Determining the developmental origins of retinoblastoma

确定视网膜母细胞瘤的发育起源

• 批准号: 9760981
• 负责人: Jacqueline Norrie
• 金额: $ 6.16万
• 依托单位: ST. JUDE CHILDREN'S RESEARCH HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-04-01 至 2022-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/9760981
• 关键词: 2 year old; Address; Affect; Age; Alleles; Animal Model; Biological Markers; Biological Models; Caring; Cell Line; Cells; Child; Childhood; Data; Development; Diagnosis; Disease; Disease model; Electron Microscopy; Excision; Eye; Eye Neoplasms; Genetic; Genetic Predisposition to Disease; Germ-Line Mutation; Goals; Human; Immune; Inherited; Life; Loss of Heterozygosity; Malignant Neoplasms; Modeling; Molecular; Mus; Mutation; Oncogenic; Organoids; Parents; Patients; Pattern; Play; Preclinical Testing; Protocols documentation; Quality of life; RB1 Gene Inactivation; RB1 gene; Reporter; Reporting; Retina; Retinal; Retinoblastoma; Role; SYK gene; Specificity; Structure; Study models; Survival Rate; System; Time; Tumor stage; Untranslated RNA; Xenograft procedure; base; cancer initiation; cell type; chemotherapy; effective therapy; exome sequencing; functional loss; human embryonic stem cell; human model; in vivo; induced pluripotent stem cell; insight; malignant neoplasm of eye; mouse model; novel; outcome forecast; research clinical testing; single cell sequencing; targeted treatment; trait; transcriptome sequencing; tumor; tumor growth; tumor initiation; tumorigenesis; virtual

Project Summary/Abstract Retinoblastoma is the most common pediatric eye cancer and is caused by the biallelic inactivation of the tumor suppressor gene RB1. There are between 200 and 300 new patients with retinoblastoma in the U.S. each year, most diagnosed before two years of age. When caught early, and with proper care, survival from retinoblastoma is relatively high. However, treatments, including enucleations, or removal of the eye, have lasting effects on the lives of the children. Furthermore, retinoblastomas with extraocular involvement have a much poorer prognosis. While animal models of retinoblastoma have been useful for preclinical and clinical testing of retinoblastoma treatment, mice with loss of RB1 never develop retinoblastoma and therefore cannot accurately recapitulate human retinoblastoma formation. This has led to gaps in our understanding of the tumor initiation, development, and cellular specificity of human retinoblastoma. The goal of this study is to use our new model of spontaneous human retinoblastoma using patient derived iPSCs to study the developmental origin(s) of retinoblastoma and characterize the mechanisms of the second hit mutation. We have generated 15 induced pluripotent stem cell (iPSC) lines derived from patients with germline mutations in RB1. Using an optimized retinal differentiation culture, we have differentiated the patient lines into retina, dissociated the organoids, injected the cells into mice to screen for tumor formation. The resulting tumors have expression patterns similar to retinoblastoma patient derived xenografts. I now have a model system to study retinoblastoma tumor formation during retinal differentiation. I will use our new retinoblastoma reporter line in H9 hESCs, SYK-GFP, and a combination of immunostaining, single cell sequencing and electron microscopy to determine cell of origin or retinoblastoma. Additionally, I will analyze the tumors over multiple rounds of differentiation to determine the mechanism of the second hit and if there is any genetic predisposition to the loss of heterozygosity that occurs in patients with germline RB1 mutations. This study will allow us, for the first time, to answer question about retinoblastoma initiation and progression. Defining the cell of origin or retinoblastoma and the mechanism of the second hit of RB1 will not only increase our depth of understanding of retinoblastoma development but can be more generally applied to the fundamental understanding of cancer initiation. Furthermore, the data provided by this study will hopefully provide insight to create more targeted therapies for children with retinoblastoma.

项目概要/摘要 视网膜母细胞瘤是最常见的儿科眼癌，是由视网膜母细胞瘤的双等位基因失活引起的 抑癌基因RB1。美国有 200 至 300 名新的视网膜母细胞瘤患者。 每年，大多数在两岁之前被诊断出来。如果及早发现并采取适当的护理，就可以生存 视网膜母细胞瘤的发病率相对较高。然而，包括摘除眼睛或摘除眼睛在内的治疗方法已经 对孩子的生活产生持久的影响。此外，累及眼外的视网膜母细胞瘤具有 预后较差。虽然视网膜母细胞瘤的动物模型可用于临床前和临床 在视网膜母细胞瘤治疗测试中，RB1 缺失的小鼠从未患上视网膜母细胞瘤，因此不能 准确地再现人类视网膜母细胞瘤的形成。这导致了我们对这个问题的理解存在差距。 人类视网膜母细胞瘤的肿瘤发生、发展和细胞特异性。 本研究的目标是使用我们的自发性人类视网膜母细胞瘤新模型，该模型使用患者衍生的 iPSC 用于研究视网膜母细胞瘤的发育起源并表征第二种视网膜母细胞瘤的机制 命中突变。我们已经生成了 15 个源自患有以下疾病的患者的诱导多能干细胞 (iPSC) 系： RB1 种系突变。使用优化的视网膜分化培养，我们对患者进行了分化 将细胞线植入视网膜，分离类器官，将细胞注射到小鼠体内以筛选肿瘤形成。这 由此产生的肿瘤具有与视网膜母细胞瘤患者来源的异种移植物相似的表达模式。我现在有一个 研究视网膜分化过程中视网膜母细胞瘤肿瘤形成的模型系统。我将使用我们的新 H9 hESC 中的视网膜母细胞瘤报告系、SYK-GFP 以及免疫染色、单细胞的组合 测序和电子显微镜以确定细胞来源或视网膜母细胞瘤。另外我来分析一下 对肿瘤进行多轮分化以确定第二次打击的机制以及是否存在 种系 RB1 突变患者中发生的杂合性丧失的任何遗传倾向。 这项研究将使我们首次回答有关视网膜母细胞瘤发生和进展的问题。 定义起源细胞或视网膜母细胞瘤以及RB1第二次打击的机制不仅会增加 我们对视网膜母细胞瘤发展的理解深度，但可以更广泛地应用于 对癌症发生的基本了解。此外，这项研究提供的数据有望 提供见解，为患有视网膜母细胞瘤的儿童创造更有针对性的治疗方法。