EZH2基因在白血病细胞耐药中的作用及机制的研究

Drug resistence of leukemic cells is the major obstacle which can influence the long survival of patients with leukemia. The mechansim of drug-resistence is very compliacted and lack effective resolution so far. Leukemic stem cells play an important role in the occurrence and development of leukemia,which is regard as the origin of drug-resistence and relapse. Some studies have presented that EZH2 can influence the activity of leukemic stem cells through trimethylation of histone H3 on Lys27 (H3K27me3).We and other studies have show that EZH2 overexpression is found in acute leukemia patients with complex karyotypes or with infiltrating outside bone marrow. These studies hint that EZH2 probably take part in drug resistence, relapse and metastasis of leukemia.So in the study, we want to observe the proliferation ability,apotosis and drug sensitivity of leukemic cells by changing the expression of EZH2.We also want to study the mechansim deeply with emphasis on the changes of NF-κB singal way. By in vivo model of AML,we also want to study the effect of downregulationg of EZH2 combination of chemotherapy drug to the survival as well as microresidual disease of leukemic mices.Through the study, We want to supply some experimental witness for if epigenetic therapy in combination of chemotherapy can probably overcome drug resistecne of leukemia.

白血病细胞耐药是影响白血病患者长期生存的主要障碍，发生机制复杂，目前尚无有效的解决方案。白血病干细胞在启动白血病发生和发展中起到关键性作用被认为是白血病耐药和复发的根源。有研究显示EZH2通过调控组蛋白H3K27甲基化特异性影响白血病干细胞的活性。我们及其他研究发现伴复杂核型异常及伴有髓外浸润的白血病患者表达高水平EZH2，提示EZH2有可能在白血病耐药、复发、转移中起重要作用。为此，本研究拟通过改变白血病耐药细胞株及原代白血病干细胞EZH2表达观察白血病细胞增殖、凋亡及药物敏感性影响，并探讨其机制，重点关注NF-κB信号通路的变化。并通过建立白血病小鼠动物模型进一步观察下调EZH2联合化疗药物对白血病小鼠生存、微小残留病变等影响。希望通过本研究为表观遗传学调控药物联合化疗克服白血病耐药提供实验依据。

EZH2和恶性肿瘤的发展密切相关，其高表达参与恶性肿瘤的发生、发展。该基因作为PRC2复合物中的核心成员，通过特异性将组蛋白H3上第27位赖氨酸残基（H3-K27）进行甲基化修饰，沉默相关靶基因的转录而发挥功能。有研究显示，EZH2在伴有髓外浸润的急性髓系白血病患者中表达较无髓外浸润的急性髓系白血病患者显著增高，提示EZH2有可能在白血病细胞侵袭中发挥重要作用。本研究旨在探讨下调EZH2表达对急性髓系白血病细胞侵袭的影响，并进一步研究其机制。研究结果显示EZH2 mRNA及蛋白表达水平在急性髓系白血病细胞株及原代细胞中均明显增高。EZH2抑制剂DZNeP作用于HL-60细胞使EZH2蛋白表达明显下调，HL-60细胞侵袭能力下降，明显低于阴性对照组；MMP-2和MMP-9基因的mRNA及蛋白表达水平明显下调。siRNA特异性干扰EZH2后HL-60细胞侵袭能力下降，明显低于未转染组及阴性对照转染组，同时MMP-2、MMP-9的mRNA水平及蛋白表达水平也明显下降。shRNA沉默 Kasumi-1细胞EZH2表达后细胞侵袭能力下降，明显低于阴性对照转染组，同时MMP-2、MMP-9的mRNA水平及蛋白表达水平也明显下降。本研究提示EZH2有可能通过调节MMP-2、MMP-9的表达参与白血病细胞的侵袭及转移，另外EZH2是否能做为判断AML患者预后的影响因素值得进一步探究。