Breast Cancer Research and Treatment

, Volume 31, Issue 1, pp 129–138 | Cite as

The estrogen receptor from a tamoxifen stimulated MCF-7 tumor variant contains a point mutation in the ligand binding domain

  • Douglas M. Wolf
  • V. Craig Jordan


The nonsteroidal antiestrogen tamoxifen (TAM) is the most commonly used endocrine treatment for all stages of breast cancer in both pre- and postmenopausal women. However, the development of resistance to the drug is common, as most patients treated with TAM eventually experience a recurrence of tumor growth. One of the potential mechanisms of treatment failure is the acquisition by the tumor of the ability to respond to TAM as a stimulatory rather than inhibitory ligand. We (Gottardis and Jordan, Cancer Res 48: 5183-5187, 1988; Wolfet al., J Natl Cancer Inst 85: 806-812, 1993) and others (Osborneet al., Eur J Cancer Clin Oncol 23: 1189-1196, 1987; Osborneet al., J Natl Cancer Inst 83: 1477-1482, 1991) have extensively described the reproducible development of TAM stimulated growth in a laboratory model system using MCF-7 human breast cancer cells grown as solid tumors in athymic mice. In this paper we report on the isolation of an estrogen receptor (ER) from a TAM stimulated tumor (MCF-7/MT2) which contains a point mutation that causes a tyrosine for aspartate substitution at amino acid 351 in the ligand binding domain. The mutant appears to the major form of ER expressed by this tumor. We also report that only wild type ER was detected in three other TAM stimulated MCF-7 tumor variants, suggesting that multiple mechanisms are possible for the development of TAM stimulated growth. The implications of these findings are discussed.

Key words

breast cancer tamoxifen drug resistance estrogen receptor mutant 


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  1. 1.
    Osborne CK, Coronado EB, Robinson JP: Human breast cancer in the athymic nude mouse: cytostatic effects of longterm antiestrogen therapy. Eur J Cancer Clin Oncol 23: 1189–1196, 1987Google Scholar
  2. 2.
    Gottardis MM, Jordan VC: Development of tamoxifenstimulated growth of MCF-7 tumors in athymic mice after long-term antiestrogen administration. Cancer Res 48: 5183–5187, 1988Google Scholar
  3. 3.
    Osborne CK, Coronado E, Allred DC, Wiebe V, DeGregorio M: Acquired tamoxifen (TAM) resistance: correlation with reduced breast tumor levels of tamoxifen and isomerization of trans-4-hydroxytamoxifen. J Natl Cancer Inst 83: 1477–1482, 1991Google Scholar
  4. 4.
    Wolf DM, Langan-Fahey SM, Parker CP, McCague R, Jordan VC: Investigation of the mechanism of tamoxifen stimulated breast tumor growth with non-isomerizable analogs of tamoxifen and its metabolites. J Natl Cancer Inst 85: 806–812, 1993Google Scholar
  5. 5.
    Osborne CK, Wiebe VJ, McGuire WL, Ciocca DR, DeGregorio M: Tamoxifen and the isomers of 4-hydroxytamoxifen in tamoxifen-resistant tumors from breast cancer patients. J Clin Oncol 10: 304–310, 1992Google Scholar
  6. 6.
    Wiebe VJ, Osborne CK, McGuire WL, DeGregorio MW: Identification of estrogenic tamoxifen metabolite(s) in tamoxifen-resistant human breast tumors. J Clin Oncol 10: 990–994, 1992Google Scholar
  7. 7.
    Wolf DM, Jordan VC: Characterization of tamoxifen stimulated MCF-7 tumor variants grown in athymic mice. Breast Cancer Res Treat (this issue)Google Scholar
  8. 8.
    Vegeto E, Allan GF, Schrader WT, Tsai M-J, McDonnell DP, O'Malley BW: The mechanism of RU486 antagonism is dependent on the conformation of the carboxy-terminal tail of the human progesterone receptor. Cell 69: 703–713, 1992Google Scholar
  9. 9.
    Wilding G, Chen M, Gelman EP: Aberrant responsesin vitro of hormone-responsive prostate cancer cells to antiandrogens. Prostate 14: 103–115, 1989Google Scholar
  10. 10.
    Veldscholte J, Ris-Stalpers C, Kuiper GGJM, Jenster G, Berrevoets C, Classen C, van Rooij HCJ, Trapman J, Brinkmann AO, Mulder E: A mutation in the ligand binding domain of the androgen receptor of human LNCaP cells affects the steroid binding characteristics and response to antiandrogens. Biochem Biophys Res Commun 173: 534, 1990Google Scholar
  11. 11.
    Olea N, Sakabe K, Soto AM, Sonnenschein C: The proliferative effect of ‘anti-androgens’ on the androgen-sensitive human prostate tumor cell line LNCaP. Endocrinology 126: 1457–1463, 1990Google Scholar
  12. 12.
    Schuurmans ALG, Bolt J, Veldscholte J, Mulder E: Stimulatory effects of antiandrogens on LNCaP human prostate tumor cell growth, EGF-receptor level and acid phosphatase secretion. J Steroid Biochem Molec Biol 37: 849–853, 1990Google Scholar
  13. 13.
    Veldscholte J, Voorhorst-Ogink MM, Bolt-de Vries J, van Rooij HCJ, Trapman J, Mulder E: Unusual specifity of the androgen receptor in the human prostate tumor cell line LNCaP: high affinity for progestagenic and estrogenic steroids. Biochim Biophys Acta 1052: 187–194, 1990Google Scholar
  14. 14.
    Jiang S-Y, Jordan VC: Growth regulation of estrogen receptor-negative breast cancer cells transfected with complementary DNAs for estrogen receptor. J Natl Cancer Inst 84: 580–591, 1992Google Scholar
  15. 15.
    Jiang S-Y, Langan-Fahey SM, Stella AL, McCague R, Jordan VC: Point mutation of estrogen receptor (ER) in the ligand binding domain changes the pharmacology of antiestrogens in ER-negative breast cancer cells stably expressing cDNA's for ER. Mol Endocrinol 6: 2167–2174, 1992Google Scholar
  16. 16.
    Jiang S-Y, Parker CJ, Jordan VC: A model to describe how a point mutation of the estrogen receptor alters the structure function relationship of antiestrogens. Breast Cancer Res Treat 26: 139–148, 1993Google Scholar
  17. 17.
    Gottardis MM, Jiang SY, Jeng MH, Jordan VC: Inhibition of tamoxifen-stimulated growth of an MCF-7 tumor variant in athymic mice by novel steroidal antiestrogens. Cancer Res 49: 4090–4093, 1989Google Scholar
  18. 18.
    Wolf DM, Jordan VC: A laboratory model to explain the sustained survival advantage observed in patients taking adjuvant tamoxifen therapy. In: Senn HJ, Goldhirsch A, Gelber RD, Turlmann B (eds) Adjuvant Therapy of Primary Breast Cancer IV. Recent Results in Cancer Research. Springer-Verlag, Berlin, 1993, Vol. 127, pp 22–33Google Scholar
  19. 19.
    Wolf DM, Arakawa RL, Friedl A, Jordan VC: Estradiol induced regression of the tamoxifen stimulated tumor variant MCF-7 TAM after prolonged exposure to tamoxifenin vivo (in preparation)Google Scholar
  20. 20.
    Orita M, Suzuki Y, Sekiya T, Hayashi K: Rapid and sensitive detection of point mutations and DNA polymorphisms using the polymerase chain reaction. Genomics 5: 874–879, 1989Google Scholar
  21. 21.
    Orita M, Iwahana H, Kanazawa H, Hayashi K, Sekiya T: Detection of polymorphisms of human DNA by gel electrophoresis as single-strand conformation polymorphisms. Proc Natl Acad Sci USA 86: 2766–2770, 1989Google Scholar
  22. 22.
    Iwahana H, Yoshimoto K, Itakura M: Detection of point mutations by SSCP of PCR-amplified DNA after endonuclease digestion. BioTechniques 12: 64–65, 1992Google Scholar
  23. 23.
    Iino Y, Wolf DM, Langan-Fahey SM, Johnson DA, Ricchio M, Thompson ME, Jordan VC: Reversible control of oestradiol-stimulated growth of MCF-7 tumors by tamoxifen in the athymic mouse. Br J Cancer 64: 1019–1024, 1991Google Scholar
  24. 24.
    Chomczynski P, Sacchi N: Single-step method of RNA isolation by acid-guanidinium thiocyanate-phenol-chloroform extraction. Anal Biochem 162: 156–159, 1987Google Scholar
  25. 25.
    Green S, Walter P, Kumar V, Krust A, Bornert J-M, Argos P, Chambon P: Human oestrogen receptor cDNA; sequence, expression and homology to v-erbA. Nature 320: 134–139, 1986Google Scholar
  26. 26.
    Tora L, Mullick A, Metzger D, Ponglikitmongkol M, Park I, Chambon P: The cloned human estrogen receptor contains a mutation which alters its hormone binding properties. EMBO J 8: 1981–1986, 1989Google Scholar
  27. 27.
    Chung CT, Miller RH: A rapid and convenient method for the preparation and storage of competent bacterial cells. Nucleic Acids Res 16: 3580, 1988Google Scholar
  28. 28.
    Fuqua SAW, Fitzgerald SD, Allred DC, Elledge RM, Nawaz Z, McDonnell DP, O'Malley BW, Greene GL, McGuire WL: Inhibition of estrogen receptor action by a naturally occurring variant in human breast tumors. Cancer Res 52: 483–486, 1992Google Scholar
  29. 29.
    Cole MP, Jones CJA, Todd IDH: A new antioestrogenic agent in late breast cancer. Br J Cancer 25: 270–275, 1971Google Scholar
  30. 30.
    Tanaka M, Abe K, Ohnami S, Adachi I, Yamaguchi K, Miyakawa S: Tamoxifen in advanced breast cancer: response rate, effect on pituitary hormone reserve and binding affinity to estrogen receptor. Jpn J Clin Oncol 8: 141–148, 1978Google Scholar
  31. 31.
    Smith IE, Harris AL, Morgan M, Ford HT, Gazet J, Harmes CL, White H, Parsons CA, Villardo A, Walsh G, McKinna JA: Tamoxifenversus aminoglutethimide in advanced breast carcinoma: a randomized cross-over trial. Br Med J 283: 1432–1434, 1981Google Scholar
  32. 32.
    Ingle JN, Ahmann DL, Green SJ, Edmonson JH, Bisel HF, Kvols LK, Nichols WC, Greagan ET, Hahn RG, Rubin J, Frytak S: Randomized clinical trial of diethylstilbestrol versus tamoxifen in postmenopausal women with advanced breast cancer. N Engl J Med 304: 16–21, 1981Google Scholar
  33. 33.
    Early Breast Cancer Trialists Collaborative Group: Systemic treatment of early breast cancer by hormonal, cytotoxic, or immune therapy: 133 randomized trials involving 31000 recurrences and 24000 deaths among 75000 women. Lancet 339: 1–15, 71-85, 1992Google Scholar
  34. 34.
    Legault-Poisson S, Jolivet J, Poisson R, Beretta-Piccoli M, Band PR: Tamoxifen-induced tumor stimulation and withdrawal response. Cancer Treat Rep 63: 1839–1841, 1979Google Scholar
  35. 35.
    Canney PA, Griffiths T, Latief TN, Priestman TJ: Clinical significance of tamoxifen withdrawal response. Lancet i: 36, 1987Google Scholar
  36. 36.
    Belani CP, Pearl P, Whitley NO, Aisner J: Tamoxifen withdrawal response. Report of a case. Archives of Internal Medicine 149: 449–450, 1989Google Scholar
  37. 37.
    Howell A, Dodwell DJ, Anderson H, Redford J: Response after withdrawal of tamoxifen and progestogens in advanced breast cancer. Ann Oncol 3: 611–617, 1992Google Scholar
  38. 38.
    Walker KJ, Price-Thomas JM, Candlish W, Nicholson RI: Influence of the antioestrogen tamoxifen on normal breast tissue. Br J Cancer 64: 764–768, 1991Google Scholar
  39. 39.
    Fritsch M, Wolf DM: Symptomatic side effects of tamoxifen therapy. In: Jordan VC (ed) Long-Term Tamoxifen Treatment for Breast Cancer. University of Wisconsin Press, Madison (in press)Google Scholar

Copyright information

© Kluwer Academic Publishers 1994

Authors and Affiliations

  • Douglas M. Wolf
    • 1
  • V. Craig Jordan
    • 2
  1. 1.Department of Human OncologyUniversity of Wisconsin Comprehensive Cancer CenterMadisonUSA
  2. 2.Department of PharmacologyUniversity of Wisconsin Comprehensive Cancer CenterMadisonUSA

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