In vivo xenograft mice experiment

All animal experiments were conducted according to the principles of the Institutional Animal Care and Use Committee at Shanghai Jiaotong University. Six-week BALB/c-nu female mice (Shanghai Slac Laboratory Animal Co., Ltd., Shanghai, China) weighing approximately 20 g were introduced to establish xenograft tumor model. 4 × 10⁶ RKO cells were subcutaneously injected into right flank of mice in 200 μl RPMI-1640 medium. When tumor volume reached about 100 mm³, mice were randomly assigned to different groups (n = 5) and treated every two days for 20 days. 4 groups were treated with UTD1 (2.5 mg/kg and 5 mg/kg, ip), paclitaxel 5 mg/kg (ip) or vehicle control. Three groups were treated with UTD1(2.5 mg/kg, ip), 5-FU (25 mg/kg, ip), or vehicle control. Tumor size was measured and mice were weighed every three days. Tumor volume was calculated by the formula: V = 0.5 × a × b², where a was the length (mm), b was the width (mm), and V was the volume of tumor. At the end of the experiment, all mice were killed and weights of tumor were recorded. To assess anticancer activity of drugs, percentage of inhibition was calculated as following: Inhibition% = (1−mean of tumor weight of treated group/mean of tumor weight of control group) × 100%.