Prostate cancer (PrCa) cells crosstalk with the tumour microenvironment by releasing small extracellular vesicles (sEVs). sEVs, as well as large extracellular vesicles (LEVs), isolated via iodixanol density gradients from PrCa cell culture media, express the epithelial-specific αvβ6 integrin, which is known to be induced in cancer. In this study, we show sEV-mediated protein transfer of αvβ6 integrin to microvascular endothelial cells (human microvascular endothelial cells 1-HMEC1); we demonstrate that de novo αvβ6 integrin expression is not caused by increased mRNA levels. Incubation of HMEC1 with sEVs isolated from PrCa PC3 cells that express the αvβ6 integrin results in a highly significant increase in the number of nodes, junctions and tubules. In contrast, incubation of HMEC1 with sEVs isolated from β6 negative PC3 cells, generated by shRNA against β6, results in a reduction in the number of nodes, junctions and tubules, a decrease in survivin levels and an increase in a negative regulator of angiogenesis, pSTAT1. Furthermore, treatment of HMEC1 with sEVs generated by CRISPR/Cas9-mediated down-regulation of β6, causes up-regulation of pSTAT1. Overall, our findings suggest that αvβ6 integrin in cancer sEVs regulates angiogenesis during PrCa progression.
BackgroundInflammatory breast cancer (IBC) is an aggressive type of advanced breast cancer with a poor prognosis. We recently found that focal adhesion kinase 1 (FAK1) is upregulated and phosphorylated (active) in IBC. In this study, we investigated the effect of CEP-37440, a dual inhibitor of FAK1 and anaplastic lymphoma kinase (ALK), using human IBC cell lines and preclinical models of IBC.MethodsCell proliferation assays were performed in the presence of several concentrations of CEP-37440 using IBC and triple-negative breast cancer non-IBC cell lines. In vitro, we studied the expression of total FAK1, phospho-FAK1 (Tyr 397), total ALK and phospho-ALK (Tyr 1604). In vivo, we tested CEP-37440 using FC-IBC02, SUM149, and SUM190 IBC xenograft mouse models.ResultsCEP-37440 at low concentration decreased the proliferation of the IBC cell lines FC-IBC02, SUM190, and KPL4, while not affecting the proliferation of normal breast epithelial cells. At higher concentration, CEP-37440 was also able to inhibit the proliferation of the IBC cell line MDA-IBC03 and the triple-negative non-IBC cell lines MDA-MB-231 and MDA-MB-468; the IBC cell line SUM149 showed a slight response to the drug. CEP-37440 decreased the cell proliferation of FC-IBC02, SUM190, and KPL4 by blocking the autophosphorylation kinase activity of FAK1 (Tyr 397). None of the cells evaluated expressed ALK. In vivo, after 7 weeks of CEP-37440 treatment, the SUM190, FC-IBC02, and SUM149 breast tumor xenografts were smaller in mice treated with 55 mg/kg bid CEP-37440 compared to the controls; the tumor growth inhibition (TGI) was 79.7 %, 33 %, and 23 %, respectively. None of the FC-IBC02 breast xenografts mice treated with CEP-37440 developed brain metastasis while 20 % of the mice in the control group developed brain metastasis. Expression array analyses in FC-IBC02 cells showed that CEP-37440 affects the expression of genes related to apoptosis, interferon signaling, and cytokines.ConclusionsCEP-37440 is effective against some IBC cells that express phospho-FAK1 (Tyr 397), and its antiproliferative activity is related to its ability to decrease phospho-FAK1. Our results suggest that combinational therapies could be more effective than using CEP-37440 as a single agent.Electronic supplementary materialThe online version of this article (doi:10.1186/s13058-016-0694-4) contains supplementary material, which is available to authorized users.
Dear Editor,Bronchial asthma is a chronic disorder of the airways characterized by airway remodelling. Airway remodelling includes bronchial epithelium damage, airway wall oedema, smooth muscle cell changes and extracellular matrix deposition. This deposition is mainly due to activation of mesenchymal cells such as fibroblasts [1]. Bronchial fibroblasts from asthmatic patients showed significant phenotypic and functional changes [2]. In mild asthma, the proliferation rate is decreased suggesting a premature senescence [3]. Cellular senescence is an irreversible proliferation arrest that can be triggered by a number of factors including ageing, DNA damage and oxidative stress. Morphological and functional changes occur during senescence and molecular mechanisms involve a complex signalling network such telomere shortening [4]. Telomeres are nucleoprotein complexes located at the ends of chromosomes in mammalian cells [5]. They are not replicated during the S phase of the cell cycle and, hence, are shortened after cell division. When telomeres reach a critical length, they induce cell cycle exit and thereby limit the cell replicative capacity.Chronic inflammation is an important feature of asthma. Production and release of cytokines and growth factors are increased in the asthmatic airways. These mediators have a direct effect on structural cells such as fibroblasts. We showed that the concomitant production of endothelin-1, TGF-b1 and PDGF-BB by the cells found in the chronic bronchial inflammatory microenvironment of asthma airways can potentially activate bronchial fibroblast proliferation suggesting that proliferation rate of asthmatic bronchial fibroblasts can contribute to their premature ageing [2]. In this study, we looked at potential changes in telomere length of bronchial fibroblasts obtained by bronchial biopsies from mild asthmatic subjects and healthy controls and assessed if there is a correlation between such length and the degree of airway hyper-responsiveness observed in these subjects. Patients and methodsSubjects were recruited from the Asthma clinic at the Institut Universitaire de Pneumologie et de Cardiologie de Qu ebec. The local ethics committee approved the study, and all subjects gave their written informed consent. The subjects with asthma were diagnosed according to the American Thoracic Society criteria. Subject characteristics are summarized in Table 1. Mild asthmatic subjects were non-smokers, atopic and used only inhaled b2-agonists on demand. None used inhaled or systemic corticosteroids. Healthy subjects were non-atopic and non-smokers with no history of asthma or allergy. Fibroblasts were isolated from all subjects and used at early passages (p2 to p4) to keep their phenotype [3]. To evaluate telomere length, genomic DNA was purified from fibroblasts using a Qiagen DNA kit and genomic-tips 20/G, following the manufacturer's instructions. Single-telomere Southern blot was performed using TeloTAGGG Telomere Length Assay (Roche, Dako, Canada). We performed telomere Q-FISH using a fl...
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