Cryosurgical Modeling: Sequence of Freezing and Cytotoxic Agent Application Affects Cell Death

Cryosurgery is now being used to eradicate a variety of tumors, including those that are found in the kidney. Because it is well known that cancer treatments are often most effective when combined with adjunctive therapies, we developed a model kidney cell culture system to determine: (1) if 5-fluorouracil (5-FU), a compound known to be a poor kidney tumor chemotherapeutic agent, can be more effective when combined with cryosurgery; and (2) how kidney cells die through freezing-induced injury that might occur during cryosurgery. The DNA isolated from kidney cells that died during an exposure to -15 degrees C showed nonrandom cleavage when separated on an agarose gel, indicating that cell death was attributable, in part, to apoptosis, whereas DNA isolated from kidney cells that died during exposure to -75 degrees C showed random cleavage, indicating that cell death was attributable, in part, to necrosis. The apoptotic protease inhibitor, caspase 1 inhibitor V, was able to prevent freezing-induced cell death, supporting the idea that apoptosis may be a mechanism of cell death in the periphery of the iceball created by cryosurgical procedures in vivo. Because 5-FU is known to induce apoptosis, this drug was used in combination with various freezing regimes to determine if the combination might be a better method of killing cells than either treatment alone. Whereas the addition of 5-FU at the same time or 2 days after freezing resulted in a synergistic lethal effect, many cells survived this combination treatment. When cells were treated with 5-FU 2 days prior to freezing, however, there was an apparent complete loss of viability. The reason may be that, by an unknown mechanism, freezing enhances the ability of 5-FU-treated cells to move more effectively through the apoptosis pathway. Kidney tumors may have robust antiapoptotic mechanisms that make them refractory to 5-FU, but these mechanisms might be overridden by freezing. In either case, the data suggest that chemotherapy may be more effective when followed by cryosurgery.