Experimental study of dual promoter baculovirus-mediated tumor-targeting radioiodine therapy

Objective: To construct a recombinant baculovirus dual expression vector containing NIS gene under the control of human telomerase reverse transcriptase (hTERT) promoter and plasminogen kringle 5 (K5) gene driven by early growth response 1 (Egrl) promoter, and to explore the feasibility of targeting both tumor and tumor vessel with combination of radioiodide and antiangiogenic therapy. Methods: The hTERT-NIS gene and Egrl-K5 gene fragments were subcloned into baculovirus vector, then packaged and aMPIified in the sf9 cells to obtain recombinant baculovirus Bac-hTERT-NIS-Egrl-K5. Bac-CMV-NIS-Egrl-K5, Bac-hTERT-0-Egrl-K5 and Bac-hTERT-NIS-Egrl-0 were constructed as controls. The expression of NIS and K5 genes in human cervix cancers cells (HeLa) was examined by Western blot and quantitative real-time PCR. Functional NIS activity was confirmed by the uptake of ¹²⁵I, the inhibition of NaClO₄ and the cytotoxicity of ¹³¹I. The apoptotic effect of ¹³¹I-induced K5 on human umbilical veins endothelial cells (HUVEC) was analyzed by an apoptosis assay using flow cytometry. Statistical analysis was performed using the analysis of variance. Results: The recombinant baculovirus Bac-hTERT-NIS-Egrl-K5 was successfully constructed. The NIS gene under the control of hTERT promoter was specifically expressed in HeLa cells. The baculovirus-infected HeLa cells showed a significant increase of ¹²⁵I uptake, which was significantly inhibited by NaClO₄(F = 199.296, P < 0.05). Furthermore, a notable decreased cell survival rate (38.3%) was found after ¹³¹I treatment. The expression of K5 gene induced by ¹³¹I was elevated in a dose or time dependent manner and result ted in obvious inhibition with cell survival rate of 30.8% in baculovirus-infected HUVEC cells, which was significantly higher than that in the control groups (11.2% and 10.9% respectively, F = 19.926, 45.409; both P < 0.05). Conclusions: A recombinant baculovirus dual expression vector containing the NIS and K5 genes has been successfully constructed. This study suggests the feasibility of a synergistic strategy of NIS-based radioiodide therapy and X5-based antiangiogenic therapy in vitro, and make it possible to perform in vivo study in the near future. (authors)