title: Fluorinated analogues as mechanistic probes in valproic acid hepatotoxicity: hepatic microvesicular steatosis and glutathione status.
abstract: It is postulated that the hepatotoxicity of valproic acid (VPA) results from the mitochondrial beta-oxidation of its cytochrome P450 metabolite, 2-propyl-4-pentenoic acid (4-ene VPA), to 2-propyl-(E)-2,4-pentadienoic acid ((E)-2,4-diene VPA) which, in the CoA thioester form, either depletes GSH or produces a putative inhibitor of beta-oxidation enzymes. In order to test this hypothesis, 2-fluoro-2-propyl-4-pentenoic acid (alpha-fluoro-4-ene VPA) which was expected to be inert to beta-oxidative metabolism was synthesized and its effect on rat liver studied in comparison with that of 4-ene VPA. Similarly, the known hepatotoxicant 4-pentenoic acid (4-PA) and 2,2-difluoro-4-pentenoic acid (F2-4-PA) were compared. Male Sprague-Dawley rats (150-180 g, 4 rats per group) were dosed ip with 4-ene VPA (0.7 mmol/kg per day), 4-PA (1.0 mmol/kg per day), or equivalent amounts of their alpha-fluorinated analogues for 5 days. Both 4-ene VPA and 4-PA induced severe hepatic microvesicular steatosis ( > 85% affected hepatocytes), and 4-ene VPA produced mitochondrial alterations. By contrast, alpha-fluoro-4-ene VPA and F2-4-PA were not observed to cause morphological changes in the liver. The major metabolite of 4-ene VPA in the rat urine and serum was the beta-oxidation product (E)-2,4-diene VPA. The N-acetylcysteine (NAC) conjugate of (E)-2,4-diene VPA was also found in the urine. Neither (E)-2,4-diene VPA nor the NAC conjugate could be detected in the rats administered alpha-fluoro-4-ene VPA. In a second set of rats (3 rats per group), total liver GSH levels were determined to be depleted to 56% and 72% of control following doses of 4-ene VPA (1.4 mmol/kg) and equivalent alpha-fluoro-4-ene VPA, respectively. Mitochondrial GSH remained unchanged in the alpha-fluoro-4-ene VPA treated group but was reduced to 68% of control in the rats administered 4-ene VPA. These results strongly support the theory that hepatotoxicity of 4-ene VPA, and possibly VPA itself, is mediated largely through beta-oxidation of 4-ene VPA to reactive intermediates that are capable of depleting mitochondrial GSH.
authors.affiliation: Division of Pharmaceutical Chemistry, Faculty of Pharmaceutical Sciences, University of British Columbia, Vancouver, Canada.
authors.forename: W
authors.initials: W
authors.lastname: Tang
authors.forename: A G
authors.initials: AG
authors.lastname: Borel
authors.forename: T
authors.initials: T
authors.lastname: Fujimiya
authors.forename: F S
authors.initials: FS
authors.lastname: Abbott
mesh_terms: D000818:Animals; D000927:Anticonvulsants / Q000633:toxicity*; D005229:Fatty Acids, Monounsaturated / Q000378:metabolism / Q000633:toxicity*; D005234:Fatty Liver / Q000139:chemically induced* / Q000473:pathology; D008401:Gas Chromatography-Mass Spectrometry; D005978:Glutathione / Q000378:metabolism*; D008099:Liver / Q000737:chemistry / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D008297:Male; D008930:Mitochondria, Liver / Q000378:metabolism; D010421:Pentanoic Acids / Q000378:metabolism / Q000633:toxicity*; D051381:Rats; D017207:Rats, Sprague-Dawley; D014635:Valproic Acid / Q000031:analogs & derivatives / Q000378:metabolism / Q000633:toxicity*
chemical_list: C096093:2,2-difluoro-4-pentenoic acid; C096095:2-fluoro-2-propyl-4-pentenoic acid; D000927:Anticonvulsants; D005229:Fatty Acids, Monounsaturated; D010421:Pentanoic Acids; C045022:2-propyl-4-pentenoic acid; D014635:Valproic Acid; C009785:4-pentenoic acid; D005978:Glutathione