Symposium Sessions
Day 1, June 10（Wed.） 15:00-15:20 Room A (5F 503)
- 1A-S1-1500
Mass Measurements for the Assessment of Critical Quality Attributes of Recombinant Adeno-Associated Virus
(UOsaka)
oYuki Yamaguchi, Susumu Uchiyama
Recombinant adeno-associated viruses (rAAVs) have emerged as an important modality for gene therapy, and the evaluation of their critical quality attributes (CQAs) is essential for ensuring therapeutic efficacy. rAAV particles consist of a capsid composed of three viral proteins, which encapsidates a therapeutic DNA genome. In addition to the percentage of full particles, VP stoichiometry and post-translational modifications may also affect rAAV-therapeutic efficacies. Here, we introduce our studies on glycosylation and deamidation of rAAV capsids analyzed by liquid chromatography-tandem mass spectrometry (LC-MS/MS), as well as a mass photometry-based method for evaluating full particles in crude cell lysate. LC-MS/MS analysis revealed distinct O-glycosylation profiles in rAAV6 capsids, and Mucin-type glycosylation were associated with altered VP stoichiometries and transduction efficiency. We also evaluated multiple quality attributes in rAAV8 and found that VP stoichiometry and deamidation were correlated with transduction efficiency, indicating that they should be considered CQAs. In addition, we developed DirectMP, a direct analytical method for quantifying the percentage of full particles in crude cell lysate. These studies highlight mass measurement techniques, including LC-MS/MS and mass photometry, as powerful tools for assessing the CQAs of rAAV-based therapeutics and supporting the development of higher-quality gene therapy products.