Objectives: To investigate the effect of adriamycin (ADM), idelalisib or ADM and their combination on cell proliferation and intracellular concentration of ADM, and to explore the reversal effect of idelalisib on drug resistance to ADM.
Methods: The K562 and K562/ADM cells were respectively treated with ADM and idelalisib at different concentrations. The 50% inhibitory concentration (IC50) and drug resistance index (RI) of ADM to the 2 kinds of cells were measured by methyl thiazolyl tetrazolium (MTT) assay. Non-cytotoxic dose (cell inhibition rate <5%) of idelalisib in the 2 kinds of cells was determined. Then the K562 and k562/ADM cells were divided into the following groups: a K562 cells + ADM group, a K562 cells + ADM + idelalisib group, a K562/ADM cells + ADM group, and a K562/ADM cells + ADM + idelalisib group. The survival rates, the intracellular ATP levels, and the relative concentration of intracellular ADM were detected by MTT method, ATP bioluminescence assay (ATP-BLA) and flow cytometry (FCM), respectively.
Results: The cell survival rates were significantly decreased in a dose-dependent manner when the cells were treated with different doses of ADM (0.001-10.000 mg/L ). The IC50 value of ADM in the K562 and K562/ADM cells were 0.2 mg/L and 1.0 mg/L, respectively. The RI value was 5. The cell survival rates were also significantly decreased in a dose-dependent manner when the cells were treated with different doses of idelalisib (1-50 μmol/L). The non-cytotoxic dose of idelalisib in the K562 and K562/ADM cells were 25 μmol/L and 15 μmol/L, respectively. The cell survival rates in the ADM+ idelalisib group was less than that in the ADM group (P<0.05)；while there was no statistical difference between the ADM+ idelalisib group and the ADM group in the K562 cells (P>0.05). The intracellular ATP level in the K562 cells was about (91.502±0.479) mmol/L, and that in the K562/ADM cells was about (24.311±0.349) mmol/L. The intracellular ATP level in the ADM+ idelalisib group in the K562/ADM cells was less than that in the ADM group (P<0.05); but there was no statistical difference between the ADM + idelalisib group and the ADM group in the K562 cells (P>0.05). The absorption of intracellular ADM in the ADM + idelalisib group in the K562/ADM cells was more than that in the ADM group (P<0.05); but there was no statistical difference in the K562 cells between the 2 groups (P>0.05). The exclusion of intracellular ADM in the ADM + idelalisib group in the K562/ADM cells was less than that in the ADM group (P<0.05 or P<0.01)；but there was no statistical difference in the K562 cells between the 2 groups (P>0.05).
Conclusions: Idelalisib exerts effect on inhibition of the proliferation in myeloid leukemia K562 and K562/ADM cells, which may partially reverse the drug resistance of K562/ADM cells to ADM. The mechanisms for the effect of idelalisib may be related to increasing the accumulation of ADM and inducing the cell apoptosis in the K562 and K562/ADM cells.
目的: 观察阿霉素(adriamycin，ADM)单药、艾德拉尼(idelalisib)单药以及ADM联合idelalisib对人髓系白血病细胞株K562和K562/ADM的增殖以及细胞内ADM相对浓度的影响，探讨idelalisib对K562和K562/ADM两种细胞株ADM耐药的逆转作用及其机制。方法: 以K562及K562/ADM细胞株为观察对象，分别以不同浓度ADM及idelalisib进行干预，计算ADM对两种细胞株的半抑制浓度(50% inhibitory concentration，IC50)及耐药指数(resistance index，RI)；计算idelalisib对于两种细胞株的无细胞毒性作用浓度(细胞抑制率<5%)。后续将K562和K562/ADM细胞株分别分为ADM组和ADM+idelalisib组。分别用MTT法、生物发光法和流式细胞术检测各组细胞存活率、细胞株内ATP和ADM水平的变化。结果: 随着ADM浓度的增加，K562及K562/ADM细胞存活率相应降低，呈剂量依赖性；ADM在K562与K562/ADM细胞株中的IC50分别为0.2 mg/L和1.0 mg/L，RI为5。随着idelalisib浓度的增加，K562及K562/ADM细胞存活率也相应降低，呈剂量依赖性；idelalisib对K562和K562/ADM细胞株的无细胞毒性体积浓度(抑制率<5%)分别约为25 μmol/L和15 μmol/L。MTT法结果显示：在K562/ADM细胞株中，ADM+idelalisib组的细胞存活率低于ADM组，差异有统计学意义(P<0.05)；在K562细胞株中，ADM+idelalisib组的细胞存活率与ADM组相比差异无统计学意义(P>0.05)。生物发光法结果显示：K562细胞株内ATP水平为(91.502±0.479) mmol/L，K562/ADM细胞株内ATP水平为(24.311±0.349) mmol/L。在K562/ADM细胞株中，ADM+idelalisib组ATP水平低于ADM组，差异有统计学意义(P<0.05)；在K562细胞株中，ADM+idelalisib组ATP水平与ADM组相比差异无统计学意义(P>0.05)。流式细胞术检测结果显示：在K562/ADM细胞株中，ADM+idelalisib组ADM吸收率高于ADM组，差异有统计学意义(P<0.05)；在K562细胞株中，ADM+idelalisib组ADM吸收率与ADM组相比差异无统计学意义(P>0.05)。在K562/ADM细胞株中，ADM+idelalisib组ADM排除率低于ADM组，差异有统计学意义(P<0.05或P<0.01)；在K562细胞株中，ADM+idelalisib组ADM排除率与ADM组相比差异无统计学意义(P>0.05)。结论: Idelalisib对体外髓系白血病细胞株K562及K562/ADM均有细胞毒性作用，并能部分逆转K562/ADM细胞株对ADM的耐药。其逆转机制可能是通过增加细胞内ADM蓄积和诱导细胞凋亡而实现的。.
Keywords: K562 cells; K562/ADM cells; idelalisib; leukemia; phosphatidylinositol 3-kinase/protein kinase B signal pathway; reverse drug resistance; tumor multiple drug resistance.