二甲双胍通过抑制IL-6/NF-κB/P-gp改善卵巢癌PARP抑制剂耐药

Metformin improves PARP inhibitor resistance in ovarian cancer by inhibiting IL-6/NF-κB/P-gp

目的:探讨二甲双胍对同源重组修复功能缺陷(HRD)阳性PARP抑制剂(PARPi)耐药卵巢癌的作用及相关机制。方法:逐步诱导法建立尼拉帕利耐药的HRD阳性SKOV3卵巢癌细胞系(SKOV3/PARPi)。白细胞介素-6(IL-6)、转染si-IL-6 RNA以及二甲双胍作用后,采用CCK-8法检测SKOV3或SKOV3/PARPi细胞增殖的变化,Western blot检测细胞内IL-6及P糖蛋白(P-gp)水平、AKT及NF-κB活性的变化,ELISA法检测细胞培养上清液中IL-6含量的变化。建立SKOV3/PARPi卵巢癌裸鼠荷瘤模型并验证二甲双胍、PARPi处理对瘤体的影响,免疫组化及Western blot检测瘤组织内P-gp蛋白的表达。结果:100μg/L IL-6作用SKOV3细胞后尼拉帕利的IC_(50)上调(P<0.001),细胞内P-gp表达增加(P<0.001)。SKOV3/PARPi细胞转染si-IL-6 RNA后尼拉帕利的IC_(50)下调(P=0.008),P-gp表达降低(P<0.001)。二甲双胍作用SKOV3/PARPi细胞后,尼拉帕利的IC_(50)下调(P<0.001);细胞内IL-6及P-gp蛋白水平、AKT及NF-κB活性均下降(P<0.001)。二甲双胍可改善SKOV3/PARPi荷瘤裸鼠尼拉帕利耐药,下调瘤组织内P-gp表达(P均<0.05)。结论:二甲双胍可通过抑制IL-6/NF-κB/P-gp改善卵巢癌PARPi耐药。

Aim:To investigate the effect and related mechanism of metformin on homologous recombination repair defect(HRD)positive PARP inhibitor(PARPi)resistant ovarian cancer.Methods:Niraparib-resistant HRD positive SKOV3 ovarian cancer cell lines(SKOV3/PARPi)were established by stepwise induction.CCK-8 assay was used to detect the proliferation of SKOV3 or SKOV3/PARPi cells treated with interleukin-6(IL-6),transfected with si-IL-6 RNA and metformin;Western blot was used to detect the levels of IL-6 and P-glycoprotein(P-gp),and the activity of AKT and NF-κB;the content of IL-6 in the cells culture medium was determined by ELISA.SKOV3/PARPi cells nude xenograft models were established and the effect of metformin and PARPi on xenograft was verified.The expression of P-gp protein in xenograft tumor tissue was detected by immunohistochemistry and Western blot.Results:After 100μg/L IL-6 was added into SKOV3 cells,the IC 50 of niraparib was significantly up-regulated(P<0.001),intracellular P-gp protein level increased significantly(P<0.001).After SKOV3/PARPi cells were transfected with si-IL-6 RNA,the IC 50 of niraparib was significantly down-regulated(P=0.008),and P-gp protein level decreased significantly(P<0.001).When treated with metformin,the IC 50 of niraparib in SKOV3/PARPi cells was significantly down-regulated(P<0.001),and the protein levels of IL-6,P-gp,and the activity of AKT,NF-κB were decreased significantly(P<0.001).Metformin reduced niraparib resistance in nude mice with SKOV3/PARPi xenografts,and down-regulated P-gp protein expression in tumor tissue(P<0.05).Conclusion:Metformin improves PARP inhibitor resistance in ovarian cancer by inhibiting IL-6/NF-κB/P-gp.