The programmed cell death (PD)-1 pathway has become an attractive therapeutic target in multiple cancers. Blocking the interaction of PD-1 with its ligands, PD-L1 and PD-L2, leads to impressive anti-tumor responses and clinical benefit in a subset of patients. However, the precise cellular and molecular mechanisms underlying this efficacy are not well understood. To better understand why PD-1/PD-L1 blockade works in some patients but not in others, it is critical to first understand how the PD-1 pathway inhibits anti-tumor immunity. Previous studies demonstrate that clinical responses to PD-1 immunotherapy positively correlate with tumor PD-L1 expression along with other predictive biomarkers such as pre-existing CD8+ T cell infiltration and mutational/neoantigen burden. This has led to the speculation that PD-L1 on tumor cells may act as a “molecular shield” to protect PD-L1+ tumor cells from T cell lysis. We sought to determine whether PD-L1 on tumor cells contributed to the suppression of T cells or simply correlated with an inflamed tumor microenvironment. Using two mouse tumor models that are sensitive to PD-1 pathway blockade, MC38 colorectal adenocarcinoma (MC38) and BRAF.PTEN melanoma (BP), we demonstrate that PD-1 pathway blockade is more effective in MC38 despite similar PD-L1 expression by both tumors. Furthermore, by comparing growth of each tumor cell line in PD-L1 or PD-1 deficient mice to wild-type (WT) mice, we found that host (non-tumor) PD-L1 plays a role in immunosuppression in BP tumors, while PD-L1 on MC38 tumor cells largely mediates suppression of anti-tumor immunity. Thus, the role of PD-L1 on the tumor is tumor-dependent. To further examine the role of MC38 PD-L1 in suppressing the anti-tumor immune response we generated PD-L1-deficient MC38 tumor cells using the CRISPR/Cas9 system. Deletion of PD-L1 on MC38 tumors cells alone leads to effective anti-tumor immunity in mice, analogous to the efficacious anti-tumor immunity seen with PD-1 pathway blockade or in PD-1 deficient mice. Further, wild-type MC38 cells outcompete PD-L1-deleted MC38 cells in vivo, demonstrating MC38-derived PD-L1 is sufficient to mediate immunosuppression. We also found that PD-L1 on MC38 cells potently inhibits CD8+ T cell cytotoxic molecule production. Taken together, our data indicate that PD-L1 expression on MC38 cells alone is sufficient to directly suppress CD8+ T cell cytotoxicity in the tumor microenvironment. These findings demonstrate a direct causal relationship between tumor-derived PD-L1 and inhibition of T cell function and establish a key mechanism by which PD-L1 on tumor cells can suppress T cells. Thus, PD-L1 on tumor cells functions to suppress anti-tumor immunity and is not just a marker of suppressed anti-tumor immunity.

Citation Format: Kathleen A. McGuire, Vikram R. Juneja, Robert T. Manguso, Martin W. LaFleur, Natalie Collins, W. Nicholas Haining, Gordon J. Freeman, Arlene H. Sharpe. Tumor PDL1 blocks CTL cytotoxicity [abstract]. In: Proceedings of the Second CRI-CIMT-EATI-AACR International Cancer Immunotherapy Conference: Translating Science into Survival; 2016 Sept 25-28; New York, NY. Philadelphia (PA): AACR; Cancer Immunol Res 2016;4(11 Suppl):Abstract nr A097.