Establishing a patient-derived hiPSC differentiation protocol to model development of neural crest cancers

建立源自患者的 hiPSC 分化方案来模拟神经嵴癌的发展

• 批准号: 2749909
• 金额: --
• 依托单位: King's College London
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2022
• 资助国家: 英国
• 起止时间: 2022 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-2749909
• 关键词: Establishing; patient; derived; hiPSC; differentiation

Our current understanding of the initial stages of many cancers is still limited. This is because current human cancer models are often based on immortalised cell lines and cells derived from late-stage tumours. While these studies are important for assessing mature tumour behaviour, they are less helpful for deciphering critical steps during tumour formation and metastasis. Because of this gap in our knowledge, it is difficult to pinpoint the causative events, which may be of potential clinical value as early markers. Here, we use human induced pluripotent stem cells to model cancer initiation and migration. We are specifically interested cancers that arisefrom embryonic cells called neural crest cells. These cancers include neuroblastoma, a rare paediatric cancer. Neural crest cells are multipotent stem cells that give rise to diverse tissues such as melanocytes, Schwann cells, craniofacial skeleton and peripheral nervous system. During embryogenesis, neural crest cells delaminate from the neural tube, undergo epithelial-mesenchymal transition and migrate long distances in a process akin to metastasis. Our ultimate aim is to establish an assay allowing us to use patient-derived hiPSCs to assess the differentiation capacity of normal and diseased hiPSCs. To achieve this, we need to define reproducible protocols for differentiation of hiPSCs to neural crest. We will profile the developmental transitions as they differentiate to neural crest lineages including sympathetic nervous system and Schwann cells, which will allow us to pinpoint pathological triggers during differentiation. n this project, we will develop an in vitro platform for modelling development transitions during neural crest development. Specific aims:1)Compare the molecular and cellular definition of human PSCs-derived cranial and trunkneural crest lineages generated fromestablished protocols (STEMCELL Technologies). 2)Generate a transgenic Phox2b reporter line in human stem cells.3)Establish a differentiation protocol for generating trunk neural crest derivatives such assympathetic neurons, enteric neurons and Schwanncells. 4)Determine the functional consequence of neuroblastoma-associated genetic variants, including ALK-F1174L, on differentiation of trunk neural crest derivatives.The overallgoal will be to understand the cellular and molecular events driving neural crest lineage segregation.This will serve as a platform for functional analysis of the developmental events underlying disease aetiology, the identification of cellular events associated with variable outcomes in neuroblastoma, and identify novel therapeutic avenues.

我们目前对许多癌症的初始阶段的了解仍然有限。这是因为目前的人类癌症模型通常基于永生化细胞系和晚期肿瘤细胞。虽然这些研究对于评估成熟肿瘤的行为很重要，但它们对于解释肿瘤形成和转移过程中的关键步骤帮助不大。由于我们知识上的这一差距，很难确定病因事件，这可能是潜在的临床价值作为早期标志物。在这里，我们使用人类诱导多能干细胞来模拟癌症的发生和迁移。我们特别感兴趣的癌症是由称为神经嵴细胞的胚胎细胞产生的。这些癌症包括神经母细胞瘤，一种罕见的儿科癌症。神经嵴细胞是一种多能干细胞，可分化为黑素细胞、雪旺细胞、颅面骨骼和周围神经系统等多种组织。在胚胎发生过程中，神经嵴细胞从神经管分层，经历上皮-间充质转化，并在类似于转移的过程中长距离迁移。我们的最终目标是建立一种测定法，允许我们使用患者来源的hiPSC来评估正常和患病hiPSC的分化能力。为了实现这一点，我们需要定义可重复的方案，用于将hiPSC分化为神经嵴。我们将描述发育转变，因为它们分化为神经嵴谱系，包括交感神经系统和许旺细胞，这将使我们能够在分化过程中查明病理触发因素。在这个项目中，我们将开发一个体外平台，用于模拟神经嵴发育过程中的发育转变。具体目的：1）比较从已建立的方案（STEMCELL Technologies）产生的人PSC来源的颅嵴和干嵴谱系的分子和细胞定义。2)在人干细胞中建立转基因Phox 2b报告细胞系。3）建立用于产生躯干神经嵴衍生物如交感神经元、肠神经元和Schwann细胞的分化方案。4)确定神经母细胞瘤相关遗传变异（包括ALK-F1174 L）对躯干神经嵴衍生物分化的功能影响。总体目标是了解驱动神经嵴谱系分离的细胞和分子事件。这将作为一个平台，用于对疾病病因学基础的发育事件进行功能分析，识别与神经母细胞瘤可变结局相关的细胞事件，并发现新的治疗途径。