Human hepatic metabolism of a novel 2-carboxyindole glycine antagonist forstroke: in vitro-in vivo correlations

1. The hepatic metabolism of 3-[-2(phen)-(carbamoyl) ethenyl]-4,6-dichloroi ndole-2-carboxylic acid (GV150526), a novel glycine antagonist for stroke, was investigated. 2. After a single intravenous administration of 800 mg GV150526 to healthy volunteers, sis metabolites were observed. The major metabolites detected i n human plasma have been shown by mass spectrometry to be glucoronides and one sulphate conjugate. 3. After incubation of GV150526 for 6 and 2 lh with human liver slices, thr ee glucuronide metabolites were observed. After incubation of GV150526 with pooled human liver microsomes, only one metabolite was observed, with the same molecular weight and HPLC retention time as the synthetic standard GV2 17053 (GV150526 hydroxylated on the para-position of the phenyl ring). 4. GV150526 hydroxylase enzyme kinetics-a step before sulphation-was determ ined using pooled human microsomes and was shown to be catalysed by cytochr ome P4502C9. Glucuronidation kinetics towards GV150526 using microsomal pre parations were also determined. Glucuronidation of GV150526 was observed wi th UGT1A1 cDNA-expressed protein, but not with UGT1A6. 5. The above enzyme kinetic data were used to calculate intrinsic clearance after scaling-up and hepatic clearance were calculated. Since GV150526 has a high plasma protein binding capacity, the effect of GV150526 binding to microsomal protein was determined. Thus, enzyme kinetic data were corrected , plotting the free (unbound) concentration of GV150526 versus enzymatic ve locities: apparent V-max did not alter significantly but apparent K-m was s imilar to 10-fold lower. Correlation of these corrected enzyme kinetic data to predict clearance with in vivo clearance of GV150526 was good when both fu(plasma) and fu(microsomes) were included in the clearance calculations.