Hedgehog−Gli2 Signaling Promotes Chemoresistance in Ovarian Cancer Cells by Regulating MDR1

Abstract

BackgroundCisplatin (DDP) resistance remains a key challenge in improving the clinical outcome of patients with ovarian cancer (OC). Gli2 overexpression can lead to DDP resistance in OC cells, but the specific underlying regulatory mechanism remains unclear. The membrane transporter encoding gene MDR1 positively regulates chemotherapy resistance in various cancer types. We evaluated MDR1 as a potential Gli2 downstream target and the contribution of the Gli2/MDR1 axis in promoting DDP resistance in OC cells.MethodsTo generate drug-resistant SKOV3/DDP cells, SKOV3 cells were grown for six months under continuous induction wherein the DDP concentration was steadily increased. Gli2 expression in OC cells with varying DDP sensitivities was detected using western blot. Cell counting kit-8 assays were used to assess the DDP sensitivity of SKOV3, SKOV3/DDP, A2780, and A2780/DDP cells and reversal of DDP resistance in SKOV3/DDP and A2780/DDP cells. Cell proliferation was analyzed using 5-ethynyl-2′-deoxyuridine (EdU) incorporation assays. The transcriptional regulation of MDR1 by Gli2 was determined using luciferase reporter assays. Finally, xenograft OC tumors were generated in nude mice, which were then treated with intraperitoneal DDP or phosphate-buffered saline (PBS) injections to investigate if Gli2 affected DDP resistance in OC in vivo.ResultsDDP-resistant SKOV3/DDP and A2780/DDP cells showed higher expression of Gli2 and MDR1 as compared with that in DDP-sensitive OC cells. Gli2 knockdown in SKOV3/DDP cells significantly reduced MDR1 expression, whereas it increased DNA damage, thereby sensitizing OC cells to DDP. Similar results were obtained after targeting Gli2 expression with the Gli-antagonist 61 inhibitor (GANT61) in SKOV3/DDP and A2780/DDP cells. In cells stably overexpressing Gli2, treatment with gradient concentrations of verapamil, an MDR1 inhibitor, significantly inhibited MDR1 expression. Our findings indicate that downregulation of MDR1 expression may reverse OC cell resistance to DDP. Moreover, dual-luciferase reporter gene assays confirmed that MDR1 is a direct downstream target of Gli2, with Gli2 positively regulating MDR1 expression. Finally, subcutaneous xenotransplantation in nude mice demonstrated that Gli2 plays a key role in regulating OC drug resistance.ConclusionsWe identified a mechanism by which Hedgehog-Gli signaling regulates OC chemoresistance by modulating MDR1 expression. Hence, Gli2 and MDR1 are potential biomarkers and therapeutic targets in patients with chemoresistant OC.

Reversal of Cisplatin Resistance in Ovarian Cancer by the Multitargeted Nanodrug Delivery System Tf-Mn-MOF@Nira@CDDP

“…Western blot analysis was performed to evaluate the expression of drug resistance-related proteins caused by the different treatments. We found that free CDDP exhibited no significant effect on MDR1 expression, resulting in multidrug resistance. − Free Nira, MCT, and MNT moderately reduced MDR1 expression, and the effect of MNCT was the most significant owing to the synergistic effect of Nira. In the SKOV3 DDP cells, the drug-loaded NPs exhibited a higher decrease in MDR1 expression than the single free drugs.…”

Baohuoside I inhibits resistance to cisplatin in ovarian cancer cells by suppressing autophagy via downregulating HIF‐1α/ATG5 axis

“…The DDP dose was increased from 50 ng/mL to a maximum of 1000 ng/mL (50, 100, 150, 200, 300, 500, 800, and 1000 ng/mL) as described in the previous study. 16 The DDP-resistant cell strain, labeled as A2780/DDP, was maintained in RPMI1640 complete medium containing 1 µg/mL DDP. During 48 h, 2.5, 5, 10, 20, or 40 μM Baohuoside I (Selleck) was added to both A2780 and A2780/ DDP cells.…”

“…After 3 days of DDP therapy (3‐day treatment cycles), the culture media was changed out for fresh, drug‐free medium and the cells were allowed to recuperate for another 3 days to a week. The DDP dose was increased from 50 ng/mL to a maximum of 1000 ng/mL (50, 100, 150, 200, 300, 500, 800, and 1000 ng/mL) as described in the previous study 16 . The DDP‐resistant cell strain, labeled as A2780/DDP, was maintained in RPMI1640 complete medium containing 1 µg/mL DDP.…”

CARD9 contributes to ovarian cancer cell proliferation, cycle arrest, and cisplatin sensitivity

“…Cancer cell line SKOV3 (ATCC) and cisplatin-resistant cell line SKOV3/DDP were cultured in McCoy’s 5a (Gibco). SKOV3/DDP was created utilizing the previously reported concentration-increasing approach [ 31 ] with cisplatin (C2210000, Merck). They were all supplemented with 10% fatal bovine serun and 1% penicillin-streptomycin (Gibco).…”