SUMMARY Pancreatic cancer is one of the most lethal malignancies due to its late diagnosis and limited response to treatment. Tractable methods to identify and interrogate pathways involved in pancreatic tumorigenesis are urgently needed. We established organoid models from normal and neoplastic murine and human pancreas tissues. Pancreatic organoids can be rapidly generated from resected tumors and biopsies, survive cryopreservation and exhibit ductal- and disease stage-specific characteristics. Orthotopically transplanted neoplastic organoids recapitulate the full spectrum of tumor development by forming early-grade neoplasms that progress to locally invasive and metastatic carcinomas. Due to their ability to be genetically manipulated, organoids are a platform to probe genetic cooperation. Comprehensive transcriptional and proteomic analyses of murine pancreatic organoids revealed genes and pathways altered during disease progression. The confirmation of many of these protein changes in human tissues demonstrates that organoids are a facile model system to discover characteristics of this deadly malignancy.

Cancer-associated fi broblasts (CAF) are major players in the progression and drug resistance of pancreatic ductal adenocarcinoma (PDAC). CAFs constitute a diverse cell population consisting of several recently described subtypes, although the extent of CAF heterogeneity has remained undefi ned. Here we use single-cell RNA sequencing to thoroughly characterize the neoplastic and tumor microenvironment content of human and mouse PDAC tumors. We corroborate the presence of myofi broblastic CAFs and infl ammatory CAFs and defi ne their unique gene signatures in vivo. Moreover, we describe a new population of CAFs that express MHC class II and CD74, but do not express classic costimulatory molecules. We term this cell population "antigenpresenting CAFs" and fi nd that they activate CD4 + T cells in an antigen-specifi c fashion in a model system, confi rming their putative immune-modulatory capacity. Our cross-species analysis paves the way for investigating distinct functions of CAF subtypes in PDAC immunity and progression. SIGNIFICANCE : Appreciating the full spectrum of fi broblast heterogeneity in pancreatic ductal adenocarcinoma is crucial to developing therapies that specifi cally target tumor-promoting CAFs. This work identifi es MHC class II-expressing CAFs with a capacity to present antigens to CD4 + T cells, and potentially to modulate the immune response in pancreatic tumors.

Pancreatic ductal adenocarcinoma (PDAC) is poorly responsive to therapies and histologically contains a paucity of neoplastic cells embedded within a dense desmoplastic stroma. Within the stroma, cancer-associated fi broblasts (CAF) secrete tropic factors and extracellular matrix components, and have been implicated in PDAC progression and chemotherapy resistance. We recently identifi ed two distinct CAF subtypes characterized by either myofi broblastic or infl ammatory phenotypes; however, the mechanisms underlying their diversity and their roles in PDAC remain unknown. Here, we use organoid and mouse models to identify TGFβ and IL1 as tumorsecreted ligands that promote CAF heterogeneity. We show that IL1 induces LIF expression and downstream JAK/STAT activation to generate infl ammatory CAFs and demonstrate that TGFβ antagonizes this process by downregulating IL1R1 expression and promoting differentiation into myofi broblasts. Our results provide a mechanism through which distinct fi broblast niches are established in the PDAC microenvironment and illuminate strategies to selectively target CAFs that support tumor growth. SIGNIFICANCE : Understanding the mechanisms that determine CAF heterogeneity in PDAC is a prerequisite for the rational development of approaches that selectively target tumor-promoting CAFs. Here, we identify an IL1-induced signaling cascade that leads to JAK/STAT activation and promotes an infl ammatory CAF state, suggesting multiple strategies to target these cells in vivo. See related commentary by Ling and Chiao, p. 173.

Summary Pancreatic cancer is a deadly malignancy that lacks effective therapeutics. We previously reported that oncogenic Kras induced the redox master regulator Nrf2/Nfe2l2 to stimulate pancreatic and lung cancer initiation. Here, we show that NRF2 is necessary to maintain pancreatic cancer proliferation by regulating mRNA translation. Specifically, loss of NRF2 led to defects in autocrine EGFR signaling and oxidation of specific translational regulatory proteins, resulting in impaired cap-dependent and cap-independent mRNA translation in pancreatic cancer cells. Combined targeting of the EGFR effector AKT and the glutathione antioxidant pathway mimicked Nrf2 ablation to potently inhibit pancreatic cancer ex vivo and in vivo, representing a promising synthetic lethal strategy for treating the disease.

Background/Aims: Mounting evidence suggests that epitranscriptional modifications regulate multiple cellular processes. N⁶-Methyladenosine (m⁶A), the most abundant reversible methylation of mRNA, has critical roles in cancer pathogenesis. However, the mechanisms and functions of long non-coding RNA (lncRNA) methylation remain unclear. Pancreatic cancer resulted in 411,600 deaths globally in 2015. By the time of pancreatic cancer diagnosis, metastasis has often occurred in other parts of the body. The present study sought to investigate lncRNA m⁶A modification and its roles in pancreatic cancer. Methods: Differential expression between cancer cells and matched normal cells was evaluated to identify candidate lncRNAs. The lncRNA KCNK15-AS1 was detected in cancer tissues and various pancreatic cells using RT-qPCR. KCNK15-AS1 was transfected into cells to explore its role in migration and invasion. Then, m⁶A RNA immunoprecipitation was performed to detect methylated KCNK15-AS1 in tissues and cells. Epithelial–mesenchymal transition (EMT) markers were used to evaluate KCNK15-AS1-mediated EMT processes. Results: KCNK15-AS1 was downregulated in pancreatic cancer tissues compared with paired adjacent normal tissues. KCNK15-AS1 inhibited migration and invasion in MIA PaCa-2 and BxPC-3 cells. Furthermore, total RNA methylation in cancer cells was significantly enriched relative to that in immortalized human pancreatic duct epithelial (HPDE6-C7) cells. In addition, the m⁶A eraser ALKBH5 was downregulated in cancer cells, which can demethylate KCNK15-AS1 and regulate KCNK15-AS1-mediated cell motility. Conclusion: Our results have revealed a novel mechanism by which ALKBH5 inhibits pancreatic cancer motility by demethylating lncRNA KCNK15-AS1, identifying a potential therapeutic target for pancreatic cancer.