Synthesis and Characterization of Aminooxy Click Chemistry-Mediated Bis -Conjugates of siRNAs and Evaluation of Their RNAi Activity

Abstract Previously, we used an aminooxy click chemistry (AOCC) strategy to introduce pairs of ligands into oligonucleotides during solid-phase synthesis by using nucleoside and non-nucleoside building blocks. However, the standard oligonucleotide synthesis conditions were incompatible with the incorporation of certain complex ligands. In this report, first, a post-oligonucleotide synthesis solution-phase AOCC approach was developed to address this issue. Different pairs of ligands were conjugated to oligonucleotides through an O-amino building block incorporated during the solid-phase synthesis. Second, a small interfering RNA with a sense strand conjugated using the AOCC strategy to target the ApoB gene using a receptor-targeting ligand (GalNAc) and a pharmacokinetic enhancer (C16 alkyl) was evaluated in mice. The bis-heteroconjugate had a longer duration of action and higher level of silencing than the siRNA carrying only the receptor-targeting ligand.