Nucleic Acids Research Advance Access published online on October 5, 2007
Nucleic Acids Research, doi:10.1093/nar/gkm770
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Targeted inhibition of the hepatitis C internal ribosomal entry site genomic RNA with oligonucleotide conjugates
1Biochimie Pharmaceutique, Inserm U835, Upres JE 2311 and 2Ingéniérie Chimique et Molécules pour le Vivant, UMR CNRS 6226, Université de Rennes 1, France
*To whom correspondence should be addressed. Tel: +33 2 23 23 48 51; Fax: +33 2 23 23 44 56; Email: bfelden{at}univ-rennes1.fr
Correspondence may also be addressed to Reynald Gillet. Tel: +33 2 23 23 45 07; Fax: +33 2 23 23 44 56; Email: rgillet{at}univ-rennes1.fr
Received July 5, 2007. Revised September 11, 2007. Accepted September 12, 2007.
Hepatitis C is a major public health concern, with an estimated 170 million people infected worldwide and an urgent need for new drug development. An attractive therapeutic approach is to prevent the ‘cap-independent’ translation initiation of the viral proteins by interfering with both the structure and function of the hepatitis C viral internal ribosomal entry site (HCV IRES). Towards this goal, we report the design, synthesis and purification of novel bi-functional molecules containing DNA or RNA antisenses attached to functional groups performing RNA hydrolysis. These 5' or 3'-coupled conjugates bind the HCV IRES with affinity and specificity and elicit targeted hydrolysis of the viral genomic RNA after short (1 h) incubation at low (500 nM) concentration at 37°C in vitro. Additional secondary cleavage sites are induced and their mapping within the RNA structure indicates that functional domains IIIb-e are excised from the IRES that, based on cryo-EM studies, becomes incapable of binding the small ribosomal subunit and initiation factor 3 (eIF3). All these molecules inhibit, in a dose-dependent manner, the ‘IRES-dependent’ translation in vitro. The 5'-coupled imidazole conjugate reduces viral protein synthesis by half at a 300 nM concentration (IC50), corresponding to a 4-fold increase of activity when compared to the naked oligonucleotide. These new conjugates are now being tested for activity on infected hepatic cell lines.