Hepatocellular carcinoma (HCC) is a commonly occurring malignant tumor, with a high incidence rate. The present study aimed to investigate the effect of knocking down the N‑glycosyltransferase‑V (GnT‑V) protein on the proliferation, migration and invasion of the human HCC drug‑resistant cell line, SMMC7721/R. SMMC7721/R cells with GnT‑V‑knockdown (SMMC‑7721/R‑GnT‑V) were constructed using the method of lentiviral transfection. The expression of GnT‑V was assessed using reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR) and western blotting. Cell proliferation was determined using an MTT assay, and the extent of cellular apoptosis was assessed using flow cytometric analysis. Additionally, the metastatic ability of the cells in vitro was analyzed using cell adhesion and invasion assays. Western blotting was used to investigate the protein expression levels of caspase‑3, caspase‑9, Bcl‑2, Bax, matrix metalloproteinase (MMP)‑2 and MMP‑9, and RT‑qPCR was used to determine the mRNA expression levels of the genes for the breast cancer resistance protein and P‑glycoprotein in the SMMC‑7721/R cells. Taken together, the results of the present study revealed that the knockdown of GnT‑V significantly suppressed the proliferation, migration and invasion (P<0.05) of the SMMC‑7721/R cells. Furthermore, the possible mechanism underlying these phenomena may be associated with the induction of mitochondria‑mediated apoptosis, inhibition of the degradation of the extracellular matrix and an enhancement of the drug-sensitivity. GnT‑V‑knockdown may therefore be used to treat drug‑resistant HCC in the future.