Abstract: Background　Human Epidermal Growth Factor Receptor 2 (HER2) is frequently overexpressed in breast cancer. Fenretinide (4-HPR), a synthetic retinoid derivative, has demonstrated potent inhibitory effects against various malignancies, including breast and prostate cancers.Objective　This study aims to investigate the impact of fenretinide on the migratory capacity of HER2-overexpressing lung cancer cells and to elucidate the underlying molecular mechanisms.Methods　The expression profile of the HER2 gene in lung cancer and its correlation with patient survival and prognosis were analyzed using The Cancer Genome Atlas (TCGA) database. The effects of Fenretinide on the proliferation and migration of the HER2-overexpressing lung adenocarcinoma cell line CALU-3 were validated via CCK-8, scratch assays, and Transwell migration assays. Furthermore, transcriptomic changes in CALU-3 cells following Fenretinide treatment were analyzed using RNA sequencing (RNA-seq). Key mechanistic targets were subsequently explored and validated through RT-qPCR. Results　HER2 is overexpressed in lung adenocarcinoma and is significantly associated with patient survival and prognosis. Fenretinide inhibited the proliferation of both A549 cells and HER2-overexpressing CALU-3 cells, but exhibited a stronger inhibitory effect on the migratory capacity of CALU-3 cells. RNA sequencing analysis revealed that the epithelial–mesenchymal transition (EMT) pathway, which is closely associated with metastasis, was suppressed following fenretinide treatment. Further RT-qPCR analysis demonstrated that the expression levels of N-cadherin, Slug, Snail, and MMP9 were significantly downregulated (P＜0.01), while E-cadherin was significantly upregulated (P＜0.05). Western blot analysis showed that the phosphorylation levels of HER2, FAK, and Src were markedly reduced in CALU-3 cells after fenretinide treatment. Conclusion　Fenretinide inhibits the activation of HER2 and its downstream FAK/Src signaling pathway and suppresses EMT, thereby reducing the migratory ability of HER2-overexpressing lung adenocarcinoma cells.