Rapid identification of antigen-specific TCRs for cancer immunotherapy

T cell receptor (TCR) can recognize peptides presented by major histocompatibility complex (MHC) molecules, referred to as HLA in humans, which enabling the targeted eradication of tumor cells expressing specific antigens. In recent years, TCR-engineered T cell (TCR-T) cell therapy has demonstrated substantial advancements in clinical trials targeting solid tumors. Notably, in August 2024, the US FDA approved the first TCR-T drug for the treatment of advanced synovial sarcoma, representing a pivotal milestone in the field. For the development of TCR-T therapy, identifying tumor-associated antigen epitopes and high functional TCRs are critical. Here, we present a comprehensive protocol outlining the process of identification of immunogenic epitopes and the efficient screening of antigenspecific TCRs from HLA transgenic mice. Additionally, the protocol also encompasses methodologies for TCR-T cell preparation and their functional evaluation in vitro. These approaches provide a robust framework for advancing the development of tumor-specific TCRs and fostering the clinical translation of TCR-T therapies.