Inhibition of cell proliferation in HCC-9204 hepatoma cells by a c-myc specific ribozyme

Abstract

A ribozyme (RZ) gene targeting c-myc mRNA was synthesized and cloned. Cleavage reaction showed that cleavage of the RZ was efficient and specific. The RZ gene-containing retrovirus vector pDOR-RZ was transfected into HCC-9204 hepatoma cells, which constitutively express high levels of c-myc using Lipofectamine. Positively transfected cells were selected using G418. In situ hybridization showed that both pDOR-RZ and pDOR vectors had been integrated into the chromosome of HCC-9204 cells. Dot blot hybridization indicated that expression of the RZ was only evident in pDOR-RZ-transfected HCC-9204 cells. Avidin-biotin complex enzyme-linked immunosorbent assay showed that c-myc expression was down-regulated. Chromatin aggregation into compact masses, cytoplasmic vacuole degeneration, and blurring of cytoplasm structure were observed by transmission electron microscopy in HCC-9204-RZ cells. These results suggest that the use of a c-myc mRNA cleaving enzyme could be most effective in tumor cells that are highly proliferative and constitutively express high levels of c-myc.