Screening of TCR-specific epitopes using peptide libraries in artificial antigen-presenting cells 3867

Description Despite the key role of T cell receptors (TCR) in the adaptive immune response, experimental validation of the interaction between activated TCRs and computationally predicted cognate antigens remains the holy grail in the development of novel molecular and cell-based therapeutics. To facilitate high-throughput screening of specific TCR-antigen interactions, we developed a universal artificial cellular system based on Jurkat-NFAT-GFP reporter cells expressing candidate-activated TCRs, and K562-HLA antigen-presenting cells (APC) expressing a library of computationally predicted peptide epitopes. For the detection of antigen-activated Jurkat cells interacting with a cognate epitope, K562 cells were engineered to express a single-chain variable fragment (scFv) that could bind IL2 released by activated Jurkat-TCR cells. When K562 APC expressing HLA-cognate epitopes are co-cultured with Jurkat-TCR reporter cells, K562 cells expressing TCR-specific epitopes are labeled with PE anti-IL2 antibody, purified by FACS, and the cognate epitope is detected by NGS. To validate the TCR-epitope library screening system, we constructed an expression library of 129 cancer-associated epitopes in K562-HLA_A02:01 cells. The K562-HLA-epitope library was co-cultured with Jurkat reporter cells expressing MART1 TCR, NY-ESO-1 TCR, or as a control, CMV TCR. By staining & FACS selection of IL2 antibody-stained APCs, we detected specific epitopes with high specificity for cancer-associated & control TCRs. Topic Categories Technological Innovations in Immunology (TECH)