Defining the stem cell origins of brain tumours containing non-neural cells

定义含有非神经细胞的脑肿瘤的干细胞起源

• 批准号: 1940968
• 金额: --
• 依托单位: University of Leeds
• 依托单位国家: 英国
• 项目类别: Studentship
• 财政年份: 2017
• 资助国家: 英国
• 起止时间: 2017 至 无数据
• 项目状态: 已结题
• 来源: https://gtr.ukri.org/projects?ref=studentship-1940968
• 关键词: Defining; stem; cell; origins; brain

A striking feature of some central nervous system (CNS) tumours is the presence of cell types indicative of mixed neural/glial and mesodermal/mesenchymal (e.g. skeletal/smooth muscle, cartilage, bone) differentiation. These "neuromesodermal" (NM) tumours affect both young children and adults and they are highly aggressive being linked to a poor prognosis. For example, a considerable fraction of gliomas, the most common adult brain tumours, display mesenchymal features with elements of bone, cartilage and muscle differentiation and some of them contain a distinct muscular sarcomatous component (gliosarcomas). Defining the cell entity driving the formation of these tumours will be pivotal for the design of effective therapeutic interventions against these malignancies. The proposed PhD project aims to examine the hypothesis that NM tumours arise from stem cells which resemble multipotent embryonic precursors, namely neuromesodermal progenitors (NMPs) or neural crest (NC) precursors. During early embryonic development NMPs give rise to spinal cord neurons as well as bone, cartilage and skeletal/smooth muscle whereas NC cells generate peripheral neurons, melanocytes and mesenchymal cell types such as smooth muscle and cartilage. We speculate that NM tumourigenesis is the result of either a mutation-driven conversion of NMPs/NC progenitors into cancer-initiating cells during embryogenesis or a "reprogramming in situ" event where corruption of the genetic/epigenetic machinery within neural/glial cells results into their conversion into NMP/NC-like cells which in turn trigger tumorigenesis within a favorable niche.Research planCurrent conventional methods for establishing brain tumour cell lines are designed to select for neural stem cell (NSC)-like cells reflecting the hypothesis that the cell-of-origin of all brain tumours is a transformed neural progenitor. Thus adherent glioma stem cell lines have been successfully established using the same conditions as those employed for the propagation of neural stem cells. However, this approach cannot be employed for the isolation of bipotent NM cells resembling NMPs/NC cells. We have recently defined the optimal culture conditions for the induction of NMPs and NC progenitors from human pluripotent stem cells (hPSCs) in vitro. Adult brain tumours will be dissociated and equal numbers of the resulting cells will be cultured in parallel in NMP, NC and neural stem cell (NSC) conditions (positive control). Successful attachment, proliferation and subsequent passaging in NMP or NC culture media would mark the presence of NM bipotent stem cells in the starting tumour. We have already obtained promising preliminary data showing that glioma-derived cells can proliferate in NMP induction conditions. The presence of bipotent cells in the resulting cell lines will be confirmed by monitoring the expression of NMP and NC markers using immunofluorescence/qPCR and in vitro differentiation assays. RNA-seq analysis will be carried out comparing the transcriptomes of NM tumour cell lines to hPSC-derived NMPs and NC cells as well as established human NSC lines to define candidate drivers of oncogenic transformation. Assessment of tumourigenic potential by xenografting/stereotactic injection into NOD-SCID mice will also be performed. If time permits we will also carry out small molecule screens aiming to identify compounds exerting cytotoxic/cytostatic effects on the NM tumour cell lines.

一些中枢神经系统（CNS）肿瘤的显著特征是存在指示混合神经/神经胶质和中胚层/间充质（例如骨骼肌/平滑肌、软骨、骨）分化的细胞类型。这些“神经中胚层”（NM）肿瘤影响幼儿和成人，并且它们具有高度侵袭性，与预后不良有关。例如，相当一部分的神经胶质瘤，最常见的成人脑肿瘤，显示具有骨、软骨和肌肉分化元素的间充质特征，其中一些包含明显的肌肉肉瘤成分（神经胶质瘤）。确定驱动这些肿瘤形成的细胞实体对于设计针对这些恶性肿瘤的有效治疗干预措施至关重要。拟议的博士项目旨在研究NM肿瘤起源于干细胞的假设，这些干细胞类似于多能胚胎前体，即神经中胚层祖细胞（NMP）或神经嵴（NC）前体。在早期胚胎发育期间，NMP产生脊髓神经元以及骨、软骨和骨骼肌/平滑肌，而NC细胞产生外周神经元、黑素细胞和间充质细胞类型如平滑肌和软骨。我们推测NM肿瘤发生是在胚胎发生期间突变驱动的NMP/NC祖细胞转化为癌症起始细胞的结果，或者是“原位重编程”事件，其中神经/神经胶质细胞内遗传/表观遗传机制的破坏导致其转化为NMP/NC。研究计划目前建立脑肿瘤细胞的常规方法设计细胞系以选择神经干细胞（NSC）样细胞，这反映了所有脑肿瘤的起源细胞是转化的神经祖细胞的假设。因此，使用与用于神经干细胞增殖的条件相同的条件，已经成功地建立了贴壁神经胶质瘤干细胞系。然而，这种方法不能用于分离类似于NMP/NC细胞的双能NM细胞。我们最近确定了体外诱导人多能干细胞（hPSC）的NMPs和NC祖细胞的最佳培养条件。将分离成人脑肿瘤，并在NMP、NC和神经干细胞（NSC）条件（阳性对照）下平行培养相同数量的所得细胞。在NMP或NC培养基中的成功附着、增殖和随后的传代将标志着起始肿瘤中NM双能干细胞的存在。我们已经获得了有希望的初步数据显示，神经胶质瘤衍生的细胞可以在NMP诱导条件下增殖。将通过使用免疫荧光/qPCR和体外分化试验监测NMP和NC标志物的表达来确认所得细胞系中双能细胞的存在。将进行RNA-seq分析，比较NM肿瘤细胞系与hPSC衍生的NMP和NC细胞以及已建立的人NSC系的转录组，以确定致癌转化的候选驱动因素。还将通过异种移植/立体定向注射至NOD-SCID小鼠中评估致瘤潜力。如果时间允许，我们还将进行小分子筛选，旨在鉴定对NM肿瘤细胞系产生细胞毒性/细胞抑制作用的化合物。