Published online 29 November 2004
Nucleic Acids Research, Vol. 32 No. 21 © Oxford University Press 2004; all rights reserved
Diversity-oriented solid-phase synthesis and biological evaluation of oligonucleotide hairpins as HIV-1 RT RNase H inhibitors
Department of Chemistry, Otto Maass Chemistry Building, McGill University, 801 Sherbrooke Street West, Montreal, QC, H3A 2K6, Canada
* To whom correspondence should be addressed. Tel: +1 514 398 7552; Fax: +1 514 398 3797; Email: masad.damha{at}mcgill.ca
Present address: Rami N. Hannoush, Department of Chemistry and Chemical Biology, Harvard University, Cambridge, MA, USA
Received September 27, 2004; Revised and Accepted October 29, 2004
The inhibitory potencies of several hairpins comprising DNA, RNA and 2',5'-linked RNA segments were assessed against the RNase H activity of the human immunodeficiency virus reverse transcriptase (HIV-1 RT), an indispensable enzyme for HIV genomic replication. The hairpin library was constructed via diversity-oriented nucleic-acid synthesis (DONAS), an approach inspired from traditional split-pool synthesis. DONAS provided access to an array of oligonucleotide hairpins possessing distinct conformational, structural and biological properties. The inhibitory potency of these compounds was highly specific towards HIV-1 RT RNase H and strongly depended on the structure of both the stem and tetraloop. Hairpins that have an overall A-type geometry are better inhibitors of RNase H activity than hairpins with ‘intermediate’ or B-type conformations, although interestingly, the inhibitory activity is quite sensitive to the nucleotide sequence in both the stem and loop regions of the hairpin. The most potent hairpins bear a 3',5'-linked rather than 2',5'-linked RNA loop, but the latter is necessary for activity of hairpins consisting of DNA stems. Inhibitory activity was essentially independent of hairpin thermal stability. The potent hairpins also demonstrated high nuclease resistance in biological media, particularly those bearing a 2',5'-linked tetraloop. These studies collectively bring into light a new class of nucleic acid aptamers that act exclusively upon the retroviral RNase H domain in vitro, and thus represent novel lead compounds for the development of specific and potent HIV-1 RT inhibitors.
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