An HLA-agnostic method for isolation of antigen-specific CD8+ T cells 3733

Abstract Description T cell receptor-engineered T cell (TCR-T) therapies are being tested in clinical trials targeting tumor-specific antigens in both solid and liquid cancers. The discovery of novel TCRs has relied on both in vivo and in vitro stimulation methods, using a range of high- and low-throughput techniques. A common approach for isolation of antigen-specific T cells is flow cytometry-based sorting of a peptide-HLA multimer positive population. However, this demands costly and often unavailable reagents, particularly when attempting to discover TCRs specific for tumor antigens presented by uncommon HLA proteins. In this study, we report a method for isolation of ex vivo expanded antigen-specific CD8+ T cells based on rate of proliferation following antigen specific stimulation. We demonstrate that treating CD8+ T cells with CellTrace dye prior to co-culture with antigen-presenting cells (APCs) allows for separation of a CellTrace negative population that largely encompasses the antigen-specific T cell compartment. We show that this approach effectively identifies antigen-specific CD8+ T cells after priming from both naïve and memory cell fractions. By using autologous T cells and APCs, our method allows for identification of antigen-specific T cells corresponding to any peptide-HLA complex without the need for HLA-specific reagents. As such, our approach could be highly useful for the development of personalized TCR-T trials, particularly for populations with a diverse HLA repertoire. Funding Sources Supported by NIH R37CA247676; T32GM122741 Topic Categories Tumor Immunology: Cellular Responses and Tumor Microevironment (TIME)