Quantitative aspects of the formation and loss of DNA interstrand crosslinks in Chinese hamster cells following treatment with cis-diamminedichloroplatinum(II) (cisplatin) I. Proportion of DNA-platinum reactions involved in DNA crosslinking

Abstract

The presence of interstrand crosslinks in the DNA of cis-diamminedichloroplatinum(II) (cisplatin)-treated Chinese hamster cells was demonstrated by the formation of a species of hybrid-labelled DNA in cells that had been density and radioactively prelabelled with a combination of [³H]thymidine and 2′-deoxy-5-bromouridine (BrdUrd) prior to treatment. The extent of crosslinking and of total binding of platinum to the DNA increased linearly with dose over a wide range of concentrations of cisplatin. Simultaneous measurement of the size of co-isolated ¹⁴C-labelled DNA from untreated cells permitted an estimate of the size of the crosslinked DNA and hence of the proportion of bound platinum involved in a crosslinking reaction. No increase in the proportion of crosslinking reactions occurred during several hours post treatment incubation of cells when crosslinks were measured by this technique, in contrast to other studies using different techniques to measure DNA interstrand crosslinking in cisplatin treated cells (see following paper, Pera, M.F., Jr., Rawlings, C.J., Shackleton, J. and Roberts, J.J. (1981) Biochim. Biophys. Acta 655, 152–166). The enforced high concentrations of cisplatin required by this method to detect crosslinks precludes studies of their possible repair.