The vailidation of defined genotoxic thresholds, leading to better invitro risk assessments of carcinogenic potential

All new chemicals are tested for their potential to cause DNA damage (genotoxicity), as this is linked to cancer. A tiered approach is used, with the chemicals first being tested in vitro. Those that induce DNA damage (test positives) are then usually tested again using animals. Some genotoxic agents give positive results in vitro at relatively high doses, but do not cause DNA damage at low doses more typical of human exposure levels. Identifying the dose below which DNA damage does not occur is important because understanding how this relates to human exposure levels can provide an opportunity to avoid using animals. This project will use an in vitro system to assess the genotoxic potential for a wide range of chemicals, helping to establish optimum testing strategies for
evaluating the effects of dose and potentially reducing animal use.

"As part of the safety assessment of new chemicals (pharmaceuticals, pesticides etc), a tiered approach to genotoxicity (DNA damaging) testing is currently recommended. This approach is designed to prevent human exposures to agents that can damage DNA, as DNA damage is closely linked to cancer development. The tiered approach involves all chemicals entering stage 1 tests where DNA damage induction is assessed in cells cultured in the laboratory. Negative results in these tests reassure the manufacturer that DNA damage is unlikely to be induced by that agent. However, any chemicals testing positive in these tests are usually then put through stage 2 tests which are carried out in animals (mice usually). This can involve thousands of such animal tests being carried out in the UK per year. The stage 2 tests are designed to overcome problems with false positive results which can occur in stage 1 tests and also to more rigorously assess the risks to human health. The false positive results in stage 1 tests can often arise due to high doses of chemicals being used. High doses of chemical have traditionally been used to ensure that DNA damaging effects are identified and it has widely been assumed that the effects are induced in a linear manner. That is, if a high dose is positive, then a low dose will also be positive.

We have recently shown how important dose can be in terms of DNA damage. We uniquely demonstrated that DNA damaging agents whilst giving positive results at high doses, are not DNA damaging at low doses. This confirms that "safe" doses exist, even for potent DNA damaging agents which are known to induce cancer. Hence, identifying these critical dose ranges is vital if we are to assess the effects of dose on DNA damage. If chemicals in stage 1 tests are only positive at high doses and not at low doses, and these positive effects were seen at doses estimated to be well above human exposures, then this would suggest that these chemicals would not be a human health risk. Importantly, the progression of these chemicals into stage 2 (animal) tests might not be necessary. Hence, by changing the testing strategy to consider the effects of dose, we could reduce the numbers of animal tests in toxicology. To add weight to the argument that DNA damaging thresholds exist and can be fully explained, we need high quality, cell-based studies of DNA damage induction for a wide range of chemicals. This application is aimed at supplying new knowledge in this field to impact on the testing strategies for DNA damage assessment with the ultimate aim of reducing the number of animal tests.
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