AUTHOR=Moshref Maryam , Lo Jerry Hung-Hao , McKay Andrew , Camperi Julien , Schroer Joseph , Ueno Norikiyo , Wang Shu , Gulati Saurabh , Tarighat Somayeh , Durinck Steffen , Lee Ho Young , Chen Dayue TITLE=Assessing a single-cell multi-omic analytic platform to characterize ex vivo-engineered T-cell therapy products JOURNAL=Frontiers in Bioengineering and Biotechnology VOLUME=12 YEAR=2024 URL=https://www.frontiersin.org/journals/bioengineering-and-biotechnology/articles/10.3389/fbioe.2024.1417070 DOI=10.3389/fbioe.2024.1417070 ISSN=2296-4185 ABSTRACT=

Genetically engineered CD8+ T cells are being explored for the treatment of various cancers. Analytical characterization represents a major challenge in the development of genetically engineered cell therapies, especially assessing the potential off-target editing and product heterogeneity. As conventional sequencing techniques only provide information at the bulk level, they are unable to detect off-target CRISPR translocation or editing events occurring in minor cell subpopulations. In this study, we report the analytical development of a single-cell multi-omics DNA and protein assay to characterize genetically engineered cell products for safety and genotoxicity assessment. We were able to quantify on-target edits, off-target events, and potential translocations at the targeting loci with per-cell granularity, providing important characterization data of the final cell product. Conclusion: A single-cell multi-omics approach provides the resolution required to understand the composition of cellular products and identify critical quality attributes (CQAs).