Abstract 1019: EphA2 receptor tyrosine kinase regulates programmed death ligand 2 (PD-L2) expression in tumor cells and inhibits immune infiltration

Abstract

Abstract Background: Given the success of both targeted and immunotherapies, there is increasing utility for identifying targeted agents that also promote anti-tumor immunity. EphA2 is a receptor tyrosine kinase that contributes to tumor growth and metastasis in various cancer types and plays a role in inflammatory processes. Previous work in our lab demonstrates EphA2 is a viable target for non-small cell lung cancer (NSCLC) and breast cancer. Here, we examine how EphA2 affects programmed death-ligand (PD-L) expression and immune response in the tumor microenvironment. Methods: Our preliminary studies suggest EphA2 regulates the expression of PD-L2 but not PD-L1 in human lung and breast cell lines. To investigate this, we induced PD-L2 expression in vitro via cytokines, overexpressed or knocked down EphA2, and measured PD ligand expression by flow cytometry. To evaluate the mechanism by which EphA2 affected PD-L2 expression, we curated the ENCODE database to identify transcription factor (TF) binding sites near the PD-L2 promoter and validated individual TF candidates with knockdown experiments. To study in vivo effects, we generated an EphA2-overexpressing murine NSCLC cell line from a primary mouse lung tumor. Both subcutaneous and tail vein injected lung tumor models were used to assess the impact of EphA2 overexpression on tumor burden and survival, as well as immune infiltration by flow cytometry. Results: In human bronchial and mammary epithelial cells, PD-L2 was induced by IFNγ and, to a lesser extent, TNFα and IL-4, and knockdown of EphA2 decreased surface expression of PD-L2. TF binding sites for Myc and p65, among others, were found in the PD-L2 promoter region. Because our lab has previously shown that Myc and YAP are downstream effectors of EphA2, we pursued these TFs for validation studies, which are ongoing. In contrast to human epithelial cells, murine counterparts do not express PD-L2, even after IFNγ stimulation. While the EphA2-overexpressing murine NSCLC cell line did not display proliferative advantage over control cells in vitro, they developed larger tumors and had worse survival in both tumor models. Analysis of lung tumor immune infiltrate revealed decreased NK and T cells in the EphA2-overexpressing tumors. Conclusion: Our studies suggest EphA2 upregulates PD-L2 in cancer cells and inhibits tumor infiltration of key lymphocytic populations. Ongoing investigations are determining the mechanisms behind these findings. Despite PD-L2's known role in immune tolerance, its impact on the tumor microenvironment is understudied compared to PD-L1. Thus, elucidation of EphA2's role in regulating PD-1 and PD-L2 interactions and immune recruitment will further our understanding of PD-1 and PD-L biology and mechanisms of immune evasion, as well as provide additional rationale for targeting EphA2 in cancer. Citation Format: Eileen Shiuan, Wenqiang Song, Deanna Edwards, Shan Wang, Sung Hoon Cho, Mark Boothby, Jin Chen. EphA2 receptor tyrosine kinase regulates programmed death ligand 2 (PD-L2) expression in tumor cells and inhibits immune infiltration [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 1019.