A Luminescence-Based Method for In Vitro Screening of Immunomodulatory Checkpoints and Therapeutics.

Early insight into the strength of antigen-specific T cell immune responses is an important aspect for the screening and development of novel immunotherapies, including immune checkpoint inhibitors. Here, we describe a simple, rapid, and cost-effective luminescence-based protocol for the assessment of antigen-specific T cell responses in vitro. This method makes use of genetically engineered Jurkat reporter cells expressing a high-affinity T cell receptor (TCR) toward a commercially available human papillomavirus 16 E7 peptide (E7-TCR), in which luciferase activity is coupled to activation of the promoter of the Nuclear Factor of Activated T Cells (NFAT) transcription factor. With this method, luminescence is generated and can be detected within 6 h only upon antigen-specific activation of the E7-TCR transgenic Jurkat cells with cognate E7 antigenic peptide. This method can be easily modified to study the impact of potential co-stimulatory and immunosuppressive molecules that may be present within the tumor microenvironment (TME) and to monitor the impact of antagonistic antibody treatment on abrogating immunosuppression. We believe this method can be further exploited as valuable tool to study factors that may affect T cell immunity in the immunosuppressive TME and the impact of molecules, cytokines, or therapeutics in triggering effective antigen-specific T cell responses.