THE EFFECTS OF 2,3,7,8-TETRACHLORODIBENZO-P-DIOXIN ON ESTROGEN METABOLISM IN MCF-7 BREAST-CANCER CELLS - EVIDENCE FOR INDUCTION OF A NOVEL 17-BETA-ESTRADIOL 4-HYDROXYLASE

Rates of microsomal 17 beta-estradiol (E(2)) hydroxylation at the C-2, -4, -6 alpha, and -15 alpha positions are each induced greater than 10-fold by treating MCF-7 breast cancer cells with 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD). The TCDD-induced activities at the C-2, -6 alpha and -15 alpha positions have been attributed to cytochrome P450 1A1 (CYP1A1); however, the low K-m 4-hydroxylase induced by TCDD appears to be a distinct enzyme. We report here that antibodies to cytochrome P450-EF (mouse CYP1B1) selectivity inhibited the C-4 hydroxylation of E, catalyzed by microsomes from TCDD-treated MCF-7 cells. Western blots probed with anti-CYP1B antibodies showed the induction of a 52 kDa microsomal protein in response to treatment with TCDD in MCF-7 cells. Western blots of microsomes from HepG2 cells did not show the TCDD-induced 52 kDa protein, and microsomes from TCDD-treated HepG2 cells did not catalyze a low K-m hydroxylation of E(2) at C-4. Cellular metabolism experiments also showed induction of both the C-2 and -4 hydroxylation pathways in TCDD-treated MCF-7 cells as evidenced by elevated 2- and 4-methoxyestradiol (MeOE(2)) formation. In contrast, TCDD-treated HepG2 cells showed 2-MeOE(2) formation predominantly over 4-MeOE(2). Northern blots of RNA isolated from untreated and TCDD-treated cells, when probed with the human CYP1B1 cDNA, showed induction of a 5.2 kb RNA in MCF-7 cells but not in HepG2 cells in response to treatment with TCDD. These results provide additional evidence for the induction by TCDD of a novel E(2) 4-hydroxylase in MCF-7 cells but not in HepG2 cells and indicate possible endocrine regulatory roles for the newly discovered group of enzymes of the CYP1B subfamily.