IN-VITRO FORMATION OF DNA-ADDUCTS BY CISPLATIN, LOBAPLATIN AND OXALIPLATIN IN CALF THYMUS DNA IN SOLUTION AND IN CULTURED HUMAN-CELLS

Two interesting representatives of a new generation of platinum-based cytostatic drugs that are currently being tested in clinical trials ar e lobaplatin [1,2-diaminomethylcyclobutane platinum(II) lactate] and o xaliplatin [1,2-diaminocyclohexane platinum (II) oxalate], Since littl e is known about the DNA adduct formation of these compounds, we studi ed their formation in DNA in vitro in calf thymus DNA and in cells, Th e major adducts formed in vitro were the Pt-GG and Pt-AG intrastrand c rosslinks. The latter adducts could be detected using a recently devel oped P-32-postlabelling method. Using both this assay and atomic absor ption spectroscopy, it was shown that there is a substantially higher rate of the in vitro adduct formation by cisplatin, compared with loba platin and oxaliplatin, Platinum concentrations required to obtain 90% cell kill during a 2 h incubation of A2780 cells were 15 mu M for cis platin and oxaliplatin and 22 mu M for lobaplatin, Using an antiserum originally raised against cisplatin-treated DNA, we were also able to detect platinum-DNA adducts induced by lobaplatin and oxaliplatin, Max imal nuclear staining for all three compounds was observed after a 4 h post-incubation period, The nuclear staining level induced by cisplat in was about 10-fold higher than after lobaplatin and oxaliplatin trea tment, GG and AG adducts, measured by P-32-postlabelling, also showed maximum levels at about 4 h after treatment, Relative GG peak levels w ere 4:1:3 for cisplatin, lobaplatin and oxaliplatin, respectively, The ratios of GG over AG intrastrand crosslinks in the A2780 cells were n ot significantly different for the various compounds, In conclusion, t he P-32-postlabelling technique has been shown to be appropriate for a dduct analysis, not only for the classical Pt compounds cisplatin and carboplatin but also for novel platinum compounds like lobaplatin and oxaliplatin. Results indicated large differences in reactivity of the latter compounds to DNA in vitro, compared with cisplatin, This differ ence was smaller in cells, suggesting enhancement of adduct formation by certain cellular mechanisms and/or compounds. From these studies, n o conclusions can be drawn with respect to the cytotoxicity of the dif ferent Pt-GG and Pt-AG intrastrand crosslinks formed by these compound s.