探討植物粗萃物對抗藥性惡性腦瘤之抑癌作用

多形性神經膠質母細胞瘤是最常見的星狀細胞瘤，占所有成人神經系統腫瘤的比例約為65％，其特性為侵襲性和惡性度極高。多形性神經膠質母細胞瘤之患者術後存活率極低，僅具有9.9~10.2個月，而整體平均存活時間僅為12~15個月。儘管結合手術治療和標準化療藥物TMZ並合併放射治療，但多形性神經膠質母細胞瘤仍然經常形成抗藥性。因此，找到一種能抑制具抗藥性之多形性神經膠質母細胞瘤的新藥是極具重要性的。本篇論文的目的是探討植物粗萃物於體外對於具抗藥性之多形性神經膠質母細胞瘤之抑癌作用及機轉。首先，我們建立了DBTRG-05MG-TMZ抗藥性細胞株，透過細胞存活率試驗證實，抗藥性細胞株對於TMZ的IC50濃度相較於DBTRG-05MG細胞多了三倍。接著透過免疫螢光染色法和西方點墨法，觀察DBTRG-05MG-TMZ抗藥性細胞株的CD133，MDR1和MGMT等抗藥性標誌之表達，發現於DBTRG-05MG-TMZ抗藥性細胞株中皆有高度的表現。之後，利用抗藥性細胞株以細胞存活率試驗篩選出兩種純化物PAT和Cedrol，其分別來自於廣藿香（Pogostemon patchouli, PPa）和雪松（Cedrus atlantica, CAt），且對於腫瘤具有良好的毒殺效果。PAT和Cedrol可誘導細胞週期運行停滯於G0 / G1 phase，並使Sub-G1 phase的百分比呈現增加的趨勢。另外，PAT和Cedrol也會同時活化外在和內在細胞凋亡途徑，並抑制抗藥性相關蛋白及體外多形性神經膠質母細胞瘤之轉移和血管生成。綜上所述結果，PAT和Cedrol在未來具有高度的潛力，能開發為治療具抗藥性之多形性神經膠質母細胞瘤之抗癌藥物。多形性神經膠質母細胞瘤是最常見的星狀細胞瘤，占所有成人神經系統腫瘤的比例約為65％，其特性為侵襲性和惡性度極高。多形性神經膠質母細胞瘤之患者術後存活率極低，僅具有9.9~10.2個月，而整體平均存活時間僅為12~15個月。儘管結合手術治療和標準化療藥物TMZ並合併放射治療，但多形性神經膠質母細胞瘤仍然經常形成抗藥性。因此，找到一種能抑制具抗藥性之多形性神經膠質母細胞瘤的新藥是極具重要性的。本篇論文的目的是探討植物粗萃物於體外對於具抗藥性之多形性神經膠質母細胞瘤之抑癌作用及機轉。首先，我們建立了DBTRG-05MG-TMZ抗藥性細胞株，透過細胞存活率試驗證實，抗藥性細胞株對於TMZ的IC50濃度相較於DBTRG-05MG細胞多了三倍。接著透過免疫螢光染色法和西方點墨法，觀察DBTRG-05MG-TMZ抗藥性細胞株的CD133，MDR1和MGMT等抗藥性標誌之表達，發現於DBTRG-05MG-TMZ抗藥性細胞株中皆有高度的表現。之後，利用抗藥性細胞株以細胞存活率試驗篩選出兩種純化物PAT和Cedrol，其分別來自於廣藿香（Pogostemon patchouli, PPa）和雪松（Cedrus atlantica, CAt），且對於腫瘤具有良好的毒殺效果。PAT和Cedrol可誘導細胞週期運行停滯於G0 / G1 phase，並使Sub-G1 phase的百分比呈現增加的趨勢。另外，PAT和Cedrol也會同時活化外在和內在細胞凋亡途徑，並抑制抗藥性相關蛋白及體外多形性神經膠質母細胞瘤之轉移和血管生成。綜上所述結果，PAT和Cedrol在未來具有高度的潛力，能開發為治療具抗藥性之多形性神經膠質母細胞瘤之抗癌藥物。

Glioblastoma multiforme (GBM) is the most common astrocytic tumor and represents about 65% of all adult nervous system tumors, which is characterized by a high aggressiveness and malignant. GBM patients with a poor survival time which are 9.9 to 10.2 months after surgery and an average survival time only 12 to 15 months. Despite combining surgical techniques and temozolomide (TMZ) standard chemotherapy plus radiotherapy for GBM, which always induced drug resistance. Therefore, it is important to find out a novel drug for against drug resistant GBM tumor. The purpose of this study is to investigate the anti-tumor effects and mechanism of plant extracts in resistant GBM tumor in vitro. First, we established DBTRG-05MG-TMZ resistant cell line, which was confirmed by MTT assay with higher IC50 concentration of TMZ for three times dose than DBTRG-05MG cell. And we verified drug resistant markers such as CD133, MDR1 and major marker MGMT in DBTRG-05MG-TMZ resistant cell line via Immunofluorescence assay and western blot, then we got high level MGMT protein, which is a major GBM drug resistant marker, in DBTRG-05MG-TMZ resistant cell line. After that, we utilized this resistant cell line to establish drug screening platform, and two pure compounds , which are PAT and Cedrol that isolated from Pogostemon patchouli (PPa) and Cedrus atlantica (CAt), with higher cytotoxicity for tumor were selected by MTT assay. PAT and Cedrol could induce cell cycle arrest at G0/G1 phase and enhance percentage of Sub-G1 phase. Moreover, the extrinsic and intrinsic cell apoptosis pathways were activated and drug resistant related proteins were inhibited by PAT and Cedrol treatment. In addition, PAT and Cedrol also inhibited metastasis and angiogenesis in vitro. All of above, this study showed the PAT and Cedrol have highly potential to develop an anti-tumor agents or adjuvants for GBM drug resistant therapy in the future.