Inhibition of Hepatitis C IRES-Mediated Gene Expression by Small Hairpin RNAs in Human Hepatocytes and Mice HEINI ILVES, a ROGER L. KASPAR, a QIAN WANG, b ATTILA A. SEYHAN, a ALEXANDER V. VLASSOV, a CHRISTOPHER H. CONTAG, b,c DEVIN LEAKE, d AND BRIAN H. JOHNSTON a,c a SomaGenics, Inc., Santa Cruz, California 95060, USA b Molecular Imaging Program at Stanford (MIPS), and Departments of Radiology, Microbiology & Immunology, Stanford University School of Medicine, Stanford, California, USA c Department of Pediatrics, Stanford University School of Medicine, Stanford, California, USA d Dharmacon RNA Technologies, LaFayette, Colorado 80026, USA ABSTRACT: The ability of small hairpin RNAs (shRNAs) to inhibit hepati- tis C virus internal ribosome entry site (HCV IRES)-dependent gene ex- pression was investigated in cultured cells and a mouse model. The results indicate that shRNAs, delivered as naked RNA or expressed from vectors, may be effective agents for the control of HCV and related viruses. KEYWORDS: hepatitis C virus; shRNA; siRNA; RNA interference Viral Hepatitis C is principally a disease of inflammation of the liver, and 70% of patients infected with the hepatitis C virus (HCV) develop chronic liver disease, including cirrhosis and hepatocellular carcinoma. HCV infec- tion afflicts 3.9 million people in the United States (175 million worldwide) and is the primary indication for liver transplants in the United States. RNA interference (RNAi)-mediated gene inhibition has been shown to robustly in- hibit gene expression in a number of mammalian systems. 1 Despite its high degree of sequence conservation, the HCV internal ribosome entry site (IRES) would appear a priori to be a poor target for RNA interference due to the high proportion of its residues involved in secondary and tertiary folding. Several groups have recently reported, however, some success targeting the HCV IRES in 293FT and Huh7 tissue culture cells (reviewed in Ref. 2). We have investigated the ability of small hairpin RNAs (shRNAs), delivered directly or expressed from pol III promotors, as well as synthetic siRNAs to Address for correspondence: Brian H. Johnston, SomaGenics, Inc., 2161 Delaware Ave., Santa Cruz, CA 95060. Voice: 831-426-7700; fax: 831-420-0685. e-mail: bjohnston@somagenics.com Ann. N.Y. Acad. Sci. 1082: 52–55 (2006). C  2006 New York Academy of Sciences. doi: 10.1196/annals.1348.060 52