Evaluation of a novel in vitro model of sonic hedgehog medulloblastoma by single cell transcriptomics

通过单细胞转录组学评估音刺猬髓母细胞瘤的新型体外模型

• 批准号: MR/V037730/1
• 负责人: Sanjeev Jacob
• 金额: $ 30.96万
• 依托单位: University of Oxford
• 依托单位国家: 英国
• 项目类别: Research Grant
• 财政年份: 2021
• 资助国家: 英国
• 起止时间: 2021 至 无数据
• 项目状态: 未结题
• 来源: https://gtr.ukri.org/projects?ref=MR%2FV037730%2F1
• 关键词: Evaluation; novel; vitro; model; sonic

CONTEXT OF THE RESEARCH: Brain cells that originate in the cerebellum give rise to the commonest childhood malignant brain tumour, termed medulloblastoma. Although cure rates of up to 70% have been reported, this is typically a devastating disease that is accompanied by considerable adverse effects resulting from multi-modal treatments including growth retardation, seizures and strokes. Like most other cancers, treatment advances crucially depend on tumour models that re-capitulate human disease, so that promising experimental treatments have a high chance of success in patients. Current medulloblastoma models are inadequate for several reasons. First, medulloblastoma cells from patients are hard to grow outside their normal environment, which prevents testing of experimental therapies. For the few tumour cell lines that have been grown ex vivo, they are deprived of their normal environment, which is an important factor in cancer progression as it influences the types of genes expressed by the tumour. Tumour gene expression in turn determines the response to treatments which explains why experimental treatments mostly fail in patients. Second, attempts to re-create the tumour environment using in vivo models of medulloblastoma, either based on genetic mouse models, or patient-derived tumour grafting into the mouse (termed PDX models) also have limitations. These models are costly, species differences limit the translation of therapies to the clinic, and in the case of PDX models, genetic divergence from the original tumour still occurs. Lastly, tumours differ genetically between patients and tumour heterogeneity is also evident within an individual. The latter discoveries have arisen from new technologies which can detect differences in the types of genes expressed at single cell resolution. Previous genetic assays of tumour cells in bulk were incapable of detecting these differences, which determine tumour growth patterns and influence treatment success. Consequently, treatment responses can be patchy across patients. Together, these shortcomings are responsible for the lack of new treatments for the last 30 years.We hypothesise that the presence of the tumour environment better simulates in vivo tumour growth conditions. To test this, we have previously coerced a type of human stem cell, called human induced pluripotent stem cells (hiPSCs), which can make any cell type in the body, to grow into miniature cerebellar structures, termed cerebellar organoids, in vitro. In a novel approach, we are currently growing human medulloblastoma cell lines within human cerebellar organoids to mimic the tumour environment. AIMS AND OBJECTIVES: I will investigate whether for one of the most prevalent genetic subtypes of medulloblastoma, termed sonic hedgehog medulloblastoma, our new model promotes tumour characteristics that better resemble human tumours. To this end, I will use the expression of genes by tumour cells as a proxy for tumour behaviour. By determining gene expression across the genome of thousands of medulloblastoma cells in our in vitro model individually, I can build a picture of their heterogeneity and determine how closely aligned our in vitro model is with its in vivo counterpart.POTENTIAL APPLICATIONS AND BENEFITS: Growing freshly obtained medulloblastoma cells on cerebellar organoids could overcome current medulloblastoma modelling difficulties and provide a new improved platform to test patient-specific therapies, including drug toxicity on normal cerebellar cells. Single cell analysis can be used to generate a cellular atlas of sonic hedgehog medulloblastoma diversity, which in turn can be used to identify patient-specific druggable molecular targets.

研究背景：起源于小脑的脑细胞会引起最常见的儿童恶性脑瘤，称为髓母细胞瘤。虽然据报道治愈率高达70%，但这是一种典型的破坏性疾病，伴随着多种治疗方法产生的相当大的不良反应，包括生长迟缓、癫痫发作和中风。像大多数其他癌症一样，治疗的进展关键依赖于使人类疾病重新屈服的肿瘤模型，因此有希望的实验性治疗在患者中有很高的成功机会。由于几个原因，目前的髓母细胞瘤模型不够充分。首先，患者的髓母细胞瘤细胞很难在正常环境外生长，这阻碍了对实验疗法的测试。对于少数在体外生长的肿瘤细胞株，它们被剥夺了正常环境，这是癌症进展的一个重要因素，因为它影响肿瘤表达的基因类型。肿瘤基因的表达反过来决定了对治疗的反应，这解释了为什么实验性治疗大多在患者身上失败。其次，使用髓母细胞瘤体内模型重建肿瘤环境的尝试也有局限性，无论是基于小鼠遗传模型，还是基于患者来源的肿瘤移植到小鼠体内(称为PDX模型)。这些模型成本高昂，物种差异限制了治疗方法在临床上的应用，而且在PDX模型的情况下，仍会发生与原始肿瘤的遗传分化。最后，不同患者的肿瘤基因不同，个体内肿瘤的异质性也很明显。后一项发现源于新技术，该技术可以检测在单细胞分辨率下表达的基因类型的差异。以前对大量肿瘤细胞的遗传分析无法检测到这些差异，这些差异决定了肿瘤的生长模式，并影响了治疗的成功。因此，患者的治疗反应可能是参差不齐的。这些缺点共同导致了过去30年来缺乏新的治疗方法。我们假设，肿瘤环境的存在更好地模拟了体内肿瘤的生长条件。为了测试这一点，我们之前强迫一种类型的人类干细胞，称为人诱导多能干细胞(HiPSCs)，它可以在体内制造任何类型的细胞，在体外生长成微型小脑结构，称为小脑器官。在一种新的方法中，我们目前正在人小脑器官中培养人髓母细胞瘤细胞系，以模拟肿瘤环境。目的和目的：我将调查一种最常见的髓母细胞瘤的遗传亚型，称为声波刺猬髓母细胞瘤，我们的新模型是否促进了与人类肿瘤更相似的肿瘤特征。为此，我将使用肿瘤细胞的基因表达作为肿瘤行为的代理。在我们的体外模型中，通过单独测定数千个髓母细胞瘤细胞的基因组基因表达，我可以建立它们的异质性图景，并确定我们的体外模型与体内模型的一致性有多紧密。潜在的应用和好处：在小脑器官上培养新鲜获得的髓母细胞瘤细胞可以克服目前髓母细胞瘤建模的困难，并提供一个新的改进平台来测试患者特定的治疗方法，包括药物对正常小脑细胞的毒性。单细胞分析可用于生成声波刺猬髓母细胞瘤多样性的细胞图谱，进而可用于确定患者特定的可用药分子靶点。