title: Report on stage III Pig-a mutation assays using benzo[a]pyrene.
abstract: Genotoxicity assays were conducted on rats treated with benzo[a]pyrene (BaP) as part of Stage III of a validation study on the Pig-a gene mutation assay. Assays were performed at the U.S. FDA-NCTR and Bayer-Germany. Starting on Day 1, groups of five 6- to 7-week-old male Fischer 344 (F344, used at FDA-NCTR) and Han Wistar rats (Bayer) were given 28 daily doses of 0, 37.5, 75, or 150 mg/kg BaP; blood was sampled on Days -1, 4, 15, 29, and 56. Pig-a mutant frequencies were determined on Days -1, 15, 29, and 56 in total red blood cells (RBCs) and reticulocytes (RETs) as RBC(CD59-) and RET(CD59-) frequencies; percent micronucleated-RETs (%MN-RET) were measured on Days 4 and 29. RBC(CD59-) and RET(CD59-) frequencies increased in a dose- and time-dependent manner, producing significant increases by Day 29 in both rat models. The responses for RETs were stronger than those for RBCs, and the responses in F344 rats were stronger than in Han Wistar rats. BaP also produced significant increases in %MN-RET frequency at Days 4 and 29, with the responses being greater in F344 than Han Wistar rats. The overall findings were consistent with those of the reference laboratory using Han Wistar rats. Finally, mutation assays performed on splenocytes from Day 56 F344 rats indicated that BaP mutant frequencies were three to fivefold higher for the Hprt gene than the Pig-a gene. The results indicate that the Pig-a RET and RBC assays are reproducible, transferable, and show promise for integrating gene mutation into 28-day repeat-dose studies.
authors: Division of Genetic and Molecular Toxicology, National Center for Toxicological Research, U.S. Food and Drug Administration, Jefferson, Arkansas 72079, USA.
authors: Javed A
authors: JA
authors: Bhalli
authors: Joseph G
authors: JG
authors: Shaddock
authors: Mason G
authors: MG
authors: Pearce
authors: Vasily N
authors: VN
authors: Dobrovolsky
authors: Xuefei
authors: X
authors: Cao
authors: Robert H
authors: RH
authors: Heflich
authors: Hans-Werner
authors: HW
authors: Vohr
mesh_terms: D000818:Animals; D001564:Benzo(a)pyrene / Q000633:toxicity*; D018957:CD59 Antigens / Q000235:genetics; D002138:Calibration; D020552:Comet Assay / Q000379:methods / Q000592:standards; D003627:Data Interpretation, Statistical; D004305:Dose-Response Relationship, Drug; D023381:Endpoint Determination; D004912:Erythrocytes / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D007041:Hypoxanthine Phosphoribosyltransferase / Q000235:genetics; D007391:International Cooperation; D007753:Laboratories / Q000592:standards; D008297:Male; D008565:Membrane Proteins / Q000235:genetics*; D015162:Micronucleus Tests / Q000379:methods / Q000592:standards; D009152:Mutagenicity Tests / Q000379:methods / Q000592:standards; D009153:Mutagens / Q000633:toxicity*; D009154:Mutation; D051381:Rats; D012015:Reference Standards; D015203:Reproducibility of Results; D012156:Reticulocytes / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D018570:Risk Assessment; D013045:Species Specificity; D013601:T-Lymphocytes / Q000187:drug effects / Q000648:ultrastructure; D013997:Time Factors
chemical_list: D018957:CD59 Antigens; D008565:Membrane Proteins; D009153:Mutagens; C080086:phosphatidylinositol glycan-class A protein; D001564:Benzo(a)pyrene; D007041:Hypoxanthine Phosphoribosyltransferase