Previously, we reported that Y_6, a new epigallocatechin gallate derivative, is efficacious in reversing doxorubicin(DOX)–mediated resistance in hepatocellular carcinoma BEL-7404/DOX cells. In this study, we evaluate...Previously, we reported that Y_6, a new epigallocatechin gallate derivative, is efficacious in reversing doxorubicin(DOX)–mediated resistance in hepatocellular carcinoma BEL-7404/DOX cells. In this study, we evaluated the efficacy of Y_6 in reversing drug resistance both in vitro and in vivo by determining its effect on the adenosine triphosphate-binding cassette protein B1 transporter(ABCB1 or P-glycoprotein, P-gp). Our results showed that Y_6 significantly sensitized cells overexpressing the ABCB1 transporter to anticancer drugs that are ABCB1 substrates. Y_6 significantly stimulated the adenosine triphosphatase activity of ABCB1. Furthermore, Y_6 exhibited a higher docking score as compared with epigallocatechin gallate inside the transmembrane domain of ABCB1. In addition, in the nude mousetumor xenograft model, Y_6(110 mg/kg, intragastric administration), in combination with doxorubicin(2 mg/kg, intraperitoneal injection), significantly inhibited the growth of BEL-7404/DOX cell xenograft tumors, compared to equivalent epigallocatechin gallate. In conclusion, Y_6 significantly reversed ABCB1-mediated multidrug resistance and its mechanisms of action may result from its competitive inhibition of the ABCB1 drug efflux function.展开更多
目的探讨汉防己甲素(Tet)联合阿霉素(DOX)体外逆转耐药型前列腺癌(DU145/DOX)细胞耐药的可行性及潜在机制。方法采用经典的DOX浓度梯度诱导法制备DU145/DOX细胞,然后采用3-(4,5-二甲基噻唑-2)-2,5-二苯基四氮唑溴盐(MTT)法测定Tet干预对...目的探讨汉防己甲素(Tet)联合阿霉素(DOX)体外逆转耐药型前列腺癌(DU145/DOX)细胞耐药的可行性及潜在机制。方法采用经典的DOX浓度梯度诱导法制备DU145/DOX细胞,然后采用3-(4,5-二甲基噻唑-2)-2,5-二苯基四氮唑溴盐(MTT)法测定Tet干预对DU145/DOX细胞生长抑制的作用,凋亡试剂盒测定联合用药对耐药细胞凋亡诱导的影响,流式细胞仪和荧光分光光度计分别测定Tet对DOX和荧光探针罗丹明123(R123)被DU145/DOX细胞摄取行为,Pgp-Glo TM分析系统测定Tet对P-糖蛋白(P-gp)ATP酶活性的影响,Western blot法测定Tet对耐药细胞膜上P-gp蛋白表达的影响,RT-PCR法测定Tet对DU145/DOX细胞MDR1 m RNA表达的影响。结果 Tet组、DOX组、DOX+Tet(1/5)组对DU145/DOX细胞的半数抑制浓度(IC50,以COX浓度计算)分别为(7.16±0.54)、(1.62±0.09)、(0.37±0.03)μM,Tet能明显增强DOX的抗肿瘤作用。DOX+Tet(1/5)组诱导DU145/DOX细胞凋亡率是DOX组的2.1倍。Tet的参与能明显提高DU145/DOX细胞内DOX和R123的累积量,其机制可能与Tet刺激细胞内P-gp ATP酶活性、降低细胞膜上P-gp蛋白表达有关。Tet可能通过降低耐药基因表达的机制,联合DOX提高对DU145/DOX细胞杀伤作用。结论 Tet与DOX体外联合使用具有逆转DU145/DOX细胞耐药的潜力,作用机制可能涉及刺激耐药泵酶活性、降低耐药蛋白表达量、下调MDR1 m RNA表达等。展开更多
基金supported by the National Natural Science Foundation of China (No. 81160532)the Open Project of Guangxi Colleges and Universities Key Laboratory of Biological Molecular Medicine Research (No. GXBMR201602, China)+1 种基金the Young and Middle-aged Teachers Foundation Ability Enhancement Project of Guangxi Colleges and Universities (No. 2018KY0102, China)US NIH (No. 1R15CA143701)
文摘Previously, we reported that Y_6, a new epigallocatechin gallate derivative, is efficacious in reversing doxorubicin(DOX)–mediated resistance in hepatocellular carcinoma BEL-7404/DOX cells. In this study, we evaluated the efficacy of Y_6 in reversing drug resistance both in vitro and in vivo by determining its effect on the adenosine triphosphate-binding cassette protein B1 transporter(ABCB1 or P-glycoprotein, P-gp). Our results showed that Y_6 significantly sensitized cells overexpressing the ABCB1 transporter to anticancer drugs that are ABCB1 substrates. Y_6 significantly stimulated the adenosine triphosphatase activity of ABCB1. Furthermore, Y_6 exhibited a higher docking score as compared with epigallocatechin gallate inside the transmembrane domain of ABCB1. In addition, in the nude mousetumor xenograft model, Y_6(110 mg/kg, intragastric administration), in combination with doxorubicin(2 mg/kg, intraperitoneal injection), significantly inhibited the growth of BEL-7404/DOX cell xenograft tumors, compared to equivalent epigallocatechin gallate. In conclusion, Y_6 significantly reversed ABCB1-mediated multidrug resistance and its mechanisms of action may result from its competitive inhibition of the ABCB1 drug efflux function.
文摘目的探讨汉防己甲素(Tet)联合阿霉素(DOX)体外逆转耐药型前列腺癌(DU145/DOX)细胞耐药的可行性及潜在机制。方法采用经典的DOX浓度梯度诱导法制备DU145/DOX细胞,然后采用3-(4,5-二甲基噻唑-2)-2,5-二苯基四氮唑溴盐(MTT)法测定Tet干预对DU145/DOX细胞生长抑制的作用,凋亡试剂盒测定联合用药对耐药细胞凋亡诱导的影响,流式细胞仪和荧光分光光度计分别测定Tet对DOX和荧光探针罗丹明123(R123)被DU145/DOX细胞摄取行为,Pgp-Glo TM分析系统测定Tet对P-糖蛋白(P-gp)ATP酶活性的影响,Western blot法测定Tet对耐药细胞膜上P-gp蛋白表达的影响,RT-PCR法测定Tet对DU145/DOX细胞MDR1 m RNA表达的影响。结果 Tet组、DOX组、DOX+Tet(1/5)组对DU145/DOX细胞的半数抑制浓度(IC50,以COX浓度计算)分别为(7.16±0.54)、(1.62±0.09)、(0.37±0.03)μM,Tet能明显增强DOX的抗肿瘤作用。DOX+Tet(1/5)组诱导DU145/DOX细胞凋亡率是DOX组的2.1倍。Tet的参与能明显提高DU145/DOX细胞内DOX和R123的累积量,其机制可能与Tet刺激细胞内P-gp ATP酶活性、降低细胞膜上P-gp蛋白表达有关。Tet可能通过降低耐药基因表达的机制,联合DOX提高对DU145/DOX细胞杀伤作用。结论 Tet与DOX体外联合使用具有逆转DU145/DOX细胞耐药的潜力,作用机制可能涉及刺激耐药泵酶活性、降低耐药蛋白表达量、下调MDR1 m RNA表达等。
