title: Ribozyme gene therapy for hepatitis C virus infection.
abstract: BACKGROUND
  The development of antiviral drugs for hepatitis C virus (HCV) infection represents a substantial challenge. Similar to human immunodeficiency virus (HIV), HCV is highly prone to mutation. It is, therefore, expected that potential HCV therapeutics currently under development, such as protease inhibitors, will suffer from the same shortcomings of HIV therapeutic drugs; the emergence of drug resistant viral mutants. Ribozymes (Rz) are enzymatic RNA molecules that can be engineered to specifically target any given RNA molecule. A therapeutic Rz can be manufactured and administered as a drug, or a Rz gene can be delivered and expressed intracellularly by gene therapy. For HCV therapeutics, we favour the gene therapy approach as delivery and in vivo expression of Rz genes will result in a constant and continuous supply of multiple intracellular Rz, offering less opportunity for the development of drug-resistant viral variants.
  
  
  OBJECTIVE
  To utilise direct intravenous injection of hepatotropic viral vectors to transfer Rz genes directly into the hepatocytes of HCV-infected patients, resulting in degradation of the HCV positive strand RNA genome, the viral mRNAs, and even the negative strand RNA replication intermediate. We plan to circumvent the emergence of drug-resistant viral mutants by targeting multiple, highly conserved HCV RNA sequences simultaneously with multiple Rz genes expressed from a single vector.
  
  
  METHODS
  Rzs targeting conserved regions of the HCV positive and negative RNAs were transcribed in vitro and used to cleave HCV target RNAs. The most effective Rzs identified were then incorporated into adeno associated viral (AAV) vectors and adenoviral (AV) vectors and tested for their ability to inhibit HCV core expression in a tissue culture model.
  
  
  RESULTS
  Several Rzs targeting highly conserved HCV sequences effectively degraded positive and negative strands of HCV RNA in vitro. Furthermore, substantial inhibition of HCV gene expression was observed in tissue culture using viral vectors to deliver and express Rz genes.
  
  
  CONCLUSIONS
  Rz gene therapy has potential for the production of anti-viral drugs directed against HCV. Initial studies employing Rz gene therapy to produced anti-viral drugs against HCV have proved successful. Rz gene therapy may be a useful approach to overcome problems associated with anti-HCV drug design, such as the emergence of drug-resistant mutants.
authors.affiliation: Immusol Inc., San Diego, CA 92121, USA.
authors.forename: P J
authors.initials: PJ
authors.lastname: Welch
authors.forename: S
authors.initials: S
authors.lastname: Yei
authors.forename: J R
authors.initials: JR
authors.lastname: Barber
mesh_terms: D000256:Adenoviridae / Q000235:genetics; D002213:Capsid / Q000378:metabolism; D015870:Gene Expression; D015316:Genetic Therapy; D005822:Genetic Vectors; D016174:Hepacivirus / Q000235:genetics* / Q000378:metabolism; D006526:Hepatitis C / Q000628:therapy*; D006801:Humans; D016337:RNA, Catalytic / Q000235:genetics* / Q000378:metabolism*
chemical_list: D016337:RNA, Catalytic