Ryo Tamamura

Secreted frizzled related protein (sFRP)-2 inhibits bone formation and promotes cell proliferation in ameloblastoma

Secreted frizzled related protein (sFRP)-2, a Wnt antagonist, was strongly expressed by both stromal and tumor cells of ameloblastoma. The aim of this study is to evaluate whether sFRP-2 secreted from tumor cells have any direct role in suppressed bone formation or not. A pre-osteoblastic cell line, KUSA/A1 cells, cultured in conditioned medium of an ameloblastoma-derived cell line (AM-1CM) was used in the study. Alkaline phosphatase (ALP) activity, alizarin red staining, mineral quantification and MTS assay was performed. Wnt-canonical pathway is a major pathway for osteoblasts. Antagonists of this pathway, sFRP-1, 2 and 3, were detected by immunohistochemistry and western blot analysis. KUSA/A1 cells cultured in AM-1CM showed high cell proliferation, low ALP activity without mineralized matrix deposition. sFRP-2 was strongly expressed in ameloblastoma tissue and AM-1 cells. After sFRP-2 depletion, the cells showed diffuse mineralization. In this study, it was confirmed that ameloblastoma cells have a major role in decreased bone formation by secreting sFRP-2 in cell culture model. Though, sFRP-2 has great effect on tumor progression, inhibition of sFRP-2s anti-bone formation activity and cell proliferative activity may reduce the invasive property of ameloblastoma and possibility of recurrence rate.

Loss of Heterozygosity at the 9p21-24 Region and Identification of BRM as a Candidate Tumor Suppressor Gene in Head and Neck Squamous Cell Carcinoma

We have analyzed allelic loss of the short arm of chromosome 9 in 39 head and neck cancers using 13 polymorphic markers and found two deletions of hot spots at 9p21 and 9p24. Loss of heterozygosity was detected at least at one locus in 28 of 39 cases (71.8%). P16, a well-known tumor suppressor gene, is considered to be a target for deletion of the 9p21 region. However, novel frequent chromosomal deletion and a candidate tumor suppressor gene, BRM at the 9p24 region, were detected. Moreover, comparison of clinicopathological variables demonstrated that loss of heterozygosity at the BRM locus was associated with a worse prognosis.

Deletion at Dickkopf (dkk)-3 locus (11p15.2) is related with lower lymph node metastasis and better prognosis in head and neck squamous cell carcinomas.

Head and neck squamous cell carcinoma (HNSCC) is a frequently occurring cancer, and despite improvement of its treatment methods, including chemotherapy, radiotherapy, and surgery, the improvement of survival remains poor. Recent advances in molecular biology of human cancer indicated various molecular abnormalities in HNSCC, including activation of oncogenes and inactivation of tumor suppressor genes (TSGs). Dickkopf (Dkk)-3 gene is known as a negative regulator of Wnt signaling and is suggested to function as TSG in several kinds of malignancies. We hypothesized that Dkk-3 might play an important role in HNSCC, too. Thus, in the current study, we analyzed allelic alteration of Dkk-3 locus (chromosome 11p15.2) by means of loss of heterozygosity (LOH) analysis. The study population consisted of 50 patients with HNSCC (mean age of 65 years old). Furthermore, we also examined the correlation between LOH findings of Dkk-3 locus with clinicopathological parameters to investigate its use as a biomarker in HNSCC. A remarkable LOH ratio (57%) was detected in the cases studied, implying that Dkk-3 is likely to be involved in HNSCC carcinogenesis. However, interestingly and in contrast to the expectations, we found that the group with LOH of Dkk-3 locus had less lymph node metastasis, and showed a favorable overall survival compared to the patients with retention of Dkk-3 area in survival analysis. These results indicate that Dkk-3 can play a role in HNSCC carcinogenesis with unknown mechanism. Moreover, allelic loss at Dkk-3 locus may also be used as a novel prognostic biomarker in HNSCC.

Allelic loss of the ING gene family loci is a frequent event in ameloblastoma

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 D2S 140 (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.

Expression and Mutation Analysis of Her2 in Head and Neck Squamous Cell Carcinoma

ABSTRACT We analyzed mutation and expression status of human epidermal growth factor receptor 2 (Her2) in head and neck squamous cell carcinoma (HNSCC) using single strand conformation polymorphism (SSCP) mutation analysis and immunohistochemistry (IHC). Mutations were absent in all 85 cases. Out of 57 cases available for IHC, Her2 protein expression was negative (0) in 40 tumors (70%). Seventeen tumors (29.8%) expressed Her2, among these 13 tumors (22.8%) showed a weak (+1) expression and 4 (7%) showed a moderate expression (+2), none showed a strong (+3) expression. There was not a significant association between expression and any of the patients’ clinical variables or prognosis. Our results suggest that Her2 may not be useful as a molecular target in HNSCC.

Stromal cells promote bone invasion by suppressing bone formation in ameloblastoma

Aims:  To study the stromal variation and role of stromal–tumour cell interaction in impaired bone formation as well as enhanced bone resorption in ameloblastoma.

Potential usage of ING family members in cancer diagnostics and molecular therapy.

The Inhibitor of Growth (ING) gene family is an emerging putative type II tumor suppressor gene (TSG). Proteins of INGs (ING1-5), critical modulator of the histone code via PHD fingers, are able to suppress cell growth and proliferation, induce apoptosis, and modulate cell cycle progression. ING proteins are involved in transcriptional regulation of genes, such as the p53-inducible gene p21. ING proteins also serve as shuttling proteins between nucleus and cytoplasm, and dysregulation of this nucleocytoplasmic traffic has been shown in some cancer cells. In cancer cells, ING mRNA levels are often lost or suppressed but the genes are rarely mutated. Recently the potential roles of ING proteins as prognostic biomarkers, detection of aggressive behavior of the tumor as well as prediction of chemo-radiotherapy response have also emerged. In this review, we summarize the up-to-date knowledge on functions of the ING proteins, the protein status in human tumors and discuss as a potential target in the molecular diagnostics and therapy of cancer.

Frequent Allelic Loss of Dkk-1 Locus (10q11.2) is Related with Low Distant Metastasis and Better Prognosis in Head and Neck Squamous Cell Carcinomas

ABSTRACT Head and neck squamous cell carcinoma (HNSCC) is a frequently occurring cancer worldwide. Dickkopf (Dkk)-1 gene is suggested to function as tumor suppressor gene (TSG) in several kinds of malignancies. In this study, we performed loss of heterozygosity (LOH) analysis of Dkk-1 and examined the correlation between LOH status and clinicopathological parameters for the first time. A pretty high LOH ratio (50%) was detected. Interestingly, in the cases with Dkk-1 retention group showed less distant metastasis and a tendency of longer disease free survival. These results indicate that Dkk-1 can play a role in HNSCC carcinogenesis and it may also be related to distant metastasis.