Enhancement of camptothecin-induced cytotoxicity with UCN-01 in breast cancer cells: abrogation of S/G(2) arrest

Purpose: To determine the ability of UCN-01 to abrogate the cell cycle arre st induced by camptothecin (CPT) in tumor cells that lack p53 function, and therefore enhance the cytotoxicity of CPT in these cells in relation to no rmal cells with wild-type p53. Methods: The responses of MDA-MB-231 and GI 101A breast cancer cells were compared to those of normal bovine endothelia l cells. Cytotoxicity was assessed by the MTT assay, and the resulting data were modeled using median-effect analysis. Inhibition of DNA synthesis was determined by loss of [H-3]thymidine incorporation, and cell cycle status was determined by flow cytometric analysis of propidium-iodide-stained nucl ei. Results: UCN-01, a specific inhibitor of protein kinase C (PKC) present ly in clinical trials, abrogated CPT-induced activation of S and G(2) check points in human MDA-MB-231 and GI 101A breast carcinoma cells, both of whic h are mutants for the p53 gene. This abrogation occurred with the use of su blethal doses (100 nM) of UCN-01 and correlated with the enhancement of CPT -induced cytotoxicity. Median-effect analysis showed that synergistic cytot oxic interactions existed between CPT and UCN-01 against these tumor cells. In normal cells, however, abrogation of the S phase arrest caused accumula tion in G(0)/G(1) phase, perhaps by the presence of wild-type p53 activity, with no change in CPT-induced cytotoxicity. Conclusion: We have shown prev iously that the cytotoxicity of CPT is correlated with cell cycle response in normal and tumor cells. Low doses of CPT arrest cells in the G(2)/M phas e and inhibit DNA synthesis, but higher doses cause arrest of cells in S ph ase. Thus modulation of events at the S and G(2) checkpoints may provide an opportunity to enhance CPT-induced cytotoxicity in tumor cells. The result s of this study indicate that UCN-01 enhances the progression of tumor cell s through S phase thus greatly increasing CPT-induced cytotoxicity. Normal cells, however, are able to arrest in G(0)/G(1) and thus avoid the increase d toxicity induced by CPT. Our findings suggest potential usefulness of com bining UCN-01 in topoisomerase I inhibitor-based drug therapy for the treat ment of breast cancer with a dysfunctional p53 gene.