Elucidating Oncogenic Mechanisms Underlying Wilms Tumor Using Kidney Organoids

使用肾脏类器官阐明肾母细胞瘤的致癌机制

基本信息

批准号：
10543184
负责人：
Matthew Jared Stevenson
金额：
$ 3.92万
依托单位：
UNIV OF NORTH CAROLINA CHAPEL HILL
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：
2021-01-01 至 2023-12-31
项目状态：
已结题

来源：
https://reporter.nih.gov/project-details/10543184
关键词：
4 year old Address Adult Affect Affinity Chromatography Alleles Automobile Driving Binding Bioinformatics Biological Models CRISPR/Cas technology Cancer Biology Cell Cycle Cell Line Cells ChIP-seq Child Childhood Chronic Chronic Kidney Failure Classification Colorectal Cancer Complex DNA DNA Binding DNA Sequence Alteration Data Data Set Development Diagnosis Engineering Environment Excision Exhibits Fetal Kidney Foundations Gene Expression Gene Expression Profiling Generations Genes Genetic Genetic Transcription Genome engineering Genomics Goals Health Heterozygote Homeodomain Proteins Human Human Engineering Immunoprecipitation In Vitro Investigation Kidney Knock-in Knowledge Laboratories Malignant - descriptor Malignant Childhood Neoplasm Malignant Neoplasms Malignant childhood renal neoplasm Malignant neoplasm of ovary Mass Spectrum Analysis Migration Assay Missense Mutation Modeling Molecular Mus Mutation NF-kappa B Nephroblastoma Nephrons Oncogenic Organoids Patients Pattern Phenotype Play Proliferating Proteins Proteomics Recurrence Relapse Research Resistance Risk Role Second Primary Cancers Specificity System Technology Therapeutic Tissue-Specific Gene Expression Tissues Training Transcriptional Regulation Work aerobic glycolysis base cancer initiation cdc Genes chemotherapy cofactor data integration experimental study genome editing high risk homeodomain human model induced pluripotent stem cell innovation insight malignant breast neoplasm man migration mutant nephrogenesis nephron progenitor new therapeutic target novel overexpression programs protein protein interaction relapse risk skills stem cells targeted treatment transcription factor transcriptome sequencing treatment strategy tumor tumorigenesis tumorigenic

项目摘要

PROJECT SUMMARY/ABSTRACT Wilms tumor is the most common pediatric kidney cancer. Blastemal-predominant tumors are classified as high-risk and have demonstrated resistance to chemotherapeutic treatments, drastically increasing the likelihood for relapse compared to other subtypes. Current treatment consists of removal of the entire affected kidney followed by chemotherapy, or vice versa. Pediatric chemotherapy increases the risk for developing secondary cancers, while kidney removal eliminates up to 50% of a patient’s functioning nephrons. Clearly, safer and more effective targeted therapeutic strategies for blastemal-predominant Wilms tumor are needed to reduce or eliminate relapse while simultaneously retaining precious functional kidney tissue to mitigate the potential for chronic health issues. The goal of my proposed research is to elucidate the oncogenic mechanisms underlying blastemal-predominant Wilms tumor formation to identify novel candidate factors for the development of targeted therapies. These tumors express markers normally restricted to the nephron progenitor niche during fetal kidney development, including the transcription factor SIX1 which plays a critical role in establishing this niche. Sequencing of blastemal-predominant tumors identified a recurring missense mutation, Q177R, at a known DNA base-contacting residue within the evolutionary-conserved DNA binding homeodomain of SIX1. I hypothesize this mutation alters direct SIX1 regulatory interactions disrupting downstream networks, driving aberrant gene expression, obstructing nephron progenitor differentiation, and promoting a pro-oncogenic state. Utilizing cutting-edge technologies including CRISPR-Cas9 genome editing and human induced pluripotent stem cell-derived kidney organoids to model blastemal-predominant Wilms tumor in vitro, I aim to: 1) identify oncogenic phenotypes resulting from SIX1-Q177R through differentiation analysis, cell cycle profiling, proliferation, and migration assays and 2) define regulatory network disruptions downstream of SIX1-Q177R through ChIP-seq, RNA-seq and immunoprecipitation-mass spectrometry. Integration of genomic and proteomic data will pinpoint target genes directly regulated by SIX1-Q177R that are potentially contributing to oncogenic phenotypes. This pioneering work will provide a substantial foundation for understanding the molecular mechanisms contributing to the development of blastemal-predominant Wilms tumor, guiding the investigation of novel targeted therapies. Furthermore, this research will contribute vital knowledge to advance our fundamental understanding of the transcriptional control of human kidney development. This research will take place in a supportive and collaborative laboratory environment under the direction of my sponsor Dr. Lori L. O’Brien, an expert in kidney development and the regulatory control of nephron progenitor cell fate. With additional training provided by my co-sponsor Dr. Albert S. Baldwin, a leader in the field of NF-kB transcriptional networks and their roles in cancer, completion of this research and execution of my training plan will strengthen conceptual and experimental research skills and enhance professional development.

项目概要/摘要肾母细胞瘤是最常见的儿童肾癌。为高风险且已表现出对化疗治疗的耐药性，从而大大增加了与其他亚型相比，复发的可能性更大。目前的治疗包括切除整个受影响的区域。儿童化疗后进行肾移植，反之亦然，会增加患肾病的风险。显然，肾脏切除会消除患者多达 50% 的功能性肾单位。对于胚层为主的肾母细胞瘤，需要更安全、更有效的靶向治疗策略减少或消除复发，同时保留宝贵的功能性肾组织，以减轻我提出的研究的目标是阐明致癌机制。潜在的胚芽为主的肾母细胞瘤形成，以确定新的候选发展因素这些肿瘤表达的标记物通常仅限于肾单位祖细胞生态位。胎儿肾脏发育，包括转录因子 SIX1，它在建立这一过程中发挥着关键作用对胚层为主的肿瘤进行测序，发现了一个反复出现的错义突变 Q177R。 SIX1 进化保守的 DNA 结合同源域内已知的 DNA 碱基接触残基。保持这种突变改变了直接的 SIX1 调节相互作用，扰乱了下游网络，驱动异常的基因表达，阻碍肾单位祖细胞分化，并促进促癌状态。利用CRISPR-Cas9基因组编辑和人类诱导多能干细胞等尖端技术细胞源性肾脏类器官在体外模拟胚层为主的肾母细胞瘤，我的目标是：1）确定通过分化分析、细胞周期分析、由 SIX1-Q177R 产生的致癌表型，增殖和迁移测定以及 2) 定义 SIX1-Q177R 下游的调控网络中断通过 ChIP-seq、RNA-seq 和免疫沉淀-质谱法整合基因组和蛋白质组。数据将查明受 SIX1-Q177R 直接调控的靶基因，这些基因可能导致致癌这项开创性的工作将为理解分子提供坚实的基础。导致胚层为主的肾母细胞瘤发生的机制，指导研究此外，这项研究将为推进我们的研究贡献重要的知识。对人类肾脏发育的转录控制的基本了解。这项研究将在支持和协作的实验室环境下进行我的赞助商 Lori L. O’Brien 博士是肾脏发育和肾单位调控方面的专家我的共同发起人 Albert S. Baldwin 博士（该领域的领导者）提供了额外的培训。 NF-kB 转录网络领域及其在癌症中的作用，完成这项研究并执行我的培训计划将加强概念和实验研究技能并促进专业发展。