Abstract A040: Multiplex fluorescent immunohistochemistry reveals immunosuppressive tumor microenvironment in COMPASS-like complex gene mutated pancreatic cancer
Authors: Zhang Shungang , Daniels Elaina , McGue Jake , Kim Hong Sun , Thomas Dafydd , Frankel Timothy , Shi Jiaqi 
Published: September 2024

Abstract
Background: Pancreatic ductal adenocarcinoma (PDAC) is highly resistant to chemoradiation and immune therapies. There is limited biomarker to identify subsets of PDAC that responds to immunotherapy or targeted therapy. We have identified a subset of PDACs (approximately 25%) carrying COMPASS-like complex (CLC) gene mutations. Analysis of the clinicopathologic and molecular features of this PDAC subset revealed that these tumors are an aggressive group of PDACs with poor or squamous histologic differentiation, increased rates of TP53 mutations, and poor patient survival compared to control PDACs. Our preliminary data showed that PDACs carrying mutations in two members of the CLC genes (KDM6A and KMT2D) have immunosuppressive tumor microenvironment (TME), suggesting that CLC genes are candidate biomarkers for immunotherapy. However, a comprehensive analysis of the immune composition, immune checkpoint expression, and the relationship between tumor cells and immune cells in CLC gene mutated PDACs compared to controls has not been performed. In this study, we characterized the immune composition in CLC gene mutated PDACs compared to paired control PDACs using tyramide signal amplification multiplex fluorescent immunohistochemistry (mfIHC). Methods: Human PDAC tissue slides were stained with 2 mfIHC panels and cell types were defined by different markers: panel 1 (PanCK, CD163, PD-L1, CD3, CD8, FoxP3) and panel 2 (PanCK, CD163, CD3, CD8, TIGIT, TIM3). Random image sections were selected in each sample (5∼8 sections per sample) and divided into wild-type (WT) group (n=57) and mutant group (n=41) and analyzed using InForm Cell Analysis software. Tissue segmentation training was performed to separate stromal and epithelial elements. Cell percentage, cell density, and nearest neighbor distance of each cell type were calculated using R program. Results: The CLC gene mutation is associated with elevated epithelial cells (ECs) and PDL1+ ECs and increased infiltration of exhausted cytotoxic T cells (TIGIT+/TIM3+ CD8 T cells). CLC gene mutated PDACs are characterized by spatial proximity between exhausted T cells (TIGIT+) and epithelial cells/antigen presenting cells (APCs). Higher number of epithelial cells is associated with reduced infiltration of helper T cells in the mutant PDAC tissue. We also observed that high numbers of APC in proximity to CD4 T cells were positively correlated with the proximity of EC and CD8 cytotoxic T cells as well as the proximity of CD4 T cells and cytotoxic T cells in the WT group but not in the CLC gene mutant group. Conclusion: Our results provide insight into the effects of CLC gene mutations on shaping the immune microenvironment of PDAC and suggest that CLC gene mutated PDACs have an immunosuppressive immune microenvironment than other PDACs and may respond differently to immunotherapy.
Citation Format: Shungang Zhang, Elaina Daniels, Jake McGue, Hong Sun Kim, Dafydd Thomas, Timothy Frankel, Jiaqi Shi. Multiplex fluorescent immunohistochemistry reveals immunosuppressive tumor microenvironment in COMPASS-like complex gene mutated pancreatic cancer [abstract]. In: Proceedings of the AACR Special Conference in Cancer Research: Advances in Pancreatic Cancer Research; 2024 Sep 15-18; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2024;84(17 Suppl_2):Abstract nr A040.