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Bicyclo[6.1.0]nonyne Phoshoramidites for Metal-Free Conjugation of Oligonucleotides

BRIAN GIBSON1, JORGE M.M. VERKADE2, NANCY S. BARTA1, JOHN C. HODGES1* , FLORIS L. VAN DELFT2,3*
*Corresponding authors
1. Berry and Associates, Inc., 2434 Bishop Circle East, Dexter, MI 48130, USA
2. SynAffix BV, Heyendaalseweg 135, 6525 AJ, Nijmegen, the Netherlands
3. Radboud University Nijmegen, Institute for Molecules and Materials, Heyendaalseweg 135, 6525 AJ, Nijmegen, the Netherlands

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 immobiliz