Utilization of the promoter region of the midkine gene as a tool to drive therapeutic genes in a tumor specific manner

摘要

The high expression of midkine (MK), a heparin-binding growth factor, is frequently observed in a variety of tumors. We have developed a system to measure MK in human serum using enzyme-linked immunoassay and confirmed usefulness of measuring MK levelin sera of patients of neuroblasloma and esophageal cancer patients as a prognostic marker. We investigated the 2.3-kb MK promoter with a reporter assay for its transactivation of the fused lueiferase gene in various tumor cell lines and found that theminimal promoter activity resides in the region of 0.3-kb (MKp-0.3kb) adjacent to the transcription initiation site. Further analysis revealed that 0.6-kb fragment (MKp-0.6kb) spanning further upstream of Mkp-0.3kb mediated ihe preferential transcriptionin immortalized calls. MKp-0.6kb was linked to HSV-TK and then this construct was transfected into both lung cancer cell line (QG-56) and normal fibroblasts. The sensitivity to GCV of QG-56 transfected with MKP-0.6-HSV-TK was increased 200- 10,000 fold(IC50 = 0.1 to 0.5mug/ml) compared with that of control. Administration of GCV into the nude mice that developed tumors with MK0.6-HSV-TK transduelion completely suppressed tumor formation. When adenoviruses harboring MKp-0.6-kb fragment 5' of E1A gene were infected into normal and immortalized fibroblasts, the latter cells were more effectively lysed. These results collectively suggest that the tumor-specific expression of therapeutic gene(s) can be achieved by the use of cis-acting MK promoter.