乳癌在全世界女性中是最常罹患的癌症。大約有70 ％的乳癌患者皆表達estrogen receptor (簡稱ER＋)。對於這一類的患者在臨床上常用的第一線用藥為tamoxifen。Tamoxifen主要藉由和雌激素(E2)競爭ER的接合，來阻止雌激素誘導癌細胞的生長。然而大約有5％的乳癌患者用tamoxifen治療5年後產生抗藥性。我們先前的研究發現用tamoxifen治療會誘導TAR RNA binding protein (TARBP2)的表達，從而使ER＋乳癌細胞降低對tamoxifen的敏感度。因此，本研究篩選來自陽明大學傳統醫藥研究所的81種中草藥萃取物，研究它們對tamoxifen誘導增加的TARBP2是否具有抑制效果，期能應用於合併治療。首先使用MTT assay確定單純加入中草藥萃取物，未對乳癌細胞(MCF-7)產生毒性。而在這些中草藥萃取物中，編號122-2和629的萃取物可顯著抑制因tamoxifen誘導的TARBP2蛋白質表現。此外還觀察到122-2和629提高乳癌細胞對tamoxifen的敏感度，並抑制因tamoxifen促進的TARBP2和SOX2蛋白質表現量。而在另一種ER＋乳癌細胞ZR75-1也觀察到類似的結果。先前研究結果顯示，p-AKT會正向調控TARBP2的蛋白質表現量，我們也證明這兩種中草藥藉由抑制p-AKT的表現量促使Merlin表現量增加，進而抑制TARBP2的蛋白質表現量。總結來說，我們鑑定出2種具有潛在性中草藥萃取物可增加乳癌細胞對tamoxifen的敏感度。

Breast cancer is the most common cancer in women worldwide. Approximately 70% of breast cancer patients are estrogen receptor expression positive (ER+). One of the most commonly used first-line therapies for ER+ breast cancer patients is tamoxifen. Tamoxifen inhibits estrogen-induced cell growth predominately by competitive blockade of the estrogen receptor. Unfortunately, about 5% of the breast cancer patients showed drug-resistance after tamoxifen treatment for five years. Our previous study demonstrated that tamoxifen induces TAR RNA-binding protein (TARBP2) expression to enhance drug resistance in ER+ breast cancer cells. In this study, we screened eighty-one herbal extracts from Institute of Traditional Medicine，National Yang-Ming University to study their effects on reducing tamoxifen-induced TARBP2 expression. We first used MTT assay to ensure that no cytotoxic effects was observed in herbal extract treatment. Among these extracts, 122-2 and 629, significantly inhibited tamoxifen-induced TARBP2 protein expression. Furthermore, we observed that either 122-2 or 629 enhances tamoxifen sensitivity and prevents tamoxifen-induced TARBP2 and SOX2 protein expression. Similar results were also observed in another ER+ breast cancer cell line, ZR75-1Previous study demonstrated that p-AKT downregulate TARBP2 protein level. We also found that either 122-2 or 629 prevents p-AKT protein expression and induces Merlin protein expression. Taken together, we identified two herbal extracts with potential in combination therapy which sensitize breast cancer cells to tamoxifen treatment.

[1]Rebecca L. Siegel, MPH ; Kimberly D. Miller, MPH ; Ahmedin Jemal, DVM, PhD. Cancer statistics, 2019. CA CANCER J CLIN 2019;69:7–34
[2]Rebecca L. Siegel, MPH1; Kimberly D. Miller, MPH ; Ahmedin Jemal, DVM, PhD Cancer statistics, 2018. CA CANCER J CLIN 2018;68:7–30
[3]2018 Taiwan Health and Welfare Report
[4]Yu X, Luo A, Liu Y, Wang S, Li Y, Shi W, Liu Z, Qu X: MiR-214 increases the sensitivity of breast cancer cells to tamoxifen and fulvestrant through inhibition of autophagy. Mol Cancer 2015; 14: 208.
[5]Lindström LS, et al. (2012) Clinically used breast cancer markers such as estrogen receptor,progesterone receptor, and human epidermal growth factor receptor 2 are unstable throughout tumor progression. J Clin Oncol 30:2601–2608.
[6]Cardoso F, Bischoff J, Brain E, Zotano AG, Luck HJ, Tjan-Heijnen VC, et al. A review of the treatment of endocrine responsive metastatic breast cancer in postmenopausal women. Cancer Treat Rev. 2013;39:457–65.
[7]Morgan LR, Jr, et al. (1976) Therapeutic use of tamoxifen in advanced breast cancer:Correlation with biochemical parameters. Cancer Treat Rep 60:1437–1443.
[8]McDonnell DP, Wardell SE, Norris JD. Oral selective estrogen receptor downregulators (SERDs), a breakthrough endocrine therapy for breast cancer. J Med Chem 2015;58:4883–7.
[9]Ma CX, Reinert T, Chmielewska I, Ellis MJ. Mechanisms of aromatase inhibitor resistance. Nat Rev Cancer2015;15:261–75.
[10]Wenwen Gu, Wenping Xu, Xiaoxi Sun, Bubing Zeng, Shuangjie Wang, Nian Dong, Xu Zhang, Chengshui Chen, Long Yang, Guowu Chen, Aijie Xin, Zhong Ni, Jian Wang & Jun Yang. Anordrin Eliminates Tamoxifen Side Effects without Changing Its Antitumor Activity. Scientific Reports volume7, Article number: 43940 (2017)
[11]Qadir MA, Kwok B, Dragowska WH, To KH, Le D, Bally MB, et al. Macroautophagy inhibition sensitizes tamoxifen-resistant breast cancer cells and enhances mitochondrial depolarization. Breast Cancer Res Treat. 2008;112:389–403.
[12] Zhong yang min zu da xue xue bao bian ji bu. : 123 [19 January 2013].
[13] Stermitz F, Lorenz P, Tawara J, Zenewicz L, Lewis K. Synergy in a medicinal plant: anti-microbial action of berberine potentiated by 5′-methoxyhydnocarpin, a multidrug pump inhibitor. Proceed Nat Acad Sci US Am. 2000;97:1433–37.
[14]Winter J, Jung S, Keller S, Gregory RI, Diederichs S.. Many roads to maturity: microRNA biogenesis pathways and their regulation. Nat Cell Biol 11: 228-234
[15]Caramuta,S.et al.,Clinical and functional impact of TARBP2 over-expression in adrenocortical carcinoma. Endocr Relat Cancer, 2013. 20(4): p. 551-64.
[16]Fu,X.,et al.The activity and expression of microRNAs in prostate cancers. Mol Biosyst, 2010. 6(12): p. 2561-72.
[17]Sand,M.et al.The miRNA machinery in primary cutaneous malignant melanoma, cutaneous malignant melanoma metastases and benign melanocytic nevi. Cell Tissue Res, 2012. 350(1): p. 119-26.
[18]Ming-Yang Wang ,Pai-Sheng Chen. et al. TARBP2-Enhanced Resistance during Tamoxifen Treatment in Breast Cancer. Cancers 2019, 11, 210; doi:10.3390/cancers11020210
[19]Hilf M, Yu VL, Sharp J, Zuravleff JJ, Korvick JA, Muder RR. Antibiotic therapy for Pseudomonas aeruginosa bacteremia: outcome correlations in a prospective study of 200 patients. Am J Med. 1989; 87 (5):540–6.
[20]Yeh WL, Lin HY, Wu HM and Chen DR: Combination treatment of tamoxifen with risperidone in breast cancer. PLoS One 9: e98805, 2014.
[21]Yi-Hua Lai, Sung-Liang Yu, Hsuan-Yu Chen, Chi-Chung Wang, Huei-Wen Chen, Jeremy J.W. Chen.(2013)The HLJ1-targeting drug-screening identified Chinese herb andrographolide that can suppress tumour growth and invasion in non-small-cell lung cancer.Carcinogenesis
[22]Chou TC: Theoretical basis, experimental design and computerized simulation of synergism and antagonism in drug combination studies. Pharmacol Rev 58: 621-681, 2006.
[23]Péter Érdi; János Tóth (1989). Mathematical Models of Chemical Reactions: Theory and Applications of Deterministic and Stochastic Models. Manchester University Press p.3.ISBN 978-0-7190-2208-1.
[24]De Pedro, N., M.R. Chica, J. Cantizani, B. Cautain and F. Vicente et al., 2013. Antiproliferative and apoptotic potential of Chinese medicinal plants against MCF-7 (luminal A), HCC1954 (HER2+) and Hs578t breast cancer cells. Phytopharmacology, 4: 454-467
[25]Martins, F.S., da Conceicao, E.C., 2015. Evaluation of extraction method on the chemical composition in Apeiba tibourbou Aubl’s extracts. Pharmacogn. Mag. 11, 368–373.
[26] L.Rizzo ,G.Longato ,A.Ruiz ,S.Tinti , A. Possenti , D. Vendramini-Costa , A. Sartoratto , G. Figueira , F. Silva and M. Eberlin , In vitro, in vivo and in silico analysis of the anticancer and estrogen-like activity of guava leaf extracts, Curr. Med. Chem., 2014, 21 , 2322 -2330
[27] Tang X, Jang SW, Wang X, Liu Z, Bahr SM, Sun SY, Brat D, Gutmann DH, Ye K. Akt phosphorylation regulates the tumour-suppressor merlin through ubiquitination and degradation. Nat Cell Biol. 2007;9:1199–1207. doi: 10.1038/ncb1641.
[28] . Lee, J. Y., H. J. Moon, W. K. Lee, H. J. Chun, C. W. Han, Y. W. Jeon, Y. Lim, Y. H. Kim, T. P. Yao, K. H. Lee, T. Y. Jun, H. K. Rha, and J. K. Kang. 2006. Merlin facilitates ubiquitination and degradation of transactivation-responsive RNA-binding protein. Oncogene 25:1143–1152.
[29] Qi W, Sun M, Kong X, Li Y, Wang X, Lv S, Ding X, Gao S, Cun J, Cai C, Wang X, Chen J, Yin A, Yang Q: Huaier extract synergizes with tamoxifen to induce autophagy and apoptosis in ER-positive breast cancer cells. Oncotarget 2016;7:26003-26015.
[30] Yeh WL, Lin HY, Wu HM, Chen DRCombination treatment of tamoxifen with risperidone in breast cancer. PLoS One 9: e98805
[31] Q. Kong, Y. Ma, J. Yu, and X. Chen, “Predicted molecular targets and pathways for germacrone, curdione, and furanodiene in the treatment of breast cancer using a bioinformatics approach,” Scientific Reports, vol. 7, no. 1, Article ID 15543, 2017.
[32] Z. Liu, F. Guo, Y. Wang et al., “BATMAN-TCM: a bioinformatics analysis tool for molecular mechanism of traditional chinese medicine,” Scientific Reports, vol. 16, Article ID 21146, 2016.
[33]Xu DP, Li Y, Meng X, Zhou T, Zhou Y, Zheng J, Zhang JJ and Li HB: Natural antioxidants in foods and medicinal plants: extraction, assessment and resources. Int J Mol Sci 2017; 18(1): E96.
[34] Xu, D.P.; Li, Y.; Meng, X.; Zhou, T.; Zhou, Y.; Zheng, J.; Zhang, J.J.; Li, H.B. Natural antioxidants in foods and medicinal plants: Extraction, assessment and resources. Int. J. Mol. Sci. 2017, 18, 96.
[35] Liu P, Cheng H, Roberts TM, Zhao JJ (2009) Targeting the phosphoinositide 3-kinase pathway in cancer. Nat Rev Drug Discov 8(8), 627–44.
[36] Sartippour MR, Pietras R, Marquez-Garban DC, Chen HW, Heber D, Henning SM, Sartippour G, Zhang L, Lu M, Weinberg O, Rao JY, Brooks MN: The combination of green tea and tamoxifen is effective against breast cancer. Carcinogenesis 2006;27:2424-2433.
[37] S. J. Ameh, O. O. Obodozie, P. C. Babalola, and K. S. Gamaniel, “Medical herbalism and herbal clinical research—a global perspective,” British Journal of Pharmaceutical Research, vol. 1, no. 4, pp. 99–123, 2011.
[38] Fu, B.; Wang, N.; Tan, H.Y.; Li, S.; Cheung, F.; Feng, Y. Multi-component herbal products in the orevention and treatment of chemotherapy-associated toxicity and side effects: A review on experimental and clinical evidences. Front. Pharmacol. 2018, 9, 1394.
[39] Lu Y, Li CS, Dong Q. Chinese herb related molecules of cancer-cell-apoptosis: a minireview of progress between Kanglaite injection and related genes. J Exp Clin Cancer Res 2008;27:31DOI: 10.1186/1756-9966-27-31
[40] Alassane Mbengue et. al., 2015. A molecular mechanism of artemisinin resistance in Plasmodium falciparum malaria. Nature. doi:10.1038/nature14412
[41] Meng J, Guo F, Xu H, Liang W, Wang C, Yang XD. Combination therapy using co-encapsulated resveratrol and paclitaxel in liposomes for drug resistance reversal in breast cancer cells in vivo. Sci Rep 2016;6:22390.