Background: HOXD9, a Hox family member, is involved in cancer growth and metastasis. But, its regulation mechanism at the molecular level particularly in colorectal cancer (CRC), is mostly unknown. Methods: The HOXD9 protein expression levels were analyzed using immunofluorescence, immunohistochemistry (IHC) assays, and western blot. The in vivo and in vitro roles of HOXD9 in CRC were determined using colony formation and EdU incorporation, CCK-8, wound scratch and transwell invasion assay, and animal models. Results: Expression of HOXD9 was higher in CRC than in matched healthy tissues. High expression of HOXD9 has significantly associated with the American Joint Committee on Cancer (AJCC) stages, tumor differentiation, lymph node metastasis, and other serious invasions, and it had a poor prognosis. In vitro, HOXD9 encouraged proliferation, movement and EMT processes in cells of CRC. Also, TGF-β1 promoted the expression of HOXD9 and this effect was dependent on the dose and downregulation of HOXD9 repressed TGF-β1 -induced EMT. In vivo, HOXD9 promoted the invasive and metastasis of CRC cells via orthotopic implantation. Conclusions: The ectopic expression of HOXD9 promoted the invasion metastasis in cells of the colorectal tumor by induction of EMT in vitro and vivo. © 2020 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.