Enzyme Inactivation by Potential Metabolites of An Aromatase-Activated Inhibitor (Mdl 18,962)

Abstract

MDL 18,962, 19-acetylenic androstenedione, is an enzyme-activated inhibitor of estrogen biosynthesis which is in Phase I clinical evaluations as a potential therapeutic agent for estrogen-dependent cancers. 19-Acetylenic analogs corresponding to the major metabolites of androstenedione were synthesized as potential metabolites of MDL 18,962. These compounds were 19-acetylenic testosterone, the product of 17 beta-hydroxy steroid oxidoreductase, 6 beta-hydroxy- and 6-oxo-19-acetylenic androstenedione, products of P450 steroid 6 beta-hydroxylase and alcohol dehydrogenase, respectively. All of these analogs showed time-dependent inactivation of human placental aromatase activity. The time-dependent Ki and t1/2 at infinite inhibitor concentration (tau 50) were 4.3 nM, 12.0 min for MDL 18,962; 28 nM, 7.8 min for 17-hydroxy analog; 13 nM, 37 min for 6 beta-hydroxy analog; and 167 nM, 6.1 min for the 6-oxo analog. The 19-acetylenic testosterone, a confirmed metabolite from primate studies, was 25% as efficient as MDL 18,962 for aromatase inactivation, while 6 beta-hydroxy- and 6-oxo analogs were 11% and 5%, respectively as efficient as their parent compound. These data indicate that first-pass metabolism of MDL 18,962 does not cause an obligatory loss of time-dependent inhibition of human aromatase activity.

The biosynthesis of steroids and triterpenoids

“…782,783 Inhibition of aromatase by R76713 has been shown to be due exclusively to the (+)-S-isomer 784 (vorozole) which is now under clinical evaluation. Structure-activity relationships of some aromatase inhibitors have been described 785,786 and several syntheses of potential new [787][788][789][790][791][792][793][794][795] and known 796 steroidal inhibitors substituted at the 4-, 6-, 7-and 19-positions have been reported; most non-steroidal inhibitors incorporate 3-and 4-pyridyl groups 656,659,664,797,798 (cf. P-450 17 ).…”

Validation of Trifluoromethylphenyl Diazirine Cholesterol Analogues As Cholesterol Mimetics and Photolabeling Reagents

“…(3β,17β)-3-(Methoxymethoxy)-androst-5-en-17-one ( 9 ) was prepared according to a literature procedure and dissolved in stirred cold (0 °C) THF/EtOH (1:4, 10 mL). NaBH 4 (76 mg, 2 mmol) was added in portions, and the reaction was allowed to warm to RT.…”

“…After 16 h, the THF was removed under reduced pressure, and the residue was purified by flash column chromatography (silica gel eluted with 25% EtOAc in hexanes) to give MQ238 (8) as a solid (115 mg, 96%): 1 (3β,17β)-3-(Methoxymethoxy)-androst-5-en-17-ol (10). (3β,17β)-3-(Methoxymethoxy)-androst-5-en-17-one (9) was prepared according to a literature procedure 34 and dissolved in stirred cold (0 °C) THF/EtOH (1:4, 10 mL). NaBH 4 (76 mg, 2 mmol) was added in portions, and the reaction was allowed to warm to RT.…”

“…Aqueous saturated NH 4 Cl (5 mL) was added, and the product was extracted into CH 2 Cl 2 (3 × 60 mL). The combined organic layers were washed with brine, dried over anhydrous Na 2 SO 4 , and filtered, and the solvent was removed under reduced pressure to give an off-white solid, which was purified by flash column chromatography (silica gel eluted with 20−35% EtOAc in hexanes) to give known steroid 13 34 as a white solid (100 mg, 99%): 1 H NMR (400 MHz, CDCl 3 ) δ 5.85 (s, 1H), 3.61 (t, J = 8.2 Hz, 1H), 2.80−0.85 (m), 0.76 (s); 13 C NMR (100 MHz, CDCl 3 ) δ 199.6, 167.4, 125.6, 81.5, 81.2, 71.9, 54.3, 50.5, 42.9, 41.6, 36.5, 36.0, 34.6, 34.5, 33.1, 31.8, 30.4, 23.3 (2 × C), 21.0, 11.1.…”

1,2,3-Thiadiazole: A Novel Heterocyclic Heme Ligand for the Design of Cytochrome P450 Inhibitors

“…Besides high specificity and affinity with the biological target, a desirable attribute of a potential therapeutic agent is its ability to function as a mechanism-based inhibitor with a low partition number (Walsh, 1982). In this connection several mechanism-based inhibitors have been developed for specific P450 enzymes (Johnston et al, 1995(Johnston et al, , 1990Warren et al, 1995;Brueggemeier, 1994;Numazawa et al, 1993;Peet et al, 1993;Lesuisse et al, 1992). P450 enzymes oxidize the acetylenic group to a reactive intermediate that can inactivate the P450 by alkylation of the protein or the heme prosthetic group (Ortiz de Montellano & Reich, 1986).…”