输和响应声波的能力刺猬平滑信号除了激活已知的 GLI1 靶基因外，tGLI1 还具有以下能力：激活不受 GLI1 调控的基因，导致迁移、侵袭和血管生成增加。我们的小鼠研究表明 tGLI1 在任何肿瘤类型的转移中发挥的作用尚不清楚。乳腺癌优先转移至大脑，相反，tGLI1 敲除选择性抑制 BCBM。tGLI1 蛋白在淋巴结转移和 BCBM 样本中过度表达，以阐明 tGLI1 的作用。促进BCBM，我们发现tGLI1高的癌细胞分泌的外泌体强烈激活星形胶质细胞，已知最丰富的脑细胞在激活时会促进肿瘤生长，并且 tGLI1 高的癌细胞由于 microRNA 可以在体外和体内与星形胶质细胞相互作用并激活星形胶质细胞，因此具有增强的能力。加载到外泌体和循环肿瘤外泌体 miRNA 中可以为远处器官做好器官特异性准备转移，我们进行了外泌体 miRNA 微阵列，然后进行了验证，发现 tGLI1-high 癌细胞在体外和体内均分泌高水平的外泌体 miR-1290 和 miR-1246。我们观察到miR-1246在任何癌症或星形胶质细胞的脑转移中发挥的作用尚不清楚。 miR-1290/1246 抑制两种转录抑制因子（FOXA2 和 SOX9）的表达，从而导致分泌睫状神经营养因子（CNTF）激活星形胶质细胞并刺激癌细胞。肿瘤 tGLI1 通过在转移微环境中启动星形胶质细胞并激活星形胶质细胞来促进脑转移我们进一步研究了tGLI1高的乳腺癌细胞。通过新的信号轴启动星形胶质细胞：肿瘤 tGLI1→外泌体 miR-1290/1246┤星形胶质细胞 FOXA2/ SOX9┤星形胶质细胞 CNTF→星形胶质细胞激活在目标 1 中，我们将确定 tGLI1 促进的程度。 BCBM 使用三种小鼠模型（细胞系来源的异种移植小鼠、PDX 和转基因小鼠）、过表达和击倒方法、两种接种途径（心内和乳腺脂肪垫注射）和人类在目标 2 中，我们将检查肿瘤 tGLI1 是否以及如何上调外泌体 miR- 1290/1246 激活转移微环境中的星形胶质细胞，并阐明 miR-1290/1246 如何抑制 FOXA2 和 SOX9 在星形胶质细胞内的表达、FOXA2 和 SOX9 如何抑制 CNTF 表达以及 CNTF 在星形胶质细胞中的作用在目标 3 中，我们将检查 miR-1290 和 miR-1246 是否至关重要。 tGLI1 介导的 BCBM 中的作用，识别其下游靶标，并阐明细胞内 miR- BCBM 中的 1290/1246 该项目可以将 tGLI1、miR-1290/1246 和 CNTF 定义为 BCBM 的新型介质。以及转移微环境中星形胶质细胞的调节因子，从而为 BCBM 提供新的机制见解。