Quercetin-Mediated Modulation of Tumor Suppressor miR-15a, miR-34a, and p53 Signaling in MCF-7 Breast Cancer Cells

Abstract

Breast cancer remains one of the most prevalent types of cancer among women globally, contributing significantly to cancer-related mortality. Despite advancements in treatment, many cases continue to exhibit resistance to chemotherapy, radiotherapy, and hormonal therapies, often resulting in drug resistance, high recurrence rates, and severe side effects. Consequently, the role of the natural food components in cancer prevention and treatment is gaining increasing attention in modern medicine. This study focuses on quercetin, a phytochemical compound, and its effects on the breast cancer cell line MCF-7. Specifically, the study has investigated changes in the expression of miR-15a and miR-34a—microRNAs (miRNAs) known to regulate gene expression at the post-transcriptional level—and the P53 gene, which is critically involved in apoptosis. The analysis was performed using quantitative polymerase chain reaction (qPCR) and Western blot techniques. The results demonstrated that quercetin treatment at concentrations of 40 µM and 80 µM led to a 1.34-fold and 2.73-fold increase in P53 gene expression, respectively. Additionally, the tumor suppressor miRNA miR-15a showed expression changes of 1.48-fold and 1.69-fold at the same quercetin concentrations. Similarly, miR-34a expression levels increased by 1.23-fold and 1.39-fold at 40 µM and 80 µM, respectively. These findings suggest that dietary phytochemicals, such as quercetin, may have therapeutic potential by modulating miRNA expression and targeting the p53 pathway. In conclusion, quercetin emerges as a promising natural therapeutic agent for breast cancer, warranting further in vivo studies and clinical trials to confirm its efficacy and explore its potential as a part of combination therapies.

Keywords

• Flavonoids
• cancer
• non-coding RNAs

MCF-7 Meme Kanseri Hücrelerinde Tümör Baskılayıcı miR-15a, miR-34a ve p53 Sinyalizasyonunun Kuersetin Aracılı Modülasyonu

Abstract

Meme kanseri, küresel olarak kadınlarda en yaygın kanser türlerinden biri olmaya devam etmekte ve kanserle ilişkili ölümlere önemli ölçüde sebep olmaktadır. Tedavideki gelişmelere rağmen, birçok vaka kemoterapi, radyoterapi ve hormonal tedavilere direnç göstermeye devam etmekte ve bu da sıklıkla ilaç direnci, yüksek tekrarlama oranları ve ciddi yan etkilerle sonuçlanmaktadır. Bu nedenle, doğal gıda bileşenlerinin kanser önleme ve tedavisindeki rolü modern tıpta giderek daha fazla ilgi görmektedir. Bu çalışma, bir fitokimyasal bileşik olan kuersetin ve meme kanseri hücre hattı MCF-7 üzerindeki etkilerine odaklanmaktadır. Çalışmada, özellikle, gen ifadesini transkripsiyon sonrası düzeyde düzenlediği bilinen mikroRNA'lar (miRNA) olan miR-15a ve miR-34a'nın ve apoptozda kritik rol oynayan P53 geninin ifadesindeki değişiklikler araştırılmıştır. Analiz, kantitatif polimeraz zincir reaksiyonu (qPCR) ve Western blot teknikleri kullanılarak gerçekleştirilmiştir. Sonuçlar, 40 µM ve 80 µM konsantrasyonlarında kuersetin tedavisinin P53 gen ekspresyonunda sırasıyla 1,34 kat ve 2,73 kat artışa yol açtığını göstermiştir. Ek olarak, tümör baskılayıcı miRNA miR-15a aynı kuersetin konsantrasyonlarında 1,48 kat ve 1,69 kat ekspresyon değişiklikleri olduğu saptanmıştır. Benzer şekilde, miR-34a ekspresyon seviyeleri sırasıyla 40 µM ve 80 µM'de 1,23 kat ve 1,39 kat artmıştır. Bu bulgular, kuersetin gibi diyet fitokimyasallarının miRNA ekspresyonunu düzenleyerek ve p53 yolunu hedefleyerek terapötik potansiyele sahip olabileceğini düşündürmektedir. Sonuç olarak, kuersetin meme kanseri için umut verici bir doğal terapötik ajan olarak ortaya çıkmakta ve etkinliğini doğrulamak ve kombinasyon terapilerinin bir parçası olarak potansiyelini keşfetmek için daha fazla in vivo çalışma ve klinik denemeyi gerektirmektedir.

Keywords

• Flavonoidler
• kanser
• kodlama yapmayan RNAlar
• P53

References

1. Adinew, G. M., Taka, E., Mendonca, P., Messeha, S.S., & Soliman, K.F.A. (2021). The anticancer effects of flavonoids through miRNAs modulations in triple-negative breast cancer. Nutrients, 13(4), 1212. https://doi.org/10.3390/nu13041212
2. Agarwal, S., Hanna, J., Sherman, M.E., Figueroa, J., & Rimm, D.L. (2015). Quantitative assessment of miR34a as an independent prognostic marker in breast cancer. British Journal of Cancer, 112(1), 61–68. https://doi.org/10.1038/bjc.2014.573
3. Aktas, H.G., & Ayan, H. (2021). Oleuropein: A Potential Inhibitor for Prostate Cancer Cell Motility by Blocking Voltage-Gated Sodium Channels. Nutrition and Cancer, 73(9), 1758–1767. https://doi.org/10.1080/01635581.2020.1807575
4. Amit, M., Takahashi, H., Dragomir, M.P., Lindemann, A., Gleber-Netto, F.O., Pickering, C.R., ..., & Myers, J.N. (2020). Loss of p53 drives neuron reprogramming in head and neck cancer. Nature, 578(7795), 449–454. https://doi.org/10.1038/s41586-020-1996-3
5. Bandi, N., Zbinden, S., Gugger, M., Arnold, M., Kocher, V., Hasan, L., ..., & Vassella, E. (2009). miR-15a and miR-16 are implicated in cell cycle regulation in a Rb-dependent manner and are frequently deleted or down-regulated in non–small cell lung cancer. Cancer Research, 69(13), 5553-5559. https://doi.org/10.1158/0008-5472.CAN-08-4277
6. Calin, G. A., Cimmino, A., Fabbri, M., Ferracin, M., Wojcik, S.E., Shimizu, M., ..., & Croce, C. M. (2008). MiR-15a and miR-16-1 cluster functions in human leukemia. Proceedings of the National Academy of Sciences, 105(13), 5166-5171. https://doi.org/10.1073/pnas.0800121105
7. Chang, T.C., Wentzel, E.A., Kent, O.A., Ramachandran, K., Mullendore, M., Lee, K.H., ... & Mendell, J.T. (2007). Transactivation of miR-34a by p53 broadly influences gene expression and promotes apoptosis. Molecular Cell, 26(5), 745-752.. https://doi.org/10.1016/j.molcel.2007.05.010
8. Chen, X., Li., H., Zhang, B., Deng, Z., & Zhang, X. (2022). The role of quercetin in cancer prevention and therapy: A comprehensive review. Phytotherapy Research, 36(1), 1–15. https://doi.org/10.1002/ptr.7286

9. Erbes, T., Hirschfeld, M., Rücker, G., Jaeger, M., Boas, J., Iborra, S., ..., & Stickeler, E. (2015). Feasibility of urinary microRNA detection in breast cancer patients and its potential as an innovative non-invasive biomarker. BMC Cancer, 15, 193.
10. Ezzati, M., Yousefi, B., Velaei, K., & Safa, A. (2020). A review on anti-cancer properties of quercetin in breast cancer. Life Sciences, 248, 117463. https://doi.org/10.1016/j.lfs.2020.117463
11. Ferdin, J., Kunej, T., & Calin, G.A. (2010). Non-coding RNAs: Identification of cancer-associated microRNAs by gene profiling. Technology in Cancer Research & Treatment, 9, 123–138. https://doi.org/10.1177/153303461000900202
12. Gilbert, E.R., & Liu, D. (2010). Flavonoids influence epigenetic-modifying enzyme activity: Structure-function relationships and the therapeutic potential for cancer. Current Medicinal Chemistry, 17(17), 1756–1768.
13. Gungormez, C., & Acar, D. (2019). Identıfıcatıon of miR-145-1, miR-21-2 And miR-92-1 Expressıon and Target Genes in Colon Cancer Stage IIIA Patients. Turkish Journal of Biochemistry. Supplement, 44, 26-28.
14. Gungormez, C. (2024). Identification of hub genes and key pathways targeted by miRNAs in pancreatic ductal adenocarcinoma: MAPK3/8/9 and TGFBR1/2. Human Gene, 39, 201267.
15. Hämäläinen, M., Nieminen, R., Vuorela, P., Heinonen, M., & Moilanen, E. (2007). Anti-inflammatory effects of flavonoids: Genistein, kaempferol, quercetin, and daidzein inhibit STAT-1 and NF-kappaB activations, whereas flavone, isorhamnetin, naringenin, and pelargonidin inhibit only NF-kappaB activation along with their inhibitory effect on iNOS expression and NO production in activated macrophages. Mediators of Inflammation, 2007, 45673. https://doi.org/10.1155/2007/45673
16. Hashemzaei, M., Delarami Far, A., Yari, A., Heravi, R.E., Tabrizian, K., Taghdisi, S.M., ..., & Rezaee, R. (2017). Anticancer and apoptosis-inducing effects of quercetin in vitro and in vivo. Oncology Reports, 38(2), 819–828. https://doi.org/10.3892/or.2017.5766
17. Hermeking, H. (2010). The miR-34 family in cancer and apoptosis. Cell Death & Differentiation, 17(2), 193–199.
18. Imani, S., Wei, C., Cheng, J., Khan, M.A., Fu, S., Yang, L., ..., & Fu, J. (2016). MicroRNA-34a targets epithelial to mesenchymal transition-inducing transcription factors (EMT-TFs) and inhibits breast cancer cell migration and invasion. Oncotarget, 7(8), 1–12. https://doi.org/10.18632/oncotarget.15214
19. Imani, S., Wu, R.C., & Fu, J. (2018). MicroRNA-34 family in breast cancer: From research to therapeutic potential. Journal of Cancer, 9(20), 3765–3775. https://doi.org/10.7150/jca.24187
20. Iqbal, J., Abbasi, B.A., Khalil, A.T., Ali, B., Mahmood, T., Kanwal, S., ..., & Ali, W. (2018). Dietary isoflavones, the modulator of breast carcinogenesis: Current landscape and future perspectives. Asian Pacific Journal of Tropical Medicine, 11(3), 186-193.
21. Kang, L., Mao, J., Tao, Y., Song, B., Ma, W., Lu, Y., ..., & Li, L. (2015). Retracted: Micro RNA‐34a suppresses the breast cancer stem cell‐like characteristics by downregulating Notch1 pathway. Cancer Science, 106(6), 700-708. https://doi.org/10.1111/cas.12656
22. Kawaguchi, T., Yan, L., Qi, Q., Peng, X., Gabriel, E.M., Young, J., ... & Takabe, K. (2017). Overexpression of suppressive microRNAs, miR-30a and miR-200c are associated with improved survival of breast cancer patients. Scientific Reports, 7(1), 15945. https://doi.org/10.1038/s41598-017-16112-y
23. Kim, G.T., Lee, S.H., Kim, J.I., & Kim, Y.M. (2014). Quercetin regulates the sestrin 2-AMPK-p38 MAPK signaling pathway and induces apoptosis by increasing the generation of intracellular ROS in a p53-independent manner. International Journal of Molecular Medicine, 33(4), 863-869.
24. Kopustinskiene, D.M., Jakstas, V., Savickas, A., & Bernatoniene, J. (2020). Flavonoids as anticancer agents. Nutrients, 12(2), 457.
25. Li, X.J., Ren, Z.J., Tang, J.H., & Yu, Q. (2019). MiR-34a: A potential therapeutic target in human cancer. Cell Death & Disease, 10(3), 193. https://doi.org/10.1038/s41419-019-1446-z
26. Li, Y., Yao, J., Han, C., Yang, J., Chaudhry, M.T., Wang, S., ..., & Yin, Y. (2020). Quercetin, inflammation and immunity. Nutrients, 12(2), 457. https://doi.org/10.3390/nu12020457
27. Mansoori, B., Mohammadi, A., Davudian, S., Shirjang, S., & Baradaran, B. (2020). The different mechanisms of cancer drug resistance: A brief review. Advanced Pharmaceutical Bulletin, 10(1), 1–8.
28. Misso, G., Di Martino, M.T., De Rosa, G., Farooqi, A.A., Lombardi, A., Campani, V., ..., & Caraglia, M. (2014). miR-34: A new weapon against cancer? Molecular Therapy - Nucleic Acids, 3, e194. https://doi.org/10.1038/mtna.2014.47
29. Muller, P.A., Vousden, K.H., & Norman, J.C. (2011). p53 and its mutants in tumor cell migration and invasion. The Journal of cell biology, 192(2), 209–218. https://doi.org/10.1083/jcb.201009059
30. Murakami, A., Ashida, H., & Terao, J. (2008). Multitargeted cancer prevention by quercetin. Cancer Letters, 269(2), 315–325. https://doi.org/10.1016/j.canlet.2008.03.046
31. O'Brien, J., Hayder, H., Zayed, Y., & Peng, C. (2018). Overview of MicroRNA Biogenesis, Mechanisms of Actions, and Circulation. Frontiers in Endocrinology, 9, 402.
32. Pang, R.T., Leung, C.O., Ye, T.M., Liu, W., Chiu, P.C., Lam, K.K., ..., & Yeung, W.S. (2010). MicroRNA-34a suppresses invasion through downregulation of Notch1 and Jagged1 in cervical carcinoma and choriocarcinoma cells. Carcinogenesis, 31(6), 1037–1044.
33. Rauf, A., Imran, M., Khan, I.A., Ur-Rehman, M., Gilani, S.A., Mehmood, Z., & Mubarak, M.S. (2018). Anticancer potential of quercetin: A comprehensive review. Phytotherapy Research, 32(11), 2109–2130. https://doi.org/10.1002/ptr.6155
34. Raver-Shapira, N., Marciano, E., Meiri, E., Spector, Y., Rosenfeld, N., Moskovits, N., ..., & Oren, M. (2007). Transcriptional activation of miR-34a contributes to p53-mediated apoptosis. Molecular Cell, 26(5), 731–743. https://doi.org/10.1016/j.molcel.2007.05.017
35. Rhman, M.A., Devnarain, N., Khan, R., & Owira, P.M.O. (2022). Synergism potentiates oxidative antiproliferative effects of naringenin and quercetin in MCF-7 breast cancer cells. Nutrients, 14(16), 3437.
36. Rodríguez-García, C., Sánchez-Quesada, C., & Gaforio, J. (2019). Dietary Flavonoids as Cancer Chemopreventive Agents: An Updated Review of Human Studies. Antioxidants (Basel, Switzerland), 8(5), 137. https://doi.org/10.3390/antiox8050137
37. Siegel, R.L., Miller, K.D., & Jemal, A. (2020). Cancer statistics, 2020. CA: A Cancer Journal for Clinicians, 70(1), 7–30. https://doi.org/10.3322/caac.21590
38. Tang, J., Ahmad, A., & Sarkar, F.H. (2012). The role of microRNAs in breast cancer migration, invasion and metastasis. International Journal of Molecular Sciences, 13(10), 13414–13437. https://doi.org/10.3390/ijms131013414
39. Tang, S.M., Deng, X.T., Zhou, J., Li, Q.P., Ge, X.X., & Miao, L. (2021). Pharmacological basis and new insights of quercetin action in respect to its anti-cancer effects. Biomedicine & Pharmacotherapy, 133, 111017. https://doi.org/10.1016/j.biopha.2020.111017
40. Tao, S.F., He, H.F., & Chen, Q. (2015). Quercetin inhibits proliferation and invasion acts by up-regulating miR-146a in human breast cancer cells. Molecular and Cellular Biochemistry, 402(1–2), 93–100.
41. Vousden, K.H., & Lane, D.P. (2007). p53 in health and disease. Nature Reviews Molecular Cell Biology, 8(4), 275–283. https://doi.org/10.1038/nrm2147
42. Wang, W., Sun, C., Mao, L., Ma, P., Liu, F., Yang, J., & Gao, Y. (2021). The biological activities, chemical stability, metabolism and delivery systems of quercetin: A review. Trends in Food Science & Technology, 109, 1–15. https://doi.org/10.1016/j.tifs.2021.01.042
43. Zhang, L., Liao, Y., & Tang, L. (2019). MicroRNA-34 family: A potential tumor suppressor and therapeutic candidate in cancer. Journal of Experimental & Clinical Cancer Research, 38(1), 53. https://doi.org/10.1186/s13046-019-1059-5
44. Zhang, J.T., Chen, J., Ruan, H.C., Li, F. X., Pang, S., Xu, Y. J., ..., & Wu, X.H. (2021). Microribonucleic Acid-15a-5p alters Adriamycin resistance in breast cancer cells by targeting cell division cycle-associated protein 4. Cancer Management and Research, 13, 8425–8434.