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Estrogen promotes the brain metastatic colonization of triple negative breast cancer cells via an astrocyte-mediated paracrine mechanism

Abstract

Brain metastases (BM) are a devastating consequence of breast cancer. BM occur more frequently in patients with estrogen receptor-negative (ER−) breast cancer subtypes; HER2 overexpressing (HER2+) tumors and triple-negative (TN) (ER−, progesterone receptor-negative (PR–) and normal HER2) tumors. Young age is an independent risk factor for the development of BM, thus we speculated that higher circulating estrogens in young, pre-menopausal women could exert paracrine effects through the highly estrogen-responsive brain microenvironment. Using a TN experimental metastases model, we demonstrate that ovariectomy decreased the frequency of magnetic resonance imaging-detectable lesions by 56% as compared with estrogen supplementation, and that the combination of ovariectomy and letrozole further reduced the frequency of large lesions to 14.4% of the estrogen control. Human BM expressed 4.2–48.4% ER+ stromal area, particularly ER+ astrocytes. In vitro, E2-treated astrocytes increased proliferation, migration and invasion of 231BR-EGFP cells in an ER-dependent manner. E2 upregulated epidermal growth factor receptor (EGFR) ligands Egf, Ereg and Tgfa mRNA and protein levels in astrocytes, and activated EGFR in brain metastatic cells. Co-culture of 231BR-EGFP cells with E2-treated astrocytes led to the upregulation of the metastatic mediator S100 Calcium-binding protein A4 (S100A4) (1.78-fold, P<0.05). Exogenous EGF increased S100A4 mRNA levels in 231BR-EGFP cells (1.40±0.02-fold, P<0.01 compared with vehicle control) and an EGFR/HER2 inhibitor blocked this effect, suggesting that S100A4 is a downstream effector of EGFR activation. Short hairpin RNA-mediated S100A4 silencing in 231BR-EGFP cells decreased their migration and invasion in response to E2-CM, abolished their increased proliferation in co-cultures with E2-treated astrocytes and decreased brain metastatic colonization. Thus, S100A4 is one effector of the paracrine action of E2 in brain metastatic cells. These studies provide a novel mechanism by which estrogens, acting through ER+ astrocytes in the brain microenvironment, can promote BM of TN breast cancers, and suggests existing endocrine agents may provide some clinical benefit towards reducing and managing BM.

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References

  1. Cheng X, Hung MC . Breast cancer brain metastases. Cancer Metastasis Rev 2007; 26: 635–643.

    Article  PubMed  Google Scholar 

  2. Brufsky AM, Mayer M, Rugo HS, Kaufman PA, Tan-Chiu E, Tripathy D et al. Central nervous system metastases in patients with HER2-positive metastatic breast cancer: incidence, treatment, and survival in patients from registHER. Clin Cancer Res 2011; 17: 4834–4843.

    Article  CAS  PubMed  Google Scholar 

  3. Kodack DP, Askoxylakis V, Ferraro GB, Fukumura D, Jain RK . Emerging strategies for treating brain metastases from breast cancer. Cancer Cell 2015; 27: 163–175.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  4. Boogerd W, Vos VW, Hart AA, Baris G . Brain metastases in breast cancer; natural history, prognostic factors and outcome. J Neurooncol 1993; 15: 165–174.

    Article  CAS  PubMed  Google Scholar 

  5. Braccini AL, Azria D, Thezenas S, Romieu G, Ferrero JM, Jacot W . Prognostic factors of brain metastases from breast cancer: Impact of targeted therapies. Breast 2013; 22: 993–998.

    Article  PubMed  Google Scholar 

  6. Evans AJ, James JJ, Cornford EJ, Chan SY, Burrell HC, Pinder SE et al. Brain metastases from breast cancer: identification of a high-risk group. Clin Oncol (R Coll Radiol) 2004; 16: 345–349.

    Article  CAS  Google Scholar 

  7. Hicks DG, Short SM, Prescott NL, Tarr SM, Coleman KA, Yoder BJ et al. Breast cancers with brain metastases are more likely to be estrogen receptor negative, express the basal cytokeratin CK5/6, and overexpress HER2 or EGFR. Am J Surg Pathol 2006. 1097–1104.

  8. Kaal ECA, CGJH Niël, Vecht CJ . Therapeutic management of brain metastasis. Lancet Neurol 2005; 4: 289–298.

    Article  PubMed  Google Scholar 

  9. Lin NU, Bellon JR, Winer EP . CNS metastases in breast cancer. J Clin Oncol 2004; 22: 3608–3617.

    Article  PubMed  Google Scholar 

  10. Tham Y-L, Sexton K, Kramer R, Hilsenbeck S, Elledge R . Primary breast cancer phenotypes associated with propensity for central nervous system metastases. Cancer 2006; 107: 696–704.

    Article  PubMed  Google Scholar 

  11. Lin NU, Vanderplas A, Hughes ME, Theriault RL, Edge SB, Wong Y-N et al. Clinicopathologic features, patterns of recurrence, and survival among women with triple-negative breast cancer in the National Comprehensive Cancer Network. Cancer 2012; 118: 5463–5472.

    Article  PubMed  Google Scholar 

  12. Lee YT . Breast carcinoma: pattern of metastasis at autopsy. J Surg Oncol 1983; 23: 175–180.

    Article  CAS  PubMed  Google Scholar 

  13. Lee SS, Ahn J-H, Kim MK, Sym SJ, Gong G, Ahn SD et al. Brain metastases in breast cancer: prognostic factors and management. Breast Cancer Res Treat 2007; 111: 523–530.

    Article  PubMed  Google Scholar 

  14. Tsukada Y, Fouad A, Pickren JW, Lane WW . Central nervous system metastasis from breast carcinoma. Autopsy study. Cancer 1983; 52: 2349–2354.

    Article  CAS  PubMed  Google Scholar 

  15. Hung MH, Liu CY, Shiau CY, Hsu CY, Tsai YF, Wang YL et al. Effect of age and biological subtype on the risk and timing of brain metastasis in breast cancer patients. PLoS ONE 2014; 9: e89389.

    Article  PubMed  PubMed Central  Google Scholar 

  16. García-Ovejero D, Veiga S, García-Segura LM, Doncarlos LL . Glial expression of estrogen and androgen receptors after rat brain injury. J Comp Neurol 2002; 450: 256–271.

    Article  PubMed  Google Scholar 

  17. Morissette M, Le Saux M, D'Astous M, Jourdain S, Al Sweidi S, Morin N et al. Contribution of estrogen receptors alpha and beta to the effects of estradiol in the brain. J Steroid Biochem Mol Biol 2008; 108: 327–338.

    Article  CAS  PubMed  Google Scholar 

  18. Pettersson K, Gustafsson J-Å . Role of estrogen receptor beta in estrogen action. Annu Rev Physiol 2001; 63: 165–192.

    Article  CAS  PubMed  Google Scholar 

  19. Razmara A, Sunday L, Stirone C, Wang XB, Krause DN, Duckles SP et al. Mitochondrial effects of estrogen are mediated by estrogen receptor α in brain endothelial cells. J Pharmacol Exp Ther 2008; 325: 782–790.

    Article  CAS  PubMed  Google Scholar 

  20. Arnold S . Estrogen suppresses the impact of glucose deprivation on astrocytic calcium levels and signaling independently of the nuclear estrogen receptor. Neurobiol Dis 2005; 20: 82–92.

    Article  CAS  PubMed  Google Scholar 

  21. Barouk S, Hintz T, Li P, Duffy AM, MacLusky NJ, Scharfman HE . 17β-estradiol increases astrocytic vascular endothelial growth factor (VEGF) in adult female rat hippocampus. Endocrinology 2011; 152: 1745–1751.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  22. Garcia-Segura LM, Azcoitia I, DonCarlos LL . Neuroprotection by estradiol. Prog Neurobiol 2001; 63: 29–60.

    Article  CAS  PubMed  Google Scholar 

  23. Garcia-Segura LM, Wozniak A, Azcoitia I, Rodriguez JR, Hutchison RE, Hutchison JB . Aromatase expression by astrocytes after brain injury: implications for local estrogen formation in brain repair. Neuroscience 1999; 89: 567–578.

    Article  CAS  PubMed  Google Scholar 

  24. Montelli S, Peruffo A, Zambenedetti P, Rossipal E, Giacomello M, Zatta P et al. Expression of aromatase P450AROM in the human fetal and early postnatal cerebral cortex. Brain Res 2012; 1475: 11–18.

    Article  CAS  PubMed  Google Scholar 

  25. Yague JG, Muñoz A, de Monasterio-Schrader P, Defelipe J, Garcia-Segura LM, Azcoitia I . Aromatase expression in the human temporal cortex. Neuroscience 2006; 138: 389–401.

    Article  CAS  PubMed  Google Scholar 

  26. Palmieri D, Bronder JL, Herring JM, Yoneda T, Weil RJ, Stark AM et al. Her-2 overexpression increases the metastatic outgrowth of breast cancer cells in the brain. Cancer Res 2007; 67: 4190–4198.

    Article  CAS  PubMed  Google Scholar 

  27. Yoneda T, Williams P, Hiraga T, Niewolna M, Nishimura R . A bone-seeking clone exhibits different biological properties from the MDA-MB-231 parental human breast cancer cells and a brain-seeking cloe in vivo and in vitro. J Bone Miner Res 2001; 16: 1486–1495.

    Article  CAS  PubMed  Google Scholar 

  28. Svendsen CN, ter Borg MG, Armstrong RJ, Rosser AE, Chandran S, Ostenfeld T et al. A new method for the rapid and long term growth of human neural precursor cells. J Neurosci Methods 1998; 85: 141–152.

    Article  CAS  PubMed  Google Scholar 

  29. Ma YJ, Berg-Von Der Emde K, Moholt-Siebert M, Hill DF, Ojeda SR . Region-Specific Regulation of Transforming Growth Factor Alpha (TGF Alpha) Gene Expression in Astrocytes of the Neuroendocrine Brain. J Neurosci. 1994; 14: 5644–5651.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  30. Lee E, Sidoryk-Wegrzynowicz M, Yin Z, Webb A, Son D-S, Aschner M . Transforming growth factor-α mediates estrogen-induced upregulation of glutamate transporter GLT-1 in rat primary astrocytes. Glia 2012; 60: 1024–1036.

    Article  PubMed  PubMed Central  Google Scholar 

  31. Gril B, Palmieri D, Bronder JL, Herring JM, Vega-Valle E, Feigenbaum L et al. Effect of lapatinib on the outgrowth of metastatic breast cancer cells to the brain. J Natl Cancer Inst 2008; 100: 1092–1103.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  32. Jenkinson SR, Barraclough R, West CR, Rudland PS . S100A4 regulates cell motility and invasion in an in vitro model for breast cancer metastasis. Br J Cancer 2004; 90: 253–262.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  33. Saleem M, Kweon MH, Johnson JJ, Adhami VM, Elcheva I, Khan N et al. S100A4 accelerates tumorigenesis and invasion of human prostate cancer through the transcriptional regulation of matrix metalloproteinase 9. Proc Natl Acad Sci USA 2006; 103: 14825–14830.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  34. Arnold S, Victor MB, Beyer C . Estrogen and the regulation of mitochondrial structure and function in the brain. J Steroid Biochem Mol Biol 2012; 131: 2–9.

    Article  CAS  PubMed  Google Scholar 

  35. Péqueux C, Raymond-Letron I, Blacher S, Boudou F, Adlanmerini M, Fouque MJ et al. Stromal estrogen receptor-α promotes tumor growth by normalizing an increased angiogenesis. Cancer Res 2012; 72: 3010–3019.

    Article  PubMed  Google Scholar 

  36. Iyer V, Klebba I, McCready J, Arendt LM, Betancur-Boissel M, Wu MF et al. Estrogen promotes ER-negative tumor growth and angiogenesis through mobilization of bone marrow-derived monocytes. Cancer Res 2012; 72: 2705–2713.

    Article  CAS  PubMed  Google Scholar 

  37. Valastyan S, Weinberg RA . Tumor metastasis: molecular insights and evolving paradigms. Cell 2011; 147: 275–292.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  38. Valiente M, Obenauf AC, Jin X, Chen Q, Zhang XH, Lee DJ et al. Serpins promote cancer cell survival and vascular co-option in brain metastasis. Cell 2014; 156: 1002–1016.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  39. Abbott NJ, Ronnback L, Hansson E . Astrocyte-endothelial interactions at the blood-brain barrier. Nat Rev Neurosci 2006; 7: 41–53.

    Article  CAS  PubMed  Google Scholar 

  40. Fidler IJ . The role of the organ microenvironment in brain metastasis. Semin Cancer Biol 2011; 21: 107–112.

    Article  PubMed  Google Scholar 

  41. Gril B, Palmieri D, Qian Y, Anwar T, Liewehr DJ, Steinberg SM et al. Pazopanib inhibits the activation of PDGFRbeta-expressing astrocytes in the brain metastatic microenvironment of breast cancer cells. Am J Pathol 2013; 182: 2368–2379.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  42. Kim S-J, Kim J-S, Park ES, Lee J-S, Lin Q, Langley RR et al. Astrocytes upregulate survival genes in tumor cells and induce protection from chemotherapy. Neoplasia 2011; 13: 286–298.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  43. Azcoitia I, Santos-Galindo M, Arevalo MA, Garcia-Segura LM . Role of astroglia in the neuroplastic and neuroprotective actions of estradiol. Eur J Neurosci 2010; 32: 1995–2002.

    Article  PubMed  Google Scholar 

  44. Dhandapani KM, Wade FM, Mahesh VB, Brann DW . Astrocyte-derived transforming growth factor-β mediates the neuroprotective effects of 17β-estradiol: involvement of nonclassical genomic signaling pathways. Endocrinology 2005; 146: 2749–2759.

    Article  CAS  PubMed  Google Scholar 

  45. Garcia-Segura LM, Torres-Aleman I, Naftolin F . Astrocytic shape and glial fibrillary acidic protein immunoreactivity are modified by estradiol in primary rat hypothalamic cultures. Brain Res Dev Brain Res 1989; 47: 298–302.

    Article  CAS  PubMed  Google Scholar 

  46. Dahlmann M, Sack U, Herrmann P, Lemm M, Fichtner I, Schlag PM et al. Systemic shRNA mediated knock down of S100A4 in colorectal cancer xenografted mice reduces metastasis formation. Oncotarget 2012; 3: 783–797.

    Article  PubMed  PubMed Central  Google Scholar 

  47. Wang L, Wang X, Liang Y, Diao X, Chen Q . S100A4 promotes invasion and angiogenesis in breast cancer MDA-MB-231 cells by upregulating matrix metalloproteinase-13. Acta Biochim Pol 2012; 59: 593–598.

    Article  CAS  PubMed  Google Scholar 

  48. Neman J, Choy C, Kowolik CM, Anderson A, Duenas VJ, Waliany S et al. Co-evolution of breast-to-brain metastasis and neural progenitor cells. Clin Exp Metastasis 2013; 30: 753–768.

    Article  CAS  PubMed  Google Scholar 

  49. Wang L, Cossette SM, Rarick KR, Gershan J, Dwinell MB, Harder DR et al. Astrocytes directly influence tumor cell invasion and metastasis in vivo. PLoS ONE 2013; 8: e80933.

    Article  PubMed  PubMed Central  Google Scholar 

  50. Fong MY, Zhou W, Liu L, Alontaga AY, Chandra M, Ashby J et al. Breast-cancer-secreted miR-122 reprograms glucose metabolism in premetastatic niche to promote metastasis. Nat Cell Biol 2015; 17: 183–194.

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  51. Yoneda T, Williams PJ, Hiraga T, Niewolna M, Nishimura R . A bone-seeking clone exhibits different biological properties from the MDA-MB-231 parental human breast cancer cells and a brain-seeking clone in vivo and in vitro. J Bone Miner Res 2001; 16: 1486–1495.

    Article  CAS  PubMed  Google Scholar 

  52. Wu P, Tarasenko YI, Gu Y, Huang LY, Coggeshall RE, Yu Y . Region-specific generation of cholinergic neurons from fetal human neural stem cells grafted in adult rat. Nat Neurosci 2002; 5: 1271–1278.

    Article  CAS  PubMed  Google Scholar 

  53. Osborne CK, Hobbs K, Clark GM . Effect of estrogens and antiestrogens on growth of human breast cancer cells in athymic nude mice. Cancer Res 1985; 45: 584–590.

    CAS  PubMed  Google Scholar 

  54. Stanczyk FZ, Mathews BW, Sherman ME . Relationships of sex steroid hormone levels in benign and cancerous breast tissue and blood: A critical appraisal of current science. Steroids 2015; 99 (Pt A): 91–102.

    Article  CAS  PubMed  Google Scholar 

  55. Frey L, Lepkin A, Schickedanz A, Huber K, Brown MS, Serkova N . ADC mapping and T1-weighted signal changes on post-injury MRI predict seizure susceptibility after experimental traumatic brain injury. Neurol Res 2014; 36: 26–37.

    Article  PubMed  Google Scholar 

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Acknowledgements

We thank the University of Colorado Cancer Center Animal Imaging Shared Resources, Tissue Culture Core, Cytometry and Cell Sorting Shared Resource and Functional Genomics Facility supported by NCI P30CA046934 and CTSA UL1TR001082 Center grants. Susan Edgerton provided de-identified tissue. This work was supported by DOD BCRP W81XWH-11-1-0101 (DMC), ACS IRG # 57-001-53 (DMC) and NCI K22CA181250 (DMC). CAS was supported by NIH R01 CA140985.

Author contributions

Conception and supervision: DMC; Development of methodology, data acquisition and analysis: DMC, CAS, VFB, TBS, PK, PSS, CTH, HC, KBK and NJS. Manuscript writing and editing: DMC, CAS and PSS. All authors read and approved manuscript.

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Sartorius, C., Hanna, C., Gril, B. et al. Estrogen promotes the brain metastatic colonization of triple negative breast cancer cells via an astrocyte-mediated paracrine mechanism. Oncogene 35, 2881–2892 (2016). https://doi.org/10.1038/onc.2015.353

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