Antisense oligonucleotides inhibit in vitro cDNA synthesis by HIV-1 reverse transcriptase

The inhibition of reverse transcription by various chemically modified antisense oligonucleotides was studied in a cell-free system, composed of an RNA template, a primer oligodeoxynucleotide, and the HIV-1 reverse transcriptase (RT). Different mechanisms of inhibition were observed depending on the chemical structure of the antisense molecule. (1) The hybridization of 2'-O-allyl oligonucleotide to the RNA template promotes a physical arrest of the polymerase. (2) The antisense effect of phosphodiester or phosphorothioate oligonucleotides is essentially due to the RNase H-mediated cleavage of the RNA. (3) A third mechanism was observed with phosphorothioate oligonucleotides that directly interact with the enzyme. Chimeric oligonucleotides, composed of an unmodified region flanked by 2'-O-methyl groups, led to less efficient inhibition than the parent unmodified oligomer, although the inhibitory mechanism was the same. No inhibitory effect was detected when alpha or methylphosphonate oligomers were used.