Objective To construct nuclear protein 1-short hairpin RNA (NUPR1-shRNA)-expressing lentiviral vector targeting NUPR1 gene, and investigate the effect of NUPR1 gene silencing on the proliferation and apoptosis of human multiple myeloma U266 cells. Methods The normal plasma cells were used as controls and the mRNA expression level of NUPR1 in myeloma cell lines U266 and RPMI8226 were detected by quantitative real-time PCR (qRT-PCR). Then the shRNA plasmid targeting NUPR1 gene was constructed and transfected into U266 cells. The transfection efficiency was detected by flow cytometry. The interference effect of NUPR1 gene was detected by qRT-PCR and Western blotting. The cell proliferation was analyzed by CCK-8 assay and trypan blue viability counting. The cell apoptosis was determined by flow cytometry and Hoechst staining. Results The mRNA expression level of NUPR1 in U266 and RPMI8226 were higher than that in the normal plasma cells. NUPR1-shRNA-expressing lentiviral vector was successfully constructed, and flow cytometry shows that the transfection efficiency was above 80%. Compared with the negative control group and the blank control group, the proliferation of U266 cells was significantly inhibited with the inhibitory rate being (58.71±1.64)%, and the apoptosis rate of U266 cells was significantly elevated in NUPR1 knockdown group. Conclusion Down-regulating the expression of NUPR1 in U266 cells can inhibit cell proliferation and promote its apoptosis.