Hepatocellular carcinoma (HCC) is one of the most common malignant tumors worldwide. Currently, the clinical strategies available for the treatment of HCC remain insufficient for the poor prognosis. Sodium/iodide symporter (NIS)-based radioiodine therapy is proposed as a promising therapeutic strategy for the treatment of HCC. However, it is difficult for HCC cells to trap iodine for the lower expression of NIS. Small activating RNA (saRNA) is a newly identified small double-stranded RNA (dsRNA) that can induce endogenous gene expression by targeting promoter sequences. Here, we designed an saRNA (saRNA-482) that targeted the NIS promoter sequences. In the cultured HepG2 cells and Hep3B cells, the expressions of NIS were upregulated after transfection of saRNA-482. In addition, the uptake ofI increased in the cultured HepG2 and Hep3B cells transfected with saRNA-482. Furthermore, the cell viabilities were significantly inhibited in the saRNA-482-transfected HepG2 and Hep3B cells after I treatment. Meanwhile, the apoptosis of saRNA-482-transfected HepG2 and Hep3B cells significantly increased afterI treatment. The results suggest that RNA activation-mediated upregulation of NIS may have an endoradiotherapeutic potential in the treatment of HCC.