title: The utility of metabolic activation mixtures containing human hepatic post-mitochondrial supernatant (S9) for in vitro genetic toxicity assessment.
abstract: In vitro genotoxicity assessment routinely employs an exogenous metabolic activation mixture to simulate mammalian metabolism. Activation mixtures commonly contain post-mitochondrial liver supernatant (i.e. S9) from chemically induced Sprague Dawley rats. Although Organization for Economic Cooperation and Development (OECD) test guidelines permit the use of other S9 preparations, assessments rarely employ human-derived S9. The objective of this study is to review and evaluate the use of human-derived S9 for in vitro genetic toxicity assessment. All available published genotoxicity assessments employing human S9 were compiled for analysis. To facilitate comparative analyses, additional matched Ames data using induced rat liver S9 were obtained for certain highly cited chemicals. Historical human and induced rat S9 quality control reports from Moltox were obtained and mined for enzyme activity and mutagenic potency data. Additional in vitro micronucleus data were experimentally generated using human and induced rat S9. The metabolic activity of induced rat S9 was found to be higher than human S9, and linked to high mutagenic potency results. This study revealed that human S9 often yields significantly lower Salmonella mutagenic potency values, especially for polycyclic aromatic hydrocarbons, aflatoxin B1 and heterocyclic amines (~3- to 350-fold). Conversely, assessment with human S9 activation yields higher potency for aromatic amines (~2- to 50-fold). Outliers with extremely high mutagenic potency results were observed in the human S9 data. Similar trends were observed in experimentally generated mammalian micronucleus cell assays, however human S9 elicited potent cytotoxicity L5178Y, CHO and TK6 cell lines. Due to the potential for reduced sensitivity and the absence of a link between enzyme activity levels and mutagenic potency, human liver S9 is not recommended for use alone in in vitro genotoxicity screening assays; however, human S9 may be extremely useful in follow-up tests, especially in the case of chemicals with species-specific metabolic differences, such as aromatic amines.
authors.affiliation: Environmental Health Science and Research Bureau, Health Canada, 50 Colombine Driveway, Tunney's Pasture 0803A, Ottawa, Ontario K1A 0K9, Canada, Department of Biology, University of Ottawa, 75 Laurier Ave E, Ottawa, Ontario K1N 6N5, Canada, julie.cox@canada.ca paul.white@hc-sc.gc.ca.
authors.forename: Julie A
authors.initials: JA
authors.lastname: Cox
authors.affiliation: Department of R&D, AstraZeneca, Alderley Park, Macclesfield, Cheshire CW11 3RN, UK and.
authors.forename: Mick D
authors.initials: MD
authors.lastname: Fellows
authors.affiliation: Shonan Research Centre, Takeda Pharmaceutical Company Limited, 26-1, Muraoka-Higashi 2-chome, Fujisawa, Kanagawa 251-8555, Japan.
authors.forename: Tsuneo
authors.initials: T
authors.lastname: Hashizume
authors.affiliation: Environmental Health Science and Research Bureau, Health Canada, 50 Colombine Driveway, Tunney's Pasture 0803A, Ottawa, Ontario K1A 0K9, Canada, Department of Biology, University of Ottawa, 75 Laurier Ave E, Ottawa, Ontario K1N 6N5, Canada, julie.cox@canada.ca paul.white@hc-sc.gc.ca.
authors.forename: Paul A
authors.initials: PA
authors.lastname: White
mesh_terms: D065767:Activation, Metabolic; D000818:Animals; D002273:Carcinogens / Q000633:toxicity; D002460:Cell Line; D002474:Cell-Free System; D004789:Enzyme Activation; D022781:Hepatocytes / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D006801:Humans; D051379:Mice; D015162:Micronucleus Tests; D009152:Mutagenicity Tests / Q000379:methods*; D009153:Mutagens / Q000633:toxicity; D051381:Rats; D012475:Salmonella / Q000187:drug effects / Q000235:genetics
chemical_list: D002273:Carcinogens; D009153:Mutagens