title: Measuring reproducibility of dose response data for the Pig-a assay using covariate benchmark dose analysis.
abstract: The reproducibility of the in vivo Pig-a gene mutation test system was assessed across 13 different Japanese laboratories. In each laboratory rats were exposed to the same dosing regimen of N-nitroso-N-ethylurea (ENU), and red blood cells (RBCs) and reticulocytes (RETs) were collected for mutant phenotypic analysis using flow cytometry. Mutant frequency dose response data were analysed using the PROAST benchmark dose (BMD) statistical package. Laboratory was used as a covariate during the analysis to allow all dose responses to be analysed at the same time, with conserved shape parameters. This approach has recently been shown to increase the precision of the BMD analysis, as well as providing a measure of equipotency. This measure of equipotency was used here to demonstrate a reasonable level of interlaboratory reproducibility. Increased reproducibility could have been achieved by increasing the number of cells scored, as this would reduce the number of zero values within the mutant frequency data. Overall, the interlaboratory trial was successful, and these findings support the transferability of the in vivo Pig-a gene mutation assay.
authors.affiliation: Swansea University Medical School, Swansea University, SA2 8PP, United Kingdom. Electronic address: g.johnson@swansea.ac.uk.
authors.forename: George E
authors.initials: GE
authors.lastname: Johnson
authors.affiliation: Drug Development Toxicology, Drug Safety Research Laboratories, Drug Discovery Research Division, Astellas Pharma Inc., 2-1-6, Kashima, Yodogawa-ku, Osaka 532-8514, Japan.
authors.forename: Mika
authors.initials: M
authors.lastname: Yamamoto
authors.affiliation: Gotemba Laboratory, BoZo Research Center Inc., Gotemba-shi, Shizuoka 412-0039, Japan.
authors.forename: Yuta
authors.initials: Y
authors.lastname: Suzuki
authors.affiliation: Preclinical Research Laboratories, Sumitomo Dainippon Pharma Co., Ltd., 3-1-98 Kasugadenaka, Konohana-ku, Osaka 554-0022, Japan.
authors.forename: Hideki
authors.initials: H
authors.lastname: Adachi
authors.affiliation: Toxicology Laboratory, Life Science Research Institute, Kumiai Chemical Industry, Co., Ltd., 3360 Kamo, Kikugawa-shi, Shizuoka 439-0031, Japan.
authors.forename: Takahiro
authors.initials: T
authors.lastname: Kyoya
authors.affiliation: Safety Assessment Department, Nonclinical Research Center, Drug Development Service Segment, LSI Medience Corporation, 14-1 Sunayama, Kamisu-shi, Ibaraki 314-0255, Japan.
authors.forename: Hironao
authors.initials: H
authors.lastname: Takasawa
authors.affiliation: Division of Genetics and Mutagenesis, National Institute of Health Sciences, 1-18-1 Kamiyoga, Setagaya-ku, Tokyo 158-8501, Japan.
authors.forename: Katsuyoshi
authors.initials: K
authors.lastname: Horibata
authors.affiliation: Quality Assurance Division, Safety Science Institute, Suntory Business Expert Limited, 8-1-1 Seikadai, Seika-cho, Soraku-gun, Kyoto 619-0284, Japan.
authors.forename: Eri
authors.initials: E
authors.lastname: Tsutsumi
authors.affiliation: Toxicology Division, The Institute of Environmental Toxicology, 4321, Uchimoriya-machi, Joso-shi, Ibaraki 303-0043, Japan.
authors.forename: Kunio
authors.initials: K
authors.lastname: Wada
authors.affiliation: Drug Safety and Pharmacokinetics Laboratories, Taisho Pharmaceutical Co., Ltd., 1-403, Yoshino-cho, Kita-ku, Saitama-shi 331-9530, Japan.
authors.forename: Ryuta
authors.initials: R
authors.lastname: Kikuzuki
authors.affiliation: Drug Safety Research Laboratories, Pharmaceutical Research Division, Takeda Pharmaceutical Company Limited, 26-1, Muraoka-Higashi 2-chome, Fujisawa, Kanagawa 251-8555, Japan.
authors.forename: Ikuma
authors.initials: I
authors.lastname: Yoshida
authors.affiliation: Pharmaceutical Development Research Laboratories, Teijin Institute for Bio-medical Research, Teijin Pharma Limited, 4-3-2 Asahigaoka, Hino-shi, Tokyo 191-8512, Japan.
authors.forename: Takafumi
authors.initials: T
authors.lastname: Kimoto
authors.affiliation: Toxicology and Pharmacokinetics Laboratories, Pharmaceutical Research Laboratories, Toray Industries Inc., 6-10-1 Tebiro, Kamakura, Kanagawa 248-8555, Japan.
authors.forename: Akihisa
authors.initials: A
authors.lastname: Maeda
authors.affiliation: Yakult Honsha Co., Ltd., 5-11 Izumi, Kunitachi-shi, Tokyo 186-8650, Japan.
authors.forename: Kazunori
authors.initials: K
authors.lastname: Narumi
mesh_terms: D000818:Animals; D004305:Dose-Response Relationship, Drug; D004912:Erythrocytes / Q000187:drug effects; D008565:Membrane Proteins / Q000235:genetics*; D009152:Mutagenicity Tests / Q000379:methods*; D009153:Mutagens / Q000633:toxicity*; D009154:Mutation; D051381:Rats; D015203:Reproducibility of Results; D012156:Reticulocytes / Q000187:drug effects
chemical_list: D008565:Membrane Proteins; D009153:Mutagens; C080086:phosphatidylinositol glycan-class A protein
keywords: Benchmark dose; Dose response; In vivo; OECD; Pig-a; Potency; Ring trial