Targeting internal ribosome entry site (IRES)-mediated translation to block hepatitis C and other RNA viruses

Department of Microbiology, Immunology and Molecular Genetics, UCLA School of Medicine, University of California-Los Angeles, 10833 Le Conte Avenue, Los Angeles, CA 90095, USA.
FEMS Microbiology Letters (Impact Factor: 2.12). 06/2004; 234(2):189-99. DOI: 10.1016/j.femsle.2004.03.045
Source: PubMed


A number of RNA-containing viruses such as hepatitis C (HCV) and poliovirus (PV) that infect human beings and cause serious diseases use a common mechanism for synthesis of viral proteins, termed internal ribosome entry site (IRES)-mediated translation. This mode of translation initiation involves entry of 40S ribosome internally to the 5' untranslated region (UTR) of viral RNA. Cap-dependent translation of cellular mRNAs, on the other hand, requires recognition of mRNA 5' cap by the translation machinery. In this review, we discuss two inhibitors that specifically inhibit viral IRES-mediated translation without interfering with cellular cap-dependent translation. We present evidence, which suggest that one of these inhibitors, a small RNA (called IRNA) originally isolated from the yeast Saccharomyces cerevisiae, inhibits viral IRES-mediated translation by sequestering both noncanonical transacting factors and canonical initiation factors required for IRES-mediated translation. The other inhibitor, a small peptide from the lupus autoantigen La (called LAP), appears to block binding of cellular transacting factors to viral IRES elements. These results suggest that it might be possible to target viral IRES-mediated translation for future development of therapeutic agents effective against a number of RNA viruses including HCV that exclusively use cap-independent translation for synthesis of viral proteins.

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Available from: Raquel Izumi, Feb 11, 2015
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    • "HCV 5′ UTR is the most conserved locus within its genome and thus efforts related to HCV RNA therapeutics have been focused on this locus [30]. Nevertheless, this region has a highly stable RNA structure and is modulated by miRNAs and RNA-binding proteins, which limits the number of accessible sites for ribozyme targeting [31]. "
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    ABSTRACT: Background Hepatitis C virus (HCV) is a human pathogen causing chronic liver disease in about 200 million people worldwide. However, HCV resistance to interferon treatment is one of the important clinical implications, suggesting the necessity to seek new therapies. It has already been shown that some forms of the catalytic RNA moiety from E. coli RNase P, M1 RNA, can be introduced into the cytoplasm of mammalian cells for the purpose of carrying out targeted cleavage of mRNA molecules. Our study is to use an engineering M1 RNA (i.e. M1GS) for inhibiting HCV replication and demonstrates the utility of this ribozyme for antiviral applications. Results By analyzing the sequence and structure of the 5′ untranslated region of HCV RNA, a putative cleavage site (C67-G68) was selected for ribozyme designing. Based on the flanking sequence of this site, a targeting M1GS ribozyme (M1GS-HCV/C67) was constructed by linking a custom guide sequence (GS) to the 3′ termini of catalytic RNA subunit (M1 RNA) of RNase P from Escherichia coli through an 88 nt-long bridge sequence. In vitro cleavage assays confirmed that the engineered M1GS ribozyme cleaved the targeted RNA specifically. Moreover, ~85% reduction in the expression levels of HCV proteins and >1000-fold reduction in viral growth were observed in supernatant of cultured cells that transfected the functional ribozyme. In contrast, the HCV core expression and viral growth were not significantly affected by a “disabled” ribozyme (i.e. M1GS-HCV/C67*). Moreover, cholesterol-conjugated M1GS ribozyme (i.e. Chol-M1GS-HCV/C67) showed almost the same bioactivities with M1GS-HCV/C67, demonstrating the potential to improve in vivo pharmacokinetic properties of M1GS-based RNA therapeutics. Conclusion Our results provide direct evidence that the M1GS ribozyme can function as an antiviral agent and effectively inhibit gene expression and multiplication of HCV.
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    • "These observations are in agreement with several reports suggesting that, beside their primary role in pol III and non-coding pol II stable RNAs biosynthesis, La homologues could be implicated in mRNA translation enhancement [for review see (5)]. For example, by binding to their 5′-UTR, the cytoplasmic La protein stimulates the internal ribosome entry site-mediated translation of viral mRNAs (32,33) as well as certain cellular mRNAs (34,35). Also, La is involved in the cap-dependent translation of 5′-terminal oligopyrimidine stretch (TOP) containing mRNAs (27). "
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    • "The approval of Vitravene as antisense drug for treatment of cytomegalovirus (CMV)-induced retinitis in AIDS patients paves the way for attempts towards finding an antisense drug that can be successfully used for treatment of HCV infected patients [6]. Several ASOs that have been designed to bind to the stem-loop structures in the HCV Internal Ribosome Entry Sites (IRES) have been effective in inhibiting HCV replication in cell-culture assays and the expression of HCV luciferase reporter gene in the livers of mice infected with recombinant vaccinia virus expressing the reporter construct [7]. Studies on HCV using ASOs have utilized antisense phosphorothioate oligonucleotides (S-ODN) that were designed as complementary to sequences present in the 5' non coding region (5'-NCR) of IRES of the viral genome. "
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