Clues to the development of mechanism-based inactivators of 3 alpha-hydroxysteroid dehydrogenase: comparison of steroidal and nonsteroidal Michael acceptors and epoxides.
A series of steroidal and nonsteroidal Michael acceptors that represent reaction products for 3 alpha-hydroxysteroid dehydrogenase were synthesized and evaluated as potential enzyme-generated inactivators. Introduction of exocyclic olefins either at C-2 or C-6 produced inhibitors with high affinity for the enzyme (0.05 to 5.0 microM). However, despite this affinity, none of these compounds produced time-dependent inactivation of the enzyme. By contrast, analogs based on 1-phenyl-2-propen-1-one were stoichiometric inactivators of the enzyme and ease of turnover of the parent latent Michael acceptor depended on the presence of an electron-withdrawing substituent at the para position. A series of steroidal and nonsteroidal epoxides in which the oxiranyl oxygen could be substituted for the 3-ketone (the acceptor carbonyl of a steroid substrate) were also synthesized and evaluated as potential mechanism-based inactivators. Steroidal 2 alpha,3 alpha-, and 3 alpha,4 alpha-epoxides as well as 3 alpha- and 3 beta-spiroepoxides did not bind to the enzyme and were unable to cause enzyme inactivation in either the presence or absence of pyridine nucleotide. In contrast, nitrostyrene oxides produced time-dependent inactivation, the rate of which was governed by the presence of an electron withdrawing group at the para position. These data indicate that the design of mechanism-based inactivators for 3 alpha-hydroxysteroid dehydrogenase requires the incorporation of electron-withdrawing groups adjacent to the latent enzyme-activated group and, as a result, the turnover and/or reactivity of these compounds is increased. Moreover, these compounds can be modeled on nonsteroids.
Penning
T M
TM
Department of Pharmacology, University of Pennsylvania School of Medicine, Philadelphia 19104-6084.
Thronton
R
R
Ricigliano
J W
JW
D015096:3-Hydroxysteroid Dehydrogenases / Q000037:antagonists & inhibitors*; D045002:3-alpha-Hydroxysteroid Dehydrogenase (B-Specific); D001665:Binding Sites; D004789:Enzyme Activation / Q000187:drug effects; D004852:Epoxy Compounds / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D007659:Ketones / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D007700:Kinetics; D015394:Molecular Structure; D013256:Steroids / Q000138:chemical synthesis / Q000737:chemistry / Q000494:pharmacology*; D013329:Structure-Activity Relationship
D004852:Epoxy Compounds; D007659:Ketones; D013256:Steroids; D015096:3-Hydroxysteroid Dehydrogenases; D045002:3-alpha-Hydroxysteroid Dehydrogenase (B-Specific)