High expression of the secreted protein dickkopf homolog 4: roles in invasion and metastasis of renal cell carcinoma and its association with Von Hippel-Lindau gene.

The aim of this study was to investigate the effects of the dickkopf homolog 4 (DKK4)/Wnt/ß-catenin signaling pathway on tumorigenesis and metastasis in clear cell renal cell carcinoma (ccRCC), as well as to elucidate the underlying mechanisms. We examined the expression of DKK4 in 30 cases of ccRCC and matched adjacent normal tissues, and investigated its correlation with clinicopathological characteristics. Stable DKK4-transfected cells were established, and DKK4 functional analyses were performed, including a T-cell factor/lymphoid enhancer factor (TCF/LEF) reporter assay, and experiments on cell viability, apoptosis, invasive capability and tumor growth in vivo. Finally, western blot analysis was performed to detect Von Hippel-Lindau (VHL) expression in 50 clinical specimens. The expression levels of the DKK4, ß-catenin and ß-catenin downstream target genes, cyclin D1 and c-myc, were determined in the these specimens, as well as in RCC4(-), T3-14(+) cell lines by qRT-PCR and western blot analysis. The same tests were also performed in human embryonic kidney (HEK)293 cells which were transfected with the pCDH-DKK4 plasmid. After 6 weeks the tumor weight significantly increased in the mice transfected with the tumor cells. DKK4 mRNA and protein expression levels were significantly upregulated (p<0.001). DKK4 was distinctly overexpressed (68.0%) in all patient tissues. VHL(-) samples accounted for 60.0% of all samples, while DKK4 expression was significantly upregulated in 50% of these samples, indicating a correlation with VHL(-) expression (r=0.403, p<0.05). We also observed reduced expression levels of cyclin D1, c-myc and ß-catenin (to a greater extent) in the VHL(-), RCC4(-) and T3-14(+) cells, as well as in the stably transfected HEK293 cells. DKK4 may be an oncogene, and its upregulated expression may be involved in the pathogenesis of ccRCC as a downstream gene of VHL. By activating other pathways apart from the Wnt/ß-catenin pathway, DKK4 may play an important role in ccRCC tumorigenesis and metastasis.