MUC4 activates HER2 signalling and enhances the motility of human ovarian cancer cells

Department of Biochemistry and Molecular Biology, University of Nebraska Medical Center, Omaha, NE 68198-5870, USA.
British Journal of Cancer (Impact Factor: 4.84). 09/2008; 99(3):520-6. DOI: 10.1038/sj.bjc.6604517
Source: PubMed


The mucin MUC4 is a high molecular weight transmembrane glycoprotein. It consists of a mucin-type subunit (MUC4alpha) and a transmembrane growth factor-like subunit (MUC4beta). The mucin MUC4 is overexpressed in many epithelial malignancies including ovarian cancer, suggesting a possible role in the pathogenesis of these cancers. In this study, we investigated the functional role of MUC4 in the human ovarian cancer cell line SKOV3. The mucin MUC4 was ectopically expressed by stable transfection, and its expression was examined by western blot and confocal microscopy analyses. The in vitro studies demonstrated an enhanced motility of MUC4-expressing SKOV3 cells compared with the vector-transfected cells. The mucin MUC4 expression was associated with apparent changes in actin organisation, leading to the formation of microspike, lammelopodia and filopodia-like cellular projections. An enhanced protein expression and activation of HER2, a receptor tyrosine kinase, was also seen, although no significant change was observed in HER-2 transcript levels in the MUC4-transfected SKOV3 cells. Reciprocal co-immunoprecipitation revealed an interaction of MUC4 with HER2. Further, the MUC4-overexpressing SKOV3 cells exhibited an increase in the phosphorylation of focal adhesion kinase (FAK), Akt and ERK, downstream effectors of HER2. Taken together, our findings demonstrate that MUC4 plays a role in ovarian cancer cell motility, in part, by altering actin arrangement and potentiating HER2 downstream signalling in these cells.

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Available from: Ajay P Singh
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    • "This binding domain may facilitate the adhesion of both cell types, and thus, has major implications for EOC tumorigenesis [37]. Interestingly, MUC4, a transmembrane mucin that is overexpressed in ovarian tumors, which has been shown to increase the motility and invasiveness of ovarian cancer cells through the induction of epithelial-to-mesenchymal transition was also elevated in our co-cultures [38] [39] [40]. To date, few studies have evaluated the pathophysiological association of MUC5AC with respect to ovarian cancer, as aberrant expression has mainly been linked to colorectal, pancreatic, and gastric carcinomas, in addition to the regulation of airway epithelial cells [33] [34]. "
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    ABSTRACT: Unlabelled: Ovarian cancer is a highly metastatic disease that is often characterized by widespread abdominal dissemination. A hallmark of ovarian cancer progression is the attachment of malignant cells to the mesothelium and the formation of invasive peritoneal implants. Therefore, delineating factors involved in cancer-peritoneal cell interaction is critical to improving patient survival, as it may lead to the discovery of novel therapeutic targets. As such, we aimed to identify proteins that participate in this interaction by comparing the secreted proteome of a co-culture model containing ovarian cancer (OVCAR-5) and mesothelial cells (LP-9), to their respective monoculture secretomes. In total, 49 proteins were differentially secreted during cancer and mesothelial cell contact. Relative mRNA expression of candidates was performed, which revealed a significant increase in MUC5AC gene expression in cancer cells cultured in three different co-culture models (OVCAR-5 and LP-9; BG-1 and LP-9; OV-90 and LP-9). An increased expression was also observed in LP-9 cells that were co-cultured with OVCAR-5 and OV-90 cancer cells. Further immunocytochemistry analysis also confirmed increased expression of MUC5AC in ovarian cancer and peritoneal co-cultures. Overall, our analysis uncovers novel molecular markers of peritoneal metastasis, which may have potential roles in regulating the progression of the disease. Biological significance: In this study, our objective was to focus on identifying novel mediators of ovarian cancer and peritoneal interaction using a mass spectrometry-based approach. Our analysis resulted in the discovery of both previously known and novel factors involved this interaction, and as such, these newly discovered proteins might have potential roles in cancer progression, such as invasion and adhesion. We believe that these findings add to our current knowledge and understanding of ovarian cancer progression, and will aid researchers in their future attempts in finding new targets of the disease.
    Full-text · Article · Apr 2014 · Journal of proteomics
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    • "FAK additionally serves as a key effector for erbB receptors, with the EGF being unable to phosphorylate the EGFR and promote migration in cells lacking FAK (Sieg et al. 2000), while heregulin/HER2 stimulation results in altered FAK phosphorylation and modification of focal contacts and cell-matrix adhesion (Vadlamudi et al. 2003). Moreover, FAK is key to the transduction of HER2 signalling in HER2-overexpressing MCF10A breast cells (Wang et al. 2005) and in MUC4-overexpressing ovarian cancer cells, which have elevated HER2 activity (Ponnusamy et al. 2008). FAK has been reported to be of prognostic significance in lung and oesophageal tumours (Itoh et al. 2004), and in breast cancer, levels of FAK (PTK2) expression correlate strongly with poor tumour differentiation and are significantly associated with HER2 overexpression (Schmitz et al. 2005). "
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    ABSTRACT: The HER2 transmembrane receptor is a well-characterized predictive marker for trastuzumab benefit and may be associated with decreased benefit from endocrine therapy use. Despite the clinical effectiveness of anti-HER2 agents, resistance represents a significant limiting factor. Focal adhesion kinase (FAK) plays an important role in HER2 signalling, mediating downstream Akt activation in addition to HER2 cross talk with other growth factor receptors. In this study, we have investigated the therapeutic potential of FAK in ER+/HER2+ breast cancer using the novel FAK-specific inhibitor, PF4554878 ('PF878'). FAK/HER2 signalling pathway activation was assessed in ER+/HER2- (MCF7, T47D) and ER+/HER2+ (BT474, MDAMB361) breast cancer cells in the presence or absence of PF878 and ± trastuzumab. The effects of PF878 on cell growth as a monotherapy and in combination with trastuzumab were assessed using MTT and coulter counting with isobologram analysis to determine synergy/additive effects. FAK activation (Y861 but not Y397) was highest in ER+/HER2+ cells that also demonstrated the greatest sensitivity to PF878. As a monotherapy, PF878 prevented heregulin-induced MDA361 cell migration but had no significant effect on cell growth. Treatment of ER+/HER2+ cells with PF878 and trastuzumab combined resulted in synergistic inhibition of cell proliferation. Underlying this was an abrogation of Akt activity and increased PARP cleavage, effects that were greatest in trastuzumab-refractory MDA361 cells. Collectively, these data support a role for FAK in ER+/HER2+ breast cancer where its targeting has the potential to improve trastuzumab response. This is particularly important in the context of ER+/HER2+, trastuzumab-refractory disease, where FAK inhibition may present an important strategy to restore trastuzumab sensitivity.
    Full-text · Article · Jul 2013 · Endocrine Related Cancer
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    • "However, aberrant expression of mucins has been reported to promote cancer development, and affects cellular growth, transformation, and invasion [7]. Aberrantly over-expressed membrane-tethered mucins, including MUC1 [8] and MUC4 [6], play diverse functional roles in several epithelial cancers, including ovarian, pancreatic [9], [10], and breast [11]. We have previously demonstrated that MUC4 enhances tumorigenicity and metastasis in pancreatic [7], [12] and ovarian [10] cancer. "
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    ABSTRACT: Current studies indicate that triple negative breast cancer (TNBC), an aggressive breast cancer subtype, is associated with poor prognosis and an early pattern of metastasis. Emerging evidence suggests that MUC4 mucin is associated with metastasis of various cancers, including breast cancer. However, the functional role of MUC4 remains unclear in breast cancers, especially in TNBCs. In the present study, we investigated the functional and mechanistic roles of MUC4 in potentiating pathogenic signals including EGFR family proteins to promote TNBC aggressiveness using in vitro and in vivo studies. Further, we studied the expression of MUC4 in invasive TNBC tissue and normal breast tissue by immunostaining. MUC4 promotes proliferation, anchorage-dependent and-independent growth of TNBC cells, augments TNBC cell migratory and invasive potential in vitro, and enhances tumorigenicity and metastasis in vivo. In addition, our studies demonstrated that MUC4 up-regulates the EGFR family of proteins, and augments downstream Erk1/2, PKC-γ, and FAK mediated oncogenic signaling. Moreover, our studies also showed that knockdown of MUC4 in TNBC cells induced molecular changes suggestive of mesenchymal to epithelial transition. We also demonstrated in this study, for the first time, that knockdown of MUC4 was associated with reduced expression of EGFR and ErbB3 (EGFR family proteins) in TNBC cells, suggesting that MUC4 uses an alternative to ErbB2 mechanism to promote aggressiveness. We further demonstrate that MUC4 is differentially over-expressed in invasive TNBC tissues compared to normal breast tissue. MUC4 mucin expression is associated with TNBC pathobiology, and its knockdown reduced aggressiveness in vitro, and tumorigenesis and metastasis in vivo. Overall, our findings suggest that MUC4 mucin promotes invasive activities of TNBC cells by altering the expression of EGFR, ErbB2, and ErbB3 molecules and their downstream signaling.
    Full-text · Article · Feb 2013 · PLoS ONE
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