Use of Freund's Complete Adjuvant (FCA) in inflammatory pain models: consequences on the metabolism and pharmacokinetics of the non-peptidic delta receptor agonist SNC80 in the rat.

This study was initiated to characterize the metabolism and pharmacokinetics of SNC80 in rats and to evaluate the impact of Freund's complete adjuvant (FCA)-induced inflammation on its body disposition. In vitro, the disappearance and intrinsic clearance (CL(int)) of SNC80 were measured following incubations in recombinant rat CYPs and in phenotyped liver microsomes from naive and 24-h FCA-treated rats. The unbound fraction (f(u)) was assessed by ultrafiltration. Based on the Clint values, in vivo blood clearance of 3.35 and 2.48 L/h/kg were predicted in naive and FCA-treated rats. In vivo, SNC80 was administered to naive and 24-h FCA-treated rats at 10 micromol/kg i.v. and 50 micromol/kg p.o. The naive animals showed high plasma clearance (3.1 L/h/kg), low renal clearance (<0.02 L/h/kg) and poor bioavailability (<4%). Following i.v. administration, plasma clearance was lower (22%) in FCA-treated vs. untreated rats. Despite the decreases in f(u) (approximately 30%) and CL(int) (approximately 40%) in vitro, in vivo the apparent bioavailability and oral clearance were not significantly different between FCA-treated and naive rats. Hepatic and possibly intestinal losses contribute to the low bioavailability of SNC80. Non-hepatic mechanisms may compensate for the decrease in plasma clearance found in FCA-treated rats, preventing an increase in the oral bioavailability of SNC80.

Identification of CYP3A4 and CYP2C8 as the major cytochrome P450 s responsible for morphine N-demethylation in human liver microsomes.

1. The aim was to identify the cytochrome P450 (CYP) enzymes responsible for the N-demethylation of morphine in vitro. 2. In human liver microsomes, normorphine formation followed Michaelis-Menten kinetics with mean Km and Vmax of 12.4 +/- 2.2 mM and 1546 +/- 121 pmol min(-1) mg(-1), respectively. In microsomes from a panel of 14 human livers phenotyped for 10 CYP enzymes, morphine N-demethylation correlated with testosterone 6beta-hydroxylation (r=0.91, p<0.001) and paclitaxel 6-alpha hydroxylation (r=0.72, p<0.001), two specific markers of CYP3A4 and CYP2C8, respectively. Normorphine formation decreased when incubated in the presence of troleandomycin or quercetin (by 46 and 33-36%, respectively), which further corroborates the contribution of CYP3A4 and CYP2C8. 3. Among eight recombinant human CYP enzymes tested, CYP3A4 and CYP2C8 exhibited the highest intrinsic clearance. More than 90% of morphine N-demethylation could be accounted for via the action of both CYP3A4 and CYP2C8. 4. The in vitro findings suggest that hepatic CYP3A4, and to a lesser extent CYP2C8, play an important role in the metabolism of morphine into normorphine.

New diarylmethylpiperazines as potent and selective nonpeptidic delta opioid receptor agonists with increased In vitro metabolic stability.

Nonpeptide delta opioid agonists are analgesics with a potentially improved side-effect and abuse liability profile, compared to classical opioids. Andrews analysis of the NIH nonpeptide lead SNC-80 suggested the removal of substituents not predicted to contribute to binding. This approach led to a simplified lead, N, N-diethyl-4-[phenyl(1-piperazinyl)methyl]benzamide (1), which retained potent binding affinity and selectivity to the human delta receptor (IC(50) = 11 nM, mu/delta = 740, kappa/delta > 900) and potency as a full agonist (EC(50) = 36 nM) but had a markedly reduced molecular weight, only one chiral center, and increased in vitro metabolic stability. From this lead, the key pharmacophore groups for delta receptor affinity and activation were more clearly defined by SAR and mutagenesis studies. Further structural modifications on the basis of 1 confirmed the importance of the N, N-diethylbenzamide group and the piperazine lower basic nitrogen for delta binding, in agreement with mutagenesis data. A number of piperazine N-alkyl substituents were tolerated. In contrast, modifications of the phenyl group led to the discovery of a series of diarylmethylpiperazines exemplified by N, N-diethyl-4-[1-piperazinyl(8-quinolinyl)methyl]benzamide (56) which had an improved in vitro binding profile (IC(50) = 0.5 nM, mu/delta = 1239, EC(50) = 3.6 nM) and increased in vitro metabolic stability compared to SNC-80.

N,N-Diethyl-4-(phenylpiperidin-4-ylidenemethyl)benzamide: a novel, exceptionally selective, potent delta opioid receptor agonist with oral bioavailability and its analogues.

The design, synthesis, and pharmacological evaluation of a novel class of delta opioid receptor agonists, N, N-diethyl-4-(phenylpiperidin-4-ylidenemethyl)benzamide (6a) and its analogues, are described. These compounds, formally derived from SNC-80 (2) by replacing the piperazine ring with a piperidine ring containing an exocyclic carbon carbon double bond, were found to bind with high affinity and exhibit excellent selectivity for the delta opioid receptor as full agonists. 6a, the simplest structure in the class, exhibited an IC(50) = 0.87 nM for the delta opioid receptors and extremely high selectivity over the mu receptors (mu/delta = 4370) and the kappa receptors (kappa/delta = 8590). Rat liver microsome studies on a selected number of compounds show these olefinic piperidine compounds (6) to be considerably more stable than SNC-80. This novel series of compounds appear to interact with delta opioid receptors in a similar way to SNC-80 since they demonstrate similar SAR. Two general approaches have been established for the synthesis of these compounds, based on dehydration of benzhydryl alcohols (7) and Suzuki coupling reactions of vinyl bromide (8), and are herewith reported.