2'-deoxy-5-azacytidine increases binding of cisplatin to DNA by a mechanism independent of DNA hypomethylation

Abstract

The chemotherapeutic agents 2'-deoxy-5-azacytidine (DAC) and cisplatin (cDDP) have been shown in vitro to be synergystic in their cytotoxicity toward human tumour cells. We have investigated possible molecular mechanisms underlying this synergy using the plasmid pSVE3 in vitro and after transfection into CMT3 cells. Increased binding of cDDP to DAC-substituted DNA generated in vivo was confirmed by flameless atomic absorption spectrophotometry (FAAS). The plasmid used in these experiments was unmethylated suggesting that DAC was effective in enhancing cDDP binding to DNA without acting as a hypomethylating agent, but by directly changing the topology of DNA. The role of DNA methylation in cDDP binding was studied using methylated and unmethylated plasmid incubated in vitro with cDDP. Restriction analyses and FAAS measurement of bound platinum indicated that methylated DNA bound more cDDP than unmethylated DNA. In addition, in vivo studies confirmed the in vitro observations since replication of methylated plasmid was inhibited to a greater extent than unmethylated plasmid.