Cholesteryl-conjugated oligonucleotides: synthesis, properties, and activity as inhibitors of replication of human immunodeficiency virus in cell culture.

Abstract

A family of oligonucleotides and phosphorothioate oligonucleotide analogues was synthesized with a cholesteryl group tethered at the 3'-terminal internucleoside link. This modification, introduced to enhance interaction of the polyanions with cell membranes, significantly increases the antiviral activity of the oligomers, as judged by inhibition of syncytia formation and expression of viral proteins p17, p24, and reverse transcriptase for human immunodeficiency virus 1 in Molt-3 cells. In the most favorable case, with a 20-mer cholesteryl-phosphorothioate derivative, complete inhibition by all assays was obtained with an oligomer concentration of 0.2 jIM. Even decamers were active, and some antiviral activity was observed for a heptanucleotide cholesteryl-phosphorothioate derivative, which binds very poorly to complementary oligonucleotides. These facts, and the finding that the activity of the phosphorothioate decamers does not correlate with a specific sequence., suggests that a mechanism other than "antisense inhibition" may be operative in these systems.

Bioconjugation of Oligonucleotides for Treating Liver Fibrosis

“…Many lipophilic molecules have been used to modify ODNs (470,471). However, among them, cholesterol is perhaps the best characterized and extensively studied in vivo (472)(473)(474). Unlike other lipophilic molecules, it has been shown that cholesteryl conjugation confers ODNs enhanced binding to lipoproteins and thereby improved cellular association and transport especially into liver tissue.…”

“…Conjugation with cholesterol can be integrated into the synthesis of ODNs using nucleoside derivatives (472,475,476). However, cholesterol can also be conjugated to post-synthetic ODNs with functionalities at ends.…”

Synthesis and anti-HIV activity of thiocholesteryl-coupled phosphodiester antisense oligonucleotides incorporated into immunoliposomes

“…Cholesterol modification has been shown to increase both the cell association and activity of phosphodiester and phosphorothioate oligonucleotides (Boutorine et al, 1989;Letsinger et al, 1989;Boutorine et al, 1993;Krieg et al, 1993). These compounds have been shown to be taken up, at least in part, by the LDL receptor (de Smidt et al, 1991;Krieg et al, 1993).…”

“…An approach to circumvent these problems has been to covalently bind native (phosphodiester) or modified (phosphorothioate) oligonucleotides to phospholipid (Shea et al, 1990) or other hydrophobic residues (Kabanov et al, 1990;Saison-Behmoaras et al, 1991), including cholesterol (Boutorine et al, 1989;Letsinger et al, 1989;Krieg et al, 1993). Cholesterol modification has been shown to increase both the cell association and activity of phosphodiester and phosphorothioate oligonucleotides (Boutorine et al, 1989;Letsinger et al, 1989;Boutorine et al, 1993;Krieg et al, 1993).…”

Specific Hammerhead Ribozyme-mediated Cleavage of Mutant N-ras mRNA in Vitro and ex Vivo

“…In the latter case, CaCl 2 or cationic liposomes have been mostly used as transfection reagents. Additional exogenous delivery methods are based on the conjugation of the oligonucleotides to polylysine compounds (22) or to lipophilic groups, like cholesterol (23,24).…”