Reduced levels of the adenomatous polyposis coli (APC) protein are associated with ceramide-induced apoptosis of colon cancer cells

Purpose: Mutations of the adenomatous polyposis coli (APC) and p53 genes are commonly found in colorectal cancers. We therefore analyzed the relative roles of APC and p53 in the induction of apoptosis of colon cancer cells by comparing the effects of the natural chemopreventive agent, C ₂-ceramide, on different human colon cancer cell lines with and without wild-type p53 and APC genes. Methods: We studied the effect of C ₂-ceramide and C₂-dihydroceramide on proliferation and/or apoptosis of colon cancer cell lines in vitro and determined the role of p53 and APC proteins in these processes. The protein and mRNA levels in colon cancer cell lines with and without treatments were determined by Western and Northern blot analysis, respectively. The cell cycle and apoptosis profiles were determined by FACS analysis and PARP-1 cleavage. Results: Our findings indicate that C₂-ceramide can induce apoptosis independently of the p53/p21(Waf-1/Cip-1) pathway. In addition, the C₂-ceramide induction of apoptosis showed a correlation with a reduction in the levels of the APC protein and mRNA. Moreover, the C₂-ceramide-induced apoptosis was blocked by pre-treatment with ZnCl₂, which stabilizes APC protein levels. Conclusions: These results suggest that C₂-ceramide treatment reduces the levels of APC protein and that the reduction in the levels of this protein plays a key role in the ability of C₂-ceramide to induce apoptosis of colon cancer cells.