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Recent studies reported that long noncoding RNAs (LncRNAs) were involved in tumorigenesis of various human cancer types, including gastric cancer (GC) through targeting microRNAs (miRNAs/miRs). The present study investigated the biological functions of LncRNA SOX2 overlapping transcript (SOX2OT)/miR-194-5p axis and its underlying mechanism in the tumor progression of GC. The results showed that relative expression of LncRNA SOX2OT was highly upregulated while the expression of miR-194-5p was down-regulated in GC tissues and cell lines (MGC-803, SGC-7901, MKN-74). Knockdown of SOX2OT inhibited cell proliferation, invasion and migration of GC cells (MGC803, MKN-74) through reducing epithelial-mesenchymal transition (EMT). Moreover, miR-194-5p was predicted to be one of the targets of SOX2OT through bioinformatics analysis and was verified by luciferase reporter assay. miR-194-5p expression was negatively regulated by SOX2OT expression in GC cells and miR-194-5p inhibitor was found to counteract the inhibitory effects of SOX2OT short hairpin (sh)RNA on cell proliferation and mobility through enhancing EMT in GC cells. Taken together, the in vitro experiments revealed that knockdown of SOX2OT inhibited cell proliferation and mobility through suppressing EMT via targeting miR-194-5p in GC. In addition, results from in vivo experiments showed that knockdown of SOX2OT suppressed GC tumor growth and matrix metalloproteinase (MMP)-2 and MMP-9 expression through inhibiting EMT. Besides that, relative expression of miR-194-5p was increased in sh-SOX2OT group compared with sh-NC group. In summary, our study elucidated that the SOX2OT/miR-194-5p axis participated in the tumor progression of GC through regulation of EMT both in vitro and in vivo. Hence, targeting the SOX2OT/miR-194-5p axis may aid in establishing novel strategies for therapy of GC. [ABSTRACT FROM AUTHOR]