Virtual cofactors for an Escherichia coli nitroreductase enzyme: Relevance to reductively activated prodrugs in antibody directed enzyme prodrug therapy (ADEPT)

• Published in: Biochemical pharmacology 1995-05, Vol.49 (11), p.1641-1647
• Main Authors: Knox, Richard J., Friedlos, Frank, Jarman, Michael, Davies, Lawrence C., Goddard, Phyllis, Anlezark, Gillian M., Melton, Roger G., Sherwood, Roger F.
• Format: Article
• Language: English
• Subjects: Animals; Antibodies; Aziridines - metabolism; bioactivation; Biological and medical sciences; bioreduction; CB 1954; Escherichia coli; Escherichia coli - enzymology; General pharmacology; Medical sciences; Mice; NAD - pharmacokinetics; NADH; NADPH; Nitroreductases - isolation & purification; Nitroreductases - metabolism; nitroreduction; Oxidation-Reduction; Pharmacology. Drug treatments; Physicochemical properties. Structure-activity relationships; Prodrugs; Pyridinium Compounds - chemical synthesis; Pyridinium Compounds - pharmacokinetics; Ribonucleosides - pharmacokinetics; Vitamin K - metabolism
• ISSN: 0006-2952; 1873-2968
• DOI: 10.1016/0006-2952(95)00077-D

A nitroreductase enzyme has been isolated from Escherichia coli that has the unusual property of being equally capable of using either NADH or NADPH as a cofactor for the reduction of its substrates which include menadione as well as 5-(aziridin-1-yl)-2,4-dinitrobenzamide (CB 1954). This property is shared with the mammalian enzyme, DT diaphorase. The nitroreductase can, like DT diaphorase, also use simple reduced pyridinium compounds as virtual cofactors. The intact NAD(P)H molecule is not required and the simplest quaternary (and therefore reducible) derivative of nicotinamide, 1-methylnicotinamide (reduced), is as effective as NAD(P)H in its ability to act as an electron donor for the nitroreductase. The structure-activity relationship is not identical to that of DT diaphorase and nicotinic acid riboside (reduced) is selective, being active only for the nitroreductase. Irrespective of the virtual cofactor used, the nitroreductase formed the same reduction products of CB 1954 (the 2-and 4-hydroxylamino derivatives in equal proportions). Nicotinic acid riboside (reduced), unlike NADH, was stable to metabolism by serum enzymes and had a plasma half-life of seven minutes in the mouse after an i.v. bolus administration. NADH had an unmeasurably short half-life. Nicotinic acid riboside (reduced) could also be produced in vivo by administration of nicotinic acid 5′- O-benzoyl riboside (reduced). These results demonstrate that the requirement for a cofactor need not be a limitation in the use of reductive enzymes in antibody directed enzyme prodrug therapy (ADEPT). It is proposed that the E. coli nitroreductase would be a suitable enzyme for ADEPT in combination with CB 1954 and a synthetic, enzyme-selective, virtual cofactor such as nicotinic acid riboside (reduced).