Universität zu Köln

Schiffer, Lina ORCID: 0000-0001-8540-4861, Brixius-Anderko, Simone ORCID: 0000-0002-3036-9285, Hannemann, Frank ORCID: 0000-0001-7991-9033, Zapp, Josef, Neunzig, Jens, Thevis, Mario and Bernhardt, Rita (2016). Metabolism of Oral Turinabol by Human Steroid Hormone-Synthesizing Cytochrome P450 Enzymes. Drug Metab. Dispos., 44 (2). S. 227 - 238. BETHESDA: AMER SOC PHARMACOLOGY EXPERIMENTAL THERAPEUTICS. ISSN 1521-009X

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Abstract

The human mitochondrial cytochrome P450 enzymes CYP11A1, CYP11B1, and CYP11B2 are involved in the biosynthesis of steroid hormones. CYP11A1 catalyzes the side-chain cleavage of cholesterol, and CYP11B1 and CYP11B2 catalyze the final steps in the biosynthesis of gluco-and mineralocorticoids, respectively. This study reveals their additional capability to metabolize the xenobiotic steroid oral turinabol (OT; 4-chlor-17 beta-hydroxy-17 alpha-methylandrosta-1,4-dien-3-on), which is a common doping agent. By contrast, microsomal steroid hydroxylases did not convert OT. Spectroscopic binding assays revealed dissociation constants of 17.7 mu M and 5.4 mu M for CYP11B1 and CYP11B2, respectively, whereas no observable binding spectra emerged for CYP11A1. Catalytic efficiencies of OT conversion were determined to be 46 min(-1) mM(-1) for CYP11A1, 741 min(-1) mM(-1) for CYP11B1, and 3338 min(-1) mM(-1) for CYP11B2, which is in the same order of magnitude as for the natural substrates but shows a preference of CYP11B2 for OT conversion. Products of OT metabolism by the CYP11B subfamily members were produced at a milligram scale with a recombinant Escherichia coli-based whole-cell system. They were identified by nuclear magnetic resonance spectroscopy to be 11 beta-OH-OT for both CYP11B isoforms, whereby CYP11B2 additionally formed 11 beta, 18-diOH-OT and 11 beta-OH-OT-18-al, which rearranges to its tautomeric form 11 beta, 18-expoxy-18-OH-OT. CYP11A1 produces six metabolites, which are proposed to include 2-OH-OT, 16-OH-OT, and 2,16-diOH-OT based on liquid chromatography-tandem mass spectrometry analyses. All three enzymes are shown to be inhibited by OT in their natural function. The extent of inhibition thereby depends on the affinity of the enzyme for OT and the strongest effect was demonstrated for CYP11B2. These findings suggest that steroidogenic cytochrome P450 enzymes can contribute to drug metabolism and should be considered in drug design and toxicity studies.

Item Type:	Journal Article
Creators:	Creators Email ORCID ORCID Put Code Schiffer, Lina UNSPECIFIED orcid.org/0000-0001-8540-4861 UNSPECIFIED Brixius-Anderko, Simone UNSPECIFIED orcid.org/0000-0002-3036-9285 UNSPECIFIED Hannemann, Frank UNSPECIFIED orcid.org/0000-0001-7991-9033 UNSPECIFIED Zapp, Josef UNSPECIFIED UNSPECIFIED UNSPECIFIED Neunzig, Jens UNSPECIFIED UNSPECIFIED UNSPECIFIED Thevis, Mario UNSPECIFIED UNSPECIFIED UNSPECIFIED Bernhardt, Rita UNSPECIFIED UNSPECIFIED UNSPECIFIED
URN:	urn:nbn:de:hbz:38-285901
DOI:	10.1124/dmd.115.066829
Journal or Publication Title:	Drug Metab. Dispos.
Volume:	44
Number:	2
Page Range:	S. 227 - 238
Date:	2016
Publisher:	AMER SOC PHARMACOLOGY EXPERIMENTAL THERAPEUTICS
Place of Publication:	BETHESDA
ISSN:	1521-009X
Language:	English
Faculty:	Unspecified
Divisions:	Unspecified
Subjects:	no entry
Uncontrolled Keywords:	Keywords Language ALDOSTERONE SYNTHASE INHIBITORS; CHOLESTEROL SIDE-CHAIN; ESCHERICHIA-COLI; SUBSTRATE-SPECIFICITY; ADRENODOXIN REDUCTASE; ELECTRON-TRANSPORT; ANABOLIC-STEROIDS; HUMAN PLASMA; EXPRESSION; PURIFICATION Multiple languages Pharmacology & Pharmacy Multiple languages
Refereed:	Yes
URI:	http://kups.ub.uni-koeln.de/id/eprint/28590