Brazilein in combination with cisplatin inhibit proliferation and migration on highly metastatic cancer cells, 4T1

Sri Handayani(1), Ratna Asmah Susidarti(2), Zalinar Udin(3), Edy Meiyanto(4), Riris Istighfari Jenie(5*)

(1) Cancer Chemoprevention Research Center, Faculty of Pharmacy, Universitas Gadjah Mada, Indonesia, 55281
(2) Faculty of Pharmacy, Universitas Gadjah Mada, Indonesia, 55281
(3) Research Center for Chemistry, Indonesian Institute of Sciences (LIPI), Indonesia
(4) Faculty of Pharmacy, Universitas Gadjah Mada, Indonesia, 55281
(5) Faculty of Pharmacy, Universitas Gadjah Mada, Indonesia, 55281
(*) Corresponding Author


Brazilein performs anti­cancer activities on several cancer cells and potentially inhibits metastasis. The aims of this study is to observe the synergistic cytotoxic and migration inhibitory effect of brazilein combined with cisplatin on 4T1 breast cancer cells. Under MTT assay, we found that brazilein revealed cytotoxic effect on 4T1 cells in a dose­dependent manner (IC50=50 ± 0.3 µM). Combination of brazilein and cisplatin showed synergistic effect (CI=0.72). Flowcytometry analysis on the cell cycle progression showed that single treatment of 25 µM brazilein induced G2/M­phase accumulation, 12.5 µM cisplatin induced S­phase accumulation, while combination of brazilein and cisplatin induced S­phase and G2/Mphase accumulation. Combination of brazilein and cisplatin induced apoptosis higher than that of the single treatments. Based on wound healing assay, 12.5 µM brazilein and its combination with 6.25 µM cisplatin inhibited cells migration. Immunoblotting and gelatin zymography analysis showed that combination of brazilein and cisplatin inhibited the expression level of Rac1 and MMP9 proteins. Based on these results, we conclude that brazilein enhanced cytotoxic activity of cisplatin and inhibited migration on 4T1 cells and potentially can be developed as an enhancing cytotoxic and antimetastasis agent.


brazilein; cisplatin; combination treatment; cytotoxic effect; cells migration

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Aggarwal, B.B., 2004. Nuclear factor-kappaB: the enemy within. Cancer Cell 6, 203–208. doi:10.1016/j.ccr.2004.09.003

Amin, H., Wani, N.A., Farooq, S., Nayak, D., Chakraborty, S., Shankar, S., Rasool, R. ur, Koul, S., Goswami, A., Rai, R., 2015. Inhibition of Invasion in Pancreatic Cancer Cells by Conjugate of EPA with β3,3-Pip-OH via PI3K/Akt/NF-kB Pathway. ACS Med. Chem. Lett. 6, 1071–1074. doi:10.1021/acsmedchemlett.5b00257

Baribeau, S., Chaudhry, P., Parent, S., Asselin, É., 2014. Resveratrol Inhibits Cisplatin-Induced Epithelial-to-Mesenchymal Transition in Ovarian Cancer Cell Lines. PLOS ONE 9, e86987. doi:10.1371/journal.pone.0086987

Brooks, S.A., Lomax-Browne, H.J., Carter, T.M., Kinch, C.E., Hall, D.M.S., 2010. Molecular interactions in cancer cell metastasis. Acta Histochem. 112, 3–25. doi:10.1016/j.acthis.2008.11.022

Chen, Q., Zheng, Y., Jiao, D., Chen, F., Hu, H., Wu, Y., Song, J., Yan, J., Wu, L., Lv, G., 2014. Curcumin inhibits lung cancer cell migration and invasion through Rac1-dependent signaling pathway. J. Nutr. Biochem. 25, 177–185. doi:10.1016/j.jnutbio.2013.10.004

Chou, T.-C., 2006. Theoretical Basis, Experimental Design, and Computerized Simulation of Synergism and Antagonism in Drug Combination Studies. Pharmacol. Rev. 58, 621–681. doi:10.1124/pr.58.3.10

Ferrari-Amorotti, G., Chiodoni, C., Shen, F., Cattelani, S., Soliera, A.R., Manzotti, G., Grisendi, G., Dominici, M., Rivasi, F., Colombo, M.P., Fatatis, A., Calabretta, B., 2014. Suppression of Invasion and Metastasis of Triple-Negative Breast Cancer Lines by Pharmacological or Genetic Inhibition of Slug Activity. Neoplasia N. Y. N 16, 1047–1058. doi:10.1016/j.neo.2014.10.006

Geng, S., Gu, L., Ju, F., Zhang, H., Wang, Y., Tang, H., Bi, Z., Yang, C., 2016. MicroRNA-224 promotes the sensitivity of osteosarcoma cells to cisplatin by targeting Rac1. J. Cell. Mol. Med. 20, 1611–1619. doi:10.1111/jcmm.12852

Hanahan, D., Weinberg, R.A., 2011. Hallmarks of Cancer: The Next Generation. Cell 144, 646–674. doi:10.1016/j.cell.2011.02.013

Hsieh, C.-Y., Tsai, P.-C., Chu, C.-L., Chang, F.-R., Chang, L.-S., Wu, Y.-C., Lin, S.-R., 2013. Brazilein suppresses migration and invasion of MDA-MB-231 breast cancer cells. Chem. Biol. Interact. 204, 105–115. doi:10.1016/j.cbi.2013.05.005

Hua, H., Li, M., Luo, T., Yin, Y., Jiang, Y., 2011. Matrix metalloproteinases in tumorigenesis: an evolving paradigm. Cell. Mol. Life Sci. 68, 3853–3868. doi:10.1007/s00018-011-0763-x

Kaur, P., Nagaraja, G.M., Zheng, H., Gizachew, D., Galukande, M., Krishnan, S., Asea, A., 2012. A mouse model for triple-negative breast cancer tumor-initiating cells (TNBC-TICs) exhibits similar aggressive phenotype to the human disease. BMC Cancer 12, 120. doi:10.1186/1471-2407-12-120

Kemper, K., Goeje, P.L. de, Peeper, D.S., Amerongen, R. van, 2014. Phenotype Switching: Tumor Cell Plasticity as a Resistance Mechanism and Target for Therapy. Cancer Res. 74, 5937–5941. doi:10.1158/0008-5472.CAN-14-1174

Kim, B.S., 2010. Brazilin Inhibits of TPA-induced MMP-9 Expression Via the Suppression of NF-κB Activation in MCF-7 Human Breast Carcinoma Cells. J. Food Hyg. Saf.

Kim, J.S., Kang, C.G., Kim, S.-H., Lee, E.-O., 2014. Rhapontigenin Suppresses Cell Migration and Invasion by Inhibiting the PI3K-Dependent Rac1 Signaling Pathway in MDA-MB-231 Human Breast Cancer Cells. J. Nat. Prod. 77, 1135–1139. doi:10.1021/np401078g

Laksmiani, N.P.L., Susidarti, R.A., Meiyanto, E., 2015. Brazilein Increases The Sensitivity of Doxorubicin on MCF-7 Resistant Doxorubicin (MCF-7/DOX) Cells Through Inhibition of HER-2 Activation. Int. J. Pharm. Pharm. Sci. 7, 525-528.

Larsson, D.E., Lövborg, H., Rickardson, L., Larsson, R., Oberg, K., Granberg, D., 2006. Identification and evaluation of potential anti-cancer drugs on human neuroendocrine tumor cell lines. Anticancer Res. 26, 4125–4129.

Latifi, A., Abubaker, K., Castrechini, N., Ward, A.C., Liongue, C., Dobill, F., Kumar, J., Thompson, E.W., Quinn, M.A., Findlay, J.K., Ahmed, N., 2011. Cisplatin treatment of primary and metastatic epithelial ovarian carcinomas generates residual cells with mesenchymal stem cell-like profile. J. Cell. Biochem. 112, 2850–2864. doi:10.1002/jcb.23199

Martin, T.A., Ye, L., Sanders, A.J., Lane, J., Jiang, W.G., 2013. Cancer Invasion and Metastasis: Molecular and Cellular Perspective. Landes Bioscience.

Mosmann, T., 1983. Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. J. Immunol. Methods 65, 55–63.

Nirmal, N.P., Rajput, M.S., Prasad, R.G.S.V., Ahmad, M., 2015. Brazilin from Caesalpinia sappan heartwood and its pharmacological activities: A review. Asian Pac. J. Trop. Med. 8, 421–430. doi:10.1016/j.apjtm.2015.05.014

Ouyang, L., Shi, Z., Zhao, S., Wang, F.-T., Zhou, T.-T., Liu, B., Bao, J.-K., 2012. Programmed cell death pathways in cancer: a review of apoptosis, autophagy and programmed necrosis. Cell Prolif. 45, 487–498. doi:10.1111/j.1365-2184.2012.00845.x

Reynolds, C.P., Maurer, B.J., 2005. Evaluating response to antineoplastic drug combinations in tissue culture models. Methods Mol. Med. 110, 173–183. doi:10.1385/1-59259-869-2:173

Ridley, A.J., 2015. Rho GTPase signalling in cell migration. Curr. Opin. Cell Biol., Cell adhesion and migration 36, 103–112. doi:10.1016/

Shen, H., Perez, R.E., Davaadelger, B., Maki, C.G., 2013. Two 4N Cell-Cycle Arrests Contribute to Cisplatin-Resistance. PLoS ONE 8, e59848. doi:10.1371/journal.pone.0059848

Tao, L., Li, J., Zhang, J., 2013. Brazilein, a compound isolated from Caesalpinia sappan Linn., induced growth inhibition in breast cancer cells via involvement of GSK-3β/β-Catenin/cyclin D1 pathway. Chem. Biol. Interact. 206, 1–5. doi:10.1016/j.cbi.2013.07.015

Tao, L., Li, J., Zhang, J., 2011. Brazilein Induced Cells Apoptosis in Human Breast Cancer MCF-7 Cells and Its Action Mechanism. J. Sun Yat-Sen Univ. Med. Sci. 32, 449–453.

Tsubaki, M., Komai, M., Fujimoto, S., Itoh, T., Imano, M., Sakamoto, K., Shimaoka, H., Takeda, T., Ogawa, N., Mashimo, K., Fujiwara, D., Mukai, J., Sakaguchi, K., Satou, T., Nishida, S., 2013. Activation of NF-κB by the RANKL/RANK system up-regulates snail and twist expressions and induces epithelial-to-mesenchymal transition in mammary tumor cell lines. J. Exp. Clin. Cancer Res. CR 32, 62. doi:10.1186/1756-9966-32-62

Valls, G., Codina, M., Miller, R.K., Valle-Pérez, B.D., Vinyoles, M., Caelles, C., McCrea, P.D., Herreros, A.G. de, Duñach, M., 2012. Upon Wnt stimulation, Rac1 activation requires Rac1 and Vav2 binding to p120-catenin. J Cell Sci 125, 5288–5301. doi:10.1242/jcs.101030

Zhang, J., Wang, Z., Hu, X., Wang, B., Wang, L., Yang, W., Liu, Y., Liu, G., Di, G., Hu, Z., Wu, J., Shao, Z., 2015. Cisplatin and gemcitabine as the first line therapy in metastatic triple negative breast cancer. Int. J. Cancer 136, 204–211. doi:10.1002/ijc.28966

Zhao, K., Wei, L., Hui, H., Dai, Q., You, Q.-D., Guo, Q.-L., Lu, N., 2014. Wogonin Suppresses Melanoma Cell B16-F10 Invasion and Migration by Inhibiting Ras-Medicated Pathways. PLOS ONE 9, e106458. doi:10.1371/journal.pone.0106458


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