Synergy of interferon-alpha and 5-fluorouracil in human renal cell carcinoma requires p53 activity.

Abstract

Immunochemical therapy combining cytokines and chemotherapeutic agents is expected to be effective for treating advanced renal cell carcinoma (RCC). We investigated the mechanism underlying the synergism of interferon-alpha (IFN-alpha) and 5-fluorouracil (5-FU) and the effect of p53 status on the synergy of the combined therapy in RCC cell lines. The synergy of IFN-alpha and 5-FU was analyzed by isobolographic analysis in five RCC cell lines. The effect of combined treatment on apoptosis induction was measured by flow cytometric analysis, Hoechst staining, and caspase activity assay; PCNA expression was investigated by Western blotting to examine the effect of combined treatment on the antiproliferative effect. We demonstrated synergy of IFN-alpha and 5-FU in five RCC cell lines with wild-type p53. IFN-alpha suppressed the proliferation of RCC cells via G1 or G2/M cell cycle arrest without inducing apoptosis, whereas 5-FU induced apoptosis in a dosage-dependent manner. IFN-alpha enhanced the apoptosis of RCC cells induced by 5-FU, whereas 5-FU did not increase the antiproliferative effect of IFN-alpha. However, the synergistic inhibition by IFN-alpha and 5-FU was abolished when the cell lines were transfected with p53 dominant-negative vector. The synergy of IFN-alpha and 5-FU requires p53 activity, suggesting that p53 status may serve as a predictive factor for response to the combination therapy. Because metastatic RCC frequently has p53 mutations, therapy restoring p53 may markedly improve the response rate of immunochemical therapy combining IFN-alpha and 5-FU.