OBJECTIVES: An amphiphathic structure of bile acids has been known to facilitate transport of various substances across biological membranes, influencing their physiological and pharmacological activity. The aim of this work was to analyze the cytotoxic effects of co-treatment with doxorubicin and bile acids on MCF-7 cell line, as well as to determine type of pharmacological interaction. METHODS: MCF-7 cells have been treated with increasing concentrations of doxorubicin, chenodeoxycholic and ursodeoxycholic acid for 48 h and cytotoxic activity was determined using colorimetric MTT assay. Cells were also co-treated with non-toxic concentrations of doxorubicin and each bile acid. Quantitative analysis of dose-effect relationship was performed according to Chou-Talalay method for drug combination assessment. RESULTS: All the agents showed a dose-dependent cytotoxic activity.  The co-incubation of cells with 0.25 μM of  doxorubicin  and non-toxic (IC20) concentrations of bile acids over 48 h resulted in statistically significant increase of cytotoxicity: co-treatment with ursodeoxycholic and chenodeoxycholic acid increased the cytotoxic effect up to 39.6% (p<0.05), and 67.9% (p<0.01), respectively, compared to  doxorubicin  treatment alone which resulted in inhibition of growth in 33.1% of cells. Quantitative analysis of dose-effect relationship resembled in combinational index (CI) value indicated that co-treatment with ursodeoxycholic acid exerted slight additive effect (CI=1.2), whereas with chenodeoxycholic acid strong synergistic activity (CI=0.18) with doxorubicin. CONCLUSIONS: These results indicate that both bile acid species modulate the cytotoxic activity of doxorubicin. Since chenodeoxycholic acid synergistically acts with doxorubicin, it may be further investigated as a novel chemo-sensitizing agent with primary aim to improve the therapeutic response to doxorubicin-containing chemotherapy regimens, while potentially decreasing its harmful effects. Acknowledgment: Supported by the Ministry of Education, Science and Technological Development, Republic of Serbia, Grant III41012.