title: Human CYP2E1 mediates the formation of glycidamide from acrylamide.
abstract: Regarding the cancer risk assessment of acrylamide (AA) it is of basic interest to know, as to what amount of the absorbed AA is metabolized to glycidamide (GA) in humans, compared to what has been observed in laboratory animals. GA is suspected of being the ultimate carcinogenic metabolite of AA. From experiments with CYP2E1-deficient mice it can be concluded that AA is metabolized to GA primarily by CYP2E1. We therefore examined whether CYP2E1 is involved in GA formation in non-rodent species with the focus on humans by using human CYP2E1 supersomes, marmoset and human liver microsomes and in addition, genetically engineered V79 cells expressing human CYP2E1 (V79h2E1 cells). Special emphasis was placed on the analytical detection of GA, which was performed by gas chromatography/mass spectrometry. The results show that AA is metabolized to GA in human CYP2E1 supersomes, in marmoset and human liver microsomes as well as in V79h2E1 cells. The activity of GA formation is highest in supersomes; in human liver it is somewhat higher than in marmoset liver. A monoclonal CYP2E1 human selective antibody (MAB-2E1) and diethyldithiocarbamate (DDC) were used as specific inhibitors of CYP2E1. The generation of GA could be inhibited by MAB-2E1 to about 80% in V79h2E1 cells and to about 90% in human and marmoset liver microsomes. Also DDC led to an inhibition of about 95%. In conclusion, AA is metabolized to GA by human CYP2E1. Overall, the present work describes (1) the application and refinement of a sensitive methodology in order to determine low amounts of GA, (2) the applicability of genetically modified V79 cell lines in order to investigate specific questions concerning metabolism and (3) the involvement, for the first time, of human CYP2E1 in the formation of GA from AA. Further studies will compare the activities of GA formation in genetically engineered V79 cells expressing CYP2E1 from different species.
authors.affiliation: Center for Experimental Toxicology, Federal Institute for Risk Assessment, Thielallee 88-92, 14195 Berlin, Germany.
authors.forename: Eva
authors.initials: E
authors.lastname: Settels
authors.forename: Ulrike
authors.initials: U
authors.lastname: Bernauer
authors.forename: Richard
authors.initials: R
authors.lastname: Palavinskas
authors.forename: Horst S
authors.initials: HS
authors.lastname: Klaffke
authors.forename: Ursula
authors.initials: U
authors.lastname: Gundert-Remy
authors.forename: Klaus E
authors.initials: KE
authors.lastname: Appel
mesh_terms: D020106:Acrylamide / Q000378:metabolism* / Q000633:toxicity; D000328:Adult; D000818:Animals; D001711:Biotransformation; D002144:Callithrix; D002273:Carcinogens / Q000378:metabolism* / Q000633:toxicity; D002460:Cell Line; D002753:Chlorzoxazone / Q000378:metabolism; D019392:Cytochrome P-450 CYP2E1 / Q000235:genetics / Q000378:metabolism*; D065691:Cytochrome P-450 CYP2E1 Inhibitors; D004050:Ditiocarb / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004852:Epoxy Compounds / Q000378:metabolism* / Q000633:toxicity; D005260:Female; D008401:Gas Chromatography-Mass Spectrometry; D006801:Humans; D006900:Hydroxylation; D007700:Kinetics; D008862:Microsomes, Liver; D008875:Middle Aged; D011994:Recombinant Proteins / Q000378:metabolism; D018570:Risk Assessment; D013045:Species Specificity; D013379:Substrate Specificity; D014162:Transfection
chemical_list: D002273:Carcinogens; D065691:Cytochrome P-450 CYP2E1 Inhibitors; D004791:Enzyme Inhibitors; D004852:Epoxy Compounds; D011994:Recombinant Proteins; D020106:Acrylamide; C071834:glycidamide; D004050:Ditiocarb; D019392:Cytochrome P-450 CYP2E1; D002753:Chlorzoxazone