The Effects of Iodoacetic acid-Induced Notch3 reduction in Human Lung Cancer Cells

根據衛生署2008 年的統計，惡性腫瘤自民國71 年起為國人死因之榜 首，而其中肺癌居癌症死因之首位。肺癌病人平均存活時間約九至十二個 月，並且整體五年存活時間約百分之十。80％ 病人在手術後兩年內會 發生癌症轉移導致死亡，少於20%病人能接受根除性手術性治 療。因此， 尋找更有效的藥物治療而且改善存活率對於肺癌病 人是必要且急需的。由之前研究結果顯示，碘乙酸(IAA)的處理 下會釋放cytochrome C 造成活化caspase 3，造成人類肺癌細胞 株的細胞核濃染之細胞凋亡， 因此可能為一個有潛力的治療藥 物。在本篇實驗中， 我們著重探討碘乙酸造成肺癌細胞死亡可 能的分子機制。研究結果顯示碘乙酸會造成藥效與劑量成正相 關的細胞死亡， 且anchorage dependent 和anchorage independent assay 的結果顯示碘乙酸會抑制肺癌細胞株的移動(migration)，而且大部分肺 癌細胞株此反應會被DTT 或NAC 抑制， 其中包括A549、 H1299、HOP62、H23 、EKVX、PC14、H1975、H441 、H460 等肺癌細胞株。根據碘乙酸microarray 結果，Notch3 基因為碘 乙酸處理下的目標基因之ㄧ 。而從RT-PCR 結果得知， 碘乙酸 處理16 或24 小時後，可造成大部份的肺癌細胞株Notch3 基因 以及其下游目標基因PbX1 mRNA dose-dependent 的下降，且此 反應亦可被DTT 或NAC 所恢復。综合上述結果，Notch3 和其 下游目標基因PbX1 可能參與在碘乙酸所造成的細胞凋亡訊息 傳遞中， 因此在肺癌治療中可能為一新且有潛力的治療策略。Lung cancer is one of the tenth leading causes of death in Taiwan. The average survival duration for patients with lung cancer is about 9-12 months and overall 5-year survival rate is about 10%. There are less than 20% lung cancer patients able to undergo curative resection. 80% of lung cancer patients undergo surgery died of metastasis within two years. More effective treatments for this devastating disease are urgently needed. Based on our previous findings, IAA induced cytochrome C release resulting in activation of caspase-3 and nuclear condensation in non-small cell lung cancer (NSCLC). In this study, we investigate possible molecular mechanism(s) of how IAA induced cell death in human lung cancer cell lines. Our results showed that IAA induced dose-dependent cell death as well as inhibited migration in anchorage dependent and anchorage independent assays. These responses could be reversed by dithiothreitol (DTT) or N-acetyl-L cystein (NAC) in most tested lung cancer cell lines, such as A549, H1299, H522, HOP62, H23, EKVX, PC14, H1975, H441 and H460. Though RT-PCR and microarray analysis, Notch-3 can be confirmed as the target of IAA. Pbx-1 is predicted as downstream oncogene of Notch-3. Cells treated with IAA for 16 or 24 hours, RT-PCR assay showed Pbx-1 mRNA suppression. However, this symptom can be reversed by DTT or NAC. As a conclusion, Notch-3 and it’s downstream, Pbx-1, involved in IAA-induced cell death, which suggesting that inactivation of Notch-3 or Pbx-1 may be a potential therapy for lung cancer