ESTROGENIC ACTIVITY IS INCREASED FOR AN ANTIESTROGEN BY A NATURAL MUTATION OF THE ESTROGEN-RECEPTOR

The estrogen receptor (ER) functions as a ligand-activated transcripti on factor which mediates the actions of estrogens and antiestrogens in target tissues. Other investigators have shown that artificial point mutations in the transcriptional activation domain AF-2 of the ligand binding domain (LED) of the ER can increase the estrogenic properties of antiestrogens, determined by transcriptional activation of estrogen -responsive reporter constructs cotransfected into cells. Although the se data provide valuable information about ER function there is no evi dence that these mutations occur naturally. We have taken a different approach and examined the naturally occurring codon 351 asp --> tyr mu tation in the LED of ER to stimulate the expression of an endogenous t arget gene. This approach avoids dependence on artificial reporter con structs and their idealized estrogen response elements (EREs). In this report we describe the regulation of transforming growth factor alpha (TGF alpha) mRNA by estradiol and the antiestrogens keoxifene and ICI 182,780 in our stable transfectants of ER-negative MDA-MB-231 breast cancer cells, which express either the wild-type (S30 cells) or codon 351 asp --> tyr mutant ER (BC-2 cells). The mutant receptor was identi fied in a tamoxifen-stimulated human breast tumor. Our results demonst rate, for the first time, that a naturally occurring mutation in the E R changes the pharmacology of the antiestrogen keoxifene by increasing estrogenic activity, and that keoxifene exhibits a gene-specific estr ogen-like effect with mutant ER but not with wild-type ER. The pure an tiestrogen ICI 182,780 maintained complete antagonistic activities in both ER transfectants, demonstrating that its action is unaffected by the mutation. (C) 1997 Elsevier Science Ltd.