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Biomedical subjects

V C Jordan

Publications and source records attributed to V C Jordan.

At least 289 records · Page 16Linked to original sources
PubMed1985

Opposing biological actions of antiestrogens in vitro and in vivo: induction of progesterone receptor in the rat and mouse uterus.

The nonsteroidal antiestrogen tamoxifen, 4-hydroxytamoxifen, and Ly117018 inhibited the estradiol-stimulated induction of progesterone receptors in primary cultures of immature rat uterine cells. This effect was found to be completely reversible with increased concentrations of estradiol. These compounds possessed no estrogenic activity. In contrast, ICI 47,699 (the cis geometric isomer of tamoxifen) and ICI 77,949 (tamoxifen without the dimethylaminoethyl side chain) were fully estrogenic, and bisphenol (4-hydroxytamoxifen without the dimethylaminoethyl side chain) possessed mixed estrogenic/antiestrogenic activity. In primary uterine cell cultures derived from mature ovariectomized mice, 4-hydroxytamoxifen was, again, nonestrogenic and inhibited the estradiol-stimulated induction of progesterone receptors. The antiestrogenic activity of 4-hydroxytamoxifen was effective against both steroidal and nonsteroidal estrogens in either rat- or mouse-derived uterine cell cultures. Using the 3-day uterine assay in vivo, 4-hydroxytamoxifen partially stimulated progesterone receptor induction in the immature rat, whereas it fully stimulated the same end point in the mature ovariectomized mouse. These results emphasize the difference between antiestrogen activity in vivo and in vitro, and also indicate that the increased agonist activity of 4-hydroxytamoxifen in the mouse compared to that in the rat in vivo is not reflected in vitro. Therefore, we have extended the model of antiestrogen action previously described in primary pituitary cell cultures to progesterone receptor induction in two murine uterine cell cultures.

Animals↗
PubMed1985

Structure-activity relationships of estrogens.

The last 50 years has seen an exponential rise in the published reports about estrogen action. The model to describe the early events in the mechanism of action of estrogens via the estrogen receptor is updated in this paper to incorporate some of the recent data on the subcellular localization of the receptor. New evidence suggests that the receptor is a nuclear protein, so it appears that estrogens must first diffuse into the nuclear compartment to initiate estrogen action via the receptor complex. This review traces the development of potent estrogenic compounds by the study of their structure-activity relationships. Studies of structure-activity relationships in vivo using Allen Doisy or 3-day uterine weight tests can provide much valuable information, but the assays suffer from the complex problems of pharmacokinetics and metabolic transformation. Studies in vitro using primary cultures of rat pituitary or uterine cells to assay the ability of a compound to induce prolactin synthesis or progesterone receptor synthesis, respectively, can provide essential information about the structural requirements for a compound to produce estrogenic effects. Nevertheless, it should be pointed out that studies in vivo are required to determine whether a compound is metabolically activated to an estrogen. Estrogen receptor binding models are presented to describe the changes in a molecule that will predict high affinity for the ligand and agonist, partial agonist and antagonist properties of the ligand-receptor complex. Most estrogenic pesticides and phytoestrogens comform to the predictions of the estrogen receptor binding model.

Animals↗
PubMed1984

Long-term tamoxifen adjuvant therapy in node-positive breast cancer: a metabolic and pilot clinical study.

One hundred twenty-four disease-free postoperative women with ipsilateral-node-positive breast carcinoma were evaluated to assess the metabolic handling and clinical side effects of adjuvant tamoxifen for periods in excess of five years. The patients received postoperative combination chemotherapy for a median of 14 months. Thirty-eight patients received no tamoxifen. Tamoxifen 10 mgs bid was administered to 86 patients during the chemotherapy duration and has been continued in 43 of these patients for up to five years after termination of chemotherapy. Serum samples obtained in 7 patients at 3 to 4-month intervals during the first 5 years revealed that levels of tamoxifen, N-desmethyltamoxifen, and metabolite Y were generally constant throughout the period of drug administration. There was no difference in side effects observed after stopping chemotherapy between the group continuing tamoxifen and the groups stopping tamoxifen or never receiving the drug. Relapse-free survival was superior with increasing duration of exposure to tamoxifen, but this observation can only be considered preliminary due to the small number of patients involved and the study design. The clinical effects observed and the failure to demonstrate the development of a host-metabolic tolerance to tamoxifen provides a rational basis for developing a clinical trial to continue adjuvant tamoxifen therapy for periods in excess of four years after chemotherapy.

Adult↗
PubMed1984

The inhibition of prolactin synthesis in GH3 rat pituitary tumor cells by monohydroxytamoxifen is associated with changes in the properties of the estrogen receptor.

E2 (1 nM) stimulated the synthesis of PRL in GH3 cells. OH TAM (100 nM) did not affect basal PRL synthesis, but completely inhibited the increase produced by 1 nM E2. [3H]E2 and [3H]OH TAM both bound to the cytosolic 8S ER and these were split into 4S subunits on sucrose gradients containing 0.4 M KCl. By comparison, ER complexes extracted from nuclei of GH3 cells cultured in media containing [3H]E2 or [3H]OH TAM both sedimented at 5S on sucrose gradients containing 0.4 M KCl. Both 4S and 5S ER complexes were recognized by the monoclonal antibody D547 which increased their sedimentation coefficients to 8-9S. In contrast, a polyclonal antibody raised to calf uterine ER in the goat, interacted with the cytosolic ER so that the binding of [3H]E2 was inhibited but the binding of [3]OH TAM was only slightly reduced. A molecular model is proposed to describe the binding of E2 and OH TAM to the ER that might contribute to an understanding of estrogen and antiestrogen action.

Animals↗
PubMed1984

An immobilized antiestrogen binds a specific uterine protein in addition to estrogen receptor proteins.

An antiestrogen affinity resin was synthesized by conjugating LY117018, a benzothiophene -derived antiestrogen, to epoxy-activated agarose. This affinity resin bound the Mr = 50 000 and 65 000 estrogen receptor proteins of rabbit uterine cytosol; in addition, it retained a protein from the cytosols of both rat and rabbit uteri that exhibited an ability to interact specifically with LY117018. The possibility that the LY117018 binding protein, which is distinct from estrogen receptors, may play a role in the antiestrogenic actions of LY117018 is discussed.

Agar↗
PubMed1984

Nuclear [3H]4-hydroxytamoxifen (4-OHTAM)- and [3H]estradiol (E2)-estrogen receptor complexes in the MCF-7 breast cancer and GH3 pituitary tumor cell lines.

Nuclear [3H]4-OHTAM-ER complexes extracted by 0.6 M KCl from the MCF-7 human breast cancer and the GH3 rat pituitary tumor cell lines, sedimented as a 5S form on sucrose gradients after 1 to 24 h exposure to ligand (20 nM). The nuclear [3H]4-OHTAM-ER from MCF-7 cells increased from 2 +/- 0.2 pmoles/mg DNA at 1 h to approximately 4 +/- 0.1 pmoles/mg DNA at 6 and 24 h. In the GH3 cells the nuclear binding of [3H]4-OHTAM increased rapidly, and there was no significant difference in the receptor levels at 1 and 6 h (10 +/- 1.3 and 8.9 +/- 1.1 pmoles/mg DNA, respectively). At 24 h there was a decrease in [3H]4-OHTAM-ER levels (7.0 +/- 0.2 pmoles/mg DNA); however, this was due primarily to an increase in DNA synthesis, which occurred in these cells by 24 h. It appears that in both cell lines there is no processing of nuclear [3H]4-OHTAM-ER, and it is not associated with changes in sedimentation coefficient of the complex. When both cell lines were incubated with [3H]E2 (20 nM), processing of the nuclear [3H]E2-ER occurred over 6 h. In the MCF-7 cells there was a decrease in receptor content within 6 h from 3.2 +/- 0.2 to 0.9 +/- 0.2 pmoles/mg DNA. The nuclear [3H]E2-ER in the GH3 cells decreased from 7.3 +/- 0.5 to 2.8 +/- 0.3 pmoles/mg DNA. Although these cell lines appeared to process the [3H]E2-ER complex, the nuclear forms of [3H]E2-ER were different.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1984

Structural requirements for the pharmacological activity of nonsteroidal antiestrogens in vitro.

The structure-activity relationships of a tamoxifen (TAM) (Z-1-(4- beta-dimethylaminoethoxyphenyl)1,2-diphenylbut-1-ene) series have been investigated. The tamoxifen derivatives were assayed in vitro by their modulation of estradiol (E2)-stimulated prolactin synthesis in primary cultures of dispersed rat pituitary gland cells. Monohydroxylation of TAM in position 4 of the stilbene ring system was found to be the optimal substitution for binding to the estrogen receptor [relative binding affinity (RBA) = 234] and to inhibit E2 (1 nM)-stimulated prolactin synthesis (IC50 7 nM) by pituitary cells in primary culture. Substitution in positions 3 and 4 to form a catechol did not decrease affinity for the estrogen receptor (RBA = 252), and potency as an antiestrogen was maintained in the prolactin assay (IC50 20 nM) as long as oxidation of the catechol was prevented. All of the hydroxylated derivatives of tamoxifen tested were estrogen antagonists; however, removal of the alkylaminoethoxy side chain from TAM produced a full estrogen agonist with low potency (20 nM). In contrast, removal of the side chain from 4-hydroxytamoxifen (4-OH TAM) produced a partial agonist. A structural analogue of 4-OH TAM, 3-[beta-dimethylaminoethoxy]-11-ethyl-12-(4-hydroxyphenyl)5,6- dihydrodibenzo[a,e]-cyclooctene (7c) had a decreased potency (IC50 16 nM) compared with 4-OH TAM (IC50 4 nM in the same experiment) as an estrogen antagonist. If the side chain was changed from a dimethylaminoethoxy to glyceryl, antagonist activity was reduced (IC50 0.8 microM). An allyl side chain produced a compound with no antiestrogenic activity at concentrations up to 1 microM. An adaptation of Belleau's macromolecular perturbation theory is suggested to explain the interaction of agonists, antagonists, and partial agonists at the ligand binding site of the estrogen receptor.

Animals↗
PubMed1984

Estrogen-stimulated prolactin synthesis in vitro. Classification of agonist, partial agonist, and antagonist actions based on structure.

An in vitro assay that depends upon the synthesis of prolactin by primary cultures of dispersed cells from immature rat pituitary cells was used to study the structural requirement for estrogen action. Two categories of estrogens were identified: full estrogens (agonists) and partial estrogens (partial agonists) with antiestrogenic actions against the effects of 0.1 nM estradiol (E2). All of the agonists [diethylstilbestrol (DES), dimethylstilbestrol (DMS), R2858, and RU16117] produced a dose-related increase in prolactin synthesis equivalent to E2, although potencies were different: E2 = DES = R2858 greater than RU16117 greater than DMS. Partial agonists [ICI 3188, tri(4-hydroxyphenyl)chloroethylene, and bisphenol] each had bis(4-hydroxyphenol) substitutions at the ethylene double bond and stimulated prolactin synthesis only to about 50% of the maximal response observed with E2. Trianisylchloroethylene was weakly active as a partial agonist, but at high concentration (10 microM) was able to decrease prolactin synthesis produced by 0.1 nM E2. Previous studies from these laboratories showed that triphenylethylene derivatives with a strategically located alkyl aminoethoxyside chain are complete E2 antagonists with no agonist activity. Two series of novel compounds were assayed to determine whether their structures would predict biological activity. LN2299, the cis geometric isomer of a triphenylbromethylene, was a full agonist with activity equivalent to zuclomiphene, the cis geometric isomer of clomiphene. Cyclofenyl was a partial agonist, but deacetylation to the diphenol increased partial agonist activity and potency. However, introduction of a single pyrrolidinoethylside chain into the deacetylated cyclofenyl increased antiestrogenic potency and completely suppressed the expression of agonist activity. Finally, LN2833, with a novel C(OH)CH3 side chain in the position normally occupied by the alkylaminoethoxyside chain of most antiestrogens, produced antiestrogen activity with no estrogen properties. Antiestrogenic potency was increased in LN2839 by a phenol in the triphenylethylene in a position equivalent to the 3-phenolic hydroxyl of E2. In general, the potency and biological properties could be predicted by reference to the structure of the molecule. Potent estrogens or antiestrogens have a phenolic hydroxyl in a position that would be equivalent to the 3-phenolic hydroxyl of E2. Partial agonist action is predicted by a 4-hydroxyphenol attached to the same carbon as the phenyl ring equivalent to the A-ring of E2.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1984

Differences between estrogen- and antiestrogen-estrogen receptor complexes from human breast tumors identified with an antibody raised against the estrogen receptor.

Radiolabeled estrogens 17 beta-[3H]estradiol and diethylstilbestrol ( [3H]DES) and the antiestrogen [3H]monohydroxytamoxifen ( [3H]MHT) all bind with high affinity to the extranuclear estrogen receptor (ER) from the MCF-7 human breast tumor cell line (Kd = 3 X 10(-10), 2 X 10(-10), and 0.63 X 10(-10) M, respectively). A polyclonal antibody raised in a goat to the calf nuclear ER selectively decreased the binding affinity and number of binding sites for 17 beta-[3H]estradiol, but did not appear to affect these binding parameters for [3H]MHT. In the presence of goat antibody, the binding of the nonsteroidal estrogen DES was so perturbed that it was not possible to quantitate the decreased number of binding sites or affinity of this compound as assessed by Scatchard saturation analysis. These results were confirmed in human breast tumor cytosols by sucrose density gradient analysis. The binding of 17 beta-[3H]-estradiol and [3H]DES to the ER was significantly reduced by preincubation with the polyclonal antibody, whereas the binding of [3H]MHT was reduced only when the tumor cytosol was preincubated with a very high concentration of antibody. At these concentrations of antibody, the binding of 17 beta-[3H]estradiol and [3H]DES to the receptor was prevented completely. In contrast, when the antibody was added to the tumor cytosol after the 3H-ligand had bound to the receptor, the binding properties of all 3H-ligands were unaffected. The [3H]MHT-ER antibody complex consistently sedimented as a higher-molecular-weight complex on sucrose density gradients than did the corresponding estrogenic complexes. The decrease in the affinity of estrogenic ligands can be explained in part by an increase in the dissociation rate at 4 degrees of these compounds from the ER. The dissociation rate of MHT was unaffected by the goat antibody. These results imply that there are important differences in the binding of antiestrogen and estrogens to the tumor cytosol ER. A ligand-binding model is proposed that may aid in the understanding of antiestrogen action.

Antibodies↗
PubMed1983

Direct and reversible inhibition of estradiol-stimulated prolactin synthesis by antiestrogens in vitro.

The antiestrogens tamoxifen and monohydroxytamoxifen inhibited the estradiol-stimulated increase in prolactin synthesis by dispersed cells in culture derived from immature rat pituitary glands. Monohydroxytamoxifen had a relative binding affinity for the estrogen receptor similar to that of estradiol, whereas tamoxifen's relative binding affinity was approximately 3%. This was consistent with the observation that monohydroxytamoxifen was 30 times more potent than tamoxifen as an antiestrogen in vitro. To avoid the possibility that tamoxifen was fractionally metabolized to monohydroxytamoxifen by the pituitary cells, the p-methyl, p-chloro, and p-fluoro derivatives of tamoxifen that are unlikely to be converted to monohydroxytamoxifen were tested for activity. The substitution did not have a detrimental effect on their ability to inhibit the binding of [3H]estradiol to either rat uterine or pituitary gland estrogen receptors. Similarly, the derivatives of tamoxifen inhibited estradiol-stimulated prolactin synthesis at concentrations that were consistent with their relative binding affinities. Although it is clearly an advantage for tamoxifen to be metabolized to the more potent antiestrogen monohydroxytamoxifen, we have shown that this is not a prerequisite for the antiestrogenic actions of tamoxifen. With the direct actions of antiestrogens established, the pituitary cell system was validated for further structure-activity relationship studies. Overall, the inhibition of estradiol-stimulated prolactin synthesis by antiestrogens is competitive and reversible with estradiol, an effect that can be explained by interactions with the estrogen receptor system.

Animals↗
PubMed1983

An estrogen receptor model to describe the regulation of prolactin synthesis by antiestrogens in vitro.

A hypothetical model of the ligand interaction with the estrogen receptor binding site has been developed to describe the structural features necessary to initiate or to inhibit prolactin synthesis in vitro. The biological potency of the binding ligands is directly related to their relative binding affinity (RBA) for the estrogen receptor. The relative potencies of antiestrogens to inhibit estradiol-stimulated prolactin synthesis was trans-monohydroxytamoxifen identical to cis-monohydroxytamoxifen identical to tamoxifen, consistent with their RBAs for uterine estrogen receptor. Similarly the relative potency of estrogens to stimulate prolactin synthesis was diethylstilbestrol identical to estradiol greater than ICI 77,949 greater than ICI 47,699 identical to zuclomiphene, consistent with their RBAs. The compound LY126412 (trioxifene without the aminoethoxy side chain) did not interact with the estrogen receptor at the concentrations tested (10(-8)--10(-6) M) or exhibit estrogenic or antiestrogenic properties using the prolactin synthesis assay. Overall, the ligand-receptor model stresses the structural requirement for high affinity binding and the critical positioning of the alkylamino-ethoxy side chain in space (in relation to the ligand-binding site on the estrogen receptor) to prevent prolactin synthesis.

Animals↗
PubMed1983

Determination of tamoxifen and metabolites in human serum by high-performance liquid chromatography with post-column fluorescence activation.

Sensitive and reproducible analyses were developed for assaying tamoxifen, monohydroxytamoxifen, N-desmethyltamoxifen, metabolite E [trans-1(4-hydroxyphenyl)1,2-diphenylbut-1-ene] and a new metabolite, metabolite Y [trans-1(4-hydroxyethoxyphenyl)-1,2-diphenylbut-1-ene] in human serum using high-performance liquid chromatography (HPLC). Three different systems were developed for specific purposes. All chromatography was performed using serum extracts made with hexane-butanol. Detection was by fluorimetry of phenanthrene derivatives formed by on-stream UV irradiation with a newly described device for post-column irradiation of the HPLC stream. This device may be of use in other HPLC systems requiring post-column photochemical reactions.

Animals↗
PubMed1983

Interaction of [3H] estradiol - and [3H] monohydroxytamoxifen-estrogen receptor complexes with a monoclonal antibody.

The aim of this study was to compare and contrast the interaction of estrogen [( 3H]17 beta-estradiol)- or antiestrogen [( 3H]monohydroxytamoxifen)-receptor complexes from human breast tumor cytosols with monoclonal antibodies raised to the human breast tumor estrogen receptor. Breast tumor cytosols containing estrogen receptor which sedimented as radiolabeled peaks in either the 8S, 8S and 4S, or 4S regions of sucrose density gradients, interacted with the monoclonal antibody D547 to produce a broad 9-10S peak, a broad 8S-10S peak, or a more discrete 8S peak, respectively. On high salt (0.4M KC1) sucrose density gradients the 4S ligand-receptor complex plus antibody produced a binding peak at approximately the 8S region of the gradient. These sedimentation studies with the monoclonal antibody D547, and similar studies with the monoclonal antibody D58, could detect no differences in the cytosolic estrogen receptor whether complexed with [3H]estradiol or with [3H]monohydroxytamoxifen. These observations were confirmed by Scatchard equilibrium saturation analysis and sucrose density gradient analysis of cytosols from the MCF-7 human breast cancer cell line. The antibody D547 interacted with 8S ER from these cytosols to produce a broad 8S-10S peak, but the antibody produced no change in the affinity or number of binding sites present in these cytosols. It seems, therefore, that the antigenic determinants recognized by these particular antibodies on the breast tumor cytosolic receptor are not significantly altered by the binding of either an estrogen or an antiestrogen to the receptor.

Antibodies, Monoclonal↗
PubMed1983

Laboratory studies to develop general principles for the adjuvant treatment of breast cancer with antiestrogens: problems and potential for future clinical applications.

The general pharmacology of tamoxifen in animals and man is reviewed with particular reference to the long-term adjuvant therapy of node-positive breast cancer. Rats with dimethylbenzanthracene (DMBA)-induced mammary carcinomata have been used extensively as a laboratory model to study hormone-dependent cancer. The administration of a 30-day course of tamoxifen (50 micrograms daily) starting 5, 15, 30, or 50 days after DMBA caused a delay in tumor appearance and decrease in the cumulative number of tumors that were induced by 200 days. Similarly, the administration of increasing doses of tamoxifen (0.2, 3, 50, and 800 micrograms daily) between 30 and 60 days after DMBA produced a dose-related delay in tumor appearance and a decrease in the cumulative number of tumors at 200 days. Since the tumors that were induced after tamoxifen still responded to ovariectomy, tamoxifen appears to act as an inhibitor of the tumor cell cycle rather than as a tumoricidal agent in this model. This principle was exemplified by comparing a short course (30 day) with a continuous course (170 day) of tamoxifen initiated 30 days after DMBA. The short course of therapy only delayed tumor appearance whereas continuous therapy maintained 90% of the animals in a tumor-free state. These data strongly support the use of long-term (up to five-year) adjuvant therapy with tamoxifen in patients as a suppressive therapy for hormone-sensitive metastases.

9,10-Dimethyl-1,2-benzanthracene↗
PubMed1983

Differential antiestrogen action in the immature rat uterus: a comparison of hydroxylated antiestrogens with high affinity for the estrogen receptor.

The non-steroidal antiestrogens LY117018 and monohydroxytamoxifen are partial agonists in the 3-day immature rat uterine weight test. Both compounds stimulated increases in uterine progesterone receptor concentration (as determined by DCC assay and SDG analysis) and luminal epithelial cell height. However, LY117018 was much less estrogenic than monohydroxytamoxifen. The study also showed that the antiestrogenic effects of LY117018 and monohydroxytamoxifen could be reversed in vivo by increasing doses of estradiol. The partial uterotrophic effect of monohydroxytamoxifen and full uterotrophic effects of estradiol were both inhibited by high doses of LY117018 at an approximate dosage ratio of 1:24, w/w. This result suggests a common mechanism for the action of estradiol and the hydroxylated antiestrogens. Since LY117018 reversed the binding of [3H]estradiol and estrogen-competable [3H]monohydroxytamoxifen binding in the rat uterus in vivo, the effects of nonsteroidal antiestrogens on gross uterine wet weight can be explained by competitive interaction with estradiol via the estrogen receptor mechanism. However, the weakly estrogenic antiestrogen LY117018 was unable to inhibit estrogen-stimulated rises in luminal epithelium cell height and progesterone receptor levels. These data suggest a differential interaction by LY117018 in the rat uterus so that only selected estrogen stimulated effects i.e.: uterine wet weight, are antagonized by this particular "antiestrogenic" dose of the drug.

Animals↗