β-lapachone induces cell cycle arrest and apoptosis in human colon cancer cells

Abstract

Background: Human colon cancers have a high frequency of p53 mutations, and cancer cells expressing mutant p53 tend to be resistant to current chemo- and radiation therapy. It is thus important to find therapeutic agents that can inhibit colon cancer cells with altered p53 status. β-Lapachone, a novel topoisomerase inhibitor, has been shown to induce cell death in human promyelocytic leukemia and prostate cancer cells through a p53-independent pathway. Here we examined the effects of β-lapachone on human colon cancer cells. Materials and Methods: Several human colon cancer cell lines, SW480, SW620, and DLD1, with mutant or defective p53, were used. The antiproliferative effects of β-lapachone were assessed by colony formation assays, cell cycle analysis, and apoptosis analysis, including annexin V staining and DNA laddering analysis. The effects on cell cycle and apoptosis regulatory proteins were examined by immunoblotting. Results: All three cell lines, SW480, SW620, and DLD1, were sensitive to β-lapachone, with an IC50 of 2 to 3 μM in colony formation assays, a finding similar to that previously reported for prostate cancer cells. However, these cells were arrested in different stages of S phase. At 24 hr post-treatment, β-lapachone induced S-, late S/G2-, and early S-phase arrest in SW480, SW620, and DLD1 cells, respectively. The cell cycle alterations induced by β-lapachone were congruous with changes in cell cycle regulatory proteins such as cyclin A, cyclin B1, cdc2, and cyclin D1. Moreover, β-lapachone induced apoptosis, as demonstrated by annexin V staining, flow cytometric analysis of DNA content, and DNA laddering analysis. Furthermore, down-regulation of mutant p53 and induction of p27 in SW480 cells, and induction of proapoptotic protein Bax in DLD1 cells may be pertinent to the anti-proliferative and apoptotic effects of β-lapachone on these cells. Conclusions: β-Lapachone induced cell cycle arrest and apoptosis in human colon cancer cells through a p53 independent pathway. For human colon cancers, which often contain p53 mutations, β-lapachone may prove to be a promising anticancer agent that can target cancer cells, especially those with mutant p53.