Evaluation of the alkaline comet assay and urinary 3-methyladenine excretion for monitoring DNA damage in melanoma patients treated with dacarbazine and tamoxifen.
Braybrooke JP., Houlbrook S., Crawley JE., Propper DJ., O'Byrne KJ., Stratford IJ., Harris AL., Shuker DE., Talbot DC.
PURPOSE: To develop, using dacarbazine as a model, reliable techniques for measuring DNA damage and repair as pharmacodynamic endpoints for patients receiving chemotherapy. METHODS: A group of 39 patients with malignant melanoma were treated with dacarbazine 1 g/m(2) i.v. every 21 days. Tamoxifen 20 mg daily was commenced 24 h after the first infusion and continued until 3 weeks after the last cycle of chemotherapy. DNA strand breaks formed during dacarbazine-induced DNA damage and repair were measured in individual cells by the alkaline comet assay. DNA methyl adducts were quantified by measuring urinary 3-methyladenine (3-MeA) excretion using immunoaffinity ELISA. Venous blood was taken on cycles 1 and 2 for separation of peripheral blood lymphocytes (PBLs) for measurement of DNA strand breaks. RESULTS: Wide interpatient variation in PBL DNA strand breaks occurred following chemotherapy, with a peak at 4 h (median 26.6 h, interquartile range 14.75-40.5 h) and incomplete repair by 24 h. Similarly, there was a range of 3-MeA excretion with peak levels 4-10 h after chemotherapy (median 33 nmol/h, interquartile range 20.4-48.65 nmol/h). Peak 3-MeA excretion was positively correlated with DNA strand breaks at 4 h (Spearman's correlation coefficient, r=0.39, P=0.036) and 24 h (r=0.46, P=0.01). Drug-induced emesis correlated with PBL DNA strand breaks (Mann Whitney U-test, P=0.03) but not with peak 3-MeA excretion. CONCLUSIONS: DNA damage and repair following cytotoxic chemotherapy can be measured in vivo by the alkaline comet assay and by urinary 3-MeA excretion in patients receiving chemotherapy.