Tamoxifen and regulation of stress-induced senescence in breast cancer cells

Introduction. Tumor cells are known not to undergo replicative aging – usually due to hyperactivation of telomerase, which restores telomere length during each cell division cycle. However, it is possible to induce aging in tumor cells through sublethal doses of cytostatics or irradiation – this is the so-called stress-induced or non-replicative senescence. Studying the mechanisms and regulatory pathways of this process is one of the important areas of modern oncology.
Aim. To investigate the mechanisms of doxorubicin-induced senescence in different breast cancer cell subtypes and to explore possible approaches to regulating non-replicative aging.
Materials and methods. The experiments were performed on in vitro cultured breast cancer cell lines MCF-7 and MDA-MB-231. Cellular senescence was assessed by β-galactosidase activity, morphological changes, and activation of the p53/p21 signaling pathway. Colorimetric assays, reporter analysis, and immunoblotting were used to evaluate the expression and activity of cellular proteins. DNA methyltransferase 3A (DNMT3A) knockdown was achieved using a standard lentiviral vector encoding antisense RNA against DNMT3A.
Results. A potentiating effect of tamoxifen on doxorubicin-induced senescence – including in estrogen-independent breast cancer cells – was demonstrated. Enhanced non-replicative senescence was observed in resistant cells characterized by constitutive suppression of DNMT3A expression. For the first time, it was shown that DNMT3A suppression – either via decitabine treatment or DNMT3A knockdown – leads to an increase and maintenance of non-replicative senescence in MCF-7 cells.
Conclusion. The findings indicate that non-replicative senescence in breast cancer cells can be enhanced and sustained in the presence of the antiestrogen tamoxifen, and underscore the key role of DNMT3A in regulating doxorubicin-induced senescence.

1. Di Micco R., Krizhanovsky V., Baker D. et al. Cellular senescence in ageing: from mechanisms to therapeutic opportunities. Nat Rev Mol Cell Biol 2021;22(2):75–95. DOI: 10.1038/s41580-020-00314-w

2. Sedrak M.S., Cohen H.J. The Aging-cancer cycle: mechanisms and opportunities for intervention. Gerontol A Biol Sci Med Sci 2023;78(7):1234–8. DOI: 10.1093/gerona/glac247

3. Razgonova M.P., Zakharenko A.M., Golokhvast K.S. et al. Telomerase and telomeres in aging theory and chronographic aging theory (review). Mol Med Rep 2020;22(3):1679–94. DOI: 10.3892/mmr.2020.11274

4. Hornsby P.J. Telomerase and the aging process. Exp Gerontol 2007;42(7):575–81. DOI: 10.1016/j.exger.2007.03.007

5. Rossiello F., Jurk D., Passos J.F. et al. Telomere dysfunction in ageing and age-related diseases. Nat Cell Biol 2022;24(2):135–47. DOI: 10.1038/s41556-022-00842-x

6. Montégut L., López-Otín C., Kroemer G. Aging and cancer. Mol Cancer 2024;23(1):106. DOI: 10.1186/s12943-024-02020-z

7. Colucci M., Sarill M., Maddalena M. et al. Senescence in cancer. Cancer Cell 2025;43(7):1204–26. DOI: 10.1016/j.ccell.2025.05.015

8. Schmitt C.A., Wang B., Demaria M. Senescence and cancer – role and therapeutic opportunities. Nat Rev Clin Oncol 2022;19(10):619–36. DOI 10.1038/s41571-022-00668-4

9. Wang L., Lankhorst L., Bernards R. Exploiting senescence for the treatment of cancer. Nat Rev Cancer 2022;22(6):340–55. DOI: 10.1038/s41568-022-00450-9

10. Guillon J., Petit C., Toutain B. et al. Chemotherapy-induced senescence, an adaptive mechanism driving resistance and tumor heterogeneity. Cell Cycle 2019;18(19):2385–97. DOI: 10.1080/15384101.2019.1652047

11. Bisht A., Avinash D., Sahu K.K. et al. A comprehensive review on doxorubicin: mechanisms, toxicity, clinical trials, combination therapies and nanoformulations in breast cancer. Drug Deliv Transl Res 2025;15(1):102–33. DOI: 10.1007/s13346-024-01648-0

12. Van der Zanden S.Y., Qiao X., Neefjes J. New insights into the activities and toxicities of the old anticancer drug doxorubicin. The FEBS J 2021;288(21):6095–111. DOI: 10.1111/febs.15583

13. Arcamone F., Cassinelli G., Fantini G. et al. Adriamycin, 14-hydroxydaunomycin, a new antitumor antibiotic from S. peucetius var. caesius. Biotechnol Bioeng 1969;11(6):1101–10. DOI 10.1002/bit.260110607

14. Cortés-Funes H., Coronado C. Role of anthracyclines in the era of targeted therapy. Cardiovasc Toxicol 2007;7(2):56–60. DOI: 10.1007/s12012-007-0015-3

15. Huang Z., Lin S., Long C. et al. Notch signaling pathway mediates Doxorubicin-driven apoptosis in cancers. Cancer Manag Res 2018;10:1439–48. DOI: 10.2147/cmar.s160315

16. Synowiec E., Hoser G., Bialkowska-Warzecha J. et al. Doxorubicin differentially induces apoptosis, expression of mitochondrial apoptosis-related genes, and mitochondrial potential in BCR-ABL1-expressing cells sensitive and resistant to imatinib. BioMed Res Int 2015;2015:673512. DOI: 10.1155/2015/673512

17. Karabicici M., Alptekin S., Fırtına Karagonlar Z. et al. Doxorubicin-induced senescence promotes stemness and tumorigenicity in EpCAM-/CD133-nonstem cell population in hepatocellular carcinoma cell line, HuH-7. Mol Oncol 2021;15(8):2185–202. DOI: 10.1002/1878-0261.12916

18. Mosmann T. Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. J Immunol Methods 1983;65(1–2):55–63. DOI: 10.1016/0022-1759(83)90303-4

19. Scherbakov A.M., Komkov A.V., Komendantova A.S. et al. Steroidal pyrimidines and dihydrotriazines as novel classes of anticancer agents against hormone-dependent breast cancer cells. Front Pharmacol 2017;8:979. DOI: 10.3389/fphar.2017.00979

20. Reid G., Hübner M.R., Métivier R. et al. Cyclic, proteasomemediated turnover of unliganded and liganded ERalpha on responsive promoters is an integral feature of estrogen signaling. Mol Cell 2003;11(3):695–707. DOI: 10.1016/s1097-2765(03)00090-x

21. Shchegolev Y., Sorokin D., Scherbakov A. et al. Upregulation of Akt/Raptor signaling is associated with rapamycin resistance of breast cancer cells. Chem Biol Inter 2020;330:109243. DOI: 10.1016/j.cbi.2020.109243

22. Mruk D.D., Cheng C.Y. Enhanced chemiluminescence (ECL) for routine immunoblotting: An inexpensive alternative to commercially available kits. Spermatogenesis 2011;1(2):121–2. DOI: 10.4161/spmg.1.2.16606

23. Bae Y.-H., Shin J.-M., Park H.-J. et al. Gain-of-function mutant p53-R280K mediates survival of breast cancer cells. Genes Genomics 2014;36(2):171–8. DOI: 10.1007/s13258-013-0154-9

24. Andreeva O.E., Sorokin D.V., Vinokurova S.V. et al. The effect of DNA methyltransferase 3A suppression in progression of the resistance phenotype in breast cancer cells. Uspekhi molekulyarnoy onkologii = Advances in Molecular Oncology 2023;10(4):149–56. DOI: 10.17650/2313-805X-2023-10-4-149-156

25. Hanahan D., Weinberg R.A. Hallmarks of cancer: the next generation. Cell 2011;144(5):646–74. DOI: 10.1016/j.cell.2011.02.013

26. Faget D.V., Ren Q., Stewart S.A. Unmasking senescence: context-dependent effects of SASP in cancer. Nat Rev Cancer 2019;19(8):439–53. DOI: 10.1038/s41568-019-0156-2

27. Niklander S.E., Lambert D.W., Hunter K.D. Senescent cells in cancer: wanted or unwanted citizens. Cells 2021;10(12):3315. DOI: 10.3390/cells10123315

28. Kadota T., Fujita Y., Yoshioka Y. et al. Emerging role of extracellular vesicles as a senescence-associated secretory phenotype: Insights into the pathophysiology of lung diseases. Mol Aspects med 2018;60:92–103. DOI: 10.1016/j.mam.2017.11.005

29. Jakhar R., Crasta K. Exosomes as emerging pro-tumorigenic mediators of the senescence-associated secretory phenotype. Int J Mol Sci 2019;20(10):2547. DOI: 10.3390/ijms20102547

30. Yamauchi S., Takahashi A. Cellular senescence: mechanisms and relevance to cancer and aging. J Biochem 2025;177(3):163–9. DOI: 10.1093/jb/mvae079

31. Shimizu K., Inuzuka H., Tokunaga F. The interplay between cell death and senescence in cancer. Sem Cancer Biol 2025;108:1–16. DOI: 10.1016/j.semcancer.2024.11.001

32. Gorgoulis V., Adams P.D., Alimonti A. et al. Cellular Senescence: defining a path forward. Cell 2019;179(4):813–27. DOI: 10.1016/j.cell.2019.10.005

33. Ewald J. A., Desotelle J.A., Wilding G. et al. Therapy-induced senescence in cancer. J Nat Cancer Inst 2010;102(20):1536–46. DOI: 10.1093/jnci/djq364

34. Semreen M.H., Alniss H., Cacciatore S. et al. GC-MS based comparative metabolomic analysis of MCF-7 and MDA-MB-231 cancer cells treated with tamoxifen and/or paclitaxel. J Proteomics 2020;225:103875. DOI: 10.1016/j.jprot.2020.103875

35. Lin H., Ai D., Liu Q. et al. Natural isoflavone glabridin targets PI3Kγ as an adjuvant to increase the sensitivity of MDA-MB-231 to tamoxifen and DU145 to paclitaxel. J Steroid Biochem Mol Biol 2024;236:106426. DOI: 10.1016/j.jsbmb.2023.106426

36. Sorokin D., Shchegolev Y., Scherbakov A. et al. Metformin restores the drug sensitivity of mcf-7 cells resistant derivates via the cooperative modulation of growth and apoptotic-related pathways. Pharmaceuticals (Basel) 2020;13(9):206. DOI: 10.3390/ph13090206