calpain and Pancreatic-Neoplasms

Sulfonylureas (SUs) are suggested to accelerate the pancreatic β-cells mass loss via apoptosis. However, little is known whether calpains mediate this process. The aim of the present study is to evaluate the involvement of calpains in SUs-induced death of human pancreatic cancer (PC) cell line 1.2B4. The cells were exposed to: glibenclamide, glimepiride and gliclazide for 72 h. The expression analysis of caspase-3 (CASP-3), TP53, calpain 1 (CAPN-1), calpain 2 (CAPN-2) and calpain 10 (CAPN-10) was detected using RT-PCR method. Intracellular Ca

Topics: Calcium; Calpain; Caspase 3; Cell Death; Cell Line, Tumor; Cell Survival; Humans; Hypoglycemic Agents; Pancreatic Neoplasms; Sulfonylurea Compounds; Tumor Suppressor Protein p53

To identify therapeutic targets for KRAS mutant pancreatic cancer, we conduct a druggable genome small interfering RNA (siRNA) screen and determine that suppression of BCAR1 sensitizes pancreatic cancer cells to ERK inhibition. Integrative analysis of genome-scale CRISPR-Cas9 screens also identify BCAR1 as a top synthetic lethal interactor with mutant KRAS. BCAR1 encodes the SRC substrate p130Cas. We determine that SRC-inhibitor-mediated suppression of p130Cas phosphorylation impairs MYC transcription through a DOCK1-RAC1-β-catenin-dependent mechanism. Additionally, genetic suppression of TUBB3, encoding the βIII-tubulin subunit of microtubules, or pharmacological inhibition of microtubule function decreases levels of MYC protein in a calpain-dependent manner and potently sensitizes pancreatic cancer cells to ERK inhibition. Accordingly, the combination of a dual SRC/tubulin inhibitor with an ERK inhibitor cooperates to reduce MYC protein and synergistically suppress the growth of KRAS mutant pancreatic cancer. Thus, we demonstrate that mechanistically diverse combinations with ERK inhibition suppress MYC to impair pancreatic cancer proliferation.

Topics: Acetamides; Apoptosis; Calpain; Cell Line, Tumor; Cell Proliferation; Crk-Associated Substrate Protein; Down-Regulation; Drug Synergism; Extracellular Signal-Regulated MAP Kinases; Gene Expression Regulation, Neoplastic; Half-Life; Humans; Microtubules; Morpholines; Mutation; Organoids; Pancreatic Neoplasms; Protein Kinase Inhibitors; Proto-Oncogene Proteins c-myc; Proto-Oncogene Proteins p21(ras); Pyridines; src-Family Kinases; Transcription, Genetic; Tubulin; Xenograft Model Antitumor Assays

Topics: Acinar Cells; Animals; Apoptosis; Ataxia Telangiectasia; Ataxia Telangiectasia Mutated Proteins; Calpain; Caspase 3; Cell Line, Tumor; Humans; Oxidative Stress; Pancreatic Neoplasms; Rats

TRIM family proteins are defined as E3 ubiquitin ligases because of their RING-finger domains. The ubiquitin-like protein interferon-stimulated gene 15 (ISG15) encodes a 15-kDa protein, that is implicated in the posttranslational modification of diverse proteins. Both TRIM29 and ISG15 play both pro-tumoral and anti-tumoral functions in cancer cells derived from different histology. In the current study, we demonstrated that correlation expression of TRIM29 and ISG15 in pancreatic ductal adenocarcinomas (PDACs). The current study demonstrated that TRIM29 knockdown destabilized ISG15 protein via promoting its processing by calpain 3 (CAPN3). Importantly, the current study found that TRIM29 knockdown suppressed cancer stem cell-like features of PDACs, which can be rescued by ISG15 independent of its conjugation function. In addition, the current study demonstrated that extracellular free ISG15 played an important role in maintenance of cancer stem cell-like features of PDACs. Thereby, the current study displayed a novel mechanism by which TRIM29 modulates ISG15 stability via CAPN3-mediated processing, and subsequently extracellular ISG15 maintains the cancer stem cell-like features of PDAC via autocrine mode of action.

Topics: Animals; Autocrine Communication; Calpain; Carcinoma, Pancreatic Ductal; Cell Line, Tumor; Cytokines; DNA-Binding Proteins; Female; Gene Knockdown Techniques; Humans; Mice; Muscle Proteins; Neoplastic Stem Cells; Pancreatic Neoplasms; Proteolysis; Transcription Factors; Ubiquitins; Xenograft Model Antitumor Assays

Pancreatic ductal adenocarcinoma (PDAC) has a devastating prognosis. The performance of clinicopathologic parameters and molecules as prognostic factors remains limited and inconsistent. The present study aimed to construct a multi-molecule biomarker panel to more accurately predict post-resectional prognosis of PDAC patients.. Firstly, a novel computational strategy integrating prognostic evidence from omics and literature on the basis of bioinformatics prediction (CIPHER) to generate the network, was designed to systematically identify potential high-confidence PDAC-related prognostic candidates. After specimens from 605 resected PDAC patients were retrospectively collected, 23 candidates were detected immunohistochemically in tissue-microarrays for the development cohort to construct a multi-molecule panel. Lastly, the panel was validated in two independent cohorts.. According to the constructed five-molecule panel, disease-specific survival (DSS) was significantly poorer in high-risk patients than in low-risk ones in development cohort (HR 2.15, 95%CI 1.51-3.05, P<0.0001; AUC 0.67). In two validation cohorts, similar significant differences between the two groups were also observed (HR 3.18 and 3.31, 95%CI 1.89-5.37 and 1.78-6.16, All P<0.0001; AUC 0.72 and 0.73). In multivariate analyses, this panel was the sole prognosticator that was significant in each cohort. Furthermore, its predictive power for long-term survival, higher than its individual constituents, could be largely enhanced by combination with traditional clinicopathological variables. Finally, adjuvant chemotherapy (ACT) correlated with better DSS only in high-risk patients, uni- and multi-variately, in all the cohorts.. The novel prognostic panel developed by a systematically network-based strategy presents strong ability in prediction of post-resectional survival of PDAC patients. Furthermore, panel-defined high-risk patients might benefit more from ACT.

Topics: Aged; Antineoplastic Combined Chemotherapy Protocols; Area Under Curve; Biomarkers, Tumor; Calpain; Carcinoma, Pancreatic Ductal; Chemotherapy, Adjuvant; Disease-Free Survival; Dishevelled Proteins; Female; Filamins; Gene Expression; Hedgehog Proteins; Humans; Immunohistochemistry; Male; Middle Aged; Pancreatectomy; Pancreatic Neoplasms; Prognosis; Retrospective Studies; Zinc Finger Protein GLI1

Pancreatic cancer (PC) is a highly aggressive and metastatic disease, with an elevated mortality rate. It is, therefore, crucial to assess factors affecting the prognosis of PC patients. Meanwhile, calpain-1 is associated with malignant tumor progression and metastasis. Thus, it is meaningful to evaluate the relationship between calpain-1 and PC.. Calpain-1 protein expression was assessed by immunohistochemistry in 96 pancreatic cancer samples and paired adjacent non-cancerous specimens. In addition, calpain-1 protein levels were assessed in six PC cell lines by western blot (WB). Next, PC cells were transfected with calpain-1 siRNA, and silencing was confirmed by WB. Finally, cell proliferation, colony formation, migration and invasion assays, and cell apoptosis analysis were performed to examine the effects of calpain-1 knockdown on proliferation, growth, apoptosis, migration, and invasion in PC cells.. The results showed that calpain-1 was overexpressed in PC tissues and cells. Meanwhile, calpain-1 overexpression was associated with tumor site (P = 0.029), metastasis (P = 0.000), and TNM stage (P = 0.000), but showed no associations with histological grade (P = 0.396), age (P = 0.809), sex (P = 1.000), and lesion size (P = 0.679). The Kaplan-Meier method demonstrated that the low calpain-1 expression group had increased overall survival (OS) compared with patients expressing high calpain-1 levels (28.7 ± 4.1 vs. 17.0 ± 2.3 months) (P = 0.005). Besides, calpain-1 in PC cells was successfully silenced by liposome-mediated RNA interference, resulting in reduced cell growth, invasion, and metastasis in PC cells, with no effect on apoptosis.. The above findings suggest that calpain-1 should be considered a potential biomarker for PC prognosis and therapy.

Topics: Apoptosis; Biomarkers, Tumor; Calpain; Carcinoma, Pancreatic Ductal; Case-Control Studies; Cell Cycle; Cell Movement; Cell Proliferation; Disease Progression; Female; Follow-Up Studies; Gene Expression Regulation, Neoplastic; Humans; Lymphatic Metastasis; Male; Middle Aged; Neoplasm Invasiveness; Pancreatectomy; Pancreatic Neoplasms; Prognosis; Survival Rate; Tumor Cells, Cultured

Desmoplasia contributes to the aggressive behavior of pancreatic cancer. However, recent clinical trials testing several antifibrotic agents on pancreatic cancer have not shown clear efficacy. Therefore, further investigation of desmoplasia-targeting antifibrotic agents by another mechanism is needed. Calpeptin, an inhibitor of calpains, suppressed fibroblast function and inhibited fibrosis. In this study, we investigated the anticancer effects of calpeptin on pancreatic cancer. We investigated whether calpeptin inhibited tumor progression using a mouse xenograft model. We used quantitative RT-PCR to evaluate the expression of calpain-1 and calpain-2 mRNA in pancreatic cancer cells (PCCs) and pancreatic stellate cells (PSCs). We also undertook functional assays, including proliferation, migration, and invasion, to evaluate the inhibitory effects of calpeptin on PCCs and PSCs. Quantitative RT-PCR indicated that PCCs and PSCs expressed calpain-2 mRNA. Calpeptin reduced tumor volume (P = 0.0473) and tumor weight (P = 0.0471) and inhibited the tumor desmoplastic reaction (P < 0.001) in xenograft tumors in nude mice. Calpeptin also inhibited the biologic functions of PCCs and PSCs including proliferation (P = 0.017), migration (P = 0.027), and invasion (P = 0.035) in vitro. Furthermore, calpeptin reduced the migration of PCCs and PSCs by disrupting the cancer-stromal interaction (P = 0.0002). Our findings indicate that calpeptin is a promising antitumor agent for pancreatic cancer, due not only to its suppressive effect on PCCs and PSCs but also its disruption of the cancer-stromal interaction.

Topics: Animals; Calpain; Cell Communication; Cell Line, Tumor; Cell Movement; Cell Proliferation; Dipeptides; Disease Models, Animal; Gene Expression; Humans; Mice; Pancreatic Neoplasms; Pancreatic Stellate Cells; RNA, Messenger; Stromal Cells; Tumor Burden; Xenograft Model Antitumor Assays

Pancreatic cancer, including cancer of the ampulla of Vater and bile duct, is very aggressive and has a poor five year survival rate; improved methods of patient stratification are required.. We assessed the expression of calpain-1, calpain-2 and calpastatin in two patient cohorts using immunohistochemistry on tissue microarrays. The first cohort was composed of 68 pancreatic adenocarcinomas and the second cohort was composed of 120 cancers of the bile duct and ampulla.. In bile duct and ampullary carcinomas an association was observed between cytoplasmic calpastatin expression and patient age (P = 0.036), and between nuclear calpastatin expression and increased tumour stage (P = 0.026) and the presence of vascular invasion (P = 0.043). In pancreatic cancer, high calpain-2 expression was significantly associated with improved overall survival (P = 0.036), which remained significant in multivariate Cox-regression analysis (hazard ratio = 0.342; 95% confidence interva l = 0.157-0.741; P = 0.007). In cancers of the bile duct and ampulla, low cytoplasmic expression of calpastatin was significantly associated with poor overall survival (P = 0.012), which remained significant in multivariate Cox-regression analysis (hazard ratio = 0.595; 95% confidence interval = 0.365-0.968; P = 0.037).. The results suggest that calpain-2 and calpastatin expression is important in pancreatic cancers, influencing disease progression. The findings of this study warrant a larger follow-up study.

Topics: Adult; Aged; Aged, 80 and over; Ampulla of Vater; Bile Duct Neoplasms; Biomarkers, Tumor; Calcium-Binding Proteins; Calpain; Carcinoma; Cohort Studies; Disease Progression; Female; Humans; Immunohistochemistry; Kaplan-Meier Estimate; Male; Middle Aged; Pancreatic Neoplasms; Proportional Hazards Models; Tissue Array Analysis

The objective of this study was to test the association between calpain-10 (CAPN10), a diabetes susceptibility gene, with risk of pancreatic cancer (PC).. DNA samples from 83 incident exocrine PC cases and 166 controls, all of whom were smokers, were genotyped for four markers of CAPN10 in a nested case-control study based on the Beta-Carotene and Retinol Efficacy Trial (CARET), a randomized chemoprevention trial of subjects at high risk of lung cancer. Controls were matched on sex, race, age, CARET intervention arm, duration of exposure to asbestos, and smoking history. Conditional logistic regression was used for statistical analyses.. The minor allele of SNP-43 (rs3792267) in intron 3 was associated with increased risk of PC with an odds ratio of 1.57 (95%CI 1.03-2.38, p = 0.035) per allele. The three markers of the highest risk haplotype had an odds ratio of 1.98 (95%CI 1.12-3.49, p = 0.019) for risk of PC compared to the most common haplotype. There was no evidence of interaction between either of these associations by diabetes status.. These results suggest that variation in CAPN10 may be associated with increased risk of PC among smokers. Thus, studies of genes associated with diabetes risk in PC are warranted in a larger population.

Topics: Aged; Calpain; Case-Control Studies; Diabetes Mellitus, Type 2; Female; Genetic Predisposition to Disease; Genotype; Humans; Male; Middle Aged; Pancreatic Neoplasms; Polymorphism, Single Nucleotide; Reverse Transcriptase Polymerase Chain Reaction; Smoking

Calpain-10 was the first gene to be identified influencing the risk of type 2 diabetes (T2D) by positioning cloning. Studies in beta-cell lines and rodent islets suggest that calpain-10 may act as a regulator of insulin secretion. However, its role in human pancreatic islets remains unclear. The aim of this study was to examine if calpain-10 expression is altered in islets from patients with T2D and if the transcript level correlates with insulin release. We also tested if polymorphisms in the CAPN10 gene are associated with gene expression and insulin secretion in vitro.. Calpain-10 mRNA expression was analysed in human pancreatic islets from 34 non-diabetic and 10 T2D multi-organ donors. CAPN10 SNP-43 and SNP-44 were genotyped and related to gene expression and insulin release in response to glucose, arginine and glibenclamide. The mRNA level of calpain-10 was elevated by 64% in pancreatic islets from patients with T2D compared with non-diabetic donors (P = 0.01). Moreover, the calpain-10 expression correlated positively with arginine-stimulated insulin release in islets from non-diabetic donors (r = 0.45, P = 0.015). However, this correlation was lost in islets from patients with T2D (r = 0.09; P = 0.8). The G/G variant of SNP-43 was associated with reduced insulin release in response to glucose (P

Topics: Arginine; Base Sequence; Calpain; Case-Control Studies; Diabetes Mellitus, Type 2; DNA Primers; Glucose; Glyburide; Humans; Insulin; Insulin Secretion; Islets of Langerhans; Pancreatic Neoplasms; Polymorphism, Single Nucleotide; Reverse Transcriptase Polymerase Chain Reaction; RNA, Messenger

Calpain-10 (CAPN10) has been identified as a diabetes susceptibility gene. Previous studies have shown that alterations in calpain activity alter both glucose uptake and insulin secretion. In this report, we investigated the role of calpain activity in the actin reorganization required for glucose-stimulated insulin secretion. In pancreatic INS-1 cells, acute exposure to a high glucose environment stimulated CAPN10 gene expression with a concomitant increase in calpain activity. However, high glucose did not significantly alter expression of the two major ubiquitously expressed calpain family members, CAPN1 and CAPN2. Furthermore, glucose stimulation resulted in the reorganization of actin and inhibition of calpain activity impaired this reorganization in INS-1 cells. Finally, we identified a 54 kDa isoform as the major CAPN10 isoform that associates with the actin cytoskeleton. Based on our findings, we propose that calpain plays a role in facilitating the actin reorganization required for glucose-stimulated insulin secretion in INS-1 cells.

Topics: Actins; Animals; Calpain; Cell Line, Tumor; Dose-Response Relationship, Drug; Glucose; Insulin; Insulin Secretion; Insulinoma; Pancreatic Neoplasms; Rats; Signal Transduction

Ordered cell cycle progression requires the expression and activation of several cyclins and cyclin-dependent kinases (Cdks). Hyperosmotic stress causes growth arrest possibly via proteasome-mediated degradation of cyclin D1. We studied the effect of hyposmotic conditions on three colonic (Caco2, HRT18, HT29) and two pancreatic (AsPC-1 and PaCa-2) cell lines. Hyposmosis caused reversible cell growth arrest of the five cell lines in a cell cycle-independent fashion, although some cell lines accumulated at the G(1)/S interface. Growth arrest was followed by apoptosis or by formation of multinucleated giant cells, which is consistent with cell cycle catastrophe. Hyposmosis dramatically decreased Cdc2, Cdk2, Cdk4, cyclin B1, and cyclin D3 expression in a time-dependent fashion, in association with an overall decrease in cellular protein synthesis. However, some protein levels remained unaltered, including cyclin E and keratin 8. Selective proteasome inhibition prevented Cdk and cyclin degradation and reversed hyposmotic stress-induced growth arrest, whereas calpain and lysosome enzyme inhibitors had no measurable effect on cell cycle protein degradation. Therefore, hyposmotic stress inhibits cell growth and, depending on the cell type, causes cell cycle catastrophe with or without apoptosis. The growth arrest is due to decreased protein synthesis and proteasome activation, with subsequent degradation of several cyclins and Cdks.

Topics: Apoptosis; Calpain; CDC2 Protein Kinase; CDC2-CDC28 Kinases; Cell Survival; Colonic Neoplasms; Cyclin B; Cyclin B1; Cyclin D1; Cyclin D3; Cyclin-Dependent Kinase 2; Cyclin-Dependent Kinase 4; Cyclin-Dependent Kinases; Cyclins; Cysteine Endopeptidases; DNA Fragmentation; Flow Cytometry; G1 Phase; Giant Cells; Humans; Keratins; Lysosomes; Microscopy, Electron; Models, Biological; Multienzyme Complexes; Osmosis; Osmotic Pressure; Pancreatic Neoplasms; Proteasome Endopeptidase Complex; Protein Serine-Threonine Kinases; Proto-Oncogene Proteins; S Phase; Time Factors; Tumor Cells, Cultured

Forty loci (16 polymorphic and 24 non-polymorphic) together with 23 cosmids isolated from a chromosome 11-specific library were used to construct a detailed genetic map of 11p13-11q13. The map was constructed by using a panel of 13 somatic cell hybrids that sub-divided this region into 19 intervals, a meiotic mapping panel of 33 multiple endocrine neoplasia type 1 (MEN1) families (134 affected and 269 unaffected members) and a mitotic mapping panel that was used to identify loss of heterozygosity in 38 MEN1-associated tumours. The results defined the most likely order of the 16 loci as being: 11pter-D11S871-(D11S288, D11S149)-11cen-CNTF-PGA-ROM1-D11S480-PYGM- SEA-D11S913-D11S970-D11S97- D11S146-INT2-D11S971-D11S533-11qter. The meiotic mapping studies indicated that the most likely location of the MEN1 gene was in the interval flanked by PYGM and D11S97, and the results of mitotic mapping suggested a possible location of the MEN1 gene telomeric to SEA. Mapping studies of the gene encoding mu-calpain (CAPN1) located CAPN1 to 11q13 and in the vicinity of the MEN1 locus. However, mutational analysis studies did not detect any germ-line CAPN1 DNA sequence abnormalities in 47 unrelated MEN1 patients and the results therefore exclude CAPN1 as the MEN1 gene. The detailed genetic map that has been constructed of the 11p13-11q13 region should facilitate the construction of a physical map and the identification of candidate genes for disease loci mapped to this region.

Topics: Animals; Base Sequence; Calpain; Chromosome Mapping; Chromosomes, Human, Pair 11; Cosmids; Female; Gastrinoma; Genetic Linkage; Germ-Line Mutation; Humans; Hybrid Cells; Insulinoma; Male; Meiosis; Mitosis; Molecular Sequence Data; Multiple Endocrine Neoplasia Type 1; Pancreatic Neoplasms; Parathyroid Neoplasms; Pedigree; Pituitary Neoplasms; Polymorphism, Genetic; Sequence Deletion