title: Evaluation of in vivo gene mutation with etoposide using Pig-a and PIGRET assays.
abstract: The Pig-a assay detects the expression of the endogenous phosphatidylinositol glycan anchor biosynthesis, class A gene (Pig-a) as a reporter of mutations and shows promise for detecting mutations in vivo. This assay requires two to four weeks to detect mutations in erythrocytes after animals are administered a single dose of test compound. In contrast, the more recently developed PIGRET assay using reticulocytes detects mutation sensitively one week after dosing, which is an advantage for conducting short-term genotoxicity studies in vivo. As part of the Japanese Environmental Mutagen Society/Mammalian Mutagenicity Study Group collaborative study, we conducted Pig-a and PIGRET assays to detect Pig-a mutations in rats treated with etoposide. The DNA topoisomerase II-inhibitor, etoposide, causes DNA cleavage and subsequent protein-linked DNA double-strand breaks, induces chromosomal aberrations and micronuclei, and is classified as 2A (probably carcinogenic to humans) by IARC. Etoposide was intravenously given once to 7-week old SD rats at doses of 0, 5, 10, and 20mg/kg. Severe myelosuppression was induced 2days after dosing in all dosing groups. We conducted Pig-a mutant analysis in Pig-a and PIGRET assays 1, 2, and 4 weeks after dosing. Neither Pig-a nor PIGRET assays showed any statistically significant increases in Pig-a mutant frequency in any of the dose groups at any of the sampling times. It was concluded that etoposide was judged negative in Pig-a and PIGRET assays of bone marrow cells, and the results of the two assays showed hardly any differences.
authors.affiliation: Drug Safety Research Laboratories, Astellas Pharma Inc. 21, Miyukigaoka, Tsukuba-shi, Ibaraki 305-8585, Japan. Electronic address: mika.yamamoto@astellas.com.
authors.forename: Mika
authors.initials: M
authors.lastname: Yamamoto
authors.affiliation: Drug Safety Research Laboratories, Astellas Pharma Inc. 21, Miyukigaoka, Tsukuba-shi, Ibaraki 305-8585, Japan.
authors.forename: Akihiro
authors.initials: A
authors.lastname: Wakata
mesh_terms: D000818:Animals; D004912:Erythrocytes / Q000187:drug effects*; D005047:Etoposide / Q000633:toxicity*; D008565:Membrane Proteins / Q000235:genetics*; D009152:Mutagenicity Tests / Q000379:methods*; D009153:Mutagens / Q000633:toxicity*; D009154:Mutation; D051381:Rats; D012156:Reticulocytes / Q000187:drug effects*
chemical_list: D008565:Membrane Proteins; D009153:Mutagens; C080086:phosphatidylinositol glycan-class A protein; D005047:Etoposide
keywords: Genotoxicity; In vivo mutation assay; Pig-a gene