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Kinetics of reaction of cis-diamminedichloroplatinum(II) with DNA.

A method based on cation exchange chromatography was developed to determine the adducts formed in the reaction of cis-diamminedichloroplatinum(II) (cis-Pt) with DNA. DNA was incubated with various concentrations of cis-Pt for various periods of time, ethanol precipitated, and enzymatically digested to nucleosides and Pt-containing oligonucleotides. The unmodified nucleosides were separated from the positively charged intra- and interstrand cis Pt adducts with a weak cation exchanger, CM-Sephadex C-25, and the adducts were further purified by HPLC. The main adduct was shown to be an intrastrand cross-link of cis-Pt bound to the N-7 atoms of two neighboring guanines. The minor adducts were intra- and interstrand cross-links of cis-Pt with adenine and guanine and an interstrand cross-link of cis-Pt with two guanines. At low levels of DNA-modification (cis-Pt:nucleotide = 1:50-1:1000) the intrastrand cross-link of cis-Pt with two guanines consisted of 60-70% of the total platination of DNA. At higher levels of DNA-modification (greater than 1:20), the amount of undigested products increased, indicating shielding of DNA by cis-Pt from nucleolytic enzymes.