Background: Programmed Death-1 Receptor (PD-1) and its ligand (PD-L1) downregulate T cell activation and suppress tumor killing. This study investigated the role of PD-L1 and tumor infiltrating lymphocytes (TILs) in response to neo-adjuvant therapy and prognosis in esophageal adenocarcinoma (EAC). Methods: Transcriptional profiling of 273 formalin fixed paraffin embedded pre-treatment endoscopic EAC biopsies was carried out using the Almac Diagnostics Xcel array and the expression levels of PD-L1 probesets corresponding to protein encoding extracted. Response was assessed by tumor regression grade (TRG; score ≤ 2 = response). Immunohistochemistry (IHC) for PD-L1 and CD8 was performed in matched resection specimens from 135 patients. All EAC patients were treated with cisplatin-based neo-adjuvant chemotherapy followed by surgical resection between 2003 and 2014 at four UK centers as part of the OCCAMS consortium. Associations between expression, protein levels and TRG were assessed by Kruskal-Wallis, Mann-Whitney Unpaired, Spearman rank correlation or chi-squared tests. Survival analysis was performed using Cox Proportional Hazards regression. Results: High PD-L1 gene expression in the pre-chemotherapy biopsies was associated with pathological response (TRG ≤ 2; p = 0.02) following neo-adjuvant chemotherapy. PD-L1 ( > 5%) was expressed in the tumor or stromal cells in 4% and 15% of resection specimens respectively. PD-L1 gene and IHC expression ( > 5%) were closely associated between the biopsies and both the tumor (p = 0.032) and stroma (p = 0.019) of the matched resection specimens. Patients with PD-L1 IHC positivity in tumor cells demonstrated improved relapse-free survival (HR 0.314; 95% CI 0.099-0.997; p = 0.049) and positive stromal PD-L1 IHC staining correlated with pathological response (p = 0.05). Biopsy gene expression of PD-L1 and CD8 was closely associated (p = 0.024) and the presence of CD8+ TILs in the microenvironment strongly correlated with tumor (p < 0.001) and stromal (p < 0.001) PD-L1 positivity. Conclusions: High PD-L1 expression in the pre-treatment biopsies in EAC is predictive of response to neo-adjuvant chemotherapy and may aid selection of conventional and immune-targeted agents.