Ethanolic Extract of Propolis from Kerman Area Triggers Apoptosis and Arrests Cell Cycle in Three Human Breast Cancer Cell Lines MDA-MB-231, SKBR and MCF-7

Document Type : Original Article


1 Ph.D. student, Department of Pathobiology School of Veterinary Medicine, Shiraz University, Shiraz, Iran

2 Associate Professor, Department of Basic Sciences, School of Veterinary Medicine, Shiraz University, Shiraz, Iran

3 Associate Professor, Department of Biotechnology, Institute of Science and High Technology and Environmental Sciences, Graduate University of Advanced Technology, Kerman, Iran

4 Professor, Department of pharmaceutics, Faculty of pharmacy and pharmaceutical sciences, Kerman University of Medical Sciences, Kerman, Iran


Background: Cancer is one of the major health problems worldwide and natural resources are being explored to develop anticancer drugs with fewer side effects. Iranian propolis contains components including flavonoids and polyphenols and has various medicinal properties. The aim of this study was to investigate the effect of Ethanolic Extract of Sirch Propolis (EESP) on three breast cancer cell lines.
Methods: The MDA-MB-231, SKBR-3 and MCF-7 cells were treated for 24 and 48 h at the presence of 1% and 10% fetal bovine serum (FBS) concentration. MTT, BrdU and flow cytometry assays were used for measuring cytotoxicity, cell proliferation and apoptosis.
Results: The highest cytotoxicity was seen on MDA-MB-231 cell at the presence of 1% and 10% FBS respectively following 48 h treatment. BrdU assay showed that treatment with 200 μg/ mL of EESP at the presence of 1% FBS for 48 h, reduced proliferation of MDA-MB-231 cell to 75% and that of MCF-7 and SKBR-3 cells to 70% and 60% respectively. Cell cycle analysis by flow cytometry showed that EESP at 200 μg/mL for 48h, induced G0/G1 phase arrest in MCF-7 and SKBR-3 cells and G2/M, S phase arrest in MDA-MB-231 cell. The cytotoxic effects of EESP were primarily found to be due to the induction of early stage apoptosis on SKBR-3 cell and early and late stage apoptosis on MCF-7 and MDA-MB-231 cells.
Conclusion:The results demonstrated that EESP is a natural anticancer mixture capable of reducing breast cancer cells proliferation and inducing cell cycle arrest and apoptosis in them.


  1. Coughlin SS, Ekwueme DU. Breast cancer as a global health concern. Cancer Epidemiol 2009; 33(5):315-8.
  2. Ferlay J, Hery C, Autier P, Sankaranarayanan R. Global burden of breast cancer. In: Li C. Breast Cancer Epidemiology. New York: Springer; 2010. p.1-19.
  3. Chen W, Zheng R, Baade PD, Zhang S, Zeng H, Bray F, et al. Cancer statistics in China, 2015. CA Cancer J Clin 2016; 66(2):115-32.
  4. Bray F, Ferlay J, Soerjomataram I, Siegel RL, Torre LA, Jemal A. Global cancer statistics 2018: globocan estimates of incidence and mortality worldwide for 36 cancers in 185 countries. CA Cancer J Clin 2018; 68(6):394-424.
  5. Mitra S, Dash R. Natural products for the management and prevention of breast cancer. Evid Based Complement Alternat Med 2018; 2018:8324696.
  6. Sanders K, Moran Z, Shi Z, Paul R, Greenlee H. Natural products for cancer prevention: clinical update 2016. Semin Oncol Nurs 2016; 32(3):215-40.
  7. Zhang QY, Wang FX, Jia KK, Kong LD. Natural product interventions for chemotherapy and radiotherapy-induced side effects. Front Pharmacol 2018; 9:1253.
  8. Newman DJ, Cragg GM. Natural products as sources of new drugs from 1981 to 2014. J Nat Prod 2016; 79(3):629-61.
  9. Pietta PG, Gardana C, Pietta AM. Analytical methods for quality control of propolis. Fitoterapia 2002; 73(Suppl 1):S7-20.
  10. Bankova V, Christov R, Kujumgiev A, Marcucci MC, Popov S. Chemical composition and antibacterial activity of Brazilian propolis. Z Naturforsch C J Biosci 1995; 50(3-4):167-72.
  11. Orsolic N, Basic I. Water-soluble derivative of propolis and its polyphenolic compounds enhance tumoricidal activity of macrophages. J Ethnopharmacol 2005; 102(1):37-45.
  12. Burdock GA. Review of the biological properties and toxicity of bee propolis (propolis). Food Chem Toxicol 1998; 36(4):347-63.
  13. Popolo A, Piccinelli LA, Morello S, Cuesta-Rubio O, Sorrentino R, Rastrelli L, et al. Antiproliferative activity of brown Cuban propolis extract on human breast cancer cells. Nat Prod Commun 2009; 4(12):1711-6.
  14. Xuan H, Li Z, Yan H, Sang Q, Wang K, He Q, et al. Antitumor activity of chinese propolis in human breast cancer MCF-7 and MDA-MB-231 cells. Evidence-Based Complementary and Alternative Medicine 2014; 2014:280120.
  15. Kabala-Dzik A, Rzepecka-Stojko A, Kubina R, Iriti M, Wojtyczka RD, Buszman E, et al. Flavonoids, bioactive components of propolis, exhibit cytotoxic activity and induce cell cycle arrest and apoptosis in human breast cancer cells MDA-MB-231 and MCF-7 – a comparative study. Cell Mol Biol (Noisy-le-grand) 2018; 64(8):1-10.
  16. Omene C, Kalac M, Wu J, Marchi E, Frenkel K, O’Connor OA. Propolis and its Active Component, Caffeic Acid Phenethyl Ester (CAPE), modulate breast cancer therapeutic targets via an epigenetically mediated mechanism of action. J Cancer Sci Ther 2013; 5(10):334-42.
  17. Padmavathi R, Senthilnathan P, Chodon D, Sakthisekaran D. Therapeutic effect of paclitaxel and propolis on lipid peroxidation and antioxidant system in 7,12 dimethyl benz(a)anthracene-induced breast cancer in female Sprague Dawley rats. Life Sci 2006; 78(24):2820-5.
  18. Seyhan MF, Yilmaz E, Timirci-Kahraman O, Saygili N, Kisakesen HI, Gazioglu S, et al. Different propolis samples, phenolic content, and breast cancer cell lines: Variable cytotoxicity ranging from ineffective to potent. IUBMB Life 2018; 71(5):619-31.
  19. Muli EM, Maingi JM, Macharia J. Antimicrobial properties of propolis and honey from the Kenyan stingless bee, Dactylurina schimidti. Apiacta 2008; 43:49-61.
  20. Yemis O, Bakkalbaşi E, Artik N. Antioxidative activities of grape (Vitis vinifera) seed extracts obtained from different varieties grown in Turkey. Int J Food Sci Technol 2009; 43(1):154-9.
  21. Barlak Y, Deger O, Colak M, Karatayli SC, Bozdayi AM, Yucesan F. Effect of Turkish propolis extracts on proteome of prostate cancer cell line. Proteome Sci 2011; 9:74.
  22. Chan GC, Cheung KW, Sze DM. The immunomodulatory and anticancer properties of propolis. Clin Rev Allergy Immunol 2013; 44(3):262-73.
  23. Frion-Herrera Y, Diaz-Garcia A, Ruiz-Fuentes J, Rodriguez-Sanchez H, Mauricio Sforcin J. Mechanisms underlying the cytotoxic effect of propolis on human laryngeal epidermoid carcinoma cells. Nat Prod Res 2018; 32(17):2085-91.
  24. Aguero MB, Svetaz L, Sanchez M, Luna L, Lima B, Lopez ML, et al. Argentinean Andean propolis associated with the medicinal plant Larrea nitida Cav. (Zygophyllaceae). HPLC-MS and GC-MS characterization and antifungal activity. Food Chem Toxicol 2011; 49(9):1970-8.
  25. Frion-Herrera Y, Diaz-Garcia A, Ruiz-Fuentes J, Rodriguez-Sanchez H, Sforcin JM. The cytotoxic effects of propolis on breast cancer cells involve PI3K/Akt and ERK1/2 pathways, mitochondrial membrane potential, and reactive oxygen species generation. Inflammopharmacology 2019; 27(5):1081-9.
  26. Kamiya T, Nishihara H, Hara H, Adachi T. Ethanol extract of Brazilian red propolis induces apoptosis in human breast cancer MCF-7 cells through endoplasmic reticulum stress. J Agric Food Chem 2012; 60(44):11065-70.
  27. Popolo A, Piccinelli AL, Morello S, Sorrentino R, Osmany CR, Rastrelli L, et al. Cytotoxic activity of nemorosone in human MCF-7 breast cancer cells. Can J Physiol Pharmacol 2011; 89(1):50-7.
  28. Zhou K, Li X, Du Q, Li D, Hu M, Yang X, et al. A CAPE analogue as novel antiplatelet agent efficiently inhibits collagen-induced platelet aggregation. Pharmazie 2014; 69(8):615-20.
  29. Seda Vatansever H, Sorkun K, Ismet Deliloglu Gurhan S, Ozdal-Kurt F, Turkoz E, Gencay O, et al. Propolis from Turkey induces apoptosis through activating caspases in human breast carcinoma cell lines. Acta Histochem 2010; 112(6):546-56.
  30. ZD Z, Cox G. MTT assay overestimates human airway smooth muscle cell number in culture. Biochem Mol Biol Int 1996; 38(3):431-6.
  31. Peters T. All About Albumin: Biochemistry, Genetics and Medical Applications. USA: Academic Press; 1995.
  32. Newman JM, DiMaria CA, Rattigan S, Steen JT, Miller KA, Eldershaw TP, et al. Relationship of MTT reduction to stimulants of muscle metabolism. Chem Biol Interact 2000; 128(2):127-40.
  33. Funk D, Schrenk HH, Frei E. Serum albumin leads to false-positive results in the XTT and the MTT assay. Biotechniques 2007; 43(2):178.
  34. Rzepecka-Stojko A, Kabala-Dzik A, Mozdzierz A, Kubina R, Wojtyczka RD, Stojko R, et al. Caffeic Acid phenethyl ester and ethanol extract of propolis induce the complementary cytotoxic effect on triple-negative breast cancer cell lines. Molecules 2015; 20(5):9242-62.
  35. Frozza CO, Ribeiro Tda S, Gambato G, Menti C, Moura S, Pinto PM, et al. Proteomic analysis identifies differentially expressed proteins after red propolis treatment in Hep-2 cells. Food Chem Toxicol 2014; 63:195-204.
  36. Ratna D, Simanjuntak P, Abdillah S, Heffen WL. Apoptosis of human breast cancer cells induced by ethylacetate extracts of Propolis. American Journal of Biochemistry and Biotechnology 2010; 6(2):84-8.
  37. Chang H, Wang Y, Yin X, Liu X, Xuan H. Ethanol extract of propolis and its constituent caffeic acid phenethyl ester inhibit breast cancer cells proliferation in inflammatory microenvironment by inhibiting TLR4 signal pathway and inducing apoptosis and autophagy. BMC Complement Altern Med 2017; 17(1):471.
  38. Noureddine H, Hage-Sleiman R, Wehbi B, Fayyad-Kazan H, Hayar S, Traboulssi M, et al. Chemical characterization and cytotoxic activity evaluation of Lebanese propolis. Biomed Pharmacother 2017; 95:298-307.
  39. Sawicka D, Car H, Borawska MH, Niklinski J. The anticancer activity of propolis. Folia Histochem Cytobiol 2012; 50(1):25-37.
  40. Ahn MR, Kumazawa S, Hamasaka T, Bang KS, Nakayama T. Antioxidant activity and constituents of propolis collected in various areas of Korea. J Agric Food Chem 2004; 52(24):7286-92.
  41. Li LK, Rola AS, Kaid FA, Ali AM, Alabsi AM. Goniothalamin induces cell cycle arrest and apoptosis in H400 human oral squamous cell carcinoma: a caspase-dependent mitochondrial-mediated pathway with downregulation of NF- kb. Arch Oral Biol 2016; 64:28-38
  42. Lirdprapamongkol K, Sakurai H, Abdelhamed S, Yokoyama S, Maruyama T, Athikomkulchai S, et al. A flavonoid chrysin suppresses hypoxic survival and metastatic growth of mouse breast cancer cells. Oncol Rep 2013; 30(5):2357-64.