1. The metabolism of a novel phosphodiesterase-IV inhibitor (V11294) was studied in human liver microsomal and cytosol preparations and in cDNA-expressed human hepatic CYP forms. 2. Human liver microsomes, but not cytosol, catalysed the NADPH-dependent metabolism of V11294 to V10331 (formed by hydroxylation of the cyclopentyl ring), V10332 (N-desethyl V11294) and V11689 (formed by hydroxylation of the isopropyl side chain). In addition, smaller amounts of a secondary metabolite V11690 (which can be formed from either V10332 or V11689) were also produced. 3. Kinetic analysis of V11294 metabolism to V10331, V10332 and V11689 in two preparations of pooled human liver microsomes revealed average K(m) = 2.5, 8.1 and 3.9 micro M, respectively. 4. The metabolism of V11294 was determined with a characterized bank of 16 individual human liver microsomal preparations employing a V11294 substrate concentration of 8 micro M (i.e. approximately the K(m) for V10332 formation and around twice the K(m) for V10331 and V11689 formation). Good correlations (r(2) = 0.570-0.903) were observed between V10331, V10332 and V11689 formation and markers of CYP3A forms. In contrast, poorer correlations (r(2) = 0.0002-0.428) were observed with markers of CYP1A2, CYP2A6, CYP2B6, CYP2C8, CYP2C9, CYP2C19, CYP2D6, CYP2E1 and CYP4A9/11. 5. Using human B-lymphoblastoid cell microsomes containing cDNA-expressed CYP forms, V11294 (8 micro M) was metabolized by cDNA-expressed CYP3A4 to V10331, V10332 and V11689, with lower amounts of V11690 also being formed. Lower rates of V11294 metabolism to some V11294 metabolites were also observed with cDNA-expressed CYP2C9, CYP2C19 and CYP2D6, whereas only very low or undetectable rates of V11294 metabolism were observed with cDNA-expressed CYP1A2, CYP2A6, CYP2B6, CYP2C8 and CYP2E1. 6. The metabolism of V11294 (8 micro M) to V10331, V10332 and V11689 was markedly inhibited by the CYP3A mechanism-based inhibitor troleandomycin. In contrast, V11294 metabolism was not significantly affected by inhibitors of CYP1A2, CYP2C9, CYP2D6 and CYP2E1 or by the CYP2C19 substrate S-mephenytoin. 7. In summary, by correlation analysis, chemical inhibition studies and the use of cDNA-expressed CYPs, V11294 metabolism in human liver to V10331, V10332 and V11689 appears to be primarily catalysed by CYP3A forms.
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[Show abstract][Hide abstract] ABSTRACT: Various phosphodiesterase (PDE) 3,4 and 5 inhibitors have been compared with glucagon for their effectiveness at increasing hepatocyte cAMP, glycogenolysis and gluconeogenesis. Preincubation of isolated hepatocytes with PDE 3 and 4 inhibitors (50 microM) for 2 h induced significant increases in cellular cAMP level. The order of effectiveness was: glucagon (78%), V11294A (42%), rolipram (40%), milrinone (36%), CDP-840 (33%), R(0) 20-1724 (31%), papaverine (27%), isobutylmethylxanthine (28%), isoliquiritigenin (25%), theophylline (22%), and amrinone (22%). The PDE 5 inhibitors dipyridamol and sildenafil had only a slight effect on cAMP levels. Glucose formation was increased as a result of increased glycogenolysis in the following order of effectiveness: glucagon (89%), V11294A (63%), rolipram (61%), milrinone (50%), CDP-840 (46%), R(0) 20-1724 (45%), sildenafil (34%), dipyridamol (31%), papaverine (30%), isobutylmethylxanthine (29%), theophylline (20%), amrinone (20%), and isoliquiritigenin (20%). Rolipram and milrinone, selective PDE 4 and PDE 3 inhibitors respectively, stimulated the gluconeogenesis of alanine, lactate + pyruvate, or fructose in hepatocytes isolated from fasted rats. On the other hand, selective cGMP specific phospodiesterase inhibitors, sildenafil and dipyridamol inhibited alanine-induced gluconeogenesis. All PDE inhibitors increased hepatocyte susceptibility to cyanide toxicity (3-4 fold) which was prevented by fructose whereas PDE 5 inhibitors did not significantly increase hepatocyte susceptibility.
[Show abstract][Hide abstract] ABSTRACT: 3-(3-Cyclopentyloxy-4-methoxy-benzyl)-8-isopropyl-adenine V11294 (1) has been identified as a lead structure, which selectively inhibits human lung PDE4 (436 nM) and is also active in a number of in vitro and in vivo models of inflammation. Here we describe the synthesis and pharmacology of a series of highly potent 8-[(benzyloxy)methyl]-substituted analogues, with potencies in the range 10-300 nM. In addition, several compounds showed interesting PDE4 subtype specificities, for example, the 3-thienyl derivative 5v, which showed 6-10 nM potency at PDE4B, D3, and D5 and a 20- to 200-fold selectivity over A and D2, respectively.
[Show abstract][Hide abstract] ABSTRACT: Cytochrome P450 (CYP) enzymes represent a superfamily of hemoproteins that are involved in the metabolism of a wide variety of endogenous and exogenous compounds. For a given CYP enzyme, kinetic properties of a substrate are usually related to substrate lipophilicity (log P or log D(7.4)). In this review, enzyme kinetic parameters (K(m), V(max), and V(max)/K(m)) of 215 CYP3A4-mediated metabolic reactions of 113 drugs in human liver microsomes were obtained from the literature, and lipophilicity values of the 113 drugs were calculated using the ACD/Labs 8.0 program. A low degree of K(m)- or (V(max)/K(m))-lipophilicity correlation, but no V(max)-lipophilicity correlation, is exhibited for the CYP3A4-mediated reactions. Overall, K(m) decreases, but V(max)/K(m) increases, with increasing substrate lipophilicity, and V(max) appears to be independent of substrate lipophilicity. In other words, a low K(m) generally confers a high V(max)/K(m) ratio for a substrate. The degree of lipophilicity-kinetics correlations is related to both reaction types (or reaction mechanisms) and regiochemical positions (or physicochemical properties) of the reaction groups of the substrates. Among the categorized CYP3A4-mediated reactions, the best lipophilicity-kinetics correlation is achieved for carbon hydroxylation, followed by N-dealkylation. No or little lipophilicity-kinetics correlations are seen for N, S-oxidation and other reactions. Within the hydroxylation group, aliphatic hydroxylation shows the best lipophilicity-kinetics correlation while hydroxylation on a carbon atom adjacent to an aromatic ring does not show any lipophilicity-kinetics correlation. The detailed structural and kinetic data sets of the human liver microsomal CYP3A4-mediated reactions represent a specialized database useful for researchers working in the area of structure-metabolism relationship modeling and analysis.
Current Drug Metabolism 05/2006; 7(3):231-49. DOI:10.2174/138920006776359329 · 2.98 Impact Factor