- 學位論文
探討ALIMTA在非小細胞肺癌之分子預測指標
Molecular Predictors of Response to ALIMTA in Non-Small Cell Lung Cancer
摘要
Pemetrexed disodium (ALIMTA®)是一種多重抗葉酸藥物，其主要可抑制腫瘤 細胞中嘌呤與嘧啶之合成，目前已做為治療非小細胞肺癌(non-small cell lung cancer, NSCLC)的一線或二線藥物。然而，目前缺乏臨床生物指標能作為評估其藥效反應之標記。本研究使用八株人類NSCLC細胞株來評估對於ALIMTA具有抗藥性與敏感性及其細胞生長與轉移分子的表現。MTT細胞毒性結果顯示，A549、TL-6、P014與P015細胞對於ALIMTA是較具為敏感性， H1975、H1650、HCC827及H1355細胞則較具抗藥性。以西方墨點法分析NSCLC細胞之內生性蛋白差異，敏感性的肺癌細胞除了A549外，其LCN2和E-cadherin的表現量皆低於具有抗藥性的細胞株。然而，A549與TL-6細胞在添加EGF，再處理ALIMTA後，其細胞存活率並不會因而提升。A549與TL-6細胞在處理ALIMTA後，其cyclin A、 pRB與pEGFR表現量皆有下降，而其LCN2、p-cyclin D、p53與p21則隨著ALIMTA劑量的提高而表現量增加。相反地，H1650與H1975細胞之cyclin A與pRB表現量有微幅上升的情形，p53、p21與LCN2則無變化，p-cyclin D的表現量則微幅下降。進一步地分析對細胞週期影響，不論是對ALIMTA敏感或是具抗藥性的肺癌細胞，ALIMTA皆造成細胞週期之S期的累積。當ALIMTA在低濃度(0.08 μM)時，即能夠有效抑制A549、TL-6和H1650細胞的移行與侵襲能力。最後，以 ELISA偵測45位NSCLC患者在接受ALIMTA治療前及治療後第一次回診抽血中LCN2的濃度，結果顯示，病患在接受ALIMTA第四週期治療即無效者，其LCN2濃度會下降，但可使用ALIMTA達四週期以上有反應療效者，則LCN2濃度則會上升。因此，ALIMTA可影響肺癌細胞之細胞週期變化及轉移能力，並且LCN2變化可以在肺癌中作為對於ALIMTA反應之預測指標。
並列摘要
Pemetrexed disodium (ALIMTA®) is a multiantifolate drug, which is able to inhibit the synthesis of purines and pyrimidines in tumor cells. ALIMTA has been now used as first- and second-line regimems for therapy of non-small cell lung cancer (NSCLC). However, there is still a lack of clinical biomarkers for predicting the therapeutic response to ALIMTA. In this study, eight human NSCLC cell lines were used to evaluate the sensitivity and resistance to ALIMTA, and the molecular expression of cellular growth and metastasis. Results of the cytotoxicity from MTT assay showed that A549, TL-6, P014, H1355 and P015 cells were sensitive to ALIMTA, but H1975, H1650 and HCC827 cells were resistant. Using Western blot analysis to detect endoqenious protein expression of NSCLC cells, sensitive cells exhibited lower expression of LCN2 and E-cadherin than resistant cells except A549. However, when A549 and TL-6 cells were treated with ALIMTA after EGF addition, their cytotoxicities were not elevated. Expression of cyclin A and pRB were slightly increased, and p53, p21 and LCN2 were not altered; The expression of p-cyclin D was slightly decreased, following increased ALIMTA doses in H1650 and H1975 cells. Futhermore, S-phase of cell cycle was accumulated under ALIMTA treatment in either sensitive or resistant lung cancer cells. The ability of cell migration and invasion were inhibited with low dose (0.08 uM) of ALIMTA in A549, TL-6, H1650 and H1975 cells. Lastly, plasma LCN2 level was analyzed in 45 NSCLC patients by ELISA before and after ALIMTA treatment. Results revealed that LCN2 were decreased in patients who had a progression disease after receiving ≤ 4 cycles of ALIMTA treatment. Therefore, ALIMTA could attect the alteration of cell cycle and the capacity of metastasis for NSCLC cells, and alteration of LCN2 may serve as potential predictor of responsiveness to ALIMTA in lung cancer.
參考文獻
延伸閱讀
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