A poor prognosis of oral squamous cell carcinoma (SCC) is partly due to the invasiveness and metastasis of the tumor. One of key elements in tumor invasion and metastasis in the degradation of extracellular matrix is tissue inhibitors of metalloproteinases (TIMPs). This study was performed to determine the expression of TIMP-1 and TIMP-2 of oral SCCs with regard to the histologic invasiveness and differentiation in 5 normal oral mucosa and 36 oral SCCs. The histologic invasiveness of oral SCCs were classified into 4 grades. The differentiation of oral SCCs was divided into 3 grades. The StreptAvidin-Biotin immunohistochemical process, using TIMP-1 and TIMP-2 monoclonal antibodies, was performed to determine the expression of TIMP-1 and TIMP-2. The expression of TIMP-1 was positive in 5 of 17 oral SCCs with weak invasiveness and was positive in 8 of 19 oral SCCs with strong invasiveness. The TIMP-1 expression did not increase significantly with respect to the invasiveness of oral SCCs (P>0.05). The expression of TIMP-2 was strongly positive in 5 out of 17 SCCs with weak invasiveness and was strongly positive in 15 of 19 SCCs with strong invasiveness. The TIMP-2 expression increased significantly with respect to the invasiveness of oral SCCs; the stronger the expression, the stronger the invasiveness (P<0.05). The expression of TIMP-1 and TIMP-2 did not increase significantly with respect to the histologic differentiation. We concluded that with respect to the invasiveness, the TIMP-2 expression increases significantly in oral SCCs but the TIMP-1 expression does not; and that with respect to the histologic differentiation, their expressions do not increase significantly. These results suggested that TIMP-2 can be used as a tool to evaluate the invasiveness of oral SCCs.

Elevated expression of survivin is strongly associated with tumorigenesis and even in human common cancers. Oral squamous cell carcinoma (OSCC) is the 7th most frequent cancer in human and responsible for more than 90% of all oral cancer. The purpose of this study is to confirm whether survivin is associated with oral carcinogenesis, expecially has a role in the development of OSCC. For the control group; 3 specimens obtained from normal oral mucosa without any inflammatory reaction were used a nd for the experimental group, specimens obtained f rom 18 sub jects of OSCC; 6 subjects from Well differentiated type OSCC; 4 subjects from Moderately differentiated type OSCC; 3 subjects from Poorly differentiated type OSCC; 3 subjects from Verrucous carcinoma: and 2 subjects from C arcinoma in situ were used. All the specimens were embedded in paraffin, sectioned 5 μm or more in thickness, and stained with hematoxylin- eosin. For immunostain, the specimens were incubated with 1;200 diluted primary antibody (anti-survivin monoclonal, Biocare Inc, USA), followed by the secondary antibody(NovoLink Polymer detection system, Novocastra Lab., UK). The bound antibodies were visualized by addition of diaminobenzidine tetrahydrochloride(DAB) for 30 minutes at room temperature. The specimens were counterstained with Mayerʼs Hematoxylin and mounted. Quantitation of immunoreactivity was performed under the light microscope with the following criteria ; Intensive reaction; +++, Moderate reaction; ++, Minimal reaction; +. Using the image analyzer(Korea Optical System), immunoreactivity of tumor cells in various field was measured and statistically analyzed with SPSS 15.0 Program. The results were as follows: Expression of survivin in OSCC was significantly increased in the nucleus and the cytoplasm of OSCC as compare to those of control group (p<0.05). Expression of survivin in the nucleus and the cytoplasm of the cells in OSCC is correlated with the cellular malignancy (p<0.05). Expression of survivin in Poorly differentiated type OSCC partly correlated to some extent to cellular malignancy (p<0.05). These results suggest that expression of survivin in OSCC is closely associated with to the development, and malignancy of the OSCC, b ut it is not enough to be used a s a marker f or the c ellular malignancy. Further studies are needed to relate the expression of survivin to cellular malignancy.

The support mechanisms that are involved in lymph node metastasis of oral squamous cell carcinoma remain largely unknown. Recent studies have demonstrated that tumor cells express chemokine receptors and use chemokines to metastasize to the target organ in many malignancies in humans There are few reports about the correlation between chemokin receptor CXCR-4 expression and clinicopathologic factors in oral squamous cell carcinomas. The object of this study was to evaluate the availabili ty of CXCR-4 expression as prognostic marker through correlation analysis of CXCR-4 expression in oral sq uamous cell carcinoma and its r elation to clinocopathologic factors and PCNA index. 80 we investigated CXCR-4 expression of 74 oral squamous cell carcinomas by immunohistochemistry. 44 out of 74 cases(59. 5%) showed CXCH-4 positive and 30 sampl es(40.5%) showed CXCH-4 negative. CXCH-4 expression showed statistically sig nificant correlation wi th lymph node metastasis(p=0.026) ‘ PCNA index (p=0.003) , survial rate(p=0.0003). From the results , it was suggested CXCR-4 oxpression might be useful a prognostic marker in oral squamous cell carClllomas

Polo-Like Kinase(PLK) is a cell cycle-regulated, cyclin-independent serine/threonine protein kinase. Recent reports have shown a critical role for PLK during tumorigenesis. To explore whether PLK plays a general role as a tumor marker of oral squamous cell carcinomas, we examined the expression of PLK mRNA and protein in oral squamous cell carcinoma cells and immortalized normal oral keratinocytes(INOK). We also investigated that PLK mRNA was expressed in specimens from 4NQO-induced SD rat tongue carcinomas using in situ RT-PCR methods. Immunocytochemically, most of the PLK was highly expressed in the nucleus of carcinoma cells, but not INOK. RT-PCR revealed PLK1 mRNA was detected in the FaDu and Hep2 cancer cells, but no detected in the INOK. In situ RT-PCR revealed PLK1 mRNA expression increased sequentially from hyperplasia to dysplasia, and squamous cell carcinoma during the malignant progression. PLK1 expression could reflect the degree of malignancy and proliferation in oral squamous cell carcinomas. Thus, in addition to being of diagnostic value, modulation of PLK1 activity in the tumors by chemotherapeutic agents or gene therapy may prove to be of therapeutic value.

The tumor suppressor gene, phosphate and tensin homologue(PTEN) has been shown to dephosphorylate the phosphatidylinositol 3-kinase(PI 3-K)-generated phosphatidylinositol(3-5)-triphosphate in vivo, thus interfering with the potentially oncogenic signals emanating from PI 3-K. Promoter hypermethylation of CpG islands has recently been shown to be an epigenetic change resulting in loss of function in some genes involved in cell cycle regulation and DNA repair. Immunohistochemal staining for monoclonal antibody 6H2.1 was performed from paraffin embedded blocks of 20 benign epithelial lesions and 40 head and neck squamous cell carcinomas(HNSCCs). Immunoreactivity was graded semiquantitatively by considering the percentage and intensity of the staining of the tumor cells. Also, this study tried to identify PTEN methylation in benign epithelial lesions(24 cases) and HNSCCs(44 cases of paraffin embedded blocks, 4 cases of frozen tissues) using methylation-specific PCR(MSP). In HNSCCs, immunoreactive scores of stage 1 and 2(12 cases, average score 85.2) were higher than those of stage 3 and 4(15 cases, 41.9) and statistically significant(P=0.017). Immunoreactive scores of moderate and poorly differentiated carcinomas(22 cases, 61.6) are more or less lower than those of well differentiated carcinoma(15 cases, 87.0) but not significant(P=0.361). Among 24 cases of benign epithelial lesions, 12 cases showed unmethylated PTEN but none methylated. In HNSCCs, 22 of 44 paraffin embedded blocks showed unmethylated PTEN but none methylated, and all 4 frozen tissue revealed unmethylated PTEN, one of which(25%) methylated. We consider that the loss of PTEN protein expression may be associated with the progression of HNSCCs and the other alteration rather than methylation may be important in the inactivation of PTEN in HNSCCs.

Expression of invasion/metastasis suppressor, E-cadherin, is reduced in many types of human carcinomas. Although somatic and germline mutations in the CDH1, which encodes the human E-cadherin, have frequently been reported in cases with diffuse gastric and lobular breast cancers, irreversible genetic inactivations are rare in other human carcinomas. Recently, it has been well documented that some genes in human cancers may be inactivated by altered CpG methylation. Herein, we determined the expression and methylation status of E-cadherin in oral squamous cell carcinoma(SCC) by immunohistochemistry and methylation-specific PCR. The expression of E-cadherin was significantly higher in the well-differentiated oral SCCs than the moderately or poorly differentiated ones. None of eight tested benign epithelial hyperplasias showed aberrant methylation, whereas five of 12 oral squamous cell carcinomas showed aberrant methylation. When we compared E-cadherin expression with methylation status, oral SCCs with normal methylation showed a higher expression of E-cadherin than those with methylation. These findings suggest that aberrant CpG methylation of CDH1 promoter region is closely associated with transcriptional inactivation and might be involved in tumor progression of the oral mucosa.

To cletermine the role of mismatch repair in the clevelopment of oral squamolls cell carcinomas (OSCCs) , the prevalence of microsatellite instability (MSI), expression of I뼈LH1 ancll띠SH2 ， ancl hypennethylation of I뻐LH1 ancl hMSH2 were explorecl. Bya panel of five markers (BAT25, BAT26 , 02S123, 05S346, ancl 017~‘250 ， the so-callecl Bethescla markers) for screening of MSI, MSI was observecl in 5 of the 15 sqllamolls cell carcinomas (33.3%). As MSI is callsecl by the clysfllnction of MMR genes, this stucly examinecl the methylation status of CpG sites in the hMLH 1 ancl hMSH2 promoters ancl the expression hMLH1 ancl hMSH2. Becallse of inappr이)riate efficiency of fonnalin-frxecl ancl paraffin-embeclclecl samples for methylation-specific PCR (MS-PCR) of hMLH1 ancl hMSH2, the role of promoter hypelmethylation 띠 the clevelopment of MSI ancl expression of hMLH1 ancl hMSH2. meùlylation was failecl to clefìne. However, loss of nllclear staining of 1ψIILH 1 ancl hMSH2 were seen in nine (69.2%) ancl four (30.7%) of 13 OSCCs, while four ancl fìve all corresponcling normal epiùlelial tisslles showecl positive nllclear stι ining ofl마ILH1 and hMSH2. These data suggest that MSI anclloss of expression of hMLH1 ancl hMSH2 play a role in the carcinc핑enesis of oral sqllamolls cell carcinomas thollgh the role of promoter hypermethylation of hMLH1 ancl hMSH2 in MSI and their expresslon IS lIncertam

p16INK4A and p15INK4B tumor suppressor genes are frequently altered in various human tumors. Hypermethylation of the promoter region of p16INK4A and p15INK4B seem to be the major mechanism of inactivation. To determine whether the change in p16INK4A and p15INK4B methylation status occur in oral squamous cell carcinomas (OSCCs) and benign oral epithelial hyperplasias, we analyzed 46 OSCCs and 20 benign oral epithelial hyperplasias by methylation-specific PCR. We also analyzed a subset of the samples for p16INK4A and p15INK4B protein expression by immunohistochemistry. The promoter region of p15INK4B was hypermethylated in 13 specimens of the 15 finally analyzed OSCCs and three specimens of the five analyzed benign oral epithelial hyperplasias. By immunohistochemical analysis, we confirmed the loss of p15INK4B expression of all hypermethylated specimens. The promoter region of p16INK4A was amplified by both an unmethylated- and a methylated-specific primers in just one OSCCs. The remaining specimens including 11 OSCCs and four benign oral epithelial hyperplasias were normally methylated. By immunohistochemistry, we analyzed the loss of p16INK4A expression in seven specimens of the 12 OSCCs and two specimens of the four benign oral epithelial hyperplasias. Except for one OSCC, however, all specimens showing loss of expression were normally methylated. These results suggest that loss of p16INK4A and p15INK4B protein expression play an important role in the development of both OSCCs and benign oral epithelial hyperplasias.