鑑選具抑制肝癌細胞侵襲與轉移有效的草藥萃取物

Identification of effective herbal extracts on inhibiting liver cancer cell invasion and metastasis

羅宇辰(Yu-Chen Luo)

指導教授 ： 李明學

國立臺灣大學/醫學院/生物化學暨分子生物學研究所/碩士(2014年)

https://doi.org/10.6342/NTU.2014.01999

摘要

肝癌是目前最惡性的癌症之一且其癌症致死率在台灣甚至是全世界都高居第三位
。肝癌高致死率主要原因是由於治療後的高復發率以及其本身高轉移能力。然而，目前尚無有效抑制肝癌轉移的藥物。因此，找尋或開發具高效能且低副作用的藥物是目前在治療肝癌上一個重要的議題。在日本、中國以及台灣，由於草藥本身低副作用且低毒性，已被廣泛的使用在治療不同疾病，或作為保健食品。所以，在治療癌症方面草藥一直被認為是最具潛力的藥物來源。在此研究結果中，發現NTU04以及MSL-UH這兩種草藥萃取物在低濃度的狀態下就能有效地抑制肝癌細胞的侵襲力。在明膠基質金屬蛋白酶活性測試 (Gelatin zymography assay) 實驗中發現NTU04能隨著濃度的增加而減低基質金屬蛋白酶-2 (MMP2) 的活性。進一步發現，NTU04所造成基質金屬蛋白酶-2的活性降低是透過抑制基質金屬蛋白酶-2的基因表現及增加其金屬蛋白酶組織抑制因子-1 (TIMP1) 的基因表現。進一步證明，NTU04透過Erk訊息傳遞路徑而影響金屬蛋白酶組織抑制因子-1的表現。為了找尋有效成分，利用中效能液相層析法將MSL-UH草藥萃取液作分層，發現在F3、F4以及F5能有效地抑制肝癌細胞的侵襲轉移能力。同時也發現，將F3、F4以及F5依混合後餵食動物也可以導致腫瘤的生長受到抑制。總而言論，NTU04以及MSL-UH草藥萃取液是透過影響基質金屬蛋白酶-2的活性，同時也調控金屬蛋白酶組織抑制因子-1的表現量，進而能有效地抑制肝癌細胞的轉移侵襲能力。而本篇研究在未來在治療肝細胞癌方面提供一種新的藥物來源，並期望其能增進對轉移性肝癌的治療效率。

關鍵字

肝癌 ； 侵襲 ； 基質金屬蛋白酶-2 ； 金屬蛋白酶組織抑制因子-1

並列摘要

Hepatocellular carcinoma (HCC) is the third common causes of cancer mortality all over the world and in Taiwan. The high mortality of HCC is often caused by high recurrence and metastasis. However, there’s currently no effective therapeutic agent that can inhibit HCC metastasis. Therefore, to identify a new drug or compound with low cytotoxicity and high efficacy is imperative for cancer therapy. 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 or compounds for cancer therapy. In the study, two herbal extracts (NTU04 and NTU43) were identified and able to inhibit HCC cell invasion with low cytotoxicity. Moreover, the results showed that NTU04 could significantly reduce matrix metalloprotease-2 (MMP-2) activities in the conditioned media of Huh7 cells at a dose-dependent manner. NTU04-reduced gelatinolytic activity of MMP-2 was at least due to down-regulation of MMP2 expression and up-regulation of tissue inhibitor of metalloprotease-1 (TIMP1) expression. Moreover, the effect of NTU04 on up-regulating TIMP-1 expression was via Erk1/2 signaling. I then used ethanol to extract a Taiwanese herb, a close species to NTU04 and named the extract as MSL-UH. MSL-UH also could inhibit HCC cell invasion with less cytotoxicity. Using liquid chromatography, MSL-UH were divided into nine fractions. The third, fourth and fifth fractions (F3, F4 and F5) can more effectively inhibit HCC cell invasion than the other six fractions. Moreover, the mixture of MSL-UH F3, F4 and F5 significantly suppressed the tumor growth of Huh7 cells in xenografted mouse model. In conclusion, NTU04 and MSL-UH can inhibit the invasion of HCC cells, partly through inhibiting MMP2 activity and up-regulating TIMP1. The data indicate that NTU04 and MSL-UH may contain effective compounds or components that can inhibit HCC cell invasion and be useful for chemoprevention or therapeutic purposes.

並列關鍵字

Hepatocellular carcinoma ； Invasion ； Matrix metalloproteinase-2 (MMP2) ； Tissue inhibitors of matrix metalloproteinases-1 (TIMP1)

參考文獻

57 Lee C-C, Huang T-S. Plasminogen Activator Inhibitor-1: The Expression, Biological Functions, and Effects on Tumorigenesis and Tumor Cell Adhesion and Migration. Journal of Cancer Molecules 2005; 1: 25-36.

1 Worldwide Cancer Mortality. Cancer Research 2013.

3 Ashwin Ananthakrishnan MD, M.P.H. et al. Epidemiology of Primary and Secondary liver cancers. SEMINARS IN INTERVENTIONAL RADIOLOGY 2006; 23: 47-63.

4 Irfan Ahmed DNL. Malignant tumours of the liver. Surgery 2009; 27: 30-37.

5 El-Serag HB, Rudolph KL. Hepatocellular carcinoma: epidemiology and molecular carcinogenesis. Gastroenterology 2007; 132: 2557-2576.