Mode of action-based risk assessment of genotoxic carcinogens

The risk assessment of chemical carcinogens is one major task in toxicology. Even though exposure has been mitigated effectively during the last decades, low levels of carcinogenic substances in food and at the workplace are still present and often not completely avoidable. The distinction between genotoxic and non-genotoxic carcinogens has traditionally been regarded as particularly relevant for risk assessment, with the assumption of the existence of no-effect concentrations (threshold levels) in case of the latter group. In contrast, genotoxic carcinogens, their metabolic precursors and DNA reactive metabolites are considered to represent risk factors at all concentrations since even one or a few DNA lesions may in principle result in mutations and, thus, increase tumour risk. Within the current document, an updated risk evaluation for genotoxic carcinogens is proposed, based on mechanistic knowledge regarding the substance (group) under investigation, and taking into account recent improvements in analytical techniques used to quantify DNA lesions and mutations as well as “omics” approaches. Furthermore, wherever possible and appropriate, special attention is given to the integration of background levels of the same or comparable DNA lesions. Within part A, fundamental considerations highlight the terms hazard and risk with respect to DNA reactivity of genotoxic agents, as compared to non-genotoxic agents. Also, current methodologies used in genetic toxicology as well as in dosimetry of exposure are described. Special focus is given on the elucidation of modes of action (MOA) and on the relation between DNA damage and cancer risk. Part B addresses specific examples of genotoxic carcinogens, including those humans are exposed to exogenously and endogenously, such as formaldehyde, acetaldehyde and the corresponding alcohols as well as some alkylating agents, ethylene oxide, and acrylamide, but also examples resulting from exogenous sources like aflatoxin B$_{1}$, allylalkoxybenzenes, 2-amino-3,8-dimethylimidazo[4,5-f] quinoxaline (MeIQx), benzo[a]pyrene and pyrrolizidine alkaloids. Additionally, special attention is given to some carcinogenic metal compounds, which are considered indirect genotoxins, by accelerating mutagenicity via interactions with the cellular response to DNA damage even at low exposure conditions. Part C finally encompasses conclusions and perspectives, suggesting a refined strategy for the assessment of the carcinogenic risk associated with an exposure to genotoxic compounds and addressing research needs

Comparative chromosome painting discloses homologous Segments in distantly related mammals

Comparative chromosome painting, termed ZOO-FISH, using DNA libraries from flow sorted human chromosomes 1,16,17 and X, and mouse chromosome 11 discloses the presence of syntenic groups in distantly related mammalian Orders ranging from primates (Homo sapiens), rodents (Mus musculus), even-toed ungulates (Muntiacus muntjak vaginalis and Muntiacus reevesi) and whales (Balaenoptera physalus). These mammalian Orders have evolved separately for 55-80 million years (Myr). We conclude that ZOO-FISH can be used to generate comparative chromosome maps of a large number of mammalian species

A mouse stock with 38 chromosomes derived from the reciprocal translocation T(7;15)33Ad.

A reciprocal translocation, T(7;15)33Ad, with presumed breakpoints in bands 7A1 and 15F3 was induced in late spermatids by injecting male (102/E1 x C3H/E1)F1 mice five times with acrylamide (50 mg/kg body weight). Outcrosses of the original semisterile T(7;15) female generated three males monosomic for the short marker 715 [Ms(715)] among a total of 15 males. The Ms(715) males sired small litters and had reduced testes weights. From inter se matings of Ms(715) animals, nullisomic progeny for chromosome 715 were obtained and mated to produce a breeding stock of mice with 38 chromosomes. For comparison, mice carrying the reciprocal translocation T(4;8), with similarly located breakpoints, were also analyzed. Fluorescent in situ hybridization (FISH) with major and minor satellite DNA probes and a telomeric DNA probe was utilized. The observed FISH signals suggest that in chromosomes 7 and 8 the breaks occurred within the pericentric heterochromatic block immediately below the centromere and in chromosomes 15 and 4 at a point near the distal telomeres. The long markers 157 and 48 are tandem fusion chromosomes. The short markers 715 and 84 also showed all appropriate FISH signals for intact chromosomes. The loss of the small chromosome 715 was compatible with survival, suggesting that no essential genes are located on the small reciprocal translocation product. The development of this tandem fusion stock is described as a laboratory example of one possible step in karyotypic evolution

Telomere Signals in Robertsonian Fusion and Fission Chromosomes: Implications for the Origin of Pseudoaneuploidy.

In situ hybridization with synthetic plant telomeric sequences resulted in labeling of all broad bean (Vicia faba) chromosomes at their ends only. Telocentric chromosomes derived by fission of the metacentric satellite chromosome of V. faba also showed signals at both of their ends, whereas the ancestral metacentric did not display signals at its primary constriction, the point of fission. As in V. faba, all acrocentric mouse chromosomes were labeled by in situ hybridization with a vertebrate telomeric probe at both ends of each chromatid exclusively. However, different metacentric Robertsonian chromosomes derived by fusion of defined acrocentrics did not show signals at their primary constrictions. The mechanism of Robertsonian rearrangement leading to a pseudoaneuploid increase or decrease in chromosome number therefore cannot consist solely of a simple fission or fusion of chromosomes without a concomitant gain or loss of chromatin material. The additional assumption of a subdetectable deletion of telomeric sequences after fusion and amplification of these sequences following fission is necessary to explain the present observations

Dose response for heritable translocations induced by acrylamide in spermatids of mice.

A heritable translocation test was carried out with acrylamide (AA) to obtain a dose-response relationship for induction of reciprocal translocations in late spermatids of the mouse. Male C3H/El mice were treated with single i.p. doses of 50 and 100 mg/kg of acrylamide and mated 7-16 days after treatment to untreated female 102/El mice. Translocation carriers among the F1 progeny were selected by a sequential procedure of fertility testing and cytogenetic analysis including G-band karyotyping to determine the chromosomes involved in the respective translocations. The translocation frequencies observed with 50 mg/kg and 100 mg/kg of AA were 0.6% and 2.7%, respectively. The historical control translocation frequency was 0.04%. Doubling dose estimates based on these and previous data are discussed