Apoptotic activity of ursolic acid may correlate with the inhibition of initiation of DNA replication

Abstract

Ursolic acid (UA), a pentacyclic triterpene acid, has been reported to exhibit anti-tumor activity. In this study, we investigated the pro-apoptotic effect of UA on HepG2 human hepatoblastoma cells. Treatment with UA decreased the viability of HepG2 cells in a concentration- and time-dependent manner. Furthermore, 30 μM of UA induced DNA fragmentation and subdiploid cells and enhanced the release of cytochrome c and the activation of caspase-3. These results suggest that UA induces cell death through apoptosis, which may be mediated by cytochrome c-dependent caspase-3 activation. In addition, cell-cycle analysis revealed that UA-treated cells were arrested predominantly in the G0 and G1 phases with a concomitant decrease in the cell population of S phase. Moreover, expression of p21WAF1, a cell-cycle regulator, was increased by UA, indicating that p21WAF1 might mediate UA-induced cell-cycle arrest. However, UA markedly inhibited SV40 DNA replication in the initiation stage in vitro and significantly reduced the DNA cleaving of topoisomerase I and the ssDNA binding activity of replication protein A. These results indicate that the inhibition of DNA replication by UA may result from blockade of the establishment of the replication fork during initiation stage, consequently contributing to UA-induced cell-cycle arrest. Taken together, we suggest that UA-induced cell-cycle arrest may be mediated by inhibition of DNA replication and the increase of p21WAF1 expression, which induces the release of cytochrome c and the activation of caspase-3, leading to apoptosis of HepG2 cells. Int. J. Cancer 87:629–636, 2000. © 2000 Wiley-Liss, Inc.