SPECIFIC-INHIBITION OF HEPATITIS-B VIRUS GENE-EXPRESSION BY AN ANTISENSE OLIGONUCLEOTIDE IN-VITRO

It was previously shown that a number of antisense oligonucleotides ag ainst hepatitis B virus (HBV) mRNAs were highly effective in inhibitio n of HBV gene expression (Yao et al., 1995). Here, using radioisotope techniques, we report a specific inhibition of HBV surface antigen (HB sAg) production in vitro by 2.2.15 cells (Hep-G2 cells transfected wit h HBV genome) by the antisense oligonucleotide 15-S-asON, a 15-mer pho sphorothioate analogue complementary to the cap site of the SPII promo ter of HBV mRNA, at a concentration of 2-5 mu mol/l. After 24 and 48 h rs of incubation of cells with 15-S-asON, the intracellular concentrat ion of the latter rose to 69.4 and 75.8 nmol/l, respectively, and the HBsAg level assayed by ELISA was reduced by 50.0% and 70.6%, respectiv ely. These results were checked by use of the radioimmunoprecipitation method: 2.2.15 cells exposed to 15-S-asON and labelled with [S-35]-me thionine for 48 hrs showed a decrease of the HBsAg level by 81.26% but almost none of the total proteins. No cytotoxicity of the 15-S-asON w as observed with regard to the cell morphology and growth. These resul ts indicate that the tested antisense oligonucleotide specifically inh ibits the HBV gene expression.