Objective To explore the mechanism of capsaicin in inhibiting the invasion and migration of colon cancer cells by upregulating tumor metastasis suppressor 1 ( MTSS1). Methods Experiment 1: 10, 25, 50 and 100 μmol / L capsaicin groups were treated with capsaicin, whereas the control group was untreated and cultured at a constant temperature for 24 hours. Experiment 2: the control group was cultured normally, the 100 μmol / L capsaicin group was treated with 100 μmol / L capsaicin, 100 μmol / L capsaicin + control siRNA and 100 μmol / L capsaicin +MTSS1 siRNA groups were treated with 50 nmol / L control siRNA and MTSS1 siRNA, respectively, based on the 100 μmol / L capsaicin group. After transfection for 4 h, 100 μmol / L capsaicin-containing culture medium was added. Cell proliferation was assessed by CCK8 assays. Cell invasion was assessed by Transwell assays. Cell migration was assessed by a scratch test. The protein levels of transient receptor potential vanilloid 1 (TRPV1), MTSS1, matrix metalloproteinase (MMP)-2, and MMP-9 were measured by Western blot. Results After treatment with capsaicin at various concentrations, compared with the control group, cell proliferation in 25, 50 and 100 μmol / L capsaicin groups was higher, the protein levels of TRPV1 and MTSS1 in 10, 25, 50 and 100 μmol / L capsaicin groups were higher, and cell invasion, the percentage of migrated cells and the protein levels of MMP-2 and MMP-9 were lower (P<0.05). After RNA interference of MTSS1 and addition of 100 μmol / L capsaicin, compared with the control group, cell invasion, the percentage of migrated cells, and the protein levels of MMP-2 and MMP-9 were lower in 100 μmol / L capsaicin, 100 μmol / L capsaicin+control siRNA, 100 μmol / L capsaicin+MTSS1 siRNA groups (P<0. 05), and the protein level of MTSS1 was higher (P<0.05). Compared with 100 μmol / L capsaicin and 100 μmol / L capsaicin+control siRNA groups, cell invasion, the percentage of migrated cells, the protein levels of MMP-2 and MMP-9 were significantly higher in the 100 μmol / L capsaicin+MTSS1 siRNA group and the level of MTSS1 protein was lower (P<0.05). Conclusions Capsaicin inhibits the invasion and migration of colon cancer cells by upregulating the expression of MTSS1, thereby exerting a protective effect against colon cancer.