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TIDES USA 2022: Oligonucleotide & Peptide Therapeutics

Hybrid Event
May 9–12, 2022

Meet Genedata Experts at TIDES USA 2022.

Don't miss the opportunity to see how Genedata solutions efficiently support the whole biopharmaceutical R&D workflow, including screening and engineering, expression and purification, characterization and analysis of biotherapeutics, cell line design and development, bioprocess optimization, and quality control of biologicals.

Recommended Oral Presentation

Semi-automated Analysis of Oligonucleotide HPLC-UV/MS Data for Chemical Process Development Using Genedata Expressionist
Tim Nagel, Ph.D., Scientist, PTDC – Analytical Research and Development / Hyphenated and Special Technologies, F. Hoffmann-La Roche, Switzerland
Wednesday, May 11, 2022 | 2:00–2:30 pm

The analysis of oligonucleotide crude samples is a highly complex and time-consuming task. We present an automated approach using Genedata Expressionist heavily simplifying the data evaluation. Most oligonucleotide therapeutics are synthesized by standard solid phase phosphoramidite chemistry. The impurity profile of the crude material coming from the synthesis is extremely complex, but in-depth understanding of the impurity profile is required for optimization of the synthesis process to obtain high-quality API. Therefore, hundreds of samples need to be analyzed during process development. These samples easily contain 50-100 impurities, which all need to be identified and quantified. Analysis is performed by ion-pairing reversed phase chromatography coupled to UV detection and low resolution (single quad) mass spectrometry. The MS data that is obtained is highly complex, due to presence of multiple charge states, adduct formation, peak overlap etc. Evaluation and reporting of one sample previously took the analyst 5-6 hours of manually data evaluation work. Ultimately, bottlenecking the process development. We present a semi-automated workflow using Genedata Expressionist facilitating evaluation and reporting of data from oligonucleotide crude samples in less than 30 min. Our workflow combines the UV and MS data to get precise and reproducible quantification of impurities. Therefore, UV and MS data need to be aligned before blank subtraction. In the following, the UV chromatogram is integrated and peaks are split into three groups: early eluting impurities, N-1/main peak/N+1 and late eluting impurities. In the next step, MS spectra are averaged based on the UV peak retention time windows. MS signals are automatically detected and annotated against a library of typical known impurities. The analyst then selects the MS signals that are used for quantification. An MS ratio giving the contribution of each impurity observed by MS to the corresponding UV peak is calculated and multiplied with the UV peak area to obtain a final result. The results are calculated and reported in an excel table that can directly be forwarded to the synthetic chemists to draw conclusions about the synthetic process. This workflow allows us to analyze more samples in shorter time with less effort for the analyst, effectively debottlenecking our process research.