- 學位論文
草藥萃取物NTU04及MSL-UH抑制胰腺癌侵襲力以及機制之研究
Herbal extracts NTU04 and MSL-UH suppress pancreatic ductal adenocarcinoma cell invasion via inhibiting MMP-2/-9 activities and matriptase
摘要
胰腺癌是目前最惡性的癌症之一，病患之五年存活率僅達6%。胰腺癌高死亡率的主因來自其較易侵襲至淋巴或轉移至肝臟，進而導致高復發率的產生。然而，目前並沒有效的治療方法能夠去抑制胰腺癌侵襲以及轉移的發生。中草藥應用在治療各種疾病上已行之有年，其擁有低毒性及低副作用等特性，使之成為現今開發癌症藥物的一個新選擇。在先前的研究當中，我們從43種草藥萃取物當中鑑選出2種草藥 (NTU04及NTU43),發現其可以抑制肝癌細胞的侵襲力。在此篇研究當中，發現NTU04可以有效地抑制胰腺癌細胞之侵襲力，部分透過降低基質金屬蛋白酶 (MMP-2/-9) 的活性以及間質蛋白酶(matriptase)的表現及其活化。進一步的研究結果顯示，MMP-2/-9 的活性下降是由於NTU04 降低MMP-2的表現量，促進其金屬蛋白酶組織抑制因子-1 (TIMP-1)的表達或直接抑制其活性而導致。另外，透過減弱以及過量表達matriptase的方法，亦證明matriptase 確實參與在NTU04 抑制細胞侵襲的機制當中。為了找尋其中之有效成分，我們利用液相層析儀分離MSL-UH，結果顯示F3,F4含有效的成分抑制胰腺癌細胞侵襲力,在動物胰臟原位腫瘤模式的實驗當中，也看到其混和物有抑制腫瘤生長以及轉移的效果。再進ㄧ步分離F3及F4，發現F3-4, F3-5, F4-2 和F4-3相較於其它的子分層擁有較好的抑制活性。另外，我們也透過核磁共振光譜儀以及質譜分析儀鑑定出F3-4 中的主要組成為表兒茶素，但進一步分析顯示表兒茶素並非F3-4中抑制侵襲力之主要活性成份。綜和以上，NTU04 可透過抑制MMP-2/-9 以及matriptase 而達到抑制胰腺癌細胞侵襲力的效果。在一步步分離MSL-UH 之後，我們將找出可能含有抑制侵襲力之活性成分之子分層並進ㄧ步鑑選出有效成分。本篇研究顯示NTU04以及MSL-UH為具有治療轉移性胰腺癌潛力之藥物，期許未來更進ㄧ步的研究可使其成為治療胰腺癌的新選擇。
並列摘要
Pancreatic ductal adenocarcinoma (PDAC) is one of the deadliest malignancies with 6% of 5-year survival rates. The high mortality of PDAC is mainly due to high lymphatic invasion incidence, liver metastasis, and consequently high recurrence. Thus, suppression of PDAC cell invasion and metastasis will be a good strategy to slow down PDAC progression and reduce the mortality of this disease. However, there’s currently no effective therapy that can inhibit PDAC cell invasion and metastasis. Herbal extracts have been used in traditional medicine for a long time and shown with low cytotoxicity and side effects. Thus, herbal extracts may serve as good sources to isolate effective components for cancer therapy. In our previous study, from 43 herbal extracts, we found that two of them (NTU04 and NTU43) were able to inhibit HCC cell invasion with low cytotoxicity. In this study, NTU04 was found to effectively suppress PDAC cell invasion via reducing MMP-2/-9 activities, matriptase’s expression and activation. The data further showed that NTU04-reduced MMP-2/9 activities were at least via down-regulating MMP-2 expression, up-regulating tissue inhibitor of metalloprotease-1 (TIMP1) as well as by direct inhibition. On the other hand, matriptase was also proved to be involved in NTU04-inhibited cell invasion by matriptase knockdown and overexpression strategies. To further identify the effective components from the herbal extracts, we purified an herbal extract (MSL-UH) from a Taiwanese herbal species close to NTU04, and used chromatography to fractionate MSL-UH. The results showed that MSL-UH also could inhibit PDAC cell invasion and the subfractions (F3, F4) exhibited more inhibitory effects on PDAC cell invasion than the other fractions. In the orthotopic pancreatic tumor xenograft model, F3,4 mixture administration also showed to suppress tumor growth and liver metastasis. Furthermore, four subfractions (F3-4, F3-5, F4-2 and F4-3) of which were fractionated from F3 and F4 exhibited more inhibitory effects on PANC-1 cell invasion than the others. We also identified that epi-catechin is the major compound in F3-4 by NMR and LC-MS/MS. However, epi-catechin was apparently not the active compound in F3-4 for inhibiting PDAC cell invasion. These results together indicate that NTU04 and MSL-UH may exhibit some potential active components and candidate agents which will be ably against malignant metastatic PDAC cancers.