Hexokinase Type II: A Novel Tumor-Specific Promoter for Gene-Targeted Therapy Differentially Expressed and Regulated in Human Cancer Cells

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Abstract
The use of tissue- or tumor-selective promoters in targeted gene therapy for cancer depends on strong and selective activity. Hexokinase type II (HK II) catalyzes the first committed step of glycolysis and is overexpressed in tumors, where it is no longer responsive to normal physiological inhibitors, e.g., glucagon. We show, in a reporter gene assay, activation of HK II in non-small cell lung carcinomas NCI-H661 and NCI-H460 at 61 and 40%, respectively, relative to the activation observed with a constitutive promoter, while it was only 0.9% in different preparations of primary normal human bronchial epithelial cells (NHBECs). Similar results were observed in a variety of normal and tumor cells. Moreover, treatment of the transfectants with glucagon did not inhibit promoter activation in the transformed H661 cells, while endogenous HK II in NHBECs is suppressed by glucagon. H460 and H661 cells infected with a recombinant adenovirus carrying an HK II/LacZ expression cassette, AdHexLacZ, demonstrated beta-galactosidase activity that correlated with the level of HK II promoter activation in these cells. Under similar conditions, no enzyme activity was observed in NHBECs. Cells were then infected with AdHexTk and treated with GCV. Our results demonstrate selectivity in toxicity, with a 10- to 100-fold increase in IC50 between lung cancer cell lines H661 and H460, respectively, and NHBECs. There was also a 100-fold increase in IC50 in NHMECs relative to breast carci- noma cell line MCF-7. In HepG2 cells, an IC50 of 1mu g/ml was observed, comparable to that of other tumor cell lines. This represents a novel use of the hexokinase type II as a selective promoter in cancer gene therapy.