Skip to main content
SpringerLink
Log in

Estradiol control of ornithine decarboxylase mRNA, enzyme activity, and polyamine levels in MCF-7 breast cancer cells: therapeutic implications

  • Report
  • Published:
Breast Cancer Research and Treatment Aims and scope Submit manuscript

Abstract

Previous studies have shown that natural polyamines - putrescine, spermidine, and spermine - play a key role in the mechanism of action of estrogens in breast cancer. Ornithine decarboxylase (ODC) is the first enzyme of the polyamine biosynthetic pathway. To examine estrogenic regulation of polyamine biosynthesis in breast cancer, we measured ODC mRNA, ODC activity, and polyamine levels in G1 synchronized MCF-7 cells. ODC mRNA and activity increased four-fold over that of cells in G1 phase between 8 to 16 h after the addition of estradiol. Polyamine levels showed a sharp increase by 8 h after the addition of estradiol and decreased by 12 h. We further examined whether synthetic homologs of putrescine or spermidine could replace natural polyamines in supporting MCF-7 cell growth. Treatment of MCF-7 cells with 1 mM difluoromethylornithine (DFMO), an inhibitor of ODC, suppressed putrescine, spermidine, and spermine levels by 74, 78, and 10%, respectively, within 48 h. Cells treated with DFMO for 48 h were supplemented with either putrescine or its homologs or spermidine or its homologs. Diaminopropane, diaminobutane (putrescine), and diaminopentane were capable of fully or partially reversing the growth inhibitory effects of DFMO, whereas diaminoethane had no significant effect. Among a series of triamines, H2N(CH2)nNH(CH2)3NH2 (where n = 2 to 8; abbreviated as APn n = 4 for spermidine, or AP4), spermidine was most effective in reversing the effects of DFMO, whereas compounds with shorter or longer methylene bridging regions were less effective. AP8 was ineffective in reversing the growth inhibitory effects of DFMO. At 10 µM concentration, AP8 also inhibited DNA synthesis by 66%, as measured by [3H]-thymidine incorporation. These data show that MCF-7 cells have a strong requirement for polyamines for their growth and that estradiol stimulates the polyamine cascade by inducing the ODC mRNA level. Our results also suggest that polyamine homologs such as AP8 might be potentially useful in breast cancer therapy.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

References

  1. Beatson GT: On the treatment of inoperable cases of carcinoma of the mamma: suggestion for a new method of treatment with illustrative cases. Lancet 2: 104–107, 1896

    Google Scholar 

  2. Kiang DT, Frenning DH, Goldman AI, Aslensa VF, Kennedy BJ: Estrogen receptors and responses to chemotherapy and hormonal therapy in advanced breast cancer. N Engl J Med 229: 1330–1334, 1984

    Google Scholar 

  3. Allegra JC, Lippman ME, Thompson EB: Relationship between the progesterone, androgen, and glucocorticoid receptors and response to endocrine therapy in metastatic breast cancer. Cancer Res 39: 1973–1979, 1979

    Google Scholar 

  4. Manni A, Pearson O: Antiestrogen-induced remission in premenopausal women with stage IV breast cancer. Cancer Treat Rep 64: 779–785, 1980

    Google Scholar 

  5. Brodie A: Aromatase and its inhibitors- An overview. J Steroid Biochem Mol Biol 40: 255–261, 1991

    Google Scholar 

  6. Santen RI: Clinical use of aromatase inhibitors in human breast carcinoma. J Steroid Biochem Mol Biol 40: 247–253, 1991

    Google Scholar 

  7. Zacharewski T, Harris M, Biegel HL, Morrison V, Merchant M, Safe S: 6-Methyl-1,3,8-trichlorodibenzofuran (MCDF) as an antiestrogen in human and rodent cancer cell lines: Evidence for the role of the Ah receptor. Toxicol Pharmacol 113: 311–318, 1992

    Google Scholar 

  8. Manni A: Polyamines and hormonal control of breast cancer growth. CRC Critical Reviews Oncogenesis 1: 163–174, 1989

    Google Scholar 

  9. Lima G, Shiu RPC: Role of polyamines in estradiol-induced growth of human breast cancer cells. Cancer Res 45: 2466–2470, 1985

    Google Scholar 

  10. Thomas T, Trend B, Butterfield JB, Jänne OA, Kiang DT: Regulation of ornithine decarboxylase gene expression in MCF-7 breast cancer cells by antiestrogens. Cancer Res 49: 5852–5857, 1989

    Google Scholar 

  11. Tabor CW, Tabor H: Polyamines. Annu Rev Biochem 53: 749–790, 1984

    Google Scholar 

  12. Pegg AE: Recent advances in the biochemistry of polyamines in eukaryotes. Biochem J 234: 249–262, 1986

    Google Scholar 

  13. Chanda R, Ganguly AK: Polyamines in relation to human breast, rectal, and squamous cell carcinoma. Cancer Lett 39: 311–318, 1988

    Google Scholar 

  14. Manni A, Wright C: Reversal of the antiproliferative effect of the antiestrogen tamoxifen by polyamines in breast cancer cells. Endocrinology 114: 836–839, 1984

    Google Scholar 

  15. Thomas T, Kiang DT: Additive growth-inhibitory effects of DL-α-difluoromethylornithine and antiestrogens on the MCF-7 breast cancer cell line. Biochem Biophys Res Commun 148: 1338–1345, 1987

    Google Scholar 

  16. Hoggard N, Green D: Polyamines and growth regulation of cultured human breast cancer cells by 17β-estradiol. Mol Cell Endocrinol 46: 71–78, 1986

    Google Scholar 

  17. Cohen FJ, Manni A, Glikman P, Bartholomew M, Demers L: Involvement of the polyamine pathway in antiestrogeninduced growth inhibition of human breast cancer. Cancer Res 48: 6819–6825, 1988

    Google Scholar 

  18. Cohen S, O'Malley BW, Stastny M: Estrogenic induction of ornithine decarboxylasein vivo and andin vitro. Science 170: 336–338, 1970

    Google Scholar 

  19. Rorke EA, Katzenellenbogen BS: Dissociated regulation of growth and ornithine decarboxylase activity by estrogen in the rat uterus. Biochem Biophys Res Commun 122: 1186–1193, 1984

    Google Scholar 

  20. Abrahamsen MS, Morris DR: Cell type specific mechanisms of regulating expression of ornithine decarboxylase gene after growth regulation. Mol Cell Biol 10: 5525–5528, 1990

    Google Scholar 

  21. White MW, Kameji T, Pegg AE, Morris DR: Increased efficiency of translation of ornithine decarboxylase mRNA in mitogen activated lymphocytes. Eur J Biochem 170: 87–92, 1987

    Google Scholar 

  22. Persson L, Holm I, Heby O: Translational regulation of ornithine decarboxylase by polyamines. FEBS Lett 205: 175–178 1988

    Google Scholar 

  23. Kendra KL, Katzenellenbogen BS: An evaluation of the involvement of polyamines in modulating MCF-7 human breast cancer cell proliferation and progesterone receptor by estrogen and antiestrogen. J Steroid Biochem 28: 123–132, 1987

    Google Scholar 

  24. Kelly K, Cochran BH, Stiles CD, Leder P: Cell specific regulation of c-myc gene by lymphocyte mitogens and platelet derived growth factor. Cell 35: 603–609, 1983

    Google Scholar 

  25. Thomas T, Kiang DT: A 22-fold increase in the relative binding affinity of estrogen receptor for poly(dA-dC) poly(dG-dT) in the presence of polyamines. Nucleic Acids Res 16: 4705–4720, 1988

    Google Scholar 

  26. Thomas T, Kiang DT: Structural alterations and stabilization of rabbit uterine estrogen receptor by natural polyamines. Cancer Res 47: 1799–1804, 1987

    Google Scholar 

  27. Thomas T, Kiang DT: Modulation of the binding of progesterone receptor to DNA by polyamines. Cancer Res 48: 1217–1222, 1988

    Google Scholar 

  28. Evans RM: The steroid and thyroid hormone receptor family. Science 240: 889–895, 1988

    Google Scholar 

  29. Wahli W, Martinez E: Superfamily of nuclear receptors: positive and negative regulators of gene expression. FASEB J 5: 2243–2249, 1991

    Google Scholar 

  30. O'Malley BW: The steroid receptor superfamily: More excitement predicted for the future. Mol Endocrinol 4: 363–369, 1990

    Google Scholar 

  31. Vedeckis WE: Nuclear receptors, transcriptional regulation and oncogenesis. Proc Soc Exptl Biol Med 199: 1–12, 1992

    Google Scholar 

  32. Thomas T: Structural specificity of polyamines in facilitating the high affinity binding of estrogen receptor to specific DNA sequences. FASEB J 3: A1192, 1989

    Google Scholar 

  33. Thomas TJ, Bloomfield VA: Ionic and structural effects on the thermal helix-coil transition of DNA complexed with natural and synthetic polyamines. Biopolymers 23: 1295–1306, 1984

    Google Scholar 

  34. Jorstad CM, Harada JJ, Morris DR: Structural specificity of the spermidine requirement of anEscherichia coli auxotroph. J Bacteriol 141: 456–463, 1980

    Google Scholar 

  35. Berthois Y, Katzenellenbogen JA, Katzenellenbogen BS: Phenol red in tissue culture media is a weak estrogen: implication concerning the study of estrogen-responsive cells in culture. Proc Natl Acad Sci USA 83: 2496–2500, 1986

    Google Scholar 

  36. Kubota M, Kamatami N, Carson DA: Biochemical and genetic analysis of the role of methylthio-adenosine phosphorylase in a murine cell line. J Biol Chem 258: 7288–7291, 1983

    Google Scholar 

  37. Kasid A, Davidson NE, Gelman EP, Lippman ME: Transcriptional control of thymidine kinase gene expression by estrogen and antiestrogen in MCF-7 human breast cancer cells. J Biol Chem 261: 5562–5567, 1986

    Google Scholar 

  38. Osborne CK, Boldt DH, Clark GM, Trent JM: Effects of tamoxifen on human breast cancer cell cycle kinetics. Cancer Res 43: 3583–3585, 1983

    Google Scholar 

  39. Maniatis T, Fritsch EF, Sambrook J: Molecular Cloning: A Laboratory Manual. Cold Spring Harbor Laboratory, 1982, pp 202–203

  40. Thomas T, Chao CC, Thomas TJ: Polyamine requirements in the expression of c-fos and cyclin B mRNAs in MCF-7 breast cancer cells. Proc Amer Assoc Cancer Res 33: 24, 1992

    Google Scholar 

  41. Hickok NJ, Seppanen PJ, Gunsalus GL, Jänne OA: Complete amino acid sequence of human ornithine decarboxylase deduced from complementary DNA. DNA 6: 179–187, 1987

    Google Scholar 

  42. Lowry OH, Rosebrough NJ, Farr AL, Randall RJ: Protein measurement with Folin phenol reagent. J Biol Chem 193: 265–275, 1951

    Google Scholar 

  43. Kabra PM, Lee HK, Lubich WP, Marton LJ: Solid phase extraction and determination of dansyl derivatives of unconjugated and acetylated polyamines. J Chromatogr Biomed Appl 380: 19–32, 1986

    Google Scholar 

  44. Burton K: A study of the conditions and mechanism of the diphenylamine reaction for the colorimetric estimation of deoxyribonucleic acid. Biochem J 62: 315–323, 1956

    Google Scholar 

  45. Singh AB, Thomas TJ, Thomas T, Singh M, Mann RA: Differential effects of polyamine homologs on the prevention of DL-α-difluoromethylornithine-mediated inhibition of malignant cell growth and normal immune response. Cancer Res 52: 1840–1847, 1992

    Google Scholar 

  46. Porter CW, Bergeron RJ: Spermidine requirement for cell proliferation in eukaryotic cells: structural specificity and quantitation. Science 219: 1083–1085, 1983

    Google Scholar 

  47. Thomas T, Thomas TJ: Regulation of cyclin B1 by estradiol and polyamines in MCF-7 breast cancer cells. Cancer Res. In Press

  48. Dubik D, Dembinski TC, Shiu RPC: Stimulation of c-myc oncogene expression associated with estrogen-induced proliferation of human breast cancer cells. Cancer Res 47: 6517–6521, 1987

    Google Scholar 

  49. Wilding G, Lippman ME, Gelman EP: Effects of steroid hormones and polypeptide growth factors on proto-oncogene c-fos expression in human breast cancer cells. Cancer Res 48: 802–805, 1988

    Google Scholar 

  50. Davis RH, Morris DR, Coffino P: Sequestered end products and enzyme regulation: The case of ornithine decarboxylase. Microbiol Reviews 56: 280–290, 1992

    Google Scholar 

  51. Murakami Y, Matsufuji S, Miyazaki Y, Hayashi S: Destabilization of ornithine decarboxylase by transfected antizyme gene expression in hepatoma tissue culture cells. J Biol Chem 267: 13138–13141, 1992

    Google Scholar 

  52. Thomas T, Kiang DT, Jänne OA, Thomas TJ: Variations in amplification and expression of the ornithine decarboxylase gene in human breast cancer cells. Breast Cancer Res Treat 19: 257–267, 1991

    Google Scholar 

  53. Russell DH, Stambrook PJ: Cell cycle specific fluctuations in adenosine 3′,5′cyclic monophosphate and polyamines of Chinese hamster cells. Proc Natl Acad Sci USA 72: 1482–1486 1975

    Google Scholar 

  54. Friedman SJ, Bellantone RA, Canellakis ES: Ornithine decarboxylase activity in synchronously growing DON C cells. Biochem Biophys Acta 261: 188–193, 1972

    Google Scholar 

  55. Goldstein GA, Heby O, Marton LJ: Biphasic stimulation of polyamine biosynthesis in primary mouse kidney cells by infection with polyoma virus: uncoupling from DNA and rRNA synthesis. Proc Natl Acad Sci USA 73: 4022–4028, 1976

    Google Scholar 

  56. Sunkara PS, Ramakrisna S, Nishioka K, Rao PN: Relationship between levels and rates of synthesis of polyamines during mammalian cell cycle. Life Sciences 29: 1497–1506, 1981

    Google Scholar 

  57. Martin RL, Ilett KF, Minchin R: Cell cycle-dependent uptake of putrescine and its importance in regulating cell cycle phase transition in cultured adult mouse hepatocytes. Hepatology 14: 1243–1250, 1991

    Google Scholar 

  58. Halmekyto M, Hyttinen J-M, Sinervirta R, Utrianen M, Myohanen S, Volpio H-M, Wahlfors J, Syrjanen S, Syrjanen K, Alhonen L, Jänne J: Transgenic mice aberrantly expressing human ornithine decarboxylase gene. J Biol Chem 266: 19746–19751, 1991

    Google Scholar 

  59. Porter CW, Cavanaugh PF, Stolowich N, Ganis B, Kelly E, Bergeron R: Biological properties of N4 and N1,N8-spermidine derivatives in cultured L1210 leukemia cells. Cancer Res 45: 2950–2957, 1985

    Google Scholar 

  60. Bernacki RJ, Bergeron RJ, Porter CW: Antitumor activity of N,N′-bis(ethyl)spermine homologues against MALME-3 Melanoma xenografts. Cancer Res 52: 2424–2430, 1992

    Google Scholar 

  61. Basu HS, Pellarin M, Feuerstein BG, Deen F, Marton LJ: Effect of N1,N14-bis(ethyl)homospermine on the growth of U-87 MG and SF-126 human brain tumor cells. Cancer Res 50: 3137–3140 1990

    Google Scholar 

  62. Basu HK, Pellarin M, Feuerstein BG, Deen DF, Marton J: Effect of N1,N14-bis-(ethyl)-homospermine (BE-444) on the growth of U251MG and SF-188 human brain tumor cells. Int J Cancer 48: 873–878, 1991

    Google Scholar 

  63. Porter CW, Miller J, Bergeron RJ: Aliphatic chain length specificity of the polyamine transport system in ascites L1210 Leukemia cells. Cancer Res 44: 126–128, 1984

    Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Environmental and Community Medicine, Environmental and Occupational Health Sciences Institute, University of Medicine and Dentistry of New Jersey-Robert Wood Johnson Medical School, 08854, Piscataway, NJ, USA

    Thresia Thomas

  2. Program in Clinical Pharmacology, Clinical Research Center, University of Medicine and Dentistry of New Jersey-Robert Wood Johnson Medical School, 08854, Piscataway, NJ, USA

    T. J. Thomas

Authors
  1. Thresia Thomas
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. T. J. Thomas
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Thomas, T., Thomas, T.J. Estradiol control of ornithine decarboxylase mRNA, enzyme activity, and polyamine levels in MCF-7 breast cancer cells: therapeutic implications. Breast Cancer Res Tr 29, 189–201 (1994). https://doi.org/10.1007/BF00665680

Download citation

  • Issue Date:

  • DOI: https://doi.org/10.1007/BF00665680

Key words

Search

Navigation