Inhibition Of Hepatitis B Virus Expression By Multi-target Ribozymes

Hepatitis B virus(HBV) , the causative agent of B-type hepatitis in human, is a small enveloped DNA virus that replicates through reverse transcription of an RNA intermediate, the terminally redundant RNA pregenome. The core protein , the most important component in formation of HBV nucleocapsid, is closely related to the replication of HBV DNA. Cleavage of the pregenomic RNA and/or mRNA of core gene, therefore, may interrupt the life cycle of HBV. Hammerhead ribozymes are short catalytic RNA molecules possessing endoun-clease activity. The enzymes specificity is mediated by variable flanking sepuences complementary to the RNA target substrate. In this report, we describe multitarget ribozymes construct that cleave a critical region of the HBV pregenomic RNA and the ribozymes expression in the HepG2 cells.By computer program, we first analysed the secondary structure of target RNA, and designed two cis —ribozymes (intramolecularly) and three trans —ribozymes (intermolecularly), which each one is specific for a different cleavage site on the mRNA of core gene. Then we constructed plasmids combining 5' and 3' trimming system (two cis—ribozymes) with each ribozyme gene or connected -type ribozyme genes containing different two ribozyme genes and a triple ribozyme gene. These plasmids were designed to release antiviral molecules (trans...