An Estrogen-Independent MCF-7 Breast Cancer Cell Line Which Contains a Novel 80-Kilodalton Estrogen Receptor-related Protein

Long-term growth of estrogen-responsive human breast cancer cell lines in estrogen-free media leads inevitably to the development of estrogen-independent growth. We have identified and characterized a unique subclone of the MCF-7 human breast cancer cell line, named MCF-7:2A, which grows maximally in the absence of endogenous estrogens but whose growth is inhibited by the antiestrogens 4-hydroxytamoxifen and ICI 164,84. The MCF-7:2A cells express high levels of estrogen receptor (ER; 477 fmol/mg protein), which can be reduced by growth in 10 nM 17β-estradiol (201 fmol/mg protein). Basal progesterone receptor synthesis is very low in the 2A cells (<1 fmol/mg protein) but can be dramatically increased by 10 nM 17β-estradiol (384 fmol/mg protein). Clearly, the pathways that control growth and estrogen-regulated genes such as the progesterone receptor are now dissociated in these cells. MCF-7:2A cells also possess two unique characteristics. First, the MCF-7:2A cells consti-tutively activate an ER-responsive luciferase reporter construct in the absence of any estrogens, and this activation can be blocked by either 4-hydroxytamoxifen or ICI 164384. This constitutive activity is not observed in the parental MCF-7 cells. Second, they express an 80-kDa protein that cross-reacts with three distinct antibodies to the ER. The MCF-7:2A cells were subjected to an additional round of limiting dilution subcloning, and 10 independent clones were all shown to express both the 66- and 80-kDa ERs as observed in the MCF-7:2A line. This confirms that both ERs are being expressed in each cell and are not the result of a mixed population of cells. While numerous ER variants have been reported previously, no ER has until now been described that is larger than the wild-type 66-kDa ER The MCF-7:2A cells provide a unique model to use in the study of ER action and the development of estrogen-independent growth in human breast cancer cells.