Induction of the Suicide HSV–TK Gene by Activation of the Egr-1 Promoter with Radioisotopes

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Abstract
In investigating new methods for the treatment of pancreatic cancer, we have explored the possibility of using a combination of radiation and gene therapies. We demonstrate herein that the early growth response gene 1 promoter (Egr-1) is sufficient to confer selective expression of the luciferase gene (Luc) in a human pancreatic tumor cell line (AsPc-1) when exposed to ionizing radiation. The Egr-1 promoter directed the radioinducible expression of luciferase, and yielded higher levels of Luc activity than that in nonirradiated lines. The radioisotopes Tc-99m, I-131, and Ga-67-citrate were selected as Egr-1 activators for their potential to accumulate in tumors. We studied Ga-67-citrate, a radioisotope employed in tumor scintigraphy, for its suitability for selective gene induction. The plasmid vector pEgr-1-Luc was transfected into AsPc-1 cells and then exposed to radioisotopes. Luciferase activity increased by 100–300 times over control. We also inserted the herpes thymidine kinase gene (TK) downstream of Egr-1 and transfected this construct into AsPc-1 cells. Ga-67-citrate and ganciclovir were added to the cells and cell survival was assessed by MTT assay. The growth of AsPc-1 cells transfected with the pEgr-TK construct was suppressed 2 days after exposure of the cells to Ga-67-citrate. The results indicate that Ga-67-citrate may be useful in combining radiation and gene therapies. A potential problem of gene therapy for cancer is nonselective induction of therapeutic genes. The goal of this study was to achieve selective gene induction using a tumor-accumulating radioisotope and a radiosensitive promoter. We found that radioisotopes can activate early growth response gene 1 (Egr-1) transcription in vitro and induce growth arrest of cancer cells transfected with a pEgr-TK construct. These findings suggest that radioisotopes may be useful for increasing selectivity of gene therapy and for the treatment of metastatic lesions.