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Identification and characterization of novel antibacterial compounds using protozoan hosts

English title Identification and characterization of novel antibacterial compounds using protozoan hosts
Applicant Scapozza Leonardo
Number 130016
Funding scheme Sinergia
Research institution Laboratoire de Biochimie Pharmaceutique Section de Pharmacie Université de Genève
Institution of higher education University of Geneva - GE
Main discipline Experimental Microbiology
Start/End 01.09.2010 - 30.11.2013
Approved amount 1'638'377.00
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Keywords (10)

Bacterial virulence; Infection; 3R principles; Non-mammalian and mammalian models; Library design; Relational database; Pathogenesis; Microfluidic-microscopy systems; Macrophages; Anti-infectiva

Lay Summary (English)

Lead
Lay summary
Bacterial infections are a major public health problem worldwide. The threat is aggravated by the evolution of drug resistance, a paucity of new antibiotics, and the emergence of previously unrecognized infectious agents. For these reasons, the identification of novel antibacterial drugs is a pressing issue. The screening of small molecule chemical libraries is the strategy of choice to identify novel compounds. A detailed understanding of bacterial virulence and the development of powerful screening tools are prerequisites to identify this new type of drugs.The general aim of this project is to discover and characterize novel antibacterial compounds with the ultimate goal to be used to prevent or treat bacterial infections. We will focus primarily on compounds inhibiting bacterial virulence, and secondarily on compounds boosting host defence or acting as antibiotics. Our project focuses on the first steps of drug-discovery: the development of adequate screening tests, the isolation of hits and the early characterization of structure-activity relationships. For the primary screens, we will use amoebae as non-mammalian infection models because they are easy to handle, robust, and cost-effective. For the validation of the hits in a mammalian system, a macrophage-based microfluidic-microscopy system will be developed to study the interaction of immune phagocytes with the studied pathogens at the single cell level. The experimental screening will make use of a highly diverse low size chemical library, which targets pathways essential for bacterial virulence and host resistance, and which covers the chemical and biological space of the compounds available for screening. The results of this study will be deposited into an on-line relational database. The project will be carried out by a consortium of 5 groups covering the necessary expertises in experimental microbiology and pharmaceutical chemistry to explore this novel approach to develop new anti-bacterial drugs. Three of them belong to the University of Geneva (Prof. Scapozza, Prof. Cosson and Dr. Soldati), one is located at the EPFL (Prof. McKinney) and the fifth one is at Ludwig-Maximilians-University in Munich (Prof. Hilbi).The long-term vision is that compounds identified in this project will be taken forward into a lead optimization phase that may eventually produce novel clinically useful anti-infective compounds. Moreover, the compounds identified will serve as valuable tools to study cellular microbiology of human pathogens.
Direct link to Lay Summary Last update: 21.02.2013

Responsible applicant and co-applicants

Employees

Publications

Publication
Targeting RNA structure in SMN2 reverses spinal muscular atrophy molecular phenotypes
Garcia-Lopez Amparo, Tessaro Francesca, Jonker Hendrik R. A., Wacker Anna, Richter Christian, Comte Arnaud, Berntenis Nikolaos, Schmucki Roland, Hatje Klas, Petermann Olivier, Chiriano Gianpaolo, Perozzo Remo, Sciarra Daniel, Konieczny Piotr, Faustino Ignacio, Fournet Guy, Orozco Modesto, Artero Ruben, Metzger Friedrich, Ebeling Martin, Goekjian Peter, Joseph Benoît, Schwalbe Harald, Scapozza Leonardo (2018), Targeting RNA structure in SMN2 reverses spinal muscular atrophy molecular phenotypes, in Nature Communications, 9(1), 2032-2032.
Antimycobacterial drug discovery using Mycobacteria-infected amoebae identifies anti-infectives and new molecular targets
Trofimov Valentin, Kicka Sébastien, Mucaria Sabrina, Hanna Nabil, Ramon-Olayo Fernando, Del Peral Laura Vela-Gonzalez, Lelièvre Joël, Ballell Lluís, Scapozza Leonardo, Besra Gurdyal S., Cox Jonathan A. G., Soldati Thierry (2018), Antimycobacterial drug discovery using Mycobacteria-infected amoebae identifies anti-infectives and new molecular targets, in Scientific Reports, 8(1), 3939-3939.
Inhibitors of Mycobacterium marinum virulence identified in a Dictyostelium discoideum host model
Ouertatani-Sakouhi Hajer, Kicka Sébastien, Chiriano Gianpaolo, Harrison Christopher F., Hilbi Hubert, Scapozza Leonardo, Soldati Thierry, Cosson Pierre (2017), Inhibitors of Mycobacterium marinum virulence identified in a Dictyostelium discoideum host model, in PLOS ONE, 12(7), e0181121-e0181121.
A microfluidic cell-trapping device for single-cell tracking of host–microbe interactions
Delincé Matthieu J., Bureau Jean-Baptiste, López-Jiménez Ana Teresa, Cosson Pierre, Soldati Thierry, McKinney John D. (2016), A microfluidic cell-trapping device for single-cell tracking of host–microbe interactions, in Lab on a Chip, 16(17), 3276-3285.
A microfluidic cell-trapping device for single-cell tracking of host-microbe interactions.
Delincé Matthieu J, Bureau Jean-Baptiste, López-Jiménez Ana Teresa, Cosson Pierre, Soldati Thierry, McKinney John D (2016), A microfluidic cell-trapping device for single-cell tracking of host-microbe interactions., in Lab on a chip, 16(17), 3276-85.
In Silico Driven Design and Synthesis of Rhodanine Derivatives as Novel Antibacterials Targeting the Enoyl Reductase InhA.
Slepikas Liudas, Chiriano Gianpaolo, Perozzo Remo, Tardy Sébastien, Kranjc Agata, Patthey-Vuadens Ophélie, Ouertatani-Sakouhi Hajer, Kicka Sébastien, Harrison Christopher F, Scrignari Tiziana, Perron Karl, Hilbi Hubert, Soldati Thierry, Cosson Pierre, Tarasevicius Eduardas, Scapozza Leonardo (2016), In Silico Driven Design and Synthesis of Rhodanine Derivatives as Novel Antibacterials Targeting the Enoyl Reductase InhA., in Journal of medicinal chemistry, XXX.
Adrenergic antagonists restrict replication of Legionella.
Harrison Christopher F, Kicka Sébastien, Kranjc Agata, Finsel Ivo, Chiriano Gianpaolo, Ouertatani-Sakouhi Hajer, Soldati Thierry, Scapozza Leonardo, Hilbi Hubert (2015), Adrenergic antagonists restrict replication of Legionella., in Microbiology (Reading, England), 161(7), 1392-406.
Amoebae-Based Screening Reveals a Novel Family of Compounds Restricting Intracellular Legionella pneumophila.
Harrison Christopher F, Chiriano Gianpaolo, Finsel Ivo, Manske Christian, Hoffmann Christine, Steiner Bernhard, Kranjc Agata, Patthey-Vuadens Ophelie, Kicka Sébastien, Trofimov Valentin, Ouertatani-Sakouhi Hajer, Soldati Thierry, Scapozza Leonardo, Hilbi Hubert (2015), Amoebae-Based Screening Reveals a Novel Family of Compounds Restricting Intracellular Legionella pneumophila., in ACS infectious diseases, 1(7), 327-38.
Formation of a pathogen vacuole according to Legionella pneumophila: how to kill one bird with many stones.
Finsel Ivo, Hilbi Hubert (2015), Formation of a pathogen vacuole according to Legionella pneumophila: how to kill one bird with many stones., in Cellular microbiology, 17(7), 935-50.
Establishment and validation of whole-cell based fluorescence assays to identify anti-mycobacterial compounds using the Acanthamoeba castellanii-Mycobacterium marinum host-pathogen system.
Kicka Sébastien, Trofimov Valentin, Harrison Christopher, Ouertatani-Sakouhi Hajer, McKinney John, Scapozza Leonardo, Hilbi Hubert, Cosson Pierre, Soldati Thierry (2014), Establishment and validation of whole-cell based fluorescence assays to identify anti-mycobacterial compounds using the Acanthamoeba castellanii-Mycobacterium marinum host-pathogen system., in PloS one, 9(1), 87834-87834.
The natural alternative: protozoa as cellular models for Legionella infection.
Hoffmann Christine, Harrison Christopher F, Hilbi Hubert (2014), The natural alternative: protozoa as cellular models for Legionella infection., in Cellular microbiology, 16(1), 15-26.
Activation of Ran GTPase by a Legionella Effector Promotes Microtubule Polymerization, Pathogen Vacuole Motility and Infection
Rothmeier Eva, Pfaffinger Gudrun, Hoffmann Christine, Harrison Christopher F., Grabmayr Heinrich, Repnik Urška, Hannemann Mandy, Wölke Stefan, Bausch Andreas R., Griffiths Gareth, Müller-Taubenberger Annette, Itzen Aymelt, Hilbi Hubert (2013), Activation of Ran GTPase by a Legionella Effector Promotes Microtubule Polymerization, Pathogen Vacuole Motility and Infection, in PLoS Pathogens, 9(9), e1003598.
Dictyostelium ACAP-A is an ArfGAP involved in cytokinesis, cell migration and actin cytoskeleton dynamics.
Dias Marco, Blanc Cédric, Thazar-Poulot Nelcy, Ben Larbi Sabrina, Cosson Pierre, Letourneur François (2013), Dictyostelium ACAP-A is an ArfGAP involved in cytokinesis, cell migration and actin cytoskeleton dynamics., in Journal of cell science, 126(Pt 3), 756-66.
Dynamic persistence of antibiotic-stressed mycobacteria.
Wakamoto Yuichi, Dhar Neeraj, Chait Remy, Schneider Katrin, Signorino-Gelo François, Leibler Stanislas, McKinney John D (2013), Dynamic persistence of antibiotic-stressed mycobacteria., in Science (New York, N.Y.), 339(6115), 91-5.
Exploring Anti-Bacterial Compounds against Intracellular Legionella
Harrison Christopher F., Kicka Sébastien, Trofimov Valentin, Berschl Kathrin, Ouertatani-Sakouhi Hajer, Ackermann Nikolaus, Hedberg Christian, Cosson Pierre, Soldati Thierry, Hilbi Hubert (2013), Exploring Anti-Bacterial Compounds against Intracellular Legionella, in PLoS ONE, 8(9), e74813.
Setting up and monitoring an infection of Dictyostelium discoideum with mycobacteria.
Arafah Sonia, Kicka Sébastien, Trofimov Valentin, Hagedorn Monica, Andreu Nuria, Wiles Siouxsie, Robertson Brian, Soldati Thierry (2013), Setting up and monitoring an infection of Dictyostelium discoideum with mycobacteria., in Methods in molecular biology (Clifton, N.J.), 983, 403-17.
Single-cell dynamics of the chromosome replication and cell division cycles in mycobacteria.
Santi Isabella, Dhar Neeraj, Bousbaine Djenet, Wakamoto Yuichi, McKinney John D (2013), Single-cell dynamics of the chromosome replication and cell division cycles in mycobacteria., in Nature communications, 4, 2470-2470.
The Legionella effector RidL inhibits retrograde trafficking to promote intracellular replication.
Finsel Ivo, Ragaz Curdin, Hoffmann Christine, Harrison Christopher F, Weber Stephen, van Rahden Vanessa A, Johannes Ludger, Hilbi Hubert (2013), The Legionella effector RidL inhibits retrograde trafficking to promote intracellular replication., in Cell host & microbe, 14(1), 38-50.
Legionella spp. outdoors: colonization, communication and persistence.
Hilbi Hubert, Hoffmann Christine, Harrison Christopher F (2011), Legionella spp. outdoors: colonization, communication and persistence., in Environmental microbiology reports, 3(3), 286-96.

Collaboration

Group / person Country
Types of collaboration
Collaboration among the group involved in this project is continuing with regular meetings Switzerland (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Publication
- Research Infrastructure
Developmental Therapeutics Program, Division of Cancer Treatment and Diagnosis, NCI, NIH United States of America (North America)
- in-depth/constructive exchanges on approaches, methods or results
Prof. Dr. Herman Spaink, Universiteit Leiden, Faculty of Sciences, Institute of Biology Netherlands (Europe)
- in-depth/constructive exchanges on approaches, methods or results
- Research Infrastructure

Scientific events



Self-organised

Title Date Place
Half-day symposium of the NEMO network 18.05.2011 Geneva, Switzerland

Knowledge transfer events

Active participation

Title Type of contribution Date Place Persons involved
Industrial partner presentation at GlaxoSmithKline Talk 16.12.2013 Tres cantos, Spain McKinney John;
Discussion with Basilea Pharmaceutica Company Talk 27.11.2013 Basilea, Switzerland Chiriano Gianpaolo; Scapozza Leonardo;


Communication with the public

Communication Title Media Place Year
Talks/events/exhibitions Amoebae as model hosts to study mechanisms of mycobacteria infection and identify novel antitubercul International 2013

Awards

Title Year
The Swiss Chemical Society and Royal Society of Chemistry Award to attend and present this work at the “Anglo-Swiss Symposium on Phenotypic Screening” held at the Roche campus on the 9th of October 2013. 2013

Associated projects

Number Title Start Funding scheme
156945 Real-Time Single-Cell Analysis of Bacterial Persistence 01.10.2014 Project funding
169386 Cell-intrinsic Mechanisms of Sensing and Defence against Mycobacteria Infection 01.10.2016 Project funding
172951 Bactericidal mechanisms in phagocytic cells 01.05.2017 Project funding
189921 An in silico and chemo-biological approach to identify anti-infective and pro-metabolic natural products 01.04.2020 Sinergia
188813 Integrated Investigations of Cell-autonomous and Altruistic Defence Mechanisms against Mycobacteria Infection 01.10.2019 Project funding
153326 Molecular mechanisms ensuring recognition, ingestion and killing of bacteria by amoebae 01.05.2014 Project funding
149390 Dissection of Innate Host-Cell Defence Mechanisms against Mycobacteria Virulence 01.10.2013 Project funding
135789 Organization and function of the endocytic pathway: a genetic analysis 01.05.2011 Project funding
132995 Cellular and Molecular Mechanisms of Mycobacterial Virulence and Host Resistance 01.10.2010 Project funding
135639 Real-Time Single-Cell Analysis of Bacterial Persistence 01.08.2011 Project funding
120056 Interaction of phagocytic cells with bacteria: a genetic approach 01.05.2008 Project funding
153200 Formation of the Legionella-containing vacuole: Effectors targeting retrograde trafficking and microtubules 01.01.2015 Project funding
116309 Cellular and molecular mechanisms of phagocytosis, relevance to host-pathogen interaction 01.08.2007 Project funding
112592 Survival of Shigella and Legionella within phagocytes 01.08.2006 SNSF Professorships

Abstract

IDENTIFICATION AND CHARACTERIZATION OF NOVEL ANTIBACTERIAL COMPOUNDS USING PROTOZOAN HOSTSSUMMARYThe general aim of this project is to discover and characterize novel antibacterial compounds with the ultimate goal to be used to prevent or treat bacterial infections. We will focus primarily on compounds inhibiting bacterial virulence, and secondarily on compounds boosting host defence or acting as antibiotics. The endpoint of the project is the identification of antibacterial compounds with the characteristics of validated hits, namely concentration-dependent antibacterial activity, an IC50 < 10 uM, 2 log bacteria-host (protozoan and mammalian) selectivity and a first structure-activity relationship (SAR).Bacterial infections are a major public health problem worldwide. The threat is aggravated by the evolution of drug resistance, a paucity of new antibiotics, and the emergence of previously unrecognized infectious agents. For these reasons, the identification of novel antibacterial drugs is a pressing issue. The screening of small molecule chemical libraries is the strategy of choice to identify novel compounds. A detailed understanding of bacterial virulence and the development of powerful screening tools are prerequisites to identify this new type of drugs.In view of the overall failure of target-based screening campaigns, it is essential to develop novel phenotype-based screens such as the ones proposed here based on pathogen-host interactions. Academia should play a leading role in exploring this research avenue. Our project focuses on the first steps of drug-discovery: the development of adequate screening tests, the isolation of hits and the early characterization of structure-activity relationships. For the primary screens, we will use amoebae as non-mammalian infection models because they are easy to handle, robust, and cost-effective. Dictyostelium discoideum and/or Acanthamoeba castellanii will be used as hosts in combination with the following human relevant pathogens: i) Klebsiella pneumoniae, Pseudomonas aeruginosa (Subproject B), ii) Legionella pneumophila (Subproject C), iii) Mycobacterium spp. (Subproject D). For the validation of the hits resulting from the protozoan screens in a mammalian system, a macrophage-based microfluidic-microscopy system will be developed to study the interaction of immune phagocytes with the above-mentioned pathogens at the single cell level (Subproject E). The experimental screening will make use of a highly diverse low size (5000 compounds) chemical library, which targets pathways essential for bacterial virulence and host resistance, and which covers the chemical and biological space of the compounds available for screening. The library will be designed starting from 8 millions compounds using ligand-based virtual screening (Subproject A), combining the knowledge on ligands known from the literature to interact with relevant host-pathogen pathways and the information from the unique scaffolds covering the chemical space. The compounds annotated with molecular descriptors and the results from the phenotypic screening will be deposited into an on-line database (Shared Results Database) to ensure the flow of the results within the different partners of this Sinergia project and to allow data mining and SAR studies. Three of the participants of this application have long-standing experience with non-mammalian host models and one with mammalian phagocytes to study the virulence of different bacterial pathogens. One participant is a specialist in pharmaceutical chemistry with long-standing experience in virtual screening and library design. This Sinergia team represents the necessary expertise in experimental microbiology and pharmaceutical chemistry, and thus, is in an ideal position to explore this novel approach to develop new anti-bacterial drugs. The long-term vision is that compounds identified in this project will be taken forward into a lead optimization phase that may eventually produce novel clinically useful anti-infective compounds. Moreover, the compounds identified will serve as valuable tools to study cellular microbiology of human pathogens.The specific aims of this proposal are to1. Develop robust and simple assays to: i) screen for new antibacterial compounds using amoebae hosts and representative bacterial pathogens, and ii) perform real-time analysis of bacteria-phagocyte interactions. 2. Design and screen a highly diverse low size chemical library to identify novel antibacterial compounds with hit characteristics in protozoan systems, which are validated in mammalian systems.3. Perform data mining and characterize candidate antibacterial compounds mechanistically. Expected valueWe expect to provide a proof of concept on a series of points. First, we will demonstrate that phenotypic screening in a simple system allows cost-effective and rapid identification of primary antibacterial compounds. Second, we expect to efficiently filter out the usual frequent hitters that can otherwise only be eliminated in laborious cellular and animal tests, such as membrane-active and cytotoxic compounds. Third, we will explore the phylogenetic conservation of bacterial virulence and host resistance mechanisms and at the same time advance the cause of 3R-compliant research aiming at reducing the need for animal experiments. Fourth, we anticipate identifying not only antibacterial compounds but also “boosters” of the cell-intrinsic defence mechanisms. Fifth, we expect to identify compounds that will advance the cellular and molecular understanding of host-pathogen interactions.
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