Cecil H. Robinson

Inactivation of aromatase in vitro by 4-hydroxy-4-androstene-3,17-dione and 4-acetoxy-4-androstene-3,17-dione and sustained effects in vivo

4-Hydroxy-4-androstene-3,17-dione (4-OHA) and 4-acetoxy-4-androstene-3,17-dione (4-AcA), in addition to being competitive inhibitors of aromatase, cause time-dependent, irreversible, loss of enzyme activity in both human placental and rat ovarian microsomes. In vivo, treatment of rats with 4-OHA also causes loss of ovarian aromatase activity. To test whether this loss of activity could have in vivo significance, rats with hormone-dependent, mammary tumors were treated with 4-OHA on alternate weeks. Tumor regression continued to occur during the weeks without treatment. These findings suggest that inactivation of aromatase is important in the mechanism of action of the compounds in vivo.

Mechanism of human placental aromatase : a new active site model

Based on Akhtars ferric peroxide mechanism and on recent studies in our own laboratory, we present a detailed proposal for aromatase action. This picture can account for the known stereochemical consequences at C-19 observed by others using isotopes of hydrogen and oxygen. The postulated process involves anchoring of the 19-hydroxymethyl and 19-oxo groups at the active site by a glutamate residue, which also serves to activate the 19-oxo group for attack by ferric peroxy species in the third oxidative step.

Synthesis and evaluation of 10β-substituted 4-estrene-3,17-diones as inhibitors of human placental microsomal aromatase

Abstract This paper describes the synthesis of a series of new 10β-substituted 4-estrene-3,17-dione analogs ( 1–8 ). These compounds, together with a number of known analogs ( 9–14 ), have been evaluated as reversible or irreversible inhibitors of human placental microsomal aromatase. The best reversible inhibitor is the 10β-vinyl compound ( 9 ). The only compounds causing irreversible inhibition of aromatase are the 10β-propargyl compound ( 1 ) and the 10β-difluoromethyl compound ( 12 ).

The Δ5-3-ketosteroid isomerase reaction: catalytic mechanism, specificity and inhibition

The Δ5-3-ketosteroid isomerase of Pseudomonas testosteroni promotes the conversion of Δ5-3-ketosteroids to the α,β-conjugated 3-ketosteroids by an unusual intramolecular rearrangement involving a cis, cis diaxial proton transfer from the 4β- to the 6β-position. The enzyme is available as a pure protein in two crystalline forms of known unit cell dimensions and crystallographic symmetry (monoclinic and hexagonal). The amino acid sequence of the protomer has been established. Many lines of evidence suggest a mechanism of isomerization involving an enolic intermediate and the participation of an acidic and a basic group of the enzyme in the catalytic process. Single histidyl and aspartyl residues may be involved in the isomerization process and a mechanism for their participation is proposed. Certain β,γ-acetylenic 5,10-seco-3-ketosteroids are converted by the isomerase to the corresponding α,β-unsaturated allene(s) which irreversibly inactivate the enzyme. A partial purification of the soluble isomerase of rat liver has been achieved. These preparations are profoundly and specifically activated by reduced glutathione. The precise mechanism of this effect remains to be established.

Novel inhibitors of lanosterol 14α-methyl demethylase, a critical enzyme in cholesterol biosynthesis

A series of novel 32-functionalised lanost-7-en-3-ols (1)–(7) has been synthesised; these compounds are all powerful inhibitors of lanosterol 14α-methyl demethylase, an important enzyme in cholesterol biosynthesis.

Novel 10β-aziridinyl steroids; inhibitors of aromatase

The novel 10β-aziridinylestr-4-ene-3,17-diones 17 and 18 and the corresponding 10β-aziridinyl-β-hydroxyestr-4-en-3-ones 19 and 20 have been synthesized from the 19-oximino-19-methyl intermediate 12b. The key reaction was the conversion of the 19-oxime 12b into the diastereoisomeric 1OR-aziridines 13 and 14 by lithium aluminium hydride (LAH). Compounds 17–20 are powerful and stereoselective inhibitors of human placental microsomal aromatase. The most potent compound was 17(Ki = 3.4 nmol dm–3). The 19R- isomers 17 and 19 are more effective than the corresponding 19S-isomers 18 and 20, respectively. Unlike the corresponding 10β-oxiranes and -thiiranes which are classical competitive inhibitors, the (19R)-aziridines 17 and 19 appear to be slow-binding inhibitors. Spectroscopic studies with microsomal aromatase preparations indicate that the inhibition process involves binding of aziridine nitrogen to the heme-iron of the enzyme.

Oxidative cyclization and fragmentation of steroidal alcohols induced by ceric ammonium nitrate

Abstract Oxidative cyclizations of steroidal alcohols (1a–1d) using ceric ammonium nitrate (CAN) in aqueous acetonitrile or aqueous acetic acid gave the corresponding cyclic ethers (2a–2d), whereas the tertiary alcohol (4) gave secosteroid (5).

Stereoselective inhibition of aromatase by novel epoxysteroids

The diastereoisomeric 10-(epoxyethyl)estr-4-ene-3,17-diones (2) and (3) have been synthesized, their conformations and configurations have been established by X-ray crystallographic analysis, and they have been shown to be powerful inhibitors of human placental aromatase.

Studies on the effect of 5α-androstane-3α, 17α-diol in the canine prostate in vivo

Abstract Adult beagle dogs, castrated one month previously, were treated with 5α-androstane-3α, 17α-diol (total dose 300 mg) over a period of one month. Examination of the prostate after treatment showed no significant change in size, weight or histological appearance from the castrate dog prostate. Subsequent administration of 5α-androstane-3α, 17β-diol (300 mg) over the same period of time resulted in restoration of the prostate size, weight and histological appearance to that of the normal intact dog prostate. It is concluded that exogenously administered 5α-androstane-3α, 17α-diol does not promote prostatic growth in the castrate beagle dog.

Stereoselective inhibition of human placental aromatase

We have synthesized the (19R)- and (19S)-isomers (2 and 3 respectively) of 10 beta-oxiranylestr-4-ene-3,17-dione. The configurations and conformations of these compounds were established by X-ray crystallographic analysis. Each of these compounds is a powerful competitive inhibitor of human placental microsomal aromatase, and stereoselectivity of inhibition was observed (Ki values for 2 and 3 were 7 and 75 nanomolar, respectively). Spectroscopic studies with purified aromatase indicate that the inhibition process involves reversible binding of oxirane oxygen to the heme iron of the enzyme. The (19R)- and (19S)-10 beta-thiiranes (6 and 7) corresponding to 2 and 3 have been synthesized from the oxiranes by a stereospecific process. The thiiranes are very effective competitive inhibitors of placental aromatase, and show even greater stereoselectivity in binding than the oxiranes (Ki values for 6 and 7 were 1 and 75 nanomolar, respectively). Spectroscopic studies with purified aromatase indicate that the inhibition process involves reversible binding of thiirane sulfur to heme iron.