The comet assay with multiple mouse organs: Comparison of comet assay results and carcinogenicity with 208 chemicals selected from the IARC monographs and USNTP carcinogenicity database

The cornet assay is a microgel electrophoresis technique for detecting DNA damage at the level of the single cell. When this technique is applied to d etect genotoxicity in experimental animals, the most important advantage is that DNA lesions can be measured in any organ, regardless of the extent of mitotic activity. The purpose of this article is to summarize the in vivo genotoxicity in eight organs of the mouse of 208 chemicals selected from In ternational Agency for Research on Cancer (IARC) Groups 1, 2A, 2B, 3, and 4 , and from the U.S. National Toxicology Program (Nm) Carcinogenicity Databa se, and to discuss the utility of the comet assay in genetic toxicology. Alkylating agents, amides, aromatic amines, azo compounds, cyclic nitro com pounds, hydrazines, halides having reactive halogens, and polycyclic aromat ic hydrocarbons were chemicals showing high positive effects, in this assay . The responses detected reflected the ability of this assay to detect the fragmentation of DNA molecules produced by DNA single strand breaks induced chemically and those derived from alkali-labile sites developed from alkyl ated bases and bulky base adducts, The mouse or rat organs exhibiting incre ased levels of DNA damage were not necessarily the target organs for carcin ogenicity. It was rare, in contrast, for the target organs not to show DNA damage. Therefore, organ-specific genotoxicity was necessary but not suffic ient for the prediction of organ-specific carcinogenicity. It would be expe cted that DNA crosslinkers would be difficult to detect by this assay, beca use of the resulting inhibition of DNA unwinding. The proportion of 10 DNA crosslinkers that was positive, however, was high in the gastrointestinal m ucosa, stomach, and colon, but less than 50% in the liver and lung. It was interesting that the genotoxicity of DNA crosslinkers could be detected in the gastrointestinal organs even though the agents were administered intrap eritoneally. Chemical carcinogens can be classified as genotoxic (Ames test-positive) an d putative nongenotoxic (Ames test-negative) carcinogens. The Ames test is generally used as a first screening method to assess chemical genotoxicity and, has provided extensive information on DNA reactivity. Out of 208 chemi cals studied, 117 are Ames test-positive rodent carcinogens, 43 are Ames te st-negative rodent carcinogens, and 30 are rodent noncarcinogens (which inc lude both Ames test-positive and negative noncarcinogens). High positive re sponse ratio (110/117) for rodent genotoxic carcinogens and a high negative response ratio (6/30) for rodent noncarcinogens were shown in the comet as say. For Ames test-negative rodent carcinogens, less than 50% were positive in the comet assay, suggesting that the assay, which detects DNA lesions, is not suitable for identifying nongenotoxic carcinogens. In the safety eva luation of chemicals, it is important to demonstrate that Ames test-positiv e agents are not genotoxic in vivo. This assay had a high positive response ratio for rodent genotoxic carcinogens and a high negative response ratio for rodent genotoxic noncarcinogens, suggesting that the comet assay can be used to evaluate the in vivo genotoxicity of in vitro genotoxic chemicals. For chemicals whose in vivo genotoxicity has been rested in multiple organ s by the comet assay, published data are summarized with unpublished data a nd compared with relevant genotoxicity and carcinogenicity data. Because it is clear that no single test is capable of detecting all relevan t genotoxic agents, the usual approach should be to carry out a battery of in vitro and vivo tests for genotoxicity. The conventional micronucleus tes t in the hematopoietic system is a simple method to assess in vivo clastoge nicity of chemicals. Its performance is related to whether a chemical reach es the hematopoietic system. Among 208 chemicals studied (including 165 rod ent carcinogens), 54 rodents carcinogens do not induce micronuclei in mouse hematopoietic system despite the positive finding with one or two in vitro tests. Forty-nine of 54 rodent carcinogens that do not induce micronuclei were positive in the comet assay, suggesting that the comet assay can be us ed as a further in vivo test apart from the cytogenetic assays in hematopoi etic cells. In this review, we provide one recommendation for the in vivo c omet assay protocol based on our own data.