TRAIL联合舒尼替尼对EGFR-TKIs抵抗的A549细胞株的抗肿瘤作用

Effect of TRAIL Combined with Sunitinib on EGFR-TKIs-resistant Human Non-Small Cell Lung Cancer Cell Line A549

[目的]探讨TRAIL与舒尼替尼(sunitinib)体外不同用药方式下对表皮生长因子受体酪氨酸激酶抑制剂(EGFR-TKI)抵抗的NSCLC A549细胞的抗肿瘤活性及其机制。[方法]实验分为对照组、TRAIL组、舒尼替尼组、TRAIL联合舒尼替尼组(T+S)、TRAIL序贯舒尼替尼组(T→S)和舒尼替尼序贯TRAIL组(S→T)。CCK8法检测TRAIL和舒尼替尼对A549细胞的生长抑制作用;流式细胞术检测TRAIL和舒尼替尼作用后细胞周期变化及诱导的细胞凋亡;Western blot检测TRAIL和舒尼替尼作用后Akt、p-Akt蛋白表达的变化。[结果]T+S组及T→S组的抗增殖作用及诱导细胞凋亡的能力明显优于TRAIL组、舒尼替尼组及S→T组(P均<0.05)。细胞周期显示,TRAIL和舒尼替尼均能使细胞周期阻滞于G0/G1期,T+S组、T→S组对G0/G1期的阻滞作用明显增强。Western blot结果显示,TRAIL单独作用于A549细胞能明显上调p-Akt的表达,而舒尼替尼能下调p-Akt的表达;T+S组、T→S组、S→T组p-Akt的表达明显减少。[结论]舒尼替尼通过抑制TRAIL诱导的PI3K/Akt通路活化,增加TRAIL诱导的A549细胞凋亡。此外,两药物对A549细胞周期的特异性阻滞作用,也是诱导凋亡增加的原因之一。

[Purpose] To investigate the efficacy of TRAIL in combination with sunitinib under three different sequence-dependent schedules on epidermal growth factor receptor tyrosine kinase inhibitors（EGFR-TKIs）-resistant human NSCLC A549 cells,and to reveal the responsible mechanisms.[Methods] Threats were divided into 6 groups：control group,TRAIL group,sunitinib group,TRAIL＋sunitinib group,TRAIL sequential sunitinib group（T→S） and S→T group. The drug sensitivity was estimated by CCK8 method. the alteration of cell cycle and apoptosis in A549 cells treated with TRAIL and sunitinib by different administration programs were analyzed by the flow cytometry（FCM）. The protein expression of Akt and p-Akt under different sequence-schedules was measured by Western blotting. [Results] Antiproliferative effects and induced apoptosis in the group of T ＋S and T→S were significantly better than those in the TRAIL group,sunitinib group and S→T group（P all 0.05）. FCM analysis showed that TRAIL and sunitinib resulted in cell cycle arrest at G0/G1 phase,while the G0/G1 phase arrest was more happen in the group of T＋S group and T→S group induced by two drugs. Western blot showed that TRAIL could induce the expression of p-Akt,and sunitinib could down regulate the expression of p-Akt;the expression of p-Akt in T ＋S group,T →S group and S→T group was significantly reduced. [Conclusion] Sunitinib enhances TRAIL-induced apoptosis by blocking the activation of PI3K/Akt signaling pathway in human NSCLC A549 cells. In addition,the two drugs resulted in cell cycle arrest at G0/G1 and can induce more cells apoptosis.