Variation of Hepatic Methotrexate 7‐Hydroxylase Activity in Animals and Humans

Abstract

SummaryThis study deals with individual and species variations in the converting activity of methotrexate (MTX) to 7-hydroxymethotrexate in animals and humans. When MTX 7-hydroxylase was assayed in six human liver cytosols, a 48-fold range of intersubject variation of the activity was observed. The variations were correlated to the concentrations of aldehyde oxidase activity in human subjects assayed with benzaldehyde as a substrate. Species differences of liver MTX 7-hydroxylase activity were also observed. The activity was highest in rabbits, followed by rats, hamsters, and monkeys but was undetectable in dogs. Strain differences of MTX 7-hydroxylase activity based on aldehyde oxidase activity were also observed in rats and mice. The results suggest that aldehyde oxidase functions as MTX 7-hydroxylase in livers of animals and humans, and the observed differences of MTX 7-hydroxylase activity are due to variations in the amount of aldehyde oxidase present. IUBMB Life, 48: 607-611, 1999

Mammalian aldehyde oxidases: genetics, evolution and biochemistry

“…This was clearly demonstrated in studies performed in rats, where marked strain and age differences in the amounts of 7-OH-MTX excreted were observed and correlated with aldehyde oxidase activity [55,122]. A similar observation was made in human livers, where there is a high degree of inter-individual variability as to methotrexate hydroxylation and this correlates with aldehyde oxidase levels [123]. 6-mercaptopurine (6-MT) is an immunosuppressant and a cytotoxic drug used for the treatment of pediatric and adult neoplastic disorders.…”

“…When methotrexate 7-hydroxylase was assayed in six human liver cytosols, a 48-fold range of inter-subject variation was observed. The variation correlated with the concentrations of aldehyde oxidase activity using benzaldehyde as a substrate [123]. Development of a non-invasive method for the measurement of aldehyde oxidase in vivo [138] lies at the basis of a further study aimed at defining age-specific and inter-individual variations in the human population.…”

“…More than two log differences between the highest activity in the and the lowest activity strain were observed [120] . Differences in the 7-hydroxylation of methotrexate and benzaldehyde by aldehyde oxidase were observed in liver cytosols of different rat strains [43,123] . Among six Wistar strains, the Slc :Wistar rats showed exceptionally low oxidation activity that was comparable to that of the F344/DuCrj strain [138,141] .…”

Construction of expression system of rabbit aldehyde oxidase cDNA for the clarification of species differences

“…Rabbit AO protein was expressed in E. coli together with monkey AO. The oxidation activities by the expressed enzymes satisfactorily reproduced the results seen in liver cytosols (16,19). These were a) the enormously high oxidation activities of rabbit AO towards cinchonidine and methotrexate compared to those of monkey and b) a few-fold higher activities of monkey AO towards (S)-RS-8359 and vanillin compared to those of rabbit.…”

“…However, ferricyanide and ethylenediaminetetraacetic acid were not added, because they were proved to be no effective on the enzyme activity itself. Methotrexate 7-hydroxylase activity was determined using the methods reported by Kitamura et al (16) other than use of an incubation time of 5 min and acetanilide as an internal standard. The oxidation activity of vanillin was measured by the method described by Panoutsopoulos et al (24), but the reaction was stopped by the addition of methanol solution of p-nitrobenzyl alcohol (internal standard) and the detection wavelength was 260 nm.…”