Effects of the modulating agent WR2721 and its main metabolites on the formation and stability of cisplatin-DNA adducts in vitro in comparison to the effects of thiosulphate and diethyldithiocarbamate

Abstract

The influence of the modulating agent WR2721, its active thiol-metabolite WR1065 and the symmetrical disulphide WR33278 on the in vitro formation and stability of cis-diamminedichloroplatinum(II) (cisplatin, CDDP)-DNA adducts was investigated and compared with the effects of the highly nucleophilic modulating agents diethyldithiocarbamate (DDTC) and thiosulphate (TS). Salmon sperm DNA (0.5 mg/mL) was incubated with 25 μg/mL (83 μM) cisplatin for 1 hr in 50 mM phosphate buffer, pH7.2 at 37° in the absence or presence of modulating agent. DDTC and TS were potent inhibitors of the platination of the DNA (95 and 89%, respectively, with 4.2 mM of modulating agent). The WR-compounds were also remarkably active in the inhibition of DNA platination. Prevention of adduct formation in the presence of 4.2 mM WR-compound decreased in the order WR1065 (74%) > WR33278 (63%) > WR2721 (51%). The prevention of CDDP-DNA adduct formation by WR1065 was strongly concentration-dependent up to 4.2 mM but at higher concentrations this protection hardly increased at all. In the presence of the modulating agents, increased levels of CDDP monofunctionally bound to a guanine residue were observed with a simultaneous decrease in the relative abundance of bifunctional adducts. All modulators were also able to reverse part of the CDDP-DNA adducts formed. After a 2-hr incubation of already platinated salmon sperm DNA with 4.2 mM of modulating agent, the removal of Pt from DNA amounted to about 43% with DDTC, 28% with WR1065 and 13–14% with TS, WR2721 and WR33278. Even CDDP bifunctionally bound to two adjacent guanines in the same DNA strand, which is considered to be a very stable adduct, was partly reversed. Our observations suggest that WR2721, especially when administered prior to or concomitantly with CDDP, can be expected to protect those tissues from CDDP-induced damage to DNA that are able to efficiently dephosphorylate WR2721 followed by uptake of the thiol metabolite WR1065. This stresses the importance of a selective formation and uptake of WR1065 by non-tumour tissues for the successful use of WR2721 as a protective agent in combination with platinum-based cancer chemotherapy.