ISOLATION OF GENES REQUIRED FOR HPV-MEDIATED ORAL CANCER

口腔鳞癌是世界上最常见的十大癌症之一。 口腔恶性肿瘤患者的总体五年预后约为50%，低于宫颈鳞状细胞癌。 宫颈癌和口腔癌的发病率在与吸烟和饮酒这两个突出的风险因素相关时都会增加。 此外，宫颈癌与HPV感染密切相关。 90%的宫颈癌携带HPV基因组的“高危”血清型。 另一方面，关于口腔癌和HPV感染之间因果关系的数据一直是可变的。 尽管HPV在口腔和生殖器粘膜癌症发展中的作用相似，但两种类型粘膜的细胞和环境特性也存在显着差异。 这可能会导致癌症发展途径的差异。为了研究细胞转化为肿瘤时基因的差异表达，我们将利用Park等人开发的体外模型系统。在该模型中，正常口腔角质形成细胞被HPV感染，成为永生化细胞。然后，这些细胞暴露于与烟草相关的致癌物质中，从而将细胞转化为肿瘤。 使用该模型系统，我们将通过mRNA差异显示（mRNA DD）分析分离和表征差异表达的基因，随着细胞进展为恶性表型。 然后，我们将通过北方分析确认mRNA DD分析，以观察各种已建立的细胞系和原代培养物中的表达模式和表达稳定性。 此外，将对患者活检进行RT-PCR和原位杂交，以确定分离的mRNA DD产物的细胞类型和阶段特异性。 从mRNA DD分析获得的上调或下调的cDNA克隆将用于表达或直接克隆，以研究其在赋予HPV介导的致癌作用中的功能。 该项目可能最终确定在肿瘤转化的某些阶段中始终存在的特定生物标志物。 然后，可以开发一种快速诊断方法，通过一种敏感、具有成本效益的程序对患者进行常规筛查。 在正常的办公室访问期间可以提供早期检测，以增加口腔癌的生存机会。

Oral squamous carcinoma is among the top ten most frequently occurring cancers in the world. The overall five year prognosis for patients with oral malignancies is about 50 percent, which is lower than the comparable squamous carcinoma of the cervix. The incidence of both cervical and oral carcinomas increase when associated with two prominent risk factors, cigarette smoking and alcohol consumption. In addition, cervical cancer is strongly correlated with HPV infection. Ninety percent of cervical cancer harbor "high-risk" serotype of HPV genome. On the other hand, data on the causal relationship between oral cancer and HPV infection have been variable. In spite of the similarities in the role of HPV in the development of cancer in the oral and genital mucosa, there are also significant differences in the cellular and the environmental properties of the two types of mucosa. This will likely result in differences in the pathways of cancer development. To investigate the differential expression of the genes as cells transforms to tumor, we will utilize an in vitro model system developed by Park et al. In this model, normal oral keratinocytes are infected with HPV to become immortalized cells. Then the cells are exposed to a tobacco-related carcinogen, which converts the cells to a tumor. Using this model system, we will isolate and characterize differentially expressed gene(s) by the mRNA differential display (mRNA DD) analysis, as the cells progress to the malignant phenotype. We will then confirm the mRNA DD analysis by the Northern analysis to observe the expression pattern and the stability of expression in various established cell lines and primary cultures. In addition, RT-PCR and in situ hybridization will be performed with patient biopsies to determine the cell type and the stage specificity of the isolated mRNA DD products. The up-or down-regulated cDNA clones obtained from the mRNA DD analysis will be used for either expression or direct cloning to study its function in conferring HPV-mediated carcinogenesis. This project may eventually identify specific biomarkers that are consistently present in certain stages of neoplastic conversion. A quick diagnostic method can then be developed to screen patients routinely by a sensitive, cost-effective procedure. Early detection can be provided during normal office visits to increase the chance of survival from oral cancer.