Oral squamous cell carcinoma (OSCC) is thought to arise as the result of cumulative genetic or epigenetic alterations in cancer-associated genes. We focused on the Dickkopf-3 (Dkk-3) gene as a candidate tumor suppressor in OSCC. Dkk-3 is a potential tumor suppressor, and its downregulation has been reported in various types of malignancies. However, our previous data demonstrated that the Dkk-3 protein was dominantly expressed in OSCC tissue, and its expression was correlated with a high incidence of metastasis and with poor prognosis. In order to explain this paradox, we performed functional analyses of the Dkk-3 gene in cancer cell lines. RT-PCR revealed that Dkk-3 mRNA expression was observed in OSCC-derived cell lines but not in gastrointestinal or colorectal adenocarcinoma‑derived cell lines. The siRNA for Dkk-3 was transfected into Dkk-3-expressing cells, and the changes in cell proliferation, invasion and migration were assessed. The knockdown of Dkk-3 mRNA by siRNA transfection did not affect cell proliferation, but it significantly decreased cell migration and invasion. To further investigate the precise mechanism that contributes to the potential oncogenic function of Dkk-3, the Wnt canonical pathway and non-canonical pathways were assessed. Western blotting demonstrated that the effect of Dkk-3 knockdown on cell migration or invasion was not caused by activation of the Wnt pathways. These data demonstrated that Dkk-3 expression in OSCC was different than that in adenocarcinomas. Dkk-3 may possess an oncogenic function that is independent of Wnt signaling.

DKK3, a member of dickkopf WNT signaling pathway inhibitor family, is believed to be a tumor suppressor, because of its reduced expression in cancer cells. However, our previous studies have revealed that DKK3 expression is predominantly observed in head and neck, and oral squamous cell carcinoma (HNSCC/OSCC). Interestingly, HNSCC/OSCC patients with DKK3 expression showed high metastasis rate and poorer survival, and siRNA-mediated knockdown of DKK3 in HNSCC-derived cancer cell lines resulted in reduced cellular migration and invasion. From these data, it is hypothesized that DKK3 might exert oncogenic function specifically to HNSCC. In the present research, DKK3 overexpression model has been established, and its influences were investigated, together with molecular mechanism studies. DKK3 expression profile in cancer cell lines was invested, including HNSCC/OSCC, esophageal, gastric, colorectal, pancreatic, prostatic and lung cancers. DKK3 overexpression was performed in HNSCC derived cells by transfection of expression plasmid. The effects of DKK3 overexpression were assessed on cellular proliferation, migration, invasion and in vivo tumor growth. The molecular mechanism of DKK3 overexpression was investing by western blotting and microarray. DKK3 overexpression significantly elevated cellular proliferation, migration and invasion, along with increased mRNA expression of cyclin D1 and c-myc. However, reporter assays did not show TCF/LEF activation, suggesting that this increased malignant property of cancer cells was not driven by Wnt/ß-catenin pathway. In investigation for the pathways/molecules in DKK3-mediated signals, western blot analyses revealed phosphorylation of Akt (S473) and c-Jun (Ser63) are elevated. Application of PI3K-kinase inhibitor, LY 294002 on HSC-3 DKK3 cells significantly decreased tumor cell proliferation, migration and invasion. From all the results, we demonstrated that DKK3 might contribute to cellular proliferation, invasion, migration, and tumor cell survival in HNSCC cells, through a mechanism other than canonical Wnt signaling pathway, which might be attributed to PI3K-Akt signaling.

Ameloblastoma is the most frequently encountered odontogenic tumor, characterized by a locally invasive behavior, frequent recurrences, and, although rare, metastatic capacity. Loss or inactivation of tumor suppressor genes (TSGs) allows cells to acquire neoplastic growth. The ING family proteins are tumor suppressors that physically and functionally interact with p53 to perform important roles in apoptosis, DNA repair, cell cycle regulation, and senescence. TP53 genetic alterations were reported to infrequently occur in ameloblastoma. Considering that other TSGs related to TP53 could be altered in this tumor, we focused our study on the ING family genes. We analyzed the loss of heterozygosity (LOH) status of the ING family (ING1‐ING5) chromosomal loci in a group of ameloblastomas by microsatellite analysis, and correlated the ING LOH status with clinicopathological characteristics. By using specific microsatellite markers, high frequency of LOH was found at the loci of each ING gene family member (33.3‐72.2%). A significant relationship was shown between LOH of D2S140 (ING5 locus) and solid tumor type (p = 0.02). LOH of ING3MS (ING3 locus) was also high in solid type tumors, showing a near significant association. In addition, a notable tendency toward higher LOH for half of the markers was observed in recurrent cases. LOH of ING family genes appears as a common genetic alteration in solid ameloblastoma. The current study provides interesting novel information regarding the potential prognostic significance of the allelic loss of the ING gene family loci in ameloblastoma tumorigenesis.

Head and neck squamous cell carcinoma (HNSCC) is one of the most frequently occurring types of cancer worldwide. We focused on the fact that the aberrant function of Wnt/β-catenin signaling is a frequent event in malignancies. Dickkopf (Dkk)-3 is a major negative regulator of Wnt/β-catenin signaling, which is a known tumor suppressor and is down-regulated in various types of cancer. However, the expression profile of the Dkk-3 protein in HNSCC has not yet been reported. The present study was conducted to investigate Dkk-3 protein expression in 90 cases of HNSCC tissue samples and HNSCC-derived cell lines. In contrast to findings available on other types of cancer, the Western blot analysis revealed that HNSCC cell lines expressed the Dkk-3 protein. In immunohistochemistry, 76 cases (84.4%) out of 90 tissue samples were Dkk-3-positive, whereas only 14 cases (15.6%) were negative. Notably, survival analysis showed that the Dkk-3 (−) group exhibited significantly longer disease-free survival (p=0.038), metastasis-free survival (p=0.013) and longer overall survival (p=0.155). The results showed that the Dkk-3 protein was dominantly expressed and may be involved in carcinogenesis and metastasis in HNSCC. Moreover, the findings suggest that the function of Dkk-3 differs depending on the tissue of origin, and that it may exert an oncogenic function in HNSCC.