细胞微泡诱导前白血病干细胞产生及其向白血病完全转化的研究

Recent reports of pre-leukemia stem cells (pre-LSCs) can provide fresh insight into the onset and relapse of acute leukemia(AL).Indificiency of models to observe complete transformation into leukemia, the distinct nature of pre-LSCs and how they transform to leukemia cells are remained to determined. We have, for the first time, successfully established a system in which microvesicles (MVs) derived from leukemia cells induce normal bone marrow hematopoietic cells to malignant transformation，leading to a full outbreak of leukemia within 20 days. And the induced leukemia cells can be passaged in vivo and vitro. This novel and specific system probably provide opportunities to observe and understand how multi-steps of clonal evolution, from normal hematopoietic stem progenitor cells(HSPC), to pre-LSC, to LSC and eventually to leukemic cells. Using this system, we will firstly determine the characteristics of pre-LSC using high-throughput parallel sequencing and bioinformatics analysis, then the key factors and pathway involved in the transformation of pre-LSCs to leukemia cells will be sorted out from the complex system that control the transformation of pre-LSCs to leukemia cells, which is next to be evaluated by lab experiments and clinical investigations. These works will contribute to clearly characterize the distinct nature of pre-LSCs and provide a key system for pre-LSC-related works and exploration of the underlying mechanisms of multi-steps pathogenesis of AL. Finally, the characterstics of pre-LSCs and sorted key node of the system will serve as markers and targets for the occurrence and recurrence of leukemia.

新近前白血病干细胞（pre-LSC）的研究，可能为急性白血病早期发生和复发提供全新机制。但由于缺乏能观察其直接向白血病完全转化的理想模型，pre-LSC的实质性特征及如何实现完全白血病转化的相关机制目前均难以明确。我们前期首次报道白血病微泡体外诱导正常造血细胞恶性转化为白血病细胞并可在体内外传代，极可能是能够展现造血干/祖细胞经pre-LSC向LSC及白血病细胞动态演进的创新体系。本项目拟采用高通量测序与我们前期建立的先进生物信息学方法，结合实验与临床验证，运用该体系来明确pre-LSC生物学特征，遴选并锁定调控其实现完全白血病转化的网络机制中的核心基因突变和异常信号通路。本项目有望为获取和研究pre-LSC、探索白血病多步骤发病机制提供创新性实验体系，为白血病的早期发生和复发提供预警标志物和精确干预靶标。

前白血病干细胞（pre-LSC）是急性白血病多步骤发病过程的起点，也是复发的关键。但目前缺乏能够直接动态观察pre-LSC向白血病完全转化的理想模型，其生物学特征和完全转化导致白血病发生的机制均不明确。我们前期发现白血病微泡（K562-MVs）能够诱导正常造血细胞恶性转化为白血病细胞，提示转化体系极可能是动态展现造血细胞经pre-LSC向白血病细胞转化的体外模型。因此，我们深入研究了K562-MVs的生物学特征并对诱导体系进行了优化，确认靶细胞为CD34+造血干细胞；对比K562细胞，对转化后白血病的细胞学特征、基因组学和蛋白质组学进行了鉴定，证实转化后的白血病细胞为全新的BCR/ABL细胞；通过对不同转化阶段造血细胞的基因组、蛋白质磷酸化检测和生物学分析，发现转化过程中促癌miRNAs的富集和PKC活性下降；干预K562-MVs内富集程度最高的miR-146b-5p后可以改变转化进程，证实促癌miRNAs的富集与K562-MVs诱导恶性转化密切相关；以MSC-MVs作为K562-MVs对照，发现MSC-MVs具有促脐血扩增的潜在应用前景，并能反映MSC的衰老状态，是鉴定MSC衰老的潜在标志物。本项目建立一种动态展示造血细胞经pre-LSC向白血病完全转化的细胞体系，构建了一株新的BCR/ABL阳性白血病细胞并找到恶性转化的可能机制，为研究pre-LSC和探索白血病多步骤发病机制提供了模型；发现了MSC-MVs促脐血扩增的潜在应用前景，为鉴定MSC衰老提供了新的潜在标志物。