The metabolism and toxicity of furosemide in the Wistar rat and CD-1 mouse: a chemical and biochemical definition of the toxicophore.
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Williams DP, Antoine DJ, Butler PJ, Jones R, Randle L, Payne A, Howard M, Gardner I, Blagg J, Park BK
The metabolism and toxicity of furosemide in the Wistar rat and CD-1 mouse: a chemical and biochemical definition of the toxicophore.
J Pharmacol Exp Ther. 2007 Sep;322(3):1208-20. doi: 10.1124/jpet.107.125302. Epub 2007 Jun 7.
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- 17556636 [ View in PubMed]
- Abstract
Furosemide, a loop diuretic, causes hepatic necrosis in mice. Previous evidence suggested hepatotoxicity arises from metabolic bioactivation to a chemically reactive metabolite that binds to hepatic proteins. To define the nature of the toxic metabolite, we examined the relationship between furosemide metabolism in CD-1 mice and Wistar rats. Furosemide (1.21 mmol/kg) was shown to cause toxicity in mice, but not rats, at 24 h, without resulting in glutathione depletion. In vivo covalent binding to hepatic protein was 6-fold higher in the mouse (1.57 +/- 0.98 nmol equivalent bound/mg protein) than rat (0.26 +/- 0.13 nmol equivalent bound/mg protein). In vivo covalent binding to mouse hepatic protein was reduced 14-fold by a predose of the cytochrome P450 (P450) inhibitor, 1-aminobenzotriazole (ABT; 0.11 +/- 0.04 nmol equivalent bound/mg protein), which also reduced hepatotoxicity. Administration of [(14)C]furosemide to bile duct-cannulated rats demonstrated turnover to glutathione conjugate (8.8 +/- 2.8%), gamma-ketocarboxylic acid metabolite (22.1 +/- 3.3%), N-dealkylated metabolite (21.1 +/- 2.9%), and furosemide glucuronide (12.8 +/- 1.8%). Furosemide-glutathione conjugate was not observed in bile from mice dosed with [(14)C]furosemide. The novel gamma-ketocarboxylic acid, identified by nuclear magnetic resonance spectroscopy, indicates bioactivation of the furan ring. Formation of gamma-ketocarboxylic acid was P450-dependent. In mouse liver microsomes, a gamma-ketoenal furosemide metabolite was trapped, forming an N-acetylcysteine/N-acetyl lysine furosemide adduct. Furosemide (1 mM, 6 h) became irreversibly bound to primary mouse and rat hepatocytes, 0.73 +/- 0.1 and 2.44 +/- 0.3 nmol equivalent bound/mg protein, respectively, which was significantly reduced in the presence of ABT, 0.11 +/- 0.03 and 0.21 +/- 0.1 nmol equivalent bound/mg protein, respectively. Furan rings are part of new chemical entities, and mechanisms underlying species differences in toxicity are important to understand to decrease the drug attrition rate.
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