Chemoresistance In Pancreatic Ductal Adenocarcinoma Research Articles

Nimbolide-encapsulated PLGA nanoparticles induces Mesenchymal-to-Epithelial Transition by dual inhibition of AKT and mTOR in pancreatic cancer stem cells

Pancreatic ductal adenocarcinoma (PDAC) is associated with poor prognosis and remains highly aggressive despite current advancements in therapies. Chemoresistance and high metastatic nature of PDAC is attributed to a small subset of stem-like cells within the tumor known as Cancer Stem Cells (CSCs). Here, we developed a strategy for targeting pancreatic CSCs through forceful induction of mesenchymal-to-epithelial transition driven by encapsulating a phytochemical Nimbolide in nanoparticles. Binding of Nimbolide with the key regulator proteins of CSCs were studied through molecular docking and molecular dynamic simulation studies, which revealed that it binds to AKT and mTOR with high affinity. Further, in vitro studies revealed that Nim NPs are capable of inducing forceful mesenchymal-to-epithelial transition of pancreatospheres that leads to loss of multidrug resistance and self-renewal properties of pancreatospheres. Our study gives a proof of concept that encapsulation of Nim in PLGA nanoparticles increases its therapeutic effect on pancreatospheres. Further, binding of Nim to AKT and mTOR negatively regulates their activity that ultimately leads to mesenchymal-to-epithelial transition of pancreatic CSCs.

Emerging Role of miR-345 and Its Effective Delivery as a Potential Therapeutic Candidate in Pancreatic Cancer and Other Cancers.

Pancreatic ductal adenocarcinoma (PDAC) is an aggressive malignancy with high mortality, poor prognosis, and palliative treatments, due to the rapid upregulation of alternative compensatory pathways and desmoplastic reaction. miRNAs, small non-coding RNAs, have been recently identified as key players regulating cancer pathogenesis. Dysregulated miRNAs are associated with molecular pathways involved in tumor development, metastasis, and chemoresistance in PDAC, as well as other cancers. Targeted treatment strategies that alter miRNA levels in cancers have promising potential as therapeutic interventions. miRNA-345 (miR-345) plays a critical role in tumor suppression and is differentially expressed in various cancers, including pancreatic cancer (PC). The underlying mechanism(s) and delivery strategies of miR-345 have been investigated by us previously. Here, we summarize the potential therapeutic roles of miR-345 in different cancers, with emphasis on PDAC, for miRNA drug discovery, development, status, and implications. Further, we focus on miRNA nanodelivery system(s), based on different materials and nanoformulations, specifically for the delivery of miR-345.

Abstract PO-098: Longitudinal profiling of pancreatic cancer patients identifies interleukin-8 as a mediator of myeloid-epithelial crosstalk

Abstract Background: Pancreatic ductal adenocarcinoma (PDAC) is the 4th leading cause of cancer-related death in the US. A key hallmark of this disease is that, while tumors initially show susceptibility to standard chemotherapeutic agents, most patients eventually develop resistance, leading to poor survival. While the mechanisms of chemoresistance are unclear, murine studies have implicated the myeloid compartment of the tumor immune microenvironment. Correlative data in human tumors supports this notion, however, mechanistic studies are lacking, thus impairing translation to the clinic. The study of human pancreatic cancer has historically been challenging due to difficulty of fresh biospecimen acquisition, patient heterogeneity, and a diverse tumor microenvironment. Moreover, the vast majority of pancreatic cancer patients do not qualify for surgical resection, further limiting tissue availability. We have overcome these difficulties by developing a pipeline to analyze human tumor samples and matched blood using high-fidelity techniques including single-cell RNA sequencing (scRNAseq) and mass cytometry (CyTOF), together with establishment of organoids from the same tumors. Notably, in this pipeline we can use small amounts of tissue from endoscopic fine needle biopsies, thus allowing us to sample tumors from patients at any disease stage. Results: We performed CyTOF on longitudinally-matched peripheral blood mononuclear cells (PBMCs) from 30 patients and single-cell RNA sequencing on 6 patients in the treatment naïve and on-treatment (FOLFIRINOX) state. CyTOF revealed distinct alterations in the myeloid population, with a shift toward CXCR2hiPD-L1hi granulocytes with FOLFIRINOX treatment over time. Analysis of PBMCs from scRNAseq showed a distinct myeloid gene signature with FOLFIRINOX and in particular highlighted interleukin-8 (IL8), a chemokine involved in myeloid cell chemotaxis that is associated with poor prognosis in pancreatic cancer. Further mapping of IL8 in tumor tissue by scRNAseq showed that it is highly expressed in subpopulations of tumor epithelial cells and tumor-infiltrating granulocytes. IL8-high tumor-infiltrating granulocytes also highly expressed VEGF and CXCR4, suggesting immunosuppressive and angiogenic roles. IL8-high tumor epithelial cells were found to have a basal-like phenotype and also expressed a network of other chemokines including CXCL1, CXCL3, CXCL5, which are known to recruit immunosuppressive myeloid cells. Conclusions: Through longitudinal and multimodal mapping using PDAC patient blood and tumor biospecimens, we have identified IL8 as a potential mediator of epithelial-myeloid crosstalk in PDAC chemoresistance and tumor aggression. Validation studies using an all-human co-culture system of PDAC patient-derived organoids and myeloid cells are currently underway. Citation Format: Eileen S. Carpenter, Samantha Kemp, Padma Kadiyala, Nina Steele, Ahmed Elhossiny, Stephanie The, Valerie Gunchick, Rémy Nicolle, Michelle Anderson, Wenting Du, Carlos Espinoza, Richard Kwon, Erik-Jan Wamsteker, Anoop Prabhu, Allison Schulman, Vaibhav Sahai, Timothy Frankel, Filip Bednar, Marina Pasca di Magliano. Longitudinal profiling of pancreatic cancer patients identifies interleukin-8 as a mediator of myeloid-epithelial crosstalk [abstract]. In: Proceedings of the AACR Virtual Special Conference on Pancreatic Cancer; 2021 Sep 29-30. Philadelphia (PA): AACR; Cancer Res 2021;81(22 Suppl):Abstract nr PO-098.

Abstract PO-066: High uptake, retention, and in vivo activity of L-Annamycin in pancreatic cancer models

Abstract Introduction: Annamycin (ANN) is a non-cardiotoxic potent topoisomerase II poison that is structurally related to doxorubicin (Dox) but it displays significantly different biological properties. ANN is formulated in multilamellar liposomes to enable effective in vivo administration as L-ANN. Pancreatic ductal adenocarcinoma (PDAC) represents a major challenge and is the deadliest cancer in decades. Pancreas drug uptake and retention remains a significant factor contributing to the chemoresistance of PDAC. Additionally, drug resistance on a cellular level further limits the therapeutic options for pancreatic cancer patients. Based on unique biological properties, we select L-ANN for detailed characterization of anti-tumor potency in preclinical PDAC models. Objective: The objective of this study was to validate the L-ANN exceptional uptake in pancreatic cancer cells and tumors and explore its high pancreas uptake to target PDACs by assessing the efficacy of L-ANN in the preclinical mouse models. Methods: The uptake, cytotoxicity, and L-ANN-induced apoptosis were studied in human MDA-PATC50 and MDA-PATC53 cell lines. L-ANN pharmacokinetics and tissue-organ distribution were analyzed in CD-1 mice after bolus administration of L-ANN, ANN or DOX. Subcutaneous and orthotopic MDA-PATC50 and MDA-PATC53 mouse models were used to assess activity of L-ANN in vivo. Results: Fluorescent microscopy and FACS analysis showed significantly higher uptake of ANN in MDA-PATC50 and MDA-PATC53 when compared to DOX-treated cells. Interestingly, ANN displayed distinct subcellular distribution with predominantly cytosolic localization. Average IC50 of ANN or DOX treated MDA-PATC50 and MDA-PATC53 cells were 25.1 nM and 41.9 nM for ANN and 32.8 nM and 130.4 nM for DOX respectively (72h exposure). Biodistribution study showed rapid L-Ann absorption in the pancreas. The Cmax of L-Ann was 15-fold higher than Dox after administration of L-ANN or Dox (both at 4 mg/kg) in pancreatic tissue. The subcutaneous models demonstrated dose-dependent reduction in tumor volumes after 3 doses of L-Ann in both PDAC models. Additionally, efficacy of L-ANN was assessed in two orthotopic PDAC models. MRI confirmed tumor volume reduction in L-Ann 3 mg/kg and 4 mg/kg doses in MDA-PATC50 model (p<0.05 for both 3 and 4 mg), and in MDA-PATC53 model (p<0.001 and p<0.0001, respectively). The median survivals (MS) were 135 and 103 days for L-Ann at 3 mg/kg and 4 mg/kg respectively (MS for vehicle was 49 days, p<0.0001). MDA-PATC50 orthotopic model demonstrated the similar results (ongoing study). Conclusion: Annamycin exhibits high and rapid uptake in the pancreatic cancer cells and in pancreas itself when compared to Dox. Subsequently, ANN appeared to be highly cytotoxic against tested pancreatic cancer cell lines and as L-ANN potently inhibits tumor growth and increase survival in experimental models of pancreatic cancer. These experiments demonstrate remarkable activity of L-ANN in PDAC models and support future clinical studies. Citation Format: Ya'an Kang, Rafal Zielinski, Roberto Cardenas Zuniga, Radjendirane Venugopal, Maria Poimenidou, Magdalena Remiszewski, Shaohua Peng, Edd Felix, Krzysztof Grela, Stanislaw Skora, Van N. Nguyen, Izabela Fokt, Waldemar Priebe. High uptake, retention, and in vivo activity of L-Annamycin in pancreatic cancer models [abstract]. In: Proceedings of the AACR Virtual Special Conference on Pancreatic Cancer; 2021 Sep 29-30. Philadelphia (PA): AACR; Cancer Res 2021;81(22 Suppl):Abstract nr PO-066.

The next "sweet" spot for pancreatic ductal adenocarcinoma: Glycoprotein for early detection.

Pancreatic ductal adenocarcinoma (PDAC) is the most common neoplastic disease of the pancreas, accounting for more than 90% of all pancreatic malignancies. As a highly lethal malignancy, PDAC is the fourth leading cause of cancer-related deaths worldwide with a 5-year overall survival of less than 8%. The efficacy and outcome of PDAC treatment largely depend on the stage of disease at the time of diagnosis. Surgical resection followed by adjuvant chemotherapy remains the only possibly curative therapy, yet 80%-90% of PDAC patients present with nonresectable PDAC stages at the time of clinical presentation. Despite our advancing knowledge of PDAC, the prognosis remains strikingly poor, which is primarily due to the difficulty of diagnosing PDAC at the early stages. Recent advances in glycoproteomics and glycomics based on mass spectrometry have shown that aberrations in protein glycosylation plays a critical role in carcinogenesis, tumor progression, metastasis, chemoresistance, and immuno-response of PDAC and other types of cancers. A growing interest has thus been placed upon protein glycosylation as a potential early detection biomarker for PDAC. We herein take stock of the advancements in the early detection of PDAC that were carried out with mass spectrometry, with special focus on protein glycosylation.

Epithelial to Mesenchymal Transition: Key Regulator of Pancreatic Ductal Adenocarcinoma Progression and Chemoresistance.

Simple SummaryPancreatic ductal adenocarcinoma’s (PDAC) dismal prognosis is associated with its aggressive biological behavior and resistance to chemotherapy. Epithelial to mesenchymal transition (EMT) has been recognized as a key driver of PDAC progression and development of drug resistance. EMT is a transient and reversible process leading to transdifferentiation of epithelial cells into a more mesenchymal phenotype. It is regulated by multiple signaling pathways that control the activity of a transcription factors network. Activation of EMT in pre-invasive stages of PDAC has been accused for early dissemination. Furthermore, it contributes to the development of intratumoral heterogeneity and drug resistance. This review summarizes the available data regarding signaling networks regulating EMT and describes the integral role of EMT in different aspects of PDAC pathogenesis.Pancreatic ductal adenocarcinoma (PDAC) is one of the deadliest malignancies, characterized by aggressive biological behavior and a lack of response to currently available chemotherapy. Emerging evidence has identified epithelial to mesenchymal transition (EMT) as a key driver of PDAC progression and a central regulator in the development of drug resistance. EMT is a reversible transdifferentiation process controlled by complex interactions between multiple signaling pathways such as TGFb, Wnt, and Notch, which converge to a network of specific transcription factors. Activation of EMT transcriptional reprogramming converts cancer cells of epithelial differentiation into a more mesenchymal phenotypic state. EMT occurrence in pre-invasive pancreatic lesions has been implicated in early PDAC dissemination. Moreover, cancer cell phenotypic plasticity driven by EMT contributes to intratumoral heterogeneity and drug tolerance and is mechanistically associated with the emergence of cells exhibiting cancer stem cells (CSCs) phenotype. In this review we summarize the available data on the signaling cascades regulating EMT and the molecular isnteractions between pancreatic cancer and stromal cells that activate them. In addition, we provide a link between EMT, tumor progression, and chemoresistance in PDAC.

CD73 induces gemcitabine resistance in pancreatic ductal adenocarcinoma: A promising target with non-canonical mechanisms

CD73, a cell surface-localized ecto-5'-nucleotidase, is the major enzymatic source of extracellular adenosine. Canonically, it plays multiple roles in cancer-related processes via its metabolite. As a druggable target, clinical trials targeting CD73 in various malignant diseases are currently ongoing. Here, we report the ecto-5'-nucleotidase-independent functions of CD73 in pancreatic ductal adenocarcinoma (PDAC). Our findings support that the elevated expression of CD73 in PDAC cells promotes gemcitabine (GEM) resistance by activating AKT. We discovered that a large amount of intracellular CD73 are localized in the endoplasmic reticulum membrane. Intracellular CD73 physically interacts with major vault protein to activate the SRC-AKT circuit. Troglitazone (TGZ) is a peroxisome proliferator-activated receptor gamma agonist that could inhibit the expression of CD73. The administration of TGZ markedly enhances sensitivity to GEM via downregulating CD73 in PDAC. Our findings support that CD73 could be targeted to overcome chemoresistance in PDAC.

CIRBP Knockdown Attenuates Tumourigenesis and Improves the Chemosensitivity of Pancreatic Cancer via the Downregulation of DYRK1B

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal malignancies worldwide with very limited treatment options. Cold-inducible RNA binding protein (CIRBP) plays promoting roles in several types of cancers, but its function remains unclear in PDAC. Here, we found that the expression of CIRBP was upregulated in PDAC tumor tissues and was significantly associated with poor prognosis. Knockdown of CIRBP in PANC-1 and SW1990 cells inhibited proliferation, migration and invasion in vitro and suppressed tumor growth in vivo. Moreover, CIRBP knockdown enhanced the antitumour effects of gemcitabine treatment in PANC-1 and SW1990 cells, whereas CIRBP overexpression exerted the opposite effects. Mechanistically, CIRBP promoted PDAC malignancy and chemoresistance via upregulation of dual-specificity tyrosine-Y-phosphorylation regulated kinase 1B (DYRK1B). Indeed, knockdown of CIRBP sensitized pancreatic tumors to gemcitabine treatment by diminishing DYRK1B expression and increasing the ratio of ERK/p38 activity. Our findings suggest that CIRBP overexpression facilitates PDAC progression and gemcitabine resistance by upregulating DYRK1B expression and inhibiting the ERK/p38 signaling pathway, highlighting CIRBP as a potential new therapeutic target for PDAC.

Post-translational down-regulation of Nrf2 and YAP proteins, by targeting deubiquitinases, reduces growth and chemoresistance in pancreatic cancer cells

The intrinsic chemoresistance of pancreatic ductal adenocarcinoma (PDAC) represents the main obstacle in treating this aggressive malignancy. It has been observed that high antioxidant levels and upregulated Nrf2 and the YAP protein expression can be involved in PDAC chemoresistance. The mechanisms of Nrf2 and YAP increase need to be clarified.We chose a panel of PDAC cell lines with diverse sensitivity to cisplatin and gemcitabine. In PANC-1 chemoresistant cells, we found a low level of oxidative stress and high levels of Nrf2 and YAP protein expressions and their respective targets. On the contrary, in CFPAC-1 chemosensitive cells, we found high levels of oxidative stress and low level of these two proteins, as well as their respective targets. In MiaPaCa-2 cells with a middle chemoresistance, we observed intermediate features. When Nrf2 and YAP were inhibited in PANC-1 cells by Ailanthone, a plant extract, we observed a reduction of viability, thus sustaining the role of these two proteins in maintaining the PDAC chemoresistance.We then delved into the mechanisms of the Nrf2 and YAP protein upregulation in chemoresistance, discovering that it was at a post-translational level since the mRNA expressions did not match the protein levels. Treatments of PANC-1 cells with the proteasome inhibitor MG-132 and the protein synthesis inhibitor cycloheximide further confirmed this observation.The expression of DUB3 and OTUD1 deubiquitinases, involved in the control of Nrf2 and YAP protein level, respectively, was also investigated. Both protein expressions were higher in PANC-1 cells, intermediate in MiaPaCa-2 cells, and lower in CFPAC-1 cells. When DUB3 or OTUD1 were silenced, both Nrf2 and YAP expressions were downregulated.Importantly, in deubiquitinase-silenced cells, we observed a great reduction of proliferation and a higher sensitivity to gemcitabine treatment, suggesting that DUB3 and OTUD1 can represent a suitable target to overcome chemoresistance in PDAC cells.

Role of epithelial-mesenchymal transition in chemoresistance in pancreatic ductal adenocarcinoma.

Pancreatic cancer (PC) is the seventh leading cause of cancer death worldwide. The vast majority of patients who have PC develop metastases, resulting in poor treatment effects. Although great progress in therapeutic approaches has been achieved in recent decades, extensive drug resistance still persists, representing a major hurdle to effective anticancer therapy for pancreatic ductal adenocarcinoma (PDAC). Therefore, there is an urgent need to better understand the drug resistance mechanisms and develop novel treatment strategies to improve patient outcomes. Numerous studies suggest that chemoresistance is closely related to epithelial-mesenchymal transition (EMT) of PDAC cells. Thus, this article summarizes the impact of EMT on PDAC from the perspective of chemotherapy resistance and discusses the possible novel applications of EMT inhibition to develop more effective drugs against PDAC.

Overcoming chemoresistance by targeting reprogrammed metabolism: the Achilles' heel of pancreatic ductal adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is one of the leading causes of cancer-related death due to its late diagnosis that removes the opportunity for surgery and metabolic plasticity that leads to resistance to chemotherapy. Metabolic reprogramming related to glucose, lipid, and amino acid metabolism in PDAC not only enables the cancer to thrive and survive under hypovascular, nutrient-poor and hypoxic microenvironments, but also confers chemoresistance, which contributes to the poor prognosis of PDAC. In this review, we systematically elucidate the mechanism of chemotherapy resistance and the relationship of metabolic programming features with resistance to anticancer drugs in PDAC. Targeting the critical enzymes and/or transporters involved in glucose, lipid, and amino acid metabolism may be a promising approach to overcome chemoresistance in PDAC. Consequently, regulating metabolism could be used as a strategy against PDAC and could improve the prognosis of PDAC.

MiRNA-3662 reverses the gemcitabine resistance in pancreatic cancer through regulating the tumor metabolism.

Gemcitabine (Gem) is one of the most commonly used chemotherapeutic drugs in treating patients with pancreatic ductal adenocarcinoma (PDAC). Acquired drug resistance against Gem presents a major clinical challenge in the chemotherapy of PDAC. It has been shown that miRNA-3662 is lowly expressed and implicated with quantities of biological processes in cancer. However, whether miRNA-3662 regulates chemoresistance in PDAC remains largely unknown. The level of miRNA-3662 in PDAC tissues was determined by real-time qPCR (RT-qPCR). Functional experiments were used to investigate the biological role of miRNA-3662 on Gem resistance of PDAC in vitro and in vivo. Fluorescence in situ hybridization (FISH), RT-qPCR, western blotting, bioinformatics analysis and luciferase reporter assay were employed to determine the precise regulation mechanisms. In this study, it was investigated that miRNA-3662 was down-regulated in PDAC clinical samples as well as cell lines. Functional assays revealed that miRNA-3662 was sufficient to inhibit Gem resistance in PDAC cells both in vitro and in vivo. Mechanistically, hypoxia-inducible factor 1ɑ (HIF-1ɑ) was one of the transcriptional target of miRNA-3662 and was up-regulated in PDAC samples. Importantly, genetic promoting of HIF-1ɑ largely compromised miR-3662-mediated chemosensitive effects. In addition, miR-3662 could impair the aerobic glycolysis in PDAC cells. This study sheds light on miRNA-3662 inhibits PDAC cell chemoresistance and aerobic glycolysis through a negative feedback loop with HIF-1ɑ. Therefore, the co-delivery of miR-3662 and Gem could be served as a promising therapeutic regimen for PDAC patients.

Metabolomic profiling of pancreatic adenocarcinoma reveals key features driving clinical outcome and drug resistance

BackgroundAlthough significant advances have been made recently to characterize the biology of pancreatic ductal adenocarcinoma (PDAC), more efforts are needed to improve our understanding and to face challenges related to the aggressiveness, high mortality rate and chemoresistance of this disease. MethodsIn this study, we perform the metabolomics profiling of 77 PDAC patient-derived tumor xenografts (PDTX) to investigate the relationship of metabolic profiles with overall survival (OS) in PDAC patients, tumor phenotypes and resistance to five anticancer drugs (gemcitabine, oxaliplatin, docetaxel, SN-38 and 5-Fluorouracil). FindingsWe identified a metabolic signature that was able to predict the clinical outcome of PDAC patients (p < 0.001, HR=2.68 [95% CI: 1.5–4.9]). The correlation analysis showed that this metabolomic signature was significantly correlated with the PDAC molecular gradient (PAMG) (R = 0.44 and p < 0.001) indicating significant association to the transcriptomic phenotypes of tumors. Resistance score established, based on growth rate inhibition metrics using 35 PDTX-derived primary cells, allowed to identify several metabolites related to drug resistance which was globally accompanied by accumulation of several diacy-phospholipids and decrease in lysophospholipids. Interestingly, targeting glycerophospholipid synthesis improved sensitivity to the three tested cytotoxic drugs indicating that interfering with metabolism could be a promising therapeutic strategy to overcome the challenging resistance of PDAC. InterpretationIn conclusion, this study shows that the metabolomic profile of pancreatic PDTX models is strongly associated to clinical outcome, transcriptomic phenotypes and drug resistance. We also showed that targeting the lipidomic profile could be used in combinatory therapies against chemoresistance in PDAC.

Cancer-associated fibroblasts-mediated ATF4 expression promotes malignancy and gemcitabine resistance in pancreatic cancer via the TGF-\u03b21/SMAD2/3 pathway and ABCC1 transactivation

Cancer-associated fibroblasts (CAFs) contribute to malignant progression and chemoresistance in pancreatic ductal adenocarcinoma (PDAC). However, little is known about the underlying mechanism. In this study, we investigated the potential role and mechanisms of activating transcription factor 4 (ATF4) in CAFs-induced malignancy and gemcitabine resistance. We demonstrated that ATF4 is overexpressed in PDAC and associated with a poor prognosis. Silencing ATF4 expression decreased proliferation, colony formation, migration, gemcitabine sensitivity, and sphere formation. Subsequently, we revealed that CAFs secrete TGF-β1 to upregulate the expression of ATF4 in PDAC cells via the SMAD2/3 pathway and induce cancer progression, cancer stemness, and gemcitabine resistance. Furthermore, we demonstrated that ATF4 directly binds to the ABCC1 promoter region to activate transcription. In summary, these data demonstrate that CAFs contribute to malignancy and gemcitabine resistance in PDAC by upregulating the expression of ATF4 via the TGF-β1/SMAD2/3 axis and highlight that ATF4 is an attractive therapeutic target for combating gemcitabine resistance in PDAC.

An integrated model of N6-methyladenosine regulators to predict tumor aggressiveness and immune evasion in pancreatic cancer.

BackgroundN6-methyladenosine (m6A) is the most abundant mRNA modification. Whether m6A regulators can determine tumor aggressiveness and risk of immune evasion in pancreatic ductal adenocarcinoma (PDAC) remains unknown.MethodsAn integrated model named “m6Ascore” is constructed based on RNA-seq data of m6A regulators in PDAC. Association of m6Ascore and overall survival is validated across several different datasets. Overlaps of m6Ascore and established molecular classifications of PDAC is examined. Immune infiltration, enriched pathways, somatic copy number alterations (SCNAs), mutation profiles and response to immune checkpoint inhibitors are compared between m6Ascore-high and m6Ascore-low tumors.Findingsm6Ascore is associated with dismal overall survival and increased tumor recurrence in PDAC as well as several other solid tumors including colorectal cancer and breast cancer. Basal-like (Squamous) PDAC has higher m6Ascore than that in the classical PDAC. Mechanism study showed m6Ascore-high tumors are characterized with reduced immune infiltration and T cells exhaustion. Meanwhile, m6Ascore is associated with genes regulating cachexia and chemoresistance in PDAC. Furthermore, distinct SCNAs patterns and mutation profiles of KRAS and TP53 are present in m6Ascore-high tumors, indicating immune evasion. m6Ascore-low tumors have higher response rates to immune checkpoint inhibitors (ICIs).InterpretationThese findings indicate m6Ascore can predict aggressiveness and immune evasion in pancreatic cancer. This model has implications for pancreatic cancer prognosis and treatment response to ICIs.FundingThis work was supported in part by National Institutes of Health (NIH) grants to M. Li (R01 CA186338, R01 CA203108, R01 CA247234 and the William and Ella Owens Medical Research Foundation) and NIH/National Cancer Institute Q39 award P30CA225520 to Stephenson Cancer Center.

Stellate Cells Aid Growth-Permissive Metabolic Reprogramming and Promote Gemcitabine Chemoresistance in Pancreatic Cancer.

Simple SummaryThe great majority, more than 90%, of patients with pancreatic ductal adenocarcinoma (PDAC) die within less than five years after detection of the disease, despite recent treatment advances. The poor prognosis is related to late diagnosis, aggressive disease progression, and tumor resistance to conventional chemotherapy. PDAC tumor tissue is characterized by dense fibrosis and poor nutrient availability. A large portion of the tumor is made up of stromal fibroblasts, the pancreatic stellate cells (PSCs), which are known to contribute to tumor progression in several ways. PSCs have been shown to act as an alternate energy source, induce drug resistance, and inhibit drug availability in tumor cells, however, the underlying exact molecular mechanisms remain unknown. In this literature review, we discuss recent available knowledge about the contributions of PSCs to the overall progression of PDAC via changes in tumor metabolism and how this is linked to therapy resistance.Pancreatic ductal adenocarcinoma (PDAC), also known as pancreatic cancer (PC), is characterized by an overall poor prognosis and a five-year survival that is less than 10%. Characteristic features of the tumor are the presence of a prominent desmoplastic stromal response, an altered metabolism, and profound resistance to cancer drugs including gemcitabine, the backbone of PDAC chemotherapy. The pancreatic stellate cells (PSCs) constitute the major cellular component of PDAC stroma. PSCs are essential for extracellular matrix assembly and form a supportive niche for tumor growth. Various cytokines and growth factors induce activation of PSCs through autocrine and paracrine mechanisms, which in turn promote overall tumor growth and metastasis and induce chemoresistance. To maintain growth and survival in the nutrient-poor, hypoxic environment of PDAC, tumor cells fulfill their high energy demands via several unconventional ways, a process generally referred to as metabolic reprogramming. Accumulating evidence indicates that activated PSCs not only contribute to the therapy-resistant phenotype of PDAC but also act as a nutrient supplier for the tumor cells. However, the precise molecular links between metabolic reprogramming and an acquired therapy resistance in PDAC remain elusive. This review highlights recent findings indicating the importance of PSCs in aiding growth-permissive metabolic reprogramming and gemcitabine chemoresistance in PDAC.

S100A14 promotes progression and gemcitabine resistance in pancreatic cancer

S100 calcium binding protein A14 (S100A14) plays an important role in the progression of several types of cancer. However, its roles in pancreatic ductal adenocarcinoma (PDAC) are largely unexplored. Here, we characterized the functional roles of S100A14 in the progression and chemoresistance of PDAC. Gene expression microarray identified that S100A14 was significantly highly expressed in four pairs of human PDAC tumor compared with corresponding non-tumor tissues genes. Quantitative reverse transcription PCR (qRT-PCR), western blotting and immunohistochemical staining (IHC) showed that S100A14 was frequently overexpressed in PDAC cell lines and tissues. Moreover, expression level of S100A14 was positively correlated to advanced cancer stages. Further, Kaplan-Meier survival analysis suggested that PDAC patients with low S100A14 expression had longer overall survival in TCGA PDAC datasets. Transient overexpressing of S100A14 promoted cell proliferation, anchorage-independent colony formation, cell migration and invasion in cell lines with low endogenous S100A14 levels, while transient silencing of S100A14 inhibited cell proliferation, anchorage-independent colony formation, cell migration and invasion in cell lines with high endogenous S100A14 levels. Persistent knockdown of S100A14 by transducing shRNAs carrying lentivirus inhibited subcutaneous tumor formation in nude mice, and sensitized the PDAC cells to gemcitabine treatment. Taken together, S100A14 exhibited oncogenic properties by promoting cell proliferation, transformation, migration and invasion, and enhanced in vivo tumor growth. More importantly, inhibition of S100A14 could effectively abrogate the cancerous properties of the PDAC cells. Our study indicated that S100A14 was a valuable target for the development of therapeutic strategy, as well as a diagnostic and prognosis biomarker for PDAC patients.

Metallothionein-1G suppresses pancreatic cancer cell stemness by limiting activin A secretion via NF-κB inhibition.

Resistance to chemotherapy is a long-standing problem in the management of cancer, and cancer stem cells are regarded as the main source of this resistance. This study aimed to investigate metallothionein (MT)-1G involvement in the regulation of cancer stemness and provide a strategy to overcome chemoresistance in pancreatic ductal adenocarcinoma (PDAC).Methods: MT1G was identified as a critical factor related with gemcitabine resistance in PDAC cells by mRNA microarray. Its effects on PDAC stemness were evaluated through sphere formation and tumorigenicity. LC-MS/MS analysis of conditional medium revealed that activin A, a NF-κB target, was a major protein secreted from gemcitabine resistant PDAC cells. Both loss-of-function and gain-of-function approaches were used to validate that MT1G inhibited NF-κB-activin A pathway. Orthotopic pancreatic tumor model was employed to explore the effects on gemcitabine resistance with recombinant follistatin to block activin A.Results: Downregulation of MT1G due to hypermethylation of its promoter is related with pancreatic cancer stemness. Secretome analysis revealed that activin A, a NF-κB target, was highly secreted by drug resistant cells. It promotes pancreatic cancer stemness in Smad4-dependent or independent manners. Mechanistically, MT1G negatively regulates NF-κB signaling and promotes the degradation of NF-κB p65 subunit by enhancing the expression of E3 ligase TRAF7. Blockade of activin A signaling with follistatin could overcome gemcitabine resistance.Conclusions: MT1G suppresses PDAC stemness by limiting activin A secretion via NF-κB inhibition. The blockade of the activin A signaling with follistatin may provide a promising therapeutic strategy for overcoming gemcitabine resistance in PDAC.

SF3B1 modulators affect key genes in metastasis and drug influx: a new approach to fight pancreatic cancer chemoresistance

Aim: Because mutations of splicing factor 3B subunit-1 (SF3B1) have been identified in 4% of pancreatic ductal adenocarcinoma (PDAC) patients, we investigated the activity of new potential inhibitors of SF3B1 in combination with gemcitabine, one of the standard drugs, in PDAC cell lines. Methods: One imidazo[2,1-b][1,3,4]thiadiazole derivative (IS1) and three indole derivatives (IS2, IS3 and IS4), selected by virtual screening from an in-house library, were evaluated by the sulforhodamine-B and wound healing assay for their cytotoxic and antimigratory activity in the PDAC cells SUIT-2, Hs766t and Panc05.04, the latter harbouring the SF3B1 mutations. The effects on the splicing pattern of proto-oncogene recepteur d’origine nantais (RON) and the gemcitabine transporter human equilibrative nucleoside transporter-1 (hENT1) were assessed by PCR, while the ability to reduce tumour volume was tested in spheroids of primary PDAC cells. Results: The potential SF3B1 modulators inhibited PDAC cell proliferation and prompted induction of cell death. All compounds showed an interesting anti-migratory ability, associated with splicing RON/ΔRON shift in SUIT-2 cells after 24 h exposure. Moreover, IS1 and IS4 potentiated the sensitivity to gemcitabine in both conventional 2D monolayer and 3D spheroid cultures, and these results might be explained by the statistically significant increase in hENT1 expression (P < 0.05 vs. untreated control cells), potentially reversing PDAC chemoresistance. Conclusion: These results support further studies on new SF3B1 inhibitors and the role of RON/hENT1 modulation to develop effective drug combinations against PDAC.

Extracellular Matrix Composition Modulates the Responsiveness of Differentiated and Stem Pancreatic Cancer Cells to Lipophilic Derivate of Gemcitabine.

Background: Pancreatic ductal adenocarcinoma (PDAC) is a highly lethal disease. Gemcitabine (GEM) is used as the gold standard drug in PDAC treatment. However, due to its poor efficacy, it remains urgent to identify novel strategies to overcome resistance issues. In this context, an intense stroma reaction and the presence of cancer stem cells (CSCs) have been shown to influence PDAC aggressiveness, metastatic potential, and chemoresistance. Methods: We used three-dimensional (3D) organotypic cultures grown on an extracellular matrix composed of Matrigel or collagen I to test the effect of the new potential therapeutic prodrug 4-(N)-stearoyl-GEM, called C18GEM. We analyzed C18GEM cytotoxic activity, intracellular uptake, apoptosis, necrosis, and autophagy induction in both Panc1 cell line (P) and their derived CSCs. Results: PDAC CSCs show higher sensitivity to C18GEM treatment when cultured in both two-dimensional (2D) and 3D conditions, especially on collagen I, in comparison to GEM. The intracellular uptake mechanisms of C18GEM are mainly due to membrane nucleoside transporters’ expression and fatty acid translocase CD36 in Panc1 P cells and to clathrin-mediated endocytosis and CD36 in Panc1 CSCs. Furthermore, C18GEM induces an increase in cell death compared to GEM in both cell lines grown on 2D and 3D cultures. Finally, C18GEM stimulated protective autophagy in Panc1 P and CSCs cultured on 3D conditions. Conclusion: We propose C18GEM together with autophagy inhibitors as a valid alternative therapeutic approach in PDAC treatment.