Novel approaches in MS characterization of highly modified synthetic RNA

In recent years, the demand for mass spectrometry (MS)-based approaches for structural characterization in quality control of oligonucleotide-based pharmaceuticals has increased increased significantly. Sense strand RNAs with >30 nucleotide building blocks and labile modifications such as N-acetylgalactosamine residues represent thereby a special challenge. We will present direct, robust and easy-to-use techniques to fully characterize the nucleotide sequence of highly modified single-stranded RNAs by top down tandem MS. This will include a case study for an N-acetylgalactosamine modified RNA sense strand. The approach can be applied on single strand level, as well as on siRNA duplex level in a single electrospray ionization MS (ESI-MS) experiment. Based on this technique, we also developed an efficient and highly sophisticated LC-MS/MS workflow for the characterization of product-related impurities in oligonucleotides such as methylation, and PS-PO and rC/rU substitution. This procedure allows us to reliably predict the modification sites in oligonucleotide impurities to support process development with important structural information.

About the speaker

Giovanni Calderisi
Project Chemist QC

Giovanni joined Bachem in Bubendorf (Switzerland) in September 2020 as an expert for mass spectrometry (MS) of oligonucleotides. In his position as a project chemist he is responsible for establishing routine workflows the for characterization of oligonucleotides and peptides by ultra-high resolution MS. Giovanni received his Ph.D. in chemistry with a focus on biomolecular MS at the University of Innsbruck, Austria.