Augmentation of Antitumor Activity of 5-fluorouracil by Interferon Alpha is Associated with Up-regulation of P27Kip1 in Human Hepatocellular Carcinoma Cells

Several clinical trials have demonstrated the effectiveness of combination therapy with 5-fluorouracil (5-FU) and IFN-alpha in colon cancer, hepatocellular carcinoma (HCC), and other malignancies. In our preliminary clinical studies, we have observed outstanding effects with this combination therapy in patients with advanced HCC. However, the underlying mechanism by which IFN-alpha modulates the effects of 5-FU is unknown. We, therefore, conducted a mechanistic study using two HCC cell lines, PLC/PRF/5 and HuH7. IFN-alpha significantly enhanced the growth inhibitory effect of 5-FU in PLC/PRF/5 cells but not in HuH7 cells, and the isobolographic analysis indicated that this effect was synergistic. Flow cytometric analysis showed a delay in the progression of G0-G1 to S phase in PLC/PRF/5, and a sustained, induction of the cyclin-dependent kinase inhibitor p27-Kip1 and down-regulation of cyclin D1 was observed. Moreover, increased expression of p27Kip1 was associated with reduced CDK-2-associated kinase activity. Another difference in the two cell types was that PLC/PRF/5 expressed abundant IFN receptors, but HuH7 did not. Apoptosis assays were not helpful in explaining the mechanism. Our results suggest that the synergistic effects of 5-FU and IFN-alpha may in part be attributable to alterations in cell cycle progression via up-regulation of p27Kip1.