IKZF1IK6亚型致急性淋巴细胞白血病耐药机制研究

Study on the Mechanism of Drug Resistance Induced by IK6 Isoform of IKZF1 in Acute Lymphoblastic Leukemia

目的:研究BIM基因在IK6亚型(IK6)及非IK6亚型(IK6)急性淋巴细胞白血病细胞耐化疗药物中的调控机制。方法:利用巢式PCR鉴定急性淋巴细胞白血病细胞Nalm6和RS4;11的IKZF1亚型;通过Alamarblue法、流式细胞术等检测适当浓度的柔红霉素下对两种细胞的活性和凋亡的影响;利用甲基化检测和实时荧光定量PCR对促凋亡基因BIM的甲基化程度和表达水平进行检测。结果:柔紅霉素对IK6^(-)Nalm6和IK6^(+)RS4;11细胞增殖均有浓度依赖的抑制作用,且对二者均有显著的致凋亡作用,相较于Nalm6,RS4;11对柔红霉素显示出更高的耐药性。柔红霉素处理后,Nalm6中的BIM甲基化水平显著性降低,BIMmRNA表达水平显著升高(P<0.05),而二者在RS4;11中未见显著性改变(P>0.05)。结论:IK6的存在可能会诱导BIM基因甲基化水平增加以抵御柔红霉素引起的BIM甲基化水平的降低从而使急性淋巴细胞白血病细胞对柔红霉素耐药。

Objective:To study the regulatory mechanism of BIM in chemotherapeutic drug resistance of IK6 isofbrm(IK6^(+))and non-IK6 isoform(IK6^(-))acute lymphoblastic leukemia cells.Methods:The IKZF1 isoform ofNalm6 and RS4;11 were identified by nested PCR,and the efiects of daunorubicin on the cell viability and apoptosis of the two kinds of cells were detected by Alamarblue and flow cytometry.Methylation detection and quantitative real-time PCR(qRT-PCR)were used to detect the methylation degree and mRNA expression level of pro-apoptotic gene BIM,respectively.Results:Daunorubicin reduced the proliferation ofIK6^(-)Nalm6 and IK6^(+)RS4;11 in a concentration-dependent manner,and significantly induced apoptosis.Compared with Nalm6,RS4;11 showed higher resistance to daunorubicin.Treatment with daunorubicin resulted in a significant decrease in BIM methylation rate and a significant increase in BIM mRNA expression in nalm6(P<0.05),whereas neither was significantly altered in RS4;11(P>0.05).Conclusion:IK6 may induce an increase in the methylation level of BIM to resist the decrease of BIM methylation level caused by daunorubicin,which makes acute lymphoblastic leukemia cells resistant to daunorubicin.