Regiospecific and stereospecific triangulation of the structures of metabolites formed by sequential metabolism at multiple prochiral centers.
Structures of in vivo secondary metabolites of a norbornane-containing drug candidate with multiple prochiral centers were triangulated, in a regio- and stereospecific fashion, using in vitro metabolism data from synthetic primary metabolites and in vivo metabolism data from the separate administration of a radiolabeled primary metabolite, [(14)C]-(S)-2-((1R,2S,4R,5S)-5-hydroxybicyclo[2.2.1]heptan-2-ylamino)-5-isopropyl-5-methylthiazol-4(5H)-one (M1). A mass balance study on the 11β hydroxysteroid dehydrogenase type 1 enzyme inhibitor [(14)C]-(S)-2-((1S,2S,4R)-bicyclo[2.2.1]heptan-2-ylamino)-5-isopropyl-5-methylthiazol-4(5H)-one (AMG 221) in rats was dosed at 2 mg/kg. Radioactivity was excreted mainly in urine. Metabolites of AMG 221 were quantified by high-performance liquid chromatography with radiometric detection and characterized by liquid chromatography-tandem mass spectrometry (LC-MS/MS). LC-MS/MS revealed at least 38 metabolites. Seven monohydroxylated metabolites mediated formation of the other 31 metabolites. Twenty-eight metabolites were identified regio- and stereo-specifically. Little parent drug was observed in urine or feces. Monohydroxy metabolite M1 was the major metabolite comprising 17 to 24% of excreted dose, and seven monohydroxy metabolites comprised 29 (male) and 37% (female) of dose. Of 11 quantifiable isobaric dihydroxy metabolites that comprised 8.3 (male) and 24% (female) of dose, 10 were identified regio- and stereospecifically by triangulation. A single trihydroxy metabolite comprised approximately 10% of dose. Complex secondary metabolism of drugs with multiple prochiral centers can be elucidated in a regio- and stereospecific fashion without NMR through synthesis and in vitro and in vivo studies on the metabolism of chiral primary oxidation products.
Pharmacokinetics and Drug Metabolism, Amgen Inc., 1201 Amgen Court West, Seattle, Washington 98119, USA.
Robert J
RJ
Greene
John A
JA
Davis
Raju
R
Subramanian
Molly R
MR
Deane
Maurice G
MG
Emery
J Greg
JG
Slatter
D043205:11-beta-Hydroxysteroid Dehydrogenase Type 1 / Q000037:antagonists & inhibitors; D000284:Administration, Oral; D000818:Animals; D001646:Bile / Q000737:chemistry; D001711:Biotransformation; D002851:Chromatography, High Pressure Liquid; D004285:Dogs; D004791:Enzyme Inhibitors / Q000097:blood / Q000737:chemistry* / Q000378:metabolism* / Q000652:urine; D005243:Feces / Q000737:chemistry; D005260:Female; D006900:Hydroxylation; D008297:Male; D008862:Microsomes, Liver / Q000378:metabolism; D015394:Molecular Structure; D010084:Oxidation-Reduction; D051381:Rats; D017207:Rats, Sprague-Dawley; D012737:Sex Factors; D013045:Species Specificity; D013237:Stereoisomerism; D053719:Tandem Mass Spectrometry; D013844:Thiazoles / Q000097:blood / Q000737:chemistry* / Q000378:metabolism* / Q000652:urine
C541878:AMG 221; D004791:Enzyme Inhibitors; D013844:Thiazoles; D043205:11-beta-Hydroxysteroid Dehydrogenase Type 1