Recently,
natural product DNA topoisomerase I inhibitors 10-hydroxycamptothecin (
HCPT) and
camptothecin (
CPT) have been shown to have
therapeutic effects in both in vitro and in vivo models of human
breast cancer. In the present study, we characterized the in vitro and in vivo apoptotic pathways induced by
HCPT and
CPT in the human
breast cancer cell lines MCF-7 and MDA-MB-468. Using various DNA fragmentation analyses and the
terminal deoxynucleotidyl transferase-mediated nick end labeling (TUNEL) assay, the apoptosis induced by
HCPT and
CPT was shown to be dose- and time-dependent. The MDA-MB-468 cells were more sensitive to both
HCPT and
CPT than MCF-7 cells.
HCPT induced apoptosis in MDA-MB-468 cells more effectively than
CPT; however, in MCF-7 cells,
CPT was more effective than
HCPT. The levels of p53 and p21WAF1/CIP1
protein increased in MCF-7 cells treated with
HCPT or
CPT in a dose- and time-dependent manner. The levels of p21WAF1/CIP1
protein also increased in a dose- and time-dependent manner in MDA-MB-468 cells treated with
HCPT or
CPT, whereas the mutated p53
protein levels had no significant change. The elevation of p53
protein levels in MCF-7 cells treated with
CPT was significantly inhibited by preincubation with DNA breaks inhibitor
aphidicolin, while the elevation of p21WAF1/CIP1
protein levels was not inhibited. The elevation of p21WAF1/CIP1 in MDA-MB-468 cells treated with
CPT was not inhibited by
aphidicolin. Using Northern blot analysis, the transcription of p21WAF1/CIP1 was shown to increase in a dose-dependent manner in MCF-7 and MDA-MB-468 cells treated with
HCPT or
CPT. These results suggest that treatment with
HCPT and
CPT results in increased levels of p21WAF1/CIP1
protein and
mRNA, and that they induce apoptosis in human
breast cancer cells through both p53-dependent and -independent pathways. These findings may be significant in further understanding the mechanisms of actions of camptothecins in the treatment of human
cancers.