自民國七十一年以來，肺癌一直是台灣女性的第一大死亡原因，已知抽煙是引起肺癌的主要病因。但是台灣女性肺癌則不到10%的患者可歸罪於抽菸，因此其他環境因子可能參與肺癌的形成。本研究室之過去研究發現肺癌患者之肺腫瘤中，有高達35.5%和41.1%的人類乳突病毒16和18型(Human papillomarvirus 16/18)的感染率。同時亦發現肺癌患者之肺腫瘤組織及周邊血液之HPV感染有70%的一致性。在原位雜交試驗中發現HPV DNA存在於血液淋巴球中，因此推測肺腫瘤組織中之HPV DNA可能來自血液循環。本計畫利用巢疊式PCR及DNA自動定序方式分析肺腫瘤組織中測得之HPV 16/18之E6、E7及L1的DNA序列，並與血液中所測得之HPV DNA序列進行比對，結果發現在10位肺腫瘤組織及血液均可測到HPV 16的肺癌患者中有4位其腫瘤組織中的HPV DNA序列與血液中不同，但序列之差異性小於2%，因此仍屬於同一亞型。而在11位肺腫瘤組織及血液均可測到HPV 18的肺癌患者中有5位其腫瘤組織中的HPV DNA序列與血液中不同，僅有一位序列之差異性大於2%，屬於不同亞型其餘4位之序列差異性均小於2%。因此推測肺腫瘤組織中之HPV 16或18與血液中之HPV可能同源。本計畫進一步以原位雜交技術(In situ hybridization; ISH)及DNA定序分析等方法分析原發性肺腫瘤以及其轉移其他器官之腫瘤組織中之HPV感染情形及同源性，結果發現兩者之HPV DNA序列具有一致性，並由原位反轉錄聚合脢連鎖反應，分析其HPV 16/18E6/E7 mRNA的表現，結果發現不論在原發性肺腫瘤或其轉移至其他器官之腫瘤組織中均可測到HPV 16/18 E6/E7 mRNA的表現，兩者具有一致性。綜合以上結果，我們推測肺腫瘤組織中之HPV可能來自血液循環且HPV 16/18E6/E7 mRNA的表現可能會增強肺腫瘤轉移之能力。 Lung cancer is the leading and second cause of cancer death in female since 1982 in Taiwan. Intriguingly, only around 10% of female lung cancer incidence in Taiwan was related to cigarette smoking. Thus, it is conceivable that environmental factors other than cigarette smoking may be associated with lung cancer development in Taiwan. The high prevalence of HPV 16/18 infections in Taiwanese lung cancer patients was reported in our recent study. Additionally, there is an approximately 70% concordance between HPV DNA detection in peripheral blood cells and lung rumor tissues of lung cancer patients. Our preliminary in situ hybridization data showed the peripheral blood leukocytes to possess HPV DNA signal. Thus, in this study, we hypothesize that HPV DNA in lung tumors may be transmitted from leukocytes in blood circulation. Nested-PCR and autosequencing were used to compared the sequence variant in L1、E6 and E7 of HPV 16/18 in lung tumors and that of leukocytes in blood circulation. Sequence variants were found in 4 of 10 lung cancer patients with present the HPV 16 signal both in tumor tissues and blood circulatuion, but the variant were <2% and classified the same subtype. Sequence variants were also found in 5 of 11 lung cancer patients with present the HPV18 signal both in tumor tissues and blood circulatuion, only I of 5 were >2%. Thus we suggested that the HPV 16/18 infection in lung tumor tissues may originate from blood circulation. To elucidate the HPV infection was involved in lung tumorigenesis, HPV DNA including L1, E6, E7 in the metastatic tumors in other organs and their primary lung tumors were detected by in situ hybridization. The DNA sequence of L1, E6, and E7 in lung tumors and metastatic tumors also analyzed and compared to understand whether HPV is the same typing. HPV 16/18 E7/E7 mRNA were also detected by in situ RT-PCR and their relationships with clinical parameters were statistically analyzed to evaluate whether HPV has an impact on the metastatic ability of the lung tumor cells will be induced by HPV. In situ hybridization data were observed the primary lung tumors and their metastatic tumors were co-infections by HPV and the HPV sequences were not different. These results suggest that HPV 16/18 infection in lung tumor may be originate from blood circulation and may be involved in lung tumorgenesis.
