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International audience; The α4β1 integrin facilitates both monocyte rolling and adhesion to the vascular endothelium and is physiologically activated by monocyte chemoattractant protein (MCP-1). The current study investigated the initial events in the adhesion of THP-1 cells to immobilized Vascular Cell Adhesion Molecule 1 (VCAM-1). Using AFM force measurements, cell adhesion was shown to be mediated by two populations of α4β1/VCAM-1 complexes. A low affinity form of α4β1 was anchored to the elastic elements of the cytoskeleton, while a higher affinity conformer was coupled to the viscous elements of the cell membrane. Within 100 ms of contact, THP-1 cells, stimulated by co-immobilized MCP-1, exhibited a tremendous increase in adhesion to VCAM-1. Enhanced cell adhesion was accompanied by a local decoupling of the cell membrane from the cytoskeleton and the formation of long membrane tethers. The tethers were individually anchored by multiple α4β1/VCAM-1 complexes that prolonged the extension of the viscous tethers. In vivo, the formation of these membrane tethers may provide the quantal structural units for the arrest of rolling monocytes within the blood vessels.

International audience; The development of targeted molecular therapies has provided remarkable advances into the treatment of human cancers. However, in most tumors the selective pressure triggered by anticancer agents encourages cancer cells to acquire resistance mechanisms. The generation of new rationally designed targeting agents acting on the oncogenic path(s) at multiple levels is a promising approach for molecular therapies. 2-phenylimidazo[2,1-b]benzothiazole derivatives have been highlighted for their properties of targeting oncogenic Met receptor tyrosine kinase (RTK) signaling. In this study, we evaluated the mechanism of action of one of the most active imidazo[2,1-b]benzothiazol-2-ylphenyl moiety-based agents, Triflorcas, on a panel of cancer cells with distinct features. We show that Triflorcas impairs in vitro and in vivo tumorigenesis of cancer cells carrying Met mutations. Moreover, Triflorcas hampers survival and anchorage-independent growth of cancer cells characterized by ''RTK swapping'' by interfering with PDGFRb phosphorylation. A restrained effect of Triflorcas on metabolic genes correlates with the absence of major side effects in vivo. Mechanistically, in addition to targeting Met, Triflorcas alters phosphorylation levels of the PI3K-Akt pathway, mediating oncogenic dependency to Met, in addition to Retinoblastoma and nucleophosmin/B23, resulting in altered cell cycle progression and mitotic failure. Our findings show how the unusual binding plasticity of the Met active site towards structurally different inhibitors can be exploited to generate drugs able to target Met oncogenic dependency at distinct levels. Moreover, the disease-oriented NCI Anticancer Drug Screen revealed that Triflorcas elicits a unique profile of growth inhibitory-responses on cancer cell lines, indicating a novel mechanism of drug action. The anti-tumor activity elicited by 2-phenylimidazo[2,1-b]benzothiazole derivatives through combined inhibition of distinct effectors in cancer cells reveal them to be promising anticancer agents for further investigation.

International audience; Neuropilins, initially characterized as neuronal receptors, act as co-receptors for cancer related growth factors and were recently involved in several signaling pathways leading to cytoskeletal organization, angiogenesis and cancer progression. Then, we sought to investigate the ability of neuropilin-2 to orchestrate epithelial-mesenchymal transition in colorectal cancer cells. Using specific siRNA to target neuropilin-2 expression, or gene transfer, we first observed that neuropilin-2 expression endows HT29 and Colo320 for xenograft formation. Moreover, neuropilin-2 conferred a fibroblastic-like shape to cancer cells, suggesting an involvement of neuropilin-2 in epithelial-mesenchymal transition. Indeed, the presence of neuropilin-2 in colorectal carcinoma cell lines was correlated with loss of epithelial markers such as cytokeratin-20 and E-cadherin and with acquisition of mesenchymal molecules such as vimentin. Furthermore, we showed by surface plasmon resonance experiments that neuropilin-2 is a receptor for transforming-growth factor-β1. The expression of neuropilin-2 on colon cancer cell lines was indeed shown to promote transforming-growth factor-β1 signaling, leading to a constitutive phosphorylation of the Smad2/3 complex. Treatment with specific TGFβ-type1 receptor kinase inhibitors restored E-cadherin levels and inhibited in part neuropilin-2-induced vimentin expression, suggesting that neuropilin-2 cooperates with TGFβ-type1 receptor to promote epithelial-mesenchymal transition in colorectal cancer cells. Our results suggest a direct role of NRP2 in epithelial-mesenchymal transition and highlight a cross-talk between neuropilin-2 and TGF-β1 signaling to promote cancer progression. These results suggest that neuropilin-2 fulfills all the criteria of a therapeutic target to disrupt multiple oncogenic functions in solid tumors.

International audience; Gliomas such as oligodendrogliomas (ODG) and glioblastomas (GBM) are brain tumours with different clinical outcomes. Histology-based classification of these tumour types is often difficult. Therefore the first aim of this study was to gain microRNA data that can be used as reliable signatures of oligodendrogliomas and glioblastomas. We investigated the levels of 282 microRNAs using membrane-array hybridisation and real-time PCR in ODG, GBM and control brain tissues. In comparison to these control tissues, 26 deregulated microRNAs were identified in tumours and the tissue levels of seven microRNAs (miR-21, miR-128, miR-132, miR-134, miR-155, miR-210 and miR-409-5p) appropriately discriminated oligodendrogliomas from glioblastomas. Genomic, epigenomic and host gene expression studies were conducted to investigate the mechanisms involved in these deregulations. Another aim of this study was to better understand glioma physiopathology looking for targets of deregulated microRNAs. We discovered that some targets of these microRNAs such as STAT3, PTBP1 or SIRT1 are differentially expressed in gliomas consistent with deregulation of microRNA expression. Moreover, MDH1, the target of several deregulated microRNAs, is repressed in glioblastomas, making an intramitochondrial-NAD reduction mediated by the mitochondrial aspartate-malate shuttle unlikely. Understanding the connections between microRNAs and bioenergetic pathways in gliomas may lead to identification of novel therapeutic targets.

We report the first successful use of irreversible electroporation for the minimally invasive treatment of aggressive cutaneous tumors implanted in mice. Irreversible electroporation is a newly developed non-thermal tissue ablation technique in which certain short duration electrical fields are used to permanently permeabilize the cell membrane, presumably through the formation of nanoscale defects in the cell membrane. Mathematical models of the electrical and thermal fields that develop during the application of the pulses were used to design an efficient treatment protocol with minimal heating of the tissue. Tumor regression was confirmed by histological studies which also revealed that it occurred as a direct result of irreversible cell membrane permeabilization. Parametric studies show that the successful outcome of the procedure is related to the applied electric field strength, the total pulse duration as well as the temporal mode of delivery of the pulses. Our best results were obtained using plate electrodes to deliver across the tumor 80 pulses of 100 μs at 0:3 Hz with an electrical field magnitude of 2500 V/cm. These conditions induced complete regression in 12 out of 13 treated tumors, (92%), in the absence of tissue heating. Irreversible electroporation is thus a new effective modality for non-thermal tumor ablation. © 2007 Al-Sakere et al.

International audience; Glioblastoma (GBM) is the most common and aggressive form of brain tumor, characterized by high migratory behavior and infiltration in brain parenchyma which render classic therapeutic approach ineffective. The migratory behaviour of GBM cells could be conditioned by a number of tissue- and glioma-derived cytokines and growth factors. Although the pro-migratory action of CXCL12 on GBM cells in vitro and in vivo is recognized, the molecular mechanisms involved are not clearly identified. In fact the signaling pathways involved in the pro-migratory action of CXCL12 may differ in individual glioblastoma and integrate with those resulting from abnormal expression and activation of growth factor receptors. In this study we investigated whether some of the receptor tyrosine kinases commonly expressed in GBM cells could cooperate with CXCL12/CXCR4 in their migratory behavior. Our results show a functional cross-talk between CXCR4 and PDGFR which appears to be essential for GBM chemotaxis.

The spatial organization of the genome in the nucleus plays a role in the regulation of gene expression. Whether co-regulated genes are subject to coordinated repositioning to a shared nuclear space is a matter of considerable interest and debate. We investigated the nuclear organization of estrogen receptor alpha (ERα) target genes in human breast epithelial and cancer cell lines, before and after transcriptional activation induced with estradiol. We find that, contrary to another report, the ERα target genes TFF1 and GREB1 are distributed in the nucleoplasm with no particular relationship to each other. The nuclear separation between these genes, as well as between the ERα target genes PGR and CTSD, was unchanged by hormone addition and transcriptional activation with no evidence for co-localization between alleles. Similarly, while the volume occupied by the chromosomes increased, the relative nuclear position of the respective chromosome territories was unaffected by hormone addition. Our results demonstrate that estradiol-induced ERα target genes are not required to co-localize in the nucleus., Author Summary Whether co-regulated genes relocalize in a coordinated fashion to a shared nuclear space is a matter of considerable interest and debate. We investigated the spatial organization of estrogen receptor alpha (ERα) target genes in three human breast epithelial cell lines, human epithelial cells (HMEC), the MCF10A normal-like diploid cell line, and the MCF-7 aneuploid tumor cell line. Nuclear positions of a subset of genes in the absence of hormone and upon addition of estradiol were assessed quantitatively using 2D and 3D in situ hybridization techniques. In our two laboratories, we find that TFF1 and GREB1 are distributed in the nucleoplasm with no particular relationship to each other. Distances between homologous and heterologous alleles of these genes and the relative nuclear position of their respective chromosome territories 2 and 21 was, contrary to a previous report, unaffected by transcription activation and hormone addition. Similar results were obtained with ERα target genes PGR and CTSD on chromosomes 11. Even in the anti-estrogen resistant LCC9 cell line, TFF1 and GREB1 and the two TFF1 alleles remained separated after exposure to estradiol. Our results thus demonstrate that estradiol-induced genes are not required to co-localize or interact in trans or in cis.