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Activation of ErbB3, EGFR and Erk is essential for growth of human breast cancer cell lines with acquired resistance to fulvestrant

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Abstract

Seven fulvestrant resistant cell lines derived from the estrogen receptor α positive MCF-7 human breast cancer cell line were used to investigate the importance of epidermal growth factor receptor (ErbB1-4) signaling. We found an increase in mRNA expression of EGFR and the ErbB3/ErbB4 ligand heregulin2 (hrg2) and a decrease of ErbB4 in all resistant cell lines. Western analyses confirmed the upregulation of EGFR and hrg2 and the downregulation of ErbB4. Elevated activation of EGFR and ErbB3 was seen in all resistant cell lines and the ErbB3 activation occurred by an autocrine mechanism. ErbB4 activation was observed only in the parental MCF-7 cells. The downstream kinases pAkt and pErk were increased in five of seven and in all seven resistant cell lines, respectively. Treatment with the EGFR inhibitor gefitinib preferentially inhibited growth and reduced the S phase fraction in the resistant cell lines concomitant with inhibition of Erk and unaltered Akt activation. In concert, inhibition of Erk with U0126 preferentially reduced growth of resistant cell lines. Treatment with ErbB3 neutralizing antibodies inhibited ErbB3 activation and resulted in a modest but statistically significant growth inhibition of two resistant cell lines. These data indicate that ligand activated ErbB3 and EGFR, and Erk signaling play important roles in fulvestrant resistant cell growth. Furthermore, the decreased level of ErbB4 in resistant cells may facilitate heterodimerization of ErbB3 with EGFR and ErbB2. Our data support that a concerted action against EGFR, ErbB2 and ErbB3 may be required to obtain complete growth suppression of fulvestrant resistant cells.

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Acknowledgements

We thank Inger Heiberg, Ole Nielsen, Hanife Dzaferi and Dorrit Lützhøft for excellent technical assistance. Also, thanks to Prof. William Gullick for the antibodies to hrg2α and hrg2β. We appreciate the financial support from; Danish Cancer Society, The Danish Medical Research Council, The Danish Cancer Research Foundation, Købmand Bernhard Rasmussen og hustru Meta Rasmussens Mindelegat, Else og Aage Grønbeck-Olsens Legat, Grosserer A.V. Lykfeldt og hustrus Legat, Fru Astrid Thaysens Legat for Lægevidenskabelig Grundforskning.

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Authors and Affiliations

  1. Department of Tumor Endocrinology, Institute of Cancer Biology, Danish Cancer Society, Strandboulevarden 49, 2100, Copenhagen, Denmark

    Thomas Frogne, Rikke V. Benjaminsen, Katrine Sonne-Hansen, Louise M. Rasmussen, Jan Stenvang & Anne E. Lykkesfeldt

  2. Department of Clinical Biochemistry, Aarhus University Hospital, Aarhus, Denmark

    Boe S. Sorensen & Ebba Nexo

  3. Department of Pathology, Odense University Hospital, Odense, Denmark

    Anne-Vibeke Laenkholm

  4. Department of Medicinal Chemistry and Molecular Pharmacology, Purdue University and Purdue Cancer Center, West Lafayette, IN, USA

    David J. Riese II

  5. Department of Tumor Biology, Institute Curie, Paris, France

    Patricia de Cremoux

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  1. Thomas Frogne
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Correspondence to Anne E. Lykkesfeldt.

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Frogne, T., Benjaminsen, R.V., Sonne-Hansen, K. et al. Activation of ErbB3, EGFR and Erk is essential for growth of human breast cancer cell lines with acquired resistance to fulvestrant. Breast Cancer Res Treat 114, 263–275 (2009). https://doi.org/10.1007/s10549-008-0011-8

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