Evaluation of a multi-endpoint assay in rats, combining the bone-marrow micronucleus test, the Comet assay and the flow-cytometric peripheral blood micronucleus test
Evaluation of a multi-endpoint assay in rats, combining the bone-marrow micronucleus test, the Comet assay and the flow-cytometric peripheral blood micronucleus test
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DOI：
10.1016/j.mrgentox.2011.02.009
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发表时间：
2011-05-18
影响因子：
1.9
通讯作者：
Kirkland, David J.
中科院分区：
文献类型：
--
作者：
Bowen, Damian E.;Whitwell, James H.;Kirkland, David J.
随着修订的ICH指南草案(ICH S2草案)的发布，有可能建立一种联合的多终点体内检测方法，以减少对体内多项检测的需求，从而减少动物的时间、成本和使用。本文介绍了一项评估试验的结果，在该试验中，对发育中的红细胞或年轻的网织红细胞进行了微核试验(观察潜在的染色体断裂和整个染色体的丢失)，并结合彗星试验(测量DNA链断裂)、胃、肝脏和血淋巴细胞。这使得可以评估各种潜在的靶组织(接触部位、新陈代谢部位和外周分布)的DNA损伤。对8种体内基因毒素(2-乙酰氨基荧苯、苯并[a]芘、多菌灵、环磷酰胺、二甲基亚硝胺、甲磺酸乙酯、乙基亚硝脲和丝裂霉素C)进行了测试，已知它们通过不同的作用模式(直接和间接作用的碎裂剂、烷化剂、基因诱变剂、交联剂和非致癌化合物)发挥作用。分别于给药后0、24、45h给雄性大鼠，末次给药后3h取骨髓、外周血(微核终点)、肝、全血、胃(彗星终点)。根据常规(急性)独立分析的现有数据，彗星和微核反应与预期一致。所有化合物至少在一个终点被检测到遗传毒性。对某些化学品(苯并[a]芘和2-乙酰氨基荧烯)的彗星终点和某些其他化学品(多菌灵和丝裂霉素C)的微核终点的独特积极反应突显了评估这两个终点的重要性。这些调查产生的数据证明了多终点设计的适用性。(C)2011爱思唯尔B.V.保留所有权利。
With the publication of revised draft ICH guidelines (Draft ICH S2), there is scope and potential to establish a combined multi-end point in vivo assay to alleviate the need for multiple in vivo assays, thereby reducing time, cost and use of animals.Presented here are the results of an evaluation trial in which the bone-marrow and peripheral blood (via MicroFlow (R) flow cytometry) micronucleus tests (looking at potential chromosome breakage and whole chromosome loss) in developing erythrocytes or young reticulocytes were combined with the Comet assay (measuring DNA strand-breakage), in stomach, liver and blood lymphocytes. This allowed a variety of potential target tissues (site of contact, site of metabolism and peripheral distribution) to be assessed for DNA damage. This combination approach was performed with minimal changes to the standard and regulatory recommended sampling times for the stand-alone assays.A series of eight in vivo genotoxins (2-acetylaminofluorene, benzo[a]pyrene, carbendazim, cyclophosphamide, dimethylnitrosamine, ethyl methanesulfonate, ethyl nitrosourea and mitomycin C), which are known to act via different modes of action (direct- and indirect-acting clastogens, alkylating agents, gene mutagens, cross-linking and aneugenic compounds) were tested. Male rats were dosed at 0,24 and 45 h, and bone marrow and peripheral blood (micronucleus endpoint), liver, whole blood and stomach (Comet endpoint) were sampled at three hours after the last dose. Comet and micronucleus responses were as expected based on available data for conventional (acute) stand-alone assays.All compounds were detected as genotoxic in at least one of the endpoints. The importance of evaluating both endpoints was highlighted by the uniquely positive responses for certain chemicals (benzo[a]pyrene and 2-acetylaminofluorene) with the Comet endpoint and certain other chemicals (carbendazim and mitomycin C) with the micronucleus endpoint.The data generated from these investigations demonstrate the suitability of the multi-endpoint design. (C) 2011 Elsevier B.V. All rights reserved.