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The combination of irinotecan and a fluoro-pyrimidine is widely accepted as a treatment for advanced colorectal cancer. However, evaluable data on the feasibility of these combinations has not been presented, and an optimal sequence for administration has not been experimentally and clinically determined. The sequential effect of a combination of 5-FU and CPT-11 in the human colon cancer cell line LoVo was evaluated by WST-8 colorimetric assay. The cytotoxicity and cell cycle distributions of each drug were analyzed by apoptosis assay and flow cytometry. Further, the potential mechanisms of the sequence-dependent effects were investigated by a microarray technique, and confirmed by Western blot analysis. The cytotoxicity of 5-FU (10, 100, 1000 microM) followed by CPT-11 (1 microM) was significantly greater than that of CPT-11 (1 microM) followed by 5-FU (10, 100, 1000 microM) (p<0.05). In cell cycle distribution, 5-FU exposure for 24 h increased the S phase fraction in a dose-dependent manner; though there was no significant difference in cell cycle distribution in 24 h CPT-11 (0.01-1 microM) exposure. Microarray analysis revealed that expressions of some apoptosis related genes such as Bcl-2 changed, and were correlated with sequence-dependent cytotoxicity of the 5-FU --> CPT-11 sequence. Western blot analysis confirmed that the Bcl-2/Bax ratio was lower after 5-FU --> CPT-11 sequence than before. The sequence-dependent cytotoxic effect may depend on the sensitizing effect of 5-FU pretreatment on CPT-11 cytotoxicity. 5-FU followed by CPT-11 administration may be an optimal sequence for IFL treatment of advanced colon cancer.

in vitro synergistic antitumor activity of a combination of 5-fluorouracil and irinotecan in human colon cancer