2026-11-092026-11-09https://scholars.csmu.edu.tw/handle/123456789/37923Lung cancer is the leading and second cause of cancer death in female and male in Taiwan, respectively. In the last two years, lung cancer has replaced liver carcinoma to be the leading cause of cancer death in Taiwanese men. The death-population cause by lung cancer is about 20% of total population of cancer death. Similar trend was also observed in western countries and Japan. Prevention of lung cancer incidence is therefore, an important study topics in human health not only in Taiwan, but also worldwide. Cigarette smoking is considered to be the most important etiological factor of lung cancer and about 90% of lung cancer worldwide can be linked to cigarette smoking. However, in Taiwan, only around 50% of lung cancer was related to cigarette smoking, particularly, less than 10% of Taiwanese women are smokers. Thus, it is conceivable that environmental factors other than cigarette smoking may be associated with lung cancer development in Taiwan, especially for women nonsmokers. It is well known that p53 and k-ras mutations are frequently found in lung cancer development. However, this fact is not applicable to explain lung tumorigenesis of Taiwanese female lung cancer due to extremely low p53 and K-ras mutation frequencies were observed. In addition, higher frequencies of p53 immunostaings were detected in female lung cancer patients compared with that of male lung cancer patients. This pattern of p53 alteration is similar to that of cervical cancer raised the possibility of HPV 16/18 infection involved in female lung cancer development. Our recent report revealed the higher prevalence of HPV 16/18 infection in female lung cancer patients compared to that of male lung cancer patients (60% vs. 24% for HPV 16, 73.3% vs. 26% for HPV 18). This result strongly suggests that HPV infection is associated with lung cancer development of nonsmoking female lung cancer patients. However, whether HPV is indeed involved in lung tumorigenesis should be further investigated. To verify the role of HPV infection in lung tumorigenesis, the relationships between E6 mRNA / p53 inactivation and E7 mRNA/ Rb inactivation should first be investigated by in situ RT-PCR and immunohistochemistry in two serial slides of lung tumor paraffin sections. Our preliminary data showed that over 80% of cases with E6 and/or E7 mRNA were accompanied with p53 and/or Rb negative immunostainings (Tables 1, 2). These results support that HPV may be involved in lung tumorigenesis. Based on the preliminary finding, we address this project to further verify the role of HPV in lung tumorigenesis. In this project, tissue cDNA microarray with laser capture microdisection (LCM) will be used to elucidate the gene expression profiles of HPV infected lung tumors subtracted from HPV non-infected lung tumors. The expression profiles compared to that of cervical cancer may provide valuable evidence to verify whether HPV is involved in lung tumorigenesis. In addition, a lung epiderminal cell line BET-1A will be transfected with full length of HPV 16/18 genome and tumorigenic activities will be examined by soft agar and tumorigenic assays. The gene expression profiles of the immmortalized cells will be examined by cDNA microarray. If the gene expression profile obtained from lung tumor is similar to that of transfected cell lines, this will further evidence that HPV is indeed involved in lung tumorigenesis. The gene expression profiles from cDNA microarray will not only show the involvement of HPV in lung tumorigenesis but also reveal different tumorigenesis pathways in lung cancer development if there is any. The integration of HPV DNA into host chromosome is the hallmark of HPV induced carcinogenesis. To elucidate the role of HPV in lung tumorigenesis, chromosome integration mapp肺癌自民國七十一年開始一直是台灣婦女和男性癌症的第一和第二大死亡原因。近兩年來，男性肺癌之死亡率已超過肝癌成為第一大死亡原因。已支抽煙是肺癌發生的最重要病因。歐美國家的肺癌發生有90%以上可用抽菸行為來解釋。但是台灣地區則有一半的肺癌發生無法以抽菸來說明，尤其台灣女性有90%以上是不抽菸者。因此不抽者罹患肺癌之病因學研究，是當前我國重要的醫藥衛生研究課題。最近本研究室發現不抽煙之女性肺癌患者有高達60%和73%的高危險性人類乳突瘤病毒 16和18型(human papillomarvirus 16/18) 的感染率。此感染率遠高於男性肺癌患者（HPV 16, 24%; HPV 18, 26%）。因此推測HPV 16/18高感染率可能與不抽菸的台灣婦女罹患肺癌有相關。但是這結果並不能證明人類乳突瘤病毒參與肺癌的形成。最近本研究室在肺腫瘤病理連續切片上，分別進行in situ RT-PCR和免疫組織化學分析，了解HPV16/18 E6和E7 mRNA是否和p53以及Rb蛋白之去活化有關，初步結果發現有80%以上有E6/E7 mRNA的腫瘤切片中的p53和Rb蛋白無法被染出，反之則p53和Rb蛋白則會被染到。這結果暗示 HPV 16/18 可能透過E6/E7致癌蛋白去活化p53和Rb蛋白之致癌路徑參與肺癌形成。為了更明確證明HPV參與肺癌形成，本計畫擬以有和沒有HPV感染之女性第一期肺腺癌之肺癌組織，用Laser capture microdissection (LCM)將肺腫瘤細胞取下，並萃取出mRNA進行cDNA microarray分析，比較分析HPV感染和不感染之肺癌祖織的基因表現輿圖中最主要差異的四大類基因，即有關調控細胞週期、訊號傳遞、免疫反應以及轉錄調控因子選出50-100種基因，製成cDNA晶片，以便檢測具有HPV感染之肺癌患者是否亦有同樣的基因表現輿圖？這結果將可確定所選出之基因是否確實是HPV感染所致。另外，我們將進行轉染實驗 (transfection experiment)，將全長HPV DNA轉植入由初級培養所得之肺上皮細胞，並以transformation 和 tumorigenic assay，以了解HPV是否會造成肺上皮細胞腫瘤化？然後再以cDNA microarray分析其基因表現輿圖，以獲知HPV是否確實參與肺癌之形成。同時可由此獲知HPV引起肺癌的路徑是否和子宮頸癌不同？這些結果將有助於預防、診斷和治療HPV引起之肺癌形成。為了能了解HPV DNA嵌入肺癌細胞染色體引起腫瘤之機轉，我們將以restriction-ligated PCR和direct sequencing分析HPV感染之肺腫瘤組織中，HPV DNA嵌入在染色體之位置及其輿圖，同時此嵌入染色體輿圖將與以FISH分析HPV感染之肺癌細胞株的HPV DNA嵌入染色體輿圖做一比較是否一致？這結果不僅能證明HPV是否參與肺癌之形成，同時還能提供許多線索作為將來探討HPV引起隻人類其他肺癌的研究基礎。肺癌人類乳突瘤病毒cDNA 晶片KeywordsLung cancerMicroarray techniqueHPVLaser capture microdissectionProject