Citations

Abstract Despite advances in surgery and chemotherapy, patients with resectable pancreatic ductal adenocarcinoma (PDAC) face high recurrence rates (40-60%). Total neoadjuvant therapy (TNT) seeks to enhance margin-negative resections (R0) and address micrometastatic disease, with potential survival benefits. Yet, TNT still results in high recurrence rates, with 1-, 2-, and 3-year disease-free survival (DFS) at 65%, 48%, and 32%, respectively. Residual cancer extent is a key prognostic factor in PDAC after TNT. Tumor response scores show favorable outcomes only for patients achieving near-complete or complete pathologic response. In a cohort of 511 PDAC patients treated with TNT and surgery (2015-2022), improved DFS was only found in those with a complete pathologic response (pCR, 7.8%). This highlights the limited use of current response scoring in stratifying post-TNT patient risk, underscoring the need to better understand disease persistence and relapse. Here we developed a quantitative method to assess residual cancer. Using a digital pathology H&E library from 2180 PDAC cases (2000-2022), an AI platform segmented tissue into cancer epithelium, stroma, and normal regions. For 202 TNT-treated cases without pCR, we calculated a ratio of cancer area to stromal area from representative slides. Modeled after the histologic tumor regression grade (HTRG) system, cases were classified into <5%, 5-50%, or >50% cancer-to-stroma ratios. However, this measure alone showed no difference in DFS, suggesting tumor burden cannot fully explain differences in clinical outcomes. We hypothesized that the spatial organization of the tumor microenvironment (TME) is a determinant of patient outcomes. Drawing from landscape ecology, which studies spatial properties of ecosystems, we applied similar principles to analyze tissue architecture. Using an AI-based two-class segmentation of whole-slide images, we assessed spatial features like patch area, perimeter, density, and Shannon diversity. Python’s ‘pylandstats’ package calculated 21 spatial metrics, reduced via stepwise Cox regression using the Akaike information criterion (AIC). The final model included mean Shannon diversity, mean and standard deviation of stromal patch areas, and stromal patch density. Kaplan-Meier analysis of 202 samples stratified model predictions into quartiles, correlating with high (mean DFS 10.85 months), medium (19.32 months), and low risk (25.92 months) groups. These findings aligned with model expectations, underscoring the potential of spatial TME analysis to predict outcomes. Citation Format: Andrea V. Maraone, Zafar Siddiqui, Luis H. Cisneros, Merih D. Toruner, Mark Truty, Cornelius Thiels, Robert R. McWilliams, Steven R. Alberts, Rondell P. Graham, Carlo Maley, Qian Shi, Martin E. Fernandez-Zapico, Ryan M. Carr. Tumor spatial configuration predicts pancreatic cancer recurrence risk after total neoadjuvant therapy [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2025; Part 1 (Regular Abstracts); 2025 Apr 25-30; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2025;85(8_Suppl_1):Abstract nr 5070.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most aggressive cancers, primarily due to its complex tumor microenvironment (TME), which drives both disease progression and therapy resistance. Understanding the molecular mechanisms governing TME dynamics is essential for developing new treatment strategies for this devastating disease. In this study, we uncover an oncogenic role for Galectin-1 (Gal1), a glycan-binding protein abundantly expressed by activated pancreatic stellate cells (PSCs), a key component of the PDAC TME that orchestrates tumor progression. Our findings reveal that Gal1 expression is elevated in the nucleus of human PSCs in both tissue samples and cultured cell lines. Using chromatin immunoprecipitation followed by sequencing analysis (ChIP-seq), we identify Gal1 occupancy at the promoters of several cancer-associated genes, including KRAS , a pivotal oncogene involved in PDAC pathogenesis. We demonstrate that Gal1 binds to the KRAS promoter, sustaining KRAS expression in PSCs, which, in turn, maintains PSC activation and promotes the secretion of protumorigenic cytokines. Mechanistically, Gal1 is required to preserve histone H3 lysine 4 monomethylation levels and to recruit the histone methyltransferase MLL1 to target promoters. Collectively, our findings define a nuclear function of Gal1 in modulating the transcriptional landscape of cancer-associated genes in PSCs within the PDAC TME, mediated through an epigenetic mechanism. These insights enhance our understanding of PDAC pathology and open potential avenues for therapeutic interventions targeting intracellular Gal1.

Abstract Purpose: KRAS G12C inhibitors can treat KRASG12C-mutant advanced colorectal cancers and pancreatic ductal adenocarcinomas (PDAC), but alterations in Kirsten rat sarcoma (KRAS), EGFR, BRAF, MAP2K1, and other genes bypass KRAS inhibition and reduce therapy efficacy. Our study evaluates the genetic landscape of candidate primary resistance alterations relevant to KRAS targeting in KRASG12C-mutant colorectal cancer and PDAC. Experimental Design: We analyzed two cohorts (national database and Mayo) of patients with advanced colorectal cancer or PDAC tested with next-generation sequencing of ctDNA via Guardant360. Cohorts were divided into three groups: KRASG12C alone (KRASG12C without a resistance gene), KRASG12C with resistance (KRASG12C and ≥1 candidate resistance gene), and KRAS not detected. Candidate resistance mutations were inferred from the reported literature. Results: Among the national (13,603 colorectal cancer and 5,016 PDAC cases) and Mayo (741 colorectal cancer and 422 PDAC cases) cohorts, resistance alterations were identified in a considerable number of KRASG12C cases (46.5% of national colorectal cancer, 16.4% of national PDAC, 53.8% of Mayo colorectal cancer, and 36.4% of Mayo PDAC). The presence of resistance alterations was associated with a trend toward worse overall survival in KRASG12C colorectal cancer (P = 0.05). Conclusions: Putative resistance alterations are prevalent in PDAC and colorectal cancer and may limit monotherapy efficacy. Identifying these alterations has potential implications in optimal patient selection for targeted therapies and the development of combination therapy strategies to overcome primary resistance.

Abstract Pancreatic cancer is a devastating malignancy in great need of new and more effective treatment approaches. In recent years, studies have indicated that nutritional interventions, particularly nutraceuticals, may provide novel avenues to modulate cancer progression. Here, our study characterizes the impact of ω-3 polyunsaturated fatty acids, eicosapentaenoic acid, and docosahexaenoic acid, as a nutraceutical intervention in pancreatic cancer using a genetically engineered mouse model driven by KrasG12D and Trp53R172H. This model closely resembles human pancreatic carcinogenesis, offering a disease relevant platform for translational research. Our findings showed that ω-3 polyunsaturated fatty acids intervention (using a diet supplemented with 6% cod liver oil) significantly reduced tumor volume as well as lung and liver metastasis and a trend toward improved survival rate compared with control treated mice. This antitumoral effect was accompanied by distinct changes in tumor membrane fatty acid profile and eicosanoids release. Furthermore, the eicosapentaenoic acid and docosahexaenoic acid intervention also reduced malignant histological parameters and induced apoptosis without affecting cell proliferation. Of note is the significant reduction in tumor fibrosis that was associated with decreased levels of Sonic Hedgehog, a major ligand controlling this cellular compartment in pancreatic cancer. All together our results demonstrate the impact of eicosapentaenoic acid and docosahexaenoic acid as antitumor regulators in pancreatic cancer, suggesting potential for ω-3 polyunsaturated fatty acids as a possible antitumoral dietary intervention. This research opens new avenues for integrating nutraceutical strategies in pancreatic cancer management.

Abstract Introduction Pancreatic adenocarcinoma (PDA) will be the second leading cause of cancer related death by 2030 and patient response to chemotherapy is poor with a strong likelihood of cancer recurrence. While past studies have investigated pre- and post-chemotherapy patient samples, there is a paucity of studies looking at fresh, patient matched samples which allow us to observe interpatient heterogeneity in chemotherapy response. Methods Here, we study the effects of chemotherapy on five different patients using sc-RNA sequencing of matched treatment naïve and post treatment tissue biopsies to compare the effects of chemotherapy, not only on the tumor cells, but also on the tumor microenvironment – a key contributor to tumor survival and proliferation. We utilized the Seurat v4 pipeline and CellChat v2 to observe transcriptomic differences in epithelial cells, myeloid cells, t-Cells, and fibroblasts and their corresponding putative interactions in pre- and post- treatment states. Results Unsurprisingly, we found insufficient tumor epithelial cells post-treatment to perform further analysis on due to the intended cytotoxic effects of chemotherapy. Therefore, we turned our focus to the tumor microenvironment, where we had sufficient numbers of stromal and immune cells, to investigate the effects of chemotherapy. We identified that fibroblasts changed the most consistently across all patients in response to chemotherapy. The fibroblasts clustered into three distinct groups: an immunomodulatory group, a myofibroblastic and neurotropic group, as well as a mixed group expressing genes from all three categories. Interestingly, the neurotropic fibroblast signature was universally enriched after chemotherapy in all five patients, despite having a small and clinically heterogeneous cohort. Conclusions Through an analysis of each fibroblast group in the treatment naïve and post treatment categories, we observed that a specific set of fibroblasts become less myofibroblastic and increasingly neurotropic following treatment, leading us to posit that fibroblasts altered by exposure to chemotherapy may potentially contribute to a ‘pro-tumor’ environment that supports recurrence. To test this in vitro, we treated patient derived fibroblasts with Gemcitabine and used RTPCR to analyze their relative changes in gene expression. We specifically probed for genes that were definitive of myofibroblasts and neurotropic fibroblasts and saw that post-chemotherapy exposure, neurotropic genes such as NEGR1 and NFIA were upregulated. This validated our scRNA conclusions that fibroblasts become increasingly neurotropic following treatment and then differentially interact with their surrounding environment. In future studies, we will further analyze the role these neurotropic fibroblasts play in chemoresponse and tumor recurrence. Matched samples are critical to studies of chemotherapy effects on PDA due to the fact that they allow us to deconvolute one aspect of PDA heterogenity - interpatient heterogeneity - which is a large limitation in studies of this disease. Citation Format: Aylin Z Henstridge, Padma Kadiyala, Ahmed M Elhossiny, Jianhua Liu, Vaibhav Sahai, Nicole Peterson, Jorge Machicado, Richard Kwon, Allison Schulman, Erik Wamsteker, George Philips, Martin Fernandez-Zapico, Mark Truty, Costas Lyssiotis, Timothy Frankel, Filip Bednar, Marina Pasca di Magliano, Eileen S Carpenter. Neurotropic fibroblast population increases following exposure to chemotherapy in pancreatic adenocarcinoma [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 A062.

Pancreatic cancer is a malignancy arising from the endocrine or exocrine compartment of this organ. Tumors from exocrine origin comprise over 90% of all pancreatic cancers diagnosed. Of these, pancreatic ductal adenocarcinoma (PDAC) is the most common histological subtype. The five-year survival rate for PDAC ranged between 5 and 9% for over four decades, and only recently saw a modest increase to ∼12–13%, making this a severe and lethal disease. Like other cancers, PDAC initiation stems from genetic changes. However, therapeutic targeting of PDAC genetic drivers has remained relatively unsuccessful, thus the focus in recent years has expanded to the non-genetic factors underlying the disease pathogenesis. Specifically, it has been proposed that dynamic changes in the epigenetic landscape promote tumor growth and metastasis. Emphasis has been given to the re-organization of enhancers, essential regulatory elements controlling oncogenic gene expression, commonly marked my histone 3 lysine 4 monomethylation (H3K4me1). H3K4me1 is typically deposited by histone lysine methyltransferases (KMTs). While well characterized as oncogenes in other cancer types, recent work has expanded the role of KMTs as tumor suppressor in pancreatic cancer. Here, we review the role and translational significance for PDAC development and therapeutics of KMTs.

Abstract Aberrant activation of GLI transcription factors has been implicated in the pathogenesis of different tumor types including pancreatic ductal adenocarcinoma. However, the mechanistic link with established drivers of this disease remains in part elusive. In this study, using a new genetically engineered mouse model overexpressing constitutively active mouse form of GLI2 and a combination of genome-wide assays, we provide evidence of a novel mechanism underlying the interplay between KRAS, a major driver of pancreatic ductal adenocarcinoma development, and GLI2 to control oncogenic gene expression. These mice, also expressing KrasG12D, show significantly reduced median survival rate and accelerated tumorigenesis compared with the KrasG12D only expressing mice. Analysis of the mechanism using RNA sequencing demonstrate higher levels of GLI2 targets, particularly tumor growth–promoting genes, including Ccnd1, N-Myc, and Bcl2, in KrasG12D mutant cells. Furthermore, chromatin immunoprecipitation sequencing studies showed that in these cells KrasG12D increases the levels of trimethylation of lysine 4 of the histone 3 (H3K4me3) at the promoter of GLI2 targets without affecting significantly the levels of other major active chromatin marks. Importantly, Gli2 knockdown reduces H3K4me3 enrichment and gene expression induced by mutant Kras. In summary, we demonstrate that Gli2 plays a significant role in pancreatic carcinogenesis by acting as a downstream effector of KrasG12D to control gene expression.

Acute pancreatitis (AP) is among the most common hospital gastrointestinal diagnoses; understanding the mechanisms underlying the severity of AP is critical for development of new treatment options for this disease. Here, we evaluate the biological function of phosphofructo-2-kinase/fructose-2,6-biphosphatase 3 (PFKFB3) in AP pathogenesis in 2 independent genetically engineered mouse models of AP. PFKFB3 was elevated in AP and severe AP (SAP), and KO of Pfkfb3 abrogated the severity of alcoholic SAP (FAEE-SAP). Using a combination of genetic, pharmacological, and molecular studies, we defined the interaction of PFKFB3 with inositol 1,4,5-trisphosphate receptor (IP3R) as a key event mediating this phenomenon. Further analysis demonstrated that the interaction between PFKFB3 and IP3R promotes FAEE-SAP severity by altering intracellular calcium homeostasis in acinar cells. Together, our results support a PFKFB3-driven mechanism controlling AP pathobiology and define this enzyme as a therapeutic target to ameliorate the severity of this condition.

Abstract Pancreatic ductal adenocarcinoma (PDAC), the most common histological subtype of pancreatic cancer, is a devastating disease predicted to be the second leading cause of cancer deaths in the next 15 years. Thus, there is an urgent need to develop new treatment approaches based on a better understanding of cellular and molecular mechanisms controlling this dismal progression. PDAC development is marked by nuclear morphological changes likely resulting in changes in chromatin organization and gene expression. Oncogenic KRAS, present in more than 90% of tumors, plays a key role in disease initiation and progression and is sufficient to induce these nuclear morphology changes. Work from our lab has identified the nuclear lamina protein Emerin as a mediator of KRAS-induced nuclear morphology changes. Further, a BioID experiment identified a KRAS-induced interaction of Emerin with a group of nuclear kinases belonging to the ribosomal S6 kinase (RSK) and mitogen- and stress- activated protein kinase (MSK) families. In silico analysis identified candidate MSK and RSK phosphorylation sites in Emerin. Thus, we hypothesize that the nuclear kinase families RSK and MSK can act as effectors of the KRAS-Emerin axis to regulate PDAC nuclear morphology. Further studies will be done to confirm if Emerin can be phosphorylated by RSK and MSK, and to validate that this relationship is dependent on oncogenic KRAS signaling and if its nuclear morphological changes are regulated by this GTPase. Overall, this work aims to define a novel signaling axis for RSK and MSK in controlling nuclear dynamics that play a role in disease progression of PDAC, which can lead to the identification of new therapeutic targets. Citation Format: Kayla LaRue, Ashley Sigafoos, Luis Flores, Brooke Tader, Martin Fernandez-Zapico. The role of nuclear kinases in pancreatic carcinogenesis [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3154.