Telomere dysfunction and telomerase reactivation in the etiology and progression of liver cancer

肝癌病因和进展中的端粒功能障碍和端粒酶再激活

基本信息

批准号：
10360832
负责人：
Luis Francisco Zirnberger Batista
金额：
$ 36.03万
依托单位：
WASHINGTON UNIVERSITY
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：
2021-12-13 至 2026-11-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10360832
关键词：
Address Biochemical Cell Lineage Cell model Cells Cicatrix Complex Copy Number Polymorphism DNA Damage DNA Repair Data Development Disease Distal Elements Etiology Event Failure Fibrosis Functional disorder Genomic Instability Goals Hepatic Stellate Cell Hepatic Tissue Hepatocyte Human Impairment In Vitro Knockout Mice Knowledge Length Link Liver Liver Failure Liver Fibrosis Malignant neoplasm of liver Methylation Modeling Molecular Mutation Pathogenesis Pathway interactions Patient-Focused Outcomes Patients Physiological Positioning Attribute Primary carcinoma of the liver cells Process Promoter Regions Protocols documentation RNA-Directed DNA Polymerase Risk Factors Role Signal Pathway Somatic Mutation System TP53 gene Telomerase Telomere Maintenance Telomere Shortening Time cell growth regulation cell injury connective tissue growth factor end stage liver disease hepatocyte nuclear factor human embryonic stem cell human pluripotent stem cell in vivo liver function loss of function mutation molecular modeling mutant novel paracrine prevent promoter response stellate cell stem cell biology stem cell differentiation targeted treatment telomere tool transcription factor

项目摘要

PROJECT SUMMARY/ABSTRACT Mutations in telomerase and telomere attrition are major risk factors for liver fibrosis and its progression to hepatocellular carcinoma (HCC). However, due to a lack of adequate models and intrinsic difficulties in studying human telomerase in physiologically relevant cells, the molecular mechanisms responsible for liver fibrosis and cancer in settings of DNA damage arising from short telomeres remain elusive. While telomerase knockout mice corroborate the importance of telomere maintenance and DNA repair for liver function, the molecular mechanisms that govern liver abnormalities in patients with damaged telomeres are still unknown. Likewise, the specific signaling pathways that trigger failure of hepatic cells following telomere shortening and accumulation of DNA damage remain to be determined. In addition, mutations in the promoter region of the telomerase reverse transcriptase component (TERT) have been described as the initial and most prevalent mutation in HCC. While these mutations have been shown to reactivate telomerase, the functional relevance of this process during failure and transformation of hepatic cells has yet to be interrogated. The focus of this proposal is to use human pluripotent stem cells as a novel platform to understand the detrimental effects of mutant telomerase, telomere shortening and accumulation of DNA damage in different hepatic cell lineages. We have previously generated isogenic hPSC lines harboring several disease-specific mutations in telomerase and have successfully derived telomerase-mutant human hepatocytes and hepatic stellate cells in vitro, following established protocols that recapitulate the in vivo development of these lineages. Here, two specific aims are proposed that utilize this platform to understand the molecular consequences of telomere erosion, DNA damage, and telomerase impairment for the function of hepatic cells, and to determine their role during early stages of transformation. In Aim 1 we will determine the role of telomere shortening and DNA damage accumulation during fibrotic failure of different hepatic cell lineages with impaired telomerase. We will determine the extent to which mitigation of DNA damage, reactivation of HNF4α, and modulation p53 prevent fibrotic triggering in telomerase-mutant hepatocytes with variable telomere lengths. As liver fibrosis and its progression to HCC are multicellular responses we will determine the role of progressive telomere shortening during the direct and the paracrine fibrotic activation of hepatic stellate cells. In Aim 2, we will investigate the molecular consequences of mutations in the TERT promoter region during progression of HCC, in settings of exacerbated DNA damage due to eroded telomeres. Specifically, we will analyze the biochemical and functional consequences of mutations in the TERT promoter region for hepatocyte function and immortalization. These studies will determine the molecular mechanisms of liver fibrosis and its progression to HCC in settings of mutant telomerase and DNA damage. Our unique tools, combined with our expertise in telomerase, DNA repair, and stem cell biology puts us in an ideal position to make a significant impact in this field.

项目摘要/摘要端粒酶和端粒损耗的突变是肝纤维化及其进展的主要危险因素到肝细胞癌（HCC）。但是，由于缺乏足够的模型和内在困难在物理相关细胞中研究人端粒酶，肝脏的分子机制短端粒引起的DNA损伤环境中的纤维化和癌症仍然难以捉摸。而端粒敲除小鼠证实了端粒维持和DNA修复对肝功能的重要性，控制端粒受损患者肝异常的分子机制仍然未知。同样，端粒缩短和 DNA损伤的积累仍有待确定。另外，在启动子区域的突变端粒酶逆转录酶成分（TERT）已被描述为初始和最普遍的 HCC中的突变。尽管这些突变已显示为重新激活端粒酶，但功能相关性在肝细胞失败和转化过程中的这一过程尚未受到质疑。该提案的重点是将人类多能干细胞用作一个新的平台，以了解突变端粒酶，端粒缩短和DNA损伤的积累的有害影响肝细胞谱系。我们以前已经产生了具有多种疾病特异性的等源性HPSC线端粒酶的突变，并成功得出端粒酶突变的人肝细胞和肝细胞遵循建立的方案，概括了这些线的体内发展，因此体外的星状细胞体外。在这里，提出了两个具体的目的，它利用该平台来了解分子后果端粒侵蚀，DNA损伤和端粒酶损伤的肝细胞功能，并确定它们在转型的早期阶段的作用。在AIM 1中，我们将确定端粒缩短和端粒酶受损的不同肝细胞谱系纤维化衰竭期间的DNA损伤积累。我们将确定DNA损伤的缓解程度，HNF4α的重新激活和调节p53防止端粒酶突变剂肝细胞的纤维化触发，其长度可变。作为肝纤维化及其向HCC的发展是多细胞反应，我们将确定进行性端粒缩短的作用在直接和甲状腺星状细胞的旁分泌纤维化激活期间。在AIM 2中，我们将调查 HCC进展过程中TERT启动子区域突变的分子后果，由于端粒侵蚀而导致的DNA损伤加剧。具体而言，我们将分析生化和功能 TERT启动子区域突变在肝细胞功能和永生化中的后果。这些研究将确定肝纤维化的分子机制及其在HCC中的发展突变端粒酶和DNA损伤的设置。我们的独特工具，结合我们在端粒酶方面的专业知识， DNA修复和干细胞生物学使我们处于在该领域产生重大影响的理想位置。