Objective To investigate the inhibitory effects of procaine on the proliferation, migration and invasion of osteosarcoma cells through the extracellular signal-regulated kinase (ERK) / mitogen-activated protein kinase (MAPK) / focal adhesion kinase (FAK) pathway. Methods Human osteosarcoma Saos-2 cells were cultured in vitro and divided into control (without drugs), procaine ( 2 μmol / L procaine), Y15 ( 10 μmol / L Y15) and procaine + Y15 ( 2 μmol / L procaine + 10 μmol / L Y15) groups. Proliferation of Saos-2 cells was assess by the CCK-8 method. Apoptosis of Saos-2 cells was detected by flow cytometry. The migration and invasion of Saos-2 cells were assessed by the transwell method. Expression of ERK/ MAPK/ FAK pathway-related proteins in Saos-2 cells was detected by Western blot. Results Compared with those of the control group, the Saos-2 cell survival rate, migrated cell number, invasive cell number and protein expression levels of p-ERK/ ERK, p-p38 MAPK/ p38 MAPK and p-FAK/ FAK in procaine, Y15 and procaine + Y15 group were significantly lower (P<0. 05) and the apoptotic rate was significantly higher (P<0. 05). Compared with those in the Y15 group, there was no significant difference in the Saos-2 cell survival rate, apoptotic rate, migrated cell number, invasive cell number, or protein expression levels of p-ERK/ ERK, p-p38 MAPK/ p38 MAPK and p-FAK/ FAK in the procaine + Y15 group ( P> 0.05). Conclusions Procaine inhibits the proliferation, migration and invasion of osteosarcoma Saos-2 cells, which may be mediated by inhibiting activation of the ERK/ MAPK/ FAK pathway.