The activation of the G-protein-coupled estrogen receptor promotes the aggressiveness of MDA-MB231 cells by targeting the IRE1α/TXNIP pathway

Maryam Mohammad-Sadeghipour , Mohammad Hadi Nematollahi, Hassan Ahmadinia, Mohammad Reza Hajizadeh , Mehdi Mahmoodi

Abstract


Background and purpose: This study investigated modulating the G protein-coupled estrogen receptor (GPER) on the IRE1α/TXNIP pathway and its role in drug resistance in MDA-MB231 cells.

Experimental approach: To determine the optimal concentrations of G1 and 4-hydroxytamoxifen (TAM), GPER expression and ERK1/2 phosphorylation were analyzed using qRT-PCR and western blotting, respectively. Cells were treated with individual concentrations of G1 (1000 nM), G15 (1000 nM), and TAM (2000 nM), as well as combinations of these treatments (G1 + G15, TAM + G15, and G1 + TAM) for 24 and 48 h. The expression levels of GPER, IRE1α, miR-17-5p, TXNIP, ABCB1, and ABCC1 genes and TXNIP protein expression were evaluated. Finally, apoptosis and cell migration were examined using flow cytometry and the wound-healing assay, respectively.

Findings/Results: Activating GPER with its specific agonist G1 and TAM significantly increased IRE1α levels in MDA-MB231 cells. IRE1α through splicing XBP1 led to unfolded protein response. In addition, decreased TXNIP gene and protein expression reduced apoptosis, increased migration, and upregulated the genes associated with drug resistance.

Conclusion and implication: Our investigation revealed that blocking the GPER/IRE1α/TXNIP pathway in MDA-MB231 cells could enhance treatment efficacy and improve chemotherapy responsiveness. The distinct unfolded protein response observed in MDA-MB231 cells may stem from the unique characteristics of these cells, which lack receptors for estrogen, progesterone, and HER2/neu hormones, possessing only the GPER receptor (ER-/PR-/HER2-/GPER+). This study introduced a new pathway in TNBC cells, indicating that targeting GPER could be crucial in comprehensive therapeutic strategies in TNBC cells.


Keywords


Breast cancer; Drug resistance; G protein-coupled estrogen receptor; miR-17-5P; Thioredoxin interacting protein; Unfolded protein response.

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References


Settacomkul R, Sangpairoj K, Phuagkhaopong S, Meemon K, Niamnont N, Sobhon P, et al. Ethanolic extract of Halymenia durvillei induced G2/M arrest and altered the levels of cell cycle regulatory proteins of MDA-MB-231 triple-negative breast cancer cells. Res Pharm Sci. 2023;18(3):279-291.DOI: 10.4103/1735-5362.371584.

Kao J, Salari K, Bocanegra M, Choi YL, Girard L, Gandhi J, et al. Molecular profiling of breast cancer cell lines defines relevant tumor models and provides a resource for cancer gene discovery. PLoS One. 2009;4(7):e6146,1-16.

DOI: 10.1371/journal.pone.0006146.

Vyshnavi H, Namboori K. Identifying potential ligand molecules EGFR mediated TNBC targeting the kinase domain-identification of customized drugs through in silico methods. Res Pharm Sci. 2023;18(2):121-137.

DOI: 10.4103/1735-5362.367792.

Derakhshan F, Reis-Filho JS. Pathogenesis of triple-negative breast cancer. Annu Rev Pathol. 2022;17:181-204.

DOI: 10.1146/annurev-pathol-042420-093238.

Li Y, Zhang H, Merkher Y, Chen L, Liu N, Leonov S, et al. Recent advances in therapeutic strategies for triple-negative breast cancer. J Hematol Oncol. 2022;15(1):121,1-30.

DOI: 10.1186/s13045-022-01341-0.

Girgert R, Emons G, Gründker C. Estrogen signaling in ERα-negative breast cancer: ERβ and GPER. Front endocrinol. 2019;9:781,1-12.

DOI: 10.3389/fendo.2018.00781.

Han N, Heublein S, Jeschke U, Kuhn C, Hester A, Czogalla B, et al. The G-protein-coupled estrogen receptor (GPER) regulates trimethylation of histone H3 at lysine 4 and represses migration and proliferation of ovarian cancer cells in vitro. Cells. 2021;10(3):619,1-23.

DOI: 10.3390/cells10030619.

Ignatov T, Claus M, Nass N, Haybaeck J, Seifert B, Kalinski T, et al. G-protein-coupled estrogen receptor GPER-1 expression in hormone receptor-positive breast cancer is associated with poor benefit of tamoxifen. Breast Cancer Res Treat. 2019;174(1):121-127.

DOI: 10.1007/s10549-018-5064-8.

Lei B, Peng W, Xu G, Wu M, Wen Y, Xu J, et al. Activation of G protein-coupled receptor 30 by thiodiphenol promotes proliferation of estrogen receptor alpha-positive breast cancer cells. Chemosphere. 2017;169:204-211.

DOI: 10.1016/j.chemosphere.2016.11.066.

Vivacqua A, Sebastiani A, Miglietta AM, Rigiracciolo DC, Cirillo F, Galli GR, et al. MiR-338-3p is regulated by estrogens through GPER in breast cancer cells and cancer-associated fibroblasts (CAFs). Cells. 2018;7(11):203,1-19.

DOI: 10.3390/cells7110203.

Liang S, Chen Z, Jiang G, Zhou Y, Liu Q, Su Q, et al. Activation of GPER suppresses migration and angiogenesis of triple negative breast cancer via inhibition of NF-κB/IL-6 signals. Cancer lett. 2017;386:12-23.

DOI: 10.1016/j.canlet.2016.11.003.

Vo DKH, Hartig R, Weinert S, Haybaeck J, Nass N. G-protein-coupled estrogen receptor (GPER)-specific agonist G1 induces ER stress leading to cell death in MCF-7 cells. Biomolecules. 2019;9(9):503,1-21.

DOI: 10.3390/biom9090503.

Weissenborn C, Ignatov T, Poehlmann A, Wege AK, Costa SD, Zenclussen AC, et al. GPER functions as a tumor suppressor in MCF-7 and SK-BR-3 breast cancer cells. J Cancer Res Clin Oncol. 2014;140(4):663-671.DOI: 10.1007/s00432-014-1598-2.

Sisinni L, Pietrafesa M, Lepore S, Maddalena F, Condelli V, Esposito F, et al. Endoplasmic reticulum stress and unfolded protein response in breast cancer: the balance between apoptosis and autophagy and its role in drug resistance. Int J Mol Sci. 2019;20(4):857,1-17.DOI: 10.3390/ijms20040857.

Adams CJ, Kopp MC, Larburu N, Nowak PR, Ali MMU. Structure and molecular mechanism of ER stress signaling by the unfolded protein response signal activator IRE1. Front Mol Biosci. 2019;6:11,1-12.DOI: 10.3389/fmolb.2019.00011.

Barua D, Gupta A, Gupta S. Targeting the IRE1-XBP1 axis to overcome endocrine resistance in breast cancer: opportunities and challenges. Cancer Lett. 2020;486:29-37.DOI: 10.1016/j.canlet.2020.05.020.

Oslowski CM, Urano F. Measuring ER stress and the unfolded protein response using mammalian tissue culture system. Methods Enzymol. 2011;490:71-92.DOI: 10.1016/B978-0-12-385114-7.00004-0.

Wu J, He GT, Zhang WJ, Xu J, Huang QB. IRE1α signaling pathways involved in mammalian cell fate determination. Cell Physiol Biochem. 2016;38(3):847-858.DOI: 10.1159/000443039.

Park JW, Lee SH, Woo GH, Kwon HJ, Kim DY. Downregulation of TXNIP leads to high proliferative activity and estrogen-dependent cell growth in breast cancer. Biochem Biophys Res Commun. 2018;498(3):566-572.DOI: 10.1016/j.bbrc.2018.03.020.

Lee JH, Jeong J, Jeong EM, Cho SY, Kang JW, Lim J, et al. Endoplasmic reticulum stress activates transglutaminase 2 leading to protein aggregation. Int J Mol Med. 2014;33(4):849-855.DOI: 10.3892/ijmm.2014.1640.21. Wei W, Chen ZJ, Zhang KS, Yang XL, Wu YM, Chen XH, et al. The activation of G protein-coupled receptor 30 (GPR30) inhibits proliferation of estrogen receptor-negative breast cancer cells in vitro and in vivo. Cell Death Dis. 2014;5(10):e1428,1-12.DOI: 10.1038/cddis.2014.398.

Yu T, Liu M, Luo H, Wu C, Tang X, Tang S, et al. GPER mediates enhanced cell viability and motility via non-genomic signaling induced by 17β-estradiol in triple-negative breast cancer cells. J Steroid Biochem Mol Biol. 2014;143:392-403.DOI: 10.1016/j.jsbmb.2014.05.003.

Wang LJ, Han SX, Bai E, Zhou X, Li M, Jing GH, et al. Dose-dependent effect of tamoxifen in tamoxifen-resistant breast cancer cells via stimulation by the ERK1/2 and AKT signaling pathways. Oncol Rep. 2013;29(4):1563-1569.DOI: 10.3892/or.2013.2245.

Shahali A, Ghanadian M, Jafari SM, Aghaei M. Mitochondrial and caspase pathways are involved in the induction of apoptosis by nardosinen in MCF-7 breast cancer cell line. Res Pharm Sci. 2018;13(1):12-21.DOI: 10.4103/1735-5362.220963.

Liu L, Liu S, Luo H, Chen C, Zhang X, He L, et al. GPR30-mediated HMGB1 upregulation in CAFs induces autophagy and tamoxifen resistance in ERα-positive breast cancer cells. Aging (Albany NY). 2021;13(12):16178-16197.DOI: 10.18632/aging.203145.

Samii B, Jafarian A, Rabbani M, Zolfaghari B, Rahgozar S, Pouraboutaleb E. The effects of Astragalus polysaccharides, tragacanthin, and bassorin on methotrexate-resistant acute lymphoblastic leukemia. Res Pharm Sci. 2023;18(4):381-391.DOI: 10.4103/1735-5362.378085.

Liu C, Liao Y, Fan S, Fu X, Xiong J, Zhou S, et al. G-protein-coupled estrogen receptor antagonist G15 decreases estrogen-induced development of non-small cell lung cancer. Oncol Res. 2019;27(3):283-292.DOI: 10.3727/096504017x15035795904677.

Masoudi F, Sharifi MR, Pourfarzam M. Investigation of the relationship between miR-33a, miR-122, erythrocyte membrane fatty acids profile, and serum lipids with components of metabolic syndrome in type 2 diabetic patients. Res Pharm Sci. 2022;17(3):242-251.DOI: 10.4103/1735-5362.343078.

Coucha M, Mohamed IN, Bartasis ML, El-Remessy AB. High fat diet dysregulates microRNA-17-5p and enhances retinal TXNIP expression: role of ER-stress. Investig Ophthalmol Vis Sci. 2015;56(7):2391.

Luo H, Yang G, Yu T, Luo S, Wu C, Sun Y, et al. GPER-mediated proliferation and estradiol production in breast cancer-associated fibroblasts. Endocr Relat Cancer. 2014;21(2):355-369.DOI: 10.1530/erc-13-0237.

Shen Y, Yang F, Zhang W, Song W, Liu Y, Guan X. The androgen receptor promotes cellular proliferation by suppression of G-protein coupled estrogen receptor signaling in triple-negative breast cancer. Cell Physiol Biochem. 2017;43(5):2047-2061.DOI: 10.1159/000484187.

Choi EH, Park SJ. TXNIP: a key protein in the cellular stress response pathway and a potential therapeutic target. Exp Mol Med. 2023;55(7): 1348-1356.DOI: 10.1038/s12276-023-01019-8.

Dong H, Adams NM, Xu Y, Cao J, Allan DSJ, Carlyle JR, et al. The IRE1 endoplasmic reticulum stress sensor activates natural killer cell immunity in part by regulating c-Myc. Nat Immunol. 2019;20(7):865-878.DOI: 10.1038/s41590-019-0388-z.

Shen L, O'Shea JM, Kaadige MR, Cunha S, Wilde BR, Cohen AL, et al. Metabolic reprogramming in triple-negative breast cancer through Myc suppression of TXNIP. Proc Natl Acad Sci U S A. 2015;112(17):5425-5430.DOI: 10.1073/pnas.1501555112.

Xu T, Ma D, Chen S, Tang R, Yang J, Meng C, et al. High GPER expression in triple-negative breast cancer is linked to pro-metastatic pathways and predicts poor patient outcomes. NPJ Breast Cancer. 2022;8(1):100,1-11.DOI: 10.1038/s41523-022-00472-4.

Li J, Yue Z, Xiong W, Sun P, You K, Wang J. TXNIP overexpression suppresses proliferation and induces apoptosis in SMMC7221 cells through ROS generation and MAPK pathway activation. Oncol Rep. 2017;37(6):3369-3376.DOI: 10.3892/or.2017.5577.

Rajaei N, Rahgouy G, Panahi N, Razzaghi-Asl N. Bioinformatic analysis of highly consumed phytochemicals as P-gp binders to overcome drug-resistance. Res Pharm Sci. 2023;18(5):505-516.DOI: 10.4103/1735-5362.383706.

Chen Y, Feng X, Yuan Y, Jiang J, Zhang P, Zhang B. Identification of a novel mechanism for reversal of doxorubicin-induced chemotherapy resistance by TXNIP in triple-negative breast cancer via promoting reactive oxygen-mediated DNA damage. Cell Death Dis. 2022;13(4):338,1-13.DOI: 10.1038/s41419-022-04783-z.

Yu T, Cheng H, Ding Z, Wang Z, Zhou L, Zhao P, et al. GPER mediates decreased chemosensitivity via regulation of ABCG2 expression and localization in tamoxifen-resistant breast cancer cells. Mol Cell Endocrinol. 2020;506:110762,1-12.DOI: 10.1016/j.mce.2020.110762.

Logue SE, McGrath EP, Cleary P, Greene S, Mnich K, Almanza A, et al. Inhibition of IRE1 RNase activity modulates the tumor cell secretome and enhances response to chemotherapy. Nat Commun. 2018;9(1):3267,1-15.DOI: 10.1038/s41467-018-05763-8.

Moh MC, Lee LH, Shen S. Cloning and characterization of hepaCAM, a novel Ig-like cell adhesion molecule suppressed in human hepatocellular carcinoma. J Hepatol. 2005;42(6): 833-841.DOI: 10.1016/j.jhep.2005.01.025.

Moh MC, Zhang T, Lee LH, Shen S. Expression of hepaCAM is downregulated in cancers and induces senescence-like growth arrest via a p53/p21-dependent pathway in human breast cancer cells. Carcinogenesis. 2008;29(12):2298-2305.DOI: 10.1093/carcin/bgn226.

Moh MC, Zhang C, Luo C, Lee LH, Shen S. Structural and functional analyses of a novel Ig-like cell adhesion molecule, hepaCAM, in the human breast carcinoma MCF7 cells. J Biol Chem. 2005;280(29):27366-27374.DOI: 10.1074/jbc.M500852200.

Nittka S, Böhm C, Zentgraf H, Neumaier M. The CEACAM1-mediated apoptosis pathway is activated by CEA and triggers dual cleavage of CEACAM1. Oncogene. 2008;27(26):3721-3728.DOI: 10.1038/sj.onc.1211033.

Ebrahimnejad A, Streichert T, Nollau P, Horst AK, Wagener C, Bamberger AM, et al. CEACAM1 enhances invasion and migration of melanocytic and melanoma cells. Am J Pathol. 2004;165(5): 1781-1787.DOI: 10.1016/s0002-9440(10)63433-5.

Liu W, Wei W, Winer D, Bamberger AM, Bamberger C, Wagener C, et al. CEACAM1 impedes thyroid cancer growth but promotes invasiveness: a putative mechanism for early metastases. Oncogene. 2007;26(19):2747-2758.DOI: 10.1038/sj.onc.1210077.

Moh MC, Shen S. The roles of cell adhesion molecules in tumor suppression and cell migration. Cell Adh Migr. 2009;3(4):334-336.DOI: 10.4161/cam.3.4.9246.


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