4-Dehydrodebrisoquine , a Novel Debrisoquine Metabolite Formed from 4-Hydroxydebrisoquine That Affects the CYP 2 D 6 Metabolic Ratio

Considerable unexplained intersubject variability in the debrisoquine metabolic ratio (urinary debrisoquine/4-hydroxydebrisoquine) exists within individual CYP2D6 genotypes. We speculated that debrisoquine was converted to as yet undisclosed metabolites. Thirteen healthy young volunteers, nine CYP2D6*1 homozygotes [extensive metabolizers (EMs)] and four CYP2D6*4 homozygotes [poor metabolizers (PMs)] took 12.8 mg of debrisoquine hemisulfate by mouth and collected 0to 8and 8to 24-h urines, which were analyzed by gas chromatography-mass spectrometry (GCMS) before and after treatment with -glucuronidase. Authentic 3,4-dehydrodebrisoquine was synthesized and characterized by GCMS, liquid chromatography-tandem mass spectrometry, and H NMR. 3,4-Dehydrodebrisoquine is a novel metabolite of debrisoquine excreted variably in 0to 24-h urine, both in EMs (3.1–27.6% of dose) and PMs (0–2.1% of dose). This metabolite is produced from 4-hydroxydebrisoquine in vitro by human and rat liver microsomes. A previously unstudied CYP2D6*1 homozygote was administered 10.2 mg of 4-hydroxydebrisoquine orally and also excreted 3,4-dehydrodebrisoquine. EMs excreted 6-hydroxydebrisoquine (0–4.8%) and 8-hydroxydebrisoquine (0–1.3%), but these phenolic metabolites were not detected in PM urine. Debrisoquine and 4-hydroxydebrisoquine glucuronides were excreted in a highly genotype-dependent manner. A microsomal activity that probably does not involve cytochrome P450 participates in the further metabolism of 4-hydroxydebrisoquine, which we speculate may also lead to the formation of 1and 3-hydroxy-debrisoquine and their ringopened products. In conclusion, this study suggests that the traditional metabolic ratio is not a true measure of the debrisoquine 4-hydroxylation capacity of an individual and thus may, in part, explain the wide intragenotype variation in metabolic ratio. Debrisoquine (3,4-dihydro-2(1H)-isoquinoline carboxamidine) sulfate was patented in the United States by Hoffmann-La Roche in 1964 (Wenner, 1964) and immediately went into clinical trials as an antihypertensive agent (Talbot, 1965; Rosendorff et al., 1968; Somers et al., 1968; Blechman et al., 1969). Surprisingly, at the time of its launch, little, if anything was known about the metabolic disposition of debrisoquine. Workers at Hoffmann-La Roche UK (Allen et al., 1975) reported on a study in which C-labeled debrisoquine was administered to rats (50 mg/kg) and a single hypertensive patient (2.6 mg, on top of 15 mg q.d.s. therapeutic dose). Debrisoquine was excreted unchanged in urine of both human and rat, together with 4-hydroxydebrisoquine as the major metabolite, and traces of the phenolic metabolites 5-, 6-, 7-, and 8-hydroxydebrisoquine. In addition, both rats and humans excrete 10 to 15% of the dose as two ring-opened acidic metabolites, presumed to arise from hydroxylation of debrisoquine in positions 1 and 3 (Fig. 1A). The nature of these metabolites was subsequently confirmed (Allen et al., 1976; Eiermann et al., 1998). For the rat, 70% of the administered radioactivity could be accounted for as debrisoquine and these seven aforementioned metabolites. In the single patient who was studied, 66% of the radioactivity was similarly accounted for (Allen et al., 1975). It was noted that there was considerable variation (7-fold) in the extent of urinary excretion of unchanged debrisoquine in 10 volunteers given 40 mg of debrisoquine sulfate by mouth and that urinary debrisoquine excretion correlated positively with hypotensive response to debrisoquine (Angelo et al., 1975). This same group performed a second C-debrisoquine study in four human volunteers, for which 71.3 to 76.7% of the radioactivity was recovered in the urine within 24 h (Angelo et al., 1976). These authors identified debrisoquine and 4-hydroxydebrisoquine in human urine, and tentatively Supported by the National Cancer Institute Intramural Research Program (F.J.G.). J.R.I. is grateful to U.S. Smokeless Tobacco Company for a grant for collaborative research. Article, publication date, and citation information can be found at http://dmd.aspetjournals.org. doi:10.1124/dmd.105.008920. ABBREVIATIONS: MR, metabolic rate; EM, extensive metabolizer; IM, intermediate metabolizer; P450, cytochrome P450; PM, poor metabolizer; UM, ultrarapid metabolizer; GCMS, gas chromatography-mass spectrometry; LC-MS/MS, liquid chromatography-tandem mass spectrometry; UPLC-TOFMS, ultra-performance liquid chromatography-coupled time-of-flight mass spectrometry. 0090-9556/06/3409-1563–1574 DRUG METABOLISM AND DISPOSITION Vol. 34, No. 9 U.S. Government work not protected by U.S. copyright 8920/3134741 DMD 34:1563–1574, 2006 Printed in U.S.A. 1563 at A PE T Jornals on Sptem er 0, 2017 dm d.aspurnals.org D ow nladed from