Parasitic Helminths

Drug Discovery in Infectious DiseasesSeries Editor Paul M. Selzer

Edited by Conor R. Caffrey

Targets, Screens, Drugs and Vaccines

Volume 3

57268File AttachmentCover.jpg

Edited by

Conor R. Caffrey

Parasitic Helminths

Titles of the Series ‘‘Drug Discovery in Infectious Diseases’’

Selzer, P. M. (ed.)

Antiparasitic and AntibacterialDrug Discovery

From Molecular Targets to Drug

Candidates

2009

ISBN: 978-3-527-32327-2

Forthcoming Topics of the Series

• Drug Discovery for Trypanosomatid Diseases

• Protein Phosphorylation in Parasites: Novel Targets for Antiparasitic Intervention

Related Titles

Gunn, A., Pitt, S. J.

Parasitology

An Integrated Approach

2012

ISBN: 978-0-470-68424-5

Scott, I., Sutherland, I.

Gastrointestinal Nematodes ofSheep and Cattle

Biology and Control

2009

ISBN: 978-1-4051-8582-0

Schwartz, E.

Tropical Diseases in Travelers

2009

ISBN: 978-1-4051-8441-0

Becker, K. (ed.)

Apicomplexan Parasites

Molecular Approaches toward Targeted

Drug Development

2011

ISBN: 978-3-527-32731-7

Edited by Conor R. Caffrey

Parasitic Helminths

Targets, Screens, Drugs and Vaccines

The Editors

Volume Editor:Dr. Conor R. CaffreySandler Center for Drug Discovery and Departmentof PathologyUniversity of California San FranciscoByers Hall 501E1700 4th StreetSan Francisco, CA 94158-2330USAConor.Caffrey@ucsf.edu

Series Editor:Prof Dr. Paul M. SelzerIntervet Innovation GmbHMSD Animal HealthMolecular Discovery SciencesZur Propstei55270 SchwabenheimGermanyPaul.Selzer@msd.de

CoverThe cover depicts a phylogenetic tree based on theligand binding regions of putative ligand-gated ionchannel genes. Nematode, platyhelminth, insect andvertebrate sequences are shown in shades of green,yellow, purple and red, respectively. Some C. elegansand human subunits are indicated and the labels forproteins involved in drug susceptibility to levamisole,monepantel and ivermectin are colored in cyan,orange and blue, respectively (see Rufener et al., PLoSPathogens (2010) 6(9):e1001091; courtesy of R.Kaminsky; see Chapter 17 for details). Left inset:freshly hatched Ascaridia galli larva (courtesy of M.Uphoff, Intervet Innovation GmbH, MSD AH; seeChapter 9 for details). Right inset: ribbon represen-tation of the crystal structure of the Schistosomamansoni Sm14 fatty acid binding protein in complexwith arachidonic acid that is shown in a space fillrepresentation (PDB 1VYG). The image was preparedby R. Marhöfer, Intervet Innovation GmbH, MSDAH and based on an original adapted from Angelucciet al., Biochemistry (2004) 43:13000-13011 that waskindly provided by M. Tendler (see Chapter 26 fordetails).

Limit of Liability/Disclaimer of Warranty: While thepublisher and author have used their best efforts inpreparing this book, they make no representations orwarranties with respect to the accuracy or complete-ness of the contents of this book and specificallydisclaim any implied warranties ofmerchantability orfitness for a particular purpose. No warranty can becreated or extended by sales representatives orwritten sales materials. The Advice and strategiescontained herein may not be suitable for your situa-tion. You should consult with a professional whereappropriate. Neither the publisher nor authors shallbe liable for any loss of profit or any other commercialdamages, including but not limited to special,incidental, consequential, or other damages.

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All rights reserved (including those of translation intoother languages). No part of this book may bereproduced in any form – by photoprinting, microfilm,or any othermeans – nor transmitted or translated intoa machine language without written permission fromthe publishers. Registered names, trademarks, etc.used in this book, even when not specifically markedas such, are not to be considered unprotected by law.

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Foreword to Parasitic Helminths: Targets, Screens,Drugs and Vaccines

Peter Hotez

The last decade has witnessed a renewed interest in neglected diseases caused byparasitic helminths, especially for the high prevalence gastrointestinal nematodeinfections, filarial infections, schistosomiasis, food-borne trematodiases and larvalcestode infections. A number of factors have contributed to this resurgent interest inhelminthic infections as global health threats:

1) There is new information suggesting that parasitic helminthiases are the mostcommon causes of infection among the bottom billion, i.e., the 1.4 billionworlds poorest peoplewho live below theWorldBankpoverty level in developingcountries of Asia, Africa, and the Americas. The major helminthiases include600–800 million people with one or more soil-transmitted helminth infection,400–600 million with schistosomiasis, more than 100 million people filarialinfections and tens of millions with food-borne trematode infections.

2) Additional studies have revealed that some of the most prevalent parasitichelminths may increase susceptibility to the big three diseases, i.e., HIV/AIDS, malaria and tuberculosis or exacerbate the morbidities of the big threediseases.

3) According to some estimates the major parasitic helminth infections togethercause a disease burdenmeasured in disability adjusted life years thatmay rival oreven exceed the big three conditions, while additional information indicates thatthese helminthiasesmay actually cause poverty through their deleterious effectson child growth and cognitive development, pregnancy outcome and agriculturalworker productivity.

The global health community has responded to this public health threat byexpanding efforts directed at mass drug administration (MDA). For example, usingeither diethylcarbamazine citrate or ivermectin together with albendazole, lymphaticfilariasis (LF) has been eliminated as a public health problem in more than20 countries, while through annual treatments with ivermectin, onchocerciasis hasbeen eliminated in Senegal andMali andmay soon be eliminated from theAmericas.Simultaneously, large scale financial support from the United States Agency forInternational Development (USAID), the British Department for InternationalDevelopment (DFID) and the non-profit Global Network for Neglected Tropical

jV

Diseases has facilitated combining LFand onchocerciasisMDAefforts withMDA forsoil-transmitted helminth infections and schistosomiasis to create rapid impactpackages of anthelmintic interventions in national programs of helminth control inmore than a dozen African countries, in addition to selected countries in Asia, LatinAmerica and the Caribbean.

The promise of MDA for parasitic helminth infections has generated excitementamong the international community that it might be possible to one day eliminateseveral helminthiases globally thereby achieving successes on this front that cannotyet be imagined for any of the big three diseases. However, there are warning signsthat MDA with currently available drugs might fail to achieve such expectations:1) high rates ofmebendazole drug failure have been reported for hookworm infectioncaused by Necator americanus and trichuriasis, i.e., two of the helminth infectionswith the greatest prevalence; 2) there is the looming specter of benzimidazole drugresistance among gastrointestinal nematodes of humans as has already occurred fornematode parasites of livestock, and 3) it has been shown that high rates of post-treatment re-infection occur for most of the major soil-transmitted helminth infec-tions, schistosomiasis, and opisthorchiasis and other food-borne trematodeinfections.

Such concerns highlight the urgent need to develop andmaintain a pipeline of newanthelmintic drugs in addition to anthelmintic vaccines to prevent infection or re-infection. Sadly, there is a glaring disconnect between the urgency for research anddevelopment (R&D) for new anthelmintic products and the global R&D budgetfor helminthiases. According to the global health think tank, Policy Cures, less than$100 million annually is spent on R&D for all human helminthiases compared tomore than $3 billion spent annually on R&D for all the other neglected infections,including the big three diseases.

This volume summarizes the work of dedicated investigators in the medical andveterinary fields who are applying the latest technologies to discover the nextgeneration of anthelmintic drugs and vaccines. Despite the difficulty in workingwith parasitic helminths in the laboratory, these investigators are overcomingsignificant hurdles in the study of the worlds most important helminths affectingmore than a billion people worldwide and countless livestock.

Their work is leading to a new generation of advances and represents the best inscience and in the pursuit of humanitarian goals.

Peter Hotez MD PhD is Dean of the National School of Tropical Medicine and Professor ofPediatrics and Molecular Virology & Microbiology, Texas Childrens Hospital and BaylorCollege of Medicine, Houston, Texas, USA

VIj Foreword to Parasitic Helminths: Targets, Screens, Drugs and Vaccines

Contents

Foreword to Parasitic Helminths: Targets, Screens, Drugs and Vaccines V

Preface XI

List of Contributors XIII

Part One Targets 1

1 Ligand-Gated Ion Channels as Targets for Anthelmintic Drugs: Past,Current, and Future Perspectives 3Kristin Lees�, Ann Sluder, Niroda Shannan, Lance Hammerland,and David Sattelle

2 How Relevant is Caenorhabditis elegans as a Model for the Analysisof Parasitic Nematode Biology? 23Lindy Holden-Dye� and Robert J. Walker

3 Integrating and Mining Helminth Genomes to Discoverand Prioritize Novel Therapeutic Targets 43Dhanasekaran Shanmugam, Stuart A. Ralph, Santiago J. Carmona,Gregory J. Crowther, David S. Roos, and Fernán Agüero�

4 Recent Progress in Transcriptomics of Key Gastrointestinal Nematodesof Animals – Fundamental Research Toward New InterventionStrategies 61Cinzia Cantacessi, Bronwyn E. Campbell, Aaron R. Jex, Ross S. Hall,Neil D. Young, Matthew J. Nolan, and Robin B. Gasser�

5 Harnessing Genomic Technologies to Explore the Molecular Biologyof Liver Flukes-Major Implications for Fundamental and Applied Research 73Neil D. Young�, Aaron R. Jex, Cinzia Cantacessi,Bronwyn E. Campbell, and Robin B. Gasser

VII

6 RNA Interference: A Potential Discovery Tool for TherapeuticTargets of Parasitic Nematodes 89Collette Britton

7 RNA Interference as a Tool for Drug Discovery in ParasiticFlatworms 105Akram A. Dadara and Patrick J. Skelly�

Part Two Screens 121

8 Mechanism-Based Screening Strategies for AnthelminticDiscovery 123Timothy G. Geary

9 Identification and Profiling of Nematicidal Compounds in VeterinaryParasitology 135Andreas Rohwer, Jürgen Lutz, Christophe Chassaing, Manfred Uphoff,Anja R. Heckeroth, and Paul M. Selzer�

10 Quantitative High-Content Screening-Based Drug Discovery againstHelmintic Diseases 159Rahul Singh

11 Use of Rodent Models in the Discovery of Novel Anthelmintics 181Rebecca Fankhauser, Linsey R. Cozzie, Bakela Nare, Kerrie Powell,Ann E. Sluder, and Lance G. Hammerland�

12 To Kill a Mocking Worm: Strategies to Improve Caenorhabditis elegansas a Model System for use in Anthelmintic Discovery 201Andrew R. Burns and Peter J. Roy�

Part Three Drugs 217

13 Anthelmintic Drugs: Tools and Shortcuts for the Long Road fromDiscovery to Product 219Eugenio L. de Hostos� and Tue Nguyen

14 Antinematodal Drugs – Modes of Action and Resistance: And WormsWill Not Come to Thee (Shakespeare: Cymbeline: IV, ii) 233Alan P. Robertson, Samuel K. Buxton, Sreekanth Puttachary,Sally M. Williamson, Adrian J. Wolstenholme, Cedric Neveu,Jacques Cabaret, Claude L. Charvet, and Richard J. Martin�

15 Drugs and Targets to Perturb the Symbiosis of Wolbachiaand Filarial Nematodes 251Mark J. Taylor�, Louise Ford, Achim Hoerauf, Ken Pfarr, Jeremy M. Foster,Sanjay Kumar, and Barton E. Slatko

VIII Contents

16 Promise of Bacillus thuringiensis Crystal Proteinsas Anthelmintics 267Yan Hu and Raffi V. Aroian�

17 Monepantel: From Discovery to Mode of Action 283Ronald Kaminsky� and Lucien Rufener

18 Discovery, Mode of Action, and Commercializationof Derquantel 297Debra J. Woods�, Steven J. Maeder, Alan P. Robertson,Richard J. Martin, Timothy G. Geary, David P. Thompson,Sandra S. Johnson, and George A. Conder

19 Praziquantel: Too Good to be Replaced? 309Livia Pica-Mattoccia� and Donato Cioli

20 Drug Discovery for Trematodiases: Challengesand Progress 323Conor R. Caffrey�, Jürg Utzinger, and Jennifer Keiser

Part Four Vaccines 341

21 Barefoot thru the Valley of Darkness: Preclinical Development of aHuman Hookworm Vaccine 343Jeffrey M. Bethony�, Maria Victoria Periago, and Amar R. Jariwala

22 Vaccines Linked to Chemotherapy: A New Approach to ControlHelminth Infections 357Sara Lustigman�, James H. McKerrow, and Maria Elena Bottazzi

23 Antifilarial Vaccine Development: Present and FutureApproaches 377Sara Lustigman�, David Abraham, and Thomas R. Klei

24 Proteases as Vaccines Against Gastrointestinal Nematode Parasitesof Sheep and Cattle 399David Knox

25 Schistosomiasis Vaccines – New Approaches to Antigen Discoveryand Promising New Candidates 421Alex Loukas�, Soraya Gaze, Mark Pearson, Denise Doolan,Philip Felgner, David Diemert, Donald P. McManus,Patrick Driguez, and Jeffrey Bethony

Contents IX

26 Sm14 Schistosoma mansoni Fatty Acid-Binding Protein:Molecular Basis for an Antihelminth Vaccine 435Miriam Tendler�, Celso Raul Romero Ramos,and Andrew J.G. Simpson

27 Mechanisms of Immune Modulation by Fasciola hepatica: Importancefor Vaccine Development and for Novel Immunotherapeutics 451Mark W. Robinson�, John P. Dalton, Sandra M. ONeill,and Sheila M. Donnelly

28 Prospects for Immunoprophylaxis Against Fasciola hepatica(Liver Fluke) 465Terry W. Spithill�, Carlos Carmona, David Piedrafita,and Peter M. Smooker

29 Vaccines Against Cestode Parasites 485Marshall W. Lightowlers�, Charles G. Gauci, Abdul Jabbar,and Cristian Alvarez

Index 505

X Contents

Preface

Parasitic helminths continue to plague the lives of billions of people, and those offarm and domestic animals. Their capacity to persist in the environment is infuri-ating, costly (health-wise and economically), and fascinating depending on onesperspective as a livestock farmer, medical provider, or research scientist. For theanimal health industry, the intrinsic capacity of helminths to resist drug pressuredrives the never-ending quest to bring new anthelmintic drugs to market. In recentyears, we have seen the fruits of that industry with the registration of new drugscontaining emodepside, monepantel, and derquantel. These drugs and other com-pounds in the pipeline are critical not just for staying ‘‘one up’’ on resistant parasitesof animals, but also for their potential to cross-over to human medicine, as hasoccurred with earlier anthelmintics that have been of immeasurable value toimproving global health. That contribution becomes all the more relevant todaygiven the increasing concerns over the continued efficacy of many first-generationanthelmintic drugs relied upon to treat human helminthiases, not least the benzi-midazoles and the ‘‘wonder drug’’ ivermectin, and the serious implications forpublic health should these drugs fail.

This volume is intended to showcase the state-of-the-art in the fields of drug andvaccine development for parasitic helminths as well as draw attention to thechallenges associated with bringing such products to market. The book is Volume3 in the series Drug Discovery in Infectious Diseases and expands on some of thethemes raised in Volume 1, Antiparasitic and Antibacterial Drug Discovery: FromMolecular Targets to Drug Candidates. Contributions from the animal health industryfigure prominently with a focus on the discovery and development of new chemicalentities. Importantly, however, the book also covers the increasingly relevant con-tribution of academia, not just in its traditional strengths of identifying new drugtargets or understanding how drug resistance arises, but also in the ways andmeansof preclinical and translational drug discovery through highly collaborative andinterdisciplinary research. Indeed, this exciting movement into the traditionaldomain of the pharmaceutical industry can be viewed as a natural consequence ofthe central importance and success of academia in the public–private consortia thatcurrently maintain dynamic drug development portfolios for other global parasiticdiseases such as malaria and the trypanosomatid diseases. The creativity andproductivity of academic scientists are highlighted in the many chapters covering

XI

the development and expansion of genomics and functional genomic tools, and theapplication of automated screening technologies to prosecute anthelmintic drugdiscovery with rigor.Finally, this volume discusses the need for, and the particular difficulties asso-

ciated with, developing anthelmintic vaccines for both humans and animals – formany the ‘‘holy grail’’ in providing the tool (including in combination with che-motherapy) to ultimately control and, hopefully, eliminate helminth diseases. Greatprogress has been made in identifying a number of candidates with proven efficacyin target animal species or that are now entering human trials, thanks in part to theestablishment of the necessary national and transnational institutional infrastruc-tures.To all of the authors, my sincere thanks for their time, insights, and patience in

contributing to an important collection of on-topic discussions.My thanks also to thebook series editor, Paul M. Selzer of Intervet Innovation GmbH, and to my collea-gues at the Sandler Center for Drug Discovery at the University of California SanFrancisco for their constructive input.

March 2012 Conor R. CaffreySan Francisco

XII Preface

List of Contributors

XIII

David AbrahamThomas Jefferson UniversityDepartment of Microbiology andImmunology233 South 10th StreetPhiladelphia, PA 19107USA

Fernán Agüero�

Universidad de San MartínInstituto de InvestigacionesBiotecnológicas25 de Mayo y FranciaSan MartínB 1650 HMP, Buenos AiresArgentinaE-mail: fernan@iib.unsam.edu.ar,fernan.aguero@gmail.com

Cristian AlvarezUniversity of MelbourneVeterinary Clinical Center250 Princes HighwayWerribeeVictoria 3030Australia

Raffi V. Aroian�

University of California San DiegoSection of Cell and DevelopmentalBiologyDivision of Biological Sciences9500 Gilman DrLa Jolla, CA 92093-0322USAE-mail: raroian@ucsd.edu

Jeffrey M. Bethony�

George Washington UniversityMedical CenterClinical Immunology LaboratoryDepartment of Microbiology,Immunology and Tropical Medicine2300 Eye Street, NWWashington, DC 20052USAE-mail: mtmjmb@gwumc.edu

and

FIOCRUZClinical Immunology LaboratoryLaboratório de Imunologia Celular eMolecularCentro de Pesquisas René RachouAv. Augusto de Lima 1715Belo HorizonteMinas Gerais 30190-002Brazil

Maria Elena BottazziNational School of Tropical MedicineSabin Vaccine Institute and TexasChildrens Center forVaccine DevelopmentSection of Pediatric Tropical MedicineBaylor College of Medicine1102 Bates St.Houston, TX 77030USA

Collette Britton�

University of GlasgowInstitute of Infection, Immunityand InflammationCollege of Medical, Veterinaryand Life SciencesBearsden RoadGlasgow G61 1QHUKE-mail: Collette.Britton@glasgow.ac.uk

Andrew R. BurnsUniversity of TorontoDepartment of Molecular Genetics andThe Donnelly Centre for Cellular andBiomolecular Research160 College Street, Rm1202TorontoOntario, M5S 1A8Canada

Samuel K. BuxtonIowa State UniversityDepartment of Biomedical SciencesAmes, IA 50011USA

Jacques CabaretINRAUR1282 Infectiologie Animale et SantéPublique37380 NouzillyFrance

Conor R. Caffrey�

University of California San FranciscoSandler Center for Drug Discovery andthe Department of Pathology1700 4th StreetSan Francisco, CA 94158-2330USAE-mail: conor.caffrey@ucsf.edu

Bronwyn E. CampbellThe University of MelbourneFaculty of Veterinary ScienceCorner Flemington Road and Park DriveParkvilleVictoria 3010Australia

Cinzia CantacessiThe University of MelbourneFaculty of Veterinary ScienceCorner Flemington Road and Park DriveParkvilleVictoria 3010Australia

Carlos CarmonaUniversidad de la RepúblicaUnidad de Biología ParasitariaInstituto de BiologíaFacultad de CienciasAv. A. Navarro 3051CP 11600 MontevideoUruguay

Santiago J. CarmonaUniversidad de San MartínInstituto de InvestigacionesBiotecnológicas25 de Mayo y FranciaSan MartínB 1650 HMP, Buenos AiresArgentina

XIV List of Contributors

Claude L. CharvetINRAUR1282 Infectiologie Animale et SantéPublique37380 NouzillyFrance

Christophe ChassaingIntervet Innovation GmbHMSD Animal HealthZur Propstei55270 SchwabenheimGermany

Donato CioliNational Research CouncilCell Biology and Neurobiology Institute32 Via RamariniMonterotondo00015 RomeItaly

George A. ConderPfizer Animal HealthVeterinary Medicine Research &Development7000 Portage RoadKalamazoo, MI 49001USA

Linsey R. CozzieMerial LtdClinical R&D Americas East115 Transtech DriveAthens, GA 30601USA

Gregory J. CrowtherUniversity of WashingtonDepartment of MedicineDivision of Allergy and InfectiousDiseases1959 NE Pacific StreetSeattle, WA 98195-7185USA

Akram A. DadaraTufts UniversityMolecular Helminthology LaboratoryDivision of Infectious DiseasesDepartment of Biomedical SciencesCummings School of VeterinaryMedicine200 Westboro RoadGrafton, MA 01536USA

John P. DaltonMcGill UniversityInstitute of Parasitology21111 Lakeshore RoadSt Anne de BellevueQuebec H9X 3V9Canada

David DiemertSabin Vaccine Development PDP723-D Ross Hall2300 Eye Street NWWashington, DC 20037USA

Sheila M. DonnellyUniversity of Technology Sydneyithree InstituteLevel 6, Building 4Corner of Thomas and Harris StreetSydneyNew South Wales 2007Australia

Denise DoolanQueensland Institute of MedicalResearchDivision of Infectious Diseases300 Herston RdBrisbaneQueensland 4006Australia

List of Contributors XV

Patrick DriguezQueensland Institute of MedicalResearchDivision of Infectious Diseases300 Herston RdBrisbaneQueensland 4006Australia

Rebecca FankhauserMerial LtdClinical R&D Americas East115 Transtech DriveAthens, GA 30601USA

Philip FelgnerUniversity of California IrvineSchool of Medicine3052 Hewitt HallIrvine, CA 92697USA

Louise FordLiverpool School of Tropical MedicineMolecular and Biochemical ParasitologyPembroke PlaceLiverpool L3 5QAUK

Jeremy M. FosterNew England BiolabsDivision of Molecular Parasitology240 County RoadIpswich, MA 01938USA

Robin B. Gasser�

The University of MelbourneFaculty of Veterinary ScienceCorner Flemington Road and Park DriveParkvilleVictoria 3010AustraliaE-mail: robinbg@unimelb.edu.au

Charles G. GauciUniversity of MelbourneVeterinary Clinical Center250 Princes HighwayWerribeeVictoria 3030Australia

Soraya GazeJames Cook UniversityQueensland Tropical Health AllianceMcGregor Rd, SmithfieldCairnsQueensland 4878Australia

Timothy G. Geary�

McGill UniversityInstitute of Parasitology21111 Lakeshore RoadSte-Anne-de-BellevueQuebec H9X 3V9CanadaE-mail: timothy.g.geary@mcgill.ca

Ross S. HallThe University of MelbourneFaculty of Veterinary ScienceCorner Flemington Road and Park DriveParkvilleVictoria 3010Australia

Lance G. Hammerland�

Merial LtdPharmaceutical R&D3239 Satellite BoulevardDuluth, GA 30096USAE-mail: lance.hammerland@merial.com

XVI List of Contributors

Anja. R. HeckerothIntervet Innovation GmbHMSD Animal HealthZur Propstei55270 SchwabenheimGermany

Achim HoeraufUniversity Clinic BonnInstitute for Medical Microbiology,Immunology, and ParasitologySigmund-Freud Strasse 2553105 BonnGermany

Lindy Holden-Dye�

University of SouthamptonCentre for Biological SciencesUniversity RoadSouthampton SO17 1BJUKE-mail: lmhd@soton.ac.uk

Eugenio L. de Hostos�

OneWorld HealthSuite 250280 Utah AvenueSouth San Francisco, CA 94080USAE-mail: edehostos@oneworldhealth.org

Peter Hotez�

National School of Tropical MedicineSabin Vaccine Institute and TexasChildren’s Center for VaccineDevelopmentSection of Pediatric Tropical MedicineBaylor College of Medicine1102 Bates St.Houston, TX 77030USAE-mail: hotez@bcm.edu

Yan HuUniversity of California San DiegoSection of Cell and DevelopmentalBiologyDivision of Biological Sciences9500 Gilman DrLa Jolla, CA 92093-0322USA

Abdul JabbarThe University of MelbourneVeterinary Clinical Center250 Princes HighwayWerribeeVictoria 3030Australia

Amar R. JariwalaGeorge Washington UniversityMedical CenterClinical Immunology LaboratoryDepartment of Microbiology,Immunology and Tropical Medicine2300 Eye Street, NWWashington, DC 20052USA

Aaron R. JexThe University of MelbourneDepartment of Veterinary ScienceCorner Flemington Road and Park DriveParkvilleVictoria 3010Australia

Sandra S. JohnsonPfizer Animal HealthVeterinary Medicine Research &Development7000 Portage RoadKalamazoo, MI 49001USA

List of Contributors XVII

Ronald Kaminsky�

NovartisCenter de Recherche Santé AnimaleRoute de la Petite Glâne1566 Saint AubinSwitzerlandE-mail: ronald.kaminsky@novartis.com

Jennifer KeiserSwiss Tropical and Public HealthInstituteDepartment of Medical Parasitology andInfection BiologyPO Box4002 BaselSwitzerland

Thomas R. KleiLouisiana State UniversitySchool of Veterinary MedicineSkip Bertman DriveBaton Rouge, LA 70803USA

David Knox�

Moredun Research InstituteParasitology DivisionBush LoanPenicuik EH26 0PZUKE-mail: dave.knox@moredun.ac.uk

Sanjay KumarNew England BiolabsDivision of Molecular Parasitology240 County RoadIpswich, MA 01938USA

Kristin Lees�

University of ManchesterFaculty of Life SciencesOxford RoadManchester M13 9PTUKE-mail: kristin.lees@manchester.ac.uk

Marshall W. Lightowlers�

University of MelbourneVeterinary Clinical Center250 Princes HighwayWerribeeVictoria 3030AustraliaE-mail: marshall@unimelb.edu.au

Alex Loukas�

James Cook UniversityQueensland Tropical Health AllianceMcGregor Rd, SmithfieldCairnsQueensland 4878AustraliaE-mail: Alex.Loukas@jcu.edu.au

Sara Lustigman�

New York Blood CenterLaboratory of Molecular ParasitologyLindsley F. Kimball Research Institute310 East 67th StreetNew York, NY 10065USAE-mail: slustigman@nybloodcenter.org

Jürgen LutzIntervet Innovation GmbHMSD Animal HealthZur Propstei55270 SchwabenheimGermany

Steven J. MaederPfizer Animal HealthVeterinary Medicine Research &Development38–42 Wharf Road West RydeSydneyNew South Wales 2114Australia

XVIII List of Contributors

Richard J. Martin�

Iowa State UniversityDepartment of Biomedical SciencesAmes, IA 50011USAE-mail: rjmartin@iastate.edu

James H. McKerrowUniversity of California San FranciscoSandler Center for Drug Discovery andDepartment of PathologyCalifornia Institute for QuantitativeBiosciences1700 4th StreetSan Francisco, CA 94158-2330USA

Donald P. McManusQueensland Institute of MedicalResearchDivision of Infectious Diseases300 Herston RdBrisbaneQueensland 4006Australia

Bakela NareScynexis Inc.Department3501C Tricenter BoulevardDurham, NC 27713USA

Cedric NeveuINRAUR1282 Infectiologie Animale et SantéPublique37380 NouzillyFrance

Tue NguyenOneWorld HealthSuite 250280 Utah AvenueSouth San Francisco, CA 94080USA

Matthew J. NolanThe University of MelbourneFaculty of Veterinary ScienceCorner Flemington Road and Park DriveParkvilleVictoria 3010Australia

Sandra M. ONeillDublin City UniversityParasite Immune Modulation GroupSchool of NursingCollins AvenueGlasnevinDublin 9Ireland

Mark PearsonJames Cook UniversityQueensland Tropical Health AllianceMcGregor Rd, SmithfieldCairnsQueensland 4878Australia

Maria Victoria PeriagoGeorge Washington University MedicalCenterClinical Immunology LaboratoryDepartment of Microbiology,Immunology and Tropical Medicine2300 Eye Street, NWWashington, DC 20052USA

List of Contributors XIX

and

FIOCRUZClinical Immunology LaboratoryLaboratório de Imunologia Celular eMolecularCentro de Pesquisas René RachouAv. Augusto de Lima 1715Belo HorizonteMinas Gerais 30190-002

Ken PfarrUniversity Clinic BonnInstitute for Medical Microbiology,Immunology, and ParasitologySigmund-Freud Strasse 2553105 BonnGermany

Livia Pica-Mattoccia�

National Research CouncilCell Biology and Neurobiology Institute32 Via RamariniMonterotondo00015 RomeItalyE-mail: lpica@ibc.cnr.it

David PiedrafitaMonash UniversityBiotechnology Research LaboratoriesWellington RoadClaytonVictoria 3800Australia

Kerrie PowellScynexis Inc.Department3501C Tricenter BoulevardDurham, NC 27713USA

Sreekanth PuttacharyIowa State UniversityDepartment of Biomedical SciencesAmes, IA 50011USA

Stuart A. RalphUniversity of MelbourneDepartment of Biochemistry andMolecular BiologyBio21 Molecular Science andBiotechnology Institute30 Flemington RoadParkvilleVictoria 3010Australia

Celso Raul Romero RamosFIOCRUZLaboratório de EsquistossomoseExperimentalAv. Brasil 4365ManguinhosRio de Janeiro 21-045-900Brazil

Alan P. RobertsonIowa State UniversityDepartment of Biomedical SciencesAmes, IA 50011USA

Mark W. Robinson�

University of Technology Sydneyithree InstituteLevel 6, Building 4Corner of Thomas and Harris StreetSydneyNew South Wales 2007AustraliaE-mail: mark.robinson@uts.edu.au

XX List of Contributors

Andreas RohwerIntervet Innovation GmbHMSD Animal HealthZur Propstei55270 SchwabenheimGermany

and

Roche Diagnostics Deutschland GmbHSandhofer Strasse 11668305 MannheimGermany

David S. RoosUniversity of PennsylvaniaDepartment of Biology and PennGenomics Institute415 South University AvePhiladelphia, PA 19104USA

Peter J. Roy�

University of TorontoDepartment of Molecular Genetics, TheDonnelly Centre for Cellular andBiomolecular Research and theCollaborative Programme inDevelopmental Biology160 College Street, Rm1202TorontoOntario, M5S 1A8CanadaE-mail: peter.roy@utoronto.ca

Lucien RufenerNovartisCenter de Recherche Santé AnimaleRoute de la Petite Glâne1566 Saint AubinSwitzerland

David SattelleUniversity of ManchesterFaculty of Life SciencesOxford RoadManchester M13 9PTUK

Dhanasekaran ShanmugamDivision of Biochemical SciencesNational Chemcial LaboratoriesDr. Homi Bhabha RoadPune, 411008India

Niroda ShannanUniversity of ManchesterFaculty of Life SciencesOxford RoadManchester M13 9PTUK

Paul M. Selzer�

Intervet Innovation GmbHMSD Animal HealthZur Propstei55270 SchwabenheimGermanyE-mail: Paul.Selzer@msd.de

Andrew J.G. SimpsonLudwig Institute for Cancer ResearchNew York Branch at Memorial Sloan-Kettering Cancer Center1275 York AvenueNew York, NY 10021USA

Rahul Singh�

San Francisco State UniversityDepartment of Computer Science1600 Holloway AvenueSan Francisco, CA 94132USAE-mail:rsingh@cs.sfusu.edu

List of Contributors XXI

Patrick J. Skelly�

Tufts UniversityMolecular Helminthology LaboratoryDivision of Infectious DiseasesDepartment of Biomedical SciencesCummings School of VeterinaryMedicine200 Westboro RoadGrafton, MA 01536USAE-mail: Patrick.Skelly@Tufts.edu

Barton E. SlatkoNew England BiolabsDivision of Molecular Parasitology240 County RoadIpswich, MA 01938USA

Ann E. SluderScynexis Inc.Department3501C Tricenter BoulevardDurham, NC 27713USA

Peter M. SmookerRMIT UniversitySchool of Applied SciencesPlenty RoadBundooraVictoria 3083Australia

Terry W. Spithill�

La Trobe UniversityDepartment of Agricultural Sciencesand Center for AgriBioscienceKingsbury DriveBundooraVictoria 3086AustraliaE-mail: t.spithill@latrobe.edu.au

Mark J. Taylor�

Liverpool School of Tropical MedicineMolecular and Biochemical ParasitologyPembroke PlaceLiverpool L3 5QAUKE-mail: mark.taylor@liverpool.ac.uk

Miriam Tendler�

FIOCRUZLaboratório de EsquistossomoseExperimentalAv. Brasil 4365ManguinhosRio de Janeiro 21-045-900BrazilE-mail: mtendler@ioc.fiocruz.br

David P. ThompsonPfizer Animal HealthVeterinary Medicine Research &Development7000 Portage RoadKalamazoo, MI 49001USA

Manfred UphoffIntervet Innovation GmbHMSD Animal HealthZur Propstei55270 SchwabenheimGermany

Jürg UtzingerSwiss Tropical and Public HealthInstituteDepartment of Epidemiology and PublicHealthPO Box4002 BaselSwitzerland

XXII List of Contributors

Robert J. WalkerUniversity of SouthamptonCentre for Biological SciencesUniversity RoadSouthampton SO17 1BJUK

Sally M. WilliamsonUniversity of GeorgiaDepartment of Infectious Diseases &Center for Tropical and EmergingGlobal DiseaseAthens, GA 30602USA

Adrian J. WolstenholmeUniversity of GeorgiaDepartment of Infectious Diseases &Center for Tropical and EmergingGlobal DiseaseAthens, GA 30602USA

Debra J. Woods�

Pfizer Animal HealthVeterinary Medicine Research &Development7000 Portage RoadKalamazoo, MI 49001USAE-mail: debra.j.woods@pfizer.com

Neil D. Young�

The University of MelbourneFaculty of Veterinary ScienceCorner Flemington Road and Park DriveParkville, Victoria 3010AustraliaE-mail: nyoung@unimelb.edu.au

List of Contributors XXIII

Part OneTargets

Parasitic Helminths: Targets, Screens, Drugs and Vaccines, First Edition. Edited by Conor R. Caffrey� 2012 Wiley-VCH Verlag GmbH & Co. KGaA. Published 2012 by Wiley-VCH Verlag GmbH & Co. KGaA.

j1

1Ligand-Gated Ion Channels as Targets for AnthelminticDrugs: Past, Current, and Future PerspectivesKristin Lees*, Ann Sluder, Niroda Shannan, Lance Hammerland, and David Sattelle

AbstractLigand-gated ion channels (LGIC) are targets for anthelmintic drugs used in humanhealth and veterinary applications. Given the diverse physiological roles of LGICs inneuromuscular function, the nervous system, and elsewhere, it is not surprising thatrandom chemical screening programs often identify drug candidates targeting thissuperfamily of transmembrane proteins. Such leads provide the basis for furtherchemical optimization, resulting in important commercial products. Currently,members of three LGIC families are known to be targeted by anthelmintics. Theseinclude the nicotinic acetylcholine receptors gating cation channels, glutamate-gatedchloride channels, and g-aminobutyric acid-gated chloride channels. The recentimpact of genomics on model invertebrates and parasitic species has been far-reaching, leading to the description of new helminth LGIC families. Among thecurrent challenges for anthelmintic drug discovery are the assessment of newlydiscovered LGICs as viable targets (validation) and circumventing resistance whenexploring further the well-established targets. Recombinant expression of helminthLGICs is not always straightforward. However, new developments in the under-standing of LGIC chaperones and automated screening technologies may holdpromise for target validation and chemical library screening on whole organismsor ex vivo preparations. Here, we describe LGIC targets for the current anthelminticsof commercial importance and discuss the potential impact of that knowledge onscreening for new compounds. In addition, we discuss some new technologies foranthelmintic drug hunting, aimed at the discovery of novel treatments to controlveterinary parasites and some neglected human diseases.

Introduction

Anthelmintic drugs are central to combating many human and veterinary disorders.One in four of the worlds population is infected with a parasitic roundworm ornematode, with infestation being particularly severe in tropical and subtropicalregions. The consequent debilitating effects on the workforce and the compounding

* Corresponding Author

j3

Parasitic Helminths: Targets, Screens, Drugs and Vaccines, First Edition. Edited by Conor R. Caffrey� 2012 Wiley-VCH Verlag GmbH & Co. KGaA. Published 2012 by Wiley-VCH Verlag GmbH & Co. KGaA.

risk of other pathogenic infections represents a considerable social and economicburden. If we add to that a very high level of roundworm infestation among theworlds farmed animals, and the devastating impact of trematode parasites in manand animals, then the need for adequate helminth control is transparent [1, 2].

The veterinary economic burden is reflected in the scale of the global animal healthdrug market (approximately US$11 billion/annum) [3]. The human health antipar-asitic drugmarket is aroundUS$0.5 billion/annum.However, it costs aroundUS$40million to develop a new drug that controls livestock nematodes, whereas it can costUS$800 million for a new drug for human use. Understandably, the cost barrier haslimited progress, but the size of the global markets for antiparasitic drugs andchemicals make their pursuit of commercial interest as well as an important humanand animal health priority.

Exciting new developments in research on vaccines targeting helminth parasitesare underway and these, undoubtedly, will make important contributions in thefuture. However, at present, chemical approaches to helminth control predominate.For example, the worlds three top-selling veterinary antiparasitic drugs (imidaclo-prid, fipronil, and ivermectin) and several others such as selamectin, levamisole,pyrantel, morantel, tribendimidine, piperazine, and amino-acetonitrile derivatives(AADs) act on Cys-loop ligand-gated ion channels (LGICs). These transmembranereceptormolecules facilitate the fast actions of neurotransmitter chemicals at nerve–nerve synapses and neuromuscular junctions (NMJs) in invertebrates. Often theyoffer rapid control of the pathogen. Much of our current knowledge of theseimportant drug targets stems from the genetic model organism and free-livingnematode, Caenorhabditis elegans, which possesses the most extensive known super-family of Cys-loop LGICs, consisting of 102 subunit-encoding genes [4]. They includecation-permeable channels gated by acetylcholine (ACh) and c-aminobutyricacid (GABA) as well as anion-selective channels gated by ACh, GABA, glutamate,5-hydroxytryptamine (5-HT), dopamine, and tyramine [5–7]. Less than half ofthe genes in the C. elegans Cys-loop LGIC superfamily have been functionallycharacterized.

Unfortunately, many of the anthelmintic drugs in current use are under threat(Table 1.1). Important compounds such as ivermectin, which have given excellentservice, are at the end of their patent life. Repeated use of effective chemicals leads tothe development of pathogen resistance. Indeed, multidrug resistance against thethree major classes of anthelmintics including macrocyclic lactones, which targetglutamate-gated chloride channels (GluCls), has become a global problem for thetreatment of gastrointestinal nematode parasites of farm animals [8–10]. Theincreasing development costs and poor return from conventional screeningapproaches are also problematic. Together, these factors bring a sense of urgencyto the development of new, effective anthelmintics.

The life of a patent has always been finite, but as the time from discovery tomarketbecomes protracted and the bar is raised for new, safer molecules with improvedspecifications on toxicity and environmental residues, the task of discovery becomesmore difficult. The introduction of generic forms of a drug has the potential to lowerthe cost of treatment andmake it availablemore widely, although this positive benefit

4j 1 Ligand-Gated Ion Channels as Targets for Anthelmintic Drugs: Past, Current, and Future Perspectives