Feasibility of a novel positive feedback effect of 131I-promoted Bac–Egr1–hNIS expression in malignant glioma through baculovirus

Abstract

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. 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. 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. 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.