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C‐ myc gene expression alone is sufficient to confer resistance to antiestrogen in human breast cancer cells
Author(s) -
Venditti Marcello,
Iwasiow Barbara,
Orr F. William,
Shiu Robert P.C.
Publication year - 2002
Publication title -
international journal of cancer
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.475
H-Index - 234
eISSN - 1097-0215
pISSN - 0020-7136
DOI - 10.1002/ijc.10269
Subject(s) - doxycycline , biology , microbiology and biotechnology , gene expression , transfection , antiestrogen , reporter gene , cancer research , mcf 7 , activator (genetics) , cell growth , cell culture , messenger rna , growth inhibition , cancer cell , estrogen receptor , gene , breast cancer , cancer , genetics , human breast , antibiotics
C‐ myc is implicated in the initiation, progression and estrogen response of breast cancer. To further investigate the role of c‐ myc in breast cancer, we have developed clonal MCF‐7 human breast cancer cell lines harboring a stably‐transfected human c‐ myc gene, whose expression was stringently controlled by the bacterial reverse tetracycline transcription activator protein. The expression of the endogenous genomic c‐ myc gene in MCF‐7 cells was abolished by the potent pure estrogen antagonist, ICI 182,780. Functional c‐Myc protein was identified by both Western immunoblotting and by its ability to transactivate a chimeric plasmid consisting of E‐box sequences upstream of the luciferase reporter gene. One MCF‐7 clone, 35im, was chosen for further characterization. C‐ myc induction by doxycycline was rapid and dose dependent; c‐ myc mRNA appeared as early as 30 min after doxycycline addition and stimulation of c‐ myc expression required as little as 50 ng/ml doxycycline, with c‐ myc mRNA levels reaching a plateau at 2.5 μg/ml doxycycline. ICI 182,780 or doxycycline (a tetracycline analog) treatment did not alter the mRNA levels of Max, the c‐ myc binding partner. As in wildtype MCF‐7 cells, the growth of clone 35im was inhibited by 1 μM or less of ICI 182,780 and stimulated by 10 nM to 1 μM 17β‐estradiol. When maintained in a complete medium containing 5% normal fetal bovine serum (FBS) and ICI 182,780, doxycycline induced cell growth by 400% in an 8‐day assay. A similar level of growth was achieved with doxycycline treatment in cells that were arrested by the use of charcoal‐stripped FBS. Doxycycline had no effect on the growth of a control MCF‐7 clone (18c). Apoptosis, assessed by caspase‐dependent cleavage of poly(ADP‐ribose) polymerase, was unchanged in clone 35im cells after treatments with doxycycline or ICI 182,780. The present study demonstrates that c‐ myc alone is sufficient to confer antiestrogen resistance in human breast cancer. Our novel c‐ myc ‐inducible MCF‐7 cell model offers a unique opportunity to study the diverse actions of the c‐ myc proto‐oncogene in human breast cancer. © 2002 Wiley‐Liss, Inc.