Immune evasion in lung metastasis of leiomyosarcoma: upregulation of EPCAM inhibits CD8? T cell infiltration

Background: Leiomyosarcomas are among the most common histological types of soft tissue sarcoma (STS), with no effective treatment available for advanced patients. Lung metastasis, the most common site of distant metastasis, is the primary prognostic factor. We analysed the immune environment targeting lung metastasis of STS to explore new targets for immunotherapy. Methods: We analysed the immune environment of primary and lung metastases in 38 patients with STS using immunohistochemistry. Next, we performed gene expression analyses on primary and lung metastatic tissues from six patients with leiomyosarcoma. Using human leiomyosarcoma cell lines, the effects of the identified genes on immune cells were assessed in vitro. Results: Immunohistochemistry showed a significant decrease in CD8? cells in the lung metastases of leiomyosarcoma. Among the genes upregulated in lung metastases, epithelial cellular adhesion molecule (EPCAM) showed the strongest negative correlation with the number of CD8? cells. Transwell assay results showed that the migration of CD8? T cells was significantly increased in the conditioned media obtained after inhibition or knock down of EPCAM. Conclusions: EPCAM was upregulated in lung metastases of leiomyosarcoma, suggesting inhibition of CD8? T cell migration. Our findings suggest that EPCAM could serve as a potential novel therapeutic target for leiomyosarcoma. Overall design: To investigate changes in gene expression in human leiomyosarcoma cell lines in which EPCAM was inhibited or knocked down, we created three clones of TYLMS-1 cells treated with tumour necrosis factor-a converting enzyme inhibitor (TAPI) + ?-secretase inhibitor (DAPT), three clones of untreated TYLMS-1 cells (dimethyl sulfoxide added), three clones of TYLMS-1 cells transfected with siEPCAM, and three clones of TYLMS-1 cells transfected with scrambled siRNA, for a total of 12 TYLMS-1 cell clones. We then performed differentially expressed genes analysis using data obtained from RNA-seq of 12 TYLMS-1 cell clones: comparing gene expression in TAPI + DAPT treated cells and untreated cells, and in siEPCAM-transfected cells and scrambled siRNA-transfected cells.