Reduction of fluoromisonidazole, a new imaging agent for hypoxia

Abstract

[ 18F]Fluoromisonidazole (1-(3-[ 18F]fluoro-2-hydroxypropyl)-2-nitroimidazole, [ 18F]FMISO) is a nitroimidazole compound that is being used as a new imaging agent for hypoxia. Because its uptake in hypoxic tissue is dependent on reduction of the nitro group on the imidazole ring, it is necessary to verify the availability of nitroreductase enzymes in a variety of tissues. FMISO reduction was studied using chemical and enzymatic reducing systems and mammalian cells. FMISO reduction by iron/HCl eliminated the absorbance peak at 325 nm caused by the nitro group. FMISO reduction by xanthine oxidase, as measured by a decrease in absorbance at 325 nm, occurred at a rate of 2.4 ± 0.3 nmol/min/ unit enzyme (mean ± SEM, N = 15). This reaction was inhibited by allopurinol. Separation of the parent drug from its reduction product following chemical and enzymatic reductions indicated that iron/ HC1 reduced the majority of the FMISO molecules present, while xanthine oxidase did not. Reduction of FMISO by NADH dehydrogenase could not be demonstrated spectrophotometrically. Measurement of the reduction of FMISO in V79 cells based on the binding of [ 3H]FMISO to cellular macromolecules was performed using a cell suspension in a three-neck flask. Hypoxic V79 cells bound [ 3H]FMISO at the rate of 0.26 ± 0.07 pmol/10 6 cells/min (N = 8). When specific inhibitors of two nitroreductase enzymes and a general inhibitor of electron transport were added to the cell suspension, no consistent, statistically significant inhibition of FMISO binding could be shown. We conclude that while inhibition of FMISO reduction by a purified nitroreductase can be shown, nitroreductase activity in cells is not inhibited so easily. This supports the hypothesis that nitroreductases are plentiful and will not limit the rate of FMISO reduction and uptake in hypoxic tumors or nonmalignant tissues.