This study was approved by the institutional review boards, and appropriate written informed consent was obtained from all patients. Immunohistochemistry Immunostaining was performed using anti-PD-L1 antibody (SP142, Roche), anti-PD-L2 antibody (NBP1C88964, Novus Biologicals), anti-PD1 antibody(315M-96, Cell marque), anti-CD3 antibody (NCL?CD3C565, Leica), anti-CD8 antibody (CRM311C, Biocare Medical), anti-MutL homolog 1 (MLH1) antibody (IR079, DAKO), anti-mutS homolog 2 (MSH2) antibody (IR085, DAKO), anti-mutS homolog 6 (MSH6) antibody(IR086, DAKO), anti-PCNA antibody(CBL407, Millipore), anti-C-met antibody (clone SP44, Roche) and anti-HER2 antibody (clone 4B5, Roche) as primary antibodies. and PCNA expression. GC patients with high level PDL1 expression exhibited better survival. GC Patients with higher T cell infiltration also showed elevated PDL1, PDL2 and PD1 Lavendustin A expression and predict favorable outcome, indicating an adaptive immune resistance mechanism may exist. The group of patients infiltrated with lower density CD3+ T cells also without PDL1 expression in tumor cells predict Lavendustin A the worst outcome in the subgroup of different PTNM stage, which may suggest an inactive immune status. These results highlights the need to assess both PDL1 expression in all tumor context and the characterization of the GC immune microenvironment. strong class=”kwd-title” KEYWORDS: gastric cancer, PDL1, PDL2, PD1, prognosis, T cells Introduction Although declining in the last few decades, Gastric cancer (GC) still remains the third most common cancer worldwide.1 It was postulated that GC, which harbored higher mutation frequency, would be more immunogenic and likely to benefit from immunotherapy, given its breakthrough effects on melanoma and lung cancer.2,3,4 Cancer cells may exhibit immune inhibition to promote tumor progression and distant metastasis.5 One key mechanism is the Programmed Cell Death 1 (PD1) / PD1 ligand 1 (PDL1) pathway. PD1 is typically expressed by activated lymphocytes, including CD8+ T cells, CD4+ T cells, natural killer (NK) T cells, B cells, activated monocytes and dendritic cells.6 It is activated by its ligands PDL1 and PDL2 to suppress antigen-stimulated lymphocyte proliferation, migration and cytokine production, ultimately resulting in attenuating effector T cells function and immunological tolerance. 6 PDL1 is constitutively expressed on T and Lavendustin A B cells, macrophages and dendritic cells, while PDL2 expression is much more typically restricted in activated DC and macrophages.7 PDL1 have also been reported to be expressed in variety tumor cells including lung, colorectal, breast and melanoma.8C11 To date, PD1 and its ligands in GC was only evaluated in small cohort of patients with controversial conclusions.12C15 These results were probably confounded by the ethnic differences, the small sample size, and/or heterogeneous patient population. In addition to PD1/PDL1, tumor infiltrating lymphocytes (TILs) reflects primary host immune response against solid tumors.16,17 A strong histological lymphocytic reaction, high density of CD3+ and CD8+ T Lavendustin A cells have been reported with better outcome in colorectal cancer and lung cancer, demonstrating an important role of T cell mediated host immunity Rabbit Polyclonal to DGKD in repressing tumor progression.12,18,19 Considering the close relationship between PD1/PDL1 pathway and T cell activation, we used a tissue microarray (TMA) including 1,014 well annotated GC specimens to investigate the expression of PD1/PDL1, quantified tumor infiltrating CD3+ and CD8+ T cells density to determine their relationships with clinicopathological features and patients’ prognosis. Results Patient characteristics A total of 1014 surgically resected FFPE primary gastric cancer sampled in tissue microarrays could be assessed for the markers, including 739 males and 275 females without any preoperative therapy (Table?1). The median follow up is 39.0 months (range: 0.2C133.9 months) and the median age at diagnosis is 61 y (range: 22C89 y). 239 cases were located at the proximal part, including the gastro-esophageal junction, 259 cases in the corpus, 468 at the distal part including antrum and 5 cases disseminated to the whole gastric structures. All the tissue samples were identified as adenocarcinoma and 78% of samples were poorly or moderate differentiated, including 8% signet ring cell type. As shown in Table?1, 569 cases (56.1%) were intestinal subtype, while 242 (23.9%) were intestinal and 184 (18.1%) were mixed type. The vast majority of tumors were classified as clinical TNM stage 2(34.7%) and stage 3(44.1%). The 1-, 3-, 5-year overall survival is 0.846 0.011, 0.613 0.016, 0.521 0.017, respectively. Table 1. Clinicopathological and molecular features according to PDL1 expression. thead th align=”center” rowspan=”1″ colspan=”1″ Lavendustin A ? /th th align=”center” rowspan=”1″ colspan=”1″ ? /th th colspan=”3″ align=”center” rowspan=”1″ PDL1 in tumor hr / /th th colspan=”3″ align=”center” rowspan=”1″ PDL1 in immune cells hr / /th th align=”center” rowspan=”1″ colspan=”1″ ? /th th align=”center” rowspan=”1″ colspan=”1″ n /th th align=”center” rowspan=”1″ colspan=”1″ Pos(%) /th th align=”center” rowspan=”1″ colspan=”1″ Neg(%) /th th align=”center” rowspan=”1″ colspan=”1″ p-value /th th align=”center” rowspan=”1″ colspan=”1″ Pos(%) /th th align=”center” rowspan=”1″ colspan=”1″ Neg(%) /th th align=”center” rowspan=”1″ colspan=”1″ p-value /th /thead Gender1014??0.140??0.437?female27541.558.5?73.126.9??male73936.463.6?75.524.5?Age1014??0.507??0.405? = 6050636.863.2?73.726.3?? 6050838.861.2?76.024.0?Localization971??0.760??0.342?Upper23938.561.5?17.66.0??Middle25935.964.1?79.920.1??Lower46839.160.9?72.627.4??whole540.060.0?60.040.0?Lauren type995??0.013??0.247?Diffuse24232.267.8?72.327.7??Intestinal56938.062.0?74.925.1??Mixed18446.253.8?79.320.7?T stage1014??0.068??0.027?T16437.562.5?82.817.2??T212449.250.8?83.116.9??T365136.363.7?72.827.2??T417535.464.6?73.726.3?N stage1007?? 0.001??0.016?N025243.756.3?75.824.2??N+75538.461.57%?77.222.8?M stage979??0.028??0.002?M091338.461.6?75.924.1??M16625.874.2?59.140.9?pTNM979?? 0.001??0.003?111844.155.9?80.519.5??228044.655.4?77.922.1??351533.866.2?73.826.2??46625.874.2?59.140.9?Vascular invasion999??0.215??0.114?neg45639.960.1?77.622.4??pos54336.163.9?73.326.7?Diameter978??0.973?? 0.001? 5cm52037.962.1?80.219.8?? = 5cm45838.062.0?70.129.9?Differentiation962??0.115??0.873?well2321.778.3?69.630.4??poorly47040.659.4?76.024.0??moderately46937.162.9?76.123.9?MMR857??0.999??0.057?deficient14240.859.2?69.730.3??proficient71540.859.2?77.222.8?EBV status967?? 0.001??0.001?neg93336.563.5?74.225.8??pos3476.523.5?100.00.0?c-Met status979?? 0.001?? 0.001?neg65533.067.0?71.128.9??pos32448.151.9?83.616.4?PCNA892?? 0.001?? 0.001?12810.789.3?57.142.9??210431.768.3?61.538.5??376040.559.5?78.421.6? Open in a separate window PDL1, PDL2 and PD1 expression PDL1 expression was evaluated in.