Oligonucleotides tethered to a short polyguanylic acid stretch are targeted to macrophages: Enhanced antiviral activity of a vesicular stomatitis virus-specific antisense oligonucleotide

The poor membrane permeability of oligonucleotides is one of the major prob lems of antisense technology. Here we report the construction of designer o ligonucleotides for targeted delivery to macrophages. The oligonucleotides tethered to a 10-mer poly(G) sequence at their 3' ends were recognized by s cavenger receptors on macrophages and were taken up about 8- to 10-fold as efficiently as those oligonucleotides that either lacked a poly(G) tail or that contained a 10-mer poly(C) tail instead of the poly(G) tail. The enhan ced uptake of poly(G) constructs was inhibited in the presence of poly(G) a nd other known ligands of the scavenger receptor. The bioefficacy of poly(G )-mediated targeting of antisense oligonucleotides (ANS) was demonstrated b y using vesicular stomatitis virus (VSV) as a model system. The ability of ANS directed against the translation initiation site of N protein mRNA of V SV to inhibit virus replication was assessed. The ANS with the 10-mer poly( G) sequences (ANS-G) brought about significant inhibition of VSV replicatio n in J774E cells (a murine monocyte/macrophage cell line) and Chinese hamst er ovary (CHO) cell transfectants expressing scavenger receptors. The ANS l acking a 10-mer poly(G) stretch were ineffective. The inhibition of VSV rep lication due to ANS-G was completely abrogated in the presence of 10-mer po ly(G), indicating that the antisense effect of the ANS G molecule was a con sequence of scavenger receptor-mediated enhanced uptake. Importantly, antis ense molecules linked exclusively by natural phosphodiester bonds were as b ioeffective as those synthesized with a mixed backbone of phosphodiester an d phosphorothioate. Taken together, these results suggest that macrophage-d irected designer ANS against infective agents may simply be obtained by add ing a short stretch of guanylic acid sequence to the desired specific ANS d uring solid-phase synthesis. This nucleic acid-based strategy, which utiliz es homogeneous preparation of ANS, may find applications in directed manipu lation of macrophage metabolism for a variety of purposes as well as in the rapy of a broad spectrum of macrophage-related disorders amenable to the an tisense approach.