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Triptolide (TPL) is a major active component isolated from the natural herb Tripteryglum wilfordii Hook. F. It has proved to possess a variety of pharmacological effects including anti-inflammatory and antitumor activities. The aim of the present study is to explore the efficiency of combination therapy with TPL and oxaliplatin (OXA) and identify the in vitro and in vivo cytotoxicity on colon cancer lines and mice model. Cell viability was measured by MTT assay and cell apoptosis rate was analyzed by FACS assay after treatment with TPL and OXA alone, and TPL combined with OXA in colon cancer cell line SW480. The results demonstrated that combination therapy of TPL and OXA could effectively inhibit the proliferation of colon cancer cell line SW480 and induce cell apoptosis of colon cancer cells. It was partly induced by inhibiting nuclear translocation of β-catenin and the expression of the target genes in cell cycle, which was detected by western blotting and real-time PCR. Moreover, in nude mice model, tumor growth was significantly suppressed in the group treated with TPL in combination with OXA. There was no obvious cytotoxity in mice analyzed by normal blood test and liver and kidney toxicity test. In conclusion, our result revealed that the combination therapy with TPL and OXA exerted synergistic antitumor effects at low concentration in colon cancer cells, with less cytotoxity, which exhibited high potency for clinical applications.