Temperature Dependence of Adriamycin-induced DNA Damage in LI 210 Cells1

We report alkaline elution experiments that reveal the temperature dependence of DNA lesions, both single-strand breaks and DNA-protein cross-links, in 1,121(1cells exposed to Adriamycin. DNA damage, which at 37°Cis equivalent to several hundred rads of ionizing radiation exposure, diminishes as the temperature of drug exposure is lowered. At all temperatures below about IST no DNA damage is detectable in LI 210 cells exposed to Adriamycin, even at relatively high doses. The low temperature inactivity is not due to a redistribution of intracellular drug since at both 37 and 0°Cthere is a high concentration of Adriamycin in both nuclear and cytoplasmic locations. The temperature profile for DNA damage parallels the profile for cytotoxicity, i.e., at low tempera ture, the drug is completely inactive as a cytotoxic agent (P. Lane, P. Vichi, D. L. Bain, and T. R. Tritton, Cancer Res., 47:4038-4042,1987). Thus, DNA breaks and cell kill appear to be correlated with one another. However, when we examined DNA lesions in nuclei isolated from 1,1210 cells we found that the low temperature inability to sustain Adriamycininduced single-strand breaks or DNA-protein cross-links was absent. In nuclei, then, the drug can provoke DNA damage at low temperature, while in whole cells it cannot. Topoisomerase II, an enzyme implicated in catalyzing DNA lesions in cells exposed to intercalating agents, retains its catalytic activity both to unknot IM DNA at 0°C, and to be induced by drug to alter the release of pBR322 supercoils, so a low temperature inactivation of this enzyme cannot explain the results. We propose that intact LI 210 cells have a regulatory factor which controls DNA damage, possibly through topoisomerase II, but which is lost when nuclei are isolated.