Dose- and time-dependent effects of doxorubicin on cytotoxicity, cell cycle and apoptotic cell death in human colon cancer cells

Abstract

The cytostatic drug doxorubicin is a well-known chemotherapeutic agent which is used in treatment of a wide variety of cancers. A key factor in the response of cancer cells to chemotherapeutic drugs is the activation of the apoptotic pathway, a pathway that is often impaired in chemoresistant colon cancer cells. The aim of the present study was to investigate the effects of doxorubicin in Hct-116 human colon carcinoma cells in order to clarify if a time/concentration range for optimal doxorubicin-induced apoptosis exists. We compared a treatment schedule were cells were bolus incubated for 3h with doxorubicin followed by 24h in drug-free medium, with a continuous doxorubicin treatment schedule for 24h. Bolus incubation was carried out to determine effects of doxorubicin accumulated during the first 3h, whereas continuous incubation allowed further (continuous) exposure to doxorubicin. We found that bolus (3h) treatment with doxorubicin resulted in a dose-dependent decrease of viable cells and concomitant increase of apoptosis. Additionally, bolus (3h) doxorubicin incubation led to phosphorylation of p53 at serine 392, induction of p21, G2 arrest and increase of proapoptotic protein Bax. In contrast, continuous (24h) treatment with doxorubicin reduced the number of living cells with no parallel raise in the amount of dead cells. Continuous (24h) treatment with 5 microM doxorubicin resulted in cell cycle arrest in G0/G1 phase that was neither accompanied by phosphorylation and activation of p53 nor enhanced expression of p21. These results suggest that doxorubicin is able to induce cell death by apoptosis only at particular dose and treatment conditions and imply a completely different cellular response following bolus or continuous exposure to doxorubicin.