Authors
Johansen, K. H., Ramanathan, K., Rodriguez-Pardo, C., Gjelstrup, A. K., Halldorsdottir, H. R., Voss, L. F., Tvingsholm, S., Grand, L. S., Dionisio, E., Skriver, A. H., Frederiksen, L. P., Bjornson, M., Svensson-Frej, M., Ormhoj, M., Hadrup, S. R.
Abstract
The engineering of autologous T cells for the expression of chimeric antigen receptors (CARs) can induce profound clinical responses in haematological malignancies, while T cell receptor-engineered T (TCR T) cells have led to durable responses to solid tumours in clinical trials. However, the clinical production of engineered T cells is exhaustive and often leads to highly differentiated and exhausted effector T cells. To circumvent this, we have developed an antigen-scaffold (Ag-scaffold) technology to preferentially expand genetically engineered T cells. Such Ag-scaffolds present cognate antigen together with stimulatory factors such as cytokines. By providing a specific and receptor-engaging stimulation to CAR/TCR-engineered T cells, the expanded product is highly enriched for engineered T cells with a favourable proliferative and efficacious phenotype. Here, we expand CRISPR/Cas9- and lentiviral-engineered TCR T and CAR T cells. We expanded TCR T-cells with Ag-scaffolds presenting peptide MHC (pMHC), and anti-CD19 CAR T cells with Ag-scaffolds presenting CD19 antigen. By applying cognate pMHC Ag-scaffolds, we achieved more than 90% antigen-specific T cells after 14 days of culture with a distinct cytotoxic, proliferative phenotypical profile. Ag-scaffold expansion enhanced initial TCR and CAR cytotoxicity; sustained control was observed after repeated rechallenges of CAR T cells. In vivo, Ag-scaffold-expanded CRISPR/Cas9-engineered anti-CD19 CAR T also showed complete tumour eradication in a B-cell lymphoma xenograft model with a low dose of CAR T cells, which was not achieved using IL2/7/15 expansion.
Preprint server:
bioRxiv
The authors list and abstract were imported from bioRxiv on 04 Nov 2025.
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