Frizzled receptor 4 (FZD4) is associated with patient survival, invasiveness and tumour formation capacity in epithelial ovarian cancer


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Nair Bonito1,Wei Dai2,Robert Brown1
1Imperial College London,2The University of Hong Kong (HKU)

Abstract

Background

Although mutations in Wnt pathway genes are rare, DNA methylation of CpG islands at multiple Wnt genes is prognostic in epithelial ovarian cancer (EOC). DNA methylation of promoter CpG island of FZD4, a Wnt Pathway receptor, is associated with shorter progression free survival (PFS) and overall survival (OS) of EOC, indicating FZD4 might play an important role in ovarian cancer progression.

Method

We analysed publically available clinical and expression data from TCGA. Followed by stable overexpression and transient knockdown of FZD4, microarray analysis, ECM assays, tumour-sphere formation assay, qPCR and Western blotting.

Results

Expression analysis of TCGA data of EOC shows that patients with higher expression of FZD4 have longer PFS and OS than those with lower expression (PFS: adjusted HR=0.63, 95% CI 0.42-0.95, p=0.026; OS: adjusted HR=0.67, 95% CI 0.45-0.99, p=0.047, n=311). Microarray analysis comparing PEA1 ovarian tumour cells overexpressing FZD4 to control transfectants identified a set of ECM-regulated genes, which are significantly down-regulated by FZD4 expression. The expression profile of this gene set is also associated with patients’ PFS and OS from the TCGA dataset (PFS: adjusted HR=0.66, 95% CI 0.45-0.97, p=0.031; OS: adjusted HR=0.63, 95% CI 0.42-0.93, p=0.022) supporting a clinically relevant functional association. In SKOV3 and PEA1 ovarian tumour cells, FZD4 overexpression decreases invasion, cell mobility, cell adhesion while FZD4 knockdown has the opposite effects. Furthermore, increasing FZD4 expression suppresses tumour-sphere formation, which is reversed by FZD4 knockdown. FZD4 overexpressing cells show a decrease in p38-MAPK activation.

Conclusion

We have identified a novel link between FZD4 expression, ovarian cancer progression and p38 activation, suggesting a model whereby FZD4 expression suppresses p38 activation of transcriptional regulation of genes involved in cell mobility and invasion while epigenetic silencing of FZD4 leads to p38 activation, increased invasion and poor prognosis.