茶多酚與抗癌藥對子宮頸癌細胞毒性協同作用

Synergistic Cytotoxicity Effects of Anticancer Drugs with Tea Polyphenol on Human Cervical Cancer Cell

蔡婉怡(Wan-Yi Tsai)

指導教授 ： 楊玲玲 ； 王靜瓊

臺北醫學大學/藥學院/生藥學研究所/碩士(2002年)

摘要

中文摘要 茶(Camellia sinensis) 在世界各國是種很普遍的飲料，近年來的研究報告指出，不論何種製法，茶中富含的多酚類具有抗氧化、清除自由基及抑制癌細胞生長等作用。目前，臨床抗癌藥物只能減輕症狀、縮小腫瘤，抑制惡性腫瘤細胞的生長及轉移，只能發揮控制症狀的效果。現今許多疾病多用雞尾酒療法治療，來提高治療效果，因此本研究希望能將茶多酚與抗子宮頸癌藥物合併使用，而達到降低化學療法藥物的劑量，提高抑制癌細胞效果。故本論文採集十五種台灣省產茶屬植物，烘乾後以70% acetone萃取，並以Folin-Ciocalteu方法分析其茶多酚總量，結果發現皆富含茶多酚，因而選用catechin, (-)-epigallocatechin gallate, (-)-epigallocatechin, (-)-epicatechin gallate, (-)-gallocatechin, (-)-gallocatechin gallate, gallic acid及tea extract from black tea等八種茶中主要之多酚類與doxorubicin, cisplatin, vincristine等三種臨床抗癌藥物，合併處理癌細胞，探討其對抑制癌細胞生長的協同作用。以DPPH及DCFH-DA方法，探討此八種多酚之抗氧化能力，發現以gallic acid作用最強，且高濃度時細胞內會有自由基產生。進而利用MTT方法檢測八種茶多酚對子宮頸癌細胞Ca-Ski之細胞毒性，亦發現gallic acid對子宮頸癌細胞Ca-Ski的毒性最強，並以其與臨床抗癌藥物合併處理時有最強的協同作用，且不會增強對正常子宮頸纖維母細胞之毒性。再以西方印蹟法、流氏細胞儀及拓撲異構酵素抑制試驗得知，gallic acid與doxorubicin合併處理癌細胞，其PARP及對拓撲異構酵素 I的抑制會隨著gallic acid的濃度增加而增加，而拓撲異構酵素 II的抑制則不明顯，細胞週期則會使由原本滯留於S及G2/M phase的細胞，變為滯留於G0/G1 phase，增加sub-G1而使細胞毒性增強。Gallic acid與cisplatin合併處理癌細胞，其PARP及對拓撲異構酵素 I及II的抑制會隨著gallic acid的濃度增加而增加，細胞週期則是非選擇性的抑制，而使細胞毒性增強。gallic acid與vincristine合併處理癌細胞，其PARP及對拓撲異構酵素 I的抑制會隨著gallic acid的濃度增加而增加，而拓撲異構酵素 II的抑制則不明顯，細胞週期則會使原本滯留於G2/M phase的細胞，變為滯留於G0/G1 phase，而使細胞毒性增強。綜合上述得知，gallic acid可增強此三種抗癌藥之細胞毒性，但隨著抗癌藥本身之作用機制不同，協同作用的強度亦有差異。

關鍵字

茶多酚 ； 抗癌藥 ； 毒性協同作用 ； 細胞週期 ； 子宮頸癌細胞 ； 正常子宮頸纖維母細胞

並列摘要

Abstract Tea is a popular drink world-wide. No matter how the tea processed, there still remain abundance of polyphenols that are antioxidative, ROS-scavenging, and antimutagenic within the tea. Anti-cancer medication , so far, are still not very effective in curing cancer, and cause lots of side effects. In the past decade, combination therapy has been employed to treat diseases in order to get better results and less side effects. We want to investigate whether the combined treatment of anticancer drug with tea polyphenol will reduce the effective dose of anticancer drug. We colleceted 15 kinds of tea from Taiwan, extracted with 70 % acetone after the leaves dried. Folin-Ciocalteu method was used to determine the content of polyphenols within the tea, 8 types of polyphenols： catechin, (-)-epigallocatechin gallate, (-)-epigallocatechin, (-)-epicatechin gallate, (-)-gallocatechin, (-)-gallocatechin gallate, gallic acid, and tea extract from black tea were used to in the experiments . The synergistic effects of these polyphenols with doxorubicin, cisplatin, and vincristin on inhibition of growth of tumor cells were then investigated. Among these polyphenols, gallic acid exhibited the strongest antioxidative activity based on DPPH and DCFH-DA methods. MTT assay revealed greater cellular toxicity of gallic acid to Ca-Ski cells and gallic acid could also enhance the anticancer effect of . When cancer cells were treated with gallic acid and doxorubicin, the inhibition of PARP and topoisomerase I increased along with increasing dose of gallic acid. Flow cytometry study revealed increased sub-G1 peak. Combined treatment of cancer cells with gallic acid and vincristine also increased the inhibition on PARP activity and topoisomerase I as the dose of gallic acid increased. Sub g1 peak also increased. Thus, the mechanism of the synergistic effect of anticancer drug with gallic acid was probably due to enhance the mechanism of anticancer drug.

並列關鍵字

tea polyphenol ； anticancer drug ； cell cycle ； Synergistic Cytotoxicity ； Ca-Ski ； Normal cervical fibroblasts

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