Modified (PNA, 2 '-O-methyl and phosphoramidate) anti-TAR antisense oligonucleotides as strong and specific inhibitors of in vitro HIV-1 reverse transcription

Natural beta-phosphodiester (16)mer and (15)mer antisense oligonucleotides targeted against the HIV-1 and HIV-2 TAR RNAs respectively were previously described as sequence-specific inhibitors of in vitro retroviral reverse tr anscription. In this work, we tested chemically modified oligonucleotide an alogues: alpha-phosphodiester, phosphorothioate, methylphosphonate, peptide nucleic acid or PNA, 2'-O-methyl and (N3'-P5') phosphoramidate versions of the 16mer anti-TAR oligonucleotide. PNA, 2'-O-methyl and (N3'-P5') phospho ramidate oligomers showed a strong inhibitory effect compared with the unmo dified 16mer, with reverse transcription inhibition (IC50) values in the na nomolar range. The inhibition was sequence-specific, as scrambled and misma tched control oligonucleotides were not able to inhibit cDNA synthesis. No direct binding of the 2'-O-methyl, PNA or (N3'-P5') phosphoramidate anti-TA R oligonucleotides to the HIV-1 reverse transcriptase was observed. The hig her T-m obtained with 2'-O-methyl, (N3'-P5') phosphoramidate and PNA molecu les concerning the annealing with the stem-loop structure of the TAR RNA, i n comparison with the P-phosphodiester oligonucleotides, is correlated with their high inhibitory effect on reverse transcription.