SR 4233 or
WIN 59075 (3-amino-1,2,4-benzotriazine-1,4-dioxide) is a novel and highly selective hypoxic cell
cytotoxin requiring reductive bioactivation for its impressive antitumour effects. Expression of appropriate
reductases will contribute to therapeutic selectivity. Here we provide more detailed information on the role of
cytochrome P450 and
cytochrome P450 reductase in
SR 4233 reduction by mouse liver microsomes. Reduction of
SR 4233 to the mono-N-
oxide SR 4317 (3-amino-1,2,4-benzotriazine-1-oxide) is
NADPH,
enzyme and
hypoxia dependent. An inhibitory antibody to
cytochrome P450 reductase decreased the microsomal
SR 4233 reduction rate by around 20%. Moreover, studies with purified rat
cytochrome P450 reductase showed unequivocally that this
enzyme was able to catalyse
SR 4233 reduction at a rate of 20-30% of that for microsomes with equivalent P450
reductase activity. Exposure to the specific
cytochrome P450 inhibitor
carbon monoxide (CO) inhibited microsomal reduction by around 70% and CO plus
reductase antibody blocked essentially all activity. Additional confirmation of
cytochrome P450 involvement was provided by the use of other P450
ligands: beta-diethylaminoethyl diphenylpropylacetate hydrochloride gave a slight stimulation while
aminopyrine, n-
octylamine and
2,4-dichloro-6-phenylphenoxyethylamine were inhibitory. Induction of
SR 4233 reduction was seen with
phenobarbitone, pregnenalone-16-alpha-carbonitrile and beta-napthoflavone, suggesting that
cytochrome P450 subfamilies IIB, IIC and IIIA may be involved. Since
cytochrome P450 and P450
reductase catalyse roughly 70 and 30%, of mouse liver microsomal
SR 4233 reduction respectively, we propose that expression of these and other
reductases in normal and tumour tissue is likely to be a major factor governing the toxicity and antitumour activity of the
drug.