It has been reported that high-mobility group box 3 is overexpressed in various cancers. This study aimed to explore its function in non-small cell lung cancer (NSCLC). A546 and H460 cell lines were used for in vivo experiments, scratch healing tests, transwell migration and invasion experiments. It was first found that HMGB3 was highly expressed in tumor tissues in the patients and associated with NSCLC stage. Silencing of HMGB3 significantly slowed the growth, proliferation and invasion of NSCLC in vitro, and repressed cell growth in vivo. Mechanistic studies suggest that the observed effects were mediated by inhibiting the expression of β-catenin/MMP7/c-Myc in Wnt pathway. Our study highlights the role of HMGB3 in NSCLC, which may provide a therapeutic target for the treatment of NSCLC.