紅麴發酵產物在癌症預防與治療中所扮演的角色

Abstract

紅麴發酵產物為深具東方特色之傳統食品，其二次代謝產物經科學研究證實具有抑制腫瘤轉移、抗發炎、降低膽固醇生成等多項作用。因此，在學理上十分適合運用於抑制腫瘤治療過程中可能引發的腫瘤轉移及組織發炎反應的輔助性治療。 本論文第一部份利用小鼠肺癌細胞 (Lewis lung carcinoma; LLC) 植入C57/BL6 小鼠所形成 LLC 荷瘤小鼠，探討給與紅麴米餵食所產生之效應。植入 5 x 105 LLC 細胞之小鼠，同時餵食 2% 紅麴米，經 14 天後與未攝食紅麴米組比較腫瘤大小，紅麴米抑制達 51.1% 之多。以紅麴乙醇萃出物 (red mold rice ethanol extract; RMRE) 進行體外侵襲與轉移試驗顯示，RMRE 可抑制透過介白質六號 (Interleukin-6; IL-6) 所活化，與侵襲及轉移能力增加相關的基質金屬蛋白酶 (matrix metalloproteinases; MMPs)，包括 MMP-2 與 MMP-9。進一步在 RMRE 合併臨床化療藥物治療的效力上發現，RMRE 不僅造成 LLC 細胞的細胞週期停止於 G1 期，同時隨著劑量提升而增加誘發其細胞凋亡現象。 第二部份探討 RMRE 抑制 LLC 荷瘤小鼠血清中血管內皮生長因子 (vascular endothelial growth factor; VEGF) 上升，同時降低荷瘤小鼠轉移的發生。結果指出 RMRE 中的 monacolin K (MK) 可能扮演著關鍵性角色，包括利用體外轉移與血管新生試驗證實 MK 抑制 LLC 細胞的轉移侵襲能力，同時經反轉錄聚合酶鏈反應 (reverse transcription polymerase chain reaction; RT-PCR) 得知，MK 藉由降解 VEGF 基因表現而抑制 LLC 細胞分泌 VEGF 誘發內皮細胞血管之生成。 第三部份以雞胚胎尿囊絨毛膜 (chorioallantoic membrane; CAM) 試驗探討 RMRE 與紅麴抗氧化物 dimerumic acid (DMA) 對於腫瘤血管新生與細胞內滲之作用。RMRE 與 DMA 隨著劑量與時間增加對人類大腸癌細胞株，SW480 與 SW620 細胞，具有劑量關係。同時經由分析雞胚 CAM 下方所含有人類 Alu 序列片段結果顯示，RMRE 可抑制於 Matrigel 包覆有 SW620 細胞之植體從雞胚 CAM 上方自發性內滲轉移至 CAM 下方的能力。含 SW620 細胞之植體於 CAM 中可增加 75.3±11.6% 之新生血管生成，然 RMRE 可隨劑量增加而顯著抑制 SW620 所誘發之 CAM 血管新生。最後經由 RT-PCR、西方墨點轉漬分析與酪蛋白酶譜分析顯示 RMRE 藉由抑制 MMP-7 活性達到降低腫瘤轉移發生。 以上顯示，利用 LLC 荷瘤小鼠或雞胚胎 CAM 等動物模式皆證實以膳食補充紅麴米方式可達到抑制腫瘤生長與轉移之發生。因此，包括 RMRE、MK、AK 與 DMA 於未來相當具潛力研究發展，使成為一無毒性、天然之化學預防膳食補充品，或作為癌症治療輔助劑。

The Monascus fermented products are of special interest because of their use in the production of a range of oriental fermented foods. The secondary metabolites of Monascus in the modern research were confirmed had the suppression tumor metastasis, anti-inflammation, the suppression cholesterol production and so on many functions. Therefore, Monascus metabolites extremely suit to utilize cancer therapy to suppress the tumor metastasis and the inflammation responded. The first section described the effects of red mold rice on Lewis lung carcinoma (LLC) bearing in syngeneic C57BL/6 mice. Oral administration of 2% red mold rice and implant 5 × 105 LLC cells in mice for 14 days significantly inhibited the tumor progress (51.1%). In vitro assays, the investigation involved the effects of red mold rice ethanol extract (RMRE) on LLC cells invasion and metastasis. RMRE interference on Interleukin-6 (IL-6) induced cell invasion and metastasis gene expressions of matrix metalloproteinases (MMPs) contained MMP-2 and MMP-9. Moreover, the assessment of chemotherapeutic efficiency combined clinical drugs used on tumor cell viabilities have showed that RMRE not only significantly arrested LLC cells G1 phase, but also induced cell apoptosis following arise the doses of RMRE treatment. The secondary part represented that oral administration with RMRE or dramatically inhibited the metastatic ability of LLC-bearing mice caused by the decline of serum vascular endothelial growth factor (VEGF) levels compared with untreated metastatic groups. These results indicated that MK was a key antimetastatic and antiangiogenesis compound in RMRE and proved that by Matrigel-coating transwell and tube-forming assays, as well as reverse transcription polymerase chain reaction (RT-PCR) illustrated the downregulation of VEGF-stimulated invasive activity in LLC cells. The third section showed RMRE and DMA significantly inhibited the proliferation of SW480 and SW620 human colorectal carcinoma cells in dose- and time-dependent manner. Moreover, SW620 cells could growth in Matrigel grafts and spontaneously intravasation from the upper to the lower of chick embryo chorioallantoic membrane (CAM) model, which detected human Alu genomic DNA by PCR amplification from the lower CAMs at RMRE-untreated group. The percentage of neovascularization was increased 75.3±11.6% by SW620 cells onplant with Matrigel grafts on the CAM model. However, addition of RMRE significantly reduced CAM neovascularization in dose-dependent effect. Finally, we have finding that RMRE and DMA effectively decreased activity of MMP-7 determined by RT-PCR, Western blotting, and casein zymography assays. In summary, using the food-based approach to cancer prevention has shown that Monascus-fermented products ingestion inhibited both the primary tumor growth and metastatic progression on both LLC-bearing mice and chick embryo CAM models in vivo. Therefore, application of RMRE, MK, AK and DMA may serve as a non-toxic natural chemopreventive or antineoplastic agent for further development of cancer adjuvant chemotherapy. We conclude that Monascus is a highly potential herb for cancer therapy especially in tumor chemotherapy, the more appropriate application worthy to be studied in the future.