Exploration of Catalytic Properties of CYP2D6 and CYP3A4 Through Metabolic Studies of Levorphanol and Levallorphan □ S

ABSTRACT: CYP2D6 and CYP3A4, two members of the cytochrome P450 superfamily of monooxygenases, mediate the biotransformation of a variety of xenobiotics. The two enzymes differ in substrate specificity and size and characteristics of the active site cavity. The aim of this study was to determine whether the catalytic properties of these isoforms, reflected by the differences observed from crystal structures and homology models, could be confirmed with experimental data. Detailed metabolite identification, reversible inhibition, and time-dependent inhibition were examined for levorphanol and levallorphan with CYP2D6 and CYP3A4. The studies were designed to provide a comparison of the orientations of substrates, the catalytic sites of the two enzymes, and the subsequent outcomes on metabolism and inhibition. The metabolite identification revealed that CYP3A4 catalyzed the formation of a variety of metabolites as a result of presenting different parts of the substrates to the heme. CYP2D6 was a poorer catalyst that led to a more limited number of metabolites that were interpreted in terms to two orientations of the substrates. The inhibition studies showed evidence for strong reversible inhibition of CYP2D6 but not for CYP3A4. Levallorphan acted as a time-dependent inhibitor on CYP3A4, indicating a productive binding mode with this enzyme not observed with CYP2D6 that presumably resulted from close interactions of the N-allyl moiety oriented toward the heme. All the results are in agreement with the large and flexible active site of CYP3A4 and the more restricted active site of CYP2D6

Dopamine receptor binding of 4-(4-chlorophenyl)-1-[4-(4-fluorophenyl)-4-oxobutyl]-1,2,3,6-tetrahydropyridine (HPTP), an intermediate metabolite of haloperidol

The neuroleptic agent haloperidol (HP) is biotransformed to metabolites such as 4-(4-chlorophenyl)-1-[4-(4-fluorophenyl)-4-oxobutyl]-1,2,3,6-tetrahydropyridine (HPTP) and 4-(4-chlorophenyl)-1-[4-(4-fluorophenyl)-4-oxobutyl]pyridinium (HPP) In this study, radioligand binding studies were performed using [H]SCH23390 as a dopamine D1 receptor ligand and [H]spiperone as a D2 ligand. K(i) values for D1 receptors were 35.8 μM and 54.9 μM for HP and HPTP, respectively Corresponding values for D2 receptors were 39.1 nM and 329.8 nM. These results indicate similar low affinities in the micromolar range for both HP and HPTP at the dopamine D1 receptor, a much higher affinity of both HP and HPTP for the D2 receptor than for the D1 receptor, and that HPTP binds to D2 receptors with a 9-fold lower affinity than HP. The data are consistent with observations in mice that HPTP is a much less potent acute neuroleptic agent than HP

Clinical and neuropathological abnormalities in baboons treated with HPTP, the tetrahydropyridine analog of haloperidol

Tardive dyskinesia (TD) is relatively common among psychiatric patients on maintenance therapy with typical neuroleptics and persists in more than 20% even after withdrawal of the medication. Such persistence suggests an underlying pathology due to neurotoxicity. We present evidence for such a neurotoxic mechanism in a baboon model of TD. Four baboons were treated chronically with the dehydration product of haloperidol, 4-(4-chlorophenyl)- 1-[4-(4-fluorophenyl)-4-oxobutyl]-1,2,3,6-tetrahydropyridine (HPTP), which is metabolized, similarly to haloperidol, to two neurotoxic pyridinium species. The animals developed orofacial dyskinesia which persisted after HPTP was ceased. Serial sections of the entire brain from the four treated animals and four vehicle-treated controls revealed volume loss in the basal forebrain and hypothalamus. Histological evaluation demonstrated a reduction in the density of magnocellular neurons in the anterior region of the nucleus basalis of Meynert (NbM). We speculate that the loss of these NbM neurons may be associated with the persistent orofacial dyskinesia observed in the HPTP- treated animals. These findings may contribute to a better understanding of neuroleptic-induced TD

Haloperidol and its tetrahydropyridine derivative (HPTP) are metabolized to potentially neurotoxic pyridinium species in the baboon

The in vivo metabolic fate of haloperidol (HP) and its tetrahydropyridine analog HPTP have been examined in the baboon to investigate the formation of potentially neurotoxic pyridinium metabolites that have been observed previously in humans. Urine samples collected from baboons treated with HPTP were shown to contain, in addition to the parent drug, the corresponding reduced HPTP (RHPTP), generated by reduction of the butyrophenone carbonyl group. RHPTP was characterized by comparison with a synthetic standard using HPLC with electrochemical detection and HPLC/MS/MS. Another compound identified by LC/MS/MS was a glucuronide metabolite of RHPTP. The HP pyridinium (HPP) and reduced pyridinium (RHPP) metabolites were shown to be present in urine from both HP and HPTP treated baboons by HPLC using fluorescence detection. The urinary excretion profile of HPP and RHPP in both groups was essentially identical and, in contrast to that observed in rodents, closely paralleled the profile found in humans treated with HP. These data in the baboon suggest that the metabolic processes involved in the production of the pyridinium metabolites of HP are similar to those in humans. Furthermore, the HPTP-treated baboon may be an appropriate model in which to study the role of pyridinium metabolites in the induction of tardive dyskinesia

Metabolic defects caused by treatment with the tetrahydropyridine analog of haloperidol (HPTP), in baboons

Mounting evidence suggests that compromised cellular energy production is a major contributor to idiopathic and drug-induced degenerative processes. Our interest in neurotoxins have prompted us to examine in the baboon the effects of HPTP, the tetrahydropyridine dehydration product of haloperidol, on urinary chemical markers that reflect defects in mitochondrial respiration. Urinary dicarboxylic acid and conjugate profiles, similar to those seen in humans with inborn errors of mitochondrial metabolism and toxin-induced Jamaican vomiting sickness (JVS) were observed in the treated baboons. We interpret these results as evidence that HPTP and/or HPTP metabolites inhibit mitochondrial respiration in the baboon and speculate that analogous effects may occur in haloperidol-treated individuals

Long-term treatment with the tetrahydropyridine analog (HPTP) of haloperidol influences dopamine ligand binding in baboon brain. An [123I]iodobenzamide (IBZM) SPECT study

Haloperidol (HP) and its tetrahydropyridine dehydration product 4-(4-chlorophenyl)-[4-(fluorophenyl)-4-oxobutyl]-1,2,3,6-tetrahydropyridine (HPTP) are both metabolized in vivo to several pyridinium metabolites with potential neurotoxic properties similar to the neurotoxin 1-methyl-4-phenylpyridinium (MPP), a metabolite of the parkinsonian-inducing agent 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine(MPTP). The effect of long-term HPTP treatment on the central nervous system of baboons (Papio ursinus) was studied using 23I]iodobenzamide (IBZM)and single photon emission computed tomography (SPECT) at 1-14 weeks after termination of HPTP treatment. Striatal dopamine receptor binding was measured semiquantitatively by calculating the IBZM count rate ratios of the basal ganglia to frontal cortex and basal ganglia to cerebellum. Relative striatal perfusion was assessed by similar 9mTc-HMPAO thexamethylpropylene amine oxime) ratios. Time activity curves of IBZM from the brain structures suggest that HPTP treatment results in a marked reduction in central dopamine ligand binding, and in particular D2-like receptor binding. Increased washout of the ligand from all the brain structures investigated was seen in the HPTP-treated animals, also consistent with reduced binding. Cerebral blood flow in the control and HPTP-treated groups was similar, indicating that this did not account for the reduced dopamine receptor binding of the BZM ligand. These data suggest that treatment with HPTP induces significant effects on dopamine receptor binding that may contribute to some of the neurological disorders in humans undergoing chronic HP treatment