三氧化二砷抑制耐药bcr-abl突变细胞株生长的机制  

Growth inhibition of arsenic trioxide in bcr-abl mutant cells and the potential mechanism

作　　者：刘金花[1] 王晓丹[2] 赵雪飞[2] 唐庆华[2] 展昭民[2] 马军[2] 李德山[1] 邱林[2] 

机构地区：[1]东北农业大学生命科学学院生物制药教研室,哈尔滨150030 [2]哈尔滨血液病肿瘤研究所

出　　处：《白血病．淋巴瘤》2010年第7期421-424,共4页Journal of Leukemia & Lymphoma

基　　金：黑龙江省自然科学基金面上资助（ZA2006-12）;哈尔滨市科技创新项目（2006RFLQS019）

摘　　要：目的 研究三氧化二砷（ATO）在体外对bcr-abl突变细胞株的生长抑制作用,并探讨其作用机制.方法 采用锥虫蓝拒染法观察ATO及ATO与丁硫氨酸亚矾胺（BFO）共同作用对bcr-abl野生型细胞株（K562、KBM5、32Dp210）和伊马替尼（IM）耐药bcr-abl突变细胞株（K562R、KBM5R、32Dp210T315I、32Dp210Q252H、32Dp210Y253H、32Dp210M351T和32Dp210E255K）的生长抑制作用;Annexin V和PI染色检测细胞凋亡,以5,5＇-双硫代对硝基苯甲酸直接比色法（DTNB比色法）测定细胞内谷胱甘肽含量（GSH）.结果 ATO对bcr-abl野生型和IM耐药细胞株均呈现剂量依赖性的生长抑制作用.且含bcr-abl点突变的耐药细胞株对ATO格外敏感.其ATO的IC50值均比其对应的bcr-abl野生型细胞株低,但无点突变的K562R细胞株的IG50值与野生型K562细胞无差异.对上述细胞株细胞内还原型GSH的含量进行检测,结果显示：含有ber-abl点突变的细胞株内GSH含量显著低于其对应的野生型细胞（均P〈0.05）.而K562和K562R细胞中GSH含量差异无统计学意义（P=0.315）.选用GSH生物合成抑制剂BSO处理各细胞株后.细胞对ATO的敏感性均显著增加.结论 与ber-abi野生型细胞株相比ATO对含有点突变的细胞株生长抑制作用更为显著,这可能与细胞内GSH的含量有关,提示ATO有可能成为治疗包括T315I在内的ber-abl突变慢性粒细胞白血病患者的新选择.Objective To investigate the effect of arsenic trioxide （ATO） on the growth inhibition of bcr-abl mutant cell lines in vitro and to explore its potential mechanism. Methods The growth inhibition of ATO on bcr-abl wild type cell lines （K562, KBM5 and 32Dp210） and imatinib（IM）-resistant cell lines （K562R, KBM5R, 32Dp210T315I, 32Dp210Q252H, 32Dp210Y253H, 32Dp210M351T and 32Dp210E255K） were measured by trypan blue exclusion. Apoptosis was assayed by AnnexinV and PI staining. Glutathione （CSH） levels were detected by DTNB colorimetry of Glutathione Assay Kit. Results ATO inhibited cell growth in both bcr-abl wild type and IM-resistant mutant type cells in a dose dependent manner. ATO significantly inhibited growth of bcr-abl point mutant cells compared with the corresponding wild type cells, and the IC50 of ATO in mutant cells was lower than that in wild type, while the IC50 in no point mutant cells K562R was not different compared with that in wild type cells K562. The GSH levels in bcr-abl point mutant cells were lower than that in the corresponding wild type cells（P =0.00106-0.0358） , but that in K562 was quite similar with K562R cells（P = 0.315）. After depletion of intracellular GSH by using BSO, the growth inhibition of ATO in both bcr-abl point mutant cells and wild type cells was significantly enhanced. Conclusion The growth inhibition of ATO on bcr-abl point mutant cells is remarkably more effective than that on wild type cells, which may be related with intracellular GSH. ATO would be a potential therapeutic select against CML with bcr-abl point mutation including the T315I mutation.

关 键 词：砷剂 生长抑制物 突变 融合蛋白质类 bcr-abl 谷胱甘肽 

分 类 号：R733.7[医药卫生—肿瘤]