吴茱萸碱对黑色素瘤细胞的耐药逆转作用

Reversal Effects of Evodiamine on Drug Resistance of Melanoma Cells

黑色素瘤是目前恶性程度最高的肿瘤之一。临床上,常采用维罗非尼(PLX4032)作为晚期患者的治疗药物,但患者很快就出现了耐药。因此,如何克服耐药、提高患者生存率成为急需解决的问题。本文选用中药吴茱萸碱(EVO)与黑色素瘤A375细胞、对PLX4032耐药的A375细胞(A375/R)进行相关研究。采用CCK-8法检测发现,用EVO的细胞非毒性浓度0.5μmol/L处理A375/R,PLX4032对A375/R细胞的半数抑制浓度(IC50)降低,逆转倍数为3.85。显示EVO能够增强黑色素瘤A375/R细胞对PLX4032的敏感性。后续实验分为对照组、EVO组、PLX组、PLX+EVO组。流式细胞仪检测结果显示,EVO组细胞凋亡率为5.88%,PLX组细胞凋亡率为17.88%,PLX+EVO组细胞凋亡率为30.28%。细胞集落结果证明,PLX4032联合EVO能够抑制A375/R细胞的克隆形成。免疫印迹法结果证明,联合用药能够上调促凋亡蛋白质Bax、胱天蛋白酶3的表达量,下调p-Akt、p-NF-κB-p65及抗凋亡蛋白Bcl-2的蛋白质水平。以上结果均表明,EVO能够有效逆转黑色素瘤A375/R细胞耐药,并且诱导细胞凋亡,抑制细胞增殖。

Melanoma is one of the most malignant tumors at present.Vemurafenib(PLX4032)is often used as a drug for the treatment of advanced patients,but patients quickly develop drug resistance.Therefore,how to overcome drug resistance and improve the survival rate of patients has become an urgent problem yet to be solved.In this paper,evodiamine(EVO),A375 cells and PLX4032-resistant A375 cells(A375/R)were used to carry out the related experiments.After A375/R cells were treated with the non-toxic concentration of EVO,the 50%inhibition rate(IC50)of PLX4032 decreased,and the reversal fold was 3.85 times more.It was shown that EVO could enhance the sensitivity of melanoma A375/R cells to PLX4032.The following experiments were divided into the control group,EVO group and PLX group,PLX+EVO group.The results of flow cytometry showed that the apoptosis rate of the EVO group was 5.88%,that of the PLX4032 group was 17.88%,and that of the PLX4032 combined with EVO group was 30.28%.The colony assay showed that PLX4032 combined with EVO could inhibit the colony formation of A375/R cells.Western blotting showed that the combination of drugs could up-regulate the expression of the apoptosis-promoting protein Bax,Cascapse-3 and down-regulate the protein levels of p-Akt,p-NF-κB-p65 and the anti-apoptosis protein Bcl-2.These results suggested that EVO can effectively reverse the drug resistance of melanoma A375/R cells,induce apoptosis and inhibit the proliferation of melanoma A375 cells.