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Background and objective To explore the function and the potential molecular mechanism of miR-218 in lung cancer cell. Methods The expression of miR-218 mRNA was determined by real-time PCR in lung cancer tissues, adjacent tissues and lung cancer cells. Transwell assay was used to detect the migration and invasion of A549 cell after transfected with Anti-miR-218 or negative control and HC4006 cell after transfected with miR-218 mimics and miR-218 negative control. Targetscan and MiRanda were used to calculate the potential targets of miR-218 and Luciferase reporter assay was performed to identify that the Robo1 was one target genes of miR-218. Transwell assay was used to detect whether miR-218 regulated the invasion of lung cancer cell transfected with anti-miR-218 or negative control via Robo1. Results The expression of miR-218 in the lung cancer tissues was significantly lower than that in the adjacent tissues (P<0.05). Inhibition of miR-218 improved the migration and invasion of A549 cell. Overexpression of miR-218 suppressed the migration and invasion of HCC4006 cell. The co-transfection of anti-miR-218 or miR-218 mimics and the Robo1 3′UTR increased or reduced the luciferase activity of Robo1 compared with the control group (P<0.05). Inhibition of miR-218 and Robo1 recovered the invaded cells of A549. Overexpression of miR-218 and inhibition of Robo1 reduced the number of the invased cells of HCC4006. These results suggested that miR-218 banded Robo1 directly and inhibited lung cancer cell invasion by targeting Robo1. Conclusion MiR-218 inhibited the migration and invasion of lung cancer cells through regulating Robo1 expression.