title: Concordance between trifluoroacetic acid and hepatic protein trifluoroacetylation after disulfiram inhibition of halothane metabolism in rats.
abstract: BACKGROUND
  Cytochrome P4502E1(CYP2E1)-mediated oxidation of halothane to a reactive intermediate (trifluoroacyl chloride) that covalently binds to hepatic proteins forming trifluoroacetylated neoantigens is believed to be the initiating event in a complex immunologic cascade culminating in antibody formation and severe hepatic necrosis ('halothane hepatitis') in susceptible patients. Trifluoroacyl chloride may also hydrolyze to the stable metabolite trifluoroacetic acid (TFA). CYP2E1 inactivation by disulfiram or its primary metabolite, diethyldithiocarbamate, inhibits human halothane oxidation to TFA in vitro and in vivo. Nevertheless, disulfiram effects on hepatic protein trifluoroacetylation by halothane in vivo are unknown. This investigation tested the hypotheses that disulfiram prevents halothane-dependent protein trifluoroacetylation in vivo, and that TFA represents a biomarker for hepatic protein trifluoroacetylation.
  
  
  METHODS
  Rats were pretreated with isoniazid (CYP2E1 induction), isoniazid followed by disulfiram (CYP2E1 inhibition), or nothing (controls), then anesthetized with halothane or nothing (controls). Plasma and urine TFA were quantified by ion HPLC; hepatic microsomal TFA-proteins were analyzed by Western blot.
  
  
  RESULTS
  CYP2E1 induction increased both TFA and TFA-protein formation compared with uninduced halothane-treated rats. Disulfiram, even after CYP2E1 induction, nearly abolished both TFA and TFA-protein formation. Pretreatments similarly affected both TFA and TFA-protein formation across all groups.
  
  
  CONCLUSIONS
  Disulfiram inhibition of CYP2E1-mediated halothane oxidation prevents hepatic protein trifluoroacetylation. Based on the concordance between TFA and TFA-protein formation, TFA appears to be a valid biomarker for TFA-protein formation. Disulfiram inhibition of human halothane oxidation in vivo, previously assessed by diminished TFA formation, probably also confers inhibition of hepatic TFA-protein formation.
authors.affiliation: Department of Anesthesiology, University of Washington, Seattle, WA 98195, USA.
authors.forename: D K
authors.initials: DK
authors.lastname: Spracklin
authors.forename: M E
authors.initials: ME
authors.lastname: Emery
authors.forename: K E
authors.initials: KE
authors.lastname: Thummel
authors.forename: E D
authors.initials: ED
authors.lastname: Kharasch
mesh_terms: D018685:Anesthetics, Inhalation / Q000378:metabolism*; D000818:Animals; D015153:Blotting, Western; D001965:Bromides / Q000378:metabolism; D002851:Chromatography, High Pressure Liquid; D065691:Cytochrome P-450 CYP2E1 Inhibitors; D004221:Disulfiram / Q000494:pharmacology*; D004790:Enzyme Induction / Q000187:drug effects; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006221:Halothane / Q000378:metabolism*; D007538:Isoniazid / Q000494:pharmacology; D008099:Liver / Q000187:drug effects / Q000378:metabolism*; D008297:Male; D010084:Oxidation-Reduction; D011506:Proteins / Q000378:metabolism*; D051381:Rats; D017207:Rats, Sprague-Dawley; D014269:Trifluoroacetic Acid / Q000378:metabolism*
chemical_list: D018685:Anesthetics, Inhalation; D001965:Bromides; D065691:Cytochrome P-450 CYP2E1 Inhibitors; D004791:Enzyme Inhibitors; D011506:Proteins; D014269:Trifluoroacetic Acid; D004221:Disulfiram; D006221:Halothane; D007538:Isoniazid