Supplementary Material for: Prognostic Significance of PD-1, PD-L1 and CD8 Gene Expression Levels in Gastric Cancer

<b><i>Introduction:</i></b> Anti-programmed cell death 1 (PD-1) therapies have shown promising clinical activity against gastric cancer (GC). We evaluated the clinical significance of immune-related gene expression in GC tissues to better understand the tumor immune microenvironment. <b><i>Methods:</i></b> <i>PD-1</i>, <i>PD-1 ligand 1 (PD-L1)</i> and <i>CD8</i> mRNA levels and clinicopathological factors, including survival, were examined by quantitative RT-PCR in 155 GC patients who underwent surgery. PD-1 and PD-L1 expression in tumor tissue from 24 GC patients was investigated by immunohistochemical analysis. <b><i>Results:</i></b> <i>PD-1</i>, <i>PD-L1</i> and <i>CD8</i> mRNA levels were significantly lower in tumor tissue than in normal tissue (<i>p</i> &lt; 0.0001, <i>p</i> &lt; 0.05, and <i>p</i> &lt; 0.0001). GC patients with low <i>PD-1</i>, <i>PD-L1</i> and <i>CD8</i> mRNA levels had significantly poorer overall survival (OS) than those with high <i>PD-1</i>, <i>PD-L1</i> and <i>CD8</i> mRNA levels, respectively (<i>p</i> &lt; 0.001, <i>p</i> &lt; 0.01 and <i>p</i> &lt; 0.05). Low <i>PD-1</i>, <i>PD-L1</i> and <i>CD8</i> mRNA levels were more significantly associated with poor prognosis in undifferentiated-type GC patients than in differentiated-type GC patients (<i>PD-1</i>: differentiated <i>p =</i> 0.0071 vs. undifferentiated <i>p =</i> 0.0024; <i>PD-L1</i>: <i>p =</i> 0.6527 vs. <i>p</i> &lt; 0.0001; <i>CD8</i>: <i>p =</i> 0.4465 vs. <i>p</i> &lt; 0.05). Multivariate analysis showed that lymph node metastasis, peritoneal dissemination, distant metastasis, low <i>PD-1</i> mRNA levels and low <i>CD8</i> mRNA levels were independent prognostic factors for worse OS (low <i>PD-1</i> mRNA level: OR 2.16, 95% CI 1.10–4.58, <i>p</i> &lt; 0.05; low <i>CD8</i> mRNA level: OR 2.55, 95% CI 1.12–6.90, <i>p</i> &lt; 0.05). <i>PD-1</i> and <i>PD-L1</i> mRNA levels in immune cells were significantly associated with PD-1 and PD-L1 protein levels (both <i>p</i> &lt; 0.05), respectively. <b><i>Conclusions:</i></b> <i>PD-1</i>, <i>PD-L1</i> and <i>CD8</i> mRNA levels may reflect antitumor immunity in GC, and low <i>PD-1</i> and <i>CD8</i> mRNA levels are potential predictive biomarkers for poor prognosis in GC patients who underwent surgery.