PDF(Pubmed)
Abstract：
The most common malignant brain tumour in children, medulloblastoma (MB), is subdivided into four clinically relevant molecular subgroups, although targeted therapy options informed by understanding of different cellular features are lacking. Here, by comparing the most aggressive subgroup (Group 3) with the intermediate (SHH) subgroup, we identify crucial differences in tumour heterogeneity, including unique metabolism-driven subpopulations in Group 3 and matrix-producing subpopulations in SHH. To analyse tumour heterogeneity, we profiled individual tumour nodules at the cellular level in 3D MB hydrogel models, which recapitulate subgroup specific phenotypes, by single cell RNA sequencing (scRNAseq) and 3D OrbiTrap Secondary Ion Mass Spectrometry (3D OrbiSIMS) imaging. In addition to identifying known metabolites characteristic of MB, we observed intra- and internodular heterogeneity and identified subgroup-specific tumour subpopulations. We showed that extracellular matrix factors and adhesion pathways defined unique SHH subpopulations, and made up a distinct shell-like structure of sulphur-containing species, comprising a combination of small leucine-rich proteoglycans (SLRPs) including the collagen organiser lumican. In contrast, the Group 3 tumour model was characterized by multiple subpopulations with greatly enhanced oxidative phosphorylation and tricarboxylic acid (TCA) cycle activity. Extensive TCA cycle metabolite measurements revealed very high levels of succinate and fumarate with malate levels almost undetectable particularly in Group 3 tumour models. In patients, high fumarate levels (NMR spectroscopy) alongside activated stress response pathways and high Nuclear Factor Erythroid 2-Related Factor 2 (NRF2; gene expression analyses) were associated with poorer survival. Based on these findings we predicted and confirmed that NRF2 inhibition increased sensitivity to vincristine in a long-term 3D drug treatment assay of Group 3 MB. Thus, by combining scRNAseq and 3D OrbiSIMS in a relevant model system we were able to define MB subgroup heterogeneity at the single cell level and elucidate new druggable biomarkers for aggressive Group 3 and low-risk SHH MB.
摘要：
儿童最常见的恶性脑瘤，髓母细胞瘤（MB），被细分为四个临床相关的分子亚群，尽管缺乏通过了解不同细胞特征而获得的靶向治疗方案。这里,通过比较最具侵略性的亚组（第3组）与中间（SHH）亚组，我们确定了肿瘤异质性的关键差异，包括第3组中独特的代谢驱动亚群和SHH中产生基质的亚群。为了分析肿瘤异质性，我们在3DMB水凝胶模型中在细胞水平上描绘了单个肿瘤结节，概括了亚组特异性表型，通过单细胞RNA测序(scRNAseq)和3DOrbiTrap二次离子质谱(3DOrbiSIMS)成像。除了鉴定已知的MB代谢物特征，我们观察到结节内和结节间异质性,并确定了亚组特异性肿瘤亚群.我们表明，细胞外基质因子和粘附途径定义了独特的SHH亚群，并由含硫物种组成独特的壳状结构，包含富含亮氨酸的小蛋白聚糖（SLRP）的组合，其中包括胶原蛋白组织者lumican。相比之下,第3组肿瘤模型的特征在于多个亚群具有显著增强的氧化磷酸化和三羧酸(TCA)循环活性。广泛的TCA循环代谢物测量揭示了非常高水平的琥珀酸盐和富马酸盐,尤其是在第3组肿瘤模型中几乎检测不到苹果酸盐水平。在患者中，高富马酸盐水平(NMR光谱)以及激活的应激反应途径和高核因子红系2相关因子2(NRF2;基因表达分析)与较差的生存率相关.基于这些发现,我们预测并证实NRF2抑制在组3MB的长期3D药物治疗测定中增加了对长春新碱的敏感性。因此，通过在相关模型系统中结合scRNAseq和3DOrbiSIMS,我们能够在单细胞水平上定义MB亚组异质性,并阐明侵袭性3组和低风险SHHMB的新的可作为药物的生物标志物.
