System analysis of p53 mutant associated cancer development with LFS patient- specific iPSCs

使用 LFS 患者特异性 iPSC 进行 p53 突变相关癌症发展的系统分析

基本信息

批准号：
9355607
负责人：
Huensuk Kim
金额：
$ 4.17万
依托单位：
ICAHN SCHOOL OF MEDICINE AT MOUNT SINAI
依托单位国家：
美国
项目类别：
财政年份：
2016
资助国家：
美国
起止时间：
2016-09-21 至 2018-08-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/9355607
关键词：
Address Agar Biological Assay CRISPR/Cas technology Cancer Model Cancer Patient Cell Differentiation process Cell Line Cell model Cells Chronic Myeloid Leukemia Clinical Clinical Trials Clone Cells Colon Carcinoma Computer Analysis Data Set Defect Development Disseminated Malignant Neoplasm Gene Expression Gene Targeting Genes H19 gene Hereditary Malignant Neoplasm Human In Vitro Knowledge Laboratories Li-Fraumeni Syndrome Malignant Neoplasms Mediating Messenger RNA Methods Modeling Molecular Molecular Profiling Mutation Oncogenes Oncogenic Organogenesis Organoids Osteoblasts Outcome Pathogenesis Pathologic Pathology Pathway interactions Patients Pharmaceutical Preparations Phase Phosphorylation Phosphotransferases Prevention Property Prostate Protein p53 Proteins Publishing Regenerative Medicine Regulation Reporting Research Research Personnel Resources Role Sampling Stem cells System Systems Analysis TP53 gene Technology Therapeutic Tissue Microarray Tissues Transcription Alteration Untranslated RNA Work cancer stem cell cancer type castration resistant prostate cancer cell behavior clinically relevant clinically significant comparative drug testing experimental study functional genomics genetic information genome editing genome integrity human disease imprint improved in vitro activity in vivo induced pluripotent stem cell insight melanoma mutant new therapeutic target novel osteoblast differentiation osteosarcoma overexpression pluripotency progenitor protein function receptor self-renewal therapeutic target three dimensional structure tool transcriptome tumor initiation tumorigenesis

项目摘要

Project Summary/Abstract Human induced pluripotent stem cells (iPSCs) not only represent a promising resource for regenerative medicine, but are also rising in their utility as a tool for studying human disease. Specifically, modeling cancer using patient specific iPSCs provides an opportunity to study human cancer pathogenesis. In our previous work in Cell 2015, `Modeling Familial Cancer with Induced Pluripotent Stem Cells', we demonstrated that iPSCs derived from human familial cancer patients of Li-Fraumeni Syndrome (LFS) mimic cancer pathogenesis in a dish. Additionally, systems analysis of LFS patient osteosarcoma development revealed that the gene H19 and its imprinted gene network, which is required for normal osteoblast differentiation, become dysregulated during LFS patient osteoblast differentiation. Although we reported the abnormal regulation of H19 and its imprinted gene network in LFS iPSC-derived osteoblasts, there is no known clinically feasible strategy to target abnormal H19 and its non-coding RNA regulation. In order to uncover clinically relevant and therapeutically targetable molecules, I continued researching our LFS iPSC-derived osteosarcoma model and identified secreted frizzled-related protein 2 (sFRP2) as an important oncogenic factor of osteosarcoma development. I further demonstrated that sFRP2 increases the phosphorylation of AXL receptor in osteoblasts, a known oncogene associated with colon cancer, melanoma, and chronic myelogenous leukemia. Additionally, using CRISPR/Cas9 gene editing technology I corrected the p53 mutation in our LFS iPSC lines. In this proposal, I will use functional and comparative analysis to identify the molecular differences of LFS- isogenic controls to wild type and the original LFS iPSCs. From this analysis, I anticipate showing that correcting the causative p53 mutation in osteosarcoma using CRISPR/Cas9 can reverse pathogenesis of osteosarcoma in an iPSC platform. Additionally, I will demonstrate the efficacy of targeting sFRP2 and the AXL receptor as potential therapeutic strategies for LFS-associated osteosarcoma. Finally, to check the clinical relevance of my approach, I will analyze the expression level of sFRP2 in human osteosarcoma samples using osteosarcoma tissue microarrays. Overall, the proposed studies will provide corrected iPSC clones that will serve as perfect isogenic controls to study the role of p53 mutation, as well as the resulting downstream changes in sFRP2 expression and AXL receptor phosphorylation, in LFS-associated osteosarcoma development. This work will not only expand our knowledge of p53 mutation mediated osteosarcoma progression, but also describe multiple potential therapeutic methods to improve clinical outcomes for p53 mutation osteosarcoma patients.

项目概要/摘要人类诱导多能干细胞（iPSC）不仅是一种有前景的再生资源医学，但作为研究人类疾病的工具，它们的实用性也在不断上升。具体来说，建立癌症模型使用患者特异性 iPSC 为研究人类癌症发病机制提供了机会。在我们之前的在 Cell 2015 的工作“用诱导多能干细胞模拟家族性癌症”中，我们证明了来自人类家族性李法美尼综合征 (LFS) 癌症患者的 iPSC 模拟癌症病机在菜中。此外，对 LFS 患者骨肉瘤发展的系统分析表明正常成骨细胞分化所需的基因 H19 及其印记基因网络变成 LFS 患者成骨细胞分化期间失调。尽管我们报道了 LFS iPSC 衍生的 H19 及其印记基因网络的异常调控成骨细胞中，尚无已知的临床可行策略针对异常 H19 及其非编码 RNA 规定。为了发现临床相关和治疗上可靶向的分子，我继续研究我们的 LFS iPSC 衍生骨肉瘤模型并鉴定出分泌性卷曲相关蛋白 2 （sFRP2）作为骨肉瘤发展的重要致癌因素。我进一步证明了 sFRP2 增加成骨细胞中 AXL 受体的磷酸化，这是一种与结肠相关的已知癌基因癌症、黑色素瘤和慢性粒细胞白血病。此外，使用 CRISPR/Cas9 基因编辑技术我纠正了我们的 LFS iPSC 系中的 p53 突变。在这个提案中，我将使用功能和比较分析来识别 LFS-的分子差异野生型和原始 LFS iPSC 的同基因对照。从这个分析中，我预计会表明使用 CRISPR/Cas9 纠正骨肉瘤中致病的 p53 突变可以逆转骨肉瘤的发病机制 iPSC 平台中的骨肉瘤。此外，我将展示针对 sFRP2 和 AXL 的功效受体作为 LFS 相关骨肉瘤的潜在治疗策略。最后，检查临床为了我的方法的相关性，我将使用以下方法分析人骨肉瘤样本中 sFRP2 的表达水平骨肉瘤组织微阵列。总体而言，拟议的研究将提供校正的 iPSC 克隆，作为完美的同基因对照研究 p53 突变的作用，以及由此产生的 sFRP2 表达和 AXL 的下游变化受体磷酸化，在 LFS 相关骨肉瘤的发展中。这项工作不仅会拓展我们的 p53 突变介导的骨肉瘤进展的知识，但也描述了多种潜在的改善 p53 突变骨肉瘤患者临床结果的治疗方法。