PT - JOURNAL ARTICLE AU - Chungang Gu AU - Markus Artelsmair AU - Charles S Elmore AU - Richard J Lewis AU - Patty Davis AU - James E Hall AU - Bruce T Dembofsky AU - Greg Christoph AU - Mark A Smith AU - Marc Chapdelaine AU - Maria Sunzel TI - Late-occurring and long-circulating metabolites of GABA<sub>Aα2,3</sub> receptor modulator AZD7325 involving metabolic cyclization and aromatization: relevance to MIST analysis and application for patient compliance AID - 10.1124/dmd.117.078873 DP - 2018 Jan 01 TA - Drug Metabolism and Disposition PG - dmd.117.078873 4099 - http://dmd.aspetjournals.org/content/early/2018/01/04/dmd.117.078873.short 4100 - http://dmd.aspetjournals.org/content/early/2018/01/04/dmd.117.078873.full AB - AZD7325 [4-amino-8-(2-fluoro-6-methoxyphenyl)-N-propylcinnoline-3-carboxamide] is a selective GABAAα2,3 receptor modulator intended for the treatment of anxiety disorders through oral administration. An interesting metabolic cyclization and aromatization pathway led to the tricyclic core of M9, i.e. 2-ethyl-7-(2-fluoro-6-methoxyphenyl)pyrimido[5,4-c]cinnolin-4(3H)-one. Further oxidative metabolism generated M10 via O-demethylation and M42 via hydroxylation. An authentic standard of M9 was synthesized to confirm the novel structure of M9 and that of M10 and M42 by liver microsomal incubation of the M9 standard. Metabolites M9, M10 and M42 were either minor or absent in plasma samples after a single dose, however, all became major metabolites in human and preclinical animal plasma after repeated doses, and circulated in humans longer than 48 h after the end of seven repeated doses. The absence of these long circulating metabolites from selected patients' plasma samples was used to demonstrate patient non-compliance as the cause of unexpected lack of drug exposure in some patients during a Phase IIb outpatient clinical study. The observation of late-occurring and long-circulating metabolites demonstrates the need to collect plasma samples at steady state after repeated doses when conducting metabolite analysis for the safety testing of drug metabolites. All 12 major non-conjugate metabolites of AZD7325 observed in human plasma at steady state were also observed in dog, rat, and mouse plasma samples collected from 3-month safety studies and at higher exposures in the animals than humans. This eliminated concern about human specific or disproportional metabolites.