AbstractNiclosamide is an antihelminthic drug. Recent studies show that niclosamide exerts antitumor activity through inhibiting multiple signals including Wnt/β‐catenin, mTORC1, signal transducer and activator of transcription 3, NF‐κB, notch signals; however, the insolubility and poor bioavailability limits its potential clinic use, the aim of the present work is to synthesize an injectable pegylated niclosamide (polyethylene glycol‐modified niclosamide) and investigate its antitumor activity in vitro and in vivo. The pegylated niclosamide (mPEG5000‐Nic) was synthesized and the chemical structure was identified by Fourier transform infrared spectra and 1H nuclear magnetic resonance spectra. The antitumor activity was evaluated in CT26 and HCT116 colon cancer cells in vitro and nude mouse xenograft model of CT26 cells in vivo. The water solubility of niclosamide in mPEG5000‐Nic was significantly increased. Niclosamide could be released from mPEG5000‐Nic nanoparticles in PBS solution. mPEG5000‐Nic inhibited the cell viability of CT26 and HCT116 cells in vitro. No animal death was observed in mice with intraperitoneal injection of mPEG5000‐Nic (equivalent to 1000 mg/kg niclosamide) within 24 hr, indicating that mPEG5000‐Nic was less toxic. In nude mouse, xenograft model of CT26 colon carcinoma, intraperitoneal injection of mPEG5000‐Nic (equivalent to niclosamide 50 mg/kg) inhibited tumor growth but had no effect on animal body weight and heart, liver, kidney, and lung weight in vivo. Meanwhile, in the same model, intraperitoneal injection of the positive clinic drug 5‐fluorouracil not only inhibited the tumor growth, but also reduced the animal body weight. Our study demonstrates that pegylated niclosamide is novel niclosamide delivery system with clinical perspective for cancer therapy.