Arnold J. Eisenfeld

Androgen Accumulation and Binding to Macromolecules in Seminal Vesicles: Inhibition by Cyproterone

Cyproterone reduces the accumulation of testosterone and dihydrotestosterone in seminal vesicles 30 minutes after intravenous administration of tritiated testosterone to castrated rats. Testosterone, added in vitro, binds to macromolecules from the supernatant fraction of the seminal vesicle homogenates; this interaction is antagonized competitively by cyproterone. Cyproterone may diminish androgenic effects by competition for binding molecules.

Evidence for estradiol binding sites in the hypothalamus—Effect of drugs

Abstract One hr after i.v. injection, the concentration of 3 H-estradiol in hypothalamus, anterior pituitary, uterus, and vagina of ovariectomized rats was lowered by prior administration of diethylstilbestrol, clomiphene, dimethylstilbestrol, U11100, or norethindrone. The concentration of 3 H-estradiol in heart, cerebrum or plasma was little, if any, reduced by these drugs. Chlormadinone and medroxyprogesterone did not decrease the concentration of 3 H-estradiol in any organ. These data support the thesis that binding sites for estradiol are present in the hypothalamus and that interaction by specific drugs with the estradiol-binding site in the hypothalamus may be a mechanism for regulation of ovulation.

Estrogen receptors and androgen receptors in the mammalian liver

An estrogen receptor and an androgen receptor are present in the mammalian liver. In the liver of the rat, the estrogen receptor concentration increases markedly at puberty and this change correlates with enhanced estrogen stimulation of plasma renin substrate synthesis. High doses of estrogen are required for nuclear binding in liver when compared to doses for the uterus. The high dose requirement appears to be predominantly due to extensive metabolism in the hepatocyte of the estrogen to inactive derivatives. Furthermore, estradiol is much weaker than ethinyl estradiol for promoting nuclear binding in the liver. This is due to extremely rapid and extensive metabolism of estradiol. In human liver the concentration of estrogen receptor is low. An androgen receptor is present in high concentration in rabbit liver and is located predominantly in the nucleus after androgen administration. High concentrations of a putative androgen receptor are also present in human liver cytosol. Preliminary studies indicate that synthetic progestins can attach to the human liver androgen receptor. To date, a progesterone receptor has not been found in the mammalian liver. Thus, it appears that extensive steroid metabolism in liver preferentially diminishes sex steroid interaction with liver receptors and that androgen receptors may mediate progestin effects in liver. These observations provide a scientific basis for improved safety of oral contraceptives. Lowering the estrogen and progestin doses in oral contraceptives will decrease the major side-effects, which are liver mediated, and still maintain the desired effects at the hypothalamic-pituitary axis and uterus. Furthermore, it is likely that by selecting which estrogen, progestin or androgen is administered as well as by utilizing a parenteral route of administration that sex steroid effects on the liver could be minimized.

Specific macromolecular binding of estradiol in the mammalian liver supernatant

Abstract The liver supernatant of adult female rats contains proteins with a high specificity for binding radioactive estradiol and other potent estrogens. Several properties of the liver macromolecules are similar to those of the presumed estrogen receptor found in the uterus. Specific estrogen binding is also present in the liver supernatant of the rabbit and green monkey. Estrogens are known to influence liver composition and function in mammals. The estrogen binding protein observed in the mammalian liver may be an estrogen receptor.

Synthesis of estradiol fatty acid esters by human breast tumors: fatty acid composition and comparison to estrogen and progesterone receptor content

The estradiol-17 beta-fatty acid esters are non-polar metabolites of estradiol, formed in many tissues, including human breast tumors. It has been shown in the rat that the synthesis of these esters is greatest in those tissues that respond to estrogen stimulation. Thus the possibility was explored that the biosynthesis of the estradiol esters in human breast tumors occurs mainly in those tumors that are estrogen sensitive; and thus that the synthesis of this family of non-polar metabolites of estradiol could be used as an additional marker for the identification of hormonally dependent tumors. However, the conversion of estradiol to the esters did not correlate with other indicators of estrogen responsiveness, the progesterone or estrogen receptors. Interestingly, the composition of the fatty acids in the estradiol-17-esters synthesized in the human tumors was markedly different from those originally identified in the bovine uterus. In the bovine uterus, the esters were predominantly unsaturated, 85%, while in this study the saturated esters were the major component. Since after systemic administration the saturated estradiol-17-esters have been found to be much longer-lived than the unsaturated esters, the biosynthesis of the relatively high proportion of saturated esters by human breast tumors may indicate a significantly prolonged duration for the estrogenic signal produced by endogenously formed estradiol esters. These esters formed and sequestered within the tumor cell, may serve as a preformed store of estradiol, which after enzymatic hydrolysis, can locally stimulate growth of tumors that are estrogen responsive.

Oral contraceptives--possible mediation of side effects via an estrogen receptor in liver.

Most of the side effects of oral contraceptives may be caused by direct estrogen interaction in the liver resulting in alteration in liver function, including the synthesis of critical plasma proteins affecting the cardiovascular system. This study has succeeded in demonstrating an estrogen receptor in the mammalian liver. The liver receptors were found to consist, at least partly, of protein, and bound estrogens with high specificity. A strong binding affinity was indicated. The steroid hormones first bind to cytoplasmic receptors, after which the cytoplasmic receptor-steroid complex translocates from the cytoplasm to the nucleus and attaches to chromatin. Estrogen binding and translocation were shown in rat liver in vitro and in vivo. Minimizing the liver interaction thus might reduce the risk of side effects, but the estrogen-receptor function in the hypothalmic-pituitary axis and in the endometrium would have to be maintained. Possible differences in receptor characteristics of the 2 organs might be exploited to reduce liver interaction and, at the same time, possibly improve therapeutic benefits. Studies of receptor characteristics in rats indicate that a higher dose of estrogen is required in the liver than in the uterus and other organs to induce at least some direct effects. There seems to be a possibility that the newer combined oral contraceptives, which contain lower amounts of estrogen, ethinyl estradiol (reduced from 50 ug to 30 ug), and progestin, might produce smaller changes in estrogen-related plasma protein and at the same time achieve contraceptive effectiveness and regular menstrual bleeding patterns. Another possibility would be the future development or selection of another and safer estrogen.

Female and male green monkey liver estrogen receptor.

Abstract Adult female and male green monkey liver cytosol contains tritiated 17β-estradiol ([ 3 H]E 2 ) binding sites. The binding of [ 3 H]E 2 to these sites is reduced substantially by diethylstilbestrol. Unlike adult male rate liver cytosol, neither male nor female green monkey liver cytosol contains moderate affinity high capacity [ 3 H]E 2 binding sites. The green monkey liver cytosol [ 3 H]E 2 sites are precipitated by ammonium sulfate at 30% of saturation. The properties of the redissolved ammonium sulfate precipitated female and male [ 3 H]E 2 binding sites are similar: they have a high [ 3 H]E 2 affinity, a low [ 3 H]E 2 capacity, are estrogen specific, and are sensitive to sulfhydryl inactivation and protease digestion. Although the female and male sites are similar, there appears to be a 2- to 3-fold sex difference in the affinity (female K d , 0.8 to 1.3 × 10 −10 M; male K d , 0.4 × 10 −10 M) and capacity (female, 2.3 to 3.7 fmoles/mg of liver; male, 1.2 fmoles/mg of liver)of the [ 3 H]E 2 binding sites. Accordingly, adult female and male green monkey liver cytosol contains putative estradiol receptors.

Preservation of steroid receptors in frozen brain and pituitary tissue: use of the cryoprotective agent, dimethylsulfoxide.

This paper examines the effects of freezing and thawing on steroid receptor concentrations in the brain and pituitary of the rat. Storage at -70 degrees C for 1-2 weeks had no detectable effect on levels of cytoplasmic estrogen receptors. However, freezing and thawing resulted in measurable losses of cytoplasmic androgen, progestin and glucocorticoid receptors. Cell nuclear receptors were measured by exchange assay after in vivo administration of non-radioactive steroids. Nuclear estrogen, androgen and progestin receptor concentrations were all reduced by freezing compared to the levels in fresh tissue. In all cases except that of cytoplasmic glucocorticoid receptors, these losses could be prevented by freezing the tissue in 10% aqueous dimethylsulfoxide.

Steroid-receptor proteins in nonepithelial malignancies of the ovary

Tissue samples from 20 patients with nonepithelial ovarian malignancies were assayed for cytosol estrogen (ERc) and in selected samples progestin-receptor (PRc) proteins. The nine germ cell malignancies assayed were low in ERc. One recurrent granulosa cell tumor had an ERc concentration of 36 fmole/mg cytosol protein (fmole/mg c.p.) while three other stromal tumors were low in ERc. Three of seven mixed mesodermal tumors had a high ERc concentration (34-91 fmole/c.p.). Two primary and one recurrent granulosa cell tumors and one endodermal sinus tumor had borderline to elevated PRc levels (18-254 fmole/c.p.). Four of seven mixed mesodermal tumors had borderline to slightly elevated PRc concentrations (13-22 fmole/c.p.). The low ERc in germ cell malignancies of the ovary is consistent with previous clinical observations that the growth of these tumors is not influenced by removal of the contralateral ovary. The high ERc levels detected in three of seven mixed mesodermal ovarian tumors and one of three granulosa cell tumors suggest that hormone-related therapy should be investigated in the management of patients with these malignancies.

Tamoxifen-induced increase in cytosol progestin receptor levels in a case of metastatic endometrial cancer

A patient with endometrial carcinoma metastatic to the groin has had serial cytosol estrogen (ERc) and progestin (PRc) receptor determinations during hormonal therapy. The primary lesion was well differentiated and a left iliac node metastasis had high ERc and borderline PRc levels. PRc levels increased substantially in a subsequently appearing right groin metastasis at three times during the clinical course when the patient was treated with the antiestrogen tamoxifen. The patient had a partial response to the first course of tamoxifen and at one later time briefly had stabilized disease on tamoxifen plus medroxyprogesterone acetate. Tamoxifen with and without progestin should be further evaluated in the treatment of endometrial carcinoma.