Adriamycin: protection from cell death by removal of extracellular drug

Adriamycin is a cytotoxic drug which has enjoyed considerable success in the treatment of cancer. This agent has a bewildering variety of biological effects both within and on the surface of cells exposed to drug, and it has proved difficult to unambiguously assign a single mechanism of action. In this report we are able to separate intracellular and extracellular actions by taking advantage of the complete lack of Adriamycin-induced cytotoxicity at low temperature. For example, cells exposed to 100 microM Adriamycin at 0 degree C are not killed by the drug, even though this concentration is orders of magnitude higher than the concentration needed to cause 100% cell death at 37 degrees C. If cells exposed to 100 microM Adriamycin at 0 degree C are shifted to fresh drug-free medium at 37 degrees C, there is a time-dependent decrease in survival. However, if the drug-free medium contains calf thymus DNA (1.5 mg/ml) to act as a reservoir for Adriamycin binding of effluxed drug, there is no ensuing cytotoxicity. Thus, the results show that no matter how much drug is present inside the cell, there must also be extracellular drug available for membrane interaction in order to initiate nuclear DNA damage and the cytotoxic cascade.