Analytical perspectives on novel oligonucleotide modalities
Nucleic Acid Insights 2026; 3(5), 339–349
DOI: 10.18609/nai.2026.041
The production of oligonucleotides has long been established for diagnostic or target validation purposes using solid‑phase oligonucleotide synthesis (SPOS) as a platform for high throughput manufacturing technology. At the same time, the SPOS process has evolved over recent decades, meaning that high‑quality molecules can now be produced on large- and commercial scale as active ingredients for RNA therapeutics following strictly controlled and well understood manufacturing processes. Initially, first‑generation antisense oligodeoxyribonucleotides (ASOs) were developed with a phosphorothioate backbone as the sole modification to prolong their systemic half‑life. As the general understanding of RNA therapeutic approaches has grown, the overall structural design of the active molecules has evolved to improve efficacy, safety, stability, or delivery. This has been achieved through chemical and structural modifications to nucleobases, sugar residues, and the phosphate backbone, or through conjugation with chemical or biological moieties. In parallel, these novel chemical and structural developments necessitated an adaptation of the analytical toolbox to analyze the resulting process-, and product‑related impurities and to enable their control by a thorough chemical manufacturing and control (CMC) strategy. A comprehensive understanding of the manufacturing process and established analytical control points throughout the process make it possible to trace process‑related impurities back to the specific process step in which they originated and to prevent their formation or limit them to an acceptable level. This knowledge can then be used to specifically optimize the process steps that are the source of impurities, in order to develop rigorous control strategies that improve product quality and safety. Given the platform‑like nature of the SPOS production process, it is generally not expected that novel modifications, chemical compounds, and conjugates will require the development of entirely new analytical tools that are not already part of the existing toolkit; rather, they will likely require molecule‑specific adaptations of existing methods.