Down regulation of estrogen receptors (ER; ER) and atypical chemokine receptors (ACKR 2; ACKR3; ACKR4) to increase 17β-estradiol (E2) levels in MCF-7 in-vitro study
Abstract
Background: Oestrogen plays a vital role in breast development and is strongly related to breast cancer. This research article delves into this paradox. Inflammation is a cancer hallmark that involve chemokine that attract inflammatory immune cells and promote breast cancer spread. E2 as a potential oestrogen can inhibit chemokine secretion, although the underlying mechanism remains unclear. Interestingly, atypical chemokine receptors (ACKRs), as anti-inflammatory G protein-independent transmembrane proteins, act as "scavengers," removing excessive chemokine's, resulting in reduced inflammation, and most strikingly, these genes are essential for normal breast development. This finding suggested that ACKRs may act as tumour suppressors. This study investigated whether a higher E2 level can influence the expression of its own receptors type and ACKRs. Method: In this research, a relative gene expression study been carried out on target genes estrogen receptors (ERa, ERb) and atypical chemokine receptors (ACKR2, ACKR3 & ACKR4) normalised with TOP1 endogenous control gene in MCF-7 breast cancer cells when treated with a higher E2 level including controls for calibration using RT-qPCR technique in designing the experimental assay. Result: Research outcome consistently revealed a down regulation in the m-RNA expression level of target genes at 18 h time point with statistical analysis done using nonparametric Mann‒Whitney's U-test confirming significant differences between the E2-treated and untreated groups (p < 0.05), supporting the hypothesis that E2 does modulates the expression of ACKR2, ACKR3 and ACKR4 via interplay with ERs. Conclusion: These findings highlight the seemingly contradictory roles of E2. While it can fuel tumour growth, it might also have anti-inflammatory effects through cross-talk with expressed ACKR genes. A study with an extended time of E2 exposure on MCF-7 is further proposed to assess its effect at cellular level and an auxiliary analysis at protein level can strengthen the possibility of ERs-ACKRs interplay. Research & development in protein receptors field are valuable for evolving novel cancer therapies.
1.Bray F, Mathieu Laversanne, Sung H, Ferlay J, Siegel RL, Soerjomataram I, et al. Global cancer statistics 2022: GLOBOCAN estimates of incidence and mortality worldwide for 36 cancers in 185 countries. Ca (Print). 2024 Apr 4;
2. Huang Y, Dong W, Li J, Zhang H, Shan Z, Teng W. Differential expression patterns and clinical significance of estrogen receptor-α and β in papillary thyroid carcinoma. BMC Cancer. 2014 May 29;14(1).
3.Klinge CM. Estrogen receptor interaction with estrogen response elements. Nucleic Acids Research. 2001 Jul 15;29(14):2905–19.
4.Rossi D, Zlotnik A. The Biology of Chemokines and their Receptors. Annual Review of Immunology. 2000 Apr;18(1):217–42.
5.Gerard C, Rollins BJ. Chemokines and disease. Nature Immunology. 2001 Feb;2(2):108–15.
6.Ali S, Lazennec G. Chemokines: novel targets for breast cancer metastasis. Cancer and Metastasis Reviews [Internet]. 2007 Aug 24 [cited 2019 Apr 30];26(3-4):401–20. Available from: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2668792/
7.Inadera H, Sekiya T, Yoshimura T, Matsushima K. Molecular Analysis of the Inhibition of Monocyte Chemoattractant Protein-1 Gene Expression by Estrogens and Xenoestrogens in MCF-7 Cells1. Endocrinology. 2000 Jan 1;141(1):50–9.
8.Zhao X, Huang Y, Huang Y, Lei P, Peng J, Wu S, et al. Transforming growth factor-β1 upregulates the expression of CXC chemokine receptor 4 (CXCR4) in human breast cancer MCF-7 cells. Acta Pharmacologica Sinica. 2010 Feb 15;31(3):347–54.
9.Bachelerie F, Ben-Baruch A, Burkhardt AM, Combadiere C, Farber JM, Graham GJ, et al. International Union of Basic and Clinical Pharmacology. [corrected]. LXXXIX. Update on the extended family of chemokine receptors and introducing a new nomenclature for atypical chemokine receptors. Pharmacological reviews [Internet]. 2014;66(1):1–79. Available from: https://www.ncbi.nlm.nih.gov/pubmed/24218476
10.Teoh PJ, Menzies FM, Hansell CAH, Clarke M, Waddell C, Burton GJ, et al. Atypical Chemokine Receptor ACKR2 Mediates Chemokine Scavenging by Primary Human Trophoblasts and Can Regulate Fetal Growth, Placental Structure, and Neonatal Mortality in Mice. The Journal of Immunology. 2014 Oct 8;193(10):5218–28.
11.Wu FY, Ou ZL, Feng LY, Luo JM, Wang LP, Shen ZZ, et al. Chemokine Decoy Receptor D6 Plays a Negative Role in Human Breast Cancer. Molecular Cancer Research. 2008 Aug 1;6(8):1276–88.
12.Wang J, Ou Z-L, Hou Y-F, Luo J-M, Shen Z-Z, Ding J, et al. Enhanced expression of Duffy antigen receptor for chemokines by breast cancer cells attenuates growth and metastasis potential. Oncogene. 2006 Jun 19;25(54):7201–11.
13.Ribas R, Ghazoui Z, Gao Q, Pancholi S, Rani A, Dunbier A, et al. Identification of chemokine receptors as potential modulators of endocrine resistance in oestrogen receptor–positive breast cancers. Breast Cancer Research. 2014 Oct;16(5).
14.Reed MJ, Purohit A. Breast Cancer and the Role of Cytokines in Regulating Estrogen Synthesis: An Emerging Hypothesis. Endocrine Reviews. 1997 Oct;18(5):701–15.
15.Roy D, Cai Q, Felty Q, Narayan S. Estrogen-induced generation of reactive oxygen and nitrogen species, gene damage, and estrogen-dependent cancers. Journal of Toxicology and Environmental Health Part B, Critical Reviews [Internet]. 2007 Jun 1;10(4):235–57. Available from: https://pubmed.ncbi.nlm.nih.gov/17620201/
16.Gest C, Joimel U, Huang L, Pritchard LL, Petit A, Dulong C, et al. Rac3 induces a molecular pathway triggering breast cancer cell aggressiveness: differences in MDA-MB-231 and MCF-7 breast cancer cell lines. BMC cancer [Internet]. 2013 Feb 6;13:63. Available from: https://pubmed.ncbi.nlm.nih.gov/23388133/
17.Green AR, Young P, Krivinskas S, Rakha EA, Claire Paish E, Powe DG, et al. The expression of ERα, ERβ and PR in lobular carcinomain situof the breast determined using laser microdissection and real-time PCR. Histopathology. 2009 Mar;54(4):419–27.
18.Livak KJ, Schmittgen TD. Analysis of Relative Gene Expression Data Using Real-Time Quantitative PCR and the 2−ΔΔCT Method. Methods. 2001 Dec;25(4):402–8.
19.Vidi PA, Bissell MJ, Lelièvre SA. Three-Dimensional Culture of Human Breast Epithelial Cells: The How and the Why. Methods in Molecular Biology. 2012;193–219.
20.Nolan T, Hands RE, Bustin SA. Quantification of mRNA using real-time RT-PCR. Nature protocols [Internet]. 2006;1(3):1559–82. Available from: https://www.ncbi.nlm.nih.gov/pubmed/17406449
21.Bustin SA, Benes V, Garson JA, Hellemans J, Huggett J, Kubista M, et al. The MIQE Guidelines: Minimum Information for Publication of Quantitative Real-Time PCR Experiments. Clinical Chemistry [Internet]. 2009 Feb 26;55(4):611–22. Available from: https://pubmed.ncbi.nlm.nih.gov/19246619/
22.Aithal MGS, Rajeswari N. Validation of Housekeeping Genes for Gene Expression Analysis in Glioblastoma Using Quantitative Real-Time Polymerase Chain Reaction. Brain Tumor Research and Treatment. 2015;3(1):24.
23.Maltseva DV, Khaustova NA, Федотов НН, Matveeva EO, Lebedev AV, M. Yu. Shkurnikov, et al. High-throughput identification of reference genes for research and clinical RT-qPCR analysis of breast cancer samples. Journal of Clinical Bioinformatics. 2013 Jan 1;3(1):13–3.
24.Moore JT, McKee DD, Slentz-Kesler K, Moore LB, Jones SA, Horne EL, et al. Cloning and Characterization of Human Estrogen Receptor β Isoforms. Biochemical and Biophysical Research Communications. 1998 Jun;247(1):75–8.
25.Ogawa S, Inoue S, Watanabe T, Hiroi H, Orimo A, Hosoi T, et al. The complete primary structure of human estrogen receptor beta (hER beta) and its heterodimerization with ER alpha in vivo and in vitro. Biochemical and Biophysical Research Communications [Internet]. 1998 Feb 4;243(1):122–6. Available from: https://pubmed.ncbi.nlm.nih.gov/9473491/
26.Leung YK ., Mak P, Hassan S, Ho SM . Estrogen receptor (ER)-beta isoforms: A key to understanding ER-beta signaling. Proceedings of the National Academy of Sciences. 2006 Aug 22;103(35):13162–7.
27.Poola I, Koduri S, Shubha Chatra, Clarke R. Identification of twenty alternatively spliced estrogen receptor alpha mRNAs in breast cancer cell lines and tumors using splice targeted primer approach. The Journal of Steroid Biochemistry and Molecular Biology. 2000 Apr 1;72(5):249–58.
28. Talashi S. Gene expression analysis of oestrogen receptors and atypical chemokine receptors in response to 17-β oestradiol in MCF-7. Zenodo; 2024.
2. Huang Y, Dong W, Li J, Zhang H, Shan Z, Teng W. Differential expression patterns and clinical significance of estrogen receptor-α and β in papillary thyroid carcinoma. BMC Cancer. 2014 May 29;14(1).
3.Klinge CM. Estrogen receptor interaction with estrogen response elements. Nucleic Acids Research. 2001 Jul 15;29(14):2905–19.
4.Rossi D, Zlotnik A. The Biology of Chemokines and their Receptors. Annual Review of Immunology. 2000 Apr;18(1):217–42.
5.Gerard C, Rollins BJ. Chemokines and disease. Nature Immunology. 2001 Feb;2(2):108–15.
6.Ali S, Lazennec G. Chemokines: novel targets for breast cancer metastasis. Cancer and Metastasis Reviews [Internet]. 2007 Aug 24 [cited 2019 Apr 30];26(3-4):401–20. Available from: https://www.ncbi.nlm.nih.gov/pmc/articles/PMC2668792/
7.Inadera H, Sekiya T, Yoshimura T, Matsushima K. Molecular Analysis of the Inhibition of Monocyte Chemoattractant Protein-1 Gene Expression by Estrogens and Xenoestrogens in MCF-7 Cells1. Endocrinology. 2000 Jan 1;141(1):50–9.
8.Zhao X, Huang Y, Huang Y, Lei P, Peng J, Wu S, et al. Transforming growth factor-β1 upregulates the expression of CXC chemokine receptor 4 (CXCR4) in human breast cancer MCF-7 cells. Acta Pharmacologica Sinica. 2010 Feb 15;31(3):347–54.
9.Bachelerie F, Ben-Baruch A, Burkhardt AM, Combadiere C, Farber JM, Graham GJ, et al. International Union of Basic and Clinical Pharmacology. [corrected]. LXXXIX. Update on the extended family of chemokine receptors and introducing a new nomenclature for atypical chemokine receptors. Pharmacological reviews [Internet]. 2014;66(1):1–79. Available from: https://www.ncbi.nlm.nih.gov/pubmed/24218476
10.Teoh PJ, Menzies FM, Hansell CAH, Clarke M, Waddell C, Burton GJ, et al. Atypical Chemokine Receptor ACKR2 Mediates Chemokine Scavenging by Primary Human Trophoblasts and Can Regulate Fetal Growth, Placental Structure, and Neonatal Mortality in Mice. The Journal of Immunology. 2014 Oct 8;193(10):5218–28.
11.Wu FY, Ou ZL, Feng LY, Luo JM, Wang LP, Shen ZZ, et al. Chemokine Decoy Receptor D6 Plays a Negative Role in Human Breast Cancer. Molecular Cancer Research. 2008 Aug 1;6(8):1276–88.
12.Wang J, Ou Z-L, Hou Y-F, Luo J-M, Shen Z-Z, Ding J, et al. Enhanced expression of Duffy antigen receptor for chemokines by breast cancer cells attenuates growth and metastasis potential. Oncogene. 2006 Jun 19;25(54):7201–11.
13.Ribas R, Ghazoui Z, Gao Q, Pancholi S, Rani A, Dunbier A, et al. Identification of chemokine receptors as potential modulators of endocrine resistance in oestrogen receptor–positive breast cancers. Breast Cancer Research. 2014 Oct;16(5).
14.Reed MJ, Purohit A. Breast Cancer and the Role of Cytokines in Regulating Estrogen Synthesis: An Emerging Hypothesis. Endocrine Reviews. 1997 Oct;18(5):701–15.
15.Roy D, Cai Q, Felty Q, Narayan S. Estrogen-induced generation of reactive oxygen and nitrogen species, gene damage, and estrogen-dependent cancers. Journal of Toxicology and Environmental Health Part B, Critical Reviews [Internet]. 2007 Jun 1;10(4):235–57. Available from: https://pubmed.ncbi.nlm.nih.gov/17620201/
16.Gest C, Joimel U, Huang L, Pritchard LL, Petit A, Dulong C, et al. Rac3 induces a molecular pathway triggering breast cancer cell aggressiveness: differences in MDA-MB-231 and MCF-7 breast cancer cell lines. BMC cancer [Internet]. 2013 Feb 6;13:63. Available from: https://pubmed.ncbi.nlm.nih.gov/23388133/
17.Green AR, Young P, Krivinskas S, Rakha EA, Claire Paish E, Powe DG, et al. The expression of ERα, ERβ and PR in lobular carcinomain situof the breast determined using laser microdissection and real-time PCR. Histopathology. 2009 Mar;54(4):419–27.
18.Livak KJ, Schmittgen TD. Analysis of Relative Gene Expression Data Using Real-Time Quantitative PCR and the 2−ΔΔCT Method. Methods. 2001 Dec;25(4):402–8.
19.Vidi PA, Bissell MJ, Lelièvre SA. Three-Dimensional Culture of Human Breast Epithelial Cells: The How and the Why. Methods in Molecular Biology. 2012;193–219.
20.Nolan T, Hands RE, Bustin SA. Quantification of mRNA using real-time RT-PCR. Nature protocols [Internet]. 2006;1(3):1559–82. Available from: https://www.ncbi.nlm.nih.gov/pubmed/17406449
21.Bustin SA, Benes V, Garson JA, Hellemans J, Huggett J, Kubista M, et al. The MIQE Guidelines: Minimum Information for Publication of Quantitative Real-Time PCR Experiments. Clinical Chemistry [Internet]. 2009 Feb 26;55(4):611–22. Available from: https://pubmed.ncbi.nlm.nih.gov/19246619/
22.Aithal MGS, Rajeswari N. Validation of Housekeeping Genes for Gene Expression Analysis in Glioblastoma Using Quantitative Real-Time Polymerase Chain Reaction. Brain Tumor Research and Treatment. 2015;3(1):24.
23.Maltseva DV, Khaustova NA, Федотов НН, Matveeva EO, Lebedev AV, M. Yu. Shkurnikov, et al. High-throughput identification of reference genes for research and clinical RT-qPCR analysis of breast cancer samples. Journal of Clinical Bioinformatics. 2013 Jan 1;3(1):13–3.
24.Moore JT, McKee DD, Slentz-Kesler K, Moore LB, Jones SA, Horne EL, et al. Cloning and Characterization of Human Estrogen Receptor β Isoforms. Biochemical and Biophysical Research Communications. 1998 Jun;247(1):75–8.
25.Ogawa S, Inoue S, Watanabe T, Hiroi H, Orimo A, Hosoi T, et al. The complete primary structure of human estrogen receptor beta (hER beta) and its heterodimerization with ER alpha in vivo and in vitro. Biochemical and Biophysical Research Communications [Internet]. 1998 Feb 4;243(1):122–6. Available from: https://pubmed.ncbi.nlm.nih.gov/9473491/
26.Leung YK ., Mak P, Hassan S, Ho SM . Estrogen receptor (ER)-beta isoforms: A key to understanding ER-beta signaling. Proceedings of the National Academy of Sciences. 2006 Aug 22;103(35):13162–7.
27.Poola I, Koduri S, Shubha Chatra, Clarke R. Identification of twenty alternatively spliced estrogen receptor alpha mRNAs in breast cancer cell lines and tumors using splice targeted primer approach. The Journal of Steroid Biochemistry and Molecular Biology. 2000 Apr 1;72(5):249–58.
28. Talashi S. Gene expression analysis of oestrogen receptors and atypical chemokine receptors in response to 17-β oestradiol in MCF-7. Zenodo; 2024.
| Files | ||
| Issue | Vol 15 No 3 (2023) | |
| Section | Original Articles | |
| Keywords | ||
| E2 – 17b-estradiol; ACKRs – atypical chemokine receptors; ERa - estrogen receptor alpha; ERb - estrogen receptor beta; RT-qPCR – real-time quantitative polymerase chain reaction. | ||
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How to Cite
1.
Talashi S. Down regulation of estrogen receptors (ER; ER) and atypical chemokine receptors (ACKR 2; ACKR3; ACKR4) to increase 17β-estradiol (E2) levels in MCF-7 in-vitro study. Basic Clin Cancer Res. 2024;15(3):197-215.
