Therapeutic Approach For Pancreatic Ductal Adenocarcinoma Research Articles

Enhancement of in-vivo cellular uptake of Coenzyme Q10 using saponin derivatives in rTALAP transgenic mice model

Pancreatic cancer remains a significant contributor to cancer-related mortality, with pancreatic ductal adenocarcinoma (PDAC) being particularly challenging to treat. Coenzyme Q10 (CoQ10), a hydrophobic antioxidant crucial for cellular energy production, holds promise in PDAC therapy. However, its limited solubility hinders efficient cellular uptake. To overcome this limitation, we developed micelle formulations incorporating CoQ10 and assessed their potential to enhance cellular delivery. In this particular study, we have utilized some of the most common saponins such as Quillaja saponin, Ginsenoside R0, and Ginsenoside Rb1 as a drug carrier in order to enhance the bioavailability and cellular uptake of CoQ10. Micelles' size and shape were characterized using DLS, TEM, and LC-MS/MS. These all saponin micelles showed better encapsulation, zeta potential, and smaller size compared to Pluronic F127 micelles. Moreover, these formulations induced a notable increase in reactive oxygen species (ROS) generation, indicative of potential apoptotic activity. Further investigations revealed that micelle treatments led to modulation of gene expression related to epithelial-mesenchymal transition (EMT) markers, with an increase in E-cadherin expression and a decrease in claudin, snail, slug, and vimentin. Additionally, in-vivo screening studies in transgenic mice demonstrated promising results as anticancer by reducing apoptosis. In conclusion, our findings suggest that CoQ10-loaded micelles, particularly those incorporating Quillaja saponin and ginsenoside derivatives, hold potential as a novel therapeutic approach for PDAC by enhancing cellular uptake, inducing ROS-mediated apoptosis, and modulating EMT markers. These findings contribute to advancing our understanding of CoQ10's role in pancreatic cancer therapy.

IRAK2-NF-κB signaling promotes glycolysis-dependent tumor growth in pancreatic cancer.

Metabolic reprogramming has emerged as a core hallmark of cancer, and cancer metabolism has long been equated with aerobic glycolysis. Moreover, hypoxia and the hypovascular tumor microenvironment (TME) are major hallmarks of pancreatic ductal adenocarcinoma (PDAC), in which glycolysis is imperative for tumor cell survival and proliferation. Here, we explored the impact of interleukin 1 receptor-associated kinase 2 (IRAK2) on the biological behavior of PDAC and investigated the underlying mechanism. The expression pattern and clinical relevance of IRAK2 was determined in GEO, TCGA and Ren Ji datasets. Loss-of-function and gain-of-function studies were employed to investigate the cellular functions of IRAK2 in vitro and in vivo. Gene set enrichment analysis, Seahorse metabolic analysis, immunohistochemistry and Western blot were applied to reveal the underlying molecular mechanisms. We found that IRAK2 is highly expressed in PDAC patient samples and is related to a poor prognosis. IRAK2 knockdown led to a significant impairment of PDAC cell proliferation via an aberrant Warburg effect. Opposite results were obtained after exogenous IRAK2 overexpression. Mechanistically, we found that IRAK2 is critical for sustaining the activation of transcription factors such as those of the nuclear factor-κB (NF-κB) family, which have increasingly been recognized as crucial players in many steps of cancer initiation and progression. Treatment with maslinic acid (MA), a NF-κB inhibitor, markedly attenuated the aberrant oncological behavior of PDAC cells caused by IRAK2 overexpression. Our data reveal a role of IRAK2 in PDAC metabolic reprogramming. In addition, we obtained novel insights into how immune-related pathways affect PDAC progression and suggest that targeting IRAK2 may serve as a novel therapeutic approach for PDAC.

Abstract 4264: Lysosome inhibition sensitizes pancreatic cancer to replication stress by aspartate depletion

Abstract Objective: Functional lysosomes are required for autophagy and macropinocytosis, the intra- and extracellular scavenging pathways cancer cells engage for nutrient acquisition. Pancreatic ductal adenocarcinoma (PDAC) tumors exhibit high basal lysosomal activity, and genetic or pharmacologic inhibition of lysosome function suppresses PDAC cell proliferation and tumor growth. However, the codependencies induced by lysosomal inhibition in PDAC have not been systematically explored. We hypothesized that identification and targeting the lysosomal inhibition-induced codependencies in PDAC cells would be an effective therapeutic strategy. Methods: A comprehensive pharmacological inhibition screen of the protein kinome was performed to identified lysosomal inhibition-induced codependency. LC-MS/MS-MRM and LC-MS methods were used to examine nucleotide biosynthesis and amino acid levels, respectively, to understand the mechanism of the identified codependency. A broad panel of PDAC cell lines, primary PDAC culture models, two- and three-dimentional culture models, a PDAC cell/stroma spheroid coculture model, and xenograft and syngeneic PDAC animal models were used to evaluate the treatment efficacy. Results: We found that replication stress response (RSR) inhibitors were synthetically lethal with chloroquine (CQ) and other lysosomal inhibitors in PDAC cells. CQ treatment reduced de novo nucleotide biosynthesis and induced replication stress. We found that CQ treatment caused mitochondrial dysfunction and depletion of aspartate, an essential precursor for de novo nucleotide synthesis, as an underlying mechanism. Supplementation with aspartate in PDAC cell culture and overexpression of the aspartate transporter SLC1A3 in xenograft PDAC tumors partially rescued the phenotypes induced by CQ. The synergy of CQ and the RSR inhibitor VE-822 was comprehensively validated in a broad panel of PDAC cell lines and primary PDAC cultures, in two- and three-dimentional PDAC cultures, in heterotypic spheroid culture with cancer-associated fibroblasts, and in in vivo xenograft and syngeneic PDAC mouse models. Conclusion: We discovered a codependency on functional lysosomes and an intact RSR pathway in PDAC, and developed the combination of CQ and RSR inhibitors as a translational therapeutic approach for PDAC. Citation Format: Shili Xu, Irmina A. Elliott, Amanda M. Dann, Stephanie S. Kim, Evan R. Abe, Woosuk Kim, Soumya Poddar, Alexandra Moore, Lei Zhou, Jennifer L. Williams, Joseph R. Capri, Razmik Ghukasyan, Cynthia Matsumura, D. Andrew Tucker, Wesley R. Armstrong, Anthony E. Cabebe, Nanping Wu, Luyi Li, Thuc M. Le, Caius G. Radu, Timothy R. Donahue. Lysosome inhibition sensitizes pancreatic cancer to replication stress by aspartate depletion [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2019; 2019 Mar 29-Apr 3; Atlanta, GA. Philadelphia (PA): AACR; Cancer Res 2019;79(13 Suppl):Abstract nr 4264.

Inhibition of endoplasmic-reticulum-stress-mediated autophagy enhances the effectiveness of chemotherapeutics on pancreatic cancer

BackgroundEndoplasmic reticulum (ER) stress and its consequent unfolded protein response (UPR) are believed to be associated with progression, survival and chemoresistance of a variety of tumor cells through multiple cellular processes, including autophagy. Therefore, the ER stress-autophagy pathway presents a potential molecular target for therapeutic intervention. The objective of this study was to evaluate the therapeutic efficacy of ER stress and autophagy modulators in the context of pancreatic ductal adenocarcinoma (PDAC).MethodsWe first targeted IRE1α, an important regulator of the UPR, through STF-083010 treatment in PDAC cell lines in vitro. Chloroquine was then used to target autophagy and an optimal combination treatment was developed using chloroquine, sunitinib and gemcitabine. Apoptosis was analyzed using TUNEL assay, autophagy was estimated using lysotracker staining and electron microscopy, and UPR was analyzed using anti-GRP78 immunostaining and XBP1 splicing. Transplantation of PDAC derived KPCP1 and Panc02 cells in mouse pancreas were performed to study treatment efficacy in vivo.ResultsSuppression of the IRE1α by STF-083010 alone resulted in increased lysosomes and reduced viability of PDAC cells. Chloroquine treatment alone inhibited downstream autophagy but was insufficient in reducing PDAC cell growth. However, combining STF-083010 and chloroquine had additive anti-tumor efficacy when used with gemcitabine. Sunitinib alone caused abnormal maturation of the autolysosomes with increased intracellular multivesicular bodies and increased apoptosis evident in PDAC cells. Sunitinib showed a synergistic effect with chloroquine in reducing in vitro PDAC cell viability and significantly increased the efficacy of gemcitabine in human and murine PDAC cell lines. The anti-proliferative effect of gemcitabine was significantly increased when used in combination with sunitinib and/or chloroquine in both in vitro and in vivo PDAC models. The addition of sunitinib and/or chloroquine to gemcitabine, resulted in a significantly increased survival of the animals without noticeably increased toxicity. Sunitinib, gemcitabine and chloroquine treated mice showed a significant reduction of GRP78 expression, reduced cell proliferation and increased apoptosis in pancreas, compatible with a tumor response.ConclusionsSunitinib combined with chloroquine reduces tumor growth through suppression of autophagy and increased apoptosis. Co-administration of modulators of ER stress-mediated autophagy with chemotherapy presents a novel therapeutic approach in PDAC.

Abstract 1135: DRD2 is critical for pancreatic cancer and promises pharmacological therapy by already established antagonists

Abstract Introduction and aims: Although the overall five-year survival of all patients with cancer stands at 63%, for pancreatic cancer patients, it is a disheartening 8% - a number that remains largely unchanged for three decades. Of the patients diagnosed with pancreatic cancer, about 85% exhibit pancreatic ductal adenocarcinoma (PDAC). Most of these patients die within 4 to 6 months after diagnosis. The poor prognosis is caused by the detection at only late stages, and lack of effective options for chemotherapy. The widely used chemotherapeutic agent gemcitabine, confers a median survival advantage of only 6 months, and resistance to therapy develops in the vast majority of patients. Given this poor prognosis of patients with PDAC, there is an urgent need to find more effective therapies. Experimental procedures: Microarrays were used to perform global gene expression profiling in 195 PDAC and 41 normal pancreatic tissue samples. Using these profiling data, we undertook an extensive analysis of PDAC transcriptome by superimposing the pathway context and interaction networks of aberrantly expressed genes to identify factors with central roles in PDAC pathways. Next, tissue microarray analysis (TMA) were used to verify the expression of the candidate target in independent set of 152 samples comprising 40 normal pancreatic tissues, 49 chronic pancreatitis sections (CP) and 63 PDAC samples. We further validated the functional relevance of the candidate molecule through RNA interference (RNAi) and pharmacological inhibition in vitro and in vivo. Results: We identified dopamine receptor D2 (DRD2) as a key modulator of cancer pathways in PDAC. DRD2 up-regulation at the protein level was validated in a large independent sample cohort. Most importantly, we found that blockade of DRD2, through RNAi or pharmacological inhibition using FDA-approved antagonists hampers the proliferative and invasive capacities of pancreatic cancer cells while modulating cAMP and endoplasmic reticulum stress pathways. Also, we observed a potent effect of DRD2 antagonists on inhibition of cancer cell proliferation using different model of primary and metastatic tumor cells derived from spontaneous pancreatic cancer mouse models and patient-derived pancreatic adenocarcinoma mouse xenograft (PDX) models. Conclusions: Our findings demonstrate that inhibiting DRD2 represents a novel therapeutic approach for PDAC. Since DRD2 inhibitors are already in the clinic for the management of schizophrenia, our results from this study could support a drug repurposing strategy to expedite clinical evaluation of these agents as novel therapy against pancreatic cancer. Citation Format: Pouria Jandaghi, Hamed S. Najafabadi, Andrea Bauer, Andreas I. Papadakis, Matteo Fassan, Anita Hall, Anie Monast, Maryam Safisamghabadi, Magnus von Knebel Doeberitz, John P. Neoptolemos, Eithne Costello, William Greenhalf, Aldo Scarpa, Bence Sipos, Daniel Auld, Mark Lathrop, Morag Park, Markus W. Büchler, Oliver Strobel, Thilo Hackert, Nathalia Giese, George Zogopoulos, Veena Sangwan, Sidong Huang, Jörg D. Hoheisel, Yaser Riazalhosseini. DRD2 is critical for pancreatic cancer and promises pharmacological therapy by already established antagonists [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 1135. doi:10.1158/1538-7445.AM2017-1135

The Development of a Novel Therapeutic Strategy to Target Hyaluronan in the Extracellular Matrix of Pancreatic Ductal Adenocarcinoma.

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal diseases to affect humans, regardless of whether patients receive multimodal therapy (including surgery, radiotherapy, and chemotherapy). This resistance to intervention is currently considered to be caused by the desmoplastic change of the extracellular matrix (ECM) in PDAC tissues, which is characterized by the accumulation of cancer-associated fibroblasts, collagen, proteoglycan, and hyaluronan. Among these ECM components, hyaluronan has attracted interest because various studies have indicated that hyaluronan-rich PDAC is correlated with the progressive properties of cancer cells, both in experimental and clinical settings. Hence, the reduction of hyaluronan in cancer tissue may represent a novel therapeutic approach for PDAC. 4-methylumbelliferone (4-MU) is a derivative of coumarin that was reported to suppress the synthesis of hyaluronan in cultured human skin fibroblasts in 1995. As an additional study, our group firstly reported that 4-MU reduced the hyaluronan synthesis of mouse melanoma cells and exerted anti-cancer activity. Subsequently, we have showed that 4-MU inhibited liver metastasis in mice inoculated with human pancreatic cancer cells. Thereafter, 4-MU has been accepted as an effective agent for hyaluronan research and is expected to have clinical applications. This review provides an overview of the interaction between PDAC and hyaluronan, the properties of 4-MU as a suppressor of the synthesis of hyaluronan, and the perspectives of PDAC treatment targeting hyaluronan.

Abstract B66: BET inhibition remodels tumor stroma and suppresses progression of human pancreatic cancer

Abstract Background and Aims: Pancreatic ductal adenocarcinoma (PDAC) is well characterized by dense fibrotic stroma with abundant cancer-associated fibroblasts (CAFs). As CAFs are activated during tumorigenesis and acquire tumor-promoting properties, activated CAFs have been implicated in PDAC progression; however, the precise mechanisms of their activation remain largely unknown. The bromodomain and extraterminal (BET) domain proteins are epigenetic reader proteins that recognize acetylated amino acid residues on histone tails and facilitate gene transcription. Recent studies have demonstrated therapeutic efficacy of BET inhibitors on various cancers including PDAC, mainly through suppression of c-myc transcription; however, how BET inhibitors suppress PDAC growth and their effects on CAFs remains largely unknown. Using patient-derived tumor xenografts (PDX) and primary CAFs, we investigated the therapeutic efficacy and dissected the underlying mechanisms of a BET inhibitor, JQ1, on human PDAC and CAFs. Methods: We established PDX lines and primary CAFs from surgically resected human PDAC specimen. For in vivo analyses, mice bearing subcutaneous tumor were treated with vehicle or JQ1. For in vitro analyses, patient-derived PDAC cells and CAFs were treated with vehicle or JQ1 and analyzed separately. To explore the pro-tumorigenic role of secretion from CAFs, PDAC cells were cultured with conditioned medium (CM) that was collected from DMSO- or JQ1- treated CAFs. Chromatin immunoprecipitation (ChIP) assay was performed to assess the binding of transcription factors and histone modifications which are associated with altered gene expression in CAFs by JQ1 treatment. Results: In vivo experiments revealed that volumes and weights of subcutaneous PDX tumors were significantly smaller in JQ1-treated mice than vehicle-treated mice. Unexpectedly, however, JQ1 exerted only minimal effects to the proliferation of PDAC cells that were isolated from PDX tumors and cultured in vitro, suggesting the involvement of cell-extrinsic mechanisms in the JQ1-mediated suppression of tumor growth in vivo. Of note, histopathological analysis of PDX tumors revealed that JQ1 treatment dramatically ameliorated desmoplastic change, with reduction in extracellular matrix (ECM) deposition and α-SMA expressing CAFs. As α-SMA expression and ECM production is a hallmark of activated CAFs, we hypothesized that JQ1 might inactivate CAFs, thereby reducing their tumor-promoting properties. To test this hypothesis, qPCR was performed to analyze gene expression in primary CAFs cultured in vitro and also in stromal cells in PDX tumors in vivo. As expectedly, JQ1 suppressed the expression of genes implicated in the properties of activated CAF, including ECM, cytokines and growth factors both in vitro and in vivo. Furthermore, when PDAC cells were cultured with CM from DMSO–treated CAFs, proliferation of PDAC cells were promoted along with activation of MAPK, AKT, and STAT3 pathways, which was abrogated when cultured with CM from JQ1-treated CAFs. Consistently, immunoblotting and immunohistochemistry of PDX tumors demonstrated that JQ1 reduced phosphorylation of ERK, AKT, and STAT3 in PDAC cells in vivo. Mechanistically, we found that JQ1 suppressed hedgehog and TGF-β/SMAD3 pathways, both of which play central roles in CAF activation, through disruption of BRD4 recruitment to the promoter regions of their target genes. Conclusions: BET proteins are critical regulators of CAF-activation in PDAC. Inactivation of CAFs by BET inhibition offers a novel therapeutic approach for PDAC. Citation Format: Keisuke Yamamoto, Keisuke Tateishi, Yotaro Kudo, Mayumi Hoshikawa, Mariko Tanaka, Takuma Nakatsuka, Hiroaki Fujiwara, Koji Miyabayashi, Ryota Takahashi, Yasuo Tanaka, Hideaki Ijichi, Yousuke Nakai, Hiroyuki Isayama, Yasuyuki Morishita, Taku Aoki, Yoshihiro Sakamoto, Kiyoshi Hasegawa, Norihiro Kokudo, Masashi Fukayama, Kazuhiko Koike.{Authors}. BET inhibition remodels tumor stroma and suppresses progression of human pancreatic cancer. [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer: Advances in Science and Clinical Care; 2016 May 12-15; Orlando, FL. Philadelphia (PA): AACR; Cancer Res 2016;76(24 Suppl):Abstract nr B66.

Necroptosis DAMPens anti-tumor immunity.

Necroptosis DAMPens anti-tumor immunity.

IL1 Receptor Antagonist Inhibits Pancreatic Cancer Growth by Abrogating NF-κB Activation

Constitutive NF-κB activation is identified in about 70% of pancreatic ductal adenocarcinoma (PDAC) cases and is required for oncogenic KRAS-induced PDAC development in mouse models. We sought to determine whether targeting IL-1α pathway would inhibit NF-κB activity and thus suppress PDAC cell growth. We determined whether anakinra, a human IL-1 receptor (rhIL-1R) antagonist, inhibited NF-κB activation. Assays for cell proliferation, migration, and invasion were performed with rhIL-1R antagonist using the human PDAC cell lines AsPc1, Colo357, MiaPaCa-2, and HPNE/K-ras(G12V)/p16sh. In vivo NF-κB activation-dependent tumorigenesis was assayed using an orthotopic nude mouse model (n = 20, 5 per group) treated with a combination of gemcitabine and rhIL-1RA. rhIL-1R antagonist treatment led to a significant decrease in NF-κB activity. PDAC cells treated with rhIL-1R antagonist plus gemcitabine reduced proliferation, migration, and invasion as compared with single gemcitabine treatment. In nude mice, rhIL-1R antagonist plus gemcitabine significantly reduced the tumor burden (gemcitabine plus rhIL-1RA vs. control, P = 0.014). We found that anakinra, an FDA-approved drug that inhibits IL-1 receptor (IL-1R), when given with or without gemcitabine, can reduce tumor growth by inhibiting IL1α-induced NF-κB activity; this result suggests that it is a useful therapeutic approach for PDAC.

GPCRs as potential therapeutic targets in pancreatic cancer‐associated fibroblasts (1066.11)

GPCRs as potential therapeutic targets in pancreatic cancer‐associated fibroblasts Shu Zhou1, Thalia McCann1, Randall French2, Andrew M. Lowy2,3, Paul A. Insel1,4 1Department of Pharmacology, 2Moores Cancer Center, 3Division of Surgical Oncology, Department of Surgery, 4Department of Medicine, UCSD, La Jolla, California Pancreatic ductal adenocarcinoma (PDAC) is characterized by a dense fibrotic stromal matrix, composed of activated fibroblasts/stellate cells, inflammatory cells and other cell types. This unique tumor microenvironment has been increasingly recognized as a key mediator of PDAC progression and drug resistance. Thus, targeting the tumor stroma may be a novel therapeutic approach for PDAC. We hypothesized that G‐protein‐coupled receptors (GPCRs) expressed by pancreatic cancer‐associated fibroblasts (CAFs) may be potential therapeutic targets for PDAC. To begin to test this hypothesis, we used an unbiased GPCRomic array approach to identify and quantify the GPCRs expressed by pancreatic CAFs. We found 125 GPCRs, whose expression was shared in CAFs obtained from the primary tumors of five patients. Among those GPCRs, 35 had at least two‐fold higher expression in the CAFs compared to normal pancreatic fibroblasts and included GPCRs that link to each of the major classes of G proteins. RT‐PCR analysis of the 10 most up‐regulated GPCRs correlated well (r2=0.88) with the data from the GPCR array. We conclude that: 1) GPCRomic array analysis can identify and quantify the profile of GPCRs expressed by pancreatic CAFs; 2) the CAF‐expressed profile differs from that of normal pancreatic fibroblasts and 3) GPCRs expressed by CAFs may have functional roles and be novel therapeutic targets for PDAC.Grant Funding Source: Supported by Cancer Therapeutics Training Program

Enhancement of the Antiproliferative Activity of Gemcitabine by Modulation of c-Met Pathway in Pancreatic Cancer

Pancreatic-ductal-adenocarcinoma (PDAC) is amongst the most lethal malignancies, mainly because of its metastatic spread and multifactorial chemoresistance. Since c-Met is a marker of pancreatic-cancer-stem-cells (CSC), playing a key role in metastasis and chemoresistance, this study evaluated the therapeutic potential of the novel c-Met/ALK inhibitor crizotinib against PDAC cells, including the Capan-1-gemcitabine-resistant cells (Capan-1-R). Crizotinib inhibited PDAC cell-growth with IC50 of 1.5 μM in Capan-1-R, and synergistically enhanced the antiproliferative and proapoptotic activity of gemcitabine, as detected by sulforhodamine-B-assay, flow cytometry and combination-index method. Capan-1-R had higher expression of the CSC markers CD44+/CD133+/CD326+, but their combined expression was significantly reduced by crizotinib, as detected by quantitative-RT-PCR and FACS-analysis. Similarly, Capan-1-R cells had significantly higher protein-expression of c-Met (≈2-fold), and increased migratory activity, which was reduced by crizotinib (e.g., > 50% reduction of cell-migration in Capan-1-R after 8-hour exposure, compared to untreated-cells), in association with reduced vimentin expression. Capan-1-R had also significantly higher mRNA expression of the gemcitabine catabolism-enzyme CDA, potentially explaining the higher CDA activity and statistically significant lower levels of gemcitabine-nucleotides in Capan-1-R compared to Capan-1, as detected by Liquid-chromatography-massspectrometry. Conversely, crizotinib significantly reduced CDA expression in both Capan-1 and Capan-1-R cells. In aggregate, these data show the ability of crizotinib to specifically target CSC-like-subpopulations, interfere with cell-proliferation, induce apoptosis, reduce migration and synergistically interact with gemcitabine, supporting further studies on this novel therapeutic approach for PDAC.

Enhancement of the Antiproliferative Activity of Gemcitabine by Modulation of c-Met Pathway in Pancreatic Cancer

Pancreatic-ductal-adenocarcinoma (PDAC) is amongst the most lethal malignancies, mainly because of its metastatic spread and multifactorial chemoresistance. Since c-Met is a marker of pancreatic-cancer-stem-cells (CSC), playing a key role in metastasis and chemoresistance, this study evaluated the therapeutic potential of the novel c-Met/ALK inhibitor crizotinib against PDAC cells, including the Capan-1-gemcitabine-resistant cells (Capan-1-R). Crizotinib inhibited PDAC cell-growth with IC50 of 1.5 μM in Capan-1-R, and synergistically enhanced the antiproliferative and proapoptotic activity of gemcitabine, as detected by sulforhodamine-B-assay, flow cytometry and combination-index method. Capan-1-R had higher expression of the CSC markers CD44+/CD133+/CD326+, but their combined expression was significantly reduced by crizotinib, as detected by quantitative-RT-PCR and FACS-analysis. Similarly, Capan-1-R cells had significantly higher protein-expression of c-Met (≈2-fold), and increased migratory activity, which was reduced by crizotinib (e.g., >50% reduction of cell-migration in Capan-1-R after 8-hour exposure, compared to untreated-cells), in association with reduced vimentin expression. Capan-1-R had also significantly higher mRNA expression of the gemcitabine catabolism-enzyme CDA, potentially explaining the higher CDA activity and statistically significant lower levels of gemcitabine-nucleotides in Capan-1-R compared to Capan-1, as detected by Liquid-chromatography-massspectrometry. Conversely, crizotinib significantly reduced CDA expression in both Capan-1 and Capan-1-R cells. In aggregate, these data show the ability of crizotinib to specifically target CSC-like-subpopulations, interfere with cell-proliferation, induce apoptosis, reduce migration and synergistically interact with gemcitabine, supporting further studies on this novel therapeutic approach for PDAC. Keywords: Pancreatic ductal adenocarcinoma, c-Met pathway, cancer stem cells, crizotinib, gemcitabine, Capan-1-gemcitabine-resistant cells (Capan-1-R), sulforhodamine-B-assay, flow cytometry, combination-index method, cell-growth