PANC-1 Cells Research Articles

The EFNA4 gene is a potential prognostic biomarker in pancreatic cancer: a bioinformatics analysis.

Pancreatic cancer is a highly aggressive malignancy with poor prognosis, and there is an urgent need to understand its molecular mechanisms for early diagnosis and treatment. Despite surgical resection being the only effective treatment, most patients are diagnosed at an advanced stage, missing the optimal window for therapy. Identifying novel biomarkers is crucial for prognostic assessment, treatment planning, and early intervention. Ephrin A4 (EFNA4), a member of the receptor tyrosine kinase family, is involved in vascular and epithelial development via regulation of cell migration and rejection. However, the role of EFNA4 in pancreatic cancer has not been reported. Therefore, our study aimed to clarify the role of EFNA4 in pancreatic cancer through bioinformatics analysis and vitro experiments. The expression of EFNA4 and its potential value as a diagnostic and prognostic biomarker in pancreatic cancer was analyzed using data from The Cancer Genome Atlas (TCGA) and the Gene Expression Profiling Interactive Analysis (GEPIA) database. According to the expression level of EFNA4, patients were divided into high expression group and low expression group, and the correlation between overall survival (OS) and disease-free survival (DFS) with different expression levels of EFNA4 and clinical parameters were analyzed. Subsequently, reverse-transcription quantitative polymerase chain reaction (RT-qPCR) was performed to detect EFNA4 expression. The proliferation, invasion, and cloning ability of the cells were detected via Cell Counting Kit 8 (CCK8), Transwell, and plate cloning assays, respectively. EFNA4 is highly expressed in pancreatic cancer, and upregulation of EFNA4 is associated with poor prognosis. In this study, EFNA4 expression was correlated with T stage and TNM (tumor-node-metastasis) stage of pancreatic cancer, and the median survival time and progression-free survival (PFS) were worse in those with high EFNA4 expression (394 days) than in those with low expression (525 days) [hazard ratio (HR): 1.47, 95% confidence interval (CI): 1.00-2.16, P=0.047]. In addition, EFNA4 was also found to be involved in the regulation of signal pathways such as cell adhesion, cyclic AMP, insulin secretion, pancreatic secretion, and protein digestion and absorption. In vitro experiments demonstrated that EFNA4 knockdown significantly inhibited the proliferation, cloning ability, and invasiveness of the PANC-1 and SW1990 pancreatic cancer cell lines. The abnormal expression of EFNA4 in pancreatic cancer is associated with poor prognosis. Knockout of EFNA4 gene could significantly inhibit the proliferation and invasion of pancreatic cancer cells. Therefore, EFNA4 may be one of the molecular targets for poor prognosis of patients with pancreatic cancer.

FFAR-mediated signaling drives migration of pancreatic cancer cells in hypoxic fibroblast co-cultures

The tumor microenvironment (TME) comprises cancer and non-cancerous stromal cells, including fibroblasts. Free fatty acids (FFAs) regulate various biological responses by binding to G protein-coupled FFA receptors (FFARs). In this study, we examined the impact of FFAR1 and FFAR4 on the cell migration of pancreatic cancer PANC-1 cells co-cultured with 3T3 fibroblast cells under hypoxic conditions. PANC-1 cells cultured at 1 % O2 exhibited elevated FFAR1 expression and decreased FFAR4 expression compared to those at 21 % O2. Cell migration of PANC-1 cells was reduced under 1 % O2 conditions. FFAR1 knockdown enhanced PANC-1 cell migration, whereas FFAR4 knockdown inhibited it. Co-culture of PANC-1 cells with 3T3 cells at 1 % O2 significantly increased FFAR4 expression, while FFAR1 expression remained unchanged. To evaluate the effects of FFAR1 and FFAR4 on PANC-1 cell migration in co-culture with 3T3 cells, we conducted a wound healing assay using the Culture-Insert 2 Well. PANC-1 and 3T3 cells were individually seeded into the two wells and incubated at both 21 % and 1 % O2 for 13 h. The cell migration of PANC-1 cells co-cultured with 3T3 cells at 1 % O2 was notably higher compared to 21 % O2. TUG-770 reduced and TUG-891 enhanced the cell migration of PANC-1 cells co-cultured with 3T3 cells under both 21 % and 1 % O2 conditions. These findings suggest that FFAR1 and FFAR4 play important roles in regulating the cell migration of PANC-1 cells co-cultured with 3T3 cells under hypoxic conditions.

Effect of SIRT7 on inhibiting the epithelial-mesenchymal transformation in pancreatic cancer cells and related mechanism

Objective: To investigate the effect and mechanism of SIRT7 in epithelial mesenchymal transformation (EMT) of pancreatic cancer cells. Methods: The pancreatic cancer cells were divided into siControl, siSIRT7, over-expression SIRT7, siSIRT7+siCOL4A1, and siSIRT7+siSLUG groups using siRNA or plasmid transfection. The proliferation, migration and invasion of pancreatic cancer cells were detected by EdU, wound healing assay and Transwell experiments, respectively. The expression of EMT and cancer stem cell (CSC) markers were detected by quantitative real-time reverse transcription polymerase chain reaction assay (qRT-PCR) and western blot. RNA sequencing (RNA-seq) in SIRT7 knockdown PANC-1 cells was performed to explore the signaling pathways and target genes regulated by SIRT7. Then the target genes directly regulated by SIRT7 were identified with quantitative chromatin immunoprecipitation experiment (q-ChIP) and chromatin immunoprecipitation polymerase chain reaction (ChIP-PCR). The expressions of SIRT7 and target genes were detected by immunohistochemical (IHC) in pancreatic cancer tissues, and the correlation between SIRT7 and target gene expression was analyzed using TCGA dataset. The correlation between expression of SIRT7 or target genes and survival was analyzed on KM-plotter website. Finally, GeneMANIA, STRING and ENCORI were used to predict SIRT7-related proteins and miRNAs. Results: EdU assay showed that the cell proliferation rates in SIRT7-overexpressed PANC-1 [(19.33±0.35)%] and BxPC-3 cells [(17.00±1.89)%] were lower than those in the control group [(31.60±1.37)% and (24.33±0.78)%, respectively, P＜0.05]. The proliferation rates of SIRT7-knockdown PANC-1 [(23.94±1.00)% and (27.08±0.97)%] and BxPC-3 cells [(22.00±1.86)% and (25.96±1.61)%] were higher than those of the siControl group [(11.80±1.86)% and (13.42±1.39)%, respectively, P＜0.05]. In PANC-1 cells, the wound healing assay showed that the relative migration rate of SIRT7-overexpression cells [(76.67±2.74)%] was lower than that of control cells [(100.00±2.13)%, P＜0.05]; the relative migration rate of cells with SIRT7 knockdown [(134.22±4.08)% and (199.82±9.20)%, respectively] was higher than that of siControl group [(102.24±3.13)%, P＜0.05]. Compared with the control group, SIRT7 overexpression decreased the number of migrated BxPC-3 cells (45.66±1.69 vs 28.33±2.62, P＜0.05); while SIRT7 knockdown increased these numbers (65.66±2.86 and 82.00±2.94 versus 33.00±0.81, P＜0.01). Transwell experiment revealed that the number of invaded cells in SIRT7 overexpression groups (16.33±2.05 and 34.66±1.69) was lower than that control groups (54.33±4.64 and 58.66±5.90, P＜0.05); with SIRT7 knockdown, the numbers of invaded PANC-1 (63.66±2.49 and 69.33±3.29) and BxPC-3 cells (134.33±3.09 and 181.66±4.02) were higher than those in control groups (35.33±2.49 and 42.00±0.81, P˂0.05). Also, SIRT7 knockdown decreased the expressions of epithelial markers and increased the expressions of mesenchymal and CSC markers. RNA-seq analysis showed that SIRT7 was involved in regulating a variety of cancer-related signaling pathways, including the pancreatic cancer pathway and the EMT pathway. Furthermore, SIRT7 could directly bind to the promoter regions of target genes, such as COL4A1 and SLUG. SIRT7 was negatively correlated with the expression and function of COL4A1 and SLUG in pancreatic cancer cells. The expressions of SIRT7, COL4A1, SLUG and SOX2 were verified in pancreatic cancer tissues by IHC. Finally, SIRT7 was predicted to be associated with many proteins and miRNAs based on GeneMANIA, STRING, and ENCORI online tools. Conclusions: SIRT7 can inhibit the EMT of pancreatic cancer cells through transcriptionally inhibiting the expression of target genes, such as COL4A1 and SLUG. Thus, SIRT7 may serve as a potential tumor suppressor gene in pancreatic cancer.

Combined effects of matrix stiffness and obesity-associated signaling directs progressive phenotype in PANC-1 pancreatic cancer cells in vitro.

Obesity is a leading risk factor of pancreatic ductal adenocarcinoma (PDAC) that contributes to poor disease prognosis and outcomes. Retrospective studies have identified this link, but interactions surrounding obesity and PDAC are still unclear. Research has shifted to contributions of fibrosis (desmoplasia) on malignancy, which involves increased deposition of collagens and other extracellular matrix (ECM) molecules and increased ECM crosslinking, all of which contribute to increased tissue stiffening. However, fibrotic stiffening is underrepresented as a model feature in current PDAC models. Fibrosis is shared between PDAC and obesity, and can be leveraged for in vitro model design, as current animal obesity models of PDAC are limited in their ability to isolate individual components of fibrosis to study cell behavior. In the current study, methacrylated type I collagen (PhotoCol®) was photo-crosslinked to pathological stiffness levels to recapitulate fibrotic ECM stiffening. PANC-1 cells were encapsulated within PhotoCol®, and the tumor-tissue constructs were prepared to represent normal (healthy) (∼600 Pa) and pathological (∼2000 Pa) tissues. Separately, human mesenchymal stem cells were differentiated into adipocytes representing lean (2D differentiation) and obese fat tissue (3D collagen matrix differentiation), and conditioned media was applied to PANC-1 tumor-tissue constructs. Conditioned media from obese adipocytes showed increased vimentin expression, a hallmark of invasiveness and progression, that was not seen after exposure to media from lean adipocytes or control media. Characterization of the obese adipocyte secretome suggested that some PANC-1 differences may arise from increased interleukin-8 and -10 compared to lean adipocytes. Additionally, high matrix stiffness associated induced an amoeboid morphology in PANC-1 cells that was not present at low stiffness. Amoeboid morphology is an accessory to epithelial-to-mesenchymal transition and is used to navigate complex ECM environments. This plasticity has greater implications for treatment efficacy of metastatic cancers. Overall, this work 1) highlights the importance of investigating PDAC-obesity interactions to study the effects on disease progression and persistence, 2) establishes PhotoCol® as a matrix material that can be leveraged to study amoeboid morphology and invasion in PDAC, and 3) emphasizes the importance of integrating both biophysical and biochemical interactions associated within both pathologies for in vitro PDAC models.

Artificial and Natural Sweeteners Biased T1R2/T1R3 Taste Receptors Transactivate Glycosylated Receptors on Cancer Cells to Induce Epithelial-Mesenchymal Transition of Metastatic Phenotype.

Understanding the role of biased taste T1R2/T1R3 G protein-coupled receptors (GPCR) agonists on glycosylated receptor signaling may provide insights into the opposing effects mediated by artificial and natural sweeteners, particularly in cancer and metastasis. Sweetener-taste GPCRs can be activated by several active states involving either biased agonism, functional selectivity, or ligand-directed signaling. However, there are increasing arrays of sweetener ligands with different degrees of allosteric biased modulation that can vary dramatically in binding- and signaling-specific manners. Here, emerging evidence proposes the involvement of taste GPCRs in a biased GPCR signaling crosstalk involving matrix metalloproteinase-9 (MMP-9) and neuraminidase-1 (Neu-1) activating glycosylated receptors by modifying sialic acids. The findings revealed that most natural and artificial sweeteners significantly activate Neu-1 sialidase in a dose-dependent fashion in RAW-Blue and PANC-1 cells. To confirm this biased GPCR signaling crosstalk, BIM-23127 (neuromedin B receptor inhibitor, MMP-9i (specific MMP-9 inhibitor), and oseltamivir phosphate (specific Neu-1 inhibitor) significantly block sweetener agonist-induced Neu-1 sialidase activity. To assess the effect of artificial and natural sweeteners on the key survival pathways critical for pancreatic cancer progression, we analyzed the expression of epithelial-mesenchymal markers, CD24, ADLH-1, E-cadherin, and N-cadherin in PANC-1 cells, and assess the cellular migration invasiveness in a scratch wound closure assay, and the tunneling nanotubes (TNTs) in staging the migratory intercellular communication. The artificial and natural sweeteners induced metastatic phenotype of PANC-1 pancreatic cancer cells to promote migratory intercellular communication and invasion. The sweeteners also induced the downstream NFκB activation using the secretory alkaline phosphatase (SEAP) assay. These findings elucidate a novel taste T1R2/T1R3 GPCR functional selectivity of a signaling platform in which sweeteners activate downstream signaling, contributing to tumorigenesis and metastasis via a proposed NFκB-induced epigenetic reprogramming modeling.

Coumarin-modified ruthenium complexes: Synthesis, characterization, and antiproliferative activity against human cancer cells.

Among ruthenium complexes studied as anticancer metallodrugs, NKP-1339, NAMI-A, RM175, and RAPTA-C have already entered clinical trials due to their potent antitumor activity demonstrated in preclinical studies and reduced toxicity in comparison with platinum drugs. Considering the advantages of ruthenium-based anticancer drugs and the cytostatic activity of organometallic complexes with triazole- and coumarin-derived ligands, we set out to synthesize Ru(II) complexes of coumarin-1,2,3,-triazole hybrids (L) with the general formula [Ru(L)(p-cymene)(Cl)]ClO4. The molecular structure of the complex [Ru(2a)(p-cymene)(Cl)]ClO4 (2aRu) was determined by single-crystal X-ray diffraction, which confirmed the coordination of the ligand to the central ruthenium(II) cation by bidentate mode of coordination. Coordination with Ru(II) resulted in the enhancement of cytostatic activity in HepG2 hepatocellular carcinoma cells and PANC-1 pancreatic cancer cells. Coumarin derivative 2a positively regulated the expression and activity of c-Myc and NPM1 in RKO colon carcinoma cells, while the Ru(II) half-sandwich complex 2cRu induced downregulation of AKT and ERK signaling in PANC-1 cells concomitant with reduced intracellular levels of reactive oxygen species. Altogether, our findings indicated that coumarin-modified half-sandwich Ru(II) complexes held potential as anticancer agents against gastrointestinal malignancies.

Cellpose as a reliable method for single-cell segmentation of autofluorescence microscopy images.

Autofluorescence microscopy uses intrinsic sources of molecular contrast to provide cellular-level information without extrinsic labels. However, traditional cell segmentation tools are often optimized for high signal-to-noise ratio (SNR) images, such as fluorescently labeled cells, and unsurprisingly perform poorly on low SNR autofluorescence images. Therefore, new cell segmentation tools are needed for autofluorescence microscopy. Cellpose is a deep learning network that is generalizable across diverse cell microscopy images and automatically segments single cells to improve throughput and reduce inter-human biases. This study aims to validate Cellpose for autofluorescence imaging, specifically from multiphoton intensity images of NAD(P)H. Manually segmented nuclear masks of NAD(P)H images were used to train new Cellpose models. These models were applied to PANC-1 cells treated with metabolic inhibitors and patient-derived cancer organoids (across 9 patients) treated with chemotherapies. These datasets include co-registered fluorescence lifetime imaging microscopy (FLIM) of NAD(P)H and FAD, so fluorescence decay parameters and the optical redox ratio (ORR) were compared between masks generated by the new Cellpose model and manual segmentation. The Dice score between repeated manually segmented masks was significantly lower than that of repeated Cellpose masks (p<0.0001) indicating greater reproducibility between Cellpose masks. There was also a high correlation (R 2 >0.9) between Cellpose and manually segmented masks for the ORR, mean NAD(P)H lifetime, and mean FAD lifetime across 2D and 3D cell culture treatment conditions. Masks generated from Cellpose and manual segmentation also maintain similar means, variances, and effect sizes between treatments for the ORR and FLIM parameters. Overall, Cellpose provides a fast, reliable, reproducible, and accurate method to segment single cells in autofluorescence microscopy images such that functional changes in cells are accurately captured in both 2D and 3D culture.

Thymoquinone affects hypoxia-inducible factor-1α expression in pancreatic cancer cells via HSP90 and PI3K/AKT/mTOR pathways.

Pancreatic cancer (PC) is associated with some of the worst prognoses of all major cancers. Thymoquinone (TQ) has a long history in traditional medical practice and is known for its anti-cancer, anti-inflammatory, anti-fibrosis and antioxidant pharmacological activities. Recent studies on hypoxia-inducible factor-1α (HIF-1α) and PC have shown that HIF-1α affects the occurrence and development of PC in many aspects. In addition, TQ could inhibit the development of renal cancer by decreasing the expression of HIF-1α. Therefore, we speculate whether TQ affects HIF-1α expression in PC cells and explore the mechanism. To elucidate the effect of TQ in PC cells and the regulatory mechanism of HIF-1α expression. Cell counting kit-8 assay, Transwell assay and flow cytometry were performed to detect the effects of TQ on the proliferative activity, migration and invasion ability and apoptosis of PANC-1 cells and normal pancreatic duct epithelial (hTERT-HPNE) cells. Quantitative real-time polymerase chain reaction and western blot assay were performed to detect the expression of HIF-1α mRNA and protein in PC cells. The effects of TQ on the HIF-1α protein initial expression pathway and ubiquitination degradation in PANC-1 cells were examined by western blot assay and co-immunoprecipitation. TQ significantly inhibited proliferative activity, migration, and invasion ability and promoted apoptosis of PANC-1 cells; however, no significant effects on hTERT-HPNE cells were observed. TQ significantly reduced the mRNA and protein expression levels of HIF-1α in PANC-1, AsPC-1, and BxPC-3 cells. TQ significantly inhibited the expression of the HIF-1α initial expression pathway (PI3K/AKT/mTOR) related proteins, and promoted the ubiquitination degradation of the HIF-1α protein in PANC-1 cells. TQ had no effect on the hydroxylation and von Hippel Lindau protein mediated ubiquitination degradation of the HIF-1α protein but affected the stability of the HIF-1α protein by inhibiting the interaction between HIF-1α and HSP90, thus promoting its ubiquitination degradation. The regulatory mechanism of TQ on HIF-1α protein expression in PC cells was mainly to promote the ubiquitination degradation of the HIF-1α protein by inhibiting the interaction between HIF-1α and HSP90; Secondly, TQ reduced the initial expression of HIF-1α protein by inhibiting the PI3K/AKT/mTOR pathway.

Novel Histone Deacetylase (HDAC) Inhibitor Induces Apoptosis and Suppresses Invasion via E-Cadherin Upregulation in Pancreatic Ductal Adenocarcinoma (PDAC).

Pancreatic ductal adenocarcinoma (PDAC) is the most lethal form of pancreatic cancer characterized by therapy resistance and early metastasis, resulting in a low survival rate. Histone deacetylase (HDAC) inhibitors showed potential for the treatment of hematological malignancies. In PDAC, the overexpression of HDAC 2 is associated with the epithelial-mesenchymal transition (EMT), principally accompanied by the downregulation of the epithelial marker E-cadherin and increased metastatic capacity. The effector cytokine transforming growth factor-β (TGF β) is known to be a major inducer of the EMT in PDAC, leading to high metastatic and invasive potential. In addition, the overexpression of HDAC 6 in PDAC is associated with reduced apoptosis. Here, we have demonstrated that a novel HDAC 2/6 inhibitor not only significantly increased E-cadherin expression in PANC-1 cells (5.5-fold) and in 3D PDAC co-culture spheroids (2.5-fold) but was also able to reverse the TGF-β-induced downregulation of E-cadherin expression. Moreover, our study indicates that the HDAC inhibitor mediated re-differentiation resulting in a significant inhibition of tumor cell invasion by approximately 60% compared to control. In particular, we have shown that the HDAC inhibitor induces both apoptosis (2-fold) and cell cycle arrest. In conclusion, the HDAC 2/6 inhibitor acts by suppressing invasion via upregulating E-cadherin mediated by HDAC 2 blockade and by inducing cell cycle arrest leading to apoptosis via HDAC 6 inhibition. These results suggest that the HDAC 2/6 inhibitor might represent a novel therapeutic strategy for the treatment of PDAC tumorigenesis and metastasis.

Chemical constituents from Cinnamomi Ramulus decoction

Six compounds were isolated from the ethyl acetate extracts of Cinnamomi Ramulus decoction by RP-18, silica gel, Sephadex LH-20 column chromatography, together with prep-HPLC methods. Based on HR-ESI-MS, MS, 1D and 2D NMR spectral analyses, the structures of the six compounds were identified as 4,5,10,11-tetrahydroxybisabol-7-ene(1), 4,5,10,11-tetrahydroxybisabolin(2), 1-phenyl-1,2,3-glycerol(3),(+)-lyoniresinol(4), benzoic acid(5), and decumbic acid(6). Compound 1 was a new bisabolene-type sesquiterpene, and compounds 2 and 3 were isolated from the Cinnamomi Ramulus for the first time. Moreover, the bisabolene-type sesquiterpene(2) was assayed for its anti-inflammatory activity and cytotoxicity to human pancreatic cancer cells(PANC-1 cells). RESULTS:: showed that compound 2 exhibited an inhibitory rate of 32.9% on nitric oxide(NO) at a dose of 40 μmol·L~(-1) and a proliferation inhibition rate of 14.5% against PANC-1 cells at a dose of 20 μmol·L~(-1). It did not demonstrate significant activity.

Modulation of purinergic signaling in endothelial cells by tumor microenvironment

Purinergic signaling plays a crucial role in vascular endothelium functions. In particular, ionotropic P2X receptors (P2XRs) are engaged in various intracellular pathways through which endothelial cells (ECs) adapt to external stimuli. However, very little is known about the impact of P2XRs on vascular remodeling during carcinogenesis.We previously demonstrated that high purinergic stimulation impairs the migratory phenotype of tumor-derived endothelial cells (TECs) but not of normal ECs.Since P2XRs are sensitive to different physical and chemical factors, we investigated the impact of tumor microenvironment (TME) on healthy ECs to verify the ability of cancer cells to affect endothelial migratory phenotype through purinergic signaling tuning. More specifically, we focused on P2XR modulation by two different types of TME, mimicking breast and pancreas cancer milieux, which show very different features in terms of vascularization and composition. ECs conditioning with both cancer cell types induced a significant upregulation of some of the most represented P2XR. However, only conditioning with MCF-7 cells and not that with PANC-1 cells was able to alter the migratory phenotype of normal ECs supporting a P2XR-mediated inhibition of cell migration. The differences observed between the two cancer cells could be due to their different proliferative potential and the subsequent different extracellular pH. In addition, in agreement with some of our previous data, the P2XR-induced inhibition of EC migration seems to be independent of calcium signals, as conditioned ECs didn't reveal any changes in the long-lasting responses evoked by purinergic agonists.Collectively, highlighting a significant P2RX modulation by TME, our data strengthen the hypothesis that purinergic signaling may play a central role in vascular remodeling during carcinogenesis. However, the molecular routes upstream and downstream of this modulation remain to be elucidated.

Combination Treatment of Biochanin A and Atorvastatin Alters Mitochondrial Bioenergetics, Modulating Cell Metabolism and Inducing Cell Cycle Arrest in Pancreatic Cancer Cells.

Pancreatic cancer is an aggressive type of cancer, with a dismally low survival rate of <5%. FDA-approved drugs like gemcitabine have shown little therapeutic success, prolonging survival by a mere six months. Isoflavones, such as biochanin A and daidzein, are known to exhibit anti-cancer activity, whereas statins reportedly have anti-proliferative effects. This study investigated the effects of combination treatment of biochanin A and atorvastatin on pancreatic cancer cells. Pancreatic cancer cells AsPC-1, PANC-1, and MIA PaCa-2 were procured from ATCC. The cell viability studies were carried out using MTT & cell count assays. Flow cytometry was used to study cell apoptosis whereas cell metabolism studies were carried out using the Seahorse Mito stress test and XF-PMP assay. The effects of treatment on cell signaling pathways & cell cycle associated proteins were investigated using western blot whereas invasiveness of cancer cells was evaluated using gelatin zymography. The combination treatment decreased the survival and enhanced pro-apoptotic responses compared to single treatments in the pancreatic cancer cells. In PANC-1 cells, the combination treatment decreased invasiveness, reduced expression of activated STAT3 and expression of critical mediators of cell cycle progression. Furthermore, the combination treatment induced a differential inhibition of respiratory complexes in the pancreatic cancer cells. The combination treatment of biochanin A and atorvastatin exerts enhanced anti-cancer effects, inducing apoptosis, down-regulating cell cycle associated proteins and invasiveness in pancreatic cancer cells and merits further investigation for new, improved treatments for pancreatic cancer.

Statement of Retraction: Propofol inhibits migration and induces apoptosis of pancreatic cancer PANC-1 cells through miR-34a-mediated E-cadherin and LOC285194 signals

Statement of Retraction: Propofol inhibits migration and induces apoptosis of pancreatic cancer PANC-1 cells through miR-34a-mediated E-cadherin and LOC285194 signals

Enhancing chemosensitivity of PANC1 pancreatic cancer cells to gemcitabine using ANGTPL4, Notch1 and NF-κβ1 siRNAs.

Aim: siRNA can silence targeted genes with lesser toxicity than therapeutic drugs. Therefore, this study aims to investigate new approaches to treatpancreatic cancer (PC) using combinations of siRNA and gemcitabine. Methods: Three genes, ANGTPL4, Notch1 and NF-κβ1, were silenced using siRNA, and their anti-proliferative effects were studied in combination with gemcitabine on pancreatic cancer cell line (PANC-1) using MTT viability assay. Results: Our results showed a significant reduction in PANC-1 cells growth upon treating cells with gemcitabine and single and combinations of siRNA sequences specific for ANGTPL4, Notch1 and NF-κβ1 genes. Conclusion: Co-transfection of gemcitabine-treated PANC-1 cells with ANGPTL4, Notch1 and NF-κβsiRNAs enhances the chemosensitivity of PANC-1 cells to gemcitabine can be a promising therapeutic approach.

Synthesis, cytotoxicity and antioxidant activity of new conjugates of N-acetyl-d-glucosamine with α-aminophosphonates

A series of the first conjugates of N-acetyl-d-glucosamine with α-aminophosphonates was synthesized using the Kabachnik-Fields reaction, the Pudovik reaction, a copper(I)-catalyzed azide-alkyne cycloaddition reaction (CuAAC) and evaluated for the in vitro cytotoxicity against human cancer cell lines M − HeLa, HuTu-80, A549, PANC-1, MCF-7, T98G and normal lung fibroblast cells WI-38. The tested conjugates, with exception of compound 21b, considered as a lead compound, were either inactive against the used cancer cells or showed moderate cytotoxicity in the range of IC50 values 33−80 μM. The lead compound 21b, being non cytotoxic against normal human cells WI-38 (IC50 = 90 μM), demonstrated good activity (IC50 = 17 μM) against breast adenocarcinoma cells (MCF-7) which to be 1.5 times higher than the activity of the used reference anticancer drug tamoxifen (IC50 = 25.0 μM). A flexible receptor molecular docking simulation showed that the cytotoxicity of the synthesized conjugates of N-acetyl-d-glucosamine with α-aminophosphonates against breast adenocarcinoma MCF-7 cell line is due to their ability to inhibit EGFR kinase domain. In addition, it was found that conjugates 22a and 22b demonstrated antioxidant activity that was not typical for α-aminophosphonates.

CCK Receptor Inhibition Reduces Pancreatic Tumor Fibrosis and Promotes Nanoparticle Delivery.

The poor prognosis for pancreatic ductal adenocarcinoma (PDAC) patients is due in part to the highly fibrotic nature of the tumors that impedes delivery of therapeutics, including nanoparticles (NPs). Our prior studies demonstrated that proglumide, a cholecystokinin receptor (CCKR) antagonist, reduced fibrosis pervading PanIN lesions in mice. Here, we further detail how the reduced fibrosis elicited by proglumide achieves the normalization of the desmoplastic tumor microenvironment (TME) and improves nanoparticle uptake. One week following the orthotopic injection of PDAC cells, mice were randomized to normal or proglumide-treated water for 3-6 weeks. Tumors were analyzed ex vivo for fibrosis, vascularity, stellate cell activation, vascular patency, and nanoparticle distribution. The histological staining and three-dimensional imaging of tumors each indicated a reduction in stromal collagen in proglumide-treated mice. Proglumide treatment increased tumor vascularity and decreased the activation of cancer-associated fibroblasts (CAFs). Additionally, PANC-1 cells with the shRNA-mediated knockdown of the CCK2 receptor showed an even greater reduction in collagen, indicating the CCK2 receptors on tumor cells contribute to the desmoplastic TME. Proglumide-mediated reduction in fibrosis also led to functional changes in the TME as evidenced by the enhanced intra-tumoral distribution of small (<12 nm) Rhodamine-loaded nanoparticles. The documented in vivo, tumor cell-intrinsic anti-fibrotic effects of CCK2R blockade in both an immunocompetent syngeneic murine PDAC model as well as a human PDAC xenograft model demonstrates that CCK2R antagonists, such as proglumide, can improve the delivery of nano-encapsulated therapeutics or imaging agents to pancreatic tumors.

PITX2 functions as a transcription factor for GPX4 and protects pancreatic cancer cells from ferroptosis

Ferroptosis inhibits tumor progression in pancreatic cancer cells, while PITX2 is known to function as a pro-oncogenic factor in various tumor types, protecting them from ferroptosis and thereby promoting tumor progression. In this study, we sought to investigate the regulatory role of PITX2 in tumor cell ferroptosis within the context of pancreatic cancer. We conducted PITX2 knockdown experiments using lentiviral infection in two pancreatic cancer cell lines, namely PANC-1 and BxPC-3. We assessed protein expression through immunoblotting and mRNA expression through RT-PCR. To confirm PITX2 as a transcription factor for GPX4, we employed Chromatin Immunoprecipitation (ChIP) and Dual-luciferase assays. Furthermore, we used flow cytometry to measure reactive oxygen species (ROS), lipid peroxidation, and apoptosis and employed confocal microscopy to assess mitochondrial membrane potential. Additionally, electron microscopy was used to observe mitochondrial structural changes and evaluate PITX2's regulation of ferroptosis in pancreatic cancer cells. Our findings demonstrated that PITX2, functioning as a transcription factor for GPX4, promoted GPX4 expression, thereby exerting an inhibitory effect on ferroptosis in pancreatic cancer cells and consequently promoting tumor progression. Moreover, PITX2 enhanced the invasive and migratory capabilities of pancreatic cancer cells by activating the WNT signaling pathway. Knockdown of PITX2 increased ferroptosis and inhibited the proliferation of PANC-1 and BxPC-3 cells. Notably, the inhibitory effect on ferroptosis resulting from PITX2 overexpression in these cells could be countered using RSL3, an inhibitor of GPX4. Overall, our study established PITX2 as a transcriptional regulator of GPX4 that could promote tumor progression in pancreatic cancer by reducing ferroptosis. These findings suggest that PITX2 may serve as a potential therapeutic target for combating ferroptosis in pancreatic cancer.

Chlorogenic Acid Inhibits Proliferation, Migration and Invasion of Pancreatic Cancer Cells via AKT/GSK-3β/β-catenin Signaling Pathway.

Chlorogenic acid (CA, United States Patent No. 10772340), a natural biologically active food ingredient, displays potent antitumor activity against a variety of cancer cells. However, the mechanism underlying its anticancer effect is not well elucidated. In the present study, we hope to dissect the mechanism underlying the anticancer effects of CA in pancreatic cancer cells. The cytotoxicity of CA in pancreatic cancer cells was determined by MTT assay. Flow cytometry was performed to evaluate the cells apoptosis, while a clonogenic assay was carried out to check the colony formation of cancer cells. Transwell assay was performed to assess the cells migration and invasion. The protein expression of AKT/GSK-3β/β-catenin signaling pathway was detected by Western Blot. Our data indicated that CA inhibited the proliferation of PANC-28 and PANC-1 cells in a dose and time-dependent manner. CA was able to inhibit colony formation, migration, and invasion ability and trigger apoptosis in PANC-28 and PANC-1 cells. Further study showed that CA down-regulated the expression of AKT, p-AKT(Thr308), p-GSK-3β(Ser9), β-catenin, N-cadherin, and vimentin while enhancing the expression of cleaved-caspase 3 and cleaved-caspase 7 in PANC-28 and PANC-1 cells. Our study provides significant evidence that CA is able to inhibit the growth of pancreatic cancer via the AKT/GSK-3β/β-catenin signaling pathway.

Paris saponin VII induces Caspase-3/GSDME-dependent pyroptosis in pancreatic ductal adenocarcinoma cells by activating ROS/Bax signaling

ObjectiveParidis Rhizoma (Chonglou in Chinese), a traditional Chinese herbal medicine, has been shown have strong anti-tumor effects. Paris saponin VII (PSVII), an active constituent isolated from Paridis Rhizoma, was demonstrated to significantly suppress the proliferation of BxPC-3 cells in our previous study. Here, we aimed to elucidate the anti-pancreatic ductal adenocarcinoma (PDAC) effect of PSVII and the underlying mechanism. MethodsCell viability was determined by CCK-8, colony formation, and cell migration assays. Cell apoptosis and reactive oxygen species (ROS) production were measured by flow cytometry with annexin V/propidine iodide (Annexin V/PI) and 2′,7′-dichlorodihydrofluorescein diacetate (DCFH-DA), respectively. Pyroptosis was evaluated by morphological features, Hoechst 33342/PI staining assay, and release of lactate dehydrogenase (LDH). JC-1 fluorescent dye was employed to measure mitochondrial membrane potential. Western blotting and reverse transcription-quantitative polymerase chain reaction (RT-qPCR) were used to determine the levels of proteins or mRNAs. The effect in vivo was assessed by a xenograft tumor model. ResultsPSVII inhibited the viability of PDAC cells (BxPC-3, PANC-1, and Capan-2 cells) and induced gasdermin E (GSDME) cleavage, as well as the simultaneous cleavage of Caspase-3 and poly (ADP-ribose) polymerase 1 (PARP). Knockdown of GSDME shifted PSVII-induced pyroptosis to apoptosis. Additionally, the effect of PSVII was significantly attenuated by Z-Asp(OMe)-Glu(OMe)-Val-Asp(OMe)-fluoromethylketone (Z-DEVD-FMK), on the induction of GSDME-dependent pyroptosis. PSVII also elevated intracellular ROS accumulation and stimulated Bax and Caspase-3/GSDME to conduct pyroptosis in PDAC cells. The ROS scavenger N-acetyl cysteine (NAC) suppressed the release of LDH and inhibited Caspase-9, Caspase-3, and GSDME cleavage in PDAC cells, ultimately reversing PSVII-induced pyroptosis. Furthermore, in a xenograft tumor model, PSVII markedly suppressed the growth of PDAC tumors and induced pyroptosis. ConclusionThese results demonstrated that PSVII exerts therapeutic effects through Caspase-3/GSDME-dependent pyroptosis and may constitute a novel strategy for preventing chemotherapeutic resistance in patients with PDAC in the future.

Anti-pancreatic cancer activity of cassane diterpenoids isolated from the seeds of Caesalpinia sappan mediated by autophagy activation via ROS/AMPK/mTORC1 pathway

Three undescribed cassane diterpenoids, caesalpanins D–F (1–3), and seven known ones were isolated from the seeds of Caesalpinia sappan. Structures and absolute configurations of 1–3 were elucidated based on the extensive spectroscopic analysis, single-crystal X-ray diffraction analysis, and ECD calculations. Structurally, compound 1 was the first example of 18-norcassane diterpenoid and 2 was a rare 20-norcassane diterpenoid having an unusual five-membered oxygen bridge between C-10/C-18. The anti-proliferative activity of 1, 3, and 4–10 against PANC-1 cells (pancreatic ductal adenocarcinoma cell line) was evaluated, and phanginin H (4) was found to exhibit anti-cancer activity with IC50 value of 18.13 ± 0.63 μM. Compound 4 inhibited PANC-1 cell growth by arresting the cell cycle at G2/M phase via regulation of cyclin-dependent kinases, and the self-renewal and metastasis of PANC-1 cells by suppressing cancer cell stemness. Furthermore, compound 4 induced ROS generation and subsequently activated autophagy, which was demonstrated by the formation of autophagic vacuoles and dynamic change of autophagic flux. The induced ROS accumulation resulted in AMPK activation and subsequently regulation of mTORC1 activity and ULK phosphorylation, indicating that 4 triggered autophagy through ROS/AMPK/mTORC1 pathway. These findings suggested that 4 might potentially be an autophagy inducer for the therapy of pancreatic cancer.