SPHK1/S1P代谢通路在慢性粒细胞白血病发病及治疗中的作用及分子机制研究

Currently, metabolic remodeling has been a new trend and hot topic in cancer area, but the role of metabolic abnormality in the pathogenesis and treatment of leukemia is still unclear. In our previous study, we conducted a plasma metabolomic analysis of our conditional BCR-ABL transgenic mice at different progression stages of chronic myeloid leukemia (CML), showing that the concentration of sphingosine-1-phosphate (S1P) and the phosphorylation of sphingosine kinase-1 (SPHK1) which catalyzed sphingosine to S1P, increased significantly along with the expression of BCR-ABL. These results indicate that the SPHK1/S1P metabolic pathway is abnormally activated during the progression of CML. Based on this observation, this study aims to explore the metabolic changes related to the SPHK1/S1P pathway during malignant transformation of CML cells, as well as the feedback effect of metabolic abnormalities on the disease progression. Furthermore, we will also elucidate the impact of SPHK1/S1P pathway on CML leukemic stem cells by disturbing it and analyze the absolute number, the quiescent state and the cancer related metabolite concentration of CML leukemic stem cells after combining treatment with inhibitors of SPHK1/S1P pathway and tyrosine kinase. Over all, this study will provide a new perspective on targeted therapy of CML leukemia stem cells.

细胞代谢重塑是当前肿瘤领域的研究热点，但目前代谢通路异常在白血病发病及治疗机制中的作用依旧很不清楚。我们课题组前期以条件性BCR-ABL转基因小鼠为研究对象，对慢性粒细胞白血病（Chronic myeloid leukemia，CML）疾病进展不同时期的血浆样本进行了代谢组学分析，发现鞘氨醇-1-磷酸（S1P）浓度及催化其产生的鞘氨醇激酶1（SPHK1）的磷酸化水平随BCR-ABL的表达而显著上调，提示SPHK1/S1P代谢通路在CML发生发展中被激活。基于此基础，本项目将探索CML细胞恶性转化过程中SPHK1/S1P通路相关代谢层面的变化，以及代谢产物异常对CML疾病进展的反馈作用；并通过干扰SPHK1/S1P通路研究其对CML白血病干细胞的作用，分析此通路抑制剂与酪氨酸激酶抑制剂联用对CML白血病干细胞数量、静息状态及其癌性代谢物浓度的影响，为CML白血病干细胞的靶向治疗提供新的方向。

鞘氨醇激酶SPHK1是鞘氨醇代谢的关键酶，其编码的蛋白催化鞘氨醇的磷酸化形成鞘氨醇-1-磷酸（S1P），我们前期在对四环素诱导的慢性粒细胞白血病小鼠模型的代谢组学研究中发现，随着发病时间的延长，小鼠血浆中S1P的浓度显著增加，但其机制并不明确。本项目以慢性粒细胞白血病细胞（K562/32D-BCR-ABL1(32D-BA)）及小鼠模型为研究对象，利用细胞生物学、分子生物学、病毒介导的小鼠骨髓移植、细胞代谢组学等技术，对BCR-ABL1融合蛋白与SPHK1的相互调控作用关系、CML疾病发生发展中LSCs细胞内SPHK1/S1P代谢通路的变化、SPHK1/S1P代谢通路对CML细胞生物学行为及对CML疾病发生发展的影响等方面进行了深入研究。结果发现：1）BCR-ABL1的表达能显著提升SPHK1的磷酸化水平，导致其激酶活性的增加，从而导致细胞内外S1P的增加；此外，BCR-ABL1还能改变SPHK1在细胞内的定位，在32D细胞中过表达BCR-ABL后膜定位的SPHK1显著增加；更为有趣的是Sphk1敲低则能反馈抑制BCR-ABL融合蛋白的表达。2）SPHK1/S1P代谢通路抑制剂显著抑制32D-BA细胞的增殖、促进细胞凋亡并损伤细胞的克隆形成能力，该作用可能通过调控PI3K-AKT-mTOR途径实现。3）体内实验证实，无论是骨髓移植构建的小鼠模型还是药物诱导的小鼠模型，抑制SPHK1/S1P通路(敲低或通路抑制剂)均能显著延长CML小鼠的生存期。综上所述，SPHK1/S1P通路在慢粒疾病的发生发展中发挥着重要作用，抑制该通路能有效减缓疾病的进程，该通路有望成为治疗慢粒的新靶点。