Publications by authors named "Philippe Delannoy"

55 Publications

-acetylated Gangliosides as Targets for Cancer Immunotherapy.

Cells 2020 03 17;9(3). Epub 2020 Mar 17.

UMR 8576 - UGSF - Unité de Glycobiologie Structurale et Fonctionnelle, CNRS, Université de Lille, F-59000 Lille, France.

-acetylation of sialic acid residues is one of the main modifications of gangliosides, and modulates ganglioside functions. -acetylation of gangliosides is dependent on sialyl--acetyltransferases and sialyl--acetyl-esterase activities. CAS1 Domain-Containing Protein 1 (CASD1) is the only human sialyl--acetyltransferases (SOAT) described until now. -acetylated ganglioside species are mainly expressed during embryonic development and in the central nervous system in healthy adults, but are re-expressed during cancer development and are considered as markers of cancers of neuroectodermal origin. However, the specific biological roles of -acetylated gangliosides in developing and malignant tissues have not been extensively studied, mostly because of the requirement of specific approaches and tools for sample preparation and analysis. In this review, we summarize our current knowledge of ganglioside biosynthesis and expression in normal and pathological conditions, of ganglioside -acetylation analysis and expression in cancers, and of the possible use of -acetylated gangliosides as targets for cancer immunotherapy.
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http://dx.doi.org/10.3390/cells9030741DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC7140702PMC
March 2020

Glycosylation changes in inflammatory diseases.

Adv Protein Chem Struct Biol 2020 26;119:111-156. Epub 2019 Nov 26.

University Lille, CNRS, UMR 8576 - UGSF - Unite de Glycobiologie Structurale et Fonctionnelle, F-59000 Lille, France.

Glycosylation is one of the most important modifications of proteins and lipids, and cell surface glycoconjugates are thought to play important roles in a variety of biological functions including cell-cell and cell-substrate interactions, bacterial adhesion, cell immunogenicity and cell signaling. Alterations of glycosylation are observed in a number of inflammatory diseases. Pro-inflammatory cytokines have been shown to modulate cell surface glycosylation by regulating the expression of glycosyltransferases and sulfotransferases involved in the biosynthesis of glycan chains, inducing the expression of specific carbohydrate antigens at the cell surface that can be recognized by different types of lectins or by bacterial adhesins, contributing to the development of diseases. Glycosylation can also regulate biological functions of immune cells by recruiting leukocytes to inflammation sites with pro- or anti-inflammatory effects. Cell surface proteoglycans provide a large panel of binding sites for many mediators of inflammation, and regulate their bio-availability and functions. In this review, we summarize the current knowledge of the glycosylation changes occurring in mucin type O-linked glycans, glycosaminoglycans, as well as in glycosphingolipids, with a particular focus on cystic fibrosis and neurodegenerative diseases, and their consequences on cell interactions and disease progression.
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http://dx.doi.org/10.1016/bs.apcsb.2019.08.008DOI Listing
January 2021

Profiling of -acetylated Gangliosides Expressed in Neuroectoderm Derived Cells.

Int J Mol Sci 2020 Jan 6;21(1). Epub 2020 Jan 6.

Univ. Lille, CNRS, UMR 8576-UGSF-Unité de Glycobiologie Structurale et Fonctionnelle, F-59000 Lille, France.

The expression and biological functions of oncofetal markers GD2 and GD3 were extensively studied in neuroectoderm-derived cancers in order to characterize their potential as therapeutic targets. Using immunological approaches, we previously identified GD3, GD2, and AcGD2 expression in breast cancer (BC) cell lines. However, antibodies specific for -acetylated gangliosides are not exempt of limitations, as they only provide information on the expression of a limited set of -acetylated ganglioside species. Consequently, the aim of the present study was to use structural approaches in order to apprehend ganglioside diversity in melanoma, neuroblastoma, and breast cancer cells, focusing on -acetylated species that are usually lost under alkaline conditions and require specific analytical procedures. We used purification and extraction methods that preserve the -acetyl modification for the analysis of native gangliosides by MALDI-TOF. We identified the expression of GM1, GM2, GM3, GD2, GD3, GT2, and GT3 in SK-Mel28 (melanoma), LAN-1 (neuroblastoma), Hs 578T, SUM 159PT, MDA-MB-231, MCF-7 (BC), and BC cell lines over-expressing GD3 synthase. Among -acetylated gangliosides, we characterized the expression of AcGM1, AcGD3, AcGD2, AcGT2, and AcGT3. Furthermore, the experimental procedure allowed us to clearly identify the position of the sialic acid residue that carries the -acetyl group on b- and c-series gangliosides by MS/MS fragmentation. These results show that ganglioside -acetylation occurs on both inner and terminal sialic acid residue in a cell type-dependent manner, suggesting different -acetylation pathways for gangliosides. They also highlight the limitation of immuno-detection for the complete identification of -acetylated ganglioside profiles in cancer cells.
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http://dx.doi.org/10.3390/ijms21010370DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6981417PMC
January 2020

Gangliosides: The Double-Edge Sword of Neuro-Ectodermal Derived Tumors.

Biomolecules 2019 07 27;9(8). Epub 2019 Jul 27.

Université de Lille, CNRS, UMR8576-UGSF-Unité de Glycobiologie Structurale et Fonctionnelle, F59000 Lille, France.

Gangliosides, the glycosphingolipids carrying one or several sialic acid residues, are mostly localized at the plasma membrane in lipid raft domains and implicated in many cellular signaling pathways mostly by interacting with tyrosine kinase receptors. Gangliosides are divided into four series according to the number of sialic acid residues, which can be also modified by -acetylation. Both ganglioside expression and sialic acid modifications can be modified in pathological conditions such as cancer, which can induce either pro-cancerous or anti-cancerous effects. In this review, we summarize the specific functions of gangliosides in neuro-ectodermal derived tumors, and their roles in reprogramming the lipidomic profile of cell membrane occurring with the induction of epithelial-mesenchymal transition.
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http://dx.doi.org/10.3390/biom9080311DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6723632PMC
July 2019

Novel Zebrafish Mono-α2,8-sialyltransferase (ST8Sia VIII): An Evolutionary Perspective of α2,8-Sialylation.

Int J Mol Sci 2019 Jan 31;20(3). Epub 2019 Jan 31.

Université de Lille, CNRS, UMR 8576-UGSF-Unité de Glycobiologie Structurale et Fonctionnelle, F-59000 Lille, France.

The mammalian mono-α2,8-sialyltransferase ST8Sia VI has been shown to catalyze the transfer of a unique sialic acid residues onto core 1 -glycans leading to the formation of di-sialylated -glycosylproteins and to a lesser extent to diSia motifs onto glycolipids like GD1a. Previous studies also reported the identification of an orthologue of the gene in the zebrafish genome. Trying to get insights into the biosynthesis and function of the oligo-sialylated glycoproteins during zebrafish development, we cloned and studied this fish α2,8-sialyltransferase homologue. In situ hybridization experiments demonstrate that expression of this gene is always detectable during zebrafish development both in the central nervous system and in non-neuronal tissues. Intriguingly, using biochemical approaches and the newly developed in vitro MicroPlate Sialyltransferase Assay (MPSA), we found that the zebrafish recombinant enzyme does not synthetize diSia motifs on glycoproteins or glycolipids as the human homologue does. Using comparative genomics and molecular phylogeny approaches, we show in this work that the human ST8Sia VI orthologue has disappeared in the ray-finned fish and that the homologue described in fish correspond to a new subfamily of α2,8-sialyltransferase named ST8Sia VIII that was not maintained in Chondrichtyes and Sarcopterygii.
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http://dx.doi.org/10.3390/ijms20030622DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6387029PMC
January 2019

Identification of 9-O-acetyl-N-acetylneuraminic acid (Neu5,9Ac) as main O-acetylated sialic acid species of GD2 in breast cancer cells.

Glycoconj J 2019 02 5;36(1):79-90. Epub 2019 Jan 5.

CNRS, UMR 8576 - UGSF - Unité de Glycobiologie Structurale et Fonctionnelle, University Lille, F-59000, Lille, France.

Mainly restricted to the nervous system in healthy adults, complex gangliosides such as GD3 and GD2 have been shown to be involved in aggressiveness and metastasis of neuro-ectoderm derived tumors such as melanoma and neuroblastoma. Interestingly, O-acetylated forms of GD2, not expressed in human peripheral nerve fibers, are highly expressed in GD2+ tumor cells. Very little information is known regarding the expression of O-acetylated disialogangliosides in breast cancer (BC) cell lines. Here, we analyzed the expression of GD2, GD3 and their O-acetylated forms O-acetyl-GD2 (OAcGD2) and O-acetyl-GD3 (OAcGD3) in BC cells. We used Hs 578T and SUM159PT cell lines, as well as cell clones over-expressing GD3 synthase derived from MDA-MB-231 and MCF-7. Using flow cytometry and immunocytochemistry/confocal microscopy, we report that BC cells express b-series gangliosides GD3 and GD2, as well as significant amounts of OAcGD2. However, OAcGD3 expression was not detected in these cells. O-acetylation of gangliosides isolated from BC cells was examined by LC-MS analysis of sialic acid DMB-derivatives. We report that the main acetylated form of sialic acid expressed in BC gangliosides is 9-O-acetyl-N-acetylneuraminic acid (Neu5,9Ac). These results highlight a close interrelationship between Neu5,9Ac and OAcGD2 expression, and suggest that OAcGD2 is synthetized from GD2 and not from OAcGD3 in BC cells.
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http://dx.doi.org/10.1007/s10719-018-09856-wDOI Listing
February 2019

Gangliosides in Cancer Cell Signaling.

Prog Mol Biol Transl Sci 2018 19;156:197-227. Epub 2018 Jan 19.

University of Lille, Villeneuve d'Ascq, France. Electronic address:

At the outer leaflet of the plasma membrane, gangliosides are found with other glycosphingolipids, phospholipids, and cholesterol in glycolipid-enriched microdomains, in which they interact with signaling molecules including receptor tyrosine kinases and signal transducers. The role of gangliosides in the regulation of signal transduction has been reported for many cases and in different cell types. The biosynthesis of gangliosides involves specific enzymes, mainly glycosyltransferases that control together with glycohydrolases, the steady state of gangliosides at the cell surface. Changes in ganglioside composition are therefore correlated with modifications of glycosyltransferases or glycohydrolases expression and result in the deregulation of cellular signals. In several types of cancers, the overexpression of disialogangliosides, such as GD3 or GD2 mainly results in the activation of cell signaling, increasing cell proliferation and migration, as well as tumor growth. In this chapter, we summarize our current knowledge of ganglioside biosynthesis, degradation, and of their role in cell signaling regulation in cancers.
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http://dx.doi.org/10.1016/bs.pmbts.2017.10.003DOI Listing
February 2019

TNF differentially regulates ganglioside biosynthesis and expression in breast cancer cell lines.

PLoS One 2018 26;13(4):e0196369. Epub 2018 Apr 26.

University of Lille, Structural and Functional Glycobiology Unit, UMR CNRS 8576, Lille, France.

Gangliosides are glycosphingolipids concentrated in glycolipid-enriched membrane microdomains. Mainly restricted to the nervous system in healthy adult, complex gangliosides such as GD3 and GD2 have been shown to be involved in aggressiveness and metastasis of neuro-ectoderm derived tumors such as melanoma and neuroblastoma. GD3 synthase (GD3S), the key enzyme that controls the biosynthesis of complex gangliosides, was shown to be over-expressed in Estrogen Receptor (ER)-negative breast cancer tumors, and associated with a decreased overall survival of patients. We previously demonstrated that GD3S expression in ER-negative breast cancer cells induced a proliferative phenotype and an increased tumor growth. In addition, our results clearly indicate that Tumor Necrosis Factor (TNF) induced GD3S over-expression in breast cancer cells via NFκB pathway. In this study, we analyzed the effect of TNF on ganglioside biosynthesis and expression in breast cancer cells from different molecular subtypes. We showed that TNF up-regulated the expression of GD3S in MCF-7 and Hs578T cells, whereas no change was observed for MDA-MB-231. We also showed that TNF induced an increased expression of complex gangliosides at the cell surface of a small proportion of MCF-7 cells. These results demonstrate that TNF differentially regulates gangliosides expression in breast cancer cell lines and establish a possible link between inflammation at the tumor site environment, expression of complex gangliosides and tumor development.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0196369PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5919650PMC
August 2018

Gangliosides: Structures, Biosynthesis, Analysis, and Roles in Cancer.

Chembiochem 2017 07 24;18(13):1146-1154. Epub 2017 Apr 24.

Université de Lille, CNRS, UMR 8576, UGSF-Unité de Glycobiologie Structurale et Fonctionnelle, 59000, Lille, France.

Gangliosides are acidic glycosphingolipids containing one or more sialic acid residues. They are essential compounds at the outer leaflet of the plasma membrane, where they interact with phospholipids, cholesterol, and transmembrane proteins, forming lipid rafts. They are involved in cell adhesion, proliferation, and recognition processes, as well as in the modulation of signal transduction pathways. These functions are mainly governed by the glycan moiety, and changes in the structures of gangliosides occur under pathological conditions, particularly in neuro-ectoderm-derived cancers. With the progress in mass spectrometry analysis of gangliosides, their role in cancer progression can be now investigated in more detail. In this review we summarize the current knowledge on the biosynthesis of gangliosides and their role in cancers, together with the recent development of cancer immunotherapy targeting gangliosides.
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http://dx.doi.org/10.1002/cbic.201600705DOI Listing
July 2017

Diverse subcellular localizations of the insect CMP-sialic acid synthetases.

Glycobiology 2017 04;27(4):329-341

Bioscience and Biotechnology Center, and Graduate School of Bioagricultural Sciences, Nagoya University, Nagoya, Japan.

The occurrence and biological importance of sialic acid (Sia) and its metabolic enzymes in insects have been studied using Drosophila melanogaster. The most prominent feature of D. melanogaster CMP-Sia synthetase (DmCSS) is its Golgi-localization, contrasted with nuclear localization of vertebrate CSSs. However, it remains unclear if the Golgi-localization is common to other insect CSSs and why it happens. To answer these questions, Aedes aegypti (mosquito) CSS (AaCSS) and Tribolium castaneum (beetle) CSS (TcCSS) were cloned and characterized for their activity and subcellular localization. Our new findings show: (1) AaCSS and TcCSS share a common overall structure with DmCSS in terms of evolutionarily conserved motifs and the absence of the C-terminal domain typical to vertebrate CSSs; (2) when expressed in mammalian and insect cells, AaCSS and TcCSS showed in vivo and in vitro CSS activities, similar to DmCSS. In contrast, when expressed in bacteria, they lacked CSS activity because the N-terminal hydrophobic region appeared to induce protein aggregation; (3) when expressed in Drosophila S2 cells, AaCSS and TcCSS were predominantly localized in the ER, but not in the Golgi. Surprisingly, DmCSS was mainly secreted into the culture medium, although partially detected in Golgi. Consistent with these results, the N-terminal hydrophobic regions of AaCSS and TcCSS functioned as a signal peptide to render them soluble in the ER, while the N-terminus of DmCSS functioned as a membrane-spanning region of type II transmembrane proteins whose cytosolic KLK sequence functioned as an ER export signal. Accordingly, the differential subcellular localization of insect CSSs are distinctively more diverse than previously recognized.
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http://dx.doi.org/10.1093/glycob/cww128DOI Listing
April 2017

Role of Cytokine-Induced Glycosylation Changes in Regulating Cell Interactions and Cell Signaling in Inflammatory Diseases and Cancer.

Cells 2016 Nov 29;5(4). Epub 2016 Nov 29.

Structural and Functional Glycobiology Unit, UMR CNRS 8576, University of Lille, Villeneuve d'Ascq 59655, France.

Glycosylation is one of the most important modifications of proteins and lipids, and cell surface glycoconjugates are thought to play important roles in a variety of biological functions including cell-cell and cell-substrate interactions, bacterial adhesion, cell immunogenicity and cell signaling. Alterations of glycosylation are observed in number of diseases such as cancer and chronic inflammation. In that context, pro-inflammatory cytokines have been shown to modulate cell surface glycosylation by regulating the expression of glycosyltransferases involved in the biosynthesis of carbohydrate chains. These changes in cell surface glycosylation are also known to regulate cell signaling and could contribute to disease pathogenesis. This review summarizes our current knowledge of the glycosylation changes induced by pro-inflammatory cytokines, with a particular focus on cancer and cystic fibrosis, and their consequences on cell interactions and signaling.
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http://dx.doi.org/10.3390/cells5040043DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5187527PMC
November 2016

TNF up-regulates ST3GAL4 and sialyl-Lewisx expression in lung epithelial cells through an intronic ATF2-responsive element.

Biochem J 2017 01 7;474(1):65-78. Epub 2016 Nov 7.

Université Lille, CNRS, UMR 8576, Unité de Glycobiologie Structurale etFonctionnelle (UGSF), F-59000 Lille, France

We have previously shown that tumor necrosis factor (TNF) induced the up-regulation of the sialyltransferase gene ST3GAL4 (α2,3-sialyltransferase gene) BX transcript through mitogen- and stress-activated kinase 1/2 (MSK1/2), extracellular signal-regulated kinase (ERK) and p38 mitogen-activated protein kinase (MAPK) signaling pathways. This up-regulation resulted in sialyl-Lewis (sLe) overexpression on high-molecular-weight glycoproteins in inflamed airway epithelium and increased the adhesion of Pseudomonas aeruginosa PAO1 and PAK strains to lung epithelial cells. In the present study, we describe a TNF-responsive element in an intronic region of the ST3GAL4 gene, whose TNF-dependent activity is repressed by ERK/p38 and MSK1/2 inhibitors. This TNF-responsive element contains potential binding sites for ETS1 and ATF2 transcription factors related to TNF signaling. We also show that ATF2 is involved in TNF responsiveness, as well as in TNF-induced ST3GAL4 BX transcript and sLe overexpression in A549 lung epithelial cells. Moreover, we show that TNF induces the binding of ATF2 to the TNF-responsive element. Altogether, these data suggest that ATF2 could be a potential target to prevent inflammation-induced P. aeruginosa binding in the lung of patients suffering from lung diseases such as chronic bronchitis or cystic fibrosis.
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http://dx.doi.org/10.1042/BCJ20160602DOI Listing
January 2017

ST6GALNAC5 Expression Decreases the Interactions between Breast Cancer Cells and the Human Blood-Brain Barrier.

Int J Mol Sci 2016 Aug 11;17(8). Epub 2016 Aug 11.

Université d'Artois (UArtois), EA2465, Laboratoire de la Barrière Hémato-Encéphalique (LBHE), Lens F-62300, France.

The ST6GALNAC5 gene that encodes an α2,6-sialyltransferase involved in the biosynthesis of α-series gangliosides, was previously identified as one of the genes that mediate breast cancer metastasis to the brain. We have shown that the expression of ST6GALNAC5 in MDA-MB-231 breast cancer cells resulted in the expression of GD1α ganglioside at the cell surface. By using a human blood-brain barrier in vitro model recently developed, consisting in CD34⁺ derived endothelial cells co-cultivated with pericytes, we show that ST6GALNAC5 expression decreased the interactions between the breast cancer cells and the human blood-brain barrier.
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http://dx.doi.org/10.3390/ijms17081309DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC5000706PMC
August 2016

Two E-selectin ligands, BST-2 and LGALS3BP, predict metastasis and poor survival of ER-negative breast cancer.

Int J Oncol 2016 Jul 13;49(1):265-75. Epub 2016 May 13.

University of Lille, INSERM, U908 - CPAC, Cell Plasticity and Cancer, Lille, Villeneuve d'Ascq, France.

Distant metastases account for the majority of cancer-related deaths in breast cancer. The rate and site of metastasis differ between estrogen receptor (ER)-negative and ER-positive tumours, and metastatic fate can be very diverse even within the ER-negative group. Characterisation of new pro-metastatic markers may help to identify patients with higher risk and improve their care accordingly. Selectin ligands aberrantly expressed by cancer cells promote metastasis by enabling interaction between circulating tumour cells and endothelial cells in distant organs. These ligands consist in carbohydrate molecules, such as sialyl-Lewis x antigen (sLex), borne by glycoproteins or glycolipids on the cancer cell surface. We have previously demonstrated that the molecular scaffold presenting sLex to selectins (e.g. glycolipid vs. glycoproteins) was crucial for these interactions to occur. Moreover, we reported that detection of sLex alone in breast carcinomas was only of limited prognostic value. However, since sLex was found to be carried by several glycoproteins in cancer cells, we hypothesized that the combination of the carbohydrate with its carriers could be more relevant than each marker independently. In this study, we addressed this question by analysing sLex expression together with two glycoproteins (BST-2 and LGALS3BP), shown to interact with E-selectin in a carbohydrate-dependent manner, in a cohort of 249 invasive breast cancers. We found both glycoproteins to be associated with distant metastasis risk and poorer survival. Importantly, concomitant high expression of BST-2 with sLex defined a sub-group of patients with ER-negative tumours displaying higher risks of liver and brain metastasis and a 3-fold decreased survival rate.
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http://dx.doi.org/10.3892/ijo.2016.3521DOI Listing
July 2016

Selection of a Relevant In Vitro Blood-Brain Barrier Model to Investigate Pro-Metastatic Features of Human Breast Cancer Cell Lines.

PLoS One 2016 9;11(3):e0151155. Epub 2016 Mar 9.

Université d'Artois (UArtois), EA2465, LBHE, Lens, F-62307, France.

Around 7-17% of metastatic breast cancer patients will develop brain metastases, associated with a poor prognosis. To reach the brain parenchyma, cancer cells need to cross the highly restrictive endothelium of the Blood-Brain Barrier (BBB). As treatments for brain metastases are mostly inefficient, preventing cancer cells to reach the brain could provide a relevant and important strategy. For that purpose an in vitro approach is required to identify cellular and molecular interaction mechanisms between breast cancer cells and BBB endothelium, notably at the early steps of the interaction. However, while numerous studies are performed with in vitro models, the heterogeneity and the quality of BBB models used is a limitation to the extrapolation of the obtained results to in vivo context, showing that the choice of a model that fulfills the biological BBB characteristics is essential. Therefore, we compared pre-established and currently used in vitro models from different origins (bovine, mice, human) in order to define the most appropriate tool to study interactions between breast cancer cells and the BBB. On each model, the BBB properties and the adhesion capacities of breast cancer cell lines were evaluated. As endothelial cells represent the physical restriction site of the BBB, all the models consisted of endothelial cells from animal or human origins. Among these models, only the in vitro BBB model derived from human stem cells both displayed BBB properties and allowed measurement of meaningful different interaction capacities of the cancer cell lines. Importantly, the measured adhesion and transmigration were found to be in accordance with the cancer cell lines molecular subtypes. In addition, at a molecular level, the inhibition of ganglioside biosynthesis highlights the potential role of glycosylation in breast cancer cells adhesion capacities.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0151155PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4784983PMC
August 2016

Preface.

Top Curr Chem 2015 ;367:v-vii

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November 2015

Challenges in Antibody Development against Tn and Sialyl-Tn Antigens.

Biomolecules 2015 Aug 11;5(3):1783-809. Epub 2015 Aug 11.

CEDOC, Chronic Diseases Research Center, NOVA Medical School/Faculdade de Ciências Médicas, Universidade NOVA de Lisboa, Campo dos Mártires da Pátria, 130, Lisboa 1169-056, Portugal.

The carbohydrate antigens Tn and sialyl-Tn (STn) are expressed in most carcinomas and usually absent in healthy tissues. These antigens have been correlated with cancer progression and poor prognosis, and associated with immunosuppressive microenvironment. Presently they are used in clinical trials as therapeutic vaccination, but with limited success due to their low immunogenicity. Alternatively, anti-Tn and/or STn antibodies may be used to harness the immune system against tumor cells. Whilst the development of antibodies against these antigens had a boost two decades ago for diagnostic use, so far no such antibody entered into clinical trials. Possible limitations are the low specificity and efficiency of existing antibodies and that novel antibodies are still necessary. The vast array of methodologies available today will allow rapid antibody development and novel formats. Following the advent of hybridoma technology, the immortalization of human B cells became a methodology to obtain human monoclonal antibodies with better specificity. Advances in molecular biology including phage display technology for high throughput screening, transgenic mice and more recently molecularly engineered antibodies enhanced the field of antibody production. The development of novel antibodies against Tn and STn taking advantage of innovative technologies and engineering techniques may result in innovative therapeutic antibodies for cancer treatment.
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http://dx.doi.org/10.3390/biom5031783DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4598775PMC
August 2015

Accumulation of GD1α Ganglioside in MDA-MB-231 Breast Cancer Cells Expressing ST6GalNAc V.

Molecules 2015 Apr 16;20(4):6913-24. Epub 2015 Apr 16.

Structural and Functional Glycobiology Unit, UMR CNRS 8576, University of Lille, 59655 Villeneuve d'Ascq, France.

α-Series gangliosides define a particular sub-class of glycosphingolipids containing sialic acid α2,6-linked to GalNAc residue that was isolated as a minor compound from the brain. The sialyltransferase ST6GalNAc V was cloned from mouse brain and showed α2,6-sialyltransferase activity almost exclusively for GM1b, to form GD1α and is considered as the main enzyme involved in the biosynthesis of α-series gangliosides. Recently, ST6GALNAC5 was identified as one of the genes over-expressed in breast cancer cell populations selected for their ability to produce brain metastasis. However, the capacity of human breast cancer cells to produce α-series gangliosides has never been clearly demonstrated. Here, we show by stable transfection and MS-MS analysis of total glycosphingolipids that ST6GALNAC5 expressing MDA-MB-231 breast cancer cells accumulate GD1α ganglioside (IV3Neu5Ac1, III6Neu5Ac1Gg4-Cer).
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http://dx.doi.org/10.3390/molecules20046913DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6272744PMC
April 2015

Sialic acid expression in the mosquito Aedes aegypti and its possible role in dengue virus-vector interactions.

Biomed Res Int 2015 22;2015:504187. Epub 2015 Mar 22.

Molecular Biology and Biotechnology Department, Biomedical Research Institute, National University of México (UNAM), 04510 México City, Mexico.

Dengue fever (DF) is the most prevalent arthropod-borne viral disease which affects humans. DF is caused by the four dengue virus (DENV) serotypes, which are transmitted to the host by the mosquito Aedes aegypti that has key roles in DENV infection, replication, and viral transmission (vector competence). Mosquito saliva also plays an important role during DENV transmission. In this study, we detected the presence of sialic acid (Sia) in Aedes aegypti tissues, which may have an important role during DENV-vector competence. We also identified genome sequences encoding enzymes involved in Sia pathways. The cDNA for Aedes aegypti CMP-Sia synthase (CSAS) was amplified, cloned, and functionally evaluated via the complementation of LEC29.Lec32 CSAS-deficient CHO cells. AedesCSAS-transfected LEC29.Lec32 cells were able to express Sia moieties on the cell surface. Sequences related to α-2,6-sialyltransferase were detected in the Aedes aegypti genome. Likewise, we identified Sia-α-2,6-DENV interactions in different mosquito tissues. In addition, we evaluated the possible role of sialylated molecules in a salivary gland extract during DENV internalization in mammalian cells. The knowledge of early DENV-host interactions could facilitate a better understanding of viral tropism and pathogenesis to allow the development of new strategies for controlling DENV transmission.
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http://dx.doi.org/10.1155/2015/504187DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4385653PMC
December 2015

Carbohydrate-to-carbohydrate interactions between α2,3-linked sialic acids on α2 integrin subunits and asialo-GM1 underlie the bone metastatic behaviour of LNCAP-derivative C4-2B prostate cancer cells.

Biosci Rep 2014 Sep 17;34(5). Epub 2014 Sep 17.

*School of Science, Technology and Engineering Management, St. Thomas University, 16401 NW 37th Avenue, Miami Gardens, FL 33054, USA.

Complex interplays among proteins, lipids and carbohydrates can alter the phenotype and are suggested to have a crucial role in tumour metastasis. Our previous studies indicated that a complex of the GSLs (glycosphingolipids), AsGM1 (asialo-GM1), which lacks α2,3-linked sialic acid, and α2β1 integrin receptors is responsible for the metastatic behaviour of C4-2B prostate cancer cells. Herein, we identified and addressed the functional significance of changes in sialylation during prostate cancer progression. We observed an increase in α2,3-linked sialic acid residues on α2 subunits of α2β1 integrin receptors, correlating with increased gene expression of α2,3-STs (sialyltransferases), particularly ST3GAL3. Cell surface α2,3-sialylation of α2 subunits was required for the integrin α2β1-dependent cell adhesion to collagen type I and the same α2,3-linked sialic acid residues on the integrin receptor were responsible for the interaction with the carbohydrate moiety of AsGM1, explaining the complex formation between AsGM1 and α2β1 integrin receptors. These results provide novel insights into the role of sialic acids in the organization and function of important membrane components in invasion and metastatic processes.
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http://dx.doi.org/10.1042/BSR20140096DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC4166120PMC
September 2014

B4GALNT2 gene expression controls the biosynthesis of Sda and sialyl Lewis X antigens in healthy and cancer human gastrointestinal tract.

Int J Biochem Cell Biol 2014 Aug 19;53:442-9. Epub 2014 Jun 19.

Structural and Functional Glycobiology Unit, UMR CNRS 8576, University Lille Nord de France, Lille1, 59655 Villeneuve d'Ascq, France. Electronic address:

The histo blood group carbohydrate Sd(a) antigen and its cognate biosynthetic enzyme B4GALNT2 show the highest level of expression in normal colon. Their dramatic down regulation previously observed in colon cancer tissues could play a role in the concomitant elevation of the selectin ligand sLe(x), involved in metastasis. However, down regulation of sLe(x) expression by B4GALNT2 has been so far demonstrated in vitro, but not in tissues. The human B4GALNT2 gene specifies at least two transcripts, diverging in the first exon, never studied in normal and cancer tissues. The long form contains a 253 nt exon 1L; the short form contains a 38 nt exon 1S. Using qPCR, we showed that cell lines and normal or cancerous colon, expressed almost exclusively the short form, while the long form was mainly expressed by the embryonic colon fibroblast cell line CCD112CoN. Immunochemistry approaches using colon cancer cells permanently expressing either B4GALNT2 cDNAs as controls, led to the observation of several protein isoforms in human normal and cancerous colon, and cell lines. We showed that tissues expressing B4GALNT2 protein isoforms were able to induce Sd(a) and to inhibit sLe(x) expression; both of which are expressed mainly on PNGase F-insensitive carbohydrate chains. Concomitant expression of B4GALNT2 and siRNA-mediated inhibition of FUT6, the major fucosyltransferase involved in sLe(x) synthesis in colon, resulted in a cumulative inhibition of sLe(x). In normal colon samples a significant relationship between sLe(x) expression and the ratio between FUT6/B4GALNT2 activities exists, demonstrating for the first time a role for B4GALNT2 in sLe(x) inhibition in vivo.
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http://dx.doi.org/10.1016/j.biocel.2014.06.009DOI Listing
August 2014

TNF induces the expression of the sialyltransferase ST3Gal IV in human bronchial mucosa via MSK1/2 protein kinases and increases FliD/sialyl-Lewis(x)-mediated adhesion of Pseudomonas aeruginosa.

Biochem J 2014 Jan;457(1):79-87

*Université Lille Nord de France, Lille 1, UGSF (Unité de Glycobiologie Structurale et Fonctionnelle), CNRS (Centre National de la Recherche Scientifique), UMR 8576, 59650 Villeneuve d'Ascq, France.

We have shown previously that the pro-inflammatory cytokine TNF (tumour necrosis factor) could drive sLe(x) (sialyl-Lewis(x)) biosynthesis through the up-regulation of the BX transcript isoform of the ST3GAL4 (ST3 β-galactoside α-2,3-sialyltransferase 4) sialyltransferase gene in lung epithelial cells and human bronchial mucosa. In the present study, we show that the TNF-induced up-regulation of the ST3GAL4 BX transcript is mediated by MSK1/2 (mitogen- and stress-activated kinase 1/2) through the ERK (extracellular-signal-regulated kinase) and p38 MAPK (mitogen-activated protein kinase) pathways, and increases sLe(x) expression on high-molecular-mass glycoproteins in inflamed airway epithelium. We also show that the TNF-induced sLe(x) expression increases the adhesion of the Pseudomonas aeruginosa PAO1 and PAK strains to lung epithelial cells in a FliD-dependent manner. These results suggest that ERK and p38 MAPK, and the downstream kinase MSK1/2, should be considered as potential targets to hamper inflammation, bronchial mucin glycosylation changes and P. aeruginosa binding in the lung of patients suffering from lung diseases such as chronic bronchitis or cystic fibrosis.
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http://dx.doi.org/10.1042/BJ20130989DOI Listing
January 2014

α-L-fucosidase inhibition by pyrrolidine-ferrocene hybrids: rationalization of ligand-binding properties by structural studies.

Chemistry 2013 Jul 5;19(29):9526-33. Epub 2013 Jun 5.

Université de Reims Champagne-Ardenne, Institut de Chimie Moléculaire de Reims, CNRS UMR 7312, UFR des Sciences Exactes et Naturelles, 51687 Reims Cedex 2, France.

Enhanced metabolism of fucose through fucosidase overexpression is a signature of some cancer types, thus suggesting that fucosidase-targetted ligands could play the role of drug-delivery vectors. Herein, we describe the synthesis of a new series of pyrrolidine-ferrocene conjugates, consisting of a L-fuco-configured dihydroxypyrrolidine as the fucosidase ligand armed with a cytotoxic ferrocenylamine moeity. Three-dimensional structures of several of these fucosidase inhibitors reveal transition-state-mimicking (3)E conformations. Elaboration with the ferrocenyl moiety results in sub-micromolar inhibitors of both bovine and bacterial fucosidases, with the 3D structure of the latter revealing electron density indicative of highly mobile alkylferrocene compounds. The best compounds show a strong antiproliferative effect, with up to 100% inhibition of the proliferation of MDA-MB-231 cancer cells at 50 μM.
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http://dx.doi.org/10.1002/chem.201301001DOI Listing
July 2013

Over-sulfated glycosaminoglycans are alternative selectin ligands: insights into molecular interactions and possible role in breast cancer metastasis.

Clin Exp Metastasis 2013 Oct 6;30(7):919-31. Epub 2013 Jun 6.

Unité de Glycobiologie Structurale et Fonctionnelle, Unité Mixte de Recherche 8576 du Centre National de la Recherche Scientifique, Institut Fédératif de Recherche 147, Université Lille Nord de France-Université des Sciences et Technologies, Bat C9 Université Lille 1, 59655, Villeneuve d'Ascq, France.

Distant metastasis account for about 90 % of cancer associated deaths, and yet the oncology field is cruelly lacking tools to accurately predict and/or prevent metastasis. Distant metastasis occurs when circulating tumor cells interact with the endothelium of distant organs and extravasate from the blood vessel into the surrounding tissue. Selectins are a family of carbohydrate receptors well depicted for their role in tumor cells extravasation. They mediate primary interactions of cancer cells with endothelial cells, as well as secondary interactions with leucocytes and platelets, which are also promoting metastasis. The cancer associated carbohydrate antigen sialyl-Lewis x (sLe(x)) has been repeatedly shown to be involved, as selectin ligand, in these interactions. However, recent studies have highlighted that glycosaminoglycans (GAGs), another class of glycans, may also serve as ligands for selectins. We report herein that cancer-associated GAGs are differentially recognized by selectins according to their density of sulfation and the pH conditions of the binding. We also show that these parameters regulate platelets-cancer cells heterotypic aggregation, supporting the idea that GAGs may have pro-metastatic function. Combining our experimental results with in depth analyses of molecular dockings, we propose a model of GAG/selectin interactions robust enough to recapitulate the differential binding of selectins to GAGs, the competition between GAGs and sLe(x) for selectin binding and the effect of sub-physiological pH on GAGs affinities towards selectins. Altogether, our data suggest GAGs to be good ligands for selectins, potentially promoting distant metastasis in a complementary way to sLe(x).
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http://dx.doi.org/10.1007/s10585-013-9592-7DOI Listing
October 2013

Estradiol represses the G(D3) synthase gene ST8SIA1 expression in human breast cancer cells by preventing NFκB binding to ST8SIA1 promoter.

PLoS One 2013 23;8(4):e62559. Epub 2013 Apr 23.

University Lille Nord de France, Lille, France.

Recent data have underlined a possible role of G(D3) synthase (GD3S) and complex gangliosides in Estrogen Receptor (ER) negative breast cancer progression. Here, we describe the main transcript of the GD3S coding gene ST8SIA1 expressed in breast tumors. We characterized the corresponding core promoter in Hs578T breast cancer cells and showed that estradiol decreases ST8SIA1 mRNA expression in ER-positive MCF-7 cells and ERα-transfected ER-negative Hs578T cells. The activity of the core promoter sequence of ST8SIA1 is also repressed by estradiol. The core promoter of ST8SIA1 contains two putative Estrogen Response Elements (ERE) that were not found to be involved in the promoter activity pathway. However, NFκB was shown to be involved in ST8SIA1 transcriptional activation and we demonstrated that estradiol prevents NFκB to bind to ST8SIA1 core promoter in ERα expressing breast cancer cells by inhibiting p65 and p50 nucleus localization. The activation of NFκB pathway in ER-negative tumors, due to the absence of estradiol signaling, might explain the overexpression of G(D3) synthase in this tumor subtype.
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http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0062559PLOS
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3633854PMC
November 2013

How Do Gangliosides Regulate RTKs Signaling?

Cells 2013 Dec 6;2(4):751-67. Epub 2013 Dec 6.

Structural and Functional Glycobiology Unit, UMR CNRS 8576, University of Sciences and Technologies of Lille, 59655 Villeneuve d'Ascq, France.

Gangliosides, the glycosphingolipids carrying one or several sialic acid residues, are located on the outer leaflet of the plasma membrane in glycolipid-enriched microdomains, where they interact with molecules of signal transduction pathways including receptors tyrosine kinases (RTKs). The role of gangliosides in the regulation of signal transduction has been reported in many cases and in a large number of cell types. In this review, we summarize the current knowledge on the biosynthesis of gangliosides and the mechanism by which they regulate RTKs signaling.
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http://dx.doi.org/10.3390/cells2040751DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3972652PMC
December 2013

Accumulation of unusual gangliosides G(Q3) and G(P3) in breast cancer cells expressing the G(D3) synthase.

Molecules 2012 Aug 10;17(8):9559-72. Epub 2012 Aug 10.

Structural and Functional Glycobiology Unit, UMR CNRS 8576, University of Sciences and Technologies of Lille, 59655 Villeneuve d'Ascq, France.

Glycosphingolipids from the ganglio-series are usually classified in four series according to the presence of 0 to 3 sialic acid residues linked to lactosylceramide. The transfer of sialic acid is catalyzed in the Golgi apparatus by specific sialyltransferases that show high specificity toward glycolipid substrates. ST8Sia I (EC 2.4.99.8, SAT-II, SIAT 8a) is the key enzyme controlling the biosynthesis of b- and c-series gangliosides. ST8Sia I is expressed at early developmental stages whereas in adult human tissues, ST8Sia I transcripts are essentially detected in brain. ST8Sia I together with b- and c-series gangliosides are also over-expressed in neuroectoderm-derived malignant tumors such as melanoma, glioblastoma, neuroblastoma and in estrogen receptor (ER) negative breast cancer, where they play a role in cell proliferation, migration, adhesion and angiogenesis. We have stably expressed ST8Sia I in MCF-7 breast cancer cells and analyzed the glycosphingolipid composition of wild type (WT) and GD3S+ clones. As shown by mass spectrometry, MCF-7 expressed a complex pattern of neutral and sialylated glycosphingolipids from globo- and ganglio-series. WT MCF-7 cells exhibited classical monosialylated gangliosides including G(M3), G(M2), and G(M1a). In parallel, the expression of ST8Sia I in MCF-7 GD3S+ clones resulted in a dramatic change in ganglioside composition, with the expression of b- and c-series gangliosides as well as unusual tetra- and pentasialylated lactosylceramide derivatives G(Q3) (II(3)Neu5Ac(4)-Gg(2)Cer) and G(P3) (II(3)Neu5Ac(5)-Gg(2)Cer). This indicates that ST8Sia I is able to act as an oligosialyltransferase in a cellular context.
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http://dx.doi.org/10.3390/molecules17089559DOI Listing
http://www.ncbi.nlm.nih.gov/pmc/articles/PMC6268691PMC
August 2012

Iminosugar-ferrocene conjugates as potential anticancer agents.

Org Biomol Chem 2012 Aug 20;10(29):5592-7. Epub 2012 Jun 20.

Université de Reims Champagne-Ardenne, Institut de Chimie Moléculaire de Reims, CNRS UMR 7312, UFR des Sciences Exactes et Naturelles, 51687 Reims Cedex 2, France.

We prepared a series of new iminosugar-ferrocene hybrids displaying potent inhibition of fucosidase (bovine kidney) and inactivation of MDA-MB-231 breast cancer cells proliferation at low micromolar concentrations. The synthetic route brought to light an unprecedented isomerisation of a 2-ethanalylpyrrolidine.
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http://dx.doi.org/10.1039/c2ob25727kDOI Listing
August 2012

TNF regulates sialyl-Lewisx and 6-sulfo-sialyl-Lewisx expression in human lung through up-regulation of ST3GAL4 transcript isoform BX.

Biochimie 2012 Sep 9;94(9):2045-53. Epub 2012 Jun 9.

Université Lille Nord de France, Lille 1, UGSF, CNRS, UMR 8576, 59650 Villeneuve d'Ascq, France.

Bronchial mucins from severely infected patients suffering from lung diseases such as chronic bronchitis or cystic fibrosis exhibit increased amounts of sialyl-Lewis(x) (NeuAcα2-3Galβ1-4[Fucα1-3]GlcNAc-R, sLe(x)) glycan structures. In cystic fibrosis, sLe(x) and its sulfated form 6-sulfo-sialyl-Lewis(x) (NeuAcα2-3Galβ1-4[Fucα1-3](HO(3)S-6)GlcNAc-R, 6-sulfo-sLe(x)) serve as receptors for Pseudomonas aeruginosa and are involved in the chronicity of airway infection. However, little is known about the molecular mechanisms regulating the changes in glycosylation and sulfation of mucins in airways. Herein, we show that the pro-inflammatory cytokine TNF increases the expression of α2,3-sialyltransferase gene ST3GAL4, both in human bronchial mucosa and in A549 lung carcinoma cells. The role of sialyltransferase ST3Gal IV in sLe(x) biosynthesis was confirmed by siRNA silencing of ST3GAL4 gene. BX is the major transcript isoform expressed in healthy bronchial mucosa and in A549 cells, and is up-regulated by TNF in both models. Bioinformatics analysis and luciferase assays have confirmed that the 2 kb genomic sequence surrounding BX exon contains a promoter region regulated by TNF-related transcription factors. These results support further work aiming at the development of anti-inflammatory strategy to reduce chronic airway infection in diseases such as cystic fibrosis.
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http://dx.doi.org/10.1016/j.biochi.2012.05.030DOI Listing
September 2012

The ganglioside G(D2) induces the constitutive activation of c-Met in MDA-MB-231 breast cancer cells expressing the G(D3) synthase.

Glycobiology 2012 Jun 1;22(6):806-16. Epub 2012 Feb 1.

Université Lille Nord de France, F-59000 Lille, France.

We have recently established and characterized cellular clones deriving from MDA-MB-231 breast cancer cells that express the human G(D3) synthase (GD3S), the enzyme that controls the biosynthesis of b- and c-series gangliosides. The GD3S positive clones show a proliferative phenotype in the absence of serum or growth factors and an increased tumor growth in severe immunodeficient mice. This phenotype results from the constitutive activation of the receptor tyrosine kinase c-Met in spite of the absence of ligand and subsequent activation of mitogen-activated protein kinase/extracellular signal-regulated kinase and phosphoinositide 3-kinase/Akt pathways. Here, we show by mass spectrometry analysis of total glycosphingolipids that G(D3) and G(D2) are the main gangliosides expressed by the GD3S positive clones. Moreover, G(D2) colocalized with c-Met at the plasma membrane and small interfering RNA silencing of the G(M2)/G(D2) synthase efficiently reduced the expression of G(D2) as well as c-Met phosphorylation and reversed the proliferative phenotype. Competition assays using anti-G(D2) monoclonal antibodies also inhibit proliferation and c-Met phosphorylation of GD3S positive clones in serum-free conditions. Altogether, these results demonstrate the involvement of the disialoganglioside G(D2) in MDA-MB-231 cell proliferation via the constitutive activation of c-Met. The accumulation of G(D2) in c-Met expressing cells could therefore reinforce the tumorigenicity and aggressiveness of breast cancer tumors.
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http://dx.doi.org/10.1093/glycob/cws049DOI Listing
June 2012