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Leishmania parasites are the causative agents of diseases ranging from self-healing cutaneous lesions known as Oriental Sore to the lethal visceral leishmaniasis known as Kala-Azar. According to WHO figures, more than 12 million humans are currently infected, with 2 million new infections per year. Leishmaniasis exists in 88 countries on four continents and is a major, compounding complication in HIV infections.
The parasites are unicellular microorganisms that are transmitted to mammals, including humans, by blood-feeding sandflies. They infect and destroy the macrophages of the mammalian immune systems, thereby causing varying forms of immune pathologies.
No vaccination exists against Leishmania infections. Protection is afforded by insect repellents, appropriate clothing, and the use of repellent-treated bed nets. First line drugs against leishmaniasis have varying efficacy depending on the species and the endemic regions, requiring the development of new therapeutic strategies.
The temperature increase upon transmission to a mammal triggers the differentiation from the promastigote to the pathogenic stage, the amastigote. Heat shock proteins play a role both in the regulation of cell fate and the intracellular survival. We aim to analyse the functions of heat shock proteins HSP100, HSP90 and their co-chaperones, their interaction with protein export mechanisms and with signal transduction pathways. We employ strategies as diverse as proteome analysis, reverse genetics, functional cloning, and electron microscopy.
In the project "TranSig" (Trans-signalling), a collaboration with the Institut Pasteur, Paris, jointly funded by the French ANS and the German DFG, we investigate the interplay of parasite heat shock proteins and signal transducting protein kinases with the innate immune responses of the mammalian host, in particular the defence mechanisms of macrophages. Both heat shock proteins and parasite protein kinases are shuttled into the host cell cytoplasm and impact on the immune response. Inside the parasite, the phosphorylation of heat shock protein h has an impact on the viability and the ability to survive inside the host cells. Understanding these processes will likely reveal new targets for therapeutic approaches (Dr A. Hombach, Katharina Bartsch MSc).
Recently, we have turned our attention to the group of the small heat shock proteins (HSP23, P23, HSP20) and found that HSP23 is essential for temperature tolerance and parasite survival in a mammalian host (Dr. A. Hombach, Julia Eick, MSc). Current efforts focus on the functions of the CPN60 (HSP60) family of heat shock proteins, and the role in viability and virulence (Henner Zirpel, MSc).
The AG Clos was one of 13 partners in a European Research Project to develop and improve drugs against Leishmaniasis, Sleeping Sickness and Chagas Disease, which ended in 02/2017.
The major objectives of this 3-year project were: i) development of drug leads which may be used in combination with a known or an investigational drug, by using a common drug discovery platform established by experts in their respective fields, and ii) the development of pharmacodynamic biomarkers enabling the proteomic profiling of compound efficacy and early identification of drug resistance.
The new platform enables high throughput screening of compound libraries, lead to candidate drugs development, proof of concept testing, and toxicology and safety testing. NMTrypI will translate drug leads into drug candidates to enter the international drug development pipelines.
The infectious diseases burden imposed by the parasites of Trypanosomatidae family represents a huge problem in people’s lives in countries where these diseases are endemic. Problems associated with existing drugs include inefficient delivery, insufficient efficacy, excessive toxicity and increasing resistance. New drugs are urgently needed now and in the foreseeable future.
The New Medicines for Trypanosomatidic Infections (NMTrypI) consortium, which also includes non EU groups from disease-endemic countries (Brazil and Sudan) is funded by the European Commission 2013 initiative for Health Innovation for SME-guided activity and promotion, and will align alongside other on-going neglected disease programmes to take advantage of synergies to enlarge the European platform in neglected disease research.
Currently, 3 lead compounds for use against Leishmaniasis are in advanced stages of characterisation and verification. BNI is involved in the in vitro testing of leads, the identification of possible resistance risks, and the identification of lead drug targets in the parasites. State of the art technologies such as functional cloning in combination with Next Generation Sequencing (Cos-Seq) but also mass spectrometric protein analysis were established for the project, widening the experimental scope of the group (Dr E. Bifeld, Julia Eick MSc).
Antimony-based drugs are still the mainstay of chemotherapy against Leishmania infections in many endemic countries. Efficacy of antimonials has been compromised by increasing numbers of resistant infections, the basis of which is not fully understood and likely involves multiple factors. By using a functional cloning strategy he have recently identified a novel antimony resistance marker, ARM58, from the parasite Leishmania braziliensis that protects the parasites against antimony-based antileishmanial compounds. Here we show that the L. infantum homologue also confers resistance against antimony but not against other anti-leishmanial drugs, and that its function depends critically on one of four conserved domains of unknown function. This critical domain requires at least two hydrophobic amino acids and is predicted to form a transmembrane structure. Overexpression of ARM58 in antimony-exposed parasites reduces the intracellular Sb accumulation by over 70% indicating a role for ARM58 in Sb extrusion pathways, but without involvement of energy-dependent transporter proteins. The protein is found in the flagellum of the parasites, but its exact function is still subject of research (N.N.).
ARM58 Overexpression Reduces Intracellular Antimony Concentration in Leishmania infantum.
Schäfer, C., Tejera Nevado, P., Zander, D., and Clos, J.
Antimicrob. Agents Chemother. 2014, 58(3):1565.
The Hsp90–Sti1 interaction is critical for Leishmania donovani proliferation in both life cycle stages
Antje Hombach, Gabi Ommen, Mareike Chrobak, and Joachim Clos
Cellular Microbiology (2013) 15(4), 585–600
Geographical sequence variability in the Leishmania major virulence factor P46
Eugenia Bifeld, Mareike Chrobak, Gabi Schönian, Ulrike Schleicher, and Joachim Clos
Infect Genet Evol 30 (2015), 195-205
A small heat shock protein is essential for thermotolerance and intracellular survival of Leishmania donovani
Antje Hombach, Gabi Ommen, Andrea MacDonald, and Joachim Clos
J Cell Science 127 (2014), 4762-4773
A novel marker, ARM58, confers antimony resistance to Leishmania spp.
Andrea Nühs, Carola Schäfer, Dorothea Zander, Leona Trübe, Paloma Tejera Nevado, Sonja Schmidt, Jorge Arevalo, Vanessa Adaui, Louis Maes, Jean-Claude Dujardin, and Joachim Clos
Int J Parasitol: Drugs and Drug Resistance 4 (2014), 37-47.
Hsp90 Inhibitors radicicol and geldanamycin have opposing effects onLeishmaniaAha1-dependent proliferation
Bartsch K, Hombach-Barrigah A and Clos J.
Cell Stress and Chaperones, accepted
Methoxylated 2'-hydroxychalcones as antiparasitic hit compounds.
Borsari C, Santarem N, Torrado J, Olías AI, Corral MJ, Baptista C, Gul S, Wolf M, Kuzikov M, Ellinger B, Reinshagen J, Linciano P, Tait A, Costantino L, Freitas-Junior LH, Moraes CB, Pascoalino B, Alcântara LMaria, Franco CH, Bertolacini CD, Fontana V, Tejera Nevado P, Clos J, Alunda JM, Cordeiro-da-Silva A, Ferrari S, Costi MP
European Journal of Medicinal Chemistry, accepted
Leishmania donovani chaperonin 10 regulates parasite internalization and intracellular survival in human macrophages
Colineau L, Clos J, Moon KM, Foster LJ and Reiner NE
Med Microbiol Immunol 206, 235-257.
Characterization of the protein tyrosine phosphatase LmPRL-1 secreted by Leishmania major via the exosome pathway.
Leitherer S, Clos J, Liebler-Tenorio EM, Schleicher U, Bogdan C and Soulat D
Infect Immun - accepted
Leishmania Heat Shock Proteins as Effectors of Immune Evasion and Virulence
Bartsch K, Eick J, Zirpel H and Clos J
Current Immunology Reviews, 2017, 13 - accepted
A versatile qPCR assay to quantify trypanosomatidic infections of host cells and tissues.
Bifeld E, Tejera Nevado P, Bartsch J, Eick J, Clos J.
Med Microbiol Immunol. doi 10.1007/s00430-016-0460-3
Profiling of flavonol derivatives for the development of anti-trypanosomatidic drugs.
Borsari C, Luciani R, Pozzi C, Pohner I, Henrich S, Trande M, Cordeiro-da-Silva A, Santarem N, Baptista C, Tait A, Di Pisa F, Dello Iacono L, Landi G, Gul S, Wolf M, Kuzikov M, Ellinger B, Reinshagen J, Witt G, Gribbon P, Kohler M, Keminer O, Behrens B, Costantino L, Tejera Nevado P, Bifeld E, Eick J, Clos J, Torrado J, Jimenez-Anton MD, Corral MJ, Alunda JM, Pellati F, Wade RC, Ferrari S, Mangani S, Costi MP
J Med Chem. doi: 10.1021/acs.jmedchem.6b00698
Joining forces: first application of a rapamycin-induced dimerizable Cre system for conditional null mutant analysis in Leishmania.
Späth GF, Clos J
Mol Microbiol doi: 100:923-927.10.1111/mmi.13374
A Telomeric Cluster of Antimony Resistance Genes on Chromosome 34 of Leishmania infantum
Tejera Nevado P, Bifeld E, Höhn K, Clos J.
Antimicrob Agents Chemother. doi: 10.1128/AAC.00544-16
Phenotypic Characterization of a Leishmania donovani Cyclophyilin 40 Null Mutant.
Wai-Lok Y, Lambertz U, Colineau L, Pescher P, MacDonald A, Zander D, Retzlaff S, Eick J, Reiner NE, Clos J, Späth GF.
J Eukaryot Microbiol. doi: 10.1111/jeu.12329
Geographical sequence variation in the Leishmania major virulence factor P46
Eugenia Bifeld, Mareike Chrobak, Dorothea Zander, Ulrike Schleicher, Gabriele Schönian, Joachim Clos
Infection, Genetics and Evolution 30 (2015) 195–205
Heat Shock Proteins of Leishmania: Parasites in the Driver's Seat
Joachim Clos and Antje Hombach
Adak, S., Datta, R. (Eds.), Leishmania – Current Biology and Control. Caister Academic Press, Norfolk, United Kingdom, pp. 17-36.
Leishmania donovani P23 protects parasites against HSP90 inhibitor-mediated growth arrest
Hombach A, Ommen G, Sattler V, Clos J
Cell Stress & Chaperones 2015, 20, 673-685
The genetics of Leishmania virulence
Bifeld, E. and Clos, J.
Med Microbiol Immunol, 204, 619-634
Co-circulation of a novel phlebovirus and Massilia virus in sandflies, Portugal
Amaro, Fatima, Ze-Ze, Libia, Alves, Maria J., Borstler, Jessica, Clos, Joachim, Lorenzen, Stephan, Becker, Stefanie Christine, Schmidt-Chanasit, Jonas, and Cadar, Daniel
Virology journal 12, 174
A Novel Marker, ARM58, Confers Antimony Resistance to Leishmania spp
Nühs A, Schäfer C, Zander D, Trübe L, Tejera Nevado P, Schmidt S, Arevalo J, Adaui V, Maes L, Dujardin J-C, Clos J
Int. J. Parasitol: Drugs and Drug Resistance (2014) 4:37-47
ARM58 Overexpression Reduces Intracellular Antimony Concentration in Leishmania infantum
Schäfer C, Tejera Nevado P, Zander D, Clos J
Antimicrob Agents Chemother (2014) 58:1565–1574
No stress - Hsp90 and the signal transduction in Leishmania
Hombach A, Clos J
Parasitology (2014) 141, 1156–1166
Cyclophilin 40-deficient Leishmania donovani fail to undergo stress-induced development of the infectious metacyclic stage
Yau W-L, Pescher P, Macdonald A, Zander D, Retzlaff S, Blisnick T, Rotureau B, Bastin P, Clos J, Späth G
Cell. Microbiol. (2014) 93:80-97
Leishmania infantum EndoG is an endo/exo-nuclease essential for parasite survival
Rico E, Oliva C, Gutierrez KJ, Alzate JF, Genes CM, Moreno D, Casanova E, Gigante A, Perez-Perez MJ, Camarasa MJ, Clos J, Gago F, Jimenez-Ruiz A
PloS one (2014) 9:e89526
A small heat shock protein is essential for thermotolerance and intracellular survival of Leishmania donovani
Hombach A, Ommen G, MacDonald A, Clos J
J. Cell Science (2014) in press
The Hsp90-Sti1 Interaction is Critical for Leishmania donovani Proliferation in Both Life Cycle Stages
Hombach A, Ommen G, Chrobak M, Clos J
Cell Microbiol (2013) 15:585-600
The loss of virulence of histone H1 overexpressing Leishmania donovani parasites is directly associated with a reduction of HSP83 rate of translation
Alexandratos A, Clos J, Samiotaki M, Efstathiou A, Panayotou G, Soteriadou K, Smirlis D
Mol Microbiol (2013) 88:1015-1031
Leishmania donovani HslV does not interact stably with HslU proteins
Chrobak M, Forster S, Meisel S, Pfefferkorn R, Forster F, Clos J
Int J Parasitol (2012) 42:329-339
Secreted virulence factors and immune evasion in visceral leishmaniasis
Lambertz U, Silverman JM, Nandan D, McMaster WR, Clos J, Foster LJ, Reiner NE
Journal of Leukocyte Biology (2012) 91:887-899
Rapid identification of Leishmania spp. in formalin-fixed, paraffin-embedded tissue samples by fluorescence in situ hybridization
Frickmann H, Alnamar Y, Essig A, Clos J, Racz P, Barth TF, Hagen RM, Fischer M, Poppert S
TM & IH (2012) 17:1117-1126
Overexpression of a single Leishmania major gene is sufficient to enhance parasite infectivity in vivo and in vitro
Reiling L, Chrobak M, Schmetz C, Clos J
Mol Microbiol (2010) 76:1175-1190
Phosphoproteome dynamics reveals heat shock protein complexes specific to the Leishmania infectious stage
Morales M, Watanabe R, Dacher M, Chafey P, Osorio y Fortéa J, Beverley S, Ommen G, Clos J, Hem S, Lenormand P, Rousselle J-C, Namane A, Spath G
Proc Natl Acad Sci U S A (2010) 107:8381-8386
The co-chaperone SGT of Leishmania donovani is essential for the parasite's viability
Ommen G, Chrobak M, Clos J
Cell Stress and Chaperones (2010) 39:541-546
An exosome-based secretion pathway is responsible for protein export from Leishmania and communication with macrophages
Silverman JM, Clos J, de'Oliveira CC, Shirvani O, Fang Y, Wang C, Foster LJ, Reiner NE
J Cell Sci (2010) 123:842-852
Leishmania exosomes modulate innate and adaptive immune responses through effects on monocytes and dendritic cells
Silverman JM, Clos J, Horakova E, Wang AY, Wiesgigl M, Kelly I, Lynn MA, McMaster WR, Foster LJ, Levings MK, Reiner NE
J Immunol (2010) 185:5011-5022
LmxMPK4, an essential mitogen-activated protein kinase of Leishmania mexicana is phosphorylated and activated by the STE7-like protein kinase LmxMKK5
von Freyend SJ, Rosenqvist H, Fink A, Melzer IM, Clos J, Jensen ON, Wiese M
Int J Parasitol (2010) 40:969-978
Characterization of a Subunit of the Outer Dynein Arm Docking Complex Necessary for Correct Flagellar Assembly in Leishmania donovani
Harder S, Thiel M, Clos J, Bruchhaus I
PLoS Negl Trop Dis (2010) 4:e586
One-step generation of double-allele gene replacement mutants in Leishmania donovani
Ommen G, Lorenz S, Clos J
Int J Parasitol (2009) 39:541-546
Leishmania infantum expresses a mitochondrial nuclease homologous to EndoG that migrates to the nucleus in response to an apoptotic stimulus
Rico E, Alzate JF, Arias AA, Moreno D, Clos J, Gago F, Moreno I, Dominguez M, Jimenez-Ruiz A
Mol Biochem Parasitol (2009) 163:28-38
Identification of a Leishmania infantum gene mediating resistance to miltefosine and SbIII
Choudhury K, Zander D, Kube M, Reinhardt R, Clos J
Int J Parasitol (2008) 38:1411-1423
Leishmania major: identification of developmentally regulated proteins in procyclic and metacyclic promastigotes
Mojtahedi Z, Clos J, Kamali-Sarvestani E
Exp Parasitol (2008) 119:422-429
Structural characterization of beta-sheeted oligomers formed on the pathway of oxidative prion protein aggregation in vitro
Redecke L, von Bergen M, Clos J, Konarev PV, Svergun DI, Fittschen UE, Broekaert JA, Bruns O, Georgieva D, Mandelkow E, Genov N, Betzel C
J Struct Biol (2007) 157:308-320
Spontaneous recovery of pathogenicity by Leishmania major hsp100-/- alters the immune response in mice
Reiling L, Jacobs T, Kroemer M, Gaworski I, Graefe S, Clos J
Infect Immun (2006) 74:6027-6036
Functional cloning as a means to identify Leishmania genes involved in drug resistance
Clos J, Choudhury K
Mini Rev Med Chem (2006) 6:123-129
Complement C3 is required for the progression of cutaneous lesions and neutrophil attraction in Leishmania major infection
Jacobs T, Andra J, Gaworski I, Graefe S, Mellenthin K, Kromer M, Halter R, Borlak J, Clos J
Med Microbiol Immunol (2005) 194:143-149
Comparative analysis of the human and chicken prion protein copper binding regions at pH 6.5
Redecke L, Meyer-Klaucke W, Koker M, Clos J, Georgieva D, Genov N, Echner H, Kalbacher H, Perbandt M, Bredehorst R, Voelter W, Betzel C
J Biol Chem (2005) 280:13987-13992
Stage-specific expression of the mitochondrial co-chaperonin of Leishmania donovani, CPN10
Zamora-Veyl FB, Kroemer M, Zander D, Clos J
Kinetoplastid Biol Dis (2005) 4:3
Oligomerization of the proteolytic products is an intrinsic property of prion proteins
Georgieva D, Koker M, Redecke L, Perbandt M, Clos J, Bredehorst R, Genov N, Betzel C
Biochem Biophys Res Commun (2004) 323:1278-1286
Synthetic human prion protein octapeptide repeat binds to the proteinase K active site
Georgieva D, Rypniewski W, Echner H, Perbandt M, Koker M, Clos J, Redecke L, Bredehorst R, Voelter W, Genov N, Betzel C
Biochem Biophys Res Commun (2004) 325:1406-1411
A Leishmania donovani gene that confers accelerated recovery from stationary phase growth arrest
Hoyer C, Zander D, Fleischer S, Schilhabel M, Kroener M, Platzer M, Clos J
Int J Parasitol (2004) 34:803-811
Developmentally induced changes of the proteome in the protozoan parasite Leishmania donovani
Bente M, Harder S, Wiesgigl M, Heukeshoven J, Gelhaus C, Krause E, Clos J, Bruchhaus I
Proteomics (2003) 3:1811-1829
Comparison of the A2 gene locus in Leishmania donovani and Leishmania major and its control over cutaneous infection
Zhang WW, Mendez S, Ghosh A, Myler P, Ivens A, Clos J, Sacks DL, Matlashewski G
J Biol Chem (2003) 278:35508-35515
Inhibition of HSP90 in Trypanosoma cruzi Induces a Stress Response but No Stage Differentiation
Graefe SE, Wiesgigl M, Gaworski I, Macdonald A, Clos J
Eukaryot Cell (2002) 1:936-943
Heat Shock Protein 90 Homeostasis Controls Stage Differentiation in Leishmania donovani.
Wiesgigl M, Clos J
Mol Biol Cell (2001) 12:3307-3316.
Leishmania and the Leishmaniases: the heat shock protein 90 of Leishmania donovani.
Wiesgigl M, Clos J
Med. Microbiol. Immunol. (2001) 190:27-31
Leishmania and the Leishmaniases: Use of genetic complementation to identify gene(s) which specify species-specific organ tropism of Leishmania.
Hoyer C, Mellenthin K, Schilhabel M, Platzer M, Clos J
Med. Microbiol. Immunol. (2001) 190:53-56
Expression and Subcellular Localization of Cpn60 Protein Family Members in Leishmania donovani
Schlueter, A., M. Wiesgigl, C. Hoyer, S. Fleischer, L. Klaholz, C. Schmetz & J. Clos
Biochim. Biophys. Acta 2000 1491: 65-74
Cross-species Homologous Recombination in Leishmania donovani Reveals the Sites of Integration
Krobitsch, S. and J. Clos
Mol. Biochem. Parasitol. 2000 107: 123-128
Uniform distribution of transcription complexes on the clpB gene locus of Leishmania donovani
Wiesgigl, M. & J. Clos
Protist 1999 150: 369-373
A novel role for 100 kD heat shock proteins in the parasiteLeishmania donovani
Krobitsch, S., and Clos, J.
Cell Stress Chaperones 1999 4, 191-198
Leishmania donovani heat shock protein 100: characterization and function in amastigote stage differentiation
Krobitsch, S., Brandau, S., Hoyer, C., Schmetz, C., Hübel, A., and Clos, J.
J Biol Chem 1998 273, 6488-6494
Chemical stress does not induce heat shock protein synthesis in Leishmania donovani
Clos, J., S. Brandau & C. and Hoyer, C
Protist 1998 149: 167-172
Leishmania major Hsp100 is required chiefly in the mammalian stage of the parasite
Hubel, A., Krobitsch, S., Horauf, A., and Clos, J.
Mol Cell Biol 1997 17, 5987-5995
Transcription of the Leishmania major Hsp70-I gene locus does not proceed through the noncoding region
Dresel, A. & J. Clos
Exp Parasitol 1997 86: 206-212
The genomic organization of the HSP83 gene locus is conserved in three Leishmania species
Hubel, A. & J. Clos
Exp Parasitol 1996 82: 225-228
A member of the ClpB family of stress proteins is expressed during heat shock in Leishmania spp
Hubel, A., Brandau, S., Dresel, A., and Clos, J.
Mol Biochem Parasitol 1995 70, 107-118
High constitutive levels of heat-shock proteins in human-pathogenic parasites of the genus Leishmania
Brandau, S., Dresel, A., and Clos, J. (). .
Biochem J 1995 310, 225-232
pJC20 and pJC40- two high-copy-number vectors for T7 RNA polymerase-dependent expression of recombinant genes in Escherichia coli
Clos, J. & S. Brandau
Prot. Expression Purif. 1994 5: 133-137
From Sep 8-10, 2015, the AG Clos, in collaboration with the Fraunhofer IME, hosted the Scientific Meeting of the European Research Initiative "New Medicines for Trypanosomatidic Infections (NMTrypI)". 31 scientists from 13 partner institutions and four members of the Scientific Advisory Board met for 3 days to discuss the progress made in the discovery of new drugs against the Neglected Tropical Infectious Diseases caused by Trypanosomatidic protozoa, namely Leishmaniasis (Kala-Azar, Oriental Sore), Chagas Disease (American Trypanosomatiasis) and HAT (Human African Trypanosomiasis a.k.a. Sleeping Sickness). The Consortium is testing synthetic compounds produced by the partners, but also natural compounds from microorganisms and traditional African healing plants.