Jeroen Vermeulen

COMMD1 disrupts HIF-1α/β dimerization and inhibits human tumor cell invasion

The gene encoding COMM domain-containing 1 (COMMD1) is a prototypical member of the COMMD gene family that has been shown to inhibit both NF-kappaB- and HIF-mediated gene expression. NF-kappaB and HIF are transcription factors that have been shown to play a role in promoting tumor growth, survival, and invasion. In this study, we demonstrate that COMMD1 expression is frequently suppressed in human cancer and that decreased COMMD1 expression correlates with a more invasive tumor phenotype. We found that direct repression of COMMD1 in human cell lines led to increased tumor invasion in a chick xenograft model, while increased COMMD1 expression in mouse melanoma cells led to decreased lung metastasis in a mouse model. Decreased COMMD1 expression also correlated with increased expression of genes known to promote cancer cell invasiveness, including direct targets of HIF. Mechanistically, our studies show that COMMD1 inhibits HIF-mediated gene expression by binding directly to the amino terminus of HIF-1alpha, preventing its dimerization with HIF-1beta and subsequent DNA binding and transcriptional activation. Altogether, our findings demonstrate a role for COMMD1 in tumor invasion and provide a detailed mechanism of how this factor regulates the HIF pathway in cancer cells.

Participation in sport: bonding and bridging as identity work

Nowadays sport is assigned a crucial role in solving social problems, especially those relating to social cohesion. Participation in sport is assumed to build relevant bonding and bridging social capital that generates reciprocal contacts and trust in others. In this essay we will present findings of two, mainly qualitative studies on participation in sport in the Netherlands. We argue that while sport indeed makes contributions to the development of social capital, bonding and bridging are much more complex and differentiated processes than is usually assumed in both social policies and social capital theory. An argument is made to view bonding and bridging as identity work.

Nuclear Kaiso Expression Is Associated with High Grade and Triple-Negative Invasive Breast Cancer

Kaiso is a BTB/POZ transcription factor that is ubiquitously expressed in multiple cell types and functions as a transcriptional repressor and activator. Little is known about Kaiso expression and localization in breast cancer. Here, we have related pathological features and molecular subtypes to Kaiso expression in 477 cases of human invasive breast cancer. Nuclear Kaiso was predominantly found in invasive ductal carcinoma (IDC) (p = 0.007), while cytoplasmic Kaiso expression was linked to invasive lobular carcinoma (ILC) (p = 0.006). Although cytoplasmic Kaiso did not correlate to clinicopathological features, we found a significant correlation between nuclear Kaiso, high histological grade (p = 0.023), ERα negativity (p = 0.001), and the HER2-driven and basal/triple-negative breast cancers (p = 0.018). Interestingly, nuclear Kaiso was also abundant in BRCA1-associated breast cancer (p<0.001) and invasive breast cancer overexpressing EGFR (p = 0.019). We observed a correlation between nuclear Kaiso and membrane-localized E-cadherin and p120-catenin (p120) (p<0.01). In contrast, cytoplasmic p120 strongly correlated with loss of E-cadherin and low nuclear Kaiso (p = 0.005). We could confirm these findings in human ILC cells and cell lines derived from conditional mouse models of ILC. Moreover, we present functional data that substantiate a mechanism whereby E-cadherin controls p120-mediated relief of Kaiso-dependent gene repression. In conclusion, our data indicate that nuclear Kaiso is common in clinically aggressive ductal breast cancer, while cytoplasmic Kaiso and a p120-mediated relief of Kaiso-dependent transcriptional repression characterize ILC.

Nuclear localization of the transcriptional coactivator YAP is associated with invasive lobular breast cancer

BackgroundYes Associated Protein (YAP) has been implicated in the control of organ size by regulating cell proliferation and survival. YAP is a transcriptional coactivator that controls cellular responses through interaction with TEAD transcription factors in the nucleus, while its transcriptional functions are inhibited by phosphorylation-dependent translocation to the cytosol. YAP overexpression has been associated with different types of cancer, such as lung, skin, prostate, ovary and liver cancer. Recently, YAP was linked to E-cadherin-dependent regulation of contact inhibition in breast cancer cells.ResultsIn this study we examined YAP protein expression and cellular localization in 237 cases of human invasive breast cancer by immunohistochemistry and related its expression to clinicopathological features and E-cadherin expression. We observed that invasive lobular carcinoma is characterized by higher expression levels of both nuclear and cytosolic YAP (p < 0.001). Nuclear YAP expression did not associate with other variables such as lymph node involvement, tumor grade, tumor size, mitotic activity or the molecular sub-types of invasive breast cancer. We observed that high nuclear and cytosolic YAP expression are associated with the E-cadherin deficient breast cancer subtype ILC (p < 0.001) and cell lines derived from human breast cancers and conditional mouse models of human lobular breast cancer.ConclusionsSince our data indicate that nuclear YAP localization is more common in breast cancers lacking functional adherens junctions, it suggests that YAP-mediated transcription may be involved in the development and progression of invasive lobular breast cancer.

Loss of p120-Catenin Induces Metastatic Progression of Breast Cancer by Inducing Anoikis Resistance and Augmenting Growth Factor Receptor Signaling

Metastatic breast cancer remains the chief cause of cancer-related death among women in the Western world. Although loss of cell-cell adhesion is key to breast cancer progression, little is known about the underlying mechanisms that drive tumor invasion and metastasis. Here, we show that somatic loss of p120-catenin (p120) in a conditional mouse model of noninvasive mammary carcinoma results in formation of stromal-dense tumors that resemble human metaplastic breast cancer and metastasize to lungs and lymph nodes. Loss of p120 in anchorage-dependent breast cancer cell lines strongly promoted anoikis resistance through hypersensitization of growth factor receptor (GFR) signaling. Interestingly, p120 deletion also induced secretion of inflammatory cytokines, a feature that likely underlies the formation of the prometastatic microenvironment in p120-negative mammary carcinomas. Our results establish a preclinical platform to develop tailored intervention regimens that target GFR signals to treat p120-negative metastatic breast cancers.

COMMD1 Promotes pVHL and O2-Independent Proteolysis of HIF-1α via HSP90/70

Background The Copper Metabolism MURR1 Domain containing 1 protein COMMD1 has been associated with copper homeostasis, NF-κB signaling, and sodium transport. Recently, we identified COMMD1 as a novel protein in HIF-1 signaling. Mouse embryos deficient for Commd1 have increased expression of hypoxia/HIF-regulated genes i.e. VEGF, PGK and Bnip3. Hypoxia-inducible factors (HIFs) are master regulators of oxygen homeostasis, which control angiogenesis, erythropoiesis, glycolysis and cell survival/proliferation under normal and pathologic conditions. Although HIF activity is mainly controlled by ubiquitination and protein degradation by the von Hippel Lindau (pVHL) tumor suppressor gene other mechanisms have recently been identified that regulate HIF signaling independently of pVHL. Principal Findings Here we characterized the mechanism by which COMMD1 regulates HIF-1α protein degradation. We show that COMMD1 competes with the chaperone heat shock protein HSP90β for binding to the NH2-terminal DNA-binding and heterodimerization domain of HIF-1α to regulate HIF-1α stability together with HSP70. Inhibition of HSP90 activity with 17-Allylamino-17-demethoxygeldanamycin (17-AAG) increased COMMD1-mediated HIF-1α degradation independent of ubiquitin and pVHL. Conclusion/Significance These data reveal a novel role for COMMD1 in conjunction with HSP90β/HSP70 in the ubiquitin and O2-independent regulation of HIF-1α.

Expression of the RNA Helicase DDX3 and the Hypoxia Response in Breast Cancer

Aims DDX3 is an RNA helicase that has antiapoptotic properties, and promotes proliferation and transformation. In addition, DDX3 was shown to be a direct downstream target of HIF-1α (the master regulatory of the hypoxia response) in breast cancer cell lines. However, the relation between DDX3 and hypoxia has not been addressed in human tumors. In this paper, we studied the relation between DDX3 and the hypoxic responsive proteins in human breast cancer. Methods and Results DDX3 expression was investigated by immunohistochemistry in breast cancer in comparison with hypoxia related proteins HIF-1α, GLUT1, CAIX, EGFR, HER2, Akt1, FOXO4, p53, ERα, COMMD1, FER kinase, PIN1, E-cadherin, p21, p27, Transferrin receptor, FOXO3A, c-Met and Notch1. DDX3 was overexpressed in 127 of 366 breast cancer patients, and was correlated with overexpression of HIF-1α and its downstream genes CAIX and GLUT1. Moreover, DDX3 expression correlated with hypoxia-related proteins EGFR, HER2, FOXO4, ERα and c-Met in a HIF-1α dependent fashion, and with COMMD1, FER kinase, Akt1, E-cadherin, TfR and FOXO3A independent of HIF-1α. Conclusions In invasive breast cancer, expression of DDX3 was correlated with overexpression of HIF-1α and many other hypoxia related proteins, pointing to a distinct role for DDX3 under hypoxic conditions and supporting the oncogenic role of DDX3 which could have clinical implication for current development of DDX3 inhibitors.

Endocytosis of EGFR requires its kinase activity and N-terminal transmembrane dimerization motif.

Summary EGFR signaling is attenuated by endocytosis and degradation of receptor–ligand complexes in lysosomes. Endocytosis of EGFR is known to be regulated by multiple post-translational modifications. The observation that prevention of these modifications does not block endocytosis completely, suggests the involvement of other mechanism(s). Recently, receptor clustering has been suggested to induce internalization of multiple types of membrane receptors. However, the mechanism of clustering-induced internalization remains unknown. We have used biparatopic antibody fragments from llama (VHHs) to induce EGFR clustering without stimulating tyrosine kinase activity. Using this approach, we have found an essential role for the N-terminal GG4-like dimerization motif in the transmembrane domain (TMD) for clustering-induced internalization. Moreover, conventional EGF-induced receptor internalization depends exclusively on this TMD dimerization and kinase activity. Mutations in this dimerization motif eventually lead to reduced EGFR degradation and sustained signaling. We propose a novel role for the TMD dimerization motif in the negative-feedback control of EGFR. The widely conserved nature of GG4-like dimerization motifs in transmembrane proteins suggests a general role for these motifs in clustering-induced internalization.

Differential expression of growth factor receptors and membrane-bound tumor markers for imaging in male and female breast cancer.

Introduction Male breast cancer accounts for 0.5–1% of all breast cancers and is generally diagnosed at higher stage than female breast cancers and therefore might benefit from earlier detection and targeted therapy. Except for HER2 and EGFR, little is known about expression of growth factor receptors in male breast cancer. We therefore investigated expression profiles of growth factor receptors and membrane-bound tumor markers in male breast cancer and gynecomastia, in comparison with female breast cancer. Methods Tissue microarrays containing 133 male breast cancer and 32 gynecomastia cases were stained by immunohistochemistry for a panel of membrane-bound targets and compared with data on 266 female breast cancers. Results Growth factor receptors were variably expressed in 4.5% (MET) up to 38.5% (IGF1-R) of male breast cancers. Compared to female breast cancer, IGF1-R and carbonic anhydrase 12 (CAXII) were more frequently and CD44v6, MET and FGFR2 less frequently expressed in male breast cancer. Expression of EGFR, HER2, CAIX, and GLUT1 was not significantly different between male and female breast cancer. Further, 48.1% of male breast cancers expressed at least one and 18.0% expressed multiple growth factor receptors. Since individual membrane receptors are expressed in only half of male breast cancers, a panel of membrane markers will be required for molecular imaging strategies to reach sensitivity. A potential panel of markers for molecular imaging, consisting of EGFR, IGF1-R, FGFR2, CD44v6, CAXII, GLUT1, and CD44v6 was positive in 77% of male breast cancers, comparable to female breast cancers. Conclusions Expression patterns of growth factor receptors and hypoxia membrane proteins in male breast cancer are different from female breast cancer. For molecular imaging strategies, a putative panel consisting of markers for EGFR, IGF1-R, FGFR2, GLUT1, CAXII, CD44v6 was positive in 77% of cases and might be considered for development of molecular tracers for male breast cancer.

HIF-1α and NOTCH signaling in ductal and lobular carcinomas of the breast.

BackgroundNOTCH signaling is involved in every step of metazoan development and maintenance of adult tissue homeostasis. It is frequently deregulated by mutations and overexpression in different cancer types including solid tumors such as breast cancer. Another common feature of solid tumors is hypoxia, which occurs due to defective or insufficient vascularization. Hypoxia-inducible factors (HIFs) are key regulators of the homeostatic response to low oxygen levels. HIF-1α is overexpressed in many solid tumors, including breast cancer. Hypoxia-induced stabilization of HIF transcription factors has been shown to lead to NOTCH activation in vitro in different contexts and tissues, causing differentiation arrest and induction of proliferation and migration.MethodsSince the link between HIF-1α and NOTCH signalling has hardly been studied, we set out to closely investigate associations between the expression of HIF-1α and NOTCH pathway members in primary and metastatic human breast cancer specimens and their prognostic value.ResultsCo-expression of NOTCH1 intracellular domain (N1ICD) and HIF-1α was associated with a high grade and a high proliferation rate in invasive breast cancer. HIF-1α expression was low in classic, but high in pleomorphic lobular cancers, which also frequently showed stromal HIF-1α expression. NOTCH1 pathway activation was prognostically unfavorable.ConclusionIn breast cancer, NOTCH pathway activation appears to be associated with a poor prognosis, but NOTCH and HIF signaling do not seem to be functionally associated.