- 學位論文
研究維生素B12及葉酸對肺癌化療藥物(愛寧達)藥效之影響
Study the effect of vitamin B12 and folic acid to Pemetrexed (Alimta) efficacy with human lung cancer cells
摘要
肺癌是全世界造成嚴重癌症死亡的原因，為ㄧ種肺部組織細胞不正常增生的疾病。主要可分為小細胞肺癌 (SCLC) 及非小細胞肺癌 (NSCLC) 兩大類。隨著近年來治療的發展，肺癌仍是無法治癒的疾病。提高病人的存活率並降低藥物毒性，一直是發展新的治療方向。因此，在研究中我們評估非小細胞肺癌化療藥物 Pemetrexed (愛寧達)同時添加維生素B12 及葉酸下提供的抗腫瘤效果。非小細胞肺癌細胞株在不同血清濃度環境下對 Pemetrexed 感受性皆不同，在10 % 血清中敏感性較差。因此，添加維生素B12 或葉酸與 Pemetrexed 共同處理後，結果顯示可以增加細胞生長被抑制的效果，並隨添加劑量增加而加強。觀察三種不同的人類非小細胞肺癌細胞株都有此相同結果。由於在低血清濃度 (2%) 下細胞對藥物敏感性佳，則維生素B12 和葉酸加強效果消失。更進一步利用流式細胞儀分析發現添加維生素B12 或葉酸能提高Pemetrexed誘導細胞凋亡的情形增加。同樣地，在其它肺癌化療藥物中添加維生素B12 或葉酸，並無增強藥效的能力，除了Methotrexate之外，顯示添加維生素B12 可加強藥效。然而，Pemetrexed可經由多重目標的調控誘導細胞凋亡，我們主要檢查可能的蛋白改變，像是 thymidylate synthase (TS)、p53 和 p21 蛋白等。結果顯示在 A549 和 H460 細胞株 (wt-p53) 中，處理 Pemetrexed 或與維生素 B12 和葉酸同時合併後會提高 p53 和其下游 p21 蛋白表現量。在 H1299 (p53-null) 細胞中，發現 Pemetrexed 不需依賴 p53 基因而誘導 p21 蛋白的表現，於三者合併處理後表現量較高。結果顯示p53基因會參與細胞凋亡，但不是主要受調控的路徑。結果發現 p21 蛋白表現的差異性較顯著，可能是經由兩條不同路徑所調控，在未來可以繼續深入探討的方向。從我們的研究發現，添加維生素B12 和/或葉酸與Pemetrexed同時使用不僅不會降低藥效而且能提供更佳的抗腫瘤效果。
並列摘要
Lung cancer is a seriously cause of cancer deaths in the worldwide, and that is a disease of uncontrolled cell growth in tissues of the lung. The main types of lung cancer are small cell lung carcinoma (SCLC) and non-small cell lung carcinoma (NSCLC). Despite recent advances in the treatment, lung cancer remains an incurable disease. This calls for an effort to develop novel therapeutics in order to increase survival of patients and decrease the tocixity of drug. Here, we evaluated the potential antitumor effect of Pemetrexed (a NSCLC chemotherapy drug) with supplementated vitamin B12 or folic acid to treat the cells. We found that NSCLC cell lines have different sensitivity for Pemetrexed treatment in various serum concentration. Pemetrexed was more resistant to the cells in 10% serum. Furthermore, we also demonstrated that cells growth inhibition of Pemetrexed treatmented cells can be increased through vitamin supplementation (folic acid and vitamin B12) with three NSCLC cell lines tested. Under 2% serum conditions, the sensitization ability by vitamin B12 and folic acid supplementation was not observed. Our data also showed that induction of apoptosis by Pemetrexed treatment can be increased with vitamin B12 or/and folic acid supplementation. The sensitization effect is specific for Pemetrexed, since there is no other chemotherapy drug efficacy can be inproved by addition of vitamin B12 or folic acid. However, Pemetrexed can induce apoptosis by the regulation of multitarget genes expression. Therefore, we checked the main protein expression, such as thymidylate synthase (TS)、p53 and p21 protein. In A549 and H460 cells with wild type of 53, up-regulation of p53 and its down stream regulated p21 protein were demonstrated by Pemetrexed treatment with or without vitamin B12 and folic acid. In H1299 cells with null type of p53, p21 protein is still induced under same treatment as A549 cells. Our data indicate that regulation of p21 expression by Pemetrexed may via two different pathways. In this study, we found vitamin B12 and folic acid supplementation is increasing but not decreasing the Pemetrexed efficacy of antitumor effect of NSCLC cells.