摘要
目的:研究Btk抑制剂PCI-32765(依鲁替尼)和酪氨酸激酶Bcr-abl抑制剂Dasatinib(达沙替尼)对急性淋巴细胞白血病(ALL)细胞(Sup-B15、RS4;11)增殖、凋亡的影响,为其用于Ph+和Ph-ALL靶向治疗提供实验依据。方法:PCI-32765和Dasatinib单药及联合用药处理Sup-B15与RS4;11细胞后,用CCK-8法检测细胞的凋亡,刘氏染色法观察对细胞形态变化,Western blot法检测PCI-32765和Dasatinib对Btk上下游信号分子表达及活性的影响。结果:PCI-32765对RS4;11和Sup-B15细胞的增殖抑制作用呈剂量依赖性,Sup-B15细胞对其敏感性高于RS4;11,IC_(50)值分别为3和8μmol/L(P<0.05)。Dasatinib对RS4;11和Sup-B15细胞的增殖抑制作用也呈剂量依赖性,IC50值分别为5μmol/L和5 nmol/L,相差1 000倍(P<0.01)。Dasatinib与PCI-32765联用后,增殖抑制作用明显增强(P<0.05)。PCI-32765和Dasatinib分别作用RS4;11和Sup-B15细胞8、12、24、36、48和72 h,PCI-32765与Dasatinib单药组及联合用药组的细胞存活率均逐渐降低,且两药具有协同作用,呈时间依赖性。PCI-32765或/和Dasatinib处理的RS4;11和Sup-B15细胞体积缩小,细胞质密度增加,核固缩、核偏位、核碎裂,但小剂量Dasatinib单药对RS4;11细胞凋亡促进不明显,联合用药组凋亡细胞增多。PCI-32765或/和Dasatinib处理SupB15细胞BCR-ABL、Btk、Lyn、Src表达水平及活性均降低,与药物浓度呈正相关;RS4;11细胞Btk、Lyn、Src表达水平及活性均降低,与药物浓度呈正相关。结论:PCI-32765或Dasatinib均可明显抑制Sup-B15和RS4;11细胞增殖,诱导其凋亡,且有协同作用。其作用机制可能与通过活化B细胞受体(B cell receptor,BCR)信号通路、促进细胞凋亡有关。
Objective：To investigate the effects of Btk inhibitor（PCI-32765） and BCR-ABL tyrosine kinase inhibitor（Dasatinib） on proliferation and apoptosis of acute lymphoblastic leukemia（ALL） cell lines（Sup-B15,RS4;11） and the possible mechanism.Methods;RS4;11 and Sup-B15 cells were treated with PCI-32765 and Dasatinib,the cell proliferation and apoptosis were detected by CCK-8,the Btk and other apoptotic proteins were detected by Western blot.Results：PCI-32765 could inhibit the proliferation of RS4;11 and Sup-B15 cells in a dose-dependent manner,Sup-B15 cells were more sensitive to PCI-32765 than RS4;11 cells,their IC50 were 3 u,mol/L and 8 pjnol/L respectively,the difference between them was statistically significant（P〈 0.05）.Dasatinib also could inhibit the proliferation of RS4;11 cells and Sup-B15 cells in a dose-dependent manner.The IC50 was 5 ｜j,mol/L and 5 nmol/L,respectively,the difference between them was statistically very significant（P〈 0.01）,and the inhibitory effect was enhanced by the combination of Damatinib with the PCI-32765（P 〈0.05）.The cell survival rate decreased gradually in PCI-32765 or Dasatinib alone group and the combination group at the different time-point（8,12,24,36,48 and 72h）,the 2 drugs showed a synergistic effect on cells in a time-dependent manner.After being treated with PCI-32765 and Dasatinib,the RS4;11 and Sup-B15 cells showed that cell shrinkage,increase of cytoplasmic density,nuclear pyknosis,deviation and karyorrhexis,and increase of the apoptotic cells in the combination group,while the promotive effect of low dosage dasatinib on apoptosis of RS4;11 cells was not strong.PCI-32765 and Dasatinib could decrease the expression and activity of BCR-ABL,Btk,Lyn,Src in Sup-B15 and RS4;11 cells.Conclusion;PCI-32765 or Dasatinib can inhibit the proliferation and induce the apoptosis of Sup-B15 and RS4;11 cells,PCI-32765 and Dasatinib displayed the synergistic effects.The possible mechanism may be related with the blocking of B cell receptor（BCR）
出处
《中国实验血液学杂志》
CAS
CSCD
北大核心
2017年第1期72-79,共8页
Journal of Experimental Hematology
基金
国家自然科学基金资助项目(81170489)
江苏省自然科学基金资助项目(BK2010287,BK20141254)
江苏省卫生厅基金资助项目(H201056)
