Assessment of the mutagenic potential of para‐chloroaniline and aniline in the liver, spleen, and bone marrow of Big Blue® rats with micronuclei analysis in peripheral blood

Splenic tumors have been reported in rat cancer bioassays with para‐chloroaniline (PCA) and aniline. Development of these tumors is hypothesized to be due to hematotoxicity via the formation of methemoglobin (MetHb) and not direct DNA reactivity. To evaluate the mode of action (MOA) for tumor formation a transgenic rodent (TGR) in vivo gene mutation assay in Big Blue® TgF344 rats was performed with parallel micronuclei analysis in peripheral blood. Male rats were gavaged daily for 28 d to 0.5, 15, and 60 mg/kg PCA and 100 mg/kg aniline, the base molecular structure of PCA. On test day 10, the 60 mg/kg PCA dose was reduced to 30 mg/kg due to toxicity. On test day 4 and 29 peripheral blood micronucleus analysis was performed and on test day 29 clinical chemistry, hematology, and MetHb measurements were taken. At study termination, on test day 31, spleen, bone marrow, and liver (control tissue) were analyzed for cII transgene mutant frequency (MF). Repeat gavage exposure to PCA and aniline for 28 d did not produce an increase in cII transgene MF in analyzed tissues. An increase in micronuclei was seen at both time points at ≥15 mg/kg PCA and 100 mg/kg aniline. At the same dose levels, significant reductions in red blood cells, increases in absolute reticulocytes (ABRET), and increased levels of MetHb were observed. Together these results support that generation of micronuclei and tumorigenicity following exposure to PCA and aniline is due to compensatory mechanisms (e.g. increased cellular turnover) and not direct DNA reactivity. Environ. Mol. Mutagen. 59:785–797, 2018. © 2018 Wiley Periodicals, Inc.