骨髓微环境通过Tim-3/HMGB1/beta-catenin通路支持CML白血病干细胞的机制与干预

Beta-catenin is required for self-renewal maintenance of leukemia stem cells (LSC) in chronic myeloid leukemia (CML), which is involved in leukemogenesis, tyrosine kinase inhibitor (TKI) resistance and disease progression. Bone marrow niche induces beta-catenin activation in CML, but the underlying mechanism is not well understood. We previously revealed that inhibition of beta-catenin restored sensitivity of TKI-resistant CML cells to nilotinib, impaired engraftment of LSC and killed LSC in vitro and in vivo; furthermore, Tim-3 defines a novel Tim-3high GMP LSC, with highest beta-catenin expression among CML stem/progenitor subsets. Tim-3 ligand HMGB1 was secreted from MSCs and macrophage in bone marrow niche. Based on the aforementioned researches, we hypothesize that Tim-3/HMGB1 pathway mediates the crosstalk between bone marrow niche and LSC, activates beta-catenin and supports LSC self-renewal. The study focus on the crosstalk between Tim-3 in LSC and HMGB1 secretion of bone marrow niche and the subsequent effect on b-catenin activation, nuclear translocation and regulation on downstream genes of beta-catenin, aiming to dissect novel mechanism of interaction between bone marrow niche and LSC and figuring out new targets for potential intervention of the interaction.

研究发现，b-catenin是CML白血病干细胞(LSC)维持自我更新所需的关键信号，参与CML发生、耐药和急变；骨髓微环境可激活LSC b-catenin表达，但机制尚不清楚。我们前期研究发现：靶向抑制b-catenin，可逆转TKI耐药、破坏LSC植入、杀伤LSC；Tim-3是CML LSC的新标记，其高表达与b-catenin激活相关；Tim-3的配体HMGB1，在骨髓微环境MSC和巨噬细胞中广泛表达。综上，我们推测：骨髓微环境细胞分泌HMGB1，与LSC表面Tim-3结合，形成Tim-3/HMGB1通路，激活b-catenin，支持LSC自我更新。本项目拟通过体内外实验，研究骨髓微环境与LSC之间的Tim-3/HMGB1通路互话对LSC b-catenin激活/核内定位/下游基因和植入功能的影响，揭示骨髓微环境支持LSC自我更新的新机制，探索干预LSC与骨髓微环境相互作用的新靶点。

白血病干细胞参与白血病发生、耐药、复发和克隆演化等重要生物学行为，骨髓微环境与白血病干细胞的互话是支持白血病干细胞自我更新的关键，探索骨髓微环境支持白血病干细胞的机制和靶向白血病干细胞的新靶点和新策略，具有重要的基础和转化意义。本项目主要针对：(1)骨髓微环境通过HMGB1/Tim-3信号通路支持白血病干细胞；（2）组蛋白去乙酰化酶抑制剂HDACi靶向白血病干细胞的新策略；（3）Tim-3作为白血病干细胞标记及免疫检测点在白血病中的预后意义进行了研究。本研究初步发现：HMGB1在髓系白血病中普遍高表达，HMGB1以胞外形式可诱导细胞中b-catenin表达和核内定位，从而支持Tim-3白血病干细胞的自我更新，提示HMGB1可介导骨髓微环境信号，通过激活胞内b-catenin信号通路，支持LSC的自我更新；我们采用HDACi处理突变/耐药的白血病细胞和干/祖细胞发现， HDACi西达本胺与酪氨酸激酶抑制剂(TKI)存在协同作用，二者可通过抑制Bcr-Abl和b-catenin信号通路，协同可逆转TKI耐药并靶向CML干/祖细胞，同时，HDACi通过SIRT1/FOXO3a抑制难治/复发AML LSC；我们通过队列研究发现，Tim-3在白血病干细胞中的表达与白血病生存显著相关，多因素分析显示，Tim-3表达是独立预后因素；同时，Tim-3作为T细胞免疫检测点和衰老耗竭表型，同样与白血病预后密切相关。本项目为解析骨髓微环境支持白血病干细胞的关键机制、探索靶向白血病干细胞的新策略和新靶点提供了新的线索和启发；对于HDACi靶向白血病干细胞、以Tim-3为靶点靶向免疫检测点和白血病干细胞，具有重要的基础和临床转化应用意义。