Background: Tumor specific antigens, such as NY-ESO-1, are emerging as key tumor immune modulating factors with great potential to be used as therapeutic targets to enhance immunotherapy efficacy and expand treatment options for GC. We sought to compare GC tumors expressing high vs low levels of NY-ESO-1 in terms of immune cell abundance in the tumor microenvironment (TME) as well as distinct molecular features and immune biomarkers. Methods: 1967 tumor samples tested with NextGen Sequencing on DNA (592 genes or WES) and RNA (WTS) by Caris Life Sciences (Phoenix, AZ) were retrospectively reviewed. The top quartile of transcripts per million was considered high while the bottom quartile was considered low NY-ISO-1 expression. Tumor mutational burden (TMB) was calculated based on somatic nonsynonymous mutations. Mismatch repair deficiency/microsatellite instability (dMMR/MSI) status was evaluated by a combination of IHC, fragment analysis and NGS of known MSI loci. Gene fusions were detected based on WTS. X², Fisher-exact, and Mann Whitney tests were used for comparison and significance adjusted for multiple testing by Benjamini-Hochberg correction (q< 0.05). Cell infiltration in the TME was estimated by quanTIseq. Gene expression profiles were analyzed for a transcriptional signature predictive of response to immunotherapy (T cell-inflamed signature: TIS). Results: The analysis was focused on primary tumors (N = 1323) in this initial study. Expression of NY-ESO-1 was lower in primary/local than metastases (Fold Change FC met vs primary: 1.60, q < 0.05). NY-ESO-1 expression did not appear to be strongly associated with distinct gene mutation profiles in GC. There were no significant differences between low and high expression of NY-ESO-1 with regards to well established immuno-oncology markers (dMMR/MSI, TMB, PD-L1). However, high NY-ESO-1 expression was positively associated with immune related gene expression including CD274, CD80, CD86, CTLA4, HAVCR2, IDO1, IFNG, LAG3, PDCD1, and PDCD1LG2 (FC low vs high: 0.56 to 0.79, q< 0.0001). High NY-ESO-1 expression was also positively associated with cell abundance in the TME including NK cells (FC = 0.87, q <0.0001), monocytes (FC = 0.29, q< 0.05), myeloid dendritic cells (FC = 0.66, q <0.0001), CD4+ non-reg T cells (FC = 0.54, q <0.0001), and CD8+ T cells (FC = 0.73, q< 0.05). Similarly, tumors with high NY-ESO-1 expression were associated with higher TIS scores (q< 0.0001). Conclusions: In our large cohort of GC, tumors expressing high NY-ESO-1 displayed a distinct landscape of immune cells in the TME and were associated with high expression of immune related genes, as well as high TIS score, which has been reported to predict benefit from anti-PD-1 treatment. The results of our analysis support an association between a more immunologically active tumor microenvironment and NY-ESO-1 gene expression which may have relevant implications on immunotherapy treatment for GC.