High-dimensional deconstruction of pancreatic ductal adenocarcinoma identifies tumor microenvironmental communities associated with survival

Abstract

Bulk and single-cell analyses of the pancreatic ductal adenocarcinoma (PDAC) tumor microenvironment (TME) have revealed a largely immunosuppressive milieu. Thus far, efforts to utilize insights from TME features to facilitate more effective therapeutics have largely failed. Here, we performed single-cell RNA sequencing (scRNA-seq) on a cohort of treatment-naive PDAC time-of-diagnosis endoscopic ultrasound-guided fine needle biopsy samples (n=22) and surgical samples (n=6), integrated with 3 public datasets (n=49), resulting in ~140,000 individual cells from 77 patients. Based on expression markers assessed by Seurat v3 and differentiation status assessed by CytoTRACE, we divided the resulting tumor cellular clusters into 5 molecular subtypes based on expression of previously reported marker genes: Basal, Mixed Basal/Classical, Classical Low, Classical High, and ADEX. We then queried these 5 tumor cell profiles, along with 15 scRNA-seq-derived tumor microenvironmental cellular profiles, in 391 bulk expression samples from 4 published datasets of localized PDAC with associated clinical metadata using CIBERSORTx. Through unsupervised clustering analysis of these 20 cell state fractions representing tumor, leukocyte and stromal cells, we identified 7 unique clustering patterns representing combinations of tumor cellular and microenvironmental cell states present in PDAC tumors. We termed these cell state patterns communities, and found them to correlate with overall survival, tumor ecotypes, and tumor cellular differentiation status. The community associated with worst overall survival contained basal tumor cells, exhausted CD4 and CD8 T cells, and was enriched for fibroblasts. In contrast, the highest overall survival was associated with a community high in immune cell enrichment. The differentiation state of tumor cells (assessed by CytoTRACE) was also correlated with survival in a dose-dependent fashion. Further, we identified a subset of PDAC samples that were significantly enriched for activated CD8 T and plasma cells that achieved a 2-year overall survival rate of 71%, suggesting we can identify PDAC patients with significantly improved prognoses and, potentially, higher sensitivity to immunotherapy. In summary, we identified novel tumor microenvironmental communities from high-dimensional analysis of PDAC RNA sequencing data that reveal new connections between tumor microenvironmental composition and patient survival that could lead to better upfront risk stratification and more personalized clinical decision-making.

Plasma cells in human pancreatic ductal adenocarcinoma secrete antibodies against self-antigens

“…The presence of effector T cells in our cohort is consistent with previous studies of human and mouse autochthonous PDAC immune microenvironments in which activated, effector T cells were found ( 16 ). A previous preprint study with single-cell sequencing and meta-analysis in PDAC also identified cytolytic CD8 + T cells and their association with improved patient outcomes ( 17 ). Current T cell checkpoint therapy (such as anti–PD-1 antibody) has not been effective in PDAC ( 1 ); thus, additional immunotherapy will be needed to enhance the preexisting T cell activity ( 5 ).…”

Hourglass, a rapid analysis framework for heterogeneous bioimaging data, identifies sex disparity in IL‐6/STAT3‐associated immune phenotypes in pancreatic cancer

“…Iterative refinement of analyses to identify confounder subsets in the patient cohort can thus be instrumental to understand the underlying biology. For instance, the two major transcriptional subtypes of PDAC, basal‐like and classical, differ in survival outcomes [27] and baseline immune microenvironments [33,34], while male PDAC patients overall exhibit higher incidence [35], poorer outcomes [36], and increased TIMP‐1‐mediated hepatic metastasis [37]. Of note, IL‐6 itself has been directly implicated in sex differences in hepatocellular carcinoma [38] and oral squamous cell carcinoma [39], and immune differences in rheumatoid arthritis [40,41].…”

Hourglass, a tool to mine bioimaging data, uncovers sex-disparities in the IL-6-associated T cell response in pancreatic tumors

“…Iterative refinement of analyses to identify confounder subsets can thus be instrumental to understand underlying biology. For instance, molecular subtypes of PDAC (29) differ in their baseline immune microenvironments (30,31) while IL-6 itself has been implicated in sex-differences in the context of hepatocellular carcinoma (32). To test whether the involvement of IL-6 in key biological patterns (immune phenotypes, TME composition) was confounded within specific PDAC patient subsets, we applied parallel subgroup comparison under otherwise identical analytical conditions across global and regional analysis levels, leveraging all steps in the Hourglass workflow (Fig.…”