SBIR-STTR Award

Hepatitis C virus (HCV) is the major cause of non-A, non-B hepatitis, which is a serious worldwide health problem. Our long term objective is to develop hairpin Ribozyme (Rz) gene therapy for the treatment of HCV infection. Immusol's Rz core technology, developed as a result of the success of its HIV program, is based upon the ability to engineer and optimize hairpin Rz genes whose transcribed RNAs can specifically degrade almost any undesirable RNA molecule. HCV is especially suited for ribozyme-mediated therapy since the viral genome is present exclusively in the form of RNA. HCV encodes only one primary transcript, thus, cleavage of any target sequence within this RNA should abolish virus expression. Immusol has generated two hairpin ribozymes target at HCV (+) strand RNA as potential therapeutic agents for HCV infection (Gene Therapy, 1996, 3: 994). Immusol also plans to target the negative strand RNA that is an essential intermediate in HCV replication. Degradation of this RNA may be more effective, since its intracellular level is lower than that of the positive strand RNA. Therefore, we propose a Phase I SBIR studies aimed at: 1) to synthesize and optimize effective ribozymes that target HCV positive- (+) and negative- (-) strand RNA; 2) to generate viral vectors (AV and AAV) that express active hairpin ribozymes; and 3) to test vector- derived ribozyme activity in reducing HCV gene expression in liver cell culture Achieving these three Aims will generate the most effective ribozyme viral vector(s) for preclinical studies. PROPOSED COMMERCIAL APPLICATION: Immusol plans to commercially develop a Rz gene therapy approach to HBV infection. We will manufacture and provide Rz-expressing viral vectors in a vial. The vectors will then be provided to medical centers around the world that will carry out the in vivo vector-mediated gene transfer into patient liver cells to combat HBV virus.

Thesaurus Terms:
antiviral agent, chemical synthesis, hepatitis C virus, ribozyme, transfection, virus RNA Adenoviridae, adeno associated virus group, enzyme activity, gene therapy, hepatitis C, transfection vector RNase protection assay, molecular cloning, tissue /cell culture, western blotting

This Phase II SBIR proposal is aimed at the development of hairpin Ribozyme (RZ) gene therapy for the treatment of Hepatitis C virus (HCV) infection. HCV is the major cause of non-A, non-B hepatitis, which is a serious worldwide health problem, and ribozymes are RNA molecules that can be engineered to cleave and inactivate other RNA molecules in a sequence-specific fashion. The investigators believe that HCV is especially suited for ribozyme-mediated therapy, because the viral genome is present exclusively in the form of RNA and because HCV encodes only one primary transcript. Therefore, the applicants reason that cleavage of any target sequence within this RNA should abolish virus expression. Immusol has generated two hairpin ribozymes targeting the HCV positive strand RNA, and in the Phase I component of this project, additional Rz were identified targeting the negative strand, an even more attractive target due to its low expression level. In this Phase II proposal, the Principal Investigator proposes the optimization of the catalytic activity and stability of the top six Rz. Hepatocyte specific promoters will be used to express the Rz as single or polycistronic transcripts and the effectiveness of these multi-ribozyme vectors will be tested in a tissue culture reporter model. Development of an HCV replication system in liver cell culture is proposed, for final testing of the ribozyme vector before pre-clinical safety evaluation in a rat toxicity study. PROPOSED COMMERCIAL APPLICATION: NOT AVAILABLE

Thesaurus Terms:
antiviral agent, enzyme activity, gene therapy, hepatitis C virus, ribozyme, technology /technique development, transfection, virus RNA Adenoviridae, adeno associated virus group, genetic promoter element, hepatitis C, liver cell, plasmid, reporter gene, transfection vector human tissue, laboratory mouse, laboratory rat, molecular cloning, tissue /cell culture