TRAIL是近年來新發現的致凋亡細胞激素，根據研究指出，TRAIL會選擇性的毒殺腫瘤細胞，而部會毒殺正常細胞。腫瘤細胞表現較多死亡接受子DR4和DR5可與TRAIL結合導致細胞發生凋亡 ; 干擾接受子DcR1和DcR2而使TRAIL無法被活化。近年來，癌症病人迅速的竄升，臨床上多以化療和電療法延長病人壽命，因而產生抗藥性和副作用等問題。因此我們想結合化療要與TRAIL的共同使用，增加對肺癌細胞的毒殺作用。在我們的研究中，首先以MTT assay、DAPI及PI染色法從十株人類非小型肺癌細胞細胞株(NSCLC)中篩選對TRAIL敏感極具抗性之細胞株，結果發現H460對TRAIL最敏感 ; 而H1299則對TRAIL最具有抗性。為了了解p53在TRAIL毒殺機制中所扮演的角色，我們利用 stable expression的方式使H1299表現wild type p53、H520表現dominant negative p53V143A ; 另外並以transient expression 的方式使Calu-1表現wild type p53、H460表現dominant negative p53V143A，研究發現p53在TRAIL毒殺機制中扮演著正調控的角色。我們選擇目前臨床上廣泛使用之化療要抗微管毒素paclitaxel、抗代謝要gemcitabine 及基化季cisplatin作為研究，結果發現任一化療藥可使細胞週期停化、激發p53及Bax，gemcitabine與cisplatin亦可促使p21的表達，並且會增加細胞表面DR4、DR5或TRAIL之表達。我們將TRAIL與化療藥物共同作用，以MTT assay 發現paclitaxel可使七株細胞產生對TRAIL之加成作用、gemcitabine可使六株細胞有加成反應、而cisplatin可使三株細胞有加成效果，同時我們以PI染色分析Sub-GI與DAPI染色觀察細胞型態上變化，進一步確認TRAIL與化療要的加成效果。最後，接種對TRAIL最敏感之H460肥癌細胞於裸鼠中，並以加成效果最好之paclitaxel與TRAIL共同治療，進一步觀察體內模式中加成治療的效果。因此，化療藥物和TRAIL的合併使用對NSCLC將可為未來一項新的治療方法。
TRAIL is a new proapoptotic cytokine. It is believed that TRAIL can selectively kill cancer cells but not normal cells. TRAIL induces apoptosis through binding its two proapoptotic receptors, DR4 and DR5. The TRAIL signaling pathway is modulated by two antiapoptotic TRAIL decoy receptors, DcR1 and DcR2. Lung cancer is the leading cause of cancer deaths in taiwan. The nunber of new case of lung cancer is still increasing. About eighty percent of all diagnosed lung cancers are non-small cell lung cancer (NSCLC). Current protocols for therapy of NSCLC include surgery, radiotherapy and chemotherapy. Intensive chemotherapy regimens using cytotoxic agents do effectively kill lung cancer cells. However, lung cancer cells are often resistant to chemotherapy and even develpo acquired chemoresistance or show multi-drug resistance as aconsequence of the previous treatment. Thus development of chemoresistance remains a major obstacle for curable lung cancer tratment. The aim of our study was to determine whether NSCLC can be sensitized by chemotherapeutic agents for TRAIL-induced apoptosis. The sensitivity of NSCLC to TRAIL-induced apoptosis was assessed by MTT assay, DAPI assay and PI staining. Our results show that H460 is most sensitive to TRAIL ; H1299 is most resistance to TRAIL. In order to investigate the role of p53 in TRAIL-induced apoptosis, the plasmids containing wild-type p53 gene were transfected into H1299 and Calu-1 cells ; also the olasmids containing mutated (p53 gene p53V143A) were transfected into H520 and H460 cells. The results show that p53 can up-regulate TRAIL-induced apoptosis in NSCLC cell lines. In this study, we used cisplatin, gemcitabine and paclitaxel as chemotherapeutic agents to investigate their effects on NSCLC cell lines. We found any one of these drugs can induce p53 andBax expression in NSCLC cell lines. Moreover, all og these drugs can induce cell surface DR4, DR5 and TRAIL expression. In the synergistics effects of chemotherapyeutic agents and TRAIL-induced apoptosis in NSCLC cell lines, paclitaxel rnhances cytotoxic effect of TRAIL in 7 to 10 cell lines. Gemcitabine and TRAIL are synergistic in 6 of 10 NSCLC cell lines. Cisplatin and TRAIL are synergistic in 3 of 10 NSCLC cell lines. Finally in vivo administration of chemotherapeutic drugs and TRAIL substantially inhibited subcutaneous tumor growth of H460 cells. Our results suggest that TRAIL is a promising anti-tumor cytokine against human NSCLC.
