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Development of a universal preparative AEX method to purify oligonucleotides

Introduction

Oligonucleotide-based therapeutics have been investigated over the last decades and their promise as a new drug modality is now being realized. The growing interest in oligonucleotides is driven by the high potential of oligonucleotides to be used in the treatments of a variety of medical conditions, the growing number of FDA approved oligonucleotide drugs, an increased focus on personalized medicine and on the development of therapies for rare diseases, and the wide adoption of nucleotide-based COVID-19 vaccines.

Oligonucleotides are short, linear sequences of DNA or RNA that are generally manufactured by chemical synthesis. Oligonucleotides are extremely susceptible to oxidation, enzymatic degradation, and clearance in vivo. Because of this, synthetic oligonucleotides are often chemically modified to improve their stability and make them resistant to extracellular and intracellular nuclease degradation. One of the original and still most widely used modifications is the phosphorothioate modification of the oligonucleotide backbone.

Due to errors during the oligonucleotide synthesis process, nucleosides may either be missing (N-X) or are attached in excess (N+X). Moreover, the chirality of the sulfur atoms in the backbone of ONs (due to the phosphorothioate modification) leads to diastereomers. To remove these impurities, the biopharma relies on chromatography during the purification process. The increased demand for oligonucleotides requires a costeffective and easy scale-up from research amounts to commercial needs.

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