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Biomedical subjects

G Prestwich

Publications and source records attributed to G Prestwich.

13 recordsLinked to original sources
PubMed1990

Chemical modification of aldehyde dehydrogenase by a vinyl ketone analogue of an insect pheromone.

A major component of the sex pheromone from the tobacco budworm moth Heliothis virescens is a C16 straight-chain aldehyde with a single unsaturation at the eleventh position. The sex pheromones are inactivated when metabolized to their corresponding acids by insect aldehyde dehydrogenase. During this investigation it was demonstrated that the C16 aldehyde is a good substrate for human aldehyde dehydrogenase (EC 1.2.1.3) isoenzymes E1 and E2 with Km and Kcat. values at pH 7.0 of 2 microM and 0.4 mumol of NADH/min per mg and of 0.6 microM and 0.24 mumol of NADH/min per mg respectively. A vinyl ketone analogue of the pheromone inhibited insect pheromone metabolism; it also inactivated human aldehyde dehydrogenase. Total inactivation of both isoenzymes was achieved at stoichiometric (equal or less than the subunit number) concentrations of vinyl ketone, incorporating 2.1-2.6 molecules/molecule of enzyme. Substrate protection was observed in the presence of the parent aldehyde and 5'-AMP. Peptide maps of tryptic digests of the E2 isoenzyme modified with 3H-labelled vinyl ketone showed that incorporation occurred into a single peptide peak. The labelled peptide of E2 isoenzyme was further purified on h.p.l.c. and sequenced. The label was incorporated into cysteine-302 in the primary structure of E2 isoenzyme, thus indicating that cysteine-302 is located in the aldehyde substrate area of the active site of aldehyde dehydrogenase. Affinity labelling of aldehyde dehydrogenase with vinyl ketones may prove to be of general utility in biochemical studies of these enzymes.

Aldehyde Dehydrogenase↗
PubMed1979

Inhibition of cell division and stimulation of progesterone receptor synthesis in rat oestrogen target tissues by non-steroidal antioestrogens.

Recent studies from this laboratory have demonstrated oestradiol-oestrogen receptors associated with the nuclear compartment of the rat uterine cell will initiate protein synthesis, as evidenced by a rise in progesterone receptor concentrations, and cell division whereas the anti-oestrogen - oestrogen receptor complex causes protein synthesis and cellular hypertrophy rather than hyperplasia. It is probable that this separation of biological activities resides in the intrinsic activity of the respective receptor complexes. We have demonstrated that caution should be exercised in the interpretation of low affinity ligand-hormone receptor interactions undertaken in vitro. Simple tests for ligand specificity for a binding protein are clearly insufficient evidence to characterise a hormone receptor complex using a conventional 15 hr technique of sucrose density gradient analysis. Oestrogens and anti-oestrogens do not seem to disrupt the subunit integrity of the cytoplasmic oestrogen receptor and it appears likely that the ligand plays a fundamental role in confering the correct biological properties to the hormone receptor complex.

Animals↗
PubMed1978

Dose-related effects of non-steroidal antioestrogens nad oestrogens on the measurement of cytoplasmic oestrogen receptors in the rat and mouse uterus.

The dose-related effects of non-steroidal antioestrogens and oestrogens on the measurement of cytoplasmic oestrogen receptors in the rat uterus have been determined. The simultaneous administration of tamoxifen or monohydroxytamoxifen and oestradiol on three consecutive days resulted in dose-dependent decreases in both the wet weight of the uterus and the number of available cytoplasmic oestrogen receptors. The oestrogenic triphenylethylenes ICI 47 699 and ICI 3188 both produced dose-dependent decreases in the number of available cytoplasmic oestrogen receptors. Increasing doses of ICI 47 699 resulted in increasing concentrations of oestrogen receptors within the nucleus. The effects of tamoxifen and oestradiol-17 beta were compared in the ovariectomized mouse; replenishment of uterine oestrogen receptors was less evident in tamoxifen-treated animals than in animals receiving oestradiol, although increases in uterine weight were similar. A single large dose of tamoxifen (50 microgram) produced a prolonged depletion of cytoplasmic oestrogen receptors whilst stimulating rises in uterine weight and DNA and protein content. The results demonstrate that the depletion of the uterine cytoplasmic oestrogen receptor pool is a function of the dose administered for any compound with the ability to translocate oestrogen receptors to the nucleus and as such is not an exclusive characteristic of non-steroidal antioestrogens.

Animals↗
PubMed1977

Binding of [3H]tamoxifen in rat uterine cytosols: a comparison of swinging bucket and vertical tube rotor sucrose density gradient analysis.

The interaction of the antioestrogen tamoxifen with rat uterine oestrogen receptor has been studied using swinging bucket and vertical tube rotor techniques of sucrose density gradient analysis. Tamoxifen inhibits the binding of [3H]oestradiol to the 8S oestrogen receptor protein whereas using the swinging bucket rotor technique of sucrose density gradient analysis [3H]-tamoxifen appears to bind to an oestradiol specific protein which sediments at 4-5S. In contrast, using rapid sucrose density gradient analysis with a vertical tube rotor [3H]tamoxifen shows oestrogen specific binding in the 8S region. It is suggested that conventional techniques of sucrose density gradient analysis are unsuited for the investigation of the low affinity ligand interactions with oestrogen receptor proteins since rapid dissociation of complexes can result in ligand binding to nonreceptor proteins.

Animals↗
PubMed1977

Potential steroidal antiestrogens.

A series of analogues of 17beta-estradiol has been synthesized and the compounds have been tested, using sucrose density gradient analysis, for their ability to compete with [6,7-3H]-17beta-estradiol for the estrogen-receptor protein from mouse uterine homogenates. Active compounds were also tested for antiuterotrophic activity in immature rats and/or mice. 3,17beta-Dihydroxy-6-phenylestra-1,3,5(10),6-tetraene (14) was the most active new compound in the in vitro test suppressing the binding of 17beta-estradiol by 34 and 87%, respectively, at molar ratios of 1 and 3.16. It was significantly more potent than the intermediate 6-oxoestradiol (4) which produced a 52% inhibition of binding at a molar ratio of 3.16. The thiosemicarbazone of 6-oxoestradiol (17) and the derived 3,17beta-dihydroxy-6-(2-imino-4-oxothiazolidinyl-1-imino)estra-1,3,5(10)-triene (19) produced, respectively, only 46 and 16% inhibition of binding at a molar ratio of 10. Introduction of a 1-methyl substituent into either 6-oxo or 6-phenyl compounds reduced affinity for the receptor significantly (compounds 5 and 15) and conversion of the 3-OH into a beta-dialkylaminoethoxy group virtually destroyed all binding activity (compounds 2, 6, 10, and 11). At a molar ratio of 10 compound 14 failed to suppress the uterine weight response of immature rats to 17beta-estradiol, whereas compound 15, at a molar ratio of 200, produced a significant increase in the uterine weight of immature rats but not of immature mice even at a molar ratio of 1000.

Animals↗
PubMed1977

A monohydroxylated metabolite of tamoxifen with potent antioestrogenic activity.

The oestrogenic and antioestrogenic properties of tamoxifen and its monohydroxylated (monohydroxytamoxifen) and dihydroxylated (dihydroxytamoxifen) metabolites have been investigated in the immature rat. Whether administered orally or subcutaneously, monohydroxytamoxifen was more active than tamoxifen as an antioestrogen. Dihydroxytamoxifen was less active than tamoxifen as an antioestrogen, but this derivative alone was unable to induce a uterotrophic response. Both metabolites of tamoxifen were potent inhibitors of the binding of [3H]oestradiol to oestrogen receptors in vitro. It is possible that the metabolites play a supportive role in the antioestrogenic activity of tamoxifen. The potent activity of monohydroxytamoxifen in vivo and in vitro suggests that this compound could be an important new tool for the subcellular investigation of oestrogenic and antioestrogenic events.

Animals↗
PubMedDate not supplied

Nonsteroidal antiestrogens: their biological effects and potential mechanisms of action.

The uterotropic and antiuterotrapic effects of a variety of structural derivatives of the nonsteroidal antiestrogen tamoxifen have been determined in the rat and the mouse. One derivative, monohydroxytamoxifen, was found to be a potent antiestrogen in the rat, with a high affinity for the estrogen receptor. Various techniques of sucrose density gradient analysis were used to demonstrate that estradiol and tamoxifen bind to the rat uterine cytoplasmic estrogen receptor. Estrogens and antiestrogens provoke the translocation of estrogen receptors to the nucleus and deplete the cytoplasmic estrogen receptor pool for short or long periods depending on the dose administered. Estradiol stimulates endometrial hyperplasia with an increase in total uterine DNA content, whereas tamoxifen stimulates endometrial hypertrophy with only a slight increase in uterine DNA content. It is concluded that the molecular shape of the ligand that binds to the estrogen receptor determines antiestrogenic activity.

Animals↗