Objective To explore the tumorigenicity and expressions of dishevelled 3 (DVL3) in HCT116 cells and HCT116 spherical cells. Methods Human colorectal tumor HCT116 cells were cultured in the serum-free culture medium for HCT116 spherical cells. Through the subcutaneous tumor experiment in nude mice and clone formation assay, we observed the tumor growth and colony formation ability of the two kinds of cells in vivo and in vitro. The Western blotting experiment was utilized to detect the expressions of DVL3 in these two kinds of cells. Results ① Colonyformation: the mean value of colony formation rate in the HCT116 cells group was 3.78%, and the mean value of fcolony formation rate in the HCT116 spherical cells group was 28.67%, which was higher in the HCT116 spherical cells group (t=21.16, P<0.05). ② Tumorigenicity in nude mice: 11 nude mice with tumor formation were observed in the HCT116 cells group, and the tumor formation rate was 55.0%; 18 nude mice with tumor formation were observed in the HCT116 spherical cells group, and the tumor formation rate was 90.0%, the tumor formation rate of the HCT116 spherical cells group was higher (P=0.039). The tumor volume of the HCT116 cells group was (92±31) mm3, and the tumor volume of HCT116 spherical cells group was (298±85) mm3, the tumor volume of the HCT116 spherical cells group was larger (t=9.27, P<0.05). ③ The expression of DVL3: the expression level of DVL3 in HCT116 cells was 0.12±0.05, and expression level of DVL3 in HCT116 spherical cells was 0.35±0.10, the expression level of DVL3 in HCT116 spherical cells was higher (t=4.31, P<0.05). Conclusions The HCT116 spherical cells have stronger colonization and tumorigenicity than the HCT116 cells. It has been speculatd that the high expression of DVL3 may be closely related with the stronger tumorigenicity.