Single-cell based elucidation of molecularly distinct GBM states and drug sensitivity

Glioblastoma heterogeneity and plasticity remain controversial, with proposed subtypes representing the average of highly heterogeneous admixtures of independent transcriptional states. Single-cell, protein-activity-based analysis identified four novel, molecularly distinct subtypes that successfully harmonize across multiple GBM datasets, including previously published bulk and single-cell profiles and single cell profiles from seven orthotopic PDX models, representative of prior subtype diversity. From these studies, GBM emerges as the plastic equilibrium of single tumor cells coexisting in two mutually-exclusive developmental states, with additional stratification provided by their proliferative potential. Consistent with these findings, all previously proposed subtypes could be recapitulated by mixtures of single cells representing these newly identified states. Most critically, drug sensitivity was predicted and experimentally confirmed as highly state-dependent, both in single-cell assays from patient-derived explants and in PDX models treated with clinically relevant drugs, suggesting that successful therapeutic strategies will likely require combinations of multiple drugs targeting these distinct tumor states. Overall design: Single-cell RNA-sequencing of control and drug-treated slice cultures of human glioma surgical specimens.

胶质母细胞瘤（Glioblastoma）的异质性与可塑性仍存在诸多争议，此前提出的各类亚型仅能代表独立转录状态高度异质性混合样本的平均特征。基于蛋白质活性的单细胞分析手段，本研究鉴定出四种全新的分子特征迥异的亚型，这些亚型可在多组胶质母细胞瘤数据集间实现良好的一致性整合，涵盖已发表的批量测序数据与单细胞转录图谱，以及来自7株原位患者来源异种移植（Patient-Derived Xenograft, PDX）模型的单细胞数据，充分覆盖了既往报道的亚型多样性。研究结果显示，胶质母细胞瘤呈现为单肿瘤细胞以两种互斥发育状态共存的可塑性平衡，而肿瘤细胞的增殖潜能可提供额外的分层依据。所有此前提出的亚型，均可通过代表新鉴定状态的单细胞混合体系得以复现，这与前述发现完全一致。最为关键的是，药物敏感性呈现高度的状态依赖性，这一点在患者来源肿瘤外植体的单细胞实验以及经临床相关药物处理的PDX模型中均得到了准确预测与实验验证，这提示有效的治疗策略大概率需要联合多种靶向不同肿瘤状态的药物。实验整体设计：对人类神经胶质瘤手术标本的对照及药物处理脑片培养物开展单细胞RNA测序。