Apigenin induces caspase-dependent apoptosis by inhibiting signal transducer and activator of transcription 3 signaling in HER2-overexpressing SKBR3 breast cancer cells

Seo,Hye‑Sook; Ku,Jin Mo; Choi,Han‑Seok; Woo,Jong‑Kyu; Jang,Bo‑Hyoung; Go,Hoyeon; Shin,Yong Cheol; Ko,Seong‑Gyu

doi:10.3892/mmr.2015.3698

Apigenin induces caspase-dependent apoptosis by inhibiting signal transducer and activator of transcription 3 signaling in HER2-overexpressing SKBR3 breast cancer cells

Authors:
- Hye‑Sook Seo
- Jin Mo Ku
- Han‑Seok Choi
- Jong‑Kyu Woo
- Bo‑Hyoung Jang
- Hoyeon Go
- Yong Cheol Shin
- Seong‑Gyu Ko
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Affiliations: Laboratory of Clinical Biology and Pharmacogenomics and Center for Clinical Research and Genomics, College of Korean Medicine, Kyung Hee University, Dongdaemun‑gu, Seoul 130‑701, Republic of Korea, College of Pharmacy, Gachon University of Medicine and Science, Yeonsu‑gu, Incheon 406‑840, Republic of Korea, Department of Oriental Medicine, Semyung University, College of Korean Medicine, Jecheon, Chungbuk 390‑711, Republic of Korea
Published online on: April 28, 2015 https://doi.org/10.3892/mmr.2015.3698
Pages: 2977-2984

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Abstract

Phytoestrogens have been demonstrated to inhibit tumor induction; however, their molecular mechanisms of action have remained elusive. The present study aimed to investigate the effects of a phytoestrogen, apigenin, on proliferation and apoptosis of the human epidermal growth factor receptor 2 (HER2)‑expressing breast cancer cell line SKBR3. Proliferation assay, MTT assay, fluorescence‑activated cell sorting analysis, western blot analysis, immunocytochemistry, reverse transcription‑polymerase chain reaction and ELISA assay were used in the present study. The results of the present study indicated that apigenin inhibited the proliferation of SKBR3 cells in a dose‑ and time‑dependent manner. This inhibition of growth was accompanied by an increase in the sub‑G0/G1 apoptotic population. Furthermore, apigenin enhanced the expression levels of cleaved caspase‑8 and ‑3, and induced the cleavage of poly(adenosine diphosphate ribose) polymerase in SKBR3 cells, confirming that apigenin promotes apoptosis via a caspase‑dependent pathway. Apigenin additionally reduced the expression of phosphorylated (p)‑janus kinase 2 and p‑signal transducer and activator of transcription 3 (STAT3), inhibited CoCl2‑induced vascular endothelial growth factor (VEGF) secretion and decreased the nuclear localization of STAT3. The STAT3 inhibitor S31‑201 decreased the cellular proliferation rate and reduced the expression of p‑STAT3 and VEGF. Therefore, these results suggested that apigenin induced apoptosis via the inhibition of STAT3 signaling in SKBR3 cells. In conclusion, the results of the present study indicated that apigenin may be a potentially useful compound for the prevention or treatment of HER2-overexpressing breast cancer.

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1	Shukla S and Gupta S: Apigenin: A promising molecule for cancer prevention. Pharm Res. 27:962–978. 2010. View Article : Google Scholar : PubMed/NCBI
2	Rezai-Zadeh K, Ehrhart J, Bai Y, Sanberg PR, Bickford P, Tan J and Shytle RD: Apigenin and luteolin modulate microglial activation via inhibition of STAT1-induced CD40 expression. J Neuroinflammation. 5:412008. View Article : Google Scholar : PubMed/NCBI
3	Moon DO, Kim MO, Choi YH, Lee HG, Kim ND and Kim GY: Gossypol suppresses telomerase activity in human leukemia cells via regulating hTERT. FEBS Lett. 582:367–373. 2008. View Article : Google Scholar
4	Nakazawa T, Yasuda T, Ueda J and Ohsawa K: Antidepressant-like effects of apigenin and 2,4,5-trimethoxycinnamic acid from Perilla frutescens in the forced swimming test. Biol Pharm Bull. 26:474–480. 2003. View Article : Google Scholar : PubMed/NCBI
5	Hashemi M, Nouri Long M, Entezari M, Nafisi S and Nowroozii H: Anti-mutagenic and pro-apoptotic effects of apigenin on human chronic lymphocytic leukemia cells. Acta Med Iran. 48:283–288. 2010.
6	Patel D, Shukla S and Gupta S: Apigenin and cancer chemoprevention: progress, potential and promise (review). Int J Oncol. 30:233–245. 2007.
7	Shukla S and Gupta S: Molecular targets for apigenin-induced cell cycle arrest and apoptosis in prostate cancer cell xenograft. Mol Cancer Ther. 5:843–852. 2006. View Article : Google Scholar : PubMed/NCBI
8	Liang YC, Huang YT, Tsai SH, Lin-Shiau SY, Chen CF and Lin JK: Suppression of inducible cyclooxygenase and inducible nitric oxide synthase by apigenin and related flavonoids in mouse macrophages. Carcinogenesis. 20:1945–1952. 1999. View Article : Google Scholar : PubMed/NCBI
9	Birt DF, Mitchell D, Gold B, Pour P and Pinch HC: Inhibition of ultraviolet light induced skin carcinogenesis in SKH-1 mice by apigenin, a plant flavonoid. Anticancer Res. 17:85–91. 1997.PubMed/NCBI
10	Seo HS, Ju JH, Jang K and Shin I: Induction of apoptotic cell death by phytoestrogens by up-regulating the levels of phospho-p53 and p21 in normal and malignant estrogen receptor α-negative breast cells. Nutrition Res. 31:139–146. 2011. View Article : Google Scholar
11	Lu HF, Chie YJ, Yang MS, Lee CS, Fu JJ, Yang JS, Tan TW, Wu SH, Ma YS, Ip SW and Chung JG: Apigenin induces caspase-dependent apoptosis in human lung cancer A549 cells through Bax- and Bcl-2-triggered mitochondrial pathway. Int J Oncol. 36:1477–1484. 2010.PubMed/NCBI
12	Wang W, Heideman L, Chung CS, Pelling JC, Koehler KJ and Birt DF: Cell-cycle arrest at G2/M and growth inhibition by apigenin in human colon carcinoma cell lines. Mol Carcinog. 28:102–110. 2000. View Article : Google Scholar : PubMed/NCBI
13	Turktekin M, Konac E, Onen HI, Alp E, Yilmaz A and Menevse S: Evaluation of the effects of the flavonoid apigenin on apoptotic pathway gene expression on the colon cancer cell line (HT29). J Med Food. 14:1107–1117. 2011. View Article : Google Scholar : PubMed/NCBI
14	Gupta S, Afaq F and Mukhtar H: Involvement of nuclear factor-kappa B, Bax and Bcl-2 in induction of cell cycle arrest and apoptosis by apigenin in human prostate carcinoma cells. Oncogene. 21:3727–3738. 2002. View Article : Google Scholar : PubMed/NCBI
15	Ruela-de-Sousa RR, Fuhler GM, Blom N, Ferreira CV, Aoyama H and Peppelenbosch MP: Cytotoxicity of apigenin on leukemia cell lines: implications for prevention and therapy. Cell Death Dis. 1:e192010. View Article : Google Scholar : PubMed/NCBI
16	Ujiki MB, Ding XZ, Salabat MR, Bentrem DJ, Golkar L, Milam B, Talamonti MS, Bell RH Hr, Iwamura T and Adrian TE: Apigenin inhibits pancreatic cancer cell proliferation through G2/M cell cycle arrest. Mol Cancer. 5:762006. View Article : Google Scholar
17	Fan TJ, Han LH, Cong RS and Liang J: Caspase family proteases and apoptosis. Acta Biochim Biophys Sin (Shanghai). 37:719–727. 2005. View Article : Google Scholar
18	Bosch M, Poulter NS, Vatovec S and Franklin-Ton VE: Initiation of programmed cell death in self-incompatibility: role for cytoskeleton modifications and several caspase-like activities. Mol Plant. 1:879–887. 2008. View Article : Google Scholar
19	Zhang A, Wu Y, Lai HWL and Yew DT: Apoptosis-a brief review. Neuroembryology. 3:47–59. 2004. View Article : Google Scholar
20	Waring P and Mullbacher A: Cell death induced by the Fas/Fas ligand pathway and its role in pathology. Immunol Cell Biol. 77:312–317. 1999. View Article : Google Scholar : PubMed/NCBI
21	Gupta S: Molecular signaling in death receptor and mitochondrial pathways of apoptosis (review). Int J Oncol. 22:15–20. 2003.
22	Green DR and Reed JC: Mitochondria and apoptosis. Science. 281:1309–1312. 1998. View Article : Google Scholar : PubMed/NCBI
23	Boulares AH, Yakovlev AG, Ivanova V, Stoica BA, Wang G, Iyer S and Smulson M: Role of poly(ADP-ribose) polymerase (PARP) cleavage in apoptosis. Caspase 3-resistant PARP mutant increases rates of apoptosis in transfected cells. J Biol Chem. 274:22932–22940. 1999. View Article : Google Scholar : PubMed/NCBI
24	Wilson CA, Cajulis EE, Green JL, Olsen TM, Chung YA, Damore MA, Dering J, Calzone FJ and Slamon DJ: HER-2 over-expression differentially alters transforming growth factor-beta responses in luminal versus mesenchymal human breast cancer cells. Breast Cancer Res. 7:R1058–R1079. 2005. View Article : Google Scholar
25	Prat A, Carey LA, Adamo B, Vidal M, Tabernero J, Cortés J, Parker JS, Perou CM and Baselga J: Molecular features and survival outcomes of the intrinsic subtypes within HER2-positive breast cancer. J Natl Cancer Inst. 106:dju1522014. View Article : Google Scholar : PubMed/NCBI
26	Joshi JP, Brown NE, Griner SE and Nahta R: Growth differentiation factor 15 (GDF15)-mediated HER2 phosphorylation reduces trastuzumab sensitivity of HER2-overexpressing breast cancer cells. Biochem Pharmacol. 82:1090–1099. 2011. View Article : Google Scholar : PubMed/NCBI
27	Favoni RE, Daga A, Malatesta P and Florio T: Preclinical studies identify novel targeted pharmacological strategies for treatment of human malignant pleural mesothelioma. Br J Pharmacol. 166:532–553. 2012. View Article : Google Scholar : PubMed/NCBI
28	Tokunaga E, Oki E, Nishida K, Koga T, Egashira A, Morita M, Kakeji Y and Maehara Y: Trastuzumab and breast cancer: Developments and current status. Int J Clin Oncol. 11:199–208. 2006. View Article : Google Scholar : PubMed/NCBI
29	Dean-Colomb W and Esteva FJ: Her2-positive breast cancer: Herceptin and beyond. Eur J Cancer. 44:2806–2812. 2008. View Article : Google Scholar : PubMed/NCBI
30	Seo HS, Choi HS, Kim SR, Choi YK, Woo SM, Shin I, Woo JK, Park SY, Shin YC and Ko SG: Apigenin induces apoptosis via extrinsic pathway, inducing p53 and inhibiting STAT3 and NFκB signaling in HER2-overexpressing breast cancer cells. Mol Cell Biochem. 366:319–334. 2012. View Article : Google Scholar : PubMed/NCBI
31	Earnshaw WC, Martins LM and Kaufmann SH: Mammalian caspases: structure, activation, substrates and functions during apoptosis. Annu Rev Biochem. 68:383–424. 1999. View Article : Google Scholar
32	Blagosklonny MV, An WG, Romanova LY, Trepel J, Fojo T and Neckers L: p53 inhibits hypoxia-inducible factor-stimulated transcription. J Biol Chem. 273:11995–11998. 1998. View Article : Google Scholar : PubMed/NCBI
33	Niu G, Wright KL, Huang M, Song L, Haura E, Turkson J, Zhang S, Wang T, Sinibaldi D, Coppola D, Heller R, Ellis LM, Karras J, Bromberg J, Pardoll D, Jove R and Yu H: Constitutive Stat3 activity up-regulates VEGF expression and tumor angiogenesis. Oncogene. 21:2000–2008. 2002. View Article : Google Scholar : PubMed/NCBI
34	Forsythe JA, Jiang BH, Iyer NV, Agani F, Leung SW, Koos RD and Semenza GL: Activation of vascular endothelial growth factor gene transcription by Hypoxia-inducible factor 1. Mol Cell Biol. 16:4604–4613. 1996.PubMed/NCBI
35	Jung JE, Lee HG, Cho IH, Chung DH, Yoon SH, Yang YM, Lee JW, Choi S, Park JW, Ye SK and Chung MH: STAT3 is a potential modulator of HIF-1-mediated VEGF expression in human renal carcinoma cells. FASEB J. 19:1296–1298. 2005.PubMed/NCBI
36	Hagemann T, Robinson SC, Schulz M, Trümper L, Balkwill FR and Binder C: Enhanced invasiveness of breast cancer cell lines upon co-cultivation with macrophages is due to TNF-alpha dependent up-regulation of matrix metalloproteases. Carcinogenesis. 25:1543–1549. 2004. View Article : Google Scholar : PubMed/NCBI
37	de la Iglesia N, Konopka G, Puram SV, Chan JA, Bachoo RM, You MJ, Levy DE, DEPinho RA and Bonni A: Identification of a PTEN-regulated STAT3 brain tumor suppressor pathway. Genes Dev. 22:449–462. 2008. View Article : Google Scholar : PubMed/NCBI
38	Lewis HD, Winter A, Murphy TF, Tripathi S, Pandey VN and Barton BE: STAT3 inhibition in prostate and pancreatic cancer lines by STAT3 binding sequence oligonucleotides: Differential activity between 5′ and 3′ ends. Mol Cancer Ther. 7:1543–1550. 2008. View Article : Google Scholar : PubMed/NCBI
39	Kortylewski M, Jove R and Yu H: Targeting STAT3 affects melanoma on multiple fronts. Cancer Metastasis Rev. 24:315–327. 2005. View Article : Google Scholar : PubMed/NCBI
40	Niu G, Bowman T, Huang M, Shivers S, Reintgen D, Daud A, Chang A, Kraker A, Jove R and Yu H: Roles of activated Src and Stat3 signaling in melanoma tumor cell growth. Oncogene. 21:7001–7010. 2002. View Article : Google Scholar : PubMed/NCBI
41	Xie TX, Huang FJ, Aldape KD, Kang SH, Liu M, Gershenwald JE, Xie K, Sawaya R and Huang S: Activation of stat3 in human melanoma promotes brain metastasis. Cancer Res. 66:3188–3196. 2006. View Article : Google Scholar : PubMed/NCBI
42	Sellers LA, Feniuk W, Humphrey PP and Lauder H: Activated G proteincoupled receptor induces tyrosine phosphorylation of STAT3 and agonist-selective serine phosphorylation via sustained stimulation of mitogen-activated protein kinase. Resultant effects on cell proliferation. J Biol Chem. 274:16423–16430. 1999. View Article : Google Scholar : PubMed/NCBI
43	Zhang Y, Turkson J, Carter-Su C, Smithgall T, Levitzki A, Kraker A, Krolewski JJ, Medveczky P and Jove R: Activation of Stat3 in v-Src-transformed fibroblasts requires cooperation of Jak1 kinase activity. J Biol Chem. 275:24935–24944. 2000. View Article : Google Scholar : PubMed/NCBI
44	Kotha A, Sekharam M, Cilenti L, Siddiquee K, Khaled A, Zervos AS, Carter B, Turkson J and Jove R: Resveratrol inhibits src and Stat3 signaling and induces the apoptosis of malignant cells containing activated Stat3 protein. Mol Cancer Ther. 5:621–629. 2006. View Article : Google Scholar : PubMed/NCBI
45	Tolaney SM and Krop IE: Mechanisms of trastuzumab resistance in breast cancer. Anticancer Agents Med Chem. 9:348–355. 2009. View Article : Google Scholar : PubMed/NCBI
46	Buzdar AU: Role of biologic therapy and chemotherapy in hormone receptor- and HER2-positive breast cancer. Ann Oncol. 20:993–999. 2009. View Article : Google Scholar : PubMed/NCBI