Selective inactivation by 21-chlorinated steroids of rabbit liver and adrenal microsomal cytochromes P-450 involved in progesterone hydroxylation.
The inactivation by 21-chlorinated steroids of rabbit liver cytochromes P-450 involved in the hydroxylation of progesterone has been investigated in intact microsomes encompassing two phenotypes of 21-hydroxylase activity, two phenotypes of 16 alpha-hydroxylase activity, and three phenotypes of 6 beta-hydroxylase activity. In liver microsomes from outbred New Zealand White male rabbits exhibiting a high content of cytochrome P-450 1, 21,21-dichloropregnenolone caused a time- and NADPH-dependent loss of 21-hydroxylase activity. This loss of activity exhibited a number of characteristics of mechanism-based inactivation, including irreversibility, saturation with increasing inhibitor concentrations, and protection by substrate, and was also documented with purified P-450 1 in a reconstituted system. 21,21-Dichloropregnenolone caused no time-dependent loss of 6 beta-hydroxylase activity in microsomes from the New Zealand White rabbits or from control or rifampicin-treated rabbits of the inbred B/J strain. In contrast, in the microsomes from the B/J rabbits, some inactivation of the 16 alpha-hydroxylase was observed (k = 0.04 min-1), regardless of the rifampicin treatment. The other two compounds tested, 21-chloropregnenolone and 21,21-dichloroprogesterone, were less effective than the dichloropregnenolone as inactivators of cytochrome P-450 1. On the other hand, 21,21-dichloroprogesterone, but not 21,21-dichloropregneolone, caused a rapid time-dependent loss of 21-hydroxylase activity in rabbit adrenal microsomes. The results indicate that the introduction of a dichloromethyl group into a substrate bearing a methyl group normally hydroxylated by only one or a few forms of cytochrome P-450 may be a rational means of designing selective inhibitors of the enzyme.
Halpert
J
J
Department of Pharmacology and Toxicology, College of Pharmacy, University of Arizona, Tucson 85721.
Jaw
J Y
JY
Balfour
C
C
Mash
E A
EA
Johnson
E F
EF
D000311:Adrenal Glands / Q000201:enzymology*; D000818:Animals; D001189:Aryl Hydrocarbon Hydroxylases; D065607:Cytochrome P-450 Enzyme Inhibitors; D003577:Cytochrome P-450 Enzyme System / Q000378:metabolism*; D006900:Hydroxylation; D007700:Kinetics; D008297:Male; D008861:Microsomes / Q000201:enzymology; D008862:Microsomes, Liver / Q000201:enzymology*; D009249:NADP / Q000494:pharmacology; D011284:Pregnenolone / Q000031:analogs & derivatives / Q000494:pharmacology; D011374:Progesterone / Q000031:analogs & derivatives / Q000378:metabolism* / Q000494:pharmacology; D011817:Rabbits; D012293:Rifampin / Q000494:pharmacology; D039181:Steroid 16-alpha-Hydroxylase; D013255:Steroid 21-Hydroxylase / Q000037:antagonists & inhibitors; D013250:Steroid Hydroxylases / Q000037:antagonists & inhibitors; D013256:Steroids / Q000494:pharmacology*
D065607:Cytochrome P-450 Enzyme Inhibitors; D013256:Steroids; C056730:21,21-dichloroprogesterone; C056731:21,21-dichloropregnenolone; C056732:21-chloropregnenolone; D011374:Progesterone; D009249:NADP; D011284:Pregnenolone; D003577:Cytochrome P-450 Enzyme System; D013250:Steroid Hydroxylases; D001189:Aryl Hydrocarbon Hydroxylases; D039181:Steroid 16-alpha-Hydroxylase; C022657:steroid hormone 6-beta-hydroxylase; D013255:Steroid 21-Hydroxylase; D012293:Rifampin