Effect of silencing tumoral carbohydrate sulfotransferase 15 on T cells in a murine model of pancreatic cancer.

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e16241 Background: Limited T cell infiltration within immunosuppressive microenvironment is still a hurdle for immune checkpoint inhibition therapies in pancreatic cancer. Carbohydrate sulfotransferase 15 (CHST15) is a proteoglycan biosynthetic enzyme and responsible for tumor matrix remodeling. To investigate the role of CHST15 on tumor immune microenvironment, we evaluated the changes in tumor immune cell profiling in a syngeneic mouse model by implanting pancreatic cancer cells with silencing of CHST15 gene. Methods: We constructed CHST15 shRNA-transfected mouse pancreatic cancer cell line KPC, confirmed repression of CHST15 protein then used as tumor-specific CHST15 knocked-down KPC (CHST15 KD). Wild-type KPC (WT) or CHST15 KD cells were implanted subcutaneously to left hind footpad of syngeneic C57BL/6J mice and the tumor growth was monitored. Kinetics of immune cell components were analyzed by immunohistochemical staining for both tumor microenvironment and tumor draining lymph node (TDLN) at weeks 1, 2, 3 and 4 after inoculation (n = 5-6 at each time point). Results: In the WT group, tumor growth was accelerated from week 2 to week 4, while the growth was almost completely inhibited in the CHST15 KD group. Quantitative analyses for immune-stained tissues at week 4 showed the significant repression of CHST15 in the CHST15 KD group. This was associated with the significant increase of CD3+, CD4+ and CD8+ T cells in tumor microenvironment, while T cells were barely detected in the WT group. A significant number of EdU+ proliferating T cells and Granzyme+ CD8+ T cells were found in the CHST15 KD group compared to the WT group. In the TDLN, EdU+ CD4+ and CD8+ T cells were diminished at week 2 from week 1 in the WT group, suggesting active T cell suppression occurred at week 2, which was prior to rapid tumor expansion. In strikingly contrast, massive proliferation of CD4+ and CD8+ T cells was observed in the CHST15 KD group from week 2. RNA expression analysis revealed that anti-tumor T cell activity-related genes including recognition of cancer cells by T cells, cancer antigen presentation, killing of cancer cells, were significantly up-regulated in both tumor and TDLN in the CHST15 KD group. These results suggest that tumoral CHST15 remotely provides T cell suppressive signal to TDLN and blockade of CHST15 reactivates T cell immunity, leading to accelerated tumor-infiltrating T cells. Conclusions: The present study demonstrated that blockade of tumoral CHST15 augments the number of activated tumor-infiltrating T cells and inhibits tumor growth in mouse pancreatic cancer. The effect of tumoral T cell accumulation is followed by boosted T cell priming in immune suppressive TDLN. These findings suggest the possibility of a new immunotherapy by enhancing T cell infiltration in refractory pancreatic cancer.