Evaluating teratoma formation risk of pluripotent stem cell-derived cell therapy products: a consensus recommendation from the Health and Environmental Sciences Institute’s International Cell Therapy Committee

Background & Aim Antigen-specific immune responses are central in the natural response towards cancer, infectious diseases and autoimmunity. Engineered transgenic T cells (TCR Ts) pose high therapeutical potential. The precise discovery and analysis of T cell epitopes and the cognate Antigen (Ag)-specific T cells is crucial for cell therapy development, verifying cellular product quality or monitor T cell immune responses in patients. Based on the individual character of Ag-specific T cell responses (MHC restriction), adaptable tools and workflows are required. These tools must be flexible and be tailored to specific cell materials as well as the patients' genetic background, plus be easily integrated into existing analytical workflows. We show that our novel MHC multimer reagents (MHC MACSimers) can advance cell therapy development from the early discovery stages until quality control (QC) of cellular products and immunomonitoring, by covering the demand for flexibility, robustness and seamless integration into common state-of-the-art TCR T development workflows. Methodology To enable the optimal integration of MHC MACSimers into drug development workflows, they feature: -high flexibility of peptide-specificity, enabling MHC peptide-loading for epitope screening and compatibility with TCR discovery workflows -compatibility with various applications, including magnetic enrichment flow cytometry sorting and multimer-duplex staining -growing ready-to-use collection of peptide-loaded MHC MACSimers for robust and reproducible QC of T cell products Results We demonstrate that MHC MACSimers can reliably detect rare virus- or cancer-specific T cells for both CD4+ and CD8+ subpopulations with a higher signal strength as compared to other MHC multimer reagents. Combining MHC MACSimers with fluorochrome-specific magnetic MicroBeads improves the discovery of peptide/MHC-specific TCRs from low-frequent T cell populations by providing highly enriched pMHC-specific T cell fractions. Additionally, Abeling with MHC MACSimers is fully reversible by REAlease® technology, improving functional downstream assays and sequential sorting. Finally, peptide-loaded MHC MACSimers reliably detect TCR Ts in QC of cell therapy products, e.g., against a common Neo-antigen. Conclusion MHC MACSimers enable Ag-specific T cell analysis along the different stages of TCR-based therapy development, covering the need for flexibility, stability, and standardization, making them ideal for integration into T cell drug development workflows.