- 學位論文
異黃酮biochanin與formononetin抑制肺癌細胞侵襲和移動及其相關機制探討
Study of inhibitory effect and its mechanism of the isoflavones biochanin and formononetin on lung cancer invasion and migration
摘要
癌症目前已是國人死亡的主要原因之一，而癌細胞轉移(metastasis)更是導致癌症病人死亡以及治療複雜度提昇的主要原因。近年來頗受醫學界重視研究的天然植物性異黃酮，其結構與女性雌激素相似，且被證實能發揮與雌激素類似的效果。異黃酮已被證實在抑制乳癌及前列腺癌細胞的增生及誘發細胞凋亡有相當的效果。但是對於抑制癌瘤細胞轉移能力的相關研究則尚未清楚，及其是否會抑制肺癌細胞的生長亦不明瞭。因此，針對肺癌細胞(A549 cells)，選用2個常見異黃酮 (biochanin A and formononetin)，探討對於抑制癌細胞增生及侵襲轉移的效果。結果發現， biochanin A在20μM的濃度下，對於肺癌細胞的侵襲/移動(invasion/migration)能力具有顯著的抑制效果(P<0.05)，且當濃度高於50 μM時，對於癌細胞的存活也有顯著的抑制效果。然而formononetin則對於肺癌細胞的侵襲/移動(invasion/migration)並無良好的抑制效果，但可有效降低癌細胞的存活。以gelatin及casein zymography的分析結果，顯示biochanin A對於肺癌細胞的基質分解酵素matrix metalloproteinase (MMP)-2及urokinase-type plasminogen activator (u-PA)之酵素活性有顯著抑制的效果，針對肺癌A549細胞進一步探討biochanin A抗癌細胞轉移的機制，發現biochanin A也會抑制細胞的貼附能力。在分子機轉的相關分析中，biochanin A會造成extracellular signal-regulated kinase 1/2 (ERK 1/2)、p38MAPK及Akt的磷酸化下降。綜合以上發現， biochanin A能有效抑制肺癌細胞的增生及侵襲的能力，且MMP-2及u-PA的活性也有相當的抑制效果。並且在動物實驗中，以缺乏免疫能力的老鼠在皮下施打A549細胞株，並再第八天的時候開始每天餵食biochanin A，發現biochanin A的組別與控制組作比較其腫瘤大小有受到明顯的抑制，在未來或許可作為抑制肺癌轉移的輔助治療。
關鍵字
異黃酮並列摘要
Metastasis of cancer cells, a multiple and intricate process, may complicate the clinical management and lead to a poor prognosis with tremendous impact to patients or communities. Isoflavones are a class of polyphenolic compounds found in soy and other legumes, and have been well characterized to have various bioactive properties. Several previous studies have established that isoflavones possess the propensity to anti-proliferation and induce apoptosis in breast and prostate cancer. Based on our knowledge, studies on the inhibitory effect of isoflavones on cancer cell invasion have been relatively less and it warrants a further study. In this study, we investigated the inhibitory effect of flavanone compounds, including biochanin A and formononetin, on the proliferation and metastasis of A549 lung cancer cells. The results showed that biochanin A and genistein significantly (P<0.05) reduced the invasion/migration of A5491 cells at the dose of 20 μM, while at the dose higher than 50 μM, they also significantly (P<0.05) deceased the cell viability of A549 cells. However, formononetin only exerted inhibitory effect on cell viability of A549 cells, but no-effect on cell invasion/migration. Gelatin and casein zymography analysis showed that biochanin A extensively inhibited the activities of MMP-2 and u-PA in A549 cells while formononetin didn’t affect these enzyme activities. In lung cancer A549 cells, treatment with biochanin A also reduced cell-matrix adhesion, and potently attenuated the phosphorylation of extracellular signal-regulated kinase 1/2 (ERK 1/2), p38MAPK and Akt. The anti-invasive effect of biochanin A was correlated with the inhibition of MMP-2, MMP-9 and u-PA activities, which may attributed to the down-regulation of ERK1/2, p38MAPK, and Akt pathway. Finally, an in vivo anti-tumor study using nude mice (BALB/c nu/nu) xenograft model by a subcutaneous inoculation of A549 cells was performed. The average tumor volume of treatment groups was statistically lower than that of the control group. In conclusion, biochanin A perturb the invasion capacities of lung cancer cells, thereby constituting an adjuvant treatment for metastasis control.