Chemoresistance Of Pancreatic Cancer Cells Research Articles

MSLN induced EMT, cancer stem cell traits and chemotherapy resistance of pancreatic cancer cells

Chemoresistance is one of the main reasons for poor prognosis of pancreatic cancer. The effects of mesothelin (MSLN) on chemoresistance in pancreatic cancer are still unclear. We aim to investigate potential roles of MSLN in chemoresistance and its relationship with proliferation, epithelial-mesenchymal transition (EMT) and cancer stemness of pancreatic cancer cells. Human pancreatic cancer cell lines ASPC-1 and Mia PaCa-2 with high and low expression of MSLN, respectively, were selected. The ASPC-1 with MSLN knockout (KO) and Mia PaCa-2 of MSLN overexpression (OE) were generated. The effects of MSLN on cell phenotypes, expression of EMT-related markers, clone formation, tumor sphere formation, and pathologic role of MSLN in tumorigenesis were detected. Sensitivity of tumor cells to gemcitabine was evaluated. The results showed that adhesion, proliferation, migration and invasion were decreased significantly in ASPC-1 with MSLN KO, whereas increased significantly in Mia PaCa-2 with MSLN OE. The size and the number of clones and tumor spheres were decreased in ASPC-1 with MSLN KO, and increased in Mia PaCa-2 with MSLN OE. In xenograft model, tumor volume was decreased (tumor grew slower) in MSLN KO group compared to control group, while increased in MSLN OE group. Mia PaCa-2 with MSLN OE had a higher IC50 of gemcitabine, while ASPC-1 with MSLN KO had a lower IC50. We concluded that MSLN could induce chemoresistance by enhancing migration, invasion, EMT and cancer stem cell traits of pancreatic cancer cells. Targeting MSLN could represent a promising therapeutic strategy for reversing EMT and chemoresistance in pancreatic cancer cells.

Synergistic Anticancer Activity of Plumbagin and Xanthohumol Combination on Pancreatic Cancer Models.

Among diverse cancers, pancreatic cancer is one of the most aggressive types due to inadequate diagnostic options and treatments available. Therefore, there is a necessity to use combination chemotherapy options to overcome the chemoresistance of pancreatic cancer cells. Plumbagin and xanthohumol, natural compounds isolated from the Plumbaginaceae family and Humulus lupulus, respectively, have been used to treat various cancers. In this study, we investigated the anticancer effects of a combination of plumbagin and xanthohumol on pancreatic cancer models, as well as the underlying mechanism. We have screened in vitro numerous plant-derived extracts and compounds and tested in vivo the most effective combination, plumbagin and xanthohumol, using a transgenic model of pancreatic cancer KPC (KrasLSL.G12D/+; p53R172H/+; PdxCretg/+). A significant synergistic anticancer activity of plumbagin and xanthohumol combinations on different pancreatic cancer cell lines was found. The combination treatment of plumbagin and xanthohumol influences the levels of B-cell lymphoma (BCL2), which are known to be associated with apoptosis in both cell lysates and tissues. More importantly, the survival of a transgenic mouse model of pancreatic cancer KPC treated with a combination of plumbagin and xanthohumol was significantly increased, and the effect on BCL2 levels has been confirmed. These results provide a foundation for a potential new treatment for pancreatic cancer based on plumbagin and xanthohumol combinations.

Evaluation of the Antitumor Effects of Platinum-Based [Pt(η1-C2H4-OR)(DMSO)(phen)]+ (R = Me, Et) Cationic Organometallic Complexes on Chemoresistant Pancreatic Cancer Cell Lines.

Pancreatic cancer is one of the most lethal malignancies with an increasing incidence and a high mortality rate, due to its rapid progression, invasiveness, and resistance to anticancer therapies. In this work, we evaluated the antiproliferative and antimigratory activities of the two organometallic compounds, [Pt(η1-C2H4-OMe)(DMSO)(phen)]Cl (1) and [Pt(η1-C2H4-OEt)(DMSO)(phen)]Cl (2), on three human pancreatic ductal adenocarcinoma cell lines with different sensitivity to cisplatin (Mia PaCa-2, PANC-1, and YAPC). The two cationic analogues showed superimposable antiproliferative effects on the tested cells, without significant differences depending on alkyl chain length (Me or Et). On the other hand, they demonstrated to be more effective than cisplatin, especially on YAPC cancer cells. For the interesting cytotoxic activity observed on YAPC, further biological assays were performed, on this cancer cell line, to evaluate the apoptotic and antimetastatic properties of the considered platinum compounds (1 and 2). The cytotoxicity of 1 and 2 compounds appeared to be related to their intracellular accumulation, which was much faster than that of cisplatin. Both 1 and 2 compounds significantly induced apoptosis and cell cycle arrest, with a high accumulation of sub-G1 phase cells, compared to cisplatin. Moreover, phenanthroline-containing complexes caused a rapid loss of mitochondria membrane potential, ΔΨM, if compared to cisplatin, probably due to their cationic and lipophilic properties. On 3D tumor spheroids, 1 and 2 significantly reduced migrated area more than cisplatin, confirming an antimetastatic ability.

STOML2 restricts mitophagy and increases chemosensitivity in pancreatic cancer through stabilizing PARL-induced PINK1 degradation

Pancreatic cancer remains one of the most lethal diseases with a relatively low 5-year survival rate, and gemcitabine-based chemoresistance occurs constantly. Mitochondria, as the power factory in cancer cells, are involved in the process of chemoresistance. The dynamic balance of mitochondria is under the control of mitophagy. Stomatin-like protein 2 (STOML2) is located in the mitochondrial inner membrane and is highly expressed in cancer cells. In this study, using a tissue microarray (TMA), we found that high STOML2 expression was correlated with higher survival of patients with pancreatic cancer. Meanwhile, the proliferation and chemoresistance of pancreatic cancer cells could be retarded by STOML2. In addition, we found that STOML2 was positively related to mitochondrial mass and negatively related to mitophagy in pancreatic cancer cells. STOML2 stabilized PARL and further prevented gemcitabine-induced PINK1-dependent mitophagy. We also generated subcutaneous xenografts to verify the enhancement of gemcitabine therapy induced by STOML2. These findings suggested that STOML2 regulated the mitophagy process through the PARL/PINK1 pathway, thereby reducing the chemoresistance of pancreatic cancer. STOML2-overexpression targeted therapy might be helpful for gemcitabine sensitization in the future.

Targeting chemoresistant senescent pancreatic cancer cells improves conventional treatment efficacy

Pancreatic cancer is one of the deadliest cancers owing to its late diagnosis and of the strong resistance to available treatments. Despite a better understanding of the disease in the last two decades, no significant improvement in patient care has been made. Senescent cells are characterized by a stable proliferation arrest and some resistance to cell death. Increasing evidence suggests that multiple lines of antitumor therapy can induce a senescent-like phenotype in cancer cells, which may participate in treatment resistance. In this study, we describe that gemcitabine, a clinically-used drug against pancreatic cancer, induces a senescent-like phenotype in highly chemoresistant pancreatic cancer cells in vitro and in xenografted tumors in vivo. The use of ABT-263, a well-described senolytic compound targeting Bcl2 anti-apoptotic proteins, killed pancreatic gemcitabine-treated senescent-like cancer cells in vitro. In vivo, the combination of gemcitabine and ABT-263 decreased tumor growth, whereas their individual administration had no effect. Together these data highlight the possibility of improving the efficacy of conventional chemotherapies against pancreatic cancer by eliminating senescent-like cancer cells through senolytic intervention. Further studies testing different senolytics or their combination with available treatments will be necessary to optimize preclinical data in mouse models before transferring these findings to clinical trials.

New insights into the role of adipocytes in pancreatic cancer progression: paving the way towards novel therapeutic targets.

Pancreatic cancer (PC) remains one of the most lethal malignancies across the world, which is due to delayed diagnosis and resistance to current therapies. The interactions between pancreatic tumor cells and their tumor microenvironment (TME) allow cancer cells to escape from anti-cancer therapies, leading to difficulties in treating PC. With endocrine function and lipid storage capacity, adipose tissue can maintain energy homeostasis. Direct or indirect interaction between adipocytes and PC cells leads to adipocyte dysfunction characterized by morphological change, fat loss, abnormal adipokine secretion, and fibroblast-like transformation. Various adipokines released from dysfunctional adipocytes have been reported to promote proliferation, invasion, metastasis, stemness, and chemoresistance of PC cells via different mechanisms. Additional lipid outflow from adipocytes can be taken into the TME and thus alter the metabolism in PC cells and surrounding stromal cells. Besides, the trans-differentiation potential enables adipocytes to turn into various cell types, which may give rise to an inflammatory response as well as extracellular matrix reorganization to modulate tumor burden. Understanding the molecular basis behind the protumor functions of adipocytes in PC may offer new therapeutic targets.

Hypoxia-elicited Exosomes Promote the Chemoresistance of Pancreatic Cancer Cells by Transferring LncROR via Hippo Signaling.

Recent studies have found that hypoxia contributes to tumor progression and drug resistance by inducing the secretion of exosomes. However, the mechanism underlying this resistance in pancreatic cancer remains to be explored. In this study, we investigated the effect of hypoxia-induced tumor-derived exosomes (Hexo) on stemness and resistance to gemcitabine in pancreatic cancer cells, as well as the molecular mechanisms involved in this process. Firstly, we discovered that hypoxia promoted stemness and induced resistance to gemcitabine in pancreatic cancer cells. Secondly, we showed that exosomes secreted by pancreatic cancer cells under normoxic or hypoxic conditions can be transfected into tumor cells. Thirdly, it was demonstrated that Hexo promotes proliferation, stemness, and resistance to gemcitabine in pancreatic cancer cells, as well as inhibits the apoptosis and cell cycle arrest induced by gemcitabine. Finally, it was verified that Hexo inactivated the Hippo/Yes-associated protein (Hippo/YAP) pathway in pancreatic cancer cells by transferring exosomal long non-coding RNA regulator of reprogramming (lncROR). In summary, the hypoxic tumor microenvironment could promote stemness and induce resistance to gemcitabine in pancreatic cancer cells. Mechanistically, Hexo enhanced stemness to promote chemoresistance in pancreatic cancer cells by transferring lncROR via Hippo signaling. Thus, exosomal lncROR may serve as a candidate target of chemotherapy for pancreatic cancer.

TYRO3 promotes chemoresistance via increased LC3 expression in pancreatic cancer

Pancreatic cancer (PC) is an aggressive malignancy with few treatment options, and improved treatment strategies are urgently required. TYRO3, a member of the TAM receptor tyrosine kinase family, is a known oncogene; however, the relationship between TYRO3 expression and PC chemoresistance remains to be elucidated. We performed gain- and loss-of-function experiments on TYRO3 to examine whether it is involved in chemoresistance in PC cells. TYRO3 knockdown decreased cell viability and enhanced apoptosis following treatment of PC cells with gemcitabine and 5-fluorouracil (5-FU). In contrast, no such effects were observed in TYRO3-overexpressing PC cells. It is known that autophagy is associated with cancer chemoresistance. We then examined effects of TYRO3 on autophagy in PC cells. TYRO3 overexpression increased LC3 mRNA levels and induced LC3 puncta in PC cells. Inhibition of autophagy by chloroquine mitigated cell resistance to gemcitabine and 5-FU. In a xenograft mouse model, TYRO3 silencing significantly increased sensitivity of the cells to gemcitabine and 5-FU. To further investigate the involvement of autophagy in patients with PC, we immunohistochemically analyzed LC3 expression in the tissues of patients who underwent pancreatectomy and compared it with disease prognosis and TYRO3 expression. LC3 expression was negatively and positively correlated with prognosis and TYRO3 expression, respectively. Furthermore, LC3- and TYRO3-positive patients had a significantly worse prognosis among patients with PC who received chemotherapy after recurrence. These results indicated that the TYRO3-autophagy signaling pathway confers PC resistance to gemcitabine and 5-FU, and could be a novel therapeutic target to resolve PC chemoresistance.

Abstract B048: Long-term moderate hyperthermia induces apoptotic signaling and reduces chemoresistance in pancreatic cancer cells in vitro

Abstract In recent years, hyperthermia (HT) is gaining popularity as a therapeutic modality that could potentiate other treatments such as chemotherapy. Although pre-clinical and clinical research do suggest a beneficial effect of moderate HT (41.5°C), more knowledge on the molecular mechanisms is required before it can be routinely implemented in the clinic. In brief, several commercially available PDAC cell lines were subjected exposed to increasing durations of HT at 41.5°C, ranging from for 6, 12, and to 24 hours. Following HT, the effects on apoptotic signaling and chemotherapy sensitivity were evaluated mainly via RT-qPCR quantification of relevant genes. We found that subjecting pancreatic ductal adenocarcinoma (PDAC) cells (BxPC-3, Capan-2) to 41.5 °C for increasing durations, leads to (1) a decrease in cell viability, (2) a decreased mRNA expression of BCL-2 and three genes associated with chemotherapy resistance RRM2, TS, ERCC1, together with (3) an increase in expression of the pro-apoptotic BAX. These effects also appear to be cell-line dependent. More specifically, exposure of BxPC-3 cells to 6, 12 and 24 h of HT led to an increase in BAX/BCL-2 ratio of respectively 1.35, 2.80 and 4.65, whereas for Capan-2 cells a distinct pattern was observed. Here, the ratio of BAX/BCL-2 changed to 0.84, 1.56 and 3.95 for the three durations. Moreover, we confirmed the increased susceptibility towards the chemotherapeutic drugs 5-fluorouracil, gemcitabine, and cisplatin by subjecting the cells to varying doses of the drugs in absence or presence of HT. In general, exposure of 5 different PDAC lines (BxPC-3, PANC-1, Capan-1, Capan-2, MIA PaCa-2) cells to 12 h of HT allowed a dose reduction of 50%, and exposure of 24 h allowed a dose reduction of 75-95% to achieve a similar cell-killing effect as that which was observed at 37 °C. Together, these results show that subjecting the exposure of PDAC cells to long durations of moderate HT induces pro-apoptotic signaling and increases their susceptibility towards commonly used chemotherapeutic drugs, and that this effect is time-dependent. Nevertheless, treatment durations of 12 and 24 h ours would not be feasible very challenging in the clinic. Therefore, future research we will attempt to uncover whether an identical or similar affect can be achieved when the total HT exposure is fractionated into several shorter intervals, as is done with radiotherapy. Lastly, it is unlikely that the effects of HT are limited to those discussed. Future research will also focus on the role of cellular stress by HT-induced reactive oxygen species and non-necrotic, non-apoptotic cell death, such as ferroptosis, in pancreatic cancer cells. Citation Format: Robin Colenbier, Costanza E. Maurici, Ivana Gorbaslieva, Eke van Zwol, Jean-Pierre Timmermans, Elisa Giovannetti, Johannes Bogers. Long-term moderate hyperthermia induces apoptotic signaling and reduces chemoresistance in pancreatic cancer cells in vitro [abstract]. In: Proceedings of the AACR Special Conference on Pancreatic Cancer; 2022 Sep 13-16; Boston, MA. Philadelphia (PA): AACR; Cancer Res 2022;82(22 Suppl):Abstract nr B048.

AHR and ELAVL1 gene interaction influence on chemoresistance of pancreatic cancer cells, in vitro

AHR and ELAVL1 gene interaction influence on chemoresistance of pancreatic cancer cells, in vitro

Evaluation of the toxicity of Caralluma europaea (C.E) extracts and their effects on apoptosis and chemoresistance in pancreatic cancer cells

Pancreatic adenocarcinoma is a disease with no effective treatment. Chemo-resistance contributes to the dismal prognosis for patients diagnosed with the disease. This study aims to evaluate the toxicity and the effect of Caralluma europaea (C.E) extracts on cancer cell survival, apoptosis, chemo-resistance, and pro-cancer pathways, in pancreatic cancer. The acute and subacute toxicities of C.E extracts were evaluated. The cytotoxic effect on pancreatic cancer cell survival and apoptosis was determined by MTT assay and DNA fragmentation. The expression of cancer stemness markers was measured using Western blot. A molecular docking was used to test the possible effects of C.E compounds in inhibiting the Hedgehog and activating caspase-3. The hydroethanolic extract’s DL50 was over 5000 mg/kg. During the subacute toxicity, only saponins extract showed some hepatic toxicity signs. Cells treated with C.E extracts combined with gemcitabine revealed an additive anti-survival activity. C.E extracts sensitized resistant MIA-PaCa-2 to gemcitabine treatment. Most of the C.E extracts downregulated the expression of cancer stemness-associated genes. Luteolin-7-O-glucoside presented the highest docking Gscore on human Smoothened. Isorhamnetin-3-O-rutinoside induced apoptosis via activation of caspase-3. C.E extracts can be considered safe in inhibiting pancreatic cancer cell survival, inducing apoptosis, and sensitizing cells to chemotherapy via Hedgehog inhibition and caspase-3 activation. Communicated by Ramaswamy H. Sarma

Predicting the chemosensitivity of pancreatic cancer cells as a personalized therapy.

Annually, approx. 4000 patients are diagnosed with pancreatic cancer in Poland, and the number of deaths is close to the number of diagnoses. Such a high morbidity/mortality ratio is caused by a high percentage of unresectable lesions (about 80%) and chemoresistance, which, among other things, is due to the specific desmoplastic environment. Currently, there are 2 main systemic treatment regimens for pancreatic cancer: FOLFIRINOX (which is a combination of folic acid, fluorouracil (5-FU), irinotecan, and oxaliplatin) and combined treatment with nab-paclitaxel plus gemcitabine (NPXL+GMC). In order to increase the effectiveness of systemic treatments for individual patients, cell lines derived from resected pancreatic tumors were developed and their chemosensitivity to various agents was examined. The hypothesis was that patients may benefit from individualization of chemotherapy. Patients with histopathologically confirmed pancreatic cancer were operated on using irreversible electroporation (IRE) procedure. After isolating and establishing individual cell lines, chemosensitivity to 5-FU, GMC and NPXL was determined using MTT assay in primary and metastatic cell cultures. Three primary cell lines were isolated for the prediction of chemosensitivity. Gemcitabine was shown to be more effective at lower doses compared to 5-FU, while NPXL was more effective than 5-FU, and both of these were less effective in metastatic cells. Pancreatic cancer cell chemoresistance was confirmed in stage IV. Determination of chemosensitivity profiles using cell lines may help in the selection of systemic treatments for individual patients. This method can be the basis for a personalized planned chemotherapeutic protocol.

Gemcitabine resistance of pancreatic cancer cells is mediated by IGF1R dependent upregulation of CD44 expression and isoform switching

Chemoresistance in pancreatic cancer cells may be caused by the expansion of inherently resistant cancer cells or by the adaptive plasticity of initially sensitive cancer cells. We investigated how CD44 isoforms switching contributed to gemcitabine resistance. Treating CD44 null/low single-cell clones with increasing amounts of gemcitabine caused an increase in expression of CD44 and development of gemcitabine resistant (GR) cells. Drug sensitivity, invasiveness, and EMT process was evaluated by MTT, Matrigel invasion assays, and western blots. Genetic knockdown and pharmacological inhibitors were used to examine the roles of CD44 and IGF1R in mediating gemcitabine resistance. CD44 promoter activity and its interactive EMT-related transcription factors were evaluated by luciferase reporter assay and chromatin immunoprecipitation assay. Kaplan–Meier curve was created by log-rank test to reveal the clinical relevance of CD44 and IGF1R expression in patients. We found silence of CD44 in GR cells partially restored E-cadherin expression, reduced ZEB1 expression, and increased drug sensitivity. The gemcitabine-induced CD44 expressing and isoform switching were associated with an increase in nuclear accumulation of phosphor-cJun, Ets1, and Egr1 and binding of these transcription factors to the CD44 promoter. Gemcitabine treatment induced phosphorylation of IGF1R and increased the expression of phosphor-cJun, Ets1, and Egr1 within 72 h. Stimulation or suppression of IGF1R signaling or its downstream target promoted or blocked CD44 promoter activity. Clinically, patients whose tumors expressed high levels of CD44/IGF1R showed a poor prognosis. This study suggests that IGF1R-dependent CD44 isoform switching confers pancreatic cancer cells to undergo an adaptive change in response to gemcitabine and provides the basis for improved targeted therapy of pancreatic cancer.

Abstract 2965: Epigenetic mechanisms involved in acquired resistance to combined chemotherapies in digestive cancer cells

Abstract Introduction. One of the challenges in the management of patients with cancer of the digestive tract is to prevent relapse. Epigenetic modifications (i.e. DNA methylation and histone marks) have been proven to be involved in the generation and maintenance of a subset of cells with stemness properties, that resist to conventional therapies. Our aim is to identify complexes of chromatin modifier enzymes (epienzymes) that are involved in acquired chemoresistance/aggressiveness of colon and pancreatic cancer cells. Experimental procedures/Methods. A meta-analysis performed on public transcriptomics data from colorectal cancer (CRC) patients cross-referenced with our own data including basal-like pancreatic cancer samples identified a limited number of epienzymes associated with aggressive subtypes of digestive cancers. Among them, the demethylase KDM4B, specific of the lysine 9 of Histone 3, and the transcriptional repressor PRDM1 where found correlated to the expression of several typical markers of cancer stemness. To confirm the role of these epienzymes in colon and pancreatic cancer cell chemoresistance, we have developed several in vivo and in vitro 2D and 3D models of acquired resistance to oxaliplatin-based combined chemotherapies, including FOLFIRINOX and FOLFOX. In this work, these models as well as patient samples have been used to study (i) the dynamic expression profile of these epienzymes (RT-qPCR, immunofluorescent multiplex staining, single-cell analysis and spatial transcriptomics), (ii) the gene regulons that are bound by these epienzymes (CUT&RUN-seq) and (iii) the functional role of these epienzymes in cancer stemness (organoid and sphere-forming assays). Results. We have shown that PRDM1 and KDM4B are expressed heterogeneously in cancer cell populations and overexpressed during specific time frames along the acquisition of resistance to oxaliplatin-based combined therapies both in colon and pancreatic cancer cells. Furthermore, we have obtained promising results regarding the binding of PRDM1 and KDM4B to key players of pluripotency/differentiation in chemoresistant colon cancer cells. Conclusions. Altogether, our data indicate that specific expression of chromatin modifier enzymes, most likely acting as epigenetic complexes, are key milestones of acquired resistance to conventional therapies. This work will pave the way for considering epienzyme-epienzyme interactions as potent drug targets that could circumvent cancer cell aggressiveness. Citation Format: Elsa Hadj Bachir, Charles Poiraud, Hugo Claux, Soumaya El Moghrabi, Sonia Paget, Belinda Duchêne, Nicolas Jonckheere, Bernadette Neve, Emmanuelle Leteurtre, Isabelle Van Seuningen, Audrey Vincent. Epigenetic mechanisms involved in acquired resistance to combined chemotherapies in digestive cancer cells [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2022; 2022 Apr 8-13. Philadelphia (PA): AACR; Cancer Res 2022;82(12_Suppl):Abstract nr 2965.

Exosomes and their roles in the chemoresistance of pancreatic cancer.

Pancreatic cancer (PC) remains one of the most lethal human malignancies worldwide. Due to the insidious onset and the rapid progression, most patients with PC are diagnosed at an advanced stage rendering them inoperable. Despite the development of multiple promising chemotherapeutic agents as recommended first-line treatment for PC, the therapeutic efficacy is largely limited by unwanted drug resistance. Recent studies have identified exosomes as essential mediators of intercellular communications during the occurrence of drug resistance. Understanding the underlying molecular mechanisms and complex signaling pathways of exosome-mediated drug resistance will contribute to the improvement of the design of new oncologic therapy regimens. This review focuses on the intrinsic connections between the chemoresistance of PC cells and exosomes in the tumor microenvironment (TME).

Sumac Induces Mitochondria‐Dependent Cellular Toxicity in Human Pancreatic Cancer Cell Lines

Pancreatic cancer is a devastating disease that kills about 57,000 Americans and 250,000 people worldwide each year. Chemoresistance, lack of symptoms, and ineffective therapy have resulted in gaps in our understanding of the molecular events underlying its etiology. We aim to identify natural small molecule compounds that are synthetic lethal in cellular models of mutant KRAS‐dependent pancreatic adenocarcinoma. Our preliminary experiments demonstrate that aqueous extracts of the Middle Eastern spice‐plant sumac induce cytotoxicity in two cultured human pancreatic cancer cell lines with KRAS gene mutations; MIA PaCa‐2 (G12C) and PANC‐1 (G12V and G12D) but not in normal fibroblasts. Pancreatic cancer patients, 90% of whom have mutations in the KRAS oncogene, do not respond well to traditional chemotherapy. We hypothesized that the phytochemical sumac added as an adjunct to the therapeutic regimen would skew the otherwise chemoresistant pancreatic cancer cells into an apoptotic phenotype. We hypothesized that sumac’s cytotoxicity was a consequence of mitochondrial targeting, specifically a derangement of mitochondrial membrane potential, leading to mitochondrial compromise, and eventually apoptosis. To test our hypothesis, we treated MIA PaCa‐2 and PANC‐1 cells in vitro with increasing concentrations of aqueous extracts of sumac (0, 0.03, 0.3, and 3.0 mg/mL) for varying time points and measured 1) mitochondrial membrane potential using TMRE assays, 2) ROS production using DHT assays, 3) ATP levels using fluorimetry, and 4) activated caspases 3 and 7. Our data show that sumac treatment 1) ameliorated ROS production, 2) abrogated mitochondrial membrane potential, 3) activated caspases 3 and 7, and 4) reduced cellular ATP levels in a dose‐dependent manner. These changes were accompanied by a dose‐dependent change in the protein expression of the energy‐sensing mitochondrial PGC‐1α, which increased in both cell lines, the anti‐apoptotic protein Bcl‐2, which decreased in the PANC‐1 cell line but not in the MIA PaCa‐2 cell line, the pro‐apoptotic Bax, and mitofusin‐1, which is responsible for the fusion of the mitochondrial outer membranes. These data suggest that sumac mediates “synthetic lethality” in pancreatic cancer cells based on the type of KRAS gene mutation. We also hypothesized that the observed ATP reduction would alter levels of SIRT3, which is major mitochondrial physiology regulating deacetylase enzyme. Immunoblot data show a paradoxical increase in SIRT3 in a dose‐dependent manner, suggesting other roles for SIRT‐3 in this pathway. Furthermore, using HPLC‐based NMR spectroscopy, we identified the active compound in the aqueous sumac extract to be [(2R, 3R, 4S, 5R)‐3, 4, 5, 6‐tetrakis [(2‐deuterio‐3, 4, 5‐trihydroxybenzoyl)oxy]oxan‐2‐yl] methyl‐2‐deuterio‐3, 4, 5‐trihydroxybenzoate. In conclusion, sumac targets hitherto unknown mitochondrial signal transduction pathways and is synthetic lethal for the KRAS mutant G12V and G12D.

High Expression of Bloom Syndrome Helicase is a Key Factor for Poor Prognosis and Advanced Malignancy in Patients with Pancreatic Cancer: A Retrospective Study.

Bloom syndrome helicase (BLM) is overexpressed in multiple types of cancers and its overexpression may induce genomic instability. This study aimed to determine the function of BLM expression in pancreatic cancer. BLM messenger RNA (mRNA) expression was analyzed using public datasets to determine its relationship with pancreatic cancer prognosis. Overall, 182 patients with pancreatic cancer who underwent radical resection at our institution were enrolled. BLM expression was evaluated by immunohistochemistry (IHC). We explored the effect of BLM on the proliferation, invasion, migration, and chemoresistance of pancreatic cancer cells via small-interfering RNAs and performed pathway analysis using gene set enrichment analysis. BLM mRNA expression was higher in tumor tissue than in normal tissue and had a prognostic effect on overall survival (OS) and recurrence-free survival. The same results were validated by IHC. Multivariate analysis showed that high BLM expression was an independent poor prognostic factor for OS (hazard ratio [HR] 1.678, p=0.029). In subgroup analysis, the effect of high BLM expression was more significant on OS in patients with younger age (HR 2.27, p=0.006), male sex (HR 2.39, p=0.002), high cancer antigen 19-9 level (HR 2.44, p=0.001), advanced tumor stage (HR 2.25, p=0.001), lymph node metastasis (HR 2.51, p=0.001), nerve invasion (HR 2.07, p=0.002), and adjuvant chemotherapy (HR 2.66, p<0.001). In vitro, BLM suppression resulted in reduced tumor proliferation, invasion, migration, and chemoresistance. Mechanistically, BLM expression may be associated with E2F1 and E2F2. BLM expression is a prognostic factor for patients with pancreatic cancer, especially in those with advanced malignancies and receiving chemotherapy.

Heat shock factor 1 inhibition sensitizes pancreatic cancer to gemcitabine via the suppression of cancer stem cell-like properties

Pancreatic cancer is a fatal disease with poor prognosis. Gemcitabine has been regarded as the mainstay of chemotherapy for pancreatic cancer; however, it is accompanied with a high rate of chemoresistance. Cancer stem cells (CSCs) are characterized by resistance to traditional chemo- and radiotherapies. We have previously reported that heat shock factor 1 (HSF1) is involved in the invasion and metastasis of pancreatic cancer, a highly conserved transcriptional factor that mediates the canonical proteotoxic stress response. Here, we investigate whether HSF1 contributes to the chemoresistance of pancreatic cancer cells caused by gemcitabine and explore the underlying mechanisms. Genetically engineered mice (LSL-KrasG12D/+; Trp53fl/+; Pdx1-Cre mice), which spontaneously develop pancreatic cancer, were used to examine the sensitivity of pancreatic cancer to gemcitabine in vivo. We found that HSF1 was enriched in sphere-forming cancer cells. Panc-1 and MiaPaCa-2 cells treated chronically with gemcitabine displayed increased transcription and expression of CSC-associated markers. In addition, gemcitabine-surviving Panc-1 and MiaPaCa-2 cells showed an increased ability to form tumorspheres. Moreover, we observed that gemcitabine treatment increased the activity and expression of HSF1, as well as transcription of its downstream targets. Finally, HSF1 inhibition significantly suppressed the expression of CSC-associated markers, augmented the cancer-killing property of gemcitabine, and increased chemosensitivity to gemcitabine in vivo. Our study reveals a novel mechanism in which HSF1 promotes the chemoresistance of pancreatic cancer to gemcitabine by modulating CSC-like properties. Targeting HSF1 could be thus a rational strategy to improve treatment outcomes.

RhoGDI2 induced malignant phenotypes of pancreatic cancer cells via regulating Snail expression.

Rho GDP dissociation inhibitor 2 (RhoGDI2) has been shown to contribute to the aggressive phenotypes of human cancers, such as tumor metastasis and chemoresistance. This study aimed to assess the effects of RhoGDI2 on tumor progression and chemoresistance in pancreatic cancer cells. The expression of RhoGDI2 in pancreatic cancer cells was detected by Western blot analysis. Gain-of-function and loss-of-function approaches were done to examine the malignant phenotypes of the RhoGDI2-expressing or RhoGDI2-depleting cells. The correlation between RhoGDI2 and Snail was also analyzed. Differential expression of RhoGDI2 protein in pancreatic cancer cell lines was identified. Gain-of-function and loss-of-function experiments showed that RhoGDI2 induced the malignant phenotypes of pancreatic cancer cells, including proliferation, migration, invasion, and gemcitabine (GEM) chemoresistance. The upregulation of RhoGDI2 stimulated the expression of Snail, resulting in the altered expression of epithelial marker E-cadherin and mesenchymal marker Vimentin, which were characteristics of the tumorigenic activity of epithelial-mesenchymal transition. The expression of RhoGDI2 and Snail was upregulated in clinical tumor samples, and higher expression of RhoGDI2 or Snail was significantly associated with poor patient survival in pancreatic ductal adenocarcinoma (PDAC). The findings indicated that RhoGDI2 promoted GEM resistance and tumor progression in pancreatic cancer and that RhoGDI2 might be a potential therapeutic target in patients with PDAC.

LncRNA ANRIL aggravates the chemoresistance of pancreatic cancer cells to gemcitabine by targeting inhibition of miR-181a and targeting HMGB1-induced autophagy.

Recent studies focus on long noncoding RNAs (lncRNA) as crucial regulators of cancer biology that contribute to essential cancer cell functions such as cell proliferation, apoptosis, and metastasis. In pancreatic cancer, several lncRNAs have been mentioned as important actors in tumorigenesis. However, the function of lncRNA ANRIL (named as ANRIL as follows) in pancreatic cancer has not been elucidated. In the present study, we show that ANRIL was up-regulated while miR-181a was down-regulated in pancreatic cancer tissues and HMGB1 was highly expressed. Knockdown of ANRIL in pancreatic cancer repressed cellular proliferation, invasion, migration, and reduced chemotherapy resistance to gemcitabine. ANRIL was negatively correlated with miR-181a, while overexpression of miR-181a could reverse the effect. For further mechanism research, we found that miR-181a aimed to HMGB1 which activated cell autophagy. Taken together, our results implicate that the ANRIL, by targeting miR-181a, activates the HMGB1-induced cell autophagy, which is thought to be critical for oncogenesis.