Purpose: Combination therapy is a promising strategy to treat cancer due to the synergistic effects. The drug and gene co-delivered systems attract more attention in the field of combination therapy. Materials and methods: In the present research, poly(ethylene glycol)-epsilon-poly(caprolactone) block copolymer was used for the co-loading of 5-Iluorouracil (5-FU) and gene. The physicochemical characteristics, in vitro and in vivo anticancer, and gene transfection efficiency were tested on colon cancer cells and tumor-bearing mice. Results: 5-FU and gene co-loaded nanocarriers had a size of 145 nm. In vivo gene delivery results showed about 60% of gene-positive cells. Tumor volume of nanocarrier groups at day 21 was around 320 mm(3), which is significantly smaller compared with free 5-FU group (852 mm(3)) and control group (1,059 mm(3)). The maximum 5-FU plasma concentration in nanocarrier groups (49 mu g/mL) was significantly greater than free 5-FU (13 mu g/mL). At 24 hours, drug level of nanocarrier groups was about 2.8 mu g/mL compared with 0.02 mu g/mL of free 5-FU. Conclusion: The resulting nanocarriers co-loaded with the anticancer drugs and genes could be considered as a promising nanomedicine for colorectal cancer therapy.