Potential Roles of Microrna Expressions and Abc (Alcoholbetalcigarette) Oral Habits on the Carcinogenesis of Oral Cancer.

At the year of 2005, the incidence of oral cancer (ORC) has become the 6th leading cancer, the 5th leading cancer in male in Taiwan, and is still increasing with years. Since this cancer usually occurs in the middle age male, at the high peak of life responsibility, it has tremendous impact of family and society. Although the habitual use of AlcoholBetalCigarette (ABC) have been reported involving the oral carcinogenesis, the molecular factors leading to oral cancer have not been well characterized. MicroRNAs (miRNAs) are a prevalent class of small single-stranded non-coading RNAs (19-25 nts). They serve widespread functions as regulatory molecules in post-transcriptional gene silencing for the regulation of cellular proliferation, differentiation, and apoptosis, which have great impacts on the malignant transformation. Apparently, the expression of miRNA profile is closely associated with cancer. The aim of this study is to investigate the potential roles of miRNA and ABC consumption habits on the carcinogenesis of ORC. Four phases of study will be designed: miRNA screening, function characterization on cells, animal investigation, and clinical study. This knowledge will provide basic information for the potential applications of these molecules in the diagnosis and treatment of oral cancer. The four parts of study are described follows. (1) miRNA screening. (a) Over 500 known human miRNAs will be profiling by microarray analysis in native cloned 10 head neck cells lines, including 4 normal keratinocytes and 10 oral cancer cells. (b) Statistically Hierarchical analysis will be performed to differentiate the normal verse cancer profiles. (c) The top 10 miRNAs with high differential frequencies and levels will be further investigated. Quantitiative RT-PCR will be used to confirm the differential expressions in 10 clinical non-cancer mucosa and 20 cancer tissues. (2) Study on cellular level. The potential roles of these miRNAs on oral carcinogenesis will be investigated. (a) Construction of miRNA expression plasmids and transfection into oral cancer cells. (b) Study the potential effect of cellular functions, including the analyses of cell growth, colony formation, migration, invasion, cell cycle status, and apoptosis. (3) Study on animal level. (a) Establishment of stable luciferase- transfectant oral cancer cell lines. (b) Establishment of xenograft oral cancer nude mice models, with in situ tumor (subcutaneous injection) and metastatic tumor (intravenous injection). (c) Examining the effect of the tumor growth and survival after miRNA delivery into the xenografted mice using In Vivo Imaging System (IVIS, Xenogen). (d) Histological investigation of apoptotic status of the xenograft tumors. (4) Study on the clinical level. (a) Evaluation of miRNA expressions by real-time quantitative RT-PCR in the clinical oral tissues, including 200 oral cancers, 100 precancer lesions, and 100 oral mucosa cells. (b) Correlation of miRNA expressions with the cliniclpathological characteristics, include TNM staging, recurrent status and patient survival, using Person Chi-square or T-test analysis. (c) The information regarding to the ABC consumptions will be acquired for all the participators: 100 normal individuals, 100 precancer lesions, and 200 oral cancer patients. (d) Determination of the individual and combined powers of the potential multiple risk factors: ABC consumption and miRNA expressions with oral cancer incidence using odds ration analysis.

Project ID:PC9709-0926
External Project ID:NSC97-2320-B182-010-MY3