Single-cell RNA-seq of human bone marrow cells by TAS-Seq

The TCF3::HLF fusion protein defines a subtype of incurable B cell acute lymphoblastic leukemia (B-ALL). Here, we identified self-reinforcing IL-1b networks in TCF3::HLF B-ALL cells using a newly-established mouse model that fully recapitulates human TCF3::HLF B-ALL, including osteolysis. We found significant upregulation of several inflammatory cytokines, including IL1B, IL6 and IFNG in the B-ALL mice. Deletion of IL1B or IL1R1 strongly inhibited the growth of the human TCF3::HLF B-ALL cells, reduced the expression of RANKL and ameliorated the destruction of bone in the transplanted mice. Genetic and epigenetic analyses identified a previously unknown regulatory region of the IL1B gene locus, where TCF3::HLF directly binds. Importantly, single cell RNA-seq profiling of TCF3::HLF B-ALL patients revealed a dramatic upregulation of IL1B at relapse compared to the time of diagnosis. These findings suggest a critical role of TCF3::HLF-IL-1b axis for the progression of TCF3::HLF B-ALL. Overall design: Cryopreserved bone marrow samples were obtained from two TCF3::HLF-positive B-ALL patients at diagnosis and at relapse. Healthy donor bone marrow mononuclear cells (BM MNC) were used as controls.

TCF3::HLF融合蛋白定义了一类无法治愈的B细胞急性淋巴细胞白血病（B-cell acute lymphoblastic leukemia, B-ALL）亚型。本研究利用全新构建的可完全复现人类TCF3::HLF阳性B-ALL病理表型（包括骨质溶解）的小鼠模型，在TCF3::HLF B-ALL细胞中鉴定出自我强化的IL-1β信号网络。我们在该B-ALL模型小鼠体内检测到包括IL1B、IL6及IFNG在内的多种炎性细胞因子表达显著上调。敲除IL1B或IL1R1可显著抑制人源TCF3::HLF B-ALL细胞的增殖，降低RANKL的表达水平，并缓解移植瘤模型小鼠的骨质破坏。遗传与表观遗传分析揭示了IL1B基因座上一处此前未被报道的调控区域，TCF3::HLF可直接结合该区域。值得注意的是，对TCF3::HLF阳性B-ALL患者的单细胞RNA测序（single cell RNA-seq, scRNA-seq）分析显示，相较于确诊阶段，患者复发时的IL1B表达量出现显著上调。上述研究结果表明，TCF3::HLF-IL-1β轴在TCF3::HLF阳性B-ALL的疾病进展中发挥关键调控作用。总体实验设计：本研究收集了2例TCF3::HLF阳性B-ALL患者在确诊时与复发时的冻存骨髓样本，并以健康供者的骨髓单个核细胞（bone marrow mononuclear cells, BM MNC）作为对照样本。