Inhibition of telomerase activity by HDV ribozyme in cancers

A pseudo-knotted HDV ribozyme (g.RZ57) directed against the RNA component of human telomerase (hTR) was designed and synthesized. An in vitro transcription plasmid and a eukaryotic expression plasmid of ribozyme were constructed. The eukaryotic expression plasmid was induced into heptocellular carcinoma 7402 cells, colon cancer HCT116 cells and L02 hepatocytes respectively. Then we determine the cleavage activity of ribozyme against human telomerase RNA component (hTR) both in vitro and in vivo, and detect telomerase activity continuously.HDV ribozyme showed a specific cleavage activity against the telomerase RNA in vitro. The maximum cleavage ratio reached about 70.4%. Transfection of HDV ribozyme into 7402 cells and colon cancer cells HCT116 led to growth arrest and the spontaneous apoptosis of cells, and the telomerase activity dropped to 10% of that before.HDV ribozyme (g.RZ57) is an effective strategy for gene therapy.Immortalized and malignant tumor cells are characterized by unlimited cell proliferation and programmed cell death (apoptosis). It has been demonstrated that malignant transformation occurs when the telomerase in normal cell is activated [1,2].Telomerase activity is found in almost all malignant tumors [3]. Human telomerase RNA (hTR) is associated with the activity of telomerase, immortalized cancer cells retain the highest level of hTR [4,5]. In recent years, hammerhead ribozymes were used to inhibit the telomerase activity by targeting the template region of telomerase RNA in malignant tumors [6,7]. Yet, there is no report about HDV ribozyme for inhibition of telomerase activity.Ribozymes are catalytic RNA molecules which can be designed to specially cleave a target RNA sequence by incorporating the flanking sequence complementary to the target[8]. Like other ribozymes, HDV ribozyme has this property. So it may have a potential application in gene therapy in which an engineered ribozyme is directed to inhibit gene expression by targeting a speci