Locked nucleic acid containing antisense oligonucleotides enhance inhibition of HIV‐1 genome dimerization and inhibit virus replication

Abstract

We have evaluated antisense design and efficacy of locked nucleic acid (LNA) and DNA oligonucleotide (ON) mix-mers targeting the conserved HIV-1 dimerization initiation site (DIS). LNA is a high affinity nucleotide analog, nuclease resistant and elicits minimal toxicity. We show that inclusion of LNA bases in antisense ONs augments the interference of HIV-1 genome dimerization. We also demonstrate the concomitant RNase H activation by six consecutive DNA bases in an LNA/DNA mix-mer. We show ON uptake via receptor-mediated transfection of a human T-cell line in which the mix-mers subsequently inhibit replication of a clinical HIV-1 isolate. Thus, the technique of LNA/DNA mix-mer antisense ONs targeting the conserved HIV-1 DIS region may provide a strategy to prevent HIV-1 assembly in the clinic.

Target Gene Knockdown by 2′,4′-BNA/LNA Antisense Oligonucleotides in Zebrafish

“…In contrast, 2¢,4¢-BNA/DNA-mixed AONs either exploit RNase H to cleave their target RNAs and/or interfere sterically with RNA to block its function. Previous studies have shown that a DNA stretch of six nucleotides in the 2¢,4¢-BNA/DNA chimeric oligomer is required to activate RNase H (Kurreck et al, 2002;Elmen et al, 2004). We therefore designed various 2¢,4¢-BNA AONs that would either activate RNase H or not (Table 1).…”

Locked Nucleic Acid: High-Affinity Targeting of Complementary RNA for RNomics

“…Various LNA oligonucleotides were transfected into HeLa cells and derivatives with a minimum length of 12 residues showed 50% inhibition using nanomolar concentration of LNAs (Arzumanov et al 2003), revealing the potential of LNA antisense oligonucleotides for in vivo targeting of RNA using non-RNase H dependent approaches. In another study, Elmén et al (2004) demonstrated that LNA/DNA mixmers enhance the inhibition of HIV-1 genome dimerization and activate RNase H, show good uptake of the LNA/DNA mixmers in a T cell line, and inhibit replication of a clinical HIV-1 isolate.…”

“…Kurreck et al (2002) investigated various LNA/DNA mixmers and gapmers and found that a gap of six DNA nucleotides is necessary for noteworthy RNase H activity, and that a gap of seven DNA nucleotides allows complete RNase H activity. Also Elmén et al (2004) demonstrated that LNA/DNA mixmers activate RNase H when the mixmer has a DNA stretch of 6 nt. In accordance, Frieden et al (2003a) concluded that a DNA gap size between 7 and 10 nt is optimal for LNA/DNA/LNA antisense gapmers.…”

Diagnostics and Therapeutics

“…However, no RNase H-mediated cleavage was observed with a fully modified 11mer LNA or with an 11mer LNA/DNA mixmer [14]. Kurreck et al [12] and Elmén et al [15] have investigated various LNA/DNA mixmers and gapmers and found that a gap of six DNA nucleotides is necessary for noteworthy RNase H activity, and that a gap of seven DNA nucleotides allows complete RNase H activity. In accordance herewith, Frieden et al [16] concluded that a DNA gap size between 7 and 10 nucleotides is optimal for LNA/DNA/LNA antisense gapmers.…”