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Overexpression of DUSP6 enhances chemotherapy-resistance of ovarian epithelial cancer by regulating the ERK signaling pathway.

ABSTRACT: Objective: DUSP6 is a negative regulator of the ERK signaling pathway and plays an important role in chemotherapy-resistance. Previously we showed that DUSP6 is overexpressed in ovarian cancer side population (SP) cells that possess cancer stem cell-like properties and are quiescent and chemotherapy-resistant. Here, we explore the effects of DUSP6 on chemotherapy-resistance by examining its regulation of the ERK signaling pathway and G0/G1 cell cycle arrest. Methods: mRNA and protein expression of DUSP6 and G0/G1 cell cycle checkpoint regulating proteins (CyclinD1, CyclinD3 and CyclinE2) was evaluated among ovarian cancer cell lines and tissue samples. Ovarian cancer cells were transiently transfected to overexpress DUSP6. After treatment with cisplatin, cell viability was measured by the MTS assay at 48 hours and the half maximal inhibitory concentration (IC50) for each cell line was calculated. Subcellular localization and cell cycle analysis were determined by using immunofluorescence and FACS, respectively. Results: SKOV3 and OVCAR8 SP cells were shown to express higher levels of DUSP6 and lower levels of CyclinD3 compared with non-SP (NSP) cells (P<0.001). Among 39 ovarian cancer tissue samples, expression of DUSP6 in the chemotherapy-resistant group (12 samples) was higher than in the chemotherapy-sensitive group (27 samples) (P<0.05). While a lower level of expression of CyclinD3 was seen in the chemotherapy-resistant group, it was not statistically different from the chemotherapy-sensitive group. HO8910 cells where shown to have higher IC50 to cisplatin than SKOV3 or OVCAR8 cells, and this correlated with higher levels of DUSP6 expression. Overexpression of DUSP6 in SKOV3 cells led to an increase in cisplatin IC50 values (P<0.05), and also markedly reduced the expression levels of phospho-ERK1/2 and CyclinD3 and to the predominance of cells in the G0/G1 phase. Conclusion: Our findings reveal an enhancement of chemotherapy-resistance and a predominance of cells in G1 cell cycle arrest in DUSP6-overexpressing ovarian cancer cells. This suggests that overexpression of DUSP6 promotes chemotherapy-resistance through the negative regulation of the ERK signaling pathway, increasing the G0/G1 phase ratio among ovarian cancer cells, and leading to cellular quiescence.

SUBMITTER: Gao Y

PROVIDER: S-EPMC7097933 | biostudies-literature |

REPOSITORIES: biostudies-literature

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Project description:BackgroundOvarian cancer has one of the highest mortality rates among all gynaecological malignancies, and increasing evidence suggests that lncRNAs are widely involved in the development of ovarian tumours. This study aimed to investigate the mechanism of the LNC00115/miR-7/ERK axis in the cisplatin resistance of ovarian cancer cells.MethodsThe expression of miR-7 and LNC00115 in ovarian cancer cell lines and tissues was detected by qRT-PCR. The ovarian cancer cell lines were constructed by overexpressing or knocking down the expression of LNC00115 or miR-7. CCK-8, transwell invasion, Western blot, immunohistochemistry, and luciferase reporter assays were carried out to identify the targets of LNC00115 and explore its roles and mechanisms in ovarian cancer. A nude mouse model was established, and the expression of LNC00115, miR-7 and ERK was detected. The changes in the tumours and body weights of the nude mice were measured.ResultsLNC00115 was upregulated in ovarian cancer tissues and cisplatin-resistant ovarian cancer cells. Moreover, LNC00115 promoted the cisplatin resistance, invasion and migration of ovarian cancer cells. LNC00115 was shown to directly target miR-7, and miR-7 was downregulated in ovarian cancer tissues and cisplatin-resistant ovarian cancer cells. miR-7 inhibited the cisplatin resistance, invasion and migration of ovarian cancer cells and directly targeted ERK. ERK was overexpressed in cisplatin-resistant ovarian cancer cells and ovarian cancer tissues. In animal experiments, overexpression of LNC00115 enhanced the cisplatin resistance of ovarian cancer cells, while miR-7 had the opposite effect. Mechanistically, LNC00115 sponged miR-7 to increase the expression of ERK, which in turn enhanced the cisplatin resistance of ovarian cancer.ConclusionOur data clarify the mechanism by which the LNC00115/miR-7/ERK axis promotes cisplatin resistance and provide a new clinical strategy for combating cisplatin resistance in ovarian cancer.

Project description:The importance of the Piwi-interacting RNA (piRNA) pathway for germ cell maintenance, genome integrity, DNA methylation and retrotransposon control raises possible roles of this pathway in cancer. Indeed aberrant expression of human PIWI orthologs and Maelstrom has been observed in various cancers. In this study we explored the expression and function of piRNA pathway genes in human ovarian cancer, based on our recent work, which showed widespread expression of piRNA pathway genes in the mammalian. Our work shows that PIWIL1 and MAEL expression is significantly increased in malignant EOC (n = 25) compared to benign tumor tissues (n = 19) and normal ovarian tissue (n = 8). The expression of PIWIL3 is lower in malignant and benign tissues when compared to normal ovary. Sequencing of PIWIL1 transcript revealed that in many tumors deletion of exon 17 leads to the introduction of a premature stop codon in the PIWI domain, likely due to a splicing error. In situ hybridization on tumor sections revealed that L1, PIWIL1, 2 and MAEL are specifically expressed in epithelial cells (cancerous cells) of EOC. Furthermore, PIWIL2 and MAEL are co-expressed in the stromal cells adjacent to tumor cells. Since PIWIL1 and MAEL are up regulated in malignant EOC and expressed in the epithelial cells, we investigated if these two genes affect invasiveness of ovarian cancer cell lines that do not normally express these genes. PIWIL1 and MAEL were transiently over expressed in the ovarian cancer cell line SKOV3, followed by real-time measurements of cell invasiveness. Surprisingly both PIWIL1 and MAEL over expression decreased the invasiveness of SKOV3 cells. Our findings support a growing body of evidence that shows that genes in this pathway are upregulated in cancer. In ovarian cancer we show for the first time that Piwil1 transcript may often be abnormal result in non functional product. In contrast to what has been observed in other cell types, we found that PIWIL1 and MAEL have a repressive effect on cell invasiveness.

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Overexpression of DUSP6 enhances chemotherapy-resistance of ovarian epithelial cancer by regulating the ERK signaling pathway.

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