Using LC-MS to Identify Clipping in Self-Assembled Nanoparticles During Vaccine Development
A hemagglutinin stabilized stem nanoparticle (HA-SS-np) that is designed to provide broad protection against influenza is being developed as a potential vaccine. During an early formulation screening study, reducing gel (rCGE) analysis indicated product degradation in a few candidate buffers at the first-week accelerated stability point, whereas no change was shown in the size exclusion chromatography (SEC) measurement. A LC-MS workflow was therefore applied to investigate the integrity of this large HA-SS-np vaccine molecule (≈ 1 MDa). Application of LC-MS was critical to rationalize the conflicting results from the rCGE and SEC assays and led to the discovery that (1) an unexpected sequence clipping in the HA-SS-np subunits occurred, explaining the atypical reducing gel profile, and (2) an undisrupted disulfide bond held the two fragments together, explaining the unchanged SEC profile. This analytical case study led to a formulation buffer redesign, which mitigated the issue.
KeywordsLC-MS Characterization Vaccine Nanoparticles Clipping Influenza
The authors would like to acknowledge the Vaccine Research Center teams within the Vaccine Production Program under NIH and thank Cindy Cai, Sashikanth Banappagari, KC Cheng, and Lisa Kueltzo for their helpful discussions, as well as Jessica Bahorich, Heather Lawlor, Kevin Carlton, and Jason Gall for their project and leadership support.
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