RE: DNA damage detected by the alkaline comet assay in the liver of mice after oral administration of tetrachloroethylene. (Mutagenesis, 25, 133-138, 2010)

Abstract

The recently published paper by Cederberg et al. (1) discusses the induction of DNA damage in an in vivo comet assay in mice dosed with tetrachloroethylene. The authors concluded that tetrachloroethylene induced significant DNA damage in the liver, but this conclusion was heavily dependent on statistical analysis. The laboratory that performed the study (Covance Laboratories Limited) concluded that tetrachloroethylene did not induce DNA damage in the liver. Cederberg et al. (1) presented and discussed the Covance opinion, but this discrepancy in the conclusions merits further discussion. It is our opinion that the paper does not accurately report the Covance evaluation criteria used to assess these data. Firstly, when assessing any genotoxicity data, including comet data, it is important to evaluate the biological relevance of the results as well as any statistical findings. In fact, most OECD guidelines state that statistical significance should not be the only criterion for conclusion of a positive result. We believe, and it is common practice, that statistical analysis should be used as an aid to interpretation and not the sole criterion. Groups of mice treated with tetrachloroethylene exhibited tail moments and tail intensities that were similar to the concurrent vehicle control group. The maximum group mean increase was only 1.43-fold above the vehicle control group and that was for tail intensity in the 2000 mg/kg group. The increase in tail moment in this group was only 1.36-fold above the vehicle control group. Although Covance had limited historical control comet data for mouse liver, a validation study on mouse liver had been performed before the tetrachloroethylene study, and all tail moment and tail intensity values for tetrachloroethylene were comparable to the validation data for the mouse liver. Covance had considerably more historical control data for rat liver, and all of the tail moment and tail intensity values for tetrachloroethylene fell within the laboratory’s rat historical control range for the liver. The variation observed within groups for tail intensity is of a similar magnitude to that observed between groups and the values observed in the vehicle and test article-treated groups overlap. This supports the opinion that the increases observed are of a magnitude considered to be due to biological variation and not indicative of DNA damage. Thus, Covance concluded that the tetrachloroethylene data did not demonstrate a biologically significant effect. The choice of statistical analysis also deserves comment. Of course, there are often many different opinions regarding the most appropriate statistical test to perform with a given data set. However, it is interesting that the expert working group of the International Workshop on Genotoxicity Tests (2) concluded the following: