GAS-Luc2 Reporter Cell Lines for Immune Checkpoint Drug Screening in Solid Tumors.

Simple Summary: Despite the recent studies emphasizing the importance of the interferon gamma receptor (IFNγR) pathway in T cell-mediated cytotoxicity against solid tumors, full human ex vivo immune checkpoint drug screening remains a challenge. We employed an engineered gamma interferon activation site response element luciferase reporter (GAS-Luc2) for immune checkpoint drug screening in diverse ex vivo T cell–solid tumor cell co-culture systems. Three GAS-Luc2 reporter tumor cell lines endogenously expressing various immune checkpoints were engineered to produce quantifiable bioluminescence signal in the presence of an immune checkpoint inhibitor where activated T cells release IFNγ. These reporter cell lines also detected paracrine IFNγ signaling for immune checkpoint-targeted ADCC drug screening. Advancement into an artificial antigen-presenting cell line (aAPC) significantly enhanced T cell signaling for improved performance in these ex vivo immune checkpoint drug screening platforms. Recent studies highlight the integral role of the interferon gamma receptor (IFNγR) pathway in T cell–mediated cytotoxicity against solid but not liquid tumors. IFNγ not only directly facilitates tumor cell death by T cells but also indirectly promotes cytotoxicity via myeloid phagocytosis in the tumor microenvironment. Meanwhile, full human ex vivo immune checkpoint drug screening remains challenging. We hypothesized that an engineered gamma interferon activation site response element luciferase reporter (GAS-Luc2) can be utilized for immune checkpoint drug screening in diverse ex vivo T cell–solid tumor cell co-culture systems. We comprehensively profiled cell surface proteins in ATCC's extensive collection of human tumor and immune cell lines, identifying those with endogenously high expression of established and novel immune checkpoint molecules and binding ligands. We then engineered three GAS-Luc2 reporter tumor cell lines expressing immune checkpoints PD-L1, CD155, or B7-H3/CD276. Luciferase expression was suppressed upon relevant immune checkpoint–ligand engagement. In the presence of an immune checkpoint inhibitor, T cells released IFNγ, activating the JAK-STAT pathway in GAS-Luc2 cells, and generating a quantifiable bioluminescent signal for inhibitor evaluation. These reporter lines also detected paracrine IFNγ signaling for immune checkpoint-targeted ADCC drug screening. Further development into an artificial antigen-presenting cell line (aAPC) significantly enhanced T cell signaling for superior performance in these ex vivo immune checkpoint drug screening platforms. [ABSTRACT FROM AUTHOR]