Mammalian mutagenicity tests

Two mammalian mutagenicity tests are generally required to support the lack of mutagenic or carcinogenic potential. Some well known tests are... 

A similar dynamic appears in this suggestion that mammalian mutagenicity testing be made part of the standard repertoire of toxicology screens [M]uta-... 

Bartsch, H., Malaveille, C., Camus, A.M., Martel-Planche, G., Brun, G., Hautefeville, A., Sabadie, N., Barbin, A., Kuroki, T., Drevon, C., Piccoli, C. and Montesano, R. Bacterial and mammalian mutagenicity tests ... 

A slight mutagenic effect of noni juice extract was observed in the Salmonella microsome assay in strain TA1537 but not in strains TA98 or TAIOO. The activity was attributed to the presence of flavonoids (Westendorf et al. 2007). No mutagenicity of noni juice was observed in the mammalian mutagenicity test with V79 Chinese hamster fibroblasts (Westendorf et al. 2007). Rats treated with a noni juice concentrate did not show DNA repair synthesis in primary rat hepatocytes, nor could DNA adducts or DNA strand breaks be observed (Westendorf et al. 2007). 

It must be emphasized that the dominant lethal assay should be applied together with other in vivo- mammalian mutagenicity tests, notably in vivo cytogenetics, karyotype analysis of bone marrow, and the host-mediated assay, in addition to ancillary submammalian tests. Such mammalian systems can be simply and practically incorporated in the course of routine toxicity testing. 

Cadmium compounds are not mutagenic in classical short term test systems, but rather exert clastogenic activity in mammalian cells. Thus, in most bacterial assays water soluble cadmium compounds were not mutagenic, and in standard mammalian mutagenicity tests effects of cadmium salts with respect to point mutations were usually weak and/or restricted to comparatively high concentrations. In contrast. 

Although in vitro mutagenicity tests suggest that some nitrofurans in general provoke a positive response, use of in vivo mammalian systems has produced equivocal or negative results. 

Physicochemical properties requked include melting/boiling point, vapor pressure, solubiUty, and flammabiUty/explosion characteristics. The toxicological studies include acute toxicity tests, oral, inhalation, and dermal skin and eye kritation skin sensiti2ation subacute toxicity, oral, inhalation, and dermal and mutagenicity tests. In vitro reverse mutation assay (Ames test) on Salmonella typhimurium and/or E.scherichia coli and mammalian cytogenic test. In vivo mouse micronucleus test. 

Ames B.N., McCann J. Yamasaki E. (1975) Methods for detecting carcinogens and mutagens with the Salmonella/mammalian microsome mutagenicity test. MutatRes, 31, 347-364. 

Ciba-Geigy Ltd. 1978a. Salmonella/mammalian-microsome mutagenicity test with TK 10 509 (REOFOS 95). Experiment No. 78-2506. Ciba-Geigy Limited, Basle, Switzerland. NTIS OTS0507280. 

Preliminary Cytotoxicity Testing. An essential first step is to carry out a preliminary study to evaluate the toxicity of the test material to the indicator cells, under the conditions of the main mutagenicity test. When selecting dose levels, the solubility of the test compound, the resulting pH of the media, and the osmolality of the test solutions all need to be considered. The latter two parameters have been known to induce false positive effects in in vitro mammalian tests (Brusick, 1986). The experimental procedure is carried out as follows. 

Chinese Hamster CHO/Hgprt System. Chinese hamster ovary (CHO) cells have 21 or 22 chromosomes with one intact X chromosome and a large acrocentric marker chromosome (Natarajan and Obe, 1982). The use of these cells in mammalian mutation experiments was first reported by Hsie et al. (1975), and was refined into a quantitative assay for mutagenicity testing by O Neill. The performance of this system has been reviewed by the USA EPA Gene-Tox Program. The experimental procedure for this assay is similar to the V79/Hgprt system already described, and for more detailed descriptions the reader is referred to Li et al. (1987). 

In vivo mammalian mutation tests are not restricted to germ cell tests. The mouse spot test described below is, again, a test used first for studying radiation-induced mutation but has also been used for screening chemicals for in vivo mutagenic potential. This test has had several proponents but compared with in vivo chromosomal assays is not widely used. 

The in vitro cytogenetic assay is a short-term mutagenicity test for detecting chromosomal damage in cultured mammalian cells. 

Arlett, C.E, Smith, D.M., Clark, G.M., Green, J.H.L., Cole, J., McGregor, D.B. and Asquith, J.C. (1989). Mammalian cell assays based upon colony formation. In UKEMS Subcommittee on Guidelines for Mutagenicity Testing. Report Part III Statistical Evaluation of Mutagenicity Test Data, (Kirkland, D.J., Ed.). Cambridge University Press, pp. 66-101. 

Dean, B.J. and Danford, N. (1984). Assays for the detection of chemically induced chromosome damage in cultures mammalian cells. In Mutagenicity Testing. A Practical Approach, (Venitt, S. and Parry, J.M., Eds.). IRL Press, Oxford, pp. 187-232. 

Natarajan, A.T. and Obe, G. (1982). Mutagenicity testing with cultured mammalian cells cytogenetic assays. In Mutagenicity, New Horizons in Genetic Toxicology, (Heddle, J.A., Ed.). Academic Press, New York, pp. 172-213. 

Mueller, D., Nelles, J., Deparde, E. and Ami, P. (1980). The activity of S-9 liver fractions from seven species in salmonella/mammalian-microsome mutagenicity tests. Mutat. Res. 70 279-300. 

Stolzenberg SJ, Mine CH. 1980. Mutagenicity of 2- and 3-carbon halogenated compounds in the Salmonella/mammalian-microsome test. Environ Mutagen 2 59-66. 

There has also been much discussion of the interpretation of the finding of hepatocellular carcinomas and adenomas in mice in the NTP (1987) study. There was a higher than usual rate of these tumors in control female mice. Because 1,4-dichlorobenzene has not been demonstrated to be mutagenic in any of the microbial or mammalian systems tested, NTP (1987) has suggested that it may act as a tumor promoter by inducing DNA replication for tissue repair processes. As discussed previously, oral administration of... 

Ames, B. N., McCann, J. and Yamasaki, E., Methods for Detecting Carcinogens and Mutagens with the Salmonella/Mammalian Microsome Mutagenicity Test , Mutat. Res. (1975) 31, 347. 

An extensive database has demonstrated that many chemicals that are positive in this test also exhibit mutagenic activity in other tests. There are, however, examples of mutagenic substances, which are not detected by this test reasons for these shortcomings can be ascribed to the specific nature of the endpoint detected, differences in metabolic activation, or differences in bioavailability. On the other hand, factors which enhance the sensitivity of the bacterial reverse mutation test can lead to an overestimation of mutagenic activity. The bacterial reverse mutation test may not be appropriate for the evaluation of certain classes of chemicals for example, highly bactericidal compounds (e.g., certain antibiotics) and those which are thought (or known) to interfere specifically with the mammalian cell replication system (e.g., some topoisomerase inhibitors and some nucleoside analogues). In such cases, mammalian mutation tests may be more appropriate. 

Before administration of a NME to man, a mutagenicity test in bacterial cells (Ames test), with and without metabolic activation, and tests for chromosomal aberrations in mammalian cells should be negative. Any positive or equivocal results will require additional tests to be performed before proceeding to man. Studies of embryo-foetal toxicity should be performed before administration of a NME to women of reproductive potential. Studies of fertility, early embryonic development and pre- and post-natal development are not required at this stage of development neither are carcinogenicity studies. 

Mammalian Microsome Mutagenicity test. The TWA breathing-zone concentrations for nitrosamines taken from six workers donating... 

There are now many different in vitro mutagenicity tests employing different strains of bacteria and also in vivo tests designed to detect mutations in higher organisms and mammalian systems. It is beyond the scope of this book to discuss these, but details can be found in other texts. 

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