We describe here further characterization of two DNA aptamers that specifically bind HCV RNA polymerase (NS5B) and inhibit its polymerase activity in vitro. Although they were obtained from the same selection procedure and contain an eleven-nucleotide consensus sequence, our results indicate that aptamers 27v and 127v use different mechanisms to inhibit HCV polymerase. While the 27v is able to compete with the RNA template for binding to the enzyme and blocks both initiation and elongation of RNA synthesis, the 127v competes poorly and inhibits initiation and post-initiation events exclusively. These results illustrate the power of the SELEX approach to select specific short DNA aptamers able to inhibit HCV NS5B by different mechanisms. We also determined that in addition to an in vitro inhibitory effect on RNA synthesis, the aptamer 27v was able to interfere with the multiplication of JFH1 HCV in Huh7 cells. The efficient cellular entry of these short DNAs and the inhibitory effect observed on human cells infected with HCV indicate that aptamers are useful tools to study HCV RNA synthesis and should become a very attractive and alternative approach in the therapy against HCV.

Bellecave P, Cazenave C, Rumi J, Staedel C, Cosnefroy O, Andreola ML, Ventura M, Tarrago-Litvak L, Astier-Gin T.

CNRS UMR 5234; INSERM U869; Université Victor Segalen Bordeaux 2, 146, rue Léo Saignat. 33076 Bordeaux cedex. France.