IFN𝛾-induced treatment of a Human Lung Adenocarcinoma Cell Line is Mediated by an Environmental Chemical Receptor (AhR) through PD-L1 and IDO Control (a549 bulk RNA)

While immunotherapy has shown efficacy in non-small cell lung cancer (NSCLC) patients, many respond only partially or not at all. One limitation in improving outcomes is the lack of a complete understanding of factors regulating immune checkpoint targets. Here, we sought to address a possible link between an environmental chemical receptor implicated in NSCLC and immune regulation, the aryl hydrocarbon receptor (AhR), and a known but counterintuitive mediator of immunosuppression, IFN𝛾, in regulation of two immune checkpoints, PD-L1 and the IDO1, in lung adenocarcinoma (LUAD). To this end we used AhR, PD-L1, and IFN𝛾R gene-edited LUAD cell lines, a syngeneic LUAD mouse model, bulk- and single cell-RNA sequencing of LUADs and tumor-infiltrating leukocytes, and existing human transcriptomic databases. The data demonstrate that: 1) the AhR regulates both PD-L1 and IDO1 in murine and human LUAD cells, 2) AhR-driven IDO1 results in production of Kyn which likely mediates an AhR→IDO1→Kyn→AhR amplification loop, 3) the previously characterized induction by IFN𝛾of PD-L1 and IDO is mediated by the AhR, 4) transplantation of LUAD cells in which the AhR is deleted results in long-term tumor immunity in approximately half of the mice; slow growing tumors in the other half exhibit significantly higher densities of CD4+ and CD8+ T cells expressing immunocompetence markers, and 5) deletion of either IFN𝛾R1 or PD-L1 does not provide the same level of immune protection as AhR deletion. The data definitively link IFN𝛾- mediated immunosuppression to the AhR and support the targeting of the AhR in the context of LUAD. The experiment is comprised of 6 samples, profiling RNA extracted from human A549 lung adenocarcinoma cells. The cells were either unmodified (naive, or "wild-type"), subjected to CRISPR with a non-targeting control sgRNA, or subjected to CRISPR with an sgRNA designed to knock out the aryl hydrocarbon receptor (AHR) gene. Each experimental group was represented in triplicate.