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Ferritin stimulates breast cancer cells through an iron-independent mechanism and is localized within tumor-associated macrophages

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Abstract

Tumor-associated macrophages play a critical role in breast tumor progression; however, it is still unclear what effector molecular mechanisms they employ to impact tumorigenesis. Ferritin is the primary intracellular iron storage protein and is also abundant in circulation. In breast cancer patients, ferritin is detected at higher levels in both serum and tumor lysates, and its increase correlates with poor clinical outcome. In this study, we comprehensively examined the distribution of ferritin in normal and malignant breast tissue at different stages in tumor development. Decreased ferritin expression in cancer cells but increased infiltration of ferritin-rich CD68-positive macrophages was observed with increased tumor histological grade. Interestingly, ferritin stained within the stroma surrounding tumors suggesting local release within the breast. In cell culture, macrophages, but not breast cancer cells, were capable of ferritin secretion, and this secretion was further increased in response to pro-inflammatory cytokines. We next examined the possible functional significance of extracellular ferritin in a breast cancer cell culture model. Ferritin stimulated the proliferation of the epithelial breast cancer cell lines MCF7 and T47D. Moreover, this proliferative effect was independent of the iron content of ferritin and did not increase intracellular iron levels in cancer cells indicating a novel iron-independent function for this protein. Together, these findings suggest that the release of ferritin by infiltrating macrophages in breast tumors may represent an inflammatory effector mechanism by which ferritin directly stimulates tumorigenesis.

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References

  1. Harrison PM, Arosio P (1996) The ferritins: molecular properties, iron storage function and cellular regulation. Biochim Biophys Acta 1275(3):161–203

    Article  PubMed  Google Scholar 

  2. Boyd D, Vecoli C, Belcher DM, Jain SK, Drysdale JW (1985) Structural and functional relationships of human ferritin H and L chains deduced from cDNA clones. J Biol Chem 260(21):11755–11761

    PubMed  CAS  Google Scholar 

  3. Lawson DM, Artymiuk PJ, Yewdall SJ, Smith JM, Livingstone JC, Treffry A, Luzzago A, Levi S, Arosio P, Cesareni G et al (1991) Solving the structure of human H ferritin by genetically engineering intermolecular crystal contacts. Nature 349(6309):541–544. doi:10.1038/349541a0

    Article  PubMed  CAS  Google Scholar 

  4. Lawson DM, Treffry A, Artymiuk PJ, Harrison PM, Yewdall SJ, Luzzago A, Cesareni G, Levi S, Arosio P (1989) Identification of the ferroxidase centre in ferritin. FEBS Lett 254(1–2):207–210

    Article  PubMed  CAS  Google Scholar 

  5. Levi S, Yewdall SJ, Harrison PM, Santambrogio P, Cozzi A, Rovida E, Albertini A, Arosio P (1992) Evidence of H- and L-chains have co-operative roles in the iron-uptake mechanism of human ferritin. Biochem J 288(Pt 2):591–596

    PubMed  CAS  Google Scholar 

  6. Todorich B, Zhang X, Connor JR (2011) H-ferritin is the major source of iron for oligodendrocytes. Glia 59(6):927–935. doi:10.1002/glia.21164

    Article  PubMed  Google Scholar 

  7. Todorich B, Zhang X, Slagle-Webb B, Seaman WE, Connor JR (2008) Tim-2 is the receptor for H-ferritin on oligodendrocytes. J Neurochem 107(6):1495–1505. doi:10.1111/j.1471-4159.2008.05678.x

    Article  PubMed  CAS  Google Scholar 

  8. Coffman LG, Parsonage D, D’Agostino R Jr, Torti FM, Torti SV (2009) Regulatory effects of ferritin on angiogenesis. Proc Natl Acad Sci USA 106(2):570–575. doi:10.1073/pnas.0812010106

    Article  PubMed  CAS  Google Scholar 

  9. Tesfay L, Huhn AJ, Hatcher H, Torti FM, Torti SV (2012) Ferritin blocks inhibitory effects of two-chain high molecular weight kininogen (HKa) on adhesion and survival signaling in endothelial cells. PLoS ONE 7(7):e40030. doi:10.1371/journal.pone.0040030

    Article  PubMed  CAS  Google Scholar 

  10. Alkhateeb A, Leitzel K, Ali SM, Campbell-Baird C, Evans M, Fuchs E, Köstler WJ, Lipton A, Connor J (2012) Elevation in serum inflammatory biomarkers predicts response to trastuzumab-containing therapy. PLoS ONE 7(12):e51379. doi:10.1371/journal.pone.0051379

  11. Jones BM, Worwood M, Jacobs A (1980) Serum ferritin in patients with cancer: determination with antibodies to HeLa cell and spleen ferritin. Clin Chim Acta 106(2):203–214

    Article  PubMed  CAS  Google Scholar 

  12. Robertson JF, Pearson D, Price MR, Selby C, Pearson J, Blamey RW, Howell A (1991) Prospective assessment of the role of five tumour markers in breast cancer. Cancer Immunol Immunother 33(6):403–410

    Article  PubMed  CAS  Google Scholar 

  13. Mannello F, Tonti GA, Medda V, Simone P, Darbre PD (2011) Analysis of aluminium content and iron homeostasis in nipple aspirate fluids from healthy women and breast cancer-affected patients. J Appl Toxicol 31(3):262–269. doi:10.1002/jat.1641

    Article  PubMed  CAS  Google Scholar 

  14. Tappin JA, George WD, Bellingham AJ (1979) Effect of surgery on serum ferritin concentration in patients with breast cancer. Br J Cancer 40(4):658–660

    Article  PubMed  CAS  Google Scholar 

  15. Jacobs A, Jones B, Ricketts C, Bulbrook RD, Wang DY (1976) Serum ferritin concentration in early breast cancer. Br J Cancer 34(3):286–290

    Article  PubMed  CAS  Google Scholar 

  16. Arosio P, Yokota M, Drysdale JW (1977) Characterization of serum ferritin in iron overload: possible identity to natural apoferritin. Br J Haematol 36(2):199–207

    Article  PubMed  CAS  Google Scholar 

  17. Weinstein RE, Bond BH, Silberberg BK (1982) Tissue ferritin concentration in carcinoma of the breast. Cancer 50(11):2406–2409

    Article  PubMed  CAS  Google Scholar 

  18. Weinstein RE, Bond BH, Silberberg BK, Vaughn CB, Subbaiah P, Pieper DR (1989) Tissue ferritin concentration and prognosis in carcinoma of the breast. Breast Cancer Res Treat 14(3):349–353

    Article  PubMed  CAS  Google Scholar 

  19. Rossiello R, Carriero MV, Giordano GG (1984) Distribution of ferritin, transferrin and lactoferrin in breast carcinoma tissue. J Clin Pathol 37(1):51–55

    Article  PubMed  CAS  Google Scholar 

  20. Jezequel P, Campion L, Spyratos F, Loussouarn D, Campone M, Guerin-Charbonnel C, Joalland MP, Andre J, Descotes F, Grenot C, Roy P, Carlioz A, Martin PM, Chassevent A, Jourdan ML, Ricolleau G (2012) Validation of tumor-associated macrophage ferritin light chain as a prognostic biomarker in node-negative breast cancer tumors: a multicentric 2004 national PHRC study. Int J Cancer 131(2):426–437. doi:10.1002/ijc.26397

    Article  PubMed  CAS  Google Scholar 

  21. Snyder AM, Neely EB, Levi S, Arosio P, Connor JR (2010) Regional and cellular distribution of mitochondrial ferritin in the mouse brain. J Neurosci Res 88(14):3133–3143. doi:10.1002/jnr.22462

    Article  PubMed  CAS  Google Scholar 

  22. Pollard JW (2004) Tumour-educated macrophages promote tumour progression and metastasis. Nat Rev Cancer 4(1):71–78. doi:10.1038/nrc1256

    Article  PubMed  CAS  Google Scholar 

  23. Simson JV, Spicer SS (1972) Ferritin particles in macrophages and in associated mast cells. J Cell Biol 52(3):536–541

    Article  PubMed  CAS  Google Scholar 

  24. Cohen LA, Gutierrez L, Weiss A, Leichtmann-Bardoogo Y, Zhang DL, Crooks DR, Sougrat R, Morgenstern A, Galy B, Hentze MW, Lazaro FJ, Rouault TA, Meyron-Holtz EG (2010) Serum ferritin is derived primarily from macrophages through a nonclassical secretory pathway. Blood 116(9):1574–1584. doi:10.1182/blood-2009-11-253815

    Article  PubMed  CAS  Google Scholar 

  25. Li JY, Paragas N, Ned RM, Qiu A, Viltard M, Leete T, Drexler IR, Chen X, Sanna-Cherchi S, Mohammed F, Williams D, Lin CS, Schmidt-Ott KM, Andrews NC, Barasch J (2009) Scara5 is a ferritin receptor mediating non-transferrin iron delivery. Dev Cell 16(1):35–46. doi:10.1016/j.devcel.2008.12.002

    Article  PubMed  CAS  Google Scholar 

  26. Breuer W, Epsztejn S, Cabantchik ZI (1995) Iron acquired from transferrin by K562 cells is delivered into a cytoplasmic pool of chelatable iron(II). J Biol Chem 270(41):24209–24215

    Article  PubMed  CAS  Google Scholar 

  27. Ruddell RG, Hoang-Le D, Barwood JM, Rutherford PS, Piva TJ, Watters DJ, Santambrogio P, Arosio P, Ramm GA (2009) Ferritin functions as a proinflammatory cytokine via iron-independent protein kinase C zeta/nuclear factor kappaB-regulated signaling in rat hepatic stellate cells. Hepatology 49(3):887–900. doi:10.1002/hep.22716

    Article  PubMed  CAS  Google Scholar 

  28. Hogemann-Savellano D, Bos E, Blondet C, Sato F, Abe T, Josephson L, Weissleder R, Gaudet J, Sgroi D, Peters PJ, Basilion JP (2003) The transferrin receptor: a potential molecular imaging marker for human cancer. Neoplasia 5(6):495–506

    PubMed  Google Scholar 

  29. Pinnix ZK, Miller LD, Wang W, D’Agostino R Jr, Kute T, Willingham MC, Hatcher H, Tesfay L, Sui G, Di X, Torti SV, Torti FM (2010) Ferroportin and iron regulation in breast cancer progression and prognosis. Sci Transl Med 2(43):43ra56. doi:10.1126/scisignal.3001127

  30. Lin EY, Li JF, Gnatovskiy L, Deng Y, Zhu L, Grzesik DA, Qian H, Xue XN, Pollard JW (2006) Macrophages regulate the angiogenic switch in a mouse model of breast cancer. Cancer Res 66(23):11238–11246. doi:10.1158/0008-5472.CAN-06-1278

    Article  PubMed  CAS  Google Scholar 

  31. DeNardo DG, Barreto JB, Andreu P, Vasquez L, Tawfik D, Kolhatkar N, Coussens LM (2009) CD4(+) T cells regulate pulmonary metastasis of mammary carcinomas by enhancing protumor properties of macrophages. Cancer Cell 16(2):91–102. doi:10.1016/j.ccr.2009.06.018

    Article  PubMed  CAS  Google Scholar 

  32. Ruffell B, Au A, Rugo HS, Esserman LJ, Hwang ES, Coussens LM (2012) Leukocyte composition of human breast cancer. Proc Natl Acad Sci USA 109(8):2796–2801. doi:10.1073/pnas.1104303108

    Article  PubMed  CAS  Google Scholar 

  33. Chen Q, Zhang XH, Massague J (2011) Macrophage binding to receptor VCAM-1 transmits survival signals in breast cancer cells that invade the lungs. Cancer Cell 20(4):538–549. doi:10.1016/j.ccr.2011.08.025

    Article  PubMed  CAS  Google Scholar 

  34. DeNardo DG, Brennan DJ, Rexhepaj E, Ruffell B, Shiao SL, Madden SF, Gallagher WM, Wadhwani N, Keil SD, Junaid SA, Rugo HS, Hwang ES, Jirstrom K, West BL, Coussens LM (2011) Leukocyte complexity predicts breast cancer survival and functionally regulates response to chemotherapy. Cancer Discov 1(1):54–67. doi:10.1158/2159-8274.CD-10-0028

    Article  PubMed  CAS  Google Scholar 

  35. Shree T, Olson OC, Elie BT, Kester JC, Garfall AL, Simpson K, Bell-McGuinn KM, Zabor EC, Brogi E, Joyce JA (2011) Macrophages and cathepsin proteases blunt chemotherapeutic response in breast cancer. Genes Dev 25(23):2465–2479. doi:10.1101/gad.180331.111

    Article  PubMed  CAS  Google Scholar 

  36. Partin AW, Criley SR, Steiner MS, Hsieh K, Simons JW, Lumadue J, Carter HB, Marshall FF (1995) Serum ferritin as a clinical marker for renal cell carcinoma: influence of tumor volume. Urology 45(2):211–217

    Article  PubMed  CAS  Google Scholar 

  37. Sato Y, Honda Y, Asoh T, Oizumi K, Ohshima Y, Honda E (1998) Cerebrospinal fluid ferritin in glioblastoma: evidence for tumor synthesis. J Neurooncol 40(1):47–50

    Article  PubMed  CAS  Google Scholar 

  38. Tran TN, Eubanks SK, Schaffer KJ, Zhou CY, Linder MC (1997) Secretion of ferritin by rat hepatoma cells and its regulation by inflammatory cytokines and iron. Blood 90(12):4979–4986

    PubMed  CAS  Google Scholar 

  39. Ghosh S, Hevi S, Chuck SL (2004) Regulated secretion of glycosylated human ferritin from hepatocytes. Blood 103(6):2369–2376. doi:10.1182/blood-2003-09-3050

    Article  PubMed  CAS  Google Scholar 

  40. Ferring-Appel D, Hentze MW, Galy B (2009) Cell-autonomous and systemic context-dependent functions of iron regulatory protein 2 in mammalian iron metabolism. Blood 113(3):679–687. doi:10.1182/blood-2008-05-155093

    Article  PubMed  CAS  Google Scholar 

  41. Singh M, Lu J, Briggs SP, McGinley JN, Haegele AD, Thompson HJ (1994) Effect of excess dietary iron on the promotion stage of 1-methyl-1-nitrosourea-induced mammary carcinogenesis: pathogenetic characteristics and distribution of iron. Carcinogenesis 15(8):1567–1570

    Article  PubMed  CAS  Google Scholar 

  42. Diwan BA, Kasprzak KS, Anderson LM (1997) Promotion of dimethylbenz[a]anthracene-initiated mammary carcinogenesis by iron in female Sprague–Dawley rats. Carcinogenesis 18(9):1757–1762

    Article  PubMed  CAS  Google Scholar 

  43. Broxmeyer HE, Williams DE, Geissler K, Hangoc G, Cooper S, Bicknell DC, Levi S, Arosio P (1989) Suppressive effects in vivo of purified recombinant human H-subunit (acidic) ferritin on murine myelopoiesis. Blood 73(1):74–79

    PubMed  CAS  Google Scholar 

  44. Cozzi A, Corsi B, Levi S, Santambrogio P, Biasiotto G, Arosio P (2004) Analysis of the biologic functions of H- and L-ferritins in HeLa cells by transfection with siRNAs and cDNAs: evidence for a proliferative role of L-ferritin. Blood 103(6):2377–2383. doi:10.1182/blood-2003-06-1842

    Article  PubMed  CAS  Google Scholar 

  45. Johnson TW, Anderson KE, Lazovich D, Folsom AR (2002) Association of aspirin and nonsteroidal anti-inflammatory drug use with breast cancer. Cancer Epidemiol Biomark Prev 11(12):1586–1591

    CAS  Google Scholar 

  46. Cotterchio M, Kreiger N, Sloan M, Steingart A (2001) Nonsteroidal anti-inflammatory drug use and breast cancer risk. Cancer Epidemiol Biomark Prev 10(11):1213–1217

    CAS  Google Scholar 

  47. Pham CG, Bubici C, Zazzeroni F, Papa S, Jones J, Alvarez K, Jayawardena S, De Smaele E, Cong R, Beaumont C, Torti FM, Torti SV, Franzoso G (2004) Ferritin heavy chain upregulation by NF-kappaB inhibits TNFalpha-induced apoptosis by suppressing reactive oxygen species. Cell 119(4):529–542. doi:10.1016/j.cell.2004.10.017

    Article  PubMed  CAS  Google Scholar 

  48. Fleming DJ, Jacques PF, Massaro JM, D’Agostino RB Sr, Wilson PW, Wood RJ (2001) Aspirin intake and the use of serum ferritin as a measure of iron status. Am J Clin Nutr 74(2):219–226

    PubMed  CAS  Google Scholar 

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Ethical Standards

The experiments presented in this manuscript comply with the current laws and standards of the USA and have been approved by the institutional review board at Penn State Hershey Medical Center.

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The authors declare that they have no conflict of interest.

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Correspondence to James R. Connor.

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10549_2012_2405_MOESM1_ESM.tif

Supplementary Fig. 1. a The L-ferritin antibody used in immunohistochemistry specifically recognized recombinant L-ferritin (rLFr) but not recombinant H-ferritin (rHFR). The L-subunit-rich human liver ferritin was used as a positive control. The appearance of the 17 kDa band in liver ferritin has been shown to be due to lysosomal cleavage of the L-ferritin protein [24]. Each lane was loaded with 0.25 μg of protein b Western blot analysis showing no phosphorylation of IKKα/β following exposure to 25nM ferritin extracted from the human liver. c Western blot analysis of total protein lysates from breast cancer cell lines shows that both responsive (i.e. MCF7) and unresponsive (i.e. MDA-MB-231) cell lines express Scara5. The expected molecular weight of Scara5 based on amino acid sequence is 53 kDa. (TIFF 2260 kb)

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Alkhateeb, A.A., Han, B. & Connor, J.R. Ferritin stimulates breast cancer cells through an iron-independent mechanism and is localized within tumor-associated macrophages. Breast Cancer Res Treat 137, 733–744 (2013). https://doi.org/10.1007/s10549-012-2405-x

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