黃體酯酮(progesterone)抑制肝癌再發之 實驗室研究

The Effect of Progesterone on Prevention of Recurrence of Hepatocellular Carcinoma –In Vitro Study

張文燦(Wen-Tsan Chang)

指導教授 ： 李金德

高雄醫學大學/醫學院/醫學研究所/碩士(2005年)

摘要

肝癌是我國男性的癌症首位，它的五年生存率為26.4%~47.4%，是預後極為不良的癌症。手術切除仍然是目前肝癌治療的優先選擇之一。而導致它不良的預後，除了低的可切除率(15-30%)之外，更棘手的是手術後高復發率，一年的再發率為35.9%~61.5%，五年累積復發率在75%~100% 之間。因此抑制肝癌的復發為提昇肝癌治療成績的當務之急。有學者術後使用干擾素或化學治療等方法，試圖抑制肝癌的術後再發，但結果似乎難令人滿意。
由於肝癌組織被發現有多量的androgen receptor，也有證據顯示testosterone及androgen receptor與肝癌的發生與復發，有密切的關係。
Progesterone 具有anti-androgen的作用。我們在臨床上，於術後給予合成的黃體酯酮，初步得到令人滿意的結果，能夠降低肝癌術後再發率。我們因此亟想探討其是否有實驗室的證據。此外epirubicin 是肝癌術後或無法切除之肝癌，常用的輔助性化學治療藥物，我們也想探討併用黃體酯酮與epirubicin是否具有加成之作用。
因此我們將具有androgen receptor之肝癌細胞株HepG2 細胞株與testosterone、progesterone及epirubicin一起培養，再以(1)3H-thymidine incorporation及MTT assay觀察HepG2 細胞株的生長情形(2)分析LDH，探測各藥物對HepG2 cell有無毒殺效果(3)並分析flow cytometry探測各藥物對HepG2 細胞株有無引起Apoptosis的情形。
我們研究顯示testosterone能夠促進HepG2 細胞株的生長。而 progesterone 能夠抑制肝癌細胞株 HepG2 的生長，但對HepG2細胞並無毒殺作用。而 epirubicin具有毒殺HepG2細胞株之作用。而progesterone能夠促進 epirubicin 對 HepG2 細胞株之毒殺作用及細胞凋亡反應，而且主要是增加epirubicin 對 HepG2 細胞株停留在細胞週期中G2/M 階段的比例。
我們推測在臨床上可運用progesterone於 (1)降低肝癌術後局部與全身復發率(2)減少epirubicin的使用劑量，降低其潛在的副作用。

關鍵字

肝癌細胞 ； 黃體酯酮

並列摘要

Hepatocellular carcinoma (HCC) is a leading cancer in man in Taiwan. It carries a very poor prognosis and a 5 –year survival rate of 26.4% to 47.4% has been reported. The partial hepatic resection is a potential curable treatment and remains the first choice of treatment. However, the resection rate of HCC is low (15-30%) and an incidence of postoperative recurrence is still very high and is the main cause of late deaths .The 1-year recurrence was 35.9% -61.5% and the cumulative 5-year recurrence rate ranging from 75% to 100%.
Undoubtedly, prevention of recurrence is the most important effort to improve the overall survival after resection of HCC. Androgen receptors are abundant in HCC tissues and strongly associated with intrahepatic recurrence of HCC has been identified. Progesterone had the anti-androgen effect was also proven. Our initial clinical data revealed that progesterone can reduce the postoperative recurrence of HCC. epirubicin was commonly administrated after liver resection of HCC.
The satisfactory clinical results prompt us to search the laboratory evidence of progesterone and study the effect of progesterone and combination of epirubicin for HCC in vitro. We added different concentration of progesterone, testosterone and epirubicin on HepG2 cell line containing androgen receptor by assessing (1)3H-thymidine incorporation and MTT assay (2) analysis of the level of LDH and (3) analysis of flow cytometery to evaluate the change of ratio of apoptosis and distribution of cell cycle of HepG2 cells.
We found the progesterone can inhibit the proliferation of HepG2 cells in the concentrations of 12.5 nM and 25 nM, but no similar consistent result was observed in the concentration of 50 nM. The progesterone had no cytotoxic effect on HepG2 cells was relealed by LDH assay. However, progesterone could enhance both the cytotoxic and apoptotic effect of epirubicin on HepG2 cells. By method of flow cytometery, the progesterone could also increase of G2/M phase arrest induced by epirubicin was also.
In conclusion, our study provides the in vitro evidence of possible benefits of combination of progesterone and epirubicin. These two drugs had the synchronous effects of cytotoxicity and apoptosis on HCC cell lines. We predict combination of these two drugs can decrease the rates of postoperative recurrence of HCC and lower the therapeutic dose of epirubicin. Then the adverse effect of chemotherapy decreases. However, it needs more clinical studies to confirm this point of view.

並列關鍵字

G2M arrest ； Progesterone ； Hepatocellular carcinoma ； epirubicin ； HepG2 cell

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