安罗替尼联合氯喹促进人非小细胞肺癌细胞系H1299凋亡

Anlotinib combined with chloroquine induces apoptosis of human non-small cell lung cancer cell line H1299

目的探讨安罗替尼和氯喹(CQ)单用及联用对人非小细胞肺癌系H1299存活率、迁移、侵袭和凋亡的影响。方法安罗替尼与CQ单用及两药联合处理H1299细胞,MTT法检测细胞的存活率;划痕愈合和Transwell小室法检测细胞迁移及侵袭能力;Hoechst 33342染色检测细胞凋亡;Western blot检测自噬标志物LC3-Ⅱ/LC3-Ⅰ和自噬底物p62蛋白表达。结果安罗替尼联合CQ组对H1299细胞存活率、侵袭及迁移抑制效应优于安罗替尼组(P<0.05);安罗替尼单用时较对照组的细胞凋亡增加,当安罗替尼联合CQ作用时细胞凋亡增加更显著。安罗替尼随浓度增加明显上调自噬标志物蛋白LC3-Ⅱ/LC3-Ⅰ和下调自噬底物蛋白p62;CQ可显著上调LC3-Ⅱ/LC3-Ⅰ和p62表达水平;与单用CQ组相比,20μmol/L安罗替尼联合CQ组更加显著上调LC3-Ⅱ/LC3-Ⅰ和下调p62。结论安罗替尼联合氯喹可显著抑制H1299细胞的存活率、迁移和侵袭,促使细胞凋亡;其机制可能通过自噬发挥抗肿瘤作用。

Objective To investigate the effects of anlotinib and chloroquine(CQ)on viability,invasion and apoptosis of H1299 cells.Methods After treatment of H1299 cells with anlotinib or/and CQ,the cell viability was measured by MTT assay;Wound-healing and Transwell chamber assay were used to evaluate cell migration and invasion abilities;Hoechst 33342 staining was adopted to test the changes of apoptosis;The protein levels of autophagy marker LC3-Ⅱ/LC3-Ⅰand autophagic substrate p62 were detected by Western blot.Results Anlotinib combined with CQ showed more obviously synergistic inhibitory effect on the viability,invasion and migration in H1299 cells than anlotinib alone(P<0.05).As compared to the untreated control cells,anlotinib alone promoted the cell apoptosis.Their combination induced an even higher apoptosis in H1299 cells.With the increase of concentration,anlotinib significantly up-regulated autophagy marker protein LC3-Ⅱ/LC3-Ⅰand down-regulated autophagicsubstrate protein p62.The expression of LC3-Ⅱ/LC3-Ⅰand p62 was significantly up-regulated by CQ.Compared with the CQ group alone,LC3-Ⅱ/LC3-Ⅰand p62 were significantly up-regulated and down-regulated in the 20μmol/L anlotinib combined with CQ group.Conclusions Combination of anlotinib and CQ remarkably inhibits the viability,migration and invasion and induces the apoptosis of H1299 cells.The potential mechanism for antitumor might be due to autophagy's action.