The in vitro sensitivity testing for four human colorectal cancer cell lines to seven chemotherapeutic drugs including CPT-11, derivative of camptothecin, and its active form SN-38 were determined. MTT assay revealed that SN-38 was the most active for all four cell lines tested and its IC50's were very close to its clinically achievable plasma concentration. Relationship between exposure time and cytocidal effect of SN-38 was also investigated using MTT assay, topoisomerase-I (Topo-I) immunoblot analysis and DNA relaxation-assay, showing that IC50 value, Topo-I protein and Topo-I activity were decreased soon after the administration of SN-38 and reached to the plateau level at 24 hours. We conclude that SN-38 is very potent for colorectal cancer and the optimal schedule of CPT-11 can be the more continuous form of administration capable of as long as 24 hours exposure of its active metabolite, SN-38.