MUC4 mucin expression in human pancreatic tumours is affected by organ environment: the possible role of TGFbeta2.
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ABSTRACT:
SUBMITTER: Choudhury A
PROVIDER: S-EPMC2409611 | biostudies-other | 2004 Feb
REPOSITORIES: biostudies-other
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Project description:Pancreatic cancer (PC) is a highly lethal malignancy with a dismal five-year survival rate. This is due to its asymptomatic nature, lack of reliable biomarkers, poor resectability, early metastasis, and high recurrence rate. Limited efficacies of current treatment modalities treatment-associated toxicity underscore the need for the development of immunotherapy-based approaches. For non-resectable, locally advanced metastatic PC, immunotherapy-based approaches including vaccines, antibody-targeted, immune checkpoint inhibition, CAR-T-cells, and adoptive T-cell transfer could be valuable additions to existing treatment modalities. Thus far, the vaccine candidates in PC have demonstrated modest immunological responses in different treatment modalities. The identification of tumor-associated antigens (TAA) and their successful implication in PC treatment is still a challenge. MUC4, a high molecular weight glycoprotein that functionally contributes to PC pathogenesis, is an attractive TAA. It is not detected in the normal pancreas; however, it is overexpressed in mouse and human pancreatic tumors. The recombinant MUC4 domain, as well as predicted immunogenic T-cell epitopes, elicited cellular and humoral anti-MUC4 response, suggesting its ulility as a vaccine candidate for PC therapy. Existence of PC-associated MUC4 splice variants, autoantibodies against overexpressed and aberrantly glycosylated MUC4 and presence of T-cell clones against the mutations present in MUC4 further reinforce its significance as a tumor antigen for vaccine development. Herein, we review the significance of MUC4 as a tumor antigen in PC immunotherapy and discuss both, the development and challenges associated with MUC4 based immunotherapy. Lastly, we will present our perspective on MUC4 antigenicity for the future development of MUC4-based PC immunotherapy.
| S-EPMC7160012 | biostudies-literature
Project description:Background and objectiveSurgery carries the best hope for cure in the treatment of cholangiocarcinoma (CC), whereas surgical outcome is not fully satisfactory. Bio-molecular markers have been used to improve tumor staging and prognosis prediction. Mucin antigen MUC4 (MUC4) has been implicated as a marker for poor survival in various tumors. However, prognostic significance of MUC4 for patients with CC remains undefined. The aim of the present meta-analysis was to investigate the association between MUC4 expression and overall survival (OS) of patients with resected CC.MethodsThe meta-analysis was conducted in adherence to the MOOSE guidelines. PubMed, Embase databases, Cochrane Library and the Chinese SinoMed were systematically searched to identify eligible studies from the initiation of the databases to April, 2016. OSs were pooled by using hazard ratio (HR) with corresponding 95% confidence interval (CI). Random effect models were utilized because of the between-study heterogeneities.ResultsFive studies reporting on 249 patients were analyzed: 94 (37.75%) were in positive or high expression group and 155 (62.25%) in negative or low expression group. The pooled HR for positive or high expression group was found to be 3.04 (95% CI 2.25-4.12) when compared with negative or low expression group with slight between-study heterogeneities (I2 3.10%, P = 0.39). The result indicated that a positive or high expression level of MUC4 was significantly related to poor survival in patients with resected CC. A commensurate result was identified by sensitivity analysis. The main limitations of the present meta-analysis were the rather small size of the studies included and relatively narrow geographical distribution of population.ConclusionThe result of this meta-analysis indicated that a positive or high expression level of MUC4 was significantly related to poor survival in patients with resected CC.
| S-EPMC4909222 | biostudies-literature
Project description:MUC4 is a type-1 transmembrane glycoprotein and is overexpressed in many carcinomas. It is a heterodimeric protein of 930 kDa, composed of a mucin-type subunit, MUC4alpha, and a membrane-bound growth factor-like subunit, MUC4beta. MUC4 mRNA contains unique 5' and 3' coding sequences along with a large variable number of tandem repeat (VNTR) domain of 7-19 kb. A direct association of MUC4 overexpression has been established with the degree of invasiveness and poor prognosis of pancreatic cancer. To understand the precise role of MUC4 in pancreatic cancer, we engineered a MUC4 complementary DNA construct, mini-MUC4, whose deduced protein (320 kDa) is comparable with that of wild-type MUC4 (930 kDa) but represents only 10% of VNTR. Stable ectopic expression of mini-MUC4 in two human pancreatic cancer cell lines, Panc1 and MiaPaCa, showed that MUC4 minigene expression follows a biosynthesis and localisation pattern similar to the wild-type MUC4. Expression of MUC4 resulted in increased growth, motility, and invasiveness of the pancreatic cancer cells in vitro. Ultra-structural examination of MUC4-transfected cells showed the presence of increased number and size of mitochondria. The MUC4-expressing cells also demonstrated an enhanced tumorigenicity in an orthotopic xenograft nude mice model, further supporting a direct role of MUC4 in inducing the cancer properties. In conclusion, our results suggest that MUC4 promotes tumorigenicity and is directly involved in growth and survival of the cancer cells.
| S-EPMC2360313 | biostudies-literature
Project description:AimsTo examine how accurately immunohistochemical markers discriminate between pancreatobiliary and intestinal-type adenocarcinomas in the pancreatic head and to explore the prognostic importance of these markers among each of these histological types.Methods and resultsHistopathological features of 114 consecutively resected adenocarcinomas of pancreatobiliary (n = 67) and intestinal (n = 47) type of differentiation were recorded according to a standardized protocol. Immunohistochemistry for cytokeratin (CK) 7, CK20, MUC1, MUC2, MUC4 and CDX2 was performed on tissue microarrays. Classification of the adenocarcinomas based on immunohistochemistry was compared with the morphological evaluation of histological type. Presence of CK7 and MUC4, and absence of CDX2, were independent predictors of pancreatobiliary versus intestinal type. Using these markers to optimize immunohistochemical classification, agreement between immunohistochemical and morphological classification was only moderate (kappa = 0.53). In pancreatobiliary differentiated tumours, MUC1 and/or MUC4 expression was an independent prognostic factor (hazard ratio 2.02, 95% confidence interval 1.02, 3.98) when adjusting for nodal involvement, vessel involvement and tumour size. In intestinally differentiated tumours, none of the markers was significantly associated with prognosis.ConclusionsAgreement between immunohistochemical and morphological classification of pancreatic head adenocarcinomas is moderate. In pancreatobiliary adenocarcinomas, MUC1 and/or MUC4 expression indicates a particularly poor prognosis.
| S-EPMC2680278 | biostudies-literature
Project description:Aberrant Wnt signaling frequently occurs in pancreatic cancer (PC) and contributes to disease progression/metastases. Likewise, the transmembrane-mucin MUC4 is expressed de novo in early pancreatic intraepithelial neoplasia (PanINs) and incrementally increases with PC progression, contributing to metastasis. To determine the mechanism of MUC4 upregulation in PC, we examined factors deregulated in early PC progression, such as Wnt/β-catenin signaling. MUC4 promoter analysis revealed the presence of three putative TCF/LEF-binding sites, leading us to hypothesize that MUC4 can be regulated by β-catenin. Immunohistochemical (IHC) analysis of rapid autopsy PC tissues showed a correlation between MUC4 and cytosolic/nuclear β-catenin expression. Knock down (KD) of β-catenin in CD18/HPAF and T3M4 cell lines resulted in decreased MUC4 transcript and protein. Three MUC4 promoter luciferase constructs, p3778, p3000, and p2700, were generated. The construct p3778, encompassing the entire MUC4 promoter, elicited increased luciferase activity in the presence of stabilized β-catenin. Mutation of the TCF/LEF site closest to the transcription start site (i.e., -2629/-2612) and furthest from the start site (i.e., -3425/-3408) reduced MUC4 promoter luciferase activity. Transfection with dominant negative TCF4 decreased MUC4 transcript and protein levels. Chromatin immunoprecipitation confirmed enrichment of β-catenin on -2629/-2612 and -3425/-3408 of the MUC4 promoter in CD18/HPAF. Functionally, CD18/HPAF and T3M4 β-catenin KD cells showed decreased migration and decreased Vimentin, N-cadherin, and pERK1/2 expression. Tumorigenicity studies in athymic nude mice showed CD18/HPAF β-catenin KD cells significantly reduced primary tumor sizes and metastases compared to scrambled control cells. We show for the first time that β-catenin directly governs MUC4 in PC.
| S-EPMC4740200 | biostudies-literature
Project description:MUC4 is a large transmembrane type I glycoprotein that is overexpressed in pancreatic cancer (PC) and has been shown to be associated with its progression and metastasis. However, the exact cellular and molecular mechanism(s) through which MUC4 promotes metastasis of PC cells has been sparsely studied. Here we showed that the nidogen-like (NIDO) domain of MUC4, which is similar to the G1-domain present in the nidogen or entactin (an extracellular matrix protein), contributes to the protein-protein interaction property of MUC4. By this interaction, MUC4 promotes breaching of basement membrane (BM) integrity, and spreading of cancer cells. These observations are corroborated with the data from our study using an engineered MUC4 protein without the NIDO domain, which was ectopically expressed in the MiaPaCa PC cells, lacking endogenous MUC4 and nidogen protein. The in vitro studies demonstrated an enhanced invasiveness of MiaPaCa cells expressing MUC4 (MiaPaCa-MUC4) compared with vector-transfected cells (MiaPaCa-Vec; P=0.003) or cells expressing MUC4 without the NIDO domain (MiaPaCa-MUC4-NIDO(?); P=0.03). However, the absence of NIDO-domain has no significant role on cell growth and motility (P=0.93). In the in vivo studies, all the mice orthotopically implanted with MiPaCa-MUC4 cells developed metastasis to the liver as compared with MiaPaCa-Vec or the MiaPaCa-MUC4-NIDO(?) group, hence, supporting our in vitro observations. Additionally, a reduced binding (P=0.0004) of MiaPaCa-MUC4-NIDO(?) cells to the fibulin-2 coated plates compared with MiaPaCa-MUC4 cells indicated a possible interaction between the MUC4-NIDO domain and fibulin-2, a nidogen-interacting protein. Furthermore, in PC tissue samples, MUC4 colocalized with the fibulin-2 present in the BM. Altogether, our findings demonstrate that the MUC4-NIDO domain significantly contributes to the MUC4-mediated metastasis of PC cells. This may be partly due to the interaction between the MUC4-NIDO domain and fibulin-2.
| S-EPMC3298579 | biostudies-literature
Project description:The MUC4 mucin is a high molecular weight, membrane-bound, and highly glycosylated protein. It is a multi-domain protein that is putatively cleaved into a large mucin-like subunit (MUC4?) and a C-terminal growth-factor like subunit (MUC4?). MUC4 plays critical roles in physiological and pathological conditions and is aberrantly overexpressed in several cancers, including those of the pancreas, cervix, breast and lung. It is also a potential biomarker for the diagnosis, prognosis and progression of several malignancies. Further, MUC4 plays diverse functional roles in cancer initiation and progression as evident from its involvement in oncogenic transformation, proliferation, inhibition of apoptosis, motility and invasion, and resistance to chemotherapy in human cancer cells. We have previously generated a monoclonal antibody 8G7, which is directed against the TR region of MUC4, and has been extensively used to study the expression of MUC4 in several malignancies. Here, we describe the generation of anti-MUC4 antibodies directed against the non-TR regions of MUC4. Recombinant glutathione-S-transferase (GST)-fused MUC4? fragments, both upstream (MUC4?-N-Ter) and downstream (MUC4?-C-Ter) of the TR domain, were used as immunogens to immunize BALB/c mice. Following cell fusion, hybridomas were screened using the aforementioned recombinant proteins ad lysates from human pancreatic cell lines. Three anti MUC4?-N-Ter and one anti-MUC4?-C-Ter antibodies were characterized by several inmmunoassays including enzyme-linked immunosorbent assay (ELISA), immunoblotting, immunofluorescene, flow cytometry and immunoprecipitation using MUC4 expressing human pancreatic cancer cell lines. The antibodies also reacted with the MUC4 in human pancreatic tumor sections in immunohistochemical analysis. The new domain-specific anti-MUC4 antibodies will serve as important reagents to study the structure-function relationship of MUC4 domains and for the development of MUC4-based diagnostics and therapeutics.
| S-EPMC3160300 | biostudies-literature
Project description:Mucins are high-molecular-weight glycoproteins and are implicated in diverse biological functions. MUC4, a member of transmembrane mucin family, is expressed in airway epithelial cells and body fluids like saliva, tear film, ear fluid, and breast milk. In addition to its normal expression, an aberrant expression of MUC4 has been reported in a variety of carcinomas. Among various potential domains of MUC4, epidermal growth factor (EGF) -like domains are hypothesized to interact with and activate the ErbB2 receptors, suggesting an intramembrane-growth factor function for MUC4. The heavily glycosylated tandem repeat domain provides the structural rigidity to the extended extracellular region. MUC4, by virtue of its extended structure, serves as a barrier for some cell-cell and cell-extracellular matrix interactions and as a potential reservoir for certain growth factors. An intricate relationship between MUC4 and growth factor signaling is also reflected in the transcriptional regulation of MUC4. The MUC4 promoter has binding sites for different transcription factors, which are responsible for the regulation of its expression in different tissues. The interferon-gamma, retinoic acid, and transforming growth factor-beta signaling pathways regulate MUC4 expression in a partially interdependent manner. Taken together, all of these features of MUC4 strongly support its role as a potential candidate for diagnostic and therapeutic applications in cancer and other diseases.
| S-EPMC2835492 | biostudies-literature
Project description:The fluorinated analog of deoxycytidine, Gemcitabine (Gemzar), is the main chemotherapeutic drug in pancreatic cancer, but survival remains weak mainly because of the high resistance of tumors to the drug. Recent works have shown that the mucin MUC4 may confer an advantage to pancreatic tumor cells by modifying their susceptibility to drugs. However, the cellular mechanism(s) responsible for this MUC4-mediated resistance is unknown. The aim of this work was to identify the cellular mechanisms responsible for gemcitabine resistance linked to MUC4 expression. CAPAN-2 and CAPAN-1 adenocarcinomatous pancreatic cancer (PC) cell lines were used to establish stable MUC4-deficient clones (MUC4-KD) by shRNA interference. Measurement of the IC50 index using tetrazolium salt test indicated that MUC4-deficient cells were more sensitive to gemcitabine. This was correlated with increased Bax/BclXL ratio and apoptotic cell number. Expression of Equilibrative/Concentrative Nucleoside Transporter (hENT1, hCNT1/3), deoxycytidine kinase (dCK), ribonucleotide reductase (RRM1/2) and Multidrug-Resistance Protein (MRP3/4/5) was evaluated by quantitative RT-PCR (qRT-PCR) and western blotting. Alteration of MRP3, MRP4, hCNT1 and hCNT3 expression was observed in MUC4-KD cells, but only hCNT1 alteration was correlated to MUC4 expression and sensitivity to gemcitabine. Decreased activation of MAPK, JNK and NF-?B pathways was observed in MUC4-deficient cells, in which the NF-?B pathway was found to have an important role in both sensitivity to gemcitabine and hCNT1 regulation. Finally, and in accordance with our in vitro data, we found that MUC4 expression was conversely correlated to that of hCNT1 in tissues from patients with pancreatic adenocarcinoma. This work describes a new mechanism of PC cell resistance to gemcitabine, in which the MUC4 mucin negatively regulates the hCNT1 transporter expression via the NF-?B pathway. Altogether, these data point out to MUC4 and hCNT1 as potential targets to ameliorate the response of pancreatic tumors to gemcitabine treatment.
| S-EPMC3936121 | biostudies-literature
Project description:MUC4 is a transmembrane mucin expressed on various epithelial surfaces, including respiratory and gastrointestinal tracts, and helps in their lubrication and protection. MUC4 is also aberrantly overexpressed in various epithelial malignancies and functionally contributes to cancer development and progression. MUC4 is putatively cleaved at the GDPH site into a mucin-like α-subunit and a membrane-tethered growth factor-like β-subunit. Due to the presence of several functional domains, the characterization of MUC4β is critical for understanding MUC4 biology. We developed a method to produce and purify multi-milligram amounts of recombinant MUC4β (rMUC4β). Purified rMUC4β was characterized by Far-UV CD and I-TASSER-based protein structure prediction analyses, and its ability to interact with cellular proteins was determined by the affinity pull-down assay. Two of the three EGF-like domains exhibited typical β-fold, while the third EGF-like domain and vWD domain were predominantly random coils. We observed that rMUC4β physically interacts with Ezrin and EGFR family members. Overall, this study describes an efficient and simple strategy for the purification of biologically-active rMUC4β that can serve as a valuable reagent for a variety of biochemical and functional studies to elucidate MUC4 function and generating domain-specific antibodies and vaccines for cancer immunotherapy.
| S-EPMC8660890 | biostudies-literature