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We aimed to investigate the role of microRNA-5100 (miRNA-5100) in oral squamous cell carcinoma (OSCC) and its underlying mechanisms. Material/Methods: The expression of miR-5100 and suppressor of cancer cell invasion (SCAI) in OSCC cell lines were examined. A luciferase reporter assay was applied to confirm the combination between miR-5100 and SCAI. Then, miR-5100 inhibitor or small hairpin RNA (shRNA)-SCAI were transfected into cells. Cell Counting Kit-8 assay was executed for testing cell proliferation ability. Flow cytometry assay was exploited for measuring cell cycle. Invasion and migration of OSCC cells were assessed using Transwell assay and wound healing assay. The expression of proteins were detected using western blotting. The results demonstrated that the level of miR-5100 was upregulated while SCAI was downregulated in OSCC cells. SCAI was verified as a direct target of miR-5100. MiR-5100 silencing suppressed proliferation of OSCC cells, increased cells in the G1 and G2 phases, and reduced those in the S phase, which was reversed after transfection with shRNA-SCAI. Moreover, miR-5100 inhibitor downregulated the expression of cyclin-dependent kinase-2 (CDK-2) and cyclinD1, accompanied by upregulation in p27 expression, whereas SCAI silencing had the opposite results. The invasion and migration abilities of OSCC cells were reduced after treatment with miR-5100 inhibitor, whereas SCAI silencing suppressed the effects of miR-5100 inhibitor on OSCC cell behaviors. These findings suggested that miR-5100 silencing inhibit proliferation, invasion and migration of OSCC cells via upregulating the expression of SCAI, which provides theoretical basis and treatment strategies for the treatment of OSCC. [ABSTRACT FROM AUTHOR]