Ibrutinib联合三氧化二砷对套细胞淋巴瘤细胞系的作用

目的:探讨Ibrutinib联合三氧化二砷（arsenic trioxide，ATO）对套细胞淋巴瘤细胞系的影响，初步研究其可能机制。方法:应用CCK-8法检测Ibrutinib联合ATO对套细胞淋巴瘤细胞系增殖的影响；流式细胞术检测Ibrutinib、ATO单药及两药联合对套细胞淋巴瘤细胞系凋亡、周期的影响；Western Blot检测周期及凋亡相关蛋白表达水平变化。结果:CCK-8结果显示两药联合对套细胞淋巴瘤细胞系增殖的抑制作用较单药更明显，两药之间具有协同作用，最佳协同浓度为Ibrutinib 5 μmol/L和ATO 0.75 μmol/L（P<0.001)；Annexin V-PE/7AAD双染流式细胞术分析结果表明未加药对照组、5 μmol/L Ibrutinib组、0.75 μmol/L ATO组、联合用药组24 h细胞凋亡率为（3.0±0.81）%、（3.5±0.91）%、（4.9±0.04）%和（20.4±0.93）%（P<0.000 1），48 h细胞凋亡率为（2.5±0.75）%、（3.6±1.82）%、（26.2±2.76）%和（76.4±9.33）%(P=0.003)；流式细胞术检测显示5 μmol/L Ibrutinib、0.75 μmol/L ATO单药及两药联合能将套细胞淋巴瘤细胞周期阻滞于S期；5 μmol/L Ibrutinib、0.75 μmol/L ATO单药及两药联合使Caspase-3、PARP、CyclinD1、Bcl-2表达水平下调，而Cleaved caspase-3、Cleaved PARP表达水平上调，且两药联合较单药更明显。结论:Ibrutinib、ATO单药及两药联合能有效抑制套细胞淋巴瘤细胞系增殖、诱导其凋亡和细胞周期阻滞，且在一定浓度范围内两药联合有协同作用。

The synergistic effect of Ibrutinib combined with arsenic trioxide on mantle cell lymphoma cell line

Objective:To investigate the synergistic effects of Ibrutinib combined with arsenic trioxide(ATO）on mantle cell lymphoma cell line and its potential mechanisms.Methods:The CCK-8 assay was used to measure the growth inhibition of cells after Ibrutinib and/or ATO.The flow cytometry was employed to detect the effect on the apoptosis and cell cycle after drug treatment.Western Blot was used to determine the changes in apoptosis and cycle-related proteins.Results:The results of CCK-8 assay showed that the combination of Ibrutinib and ATO had a synergistic effect on growth in cells and the optimal synergy concentration was Ibrutinib 5 μmol/L and ATO 0.75 μmol/L.Flow cytometry analysis confirmed that the apoptosis rate at 24 h was in the untreated group （3.0±0.81）%，Ibrutinib 5 μmol/L （3.5±0.91）%，ATO 0.75 μmol/L （4.9±0.04）% and the combination group （20.4±0.93）%(P<0.000 1)，and apoptosis rate at 48 h was in the untreated group （2.5±0.75）%，Ibrutinib 5 μmol/L （3.6±1.82）%，ATO 0.75 μmol/L （26.2±2.76）% and the combination group （76.4±9.33）%（P=0.003).Also，in 5 μmol/L Ibrutinib，0.75 μmol/L ATO，combination group，cell cycle distribution analysis showed that the cell cycle was arrested in the S phase.The protein expression levels of Caspase-3，PARP，CyclinD1 and Bcl-2 decreased.In contrast，the protein expression level of Cleaved caspase-3，Cleaved PARP increased.The combination group was superior to single drug group.Conclusion:Ibrutinib combined with ATO can effectively inhibit the proliferation and induce apoptosis and cell cycle blockage of cells than single agent.