Molecular and Cellular Pharmacology
Stereoselective Biotransformation of the Selective Serotonin Reuptake Inhibitor Citalopram and Its Demethylated Metabolites by Monoamine Oxidases in Human Liver

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Abstract

Citalopram (CIT) is an antidepressive drug of the group of selective serotonin reuptake inhibitors (SSRIs). The tertiary amine CIT is given as a racemic drug, but its pharmacological activity resides mainly in S-CIT. CIT is metabolised by cytochrome P450 (CYP) to N-demethylcitalopram (DCIT) and N-didemethylcitalopram (DDCIT). The citalopram propionic acid derivative (CIT-PROP) is another, but pharmacologically inactive, metabolite, the formation of which has been poorly characterised but is postulated to occur by deamination of CIT, DCIT and/or DDCIT. The aim of the present investigation was to study the formation of the enantiomers of CIT-PROP from CIT and its two N-demethylated metabolites, DCIT and DDCIT, in an in vitro incubation system (microsomal and cytosolic fractions) obtained from human livers. The production of CIT-PROP was measured by a stereospecific HPLC method. Incubation of rac-CIT, rac-DCIT and rac-DDCIT (500 μM each, separately) in the presence (or absence) of NADP showed that CIT-PROP formation was substrate-dependent and essentially NADP-independent. Monoamine oxidases (MAO) type A and B and aldehyde oxidase were identified as the probable enzymes involved in the formation of CIT-PROP from CIT, DCIT and DDCIT. Indeed, the irreversible monoamine oxidase type A inhibitor clorgyline and the irreversible monoamine oxidase type B inhibitor selegiline (both at 0.5 μM in the incubation mixture) inhibited CIT-PROP formation, depending on the substrate, up to 70% and 88%, respectively. The participation of aldehyde oxidase in the subsequent step is suggested by the inhibition caused by menadione (50 μM) in CIT-PROP formation. Preliminary experiments suggest the presence of four unknown metabolites, probably products of deamination, which were detected in plasma and urine samples of patients treated with CIT as well as in in vitro biotransformations. Their presence confirms the importance of deamination in the biotransformation of CIT and its demethylated metabolites, especially in the brain where, in contrast to the liver, the role of cytochrome P450 appears to be low.

Section snippets

Chemicals

The following drugs were gifts: The racemates and enantiomers (purity >99%) of CIT hydrobromide or oxalate, DCIT hydrochloride or fumarate, DDCIT L-tartrate or fumarate, citalopram N-oxide (CIT-N-oxide) and CIT-PROP (Lundbeck AS, Copenhagen, Denmark), the IS S-Flurbi (Boots Limited, Nottingham, U.K.), ketoconazole (Prof. P. Dayer, Geneva, Switzerland), and omeprazole (Astra, Dietikon, Switzerland).

CIT, its metabolites and S-Flurbi were dissolved in methanol at a concentration of 1 mg/mL of base

Production of CIT-PROP from CIT, DCIT and DDCIT

In preliminary experiments, S-enantiomers of CIT, DCIT and DDCIT were incubated for 120 min in the presence of an NADPH-regenerating system (NADP). CIT-PROP formation by the human liver microsomal fraction was observed for all substrates. S-CIT-PROP production was linear at least until 60 min and with microsomal protein concentrations ranging from 50 to 200 μg with or without cytosolic proteins (100 μg) in the incubation mixture (data not shown). The cytosolic fraction alone was unable to

Discussion

Several clinical pharmacokinetic studies showed the presence of CIT-PROP in the blood of patients treated with CIT (for a review, cf. 8, 27). This study demonstrates the stereoselective in vitro formation of CIT-PROP from the three racemic amines CIT, DCIT and DDCIT in human liver extracts (Table 1). CIT-PROP production was highest using a mixture of microsomal and cytosolic fractions. In the presence of NADP, deamination was observed with substrate selectivity and stereoselectivity, the S/R

Overview and Perspectives

Taking into account that CYP content in human brain is very low 33, 34, 35, in situ deamination of CIT and its demethylated metabolites by MAO should be the main biotransformation in this target organ. It could be very interesting to investigate more intensively this biochemical pathway from a pharmacokinetic as well as pharmacodynamic point of view.

No data are available concerning CIT brain concentrations under therapeutic conditions. It is, however, of interest that chronic oral

Acknowledgements

This study was financed by the Fonds national suisse de la recherche scientifique, FNSRS (32-27579.89, 32-42-42076.94), by the OFES (COST B1), the Lundbeck Foundation, and the Société Académique Vaudoise. We are grateful to Drs. Claus Selch Larsen and Nils Mørk (Lundbeck, Copenhagen, Denmark) and to Dr. Marianne Reymond, CHUV, Lausanne, Switzerland, for providing us with the enantiomers of CIT and of its metabolites, and for rat brain tissues, respectively. We thank Christine Beedham for her

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