Abstract 4392: Tumor suppressor miR-339-5p regulates PD-L1 expression in lung cancer

Abstract

Abstract The PD1/PD-L1 (programmed death 1/programmed death 1 ligand 1) pathway is a central mechanism by which tumors escape host immunity. PD1 is a B7 receptor and is expressed by immune effector cells, including T cells, B cells, and natural killer cells. The predominant ligand for the PD1 receptor is PD-L1. Tumor expression of PD-L1 is thought to promote T-cell tolerance by binding PD1 on activated T cells and suppressing their capacity to secrete stimulatory cytokines. Tumor expression of PD-L1 has also been shown to directly inhibit T-cell-mediated lysis by inactivating tumor-reactive cytotoxic T lymphocytes. Targeting immune checkpoints that promote immune suppression such as PD1/PD-L1 has proven effective for suppressing tumor growth. Thus an important area of focus is on clarifying the mechanisms of cellular sensitivity and resistance to PD1/PD-L1 blockade and identifying biomarkers of response and survival with such therapy. MicroRNAs (miRNAs) are small RNAs that regulate virtually all processes in the cell, including immune response. However, there are very few studies linking miRNAs regulation to immune modulators. In this study, we aimed to identify miRNAs that regulate the immunoregulator PD-L1 in lung cancer. By using three different prediction programs (targetscan; MirWalk; miRNAbodymap), we identified a list of twenty miRNAs that potentially regulate PD-L1. We then selected the miRNAs that inversely correlated with PD-L1 expression in 532 patients with non-small lung cancer (NSCLC) from The Cancer Genome Atlas data (TCGA). Next, we overexpressed these miRNAs in NSCLC cells and analyzed the expression of PD-L1 by quantitative PCR and western blotting. Among the miRNAs selected, enforced overexpression of miR-339-5p decreased the expression of PD-L1 mRNA and protein compared with scrambled control. To investigate whether miR-339-5p interacts directly with the putative target gene PD-L1, we co-transfected 3′ UTR luciferase reporter vectors with miR-339-5p precursors and used luciferase assays to confirm that miR-339-5p regulates PD-L1 expression. Specifically, luciferase activity was reduced in cells transfected with miR-339-5p, PD-L1 3′ UTR constructs compared with scrambled. Mutation of miR-339-5p interaction sites rescued the luciferase activity, thus confirming that miR-339-5p directly interacts with the PD-L1 3´ UTRs. Taken together, these findings suggest that miR-339-5p is a new regulator of PD-L1 expression in NSCLC and may possibly be used as immunotherapy or as biomarker in future clinical approaches. Citation Format: Rachel Heymach, Lixia Diao, Lauren Averett Byers, Monique Nilsson, Jing Wang, Maria A. Cortez. Tumor suppressor miR-339-5p regulates PD-L1 expression in lung cancer. [abstract]. In: Proceedings of the 105th Annual Meeting of the American Association for Cancer Research; 2014 Apr 5-9; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2014;74(19 Suppl):Abstract nr 4392. doi:10.1158/1538-7445.AM2014-4392