NHERF1/EBP50 in Breast Cancer: Clinical Perspectives

Na(+)/H(+) exchanger regulatory factor 1 (NHERF1) is a postsynaptic density 95/disc-large/zona occludens (PDZ) domain-containing protein that recruits membrane receptors/transporters and cytoplasmic signaling proteins into functional complexes. NHERF1 expression has been demonstrated to be altered in breast cancer, but its role in mammary cancerogenesis and progression remains still undefined. In this paper, we review what is known on the pathological role and the potential clinical application of NHERF1 protein in breast cancer. Recent evidence shows that an increased cytoplasmic expression of NHERF1 suggests a key role of its localization/compartmentalization in defining cancerogenesis, progression, and invasion. NHERF1 overexpression is associated with increasing tumor cytohistological grade, aggressive clinical behavior, unfavorable prognosis, and increased tumor hypoxia. Moreover, NHERF1 co-localizes with the oncogenic receptor HER2/neu in HER2/neu-overexpressing carcinoma and in distant metastases. These data make NHERF1 also a potential candidate of clinical relevance for anti-HER2/neu therapy

The scaffolding protein NHERF1 sensitizes EGFR-dependent tumor growth, motility and invadopodia function to gefitinib treatment in breast cancer cells.

Triple negative breast cancer (TNBC) patients cannot be treated with endocrine therapy or targeted therapies due to lack of related receptors. These patients overexpress EGFR but are resistant to Tyrosine Kinase Inhibitors (TKIs) and anti-EGFR therapies. Mechanisms suggested for resistance to TKIs include EGFR independence, mutations and alterations in EGFR and in its downstream signalling pathways. Ligand-induced endocytosis and degradation of EGFR play important roles in the down-regulation of the EGFR signal suggesting that its activity could be regulated by targeting its trafficking. Evidence in normal cells showing that the scaffolding protein Na+/H+ Exchanger Regulatory Factor 1 (NHERF1) can associate with EGFR to regulate its trafficking, led us to hypothesize that NHERF1 expression levels could regulate EGFR trafficking and functional expression in TNBC cells and, in this way, modulate its role in progression and response to treatment. We investigated the subcellular localization of NHERF1 and its interaction with EGFR in a metastatic basal like TNBC cell model, MDA-MB-231, and the role of forced NHERF1 overexpression and/or stimulation with EGF on the sensitivity to EGFR specific TKI treatment with gefitinib. Stimulation with EGF induces an interaction of NHERF1 with EGFR to regulate its localization, degradation and function. NHERF1 overexpression is sufficient to drive its interaction with EGFR in non-stimulated conditions, inhibits EGFR degradation and increases its retention time in the plasma membrane. Importantly, NHERF1 overexpression strongly sensitized the cell to the pharmacological inhibition by gefitinib of EGFR-driven growth, motility and invadopodia-dependent ECM proteolysis. The further determination of how the NHERF1-EGFR interaction is regulated may improve our understanding of TNBC resistance to the action of existing anticancer drugs

Crocus sativus L. Ecotypes from Mediterranean Countries: Phenological, Morpho-Productive, Qualitative and Genetic Traits

The characterization of C. sativus ecotypes is of great interest for preserving them from a possible genetic erosion due to the decrease of European cultivation surface. In this study, we evaluated four ecotypes from Italy (Sardinia and Abruzzo), Spain (Castilla-La Mancha), and Greece (Kozani) in order to detect the existence of variability and promote the biodiversity of this crop. Thirty-one traits related to saffron flowering, flower morphology, production of spice and daughter corms, vegetative development (leaf and corm traits), and spice quality, were evaluated. In addition, a genetic analysis through three PCR-based approaches, SSRs, RAPD, and SRAP was assessed. Results highlighted a phenotypic variation among ecotypes during two consecutive years. All the studied parameters were influenced by the ecotype except for the stamen length, color coordinates of tepals, leaf length, and leaf number per plant. Sardinia had a longer flowering interval, earlier flowering, and higher spice yield and quality than the other corm origins. The maximum values of morphological traits, such as stigma length, dry weight of stigmas, tepals, flowers and leaves, leaf area, and daughter corm weight were observed in the Abruzzo ecotype. Principal component analysis (PCA) showed a clear separation among ecotypes, in which Sardinia and Spain showed more similarities than Abruzzo and Kozani. Significant negative correlation was found between days to flower with stigma yield and quality. However, we could not find molecular markers discriminating among corm origins. In conclusion, this study suggests the importance of C. sativus ecotypes as precious source of biodiversity and bioactive compounds, and of their enhancement as fundamental prerequisite for a sustainable development strategy and as an agricultural diversification opportunity for growers

Acoustic characteristics evaluation of an innovative metamaterial obtained through 3D printing technique

The reduction of interior noise level in the transportation sector is a big problem to cope with in view to increase the comfort of passengers. For this reason a great emphasis from the research community is devoted to develop new technology which are able to satisfy the mechanical requirements with concrete benefits from the acoustic point of view. Currently, it does not exist a solution for wideband range of frequency. Indeed, porous materials are characterized by outstanding dissipation in the high frequency range but they exhibit poor performance in the low and medium frequency range, where instead resonant cavities systems have the best performances but with narrow-band sound absorption. For this reason, the design and development of new materials which offers a good acoustic absorption over a wide range of frequencies is requested. In this paper, a hybrid metamaterial is designed, by coupling resonant cavities with micro-porous material and obtained through additive manufacturing technique which enables to model complex geometries that could not be feasible with classical manufacturing. Numerical and experimental studies have been conducted on the manufactured samples of PLA, with an interesting focus on the effect of each parameter which affects the absorption properties

Increased demand for FAD synthesis in differentiated and stem pancreatic cancer cells is accomplished by modulating FLAD1 gene expression: the inhibitory effect of Chicago Sky Blue

FLAD1, along with its FAD synthase (FADS, EC 2.7.7.2) product, is crucial for flavin homeostasis and, due to its role in the mitochondrial respiratory chain and nuclear epigenetics, is closely related to cellular metabolism. Therefore, it is not surprising that it could be correlated with cancer. To our knowledge, no previous study has investigated FLAD1 prognostic significance in pancreatic ductal adenocarcinoma (PDAC). Thus, in the present work, the FAD synthesis process was evaluated in two PDAC cell lines: (a) PANC‐1‐ and PANC‐1‐derived cancer stem cells (CSCs), presenting the R273H mutation in the oncosuppressor p53, and (b) MiaPaca2 and MiaPaca2‐derived CSCs, presenting the R248W mutation in p53. As a control, HPDE cells expressing wt‐p53 were used. FADS expression/activity increase was found with malignancy and even more with stemness. An increased FAD synthesis rate in cancer cell lines is presumably demanded by the increase in the FAD‐dependent lysine demethylase 1 protein amount as well as by the increased expression levels of the flavoprotein subunit of complex II of the mitochondrial respiratory chain, namely succinate dehydrogenase. With the aim of proposing FADS as a novel target for cancer therapy, the inhibitory effect of Chicago Sky Blue on FADS enzymatic activity was tested on the recombinant 6His‐hFADS2 (IC50 = 1.2 μm) and PANC‐1‐derived CSCs' lysate (IC50 = 2–10 μm). This molecule was found effective in inhibiting the growth of PANC‐1 and even more of its derived CSC line, thus assessing its role as a potential chemotherapeutic drug

Correctors of mutant CFTR enhance subcortical cAMP-PKA signaling through modulating ezrin phosphorylation and cytoskeleton organization

The most common mutation of the cystic fibrosis transmembrane regulator (CFTR) gene, F508del, produces a misfolded protein resulting in its defective trafficking to the cell surface and an impaired chloride secretion. Pharmacological treatments partially rescue F508del CFTR activity either directly by interacting with the mutant protein and/or indirectly by altering the cellular protein homeostasis. Here, we show that the phosphorylation of ezrin together with its binding to phosphatidylinositol-4,5-bisphosphate (PIP2) tethers the F508del CFTR to the actin cytoskeleton, stabilizing it on the apical membrane and rescuing the sub-membrane compartmentalization of cAMP and activated PKA. Both the small molecules trimethylangelicin (TMA) and VX-809, which act as 'correctors' for F508del CFTR by rescuing F508del-CFTR-dependent chloride secretion, also restore the apical expression of phosphorylated ezrin and actin organization and increase cAMP and activated PKA submembrane compartmentalization in both primary and secondary cystic fibrosis airway cells. Latrunculin B treatment or expression of the inactive ezrin mutant T567A reverse the TMA and VX-809-induced effects highlighting the role of corrector-dependent ezrin activation and actin re-organization in creating the conditions to generate a sub-cortical cAMP pool of adequate amplitude to activate the F508del-CFTR-dependent chloride secretion

Gene expression signature induced by grape intake in healthy subjects reveals wide-spread beneficial effects on peripheral blood mononuclear cells

Abstract Using a transcriptomic approach, we performed a pilot study in healthy subjects to evaluate the changes in gene expression induced by grape consumption. Blood from twenty subjects was collected at baseline (T0), after 21 days of grape-rich diet (T1) and after one-month washout (T2). Gene expression profiling of peripheral blood mononuclear cells from six subjects identified 930 differentially expressed transcripts. Gene functional analysis revealed changes (at T1 and/or T2) suggestive of antithrombotic and anti-inflammatory effects, confirming and extending previous finding on the same subjects. Moreover, we observed several other favourable changes in the transcription of genes involved in crucial processes such as immune response, DNA and protein repair, autophagy and mitochondrial biogenesis. Finally, we detected significant changes in many long non-coding RNAs genes, whose regulatory functions are being increasingly appreciated. Altogether, our data suggest that a grape diet may exert its beneficial effects by targeting different strategic pathways

A Novel NHE1-Centered Signaling Cassette Drives Epidermal Growth Factor Receptor–Dependent Pancreatic Tumor Metastasis and Is a Target for Combination Therapy

Pancreatic ductal adenocarcinoma (PDAC) is one of the most lethal cancers principally because of early invasion and metastasis. The epidermal growth factor receptor (EGFR) is essential for PDAC development even in the presence of Kras, but its inhibition with erlotinib gives only a modest clinical response, making the discovery of novel EGFR targets of critical interest. Here, we revealed by mining a human pancreatic gene expression database that the metastasis promoter Na+/H+ exchanger (NHE1) associates with the EGFR in PDAC. In human PDAC cell lines, we confirmed that NHE1 drives both basal and EGF-stimulated three-dimensional growth and early invasion via invadopodial extracellular matrix digestion. EGF promoted the complexing of EGFR with NHE1 via the scaffolding protein Na +/H + exchanger regulatory factor 1, engaging EGFR in a negative transregulatory loop that controls the extent and duration of EGFR oncogenic signaling and stimulates NHE1. The specificity of NHE1 for growth or invasion depends on the segregation of the transient EGFR/Na +/H + exchanger regulatory factor 1/NHE1 signaling complex into dimeric subcomplexes in different lipid raftlike membrane domains. This signaling complex was also found in tumors developed in orthotopic mice. Importantly, the specific NHE1 inhibitor cariporide reduced both three-dimensional growth and invasion independently of PDAC subtype and synergistically sensitized these behaviors to low doses of erlotinib