Abstract

Phosphoramidite building blocks based on a bicyclo[6.1.0]nonyne (BCN) core, are developed and readily integrated into solid phase synthesis of oligonucleotides. Strategies are reported for the preparation of oligonucleotides containing either a 5′-conjugated or an internal BCN, and a lipophilicity comparison is made with a dibenzoannulated cyclooctyne. It is demonstrated that smooth and efficient conjugation to azido-containing functional groups takes place. The reported building blocks and strategy pave the way for the preparation of a wide variety of oligonucleotide conjugates based on readily available building blocks.

INTRODUCTION

Synthetic DNA and RNA oligonucleotides (ONs) are key tools in a broad variety of diagnostic and therapeutic applications, including microarray technology (1), antisense and gene-silencing therapies (2), nanotechnology (3) and materials sciences (4). Generally, such applications require the introduction of a suitable handle in an oligonucleotide to enable selective conjugation to a functionality of interest (5). For example, attachment of a cell-penetrating ligand is the most commonly applied strategy to tackle the low internalization rate of ONs into target cells (2), which is the main bottle-neck in oligonucleotide-based therapeutics (antisense, siRNA). Similarly, the preparation of oligonucleotide-based microarrays requires the selective immobilization of ONs to a suitable solid surface, e.g. glass. Conventional post-synthetic labeling protocols, based on amide bond formation or sulfide-based chemistry (5c,6), typically suffer from low yield, prolonged reaction time and often require a high concentration of the biomolecule in combination with a large excess of coupling partner. One promising alternative to the traditional conjugation techn ...