Fig. 1
Schematic representation of the UniCAR system targeting the FAP antigen using different αFAP TM formats. a UniCAR T-cells consist of an extracellular single-chain fragment variable (scFv) directed to the peptide epitope E5B9, the CD28 transmembrane and intracellular signaling domains, and the CD3 zeta signaling portion. For UniCAR T-cell redirection to target FAP, an intermediary target module (TM, in this case αFAP TM) is required that is composed of a scFv that recognizes FAP and the epitope E5B9 that interacts with the UniCAR. Different formats of such TMs, e.g. scFv-based TMs (αFAP-scFv TM) and also TMs integrating the backbone of a human IgG4 (αFAP-IgG4 TM) can be combined with the UniCAR system. LP, leader peptide; VH, variable domain of the antibody heavy chain; VL, variable domain of the antibody light chain; Fc, fragment crystallizing; His, hexa-histidine tag. b and c TMs were purified via their His-tag from cell culture supernatants of producer cell lines genetically modified to permanently express the respective recombinant protein. Purified TMs were analyzed using SDS-PAGE followed by Quick Coomassie Stain (b) or immunoblotting followed by TM detection with the anti-La mAb 5B9 (c)