米托坦逆转P-gp介导的人卵巢癌细胞的耐药作用及机制探讨

Reversal effect of mitotane on the drug-resistance mediated by P-gp in Human ovarian cancer cells

目的探讨米托坦对人卵巢癌细胞耐药的逆转作用及机制。方法采用10.0μM的阿霉素(ADM)体外高浓度反复间歇诱导法建立人卵巢癌细胞OVCAR-3/ADM耐药模型,观察其对紫杉醇、米托蒽醌、长春新碱、丝裂霉素、顺铂的耐药情况;MTT法检测不同浓度米托坦以及不同浓度米托坦联合ADM对OVCAR-3/ADM的生长抑制作用;Western blot法检测OVCAR-3/ADM的P-gp蛋白表达情况。结果 OVCAR-3的IC50为10.0μM;OVCAR-3/ADM的IC50为164.65μM,耐药指数为16.3。OVCAR-3/ADM对上述六种化疗药均产生了耐药性。10.0μM的ADM对OVCAR-3/ADM细胞的生长抑制率为4.78%。米托坦在高浓度时(≥30.0μM)时才表现出强的抑制肿瘤细胞生长作用,而米托坦与ADM联合应用时在米托坦浓度大于1μM时便可呈剂量依赖性抑制肿瘤细胞的生长,且抑制率明显高于单纯应用米托坦组及ADM组,100.0μM米托坦+10.0μM ADM组细胞生长抑制率最高,达到50.24%,与ADM组比较,经米托坦处理过的OVCAR-3/ADM的P-gp蛋白表达下降。结论 OVCAR-3/ADM为获得性多药耐药细胞系,米托坦可增强该细胞系对ADM的敏感性,其逆转耐药的机制可能与下调OVCAR-3/ADM的P-gp蛋白表达有关。

Objective To investigate the reversal effect and mechanism of mitotane on human ovarian cancer cells. Methods We established adriamycin （ADM）-resistant human ovarian caner cell line OVCAR-3/ADM by using high-dose and interval-inactivation methods in vitro to observe its resistance characteristics. The inhibitory rate of cell growth was measured by MTT assay at different drug concentrations, and the expression of P-glycoprotein （P-gp） was detected by Western blotting. Results IC50 value of ADM to OVCAR-3 was 10.0μM while it to OVCAR-3/ADM was 164.65 μM, with a resistant index of 16.3. The OVCAR-3/ADM cells had drug-resistance to paclitaxel, mitoxantrone, vincristine, mit- omycin and eisplatin. The growth inhibition rate of 10.0μM ADM on OVCAR-3/ADM cells was only 4.78%. Mitotane ex- pressed strong inhibition of tumor cell growth at high concentrations （≥30.0 μM） , the inhibitory rate of combined use of mitotane （ 〉 1 μM） and ADM was significantly higher than the mitotane group and ADM group and in a dose-dependent manner. The cell growth inhibitory rate in 100 μM mitotane ＋ 10.0μM ADM group was the highest （50.24%）. Com- pared with ADM group, the expression of P-gp in OVCAR-3/ADM cells treated by mitotane was obviously decreased. Con- clusions OVCAR-3/ADM is an acquired multidrug-resistant cell line, and mitotane enhances the sensitivity of human o- varian cancer cells to adriamycin. The mechanism may be associated with the down-regulation of P-gp expression.