Poor Survival In Pancreatic Ductal Adenocarcinoma Patients Research Articles

Abstract C067: Tuft cells transdifferentiate to neural-like progenitor cells in the progression of pancreatic cancer

Abstract Pancreatic ductal adenocarcinoma (PDA) is partly initiated through the transdifferentiation of acinar cells to metaplastic ducts that act as precursors of neoplasia and cancer. Tuft cells are solitary chemosensory cells not found in the normal pancreas but arise in metaplasia and neoplasia, diminishing as neoplastic lesions progress to carcinoma. Metaplastic tuft cells (mTCs) function to suppress tumor progression through communication with the tumor microenvironment, but their fate during progression is unknown. To determine the fate of mTCs during PDA progression, we have created a lineage tracing model that uses a tamoxifen-inducible tuft-cell specific Pou2f3CreERT/+ driver to induce transgene expression, including the lineage tracer tdTomato or the oncogene Myc. mTC lineage trace models of pancreatic neoplasia and carcinoma were used to follow mTC fate. We found that mTCs, in the carcinoma model, transdifferentiate into neural-like progenitor cells (NRPs), a cell type associated with poor survival in PDA patients. Using conditional knock-out and overexpression systems, we found that Myc activity in mTCs is necessary and sufficient to induce this Tuft-to- Neuroendocrine-Transition (TNT). Citation Format: Daniel J. Salas-Escabillas, Megan T. Hoffman, Sydney M. Brender, Jacee S. Moore, Hui-Ju Wen, Simone Benitz, Erick T. Davis, Daniel Long, Allison Wombwell, Nina G. Steele, Brittany Allen-Petersen, Rosalie C. Sears, Ichiro Matsumoto, Kathleen E. DelGiorno, Howard C. Crawford. Tuft cells transdifferentiate to neural-like progenitor cells in the progression of 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 C067.

Dinaciclib exerts a tumor-suppressing effect via β-catenin/YAP axis in pancreatic ductal adenocarcinoma.

Dinaciclib, a cyclin-dependent kinase-5 (CDK5) inhibitor, has significant anti-tumor properties. However, the precise mechanism of dinaciclib requires further investigation. Herein, we investigated the anti-tumor functions and molecular basis of dinaciclib in pancreatic ductal adenocarcinoma (PDAC). PDAC and matched para-carcinoma specimens were collected from the patients who underwent radical resection. Immunohistochemistry was performed to assess CDK5 expression. Cell proliferation ability, migration, and invasion were measured using Cell Counting Kit-8, wound healing, and transwell assay, respectively. The cell cycle and apoptosis were assessed using flow cytometry. Gene expression was examined using RNA-seq and quantitative real-time PCR. Protein expression of proteins was measured by western blot analysis and immunofluorescence microscopy. Tumor-bearing mice were intraperitoneally injected with dinaciclib. CDK5 is highly expressed in PDAC. The expression level of CDK5 was significantly related to tumor size, T stage, and the American Joint Committee on Cancer stage. High CDK5 expression can predict poor survival in PDAC patients. In addition, the expression level of CDK5 might be an independent prognostic factor for PDAC patients. Dinaciclib inhibits the growth and motility of PDAC cells and induces apoptosis and cell cycle arrest in the G2/M phase. Mechanistically, dinaciclib down-regulated yes-associated protein (YAP) mRNA and protein expression by reducing β-catenin expression. Moreover, dinaciclib significantly inhibited PDAC cell growth in vivo . Our findings reveal a novel anti-tumor mechanism of dinaciclib in which it decreases YAP expression by down-regulating β-catenin at the transcriptional level rather than by activating Hippo pathway-mediated phosphorylation-dependent degradation.

DDX18 drives tumor immune escape through transcription-activated STAT1 expression in pancreatic cancer.

Pancreatic ductal adenocarcinoma (PDAC) resists to current treatments due to its inherent tumor heterogeneity, therapy-resistant cancer stem/initiating cells survival, and immune evasion in the immunosuppressive tumor microenvironment (TME). Here, the results show that clinical PDAC and adjacent tissues undergo distinct chromatin remodeling. Multiple omics analysis revealed DEAD-box RNA helicase 18 (DDX18), a carcinogenic gene with similar H3K4me3 profile, is up-regulated and correlates with poor survival in PDAC patients. We validated that DDX18 deposits on the STAT1 promoter region and counteracts H3K27me3 deposition on the STAT1 promoter sequence by modulating the formation of the PRC2 complex to up-regulate the expression of STAT1, which results in the up-regulation of PD-L1 expression, T lymphocyte accumulation and overactivation in the highly desmoplastic and immunosuppressive pancreatic TME. DDX18-STAT1 axis inhibition also affects stemness of cancer cells, epithelial-mesenchymal transition (EMT) and disrupts the immunosuppressive TME simultaneously, producing sustained remissions of aggressive PDAC by synergizing with anti-PD-L1 therapy. Combining DDX18 inhibition with anti-PD-L1 immunochemotherapy to treat PDAC patients will pave a new way for clinical treatment of patients with PDAC. This study found that clinical PDAC and adjacent pancreatic tissues undergo distinct chromatin remodeling featured by the upregulation of DEAD-box RNA helicase 18 (DDX18). We further validated that DDX18 deposits on the STAT1 promoter region and counteracts H3K27me3 deposition on the STAT1 promoter by modulating the formation of the PRC2 complex to up-regulate the expression of STAT1. DDX18-STAT1 axis enhances the stemness of cancer cells, the upregulation of PD-L1 expression, T lymphocyte accumulation and overactivation in the highly desmoplastic and immunosuppressive pancreatic TME.

The Tango between Cancer-Associated Fibroblasts (CAFs) and Immune Cells in Affecting Immunotherapy Efficacy in Pancreatic Cancer.

The lack of response to therapy in pancreatic ductal adenocarcinoma (PDAC) patients has contributed to PDAC having one of the lowest survival rates of all cancer types. The poor survival of PDAC patients urges the exploration of novel treatment strategies. Immunotherapy has shown promising results in several other cancer types, but it is still ineffective in PDAC. What sets PDAC apart from other cancer types is its tumour microenvironment (TME) with desmoplasia and low immune infiltration and activity. The most abundant cell type in the TME, cancer-associated fibroblasts (CAFs), could be instrumental in why low immunotherapy responses are observed. CAF heterogeneity and interactions with components of the TME is an emerging field of research, where many paths are to be explored. Understanding CAF-immune cell interactions in the TME might pave the way to optimize immunotherapy efficacy for PDAC and related cancers with stromal abundance. In this review, we discuss recent discoveries on the functions and interactions of CAFs and how targeting CAFs might improve immunotherapy.

Abstract B009: Multi-omics analysis of glutathione S-transferase P1 (GSTP1) knockdown model suggests role in pancreatic ductal adenocarcinoma (PDAC) cell metabolism

Abstract GSTP1 belongs to a large family of detoxification enzymes essential for cellular homeostasis in healthy cells. GSTP1 is overexpressed in many cancers and is associated with poor survival in PDAC patients. Studies in glioblastoma and breast cancer have suggested crucial roles of GST family members in metabolism through increased lactate dehydrogenase A and increased GAPDH activity, respectively. The mechanisms underlying the action of GSTP1 in PDAC metabolism and cell proliferation are not well studied. To begin characterizing the diverse functions of GSTP1 in cancer cells, we knocked down GSTP1 expression in three different PDAC cell lines. We revealed unique changes in the global transcriptomic, proteomic, and metabolomic signatures associated with GSTP1 knockdown compared to control cells through multi-omics techniques, real-time ATP production assays, and comprehensive hydrophilic metabolite and phospholipid panels. Through preliminary transcriptomic and proteomic analyses, we found 461 genes and 57 proteins differentially expressed in our GSTP1 knockdown MIA PaCa-2 cells. Among these, 41 were similarly upregulated or downregulated at the mRNA and protein levels. Pathway analysis of these changes revealed profound effects on energy production and lipid metabolism in GSTP1 knockdown cells. Hydrophilic metabolite and phospholipid panels and real-time ATP production assays supported our findings. Our results indicate that GSTP1 knockdown may directly influence mitochondrial dysfunction and lipid peroxidation pathways through associated reductions in ALDH7A1, GLUT3, and SLC7A11 levels. Continued research to further elucidate precise mechanisms and understand GSTP1’s role in driving cancer cell metabolism will aid in establishing GSTP1 targeting as a therapeutic opportunity for PDAC treatment. Citation Format: Jenna Duttenhefner, Katie M. Reindl. Multi-omics analysis of glutathione S-transferase P1 (GSTP1) knockdown model suggests role in pancreatic ductal adenocarcinoma (PDAC) cell metabolism [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer; 2022 Sep 13-16; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2022;82(22 Suppl):Abstract nr B009.

PLA2G2A Phospholipase Promotes Fatty Acid Synthesis and Energy Metabolism in Pancreatic Cancer Cells with K-ras Mutation.

Oncogenic K-ras is often activated in pancreatic ductal adenocarcinoma (PDAC) due to frequent mutation (>90%), which drives multiple cellular processes, including alterations in lipid metabolism associated with a malignant phenotype. However, the role and mechanism of the altered lipid metabolism in K-ras-driven cancer remains poorly understood. In this study, using human pancreatic epithelial cells harboring inducible K-rasG12D (HPNE/K-rasG12D) and pancreatic cancer cell lines, we found that the expression of phospholipase A2 group IIA (PLA2G2A) was upregulated by oncogenic K-ras. The elevated expression of PLA2G2A was also observed in pancreatic cancer tissues and was correlated with poor survival of PDAC patients. Abrogation of PLA2G2A by siRNA or by pharmacological inhibition using tanshinone I significantly increased lipid peroxidation, reduced fatty acid synthase (FASN) expression, and impaired mitochondrial function manifested by a decrease in mitochondrial transmembrane potential and a reduction in ATP production, leading to the inhibition of cancer cell proliferation. Our study suggests that high expression of PLA2G2A induced by oncogenic K-ras promotes cancer cell survival, likely by reducing lipid peroxidation through its ability to facilitate the removal of polyunsaturated fatty acids from lipid membranes by enhancing the de novo fatty acid synthesis and energy metabolism to support cancer cell proliferation. As such, PLA2G2A might function as a downstream mediator of K-ras and could be a potential therapeutic target.

Co‐expression of Zeb‐1 and K17 provides new insights into the role of EMT in pancreatic cancer progression

INTRODUCTIONDespite improvements in screening and therapeutic options, pancreatic ductal adenocarcinoma (PDAC) has the third‐worst mortality rate, highlighting the need for better diagnostic and therapeutic approaches. Epithelial‐mesenchymal transition (EMT) has been conferred with a greater likelihood of metastasis in multiple cancers, specifically identified through markers like Zeb‐1 and Twist‐1. Zeb‐1 (Zinc Finger E‐Box Binding Homeobox 1) was shown to repress E‐cadherin expression, consequently inducing EMT.Our lab has previously identified K17 (Keratin 17) as an independent prognostic biomarker of poor outcomes in PDAC patients. While increasing evidence has suggested that EMT drives pancreatic cancer migration and metastasis, there have not been any well‐established connections between K17 expression and EMT. This study aims to provide new insights into the role of K17 in EMT by evaluating the co‐expression of K17 and Zeb‐1 in human PDAC biopsies. We hypothesized that tumor cells co‐express K17 and Zeb‐1 as a hallmark of EMT in PDAC. In addition, our study aims to establish a novel method of scoring Zeb‐1 expression in stromal fibroblasts via immunofluorescence to remove variability in the stromal constituency.METHODSPDAC tissues obtained from patients who underwent surgical resection between 2013 and 2017 at Stony Brook University (n=20) were selected for serial sectioning. Double immunofluorescence and single immunohistochemical staining were achieved by incubation with Zeb‐1 and K17 antibodies followed by incubation of fluorescent or biotin conjugated secondary antibodies, respectively. Histologic examination was performed using immunofluorescence staining along with H&E and immunohistochemical staining (IHC) to determine concordance.RESULTS● IHC for Zeb‐1 was detected predominantly in the nuclei of centroacinar cells, benign proliferative ducts, and stromal fibroblasts. Focal nuclear staining in tumor epithelial was also detected in some cases.● High levels of K17 and but decreased expression of Zeb‐1 were detected in diffusely infiltrative cells with morphologic features of EMT.● Both high Zeb‐1 and high K17 protein expression were associated with poor survival of PDAC patients.● Using confocal immunofluorescence microscopy, we detected co‐localization of Zeb‐1 and K17 in PDAC.● We quantified Zeb‐1 protein expression in PDAC samples from 20 patients and confirmed significantly higher expression in the tumor‐associated stroma of high‐grade tumors compared to stromal fibroblasts in areas devoid of tumor.CONCLUSIONOur results provide evidence that K17 is upregulated in tumor epithelial cells that show morphologic immunohistochemical properties of EMT. The loss of Zeb‐1 in diffusely infiltrative, K17‐positive tumor cells suggests that EMT upregulates K17, driving tumor aggression. Our results suggest that K17 may enhance PDAC aggression by mediating EMT. Hence, novel therapeutic agents targeting K17 may improve clinical care for this biologically distinct patient subgroup.

The MYEOV-MYC association promotes oncogenic miR-17/93-5p expression in pancreatic ductal adenocarcinoma

Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal malignancy worldwide. As metastasis and malignant progression are primarily responsible for the poor clinical outcomes of PDAC, identifying key genes involved in these processes and the underlying molecular mechanisms of PDAC is vital. In this study, by analyzing TCGA PDAC data and matched GTEx data, we found that MYEOV expression is associated with poor survival in PDAC patients and higher in carcinoma tissues than in healthy tissues. Elevated levels of MYEOV led to enhanced cell proliferation, invasion and migration in vitro and in vivo. Transcriptome analysis results revealed that MYEOV mediates global alterations in gene expression profiles in PDAC cells. MiRNA-seq analysis showed that MYEOV regulates the expression levels of miR-17-5p and miR-93-5p, and its depletion resulted in reduced cell proliferation, invasion and migration, as observed in MYEOV-knockdown PDAC cells. These effects are likely due to the ability of MYEOV to regulate enrichment of the transcription factor MYC at the gene promoter regions of the two miRNAs. Furthermore, we identified a complex containing MYEOV and MYC in the nucleus, providing additional evidence for the association of MYEOV with MYC. Taken together, our results suggest that MYEOV promotes oncogenic miR-17/93-5p expression by associating with MYC, contributing to PDAC progression.

Targeting Pin1 renders pancreatic cancer eradicable by synergizing with immunochemotherapy

Pancreatic ductal adenocarcinoma (PDAC) is characterized by notorious resistance to current therapies attributed to inherent tumor heterogeneity and highly desmoplastic and immunosuppressive tumor microenvironment (TME). Unique proline isomerase Pin1 regulates multiple cancer pathways, but its role in the TME and cancer immunotherapy is unknown. Here, we find that Pin1 is overexpressed both in cancer cells and cancer-associated fibroblasts (CAFs) and correlates with poor survival in PDAC patients. Targeting Pin1 using clinically available drugs induces complete elimination or sustained remissions of aggressive PDAC by synergizing with anti-PD-1 and gemcitabine in diverse model systems. Mechanistically, Pin1 drives the desmoplastic and immunosuppressive TME by acting on CAFs and induces lysosomal degradation of the PD-1 ligand PD-L1 and the gemcitabine transporter ENT1 in cancer cells, besides activating multiple cancer pathways. Thus, Pin1 inhibition simultaneously blocks multiple cancer pathways, disrupts the desmoplastic and immunosuppressive TME, and upregulates PD-L1 and ENT1, rendering PDAC eradicable by immunochemotherapy.

Multiplex Patient-Based Drug Response Assay in Pancreatic Ductal Adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDA) is an extremely lethal malignancy arising from the pancreas. The treatment of PDA is complicated by ineffective treatments and a lack of biomarkers predictive of treatment success. We have designed a patient-derived organoid (PDO) based high-throughput drug screening assay to model treatment response to a variety of conventional and investigational treatments for PDA. Consecutive patients undergoing endoscopic ultrasound-guided fine-needle biopsy for tissue diagnosis of PDA at Rush University Medical Center were offered to participate in the study. Biopsies were immediately processed to develop organoids. Fifteen PDOs were screened for sensitivity to 18 compounds, including conventional PDA chemotherapies and FDA-approved investigational targeted therapies in cancer using Cell-titer GLO 3D (Promega) cell viability assay. The area under the curve (AUC) was calculated and normalized to the maximum area under the curve to generate a normalized AUC between 0 and 1. Molecular profiling of PDOs was conducted using RNA-seq. Human PDA transcriptomic was extracted from The Cancer Genome Atlas (TCGA). The drug response curves were reproducible. We observed variation in response to conventional therapies overall as well as among individual patients. There were distinct transcriptome signatures associated with response to the conventional chemotherapeutics in PDA. The transcriptomic profile of overall resistance to conventional therapies in our study was associated with poor survival in PDA patients in TCGA. Our pathway analysis for targeted drugs revealed a number of predictors of response associated with the mechanism of action of the tested drug. The multiplex organoid-based drug assay could be used in preclinical to inform patient stratification and therapeutic selection in PDA. When combined with omics data, ex vivo response to treatment could help identify gene signatures associated with response to novel therapies.

Low expression of tRF-Pro-CGG predicts poor prognosis in pancreatic ductal adenocarcinoma.

ABSTRACTBackground & AimstRFs (tRNA‐derived RNA fragments) have been reported to facilitate cancer progression in multiple cancers. However, their role in pancreatic ductal adenocarcinoma (PDAC) remains to be determined. In this study, we mainly investigated the expression of tRF‐Pro‐CGG in pancreatic ductal adenocarcinoma and evaluated its relationship with the clinicopathology and survival time of patients.Methods37 cases of pancreatic ductal adenocarcinoma, and 15 cases of normal pancreatic tissues were collected which were resected by surgery from January 2017 to June 2020 from the Department of Hepatobiliary and Pancreatic surgery of Changzhou second people's Hospital. The expression of tRF‐Pro‐CGG in paraffin‐embedded tissues was detected by fluorescence in situ hybridization (FISH). The clinical data including age, sex, tumor location, tumor diameter, tumor clinical stage (TNM stage), depth of invasion, regional lymph node metastasis, serum CA199, and serum CEA were collected and analyzed retrospectively, whether the expression tRF‐Pro‐CGG was correlation with the pathological parameters and clinical outcomes of patients.ResultsThe expression level of tRF‐Pro‐CGG was significantly downregulated in PDAC and associated with an advanced TNM stage (P=0.000) and the N stage (P=0.000) of patients. More importantly, low tRF‐Pro‐CGG expression predicted poor survival in PDAC patients (P=0.003).ConclusionsTRF‐Pro‐CGG is under‐expressed in PDAC and is associated with short clinical survival and poor prognosis. tRF‐Pro‐CGG is an independent prognostic factor, which highlights its role as a potential biomarker for PDAC progression and therapy.

KLHDC7B-DT aggravates pancreatic ductal adenocarcinoma development via inducing cross-talk between cancer cells and macrophages.

Tumor microenvironment (TME) exerts key roles in pancreatic ductal adenocarcinoma (PDAC) development. However, the factors regulating the cross-talk between PDAC cells and TME are largely unknown. In the present study, we identified a long noncoding RNA (lncRNA) KLHDC7B divergent transcript (KLHDC7B-DT), which was up-regulated in PDAC and correlated with poor survival of PDAC patients. Functional assays demonstrated that KLHDC7B-DT enhanced PDAC cell proliferation, migration, and invasion. Mechanistically, KLHDC7B-DT was found to directly bind IL-6 promoter, induce open chromatin structure at IL-6 promoter region, activate IL-6 transcription, and up-regulate IL-6 expression and secretion. The expression of KLHDC7B-DT was positively correlated with IL-6 in PDAC tissues. Via inducing IL-6 secretion, KLHDC7B-DT activated STAT3 signaling in PDAC cells in an autocrine manner. Furthermore, KLHDC7B-DT also activated STAT3 signaling in macrophages in a paracrine manner, which induced macrophage M2 polarization. KLHDC7B-DT overexpressed PDAC cells-primed macrophages promoted PDAC cell proliferation, migration, and invasion. Blocking IL-6/STAT3 signaling reversed the effects of KLHDC7B-DT on macrophage M2 polarization and PDAC cell proliferation, migration, and invasion. In conclusion, KLHDC7B-DT enhanced malignant behaviors of PDAC cells via IL-6-induced macrophage M2 polarization and IL-6-activated STAT3 signaling in PDAC cells. The cross-talk between PDAC cells and macrophages induced by KLHDC7B-DT represents potential therapeutic target for PDAC.

Upregulated GDF-15 expression facilitates pancreatic ductal adenocarcinoma progression through orphan receptor GFRAL.

Growth and differentiation factor 15 (GDF-15) has been studied as an important hallmark of cancer. However, the receptor of GDF-15 in pancreatic cancer cell remains unclear. Here, we investigated its biological effects in pancreatic ductal adenocarcinoma (PDAC). We found that aberrant GDF-15 expression positively correlated with poor survival of PDAC patients. GDF-15 protein enhanced tumor cell proliferation in two pancreatic cancer lines, AsPC-1 and BxPC-3. Knockdown GDF-15 attenuated its biological function in vitro and reduced PDAC cell tumorigenesis upon xenotransplantation into nude mice. Moreover, we identified that glial-derived neurotropic factor family receptor α-like (GFRAL) was upregulated in PDAC tissues and positively correlated with GDF-15 expression. High GFRAL expression was significantly associated with poor survival in PDAC patients. Furthermore, we identified that the biological effects of GDF-15 are mediated by its receptor GFRAL which is present in PDAC cells. After overexpression GFRAL in pancreatic cancer cells, the effect of GDF-15 was significantly enhanced. Overall, our findings demonstrated that the GDF-15 secreted by PDAC cells, binds to GFRAL, itself localized in PDAC cells, to promote cancer cell growth and metastasis through the GDF-15/GFRAL signaling pathway.

Abstract 6041: LAMC2 as a therapeutic target in pancreatic ductal adenocarcinoma

Abstract BACKGROUND. Pancreatic cancer stands as one of the deadliest tumors, in part due to the limited efficacy of currently existing therapies. Pancreatic ductal adenocarcinoma (PDAC) represents the predominant subtype (around 85% of total cases), where KRAS is the most frequently mutated oncogene. Tumor microenvironment in PDAC is characterized by a desmoplastic stroma which is composed, among other molecules, of collagens of different types, fibronectin and laminins. A member of the laminin family, LAMININ γ2 or LAMC2, was previously identified by our group as part of a cross-tumors KRAS signature whose high expression was a marker of poor survival in PDAC patients. Given the relevant functional implication of other members of the cross-tumors KRAS signature in PDAC, we aimed to investigate the role of LAMC2 in this tumor type. METHODS. A meta-analysis of several human PDAC data sets (n=6) and survival analysis of the TCGA data were performed to query the expression levels and prognosis significance of LAMC2. Human and mouse PDAC cell lines as well as a mouse model of PDAC (Kras, Tp53f/f, Ptf1aCre) were used to assess LAMC2 expression. In vitro (2D and 3D organoids) and in vivo models derived from human and mouse PDAC cell lines or primary tumors were used to define the functional role of LAMC2 in PDAC. Lastly, cellular and molecular analysis were deployed to dissect the mechanism of action of LAMC2 in PDAC. RESULTS. At the clinical level, we have observed that LAMC2 is overexpressed in human PDAC patients with regard to normal tissue and that its high levels identify the group of patients with the worst survival. The overexpression of LAMC2 was recapitulated in human and mouse cell lines. Genetic depletion of LAMC2 had an adverse effect in 2D proliferation, clonogenic efficiency, 3D organoid growth, and PDAC-based xenograft/allograft tumor development. This deleterious effect is driven by an induction in apoptosis and a decrease in S phase consistent across species. Mechanistically, LAMC2 is mainly linked to the KRAS pathway through ERK1/2 and is transcriptionally regulated by the transcription factor FOSL1/AP1, which was also a member of the cross-tumors signature. These data suggest that LAMC2 is involved in the control of different functions related to the PDAC phenotype induced by KRAS. CONCLUSION. LAMC2 is a molecular target in PDAC with clinical implications based on the potential development of new therapeutic and diagnostic strategies for PDAC patients. Citation Format: Oihane Erice, Ester Blanco, Karmele Valencia, Elizabeth Guruceaga, Haritz Moreno, Fernando Lecanda, Vincenzo Corbo, Daniel Ölund, Mariano Ponz-Sarvise, Vicent Silve. LAMC2 as a therapeutic target in pancreatic ductal adenocarcinoma [abstract]. In: Proceedings of the Annual Meeting of the American Association for Cancer Research 2020; 2020 Apr 27-28 and Jun 22-24. Philadelphia (PA): AACR; Cancer Res 2020;80(16 Suppl):Abstract nr 6041.

Reduced mean platelet volume levels predict shorter survival in patients with resectable pancreatic ductal adenocarcinoma and type 2 diabetes

BackgroundType 2 diabetes mellitus (T2DM) increased the risk of developing pancreatic cancer. Pancreatic ductal adenocarcinoma (PDAC) is the most common neoplastic disease originating from the pancreas. Increasing evidence indicates that platelets activation plays a prominent role in tumor and T2DM. Mean platelet volume (MPV) is an indicator of activated platelets and is altered in several cancers. The current study aimed to evaluate the prognostic role of MPV in resectable PDAC patients with T2DM.MethodsEight hundred and three patients with PDAC were included in this retrospective study. We determined the optimal cutoff value of MPV for 5-year overall survival (OS) using the receiver operating characteristic (ROC) method. The associations between MPV levels and clinical characteristics were analyzed. Kaplan-Meier survival analysis and Cox’s proportional hazard regression model were used to evaluate the prognostic value of MPV for OS.ResultsCompared to the PDAC patients without T2DM, MPV levels were significantly higher in the PDAC patients with T2DM. Moreover, MPV was significantly associated with the differentiation between T2DM and non-T2DM. In addition, Kaplan-Meier analysis found that patients with low MPV levels had a poorer 5-year OS than patients with high MPV levels in diabetic patients. Multivariate analyses revealed that MPV was an independent prognostic factor for OS in patients with T2DM. However, the independent prognostic role of MPV was not observed in patients without T2DM.ConclusionMPV independently predicts poor survival in PDAC patients with T2DM. Prospective studies are required to confirm the role of MPV in PDAC.

Inflammatory IFIT3 renders chemotherapy resistance by regulating post-translational modification of VDAC2 in pancreatic cancer.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal cancers worldwide and effective therapy remains a challenge. IFIT3 is an interferon-stimulated gene with antiviral and pro-inflammatory functions. Our previous work has shown that high expression of IFIT3 is correlated with poor survival in PDAC patients who receive chemotherapy suggesting a link between IFIT3 and chemotherapy resistance in PDAC. However, the exact role and molecular mechanism of IFIT3 in chemotherapy resistance in PDAC has been unclear.Methods: A group of transcriptome datasets were downloaded and analyzed for the characterization of IFIT3 in PDAC. Highly metastatic PDAC cell line L3.6pl and patient-derived primary cell TBO368 were used and IFIT3 knockdown and the corresponding knockin cells were established for in vitro studies. Chemotherapy-induced apoptosis, ROS production, confocal immunofluorescence, subcellular fractionation, chromatin-immunoprecipitation, co-immunoprecipitation and mass spectrometry analysis were determined to further explore the biological role of IFIT3 in chemotherapy resistance of PDAC.Results: Based on PDAC transcriptome data, we show that IFIT3 expression is associated with the squamous molecular subtype of PDAC and an increase in inflammatory response and apoptosis pathways. We further identify a crucial role for IFIT3 in the regulation of mitochondria-associated apoptosis during chemotherapy. Knockdown of IFIT3 attenuates the chemotherapy resistance of PDAC cells to gemcitabine, paclitaxel, and FOLFIRINOX regimen treatments, independent of individual chemotherapy regimens. While IFIT3 overexpression was found to promote drug resistance. Co-immunoprecipitation identified a direct interaction between IFIT3 and the mitochondrial channel protein VDAC2, an important regulator of mitochondria-associated apoptosis. It was subsequently found that IFIT3 regulates the post-translational modification-O-GlcNAcylation of VDAC2 by stabilizing the interaction of VDAC2 with O-GlcNAc transferase. Increased O-GlcNAcylation of VDAC2 protected PDAC cells from chemotherapy induced apoptosis.Conclusions: These results effectively demonstrate a central mechanism by which IFIT3 expression can affect chemotherapy resistance in PDAC. Targeting IFIT3/VDAC2 may represent a novel strategy to sensitize aggressive forms of pancreatic cancer to conventional chemotherapy regimens.

Elucidating the roles of ASPM isoforms reveals a novel prognostic marker for pancreatic cancer.

Pancreatic ductal adenocarcinoma (PDAC) is one of the deadliest cancers worldwide. Late diagnosis, desmoplastic tissue and intrinsic resistance to therapy are among the main reasons for its aggressive phenotype. In addition, it is now appreciated that cancer stem cells-a rare subpopulation of tumor cells highly resistant to therapy-are crucial players in PDAC initiation, progression and resistance to therapy. In a recent article in The Journal of Pathology, Hsu et al elucidated the specific roles of abnormal spindle-like, microcephaly-associated protein (ASPM) isoforms in PDAC. The authors reported that ASPM isoform I (ASPM-iI) is mainly expressed in the cytoplasm of PDAC cells. Its expression is associated with the Wnt signaling pathway, which promotes stemness and maintains the cancer stem cell niche. Clinically, expression of ASPM-iI correlates with poor survival in PDAC patients. Thus, this study revealed a novel prognostic marker as well as a potential therapeutic target for PDAC. © 2019 Pathological Society of Great Britain and Ireland. Published by John Wiley & Sons, Ltd.

Abstract 4328: Galectin-3 mediates tumor-stromal interaction by activating pancreatic stellate cell through integrinβ1/ILK/NF-κB signaling

Abstract Background: Pancreatic ductal adenocarcinoma (PDAC) is a highly aggressive tumor with poor survival in PDAC patients. Proinflammatory cytokines/chemokines play an essential role in tumor microenvironments associated with its invasive and aggressive phenotype. Galectin-3 (Gal-3), a member of a family of β-galactoside-specific lectins, has been found to be involved in tumor progression and tissue fibrosis. However, the precise molecular mechanisms of Gal-3 mediate tumor-stromal interaction and abundant cytokine production is unknown. Methods: The effects of Gal-3 on secretion of cytokines/chemokines from stromal cells (HPSC) were assessed by cytokine array. HPSC proliferation and invasion were assessed by the MTS and Matrigel invasion assays. Co-culture and Conditional medium from PDAC tumor cells with genetic alterations of Gal-3 levels were used to study the interaction between tumor cell and stromal cells. IL-8 production in both tumor cells and stromal cells was confirmed by ELISA. Transcriptional activities of NF-κB and IL-8 were determined by transient transfection. An IKB mutant construct (IKBM), Mutation of the IL-8 promoter at the NF-κB site, and inhibitors of NF-κB (Bay11), ILK and neutral antibody of Integrinβ1 were used to determine the signaling pathways evoked by Gal-3. Results: Gal-3 expression is in both tumor cells and stromal cells of human and genetic mouse model. Secretion of cytokines such as IL-8, GM-CSF, CXCL1, CCL2 and IL6 were significantly increased in HPSC cells treated with rGal-3 which is concomitant with activation of HPSC. Conditioned medium or co-culture from Panc-1 cells in which Gal-3 was down-regulated inhibited the growth and invasion capacity of HPSC cells compared with the vector controls. rGal-3 dramatically induced production and secretion of IL-8 in HPSCs. Mechanistically, Gal-3 stimulates IL-8 secretion at the level of transcription and through Integrinβ1/ILK/NF-kB signaling. rGal-3 strongly stimulated integrin β1 expression and NF-κB transcriptional activity; and inhibitors of ILK inhibitor or NF-κB or neutral antibody of integrin β1 blocked IL-8 transcriptional activity and its secretion. NF-κB mutant construct IKBM or mutation of the NF-κB site in the IL-8 promoter completely block the induction of IL-8 activity by Gal-3. GEO data and our own patients’ cohort analysis of pancreatic tumor samples demonstrated that the level of Gal-3, IL-8 and other cytokines is significantly higher in tumor tissues than those of normal and pancreatitis. Conclusion: Collectively, these data suggest that Gal-3 secreted from tumor cells activate HPSC cells and induce many inflammatory cytokines including IL-8 production via Integrinβ1/ILK/NF-κB signaling. Thus, Gal-3 may play a critical role in mediating tumor-stromal interaction and remodeling tumor associated microenvironments. Citation Format: Wei Zhao, Yan Xu, Guha Sushovan, Rosa Hwang, Nobuo Ochi, Margarete Hafley, Robert Bresalier, Craig Logsdon, Zhenning Wang, Jaffer Ajani, Shumei Song. Galectin-3 mediates tumor-stromal interaction by activating pancreatic stellate cell through integrinβ1/ILK/NF-κB signaling [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2017; 2017 Apr 1-5; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2017;77(13 Suppl):Abstract nr 4328. doi:10.1158/1538-7445.AM2017-4328

Inhibition of Neddylation Modification Sensitizes Pancreatic Cancer Cells to Gemcitabine

Pancreatic ductal adenocarcinoma (PDAC) is the fourth leading cause of cancer death in the USA with a 5-year survival rate less than 3% to 5%. Gemcitabine remains as a standard care for PDAC patients. Although protein neddylation is abnormally activated in many human cancers, whether neddylation dysregulation is involved in PDAC and whether targeting neddylation would sensitize pancreatic cancer cells to gemcitabine remain elusive. Here we report that high expression of neddylation components, NEDD8 and NAE1, are associated with poor survival of PDAC patients. Blockage of neddylation by MLN4924, a small molecule inhibitor targeting this modification, significantly sensitizes pancreatic cancer cells to gemcitabine, as evidenced by reduced growth both in monolayer culture and soft agar, reduced clonogenic survival, decreased invasion capacity, increased apoptosis, G2/M arrest, and senescence. Importantly, combinational treatment of MLN4924-gemcitabine near completely suppressed in vivo growth of pancreatic cancer cells. Mechanistically, accumulation of NOXA, a pro-apoptotic protein and ERBIN, a RAS signal inhibitor, appears to play, at least in part, a causal role in MLN4924 chemo-sensitization. Our study demonstrates that neddylation modification is a valid target for PDAC, and provides the proof-of-concept evidence for future clinical trial of MLN4924-gemcitabine combination for the treatment of pancreatic cancer patients.

Increased semaphorin 3c expression promotes tumor growth and metastasis in pancreatic ductal adenocarcinoma by activating the ERK1/2 signaling pathway

Pancreatic cancer is characterized by neural alterations and aberrant expression of neural-specific factors. Semaphorins have been recognized as key contributors in axon guidance, the immune response and tumor progression. Recent studies have suggested the involvement of Semaphorin 3c (sema3c) in tumorigenesis and metastasis in numerous types of cancer, however, the clinical significance of sema3c and its role in the growth and metastasis of pancreatic ductal adenocarcinoma (PDAC) remain unclear. In this study, we found that aberrant sema3c expression was positively associated with a particular tumor stage and correlated with poor survival of PDAC patients. Knockdown of sema3c attenuated proliferation, migration, invasion and epithelial–mesenchymal transition (EMT) in a pancreatic cancer cell line and reduced PDAC cell tumorigenesis upon xenotransplantation into NOD/SCID mice. Overexpression of sema3c produced the opposite effects and promoted the extracellular signal-regulated kinase (ERK)1/2 signaling pathway. Overall, our findings demonstrate that aberrant expression of sema3c is correlated with poor prognosis of PDAC patients and promotes tumor growth and metastasis by activating ERK1/2 signaling pathway.