Combined RNA Interference of Hexokinase II and 131I-Sodium Iodide Symporter Gene Therapy for Anaplastic Thyroid Carcinoma

Abstract

The purpose of this study was to investigate the enhanced therapeutic effect of the combined use of shRNA (small hairpin RNA) therapy for the hexokinase II (HKII) gene and (131)I human sodium iodide symporter (hNIS) as a gene therapy for in vitro and in vivo treatment of anaplastic thyroid carcinoma cells (ARO) in an animal model. A recombinant lentivirus containing a plasmid with the hNIS gene driven by phosphoglycerate kinase promoter and green fluorescent protein (GFP) linked with an internal ribosome entry site sequence was produced. ARO cells were transfected with the virus and sorted by fluorescent activated cell sorting using GFP (ARO-NG). The messenger RNA expression of hNIS and GFP were evaluated with reverse-transcriptase polymerase chain reaction, and the function of hNIS was verified by (125)I uptake. The lentiviral vector expressing shRNA against HKII (Lenti-HKII shRNA) was constructed and used to infect ARO-NG cells. The effect of Lenti-HKII shRNA was evaluated by reverse-transcriptase polymerase chain reaction, (18)F-FDG uptake, and HK activity. An in vitro clonogenic assay was performed after Lenti-HKII shRNA therapy, (131)I therapy, and a combined therapy. The therapies were also applied in vivo to an animal model with an ARO-NG xenograft, and the effects were assessed with caliper measurements and (18)F-FDG PET. ARO-NG cells showed an (125)I uptake 76-fold higher than the parent ARO cells. Compared with the uninfected ARO-NG cells, ARO-NG cells infected with Lenti-HKII shRNA had lower HKII messenger RNA expression, lower (18)F-FDG uptake, and HK activity. The proliferation of ARO-NG cells was inhibited by (131)I and Lenti-HKII shRNA therapies and further inhibited by the combined (131)I and Lenti-HKII shRNA therapy. Both the Lenti-HKII shRNA therapy and the (131)I therapy inhibited in vivo tumor growth in the tumor xenograft model. The combined Lenti-HKII shRNA and (131)I therapy resulted in a further decrease of tumor growth. Our results suggest that the combined HKII shRNA and (131)I therapy has a stronger antitumor effect than either the (131)I therapy or the HKII shRNA alone. Therefore, this combined therapy could be used as a powerful strategy for treating anaplastic thyroid carcinoma.