2026-11-092026-11-09https://scholars.csmu.edu.tw/handle/123456789/38373Lung cancer is the leading cause of cancer death in Taiwanese women since two decades. However, over 90 and 70% of female lung cancer is nonsmokers and adenocarcinoma (ADC). Based on pathological observation in our cases, acinar and papillary subtypes were more frequently found in nonsmoking female ADC, and bronchioloalveolar carcinoma was rarely observed. Most of female cases were relatively younger than male, and female had a poor prognosis due to difficult early detection. Unfortunately, the etiological factors were still unknown although some epidemiological reports indicated that some factors including exposure of cooking oil fumes, chronic pulmonary diseases, and a short menstrual cycle were associated with lung cancer development of Chinese women including Taiwan. Therefore, it is necessary to investigate the etiological factor of nonsmoking female ADC for the prevention, early detection and treatment of this disease. In our previous studies, a high prevalence of HPV 16/18 infections was found in nonsmoking female lung cancer in Taiwan, not in male cases and the detection of HPV DNA in blood circulation may act as a prominent risk marker of lung cancer in Taiwan. cDNA microarray data from our previous projects indicated that cell-cycle related- and immune related-gene expression profiles were quite similar with those of cervical keritinocytes transformed by transfection of HPV 16 E6/E7 and HPV 16 E6/E7 transfected lung ADC CL-3 cell. Additionally, RS-PCR results showed that HPV 16/18 DNA was indeed integrated into chromosome 1, 12, 17, 18 and X in HPV infected lung tumor tissues. These results seem to suggest that HPV may play a certain extent in lung ADC development of Taiwanese women nonsmokers. Due to HPV DNA integration seems to be random in HPV infected lung cancer. In this project, genome-wide analysis will be performed to investigate the potential loci occurred loss of heterozygosity (LOH) in whole genome from comparison between HPV infected and non-infected lung tumors with the same ADC subtypes. The potential tumor suppressor genes (TSGs) involved in HPV associated lung tumorigenesis will be found in the LOH loci. For the known TSGs, RT-PCR and immunohistochemistry will be used to confirm the gene inactivation through LOH in nonsmoking female ADC tumor samples. For unkown TSGs, two-step total gene synthesis method will be performed to identify the novel TSGs involved in HPV infected lung tumorigenesis. Moreover, the specific antibody will be developed for detection of novel TSGs to confirm LOH results. The LOH results from this project will be compared with the data of cDNA microarray and RS-PCR from our present project and hypermethylation profile data from another project which is being under reviewed. The data from genome, transcription, and protein levels will be helpful to verify which LOH may play a crucial role in HPV infected lung tumorigenesis. These results will be not only to reveal the potential tumorgenic pathway from these candidate TSGs but also to better understand the molecular pathogenesis of HPV-associated lung ADC among nonsmokers. These results will provide efficient strategies in prevention, early detection, and clinical treatment in lung ADC, especially in HPV infected lung ADC tumorigenesis.肺癌多年來一直是台灣女性之第一大癌症死因。尤其是台灣女性肺癌患者又大多為不抽煙者。在肺癌病理分類上主要是肺腺癌，女性肺癌又以acinar和papillary subtypes為主。女性被發現罹患肺癌時，大多已發生腫瘤轉移，其預後都較差。雖然過去有些流行病學報告指出二手煙，油煙暴露以及慢性肺疾病與台灣女性肺癌發生有關，但其致病因子大多不清楚。事實上，肺腺癌之致病因子就不清楚，尤其是不抽煙之肺腺癌。本研究室發現高危險人類乳突病毒16/18型 (human papillomavirus, HPV 16/18)感染與台灣女性肺癌發生有關，又血液循環中有HPV DNA者罹患肺癌之為顯示沒有者之75倍。這些結果顯示HPV有可能與台灣女性肺癌形成有關。過去研究已知， HPV 16/18 引起人類腫瘤之主要分子機轉是透過研究子宮頸癌而獲得。一是經由E6和E7致癌蛋白將p53與Pb 蛋白分解，使細胞增生而腫瘤化。另外一是HPV DNA會嵌入宿主染色體之common fragile sites (CFS)，而引起染色體不穩定而發生LOH，終致引起腫瘤。在先前計畫之cDNA microarray與RS-PCR的結果已獲知，HPV感染之肺腫瘤參與細胞週期與DNA合成以及免疫相關之基因表現輿圖與E6與E7 transformed keritinocytes以及肺癌細胞轉殖 E6, E7之表現輿圖相似。這顯示HPV感染確實會改變肺腫瘤細胞基因之表現。RS-PCR結果發現HPV 16/18 DNA會嵌入到染色體1, 12, 17, 18和X中，顯示HPV DNA 嵌入到肺腫瘤細胞可能是隨機性的，因此欲找到HPV嵌入引起之染色體LOH，可能需要以genome-wide全面篩檢方式，較能找到參與HPV感染引起之肺腫瘤化的TSGs。本計畫擬genomic-wide 分析人類乳突病毒感染之不抽菸女性肺腺癌主要發生異質性基因座缺失 (loss of heterozygosity, LOH)之染色體區域，並希望能經由發生LOH位置，找到並確定主要參與之已知與未知之抑癌基因 (tumor suppressor genes, TSGs)。本計畫擬由病理專家確定HPV有或沒有感染之女性肺腺癌之主要subtypes做研究標本，以LCM將腫瘤細胞取下萃取純化出DNA，另外以該病患之週邊血液淋巴球做正常組DNA，用來進行Genome-wide 分析以找出HPV感染之肺腺癌較常發生LOH之染色體區域，並推測可能之已知TSGs。然後以RT-PCR以及免疫組織化學分析法 (immunohistochemistry)了解不抽煙女性肺腫瘤中，該基因在mRNA和蛋白層次是否確實沒有表達？若LOH之區域為未知之TSGs則將以two-step total gene synthesis method找出並確定該未知TSGs，當然這位之基因將同樣以RT-PCR方法了解其mRNA是否沒有表現？同時將發展其抗體，做免疫組織化學分析，已確定其蛋白之表現與否？因此本計畫將在染色體層次全面分析HPV感染有關之染色體不穩定區域，以便與過去cDNA microarray和RS-PCR5 之結果以及現在正在申請中之 HPV感染肺腺癌之hypermethylation profiles 研究計畫之結果分別在基因，mRNA以及蛋白等三個層次做一較完整之比較分析，以了解那些染色體區域之LOH所造成之TSGs去活化，才最有可能參與HPV感染之肺腺癌形成。將這些TSGs串聯起來，即可推測出HPV感染之肺腺癌主要是經由何種腫瘤化路徑形成肺腺癌。因此本計畫不僅能了解不抽煙肺腺癌形成之可能腫瘤化路徑，以及HPV感染引起之肺腺癌之腫瘤化路徑是否不同於未感染者，這些結果將有助於早期診斷不抽煙之肺腺癌，以預防並減少肺腺癌之發生。肺腺癌人類乳突病毒異質性基因座缺失女性Lung adenocarcinomaHuman papillomavirusLoss of heterozygosityfemaleProject