Metabolism of LY654322, a Growth Hormone Secretagogue, to an Unusual Diimidazopyridine Metabolite

• Published in: Drug metabolism and disposition Vol. 39; no. 5; pp. 740 - 749
• Main Authors: BOREL, Anthony G, JONES, Timothy M, BARBUCH, Robert J, JACKSON, David A, KULANTHAIVEL, Palaniappan, MATTIUZ, Edward, KLIMKOWSKI, Valentine J, WHEELER, William J, RENER, Gregory A
• Format: Journal Article
• Language: English
• Published: Bethesda, MD American Society for Pharmacology and Experimental Therapeutics 01.05.2011
• Subjects: Animals; Biological and medical sciences; Dipeptides - blood; Dipeptides - chemistry; Dipeptides - metabolism; Dipeptides - pharmacokinetics; Dipeptides - pharmacology; Dogs; Female; Heterocyclic Compounds, 3-Ring - analysis; Heterocyclic Compounds, 3-Ring - chemistry; Human Growth Hormone - metabolism; Imidazoles - blood; Imidazoles - chemistry; Imidazoles - metabolism; Imidazoles - pharmacokinetics; Imidazoles - pharmacology; Male; Medical sciences; Pharmacology. Drug treatments; Pyridines - analysis; Pyridines - chemistry; Rats; Rats, Inbred F344; Receptors, Ghrelin - agonists; Receptors, Ghrelin - metabolism; Somatotropin; Somatropin; Adenohypophyseal hormone; Metabolite; Neurohypophyseal hormone; Exploration; Pharmacokinetics; Metabolism; Quantitative analysis
• ISSN: 0090-9556; 1521-009X
• DOI: 10.1124/dmd.110.037598

2-Methylalanyl-N-{1-[(1R)-1-(4-fluorophenyl)-1-methyl-2-oxo-2-pyrrolidin-1-ylethyl]-1H-imidazol-4-yl}-5-phenyl-D-norvalinamide (LY654322) was rapidly cleared in rats and dogs by renal excretion of parent and metabolism (oxidative and hydrolytic). Among the metabolites identified in the urine of rats and dogs was M25, which was structurally unusual. Indeed, the characterization of M25 and investigation into its disposition relied on the convergence of diverse analytical methodologies. M25 eluted after the parent on reverse-phase chromatography with an MH(+) at m/z 598 (parent + 35 Da). Given its increased lipophilicity and its mass difference compared with the parent, it was evident that M25 was not a phase 2 conjugate. Subsequent liquid chromatography with multiple-stage tandem mass spectrometry and accurate mass experiments identified the structure of M25 as having two replicates of the 1-(4-fluorophenyl)-1-methyl-2-oxo-2-pyrrolidinyl substructure flanking a central aromatic core of composition C(7)H(3)N(5) that was refractory to fragmentation. Compared with the UV spectrum of the parent (λ(max) = 213 nm), M25 displayed a bathochromic shift (λ(max) = 311 nm), which substantiated extensive conjugation within the central core. Subsequent NMR analysis of M25 isolated from dog urine coupled with molecular modeling revealed the structure to be consistent with a diimidazopyridine core with two symmetrically substituted 1-(4-fluorophenyl)-1-methyl-2-oxo-2-pyrrolidinyl moieties. Using a structural analog with a chromophore similar to M25, LC-UV was used to quantitate M25 and determine its urinary disposition. The formation of M25 appears consistent with hydrolysis of LY654322 to an aminoimidazole, dimerization of the latter with the loss of NH(3), C-formylation, and subsequent ring closure and aromatization with loss of H(2)O.