2-Methoxyestradiol inhibits the proliferation and migration and reduces the radioresistance of nasopharyngeal carcinoma CNE-2 stem cells via NF-κB/HIF-1 signaling pathway inactivation and EMT reversal

Wu,Shun-Long; Li,Ya-Jun; Liao,Kui; Shi,Lei; Zhang,Na; Liu,Shuang; Hu,Yao-Yao; Li,Shao-Lin; Wang,Ying

doi:10.3892/or.2016.5319

2-Methoxyestradiol inhibits the proliferation and migration and reduces the radioresistance of nasopharyngeal carcinoma CNE-2 stem cells via NF-κB/HIF-1 signaling pathway inactivation and EMT reversal

Authors:
- Shun-Long Wu
- Ya-Jun Li
- Kui Liao
- Lei Shi
- Na Zhang
- Shuang Liu
- Yao-Yao Hu
- Shao-Lin Li
- Ying Wang
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Affiliations: Department of Oncology, The First Affiliated Hospital of Chongqing Medical University, Chongqing 400016, P.R. China, Zunyi Medical College, Zunyi, Guizhou 563000, P.R. China, Department of Oncology, Affiliated Hospital of Zunyi Medical College, Zunyi, Guizhou 563000, P.R. China, Department of Oncology and Radiation Medicine, College of Basic Medicine, Chongqing Medical University, Chongqing 400016, P.R. China, Department of Radiotherapy, Chongqing Cancer Institute, Chongqing 400030, P.R. China
Published online on: December 14, 2016 https://doi.org/10.3892/or.2016.5319
Pages: 793-802

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Abstract

Accumulating evidence indicates that cancer stem cells (CSCs) are a source of resistance to radiation therapy (RT); however, the mechanism of this resistance remains unclear. 2-Methoxyestradiol (2-ME2) is a metabolic product of estrogen in the body. Recent studies have found that 2-ME2 regulates the activation of transcription factors, including nuclear factor (NF)-κB/hypoxia-inducible factor-1 (HIF-1), thus contributing to tumor cell apoptosis and chemosensitivity. Therefore, 2-ME2 is being studied as a potential anticancer drug. The purpose of this study was to determine the effect and mechanism by which 2-ME2 inhibits nasopharyngeal carcinoma CNE-2 stem-like cell (NPCSC) proliferation and migration and reduces NPCSC radioresistance. This study has important significance for reducing the radioresistance of these cells to improve the cure rate of NPC. First, the NPCSCs were collected in a serum-free culture system and then identified by relevant experiments. The NPCSCs were treated with 2-ME2 (0-8 µM) combined with X-ray exposure and Cell Counting Kit-8 (CCK-8), Transwell assay, colony formation assay, western blot analysis, RT-PCR, flow cytometry and RNA interference technology were used to explore the effect and mechanism of 2-ME2 on NPCSCs. The results showed that the microspheres collected in the serum‑free culture system possessed CSC traits and radioresistance. 2-ME2 obviously inhibited NPCSC growth and migration and reduced NPCSC radioresistance. 2-ME2 decreased NF-κB p65 and HIF-1α protein expression, downregulated NF-κB p65 nuclear localization, and reversed epithelial-mesenchymal transition (EMT). NF-κB p65 knockdown reduced HIF-1α expression, reversed EMT, and enhanced the suppressive effect of 2-ME2 on NPCSCs. Collectively, these data indicate that 2-ME2 inhibits NPCSC proliferation and migration and reduces the radioresistance of NPCSCs via NF-κB/HIF-1 signaling pathway inactivation and EMT reversal.

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1	Peng G, Wang T, Yang KY, Zhang S, Zhang T, Li Q, Han J and Wu G: A prospective, randomized study comparing outcomes and toxicities of intensity-modulated radiotherapy vs. conventional two-dimensional radiotherapy for the treatment of nasopharyngeal carcinoma. Radiother Oncol. 104:286–293. 2012. View Article : Google Scholar : PubMed/NCBI
2	Wang R, Wu F, Lu H, Wei B, Feng G, Li G, Liu M, Yan H, Zhu J, Zhang Y, et al: Definitive intensity-modulated radiation therapy for nasopharyngeal carcinoma: Long-term outcome of a multicenter prospective study. J Cancer Res Clin Oncol. 139:139–145. 2013. View Article : Google Scholar : PubMed/NCBI
3	Yang CF, Peng LX, Huang TJ, Yang GD, Chu QQ, Liang YY, Cao X, Xie P, Zheng LS, Huang HB, et al: Cancer stem-like cell characteristics induced by EB virus-encoded LMP1 contribute to radioresistance in nasopharyngeal carcinoma by suppressing the p53-mediated apoptosis pathway. Cancer Lett. 344:260–271. 2014. View Article : Google Scholar : PubMed/NCBI
4	Wang WJ, Wu SP, Liu JB, Shi YS, Huang X, Zhang QB and Yao KT: MYC regulation of CHK1 and CHK2 promotes radioresistance in a stem cell-like population of nasopharyngeal carcinoma cells. Cancer Res. 73:1219–1231. 2013. View Article : Google Scholar : PubMed/NCBI
5	Wei P, Niu M, Pan S, Zhou Y, Shuai C, Wang J, Peng S and Li G: Cancer stem-like cell: A novel target for nasopharyngeal carcinoma therapy. Stem Cell Res Ther. 5:442014. View Article : Google Scholar : PubMed/NCBI
6	Zhu Y, Liu H, Xu L, An H, Liu W, Liu Y, Lin Z and Xu J: p21-activated kinase 1 determines stem-like phenotype and sunitinib resistance via NF-κB/IL-6 activation in renal cell carcinoma. Cell Death Dis. 6:e16372015. View Article : Google Scholar : PubMed/NCBI
7	Vinogradov S and Wei X: Cancer stem cells and drug resistance: The potential of nanomedicine. Nanomedicine (Lond). 7:597–615. 2012. View Article : Google Scholar : PubMed/NCBI
8	Meijer TW, Kaanders JH, Span PN and Bussink J: Targeting hypoxia, HIF-1, and tumor glucose metabolism to improve radiotherapy efficacy. Clin Cancer Res. 18:5585–5594. 2012. View Article : Google Scholar : PubMed/NCBI
9	Scholz CC, von Kriegsheim A, Tambuwala MM, Hams E, Cheong A, Bruning U, Fallon PG, Cummins EP and Taylor CT: Prolyl Hydroxylase 1 (PHD1) and Factor Inhibiting HIF (FIH) regulate IL-1β-induced NF-κB activity linking key hypoxic and inflammatory signaling pathways. FASEB J. 27:717–719. 2013.
10	Wong JH, Lui VW, Umezawa K, Ho Y, Wong EY, Ng MH, Cheng SH, Tsang CM, Tsao SW and Chan AT: A small molecule inhibitor of NF-kappaB, dehydroxymethylepoxyquinomicin (DHMEQ), suppresses growth and invasion of nasopharyngeal carcinoma (NPC) cells. Cancer Lett. 287:23–32. 2010. View Article : Google Scholar : PubMed/NCBI
11	Kan R, Shuen WH, Lung HL, Cheung AK, Dai W, Kwong DL, Ng WT, Lee AW, Yau CC, Ngan RK, et al: NF-κB p65 subunit is modulated by Latent Transforming Growth Factor-β Binding Protein 2 (LTBP2) in nasopharyngeal carcinoma HONE1 and HK1 cells. PLoS One. 10:e01272392015. View Article : Google Scholar : PubMed/NCBI
12	Li CW, Xia W, Huo L, Lim SO, Wu Y, Hsu JL, Chao CH, Yamaguchi H, Yang NK, Ding Q, et al: Epithelial-mesenchymal transition induced by TNF-α requires NF-κB-mediated transcriptional upregulation of Twist1. Cancer Res. 72:1290–1300. 2012. View Article : Google Scholar : PubMed/NCBI
13	Zhao D, Tang XF, Yang K, Liu JY and Ma XR: Over-expression of integrin-linked kinase correlates with aberrant expression of Snail, E-cadherin and N-cadherin in oral squamous cell carcinoma: Implications in tumor progression and metastasis. Clin Exp Metastasis. 29:957–969. 2012. View Article : Google Scholar : PubMed/NCBI
14	Kambhampati S, Rajewski RA, Tanol M, Haque I, Das A, Banerjee S, Jha S, Burns D, Borrego-Diaz E, Van Veldhuizen PJ, et al: A second-generation 2-Methoxyestradiol prodrug is effective against Barrett's adenocarcinoma in a mouse xenograft model. Mol Cancer Ther. 12:255–263. 2013. View Article : Google Scholar : PubMed/NCBI
15	Zhang XY, Zhan R, Huang HB and Yang T: Mechanism underlying 2-methoxyestradiol inducing apoptosis of K562 cells. Zhongguo Shi Yan Xue Ye Xue Za Zhi. 17:340–344. 2009.(In Chinese). PubMed/NCBI
16	Kumar AP, Garcia GE, Orsborn J, Levin VA and Slaga TJ: 2-Methoxyestradiol interferes with NFκB transcriptional activity in primitive neuroectodermal brain tumors: Implications for management. Carcinogenesis. 24:209–216. 2003. View Article : Google Scholar : PubMed/NCBI
17	Parrondo R, de las Pozas A, Reiner T, Rai P and Perez-Stable C: NF-κB activation enhances cell death by antimitotic drugs in human prostate cancer cells. Mol Cancer. 9:1822010. View Article : Google Scholar : PubMed/NCBI
18	Semenza GL: Hypoxia-inducible factors: Mediators of cancer progression and targets for cancer therapy. Trends Pharmacol Sci. 33:207–214. 2012. View Article : Google Scholar : PubMed/NCBI
19	Marie-Egyptienne DT, Lohse I and Hill RP: Cancer stem cells, the epithelial to mesenchymal transition (EMT) and radioresistance: Potential role of hypoxia. Cancer Lett. 341:63–72. 2013. View Article : Google Scholar : PubMed/NCBI
20	Conley SJ, Baker TL, Burnet JP, Thiesen RL, Lazarus D, Peters CG, Clouthier SG, Eliasof S and Wicha MS: CRLX101, an investigational camptothecin-containing nanoparticle-drug conjugate, reverses the HIF-1α-mediated increase in cancer stem cells caused by bevacizumab in a preclinical model of triple-negative breast cancer. Cancer Res. 75:(Suppl 15). 13842015. View Article : Google Scholar
21	Kazi AA, Shah P, Schech A, Sabnis G, Chumsri S and Brodie A: Inhibition of non-hypoxic HIF-1 expression in letrozole-resistant breast cancer cells reduces their cancer stem cell characteristics. Cancer Res. 73:(Suppl 8). 952013. View Article : Google Scholar
22	Gammon L and Mackenzie IC: Roles of hypoxia, stem cells and epithelial-mesenchymal transition in the spread and treatment resistance of head and neck cancer. J Oral Pathol Med. 45:77–82. 2016. View Article : Google Scholar : PubMed/NCBI
23	Ricker JL, Chen Z, Yang XP, Pribluda VS, Swartz GM and Van Waes C: 2-methoxyestradiol inhibits hypoxia-inducible factor 1α, tumor growth, and angiogenesis and augments paclitaxel efficacy in head and neck squamous cell carcinoma. Clin Cancer Res. 10:8665–8673. 2004. View Article : Google Scholar : PubMed/NCBI
24	Li YJ, Wu SL, Lu SM, Chen F, Guo Y, Gan SM, Shi YL, Liu S and Li SL: (−)-Epigallocatechin-3-gallate inhibits nasopharyngeal cancer stem cell self-renewal and migration and reverses the epithelial-mesenchymal transition via NF-κB p65 inactivation. Tumour Biol. 36:2747–2761. 2015. View Article : Google Scholar : PubMed/NCBI
25	Fan X, Liu S, Su F, Pan Q and Lin T: Effective enrichment of prostate cancer stem cells from spheres in a suspension culture system. Urol Oncol. 30:314–318. 2012. View Article : Google Scholar : PubMed/NCBI
26	Chen SF, Chang YC, Nieh S, Liu CL, Yang CY and Lin YS: Nonadhesive culture system as a model of rapid sphere formation with cancer stem cell properties. PLoS One. 7:e318642012. View Article : Google Scholar : PubMed/NCBI
27	Kang SH, Cho HT, Devi S, Zhang Z, Escuin D, Liang Z, Mao H, Brat DJ, Olson JJ, Simons JW, et al: Antitumor effect of 2-methoxyestradiol in a rat orthotopic brain tumor model. Cancer Res. 66:11991–11997. 2006. View Article : Google Scholar : PubMed/NCBI
28	Chua YS, Chua YL and Hagen T: Structure activity analysis of 2-methoxyestradiol analogues reveals targeting of microtubules as the major mechanism of antiproliferative and proapoptotic activity. Mol Cancer Ther. 9:224–235. 2010. View Article : Google Scholar : PubMed/NCBI
29	Maier HJ, Schmidt-Strassburger U, Huber MA, Wiedemann EM, Beug H and Wirth T: NF-κB promotes epithelial-mesenchymal transition, migration and invasion of pancreatic carcinoma cells. Cancer Lett. 295:214–228. 2010. View Article : Google Scholar : PubMed/NCBI
30	Liu M, Sakamaki T, Casimiro MC, Willmarth NE, Quong AA, Ju X, Ojeifo J, Jiao X, Yeow WS, Katiyar S, et al: The canonical NF-κB pathway governs mammary tumorigenesis in transgenic mice and tumor stem cell expansion. Cancer Res. 70:10464–10473. 2010. View Article : Google Scholar : PubMed/NCBI
31	Lin Y, Bai L, Chen W and Xu S: The NF-κB activation pathways, emerging molecular targets for cancer prevention and therapy. Expert Opin Ther Targets. 14:45–55. 2010. View Article : Google Scholar : PubMed/NCBI
32	Ray G, Dhar G, Van Veldhuizen PJ, Banerjee S, Saxena NK, Sengupta K and Banerjee SK: Modulation of cell-cycle regulatory signaling network by 2-methoxyestradiol in prostate cancer cells is mediated through multiple signal transduction pathways. Biochemistry. 45:3703–3713. 2006. View Article : Google Scholar : PubMed/NCBI
33	Siebert AE, Sanchez AL, Dinda S and Moudgil VK: Effects of estrogen metabolite 2-methoxyestradiol on tumor suppressor protein p53 and proliferation of breast cancer cells. Syst Biol Reprod Med. 57:279–287. 2011. View Article : Google Scholar : PubMed/NCBI
34	Pasquier E, André N and Braguer D: Targeting microtubules to inhibit angiogenesis and disrupt tumour vasculature: Implications for cancer treatment. Curr Cancer Drug Targets. 7:566–581. 2007. View Article : Google Scholar : PubMed/NCBI
35	Moeller BJ, Cao Y, Li CY and Dewhirst MW: Radiation activates HIF-1 to regulate vascular radiosensitivity in tumors: Role of reoxygenation, free radicals, and stress granules. Cancer Cell. 5:429–441. 2004. View Article : Google Scholar : PubMed/NCBI
36	Walmsley SR, Print C, Farahi N, Peyssonnaux C, Johnson RS, Cramer T, Sobolewski A, Condliffe AM, Cowburn AS, Johnson N, et al: Hypoxia-induced neutrophil survival is mediated by HIF-1α-dependent NF-κB activity. J Exp Med. 201:105–115. 2005. View Article : Google Scholar : PubMed/NCBI
37	Taylor CT: Interdependent roles for hypoxia inducible factor and nuclear factor-κB in hypoxic inflammation. J Physiol. 586:4055–4059. 2008. View Article : Google Scholar : PubMed/NCBI