High sensitivity of human epidermal keratinocytes (HaCaT) to topoisomeraseinhibitors

In the panorama of the numerous established cell lines, the human keratinoc yte line HaCaT has a very interesting feature, having a close similarity in functional competence to normal keratinocytes. This cell line has been use d in many studies as a paradigm for epidermal cells and therefore we select ed HaCaT as a cell model for investigating the activity of three antitopois omerase drugs (Camptothecin, Doxorubicin, Ciprofloxacin) on in vitro cell g rowth. The effect was evaluated both by a 24-h cytotoxicity test and by a 7 -day antiproliferation assay, in which the cell viability was assessed by a n MTT (3-(4,5-dimethyl-2-thiazolyl) 2,5-diphenil-2-H-tetrazolium bromide) t est. DNA topoisomerase I was also partially purified from a nuclear extract of HaCaT cells, the level of topo I catalytic activity was measured by a p BR322 DNA relaxation assay and then the in vitro effect of antitopoisomeras e drugs on the target enzyme was also assessed. The results indicated that the in vitro sensitivity of human epidermal HaCa T cells to antitopoisomerase drugs is comparable to that of many human tumo ur cell lines. HaCaT cells express a high level of topoisomerase I activity that is significantly inhibited by both Camptothecin and Doxorubicin and t o a minor degree by Ciprofloxacin. A high correlation between the cell sens itivity to the antitopoisomerase I drug measured by the MTT test and the in vitro direct inhibition of HaCaT topoisomerase I was observed, suggesting that HaCaT cells can represent a very interesting model both for studying c ellular pharmacokinetics of antineoplastic drugs on keratinocytes and for p redicting possible secondary effects, exerted by these drugs on cutaneous c ells, during treatment with chemotherapy.