Occupational cancer has been associated with chronic exposure to cytotoxic agents (chemicals, radiation) in the workplace. In an attempt to formally link these malignancies to specific causative agents, a large number of in vitrostudies have been carried out, in which cells in culture were treated with a range of putative causative agents. However, most of these studies used only single treatments with the respective substances, and may thus be poor models of chronic exposure. Here, we describe the comparison of treating human cultured cells with either a single or repeated doses MOCA [4,4’-methylene bis(2-chloroaniline)], an aromatic amine suspected of causing occupational bladder cancer. To this end, we evaluated its cytotoxicity and its effect on cellular proliferation and on the survival signaling pathways ERK and Akt, as well as on the expression level of cytochrome P450 (CYP) that metabolically activates MOCA.

We show that the cytotoxicity induced by repetitive MOCA treatment was lower than that caused by the same dose used singly, which agrees with our finding that ERK activation by a single MOCA administration required higher drug concentrations than by repetitive treatment. This may provide chronically exposed cells with a survival advantage, which could allow them to acquire further mutations necessary for transformation to malignancy. The latter results demonstrate the potential of our assay in occupational hygiene research, given that it appears able to reveal clear differences between chronic and acute treatments in a simple in vitro setting. We are currently attempt ing to optimize the assay conditions further and search for additional biomarkers; as Akt and CYP1A1 levels varied already in the negative controls, the latter markers are unlikely to provide a reliable readout in an assay of this type.