Purpose: This study was performed to identify the detailed mechanisms by which miR-296-3p functions as a tumor suppressor to prevent lung adenocarcinoma (LADC) cell growth, metastasis, and chemoresistance.Experimental Design: The miR-296-3p expression was examined by real-time PCR and in situ hybridization. MTT, EdU incorporation, Transwell assays, and MTT cytotoxicity were respectively performed for cell proliferation, metastasis, and chemoresistance; Western blotting was performed to analyze the pathways by miR-296-3p and HDGF/DDX5 complex. The miRNA microarray and luciferase reporter assays were respectively used for the HDGF-mediated miRNAs and target genes of miR-296-3p. The ChIP, EMSA assays, and coimmunoprecipitation combined with mass spectrometry and GST pull-down were respectively designed to analyze the DNA-protein complex and HDGF/DDX5/β-catenin complex.Results: We observed that miR-296-3p not only controls cell proliferation and metastasis, but also sensitizes LADC cells to cisplatin (DDP) in vitro and in vivo Mechanistic studies demonstrated that miR-296-3p directly targets PRKCA to suppress FAK-Ras-c-Myc signaling, thus stimulating its own expression in a feedback loop that blocks cell cycle and epithelial-mesenchymal transition (EMT) signal. Furthermore, we observed that suppression of HDGF-β-catenin-c-Myc signaling activates miR-296-3p, ultimately inhibiting the PRKCA-FAK-Ras pathway. Finally, we found that DDX5 directly interacts with HDGF and induces β-catenin-c-Myc, which suppresses miR-296-3p and further activates PRKCA-FAK-Ras, cell cycle, and EMT signaling. In clinical samples, reduced miR-296-3p is an unfavorable factor that inversely correlates with HDGF/DDX5, but not PRKCA.Conclusions: Our study provides a novel mechanism that the miR-296-3p-PRKCA-FAK-Ras-c-Myc feedback loop modulated by HDGF/DDX5/β-catenin complex attenuates cell growth, metastasis, and chemoresistance in LADC. Clin Cancer Res; 23(20); 6336-50. ©2017 AACR. ©2017 American Association for Cancer Research.