Drug Resistance In Pancreatic Cancer Research Articles

MiR-331-3p Links to Drug Resistance of Pancreatic Cancer Cells by Activating WNT/β-Catenin Signal via ST7L.

Background:Pancreatic cancer is an aggressive type of cancer with poor prognosis, short survival rate, and high mortality. Drug resistance is a major cause of treatment failure in the disease. MiR-331-3p has been reported to play an important role in several cancers. We previously showed that miR-331-3p is upregulated in pancreatic cancer and promotes pancreatic cancer cell proliferation and epithelial-to-mesenchymal transition–mediated metastasis by targeting ST7L. However, it is uncertain whether miR-331-3p is involved in drug resistance.Methods:We investigated the relationship between miR-331-3p and pancreatic cancer drug resistance. As part of this, microRNA mimics or inhibitors were transfected into pancreatic cancer cells. Quantitative polymerase chain reaction was used to detect miR-331-3p expression, and flow cytometry was used to detect cell apoptosis. The Cell Counting Kit-8 assay was used to measure the IC50 values of gemcitabine in pancreatic cancer cells. The expression of multidrug resistance protein 1, multidrug resistance-related protein 1, breast cancer resistance protein, β-Catenin, c-Myc, Cyclin D1, Bcl-2, and Caspase-3 was evaluated by Western blotting.Results:We confirmed that miR-331-3p is upregulated in gemcitabine-treated pancreatic cancer cells and plasma from chemotherapy patients. We also confirmed that miR-331-3p inhibition decreased drug resistance by regulating cell apoptosis and multidrug resistance protein 1, multidrug resistance-related protein 1, and breast cancer resistance protein expression in pancreatic cancer cells, whereas miR-331-3p overexpression had the opposite effect. We further demonstrated that miR-331-3p effects in drug resistance were partially reversed by ST7L overexpression. In addition, overexpression of miR-331-3p activated Wnt/β-catenin signaling in pancreatic cancer cells, and ST7L overexpression restored activation of Wnt/β-catenin signaling.Conclusions:Taken together, our data demonstrate that miR-331-3p contributes to drug resistance by activating Wnt/β-catenin signaling via ST7L in pancreatic cancer cells. These data provide a theoretical basis for new targeted therapies in the future.

Wnt/β-Catenin Signaling: The Culprit in Pancreatic Carcinogenesis and Therapeutic Resistance.

Pancreatic ductal adenocarcinoma (PDAC) is responsible for 7.3% of all cancer deaths. Even though there is a steady increase in patient survival for most cancers over the decades, the patient survival rate for pancreatic cancer remains low with current therapeutic strategies. The Wnt/β-catenin pathway controls the maintenance of somatic stem cells in many tissues and organs and is implicated in pancreatic carcinogenesis by regulating cell cycle progression, apoptosis, epithelial-mesenchymal transition (EMT), angiogenesis, stemness, tumor immune microenvironment, etc. Further, dysregulated Wnt has been shown to cause drug resistance in pancreatic cancer. Although different Wnt antagonists are effective in pancreatic patients, limitations remain that must be overcome to increase the survival benefits associated with this emerging therapy. In this review, we have summarized the role of Wnt signaling in pancreatic cancer and suggested future directions to enhance the survival of pancreatic cancer patients.

Quercetin facilitates cell death and chemosensitivity through RAGE/PI3K/AKT/mTOR axis in human pancreatic cancer cells

The triggering of gemcitabine (GEM) drug resistance in pancreatic cancer by the receptor for advanced glycation end products (RAGE) has been demonstrated. Hence, finding a safe and effective adjuvant for preventing pancreatic cancer progression is imperative. Quercetin is a flavonoid that is abundant in apples, grapes, red raspberry, and onions and has been reported to inhibit RAGE. This research aimed to investigate the mechanisms of quercetin in regulating cell death and enhancing drug effects through RAGE reduction, especially in GEM-resistant pancreatic cancer cells. Our results showed that silencing RAGE expression by RAGE-specific siRNA transfection significantly increased cell death by apoptosis, autophagy and GEM-induced cytotoxicity by suppressing the PI3K/AKT/mTOR axis in MIA Paca-2 and MIA Paca-2 GEMR cells (GEM-resistant cells). Notably, quercetin showed a dramatic effect similar to RAGE silencing that effectively attenuated RAGE expression to facilitate cell cycle arrest, autophagy, apoptosis, and GEM chemosensitivity in MIA Paca-2 GEMR cells, suggesting that an additional reaction occurred under combined quercetin and GEM treatment. In conclusion, the results demonstrated that the molecular mechanisms of quercetin in regulating apoptosis and autophagy-related pathways and increasing GEM chemosensitivity in pancreatic cancer cells involved inhibition of RAGE expression.

Abstract 3628: Comparison of CD44 expression in pancreatic cancer, pancreatic stem and normal pancreatic cells: Development of CLENs for tumor targeting and therapy using cell models

Abstract Purpose: The incidence of drug resistance in pancreatic cancer correlates with poor prognosis, increased disease progression and unfavorable treatment outcomes. CD44, a surface membrane proteoglycan has been identified to be directly involved in pancreatic cancer drug resistance and metastasis. As such, the aim of this research was to screen various pancreatic cells for their expression of CD44, and to begin the early design and development of cell membrane lipid-extracted nanoliposmes (CLENs) for enhanced targeting and therapy. Methods: Three pancreatic cancer cells- Panc-1, MiaPaCa 2, MS1 VEGF and a normal cell line- hTERT- were employed for this study. Stem cell, Mia PaCa-2, served as a positive control for screening purposes. Four liposome compositions were prepared using various ratios of DOPC, Chol, DPPE-PEG5000, and cellular membrane lipid-extracted (LE) was derived from target cell, Panc 1. To evaluate CD44 glycoprotein expression, cells were seeded in 48-well plates and incubated overnight. Fluorescence measurements were determined at excitation wavelength of 485/25 and emission wavelength at 528/20 nm. G44-26 monoclonal antibody was used for CLENs antibody conjugation studies. A Sephadex G-25 column was used for separation of bound and unbound antibody material, and fluorescence detection was used for cell culture analysis. Fluorescence detection was performed at excitation and emission wavelengths of 530/25 and 590/25, respectively. Results: The relative level of CD44 expression for each of the cell lines was as follows. Panc1>Mia PaCa-2 = hTERT>MS1-VEGF. There was a statistically significant difference between Panc-1 and Mia Paca 2 (P<0.001), between hTERT and MS1 VEGF (P<0.05), and between Panc-1 and hTERT (P<0.0001). The inclusion of Panc1-LE in combination with additional liposome components did not increase cellular uptake significantly. The conjugation of G44-26 monoclonal antibody to the liposomes increased average size of all 4 preparations. Average size for CLENs and G44-26 monoclonal antibody -CLENs increased from 140.75 ± 3.22 nm to 250.23 ± 15.31 nm. Conclusion: Future studies are required to demonstrate the effect of G44-26 monoclonal antibody liposome conjugates to further exploit the optimal expression of CD44 for pancreatic drug targeting. Citation Format: Dotun D. Adegunle, Eugene Boakye Ansah, Danny Nguyen, Kanchi Sanghvi, Drew Goodrich, Robert Campbell. Comparison of CD44 expression in pancreatic cancer, pancreatic stem and normal pancreatic cells: Development of CLENs for tumor targeting and therapy using cell models [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 3628.

Oridonin overcomes the gemcitabine resistant PANC-1/Gem cells by regulating GST pi and LRP/1 ERK/JNK signalling.

Background: Chemotherapy remains a primary treatment method for advanced pancreatic cancer. However, chemotherapy resistance can influence the therapeutic effect of pancreatic cancer. The resistance mechanism of chemotherapeutic agents such as gemcitabine, which is an agent typically used to treat pancreatic cancer, is complicated and can be influenced by genes and the environment. Oridonin is a tetracyclic diterpenoid compound extracted from the traditional Chinese herb Rabdosia labtea. Oridonin may overcome drug resistance in pancreatic cancer, but researching pancreatic cancer drug resistance of chemotherapy by oridonin is not completely understood.Purpose: The present study aimed to assess the impact of oridonin on multidrug resistance proteins, apoptosis-associated proteins and energy metabolism in gemcitabine-resistant PANC-1 (PANC-1/Gem) pancreatic cancer cells.Methods: Gemcitabine resistance in PANC-1/Gem cells was induced using a concentration gradient of gemcitabine. Cell Counting Kit-8 assays were used to detect the impact of gemcitabine and oridonin on the proliferation of PANC-1 and PANC-1/Gem cells. Western blot analysis and immunofluorescence were used to detect the expression of multidrug resistance proteins, apoptosis-associated proteins and low-density lipoprotein receptor protein 1 (LRP1) proteins in PANC-1/Gem cells. The effects of gemcitabine and oridonin on PANC-1/Gem cells apoptosis were detected using flow cytometry. Animal xenograft tumor assays were used to detect the effect of gemcitabine and oridonin on pancreatic cancer in vivo. Furthermore, the ATP Assay kit was used to determine the effects of gemcitabine and oridonin on ATP levels in PANC-1/Gem cells. Immunofluorescence assays were used to detect the effects of gemcitabine and oridonin on the expression of low-density lipoprotein receptor protein 1 (LRP1) in PANC-1/Gem cells. In addition, LRP1 expression was knocked down in PANC-1/Gem cells via lentiviral vector-mediated RNA silencing. Clone formation assays and Western blot analysis were used to detect the effect of LRP1 knockdown on the proliferation of PANC-1/Gem cells.Results: The present results demonstrate that oridonin overcomes PANC-1/Gem cells gemcitabine reistance by regulating GST pi and LRP1/ERK/JNK signaling.Conclusion: In conclusion, the present study indicated that oridonin could overcome gemcitabine resistance in PANC-1/Gem cells by regulating GST pi and LRP1/ ERK/JNK signaling, inducing cell apoptosis. Therefore, oridonin with gemcitabine may be a promising preoperative treatment for patients who suffer from pancreatic cancer.

Abstract 1283: Targeting c-MYC and MAPK pathway to overcome pancreatic cancer drug resistance

Abstract Background: Acquired resistance to systemic chemotherapy is the main complication in pancreatic ductal adenocarcinoma (PDAC) treatment. Although there are studies focused on gemcitabine resistance mechanisms, our understanding of the mechanisms of nab-paclitaxel (n-PTX) treatment failure remains extremely limited. To enhance the use of properly powered patient-derived platforms, we adopted the conditionally reprogrammed (CR) cell culture technique in order to develop both parental and nab-PTX-resistant cells. The CR approach allowed us to identify the critical role of c-MYC and ERK in the PDAC drug response. Small molecule activators of PP2A (SMAPS) have showed activity in inhibiting lung KRAS-mutant tumor growth. We used SMAPS as new therapeutic agents in PDAC, for its ability to alter c-MYC activity through PP2A dysregulation and enhance PDAC sensitivity to n-PTX. Methods: Long-term cultures of PDAC CRs were established from treatment-naive PDAC patients’ biopsies, and used to generate drug-resistant cells. Zebrafish and mouse model were used to test the cells’ ability to form tumors and to verify the drug resistance in vivo. Molecular analyses were used to characterize the drug-resistant cells and to identify key pathways involved in the drug resistance evolution. Genomic and chemical alterations of the key proteins were used to confirm the involvement in the drug resistance mechanism. We regulated the expression of c-MYC and ERK using SMAPS as a new targeting agent and trametenib to verify the direct correlation between c-MYC and ERK and the drug resistance mechanism. Results: Using the credentialed KRAS-mutant CR cultures, we generated n-PTX-resistant cell lines. The parental and nab-PTX resistant cells were subjected to subcutaneous injections in nude mice, and formed tumors in 2-3 weeks. Histological evaluation showed that the CRs self-assembled into ductal structures, surrounded by a desmoplastic stromal microenvironment that faithfully recapitulates human PDAC. Resistant profiles were verified both in mouse and Zebrafish model. RNA microarrays identified a sustained induction of a pro-inflammatory pathway leading to c-MYC overexpression. c-MYC silencing and overexpression confirmed the role of c-MYCin the evolution of nab-PTX resistance. Treatment of the resistant CRs with either trametenib or with SMAPS resulted in enhanced sensitivity to nab-PTX. We furtherverified that the enhanced sensitivity was commensurate with a reduction in p-Erk and c-Myc. Conclusion: The CR methodology addresses the need for a reliable method for generating primary cell lines on a single patient basis. The ability to rapidly model in vitro, and verify in vivo, that the overexpression of c-MYC contributes to the development of n-PTX resistance is a significant advancement in the field. Our data showed that SMAPs or trametanib overcome a significant component of the n-PTX resistance providing new hope for refractory PDAC. Citation Format: Erika Maria Parasido, George S. Avetian, Jonathan Brody, Jordan Winter, Eric Londin, Michael Pishvaian, Eric Glasgow, Stephen Byers, Goutham Narla, Christopher Albanese. Targeting c-MYC and MAPK pathway to overcome pancreatic cancer drug resistance [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 1283.

Abstract 3621: Targeted hyaluronic acid nanoparticles improve treatment response in pancreatic cancer

Abstract Pancreatic cancer has poor response to chemotherapy with desmoplastic stroma identified as a delivery barrier. We have developed a nanoparticle (NP) drug system using an engineered ligand of receptor binding region of urokinase plasminogen activator (ATF) and the catalytic domain of metalloprotease (MMP14) carrying SN38 (CPT-11 analog). Hyaluronic acid (HA), a naturally occurring protein was chosen as the backbone to produce a biocompatible NP. HA can also bind to CD44, highly expressed in epithelial cancers. Dual uPAR, CD44 targeting allows for targeted delivery and facilitates receptor mediated endocytosis of NP-drug complex. MMP14 activity degrades extracellular matrix to deplete the stromal barrier. SN38 was encapsulated and recombinant ATFmmp14 conjugated to surface of self-assembled hyaluronic acid spheres (200nm) to form the complete particle (HANP). Cytotoxicity assays were conducted to determine IC50 in cell lines. Patient derived xenograft (PDX) model of a drug resistant pancreatic cancer was used for efficacy studies. Orthotopically implanted tumors were treated with HANP (10mg/kg SN38 dosage) weekly for 6 weeks via tail vein and overall survival compared to conventional therapy. HANP in-vitro cytotoxicity was greater than conventional Irinotecan (>80x) and liposomal irinotecan (>900x) in a PDX derived cell line (Table 1). In-vivo efficacy study demonstrated a significant improvement in survival of PDX bearing mice with HANP (n=9) (median survival 50 days), compared to FOLFIRINOX (n=9) and Gemcitabine-nab-Paclitaxel (n=9) (median survival 37 days each) and no treatment (n=13) (median survival 22 days). Combination of HANP with a modified-FOLFIRINOX regimen (n=9) led to median survival >72 days (p<0.001). HANP alone or in combination with a modified FOLFIRINOX led to significantly improved survival in a pancreatic cancer PDX model. Further studies are underway to evaluate preclinical PD/PK for eventual translation as targeted therapy for pancreatic cancer. In-vitro cytoxicity assay for IC50 determinationDrugIC50 (uM)HANP0.01SN-380.012Irinotecan0.8Liposomal Irinotecan9 Citation Format: Mohammad Raheel Jajja, Lei Zhu, Dazhi Wang, Charles A. Staley, Bassel El-Rayes, David A. Kooby, Lily Yang. Targeted hyaluronic acid nanoparticles improve treatment response in pancreatic cancer [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 3621.

A randomized clinical trial of chemotherapy with gemcitabine/cisplatin/nabpaclitaxel with or without the AXL inhibitor bemcentinib (BGB324) for patients with advanced pancreatic cancer.

TPS473 Background: The Axl pathway coordinately mediates immune evasion and drug resistance in pancreatic cancer. Systemic Axl inhibition can enhance the efficacy of cancer therapy by blocking tumor cell proliferation, survival and drug resistance associated with epithelial-mesenchymal transition (EMT), and targeting innate immune suppression in the tumor microenvironment. Bemcentinib (BGB324) is a first in class, selective oral inhibitor of Axl. Our group has shown that bemcentinib therapy, in combination with gemcitabine, improved survival in multiple preclinical models of pancreatic cancer. Methods: This is a multicenter, randomized, phase 1b/2 clinical trial of nab-paclitaxel/gemcitabine/cisplatin with or without bemcentinib. Patients with metastatic pancreatic cancer, good performance status and preserved liver, kidney and hematologic function are eligible. The treatment schedule is as follows: Bemcentinib 100 or 200 mg daily, nab-paclitaxel 125 mg/m2, gemcitabine 1000 mg/m2 and cisplatin 25 mg/m2 intravenously on D1, 8 every 21 days. 3 -12 patients will be recruited in part 1 following a modified 3+3 dose finding scheme. Part 2 of the study is a 1:1 randomized phase 2 design enrolling 62 patients. The primary objective is to determine complete response rate. Secondary end points are overall response rate, PFS and adverse events. A parallel biomarker study will accompany the trial analyzing blood and tissue samples to determine the effect of chemotherapy and bemcentinib on 1) Axl pathway activity in tumor tissue, 2) changes in immune landscape including upregulation of immune cytokines, and immune cell infiltration into the tumor, 3) apoptosis and decreased proliferation of tumor and 4) to identify predictive biomarkers of response. Clinical trial information: NCT03649321.

Increased miR-142 and decreased DJ-1 enhance the sensitivity of pancreatic cancer cell to adriamycin.

Phosphatidylinositol-3 kinase (PI3K)/protein kinase B (AKT) signaling pathway is related to tumorigenesis by up-regulating survivin. Phosphatase and tensin homologue deleted on chromosome ten (PTEN) can suppress PI3K/AKT signaling pathway, while DJ-1 is the negative regulator of PTEN. DJ-1 up-regulation is closely correlated with the occurrence, progression, and drug resistance of pancreatic cancer. MicroRNA-142 (MiR-142) is significantly declined in pancreatic cancer tissue. Bioinformatics analysis demonstrated that complementary binding site exists between miR-142 and DJ-1. This investigation, therefore, aimed to study the role of miR-142 in the regulation of DJ-1-PTEN/PI3K/AKT/Survivin signaling pathway as well as in pancreatic cancer cell proliferation, apoptosis, and adriamycin (ADM) resistance. Dual luciferase assay was performed to assess the targeted relationship between miR-142 and DJ-1. MiR-142, DJ-1, and PTEN expressions in SW1990 cells and drug-resistant SW1990/ADM cells were compared. SW1990/ADM cells were divided into five groups, including mimic-NC, miR-142 mimic, small interfere normal control (si-NC), si-DJ-1, and miR-142 mimic + si-DJ-1 groups. DJ-1, PTEN, phosphorylated-AKT (p-AKT), and Survivin expressions were tested. Cell apoptosis was determined by flow cytometry. Cell proliferation was evaluated by EdU staining. MiR-142 targeted inhibited DJ-1 expression. MiR-142, PTEN, and cell apoptosis significantly down-regulated, while DJ-1, p-AKT, Survivin, and cell proliferation significantly elevated in SW1990/ADM cells compared with SW1990 cells. MiR-142 mimics and/or si-DJ-1 transfection markedly reduced DJ-1, p-AKT, and Survivin expressions enhanced PTEN level, attenuated cell proliferation, enhanced cell apoptosis, and weakened ADM resistance. MiR-142 over-expression weakened ADM resistance in pancreatic cancer cells by targeting DJ-1 to enhance PTEN expression and attenuate PI3K/AKT signaling pathway activity.

The role of GLI-SOX2 signaling axis for gemcitabine resistance in pancreatic cancer

Pancreatic cancer, mostly pancreatic ductal adenocarcinomas (PDAC), is one of the most lethal cancers, with a dismal median survival around 8 months. PDAC is notoriously resistant to chemotherapy. Thus far, numerous attempts using novel targeted therapies and immunotherapies yielded limited clinical benefits for pancreatic cancer patients. It is hoped that delineating the molecular mechanisms underlying drug resistance in pancreatic cancer may provide novel therapeutic options. Using acquired gemcitabine resistant pancreatic cell lines, we revealed an important role of the GLI-SOX2 signaling axis for regulation of gemcitabine sensitivity in vitro and in animal models. Down-regulation of GLI transcriptional factors (GLI1 or GLI2), but not SMO signaling inhibition, reduces tumor sphere formation, a characteristics of tumor initiating cell (TIC). Down-regulation of GLI transcription factors also decreased expression of TIC marker CD24. Similarly, high SOX2 expression is associated with gemcitabine resistance whereas down-regulation of SOX2 sensitizes pancreatic cancer cells to gemcitabine treatment. We further revealed that elevated SOX2 expression is associated with an increase in GLI1 or GLI2 expression. Our ChIP assay revealed that GLI proteins are associated with a putative Gli binding site within the SOX2 promoter, suggesting a more direct regulation of SOX2 by GLI transcription factors. The relevance of our findings to human disease was revealed in human cancer specimens. We found that high SOX2 protein expression is associated with frequent tumor relapse and poor survival in stage II PDAC patients (all of them underwent gemcitabine treatment), indicating that reduced SOX2 expression or down-regulation of GLI transcription factors may be effective in sensitizing pancreatic cancer cells to gemcitabine treatment.

An ABCG2 non-substrate anticancer agent FL118 targets drug-resistant cancer stem-like cells and overcomes treatment resistance of human pancreatic cancer

BackgroundPancreatic cancer is a deadly disease with a very low 5-year patient survival rate of 6–8%. The major challenges of eliminating pancreatic cancer are treatment resistance and stromal barriers to optimal drug access within the tumor. Therefore, effective molecular targeting drugs with high intra-tumor access and retention are urgently needed for managing this devastating disease in the clinic.MethodsThis study has used the following in vitro and in vivo techniques for the investigation of exceptional anticancer drug FL118’s efficacy in treatment of resistant pancreatic cancer: cell culture; immunoblotting analysis to test protein expression; DNA sub-G1 flow cytometry analyses to test cell death; MTT assay to test cell viability; pancreatic cancer stem cell assays (fluorescence microscopy tracing; matrigel assay; CD44-positive cell colony formation assay); human luciferase-labeled pancreatic tumor orthotopic animal model in vivo imaging; pancreatic cancer patient-derived xenograft (PDX) animal models; and toxicology studies with immune-competent BALB/cj mice and beagle dogs.ResultsOur studies found that FL118 alone preferentially killed cisplatin-resistant cancer cells, while a combination of FL118 with cisplatin synergistically killed resistant pancreatic cancer cells and reduced spheroid formation of treatment-resistant pancreatic cancer stem-like cells. Furthermore, using in vivo-imaging, we found that FL118 in combination with cisplatin strongly inhibited both drug-resistant pancreatic xenograft tumor growth and metastasis. In PDX model, we demonstrated that FL118 alone effectively eliminated PDX tumors, while FL118 in combination with gemcitabine eliminated PDX tumors that showed relative resistance (less sensitivity) to treatment with FL118. These FL118 efficacy results are consistent with our molecular-targeting data showing that FL118 inhibited the expression of multiple antiapoptotic proteins (survivin, Mcl-1, XIAP, cIAP2) and ERCC6, a critical regulator of DNA repair, in treatment-resistant pancreatic stem-like cancer cells. Furthermore, FL118 toxicity studies in BALB/cj mice and beagle dogs indicated that FL118 exhibits favorable hematopoietic and biochemical toxicities.ConclusionTogether, our studies suggest that FL118 is a promising anticancer drug for further clinical development to effectively treat drug-resistant pancreatic cancer alone or in combination with other pancreatic cancer chemotherapeutic drugs.

Identification of lncRNAs and Their Functional Network Associated with Chemoresistance in SW1990/GZ Pancreatic Cancer Cells by RNA Sequencing.

Drug resistance is a major problem in the treatment of pancreatic cancer (PC). Long noncoding (lnc)RNAs modulate a variety of cellular processes. This study was carried out to identify lncRNAs that are differentially expressed in drug-resistant PC by next-generation RNA sequencing. We identified 205 differentially expressed lncRNAs (DELs) and 847 differentially expressed mRNAs (DEMs) in a comparison of gemcitabine-resistant and -sensitive SW1990 human PC cells. The expression levels of 12 randomly selected lncRNAs were confirmed by quantitative real-time PCR. A total of 55 and 99 DEMs were predicted to be targeted by the DELs through cis and trans mechanisms, respectively. The DEMs were significantly enriched in the Gene Ontology terms cell part, binding, and cellular processes, and a Kyoto Encyclopedia of Genes and Genomes analysis showed that they were associated with metabolic pathway, pathways in cancer, insulin resistance, microRNAs in cancer, and phosphotidylinositol 3-kinase-Akt signaling pathway. A coexpression network revealed a hub comprising lncRNAs (MIR210HG, SNHG1, and LOC729970) and mRNAs (RAB3D, DDX17, and SPNS2) that presumably mediate drug resistance in PC. The identified lncRNAs can serve as diagnostic or prognostic biomarkers and therapeutic targets for PC.

Micro-RNA-21 Regulates Cancer-Associated Fibroblast-Mediated Drug Resistance in Pancreatic Cancer.

Pancreatic ductal adenocarcinoma (PDAC) is one of the leading causes of cancer deaths due to its highly aggressive biological nature and resistance to chemotherapy. Previous studies indicate that miR-21 is an important regulator in the activation of cancer-associated fibroblasts (CAFs). However, whether miR-21 in CAFs would regulate PDAC’s tumor microenvironment and lead to drug resistance remain unknown. In this study, we evaluated the relationship between CAF activation, miR-21 expression, and drug resistance using tumor samples from PDAC patients. We changed the miR-21 expression level in CAFs and tested its roles in regulating the function of CAFs. In addition, we explored the roles of miR-21 in CAFs in the development of PDAC using an animal model. We found that PDAC patients who were resistant to gemcitabine treatment tended to have higher miR-21 expression and more activated CAFs. An in vitro study showed that CAFs with high miR-21 expression had elevated MMP-3, MMP-9, PDGF, and CCL-7 expression and promoted the invasion of PDAC cell lines. miR-21 overexpression also contributed to the activation of CAFs by regulating the PDCD4 gene. The in vivo study showed that upregulating miR-21 in CAFs promoted PDAC desmoplasia and increased its drug resistance to gemcitabine treatment, but downregulating miR-21 in CAFs suppressed desmoplasia and enhanced the effect of gemcitabine. We concluded that miR-21 promoted the activation of CAFs and contributed to the drug resistance of PDAC.

Abstract 2697: Alterations in the HSP70 interactome are induced by overexpression of MUC1 in pancreatic cancer

Abstract Heat Shock Protein 70 (HSP70) is an evolutionarily well-conserved molecular chaperone involved in several cellular processes such as folding of proteins, modulating protein-protein interactions and transport of proteins across membranes. HSP70 works synergistically with a series of chaperones, co-chaperones, adaptors, and folding enzymes known as the HSP70 “interactome”. Previous work has demonstrated that HSP70 interactome changes can be promoted by overexpression of oncoproteins. An important “client” of HSP70 is the oncoprotein MUC1. MUC1 is overexpressed and promotes drug resistance in pancreatic cancer. We applied quantitative affinity-purification mass spectrometry (AP-MS) to characterize the HSP70 interactome differences between PC3/NEO and PC3/MUC1 cell lines. 439 HSP70-interacting proteins were identified and quantified via LC-MS/MS. Interestingly while 53% of HSP70 interactions remained constant upon MUC1 overexpression, 29% significantly increased and 18% decreased. Many of the altered proteins were key co-chaperones that regulate and maintain HSP70 activity. High levels of MUC1 promoted HSP70 association with 15 co-chaperones (HSPA5, DNAJB1, STIP1, TXNDC5, HSPB1, CCT2, TCP1, P4HB, CCT6A, CCT8, PDIA3 and HSP90B). Conversely, MUC1 overexpression triggered the loss of HSP70 interaction with only 2 co-chaperones (PPIB and DNAJA2). We are currently validating these altered interactions using an array of biochemical techniques. Our hope is that by understanding these HSP70 interactome changes, we may be able to uncover novel ways to inhibit MUC1 function in pancreatic cancer. Citation Format: . Nitika, Donald J. Wolfgeher, Pinku Mukherjee, Andrew W. Truman. Alterations in the HSP70 interactome are induced by overexpression of MUC1 in pancreatic cancer [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2018; 2018 Apr 14-18; Chicago, IL. Philadelphia (PA): AACR; Cancer Res 2018;78(13 Suppl):Abstract nr 2697.

Microarray analysis of circular RNA expression profiles associated with gemcitabine resistance in pancreatic cancer cells

Pancreatic cancer (PC) is one of the most malignant tumors of the digestive system due to its rapid progression, metastasis and resistance to chemotherapy. Gemcitabine (GEM) chemotherapy is the first‑choice treatment for advanced PC. However, the effect of GEM‑based chemotherapy on PC is limited due to the development of chemoresistance, and the molecular mechanisms underlying this resistance have yet to be investigated. Circular RNAs (circRNAs), which can function as microRNA sponges, have been found to be involved in the development of several types of cancer. However, research on circRNAs in PC drug resistance is limited. In the present study, the GEM‑resistant PC cell line, SWl990/GZ, was successfully established by treating parental SWl990 cells invitro with increasing concentrations of GEM in culture medium intermittently for 10months. By analyzing the expression profiles of circRNAs in microarray between SWl990/GZ and parental SW1990 cells, we identified 26upregulated and 55downregulated circRNAs (fold change≥2 and P<0.05) among 12,866 detected circRNAs in SWl990/GZ compared with SW1990 cells. Furthermore, the changes in the expression of six representative circRNAs was validated by reverse transcription‑quantitative PCR. In addition, Kyoto Encyclopedia of Genes and Genomes pathway analysis and Gene Ontology analysis were performed. These analyses revealed that the dysregulated circRNAs regulated several cancer‑related pathways, such as the mitogen‑activated protein kinase (MAPK) and mammalian target of rapamycin (mTOR) signaling pathways, and may be involved in the biological process of the regulation of chemoresistance, including nucleic acid metabolic process and cellular response to stress. The present study undertook a comprehensive expression analysis and revealed the functional profiles of differentially expressed circRNAs associated with GEM‑resistance in PC, thereby indicating the possible participation of these dysregulated circRNAs in the development of chemoresistance and providing novel potential therapeutic targets for PC.

Resveratrol-Induced Downregulation of NAF-1 Enhances the Sensitivity of Pancreatic Cancer Cells to Gemcitabine via the ROS/Nrf2 Signaling Pathways.

NAF-1 (nutrient-deprivation autophagy factor-1), which is an outer mitochondrial membrane protein, is known to play important roles in calcium metabolism, antiapoptosis, and antiautophagy. Resveratrol, a natural polyphenolic compound, is considered as a potent anticancer agent. Nevertheless, the molecular mechanisms underlying the effects of resveratrol and NAF-1 and their mediation of drug resistance in pancreatic cancer remain unclear. Here, we demonstrate that resveratrol suppresses the expression of NAF-1 in pancreatic cancer cells by inducing cellular reactive oxygen species (ROS) accumulation and activating Nrf2 signaling. In addition, the knockdown of NAF-1 activates apoptosis and impedes the proliferation of pancreatic cancer cells. More importantly, the targeting of NAF-1 by resveratrol can improve the sensitivity of pancreatic cancer cells to gemcitabine. These results highlight the significance of strategies that target NAF-1, which may enhance the efficacy of gemcitabine in pancreatic cancer therapy.

Abstract A074: Killing late-stage cancer cells by coupling DNA damage induction with MAPK pathway activation

Abstract Acquired resistance of metastatic melanoma (MM) tumors to BRAF V600E inhibitors (BRAFis) is commonplace in the clinic. Habitual relapse of patients contributes to &amp;lt;20% 5-year survival rates in MM. We previously identified serine synthesis as a critical mechanism of late-stage cancer cell resistance to BRAFis. Pretreatment with DNA-damaging agent and nucleoside analog gemcitabine resensitized drug-resistant cancer cells to BRAFis dabrafenib (Tafinlar) and vemurafenib (Zelboraf). The combination treatments were effective against BRAF wild type cancer cells, potentially expanding the clinical reach of BRAFis. In this study, we identify two additional DNA-damaging agents that resensitized drug-resistant MM and pancreatic cancer cells to BRAFis dabrafenib and encorafenib. We identify the antifolate and DNA-damaging agent methotrexate (MTX), and the topoisomerase 1 inhibitor and DNA-damaging agent camptothecin (Camp) as sensitizers of late-stage, drug-resistant cancer cells to BRAFis dabrafenib and encorafenib (LGX818). Importantly, we show a novel positive correlation between DNA repair delay and and allosteric MAPK activation via BRAFi treatment in BRAF wild type cells. Cells arrest in G1/S following DNA-damaging agent + BRAFi combination treatments and ultimately, cell death is triggered via apoptosis, presumably due to DNA damage repair delays. Additionally, we have identified activating KRAS codon 12 mutations as critical to the efficacy of combination treatments. In summary, we have elucidated a novel method of cell death induction via DNA-damaging agent + BRAFi combination treatments that has the potential to significantly impact late-stage MM and pancreatic cancer therapy in the clinic. Citation Format: Vikram Bhattacharjee. Killing late-stage cancer cells by coupling DNA damage induction with MAPK pathway activation [abstract]. In: Proceedings of the AACR-NCI-EORTC International Conference: Molecular Targets and Cancer Therapeutics; 2017 Oct 26-30; Philadelphia, PA. Philadelphia (PA): AACR; Mol Cancer Ther 2018;17(1 Suppl):Abstract nr A074.

Downregulation of MicroRNA-455-3p Links to Proliferation and Drug Resistance of Pancreatic Cancer Cells via Targeting TAZ.

Drug resistance is a major cause of treatment failure in pancreatic cancer. The limited evidence indicates the involvement of miR-455-3p in chemotherapy resistance of cancer. Here we observed by qPCR that miR-455-3p was significantly decreased in pancreatic cancer tissues and cell lines. We then confirmed that the inhibition of miR-455-3p increased cell proliferation and gemcitabine resistance of pancreatic cancer, whereas forced overexpression of miR-455-3p had the opposite effect. Furthermore, we demonstrated that TAZ, which is associated with drug resistance of pancreatic cancer, is a new direct downstream target of miR-455-3p. Our present study suggests that miR-455-3p contributes to cell proliferation and drug resistance in pancreatic cancer cells via targeting TAZ.

TLR2 and TLR4 predict favourable prognosis in early pancreatic cancer

Pancreatic ductal adenocarcinoma (PDAC) is one of the most common causes of cancer-related death all over the world. This disease is difficult to treat and patients have an overall 5-year survival rate of less than 5%. Although two drugs, gemcitabine (GEM) and 5-fluorouracil (5-FU) have been shown to improve the survival rate of patients systematically, they do not increase general survival to a clinically acceptable degree. Lack of ideal clinical response of pancreatic cancer patients to chemotherapy is likely to be due to intrinsic and acquired chemoresistance of tumor cells. Various mechanisms of drug resistance have been investigated in pancreatic cancer, including genetic and epigenetic changes in particular genes or signaling pathways. In addition, evidence suggests that microRNAs (miRNAs) play significant roles as key regulators of gene expression in many cellular processes, including drug resistance. Understanding underlying genes and mechanisms of drug resistance in pancreatic cancer is critical to develop new effective treatments for this deadly disease. This review illustrates the genes and miRNAs involved in resistance to gemcitabine in pancreatic cancer.

Epithelial-specific E3 ligase CBLc regulates differentiation and motility of pancreatic cancer cells

Pancreatic ductal adenocarcinoma (PDAC) is one of the most common causes of cancer-related death all over the world. This disease is difficult to treat and patients have an overall 5-year survival rate of less than 5%. Although two drugs, gemcitabine (GEM) and 5-fluorouracil (5-FU) have been shown to improve the survival rate of patients systematically, they do not increase general survival to a clinically acceptable degree. Lack of ideal clinical response of pancreatic cancer patients to chemotherapy is likely to be due to intrinsic and acquired chemoresistance of tumor cells. Various mechanisms of drug resistance have been investigated in pancreatic cancer, including genetic and epigenetic changes in particular genes or signaling pathways. In addition, evidence suggests that microRNAs (miRNAs) play significant roles as key regulators of gene expression in many cellular processes, including drug resistance. Understanding underlying genes and mechanisms of drug resistance in pancreatic cancer is critical to develop new effective treatments for this deadly disease. This review illustrates the genes and miRNAs involved in resistance to gemcitabine in pancreatic cancer.