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820 results on '"MESH: Cell Line, Tumor"'

1. Discoidin Domain Receptor 2 orchestrates melanoma resistance combining phenotype switching and proliferation

Author

Margaux Sala, Nathalie Allain, Mélanie Moreau, Arnaud Jabouille, Elodie Henriet, Aya Abou-Hammoud, Arnaud Uguen, Sylvaine Di-Tommaso, Cyril Dourthe, Anne-Aurélie Raymond, Jean-William Dupuy, Emilie Gerard, Nathalie Dugot-Senant, Benoit Rousseau, Jean-Phillipe Merlio, Anne Pham-Ledart, Béatrice Vergier, Sophie Tartare-Deckert, Violaine Moreau, Frédéric Saltel, BoRdeaux Institute in onCology (Inserm U1312 - BRIC), Université de Bordeaux (UB)-Institut National de la Santé et de la Recherche Médicale (INSERM), Johns Hopkins University (JHU), Microbiologie Fondamentale et Pathogénicité (MFP), Université Bordeaux Segalen - Bordeaux 2-Centre National de la Recherche Scientifique (CNRS), Hôpital Morvan - CHRU de Brest (CHU - BREST ), TBM-Core [Bordeaux] (CNRS UMS 3427 - INSERM US 005), Université de Bordeaux (UB)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Université de Bordeaux (UB), CHU Bordeaux [Bordeaux], Bordeaux Research In Translational Oncology [Bordeaux] (BaRITOn), Université de Bordeaux (UB)-CHU Bordeaux [Bordeaux]-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre méditerranéen de médecine moléculaire (C3M), Université Nice Sophia Antipolis (1965 - 2019) (UNS), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Côte d'Azur (UCA), and TARTARE-DECKERT, Sophie

Subjects
Proto-Oncogene Proteins B-raf, MESH: Proto-Oncogene Proteins B-raf, Cancer Research, MESH: Cell Line, Tumor, MESH: Humans, MESH: Melanoma, [SDV]Life Sciences [q-bio], MESH: Phenotype, MESH: Drug Resistance, Neoplasm, [SDV] Life Sciences [q-bio], Discoidin Domain Receptor 2, Phenotype, Drug Resistance, Neoplasm, Cell Line, Tumor, MESH: Cell Proliferation, MESH: Discoidin Domain Receptor 2, Genetics, Humans, MESH: Protein Kinase Inhibitors, Melanoma, Protein Kinase Inhibitors, Molecular Biology, Cell Proliferation
Abstract

International audience; Combined therapy with anti-BRAF plus anti-MEK is currently used as first-line treatment of patients with metastatic melanomas harboring the somatic BRAF V600E mutation. However, the main issue with targeted therapy is the acquisition of tumor cell resistance. In a majority of resistant melanoma cells, the resistant process consists in epithelial-to-mesenchymal transition (EMT). This process called phenotype switching makes melanoma cells more invasive. Its signature is characterized by MITF low, AXL high, and actin cytoskeleton reorganization through RhoA activation. In parallel of this phenotype switching phase, the resistant cells exhibit an anarchic cell proliferation due to hyper-activation of the MAP kinase pathway. We show that a majority of human melanoma overexpress discoidin domain receptor 2 (DDR2) after treatment. The same result was found in resistant cell lines presenting phenotype switching compared to the corresponding sensitive cell lines. We demonstrate that DDR2 inhibition induces a decrease in AXL expression and reduces stress fiber formation in resistant melanoma cell lines. In this phenotype switching context, we report that DDR2 control cell and tumor proliferation through the MAP kinase pathway in resistant cells in vitro and in vivo. Therefore, inhibition of DDR2 could be a new and promising strategy for countering this resistance mechanism.

Published
2022

2. NSs amyloid formation is associated with the virulence of Rift Valley fever virus in mice

Author

Susann Kummer, Hans-Georg Kräusslich, Psylvia Léger, Steeve Boulant, Jana Koch, Pierre-Yves Lozach, Megan L. Stanifer, Qilin Xin, Michèle Bouloy, Robin Burk, Carmen Nussbaum-Krammer, Carole Tamietti, Marie Flamand, Karsten Richter, Eliana Nachman, Xavier Montagutelli, Heidelberg University Hospital [Heidelberg], Center for Molecular Biology - Zentrum für Molekulare Biologie [Heidelberg, Germany] (ZMBH), Universität Heidelberg [Heidelberg] = Heidelberg University, German Cancer Research Center - Deutsches Krebsforschungszentrum [Heidelberg] (DKFZ), Virologie Structurale - Structural Virology, Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), Infections Virales et Pathologie Comparée - UMR 754 (IVPC), École Pratique des Hautes Études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Génétique Moléculaire des Bunyavirus, Institut Pasteur [Paris] (IP), Génétique de la souris - Mouse Genetics, This work was supported by grants from CellNetworks Research Group funds, Heidelberg, and from the Deutsche Forschungsgemeinschaft (DFG, LO-2338/1-1 and LO-2338/3-1). It was also supported by a Chinese Scholarship Council fellowship to Q.X, Universität Heidelberg [Heidelberg], Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), École pratique des hautes études (EPHE), and Institut Pasteur [Paris]

Subjects
0301 basic medicine, Rift Valley Fever, General Physics and Astronomy, Protein aggregation, Viral Nonstructural Proteins, MESH: Virulence, Virulence factor, chemistry.chemical_compound, Mice, MESH: Chlorocebus aethiops, Chlorocebus aethiops, MESH: Rift Valley Fever, MESH: Microscopy, Confocal, MESH: Animals, lcsh:Science, Multidisciplinary, Microscopy, Confocal, MESH: Protein Aggregation, Pathological, Virulence, Virus structures, 3. Good health, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, MESH: Microscopy, Electron, Transmission, MESH: Rift Valley fever virus, Thioflavin, Encephalitis, MESH: Cell Nucleus, Amyloid, MESH: Cell Line, Tumor, Virulence Factors, Prions, Science, MESH: Vero Cells, Amyloidogenic Proteins, macromolecular substances, Biology, Fibril, Protein Aggregation, Pathological, General Biochemistry, Genetics and Molecular Biology, Article, 03 medical and health sciences, Microscopy, Electron, Transmission, Cell Line, Tumor, mental disorders, medicine, Animals, Humans, Vero Cells, MESH: Mice, MESH: Virulence Factors, Cell Nucleus, MESH: Amyloid, MESH: Amyloidogenic Proteins, MESH: Humans, 030102 biochemistry & molecular biology, General Chemistry, medicine.disease, Rift Valley fever virus, Virology, 030104 developmental biology, chemistry, MESH: HeLa Cells, Vero cell, MESH: Viral Nonstructural Proteins, lcsh:Q, HeLa Cells
Abstract

Amyloid fibrils result from the aggregation of host cell-encoded proteins, many giving rise to specific human illnesses such as Alzheimer’s disease. Here we show that the major virulence factor of Rift Valley fever virus, the protein NSs, forms filamentous structures in the brain of mice and affects mortality. NSs assembles into nuclear and cytosolic disulfide bond-dependent fibrillary aggregates in infected cells. NSs structural arrangements exhibit characteristics typical for amyloids, such as an ultrastructure of 12 nm-width fibrils, a strong detergent resistance, and interactions with the amyloid-binding dye Thioflavin-S. The assembly dynamics of viral amyloid-like fibrils can be visualized in real-time. They form spontaneously and grow in an amyloid fashion within 5 hours. Together, our results demonstrate that viruses can encode amyloid-like fibril-forming proteins and have strong implications for future research on amyloid aggregation and toxicity in general., Rift Valley fever virus (RVFV) can cause severe diseases in humans, including encephalitis. Here the authors show that NSs, the major virulence factor of RVFV, is an amyloidogenic protein forming fibrils in infected mouse brains and causing increased mortality in mice.

Published
2020

3. Menin inhibition suppresses castration-resistant prostate cancer and enhances chemosensitivity

Author

Cherif, Chaïma, Nguyen, Dang Tan, Paris, Clément, Le, Thi Khanh, Sefiane, Thibaud, Carbuccia, Nadine, Finetti, Pascal, Chaffanet, Max, Kaoutari, Abdessamad El, Vernerey, Julien, Fazli, Ladan, Gleave, Martin, Manai, Mohamed, Barthélémy, Philippe, Birnbaum, Daniel, Bertucci, François, Taïeb, David, Rocchi, Palma, Centre de Recherche en Cancérologie de Marseille (CRCM), Aix Marseille Université (AMU)-Institut Paoli-Calmettes, Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), University of British Columbia (UBC), University of Tunis El Manar, Acides Nucléiques : Régulations Naturelle et Artificielle (ARNA), Université de Bordeaux (UB)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), and Rocchi, Palma

Subjects
Male, congenital, hereditary, and neonatal diseases and abnormalities, endocrine system, MESH: Cell Line, Tumor, MESH: Humans, endocrine system diseases, MESH: Genomics, [SDV]Life Sciences [q-bio], Genomics, Hormone receptors, urologic and male genital diseases, Article, MESH: Male, MESH: Proto-Oncogene Proteins, [SDV] Life Sciences [q-bio], Prostatic Neoplasms, Castration-Resistant, Cell Line, Tumor, Proto-Oncogene Proteins, MESH: Cell Proliferation, MESH: Prostatic Neoplasms, Castration-Resistant, Animals, Humans, MESH: Animals, Cancer genetics, Cell Proliferation
Abstract

International audience; Abstract Disease progression and therapeutic resistance of prostate cancer (PC) are linked to multiple molecular events that promote survival and plasticity. We previously showed that heat shock protein 27 (HSP27) acted as a driver of castration-resistant phenotype (CRPC) and developed an oligonucleotides antisense (ASO) against HSP27 with evidence of anti-cancer activity in men with CRPC. Here, we show that the tumor suppressor Menin ( MEN1 ) is highly regulated by HSP27. Menin is overexpressed in high-grade PC and CRPC. High MEN1 mRNA expression is associated with decreased biochemical relapse-free and overall survival. Silencing Menin with ASO technology inhibits CRPC cell proliferation, tumor growth, and restores chemotherapeutic sensitivity. ChIP-seq analysis revealed differential DNA binding sites of Menin in various prostatic cells, suggesting a switch from tumor suppressor to oncogenic functions in CRPC. These data support the evaluation of ASO against Menin for CRPC.

Published
2022

4. Dual-drug loaded nanomedicine hydrogel as a therapeutic platform to target both residual glioblastoma and glioma stem cells

Author

Elia Bozzato, Nikolaos Tsakiris, Adrien Paquot, Giulio G. Muccioli, Chiara Bastiancich, Véronique Préat, Institut de neurophysiopathologie (INP), and Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS)

Subjects
MESH: Hydrogels, Local delivery, MESH: Cell Line, Tumor, Curcumin, [SDV]Life Sciences [q-bio], Pharmaceutical Science, [SDV.CAN]Life Sciences [q-bio]/Cancer, Cell Line, MESH: Glioma, MESH: Curcumin, Cell Line, Tumor, Glioma stem cells, Humans, MESH: Humans, Tumor, Gemcitabine, Glioblastoma, Hydrogel, Salinomycin, Nanomedicine, Hydrogels, Lipids, Neoplastic Stem Cells, Brain Neoplasms, Glioma, MESH: Glioblastoma, MESH: Lipids, MESH: Neoplastic Stem Cells, MESH: Nanomedicine, MESH: Brain Neoplasms
Abstract

Glioblastoma (GBM) recurrences are inevitable, and mainly originate from residual tumor cells and the presence of glioma stem cells (GSC) around the resection cavity borders. We previously showed that the local treatment of GBM with nanomedicine-based Lauroyl-gemcitabine lipid nanocapsules (GemC12-LNC) hydrogel delayed tumor onset in various preclinical models and can be used as a scaffold to deliver multiple drugs. However, it does not inhibit tumor relapse in the long-term. In this work, we aim at encapsulating an anti-GSC molecule in the GemC12-LNC hydrogel to eliminate both GBM cells and GSC. We performed a screening on GBM cell lines (GL261 and U-87 MG) and patient-derived GSC (GBM9) to select the anti-GSC molecule that could act synergically with GemC12. Based on our results, salinomycin (Sal) and curcumin (Cur) were selected for further development. Both GemC12-Sal-LNC and GemC12-Cur LNC showed similar size (55 nm), zeta potential (- 2 mV) and viscoelastic properties compared to the GemC12-LNC hydrogel. Encapsulation efficiency was above 95 %. Moreover, the GemC12-Sal-LNC hydrogel was stable for at least 6 months and released both drugs over 30 days in vitro. Both hydrogels inhibited the growth of GL261 and U-87 MG spheroids. Flow cytometry analysis showed that Sal reduced the GSC population in GL261 and U-87 MG cells. Our results show that the co-encapsulation of Sal in the GemC12-LNC hydrogel can reduce both GBM cells and GSC, and therefore might be promising to avoid the onset of GBM recurrences.

Published
2022

5. MAD2L2 dimerization and TRIP13 control shieldin activity in DNA repair

Author

de Krijger, Inge, Föhr, Bastian, Pérez, Santiago Hernández, Vincendeau, Estelle, Serrat, Judit, Thouin, Alexander Marc, Susvirkar, Vivek, Lescale, Chloé, Paniagua, Inés, Hoekman, Liesbeth, Kaur, Simranjeet, Altelaar, Maarten, Deriano, Ludovic, Faesen, Alex C, Jacobs, Jacqueline J L, Afd Biomol.Mass Spect. and Proteomics, Biomolecular Mass Spectrometry and Proteomics, Netherlands Cancer Institute (NKI), Antoni van Leeuwenhoek Hospital, Max Planck Institute for Biophysical Chemistry (MPI-BPC), Max-Planck-Gesellschaft, Université Paris Diderot, Sorbonne Paris Cité, Paris, France, Université Paris Diderot - Paris 7 (UPD7), Intégrité du génome, immunité et cancer - Genome integrity, Immunity and Cancer, Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Physiopathologie du système immunitaire (Inserm U1223), Utrecht Institute for Pharmaceutical Sciences (UIPS), Utrecht University [Utrecht], This work was supported by project grant 10999/2017-1 from the Dutch Cancer Society (KWF) to J.J.L.J., by the Institut Pasteur to L.D., by the Institut National Du Cancer (INCA_13852) to L.D., by the Ligue Nationale Contre le Cancer (Equipe labellisée 2019) to L.D. The Proteomics Facility of the Netherlands Cancer Institute was supported by the X-omics Initiative, partially funded by the Dutch Research Council (NWO)(project 184.034.019). E.V. received funding from Université de Paris, Sorbonne Paris Cité. A.C.F. is grateful for generous core funding by the Max Planck Society., Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Cité (UPCité), Université Sorbonne Paris Cité (USPC), Afd Biomol.Mass Spect. and Proteomics, Biomolecular Mass Spectrometry and Proteomics, Institut Pasteur [Paris], Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Pasteur [Paris], and Lescale, Chloé

Subjects
DNA Repair, Chemistry(all), ATPase, MESH: DNA Breaks, Double-Stranded, Gene Expression, General Physics and Astronomy, Cell Cycle Proteins, [SDV.GEN] Life Sciences [q-bio]/Genetics, [SDV.BBM.BM] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, Biochemistry, Piperazines, MESH: Recombinant Proteins, Mice, chemistry.chemical_compound, 0302 clinical medicine, DNA Breaks, Double-Stranded, MESH: Animals, 0303 health sciences, Multidisciplinary, biology, Chemistry, MESH: Protein Multimerization, DNA damage and repair, MESH: DNA, Recombinant Proteins, 3. Good health, Cell biology, DNA-Binding Proteins, MESH: ATPases Associated with Diverse Cellular Activities, MESH: HEK293 Cells, Mad2 Proteins, MESH: Phthalazines, MESH: Mad2 Proteins, Protein Binding, MESH: Cell Line, Tumor, MESH: Gene Expression, DNA repair, Science, Physics and Astronomy(all), Article, Chromosomes, General Biochemistry, Genetics and Molecular Biology, Cell Line, 03 medical and health sciences, MESH: Cell Cycle Proteins, Cell Line, Tumor, Animals, Humans, MESH: Protein Binding, Protein Interaction Domains and Motifs, MESH: Mice, 030304 developmental biology, MESH: Protein Interaction Domains and Motifs, [SDV.GEN]Life Sciences [q-bio]/Genetics, Binding Sites, MESH: Humans, TRIP13, Biochemistry, Genetics and Molecular Biology(all), [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, DNA, General Chemistry, DNA Repair Pathway, Fibroblasts, MESH: Cell Line, HEK293 Cells, MESH: Binding Sites, MESH: Cisplatin, MESH: Fibroblasts, MESH: HeLa Cells, biology.protein, ATPases Associated with Diverse Cellular Activities, Phthalazines, Cisplatin, Protein Multimerization, 030217 neurology & neurosurgery, Function (biology), MESH: DNA-Binding Proteins, HeLa Cells, Genetics and Molecular Biology(all)
Abstract

MAD2L2 (REV7) plays an important role in DNA double-strand break repair. As a member of the shieldin complex, consisting of MAD2L2, SHLD1, SHLD2 and SHLD3, it controls DNA repair pathway choice by counteracting DNA end-resection. Here we investigated the requirements for shieldin complex assembly and activity. Besides a dimerization-surface, HORMA-domain protein MAD2L2 has the extraordinary ability to wrap its C-terminus around SHLD3, likely creating a very stable complex. We show that appropriate function of MAD2L2 within shieldin requires its dimerization, mediated by SHLD2 and accelerating MAD2L2-SHLD3 interaction. Dimerization-defective MAD2L2 impairs shieldin assembly and fails to promote NHEJ. Moreover, MAD2L2 dimerization, along with the presence of SHLD3, allows shieldin to interact with the TRIP13 ATPase, known to drive topological switches in HORMA-domain proteins. We find that appropriate levels of TRIP13 are important for proper shieldin (dis)assembly and activity in DNA repair. Together our data provide important insights in the dependencies for shieldin activity., MAD2L2 — a member of the shieldin complex — is known to play important roles in DNA repair. Here the authors demonstrate how MAD2L2 dimerization mediated through SHLD2 participates in shieldin assembly and function.

Published
2021

6. TMPRSS2:ERG gene fusion expression regulates bone markers and enhances the osteoblastic phenotype of prostate cancer bone metastases

Author

Rachel Deplus, Martine Duterque-Coquillaud, Anne Flourens, Edith Bonnelye, Nathalie Vanpouille, Tian V. Tian, Philippe Clézardin, Anais Fradet, Carine Delliaux, Xavier Leroy, Yvan de Launoit, Mathilde Bouchet, Arnauld Villers, Mécanismes de tumorigenèse et thérapies ciblées, Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Institut de Recherches Cliniques de Montréal (IRCM), Université de Montréal (UdeM), Universitat Pompeu Fabra [Barcelona] (UPF), Physiopathologie, diagnostic et traitements des maladies osseuses / Pathophysiology, Diagnosis & Treatments of Bone Diseases (LYOS), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), Pôle de Biologie Pathologie Génétique [CHU Lille], This work was in part supported by the Centre national de la recherche scientifique (CNRS), Ligue nationale contre le Cancer (Comité du Pas-de-Calais), Institut national du cancer (INCa_4419), Institut Pasteur de Lille, Conseil Régional du Nord-Pas-de-Calais and Fondation pour la Recherche Médicale (FRM), Conseil Régional du Nord-Pas-de-Calais, Association pour la Recherche sur le Cancer (ARC), MINECO (Juan de la Cierva Postdoctoral Fellowship FJCI-2014-22946), ARTP (Association de Recherche sur les tumeurs de prostate)., Mécanismes de la Tumorigénèse et Thérapies Ciblées (M3T) - UMR 8161 (M3T), Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), Center for Genomic Regulation (CRG-UPF), CIBER de Epidemiología y Salud Pública (CIBERESP), Service d'urologie, Hôpital Huriez-Centre Hospitalier Régional Universitaire [Lille] (CHRU Lille), We would like to thank M. Tardivel, A. Bongiovanni and E. Werkmeister from the BioImaging Center Lille Nord de France (BICeL) for their technical assistance, M. Canouil for his statistics advice, and and M. Vernier for his bioinformatic advice and stimulating discussions. We would also like to thank the local Tumor Tissue Bank (Tumorothèque), Regional Reference Oncology Center (CRRC) (Head, Pr. M.C. Copin) in Lille, France.

Subjects
Male, 0301 basic medicine, Cancer Research, Oncogene Proteins, Fusion, [SDV]Life Sciences [q-bio], MESH: Gene Expression Regulation, Neoplastic, Mice, SCID, urologic and male genital diseases, Fusion gene, MESH: Oncogene Proteins, Fusion/metabolism, Metastasis, Prostate cancer, 0302 clinical medicine, MESH: Animals, MESH: Mice, SCID, MESH: Osteoblasts/pathology, ComputingMilieux_MISCELLANEOUS, MESH: Prostatic Neoplasms/genetics, Endothelin-1, Bone metastasis, MESH: Prostatic Neoplasms/pathology, Tumor Burden, 3. Good health, Gene Expression Regulation, Neoplastic, Phenotype, Osteoblastic lesions, Oncology, 030220 oncology & carcinogenesis, PC-3 Cells, MESH: Bone Neoplasms/metabolism, MESH: Bone Neoplasms/secondary, MESH: PC-3 Cells, MESH: Endothelin-1/metabolism, Erg, MESH: Alkaline Phosphatase/metabolism, MESH: Cell Line, Tumor, Transplantation, Heterologous, MESH: Biomarkers, Tumor/genetics, MESH: Tumor Burden/genetics, Bone Neoplasms, [SDV.CAN]Life Sciences [q-bio]/Cancer, Biology, MESH: Phenotype, TMPRSS2, MESH: Oncogene Proteins, Fusion/genetics, 03 medical and health sciences, Transcriptional regulation, Cell Line, Tumor, Biomarkers, Tumor, medicine, Animals, Humans, MESH: Transplantation, Heterologous, MESH: Biomarkers, Tumor/metabolism, Osteoblasts, MESH: Alkaline Phosphatase/genetics, MESH: Humans, MESH: Prostatic Neoplasms/metabolism, Prostatic Neoplasms, Gene rearrangement, Alkaline Phosphatase, MESH: Bone Neoplasms/genetics, MESH: Osteoblasts/metabolism, medicine.disease, MESH: Male, Collagen Type I, alpha 1 Chain, TMPRSS2:ERG, 030104 developmental biology, MESH: Endothelin-1/genetics, Cancer research, Ectopic expression
Abstract

International audience; Abstract : Prostate cancers have a strong propensity to metastasize to bone and promote osteoblastic lesions. TMPRSS2:ERG is the most frequent gene rearrangement identified in prostate cancer, but whether it is involved in prostate cancer bone metastases is largely unknown. We exploited an intratibial metastasis model to address this issue and we found that ectopic expression of the TMPRSS2:ERG fusion enhances the ability of prostate cancer cell lines to induce osteoblastic lesions by stimulating bone formation and inhibiting the osteolytic response. In line with these in vivo results, we demonstrate that the TMPRSS2:ERG fusion protein increases the expression of osteoblastic markers, including Collagen Type I Alpha 1 Chain and Alkaline Phosphatase, as well as Endothelin-1, a protein with a documented role in osteoblastic bone lesion formation. Moreover, we determined that the TMPRSS2:ERG fusion protein is bound to the regulatory regions of these genes in prostate cancer cell lines, and we report that the expression levels of these osteoblastic markers are correlated with the expression of the TMPRSS2:ERG fusion in patient metastasis samples. Taken together, our results reveal that the TMPRSS2:ERG gene fusion is involved in osteoblastic lesion formation induced by prostate cancer cells.

Published
2018

7. Alternative DNA Structures In Vivo : Molecular Evidence and Remaining Questions

Author

Guy-Franck Richard, Lucie Poggi, Collège Doctoral, Sorbonne Université (SU), Génétique des génomes - Genetics of Genomes (UMR 3525), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), and Work in our laboratory is generously supported by the Institut Pasteur and by the Centre National de la Recherche Scientifique (CNRS). L.P. was supported by the Fondation Blanchecape and the Association Française contre les Myopathies (AFM).

Subjects
MESH: Sequence Inversion, MESH: Trinucleotide Repeats, MESH: Cell Line, Tumor, Inverted repeat, [SDV]Life Sciences [q-bio], DNA hairpin, MESH: Base Pairing, Computational biology, Biology, G-quadruplex, Microbiology, 03 medical and health sciences, chemistry.chemical_compound, 0302 clinical medicine, MESH: Chromosome Fragile Sites, MESH: Animals, Molecular Biology, 030304 developmental biology, secondary structures, 0303 health sciences, MESH: Humans, Chromosomal fragile site, MESH: DNA, palindromes, mismatch repair, MESH: DNA Mismatch Repair, Infectious Diseases, Minisatellite, chemistry, G quadruplex, trinucleotide repeats, MESH: Minisatellite Repeats, MESH: HeLa Cells, cruciform, DNA mismatch repair, fragile sites, 030217 neurology & neurosurgery, DNA, Function (biology), MESH: G-Quadruplexes, Triple helix
Abstract

International audience; Duplex DNA naturally folds into a right-handed double helix in physiological conditions. Some sequences of unusual base composition may nevertheless form alternative structures, as was shown for many repeated sequences in vitro. However, evidence for the formation of noncanonical structures in living cells is difficult to gather. It mainly relies on genetic assays demonstrating their function in vivo or through genetic instability reflecting particular properties of such structures. Efforts were made to reveal their existence directly in a living cell, mainly by generating antibodies specific to secondary structures or using chemical ligands selected for their affinity to these structures. Among secondary structure-forming DNAs are G-quadruplexes, human fragile sites containing minisatellites, AT-rich regions, inverted repeats able to form cruciform structures, hairpin-forming CAG/CTG triplet repeats, and triple helices formed by homopurine-homopyrimidine GAA/TTC trinucleotide repeats. Many of these alternative structures are involved in human pathologies, such as neurological or developmental disorders, as in the case of trinucleotide repeats, or cancers triggered by translocations linked to fragile sites. This review will discuss and highlight evidence supporting the formation of alternative DNA structures in vivo and will emphasize the role of the mismatch repair machinery in binding mispaired DNA duplexes, triggering genetic instability.

Published
2021

8. Inhibition of InsP3R with Xestospongin B Reduces Mitochondrial Respiration and Induces Selective Cell Death in T Cell Acute Lymphoblastic Leukemia Cells

Author

Daniela Sauma, Alenka Lovy, Galdo Bustos, Ulises Ahumada-Castro, César Cárdenas, Jordi Molgó, Pablo Cruz, Universidad Mayor [Santiago de Chile], Service d'Ingénierie Moléculaire pour la Santé (ex SIMOPRO) (SIMoS), Médicaments et Technologies pour la Santé (MTS), Université Paris-Saclay-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE)-Université Paris-Saclay-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Universidad de Chile = University of Chile [Santiago] (UCHILE), University of California [Santa Barbara] (UC Santa Barbara), University of California (UC), and This research was funded by FONDECYT #1200255, 1180385, CONICYT/FONDAP # 15150012 and CONICYT Doctoral Fellowship Program funds PC (#21180306)

Subjects
MESH: Cell Death, 0301 basic medicine, Mitochondrion, Precursor T-Cell Lymphoblastic Leukemia-Lymphoma, bioenergetics, Jurkat cells, lcsh:Chemistry, chemistry.chemical_compound, MESH: Macrocyclic Compounds, 0302 clinical medicine, Inositol 1,4,5-Trisphosphate Receptors, Inositol, Receptor, Oxazoles, lcsh:QH301-705.5, Spectroscopy, Cell Death, MESH: Oxazoles, [SDV.MHEP.HEM]Life Sciences [q-bio]/Human health and pathology/Hematology, General Medicine, Computer Science Applications, Mitochondria, MESH: Precursor T-Cell Lymphoblastic Leukemia-Lymphoma, MESH: Leukocytes, Mononuclear, medicine.anatomical_structure, 030220 oncology & carcinogenesis, T-ALL, Programmed cell death, MESH: Cell Line, Tumor, Macrocyclic Compounds, MESH: Mitochondria, T cell, Cell Respiration, [SDV.CAN]Life Sciences [q-bio]/Cancer, Catalysis, Article, Inorganic Chemistry, 03 medical and health sciences, MESH: Inositol 1,4,5-Trisphosphate Receptors, Cell Line, Tumor, medicine, Humans, cancer, Physical and Theoretical Chemistry, Molecular Biology, MESH: Humans, calcium, Endoplasmic reticulum, Organic Chemistry, 030104 developmental biology, chemistry, lcsh:Biology (General), lcsh:QD1-999, Cell culture, MESH: Biomarkers, [SDV.SP.PHARMA]Life Sciences [q-bio]/Pharmaceutical sciences/Pharmacology, Cancer research, Leukocytes, Mononuclear, MESH: Cell Respiration, metabolism, Biomarkers
Abstract

International audience; T-cell acute lymphoblastic leukemia (T-ALL) is an aggressive hematological malignancy whose chemoresistance and relapse persist as a problem despite significant advances in its chemotherapeutic treatments. Mitochondrial metabolism has emerged as an interesting therapeutic target given its essential role in maintaining bioenergetic and metabolic homeostasis. T-ALL cells are characterized by high levels of mitochondrial respiration, making them suitable for this type of intervention. Mitochondrial function is sustained by a constitutive transfer of calcium from the endoplasmic reticulum to mitochondria through the inositol 1,4,5-trisphosphate receptor (InsP3R), making T-ALL cells vulnerable to its inhibition. Here, we determine the bioenergetic profile of the T-ALL cell lines CCRF-CEM and Jurkat and evaluate their sensitivity to InsP3R inhibition with the specific inhibitor, Xestospongin B (XeB). Our results show that T-ALL cell lines exhibit higher mitochondrial respiration than non-malignant cells, which is blunted by the inhibition of the InsP3R. Prolonged treatment with XeB causes T-ALL cell death without affecting the normal counterpart. Moreover, the combination of XeB and glucocorticoids significantly enhanced cell death in the CCRF-CEM cells. The inhibition of InsP3R with XeB rises as a potential therapeutic alternative for the treatment of T-ALL.

Published
2021

9. Instability of the NS1 Glycoprotein from La Reunion 2018 Dengue 2 Virus (Cosmopolitan-1 Genotype) in Huh7 Cells Is Due to Lysine Residues on Positions 272 and 324

Author

Philippe Desprès, Olivier Diaz, Pierre-Olivier Vidalain, Vincent Lotteau, Eva Ogire, Marjolaine Roche, Processus Infectieux en Milieu Insulaire Tropical (PIMIT), Université de La Réunion (UR)-Institut National de la Santé et de la Recherche Médicale (INSERM)-IRD-Centre National de la Recherche Scientifique (CNRS), Centre International de Recherche en Infectiologie - UMR (CIRI), École normale supérieure - Lyon (ENS Lyon)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Vidalain, Pierre-Olivier, Centre National de la Recherche Scientifique (CNRS)-IRD-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de La Réunion (UR), Centre International de Recherche en Infectiologie (CIRI), École normale supérieure de Lyon (ENS de Lyon)-Université Claude Bernard Lyon 1 (UCBL), and Université de Lyon-Université de Lyon-Université Jean Monnet - Saint-Étienne (UJM)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)

Subjects
0301 basic medicine, Serotype, viruses, MESH: Dengue, Dengue virus, Viral Nonstructural Proteins, medicine.disease_cause, soluble viral protein, MESH: Dengue Virus, Tanzania, Dengue fever, lcsh:Chemistry, MESH: Recombinant Proteins, MESH: Genotype, MESH: Hepatocytes, Dengue, multimeric viral protein, flavivirus, Genotype, lcsh:QH301-705.5, Antigens, Viral, Spectroscopy, chemistry.chemical_classification, [SDV.MP.VIR] Life Sciences [q-bio]/Microbiology and Parasitology/Virology, biology, MESH: Protein Multimerization, Protein Stability, virus diseases, General Medicine, MESH: Amino Acid Substitution, Recombinant Proteins, 3. Good health, Computer Science Applications, Flavivirus, MESH: HEK293 Cells, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, MESH: Reunion, MESH: Antigens, Viral, MESH: Cell Line, Tumor, 030106 microbiology, Serogroup, Transfection, Arbovirus, Article, Catalysis, Virus, Inorganic Chemistry, 03 medical and health sciences, MESH: Tanzania, MESH: Protein Stability, Cell Line, Tumor, medicine, Humans, nonstructural protein 1, MESH: Lysine, Physical and Theoretical Chemistry, Epidemics, Molecular Biology, MESH: Epidemics, MESH: Humans, MESH: Transfection, Lysine, Organic Chemistry, MESH: Serogroup, biochemical phenomena, metabolism, and nutrition, Dengue Virus, biology.organism_classification, medicine.disease, Virology, 030104 developmental biology, arbovirus, HEK293 Cells, lcsh:Biology (General), lcsh:QD1-999, chemistry, Amino Acid Substitution, Hepatocytes, MESH: Viral Nonstructural Proteins, recombinant viral protein, Protein Multimerization, Glycoprotein, hepatoma cells, Reunion
Abstract

La Reunion island in the South West Indian Ocean is now endemic for dengue following the introduction of dengue virus serotype 2 (DENV-2) cosmopolitan-I genotype in 2017. DENV-2 infection causes a wide spectrum of clinical manifestations ranging from flu-like disease to severe dengue. The nonstructural glycoprotein 1 (NS1) has been identified as playing a key role in dengue disease severity. The intracellular NS1 exists as a homodimer, whereas a fraction is driven towards the plasma membrane or released as a soluble hexameric protein. Here, we characterized the NS1 glycoproteins from clinical isolates DES-14 and RUN-18 that were collected during the DENV-2 epidemics in Tanzania in 2014 and La Reunion island in 2018, respectively. In relation to hepatotropism of the DENV, expression of recombinant DES-14 NS1 and RUN-18 NS1 glycoproteins was compared in human hepatoma Huh7 cells. We observed that RUN-18 NS1 was poorly stable in Huh7 cells compared to DES-14 NS1. The instability of RUN-18 NS1 leading to a low level of NS1 secretion mostly relates to lysine residues on positions 272 and 324. Our data raise the issue of the consequences of a defect in NS1 stability in human hepatocytes in relation to the major role of NS1 in the pathogenesis of the DENV-2 infection.

Published
2020

10. p27 controls Ragulator and mTOR activity in amino acid-deprived cells to regulate the autophagy–lysosomal pathway and coordinate cell cycle and cell growth

Author

Ada Nowosad, Caroline Callot, Carine Joffre, Arnaud Besson, Stéphane Manenti, Patrice Codogno, Pauline Jeannot, Justine Creff, Renaud T. Perchey, and Centre National de la Recherche Scientifique (CNRS)

Subjects
MESH: Signal Transduction, MESH: Amino Acids, MESH: Cell Line, Tumor, MESH: Starvation, MESH: Cell Cycle, mTORC1, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, 03 medical and health sciences, 0302 clinical medicine, Lysosome, MESH: Cell Proliferation, medicine, MESH: Autophagy, PI3K/AKT/mTOR pathway, MESH: TOR Serine-Threonine Kinases, 030304 developmental biology, 0303 health sciences, MESH: Humans, Cell growth, Chemistry, Autophagy, Cell Biology, Cell cycle, Ragulator complex, Cell biology, MESH: Cell Line, medicine.anatomical_structure, MESH: Proliferating Cell Nuclear Antigen, 030220 oncology & carcinogenesis, MESH: HEK293 Cells, MESH: HeLa Cells, TFEB, biological phenomena, cell phenomena, and immunity, MESH: Lysosomes
Abstract

International audience; Autophagy is a catabolic process whereby cytoplasmic components are degraded within lysosomes, allowing cells to maintain energy homeostasis during nutrient depletion. Several studies reported that the CDK inhibitor p27Kip1 promotes starvation-induced autophagy by an unknown mechanism. Here we find that p27 controls autophagy via an mTORC1-dependent mechanism in amino acid-deprived cells. During prolonged starvation, a fraction of p27 is recruited to lysosomes, where it interacts with LAMTOR1, a component of the Ragulator complex required for mTORC1 activation. Binding of p27 to LAMTOR1 prevents Ragulator assembly and mTORC1 activation, promoting autophagy. Conversely, p27-/- cells exhibit elevated mTORC1 signalling as well as impaired lysosomal activity and autophagy. This is associated with cytoplasmic sequestration of TFEB, preventing induction of the lysosomal genes required for lysosome function. LAMTOR1 silencing or mTOR inhibition restores autophagy and induces apoptosis in p27-/- cells. Together, these results reveal a direct coordinated regulation between the cell cycle and cell growth machineries.

Published
2020

11. Stealth fluorescence labeling for live microscopy imaging of mRNA delivery

Author

Elin K. Esbjörner, Jesper R. Nilsson, Audrey Gallud, Marcel Hollenstein, Cécile Gasse, Emanuele Celauro, Anders Dahlén, Fabienne Levi-Acobas, Tom Baladi, Ivo Sarac, L. Marcus Wilhelmsson, Chalmers University of Technology [Gothenburg, Sweden], AstraZeneca, Génomique métabolique (UMR 8030), Genoscope - Centre national de séquençage [Evry] (GENOSCOPE), Université Paris-Saclay-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université Paris-Saclay-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Université d'Évry-Val-d'Essonne (UEVE)-Centre National de la Recherche Scientifique (CNRS), Chimie bioorganique des acides nucléiques - Bioorganic chemistry of nucleic acids, Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), This work was conducted as part of the FoRmulaEx research center for nucleotide delivery and with associated financial support to E.K.E. and L.M.W. from the Swedish Foundation for Strategic Research (SSF, Grant No. IRC15-0065). F.L.-A., I.S., and M.H. acknowledge funding from Institut Pasteur., Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS)-Université d'Évry-Val-d'Essonne (UEVE), and Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS)

Subjects
MESH: Cytosine, translation, [CHIM.THER]Chemical Sciences/Medicinal Chemistry, fluorescence labeling, Biochemistry, 01 natural sciences, law.invention, Green fluorescent protein, Histones, chemistry.chemical_compound, Colloid and Surface Chemistry, law, MESH: COVID-19, nucleobase triphosphate, MESH: Histones, 0303 health sciences, Molecular Structure, medicine.diagnostic_test, [CHIM.ORGA]Chemical Sciences/Organic chemistry, Chemistry, Electroporation, Translation (biology), Transfection, MESH: Fluorescent Dyes, Molecular Imaging, Cell biology, [CHIM.THEO]Chemical Sciences/Theoretical and/or physical chemistry, MESH: Spectrometry, Fluorescence, Cytosine, MESH: Cell Line, Tumor, mRNA, Green Fluorescent Proteins, MESH: Molecular Structure, 010402 general chemistry, Fluorescence, Article, Catalysis, Flow cytometry, 03 medical and health sciences, MESH: Green Fluorescent Proteins, Confocal microscopy, Cell Line, Tumor, medicine, Humans, [CHIM]Chemical Sciences, RNA, Messenger, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biochemistry [q-bio.BM], [SDV.IB.BIO]Life Sciences [q-bio]/Bioengineering/Biomaterials, Fluorescent Dyes, MESH: RNA, Messenger, 030304 developmental biology, Messenger RNA, MESH: Humans, MESH: Molecular Imaging, MESH: Fluorescence, RNA, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, General Chemistry, COVID-19 Drug Treatment, 0104 chemical sciences, live cell imaging, Spectrometry, Fluorescence, drug delivery, Nucleic acid
Abstract

International audience; Methods for tracking RNA molecules inside living cells without perturbing their natural interactions and functions are critical within biology and, in particular, to facilitate studies of therapeutic RNA delivery. We present a stealth labeling approach that can efficiently, and with high fidelity, generate RNA transcripts of any length, through enzymatic incorporation of the triphosphate of tC O-a fluorescent tricyclic cytosine analogue. We demonstrate this by incorporation of tC O in up to 100% of the natural cytosine positions of a 1.2 kb mRNA encoding for the histone H2B fused to GFP (H2B:GFP). Spectroscopic characterization of this mRNA shows that the incorporation rate of tC O is similar to cytosine, which not only indicates tC O 's excellent nucleotide analogue properties, but also the possibility for efficient labeling and controlled tuning of labeling ratios for different applications. Using live cell confocal microscopy and flow cytometry, we show that the tC Olabeled mRNA is efficiently and correctly translated into H2B:GFP inside human cells. Hence, we not only pioneer the use of fluorescent base analogue labeling of nucleic acids in live-cell microscopy but also, importantly, show that the resulting transcript can be correctly translated. Moreover, the spectral properties of our transcripts and their translation product allow for their straightforward, simultaneous visualization in live cells. Finally, we find that transfected tC O-labeled RNA, unlike the corresponding state-of-the-art fluorescently labeled RNA, translates equally efficient as its natural counterpart, hence representing a methodology for studying natural, unperturbed processing of mRNA used in RNA therapeutics as well as in vaccines, like the ones currently under development against SARS-CoV-2.

Published
2020

12. Retinoic Acid Inducible Gene I and Protein Kinase R, but Not Stress Granules, Mediate the Proinflammatory Response to Yellow Fever Virus

Author

Maxime Chazal, Sarah Lesage, Takashi Fujita, Ségolène Gracias, Eliane F. Meurs, Laura Sinigaglia, Frédéric Tangy, Daniela Bruni, Felix Streicher, Guillaume Beauclair, Salomé Bourgeau, Nolwenn Jouvenet, Département de Virologie - Department of Virology, Institut Pasteur [Paris] (IP), Centre National de la Recherche Scientifique (CNRS), Signalisation antivirale - Virus sensing and signaling, Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), Université Paris Cité (UPCité), Génomique virale et vaccination, Institut de recherche en santé, environnement et travail (Irset), 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 ), Kyoto University, Hépacivirus et Immunité innée, ANR-16-CE15002501, Agence Nationale de la Recherche, ANR-16-CE15-0025,Viro-Storm,Mécanismes de production incontrôlée de cytokines au cours de l'infection virale(2016), Institut Pasteur [Paris], Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), Université de Paris (UP), 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 Kyoto University [Kyoto]

Subjects
Small interfering RNA, MESH: Poly-ADP-Ribose Binding Proteins, viruses, MESH: DNA Helicases, MESH: DEAD Box Protein 58, MESH: Hepatocytes, eIF-2 Kinase, flavivirus, MESH: RNA, Small Interfering, Stress granule assembly, MESH: Animals, RNA, Small Interfering, Receptors, Immunologic, MESH: Carcinoma, Hepatocellular, Poly-ADP-Ribose Binding Proteins, innate immunity, MESH: Haplorhini, 0303 health sciences, MESH: Cytokines, MESH: RNA Helicases, 030302 biochemistry & molecular biology, RNA-Binding Proteins, Haplorhini, liver inflammation, 3. Good health, Cell biology, Virus-Cell Interactions, interferons, RNA Recognition Motif Proteins, MESH: RNA, Viral, Gene Knockdown Techniques, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, MESH: T-Cell Intracellular Antigen-1, Cytokines, DEAD Box Protein 58, RNA, Viral, Yellow fever virus, RNA Helicases, stress granules, MESH: Cell Line, Tumor, Carcinoma, Hepatocellular, Immunology, Biology, Microbiology, Virus, Proinflammatory cytokine, Cell Line, MESH: eIF-2 Kinase, 03 medical and health sciences, Stress granule, Virology, Cell Line, Tumor, Animals, Humans, 030304 developmental biology, MESH: Adaptor Proteins, Signal Transducing, Adaptor Proteins, Signal Transducing, MESH: Humans, MESH: Transcriptome, pattern recognition receptors, DNA Helicases, RNA virus, biology.organism_classification, Protein kinase R, MESH: Gene Knockdown Techniques, MESH: Yellow fever virus, MESH: Cell Line, T-Cell Intracellular Antigen-1, MESH: RNA-Binding Proteins, Insect Science, MESH: RNA Recognition Motif Proteins, Hepatocytes, Cytokine secretion, Transcriptome
Abstract

International audience; Yellow fever virus (YFV) is an RNA virus primarily targeting the liver. Severe YF cases are responsible for hemorrhagic fever, plausibly precipitated by excessive proinflammatory cytokine response. Pathogen recognition receptors (PRRs), such as the cytoplasmic retinoic acid inducible gene I (RIG-I)-like receptors (RLRs), and the viral RNA sensor protein kinase R (PKR), are known to initiate a proinflammatory response upon recognition of viral genomes. Here, we sought to reveal the main determinants responsible for the acute cytokine expression occurring in human hepatocytes following YFV infection. Using a RIG-I-defective human hepatoma cell line, we found that RIG-I largely contributes to cytokine secretion upon YFV infection. In infected RIG-I-proficient hepatoma cells, RIG-I was localized in stress granules. These granules are large aggregates of stalled translation preinitiation complexes known to concentrate RLRs and PKR and are so far recognized as hubs orchestrating RNA virus sensing. Stable knockdown of PKR in hepatoma cells revealed that PKR contributes to both stress granule formation and cytokine induction upon YFV infection. However, stress granule disruption did not affect the cytokine response to YFV infection, as assessed by small interfering RNA (siRNA)-knockdown-mediated inhibition of stress granule assembly. Finally, no viral RNA was detected in stress granules using a fluorescence hybridization approach coupled with immunofluorescence. Our findings suggest that both RIG-I and PKR mediate proinflammatory cytokine induction in YFV-infected hepatocytes, in a stress granule-independent manner. Therefore, by showing the uncoupling of the cytokine response from the stress granule formation, our model challenges the current view in which stress granules are required for the mounting of the acute antiviral response. Yellow fever is a mosquito-borne acute hemorrhagic disease caused by yellow fever virus (YFV). The mechanisms responsible for its pathogenesis remain largely unknown, although increased inflammation has been linked to worsened outcome. YFV targets the liver, where it primarily infects hepatocytes. We found that two RNA-sensing proteins, RIG-I and PKR, participate in the induction of proinflammatory mediators in human hepatocytes infected with YFV. We show that YFV infection promotes the formation of cytoplasmic structures, termed stress granules, in a PKR- but not RIG-I-dependent manner. While stress granules were previously postulated to be essential platforms for immune activation, we found that they are not required for the production of proinflammatory mediators upon YFV infection. Collectively, our work uncovered molecular events triggered by the replication of YFV, which could prove instrumental in clarifying the pathogenesis of the disease, with possible repercussions for disease management.

Published
2020

13. Design, synthesis, and biological evaluation of dual targeting inhibitors of histone deacetylase 6/8 and bromodomain BRPF1

Author

Manfred Jung, Karin Schmidtkunz, Elizabeth R Morales, Stefan Günther, Wolfgang Sippl, Martin Hügle, Frank Erdmann, Patrik Zeyen, Ehab Ghazy, Dina Robaa, Christophe Romier, Matthias Schmidt, Daniel Herp, Martin-Luther-Universität Halle Wittenberg (MLU), Albert-Ludwigs-Universität Freiburg, Institut de Génétique et de Biologie Moléculaire et Cellulaire (IGBMC), Université de Strasbourg (UNISTRA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), and Romier, Christophe

Subjects
Bromodomain, [CHIM.THER]Chemical Sciences/Medicinal Chemistry, MESH: Drug Design, Histone Deacetylase 6, 01 natural sciences, MESH: Structure-Activity Relationship, Drug Discovery, MESH: Molecular Dynamics Simulation, MESH: Histone Deacetylase Inhibitors, 0303 health sciences, biology, Chemistry, Acetylation, General Medicine, Hydroxamic acids, 3. Good health, Cell biology, DNA-Binding Proteins, Molecular Docking Simulation, Leukemia, Myeloid, Acute, Histone, MESH: Repressor Proteins, Epigenetics, MESH: Leukemia, Myeloid, Acute, MESH: Acetylation, MESH: Cell Line, Tumor, [SDV.SP.MED] Life Sciences [q-bio]/Pharmaceutical sciences/Medication, [CHIM.THER] Chemical Sciences/Medicinal Chemistry, Antineoplastic Agents, [SDV.CAN]Life Sciences [q-bio]/Cancer, Molecular Dynamics Simulation, Histone Deacetylases, 03 medical and health sciences, Structure-Activity Relationship, [SDV.SP.MED]Life Sciences [q-bio]/Pharmaceutical sciences/Medication, [SDV.CAN] Life Sciences [q-bio]/Cancer, Cell Line, Tumor, MESH: Molecular Docking Simulation, Humans, Dual targeting inhibitors, 030304 developmental biology, Adaptor Proteins, Signal Transducing, MESH: Adaptor Proteins, Signal Transducing, Pharmacology, Acute myeloid leukemia, MESH: Humans, 010405 organic chemistry, Organic Chemistry, HDAC8, HDAC6, 0104 chemical sciences, Histone Deacetylase Inhibitors, Repressor Proteins, MESH: Histone Deacetylases, Docking (molecular), PHD finger, BRPF1, Drug Design, biology.protein, MESH: Antineoplastic Agents, MESH: Histone Deacetylase 6, MESH: DNA-Binding Proteins
Abstract

International audience; Histone modifying proteins, specifically histone deacetylases (HDACs) and bromodomains, have emerged as novel promising targets for anticancer therapy. In the current work, based on available crystal structures and docking studies, we designed dual inhibitors of both HDAC6/8 and the bromodomain and PHD finger containing protein 1 (BRPF1). Biochemical and biophysical tests showed that compounds 23a,b and 37 are nanomolar inhibitors of both target proteins. Detailed structure-activity relationships were deduced for the synthesized inhibitors which were supported by extensive docking and molecular dynamics studies. Cellular testing in acute myeloid leukemia (AML) cells showed only a weak effect, most probably because of the poor permeability of the inhibitors. We also aimed to analyse the target engagement and the cellular activity of the novel inhibitors by determining the protein acetylation levels in cells by western blotting (tubulin vs histone acetylation), and by assessing their effects on various cancer cell lines.

Published
2020

14. Regulation of ALT-associated homology-directed repair by polyADP-ribosylation

Author

Ian D. Waddell, Robert W. Sobol, Kate M. Smith, Justin L. Roncaioli, Felipe da Veiga Leprevost, Song My Hoang, Dattatreya Mellacharevu, Dominique Ray-Gallet, Michelle L. Lynskey, Roderick J. O’Sullivan, Ragini Bhargava, Jonathan Barroso-González, Nicole Kaminski, Geneviève Almouzni, Anne R. Wondisford, Jianfeng Li, Donald J. Ogilvie, Alexey I. Nesvizhskii, Dominic I. James, Callen T. Wallace, Simon C. Watkins, Laura García-Expósito, University of Pittsburgh School of Medicine, Pennsylvania Commonwealth System of Higher Education (PCSHE), University of Manchester [Manchester], University of Michigan [Ann Arbor], University of Michigan System, University of South Alabama, Dynamique du noyau [Institut Curie], Institut Curie [Paris]-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS), and HAL-SU, Gestionnaire

Subjects
MESH: Signal Transduction, Poly Adenosine Diphosphate Ribose, Cell Cycle Proteins, Histones, Poly ADP Ribosylation, 0302 clinical medicine, Structural Biology, MESH: RNA, Small Interfering, RNA, Small Interfering, MESH: Histones, 0303 health sciences, PARG, MESH: X-linked Nuclear Protein, biology, MESH: Histone Chaperones, DNA, Neoplasm, MESH: Transcription Factors, MESH: Gene Expression Regulation, Neoplastic, Telomere, Chromatin, Cell biology, Gene Expression Regulation, Neoplastic, MESH: Recombinational DNA Repair, Histone, MESH: Poly Adenosine Diphosphate Ribose, MESH: Epithelial Cells, [SDV.BBM.GTP] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], Poly(ADP-ribose) Polymerases, Signal Transduction, G2 Phase, X-linked Nuclear Protein, MESH: Cell Line, Tumor, Glycoside Hydrolases, DNA repair, DNA damage, MESH: DNA, Neoplasm, Article, MESH: Chromatin, MESH: Telomere Homeostasis, Homology directed repair, 03 medical and health sciences, MESH: Cell Cycle Proteins, Cell Line, Tumor, [SDV.BBM.GTP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], MESH: Glycoside Hydrolases, Humans, Histone Chaperones, Molecular Biology, ATRX, 030304 developmental biology, MESH: DNA Damage, MESH: Humans, MESH: Poly ADP Ribosylation, MESH: Poly(ADP-ribose) Polymerases, Recombinational DNA Repair, Telomere Homeostasis, Epithelial Cells, MESH: G2 Phase, MESH: Protein Processing, Post-Translational, MESH: HeLa Cells, biology.protein, MESH: Telomere, Protein Processing, Post-Translational, 030217 neurology & neurosurgery, DNA Damage, HeLa Cells, Transcription Factors
Abstract

International audience; The synthesis of poly(ADP-ribose) (PAR) reconfigures the local chromatin environment and recruits DNA-repair complexes to damaged chromatin. PAR degradation by poly(ADP-ribose) glycohydrolase (PARG) is essential for progression and completion of DNA repair. Here, we show that inhibition of PARG disrupts homology-directed repair (HDR) mechanisms that underpin alternative lengthening of telomeres (ALT). Proteomic analyses uncover a new role for poly(ADP-ribosyl)ation (PARylation) in regulating the chromatin-assembly factor HIRA in ALT cancer cells. We show that HIRA is enriched at telomeres during the G2 phase and is required for histone H3.3 deposition and telomere DNA synthesis. Depletion of HIRA elicits systemic death of ALT cancer cells that is mitigated by re-expression of ATRX, a protein that is frequently inactivated in ALT tumors. We propose that PARylation enables HIRA to fulfill its essential role in the adaptive response to ATRX deficiency that pervades ALT cancers.

Published
2020

15. High Sensitivity of Circulating Tumor Cells Derived from a Colorectal Cancer Patient for Dual Inhibition with AKT and mTOR Inhibitors

Author

Marie-Therese Haider, Laure Cayrefourcq, Nico Hinz, Klaus Pantel, Catherine Alix-Panabières, Daniel J Smit, Manfred Jücker, Universitaetsklinikum Hamburg-Eppendorf = University Medical Center Hamburg-Eppendorf [Hamburg] (UKE), Centre Hospitalier Régional Universitaire [Montpellier] (CHRU Montpellier), Aide à la Décision pour une Médecine Personnalisé - Laboratoire de Biostatistique, Epidémiologie et Recherche Clinique - EA 2415 (AIDMP), Université Montpellier 1 (UM1)-Université de Montpellier (UM), Laboratoire Cellules Circulantes Rares Humaines [CHRU Montpellier] (LCCRH), and Centre Hospitalier Régional Universitaire [Montpellier] (CHRU Montpellier)-CHU Saint-Eloi

Subjects
0301 basic medicine, MAPK/ERK pathway, MESH: Signal Transduction, AKT1, Apoptosis, AKT2, Phosphatidylinositol 3-Kinases, 0302 clinical medicine, Circulating tumor cell, MESH: Protein Kinase Inhibitors, MK2206, Phosphorylation, lcsh:QH301-705.5, MESH: Inhibitory Concentration 50, Chemistry, TOR Serine-Threonine Kinases, drug sensitivity assays, General Medicine, personalized medicine, MESH: Gene Expression Regulation, Neoplastic, Neoplastic Cells, Circulating, targeted therapy, 3. Good health, Gene Expression Regulation, Neoplastic, 030220 oncology & carcinogenesis, MESH: HEK293 Cells, PI3K/AKT/mTOR pathway, Signal transduction, CTCs, Colorectal Neoplasms, Heterocyclic Compounds, 3-Ring, Signal Transduction, MESH: Cell Line, Tumor, Antineoplastic Agents, [SDV.CAN]Life Sciences [q-bio]/Cancer, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, circulating tumor cells, MESH: Everolimus, MESH: Neoplastic Cells, Circulating, Article, MESH: Cell Adhesion, Inhibitory Concentration 50, 03 medical and health sciences, colorectal carcinoma, Cell Line, Tumor, MESH: Cell Proliferation, Cell Adhesion, Humans, Everolimus, Protein Kinase Inhibitors, Protein kinase B, RAD001, MESH: TOR Serine-Threonine Kinases, Cell Proliferation, MESH: Heterocyclic Compounds, 3-Ring, MESH: Humans, MESH: Phosphorylation, Cell growth, MESH: Proto-Oncogene Proteins c-akt, AKT, dual targeting, MESH: Apoptosis, [SDV.MHEP.HEG]Life Sciences [q-bio]/Human health and pathology/Hépatology and Gastroenterology, HEK293 Cells, 030104 developmental biology, lcsh:Biology (General), MESH: Phosphatidylinositol 3-Kinases, Cancer research, MESH: Antineoplastic Agents, Proto-Oncogene Proteins c-akt, MESH: Colorectal Neoplasms
Abstract

International audience; Circulating tumor cells (CTCs) are cells shed from the primary tumor into the bloodstream. While many studies on solid tumor cells exist, data on CTCs are scarce. The mortality of cancer is mostly associated with metastasis and recent research identified CTCs as initiators of metastasis. The PI3K/AKT/mTOR signaling pathway is an intracellular pathway that regulates essential functions including protein biosynthesis, cell growth, cell cycle control, survival and migration. Importantly, activating oncogenic mutations and amplifications in this pathway are frequently observed in a wide variety of cancer entities, underlining the significance of this signaling pathway. In this study, we analyzed the functional role of the PI3K/AKT/mTOR signaling pathway in the CTC-MCC-41 line, derived from a patient with metastatic colorectal cancer. One striking finding in our study was the strong sensitivity of this CTC line against AKT inhibition using MK2206 and mTOR inhibition using RAD001 within the nanomolar range. This suggests that therapies targeting AKT and mTOR could have been beneficial for the patient from which the CTC line was isolated. Additionally, a dual targeting approach of AKT/mTOR inside the PI3K/AKT/mTOR signaling pathway in the colorectal CTCs showed synergistic effects in vitro. Depending on the phenotypical behavior of CTC-MCC-41 in cell culture (adherent vs. suspension), we identified altered phosphorylation levels inside the PI3K/AKT/mTOR pathway. We observed a downregulation of the PI3K/AKT/mTOR signaling pathway, but not of the RAS/RAF/MAPK pathway, in CTCs growing in suspension in comparison to adherent CTCs. Our results highlight distinct functions of AKT isoforms in CTC-MCC-41 cells with respect to cell proliferation. Knockdown of AKT1 and AKT2 leads to significantly impaired proliferation of CTC-MCC-41 cells in vitro. Therefore, our data demonstrate that the PI3K/AKT/mTOR signaling pathway plays a key role in the proliferation of CTC-MCC-41.

Published
2020

16. A Potent Anti-Malarial Human Monoclonal Antibody Targets Circumsporozoite Protein Minor Repeats and Neutralizes Sporozoites in the Liver

Author

Annie S. P. Yang, Carolina Barillas-Mury, Marlon Dillon, Rachel Vistein, Nicole Cavett, David Baker, Arne Schön, Monica W. Gerber, Robert W. Sauerwein, Reid B. Ballard, James O’Connor, Barbara J. Flynn, Alvaro Molina-Cruz, Jorgen Nelson, Fidel Zavala, Lawrence T. Wang, Robert A. Seder, Neville K. Kisalu, Amanda Fabra-García, Joseph R. Francica, Lais Pereira, Rogerio Amino, Bryan T. Mayer, Ian A. Cockburn, Neil P. King, Marie Pancera, Rosemarie D. Mason, Yevel Flores-Garcia, Raphael Gottardo, Azza H. Idris, Nicholas K. Hurlburt, National Institutes of Health [Bethesda] (NIH), Johns Hopkins Bloomberg School of Public Health [Baltimore], Johns Hopkins University (JHU), Australian National University (ANU), Medical School [Australian National University - ANU], Fred Hutchinson Cancer Research Center [Seattle] (FHCRC), Radboud University Medical Center [Nijmegen], University of Washington [Seattle], Infection et Immunité paludéennes - Malaria Infection and Immunity, Institut Pasteur [Paris] (IP), This work was supported by the National Institute of Allergy and Infectious Diseases, the National Cancer Institute (HHSN261200800001E to A.S.), the Dutch Research Council (NWO) talent program veni (VI.Veni.192.171 to A.S.P.Y.), the Bill & Melinda Gates Foundation for N.K.H and M.P. (OPP1156262) and Y.F.-G. and F.Z., who also thank the Bloomberg Philanthropies for continued support., and Institut Pasteur [Paris]

Subjects
Male, 0301 basic medicine, sporozoites, [SDV]Life Sciences [q-bio], Protozoan Proteins, lnfectious Diseases and Global Health Radboud Institute for Molecular Life Sciences [Radboudumc 4], Antibodies, Protozoan, MESH: Sporozoites, Neutralization, MESH: Antibodies, Monoclonal, MESH: Antibodies, Neutralizing, MESH: Hepatocytes, Epitopes, Mice, 0302 clinical medicine, Immunology and Allergy, MESH: Animals, MESH: Protozoan Proteins, MESH: Plasmodium falciparum, MESH: Middle Aged, biology, Antibodies, Monoclonal, Translation (biology), NVDP minor repeats, MESH: Malaria Vaccines, Middle Aged, 3. Good health, Circumsporozoite protein, Infectious Diseases, [SDV.MP]Life Sciences [q-bio]/Microbiology and Parasitology, Liver, MESH: Young Adult, 030220 oncology & carcinogenesis, MESH: HEK293 Cells, malaria vaccines, Female, Antibody, circumsporozoite protein, Adult, Subdominant, MESH: Cell Line, Tumor, MESH: Epitopes, Adolescent, medicine.drug_class, Immunology, Plasmodium falciparum, MESH: Malaria, Monoclonal antibody, Cell Line, Antimalarials, Young Adult, 03 medical and health sciences, All institutes and research themes of the Radboud University Medical Center, MESH: Mice, Inbred C57BL, Cell Line, Tumor, medicine, Animals, Humans, MESH: Antibodies, Protozoan, MESH: Mice, MESH: Adolescent, passive transfer, MESH: Humans, Isothermal titration calorimetry, MESH: Adult, biology.organism_classification, Antibodies, Neutralizing, Virology, MESH: Antimalarials, MESH: Male, Malaria, MESH: Cell Line, Mice, Inbred C57BL, HEK293 Cells, 030104 developmental biology, Hepatocytes, biology.protein, Monoclonal antibodies, MESH: Female, MESH: Liver
Abstract

International audience; Discovering potent human monoclonal antibodies (mAbs) targeting the Plasmodium falciparum circumsporozoite protein (PfCSP) on sporozoites (SPZ) and elucidating their mechanisms of neutralization will facilitate translation for passive prophylaxis and aid next-generation vaccine development. Here, we isolated a neutralizing human mAb, L9 that preferentially bound NVDP minor repeats of PfCSP with high affinity while cross-reacting with NANP major repeats. L9 was more potent than six published neutralizing human PfCSP mAbs at mediating protection against mosquito bite challenge in mice. Isothermal titration calorimetry and multiphoton microscopy showed that L9 and the other most protective mAbs bound PfCSP with two binding events and mediated protection by killing SPZ in the liver and by preventing their egress from sinusoids and traversal of hepatocytes. This study defines the subdominant PfCSP minor repeats as neutralizing epitopes, identifies an in vitro biophysical correlate of SPZ neutralization, and demonstrates that the liver is an important site for antibodies to prevent malaria.

Published
2020

17. Demethylation by low-dose 5-aza-2′-deoxycytidine impairs 3D melanoma invasion partially through miR-199a-3p expression revealing the role of this miR in melanoma

Author

Chantal Etievant, Audrey Delmas, Paola B. Arimondo, Arnaud Pillon, Gilles Favre, Joëlle Riond, Jörg Tost, Arnaud Carrier, Florence Busato, Cécile Desjobert, Antoine Daunay, Diego M. Marzese, Pharmacochimie de la Régulation Epigénétique du Cancer (ETaC), PIERRE FABRE-Centre National de la Recherche Scientifique (CNRS), Centre de biologie du développement (CBD), Centre National de la Recherche Scientifique (CNRS)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées, Centre d'Etude du Polymorphisme Humain (CEPH), Université Paris Diderot - Paris 7 (UPD7)-Institut Universitaire d'Hématologie (IUH), Université Paris Diderot - Paris 7 (UPD7)-Fondation Jean Dausset, Centre National de Génotypage (CNG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Laboratoires Pierre Fabre, Laboratoire d'Epigénétique et d'Environnement [Evry] (CNG - CEA), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de Genotypage-Institut de Génomique [Evry], Centre de Recherches en Cancérologie de Toulouse (CRCT), Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS), PIERRE FABRE-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Université de Toulouse (UT)-Université de Toulouse (UT)-Centre National de la Recherche Scientifique (CNRS)-Centre de Biologie Intégrative (CBI), Université de Toulouse (UT)-Université de Toulouse (UT)-Centre National de la Recherche Scientifique (CNRS)-Centre National de la Recherche Scientifique (CNRS), Université de Toulouse (UT)-Université de Toulouse (UT)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), John Wayne Cancer Institute [Santa Monica, CA, USA], Chimie biologique épigénétique - Epigenetic Chemical Biology (EpiCBio), Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS), This work was supported to P.B.A. by Centre National de la Recherche Scientifique (CNRS) [ATIP], Région Midi Pyrenées [Equipe d’Excellence and FEDER CNRS/Région Midi Pyrenées], Fondation InNaBioSanté, and PlanCancer2014 (N°EPIG201401)., Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre de Biologie Intégrative (CBI), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS)-Centre National de la Recherche Scientifique (CNRS), Fondation Jean Dausset-Institut Universitaire d'Hématologie (IUH), Université Paris Diderot - Paris 7 (UPD7)-Université Paris Diderot - Paris 7 (UPD7), Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS)-Centre de Biologie Intégrative (CBI), and Riond, Joelle

Subjects
0301 basic medicine, Lung Neoplasms, [SDV]Life Sciences [q-bio], lcsh:Medicine, MESH: Spheroids, Cellular, Transcriptome, Mice, chemistry.chemical_compound, 0302 clinical medicine, MESH: DNA Methylation, MESH: Up-Regulation, MESH: Sequence Analysis, RNA, Gene Regulatory Networks, MESH: Animals, Promoter Regions, Genetic, Melanoma, Genetics (clinical), MESH: Gene Regulatory Networks, DNA methylation, microRNA, MESH: Gene Expression Regulation, Neoplastic, Up-Regulation, 3. Good health, Gene Expression Regulation, Neoplastic, [SDV] Life Sciences [q-bio], MESH: Cell Survival, 030220 oncology & carcinogenesis, Female, Deoxycytidine, Epigenetics, MESH: Cell Line, Tumor, lcsh:QH426-470, Cell Survival, MESH: Melanoma, education, Biology, Decitabine, 03 medical and health sciences, MESH: Gene Expression Profiling, In vivo, Cell Line, Tumor, Spheroids, Cellular, MESH: Promoter Regions, Genetic, Genetics, medicine, Animals, Humans, Neoplasm Invasiveness, Molecular Biology, MESH: Mice, neoplasms, MESH: Humans, MESH: Decitabine, Sequence Analysis, RNA, Research, Gene Expression Profiling, lcsh:R, MESH: Neoplasm Invasiveness, medicine.disease, Demethylating agent, MESH: Lung Neoplasms, lcsh:Genetics, MicroRNAs, 030104 developmental biology, chemistry, Cancer research, Melanoma aggressiveness, MESH: Female, MESH: MicroRNAs, Neoplasm Transplantation, MESH: Neoplasm Transplantation, Developmental Biology
Abstract

Background Efficient treatments against metastatic melanoma dissemination are still lacking. Here, we report that low-cytotoxic concentrations of 5-aza-2′-deoxycytidine, a DNA demethylating agent, prevent in vitro 3D invasiveness of metastatic melanoma cells and reduce lung metastasis formation in vivo. Results We unravelled that this beneficial effect is in part due to MIR-199A2 re-expression by promoter demethylation. Alone, this miR showed an anti-invasive and anti-metastatic effect. Throughout integration of micro-RNA target prediction databases with transcriptomic analysis after 5-aza-2′-deoxycytidine treatments, we found that miR-199a-3p downregulates set of genes significantly involved in invasion/migration processes. In addition, analysis of data from melanoma patients showed a stage- and tissue type-dependent modulation of MIR-199A2 expression by DNA methylation. Conclusions Thus, our data suggest that epigenetic- and/or miR-based therapeutic strategies can be relevant to limit metastatic dissemination of melanoma. Electronic supplementary material The online version of this article (10.1186/s13148-018-0600-2) contains supplementary material, which is available to authorized users.

Published
2019

18. GLUT1 protects prostate cancer cells from glucose deprivation-induced oxidative stress

Author

Rosa M. Sainz, Sandrina Kinet, Pedro Gonzalez-Menendez, Ivan Gonzalez-Pola, Juan C. Mayo, Aida Rodriguez-Garcia, Alejandro Alvarez-Artime, David Hevia, Naomi Taylor, Rebeca Alonso-Arias, Institut de Génétique Moléculaire de Montpellier (IGMM), and Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM)

Subjects
Male, 0301 basic medicine, [SDV]Life Sciences [q-bio], Glucose uptake, Clinical Biochemistry, urologic and male genital diseases, medicine.disease_cause, Biochemistry, lcsh:QH301-705.5, MESH: Superoxide Dismutase, MESH: Glutathione, lcsh:R5-920, Glucose Transporter Type 1, Prostate cancer, MESH: Oxidative Stress, biology, Chemistry, Prostate, MESH: Gene Expression Regulation, Neoplastic, Glutathione, Gene Expression Regulation, Neoplastic, MESH: Glucose, Prostatic Neoplasms, Castration-Resistant, Androgen receptor, Receptors, Androgen, Glucose deprivation, MESH: Hydrogen Peroxide, MESH: Receptors, Androgen, lcsh:Medicine (General), medicine.medical_specialty, Programmed cell death, MESH: Cell Line, Tumor, MESH: Prostate, 03 medical and health sciences, Downregulation and upregulation, Cell Line, Tumor, MESH: Cell Proliferation, Internal medicine, LNCaP, medicine, Humans, Cell Proliferation, MESH: Humans, Superoxide Dismutase, Organic Chemistry, Glucose transporter, Hydrogen Peroxide, Glut1, MESH: Male, Glucose, 030104 developmental biology, Endocrinology, lcsh:Biology (General), Oxidative stress, MESH: Prostatic Neoplasms, Castration-Resistant, Cancer cell, biology.protein, GLUT1, MESH: Glucose Transporter Type 1
Abstract

Glucose, chief metabolic support for cancer cell survival and growth, is mainly imported into cells by facilitated glucose transporters (GLUTs). The increase in glucose uptake along with tumor progression is due to an increment of facilitative glucose transporters as GLUT1. GLUT1 prevents cell death of cancer cells caused by growth factors deprivation, but there is scarce information about its role on the damage caused by glucose deprivation, which usually occurs within the core of a growing tumor. In prostate cancer (PCa), GLUT1 is found in the most aggressive tumors, and it is regulated by androgens. To study the response of androgen-sensitive and insensitive PCa cells to glucose deprivation and the role of GLUT1 on survival mechanisms, androgen-sensitive LNCaP and castration-resistant LNCaP-R cells were employed. Results demonstrated that glucose deprivation induced a necrotic type of cell death which is prevented by antioxidants. Androgen-sensitive cells show a higher resistance to cell death triggered by glucose deprivation than castration-resistant cells. Glucose removal causes an increment of H2O2, an activation of androgen receptor (AR) and a stimulation of AMP-activated protein kinase activity. In addition, glucose removal increases GLUT1 production in androgen sensitive PCa cells. GLUT1 ectopic overexpression makes PCa cells more resistant to glucose deprivation and oxidative stress-induced cell death. Under glucose deprivation, GLUT1 overexpressing PCa cells sustains mitochondrial SOD2 activity, compromised after glucose removal, and significantly increases reduced glutathione (GSH). In conclusion, androgen-sensitive PCa cells are more resistant to glucose deprivation-induced cell death by a GLUT1 upregulation through an enhancement of reduced glutathione levels. Keywords: Glut1, Prostate cancer, Glucose deprivation, Androgen receptor, Glutathione, Oxidative stress

Published
2018

19. Repairing folding-defective α-sarcoglycan mutants by CFTR correctors, a potential therapy for limb-girdle muscular dystrophy 2D

Author

Isabelle Richard, Chiara Fecchio, Roberta Sacchetto, Marcello Carotti, Elisa Bianchini, Michela Soardi, Sara F Henriques, Chiara Gomiero, Justine Marsolier, Dorianna Sandonà, Romeo Betto, Université Paris-Saclay, Universita degli Studi di Padova, CNR Institute of Neuroscience, National Research Council [Italy] (CNR), Approches génétiques intégrées et nouvelles thérapies pour les maladies rares (INTEGRARE), École pratique des hautes études (EPHE)-Université d'Évry-Val-d'Essonne (UEVE)-GENETHON 3-Institut National de la Santé et de la Recherche Médicale (INSERM), École pratique des hautes études (EPHE), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Université d'Évry-Val-d'Essonne (UEVE)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Généthon, Università degli Studi di Padova = University of Padua (Unipd), and École Pratique des Hautes Études (EPHE)

Subjects
0301 basic medicine, [SDV.BIO]Life Sciences [q-bio]/Biotechnology, MESH: Muscle Contraction, Cystic Fibrosis Transmembrane Conductance Regulator, MESH: Proof of Concept Study, Missense mutation, Muscular dystrophy, Genetics (clinical), MESH: Cystic Fibrosis Transmembrane Conductance Regulator, MESH: Muscle, Skeletal, biology, Myogenesis, MESH: Sarcoglycanopathies, Articles, General Medicine, musculoskeletal system, 3. Good health, Cell biology, small molecules, Sarcoglycan, MESH: HEK293 Cells, Dystrophin, MESH: Muscle, Striated, Muscle Contraction, musculoskeletal diseases, congenital, hereditary, and neonatal diseases and abnormalities, MESH: Cell Line, Tumor, CFTR corrector, Mutation, Missense, Proof of Concept Study, 03 medical and health sciences, pharmacological treatment, Cell Line, Tumor, Sarcoglycans, sarcoglycan, Sarcoglycanopathies, Genetics, medicine, Humans, MESH: Sarcoglycans, Muscle, Skeletal, Molecular Biology, SGCA, MESH: Mutation, Missense, MESH: Humans, Sarcolemma, Cell Membrane, sarcoglycan,LGMD2D,CFTR corrector,myogenic cells,myotubes,small molecules,pharmacological treatment, Dystrophy, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, medicine.disease, Muscle, Striated, HEK293 Cells, 030104 developmental biology, myotubes, [SDV.GEN.GH]Life Sciences [q-bio]/Genetics/Human genetics, biology.protein, LGMD2D, myogenic cells, MESH: Cell Membrane
Abstract

International audience; Limb-girdle muscular dystrophy type 2D (LGMD2D) is a rare autosomal-recessive disease, affecting striated muscle, due to mutation of SGCA, the gene coding for α-sarcoglycan. Nowadays, more than 50 different SGCA missense mutations have been reported. They are supposed to impact folding and trafficking of α-sarcoglycan because the defective polypeptide, although potentially functional, is recognized and disposed of by the quality control of the cell. The secondary reduction of α-sarcoglycan partners, β-, γ- and δ-sarcoglycan, disrupts a key membrane complex that, associated to dystrophin, contributes to assure sarcolemma stability during muscle contraction. The complex deficiency is responsible for muscle wasting and the development of a severe form of dystrophy. Here, we show that the application of small molecules developed to rescue ΔF508-CFTR trafficking, and known as CFTR correctors, also improved the maturation of several α-sarcoglycan mutants that were consequently rescued at the plasma membrane. Remarkably, in myotubes from a patient with LGMD2D, treatment with CFTR correctors induced the proper re-localization of the whole sarcoglycan complex, with a consequent reduction of sarcolemma fragility. Although the mechanism of action of CFTR correctors on defective α-sarcoglycan needs further investigation, this is the first report showing a quantitative and functional recovery of the sarcoglycan-complex in human pathologic samples, upon small molecule treatment. It represents the proof of principle of a pharmacological strategy that acts on the sarcoglycan maturation process and we believe it has a great potential to develop as a cure for most of the patients with LGMD2D.

Published
2018

20. Pharmacological Investigation of CC-LAAO, an L-Amino Acid Oxidase from Cerastes cerastes Snake Venom

Author

Zaineb Abdelkafi-Koubaa, Ines ELBini-Dhouib, Soumaya Souid, Jed Jebali, Raoudha Doghri, Najet Srairi-Abid, Khadija Essafi-Benkhadir, Olivier Micheau, Naziha Marrakchi, Institut Pasteur de Tunis, Réseau International des Instituts Pasteur (RIIP), Université de Tunis El Manar (UTM), University of Louisiana, Salah Azaiez Institute, Lipides - Nutrition - Cancer [Dijon - U1231] (LNC), Université de Bourgogne (UB)-Institut National de la Santé et de la Recherche Médicale (INSERM)-AgroSup Dijon - Institut National Supérieur des Sciences Agronomiques, de l'Alimentation et de l'Environnement, The authors are grateful to the Ministry of Higher Education and Scientific Research of Tunisia for financial support (LR20IPT01)., Micheau, Olivier, Laboratoire d'Epidémiologie et d'Ecologie parasitaire, Institut Pasteur de Tunis, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP), and Laboratoire d'Epidémiologie Médicale, Institut Pasteur de Tunis

Subjects
MESH: L-Lactate Dehydrogenase, MESH: Cell Line, Tumor, glioblastoma cells, Health, Toxicology and Mutagenesis, MESH: Viperidae, MESH: Creatinine, Toxicology, 03 medical and health sciences, SV-LAAOs, envenomation, toxicity, apoptosis, MESH: L-Amino Acid Oxidase, [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, MESH: Animals, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, MESH: Mice, 030304 developmental biology, 0303 health sciences, MESH: Humans, 030302 biochemistry & molecular biology, MESH: Chick Embryo, MESH: Male, 3. Good health, MESH: Edema, MESH: Cell Survival, MESH: Alanine Transaminase, Medicine, MESH: Viper Venoms, MESH: Hemorrhage
Abstract

Snake venom proteins, which are responsible for deadly snakebite envenomation, induce severe injuries including neurotoxicity, myotoxicity, cardiotoxicity, hemorrhage, and the disruption of blood homeostasis. Yet, many snake-venom proteins have been developed as potential drugs for treating human diseases due to their pharmacological effects. In this study, we evaluated the use of, an L-amino acid oxidase isolated from Cerastes cerastes snake venom CC-LAAO, as a potential anti-glioblastoma drug, by investigating its in vivo and in vitro pharmacological effects. Our results showed that acute exposure to CC-LAAO at 1 and 2.5 µg/mL does not induce significant toxicity on vital organs, as indicated by the murine blood parameters including aspartate transaminase (AST), alanine transaminase (ALT), lactate dehydrogenase (LDH) activities, and creatinine levels. The histopathological examination demonstrated that only at high concentrations did CC-LAAO induce inflammation and necrosis in several organs of the test subjects. Interestingly, when tested on human glioblastoma U87 cells, CC-LAAO induced a dose-dependent apoptotic effect through the H2O2 generated during the enzymatic reaction. Taken altogether, our data indicated that low concentration of CC-LAAO may be safe and may have potential in the development of anti-glioblastoma agents.

Published
2021

21. Optimization of Advanced Live-Cell Imaging through Red/Near-Infrared Dye Labeling and Fluorescence Lifetime-Based Strategies

Author

Komuro, Yutaro, Galas, Ludovic, Morozov, Yury, Fahrion, Jennifer, Raoult, Emilie, Lebon, Alexis, Tilot, Amanda, Kikuchi, Shin, Ohno, Nobuhiko, Vaudry, David, Rakic, Pasko, Komuro, Hitoshi, Plate-Forme de Recherche en Imagerie Cellulaire de Haute-Normandie (PRIMACEN), Université de Rouen Normandie (UNIROUEN), Normandie Université (NU)-Normandie Université (NU)-Institute for Research and Innovation in Biomedicine (IRIB), Normandie Université (NU)-Normandie Université (NU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université de Rouen Normandie (UNIROUEN), Normandie Université (NU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-High-tech Research Infrastructures for Life Sciences (HeRacLeS), Normandie Université (NU)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Institut National de la Santé et de la Recherche Médicale (INSERM), Département de Pathologie [CHU Caen], Université de Caen Normandie (UNICAEN), Normandie Université (NU)-Normandie Université (NU)-CHU Caen, Normandie Université (NU)-Tumorothèque de Caen Basse-Normandie (TCBN)-Tumorothèque de Caen Basse-Normandie (TCBN), Imagerie et Stratégies Thérapeutiques des pathologies Cérébrales et Tumorales (ISTCT), Normandie Université (NU)-Normandie Université (NU)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), Lerner Research Institute [Cleveland, OH, USA], Cleveland Clinic, Lebon, Alexis, Department of Neurosciences [Lerner Research Institute, Cleveland Clinic], Cleveland Clinic-Cleveland Clinic, Department of Neuroscience, Yale University School of Medicine, Yale School of Medicine [New Haven, Connecticut] (YSM), Department of Genomic Medicine [Lerner Research Institute, Cleveland Clinic], Division of Neurobiology and Bioinformatics, National Institute for Physiological Sciences, Aichi 444-8787, Japan, National Institute for Physiological Sciences, School of Medicine [Jichi Medical University, Japan], Jichi Medical University [Tochigi-Ken, Japan], Différenciation et communication neuronale et neuroendocrine (DC2N), Normandie Université (NU)-Normandie Université (NU)-Institut National de la Santé et de la Recherche Médicale (INSERM), Kavli Institute for Neuroscience, Yale School of Medicine, and Yale School of Medicine [New Haven, Connecticut] (YSM)-Yale School of Medicine [New Haven, Connecticut] (YSM)

Subjects
MESH: Photons, [SDV]Life Sciences [q-bio], MESH: Microscopy, Fluorescence, live-cell imaging, law.invention, law, Microscopy, Image Processing, Computer-Assisted, MESH: Microscopy, Confocal, Stimulated emission, Biology (General), MESH: Fluorescein, Spectroscopy, Microscopy, Confocal, STED microscopy, Equipment Design, General Medicine, MESH: Fluorescent Dyes, MESH: Image Processing, Computer-Assisted, Fluorescence, Computer Science Applications, Numerical aperture, STED, [SDV] Life Sciences [q-bio], Chemistry, confocal, Optoelectronics, Fluorescein, MESH: Infrared Rays, FLIM, Photomultiplier, MESH: Cell Line, Tumor, Materials science, QH301-705.5, Infrared Rays, τ separation, MESH: Rhodamines, Article, Catalysis, Inorganic Chemistry, Confocal microscopy, Cell Line, Tumor, fluorescence lifetime, Humans, Physical and Theoretical Chemistry, QD1-999, Molecular Biology, Fluorescent Dyes, Photons, MESH: Humans, Rhodamines, business.industry, MESH: Fluorescence, Organic Chemistry, FLIM-STED, Photon counting, Microscopy, Fluorescence, business, sample preservation, MESH: Equipment Design
Abstract

Fluorescence microscopy is essential for a detailed understanding of cellular processes, however, live-cell preservation during imaging is a matter of debate. In this study, we proposed a guide to optimize advanced light microscopy approaches by reducing light exposure through fluorescence lifetime (τ) exploitation of red/near-infrared dyes. Firstly, we characterized key instrumental elements which revealed that red/near-infrared laser lines with an 86x (Numerical Aperture (NA) = 1.2, water immersion) objective allowed high transmission of fluorescence signals, low irradiance and super-resolution. As a combination of two technologies, i.e., vacuum tubes (e.g., photomultiplier) and semiconductor microelectronics (e.g., avalanche photodiode), type S, X and R of hybrid detectors (HyD-S, HyD-X and HyD-R) were particularly adapted for red/near-infrared photon counting and τ separation. Secondly, we tested and compared lifetime-based imaging including coarse τ separation for confocal microscopy, fitting and phasor plot analysis for fluorescence lifetime microscopy (FLIM), and lifetimes weighting for enhanced stimulated emission depletion (STED) nanoscopy, in light of red/near-infrared multiplexing. Mainly, we showed that the choice of appropriate imaging approach may depend on fluorochrome number, together with their spectral/lifetime characteristics and STED compatibility. Photon-counting mode and sensitivity of HyDs together with phasor plot analysis of fluorescence lifetimes enabled the flexible and fast imaging of multi-labeled living H28 cells. Therefore, a combination of red/near-infrared dyes labeling with lifetime-based strategies offers new perspectives for live-cell imaging by enhancing sample preservation through acquisition time and light exposure reduction.

Published
2021

22. STAT2 is an essential adaptor in USP18-mediated suppression of type I interferon signaling

Author

Frédéric Colland, Kei-ichiro Arimoto, Ming Yan, Dong-Er Zhang, Sandra Pellegrini, Stephan Wilmes, Yue Zhang, Jacob Piehler, Samuel A Stoner, Sayuri Miyauchi, Jürgen J. Heinisch, Sara Löchte, Christoph Burkart, Zhi Li, Jun-Bao Fan, University of California [San Diego] (UC San Diego), University of California, Fachhochschule Osnabrück, Signalisation des Cytokines - Cytokine Signaling, Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), Hybrigenics [Paris], Hybrigenics, This study was supported by NIH R01HL091549 and R01CA177305 to D.-E.Z. and SFB 944 from the DFG to J.P. and J.J.H., University of California (UC), Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), and Li, Zhi

Subjects
0301 basic medicine, MESH: Signal Transduction, MESH: Interferon Type I, MESH: Feedback, Physiological, Receptor, Interferon alpha-beta, Medical and Health Sciences, Interferon alpha-beta, MESH: Mutant Proteins, Mice, 0302 clinical medicine, Structural Biology, Interferon, MESH: STAT2 Transcription Factor, MESH: Animals, STAT2, Receptor, MESH: Endopeptidases, Feedback, Physiological, Tumor, biology, MESH: Immunoblotting, Effector, Biological Sciences, 3. Good health, Cell biology, 030220 oncology & carcinogenesis, Interferon Type I, MESH: Protein Domains, Signal transduction, Ubiquitin Thiolesterase, medicine.drug, Protein Binding, Signal Transduction, Cell signaling, MESH: Cell Line, Tumor, Physiological, Immunoblotting, Biophysics, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, MESH: Two-Hybrid System Techniques, Article, Cell Line, Feedback, 03 medical and health sciences, Immune system, Protein Domains, Cell Line, Tumor, Two-Hybrid System Techniques, Endopeptidases, [SDV.BBM] Life Sciences [q-bio]/Biochemistry, Molecular Biology, medicine, MESH: Protein Binding, MESH: Receptor, Interferon alpha-beta, Animals, Humans, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, [SDV.BC] Life Sciences [q-bio]/Cellular Biology, Molecular Biology, MESH: Mice, MESH: Humans, STAT2 Transcription Factor, 030104 developmental biology, Chemical Sciences, Cancer research, biology.protein, Mutant Proteins, Interferon type I, Developmental Biology
Abstract

International audience; Type I interferons (IFNs) are multifunctional cytokines that regulate immune responses and cellular functions but also can have detrimental effects on human health. A tight regulatory network therefore controls IFN signaling, which in turn may interfere with medical interventions. The JAK-STAT signaling pathway transmits the IFN extracellular signal to the nucleus, thus resulting in alterations in gene expression. STAT2 is a well-known essential and specific positive effector of type I IFN signaling. Here, we report that STAT2 is also a previously unrecognized, crucial component of the USP18-mediated negative-feedback control in both human and mouse cells. We found that STAT2 recruits USP18 to the type I IFN receptor subunit IFNAR2 via its constitutive membrane-distal STAT2-binding site. This mechanistic coupling of effector and negative-feedback functions of STAT2 may provide novel strategies for treatment of IFN-signaling-related human diseases.

Published
2017

23. Targetting αvβ3 and α5β1 integrins with Ecballium elaterium (L.) A. Rich. seed oil

Author

Imen Touihri-Barakati, Belgacem Hanchi, José Luis, Abdennacer Boulila, Olfa Kallech-Ziri, Karim Hosni, Naziha Marrakchi, Khaoula Khwaldia, Laboratoire des Substances Naturelles (LR02INRAP10), Institut National de Recherche et d'Analyse Physico-chimique (INRAP-Tunisie), Faculté de Médecine de Tunis, Université Tunis El Manar (UTM), Laboratoire des Venins et Biomolécules Thérapeutiques - Laboratory of Venoms and Therapeutic Biomolecules (LR11IPT08), Institut Pasteur de Tunis, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP), Faculté des Sciences de Tunis, Université de Tunis [Tunis], Centre de Recherches en Oncologie biologique et Oncopharmacologie (CRO2), Aix Marseille Université (AMU)- Hôpital de la Timone [CHU - APHM] (TIMONE)-Institut National de la Santé et de la Recherche Médicale (INSERM), and This research was supported by Ministry of High Education and Scientific Research of Tunisia (MERST) and the Institut National de la Santé et de la Recherche Médicale (INSERM) and by grants from INCa (Institut National du Cancer) and ARCUS (Action en Région de Coopération Universitaire et Scientifique).

Subjects
0301 basic medicine, MESH: Plant Oils/pharmacology, Angiogenesis, [SDV]Life Sciences [q-bio], MESH: Cucurbitaceae/chemistry, Pharmacology, MESH: Glioma/pathology, MESH: Cell Movement/drug effects, Glioma cell line, MESH: Cell Proliferation/drug effects, 0302 clinical medicine, MESH: Glioma/drug therapy, Cell Movement, MESH: Brain Neoplasms/blood supply, Neovascularization, Pathologic, biology, Brain Neoplasms, Integrin beta3, Glioma, General Medicine, 3. Good health, Biochemistry, 030220 oncology & carcinogenesis, Seeds, Adhesion, αvβ3 and α5β1 integrins, Seed oil, Integrin alpha5beta1, MESH: Cell Line, Tumor, Integrin, Time-Lapse Imaging, 03 medical and health sciences, MESH: Cell Adhesion/drug effects, MESH: Neovascularization, Pathologic/drug therapy, Cell Line, Tumor, Cell Adhesion, Humans, Plant Oils, MESH: Brain Neoplasms/drug therapy, MESH: Seeds/chemistry, MESH: Time-Lapse Imaging, MESH: Integrin beta3/metabolism, Cell adhesion, Cell Proliferation, MESH: Humans, MESH: Integrin alpha5beta1/metabolism, MESH: Brain Neoplasms/pathology, Cell growth, Elaterium, Ecballium elaterium, MESH: Plant Oils/therapeutic use, biology.organism_classification, MESH: Glioma/blood supply, Fibronectin, Cucurbitaceae, 030104 developmental biology, Cell culture, biology.protein
Abstract

International audience; In the present study, the effect of Ecbalium elaterium seed oil on adhesion, migration and proliferation of human brain cancer cell line (U87) was determined. Treatment of U87 cell line with the seed oil resulted in strong inhibition of their adhesion to fibrinogen (Fg), fibronectin (Fn). It also reduced their migration and proliferation in a dose-dependent manner without being cytotoxic. Concomitantly, by using Matrigel™ assays, the oil significantly inhibited angiogenesis. The anti- tumor effect of the oil is specifically mediated by αvβ3 and α5β1 integrins. The presence of integrin antagonists in seed oil from E. elaterium could be used for the development of anticancer drugs with targeted "multi-modal" therapies combining anti-adhesif, antiproliferative, antimetastasic and anti-angiogenic, approaches.

Published
2016

24. Integrative proteomic and phosphoproteomic profiling of prostate cell lines

Author

Maria Katsogiannou, Jean Baptiste Boyer, Luc Camoin, Laurence Calzone, Anaïs Baudot, Stéphane Audebert, Elisabeth Remy, Palma Rocchi, Alberto Valdeolivas, Centre de Recherche en Cancérologie de Marseille (CRCM), Aix Marseille Université (AMU)-Institut Paoli-Calmettes, Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Theories and Approaches of Genomic Complexity (TAGC), Aix Marseille Université (AMU)-Institut National de la Santé et de la Recherche Médicale (INSERM), Institut Mathématiques de Luminy (IML), Université de la Méditerranée - Aix-Marseille 2-Centre National de la Recherche Scientifique (CNRS), Centre de recherche de l'Institut Curie [Paris], Institut Curie [Paris], Marseille medical genetics - Centre de génétique médicale de Marseille (MMG), Institut de Mathématiques de Marseille (I2M), Centre National de la Recherche Scientifique (CNRS)-École Centrale de Marseille (ECM)-Aix Marseille Université (AMU), Hôpital Saint-Joseph [Marseille], Aix Marseille Université (AMU)-École Centrale de Marseille (ECM)-Centre National de la Recherche Scientifique (CNRS), ProGeLife [Marseille], Mines Paris - PSL (École nationale supérieure des mines de Paris), Université Paris sciences et lettres (PSL), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Paoli-Calmettes, Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Aix Marseille Université (AMU), and Rocchi, Palma

Subjects
0301 basic medicine, Proteomics, Male, MESH: Neoplasm Proteins, Proteomes, [SDV]Life Sciences [q-bio], Cell Membranes, Proteomic databases, Biochemistry, Mass Spectrometry, Androgen deprivation therapy, Prostate cancer, Database and Informatics Methods, 0302 clinical medicine, Stable isotope labeling by amino acids in cell culture, Membrane proteins, Transcriptional regulation, Medicine and Health Sciences, Post-Translational Modification, Phosphorylation, ComputingMilieux_MISCELLANEOUS, DU145 cells, 0303 health sciences, Multidisciplinary, MESH: Proteomics, Phosphoproteomics, Prostate Diseases, Prostate, MESH: Gene Expression Regulation, Neoplastic, 3. Good health, Neoplasm Proteins, Gene Expression Regulation, Neoplastic, [SDV] Life Sciences [q-bio], Oncology, 030220 oncology & carcinogenesis, Proteome, Cell lines, Medicine, Cellular Structures and Organelles, Biological cultures, Research Article, MESH: Cell Line, Tumor, Urology, Science, [SDV.CAN]Life Sciences [q-bio]/Cancer, Computational biology, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Biology, MESH: Phosphoproteins, MESH: Prostate, 03 medical and health sciences, DU145, Cell Line, Tumor, LNCaP, medicine, Gene Expression and Vector Techniques, Humans, Molecular Biology Techniques, Molecular Biology, 030304 developmental biology, MESH: Mass Spectrometry, Molecular Biology Assays and Analysis Techniques, MESH: Humans, Biology and Life Sciences, Proteins, Cancers and Neoplasms, Prostatic Neoplasms, Cell Biology, medicine.disease, Phosphoproteins, MESH: Male, Research and analysis methods, Genitourinary Tract Tumors, 030104 developmental biology, Biological Databases, MESH: Prostatic Neoplasms, MESH: Biomarkers, Protein expression, Biomarkers
Abstract

BackgroundProstate cancer is a major public health issue, mainly because patients relapse after androgen deprivation therapy. Proteomic strategies, aiming to reflect the functional activity of cells, are nowadays among the leading approaches to tackle the challenges not only of better diagnosis, but also of unraveling mechanistic details related to disease etiology and progression.MethodsWe conducted here a large SILAC-based Mass Spectrometry experiment to map the proteomes and phosphoproteomes of four widely used prostate cell lines, namely PNT1A, LNCaP, DU145 and PC3, representative of different cancerous and hormonal status.ResultsWe identified more than 3000 proteins and phosphosites, from which we quantified more than 1000 proteins and 500 phosphosites after stringent filtering. Extensive exploration of this proteomics and phosphoproteomics dataset allowed characterizing housekeeping as well as cell-line specific proteins, phosphosites and functional features of each cell line. In addition, by comparing the sensitive and resistant cell lines, we identified protein and phosphosites differentially expressed in the resistance context. Further data integration in a molecular network highlighted the differentially expressed pathways, in particular migration and invasion, RNA splicing, DNA damage repair response and transcription regulation.ConclusionsOverall, this study proposes a valuable resource toward the characterization of proteome and phosphoproteome of four widely used prostate cell lines and reveals candidates to be involved in prostate cancer progression for further experimental validation.

Published
2019

25. Hormonal and spatial control of SUMOylation in the human and mouse adrenal cortex

Author

T. Dumontet, Antoine Martinez, Bruno Ragazzon, Anne-Marie Lefrançois-Martinez, Jean-Christophe Pointud, Nathanaëlle Montanier, Jérôme Bertherat, Igor Tauveron, Florence Roucher-Boulez, Isabelle Sahut-Barnola, Annabel Berthon, Damien Dufour, Pierre Val, Cyril Djari, M. Batisse-Lignier, Institut Cochin (IC UM3 (UMR 8104 / U1016)), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Descartes - Paris 5 (UPD5)-Centre National de la Recherche Scientifique (CNRS), Interactions génétiques et cellulaires au cours de la différenciation, Université d'Auvergne - Clermont-Ferrand I (UdA)-Institut National de la Santé et de la Recherche Médicale (INSERM), Génétique, Reproduction et Développement (GReD), Centre National de la Recherche Scientifique (CNRS)-Université Clermont Auvergne [2017-2020] (UCA [2017-2020])-Institut National de la Santé et de la Recherche Médicale (INSERM), Service d’endocrinologie diabétologie et maladies métaboliques, CHU Clermont-Ferrand, CHU Lyon, Unité de Nutrition Humaine (UNH), Institut National de la Recherche Agronomique (INRA)-Université Clermont Auvergne [2017-2020] (UCA [2017-2020]), Génétique, Reproduction et Développement (GReD ), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Clermont Auvergne [2017-2020] (UCA [2017-2020]), Service d’Endocrinologie Diabète et Maladies Métaboliques [CHU Clermont-Ferrand], Pôle RHEUNNIRS [CHU Clermont-Ferrand], CHU Gabriel Montpied [Clermont-Ferrand], CHU Clermont-Ferrand-CHU Clermont-Ferrand-CHU Gabriel Montpied [Clermont-Ferrand], CHU Clermont-Ferrand-CHU Clermont-Ferrand, Centre National de la Recherche Scientifique (CNRS)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM), CHU Clermont-Ferrand, Centre de référence des maladies rares de la surrénale ( CHU Cochin [AP-HP]), Hôpital Cochin [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Génétique, Reproduction et Développement - Clermont Auvergne (GReD), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Clermont Auvergne (UCA)-Centre National de la Recherche Scientifique (CNRS), Institut National de la Recherche Agronomique (INRA)-Université d'Auvergne - Clermont-Ferrand I (UdA)-Clermont Université, Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Clermont Auvergne [2017-2020] (UCA [2017-2020])-Centre National de la Recherche Scientifique (CNRS), ANR-16-IDEX-0001,CAP 20-25,CAP 20-25(2016), and Clermont Université-Université d'Auvergne - Clermont-Ferrand I (UdA)-Institut National de la Recherche Agronomique (INRA)

Subjects
0301 basic medicine, MESH: Neoplasm Proteins, MESH: Signal Transduction, MESH: beta Catenin, SUMO protein, MESH: Adrenal Cortex Neoplasms, Biochemistry, MESH: Mice, Knockout, Dexamethasone, MESH: Zona Fasciculata, Mice, 0302 clinical medicine, Cortex (anatomy), PKA, MESH: Animals, Cycloheximide, MESH: Cycloheximide, Wnt Signaling Pathway, beta Catenin, ComputingMilieux_MISCELLANEOUS, Mice, Knockout, Adrenal cortex, MESH: Sumoylation, MESH: Wnt Signaling Pathway, [SDV.MHEP.EM]Life Sciences [q-bio]/Human health and pathology/Endocrinology and metabolism, PIAS3.β-catenin, Cell biology, Neoplasm Proteins, medicine.anatomical_structure, MESH: Dexamethasone, Dactinomycin, Female, Zona Glomerulosa, Signal transduction, Biotechnology, Endocrine gland, Signal Transduction, MESH: Colforsin, MESH: Cell Line, Tumor, MESH: Mice, Transgenic, MESH: Zona Glomerulosa, MESH: Delayed-Action Preparations, Repressor, Mice, Transgenic, [SDV.CAN]Life Sciences [q-bio]/Cancer, MESH: Cyclic AMP-Dependent Protein Kinases, Biology, 03 medical and health sciences, MESH: Carney Complex, Adrenocorticotropic Hormone, Cell Line, Tumor, [SDV.BBM.GTP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], Genetics, medicine, Animals, Humans, mouse models, Carney Complex, Molecular Biology, Psychological repression, MESH: Mice, MESH: Adrenocorticotropic Hormone, MESH: Humans, Colforsin, Sumoylation, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, endocrine diseases, Cyclic AMP-Dependent Protein Kinases, Adrenal Cortex Neoplasms, MESH: Dactinomycin, [SDV.GEN.GA]Life Sciences [q-bio]/Genetics/Animal genetics, 030104 developmental biology, SENP1/2, [SDV.BDD.EO]Life Sciences [q-bio]/Development Biology/Embryology and Organogenesis, Delayed-Action Preparations, MESH: Protein Processing, Post-Translational, Adrenal Cortex, MESH: Adrenal Cortex, Zona Fasciculata, Protein Processing, Post-Translational, MESH: Female, 030217 neurology & neurosurgery, Homeostasis
Abstract

SUMOylation is a highly conserved and dynamic post-translational mechanism primarily affecting nuclear programs for adapting organisms to stressful challenges. Alteration of SUMOylation cycles leads to severe developmental and homeostatic defects and malignancy, but signals coordinating SUMOylation are still unidentified. The adrenal cortex is a zonated endocrine gland that controls body homeostasis and stress response. Here, we show that in human and in mouse adrenals, SUMOylation follows a decreasing centripetal gradient that mirrors cortical differentiation flow and delimits highly and weakly SUMOylated steroidogenic compartments, overlapping glomerulosa, and fasciculata zones. Activation of PKA signaling by acute hormonal treatment, mouse genetic engineering, or in Carney complex results in repression of small ubiquitin-like modifier (SUMO) conjugation in the inner cortex by coordinating expression of SUMO pathway inducers and repressors. Conversely, genetic activation of canonical wingless-related integration site signaling maintains high SUMOylation potential in the outer neoplastic cortex. Thus, SUMOylation is tightly regulated by signaling pathways that orchestrate adrenal zonation and diseases.-Dumontet, T., Sahut-Barnola, I., Dufour, D., Lefrancois-Martinez, A.-M., Berthon, A., Montanier, N., Ragazzon, B., Djari, C., Pointud, J.-C., Roucher-Boulez, F., Batisse-Lignier, M., Tauveron, I., Bertherat, J., Val, P., Martinez, A. Hormonal and spatial control of SUMOylation in the human and mouse adrenal cortex.

Published
2019

26. Heterogeneous expression of Pil3 pilus is critical for Streptococcus gallolyticus translocation across polarized colonic epithelial monolayers

Author

Mariana Martins, Shaynoor Dramsi, Laurence du Merle, Patrick Trieu-Cuot, Biologie des Bactéries pathogènes à Gram-positif - Biology of Gram-Positive Pathogens (BBPG+), Institut Pasteur [Paris]-Centre National de la Recherche Scientifique (CNRS), This work was supported by the French Government’s 'Investissement d’Avenir' program Laboratoire d’Excellence 'Integrative Biology of Emerging Infectious Diseases' Grant ANR-10-LABX-62-IBEID. Mariana Martins was funded by the PPU program and the ARC foundation., We sincerely thank Tarek Msadek for the critical reading of the manuscript, ANR-10-LABX-0062,IBEID,Integrative Biology of Emerging Infectious Diseases(2010), and Institut Pasteur [Paris] (IP)-Centre National de la Recherche Scientifique (CNRS)

Subjects
0301 basic medicine, [SDV]Life Sciences [q-bio], Mutant, Chromosomal translocation, Pilus, Bacterial Adhesion, 0303 health sciences, education.field_of_study, Chemistry, Human gastrointestinal tract, S. bovis, MESH: Genetic Heterogeneity, Cell biology, Infectious Diseases, medicine.anatomical_structure, MESH: Epithelial Cells, Paracellular transport, MESH: Caco-2 Cells, MESH: Bacterial Translocation, MESH: Cell Line, Tumor, MESH: Mutation, Dependent manner, Streptococcus gallolyticus, Colon, Confocal, 030106 microbiology, Immunology, Population, Translocation, Biology, Microbiology, MESH: Fimbriae, Bacterial, 03 medical and health sciences, Genetic Heterogeneity, Immune system, Cell Line, Tumor, medicine, Humans, MESH: Streptococcus gallolyticus, MESH: Bacterial Adhesion, education, 030304 developmental biology, MESH: Humans, 030306 microbiology, Pil3 pilus, Epithelial Cells, In vitro, 030104 developmental biology, Bacterial Translocation, Fimbriae, Bacterial, Mutation, Caco-2 Cells
Abstract

Streptococcus gallolyticus subspecies gallolyticus (Sgg) is an opportunistic pathogen responsible for septicaemia and endocarditis in elderly persons. Sgg is also a commensal of the human gastrointestinal tract. Here we demonstrate that Sgg strain UCN34 translocates across tight intestinal barriers in vitro in a Pil3-dependent manner. Confocal images of UCN34 passage across human colonic cells reveals that Sgg utilizes a paracellular pathway. Pil3 was previously shown to be expressed heterogeneously and WT UCN34 consists of about 90% of Pil3low and 10% of Pil3high cells. We found that both the Δpil3 mutant and the Pil3+ overexpressing variant could not translocate across Caco-2 and T84 barriers. Interestingly, combining live Δpil3 mutant cells with fixed Pil3+ variants in a 10:1 ratio (mimicking UCN34 WT population) allowed efficient translocation of the Δpil3 mutant. These experiments demonstrate that heterogeneous expression of Pil3 plays a key role in optimal translocation of Sgg across the intestinal barrier.ABSTRACT IMPORTANCEStreptococcus gallolyticus subsp. gallolyticus (Sgg) is an opportunistic pathogen responsible for septicemia and infective endocarditis in elderly persons. Sgg is a commensal of the rumen of herbivores and transmission to humans most probably occurs through the oral route. In this work, we have studied how this bacterium crosses the intestinal barrier using well-known in vitro models. Confocal microscopy images revealed that Sgg UCN34 can traverse the epithelial monolayer in between adjacent cells. We next showed that passage of Sgg from the apical to the basolateral compartment is dependent on the heterogenous expression of the Pil3 pilus at the bacterial surface. We hypothesize that Pil3high cocci adhere firmly to epithelial cells to activate transient opening of tight junctions thereby allowing the traversal of Pil3low bacteria.

Published
2019

27. Analysis of mRNA, miRNA, and DNA in Bone Cells by RT-qPCR and In Situ Hybridization

Author

Brice Moukengue, Jérôme Amiaud, Francois Lamoureux, Camille Jacques, Benjamin Ory, Céline Charrier, Sarcomes osseux et remodelage des tissus calcifiés - Phy-Os [Nantes - INSERM U1238] (Phy-Os), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Bretagne Loire (UBL)-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), Equipe Labellisée LIGUE 2012 [Nantes], Physiologie neurovégétative - PNV (PNP), Université Paul Cézanne - Aix-Marseille 3-Institut National de la Recherche Agronomique (INRA)-Centre National de la Recherche Scientifique (CNRS), Physiopathologie de la résorption osseuse et thérapie des tumeurs osseuses primitives, Université de Nantes (UN)-IFR26-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)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Bretagne Loire (UBL)-Centre hospitalier universitaire de Nantes (CHU Nantes)

Subjects
0301 basic medicine, MESH: Xenograft Model Antitumor Assays, Histology, MESH: Cell Line, Tumor, mRNA, [SDV]Life Sciences [q-bio], Chromogenic in situ hybridization, In situ hybridization, 03 medical and health sciences, chemistry.chemical_compound, MESH: Gene Expression Profiling, 0302 clinical medicine, MESH: In Situ Hybridization, MESH: Oligonucleotides, Bone sarcoma, MESH: Reverse Transcriptase Polymerase Chain Reaction, Bone cell, MESH: Mice, Nude, Digoxigenin, MESH: Animals, Locked nucleic acid, CISH, Bone, MESH: Mice, miRNA, MESH: RNA, Messenger, LNA probes, MESH: Humans, Chemistry, MESH: Real-Time Polymerase Chain Reaction, RT-qPCR, MESH: Digoxigenin, MESH: DNA, DNA, MESH: Bone and Bones, Molecular biology, Reverse transcriptase, 3. Good health, Primers, MESH: Histocytological Preparation Techniques, 030104 developmental biology, Real-time polymerase chain reaction, 030220 oncology & carcinogenesis, Gene expression, MESH: MicroRNAs
Abstract

International audience; The aim of this chapter is to describe a method used to evaluate gene expression and microRNAs (miRNAs) in bone cells or tissue using Reverse transcription and quantitative Polymerase Chain Reaction (RT-qPCR), and a method to assess chromogenic in situ hybridization (CISH) on Formalin Fixed Paraffin Embedded (FFPE ) mouse bone tissue to detect both DNA and mRNA transcripts using the double digoxigenin (DIG) locked nucleic acid (LNA™) probes .

Published
2019

28. PD-L1 microSPECT/CT Imaging for Longitudinal Monitoring of PD-L1 Expression in Syngeneic and Humanized Mouse Models for Cancer

Author

Heskamp, Sandra, Wierstra, Peter, Molkenboer-Kuenen, Janneke, Sandker, Gerwin, Thordardottir, Soley, Cany, Jeannette, Olive, Daniel, BUSSINK, Johan, Boerman, Otto, Dolstra, Harry, Aarntzen, Erik, Hobo, Willemijn, Molkenboer-Kuenen, Janneke D.M., Aarntzen, Erik H.J.G., Radboud University Medical Center [Nijmegen], Biothérapies Hépatiques, Université de Nantes (UN)-Institut National de la Santé et de la Recherche Médicale (INSERM), Centre de Recherche en Cancérologie de Marseille (CRCM), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Paoli-Calmettes, Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Aix Marseille Université (AMU), Department of Nuclear Medecine (Radboud University Medical Centre), Department of Laboratory Medicine, Radboud university [Nijmegen]-Nijmegen Centre for Molecular Life Sciences-Nijmegen Medical Centre [Nijmegen], Aix Marseille Université (AMU)-Institut Paoli-Calmettes, Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), and Radboud University [Nijmegen]-Nijmegen Centre for Molecular Life Sciences-Nijmegen Medical Centre [Nijmegen]

Subjects
0301 basic medicine, Cancer Research, Cancer development and immune defence Radboud Institute for Molecular Life Sciences [Radboudumc 2], [SDV]Life Sciences [q-bio], lnfectious Diseases and Global Health Radboud Institute for Molecular Life Sciences [Radboudumc 4], B7-H1 Antigen, MESH: Antibodies, Monoclonal, Tumours of the digestive tract Radboud Institute for Health Sciences [Radboudumc 14], 0302 clinical medicine, Neoplasms, MESH: B7-H1 Antigen, MESH: Animals, MESH: Neoplasms, MESH: Indium Radioisotopes, 0303 health sciences, Mice, Inbred BALB C, biology, Indium Radioisotopes, Antibodies, Monoclonal, Hematology, 3. Good health, medicine.anatomical_structure, Oncology, 030220 oncology & carcinogenesis, Pd l1 expression, Female, MESH: Single Photon Emission Computed Tomography Computed Tomography, Lymph, Antibody, Rare cancers Radboud Institute for Health Sciences [Radboudumc 9], Single Photon Emission Computed Tomography Computed Tomography, MESH: Cell Line, Tumor, Immunology, MESH: Mice, Inbred BALB C, Spleen, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Rare cancers Radboud Institute for Molecular Life Sciences [Radboudumc 9], 03 medical and health sciences, Immune system, Downregulation and upregulation, MESH: Mice, Inbred C57BL, PD-L1, Cell Line, Tumor, medicine, Animals, Humans, 030304 developmental biology, MESH: Humans, business.industry, Cancer, medicine.disease, Mice, Inbred C57BL, Disease Models, Animal, 030104 developmental biology, Humanized mouse, biology.protein, Cancer research, Ct imaging, MESH: Disease Models, Animal, business, Nanomedicine Radboud Institute for Molecular Life Sciences [Radboudumc 19], MESH: Female
Abstract

Contains fulltext : 202258.pdf (Publisher’s version ) (Closed access) Antibodies that block the interaction between programmed death ligand 1 (PD-L1) and PD-1 have shown impressive responses in subgroups of patients with cancer. PD-L1 expression in tumors seems to be a prerequisite for treatment response. However, PD-L1 is heterogeneously expressed within tumor lesions and may change upon disease progression and treatment. Imaging of PD-L1 could aid in patient selection. Previously, we showed the feasibility to image PD-L1(+) tumors in immunodeficient mice. However, PD-L1 is also expressed on immune cell subsets. Therefore, the aim of this study was to assess the potential of PD-L1 micro single-photon emission tomography/computed tomography (microSPECT/CT) using radiolabeled PD-L1 antibodies to (i) measure PD-L1 expression in two immunocompetent tumor models (syngeneic mice and humanized mice harboring PD-L1 expressing immune cells) and (ii) monitor therapy-induced changes in tumor PD-L1 expression. We showed that radiolabeled PD-L1 antibodies accumulated preferentially in PD-L1(+) tumors, despite considerable uptake in certain normal lymphoid tissues (spleen and lymph nodes) and nonlymphoid tissues (duodenum and brown fat). PD-L1 microSPECT/CT imaging could also distinguish between high and low PD-L1-expressing tumors. The presence of PD-L1(+) immune cells did not compromise tumor uptake of the human PD-L1 antibodies in humanized mice, and we demonstrated that radiotherapy-induced upregulation of PD-L1 expression in murine tumors could be monitored with microSPECT/CT imaging. Together, these data demonstrate that PD-L1 microSPECT/CT is a sensitive technique to detect variations in tumor PD-L1 expression, and in the future, this technique may enable patient selection for PD-1/PD-L1-targeted therapy.

Published
2019

29. Lipid nanocapsules decorated and loaded with cannabidiol as targeted prolonged release carriers for glioma therapy: In vitro screening of critical parameters

Author

Victor Sebastian, Jean-Pierre Benoit, Juan Aparicio-Blanco, Ana I. Torres-Suárez, Micro et Nanomédecines Translationnelles (MINT), and Université d'Angers (UA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)

Subjects
Cannabinoid receptor, Pharmaceutical Science, Lipid-based carriers, 02 engineering and technology, Pharmacology, 030226 pharmacology & pharmacy, 0302 clinical medicine, MESH: Nanocapsules, Cannabidiol, MESH: Inhibitory Concentration 50, Chemistry, MESH: Glioblastoma, Glioma targeting, MESH: Drug Screening Assays, Antitumor, General Medicine, 021001 nanoscience & nanotechnology, Lipids, surgical procedures, operative, MESH: Cannabidiol, Lipophilicity, 0210 nano-technology, Biotechnology, medicine.drug, MESH: Cell Line, Tumor, Lipid nanocapsules, MESH: Delayed-Action Preparations, Antineoplastic Agents, digestive system, Excipients, 03 medical and health sciences, Inhibitory Concentration 50, Nanocapsules, [SDV.SP.MED]Life Sciences [q-bio]/Pharmaceutical sciences/Medication, In vivo, Glioma, Cell Line, Tumor, medicine, Humans, MESH: Particle Size, Particle Size, IC50, Cannabis, MESH: Excipients, MESH: Humans, Cannabinoids, medicine.disease, MESH: Lipids, In vitro, digestive system diseases, Delayed-Action Preparations, MESH: Cannabis, MESH: Antineoplastic Agents, Drug Screening Assays, Antitumor, Extended-release, Glioblastoma
Abstract

International audience; The therapeutic potential of cannabinoids has been truly constrained heretofore due to their strong psychoactive effects and their high lipophilicity. In this context, precisely due to the lack of psychoactive properties, cannabidiol (CBD), the second major component of Cannabis sativa, arises as the phytocannabinoid with the most auspicious therapeutic potential. Hence, the incorporation of CBD in lipid nanocapsules (LNCs) will contribute to overcome the dosing problems associated with cannabinoids. Herein, we have prepared LNCs decorated and loaded with CBD for glioma therapy and screened in vitro their critical parameters. On the one hand, we have encapsulated CBD into the oily core of LNCs to test their in vitro efficacy as extended-release carriers against the human glioblastoma cell line U373MG. The in vitro antitumor effect was highly dependent on the size of LNCs due to its pivotal role in the extent of CBD release. Effectively, a comparison between two differently-sized LNCs (namely, 20-nm and 50-nm sized carriers) showed that the smaller LNCs reduced by 3.0-fold the IC50 value of their 50-nm sized counterparts. On the other hand, to explore the potential of this phytocannabinoid to target any of the cannabinoid receptors overexpressed in glioma cells, we decorated the LNCs with CBD. This functionalization strategy enhanced the in vitro glioma targeting by 3.4-fold in comparison with their equally-sized undecorated counterparts. Lastly, the combination of CBD-loading with CBD-functionalization further reduced the IC50 values. Hence, the potential of these two strategies of CBD incorporation into LNCs deserves subsequent in vivo evaluation.

Published
2019

30. Preventing Calpain Externalization by Reducing ABCA1 Activity with Probenecid Limits Melanoma Angiogenesis and Development

Author

Laurent Baud, Guillaume Hanouna, Joëlle Perez, Jean-Philippe Haymann, Emmanuel Letavernier, Sophie Vandermeersch, Ellie Tang, Gestionnaire, Hal Sorbonne Université, Des Maladies Rénales Rares aux Maladies Fréquentes, Remodelage et Réparation, Institut National de la Santé et de la Recherche Médicale (INSERM)-Sorbonne Université (SU), Departement Hospitalo- Universitaire - Inflammation, Immunopathologie, Biothérapie [Paris] (DHU - I2B), Institut National de la Santé et de la Recherche Médicale (INSERM)-CHU Pitié-Salpêtrière [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-CHU Trousseau [APHP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS)-CHU Saint-Antoine [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU), CHU Tenon [AP-HP], CHU Trousseau [APHP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-CHU Saint-Antoine [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-CHU Pitié-Salpêtrière [AP-HP], Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), Sorbonne Université (SU)-CHU Pitié-Salpêtrière [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-CHU Saint-Antoine [AP-HP], and Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-Sorbonne Université (SU)-CHU Trousseau [APHP]

Subjects
0301 basic medicine, Male, Skin Neoplasms, MESH: Calpain, Angiogenesis, [SDV]Life Sciences [q-bio], Melanoma, Experimental, MESH: Probenecid, MESH: Calcium-Binding Proteins, Biochemistry, chemistry.chemical_compound, Mice, 0302 clinical medicine, MESH: Animals, Calpastatin, biology, Neovascularization, Pathologic, Calpain, Probenecid, Melanoma, [SDV] Life Sciences [q-bio], MESH: Melanoma, Experimental, Vascular endothelial growth factor, 030220 oncology & carcinogenesis, medicine.drug, ATP Binding Cassette Transporter 1, MESH: Cell Line, Tumor, MESH: Mice, Transgenic, Mice, Transgenic, Dermatology, 03 medical and health sciences, MESH: Cell Proliferation, Cell Line, Tumor, medicine, Extracellular, Animals, Humans, MESH: ATP Binding Cassette Transporter 1, MESH: Mice, Molecular Biology, Cell Proliferation, MESH: Humans, MESH: Skin Neoplasms, Calcium-Binding Proteins, Cell Biology, medicine.disease, MESH: Male, Fibronectin, 030104 developmental biology, chemistry, biology.protein, Cancer research, MESH: Neovascularization, Pathologic
Abstract

International audience; Calpains, intracellular proteases specifically inhibited by calpastatin, play a major role in neoangiogenesis involved in tumor invasiveness and metastasis. They are partly exteriorized via the ATP-binding cassette transporter A1(ABCA1) transporter, but the importance of this process in tumor growth is still unknown. The aim of our study was to investigate the role of extracellular calpains in a model of melanoma by blocking their extracellular activity or exteriorization. In the first approach, a B16-F10 model of melanoma was developed in transgenic mice expressing high extracellular levels of calpastatin. In these mice, tumor growth was inhibited by ∼ 3-fold compared with wild-type animals. In vitro cytotoxicity assays and in vivo tumor studies have demonstrated that this protection was associated with a defect in tumor neoangiogenesis. Similarly, in wild-type animals given probenecid to blunt ABCA1 activity, melanoma tumor growth was inhibited by ∼ 3-fold. Again, this response was associated with a defect in neoangiogenesis. In vitro studies confirmed that probenecid limited endothelial cell migration and capillary formation from vascular explants. The observed reduction in fibronectin cleavage under these conditions is potentially involved in the response. Collectively, these studies demonstrate that probenecid, by blunting ABCA1 activity and thereby calpain exteriorization, limits melanoma tumor neoangiogenesis and invasiveness.

Published
2018

31. Histone H4K20 methylation mediated chromatin compaction threshold ensures genome integrity by limiting DNA replication licensing

Author

Eric Julien, Fanny Izard, Klaus Hansen, Jens Vilstrup Johansen, Mads Lerdrup, David Llères, Birthe Fahrenkrog, Muhammad Shoaib, David Walter, Claus Storgaard Sørensen, J. Julian Blow, Peter J. Gillespie, University of Copenhagen = Københavns Universitet (KU), University of Dundee, Institut de Recherche en Cancérologie de Montpellier (IRCM - U1194 Inserm - UM), CRLCC Val d'Aurelle - Paul Lamarque-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Montpellier (UM), Université libre de Bruxelles (ULB), Institut de Génétique Moléculaire de Montpellier (IGMM), Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM), Herrada, Anthony, University of Copenhagen = Københavns Universitet (UCPH), and Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS)

Subjects
DNA Replication, 0301 basic medicine, MESH: Cell Line, Tumor, Cell division, Science, General Physics and Astronomy, MESH: DNA Replication, MESH: Flow Cytometry, MESH: Microscopy, Fluorescence, Article, General Biochemistry, Genetics and Molecular Biology, MESH: Chromatin, Histones, Histone H4, 03 medical and health sciences, Cell Line, Tumor, [SDV.BBM.GTP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], MESH: RNA, Small Interfering, Humans, Chimie, RNA, Small Interfering, lcsh:Science, Mitosis, MESH: DNA Damage, MESH: Histones, MESH: Humans, Multidisciplinary, biology, Chemistry, Physique, DNA replication, General Chemistry, Astronomie, Flow Cytometry, Chromatin, Cell biology, Technologie de l'environnement, contrôle de la pollution, 030104 developmental biology, Histone, Microscopy, Fluorescence, Mitotic exit, biology.protein, [SDV.BBM.GTP] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Genomics [q-bio.GN], Origin recognition complex, lcsh:Q, DNA Damage
Abstract

The decompaction and re-establishment of chromatin organization immediately after mitosis is essential for genome regulation. Mechanisms underlying chromatin structure control in daughter cells are not fully understood. Here we show that a chromatin compaction threshold in cells exiting mitosis ensures genome integrity by limiting replication licensing in G1 phase. Upon mitotic exit, chromatin relaxation is controlled by SET8-dependent methylation of histone H4 on lysine 20. In the absence of either SET8 or H4K20 residue, substantial genome-wide chromatin decompaction occurs allowing excessive loading of the origin recognition complex (ORC) in the daughter cells. ORC overloading stimulates aberrant recruitment of the MCM2-7 complex that promotes single-stranded DNA formation and DNA damage. Restoring chromatin compaction restrains excess replication licensing and loss of genome integrity. Our findings identify a cell cycle-specific mechanism whereby fine-tuned chromatin relaxation suppresses excessive detrimental replication licensing and maintains genome integrity at the cellular transition from mitosis to G1 phase., SCOPUS: ar.j, info:eu-repo/semantics/published

Published
2018

32. RK, the first scorpion peptide with dual disintegrin activity on α1β1 and αvβ3 integrins

Author

Hazem Ben Mabrouk, Oussema Khamessi, Michel De Waard, Mejdoub Hafedh, Houcemeddine Othman, R. Kharrat, Najet Srairi-Abid, Naziha Marrakchi, Rym ElFessi-Magouri, Laboratoire des Venins et Biomolécules Thérapeutiques - Laboratory of Venoms and Therapeutic Biomolecules (LR11IPT08), Institut Pasteur de Tunis, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP), unité de recherche de l'institut du thorax UMR1087 UMR6291 (ITX), Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université de Nantes - UFR de Médecine et des Techniques Médicales (UFR MEDECINE), Université de Nantes (UN)-Université de Nantes (UN), Smartox Biotechnology, USCR Séquenceur de Protéines, Faculté des Sciences de Sfax, Université de Sfax - University of Sfax-Université de Sfax - University of Sfax, This work was supported by grants from the Tunisian Ministry of Higher Education and Scientific Research (LR11IPT08) and Institut Pasteur of Tunisia (approval number: LNFP/Pro 152012)., and We thank Pr Hechmi Louzir, head of Institut Pasteur de Tunis for his continued interest in this study and for his support.

Subjects
0301 basic medicine, MESH: Cell Line, Tumor, Integrin, Molecular modeling, Venom, Peptide, MESH: Amino Acid Sequence, MESH: Disintegrins/metabolism, Biochemistry, MESH: Integrin alphaVbeta3/metabolism, 03 medical and health sciences, 0302 clinical medicine, MESH: Protein Conformation, MESH: Cell Adhesion/drug effects, Structural Biology, RK peptide, Disintegrin, MESH: Integrin alpha1beta1/metabolism, MESH: Molecular Docking Simulation, MESH: Animals, MESH: Molecular Dynamics Simulation, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Cell adhesion, Receptor, Molecular Biology, MESH: Mice, chemistry.chemical_classification, MESH: Humans, biology, Chemistry, MESH: Disintegrins/chemistry, General Medicine, biology.organism_classification, MESH: Male, Cell biology, 030104 developmental biology, Snake venom, biology.protein, Adhesion, MESH: Disintegrins/pharmacology, MESH: Scorpion Venoms/chemistry, Buthus occitanus, 030217 neurology & neurosurgery
Abstract

International audience; the first scorpion peptide with dual disintegrin activity on α1β1 and αvβ3 integrins. Abstract: Scorpion peptides are well known for their pharmaceutical potential on different targets. These include mainly the ion channels which were found to be highly expressed in many diseases, including cancer, auto-immune pathologies and Alzheimer. So far, however, the disintegrin activity had only been characterized for snake venom molecules. Herein, we present the first short peptide, purified from the venom of Buthus occitanus tunetanus, (termed RK) able to inhibit the cell adhesion of Glioblastoma, Melanoma and Rat pheochromocytoma to different extracellular matrix (ECM) receptors. Anti-integrin antibody assay suggests that RK interacts with both α1β1 and αvβ3 with a more pronounced effect for the former. The examination of the primary structure of RK suggests the involvement of two motifs: KSS, analogue to KTS which was characterized for α1β1 Snake venom disintegrins, and ECD, analogue to RGD which was found to be active on αvβ3. To assess their roles in the disintegrin activity of RK, we conducted a computational analysis. The molecular docking study shows that RK involves mainly two segments to interact with the α1β1 integrin, but the peptide does not implicate the KSS motif in the interaction. The molecular modeling study, suggests the key contribution of the ECD segment in the interaction with αvβ3 integrin.

Published
2018

33. Lactate Dehydrogenase B Controls Lysosome Activity and Autophagy in Cancer

Author

Thibaut Vazeille, Pierre Sonveaux, Vincent F. Van Hée, Paolo E. Porporato, Tamara Copetti, Morgane Tardy, Lucie Brisson, Piotr Bański, Caroline Bouzin, Pierre Danhier, Raphaël Frédérick, Jorge Falces, Carine Michiels, Martina Sboarina, Coralie Dethier, Marie Joséphine Fontenille, Université Catholique de Louvain = Catholic University of Louvain (UCL), Louvain Drug Research Institute [Bruxelles, Belgique] (LDRI), and Université de Namur [Namur] (UNamur)

Subjects
0301 basic medicine, MESH: L-Lactate Dehydrogenase, Cancer Research, MESH: Cell Line, Tumor, [SDV.CAN]Life Sciences [q-bio]/Cancer, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, Oxidative phosphorylation, Biology, Cell Line, Mice, 03 medical and health sciences, Cell Line, Tumor, Neoplasms, MESH: Cell Proliferation, Lysosome, Autophagy, Human Umbilical Vein Endothelial Cells, medicine, Animals, Humans, MESH: Autophagy, Cell Proliferation, Isoenzymes, L-Lactate Dehydrogenase, Lysosomes, MESH: Animals, MESH: Neoplasms, Glycolysis, MESH: Human Umbilical Vein Endothelial Cells, MESH: Mice, Tumor, MESH: Humans, Cancer, Cell Biology, medicine.disease, 3. Good health, Cell biology, 030104 developmental biology, medicine.anatomical_structure, Oncology, Biochemistry, Tumor progression, Cancer cell, MESH: Isoenzymes, NAD+ kinase, MESH: Lysosomes
Abstract

Metabolic adaptability is essential for tumor progression and includes cooperation between cancer cells with different metabolic phenotypes. Optimal glucose supply to glycolytic cancer cells occurs when oxidative cancer cells use lactate preferentially to glucose. However, using lactate instead of glucose mimics glucose deprivation, and glucose starvation induces autophagy. We report that lactate sustains autophagy in cancer. In cancer cells preferentially to normal cells, lactate dehydrogenase B (LDHB), catalyzing the conversion of lactate and NAD+ to pyruvate, NADH and H+, controls lysosomal acidification, vesicle maturation, and intracellular proteolysis. LDHB activity is necessary for basal autophagy and cancer cell proliferation not only in oxidative cancer cells but also in glycolytic cancer cells.

Published
2016

34. The anti-HER3 (ErbB3) therapeutic antibody 9F7-F11 induces HER3 ubiquitination and degradation in tumors through JNK1/2- dependent ITCH/AIP4 activation

Author

Christel Larbouret, Marie Alix Poul, Thierry Chardès, André Pèlegrin, Gerry Melino, Philippe Mondon, Christophe Le Clorennec, Yassamine Lazrek, Olivier Dubreuil, Institut de Recherche en Cancérologie de Montpellier (IRCM - U1194 Inserm - UM), CRLCC Val d'Aurelle - Paul Lamarque-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Montpellier (UM), Institut Pasteur de la Guyane, Réseau International des Instituts Pasteur (RIIP), Millegen SA, MILEGEN-Immeuble BIOSTEP, GamaMabs Pharma SA [Toulouse] ( Centre Pierre Potier), Oncopole de Toulouse-Centre Pierre Potier [Toulouse], LFB Biotechnologies [Lille, France], Università degli Studi di Roma Tor Vergata [Roma], University of Leicester, and Herrada, Anthony

Subjects
0301 basic medicine, Cell cycle checkpoint, Receptor, ErbB-3, MESH: Down-Regulation, MESH: Antibodies, Monoclonal, Antibodies, Monoclonal, Murine-Derived, 0302 clinical medicine, Ubiquitin, antibody, Medicine, skin and connective tissue diseases, [SDV.MHEP] Life Sciences [q-bio]/Human health and pathology, biology, treatment, Antibodies, Monoclonal, 3. Good health, Ubiquitin ligase, ITCH/AIP4, MESH: Receptor, ErbB-3, Oncology, MESH: Repressor Proteins, 030220 oncology & carcinogenesis, MESH: Mitogen-Activated Protein Kinase 9, Phosphorylation, MESH: Mitogen-Activated Protein Kinase 8, Research Paper, MESH: Cell Line, Tumor, MAP Kinase Signaling System, Ubiquitin-Protein Ligases, Down-Regulation, [SDV.CAN]Life Sciences [q-bio]/Cancer, Antineoplastic Agents, 03 medical and health sciences, [SDV.CAN] Life Sciences [q-bio]/Cancer, HER3, Cell Line, Tumor, cancer, Humans, Mitogen-Activated Protein Kinase 9, Mitogen-Activated Protein Kinase 8, Settore BIO/10, Protein kinase B, PI3K/AKT/mTOR pathway, MESH: Humans, business.industry, MESH: MAP Kinase Signaling System, Ubiquitination, MESH: Ubiquitin-Protein Ligases, body regions, Repressor Proteins, 030104 developmental biology, USP9X, MESH: Antibodies, Monoclonal, Murine-Derived, Cancer cell, Immunology, biology.protein, Cancer research, MESH: Ubiquitination, MESH: Antineoplastic Agents, business, [SDV.MHEP]Life Sciences [q-bio]/Human health and pathology
Abstract

International audience; We characterized the mechanism of action of the neuregulin-non-competitive anti-HER3 therapeutic antibody 9F7-F11 that blocks the PI3K/AKT pathway, leading to cell cycle arrest and apoptosis in vitro and regression of pancreatic and breast cancer in vivo. We found that 9F7-F11 induces rapid HER3 down-regulation. Specifically, 9F7-F11-induced HER3 ubiquitination and degradation in pancreatic, breast and prostate cancer cell lines was driven mainly by the itchy E3 ubiquitin ligase (ITCH/AIP4). Overexpression of the ITCH/AIP4 inhibitor N4BP1 or small-interfering RNA-mediated knockdown of ITCH/AIP4 inhibited HER3 ubiquitination/degradation and PI3K/AKT signaling blockade induced by 9F7-F11. Moreover, 9F7-F11-mediated JNK1/2 phosphorylation led to ITCH/AIP4 activation and recruitment to HER3 for receptor ubiquitination and degradation. ITCH/AIP4 activity was activated by the deubiquitinases USP8 and USP9X, as demonstrated by RNA interference. Taken together, our results suggest that 9F7-F11-induced HER3 ubiquitination and degradation in cancer cells mainly occurs through JNK1/2-dependent ITCH/AIP4 activation.

Published
2016

35. The oncogenic and druggable hPG80 (Progastrin) is overexpressed in multiple cancers and detected in the blood of patients

Author

Vahan Kepenekian, Veronique Saywell, Laurent Villeneuve, Benoit You, Pierre Liaud, Berengere Vire, Frederic Mercier, Delphine Maucort-Boulch, Winston Tan, Alexandre Prieur, Julien Soulé, Maud Flacelière, Philippe Elies, Manish Kohli, Olivier Glehen, Carole Langlois-Jacques, Fanny Belouin, Dominique Joubert, Thibault Mazard, E. Assenat, Marc Ychou, Lea Payen, Eric Morency, Marie Dupuy, Service d'Oncologie Médicale [Centre hospitalier Lyon Sud - HCL], Centre Hospitalier Lyon Sud [CHU - HCL] (CHLS), Hospices Civils de Lyon (HCL)-Hospices Civils de Lyon (HCL), Eurobiodev, Institut de Génétique Moléculaire de Montpellier (IGMM), Université de Montpellier (UM)-Centre National de la Recherche Scientifique (CNRS), Centre Hospitalier Régional Universitaire [Montpellier] (CHRU Montpellier), Laboratoire d'InfoRmatique en Image et Systèmes d'information (LIRIS), Université Lumière - Lyon 2 (UL2)-École Centrale de Lyon (ECL), Université de Lyon-Université de Lyon-Université Claude Bernard Lyon 1 (UCBL), 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)-Centre National de la Recherche Scientifique (CNRS), Laboratoire de Biométrie et Biologie Evolutive - UMR 5558 (LBBE), Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-Université de Lyon-Institut National de Recherche en Informatique et en Automatique (Inria)-VetAgro Sup - Institut national d'enseignement supérieur et de recherche en alimentation, santé animale, sciences agronomiques et de l'environnement (VAS)-Centre National de la Recherche Scientifique (CNRS), Université de Lyon, Université de Bretagne Occidentale - UFR Médecine et Sciences de la Santé (UBO UFR MSS), Université de Brest (UBO), Institut du Cancer de Montpellier (ICM), Institut de Recherche en Cancérologie de Montpellier (IRCM - U1194 Inserm - UM), CRLCC Val d'Aurelle - Paul Lamarque-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Montpellier (UM), Dpt of Oncology [Rochester], Mayo Clinic, Hospices Civils de Lyon (HCL), ECS Progastrin, Salvy-Córdoba, Nathalie, Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM), Institut National des Sciences Appliquées de Lyon (INSA Lyon), Université de Lyon-Institut National des Sciences Appliquées (INSA)-Université de Lyon-Institut National des Sciences Appliquées (INSA)-Centre National de la Recherche Scientifique (CNRS)-Université Claude Bernard Lyon 1 (UCBL), Université de Lyon-École Centrale de Lyon (ECL), and Université de Lyon-Université Lumière - Lyon 2 (UL2)

Subjects
Male, [SDV.MHEP.HEM] Life Sciences [q-bio]/Human health and pathology/Hematology, 0301 basic medicine, MESH: Protein Precursors, Research paper, MESH: Oncogenes, Antibodies, Neoplasm, Colorectal cancer, Druggability, lcsh:Medicine, MESH: Spheroids, Cellular, MESH: Gastrins, Neutralization, Cohort Studies, MESH: Aged, 80 and over, 0302 clinical medicine, Neoplasms, Medicine, MESH: Neoplasms, MESH: Cohort Studies, MESH: Organ Specificity, Cancer, Aged, 80 and over, MESH: Aged, lcsh:R5-920, MESH: Middle Aged, MESH: Kinetics, Progastrin, [SDV.MHEP.HEM]Life Sciences [q-bio]/Human health and pathology/Hematology, MESH: Gene Expression Regulation, Neoplastic, General Medicine, Middle Aged, 3. Good health, Peritoneal carcinomatosis, Gene Expression Regulation, Neoplastic, Organ Specificity, MESH: Young Adult, 030220 oncology & carcinogenesis, Immunohistochemistry, Female, lcsh:Medicine (General), Adult, MESH: Cell Line, Tumor, Monitoring, Blood biomarker, Antineoplastic Agents, [SDV.CAN]Life Sciences [q-bio]/Cancer, General Biochemistry, Genetics and Molecular Biology, Young Adult, 03 medical and health sciences, [SDV.CAN] Life Sciences [q-bio]/Cancer, Cancer stem cell, Cell Line, Tumor, Spheroids, Cellular, MESH: Cell Proliferation, Gastrins, Humans, RNA, Messenger, Protein Precursors, Gene, Aged, Cell Proliferation, MESH: RNA, Messenger, MESH: Humans, hPG80, business.industry, lcsh:R, MESH: Adult, Oncogenes, medicine.disease, MESH: Male, Kinetics, MESH: Antibodies, Neoplasm, 030104 developmental biology, Cancer research, MESH: Antineoplastic Agents, Therapy, business, MESH: Female
Abstract

Background: In colorectal cancer, hPG80 (progastrin) is released from tumor cells, promotes cancer stem cells (CSC) self-renewal and is detected in the blood of patients. Because the gene GAST that encodes hPG80 is a target gene of oncogenic pathways that are activated in many tumor types, we hypothesized that hPG80 could be expressed by tumors from various origins other than colorectal cancers, be a drug target and be detectable in the blood of these patients. Methods: hPG80 expression was monitored by fluorescent immunohistochemistry and mRNA expression in tumors from various origins. Cancer cell lines were used in sphere forming assay to analyze CSC self-renewal. Blood samples were obtained from 1546 patients with 11 different cancer origins and from two retrospective kinetic studies in patients with peritoneal carcinomatosis or hepatocellular carcinomas. These patients were regularly sampled during treatments and assayed for hPG80. Findings: We showed that hPG80 was present in the 11 tumor types tested. In cell lines originating from these tumor types, hPG80 neutralization decreased significantly CSC self-renewal by 28 to 54%. hPG80 was detected in the blood of patients at significantly higher concentration than in healthy blood donors (median hPG80: 4.88 pM versus 1.05 pM; p

Published
2020

36. Assessment of the targeting specificity of a fluorescent albumin conceived as a preclinical agent of the liver function

Author

Michel Bessodes, Pascal Houzé, Johanne Seguin, Katia Lemdani, Nathalie Mignet, Rabah Gahoual, Daniel Scherman, Hugo Salmon, Unité de Technologies Chimiques et Biologiques pour la Santé (UTCBS - UM 4 (UMR 8258 / U1022)), Ecole Nationale Supérieure de Chimie de Paris - Chimie ParisTech-PSL (ENSCP), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Unité de pharmacologie chimique et génétique et d'imagerie (UPCGI - UMR 8151 / UMR_S 1022 ), Centre de recherche de Vitry-Alfortville, Rhone Poulenc SA, Laboratoire de Neurochimie, Hospices Civils de Lyon (HCL), Mignet, Nathalie, Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Université Paris Descartes - Paris 5 (UPD5), Université Sorbonne Paris Cité (USPC), Université Paris sciences et lettres (PSL), Service de biochimie métabolique [CHU Necker], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP)-CHU Necker - Enfants Malades [AP-HP], Assistance publique - Hôpitaux de Paris (AP-HP) (AP-HP), CHU Necker - Enfants Malades [AP-HP], Université Paris Descartes - Faculté de Pharmacie de Paris (UPD5 Pharmacie), Université Paris sciences et lettres (PSL)-Université Paris sciences et lettres (PSL)-Université Paris Descartes - Paris 5 (UPD5)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Institut de Chimie du CNRS (INC), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Paris Descartes - Paris 5 (UPD5)-Ecole Nationale Supérieure de Chimie de Paris- Chimie ParisTech-PSL (ENSCP), Ecole Nationale Supérieure de Chimie de Paris- Chimie ParisTech-PSL (ENSCP), Paris Sciences et Lettres (PSL), Assistance publique - Hôpitaux de Paris (AP-HP) (APHP)-CHU Necker - Enfants Malades [AP-HP], and PSL Research University (PSL)

Subjects
0301 basic medicine, [SDV]Life Sciences [q-bio], Contrast Media, Lactose, [CHIM.THER]Chemical Sciences/Medicinal Chemistry, [SPI.MAT]Engineering Sciences [physics]/Materials, chemistry.chemical_compound, Mice, MESH: Liver Neoplasms, [CHIM] Chemical Sciences, General Materials Science, Tissue Distribution, MESH: Animals, Mice, Inbred BALB C, medicine.diagnostic_test, Liver Neoplasms, Optical Imaging, [CHIM.MATE]Chemical Sciences/Material chemistry, Carbocyanines, Human serum albumin, Fluorescence, [SDV.BBM.BP]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biophysics, [SDV] Life Sciences [q-bio], medicine.anatomical_structure, Biochemistry, Liver, Hepatocyte, [PHYS.PHYS.PHYS-MED-PH]Physics [physics]/Physics [physics]/Medical Physics [physics.med-ph], Female, MESH: Carbocyanines, medicine.drug, MESH: Cell Line, Tumor, MESH: Mice, Inbred BALB C, MESH: Serum Albumin, Context (language use), [SDV.CAN]Life Sciences [q-bio]/Cancer, Flow cytometry, 03 medical and health sciences, [CHIM.ANAL]Chemical Sciences/Analytical chemistry, Cell Line, Tumor, MESH: Contrast Media, medicine, MESH: Transplantation, Homologous, Animals, Humans, Transplantation, Homologous, [CHIM]Chemical Sciences, MESH: Lactose, [PHYS.PHYS.PHYS-INS-DET]Physics [physics]/Physics [physics]/Instrumentation and Detectors [physics.ins-det], MESH: Tissue Distribution, MESH: Mice, Serum Albumin, MESH: Optical Imaging, MESH: Humans, Albumin, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, 030104 developmental biology, chemistry, Liver function, MESH: Female, MESH: Liver
Abstract

International audience; In the context of increasing liver diseases, no contrast agent is currently available in Europe and the United States to directly assess the liver function. Only neolactosylated human serum albumin is being clinically used in Asia. In order to perform preclinical studies in the context of liver diseases, we conceived a fluorescent lactosylated albumin for the quantification of liver functional cells (l-Cyal). Precise characterization was achieved in order to determine the amounts of lactose and Cyanine 5 (Cy5) coupled to the albumin. In addition, potential aggregation was characterized by asymmetrical flow field-flow fractionation hyphenated to multi-angle light scattering (AF4-MALS). The optimal functionalized albumin exhibited a mass greater than 87 kDa which corresponds to the addition of 34 lactose moieties per protein and 1-2 Cy5 labels. Also, no significant formation of aggregates could be identified due to the modification of the native albumin. In healthy mice, the accumulation of l-Cyal in the liver and its selectivity for hepatocyte cells were shown by optical imaging and flow cytometry. Administration of l-Cyal to mice bearing liver metastases showed a reduced signal in the liver related to a decrease in the number of hepatocytes. The l-Cyal bioimaging contrast agent could be particularly useful for assessing the state of liver related diseases.

Published
2018

37. Cell polarity and adherens junction formation inhibit epithelial Fas cell death receptor signaling

Author

Gagnoux-Palacios, Laurent, Awina, Hala, Audebert, Stéphane, Rossin, Aurélie, Mondin, Magali, Borgese, Franck, Planas-Botey, Carlota, Mettouchi, Amel, Borg, Jean-Paul, Hueber, Anne-Odile, Institut de Biologie Valrose (IBV), Université Nice Sophia Antipolis (1965 - 2019) (UNS), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS)-Université Côte d'Azur (UCA), Centre de Recherche en Cancérologie de Marseille (CRCM), Aix Marseille Université (AMU)-Institut Paoli-Calmettes, Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Institute of Developmental Biology and Cancer (IBDC), COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Centre National de la Recherche Scientifique (CNRS)-Université Côte d'Azur (UCA), Toxines bactériennes - Bacterial Toxins, Institut Pasteur [Paris] (IP)-Université Paris Diderot - Paris 7 (UPD7)-Centre National de la Recherche Scientifique (CNRS), Institut Paoli-Calmettes, Fédération nationale des Centres de lutte contre le Cancer (FNCLCC), This work was supported by institutional funding from Inserm, Centre National de la Recherche Scientifique, and by grants from the Association pour la Recherche sur le Cancer and the LABEX SIGNALIFE program (ANR-11-LABX-0028-01). J.-P. Borg’s laboratory is supported by La Ligue Nationale Contre le Cancer (Label Ligue) and Ruban Rose Award. J.-P. Borg is a scholar of Institut Universitaire de France. Proteomic analyses were done using the mass spectrometry facility of Aix Marseille Université Proteomics labeled 'Plateforme technologique de l’Université Aix-Marseille,' supported by Infrastructures Biologie Santé et Agronomie, the Cancéropôle PACA, the Provence-Alpes-Côte d'Azur Region, the Institut Paoli-Calmettes, and the Centre de Recherche en Cancérologie de Marseille., ANR-11-LABX-0028,SIGNALIFE,Réseau d'Innovation sur les Voies de Signalisation en Sciences de la Vie(2011), Université Nice Sophia Antipolis (... - 2019) (UNS), Université Côte d'Azur (UCA)-Université Côte d'Azur (UCA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Centre National de la Recherche Scientifique (CNRS), Centre National de la Recherche Scientifique (CNRS)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Paoli-Calmettes, Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Fédération nationale des Centres de lutte contre le Cancer (FNCLCC)-Aix Marseille Université (AMU), Université Côte d'Azur (UCA)-Université Côte d'Azur (UCA)-Centre National de la Recherche Scientifique (CNRS), Institut Pasteur [Paris]-Université Paris Diderot - Paris 7 (UPD7)-Centre National de la Recherche Scientifique (CNRS), ANR: 11-LABX-0028,SIGNALIFE,Réseau d'Innovation sur les Voies de Signalisation en Sciences de la Vie(2011), and COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-COMUE Université Côte d'Azur (2015-2019) (COMUE UCA)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université Côte d'Azur (UCA)-Centre National de la Recherche Scientifique (CNRS)

Subjects
Proteomics, Fas Ligand Protein, MESH: Cell Line, Tumor, MESH: Adaptor Proteins, Signal Transducing/metabolism, [SDV]Life Sciences [q-bio], MESH: Epithelial Cells/cytology, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, MESH: Cadherins/genetics, MESH: fas Receptor/genetics, Discs Large Homolog 1 Protein, MESH: Membrane Proteins/genetics, MESH: Adaptor Proteins, Signal Transducing/genetics, Protein Domains, Antigens, CD, Cell Line, Tumor, Report, MESH: Multiprotein Complexes/genetics, Humans, MESH: Membrane Proteins/metabolism, fas Receptor, MESH: Multiprotein Complexes/metabolism, Research Articles, MESH: fas Receptor/metabolism, Adaptor Proteins, Signal Transducing, MESH: Humans, MESH: Adherens Junctions/genetics, MESH: Cadherins/metabolism, MESH: Proteomics, Cell Polarity, Membrane Proteins, Epithelial Cells, Adherens Junctions, Cadherins, MESH: Fas Ligand Protein/genetics, MESH: Fas Ligand Protein/metabolism, MESH: Epithelial Cells/metabolism, Multiprotein Complexes, MESH: Antigens, CD/genetics, MESH: Protein Domains, MESH: Cell Polarity
Abstract

Control of epithelial cell death is crucial to maintaining tissue integrity. Gagnoux-Palacios et al. show that cell polarity and adherens junction formation prevent proapoptotic signals emanating from the Fas death receptor. Therefore, Fas-dependent cell death contributes to the elimination of nonpolarized or nonadherent cells from human epithelia., Finely tuned regulation of epithelial cell death maintains tissue integrity and homeostasis. At the cellular level, life and death decisions are controlled by environmental stimuli such as the activation of death receptors. We show that cell polarity and adherens junction formation prevent proapoptotic signals emanating from the Fas death receptor. Fas is sequestered in E-cadherin actin-based adhesion structures that are less able to induce downstream apoptosis signaling. Using a proteomic-based approach, we find that the polarity molecule Dlg1 interacts with the C-terminal PDZ-binding site in Fas and that this interaction decreases formation of the death-inducing complex upon engagement with Fas ligand (FasL), thus acting as an additional cell death protection mechanism. We propose that E-cadherin and Dlg1 inhibit FasL-induced cell death by two complementary but partially independent mechanisms that help to maintain epithelial homeostasis by protecting normal polarized epithelia from apoptosis. When polarity is lost, the Fas–cadherin–Dlg1 antiapoptotic complex is disrupted, and FasL can promote the elimination of compromised nonpolarized cells.

Published
2018

38. High stromal transforming growth factor β-induced expression is a novel marker of progression and poor prognosis in gastric cancer

Author

Kyoichi Ogata, Andrei Turtoi, Takehiko Yokobori, Masaki Suzuki, Ken Shirabe, Tadashi Handa, Hiroyuki Kuwano, Masakazu Yashiro, Tetsunari Oyama, Navchaa Gombodorj, Department of General Surgical Science [Maebashi, Japan], Gunma University Graduate School of Medicine [Maebashi, Japan], Department of Innovative Cancer Immunotherapy [Maebashi, Japan], Department of Surgical Oncology MolecularOncology and Therapeutics [Osaka, Japan] (Osaka City), University Graduate School of Medicine [Osaka, Japan], Institut de Recherche en Cancérologie de Montpellier (IRCM - U1194 Inserm - UM), CRLCC Val d'Aurelle - Paul Lamarque-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Montpellier (UM), Institut du Cancer de Montpellier (ICM), Department of Diagnostic Pathology [Maebashi, Japan], Department of Hepatobiliary and Pancreatic Surgery [Maebashi, Japan], Gunma University Graduate Schoolof Medicine [Maebashi, Japan], This studywas funded by Grants‐in‐Aid for Scientific Research from the JapanSociety for the Promotion of Science (JSPS) (grant nos JP26461969,JP15K10085, and JP17K19893)., and Herrada, Anthony

Subjects
Male, 0301 basic medicine, MESH: Extracellular Matrix Proteins, cancer-associated fibroblast, Extracellular matrix, 0302 clinical medicine, Cell Movement, Transforming Growth Factor beta, MESH: RNA, Small Interfering, Tumor Cells, Cultured, Medicine, RNA, Small Interfering, MESH: Cell Movement, MESH: Aged, Extracellular Matrix Proteins, Predictive marker, [SDV.MHEP] Life Sciences [q-bio]/Human health and pathology, MESH: Middle Aged, General Medicine, MESH: Stomach Neoplasms, Middle Aged, Prognosis, Immunohistochemistry, 3. Good health, Oncology, Gene Knockdown Techniques, 030220 oncology & carcinogenesis, Disease Progression, Female, MESH: Disease Progression, MESH: Biomarkers, Tumor, Stromal cell, MESH: Cell Line, Tumor, [SDV.CAN]Life Sciences [q-bio]/Cancer, MESH: Prognosis, MESH: Coculture Techniques, 03 medical and health sciences, Downregulation and upregulation, [SDV.CAN] Life Sciences [q-bio]/Cancer, Stomach Neoplasms, Cell Line, Tumor, Biomarkers, Tumor, stroma, Humans, MESH: Tumor Cells, Cultured, MESH: Transforming Growth Factor beta, Aged, MESH: Humans, business.industry, gastric cancer, Cancer, MESH: Immunohistochemistry, medicine.disease, MESH: Gene Knockdown Techniques, Coculture Techniques, eye diseases, transforming factor β-induced, MESH: Male, 030104 developmental biology, Cancer research, Surgery, Stromal Cells, MESH: Stromal Cells, business, MESH: Female, [SDV.MHEP]Life Sciences [q-bio]/Human health and pathology, TGFBI, Transforming growth factor
Abstract

International audience; BACKGROUND AND OBJECTIVES:Transforming growth factor β-induced (TGFBI) protein is a secreted extracellular matrix protein with conflicting roles in cancer, acting as a tumour suppressor and a promoter, which appears to be tissue specific. The role of TGFBI in gastric cancer (GC) remains unclear, which we aimed to investigate using the clinical samples as well as an in vitro coculture model of GC.METHODS:The clinical significance of TGFBI was assessed in 208 GC samples using immunohistochemistry. Molecular function of TGFBI in the GC cells was examined by small interfering RNA-mediated TGFBI downregulation in the gastric fibroblasts cocultured with the GC cells.RESULTS:TGFBI expression was localised mainly in the cancer stroma and not in the noncancerous gastric tissue or the GC cells. High TGFBI expression was significantly associated with poor prognosis and cancer progression. Downregulation of TGFBI in the cocultured gastric fibroblasts inhibited the invasion and migration abilities of the GC cells.CONCLUSIONS:High stromal TGFBI expression might be a useful predictive marker for poor prognosis in GC patients. Furthermore, TGFBI in the cancer stromal cells is a promising target for GC treatment.

Published
2018

39. A Quinoline-Based DNA Methyltransferase Inhibitor as a Possible Adjuvant in Osteosarcoma Therapy

Author

Roberta Mazzone, Piero Picci, Katia Scotlandi, Michela Pasello, Patrizia Nanni, Giordano Nicoletti, Maria Cristina Manara, Antonello Mai, Clara Guerzoni, Camilla Cristalli, Pier Luigi Lollini, Lorena Landuzzi, Sergio Valente, Giulia Stazi, Paola B. Arimondo, Clemens Zwergel, Istituto Ortopedico Rizzoli [Bologna, Italy], Università degli Studi di Roma 'La Sapienza' = Sapienza University [Rome], Centre National de la Recherche Scientifique (CNRS), Department of Medicinal Chemistry and Technologies, Institut Pasteur, Fondation Cenci Bolognetti - Istituto Pasteur Italia, Fondazione Cenci Bolognetti, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Università degli Studi di Roma 'La Sapienza' = Sapienza University [Rome], This work was funded by AIRC (IG 18451 to K. Scotlandi, IG 19162 to A. Mai), the Italian Ministry of Health (TRANSCAN_ project TORPEDO_ER-2015-23604059 to K. Scotlandi), Progetto Ateneo Sapienza (to A. Mai), PRIN 2016 (prot 20152TE5PK to A. Mai), and the NIH (R01GM114306 to A. Mai)., The authors thank Cristina Ghinelli for editing the manuscript., Manara, Maria Cristina, Valente, Sergio, Cristalli, Camilla, Nicoletti, Giordano, Landuzzi, Lorena, Zwergel, Clemen, Mazzone, Roberta, Stazi, Giulia, Arimondo, Paola B, Pasello, Michela, Guerzoni, Clara, Picci, Piero, Nanni, Patrizia, Lollini, Pier-Luigi, Mai, Antonello, and Scotlandi, Katia

Subjects
0301 basic medicine, Male, patient-derived xenograft (PDX), MESH: Osteosarcoma/genetics, MESH: Antineoplastic Combined Chemotherapy Protocols/therapeutic use, Cellular differentiation, DNA Methyltransferase Inhibitor, [CHIM.THER]Chemical Sciences/Medicinal Chemistry, chemotherapy, MESH: Mice, Knockout, 0302 clinical medicine, MESH: Osteosarcoma/drug therapy, Antineoplastic Combined Chemotherapy Protocols, MESH: Aminoquinolines/administration & dosage, MESH: Animals, MESH: Quinolines/chemistry, Enzyme Inhibitors, MESH: DNA (Cytosine-5-)-Methyltransferase 1/metabolism, Mice, Knockout, DNA methylation, Chemistry, 3. Good health, Tumor Burden, MESH: Benzamides/pharmacology, Gene Expression Regulation, Neoplastic, 030220 oncology & carcinogenesis, Benzamides, oncology, Aminoquinolines, Quinolines, MESH: Cisplatin/administration & dosage, MESH: Bone Neoplasms/metabolism, Osteosarcoma, MESH: Doxorubicin/administration & dosage, epigenetic, MESH: Bone Neoplasms/genetics, medicine.drug, DNA (Cytosine-5-)-Methyltransferase 1, musculoskeletal diseases, MESH: Aminoquinolines/pharmacology, MESH: Cell Line, Tumor, DNA damage, MESH: Gene Expression Regulation, Neoplastic/drug effects, MESH: Tumor Burden/genetics, MESH: Xenograft Model Antitumor Assays, Bone Neoplasms, [SDV.CAN]Life Sciences [q-bio]/Cancer, 03 medical and health sciences, Cell Line, Tumor, osteosarcoma, medicine, Animals, Humans, Doxorubicin, MESH: DNA (Cytosine-5-)-Methyltransferase 1/antagonists & inhibitors, MESH: Enzyme Inhibitors/chemistry, Cisplatin, MESH: Humans, Nucleoside inhibitor, medicine.disease, MESH: Enzyme Inhibitors/pharmacology, Xenograft Model Antitumor Assays, MESH: Male, MESH: Enzyme Inhibitors/administration & dosage, MESH: Bone Neoplasms/drug therapy, 030104 developmental biology, MESH: Osteosarcoma/metabolism, Cancer research, cancer research, MESH: DNA (Cytosine-5-)-Methyltransferase 1/genetics, MESH: Benzamides/administration & dosage, MESH: Tumor Burden/drug effects
Abstract

The identification of new therapeutic strategies against osteosarcoma, the most common primary bone tumor, continues to be a primary goal to improve the outcomes of patients refractory to conventional chemotherapy. Osteosarcoma originates from the transformation of mesenchymal stem cells (MSC) and/or osteoblast progenitors, and the loss of differentiation is a common biological osteosarcoma feature, which has strong significance in predicting tumor aggressiveness. Thus, restoring differentiation through epigenetic reprogramming is potentially exploitable for therapeutic benefits. Here, we demonstrated that the novel nonnucleoside DNMT inhibitor (DNMTi) MC3343 affected tumor proliferation by blocking osteosarcoma cells in G1 or G2–M phases and induced osteoblastic differentiation through the specific reexpression of genes regulating this physiologic process. Although MC3343 has a similar antiproliferative effect as 5azadC, the conventional FDA-approved nucleoside inhibitor of DNA methylation, its effects on cell differentiation are distinct. Induction of the mature osteoblast phenotype coupled with a sustained cytostatic response was also confirmed in vivo when MC3343 was used against a patient-derived xenograft (PDX). In addition, MC3343 displayed synergistic effects with doxorubicin and cisplatin (CDDP), two major chemotherapeutic agents used to treat osteosarcoma. Specifically, MC3343 increased stable doxorubicin bonds to DNA, and combined treatment resulted in sustained DNA damage and increased cell death. Overall, this nonnucleoside DNMTi is an effective novel agent and is thus a potential therapeutic option for patients with osteosarcoma who respond poorly to preadjuvant chemotherapy. Mol Cancer Ther; 17(9); 1881–92. ©2018 AACR.

Published
2018

40. Shieldin complex promotes DNA end-joining and counters homologous recombination in BRCA1-null cells

Author

Stephen P. Jackson, Mareike Herzog, Alejandra Bruna, Luca Pellegrini, Violeta Serra, Mark J. O'Connor, Zhongwu Lai, Chloé Lescale, Jacqueline J.L. Jacobs, Fengtang Yang, Jonathan Lam, Matylda Sczaniecka-Clift, Abigail Shea, Carlos Caldas, Matthias Ostermaier, Gabriel Balmus, Julia Coates, Wenming Wei, Inge de Krijger, Yaron Galanty, Mukerrem Demir, Ludovic Deriano, Petra Beli, Domenic Pilger, Harveer Dev, Rimma Belotserkovskaya, Alistair Martin, Beiyuan Fu, Ting-Wei Will Chiang, Qian Wu, Dev, Harveer [0000-0003-2874-6894], Yang, Fengtang [0000-0002-3573-2354], Balmus, Gabriel [0000-0003-2872-4468], Serra, Violeta [0000-0001-6620-1065], Beli, Petra [0000-0001-9507-9820], Pellegrini, Luca [0000-0002-9300-497X], Deriano, Ludovic [0000-0002-9673-9525], Jacobs, Jacqueline JL [0000-0002-7704-4795], Jackson, Stephen P [0000-0001-9317-7937], Apollo - University of Cambridge Repository, Wellcome Trust/Cancer Research UK Gurdon Institute, University of Cambridge [UK] (CAM), Department of Biochemistry [Cambridge], Cambridge University Hospitals - NHS (CUH), Intégrité du génome, immunité et cancer - Genome integrity, Immunity and Cancer, Institut Pasteur [Paris] (IP), Netherlands Cancer Institute (NKI), Antoni van Leeuwenhoek Hospital, Cancer Research UK Cambridge Institute [Cambridge, Royaume-Uni] (CRUK), Institute of Molecular Biology (IMB), Johannes Gutenberg - Universität Mainz = Johannes Gutenberg University (JGU), Wellcome Trust Sanger Institute [Hinxton, UK], AstraZeneca US [Waltham, USA], AstraZeneca [Cambridge, UK], The Wellcome Trust Sanger Institute [Cambridge], Vall d'Hebron Institute of Oncology [Barcelone] (VHIO), Vall d'Hebron University Hospital [Barcelona], The SPJ lab is largely funded by a Cancer Research UK (CRUK) Program Grant, C6/A18796, and a Wellcome Trust (WT) Investigator Award, 206388/Z/17/Z. Core infrastructure funding was provided by CRUK grant C6946/A24843 and WT grant WT203144. S.P.J. receives a salary from the University of Cambridge. H.D. is funded by WT Clinical Fellowship 206721/Z/17/Z. TWC was supported by a Cambridge International Scholarship. D.P. is funded by Cancer Research UK studentship C6/A21454. The P.B. lab is supported by the Emmy Noether Program (BE 5342/1-1) from the German Research Foundation and a Marie Curie Career Integration Grant from the European Commission (630763). The L.P. lab is funded by the WT (investigator award 104641/Z/14/Z) and the Medical Research Council (project grant MR/N000161/1). The C.C. lab was supported with funding from CRUK. The J.J. lab was supported by the European Research Council grant ERC-StG 311565, The Dutch Cancer Society (KWF) grant KWF 10999, and the Netherlands Organization for Scientific Research (NWO) as part of the National Roadmap Large-scale Research Facilities of the Netherlands, Proteins@Work (project no. 184.032.201 to the Proteomics Facility of the Netherlands Cancer Institute). The L.D. lab is funded by the Institut Pasteur, the Institut National du Cancer (no. PLBIO16-181) and the European Research Council (starting grant agreement no. 310917). W.W. is part of the Pasteur–Paris University (PPU) International PhD program and this project received funding from the CNBG company, China. Q.W. is funded by the Wellcome Trust (200814/Z/16/Z ). The V.S. lab work was funded by the Instituto de Salud Carlos III (ISCIII), an initiative of the Spanish Ministry of Economy and Innovation partially supported by European Regional Development FEDER Funds (PI17-01080 to VS), the European Research Area-NET, Transcan-2 (AC15/00063), a non-commercial research agreement with AstraZeneca UK, and structural funds from the Agència de Gestió d’Ajuts Universitaris i de Recerca (AGAUR, 2017 SGR 540) and the Orozco Family. V.S. received a salary and travel support to C.C.’s lab from ISCIII (CP14/00228, MV15/00041) and the FERO Foundation., The authors thank all S.P.J. laboratory members for support and advice, and Cambridge colleagues N. Lawrence for OMX super-resolution microscopy support and R. Butler for help with computational image analyses and programming. The authors also thank S. Selivanova and S. Hough for help with plasmid amplification, sample preparation and tissue culture maintenance, K. Dry for extensive editorial assistance, F. Muñoz-Martinez for assistance with CRISPR–Cas9 knockout generation, L. Radu for assistance with protein purification, C. Lord (Institute of Cancer Research, London) for SUM149PT cells, D. Durocher (University of Toronto, Canada) for U2OS LacSceIII cells, F. Alt (Harvard University, USA) for CH12F3 cells and 53bp1 knockout CH12F3 cell clones, T. Honjo (Kyoto University, Japan) for permission to use the CH12F3 cell line, and J. Serrat in the Jacobs lab for technical assistance, Institut Pasteur [Paris], and Johannes Gutenberg - Universität Mainz (JGU)

Subjects
MESH: DNA Breaks, Double-Stranded, RAD51, Cell Cycle Proteins, Poly (ADP-Ribose) Polymerase Inhibitor, MESH: Recombinational DNA Repair, Mice, MESH: Animals, DNA Breaks, Double-Stranded, skin and connective tissue diseases, Cancer, Telomere-binding protein, Ovarian Neoplasms, MESH: Breast Neoplasms / metabolism, MESH: Telomere-Binding Proteins / metabolism, 3. Good health, Cell biology, MESH: HEK293 Cells, MESH: Proteins / genetics, MESH: Telomere-Binding Proteins / genetics, MESH: Tumor Suppressor p53-Binding Protein 1 / metabolism, MESH: Xenograft Model Antitumor Assays, Telomere-Binding Proteins, MESH: Ovarian Neoplasms / drug therapy, Bone Neoplasms, MESH: Ovarian Neoplasms / metabolism, Article, 03 medical and health sciences, MESH: Cell Cycle Proteins, MESH: Bone Neoplasms / metabolism, Humans, MESH: Osteosarcoma / metabolism, [SDV.GEN]Life Sciences [q-bio]/Genetics, MESH: Humans, MESH: Tumor Suppressor p53-Binding Protein 1 / genetics, Dose-Response Relationship, Drug, HEK 293 cells, Proteins, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, DNA, MESH: BRCA1 Protein / deficiency, 030104 developmental biology, Multiprotein Complexes, MESH: Mad2 Proteins / metabolism, MESH: Breast Neoplasms / genetics, MESH: Bone Neoplasms / drug therapy, Cisplatin, Homologous recombination, MESH: Osteosarcoma / genetics, MESH: Female, 0301 basic medicine, DNA End-Joining Repair, MESH: Proteins / metabolism, MESH: Dose-Response Relationship, Drug, chemistry.chemical_compound, MESH: Osteosarcoma / pathology, MESH: Breast Neoplasms / pathology, Homologous Recombination, Polymerase, MESH: Breast Neoplasms / drug therapy, Osteosarcoma, biology, Chemistry, BRCA1 Protein, DNA damage and repair, MESH: Poly(ADP-ribose) Polymerase Inhibitors / pharmacology, MESH: Bone Neoplasms / genetics, DNA-Binding Proteins, MESH: Bone Neoplasms / pathology, Mad2 Proteins, Female, MESH: Ovarian Neoplasms / genetics, Tumor Suppressor p53-Binding Protein 1, MESH: Cisplatin / pharmacology, MESH: Cell Line, Tumor, Lymphocytes, Null, [SDV.CAN]Life Sciences [q-bio]/Cancer, Breast Neoplasms, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, MESH: BRCA1 Protein / genetics, Poly(ADP-ribose) Polymerase Inhibitors, Cell Line, Tumor, MESH: Drug Resistance, Neoplasm* / genetics, MESH: Mad2 Proteins / genetics, MESH: Ovarian Neoplasms / pathology, Animals, MESH: Mice, MESH: Osteosarcoma / drug therapy, Oligonucleotide, Protective Devices, Recombinational DNA Repair, Cell Biology, MESH: Multiprotein Complexes, Xenograft Model Antitumor Assays, HEK293 Cells, Drug Resistance, Neoplasm, biology.protein, MESH: DNA End-Joining Repair, MESH: DNA-Binding Proteins
Abstract

International audience; BRCA1 deficiencies cause breast, ovarian, prostate and other cancers, and render tumours hypersensitive to poly(ADP-ribose) polymerase (PARP) inhibitors. To understand the resistance mechanisms, we conducted whole-genome CRISPR-Cas9 synthetic-viability/resistance screens in BRCA1-deficient breast cancer cells treated with PARP inhibitors. We identified two previously uncharacterized proteins, C20orf196 and FAM35A, whose inactivation confers strong PARP-inhibitor resistance. Mechanistically, we show that C20orf196 and FAM35A form a complex, 'Shieldin' (SHLD1/2), with FAM35A interacting with single-stranded DNA through its C-terminal oligonucleotide/oligosaccharide-binding fold region. We establish that Shieldin acts as the downstream effector of 53BP1/RIF1/MAD2L2 to promote DNA double-strand break (DSB) end-joining by restricting DSB resection and to counteract homologous recombination by antagonizing BRCA2/RAD51 loading in BRCA1-deficient cells. Notably, Shieldin inactivation further sensitizes BRCA1-deficient cells to cisplatin, suggesting how defining the SHLD1/2 status of BRCA1-deficient tumours might aid patient stratification and yield new treatment opportunities. Highlighting this potential, we document reduced SHLD1/2 expression in human breast cancers displaying intrinsic or acquired PARP-inhibitor resistance.

Published
2018

41. Fatty acid metabolism complements glycolysis in the selective regulatory T cell expansion during tumor growth

Author

Giuseppe Matarese, Nico Mitro, Massimiliano Pagani, Martina Severa, Chiara Focaccetti, Yu Wei, Eliana M. Coccia, Stefano Miacci, Silvia Piconese, Eleonora Timperi, Claudio Procaccini, Giuseppe Danilo Norata, Ilenia Pacella, Valeria Ranzani, Vincenzo Barnaba, Fabiana Rizzo, Grazisa Rossetti, Fabrizia Bonacina, Ezio Giorda, Deriggio Faicchia, Università degli Studi di Roma 'La Sapienza' = Sapienza University [Rome] (UNIROMA), Consiglio Nazionale delle Ricerche [Napoli] (CNR), Istituto Superiore di Sanità (ISS), Università degli Studi di Milano = University of Milan (UNIMI), Curtin University [Perth], Planning and Transport Research Centre (PATREC), Istituto Nazionale Genetica Molecolare [Milano] (INGM), Istituto di Ricovero e Cura a Carattere Scientifico (IRCCS), Pathogenèse des Virus de l'Hépatite B (PVHB), Institut Pasteur [Paris] (IP)-Institut National de la Santé et de la Recherche Médicale (INSERM), University of Naples Federico II = Università degli studi di Napoli Federico II, Institut Pasteur, Fondation Cenci Bolognetti - Istituto Pasteur Italia, Fondazione Cenci Bolognetti, Réseau International des Instituts Pasteur (RIIP), Istituto Italiano di Tecnologia (IIT), This work was supported by Associazione Italiana per la Ricerca sul Cancro Grants IG-2014 15199 and IG-2017 19939 (to V.B.) and IG-2017 19784 (to S.P.), Ministry of Education, University and Research (MIUR) Grants RF-2010-2310438 and RF 2010-2318269, Fondazione Italiana Sclerosi Multipla Grants code 2015/R/04 (to V.B.), code 2013/R/9 (to E.M.C.), code 2016/R/18 (to G.M.), and code 2014/R/19 (to M.S.), MIUR Progetti di Ricerca di Interesse Nazionale Grant 2010-2011 protocol 2010LC747T_004, MIUR Fondo per gli Investimenti della Ricerca di Base 2011/13 Grant no. RBAP10TPXK, Istituto Pasteur Italia–Fondazione Cenci Bolognetti Grant 2014-2016, International Network Institut Pasteur Programmes Transversaux de Recherche Grant 20-16, Fondazione Roma Grant for Biomedical Research NCDS-2013-000000345, the European Union’s Seventh Framework Program (FP7) under Grant Agreement 259743 (MODHEP consortium) (to Y.W.), Fondazione Cariplo Grant 2016-0852 (to G.D.N.), Ministero della Salute Grants GR-2011-02346974 (to G.D.N.) and GR-2013-02355011 (to F.B.), and European Union IDEAS Programme European Research Council Starting Grant menTORingTregs 310496 and Telethon Grant GGP17086 (to G.M.). C.F. was supported by a 2015 Fellowship from Fondazione Veronesi., European Project: 259743,EC:FP7:HEALTH,FP7-HEALTH-2010-two-stage,MODHEP(2011), European Project: 310496,EC:FP7:ERC,ERC-2012-StG_20111109,MENTORINGTREGS(2013), Pacella, Ilenia, Procaccini, Claudio, Focaccetti, Chiara, Miacci, Stefano, Timperi, Eleonora, Faicchia, Deriggio, Severa, Martina, Rizzo, Fabiana, Coccia, Eliana Marina, Bonacina, Fabrizia, Mitro, Nico, Norata, Giuseppe Danilo, Rossetti, Grazisa, Ranzani, Valeria, Pagani, Massimiliano, Giorda, Ezio, Wei, Yu, Matarese, Giuseppe, Barnaba, Vincenzo, Piconese, Silvia, Istituto Superiore di Sanita [Rome], Systems biology of liver cancer: an integrative genomic-epigenomic approach - MODHEP - - EC:FP7:HEALTH2011-01-01 - 2016-06-30 - 259743 - VALID, Unravelling paradoxes in regulatory T cell biology: the molecular basis for an mTOR-dependent oscillatory metabolic switch controlling immune tolerance and autoimmunity - MENTORINGTREGS - - EC:FP7:ERC2013-05-01 - 2018-04-30 - 310496 - VALID, Università degli Studi di Roma 'La Sapienza' = Sapienza University [Rome], Università degli Studi di Milano [Milano] (UNIMI), Institut Pasteur [Paris]-Institut National de la Santé et de la Recherche Médicale (INSERM), and Università degli studi di Napoli Federico II

Subjects
0301 basic medicine, MESH: Neoplasm Proteins, MESH: Oxidation-Reduction, Treg, fatty acid synthesis, glycolysis, ox40, tumor microenvironment, Glycolysi, T-Lymphocytes, Type I, Settore MED/04, T-Lymphocytes, Regulatory, Transgenic, chemistry.chemical_compound, Mice, 0302 clinical medicine, Neoplasms, MESH: Tumor Microenvironment, Glycolysis, MESH: Animals, Tumor, Multidisciplinary, Chemistry, Fatty Acids, hemic and immune systems, Regulatory, Cell biology, Neoplasm Proteins, MESH: Fatty Acids, Fatty Acid Synthase, Type I, medicine.anatomical_structure, MESH: Neoplasms, Experimental, PNAS Plus, Fatty Acid Synthase, 030220 oncology & carcinogenesis, MESH: Glycolysis, [SDV.IMM]Life Sciences [q-bio]/Immunology, MESH: Fatty Acid Synthase, Type I, Fatty acid synthesis, Ox40, Tumor microenvironment, Animals, Cell Line, Tumor, Humans, Mice, Transgenic, Neoplasms, Experimental, Oxidation-Reduction, Tumor Microenvironment, Intracellular, MESH: Cell Line, Tumor, [SDV.IMM] Life Sciences [q-bio]/Immunology, Regulatory T cell, MESH: Mice, Transgenic, T cell, chemical and pharmacologic phenomena, Cell Line, 03 medical and health sciences, Experimental, Fatty acid synthesi, medicine, MESH: Mice, MESH: Humans, Fatty acid metabolism, MESH: T-Lymphocytes, Regulatory, Lipid metabolism, 030104 developmental biology
Abstract

International audience; The tumor microenvironment restrains conventional T cell (Tconv) activation while facilitating the expansion of Tregs. Here we showed that Tregs' advantage in the tumor milieu relies on supplemental energetic routes involving lipid metabolism. In murine models, tumor-infiltrating Tregs displayed intracellular lipid accumulation, which was attributable to an increased rate of fatty acid (FA) synthesis. Since the relative advantage in glucose uptake may fuel FA synthesis in intratumoral Tregs, we demonstrated that both glycolytic and oxidative metabolism contribute to Tregs' expansion. We corroborated our data in human tumors showing that Tregs displayed a gene signature oriented toward glycolysis and lipid synthesis. Our data support a model in which signals from the tumor microenvironment induce a circuitry of glycolysis, FA synthesis, and oxidation that confers a preferential proliferative advantage to Tregs, whose targeting might represent a strategy for cancer treatment.

Published
2018

42. Multi-target-directed ligands for treating Alzheimer's disease: Butyrylcholinesterase inhibitors displaying antioxidant and neuroprotective activities

Author

Janko Kos, Anja Pišlar, Xavier Brazzolotto, Marko Jukič, Stanislav Gobec, Jacques-Philippe Colletier, Nicolas Coquelle, Simon Žakelj, Florian Nachon, Janez Mravljak, Damijan Knez, Matej Sova, Faculty of Pharmacy, University of Ljubljana, Institut de biologie structurale (IBS - UMR 5075 ), Centre National de la Recherche Scientifique (CNRS)-Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Direction de Recherche Fondamentale (CEA) (DRF (CEA)), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA), Toxicology, Centre de Recherches du Service de Santé des Armées, Institut de Recherche Biomédicale des Armées [Brétigny-sur-Orge] (IRBA), Université Grenoble Alpes [2016-2019] (UGA [2016-2019])-Institut de Recherche Interdisciplinaire de Grenoble (IRIG), Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Commissariat à l'énergie atomique et aux énergies alternatives (CEA)-Centre National de la Recherche Scientifique (CNRS), and Institut de Recherche Biomédicale des Armées (IRBA)

Subjects
0301 basic medicine, Models, Molecular, Antioxidant, DPPH, Multi-target-directed ligands, medicine.medical_treatment, MESH: Drug Design, Ligands, Antioxidants, chemistry.chemical_compound, 0302 clinical medicine, 8-hydroxyquinoline, Piperidines, Drug Discovery, MESH: Ligands, Butyrylcholinesterase, chemistry.chemical_classification, MESH: Butyrylcholinesterase, [SDV.BBM.BS]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Structural Biology [q-bio.BM], General Medicine, MESH: Neuroprotective Agents, Alzheimer's disease, MESH: Amyloid beta-Peptides, 3. Good health, MESH: Piperidines, Neuroprotective Agents, Biochemistry, Efflux, MESH: Caco-2 Cells, MESH: Cholinesterase Inhibitors, Intracellular, MESH: Models, Molecular, MESH: Cell Line, Tumor, Neuroprotection, 03 medical and health sciences, Alzheimer Disease, Cell Line, Tumor, medicine, Humans, Chelation, Pharmacology, Reactive oxygen species, MESH: Humans, Amyloid beta-Peptides, Organic Chemistry, MESH: Antioxidants, 030104 developmental biology, chemistry, Drug Design, Cholinesterase Inhibitors, Caco-2 Cells, 030217 neurology & neurosurgery, MESH: Alzheimer Disease
Abstract

International audience; The limited clinical efficacy of current symptomatic treatment and minute effect on progression of Alzheimer's disease has shifted the research focus from single targets towards multi-target-directed ligands. Here, a potent selective inhibitor of human butyrylcholinesterase was used as the starting point to develop a new series of multifunctional ligands. A focused library of derivatives was designed and synthesised that showed both butyrylcholinesterase inhibition and good antioxidant activity as determined by the DPPH assay. The crystal structure of compound 11 in complex with butyrylcholinesterase revealed the molecular basis for its low nanomolar inhibition of butyrylcholinesterase (Ki = 1.09 ± 0.12 nM). In addition, compounds 8 and 11 show metal-chelating properties, and reduce the redox activity of chelated Cu2+ ions in a Cu-ascorbate redox system. Compounds 8 and 11 decrease intracellular levels of reactive oxygen species, and are not substrates of the active efflux transport system, as determined in Caco2 cells. Compound 11 also protects neuroblastoma SH-SY5Y cells from toxic Aβ1-42 species. These data indicate that compounds 8 and 11 are promising multifunctional lead ligands for treatment of Alzheimer's disease.

Published
2018

43. Functional role of Kv1.1 and Kv1.3 channels in the neoplastic progression steps of three cancer cell lines, elucidated by scorpion peptides

Author

Wassim Moslah, Soumaya Souid, Jed Jebali, Dorra Aissaoui, Houcemeddine Othman, Mohamed Elayeb, Saoussen Mlayah-Bellalouna, José Luis, Zaineb Abdelkafi-Koubaa, Khadija Essafi-Benkhadir, Naziha Marrakchi, Najet Srairi-Abid, Laboratoire des Venins et Biomolécules Thérapeutiques - Laboratory of Venoms and Therapeutic Biomolecules (LR11IPT08), Institut Pasteur de Tunis, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP), Institut National des Sciences Appliquées et de Technologie - Carthage (INSAT Carthage), Université de Tunis El Manar (UTM), Laboratoire d'Epidémiologie Moléculaire et de Pathologie Expérimentale Appliquée aux Maladies Infectieuses (LR11IPT04), Université de Tunis El Manar (UTM)-Institut Pasteur de Tunis, Institut de neurophysiopathologie (INP), Aix Marseille Université (AMU)-Centre National de la Recherche Scientifique (CNRS), This work was supported by the Tunisian Ministry of Higher Education and Scientific Research (LR11IPT08) and the research grant Partenariat Hubert Curien-Utique (PHC-Utique) [code 17G 0811]., We would like to thank Daniel Lafitte and Claude Villard (INSERM UMR 911-CRO2. Faculty of Pharmacy, Marseille, France) for mass spectrometry studies, Maram Morjan, Thouraya Chagour and Amene Allah Ellafi (LVMT, Pasteur Institute of Tunis) for their technical help., and Centre National de la Recherche Scientifique (CNRS)-Aix Marseille Université (AMU)

Subjects
0301 basic medicine, Carcinogenesis, MESH: Scorpions/chemistry, [SDV]Life Sciences [q-bio], Proliferation, Venom, MESH: Neoplasms/drug therapy, Biochemistry, MESH: Peptides/pharmacology, 0302 clinical medicine, MESH: Cell Proliferation/drug effects, Structural Biology, Neoplasms, Neoplastic progression, MESH: Animals, U87, MESH: Potassium Channel Blockers/pharmacology, MESH: Kv1.3 Potassium Channel/genetics, Cancer, Kv1.3 Potassium Channel, biology, Chemistry, General Medicine, MESH: Shaker Superfamily of Potassium Channels/genetics, 3. Good health, Gene Expression Regulation, Neoplastic, 030220 oncology & carcinogenesis, Scorpion peptide, biological phenomena, cell phenomena, and immunity, MESH: Cell Line, Tumor, MESH: Carcinogenesis/drug effects, Scorpion, MESH: Gene Expression Regulation, Neoplastic/drug effects, Scorpion Venoms, Adhesion and migration, [SDV.CAN]Life Sciences [q-bio]/Cancer, [SDV.BC]Life Sciences [q-bio]/Cellular Biology, complex mixtures, Scorpions, MESH: Peptides/chemistry, 03 medical and health sciences, Breast cancer, biology.animal, Cell Line, Tumor, Homologous chromosome, medicine, Potassium Channel Blockers, Animals, Humans, natural sciences, [SDV.BBM]Life Sciences [q-bio]/Biochemistry, Molecular Biology, Amino Acid Sequence, Potassium channel subtypes, Molecular Biology, Cell Proliferation, MESH: Amino Acid Sequence/genetics, MESH: Potassium Channel Blockers/chemistry, MESH: Humans, MESH: Scorpion Venoms/pharmacology, urogenital system, MESH: Potassium/metabolism, medicine.disease, 030104 developmental biology, nervous system, Cell culture, MESH: Scorpion Venoms/chemistry, Cancer research, Potassium, Shaker Superfamily of Potassium Channels, MESH: Neoplasms/genetics, Peptides
Abstract

International audience; Voltage-gated potassium (Kv) channels are known to play a pivotal role in the progression of various cancer types and considered as new targets for designing anti-cancer therapy. However, the fact that many Kv channels are expressed in different cell lines makes it difficult to ascribe a functional role for a given Kv channel on a specific aspect of the tumorogenesis. In this work, we showed that although both Kv1.1 and Kv1.3 channels are expressed in U87 (glioblastoma), MDA-MB-231 (breast cancer) and LS174 (colon adenocarcinoma) cells, these respond differently to KAaH1 or KAaH2, two homologous Kv1 blockers from scorpion venom. KAaH1 is active on Kv1.1 and Kv1.3 and was found to inhibit migration and adhesion of U87 cells whereas KAaH2 which is slightly active only on Kv1.1 channel, inhibits their proliferation via the EGFR signaling pathway. The correlation between the electro-physiological activity of the scorpion peptides and their anti-migratory effects suggests the involvement of the Kv1.1 and Kv1.3 channels in the mobility of the three cancer cell lines. Our results showed that besides they can elucidate the implication of Kv1.1 and Kv1.3 channels in molecular mechanisms of neoplastic progression, KAaH1 and KAaH2 may be used as therapeutic tools against glioblastoma.

Published
2018

44. Macrophage scavenger receptor SR-AI contributes to the clearance of von Willebrand factor

Author

Wohner, Nikolett, Muczynski, Vincent, Mohamadi, Amel, Legendre, Paulette, Proulle, Valérie, Ayme, Gabriel, Christophe, Olivier, Lenting, Peter, Denis, Cécile, Casari, Caterina, Institut National de la Santé et de la Recherche Médicale (INSERM), Université Paris-Sud - Paris 11 (UP11), AP-HP Hôpital Bicêtre (Le Kremlin-Bicêtre), and AYME, Gabriel

Subjects
[SDV.SP.MED] Life Sciences [q-bio]/Pharmaceutical sciences/Medication, MESH: Cell Line, Tumor, [SDV.BIO]Life Sciences [q-bio]/Biotechnology, [SDV.IMM] Life Sciences [q-bio]/Immunology, [SDV.BBM.BM] Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, [SDV.SP.PG] Life Sciences [q-bio]/Pharmaceutical sciences/Galenic pharmacology, Article, Cell Line, Mice, MESH: Mutant Proteins, [SDV.SP.MED]Life Sciences [q-bio]/Pharmaceutical sciences/Medication, Cell Line, Tumor, von Willebrand Factor, Animals, Humans, MESH: Protein Binding, MESH: Animals, MESH: von Willebrand Factor, MESH: Receptors, Scavenger, [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], MESH: Mice, Receptors, Scavenger, Hemostasis, MESH: Humans, Macrophages, Scavenger Receptors, Class A, MESH: Macrophages, [SDV.BBM.BM]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Molecular biology, [SDV.IMM.IMM]Life Sciences [q-bio]/Immunology/Immunotherapy, [SDV.BIO] Life Sciences [q-bio]/Biotechnology, MESH: Cell Line, MESH: Scavenger Receptors, Class A, [SDV.BBM.BC]Life Sciences [q-bio]/Biochemistry, Molecular Biology/Biomolecules [q-bio.BM], [SDV.SP.PG]Life Sciences [q-bio]/Pharmaceutical sciences/Galenic pharmacology, [SDV.SP.PHARMA] Life Sciences [q-bio]/Pharmaceutical sciences/Pharmacology, [SDV.SP.PHARMA]Life Sciences [q-bio]/Pharmaceutical sciences/Pharmacology, [SDV.IMM]Life Sciences [q-bio]/Immunology, Mutant Proteins, [SDV.IMM.IMM] Life Sciences [q-bio]/Immunology/Immunotherapy, Protein Binding
Abstract

International audience; Previously, we found that LDL-receptor related protein-1 on macrophages mediated shear stress-dependent clearance of von Willebrand factor. In control experiments, however, we observed that von Willebrand factor also binds to macrophages independently of this receptor under static conditions, suggesting the existence of additional clearance-receptors. In search for such receptors, we focused on the macrophage-specific scavenger-receptor SR-AI. von Willebrand factor displays efficient binding to SR-AI (half-maximum binding 14±5 nM). Binding is calcium-dependent and is inhibited by 72±4% in the combined presence of antibodies against the A1- and D4-domains. Association with SR-AI was confirmed in cell-binding experiments. In addition, binding to bone marrow-derived murine SR-AI-deficient macrophages was strongly reduced compared to binding to wild-type murine macrophages. Following expression via hydrodynamic gene transfer, we determined ratios for von Willebrand factor-propeptide over von Willebrand factor-antigen, a marker of von Willebrand factor clearance. Propeptide/antigen ratios were significantly reduced in SR-AI-deficient mice compared to wild-type mice (0.6±0.2 versus 1.3±0.3; P

Published
2018

45. Exploration of the role of the virulence factor ElrA during Enterococcus faecalis cell infection

Author

Stéphane Gaubert, Jean-Marie Herry, Romain Briandet, Julien Deschamps, Pascale Serror, Natalia Nunez, Yu Wei, Thomas Baranek, Mustapha Si-Tahar, Aurélie Derré-Bobillot, Cristel Archambaud, Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), MICrobiologie de l'ALImentation au Service de la Santé (MICALIS), Institut National de la Recherche Agronomique (INRA)-AgroParisTech, Département de Virologie - Department of Virology, Institut Pasteur [Paris] (IP), Centre d’Etude des Pathologies Respiratoires (CEPR), UMR 1100 (CEPR), Université de Tours (UT)-Institut National de la Santé et de la Recherche Médicale (INSERM), N.N. was supported by a fellowship from the French Ministère de l’Enseignement Supérieur et de la Recherche. P.S. thanks the French-Brazilian USP COFECUB project Uc Me 147/13 for traveling support., Institut Pasteur [Paris], AgroParisTech-Université Paris-Saclay-Institut National de Recherche pour l’Agriculture, l’Alimentation et l’Environnement (INRAE), Institut Pasteur de Shanghai, Académie des Sciences de Chine - Chinese Academy of Sciences (IPS-CAS), Réseau International des Instituts Pasteur (RIIP), Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Tours, and Wei, Yu

Subjects
0301 basic medicine, [SDV]Life Sciences [q-bio], Cell, lcsh:Medicine, MESH: Virulence, Virulence factor, Mice, Enterococcus faecalis, MESH: RAW 264.7 Cells, Macrophage, MESH: Animals, lcsh:Science, MESH: Bacterial Proteins, Multidisciplinary, Virulence, biology, Hep G2 Cells, medicine.anatomical_structure, MESH: Caco-2 Cells, MESH: Enterococcus faecalis, MESH: Cell Line, Tumor, Virulence Factors, 030106 microbiology, MESH: Hep G2 Cells, Article, Cell Line, Microbiology, 03 medical and health sciences, Bacterial Proteins, Leucine, Cell Line, Tumor, medicine, Extracellular, Animals, Humans, MESH: Staphylococcal Protein A, Staphylococcal Protein A, MESH: Mice, Gram-Positive Bacterial Infections, MESH: Virulence Factors, MESH: Humans, Macrophages, lcsh:R, MESH: Macrophages, biology.organism_classification, [SDV.MP.BAC]Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology, MESH: Cell Line, FHL2, RAW 264.7 Cells, MESH: Leucine, MESH: HeLa Cells, biology.protein, lcsh:Q, MESH: Gram-Positive Bacterial Infections, Caco-2 Cells, [SDV.MP.BAC] Life Sciences [q-bio]/Microbiology and Parasitology/Bacteriology, Protein A, HeLa Cells
Abstract

Enterococcus faecalis, an organism generally not pathogenic for healthy humans, has the potential to cause disease in susceptible hosts. While it seems to be equipped to interact with and circumvent host immune defense, most of the molecular and cellular mechanisms underlying the enterococcal infectious process remain elusive. Here, we investigated the role of the Enterococcal Leucine Rich protein A (ElrA), an internalin-like protein of E. faecalis also known as a virulence factor. ElrA was previously shown to prevent adhesion to macrophages. We show that ElrA does not inhibit the basic phagocytic process, but is able to prevent sensing and migration of macrophages toward E. faecalis. Presence or absence of FHL2, a eukaryotic partner of ElrA, does not affect the ElrA-dependent mechanism preventing macrophage migration. However, we highlight a partial contribution of FHL2 in ElrA-mediated virulence in vivo. Our results indicate that ElrA plays at least a dual role of which anti-phagocytic activity may contribute to dissemination of extracellular E. faecalis during infection.

Published
2018

46. Tyrosine-1 of RNA Polymerase II CTD Controls Global Termination of Gene Transcription in Mammals

Author

Amal Zine El Aabidine, Dirk Eick, David C. Martin, Cyril Esnault, Yousra Yahia, Jean-Christophe Andrau, Nilay Shah, Roland Schüller, Stefan Krebs, Tim-Michael Decker, Helmut Blum, Axel Imhof, Muhammad Ahmad Maqbool, Ignasi Forné, Institut de Génétique Moléculaire de Montpellier (IGMM), and Centre National de la Recherche Scientifique (CNRS)-Université de Montpellier (UM)

Subjects
0301 basic medicine, Transcription, Genetic, viruses, [SDV]Life Sciences [q-bio], RNA polymerase II, read-through transcription, MESH: Tyrosine, MESH: RNA, Small Nuclear, chemistry.chemical_compound, 0302 clinical medicine, divergent transcription, Transcription (biology), RNA polymerase, RNA, Small Nuclear, Promoter Regions, Genetic, biology, Chromatin, Cell biology, Tyrosine1, RNA splicing, MESH: Cell Line, Tumor, MESH: Mutation, promoter-proximal pausing, MESH: Chromatin, 03 medical and health sciences, MESH: Transcription Termination, Genetic, Cell Line, Tumor, MESH: Promoter Regions, Genetic, Humans, RNA, Messenger, Enhancer, Molecular Biology, Gene, transcription termination, MESH: RNA, Messenger, MESH: Humans, MESH: Transcription, Genetic, Promoter, Cell Biology, CTD, MESH: RNA Polymerase II, 030104 developmental biology, chemistry, Transcription Termination, Genetic, Mutation, biology.protein, Tyrosine, 030217 neurology & neurosurgery
Abstract

International audience; The carboxy-terminal domain (CTD) of RNA polymerase (Pol) II is composed of a repetition of YSPTSPS heptads and functions as a loading platform for protein complexes that regulate transcription, splicing, and maturation of RNAs. Here, we studied mammalian CTD mutants to analyze the function of tyrosine1 residues in the transcription cycle. Mutation of 3/4 of the tyrosine residues (YFFF mutant) resulted in a massive read-through transcription phenotype in the antisense direction of promoters as well as in the 3' direction several hundred kilobases downstream of genes. The YFFF mutant shows reduced Pol II at promoter-proximal pause sites, a loss of interaction with the Mediator and Integrator complexes, and impaired recruitment of these complexes to chromatin. Consistent with these observations, Pol II loading at enhancers and maturation of snRNAs are altered in the YFFF context genome-wide. We conclude that tyrosine1 residues of the CTD control termination of transcription by Pol II.

Published
2018

47. Lipase inhibitor orlistat prevents hepatitis B virus infection by targeting an early step in the virus life cycle

Author

Ulrike Protzer, Alma Zernecke, Jochen M. Wettengel, Almut Glinzer, Katrin Singethan, Knud Esser, Julie Lucifora, Technische Universität Munchen - Université Technique de Munich [Munich, Allemagne] (TUM), German Center for Infection Research, Partnersite Munich (DZIF), and Technische Universität München [München] (TUM)

Subjects
0301 basic medicine, MESH: Virus Internalization, medicine.disease_cause, MESH: Hepatocytes, MESH: Lipase, Mice, 0302 clinical medicine, [SDV.MHEP.MI]Life Sciences [q-bio]/Human health and pathology/Infectious diseases, MESH: Animals, Pathogen, MESH: Lipid Metabolism, Chemistry, virus diseases, Hepatitis B, MESH: Apolipoproteins E, MESH: Lipoproteins, 3. Good health, MESH: Hepatitis B virus, Lipase inhibitors, [SDV.MP.VIR]Life Sciences [q-bio]/Microbiology and Parasitology/Virology, 030211 gastroenterology & hepatology, MESH: Antiviral Agents, Hepatitis B virus, MESH: Cell Line, Tumor, Lipoproteins, Primary Cell Culture, Antiviral Agents, Virus, MESH: Primary Cell Culture, 03 medical and health sciences, Apolipoproteins E, Viral life cycle, Viral entry, Virology, Cell Line, Tumor, medicine, Animals, Humans, MESH: Mice, Pharmacology, Orlistat, MESH: Humans, MESH: Hepatitis B, [SDV.MHEP.HEG]Life Sciences [q-bio]/Human health and pathology/Hépatology and Gastroenterology, Lipase, Virus Internalization, Lipid Metabolism, digestive system diseases, MESH: Orlistat, 030104 developmental biology, Hepatocytes, [SDV.SP.PHARMA]Life Sciences [q-bio]/Pharmaceutical sciences/Pharmacology, Hepatic lipase, Lipoprotein
Abstract

International audience; Hepatitis B Virus (HBV) is a strictly hepatotropic pathogen which is very efficiently targeted to the liver and into its host cell, the hepatocyte. The sodium taurocholate co-transporting polypeptide (NTCP) has been identified as a key virus entry receptor, but the early steps in the virus life cycle are still only barely understood. Here, we investigated the effect of lipase inhibition and lipoprotein uptake on HBV infection using differentiated HepaRG cells and primary human hepatocytes. We found that an excess of triglyceride rich lipoprotein particles in vitro diminished HBV infection and a reduced hepatic virus uptake in vivo if apolipoprotein E is lacking indicating virus transport along with lipoproteins to target hepatocytes. Moreover, we showed that HBV infection of hepatocytes was inhibited by the broadly active lipase inhibitor orlistat, approved as a therapeutic agent which blocks neutral lipid hydrolysis activity. Orlistat treatment targets HBV infection at a post-entry step and inhibited HBV infection during virus inoculation strongly in a dose-dependent manner. In contrast, orlistat had no effect on HBV gene expression or replication or when added after HBV infection. Taken together, our data indicate that HBV connects to the hepatotropic lipoprotein metabolism and that inhibition of cellular hepatic lipase(s) may allow to target early steps of HBV infection.

Published
2018

48. ETV4 transcription factor and MMP13 metalloprotease are interplaying actors of breast tumorigenesis

Author

Dumortier, Mandy, Ladam, Franck, Damour, Isabelle, Vacher, Sophie, Bièche, Ivan, Marchand, Nathalie, De Launoit, Yvan, Tulasne, David, Chotteau-Lelièvre, Anne, de Launoit, Yvan, Mécanismes de la Tumorigénèse et Thérapies Ciblées - UMR 8161 (M3T), Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Institut National de la Santé et de la Recherche Médicale (INSERM)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), University of Massachusetts Medical School [Worcester] (UMASS), University of Massachusetts System (UMASS), Institut Curie [Paris], Institut de biologie de Lille - UMS 3702 (IBL), Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Université de Lille-Centre National de la Recherche Scientifique (CNRS), This work was supported by the Centre national de la recherche scientifique (CNRS), the Institut Pasteur de Lille, and Institut National de la Santé et de la Recherche Médicale (INSERM), as well as by grants from the Ligue contre le Cancer, comité Aisne and the Association pour la Recherche sur le Cancer., We thank Martine Duterque-Coquillaud, Anne Flourens, and Antonino Bongiovanni for their precious help in IHC experiments. We thank the Microscopy-Imaging-Cytometry Facility of the BioImaging Center Lille Nord-de-France for access to instruments and technical advice., Mécanismes de tumorigenèse et thérapies ciblées, Mécanismes de la Tumorigénèse et Thérapies Ciblées (M3T) - UMR 8161 (M3T), Institut Curie, Institut de biologie de Lille - IBL (IBLI), Université de Lille, Sciences et Technologies-Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Université de Lille, Droit et Santé-Centre National de la Recherche Scientifique (CNRS), Université de Lille-Institut Pasteur de Lille, Réseau International des Instituts Pasteur (RIIP)-Réseau International des Instituts Pasteur (RIIP)-Centre National de la Recherche Scientifique (CNRS), and Département de Génétique [Institut Curie, Paris] (Unité de Pharmacogénomique)

Subjects
Carcinogenesis, [SDV]Life Sciences [q-bio], Mice, Breast cancer, MESH: Aged, 80 and over, Cell Movement, MESH: Animals, ComputingMilieux_MISCELLANEOUS, MESH: Breast Neoplasms/pathology, Aged, 80 and over, MESH: Carcinogenesis/genetics, MESH: Aged, MESH: Middle Aged, MESH: Cell Movement/genetics, MMP13, MESH: Gene Expression Regulation, Neoplastic, Middle Aged, Prognosis, lcsh:Neoplasms. Tumors. Oncology. Including cancer and carcinogens, MESH: Neoplasm Invasiveness/genetics, MESH: Breast Neoplasms/genetics, Gene Expression Regulation, Neoplastic, [SDV] Life Sciences [q-bio], MESH: Proto-Oncogene Proteins/genetics, Female, Adenovirus E1A Proteins, MESH: Matrix Metalloproteinase 13/genetics, Research Article, Adult, MESH: Xenograft Model Antitumor Assays, MESH: Cell Line, Tumor, Breast Neoplasms, [SDV.CAN]Life Sciences [q-bio]/Cancer, lcsh:RC254-282, MESH: Prognosis, Cell Line, Tumor, Proto-Oncogene Proteins, Matrix Metalloproteinase 13, Animals, Humans, Neoplasm Invasiveness, MESH: Mice, Aged, Cell Proliferation, MESH: Humans, Proto-Oncogene Proteins c-ets, ETV4, MESH: Cell Proliferation/genetics, MESH: Adult, Xenograft Model Antitumor Assays, Tumorigenesis, MESH: Adenovirus E1A Proteins/genetics, MESH: Neoplasm Invasiveness/pathology, Transcription factor, MESH: Female
Abstract

Background The ETS transcription factor ETV4 is involved in the main steps of organogenesis and is also a significant mediator of tumorigenesis and metastasis, such as in breast cancer. Indeed, ETV4 is overexpressed in breast tumors and is associated with distant metastasis and poor prognosis. However, the cellular and molecular events regulated by this factor are still misunderstood. In mammary epithelial cells, ETV4 controls the expression of many genes, MMP13 among them. The aim of this study was to understand the function of MMP13 during ETV4-driven tumorigenesis. Methods Different constructs of the MMP13 gene promoter were used to study the direct regulation of MMP13 by ETV4. Moreover, cell proliferation, migration, invasion, anchorage-independent growth, and in vivo tumorigenicity were assayed using models of mammary epithelial and cancer cells in which the expression of MMP13 and/or ETV4 is modulated. Importantly, the expression of MMP13 and ETV4 messenger RNA was characterized in 456 breast cancer samples. Results Our results revealed that ETV4 promotes proliferation, migration, invasion, and anchorage-independent growth of the MMT mouse mammary tumorigenic cell line. By investigating molecular events downstream of ETV4, we found that MMP13, an extracellular metalloprotease, was an ETV4 target gene. By overexpressing or repressing MMP13, we showed that this metalloprotease contributes to proliferation, migration, and anchorage-independent clonogenicity. Furthermore, we demonstrated that MMP13 inhibition disturbs proliferation, migration, and invasion induced by ETV4 and participates to ETV4-induced tumor formation in immunodeficient mice. Finally, ETV4 and MMP13 co-overexpression is associated with poor prognosis in breast cancer. Conclusion MMP13 potentiates the effects of the ETV4 oncogene during breast cancer genesis and progression. Electronic supplementary material The online version of this article (10.1186/s13058-018-0992-0) contains supplementary material, which is available to authorized users.

Published
2018

49. 8-Aryl-6-chloro-3-nitro-2-(phenylsulfonylmethyl)imidazo[1,2-a] pyridines as potent antitrypanosomatid molecules bioactivated by type 1 nitroreductases

Author

Maxime D. Crozet, Alan H. Fairlamb, Geneviève Pratviel, Alexis Valentin, Pascal Rathelot, Alix Sournia-Saquet, Clotilde Boudot, Anita Cohen, Patrice Vanelle, Julien Pedron, Jean-Luc Stigliani, Sandra Bourgeade-Delmas, Susan Wyllie, Pierre Verhaeghe, Nadine Azas, Caroline Castera-Ducros, Cyril Fersing, Alain Moreau, Sébastien Hutter, Nicolas Primas, Bertrand Courtioux, Institut de Chimie Radicalaire (ICR), Aix Marseille Université (AMU)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Neuroépidémiologie Tropicale (NET), CHU Limoges-Institut d'Epidémiologie Neurologique et de Neurologie Tropicale-Institut National de la Santé et de la Recherche Médicale (INSERM)-Institut Génomique, Environnement, Immunité, Santé, Thérapeutique (GEIST), Université de Limoges (UNILIM)-Université de Limoges (UNILIM), Laboratoire de chimie de coordination (LCC), Institut National Polytechnique (Toulouse) (Toulouse INP), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Institut de Chimie de Toulouse (ICT-FR 2599), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche pour le Développement (IRD)-Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche pour le Développement (IRD)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS), Infections Parasitaires : Transmission, Physiopathologie et Thérapeutiques (IP-TPT), Service de Santé des Armées-Assistance Publique - Hôpitaux de Marseille (APHM)-Aix Marseille Université (AMU)-Institut de Recherche pour le Développement (IRD), Aix-Marseille Université - Faculté de pharmacie (AMU PHARM), Aix Marseille Université (AMU), Pharmacochimie et Biologie pour le Développement (PHARMA-DEV), Université Toulouse III - Paul Sabatier (UT3), Université Fédérale Toulouse Midi-Pyrénées-Université Fédérale Toulouse Midi-Pyrénées-Institut de Chimie de Toulouse (ICT-FR 2599), Université Fédérale Toulouse Midi-Pyrénées-Institut de Chimie du CNRS (INC)-Institut National Polytechnique (Toulouse) (Toulouse INP), Université Fédérale Toulouse Midi-Pyrénées-Centre National de la Recherche Scientifique (CNRS)-Institut de Recherche pour le Développement (IRD)-Institut de Chimie du CNRS (INC)-Institut de Recherche pour le Développement (IRD), Department of Biochemistry, Medical Sciences Institute, Dundee University, Dundee, Institut de Chimie de Toulouse (ICT), Institut de Recherche pour le Développement (IRD)-Université Toulouse III - Paul Sabatier (UT3), Université de Toulouse (UT)-Université de Toulouse (UT)-Institut de Chimie du CNRS (INC)-Centre National de la Recherche Scientifique (CNRS)-Institut National Polytechnique (Toulouse) (Toulouse INP), Université de Toulouse (UT)-Institut de Recherche pour le Développement (IRD)-Université Toulouse III - Paul Sabatier (UT3), Université de Toulouse (UT)-Centre National de la Recherche Scientifique (CNRS), Institut de Recherche pour le Développement (IRD)-Aix Marseille Université (AMU)-Assistance Publique - Hôpitaux de Marseille (APHM)-Service de Santé des Armées, Institut de Recherche pour le Développement (IRD)-Institut de Chimie de Toulouse (ICT), and Université de Toulouse (UT)

Subjects
Models, Molecular, 0301 basic medicine, SARs, Redox potentials, HepG2 cytotoxicity, Crystallography, X-Ray, MESH: Parasitic Sensitivity Tests, MESH: Dose-Response Relationship, Drug, chemistry.chemical_compound, MESH: Structure-Activity Relationship, Parasitic Sensitivity Tests, In vitro activity, Drug Discovery, Nitroheterocycles, Moiety, MESH: Trypanocidal Agents, Imidazo[1, Leishmania, 2-a]pyridine, Molecular Structure, MESH: Drug Screening Assays, Antitumor, Hep G2 Cells, General Medicine, Trypanocidal Agents, 3. Good health, MESH: Nitroreductases, MESH: Cell Survival, Imidazo[1,2-a]pyridine, Pharmacophore, MESH: Models, Molecular, Trypanosoma, MESH: Cell Line, Tumor, Cell Survival, Stereochemistry, Antiparasitic, medicine.drug_class, Trypanosoma brucei brucei, 030106 microbiology, MESH: Molecular Structure, Substituent, Antineoplastic Agents, MESH: Hep G2 Cells, Article, Structure-Activity Relationship, 03 medical and health sciences, Cell Line, Tumor, MESH: Cell Proliferation, medicine, Humans, Amastigote, Cell Proliferation, Pharmacology, MESH: Humans, MESH: Leishmania donovani, Dose-Response Relationship, Drug, Aryl, Organic Chemistry, MESH: Trypanosoma brucei brucei, Nitroreductases, MESH: Crystallography, X-Ray, 030104 developmental biology, chemistry, Nitro, Suzuki-Miyaura cross-coupling reaction, MESH: Antineoplastic Agents, [SDV.SPEE]Life Sciences [q-bio]/Santé publique et épidémiologie, Drug Screening Assays, Antitumor, Leishmania donovani, Fexinidazole
Abstract

International audience; Based on a previously identified antileishmanial 6,8-dibromo-3-nitroimidazo[1,2-a]pyridine derivative, a Suzuki-Miyaura coupling reaction at position 8 of the scaffold was studied and optimized from a 8-bromo-6-chloro-3-nitroimidazo[1,2-a]pyridine substrate. Twenty-one original derivatives were prepared, screened in vitro for activity against L. infantum axenic amastigotes and T. brucei brucei trypomastigotes and evaluated for their cytotoxicity on the HepG2 human cell line. Thus, 7 antileishmanial hit compounds were identified, displaying IC50 values in the 1.1-3 μM range. Compounds 13 and 23, the 2 most selective molecules (SI = >18 or >17) were additionally tested on both the promastigote and intramacrophage amastigote stages of L. donovani. The two molecules presented a good activity (IC50 = 1.2-1.3 μM) on the promastigote stage but only molecule 23, bearing a 4-pyridinyl substituent at position 8, was active on the intracellular amastigote stage, with a good IC50 value (2.3 μM), slightly lower than the one of miltefosine (IC50 = 4.3 μM). The antiparasitic screening also revealed 8 antitrypanosomal hit compounds, including 14 and 20, 2 very active (IC50 = 0.04-0.16 μM) and selective (SI = >313 to 550) molecules toward T. brucei brucei, in comparison with drug-candidate fexinidazole (IC50 = 0.6 & SI > 333) or reference drugs suramin and eflornithine (respective IC50 = 0.03 and 13.3 μM). Introducing an aryl moiety at position 8 of the scaffold quite significantly increased the antitrypanosomal activity of the pharmacophore. Antikinetoplastid molecules 13, 14, 20 and 23 were assessed for bioactivation by parasitic nitroreductases (either in L. donovani or in T. brucei brucei), using genetically modified parasite strains that over-express NTRs: all these molecules are substrates of type 1 nitroreductases (NTR1), such as those that are responsible for the bioactivation of fexinidazole. Reduction potentials measured for these 4 hit compounds were higher than that of fexinidazole (-0.83 V), ranging from -0.70 to -0.64 V.

Published
2018

50. Diindolylmethane and its halogenated derivatives induce protective autophagy in human prostate cancer cells via induction of the oncogenic protein AEG-1 and activation of AMP-activated protein kinase (AMPK)

Author

Hossam M. Draz, Alexander A. Goldberg, J. Thomas Sanderson, Stephen Safe, Vladimir I. Titorenko, Emma S. Tomlinson Guns, National Research Centre - NRC (EGYPT), Institut Armand Frappier (INRS-IAF), Institut National de la Recherche Scientifique [Québec] (INRS)-Réseau International des Instituts Pasteur (RIIP), McGill University = Université McGill [Montréal, Canada], Concordia University [Montreal], University of British Columbia (UBC), Texas A&M University [College Station], and This work was funded by an operating grant from the Canadian Institutes of Health Research (CIHR grant no. MOP-115019) to JTS, EG and SS. HD received a scholarship from the Fonds de Recherche du Québec - Santé (FRQS). All authors declare to have no conflicts of interest.

Subjects
Male, 0301 basic medicine, AMPK, MESH: Signal Transduction, Indoles, Halogenation, genetic structures, Carcinogenesis, [SDV]Life Sciences [q-bio], Apoptosis, AMP-Activated Protein Kinases, 0302 clinical medicine, AMP-activated protein kinase, MESH: RNA, Small Interfering, MESH: AMP-Activated Protein Kinases, RNA, Small Interfering, MESH: Indoles, Prostate cancer, biology, Chemistry, LNCaP, RNA-Binding Proteins, MESH: Carcinogenesis, 3. Good health, Cell biology, 030220 oncology & carcinogenesis, MESH: Cell Adhesion Molecules, AEG-1, Signal Transduction, Atg1, MESH: Cell Line, Tumor, Diindolylmethane, 03 medical and health sciences, Cell Line, Tumor, Autophagy, Humans, MESH: Autophagy, C42B, Protein kinase A, MESH: Halogenation, MESH: Humans, MESH: Apoptosis, Membrane Proteins, Prostatic Neoplasms, Cell Biology, MESH: Male, 030104 developmental biology, MESH: Prostatic Neoplasms, Cancer cell, biology.protein, sense organs, Cell Adhesion Molecules
Abstract

International audience; 3,3'-Diindolylmethane (DIM) and its synthetic halogenated derivatives 4,4'-Br2- and 7,7'-Cl2DIM (ring-DIMs) have recently been shown to induce protective autophagy in human prostate cancer cells. The mechanisms by which DIM and ring-DIMs induce autophagy have not been elucidated. As DIM is a mitochondrial ATP-synthase inhibitor, we hypothesized that DIM and ring-DIMs induce autophagy via alteration of intracellular AMP/ATP ratios and activation of AMP-activated protein kinase (AMPK) signaling in prostate cancer cells. We found that DIM and ring-DIMs induced autophagy was accompanied by increased autophagic vacuole formation and conversion of LC3BI to LC3BII in LNCaP and C42B human prostate cancer cells. DIM and ring-DIMs also induced AMPK, ULK-1 (unc-51-like autophagy activating kinase 1; Atg1) and acetyl-CoA carboxylase (ACC) phosphorylation in a time-dependent manner. DIM and the ring-DIMs time-dependently induced the oncogenic protein astrocyte-elevated gene 1 (AEG-1) in LNCaP and C42B cells. Downregulation of AEG-1 or AMPK inhibited DIM- and ring-DIM-induced autophagy. Pretreatment with ULK1 inhibitor MRT 67307 or siRNAs targeting either AEG-1 or AMPK potentiated the cytotoxicity of DIM and ring-DIMs. Interestingly, downregulation of AEG-1 induced senescence in cells treated with overtly cytotoxic concentrations of DIM or ring-DIMs and inhibited the onset of apoptosis in response to these compounds. In summary, we have identified a novel mechanism for DIM- and ring-DIM-induced protective autophagy, via induction of AEG-1 and subsequent activation of AMPK. Our findings could facilitate the development of novel drug therapies for prostate cancer that include selective autophagy inhibitors as adjuvants.

Published
2017

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