Skip to main content

Oleuropein is a natural inhibitor of PAI-1-mediated proliferation in human ER-/PR- breast cancer cells



Elevated expression of PAI-1 has been widely linked with adverse outcomes in a variety of human cancers, such as breast, gastric and ovarian cancers, rendering PAI-1 a prognostic biomarker. As a result, several chemical inhibitors are currently being developed against PAI-1; however, the clinical setting where they might confer survival benefits has not yet been elucidated.


RNA sequencing data analysis from the TCGA/GTEx cancer portals (n = 3607 samples). In silico molecular docking analyses to predict functional macromolecule interactions. ER-/PR- (MDA-MB-231) and ER+/PR+ (MCF-7) breast cancer cell lines implemented to assess the effect of oleuropein as a natural inhibitor of PAI-1-mediated oncogenic proliferation.


We show that high PAI-1 levels inversely correlate with ER and PR expressions in a wide panel of estrogen/progesterone-responsive human malignancies. By implementing an in silico molecular docking analysis, we identify oleuropein, a phenolic component of olive oil, as a potent PAI-1-binding molecule displaying increased affinity compared to the other olive oil constituents. We demonstrate that EVOO or oleuropein treatment alone may act as a natural PAI-1 inhibitor by incrementally destabilising PAI-1 levels selectively in ER-/PR- breast cancer cells, accompanied by downstream caspase activation and cell growth inhibition. In contrast, ER+/PR+ breast cancer cells, where PAI-1 expression is absent or low, do not adequately respond to treatment.


Our study demonstrates an inverse correlation between PAI-1 and ESR1/PGR levels, as well as overall patient survival in estrogen/progesterone-responsive human tumours. With a focus on breast cancer, our data identify oleuropein as a natural PAI-1 inhibitor and suggest that oleuropein-mediated PAI-1 destabilisation may confer clinical benefit only in ER-/PR- tumours.

This is a preview of subscription content, access via your institution.

Fig. 1
Fig. 2
Fig. 3
Fig. 4


  1. 1.

    Li S et al (2018) Plasminogen activator inhibitor-1 in cancer research. Biomed Pharmacother 105:83–94

    CAS  PubMed  Google Scholar 

  2. 2.

    Duffy MJ et al (2014) uPA and PAI-1 as biomarkers in breast cancer: validated for clinical use in level-of-evidence-1 studies. Breast Cancer Res 16(4):428

    PubMed  PubMed Central  Google Scholar 

  3. 3.

    Harbeck N et al (2013) Ten-year analysis of the prospective multicentre Chemo-N0 trial validates American Society of Clinical Oncology (ASCO)-recommended biomarkers uPA and PAI-1 for therapy decision making in node-negative breast cancer patients. Eur J Cancer 49(8):1825–35

    CAS  PubMed  Google Scholar 

  4. 4.

    Ferroni P et al (2014) Plasma plasminogen activator inhibitor-1 (PAI-1) levels in breast cancer - relationship with clinical outcome. Anticancer Res 34(3):1153–61

    PubMed  Google Scholar 

  5. 5.

    Placencio VR, DeClerck YA (2015) Plasminogen activator inhibitor-1 in cancer: rationale and insight for future therapeutic testing. Cancer Res 75(15):2969–74

    CAS  PubMed  PubMed Central  Google Scholar 

  6. 6.

    Bajou K et al (2008) Plasminogen activator inhibitor-1 protects endothelial cells from FasL-mediated apoptosis. Cancer Cell 14(4):324–34

    CAS  PubMed  PubMed Central  Google Scholar 

  7. 7.

    Fang H, Placencio VR, DeClerck YA (2012) Protumorigenic activity of plasminogen activator inhibitor-1 through an antiapoptotic function. J Natl Cancer Inst 104(19):1470–84

    CAS  PubMed  PubMed Central  Google Scholar 

  8. 8.

    Henderson BE, Feigelson HS (2000) Hormonal carcinogenesis. Carcinogenesis 21(3):427–33

    CAS  PubMed  Google Scholar 

  9. 9.

    Jordan VC (2007) Chemoprevention of breast cancer with selective oestrogen-receptor modulators. Nat Rev Cancer 7(1):46–53

    CAS  PubMed  Google Scholar 

  10. 10.

    Trabert B, et al (2020) Progesterone and Breast Cancer. Endocr Rev 41(2)

  11. 11.

    Chen S et al (2017) The positivity of estrogen receptor and progesterone receptor may not be associated with metastasis and recurrence in epithelial ovarian cancer. Sci Rep 7(1):16922

    PubMed  PubMed Central  Google Scholar 

  12. 12.

    Syed V et al (2001) Expression of gonadotropin receptor and growth responses to key reproductive hormones in normal and malignant human ovarian surface epithelial cells. Cancer Res 61(18):6768–76

    CAS  PubMed  Google Scholar 

  13. 13.

    Cancer Genome Atlas Research N, et al (2013) Integrated genomic characterization of endometrial carcinoma. Nature 497(7447):67-73.

  14. 14.

    Di Zazzo E et al (2018) Estrogens and their receptors in prostate cancer: therapeutic implications. Front Oncol 8:2

    PubMed  PubMed Central  Google Scholar 

  15. 15.

    Yu Y et al (2013) Expression and function of the progesterone receptor in human prostate stroma provide novel insights to cell proliferation control. J Clin Endocrinol Metab 98(7):2887–96

    CAS  PubMed  PubMed Central  Google Scholar 

  16. 16.

    Ishibashi H et al (2005) Progesterone receptor in non-small cell lung cancer–a potent prognostic factor and possible target for endocrine therapy. Cancer Res 65(14):6450–8

    CAS  PubMed  Google Scholar 

  17. 17.

    Siegfried JM (2014) Smoking out reproductive hormone actions in lung cancer. Mol Cancer Res 12(1):24–31

    CAS  PubMed  PubMed Central  Google Scholar 

  18. 18.

    Caiazza F et al (2015) Estrogen receptors and their implications in colorectal carcinogenesis. Front Oncol 5:19

    PubMed  PubMed Central  Google Scholar 

  19. 19.

    Dunnwald LK, Rossing MA, Li CI (2007) Hormone receptor status, tumor characteristics, and prognosis: a prospective cohort of breast cancer patients. Breast Cancer Res 9(1):R6

    PubMed  PubMed Central  Google Scholar 

  20. 20.

    Hua H et al (2018) Mechanisms for estrogen receptor expression in human cancer. Exp Hematol Oncol 7:24

    PubMed  PubMed Central  Google Scholar 

  21. 21.

    Louie MC, Sevigny MB (2017) Steroid hormone receptors as prognostic markers in breast cancer. Am J Cancer Res 7(8):1617–1636

    CAS  PubMed  PubMed Central  Google Scholar 

  22. 22.

    Yu CP et al (2013) Estrogen inhibits renal cell carcinoma cell progression through estrogen receptor-beta activation. PLoS One 8(2):e56667

    CAS  PubMed  PubMed Central  Google Scholar 

  23. 23.

    Zhai Y et al (2010) Loss of estrogen receptor 1 enhances cervical cancer invasion. Am J Pathol 177(2):884–95

    CAS  PubMed  PubMed Central  Google Scholar 

  24. 24.

    Lee P et al (2005) Expression of progesterone receptor is a favorable prognostic marker in ovarian cancer. Gynecol Oncol 96(3):671–7

    CAS  PubMed  Google Scholar 

  25. 25.

    Look MP et al (2002) Pooled analysis of prognostic impact of urokinase-type plasminogen activator and its inhibitor PAI-1 in 8377 breast cancer patients. J Natl Cancer Inst 94(2):116–28

    CAS  PubMed  Google Scholar 

  26. 26.

    Smith LH et al (2004) Differential and opposing regulation of PAI-1 promoter activity by estrogen receptor alpha and estrogen receptor beta in endothelial cells. Circ Res 95(3):269–75

    CAS  PubMed  Google Scholar 

  27. 27.

    Brooks TD et al (2004) XR5967, a novel modulator of plasminogen activator inhibitor-1 activity, suppresses tumor cell invasion and angiogenesis in vitro. Anticancer Drugs 15(1):37–44

    CAS  PubMed  Google Scholar 

  28. 28.

    Giacoia EG et al (2014) PAI-1 leads to G1-phase cell-cycle progression through cyclin D3/cdk4/6 upregulation. Mol Cancer Res 12(3):322–34

    CAS  PubMed  PubMed Central  Google Scholar 

  29. 29.

    Lee YC et al (2016) Plasminogen activator inhibitor-1 as regulator of tumor-initiating cell properties in head and neck cancers. Head Neck 38(Suppl 1):E895-904

    PubMed  Google Scholar 

  30. 30.

    Masuda T et al (2013) SK-216, an inhibitor of plasminogen activator inhibitor-1, limits tumor progression and angiogenesis. Mol Cancer Ther 12(11):2378–88

    CAS  PubMed  Google Scholar 

  31. 31.

    Gouda MM, Prabhu A, Bhandary YP (2018) Curcumin alleviates IL-17A-mediated p53-PAI-1 expression in bleomycin-induced alveolar basal epithelial cells. J Cell Biochem 119(2):2222–2230

    CAS  PubMed  Google Scholar 

  32. 32.

    Wang X et al (2017) Oxymatrine inhibits the migration of human colorectal carcinoma RKO cells via inhibition of PAI-1 and the TGF-beta1/Smad signaling pathway. Oncol Rep 37(2):747–753

    CAS  PubMed  Google Scholar 

  33. 33.

    Papaspyropoulos A et al (2020) Modeling and targeting alzheimer’s disease with organoids. Front Pharmacol 11:396

  34. 34.

    Tzekaki EE et al (2020) Restoration of BMI1 levels after the administration of early harvest extra virgin olive oil as a therapeutic strategy against Alzheimer’s disease. Exp Gerontol 144:111178

  35. 35.

    Angeloni C et al (2017) Bioactivity of olive oil phenols in neuroprotection. Int J Mol Sci 18(11)

  36. 36.

    Corominas-Faja B et al (2018) Extra-virgin olive oil contains a metabolo-epigenetic inhibitor of cancer stem cells. Carcinogenesis 39(4):601–613

    CAS  PubMed  PubMed Central  Google Scholar 

  37. 37.

    Rossi M et al (2017) Protection by extra virgin olive oil against oxidative stress in vitro and in vivo.Chemical and biological studies on the health benefits due to a major component of the Mediterranean diet. PLoS One 12(12):e0189341

    PubMed  PubMed Central  Google Scholar 

  38. 38.

    Piroddi M et al (2017) Nutrigenomics of extra-virgin olive oil: A review. Biofactors 43(1):17–41

    CAS  PubMed  Google Scholar 

  39. 39.

    Han J et al (2009) Anti-proliferative and apoptotic effects of oleuropein and hydroxytyrosol on human breast cancer MCF-7 cells. Cytotechnology 59(1):45–53

    CAS  PubMed  PubMed Central  Google Scholar 

  40. 40.

    Elamin MH et al (2013) Olive oil oleuropein has anti-breast cancer properties with higher efficiency on ER-negative cells. Food Chem Toxicol 53:310–6

    CAS  PubMed  Google Scholar 

  41. 41.

    Akl MR et al (2014) Olive phenolics as c-Met inhibitors: (-)-Oleocanthal attenuates cell proliferation, invasiveness, and tumor growth in breast cancer models. PLoS One 9(5):e97622

    PubMed  PubMed Central  Google Scholar 

  42. 42.

    Sachs N et al (2019) Long-term expanding human airway organoids for disease modeling. EMBO J 38(4)

  43. 43.

    Wiener DJ et al (2018) Establishment and characterization of a canine keratinocyte organoid culture system. Vet Dermatol 29(5):375–e126

  44. 44.

    Tang Z et al (2017) GEPIA: a web server for cancer and normal gene expression profiling and interactive analyses. Nucleic Acids Res 45(W1):W98–W102

    CAS  PubMed  PubMed Central  Google Scholar 

  45. 45.

    Tang Z et al (2019) GEPIA2: an enhanced web server for large-scale expression profiling and interactive analysis. Nucleic Acids Res 47(W1):W556–W560

    CAS  PubMed  PubMed Central  Google Scholar 

  46. 46.

    Papaspyropoulos A et al (2018) RASSF1A uncouples Wnt from Hippo signalling and promotes YAP mediated differentiation via p73. Nat Commun 9(1):424

    PubMed  PubMed Central  Google Scholar 

  47. 47.

    Papaneophytou CP et al (2012) Flagellin gene (fliC) of Thermus thermophilus HB8: characterization of its product and involvement to flagella assembly and microbial motility. Appl Microbiol Biotechnol 94(5):1265–77

    CAS  PubMed  Google Scholar 

  48. 48.

    Andreadou E et al (2017) Rhamnolipids, microbial virulence factors, in Alzheimer’s disease. J Alzheimers Dis 59(1):209–222

    CAS  PubMed  Google Scholar 

  49. 49.

    Dai X et al (2017) Breast cancer cell line classification and its relevance with breast tumor subtyping. J Cancer 8(16):3131–3141

    PubMed  PubMed Central  Google Scholar 

  50. 50.

    Uhlen M, et al (2017) A pathology atlas of the human cancer transcriptome. Science 357(6352)

  51. 51.

    Kawarada Y et al (2016) TGF-beta induces p53/Smads complex formation in the PAI-1 promoter to activate transcription. Sci Rep 6:35483

    CAS  PubMed  PubMed Central  Google Scholar 

  52. 52.

    Kortlever RM, Higgins PJ, Bernards R (2006) Plasminogen activator inhibitor-1 is a critical downstream target of p53 in the induction of replicative senescence. Nat Cell Biol 8(8):877–84

    CAS  PubMed  PubMed Central  Google Scholar 

  53. 53.

    Kunz C et al (1995) Differential regulation of plasminogen activator and inhibitor gene transcription by the tumor suppressor p53. Nucleic Acids Res 23(18):3710–7

    CAS  PubMed  PubMed Central  Google Scholar 

  54. 54.

    Shetty S et al (2008) Regulation of plasminogen activator inhibitor-1 expression by tumor suppressor protein p53. J Biol Chem 283(28):19570–80

    CAS  PubMed  PubMed Central  Google Scholar 

  55. 55.

    Mu XC, Higgins PJ (1995) Differential growth state-dependent regulation of plasminogen activator inhibitor type-1 expression in senescent IMR-90 human diploid fibroblasts. J Cell Physiol 165(3):647–57

    CAS  PubMed  Google Scholar 

  56. 56.

    Wang S et al (2013) PAI-1 4G/5G polymorphism contributes to cancer susceptibility: evidence from meta-analysis. PLoS One 8(2):e56797

    CAS  PubMed  PubMed Central  Google Scholar 

  57. 57.

    Goldsmith CD, et al (2018) The olive biophenols oleuropein and hydroxytyrosol selectively reduce proliferation, influence the cell cycle, and induce apoptosis in pancreatic cancer cells. Int J Mol Sci 19(7)

  58. 58.

    LeGendre O, Breslin PA, Foster DA (2015) (-)-Oleocanthal rapidly and selectively induces cancer cell death via lysosomal membrane permeabilization. Mol Cell Oncol 2(4):e1006077

    PubMed  PubMed Central  Google Scholar 

  59. 59.

    Menendez JA et al (2007) Olive oil’s bitter principle reverses acquired autoresistance to trastuzumab (Herceptin) in HER2-overexpressing breast cancer cells. BMC Cancer 7:80

    PubMed  PubMed Central  Google Scholar 

Download references


We would like to thank Yanni’s Olive Grove Company in Potidea Chalkidiki, Greece for providing the early harvest EVOO.


This research is co-financed by Greece and the European Union (European Social Fund- ESF) through the Operational Program «Human Resources Development, Education and Lifelong Learning» in the context of the project ‘Reinforcement of Postdoctoral Researchers - 2nd Cycle’ (grant number: 2019-050-0503-18066), implemented by the State Scholarships Foundation (ΙΚΥ).

Author information




EET performed experiments, analysed and interpreted data. G.G. performed the in silico data analysis. SNL and MPT assisted with experimental equipment. AAP contributed towards data interpretation and co-supervised the project. AP designed the study, performed experiments, analysed and interpreted data, wrote the manuscript and supervised the project.

Corresponding author

Correspondence to Angelos Papaspyropoulos.

Ethics declarations

Conflict of interest

All authors declare no conflict of interest.

Ethical approval

This article does not contain any studies with human participants or animals performed by any of the authors.

Additional information

Publisher's Note

Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.

Supplementary Information

Below is the link to the electronic supplementary material.


Supplementary Fig. 1. Densitometry providing quantification for Western blots. (a) Quantification of PAI-1 and CASP-8 levels based on β-actin expression for Fig. 3a. (b) Same as (a), for Fig. 3c. (c) Same as (a), for Fig. 3e. **P<0.01 and ***P<0.001, respectively, of Student’s t-test; n.s.: non-significant. Error bars indicate s.e.m. Data shown are representative of at least 3 independent experiments. Supplementary Fig. 2 PAI-1 and TP53 correlation in estrogen/progesterone-responsive tumours. (a) Correlation analysis between PAI-1 and TP53 levels expressed in transcripts per million (TPM) across 9 estrogen/progesterone-responsive human tumour types (BRCA, COAD, LUAD, LUSC, OV, PRAD, READ, UCEC, UCS; n indicates total number of tumour samples). RNA seq data were retrieved from the TCGA and GTEx databases and analysed using the GEPIA and GEPIA2 online tools. (b) Overall survival data of cancer patients in (a) based on TP53 expression levels. TP53 level changes do not confer a significant effect on survival (p=0.58). See also Fig 1. Data were retrieved from the TCGA and GTEx databases using the GEPIA online tool(pptx 335 kb)


Supplementary Table 1 Characterisation of the phenolic and non-phenolic content of EVOO used in in vitro experiments (pptx 73 kb)

Rights and permissions

Reprints and Permissions

About this article

Verify currency and authenticity via CrossMark

Cite this article

Tzekaki, E.E., Geromichalos, G., Lavrentiadou, S.N. et al. Oleuropein is a natural inhibitor of PAI-1-mediated proliferation in human ER-/PR- breast cancer cells. Breast Cancer Res Treat 186, 305–316 (2021).

Download citation


  • Plasminogen activator inhibitor-1 (PAI-1)
  • Extra virgin olive oil (EVOO)
  • Oleuropein
  • ER/PR-responsive human cancer
  • MDA-MB-231
  • MCF-7