Inhibition of HIV-1 Replication In Vitro and in Human Infected Cells by Modified Antisense Oligonucleotides Targeting the tRNA Lys3 /RNA Initiation Complex

Abstract

The untranslated 5 ′ leader region of the human immunodeficiency virus type 1 (HIV-1) RNA plays an essential role in retroviral replication. It is the first retrotranscribed RNA region, primed from a cellular tRNA^Lys3 partially annealed to the HIV-1 primer binding site (PBS). The structural and functional features of the HIV-1 reverse transcription initiation complex have been thoroughly studied. In this work, we used chemically modified antisense oligonucleotides (AS-ODN) as competitors of the natural tRNA^Lys3 primer for the PBS region. Modified 2′-O-methyl AS-ODN were able to inhibit in vitro HIV-1 reverse transcription and displace the tRNA^Lys3 previously annealed to the PBS. The destabilization of the initiation complex by 2′-O-methyl ODN was a sequence-specific process. We further demonstrated the importance of an anchor region contiguous to the PBS in the annealing of the antisense molecule, allowing the displacement of tRNA^Lys3. The 20-mer 2′-O-methyl molecules were also able to inhibit viral replication in HIV-1-human infected cells, either by blocking cDNA synthesis during the early phase or by interfering with the annealing of the tRNA^Lys3 primer to the PBS during the late phase of the viral cycle. Thus, the highly conserved retroviral initiation complex was shown to be a promising target when using the antisense strategy.