Effects of short RNA structural analogues against hepatitis C virus genotypes 2, 3 and 4 in replicon cells

Objective To determine whether computer‐predicted short RNA structural analogues could inhibit hepatitis C virus (HCV) genotype 2a, 3a and 4a replication in cultured cells. Methods Short RNA sequences, X12, X12a and X12b, designed to be identical in secondary structure to the X region in the 3′‐untranslated region (3′‐UTR) of the HCV 1b genome, as well as shorter stem‐loop components of X region, were inserted into a plasmid and transfected into separate Huh7.5 human hepatoma cells stably transfected with subgenomic replicons for genotypes 2a, 3a and 4a. All replicons included a firefly luciferase reporter gene. After 48 h of plasmid transfection, the inhibition of HCV replication was determined by HCV RNA isolation and quantification by real‐time polymerase chain reaction and luciferase assays. Results All the secondary structural analogues to genotype 1b X region cross‐inhibited genotype 2a, 3a and 4a replicons. The maximum inhibition by genotype 1b X region structural analogues was obtained against genotype 2a cells in which X12, X12a and X12b inhibited replication by 30%, 63% and 72%, respectively (P