4-fluoroestradiol and 2-fluoroestradiol

Fluorination preventing metabolic hydroxylation of 17β-estradiol (E

Topics: Animals; Breast Neoplasms; Cell Transformation, Neoplastic; Estradiol; Female; Organ Size; Rats, Inbred ACI; Risk Assessment; Uterus

17 beta-Estradiol is known to induce kidney tumors in male Syrian hamsters when administered chronically, whereas 4-fluoroestradiol does so only after an extended induction period and 2-fluoroestradiol is not carcinogenic; both fluorinated analogs are hormonally active. Because C-4 and C-2 hydroxylations are, respectively, minor and major routes of estrogen metabolism in vivo, these observations suggest mediation of tumorigenesis by catecholestrogen metabolites. However, the analogs were reported to undergo oxidative defluorination in vitro. We have determined the metabolic fates of estradiol, 2-fluoroestradiol, and 4-fluoroestradiol in male hamsters. [6,7-3H]Estradiol was principally C-2 hydroxylated when given intravenously at either 0.1 mumol/kg or 50 mumol/kg; 2-hydroxyestradiol was eliminated in bile and urine, largely as a glucuronide, without undergoing extensive deactivation via O-methylation. Alicyclic alcohol metabolites were minor products. [6,7-3H]2-Fluoroestradiol underwent either glucuronylation or sequential dehydrogenation and alicyclic hydroxylation followed by glucuronylation but neither oxidative defluorination nor compensatory C-4 hydroxylation. [6,7-3H]4-Fluoroestradiol was also considerably dehydrogenated to the keto form and glucuronylated. Nevertheless, only 4-fluoroestradiol yielded appreciable quantities of C-2 hydroxylated metabolite at the lower dose; methylation was an insignificant pathway. No defluorinated products were observed. Dehydrogenation of both analogs and alicyclic hydroxylation of the 2-fluoroestrone metabolite were less extensive at the higher dose; all of the polar metabolites of 2-fluoroestradiol in bile, although not those in urine, declined to trace amounts. C-2 hydroxylation of 4-fluoroestradiol was greater at this dose. Thus, the rank order of catechol formation from estradiol and its fluoro analogs observed in vivo, unlike that found in microsomal incubations, was consistent with the hypothesis that catechols mediate estrogen-dependent renal carcinogenesis in hamsters.

Topics: Animals; Bile; Carcinogens; Cricetinae; Estradiol; Estrogens, Catechol; Hydroxylation; Kidney Neoplasms; Male; Mesocricetus; Tritium

In male Syrian hamsters, an animal model for estradiol-induced carcinogenesis, 2-fluoroestradiol was not carcinogenic, whereas 4-fluoroestradiol induced kidney tumors after a prolonged latency period, compared with estradiol (100% tumor incidence), when the compounds were administered to hamsters in hormonally equipotent doses. Catechol estrogen metabolites have previously been postulated to mediate this estrogen-induced kidney carcinogenesis. To examine this proposed mechanism of tumor induction by estrogens, we investigated the conversion of 2- and 4-fluoroestradiol to catechol metabolites by kidney and liver microsomes of hamsters and the further conversion to methyl ethers by catechol-O-methyltransferase, and we compared the values with those obtained with nonfluorinated estrogens as substrates. The rates of conversion of 2-fluoroestradiol to 2-hydroxyestradiol and 2-fluoro-4-hydroxyestradiol by hepatic microsomes were 30-50% lower than corresponding rates with estradiol as substrate. With renal microsomes the rate of 4-hydroxylation was 10 times faster than that of estradiol, whereas 2-hydroxylation was at best marginal. With 4-fluoroestradiol as substrate the rate of 2-hydroxylation by hepatic microsomes was enhanced 5-fold, compared with values for estradiol, but 4-hydroxyestradiol formation was almost eliminated. In contrast, the conversion of this substrate to 4-fluoro-2-hydroxyestradiol by kidney microsomes occurred at a rate 15 times faster than 2-hydroxylation of estradiol, whereas 4-hydroxyestradiol formation proceeded at a rate of 315 pmol/mg of protein/min. Except for the decrease in both 2- and 4-hydroxylation of 2-fluoroestradiol by liver microsomes, fluorine substitution of estrogenic phenols enhanced microsome-mediated aromatic hydroxylation at sites unoccupied by substituents. At pH 7.5, the highest rates of catechol-O-methyltransferase-mediated methylation were observed with the catechol metabolites of 2-fluoroestradiol, 2-fluoro-4-hydroxyestradiol and 2-hydroxyestradiol (3780 and 2960 pmol/mg of protein/min, respectively). Lower rates were found with those of 4-fluoroestradiol, 4-fluoro-2-hydroxyestradiol and 4-hydroxyestradiol (1670 and 470 pmol/mg of protein/min, respectively). These data are consistent with the postulate that catechol metabolites of estrogens are reactive intermediates in estrogen-induced carcinogenesis. For the noncarcinogenic 2-fluoroestradiol, a high metabolic flux was observed through a pathway of renal 2-fluo

Topics: Animals; Carcinogens; Catechol O-Methyltransferase; Cricetinae; DNA; Estradiol; Estrogens, Catechol; Hydroxylation; In Vitro Techniques; Kidney; Male; Mesocricetus; Methylation; Microsomes; Microsomes, Liver

2-Fluoro-[6,7-3H]17 beta-oestradiol([3H]2-FE2) and 4-fluoro-[6,7-3H]17 beta-oestradiol([3H]4-FE2) were synthesized by the fluorination and reduction of [3H]oestrone and purified by HPLC. [3H]2-FE2 and [3H]4-FE2 (72.5 micrograms/kg; 0.25 mumol/kg) were administered i.v. to anaesthetized female and male Wistar rats (N = 4) with biliary cannulae. Bile was collected for 6 hr. Female rats administered [3H]2-FE2 excreted 85% of the dose into bile over 6 hr whilst male rats excreted 77%. After the administration of [3H]4-FE2, female and male rats excreted 72 and 83% of dose into bile over 6 hr, respectively. The biliary metabolites were glucuronides in all cases. The principal metabolite of [3H]2-FE2 liberated from biliary conjugates by beta-glucuronidase was 2-fluoroestrone in both female rats (64% of dose) and male rats (57%). No 2-hydroxylated, i.e. oxidatively defluorinated, metabolites were detected in either sex. In contrast, 2-hydroxylation of [3H]4-FE2 did occur, but only in female rats: 2-hydroxy-4-fluoro-oestrone (22%) and 2-methoxy-4-fluoroestrone (17%) were identified as biliary aglycones. However, the major metabolite was 4-fluoroestrone (4FE1; 38%). In male rats, 4-FE1 and 4-fluoro D-ring-oxygenated products were the principal biliary aglycones. The differences in metabolism between the two fluoro analogues and oestradiol are discussed with particular reference to the possible involvement of 2- and 4-hydroxy (catechol) oestrogens in oestrogen toxicity.

Topics: Animals; Bile; Biotransformation; Chromatography, High Pressure Liquid; Estradiol; Female; Glucuronates; Inactivation, Metabolic; Male; Mass Spectrometry; Rats; Rats, Inbred Strains

Catechol estradiol can induce implantation of the embryo in a progesterone-primed uterus, but we do not know whether conversion of estrogen to a catechol is essential for implantation. The present study examined the ability of fluorinated estradiols that have a reduced capability of catechol formation to induce implantation. Delayed implantation in rats that were hypophysectomized on the third day postcoitum was maintained by daily injection of progesterone. On the fifth day of progesterone treatment they were injected intravenously with estradiol-17 beta (E2), or various doses of 2-fluoroestradiol-17 beta (2-F1-E2) or 4-fluoroestradiol-17 beta (4-F1-E2) and examined 24 hr later for evidence of initiation of implantation. All animals treated with 25 ng of E2 showed normal numbers of implantation sites, as did those receiving 60 ng of 4-F1-E2. In contrast, 2-F1-E2 failed to initiate implantation with doses as high as 300 ng per animal; there were only single sites in two of eight rats treated with 500 ng. Initiation of implantation was not correlated with lack of uterotropic estrogenicity. The results suggest that formation of catechol estrogen may be an important step in mediating estrogen function for implantation of the embryo.

Topics: Animals; Embryo Implantation; Embryo Implantation, Delayed; Estradiol; Estrogens, Catechol; Female; Organ Size; Pregnancy; Rats; Uterus

In an attempt to define pharmacological probes with which to test the role of catechol oestrogen formation in the central nervous system, five oestrogens (oestradiol-17 beta, oestradiol-17 alpha, 4-fluoro-oestradiol, 2-fluoro-oestradiol and moxestrol (11 beta-methoxy-17 alpha-ethynyloestradiol) were studied for binding to oestrogen receptors and conversion to catechol metabolites. Binding to cytosol oestrogen receptors was measured in the hypothalamus-preoptic area-amygdala (HPA), pituitary gland and uterus of ovariectomized rats. Conversion to catechol oestrogens was tested in microsomes from the HPA, pituitary gland and liver, using a catechol-O-methyltransferase-coupled radioenzymatic assay. Oestradiol-17 alpha was the only weak oestrogen receptor ligand. Binding affinities of the other compounds tested were much higher and comparable to those of oestradiol-17 beta. In contrast, oestradiol-17 alpha was rapidly converted to catechol metabolites, while moxestrol was a relatively poor substrate for catechol oestrogen formation. 4-Fluoro-oestradiol could be 2-hydroxylated but not 4-hydroxylated. 2-Fluoro-oestradiol exhibited impaired 2-hydroxylation but normal 4-hydroxylation.

Topics: Animals; Central Nervous System; Chromatography, High Pressure Liquid; Cytosol; Estradiol; Estrogens, Catechol; Ethinyl Estradiol; Female; Hydroxylation; Ovariectomy; Radioligand Assay; Rats; Rats, Inbred Strains; Receptors, Estrogen

The role of catechol oestrogen formation in the mechanism by which circulating oestrogens facilitate gonadotrophin release and female sexual behaviour was explored in adult female rats. The effects of oestradiol-17 beta were compared with those of a group of oestrogens with either a reduced affinity for oestrogen receptors (oestradiol-17 alpha) or a reduced ability to act as substrates for catechol oestrogen formation (2-fluoro-oestradiol, 4-fluoro-oestradiol and moxestrol (11 beta-methoxy-17 alpha-ethynyloestradiol]. Rats were ovariectomized on the evening of dioestrus day 1 of the 4-day oestrous cycle and implanted s.c. 12 h later with infusion pumps containing either one of the test oestrogens or vehicle alone. Infusion rates for oestradiol-17 beta, moxestrol, 2-fluoro-oestradiol and 4-fluoro-oestradiol were adjusted to give concentrations of nuclear oestrogen receptors in the brain and pituitary gland within the range of those found in intact female rats during pro-oestrus. Oestradiol-17 alpha was infused at the same and at a tenfold higher rate than that of oestradiol-17 beta; neither of these treatments with oestradiol-17 alpha significantly increased brain or pituitary gland nuclear oestrogen receptor levels. On the day after the pump was implanted, samples of tail vein blood were withdrawn at 12.00, 14.00, 16.00 and 18.00 h for LH assay. All animals were then injected s.c. with 1 mg progesterone in propylene glycol, and tested for feminine sexual behaviour 5 h later. Oestradiol-17 beta, moxestrol, 2-fluoro-oestradiol and 4-fluoro-oestradiol all elicited pronounced LH surges and facilitated progesterone-triggered proceptive and lordosis behaviours.(ABSTRACT TRUNCATED AT 250 WORDS)

Topics: Animals; Central Nervous System; Estradiol; Estrogens; Estrogens, Catechol; Ethinyl Estradiol; Female; Luteinizing Hormone; Ovariectomy; Progesterone; Rats; Rats, Inbred Strains; Sexual Behavior, Animal