K-ras致活性突變在非小細胞肺癌的基因群表現模式

Gene Expression Profiles of K-ras Activated Mutation in Non-Small Cell Lung Cancer

王程遠(Cheng-Yuan Wang)

指導教授 ： 黃明賢

共同指導教授 ： 林?茹

高雄醫學大學/醫學院/醫學研究所/碩士(2006年)

摘要

文獻報導與肺癌病人預後有關並且會影響用藥選擇的分子生物標記有三種：(1) RAS家族的變異(2)上皮成長因子接受器(epidermal growth factor receptor, EGFR)基因的變異(3)是否擁有神經內分泌分化的標記。再者抽煙人口逐年上升，肺癌發生率亦逐年上升，且K-ras突變與抽煙強烈相關。K-ras突變者對於放射線治療、化療、標的療法皆反應不好，且預後較野生型K-ras為差，可見K-ras突變在肺癌拌演日漸重要的角色。本研究的目的在於探討使用基因陣列尼膜片取代傳統定序方式來偵測肺癌病人K-ras是否突變的可行性，及K-ras突變在肺癌造成的特異性活化基因群及其與臨床資料的關係。本研究共收集30組成對的肺癌病人腫瘤組織及血液檢體，以直接核酸定序結果發現30個肺癌病人腫瘤組織共有11個有K-ras突變，突變率為36.7%，突變的位置集中在譯碼12及13。其中譯碼12佔所有突變點的81.9%，譯碼13佔18.1%。以基因陣列尼龍膜片偵測血液中K-ras致癌基因活化情形也發現有11個檢體呈陽性反應，與腫瘤組織直接定序的結果比較,總計其敏感度為81.8%。特異度為89.5%，精確度為86.7%，一致性kappa值為0.713。男性肺癌K-ras突變率為21.4%，女性肺癌K-ras突變率為50%；初期肺癌(第一期+第二期) K-ras突變率為21.4%，末期肺癌(第三期及第四期) K-ras突變率為50%。可看出K-ras突變在女性及末期肺癌較常見的趨勢，但未達統計學上顯著的差別。基因陣列尼龍膜片陽性率在男性肺癌為21.4%，在女性肺癌為50%；在初期肺癌(第一期及第二期)為28.6%，在末期肺癌(第三期及第四期)為43.8%。可看出基因陣列尼龍膜片陽性率在女性及末期肺癌有較常見的趨勢，但未達統計學上顯著的差別。TBX19的過度表現在初期肺癌(第一期及第二期)為7.1%，在末期肺癌(第三期及第四期)為43.8%，在統計上有顯著的差別(p <0.05)。分析個別基因與K-ras突變的關係發現有四個基因的過度表現在K-ras突變型及K-ras野生型有顯著的差異：BLC2(p <0.01)，E2F4(p <0.05)，MMP1(p <0.05)，TBX19(p <0.01)。提供吾人往後深入研究K-ras突變在肺癌致癌路徑拌演的角色的方向。

關鍵字

非小細胞肺癌 ； k-ras突變 ； 基因群表現模式

並列摘要

The biomarkers of lung cancer that can significantly affect the prognosis of the patients and the effectiveness of anti-cancer drug therapy are (1) K-ras mutation (2) EGFR mutation (3) biomarkers of neuroendocrine features. From literature review, we know that the prevalence of lung cancer is increasing, the prevalence of smoking is increasing too, and K-ras mutation is significantly correlated with the smoking status. So the role of K-ras mutation is lung cancer carcinogenesis is becoming more and more significant. Our study focused on the feasibility of predicting K-ras mutational status by analyzing the gene expression profile of peripheral blood by membrane array, and the roles of K-ras mutation in lung cancer carcinogenesis. We sieve out 28 genes that most related to the mutated K-ras associated carcinogenetic pathway by means of comparative genome hybridization and bioinformatics. Then we constructed the membrane array including these 28 genes. We collected 30 coupled lung cancer tumor tissue and peripheral blood samples. By direct sequencing of the tumor tissue, the K-ras mutation rate is 36.7% (11/30), the hot spots of K-ras mutation are codon 12 (81.9%) and codon 13 (18.1%). There are 11 patients have positive results on membrane array on peripheral blood analysis, compared with the results of tumor tissue direct sequencing, the sensitivity, specificity and accuracy of the membrane array are 81.8%, 89.5%, 86.7% respectively. The kappa statistic is 0.713, revealed a good correlation between membrane array and direct sequencing. The K-ras mutation rate of the male and the female are 21.4% and 50% respectively. The K-ras mutation rate in early stage(stage I+II) and late stage lung cancer(stage III+IV) are 21.4% and 50% respectively. The membrane array positive rate of the male and female are 21.4% and 50% respectively, the membrane array positive rate in early stage and late stage lung cancer are 28.6% and 43.8% respectively. The K-ras mutation rate and membrane array positive rate is higher in women and in late stage lung cancer, but not statistically significant. TBX19 is over-expressed more commonly in late stage lung cancer (43.8%) than in early stage lung cancer (7.1%), and is statistically significant (p<0.05). Besides, we find out 4 genes that their over-expression is significantly correlated with K-ras mutation. These four genes include BCL2 (p<0.001), E2F4 (p<0.001), MMP1 (p<0.05), TBX19 (p<0.001). The next step of our study is to clarify the relationship between the four genes and the mutated K-ras in lung cancer carcinogenesis.

並列關鍵字

K-ras mutation ； non-small cell lung cancer ； gene expression profiles

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