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1. hnRNP‐A1 binds to the IRES of MELOE‐1 antigen to promote MELOE‐1 translation in stressed melanoma cells

Author

Maud Charpentier, Emilie Dupré, Agnès Fortun, Floriane Briand, Mike Maillasson, Emmanuelle Com, Charles Pineau, Nathalie Labarrière, Catherine Rabu, and François Lang

Subjects
ER stress, IRES, ITAF, long noncoding RNA, melanoma, tumor antigens, Neoplasms. Tumors. Oncology. Including cancer and carcinogens, RC254-282
Abstract

The major challenge in antigen‐specific immunotherapy of cancer is to select the most relevant tumor antigens to target. To this aim, understanding their mode of expression by tumor cells is critical. We previously identified a melanoma‐specific antigen, melanoma‐overexpressed antigen 1 (MELOE‐1)—coded for by a long noncoding RNA—whose internal ribosomal entry sequence (IRES)‐dependent translation is restricted to tumor cells. This restricted expression is associated with the presence of a broad‐specific T‐cell repertoire that is involved in tumor immunosurveillance in melanoma patients. In the present work, we explored the translation control of MELOE‐1 and provide evidence that heterogeneous nuclear ribonucleoprotein A1 (hnRNP‐A1) binds to the MELOE‐1 IRES and acts as an IRES trans‐activating factor (ITAF) to promote the translation of MELOE‐1 in melanoma cells. In addition, we showed that endoplasmic reticulum (ER) stress induced by thapsigargin, which promotes hnRNP‐A1 cytoplasmic translocation, enhances MELOE‐1 translation and recognition of melanoma cells by a MELOE‐1‐specific T‐cell clone. These findings suggest that pharmacological stimulation of stress pathways may enhance the efficacy of immunotherapies targeting stress‐induced tumor antigens such as MELOE‐1.

Published
2022
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2. Selective Targeting of IL-15Rα Is Sufficient to Reduce Inflammation

Author

Dihia Meghnem, Mike Maillasson, Isabelle Barbieux, Sébastien Morisseau, Dalloba Keita, Yannick Jacques, Agnès Quéméner, and Erwan Mortier

Subjects
interleukin, receptor, IL-2, inhibition, homeostasis, Immunologic diseases. Allergy, RC581-607
Abstract

Cytokines are crucial molecules for maintaining the proper functioning of the immune system. Nevertheless, a dysregulation of cytokine expression could be involved in the pathogenesis of autoimmune diseases. Interleukin (IL)-15 is a key factor for natural killer cells (NK) and CD8 T cells homeostasis, necessary to fight cancer and infections but could also be considered as a pro-inflammatory cytokine involved in autoimmune inflammatory disease, including rheumatoid arthritis, psoriasis, along with tumor necrosis factor alpha (TNF-α), IL-6, and IL-1β. The molecular mechanisms by which IL-15 exerts its inflammatory function in these diseases are still unclear. In this study, we generated an IL-15-derived molecule called NANTIL-15 (New ANTagonist of IL-15), designed to selectively inhibit the action of IL-15 through the high-affinity trimeric IL-15Rα/IL-2Rβ/γc receptor while leaving IL-15 signaling through the dimeric IL-2Rβ/γc receptor unaffected. Administrating of NANTIL-15 in healthy mice did not affect the IL-15-dependent cell populations such as NK and CD8 T cells. In contrast, we found that NANTIL-15 efficiently reduced signs of inflammation in a collagen-induced arthritis model. These observations demonstrate that the inflammatory properties of IL-15 are linked to its action through the trimeric IL-15Rα/IL-2Rβ/γc receptor, highlighting the interest of selectively targeting this receptor.

Published
2022
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3. Two-Chains Tissue Plasminogen Activator Unifies Met and NMDA Receptor Signalling to Control Neuronal Survival

Author

Elodie Hedou, Sara Douceau, Arnaud Chevilley, Alexandre Varangot, Audrey M. Thiebaut, Hortense Triniac, Isabelle Bardou, Carine Ali, Mike Maillasson, Tiziana Crepaldi, Paolo Comoglio, Eloïse Lemarchand, Véronique Agin, Benoit D. Roussel, and Denis Vivien

Subjects
neuronal death, tissue-type plasminogen activator, NMDA receptor, MET receptor, Biology (General), QH301-705.5, Chemistry, QD1-999
Abstract

Tissue-type plasminogen activator (tPA) plays roles in the development and the plasticity of the nervous system. Here, we demonstrate in neurons, that by opposition to the single chain form (sc-tPA), the two-chains form of tPA (tc-tPA) activates the MET receptor, leading to the recruitment of N-Methyl-d-Aspartate receptors (NMDARs) and to the endocytosis and proteasome-dependent degradation of NMDARs containing the GluN2B subunit. Accordingly, tc-tPA down-regulated GluN2B-NMDAR-driven signalling, a process prevented by blockers of HGFR/MET and mimicked by its agonists, leading to a modulation of neuronal death. Thus, our present study unmasks a new mechanism of action of tPA, with its two-chains form mediating a crosstalk between MET and the GluN2B subunit of NMDARs to control neuronal survival.

Published
2021
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4. Functional characterization of a chimeric soluble Fas ligand polymer with in vivo anti-tumor activity.

Author

Sophie Daburon, Christel Devaud, Pierre Costet, Aurore Morello, Laure Garrigue-Antar, Mike Maillasson, Nathalie Hargous, Delphine Lapaillerie, Marc Bonneu, Julie Dechanet-Merville, Patrick Legembre, Myriam Capone, Jean-François Moreau, and Jean-Luc Taupin

Subjects
Medicine, Science
Abstract

Binding of ligand FasL to its receptor Fas triggers apoptosis via the caspase cascade. FasL itself is homotrimeric, and a productive apoptotic signal requires that FasL be oligomerized beyond the homotrimeric state. We generated a series of FasL chimeras by fusing FasL to domains of the Leukemia Inhibitory Factor receptor gp190 which confer homotypic oligomerization, and analyzed the capacity of these soluble chimeras to trigger cell death. We observed that the most efficient FasL chimera, called pFasL, was also the most polymeric, as it reached the size of a dodecamer. Using a cellular model, we investigated the structure-function relationships of the FasL/Fas interactions for our chimeras, and we demonstrated that the Fas-mediated apoptotic signal did not solely rely on ligand-mediated receptor aggregation, but also required a conformational adaptation of the Fas receptor. When injected into mice, pFasL did not trigger liver injury at a dose which displayed anti-tumor activity in a model of human tumor transplanted to immunodeficient animals, suggesting a potential therapeutic use. Therefore, the optimization of the FasL conformation has to be considered for the development of efficient FasL-derived anti-cancer drugs targeting Fas.

Published
2013
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5. hnRNP‐A1 binds to the IRES of MELOE‐1 antigen to promote MELOE‐1 translation in stressed melanoma cells

Author

Emmanuelle Com, François Lang, Charles Pineau, Catherine Rabu, Nathalie Labarrière, Agnès Fortun, Emilie Dupré, Floriane Briand, Maud Charpentier, and Mike Maillasson

Subjects
Cancer Research, Heterogeneous Nuclear Ribonucleoprotein A1, medicine.medical_treatment, Internal Ribosome Entry Sites, Biology, 03 medical and health sciences, 0302 clinical medicine, Antigen, IRES, Antigens, Neoplasm, melanoma, Genetics, medicine, Humans, long noncoding RNA, RC254-282, Research Articles, 030304 developmental biology, 0303 health sciences, Melanoma, Endoplasmic reticulum, Neoplasms. Tumors. Oncology. Including cancer and carcinogens, Translation (biology), General Medicine, Immunotherapy, medicine.disease, ITAF, Neoplasm Proteins, 3. Good health, Cell biology, Internal ribosome entry site, tumor antigens, Oncology, Protein Biosynthesis, 030220 oncology & carcinogenesis, Unfolded protein response, Molecular Medicine, ER stress, Ribosomes, Research Article
Abstract

The major challenge in antigen‐specific immunotherapy of cancer is to select the most relevant tumor antigens to target. To this aim, understanding their mode of expression by tumor cells is critical. We previously identified a melanoma‐specific antigen, melanoma‐overexpressed antigen 1 (MELOE‐1)—coded for by a long noncoding RNA—whose internal ribosomal entry sequence (IRES)‐dependent translation is restricted to tumor cells. This restricted expression is associated with the presence of a broad‐specific T‐cell repertoire that is involved in tumor immunosurveillance in melanoma patients. In the present work, we explored the translation control of MELOE‐1 and provide evidence that heterogeneous nuclear ribonucleoprotein A1 (hnRNP‐A1) binds to the MELOE‐1 IRES and acts as an IRES trans‐activating factor (ITAF) to promote the translation of MELOE‐1 in melanoma cells. In addition, we showed that endoplasmic reticulum (ER) stress induced by thapsigargin, which promotes hnRNP‐A1 cytoplasmic translocation, enhances MELOE‐1 translation and recognition of melanoma cells by a MELOE‐1‐specific T‐cell clone. These findings suggest that pharmacological stimulation of stress pathways may enhance the efficacy of immunotherapies targeting stress‐induced tumor antigens such as MELOE‐1., Our study demonstrates that reticular stress in melanoma cells promoted the translocation of the IRES transacting factor hnRPN‐A1 to the cytoplasm. By binding to the IRES site of meloe mRNA, hnRPN‐A1 promoted the translation of the melanoma‐specific antigen MELOE‐1 and led to the expression of a new immunogenic HLA/peptide complex at the cell surface. Altogether, our data suggest that pharmacological stimulation of stress pathways may enhance the efficacy of T‐cell based immunotherapies in targeting stress‐induced tumor antigens, such as MELOE‐1 in patients with melanoma.

Published
2021

7. IL-15Rα membrane-anchorage either in cis or in trans is required for stabilization of IL-15 and optimal signaling

Author

Jean-Yves Le Questel, Rui P. Sousa, Johann Dion, Agnès Quéméner, Kilian Trillet, Isabelle Barbieux, Adèle D. Laurent, Erwan Mortier, Isabelle Leray, Marie Frutoso, Mike Maillasson, Yannick Jacques, Sébastien Morisseau, Immunobiology of Human αβ and γδ T Cells and Immunotherapeutic Applications (CRCINA-ÉQUIPE 1), Centre de Recherche en Cancérologie et Immunologie Nantes-Angers (CRCINA), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), Université de Nantes (UN)-Université de Nantes (UN)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Centre National de la Recherche Scientifique (CNRS)-Université d'Angers (UA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), Université de Nantes (UN)-Université de Nantes (UN)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Centre National de la Recherche Scientifique (CNRS)-Université d'Angers (UA), LabEx IGO 'Immunotherapy, Graft, Oncology' [Nantes], Centre hospitalier universitaire de Nantes (CHU Nantes), Chimie Et Interdisciplinarité : Synthèse, Analyse, Modélisation (CEISAM), Université de Nantes - UFR des Sciences et des Techniques (UN UFR ST), Université de Nantes (UN)-Université de Nantes (UN)-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC), Structure fédérative de recherche François Bonamy (SFR François Bonamy), Université de Nantes (UN)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche en Santé de l'Université de Nantes (IRS-UN), This work was also supported by the CNRS, Inserm, the University of Nantes, and realized in the context of the IHU-Cesti (ANR-10-IBHU-005) project which received French government financial support managed by the National Research Agency and supported by Nantes Métropole and Pays de la Loire Région. The MD simulations were partly carried out using HPC resources from the CCIPL (Centre de Calcul Intensif des Pays de la Loire)., ANR-10-IBHU-0005,CESTI (TSI-IHU),Centre Européen des Sciences de la Transplantation et de l'Immunothérapie (TSI-IHU)(2010), Bernardo, Elizabeth, Instituts Hospitalo-Universitaires B - Centre Européen des Sciences de la Transplantation et de l'Immunothérapie (TSI-IHU) - - CESTI (TSI-IHU)2010 - ANR-10-IBHU-0005 - IBHU - VALID, Université d'Angers (UA)-Université de Nantes (UN)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Université d'Angers (UA)-Université de Nantes (UN)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Centre hospitalier universitaire de Nantes (CHU Nantes), LabEX IGO Immunothérapie Grand Ouest, Nantes Université (Nantes Univ), and Université de Nantes (UN)-Université de Nantes (UN)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)

Subjects
[SDV.IMM] Life Sciences [q-bio]/Immunology, Interaction, medicine.medical_treatment, Cell, T cells, [SDV.CAN]Life Sciences [q-bio]/Cancer, NK cells, Biology, 03 medical and health sciences, 0302 clinical medicine, Immune system, [SDV.CAN] Life Sciences [q-bio]/Cancer, Contact, medicine, Receptor, Cytokine, 030304 developmental biology, 0303 health sciences, Interleukin, Cell Biology, Cell biology, medicine.anatomical_structure, Membrane, Interleukin 15, [SDV.IMM]Life Sciences [q-bio]/Immunology, Trans-acting, 030217 neurology & neurosurgery
Abstract

Interleukin (IL)-15 plays an important role in the communication between immune cells. It delivers its signal through different modes involving three receptor chains: IL-15Rα, IL-2Rβ and IL-2Rγc. The combination of the different chains result in the formation of IL-15Rα/IL-2Rβ/γc trimeric or IL-2Rβ/γc dimeric receptors. In this study, we have investigated the role of the IL-15Rα chain in stabilizing the cytokine in the IL-2Rβ/γc dimeric receptor. By analyzing the key amino acid residues of IL-15 facing IL-2Rβ, we provide evidence of differential interfaces in the presence or in the absence of membrane-anchored IL-15Rα. Moreover, we found that the anchorage of IL-15Rα to the cell surface regardless its mode of presentation – i.e. cis or trans – is crucial for complete signaling. These observations show how the cells can finely modulate the intensity of cytokine signaling through the quality and the level of expression of the receptor chains.

Published
2019

8. Rational modification, synthesis and biological evaluation of N-substituted phthalazinone derivatives designed to target interleukine-15 protein

Author

Monique Mathé-Allainmat, Jacques Lebreton, Jimmy Smadja, Laurence Arzel, Agnès Quéméner, Erwan Mortier, Benoit Sicard, Didier Dubreuil, Aurélien Leray, Mike Maillasson, Centre de Recherche en Cancérologie et Immunologie Nantes-Angers (CRCINA), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), Université de Nantes (UN)-Université de Nantes (UN)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Centre National de la Recherche Scientifique (CNRS)-Université d'Angers (UA), Immunobiology of Human αβ and γδ T Cells and Immunotherapeutic Applications (CRCINA-ÉQUIPE 1), Université de Nantes (UN)-Université de Nantes (UN)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Centre National de la Recherche Scientifique (CNRS)-Université d'Angers (UA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), LabEx IGO 'Immunotherapy, Graft, Oncology' [Nantes], Chimie Et Interdisciplinarité : Synthèse, Analyse, Modélisation (CEISAM), Centre National de la Recherche Scientifique (CNRS)-Université de Nantes - UFR des Sciences et des Techniques (UN UFR ST), Université de Nantes (UN)-Université de Nantes (UN), Plateforme IMPACT Nantes (CRCINA-IMPACT), Centre de Recherche en Cancérologie Nantes-Angers (CRCNA), Centre Hospitalier Universitaire d'Angers (CHU Angers), PRES Université Nantes Angers Le Mans (UNAM)-PRES Université Nantes Angers Le Mans (UNAM)-Hôtel-Dieu de Nantes-Institut National de la Santé et de la Recherche Médicale (INSERM)-Hôpital Laennec-Centre National de la Recherche Scientifique (CNRS)-Faculté de Médecine d'Angers-Centre hospitalier universitaire de Nantes (CHU Nantes)-Centre Hospitalier Universitaire d'Angers (CHU Angers), PRES Université Nantes Angers Le Mans (UNAM)-PRES Université Nantes Angers Le Mans (UNAM)-Hôtel-Dieu de Nantes-Institut National de la Santé et de la Recherche Médicale (INSERM)-Hôpital Laennec-Centre National de la Recherche Scientifique (CNRS)-Faculté de Médecine d'Angers-Centre hospitalier universitaire de Nantes (CHU Nantes), Université d'Angers (UA)-Université de Nantes (UN)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Centre hospitalier universitaire de Nantes (CHU Nantes), Université d'Angers (UA)-Université de Nantes (UN)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Université d'Angers (UA)-Université de Nantes (UN)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Centre hospitalier universitaire de Nantes (CHU Nantes), LabEX IGO Immunothérapie Grand Ouest, Nantes Université (Nantes Univ), Université de Nantes - UFR des Sciences et des Techniques (UN UFR ST), Université de Nantes (UN)-Université de Nantes (UN)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Mortier, Erwan, and Université de Nantes (UN)-Université de Nantes (UN)-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC)

Subjects
[SDV.IMM] Life Sciences [q-bio]/Immunology, medicine.medical_treatment, T cell, Clinical Biochemistry, Pharmaceutical Science, autoimmune disease, 01 natural sciences, Biochemistry, Cell Line, Structure-Activity Relationship, Drug Discovery, medicine, Humans, Receptor, Molecular Biology, Interleukin-15, Virtual screening, 010405 organic chemistry, Chemistry, Spectrum Analysis, Organic Chemistry, Interleukin, Protein-protein interaction PPI, inflammatory disease, 0104 chemical sciences, 3. Good health, Cell biology, inhibitor, 010404 medicinal & biomolecular chemistry, Cytokine, medicine.anatomical_structure, Interleukin 15, Phthalazines, [SDV.IMM]Life Sciences [q-bio]/Immunology, Molecular Medicine, IL-15Rbeta, Pharmacophore, CD8
Abstract

International audience; Interleukin (IL)-15 is a pleiotropic cytokine structurally close to IL-2 and sharing with the IL-2Rβ and γc receptor (R) subunits. IL-15 plays important roles in innate and adaptative immunity, supporting the activation and proliferation of NK, NK-T, and CD8 + T cells. Over-expression of IL-15 has been shown to participate to the development of inflammatory and autoimmune diseases and diverse T cell malignancies. This study is in continuity of our previous work through which a family of small-molecule inhibitors impeding IL-15/IL-2Rβ interaction with sub-micromolar activity has been identified using pharmacophore-based virtual screening and hit optimization methods. With the aim to improve the efficacy and selectivity of our lead inhibitor, specific modifications have been introduced on the basis of optimized SAR and modelisation. The new series of compounds generated have been evaluated for their capacity to inhibit the proliferation as well as the downstream signaling of IL-15-dependent cells and to bind to IL-15.

Published
2021

9. Discovery of a Small-Molecule Inhibitor of Interleukin 15: Pharmacophore-Based Virtual Screening and Hit Optimization

Author

Yannick Jacques, Monique Mathé-Allainmat, Laurence Arzel, Mike Maillasson, Erwan Mortier, Jacques Lebreton, Romy Vomiandry, Didier Dubreuil, Agnès Quéméner, Benoit Sicard, Centre de Recherche en Cancérologie et Immunologie Nantes-Angers (CRCINA), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), Université de Nantes (UN)-Université de Nantes (UN)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Centre National de la Recherche Scientifique (CNRS)-Université d'Angers (UA), Chimie Et Interdisciplinarité : Synthèse, Analyse, Modélisation (CEISAM), Université de Nantes - UFR des Sciences et des Techniques (UN UFR ST), Université de Nantes (UN)-Université de Nantes (UN)-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC), This work was funded by the programs CIMATH2 (Ciblage Moléculaire et Applications Thérapeutiques 2) and PIRAMID (Protein‐protein Interactions in Rational Approaches for Medicinal Innovative Drugs) supported by La Région Pays de la Loire. R.V. was supported by a fellowship from La Région Pays de la Loire. We thank CHEM)Symbiose and IMPACT facilities for technical assistance., Bernardo, Elizabeth, Université d'Angers (UA)-Université de Nantes (UN)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Centre hospitalier universitaire de Nantes (CHU Nantes), and Université de Nantes (UN)-Université de Nantes (UN)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)

Subjects
0301 basic medicine, medicine.medical_treatment, [SDV.CAN]Life Sciences [q-bio]/Cancer, 01 natural sciences, Cell Line, Small Molecule Libraries, Mice, Structure-Activity Relationship, 03 medical and health sciences, [SDV.CAN] Life Sciences [q-bio]/Cancer, Drug Discovery, medicine, Animals, Humans, Receptor, Cell Proliferation, Interleukin-15, Virtual screening, 010405 organic chemistry, Chemistry, Interleukin, Stereoisomerism, Triazoles, Small molecule, 0104 chemical sciences, 3. Good health, Cell biology, Interleukin-2 Receptor beta Subunit, Molecular Docking Simulation, 030104 developmental biology, Cytokine, Interleukin 15, Immunology, Phthalazines, Molecular Medicine, Pharmacophore, Databases, Chemical, CD8
Abstract

International audience; Interleukin (IL)-15 is a pleiotropic cytokine, which is structurally close to IL-2 and shares with it the IL-2 β and γ receptor (R) subunits. By promoting the activation and proliferation of NK, NK-T and CD8+ T cells, IL-15 plays important roles in innate and adaptative immunity. Moreover, the association of high levels of IL-15 expression with inflammatory and autoimmune diseases has led to the development of various antagonistic approaches targeting IL- 15. This study is an original approach aimed at discovering small-molecule inhibitors impeding IL-15/IL-15R interaction. A pharmacophore and docking-based virtual screening of compound libraries led to the selection of 240 high-scoring compounds, 36 of which were found to bind IL-15, to inhibit the binding of IL-15 to the IL-2Rβ chain and/or the proliferation of IL-15-dependent cells. One of them was selected as a hit, optimized by a structure-activity relationship approach, leading to the first small-molecule IL-15 inhibitor with sub-micromolar activity.

Published
2017

10. IL-15Rα membrane anchorage in either

Author

Agnès, Quéméner, Sébastien, Morisseau, Rui P, Sousa, Kilian, Trillet, Mike, Maillasson, Isabelle, Leray, Yannick, Jacques, Johann, Dion, Isabelle, Barbieux, Marie, Frutoso, Adèle D, Laurent, Jean-Yves, Le Questel, and Erwan, Mortier

Subjects
Interleukin-15, Interleukin-2 Receptor beta Subunit, Models, Molecular, Epitopes, Binding Sites, Receptors, Interleukin-15, Multiprotein Complexes, Humans, Crystallography, X-Ray, Cell Line, Signal Transduction
Abstract

Interleukin (IL)-15 plays an important role in the communication between immune cells. It delivers its signal through different modes involving three receptor chains: IL-15Rα, IL-2Rβ and IL-2Rγc. The combination of the different chains result in the formation of IL-15Rα/IL-2Rβ/γc trimeric or IL-2Rβ/γc dimeric receptors. In this study, we have investigated the role of the IL-15Rα chain in stabilizing the cytokine in the IL-2Rβ/γc dimeric receptor. By analyzing the key amino acid residues of IL-15 facing IL-2Rβ, we provide evidence of differential interfaces in the presence or in the absence of membrane-anchored IL-15Rα. Moreover, we found that the anchorage of IL-15Rα to the cell surface regardless its mode of presentation - i.e.

Published
2019

11. Multivalent Thiosialosides and Their Synergistic Interaction with Pathogenic Sialidases

Author

Benoît Roubinet, Charles Tellier, Julien Bernard, Clarisse Bridot, Coralie Assailly, Franck Daligault, Clémence Petitot, Cyrille Grandjean, Julie Bouckaert, Blanka Didak, Amélie Saumonneau, Xibo Yan, Mike Maillasson, Sébastien G. Gouin, David Deniaud, Ralf Blossey, Yoan Brissonnet, Ludovic Landemarre, Chimie Et Interdisciplinarité : Synthèse, Analyse, Modélisation (CEISAM), Université de Nantes - UFR des Sciences et des Techniques (UN UFR ST), Université de Nantes (UN)-Université de Nantes (UN)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Unité de fonctionnalité et ingénierie de protéines (UFIP), Université de Nantes (UN)-Université de Nantes (UN)-Centre National de la Recherche Scientifique (CNRS), Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS), IMPACT - Interactions Moléculaires Puces ACTivités, Centre de Recherche en Cancérologie et Immunologie Nantes-Angers (CRCINA), Université d'Angers (UA)-Université de Nantes (UN)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Université d'Angers (UA)-Université de Nantes (UN)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Centre hospitalier universitaire de Nantes (CHU Nantes), Immunobiology of Human αβ and γδ T Cells and Immunotherapeutic Applications (CRCINA-ÉQUIPE 1), Chimie Organique et Bioorganique : Réactivité et Analyse (COBRA), Institut de Chimie Organique Fine (IRCOF), Université de Rouen Normandie (UNIROUEN), Normandie Université (NU)-Normandie Université (NU)-Institut national des sciences appliquées Rouen Normandie (INSA Rouen Normandie), Institut National des Sciences Appliquées (INSA)-Normandie Université (NU)-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS)-Université de Rouen Normandie (UNIROUEN), Institut National des Sciences Appliquées (INSA)-Normandie Université (NU)-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS)-Institut Normand de Chimie Moléculaire Médicinale et Macromoléculaire (INC3M), Université de Caen Normandie (UNICAEN), Normandie Université (NU)-Normandie Université (NU)-École Nationale Supérieure d'Ingénieurs de Caen (ENSICAEN), Normandie Université (NU)-Université Le Havre Normandie (ULH), Normandie Université (NU)-Université de Rouen Normandie (UNIROUEN), Normandie Université (NU)-Institut national des sciences appliquées Rouen Normandie (INSA Rouen Normandie), Institut National des Sciences Appliquées (INSA)-Normandie Université (NU)-Institut National des Sciences Appliquées (INSA)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Université de Caen Normandie (UNICAEN), Normandie Université (NU)-École Nationale Supérieure d'Ingénieurs de Caen (ENSICAEN), Normandie Université (NU)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Centre National de la Recherche Scientifique (CNRS), GLYcoDiag, Ingénierie des Matériaux Polymères (IMP), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Université Jean Monnet - Saint-Étienne (UJM)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Unité de Biotechnologie, Biocatalyse et Biorégulation (U3B), Université de Nantes (UN)-Centre National de la Recherche Scientifique (CNRS), University of Bern, ANR-17-CE07-0028,HICARE,Inhibiteurs hétérovalents de sialidases et trans-sialidases(2017), Université de Nantes (UN)-Université de Nantes (UN)-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC), Institut National de la Recherche Agronomique (INRA)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), Université de Nantes (UN)-Université de Nantes (UN)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Centre National de la Recherche Scientifique (CNRS)-Université d'Angers (UA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), Université de Nantes (UN)-Université de Nantes (UN)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Centre National de la Recherche Scientifique (CNRS)-Université d'Angers (UA), Institut Normand de Chimie Moléculaire Médicinale et Macromoléculaire (INC3M), Institut de Chimie du CNRS (INC)-École Nationale Supérieure d'Ingénieurs de Caen (ENSICAEN), Institut National des Sciences Appliquées (INSA)-Normandie Université (NU)-Institut National des Sciences Appliquées (INSA)-Université Le Havre Normandie (ULH), Normandie Université (NU)-Centre National de la Recherche Scientifique (CNRS)-Université de Caen Normandie (UNICAEN), Normandie Université (NU)-Institut de Chimie du CNRS (INC)-École Nationale Supérieure d'Ingénieurs de Caen (ENSICAEN), Normandie Université (NU)-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie Organique Fine (IRCOF), Institut National des Sciences Appliquées (INSA)-Normandie Université (NU)-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS)-Centre National de la Recherche Scientifique (CNRS), GLYcoDIAG, Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Université Jean Monnet [Saint-Étienne] (UJM)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Université de Nantes - Faculté des Sciences et des Techniques, Transport des assimilats (TA), Université de Poitiers-Centre National de la Recherche Scientifique (CNRS), Unité de Glycobiologie Structurale et Fonctionnelle - UMR 8576 (UGSF), Université de Lille-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Recherche Agronomique (INRA), Centre de recherche de Cancérologie et d'Immunologie / Nantes - Angers (CRCINA), Université d'Angers (UA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), Université de Nantes (UN)-Université de Nantes (UN)-Université d'Angers (UA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), Université de Nantes (UN)-Université de Nantes (UN), Immunobiology of Human αβ and γδ T cells and Immunotherapeutic Applications (CRCINA - Département INCIT - Equipe 1), Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie Organique Fine (IRCOF), Institut National des Sciences Appliquées (INSA)-Normandie Université (NU)-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS), Institut de Recherche Interdisciplinaire [Villeneuve d'Ascq] (IRI), Université de Lille, Sciences et Technologies-Université de Lille, Droit et Santé-Centre National de la Recherche Scientifique (CNRS), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Institut National des Sciences Appliquées (INSA)-Université Jean Monnet [Saint-Étienne] (UJM)-Centre National de la Recherche Scientifique (CNRS), Université de Lille, CNRS, Unité de Glycobiologie Structurale et Fonctionnelle UMR 8576 [UGSF], and Unité de Glycobiologie Structurale et Fonctionnelle (UGSF) - UMR 8576

Subjects
chemistry.chemical_classification, 010405 organic chemistry, Glycoconjugate, Organic Chemistry, enzymes, glycoclusters, glycosidases, inhibition, sialidases, Virulence, Pathogenic bacteria, General Chemistry, 010402 general chemistry, medicine.disease_cause, Sialidase, 01 natural sciences, Catalysis, 0104 chemical sciences, Enzyme, Biochemistry, chemistry, medicine, [CHIM]Chemical Sciences, Catalytic efficiency, Eukaryotic cell
Abstract

International audience; Sialidases (SAs) hydrolyze sialyl residues from glycoconjugates of the eukaryotic cell surface and are virulence factors expressed by pathogenic bacteria, viruses, and parasites. The catalytic domains of SAs are often flanked with carbohydrate-binding module(s) previously shown to bind sialosides and to enhance enzymatic catalytic efficiency. Herein, non-hydrolyzable multivalent thiosialosides were designed as probes and inhibitors of V. cholerae, T. cruzi, and S. pneumoniae (NanA) sialidases. NanA was truncated from the catalytic and lectinic domains (NanA-L and NanA-C) to probe their respective roles upon interacting with sialylated surfaces and the synthetically designed di- and polymeric thiosialosides. The NanA-L domain was shown to fully drive NanA binding, improving affinity for the thiosialylated surface and compounds by more than two orders of magnitude. Importantly, each thiosialoside grafted onto the polymer was also shown to reduce NanA and NanA-C catalytic activity with efficiency that was 3000-fold higher than that of the monovalent thiosialoside reference. These results extend the concept of multivalency for designing potent bacterial and parasitic sialidase inhibitors.

Published
2019

12. A novel, smaller scaffold for Affitins: Showcase with binders specific for EpCAM

Author

Valentina Kalichuk, Véronique Préat, Gonzalo Obal, Frédéric Pecorari, Mike Maillasson, Federico Carrión, Barbara Mouratou, Ghislaine Béhar, Axelle Renodon-Cornière, Nuclear Oncology (CRCINA-ÉQUIPE 13), Centre de Recherche en Cancérologie et Immunologie Nantes-Angers (CRCINA), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), Université de Nantes (UN)-Université de Nantes (UN)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Centre National de la Recherche Scientifique (CNRS)-Université d'Angers (UA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), Université de Nantes (UN)-Université de Nantes (UN)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Centre National de la Recherche Scientifique (CNRS)-Université d'Angers (UA), Advanced Drug Delivery and Biomaterials [Brussels, Belgium], Université Catholique de Louvain = Catholic University of Louvain (UCL)-Louvain Drug Research Institute (LDRI), Protein Biophysics [Montevideo] (UBP), Institut Pasteur de Montevideo, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP), IMPACT - Interactions Moléculaires Puces ACTivités, European Erasmus Mundus Joint Doctorate in nanomedicine and pharmaceutical innovation, Nanofar, Bernardo, Elizabeth, and Université d'Angers (UA)-Université de Nantes (UN)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Université d'Angers (UA)-Université de Nantes (UN)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Centre hospitalier universitaire de Nantes (CHU Nantes)

Subjects
0301 basic medicine, Affitin, Archaeal Proteins, [SDV]Life Sciences [q-bio], Bioengineering, Protein Engineering, Applied Microbiology and Biotechnology, 03 medical and health sciences, chemistry.chemical_compound, Affinity chromatography, EpCAM protein engineering, Humans, Aho7c, Thermostability, chemistry.chemical_classification, 030102 biochemistry & molecular biology, biology, ribosome display, Circular Dichroism, Protein engineering, biology.organism_classification, Epithelial Cell Adhesion Molecule, Recombinant Proteins, 3. Good health, Sulfolobus, Amino acid, [SDV] Life Sciences [q-bio], DNA-Binding Proteins, 030104 developmental biology, Biochemistry, chemistry, Sac7d, Ribosome display, Biotechnology, Acidianus, Protein Binding
Abstract

International audience; Affitins are highly stable engineered affinity proteins, originally derived from Sac7d and Sso7d, two 7 kDa DNA-binding polypeptides from Sulfolobus genera. Their efficiency as reagents for intracellular targeting, enzyme inhibition, affinity purification, immunolocalization, and various other applications has been demonstrated. Recently, we have characterized the 7 kDa DNA-binding family, and Aho7c originating from Acidianus hospitalis was shown to be its smallest member with thermostability comparable to those of Sac7d and Sso7d. Here, after four rounds of selection by ribosome display against the human recombinant Epithelial Cell Adhesion Molecule (hrEpCAM), we obtained novel Aho7c-based Affitins. The binders were expressed in soluble form in Escherichia coli, displayed high stability (up to 74°C; pH 0-12) and were shown to be specific for the hrEpCAM extracellular domain with picomolar affinities (KD = 110 pM). Thus, we propose Aho7c as a good candidate for the creation of Affitins with a 10% smaller size than the Sac7d-based ones (60 vs. 66 amino acids).

Published
2017

13. Enriching a cellulose hydrogel with a biologically active marine exopolysaccharide for cell-based cartilage engineering

Author

Sarah Beck-Cormier, Gildas Rethore, Borhane H. Fellah, Sylvia Colliec-Jouault, Claire Vinatier, Pierre Weiss, Yannick Jacques, Emilie Rederstorff, Eva Mathieu, Jérôme Guicheux, Mike Maillasson, and Sophie Sourice

Subjects
0301 basic medicine, Chemistry, Cartilage, technology, industry, and agriculture, Biomedical Engineering, Type II collagen, Medicine (miscellaneous), Chondrogenesis, Cell biology, body regions, Biomaterials, Extracellular matrix, 03 medical and health sciences, 030104 developmental biology, medicine.anatomical_structure, Tissue engineering, Self-healing hydrogels, medicine, Cartilaginous Tissue, Aggrecan, Biomedical engineering
Abstract

The development of biologically and mechanically competent hydrogels is a prerequisite in cartilage engineering. We recently demonstrated that a marine exopolysaccharide, GY785, stimulates the in vitro chondrogenesis of adipose stromal cells. In the present study, we thus hypothesized that enriching our silated hydroxypropyl methylcellulose hydrogel (Si-HPMC) with GY785 might offer new prospects in the development of scaffolds for cartilage regeneration. The interaction properties of GY785 with growth factors was tested by surface plasmon resonance (SPR). The biocompatibility of Si-HPMC/GY785 towards rabbit articular chondrocytes (RACs) and its ability to maintain and recover a chondrocytic phenotype were then evaluated in vitro by MTS assay, cell counting and qRT-PCR. Finally, we evaluated the potential of Si-HPMC/GY785 associated with RACs to form cartilaginous tissue in vivo by transplantation into the subcutis of nude mice for 3 weeks. Our SPR data indicated that GY785 was able to physically interact with BMP-2 and TGFβ. Our analyses also showed that three-dimensionally (3D)-cultured RACs into Si-HPMC/GY785 strongly expressed type II collagen (COL2) and aggrecan transcripts when compared to Si-HPMC alone. In addition, RACs also produced large amounts of extracellular matrix (ECM) containing glycosaminoglycans (GAG) and COL2. When dedifferentiated RACs were replaced in 3D in Si-HPMC/GY785, the expressions of COL2 and aggrecan transcripts were recovered and that of type I collagen decreased. Immunohistological analyses of Si-HPMC/GY785 constructs transplanted into nude mice revealed the production of a cartilage-like extracellular matrix (ECM) containing high amounts of GAG and COL2. These results indicate that GY785-enriched Si-HPMC appears to be a promising hydrogel for cartilage tissue engineering. Copyright © 2015 John Wiley & Sons, Ltd.

Published
2015

14. Switching an anti-IgG binding site between archaeal extremophilic proteins results in Affitins with enhanced pH stability

Author

Ghislaine Béhar, Sabino Pacheco, Barbara Mouratou, Mike Maillasson, and Frédéric Pecorari

Subjects
Hot Temperature, Affitin, Archaeal Proteins, Bioengineering, Biology, Protein Engineering, medicine.disease_cause, Applied Microbiology and Biotechnology, chemistry.chemical_compound, Escherichia coli, medicine, Thermal stability, Binding site, Thermostability, Binding Sites, Wild type, General Medicine, Protein engineering, Hydrogen-Ion Concentration, Antibodies, Anti-Idiotypic, DNA-Binding Proteins, Biochemistry, chemistry, IgG binding, Immunoglobulin G, Mutation, Biophysics, Biotechnology
Abstract

As a useful reagent for biotechnological applications, a scaffold protein needs to be as stable as possible to ensure longer lifetimes. We have developed archaeal extremophilic proteins from the “7 kDa DNA-binding” family as scaffolds to derive affinity proteins (Affitins). In this study, we evaluated a rational structure/sequence-guided approach to stabilize an Affitin derived from Sac7d by transferring its human IgG binding site onto the framework of the more thermally stable Sso7d homolog. The chimera obtained was functional, well expressed in Escherichia coli, but less thermally stable than the original Affitin (T(m) = 74.2 °C vs. T(m) = 80.4 °C). Two single mutations described as thermally stabilizing wild type Sso7d were introduced into chimeras. Only the double mutation nearly restored thermal stability (T(m) = 76.9 °C). Interestingly, the chimera and its double mutant were stable from pH 0 up to at least pH 13. Our results show that it is possible to increase further the stability of Affitins toward alkaline conditions (+2 pH units) while conserving their advantageous properties. As Affitins are based on a growing family of homologs from archaeal extremophiles, we conclude that this approach offers new potential for their improvement, which will be useful in demanding biotechnological applications.

Published
2014

15. Cutting Edge: Differential Fine-Tuning of IL-2- and IL-15- Dependent Functions by targeting their Common IL-2/15Rβ/γc Receptor

Author

Sarah Khaddage, Markus Hildinger, Dihia Meghnem, Mike Maillasson, Isabelle Barbieux, Marie Frutoso, Isabelle Leray, Yannick Jacques, Sébastien Morisseau, Kilian Trillet, Erwan Mortier, Agnès Quéméner, Bernardo, Elizabeth, Instituts Hospitalo-Universitaires B - Centre Européen des Sciences de la Transplantation et de l'Immunothérapie (TSI-IHU) - - CESTI (TSI-IHU)2010 - ANR-10-IBHU-0005 - IBHU - VALID, Centre de Recherche en Cancérologie et Immunologie Nantes-Angers (CRCINA), Université d'Angers (UA)-Université de Nantes (UN)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Centre hospitalier universitaire de Nantes (CHU Nantes), Centre hospitalier universitaire de Nantes (CHU Nantes), Evitria [Schlieren, Suisse], Infors AG [Bottmingen, Suisse], This work was supported by CNRS, Inserm, Inserm Transfert (R13079NS) and was realized in the context of the Institut-Hospitalo Universitaire–Centre Europeen des Sciences de la Transplantation et Immunotherapies (ANR-10-IBHU-005) project, which received French government financial support managed by the National Research Agency and supported by Nantes Metropole and Pays de la Loire Region. D.M. was supported by a specific thesis allocation from the Institut-Hospitalo Universitaire–Centre Europeen des Sciences de la Transplantation et Immunotherapies., ANR-10-IBHU-0005,CESTI (TSI-IHU),Centre Européen des Sciences de la Transplantation et de l'Immunothérapie (TSI-IHU)(2010), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), and Université de Nantes (UN)-Université de Nantes (UN)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Centre National de la Recherche Scientifique (CNRS)-Université d'Angers (UA)

Subjects
0301 basic medicine, Interleukin 2, Regulatory T cell, Immunology, [SDV.CAN]Life Sciences [q-bio]/Cancer, Biology, CD8-Positive T-Lymphocytes, Lymphocyte Activation, T-Lymphocytes, Regulatory, Cell Line, 03 medical and health sciences, Mice, Immune system, Interleukin-15 Receptor alpha Subunit, [SDV.CAN] Life Sciences [q-bio]/Cancer, medicine, Immunology and Allergy, Animals, Humans, Interleukin-7 receptor, Receptor, Common gamma chain, Cell Proliferation, Interleukin-15, Interleukin-2 Receptor alpha Subunit, Cell biology, Interleukin-2 Receptor beta Subunit, Killer Cells, Natural, 030104 developmental biology, medicine.anatomical_structure, Interleukin 15, Interleukin-2, CD8, medicine.drug, Interleukin Receptor Common gamma Subunit, Protein Binding, Signal Transduction
Abstract

Interleukin 2 and IL-15 are two closely related cytokines, displaying important functions in the immune system. They share the heterodimeric CD122/CD132 receptor to deliver their signals within target cells. Their specificity of action is conferred by their α receptor chains, IL-2Rα and IL-15Rα. By combining an increased affinity for CD122 and an impaired recruitment of CD132, we have generated an original molecule named IL-2Rβ/γ (CD122/CD132) inhibitor (BiG), targeting the CD122/CD132 receptor. BiG efficiently inhibited IL-15– and IL-2–dependent functions of primary cells, including CD8 T and NK cells, in vitro and in vivo. We also report a differential dynamic of action of these cytokines by highlighting a major role played by the IL-2Rα receptor. Interestingly, due to the presence of IL-2Rα, BiG had no impact on IL-2–dependent regulatory T cell proliferation. Thus, by acting as a fine switch in the immune system, BiG emphasizes the differential roles of these two cytokines.

Published
2017

16. Highly potent anti-CD20-RLI immunocytokine targeting established human B lymphoma in SCID mouse

Author

Séverine Loisel, Yannick Jacques, Béatrice Clémenceau, Véronique Solé, Mike Maillasson, Anne Bessard, Laure Garrigue-Antar, Géraldine Teppaz, Marie Vincent, David Bechard, Agnès Quéméner, Laurie Lajoie, Gilles Thibault, Centre de Recherche en Cancérologie Nantes-Angers (CRCNA), Centre Hospitalier Universitaire d'Angers (CHU Angers), PRES Université Nantes Angers Le Mans (UNAM)-PRES Université Nantes Angers Le Mans (UNAM)-Hôtel-Dieu de Nantes-Institut National de la Santé et de la Recherche Médicale (INSERM)-Hôpital Laennec-Centre National de la Recherche Scientifique (CNRS)-Faculté de Médecine d'Angers-Centre hospitalier universitaire de Nantes (CHU Nantes), Génétique, immunothérapie, chimie et cancer (GICC), UMR 7292 CNRS [2012-2017] (GICC UMR 7292 CNRS), Université de Tours (UT)-Centre National de la Recherche Scientifique (CNRS), Immunologie et Pathologie (EA2216), Université de Brest (UBO)-IFR148, Cytune Pharma SAS, Université de Tours-Centre National de la Recherche Scientifique (CNRS), and Garrigue-Antar, Laure

Subjects
Male, Interleukin 2, Cellular immunity, Immunoconjugates, Lymphoma, B-Cell, immunocytokine, medicine.drug_class, Recombinant Fusion Proteins, [SDV]Life Sciences [q-bio], Immunology, Antineoplastic Agents, lymphoma, interleukin-15, [SDV.CAN]Life Sciences [q-bio]/Cancer, CHO Cells, Mice, SCID, Biology, Monoclonal antibody, Antibodies, Monoclonal, Murine-Derived, Mice, Cricetulus, [SDV.CAN] Life Sciences [q-bio]/Cancer, Antigen, Cricetinae, Report, medicine, Animals, Humans, Immunology and Allergy, CD20, B cell malignancies, Antibody-dependent cell-mediated cytotoxicity, Severe combined immunodeficiency, medicine.disease, Xenograft Model Antitumor Assays, 3. Good health, [SDV] Life Sciences [q-bio], Interleukin 15, biology.protein, Interleukin-2, cancer therapy, Female, Antibody, Rituximab, medicine.drug
Abstract

International audience; Rituximab (RTX), a chimeric IgG1 monoclonal antibody directed against the CD20 antigen, has revolutionized the treatment of B-cell alignancies. Nevertheless, the relapsed/refractory rates are still high. One strategy to increase the clinical effectiveness of RTX is based on antibody-cytokine fusion protein (immunocytokine; ICK) vectorizing together at the tumor site the antibody effector activities and the cytokine co-signal required for the generation of cytotoxic cellular immunity. Such ICKs linking various antibody formats to interleukin (IL)-2 are currently being investigated in clinical trials and have shown promising results in cancer therapies. IL-15, a structurally-related cytokine, is now considered as having a better potential than IL-2 in antitumor immunotherapeutic strategies. We have previously engineered the fusion protein RLI, linking a soluble form of human IL-15Rα-sushi+ domain to human IL-15. Compared with IL-15, RLI displayed better biological activities in vitro and higher antitumor effects in vivo in murine and human cancer models. In this study, we investigated the advantages of fusing RLI to RTX. Anti-CD20-RLI kept its binding capacity to CD20, CD16 and IL-15 receptor and therefore fully retained both antibody effector functions (ADCC and CDC), and the cytokine potential of RLI. In a severe combined immunodeficiency (SCID) mouse model of disseminated residual lymphoma, anti-CD20-RLI was found to induce long-term survival of 90% of mice up to at least 120 days whereas RLI and RTX, alone or in combination, just delayed the disease onset (100% of death at 28, 40 and 51 days respectively). These findings suggest that such ICK could improve the clinical efficacy of RTX, particularly in patients with refractory B-cell lymphoma.

Published
2014

17. Tumor targeting of the IL-15 superagonist RLI by an anti-GD2 antibody strongly enhances its antitumor potency

Author

Mike Maillasson, Géraldine Teppaz, Yannick Jacques, Laure Garrigue-Antar, Véronique Solé, Denis Cochonneau, Stéphane Birklé, Agnès Quéméner, Marie Vincent, and Anne Bessard

Subjects
0303 health sciences, Cancer Research, medicine.medical_treatment, Context (language use), Immunotherapy, Biology, Pharmacology, 3. Good health, 03 medical and health sciences, 0302 clinical medicine, Cytokine, Therapeutic index, Oncology, Antigen, Interleukin 15, In vivo, 030220 oncology & carcinogenesis, medicine, Cytotoxicity, 030304 developmental biology
Abstract

Immunocytokines (ICKs) targeting cytokines to the tumor environment using antibodies directed against a tumor-associated antigen often have a higher therapeutic index than the corresponding unconjugated cytokines. Various ICKs displaying significant antitumoral effects in several murine tumor models have already been developed, and some of them, in particular interleukin (IL)-2-based ICKs, are in Phase II clinical trials. Although sharing common biological activities with IL-2 in vitro, IL-15 is now considered as having a better potential in antitumor immunotherapeutical strategies and has been shown to be less toxic than IL-2 in preclinical studies. We previously developed the fusion protein RLI, linking a soluble form of human IL-15Rα-sushi+ domain to human IL-15. RLI showed better biological activities than IL-15 in vitro as well as higher antitumoral effects in vivo in murine and human cancer models. Here, we investigated, in the context of an ICK, the effect of associating RLI with an antibody targeting the GD2 ganglioside, a validated tumoral target expressed on many neurectodermal tumors. Anti-GD2-RLI fully retained the cytokine potential of RLI and the antibody effector functions (antibody-dependent cellular cytotoxicity and complement-dependent cytotoxicity). It displayed strong antitumor activities in two syngeneic cancer models in immunocompetent mice (subcutaneous EL4 and metastatic NXS2). Its therapeutic potency was higher than those of RLI and anti-GD2 alone or in combination. We suggest that this is related to its bifunctional (cytokine and antibody) nature.

Published
2013

18. IL-34 and M-CSF form a novel heteromeric cytokine and regulate the M-CSF receptor activation and localization

Author

Régis Brion, Stéphane Téletchéa, Bénédicte Brulin, Mike Maillasson, Dominique Heymann, Aude I. Segaliny, Céline Charrier, Physiopathologie des Adaptations Nutritionnelles (PhAN), Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), Université de Nantes (UN)-Université de Nantes (UN)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Centre hospitalier universitaire de Nantes (CHU Nantes), Plateforme IMPACT Nantes (CRCINA-IMPACT), Centre de Recherche en Cancérologie et Immunologie Nantes-Angers (CRCINA), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), Université de Nantes (UN)-Université de Nantes (UN)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Centre National de la Recherche Scientifique (CNRS)-Université d'Angers (UA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), Université de Nantes (UN)-Université de Nantes (UN)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Centre National de la Recherche Scientifique (CNRS)-Université d'Angers (UA), Centre de Recherche en Cancérologie Nantes-Angers (CRCNA), Centre Hospitalier Universitaire d'Angers (CHU Angers), PRES Université Nantes Angers Le Mans (UNAM)-PRES Université Nantes Angers Le Mans (UNAM)-Hôtel-Dieu de Nantes-Institut National de la Santé et de la Recherche Médicale (INSERM)-Hôpital Laennec-Centre National de la Recherche Scientifique (CNRS)-Faculté de Médecine d'Angers-Centre hospitalier universitaire de Nantes (CHU Nantes), Unité de fonctionnalité et ingénierie de protéines (UFIP), Université de Nantes - UFR des Sciences et des Techniques (UN UFR ST), Université de Nantes (UN)-Université de Nantes (UN)-Centre National de la Recherche Scientifique (CNRS), and This study was supported by the Region des Pays de la Loire (CIMATH II research project) and by the Ligue Nationale Contre le Cancer (Equipe LIGUE 2012).

Subjects
Macrophage colony-stimulating factor, Macrophage-Colony Stimulating Factor, Cell Survival, medicine.medical_treatment, Immunology, Molecular modeling, Receptor, Macrophage Colony-Stimulating Factor, [SDV.CAN]Life Sciences [q-bio]/Cancer, Interleukin-17 receptor, Proximity ligation assay, cFMS trafficking, Biology, Biochemistry, Monocytes, 03 medical and health sciences, 0302 clinical medicine, Cell Movement, Cell Line, Tumor, medicine, Immunology and Allergy, Humans, Phosphorylation, Receptor, Molecular Biology, 030304 developmental biology, Cell Proliferation, 0303 health sciences, Interleukins, Macrophage Colony-Stimulating Factor, Cell Differentiation, Hematology, Molecular biology, Interleukin-34, Cell biology, Molecular Docking Simulation, Cytokine, HEK293 Cells, Heteromeric cytokine, 030220 oncology & carcinogenesis, Interleukin 34, Cytokines, Signal transduction, Protein Multimerization, Signal Transduction
Abstract

International audience; Interleukin-34 (IL-34) is a newly-discovered homodimeric cytokine that regulates, like Macrophage Colony-Stimulating Factor (M-CSF), the differentiation of the myeloid lineage through M-CSF receptor (M-CSFR) signaling pathways. To date, both cytokines have been considered as competitive cytokines with regard to the M-CSFR. The aim of the present work was to study the functional relationships of these cytokines on cells expressing the M-CSFR. We demonstrate that simultaneous addition of M-CSF and IL-34 led to a specific activation pattern on the M-CSFR, with higher phosphorylation of the tyrosine residues at low concentrations. Similarly, both cytokines showed an additive effect on cellular proliferation or viability. In addition, BIAcore experiments demonstrated that M-CSF binds to IL-34, and molecular docking studies predicted the formation of a heteromeric M-CSF/IL-34 cytokine. A proximity ligation assay confirmed this interaction between the cytokines. Finally, co-expression of the M-CSFR and its ligands differentially regulated M-CSFR trafficking into the cell. This study establishes a new foundation for the understanding of the functional relationship between IL-34 and M-CSF, and gives a new vision for the development of therapeutic approaches targeting the IL-34/M-CSF/M-CSFR axis.

Published
2015

19. Glycosaminoglycans inhibit the adherence and the spreading of osteoclasts and their precursors: Role in osteoclastogenesis and bone resorption

Author

Gaëtan Jego, Carmen Ruiz-Velasco, Laurence Duplomb, Françoise Rédini, Nijole Gasiunas, Annamaria Naggi, Dominique Heymann, Marc Baud'huin, Céline Charrier, John T. Gallagher, Mike Maillasson, and Marc Padrines

Subjects
musculoskeletal diseases, medicine.medical_specialty, Histology, Blotting, Western, Lipopolysaccharide Receptors, Gene Expression, Osteoclasts, Polymerase Chain Reaction, Bone resorption, Cell Line, Pathology and Forensic Medicine, Bone remodeling, Glycosaminoglycan, Mice, 03 medical and health sciences, 0302 clinical medicine, Sulfation, Osteoclast, Internal medicine, Cell Adhesion, medicine, Animals, Humans, RNA, Messenger, Bone Resorption, Glycosaminoglycans, 030304 developmental biology, 0303 health sciences, biology, CD11 Antigens, Heparin, Chemistry, Macrophages, RANK Ligand, Cell Differentiation, Cell Biology, General Medicine, In vitro, Cell biology, medicine.anatomical_structure, Endocrinology, RANKL, 030220 oncology & carcinogenesis, biology.protein, Bone Remodeling, medicine.drug
Abstract

The bone microenvironment (e.g. glycosaminoglycans (GAGs), growth factors) plays a major role in bone resorption, especially in the formation of osteoclasts which differentiate from the hematopoietic lineage in the presence of RANKL. Previous studies revealed that GAGs may influence osteoclastogenesis, but data are very controversial, some studies showing an inhibitory effect of GAGs on osteoclastic differentiation whereas others demonstrated a stimulatory effect. To clarify their activities, we investigated the effect of 5 families of GAGs in three different models of human/mouse osteoclastogenesis. The present data revealed that heparin inhibited osteoclastogenesis in these three models, which was confirmed by a decrease in mRNA expression of osteoclastic markers and by an inhibition of the bone resorption capacity. We also demonstrated in RAW 264.7 cells that other families of GAGs different from heparin inhibited RANKL-induced osteoclastogenesis, and that this inhibition was dependent on the length and the level of sulfation of GAGs. In the present work, heparin did not bind to RANKL and did not modulate RANKL signaling. Heparin acted at 2 distinct steps of osteoclastogenesis from human CD14(+) cells: first, heparin strongly decreased the adherence of osteoclast precursors, and secondly inhibited osteoclasts to spread and to be active. Furthermore, the second action of heparin was reversible as the removal of heparin at the end of the culture time allowed the condensed cells to spread out and showed the formation of morphological active osteoclasts. The present work clearly evidences that GAGs inhibit osteoclastogenesis in vitro and strengthens the therapeutic interest of defined GAGs in osteolytic diseases.

Published
2011

20. Phosphorothioate-Modified TLR9 Ligands Protect Cancer Cells against TRAIL-Induced Apoptosis

Author

Régis Bataille, Mike Maillasson, David Chiron, Gaëtan Jego, and Catherine Pellat-Deceunynck

Subjects
CpG Oligodeoxynucleotide, Lexatumumab, Immunology, Phosphorothioate Oligonucleotides, Apoptosis, Breast Neoplasms, Biology, Ligands, TNF-Related Apoptosis-Inducing Ligand, Immune system, Adjuvants, Immunologic, Cell Line, Tumor, medicine, Humans, Immunology and Allergy, TLR9, hemic and immune systems, respiratory system, Molecular biology, Oligodeoxyribonucleotides, Toll-Like Receptor 9, Colonic Neoplasms, Cancer cell, Cancer research, CpG Islands, Female, Signal transduction, Multiple Myeloma, Mapatumumab, medicine.drug
Abstract

Hypomethylated CpG oligodeoxynucleotides (CpG ODNs) target TLR9 expressed by immune cells and are currently being evaluated as adjuvants in clinical trials. However, TLR signaling can promote some tumor growth and immune evasion, such as in multiple myeloma (MM). Therefore, deciphering the effects of CpG ODNs on cancer cells will help in preventing these adverse effects and in designing future clinical trials. TLR activation induces multiple signaling pathways, notably NF-κB that has been involved in the resistance to TRAIL. Thus, we wondered if CpG ODNs could modulate TRAIL-induced apoptosis in different models of tumors. Here, we show that TLR9+ (NCI-H929, NAN6, KMM1) and TLR9− MM cells (MM1S) were protected by CpG ODNs against recombinant TRAIL-induced apoptosis. By using two fully human, agonist mAbs directed against TRAIL receptors DR4 and DR5 (mapatumumab and lexatumumab, respectively), we show that the protection was restricted to DR5-induced apoptosis. Similar results were observed for two colon cancer (C45 and Colo205) and two breast cancer cell lines (HCC1569 and Cal51). The protection of CpG ODNs was mediated by its nuclease-resistant phosphorothioate backbone independent of TLR9. We next demonstrated by surface plasmon resonance that phosphorothioate-modified CpG ODNs directly bound to either TRAIL or lexatumumab and then decreased their binding to DR5. Finally, NK cell lysis of a DR5-sensitive MM cell line (NCI-H929) through TRAIL was partially inhibited by phosphorothioate-modified CpG ODNs. In conclusion, our results suggest that the phosphorothioate modification of CpG ODNs could dampen the clinical efficacy of CpG ODN-based adjuvants by altering TRAIL/TRAIL receptor interaction.

Published
2009

21. Affinity transfer to the archaeal extremophilic Sac7d protein by insertion of a CDR

Author

Frédéric Pecorari, Ghislaine Béhar, Sabino Pacheco, Mike Maillasson, and Barbara Mouratou

Subjects
Scaffold protein, Affitin, Camelus, Archaeal Proteins, Molecular Sequence Data, Bioengineering, Biology, Protein Engineering, Biochemistry, chemistry.chemical_compound, Animals, Thermal stability, Amino Acid Sequence, Molecular Biology, Flat surface, Protein Stability, Structural integrity, DNA, Complementarity Determining Regions, DNA-Binding Proteins, Directed mutagenesis, chemistry, Biophysics, Muramidase, Chickens, Biotechnology, Protein Binding
Abstract

Artificially transforming a scaffold protein into binders often consists of introducing diversity into its natural binding region by directed mutagenesis. We have previously developed the archaeal extremophilic Sac7d protein as a scaffold to derive affinity reagents (Affitins) by randomization of only a flat surface, or a flat surface and two short loops with natural lengths. Short loops are believed to contribute to stability of extremophilic proteins, and loop extension has been reported detrimental for the thermal and chemical stabilities of mesophilic proteins. In this work, we wanted to evaluate the possibility of designing target-binding proteins based on Sac7d by using a complementary determining region (CDR). To this aim, we inserted into three different loops a 10 residues CDR from the cAb-Lys3 anti-lysozyme camel antibody. The chimeras obtained were as stable as wild-type (WT) Sac7d at extreme pH and their structural integrity was supported. Chimeras were thermally stable, but with T(m)s from 60.9 to 66.3°C (cf. 91°C for Sac7d) which shows that loop extension is detrimental for thermal stability of Sac7d. The loop 3 enabled anti-lysozyme activity. These results pave the way for the use of CDR(s) from antibodies and/or extended randomized loop(s) to increase the potential of binding of Affitins.

Published
2014

22. Syndecan-1 regulates the biological activities of interleukin-34

Author

Dominique Heymann, Aude I. Segaliny, Michel Chérel, Régis Brion, Benoit Le Goff, Yannick Jacques, Mike Maillasson, and Erwan Mortier

Subjects
Cell type, medicine.medical_treatment, Receptor, Macrophage Colony-Stimulating Factor, Protein tyrosine phosphatase, Biology, Models, Biological, Syndecan 1, Cell Line, Co-receptor chondroitin sulphate, chemistry.chemical_compound, M-CSFR, Cell Movement, medicine, Chondroitin, Humans, Myeloid Cells, Phosphorylation, RNA, Small Interfering, Molecular Biology, Receptor-Like Protein Tyrosine Phosphatases, Class 5, Interleukins, Macrophage Colony-Stimulating Factor, HEK 293 cells, Chondroitin Sulfates, Cytokine bioavailability, Cell migration, Cell Biology, Interleukin-34, Molecular biology, carbohydrates (lipids), Cytokine, chemistry, Interleukin 34, Syndecan-1, M2a macrophage, Protein Binding
Abstract

IL-34 is a challenging cytokine sharing functional similarities with M-CSF through M-CSFR activation. It also plays a singular role that has recently been explained in the brain, through a binding to the receptor protein tyrosine phosphatase RPTPβ/ζ. The aim of this paper was to look for alternative binding of IL-34 on other cell types. Myeloid cells (HL-60, U-937, THP-1) were used as cells intrinsically expressing M-CSFR, and M-CSFR was expressed in TF-1 and HEK293 cells. IL-34 binding was studied by Scatchard and binding inhibition assays, using 125I-radiolabelled cytokines, and surface plasmon resonance. M-CSFR activation was analysed by Western blot after glycosaminoglycans abrasion, syndecan-1 overexpression or repression and addition of a blocking anti-syndecan antibody. M-CSF and IL-34 induced different patterns of M-CSFR phosphorylations, suggesting the existence of alternative binding for IL-34. Binding experiments and chondroitinase treatment confirmed low affinity binding to chondroitin sulphate chains on cells lacking both M-CSFR and RPTPβ/ζ. Amongst the proteoglycans with chondroitin sulphate chains, syndecan-1 was able to modulate the IL-34-induced M-CSFR signalling pathways. Interestingly, IL-34 induced the migration of syndecan-1 expressing cells. Indeed, IL-34 significantly increased the migration of THP-1 and M2a macrophages that was inhibited by addition of a blocking anti-syndecan-1 antibody. This paper provides evidence of alternative binding of IL-34 to chondroitin sulphates and syndecan-1 at the cell surface that modulates M-CSFR activation. In addition, IL-34-induced myeloid cell migration is a syndecan-1 dependent mechanism.

Published
2014

24. Chondroitin sulfate chains are co-receptors for interleukin- 34

Author

Erwan Mortier, Mike Maillasson, Yannick Jacques, Dominique Heymann, Benoit Le Goff, Régis Brion, and Aude I. Segaliny

Subjects
chemistry.chemical_compound, chemistry, Biochemistry, Interleukin 34, General Medicine, Chondroitin sulfate, Receptor
Published
2014

25. Influence of pegylation and hapten location at the surface of radiolabelled liposomes on tumour immunotargeting using bispecific antibody

Author

P. Le Saëc, Marie Mougin-Degraef, M. Frindel, Jean-François Gestin, Catherine Maurel, Holisoa Rajerison, A. Rauscher, Jacques Barbet, Mike Maillasson, and Alain Faivre-Chauvet

Subjects
Cancer Research, medicine.medical_treatment, Adenocarcinoma, Polyethylene Glycols, Mice, In vivo, Cell Line, Tumor, PEG ratio, Antibodies, Bispecific, medicine, Animals, Humans, Radiology, Nuclear Medicine and imaging, Pretargeting, Liposome, Chemistry, Cell Membrane, Radioimmunotherapy, Molecular biology, Monoclonal, Liposomes, PEGylation, Cancer research, Molecular Medicine, Female, Colorectal Neoplasms, Hapten, Haptens
Abstract

Introduction This paper proposes liposomes as a potential new tool for radioimmunotherapy in solid tumours with a two step targeting system. Tumour pretargeting is obtained by using a monoclonal bispecific antibody (BsmAb, anti CEA x anti-DTPA-In) and pegylated liposomes containing lipid-hapten (DSPE-DTPA-In or DSPE-PEG-DTPA-In). To optimise at the same time in vivo behaviour and specific targeting, the study focuses on the liposome formulation in order to determine more precisely the role of pegylation on both the blood half-life and the specific recognition with the BsmAb. Methods Different liposome formulations containing two PEG length (1000 and 2000) in varying amount (1.5–6 mol%) were prepared with DTPA directly coupled to DSPE or at the end of the PEG chain (DSPE-DTPA or DSPE-PEG-DTPA). Liposomes were immobilized on an L1 chip to measure by SPR (Surface Plasmon Resonance) the effect of pegylation on the BsmAb recognition of the DTPA-In hapten. Pharmacokinetic studies were performed in mice. Tumour targeting was studied in nude mice xenografted with human colorectal adenocarcinoma cells that express CEA, and doubly radiolabelled liposomes (with 111In and 125I) injected 24 h after the BsmAb. Results The best in vitro apparent dissociation constant was obtained with liposomes bearing DTPA at the end of the PEG chain (KD = 6.3 nM), which showed significant specific tumour uptake after BsmAb injection (8.6 ± 2.4% ID/g at 24 h versus 4.5 ± 0.5%ID/g for passive targeting, α = 0.01). All tumour/organ ratios were superior to 1 at 24 h for this formulation, except for the spleen. Conclusion The feasibility of specific tumour targeting in mice with a BsmAb and radiolabelled liposomes was demonstrated and the interest of SPR to predict their targeting performance in vivo was highlighted. This original and new approach provides promising prospects for the radioimmunotherapy of solid tumours.

Published
2013

26. Osteoprotegerin: multiple partners for multiple functions

Author

Dominique Heymann, Marc Baud'huin, Stéphane Téletchéa, Marie-Françoise Heymann, Laurence Duplomb, Carmen Ruiz-Velasco, Mike Maillasson, Francois Lamoureux, and Françoise Rédini

Subjects
musculoskeletal diseases, Cell Survival, Endocrinology, Diabetes and Metabolism, Immunology, Osteoclasts, General Biochemistry, Genetics and Molecular Biology, TNF-Related Apoptosis-Inducing Ligand, Osteoprotegerin, Immunology and Allergy, Animals, Humans, Cell adhesion, Receptor, Cell Proliferation, biology, Activator (genetics), Cell growth, Chemistry, RANK Ligand, Cell Differentiation, In vitro, Cell biology, Biochemistry, RANKL, biology.protein, Decoy
Abstract

Osteoprotegerin (OPG) is an essential secreted protein in bone turnover due to its role as a decoy receptor for the Receptor Activator of Nuclear Factor-kB ligand (RANKL) in the osteoclasts, thus inhibiting their differentiation. However, there are additional ligands of OPG that confer various biological functions. OPG can promote cell survival, cell proliferation and facilitates migration by binding TNF-related apoptosis inducing ligand (TRAIL), glycosaminoglycans or proteoglycans. A large number of in vitro, pre-clinical and clinical studies provide evidences of OPG involvement in vascular, bone, immune and tumor biology. This review describes an overview of the different OPG ligands regulating its biological functions.

Published
2013

27. Tumor targeting of the IL-15 superagonist RLI by an anti-GD2 antibody strongly enhances its antitumor potency

Author

Marie, Vincent, Anne, Bessard, Denis, Cochonneau, Géraldine, Teppaz, Véronique, Solé, Mike, Maillasson, Stéphane, Birklé, Laure, Garrigue-Antar, Agnès, Quéméner, Yannick, Jacques, Centre de Recherche en Cancérologie Nantes-Angers (CRCNA), Centre Hospitalier Universitaire d'Angers (CHU Angers), PRES Université Nantes Angers Le Mans (UNAM)-PRES Université Nantes Angers Le Mans (UNAM)-Hôtel-Dieu de Nantes-Institut National de la Santé et de la Recherche Médicale (INSERM)-Hôpital Laennec-Centre National de la Recherche Scientifique (CNRS)-Faculté de Médecine d'Angers-Centre hospitalier universitaire de Nantes (CHU Nantes), and QUEMENER, Agnès

Subjects
Male, [SDV.IMM] Life Sciences [q-bio]/Immunology, Antibodies, Neoplasm, Mice, Inbred A, Recombinant Fusion Proteins, [SDV]Life Sciences [q-bio], [SDV.CAN]Life Sciences [q-bio]/Cancer, Lymphoma, T-Cell, Mice, Neuroblastoma, Interleukin-15 Receptor alpha Subunit, [SDV.CAN] Life Sciences [q-bio]/Cancer, Antigens, Neoplasm, Cell Line, Tumor, Gangliosides, Animals, Humans, Neoplasm Metastasis, Cell Proliferation, Interleukin-15, Liver Neoplasms, Antibody-Dependent Cell Cytotoxicity, Antibodies, Monoclonal, Mice, Inbred C57BL, [SDV] Life Sciences [q-bio], [SDV.IMM]Life Sciences [q-bio]/Immunology, Female, Immunotherapy, Protein Binding
Abstract

International audience; Immunocytokines (ICKs) targeting cytokines to the tumor environment using antibodies directed against a tumor-associated antigen often have a higher therapeutic index than the corresponding unconjugated cytokines. Various ICKs displaying significant antitumoral effects in several murine tumor models have already been developed, and some of them, in particular interleukin (IL)-2-based ICKs, are in Phase II clinical trials. Although sharing common biological activities with IL-2 in vitro, IL-15 is now considered as having a better potential in antitumor immunotherapeutical strategies and has been shown to be less toxic than IL-2 in preclinical studies. We previously developed the fusion protein RLI, linking a soluble form of human IL-15Rα-sushi+ domain to human IL-15. RLI showed better biological activities than IL-15 in vitro as well as higher antitumoral effects in vivo in murine and human cancer models. Here, we investigated, in the context of an ICK, the effect of associating RLI with an antibody targeting the GD2 ganglioside, a validated tumoral target expressed on many neurectodermal tumors. Anti-GD2-RLI fully retained the cytokine potential of RLI and the antibody effector functions (antibody-dependent cellular cytotoxicity and complement-dependent cytotoxicity). It displayed strong antitumor activities in two syngeneic cancer models in immunocompetent mice (subcutaneous EL4 and metastatic NXS2). Its therapeutic potency was higher than those of RLI and anti-GD2 alone or in combination. We suggest that this is related to its bifunctional (cytokine and antibody) nature.

Published
2013

28. Tolerance of the archaeal Sac7d scaffold protein to alternative library designs: characterization of anti-immunoglobulin G Affitins

Author

Ghislaine Béhar, Barbara Mouratou, Marco Bellinzoni, Xuemei He, Pedro M. Alzari, Mike Maillasson, Lauranne Paillard-Laurance, Frédéric Pecorari, Unité de Biotechnologie, Biocatalyse et Biorégulation (U3B), Université de Nantes (UN)-Centre National de la Recherche Scientifique (CNRS), Microbiologie structurale - Structural Microbiology (Microb. Struc. (UMR_3528 / U-Pasteur_5)), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS)-Université Paris Cité (UPCité), Plateforme IMPACT Nantes (CRCINA-IMPACT), Centre de Recherche en Cancérologie et Immunologie Nantes-Angers (CRCINA), Université d'Angers (UA)-Université de Nantes (UN)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Université d'Angers (UA)-Université de Nantes (UN)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Centre hospitalier universitaire de Nantes (CHU Nantes), Bio-Rad [Hercules, CA], This work was supported by the ‘La Région des Pays de la Loire’ and by Bio-Rad., Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS)-Université de Paris (UP), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), Université de Nantes (UN)-Université de Nantes (UN)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Centre National de la Recherche Scientifique (CNRS)-Université d'Angers (UA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), Université de Nantes (UN)-Université de Nantes (UN)-Centre hospitalier universitaire de Nantes (CHU Nantes)-Centre National de la Recherche Scientifique (CNRS)-Université d'Angers (UA), and de TARRAGON, Marie

Subjects
Scaffold protein, Scaffold, MESH: Antibodies, Anti-Idiotypic, Ligands, Protein Engineering, Biochemistry, Epitope, Epitopes, MESH: Protein Structure, Tertiary, Protein structure, MESH: Ligands, [INFO.INFO-BI] Computer Science [cs]/Bioinformatics [q-bio.QM], Genetics, MESH: Immunoglobulin G, 0303 health sciences, [PHYS.PHYS.PHYS-BIO-PH] Physics [physics]/Physics [physics]/Biological Physics [physics.bio-ph], [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Protein Stability, 030302 biochemistry & molecular biology, MESH: Archaeal Proteins, Antibodies, Anti-Idiotypic, DNA-Binding Proteins, MESH: Protein Engineering, MESH: Archaea, Biotechnology, MESH: Epitopes, [SDV.BBM.BS] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], Archaeal Proteins, [PHYS.PHYS.PHYS-BIO-PH]Physics [physics]/Physics [physics]/Biological Physics [physics.bio-ph], Mutagenesis (molecular biology technique), Bioengineering, Computational biology, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Biology, 03 medical and health sciences, Molecular recognition, Peptide Library, MESH: Protein Stability, [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, [CHIM.CRIS]Chemical Sciences/Cristallography, Humans, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, [CHIM.CRIS] Chemical Sciences/Cristallography, Peptide library, [SDV.BC] Life Sciences [q-bio]/Cellular Biology, Molecular Biology, 030304 developmental biology, MESH: Humans, Protein engineering, Archaea, Protein Structure, Tertiary, Immunoglobulin G, MESH: Peptide Library, [INFO.INFO-BI]Computer Science [cs]/Bioinformatics [q-bio.QM], Ribosomes, MESH: Ribosomes, MESH: DNA-Binding Proteins
Abstract

International audience; Engineered protein scaffolds have received considerable attention as alternatives to antibodies in both basic and applied research, as they can offer superior biophysical properties often associated with a simpler molecular organization. Sac7d has been demonstrated as an effective scaffold for molecular recognition. Here, we used the initial L1 'flat surface' library constructed by randomization of 14 residues, to identify ligands specific for human immunoglobulin G. To challenge the plasticity of the Sac7d protein scaffold, we designed the alternative L2 'flat surface & loops' library whereof only 10 residues are randomized. Representative binders (Affitins) of the two libraries exhibited affinities in the low nanomolar range and were able to recognize different epitopes within human immunoglobulin G. These Affitins were stable up to pH 12 while largely conserving other favorable properties of Sac7d protein, such as high expression yields in Escherichia coli, solubility, thermal stability up to 80.7°C, and acidic stability (pH 0). In agreement with our library designs, mutagenesis study revealed two distinct binding areas, one including loops. Together, our results indicate that the Sac7d scaffold tolerates alternative library designs, which further expands the diversity of Affitins and may provide a general way to create tailored affinity tools for demanding applications.

Published
2013

29. Functional characterization of a chimeric soluble Fas ligand polymer with in vivo anti-tumor activity

Author

Mike Maillasson, Delphine Lapaillerie, Patrick Legembre, Aurore Morello, Jean-Luc Taupin, Laure Garrigue-Antar, Sophie Daburon, Marc Bonneu, Myriam Capone, Pierre Costet, Julie Déchanet-Merville, Jean-François Moreau, Christel Devaud, Nathalie Hargous, Composantes innées de la réponse immunitaire et différenciation (CIRID), Université Bordeaux Segalen - Bordeaux 2-Centre National de la Recherche Scientifique (CNRS), Animalerie spécialisée, Université Bordeaux Segalen - Bordeaux 2, Croissance cellulaire, réparation et régénération tissulaires (CRRET), Université Paris-Est Créteil Val-de-Marne - Paris 12 (UPEC UP12)-Centre National de la Recherche Scientifique (CNRS), Centre de Recherche en Cancérologie Nantes-Angers (CRCNA), Centre Hospitalier Universitaire d'Angers (CHU Angers), PRES Université Nantes Angers Le Mans (UNAM)-PRES Université Nantes Angers Le Mans (UNAM)-Hôtel-Dieu de Nantes-Institut National de la Santé et de la Recherche Médicale (INSERM)-Hôpital Laennec-Centre National de la Recherche Scientifique (CNRS)-Faculté de Médecine d'Angers-Centre hospitalier universitaire de Nantes (CHU Nantes), Centre génomique fonctionnelle, Récepteur de mort et échappement tumoral, Institut de recherche en santé, environnement et travail (Irset), Université d'Angers (UA)-Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-École des Hautes Études en Santé Publique [EHESP] (EHESP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique )-Université d'Angers (UA)-Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-École des Hautes Études en Santé Publique [EHESP] (EHESP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique ), and Université d'Angers (UA)-Université de Rennes (UR)-École des Hautes Études en Santé Publique [EHESP] (EHESP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique )-Université d'Angers (UA)-Université de Rennes (UR)-École des Hautes Études en Santé Publique [EHESP] (EHESP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique )

Subjects
Anatomy and Physiology, Mouse, Oncogene Proteins, Fusion, lcsh:Medicine, Apoptosis, Leukemia inhibitory factor receptor, Ligands, Biochemistry, Fas ligand, Jurkat Cells, Mice, 0302 clinical medicine, MESH: Genetic Vectors, Immune Physiology, Neoplasms, Molecular Cell Biology, Basic Cancer Research, MESH: Jurkat Cells, MESH: Ligands, MESH: Animals, MESH: Neoplasms, lcsh:Science, Receptor, 0303 health sciences, Immune System Proteins, Multidisciplinary, Cell Death, hemic and immune systems, Animal Models, Fas receptor, Recombinant Proteins, Cell biology, MESH: Antigens, CD95, Oncology, 030220 oncology & carcinogenesis, Cytokines, Medicine, biological phenomena, cell phenomena, and immunity, Research Article, Fas Ligand Protein, Receptors, OSM-LIF, Recombinant Fusion Proteins, Genetic Vectors, chemical and pharmacologic phenomena, [SDV.CAN]Life Sciences [q-bio]/Cancer, Biology, Transfection, 03 medical and health sciences, Chimera (genetics), Model Organisms, Cell surface receptor, MESH: Recombinant Fusion Proteins, Animals, Humans, fas Receptor, MESH: Mice, 030304 developmental biology, Receptor Aggregation, MESH: Receptors, OSM-LIF, MESH: Humans, MESH: Transfection, MESH: Apoptosis, lcsh:R, Proteins, MESH: Fas Ligand Protein, lcsh:Q, MESH: Oncogene Proteins, Fusion
Abstract

International audience; Binding of ligand FasL to its receptor Fas triggers apoptosis via the caspase cascade. FasL itself is homotrimeric, and a productive apoptotic signal requires that FasL be oligomerized beyond the homotrimeric state. We generated a series of FasL chimeras by fusing FasL to domains of the Leukemia Inhibitory Factor receptor gp190 which confer homotypic oligomerization, and analyzed the capacity of these soluble chimeras to trigger cell death. We observed that the most efficient FasL chimera, called pFasL, was also the most polymeric, as it reached the size of a dodecamer. Using a cellular model, we investigated the structure-function relationships of the FasL/Fas interactions for our chimeras, and we demonstrated that the Fas-mediated apoptotic signal did not solely rely on ligand-mediated receptor aggregation, but also required a conformational adaptation of the Fas receptor. When injected into mice, pFasL did not trigger liver injury at a dose which displayed anti-tumor activity in a model of human tumor transplanted to immunodeficient animals, suggesting a potential therapeutic use. Therefore, the optimization of the FasL conformation has to be considered for the development of efficient FasL-derived anti-cancer drugs targeting Fas.

Published
2013

30. Effects of a sulfated exopolysaccharide produced by Altermonas infernus on bone biology

Author

Mike Maillasson, Corinne Sinquin, Marc Padrines, Marc Baud'huin, C. Ruiz Velasco, Sylvia Colliec-Jouault, Dominique Heymann, Physiopathologie des Adaptations Nutritionnelles (PhAN), Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), Université de Nantes (UN)-Université de Nantes (UN)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Institut Français de Recherche pour l'Exploitation de la Mer - Atlantique (IFREMER Atlantique), Institut Français de Recherche pour l'Exploitation de la Mer (IFREMER), Centre de Recherche en Cancérologie Nantes-Angers (CRCNA), Centre Hospitalier Universitaire d'Angers (CHU Angers), PRES Université Nantes Angers Le Mans (UNAM)-PRES Université Nantes Angers Le Mans (UNAM)-Hôtel-Dieu de Nantes-Institut National de la Santé et de la Recherche Médicale (INSERM)-Hôpital Laennec-Centre National de la Recherche Scientifique (CNRS)-Faculté de Médecine d'Angers-Centre hospitalier universitaire de Nantes (CHU Nantes), Centre hospitalier universitaire de Nantes (CHU Nantes), This work was supported by the Région des Pays de la Loire [Program entitled 'Ciblage Moléculaire et Applications Thérapeutique' (CIMATH)] and by the ANR 2007 INSERM Pathophysiology of Human Diseases project N° R07196NS. Carmen RUIZ VELASCO received a fellowship from the Région des Pays de la Loire., Heymann, D, and Institut National de la Recherche Agronomique (INRA)-Université de Nantes (UN)

Subjects
Swine, Osteoporosis, Apoptosis, MESH: Rats, Sprague-Dawley, heparin, Biochemistry, Bone remodeling, Rats, Sprague-Dawley, Mice, 0302 clinical medicine, Bone cell, Carbohydrate Conformation, MESH: Animals, MESH: Carbohydrate Conformation, MESH: Swine, 0303 health sciences, [SDV.MHEP.RSOA] Life Sciences [q-bio]/Human health and pathology/Rhumatology and musculoskeletal system, biology, Chemistry, Sulfates, MESH: Bone Marrow Cells, Cell biology, medicine.anatomical_structure, [SDV.MHEP.RSOA]Life Sciences [q-bio]/Human health and pathology/Rhumatology and musculoskeletal system, RANKL, 030220 oncology & carcinogenesis, exopolysaccharide, Cancellous bone, Stromal cell, MESH: Alteromonas, MESH: Rats, Bone Marrow Cells, Bone resorption, Article, 03 medical and health sciences, Osteoclast, Polysaccharides, MESH: Cell Proliferation, medicine, glycosaminoglycan, Animals, Humans, Bone, MESH: Mice, bone remodeling, 030304 developmental biology, Cell Proliferation, MESH: Humans, Heparin, MESH: Apoptosis, medicine.disease, Rats, MESH: Polysaccharides, Glycosaminoglycan, biology.protein, bone metabolism, Stromal Cells, MESH: Stromal Cells, Alteromonas, MESH: Sulfates
Abstract

International audience; The growth and differentiation of bone cells is controlled by various factors, which can be modulated by heparan sulfates. Here, we investigated the effects of an oversulfated exopolysaccharide (OS-EPS) on the bone. We compared the effect of this compound with that of a native EPS. Long-term administration of OS-EPS causes cancellous bone loss in mice due, in part, to an increase in the number of osteoclasts lining the trabecular bone surface. No significant difference in cancellous bone volume was found between EPS-treated mice and age-matched control mice, underlying the importance of sulfation in trabecular bone loss. However, the mechanism sustaining this osteoporosis was unclear. To clarify OS-EPS activities, we investigated the effect of OS-EPS on osteogenesis. Our results demonstrated that OS-EPS inhibited osteoclastogenesis in two cell models. Using the surface plasmon resonance technique, we revealed that OS-EPS can form a hetero-molecular complex OS-EPS/receptor activator of NF-κB ligand (RANKL)/RANK and that RANK had a higher affinity for RANKL pre-incubated with OS-EPS than for RANKL alone, which would be in favor of an increase in bone resorption. However, in vitro, OS-EPS inhibited the early steps of osteoclast precursor adhesion and therefore inhibited the cell fusion step. In addition, we showed that OS-EPS reduced proliferation and accelerated osteoblastic differentiation, leading to strong inhibition of mineralized nodule formation, which would be in favor of an increase in bone resorption. Taken together, these data show different levels of bone resorption regulation by EPSs, most of them leading to proresorptive effects.

Published
2011

31. Evolution of interleukin-15 for higher E. coli expression and solubility

Author

Véronique Solé, Mike Maillasson, Yannick Jacques, Agnès Quéméner, Alain Defontaine, Charles Tellier, Ghislaine Béhar, Unité de Biotechnologie, Biocatalyse et Biorégulation (U3B), Université de Nantes (UN)-Centre National de la Recherche Scientifique (CNRS), Centre de Recherche en Cancérologie Nantes-Angers (CRCNA), Centre Hospitalier Universitaire d'Angers (CHU Angers), PRES Université Nantes Angers Le Mans (UNAM)-PRES Université Nantes Angers Le Mans (UNAM)-Hôtel-Dieu de Nantes-Institut National de la Santé et de la Recherche Médicale (INSERM)-Hôpital Laennec-Centre National de la Recherche Scientifique (CNRS)-Faculté de Médecine d'Angers-Centre hospitalier universitaire de Nantes (CHU Nantes), and QUEMENER, Agnès

Subjects
Chloramphenicol O-Acetyltransferase, Sushi domain, Cytoplasm, [SDV.BIO]Life Sciences [q-bio]/Biotechnology, T-Lymphocytes, Mutant, Gene Expression, Bioengineering, [SDV.BBM.BM] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, medicine.disease_cause, Biochemistry, Cell Line, Chloramphenicol acetyltransferase, 03 medical and health sciences, 0302 clinical medicine, Interleukin-15 Receptor alpha Subunit, Escherichia coli, medicine, Humans, Solubility, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], [SDV.BBM.BC] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Molecular Biology, Cell Proliferation, 030304 developmental biology, Interleukin-15, 0303 health sciences, Chemistry, Chloramphenicol, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, Directed evolution, Molecular biology, [SDV.BIO] Life Sciences [q-bio]/Biotechnology, Cell culture, 030220 oncology & carcinogenesis, Mutation, Directed Molecular Evolution, Biotechnology, medicine.drug
Abstract

International audience; Directed evolution was used to generate IL-15 mutants with increased solubility and cytoplasmic over-expression in Escherichia coli. A protein solubility selection method was used in which the IL-15 gene was expressed as an N-terminal fusion to chloramphenicol acetyltransferase (CAT) as reporter protein. Clones that grew in the presence of high concentrations of chloramphenicol were then screened by ELISA to assay the binding activity of the IL-15-CAT fusion to the IL-15Ra Sushi domain. Two variants of IL-15, M38 and M253, containing five mutations and one mutation respectively, were selected with a dramatic improvement in solubility; the soluble concentration in cell culture was 12-to 18-fold higher, respectively, than for WT IL-15. Characterization of their binding to IL-15Ra and their ability to stimulate the T-cell growth response showed that M38 binds as strongly as native IL-15 to IL-15Ra and acts as an effective agonist of IL-15.

Published
2011

32. Antibody-Hapten Recognition at the Surface of Functionalized Liposomes Studied by SPR: Steric Hindrance of Pegylated Phospholipids in Stealth Liposomes Prepared for Targeted Radionuclide Delivery

Author

Mike Maillasson, Marie Mougin-Degraef, Yannick Jacques, Jacques Barbet, Alain Faivre-Chauvet, Jean-François Gestin, Patricia Remaud-Le Saëc, Eliot P. Botosoa, Centre de Recherche en Cancérologie Nantes-Angers (CRCNA), Centre Hospitalier Universitaire d'Angers (CHU Angers), PRES Université Nantes Angers Le Mans (UNAM)-PRES Université Nantes Angers Le Mans (UNAM)-Hôtel-Dieu de Nantes-Institut National de la Santé et de la Recherche Médicale (INSERM)-Hôpital Laennec-Centre National de la Recherche Scientifique (CNRS)-Faculté de Médecine d'Angers-Centre hospitalier universitaire de Nantes (CHU Nantes), Plateforme Protéomique-Biogenouest (PPB), Institut de recherche en santé, environnement et travail (Irset), Université d'Angers (UA)-Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-École des Hautes Études en Santé Publique [EHESP] (EHESP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique )-Université d'Angers (UA)-Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-École des Hautes Études en Santé Publique [EHESP] (EHESP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique )-Proteomics Core Facility (Protim), Université de Rennes 1 (UR1), Université de Rennes (UNIV-RENNES)-Université de Rennes (UNIV-RENNES)-Plateforme Génomique Santé Biogenouest®-Plateforme Génomique Santé Biogenouest®, This work was supported in part by the EC through the FP7 collaborative project TARCC, the French Agence Nationale de la Recherche through the PCV grant VecRIT, and by INCa through the Action Concertee Incitative 2007 'Innovative delivery systems for cancer radionuclide therapy.' The authors also thank Dr. Cedric Gaillard from 'Plate-forme RIO BIBS INRA de Nantes' for all AFM experiments., ANR-07-PCVI-0019,VecRIT,Vectorisation de radioéléments pour la radioimmunothérapie des tumeurs solides de petite taille(2007), European Project: 201962,EC:FP7:HEALTH,FP7-HEALTH-2007-A,TARCC(2008), Université d'Angers (UA)-Université de Rennes (UR)-École des Hautes Études en Santé Publique [EHESP] (EHESP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique )-Université d'Angers (UA)-Université de Rennes (UR)-École des Hautes Études en Santé Publique [EHESP] (EHESP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Structure Fédérative de Recherche en Biologie et Santé de Rennes ( Biosit : Biologie - Santé - Innovation Technologique )-Proteomics Core Facility (Protim), Université de Rennes (UR)-Plateforme Génomique Santé Biogenouest®-Plateforme Génomique Santé Biogenouest®, Bernardo, Elizabeth, Physique et chimie du vivant - Vectorisation de radioéléments pour la radioimmunothérapie des tumeurs solides de petite taille - - VecRIT2007 - ANR-07-PCVI-0019 - PCVI - VALID, and Targeting alpha-particle emitting radionuclides to combat cancer - TARCC - - EC:FP7:HEALTH2008-01-01 - 2011-06-30 - 201962 - VALID

Subjects
Steric effects, 0303 health sciences, Liposome, Article Subject, Chemistry, lcsh:RS1-441, [SDV.CAN]Life Sciences [q-bio]/Cancer, 02 engineering and technology, 021001 nanoscience & nanotechnology, Receptor–ligand kinetics, lcsh:Pharmacy and materia medica, 03 medical and health sciences, [SDV.CAN] Life Sciences [q-bio]/Cancer, PEG ratio, Immunology, Biophysics, Moiety, Surface plasmon resonance, 0210 nano-technology, Lipid bilayer, Hapten, 030304 developmental biology, Research Article
Abstract

International audience; Targeted PEGylated liposomes could increase the amount of drugs or radionuclides delivered to tumor cells. They show favorable stability and pharmacokinetics, but steric hindrance of the PEG chains can block the binding of the targeting moiety. Here, specific interactions between an antihapten antibody (clone 734, specific for the DTPA-indium complex) and DTPA-indium-tagged liposomes were characterized by surface plasmon resonance (SPR). Non-PEGylated liposomes fused on CM5 chips whereas PEGylated liposomes did not. By contrast, both PEGylated and non-PEGylated liposomes attached to L1 chips without fusion. SPR binding kinetics showed that, in the absence of PEG, the antibody binds the hapten at the surface of lipid bilayers with the affinity of the soluble hapten. The incorporation of PEGylated lipids hinders antibody binding to extents depending on PEGylated lipid fraction and PEG molecular weight. SPR on immobilized liposomes thus appears as a useful technique to optimize formulations of liposomes for targeted therapy.

Published
2011

33. Definition and Characterization of an Inhibitor for Interleukin-31

Author

Sylvie Chevalier, Laurence Preisser, Mike Maillasson, Yannick Jacques, Caroline Diveu, Linda Grimaud, Emilie Venereau, Josy Froger, Elisa Ravon, Hugues Gascan, Yannic Danger, Laure Garrigue-Antar, Cytokines : structure, signalisation et prolifération tumorale, Université d'Angers (UA)-Institut National de la Santé et de la Recherche Médicale (INSERM), PRES Université Nantes Angers Le Mans (UNAM), Institut de Recherche en Horticulture et Semences (IRHS), Université d'Angers (UA)-Institut National de la Recherche Agronomique (INRA)-AGROCAMPUS OUEST, Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro)-Institut national d'enseignement supérieur pour l'agriculture, l'alimentation et l'environnement (Institut Agro), Centre de Recherche en Cancérologie Nantes-Angers (CRCNA), Centre Hospitalier Universitaire d'Angers (CHU Angers), PRES Université Nantes Angers Le Mans (UNAM)-PRES Université Nantes Angers Le Mans (UNAM)-Hôtel-Dieu de Nantes-Institut National de la Santé et de la Recherche Médicale (INSERM)-Hôpital Laennec-Centre National de la Recherche Scientifique (CNRS)-Faculté de Médecine d'Angers-Centre hospitalier universitaire de Nantes (CHU Nantes), Recherches en cancérologie, Université de Nantes (UN)-IFR26-Institut National de la Santé et de la Recherche Médicale (INSERM), Chercheur indépendant, AGROCAMPUS OUEST-Institut National de la Recherche Agronomique (INRA)-Université d'Angers (UA), Centre de Recherche en Cancérologie / Nantes - Angers (CRCNA), Centre hospitalier universitaire de Nantes (CHU Nantes)-Faculté de Médecine d'Angers-Centre Hospitalier Universitaire d'Angers (CHU Angers), and PRES Université Nantes Angers Le Mans (UNAM)-PRES Université Nantes Angers Le Mans (UNAM)-Centre National de la Recherche Scientifique (CNRS)-Hôpital Laennec-Institut National de la Santé et de la Recherche Médicale (INSERM)-Hôtel-Dieu de Nantes

Subjects
Recombinant Fusion Proteins, Mice, Transgenic, Biology, Polymerase Chain Reaction, Biochemistry, Cell Line, Dermatitis, Atopic, Mice, 03 medical and health sciences, Cell surface receptor, Animals, Humans, Immunoprecipitation, Phosphorylation, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], Receptor, Molecular Biology, ComputingMilieux_MISCELLANEOUS, Cell Proliferation, 030304 developmental biology, Common gamma chain, 0303 health sciences, Interleukins, 030302 biochemistry & molecular biology, Oncostatin M receptor, Receptors, Oncostatin M, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, Receptors, Interleukin, Cell Biology, Molecular biology, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], Interleukin 10, Interleukin 31, Haplotypes, Interleukin-21 receptor, Signal transduction, [SDV.MHEP.DERM]Life Sciences [q-bio]/Human health and pathology/Dermatology
Abstract

Interleukin-31 (IL-31) is a recently described T cell-derived cytokine, mainly produced by T helper type 2 cells and related to the IL-6 cytokine family according to its structure and receptor. IL-31 is the ligand for a heterodimeric receptor composed of a gp130-like receptor (GPL) associated with the oncostatin M receptor (OSMR). A link between IL-31 and atopic dermatitis was shown by studying the phenotype of IL-31 transgenic mice and IL-31 gene haplotypes in patients suffering from dermatitis. In this study, we generated a potent IL-31 antagonist formed by external portions of OSMR and GPL fused with a linker. This fusion protein, OSMR-L-GPL, consisting of 720 amino acids, counteracted the binding of IL-31 to its membrane receptor complex and the subsequent signaling events involving the STATs and MAPK pathways. Neutralizing effects were found in IL-31-sensitive cell lines, including brain-derived cells and primary cultures of keratinocytes.

Published
2010

34. Factor VIII-von Willebrand Factor Complex Inhibits Osteoclastogenesis and Controls Cell Survival

Author

Marc Fouassier, Marc Baud'huin, Stéphane Téletchéa, Dominique Heymann, Céline Charrier, Mike Maillasson, and Laurence Duplomb

Subjects
musculoskeletal diseases, medicine.medical_specialty, congenital, hereditary, and neonatal diseases and abnormalities, Platelet Aggregation, Cell Survival, Osteoclasts, Apoptosis, Bone Neoplasms, Enzyme-Linked Immunosorbent Assay, Sarcoma, Ewing, 030204 cardiovascular system & hematology, Biochemistry, TNF-Related Apoptosis-Inducing Ligand, 03 medical and health sciences, Mice, 0302 clinical medicine, Molecular Basis of Cell and Developmental Biology, Osteoprotegerin, Von Willebrand factor, Internal medicine, hemic and lymphatic diseases, von Willebrand Factor, medicine, Animals, Humans, Receptor, Molecular Biology, Cells, Cultured, 030304 developmental biology, Cell Proliferation, 0303 health sciences, Osteosarcoma, Factor VIII, biology, Chemistry, RANK Ligand, Cell Differentiation, Cell Biology, Surface Plasmon Resonance, Ligand (biochemistry), Endocrinology, Coagulation, RANKL, Cancer research, biology.protein, Binding domain, circulatory and respiratory physiology
Abstract

Factor VIII-von Willebrand factor (FVIII·vWF) complex, a molecule involved in coagulation, can be physically associated with osteoprotegerin (OPG). OPG is an anti-osteoclastic protein and a soluble receptor for the proapoptotic protein TRAIL (tumor necrosis factor-related apoptosis-inducing ligand), suggesting a potential role of FVIII·vWF complex in bone and cancer biology. We, thus, assessed the effects of FVIII·vWF complex on osteoclastogenesis and cell survival. We first evidenced that FVIII·vWF complex inhibited RANKL-induced osteoclastogenesis and enhanced the inhibitory effect of OPG. Interestingly, we revealed by surface plasmon resonance that FVIII·vWF complex bound to RANKL, whereas recombinant FVIII and vWF did not. By modeling, we showed that the OPG binding domain to the A1 domain of vWF was closely located and partially overlapped to its binding site to RANKL. Then, we demonstrated that FVIII·vWF complex cancelled the inhibitory activity of OPG on TRAIL-induced apoptosis and characterized interactions between these molecules. The present work evidenced a direct activity of FVIII·vWF complex on osteoclasts and on induced cell apoptosis, pointing out its potential involvement in physiological bone remodeling or in bone damages associated with severe hemophilia and cancer development.

Published
2009

36. Glycosaminoglycans inhibit rankl-induced osteoclastogenesis

Author

Marc Baud'huin, Dominique Heymann, Mike Maillasson, Françoise Rédini, C. Ruiz Velasco, Gaëtan Jego, Céline Charrier, and Laurence Duplomb

Subjects
Glycosaminoglycan, Histology, biology, Physiology, Chemistry, RANKL, Endocrinology, Diabetes and Metabolism, Cancer research, biology.protein
Published
2009

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