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Epithelial-to-mesenchymal transition (EMT) confers cancer cells the ability of invasion and metastasis. However, how does EMT contribute to evasion of immune surveillance is unclear, especially in salivary adenoid cystic carcinoma (SACC). In this study, we investigated the molecular link between EGF-induced EMT and the immune checkpoint ligand programmed death-ligand 1 (PD-L1) by immunoprecipitation (IP) and Westernblot analysis. Cell migration and invasion activity was assayed by transwell assay. Immunohistochemical (IHC) staining analysis was performed for measurement of EMT markers and PD-L1 expression levels in tumor tissues. We found that EGF-induced EGFR activation stabilized Snail expression and induced EMT in SACC. Interestingly, EGFR activation induced simultaneously both EMT and PD-L1 in SACC. Importantly, knockdown of Snail greatly suppressed EGF-induced EMT, but not EGF-induced PD-L1 expression; whereas knockdown of c-Myc strongly repressed PD-L1 expression, but not snail expression and EMT. The molecular link is strongly supported by robust correlations between the EMT markers and PD-L1 expression in human cancer samples.These results suggest that EGFR activated EMT and PD-L1 via two distinct mechanisms. EGFR activation induced EMT and PD-L1 expression in SACC. Snail is required for EGF-induced EMT, but not PD-L1 expression; whereas c-Myc is required for EGFR-mediated PD-L1 upregulation but not EMT. Thus, targeting activated EGFR may inhibit both EMT and PD-L1, which may potentiate the therapeutic effect of PD-L1-based immunotherapy, especially in the malignant subgroups of SACC patients with activated EGFR.

Defects in apoptotic pathway contribute to development and progression of oral cancer. Survivin, a member of the inhibitors of apoptosis protein (IAP) family, is increased in many types of cancers. However, it is unclear whether increased survivin is associated with oral squamous cell carcinomas (OSCC), and what mechanisms may involve in. In this study, we examined survivin expression in OSCC compared with normal oral tissues via immunohistochemical staining. The results showed that, not only total survivin is increased in OSCCs, but also the subcellular location of survivin is changed in OSCCs compared with normal oral tissues. In most of normal oral tissues, survivin staining was either negative, or cytoplasmic positive/nuclear negative; whereas in most of OSCC tissues, survivin staining was nuclear positive. Statistic analysis indicates that nuclear survivin, rather than total or cytoplasmic one, correlates with tumor TNM stage and differentiation grade. Consistently, in vitro analysis showed that survivin is in cytoplasm in normal human oral kinotinocyte (HOK) cells; whereas it is in nucleus in OSCC HN6 cells. Importantly, treatment of HOK cells with HDAC inhibitor Trichostatin A (TSA) induces survivin acetylation and promotes its nuclear localization. Moreover, nuclear survivin in OSCC cells was acetylated at K129 in its C-terminal, suggesting that the acetylation is important for nuclear location of survivin. Our study demonstrates that it is nuclear survivin, rather than total or cytoplasmic one, associates with TNM stage and tumor grade of OSCC. Thus, we propose nuclear survivin as a prognostic marker for the progression of OSCC.