Feasibility of a novel positive feedback effect of 131I-promoted Bac-Egr1-hNIS expression in malignant glioma through baculovirus: a comparative study with Bac-CMV-hNIS.

摘要

OBJECTIVE: Increased expression of sodium iodide symporter (NIS) is required for reporter gene imaging and effective radioiodine treatment of tumor. As the early-growth response-1 (Egr1) promoter is activated by radioisotopes, the existence of a positive feedback effect of I-promoted Egr1-hNIS expression is possible. Compared with a widely used cytomegalovirus (CMV) promoter, we investigated a possible increased activity of I-stimulated human NIS (hNIS) transgene expression in malignant glioma using a baculovirus vector containing the Egr1 promoter. METHODS: Recombinant baculovirus (Bac-CMV-hNIS) encoding the hNIS gene under the control of the CMV promoter and Bac-Egr1-hNIS encoding the hNIS gene under the control of the radiation-inducible Egrl promoter were constructed. After human malignant glioma U87 cells were transfected with Bac-CMV-hNIS or Bac-Egr1-hNIS, stimulated with or without I, the expression of the hNIS protein was detected by immunofluorescence and a flow cytometry test. The uptake and efflux of iodine were determined after the incubation of the transfected cells with I. RESULTS: Immunocytochemical staining and flow cytometry test showed a lower hNIS protein expression in U87 cells transfected with Bac-Egr1-hNIS (even after I stimulation) compared with U87 cells transfected with Bac-CMV-hNIS. Bac-CMV-hNIS-transfected U87 cells accumulated up to approximately 25.8 times more I than nontransfected cells, whereas Bac-Egr1-hNIS-transfected U87 cells accumulated up to approximately 3.5 and 14.2 times more I pre-stimulation and post-stimulation. However, rapid efflux of radioactivity was observed in both groups, with 50% lost during the first 2 min after the I-containing medium was replaced by a nonradioactive medium. CONCLUSION: Our results indicated that an improved transgene expression of I-stimulated hNIS in U87-malignant glioma cells using a baculovirus vector containing the Egr1 promoter is possible, but the expression level is lower than that of Bac-CMV-hNIS-transfected U87 cells. However, it might be an approach to improve the specificity of gene therapy using radiosensitive promoters to activate hNIS gene expression selectively in the radiation field.