Combination of nigericin with cisplatin enhances the inhibitory effect of cisplatin on epithelial ovarian cancer metastasis by inhibiting slug expression via the Wnt/β‑catenin signalling pathway

Abstract Background: KIAA1456 is effective in the inhibition of tumorigenesis. We previously confirmed that KIAA1456 inhibits cell proliferation and metastasis in epithelial ovarian tumours. In the current study, the specific molecular mechanisms and clinical significance of KIAA1456 underlying the repression of epithelial ovarian cancer were investigated.Methods: Immunohistochemistry was used to evaluate the protein expression of KIAA1456 and SSX1 in epithelial ovarian tumours and normal ovarian tissues. The relationship of KIAA1456 and SSX1 with overall survival of patients with epithelial ovarian cancer was analysed with Kaplan–Meier survival curve and log-rank tests. KIAA1456 was overexpressed and silenced in HO8910PM cells with a lentivirus. The anticancer activity of KIAA1456 was tested by CCK8, plate clone formation assay, flow cytometry, wound healing assay and Transwell invasion assay. Xenograft tumour models were used to investigate the effects of KIAA1456 on tumour growth in vivo. Bioinformatics analyses of microarray profiling indicated that SSX1 and the PI3K/AKT signalling pathway were differentially expressed in KIAA1456-overexpressing and control cells. Therefore, the biological function of HO8910PM cotransfected with KIAA1456- and SSX1-overexpressing cells was detected to validate the rescue effect of SSX1. The downstream factors of PI3K/AKT that are related to cell growth and apoptosis, including p-AKT, PCNA, MMP9, CyclinD1 and Bcl-2, were detected by Western blot analysis.Results: KIAA1456 expression was lower in epithelial ovarian tumours than in normal ovarian tissues. Its expression level negatively correlated with pathological grade. Pearson’s correlation analysis showed that KIAA1456 negatively correlated with SSX1 expression. The overexpression of KIAA1456 in HO8910PM cells inhibited proliferation, migration and invasion and promoted apoptosis. By contrast, the silencing of KIAA1456 resulted in the opposite behaviour. A xenograft tumour experiment showed that KIAA1456 overexpression inhibited tumour growth in vivo. Mechanistically, the overexpression of KIAA1456 inhibited SSX1 expression and AKT phosphorylation in HO8910PM cells, causing the inactivation of the AKT signalling pathway and eventually reducing the expression of PCNA, CyclinD1, MMP9 and Bcl2. Similarly, the silencing of KIAA1456 resulted in the opposite behaviour. Finally, SSX1 overexpression could partially reverse the KIAA1456-induced biological effect.Conclusion: KIAA1456 may serve as a tumour suppressor via the inactivation of SSX1 and the AKT pathway, providing a promising therapeutic target for epithelial ovarian cancers.

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AMPK-independent LKB1 activity is required for efficient epithelial ovarian cancer metastasis

10.1101/591073 ◽

2019 ◽

Author(s):

Adrian Buensuceso ◽

Yudith Ramos Valdes ◽

Gabriel E. DiMattia ◽

Trevor G. Shepherd

Keyword(s):

Ovarian Cancer ◽

Epithelial Ovarian Cancer ◽

Cell Survival ◽

Cell Lines ◽

Cancer Metastasis ◽

Metastatic Potential ◽

Metastatic Tumour ◽

Ampk Signaling ◽

Dual Specificity ◽

Liver Kinase B1

ABSTRACTEpithelial ovarian cancer (EOC) spreads by direct dissemination of malignant cells and multicellular clusters, known as spheroids, into the peritoneum followed by implantation and growth on abdominal surfaces. Using a spheroid model system of EOC metastasis, we discovered that Liver kinase B1 (LKB1), encoded by theSTK11gene, and its canonical substrate AMP-activated protein kinase (AMPK) are activated in EOC spheroids, yet only LKB1 is required for cell survival. We have now generatedSTK11-knockout cell lines using normal human FT190 cells and three EOC cell lines, OVCAR8, HeyA8, and iOvCa147.STK11KO did not affect growth and viability in adherent culture, but it decreased anchorage-independent growth of EOC cells. EOC spheroids lacking LKB1 had markedly impaired growth and viability, whereas there was no difference in normal FT190 spheroids. To test whether LKB1 loss affects EOC metastasis, we performed intraperitoneal injections of OVCAR8-, HeyA8-, and iOvCa147-STK11KO cells, and respective controls. LKB1 loss exhibited a dramatic reduction on tumour burden and metastatic potential; in particular, OVCAR8-STK11KO tumours had evidence of extensive necrosis, apoptosis and hypoxia. Interestingly, LKB1 loss did not affect AMPKα phosphorylation in EOC spheroids and tumour xenografts, indicating that LKB1 signaling to support EOC cell survival in spheroids and metastatic tumour growth occurs via other downstream mediators. We identified the dual-specificity phosphatase DUSP4 as a commonly upregulated protein due to LKB1 loss; indeed,DUSP4knockdown in HeyA8-STK11KO cells restored spheroid formation and viability. Our results strongly indicate that intact LKB1 activity independent of downstream AMPK signaling is required during EOC metastasis.

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