In vivo and in vitro inhibitory effects of acrylamide on DNA topoisomerase II

Acrylamide (AA), a chemical produced in several foodstuffs when cooked at a high temperature, is considered a probable human carcinogen, but the molecular mechanism underlying its genotoxicity has not fully known. Numerous authors have reported the induction by AA of DNA double strand breaks and sister chromatid exchange (SCE); we here confirmed the acrylamide capability of damaging DNA by utilizing Comet assay, which showed a dose-dependent increase of tail lenght, in metabolically non competent V79 Chinese hamster cells. Moreover, we observed that Acrylamide (AA) was able to antagonize in vivo the citotoxicity of well know poison etoposide; this suggested that topoisomerase II activity was affected by AA. The hypothesis was confirmed in in vitro tests: we observed, in fact, a strong inhibitory effect of AA against topoisomerase II in the kDNA assay and in topoisomerase II-mediated supercoiled DNA relaxation assay. In particular, this latter inhibition was not accompanied by stabilization of a covalent topoisomerase II-DNA intermediate. In order to characterize AA target(s), we pretreated pBR322 with AA and observed that this substrate became quickly incompetent in the topoisomerase II catalytic assay. These results suggest that DNA could be a target for Acrylamide-induced inhibition of topoisomerase II. Furthermore, preliminary results seem to indicate the possibility that another mode of action of acrylamide is related to its affinity for topoisomerase II sulphydryl groups, according to recent evidences reporting that thiol-reactive compounds can induce DNA damage through a nongenotoxic mechanism, such as the thiolation of the nuclear protein topoisomerase II.