GAS-LUC2 Reporter Cell Lines for Immune Checkpoint Drug Screening in Solid Tumors

Authors: Hyeyoun Chang, PhD; Kevin Tyo, PhD; John Foulke, MS; Luping Chen, BS; Fang Tian, PhD;* Zhizhan Gu, PhD*
Immuno-Oncology Group, Cell Biology R&D, ATCC, Gaithersburg MD 20877

This study demonstrates the use of three robust, responsive, and reproducible GAS-Luc2 reporter solid tumor cell lines for the assessment of T cell and other immune/tumor microenvironmental cell-mediated immune responses triggered by PD-1/PD-L1, CD-155/TIGIT, or B7-H3 (CD276) immune checkpoints.

Cancer immunotherapy targeting immune checkpoints, such as immune checkpoint inhibitors (ICIs), antibody-dependent cellular cytotoxicity (ADCC), and antibody-drug conjugates (ADCs), have shown tremendous success in the treatment of solid tumors including skin, lung, breast, renal, and liver cancers. However, the built-in complexity of immunological models and the variable drug responses among different cancer types have been challenging the development and application of these novel immunotherapies. To facilitate large-scale drug discovery for this growing class of immunomodulators, we conducted a comprehensive cell surface protein profiling of ATCC’s vast portfolio of human tumor and immune cell lines for established and novel immune checkpoint molecules, as well as their binding ligands. Based on this protein profiling data, we generated three immune checkpoint reporter cell lines, HCC827-GAS-Luc2, MG-63-GAS-Luc2, and NCI-H1650-GAS-Luc2, that endogenously express high levels of programmed death-ligand 1 (PD-L1), cluster of differentiation 155 (CD155), and B7 homolog 3 protein (B7-H3/CD276), respectively. These reporter cell lines were engineered to contain a gamma interferon activation site (GAS)–response element upstream of a luciferase gene. The luciferase expression is suppressed when the relevant immune checkpoint marker on the cancer cells binds to the corresponding checkpoint protein on T cells. In the presence of the relevant immune checkpoint inhibitor, the GAS-Luc2 reporter cell senses the IFN-γ from the activated T cells to produce a luciferase expression–based bioluminescent signal, which can be readily detected and quantified to evaluate the efficacy, potency, and dynamics of the checkpoint inhibitor. In addition to drug screening for immune checkpoint inhibitors, these GAS-Luc2 reporter tumor cell lines are also proven to be effective in detecting paracrine IFN-γ signaling for immune checkpoint targeted ADCC drug screening.