Table Of ContentEdited by
Timo Ja¨ger, Oliver Koch, and Leopold Floh´e
Trypanosomatid Diseases
Titles of the Series ‘‘Drug Discovery in Infectious Diseases’’
Selzer,P.M. (ed.) Conor R.Caffrey (ed.)
AntiparasiticandAntibacterial ParasiticHelminths
DrugDiscovery Targets,Screens,DrugsandVaccines
FromMolecularTargetstoDrug 2012
Candidates
ISBN978-3-527-33059-1
2009
ISBN:978-3-527-32327-2
Becker,K.(ed.)
ApicomplexanParasites
MolecularApproachestowardTargeted
DrugDevelopment
2011
ISBN:978-3-527-32731-7
Forthcoming Topics of the Series
(cid:129)Protein Phosphorylation inParasites:Novel Targets for Antiparasitic Intervention
Related Titles
Lucius,R.,Loos-Frank,B.,Grencis,R.K., Zajac,A. M.,Conboy, G.A.(eds.)
Striepen, B.,Poulin,R. (eds.)
VeterinaryClinicalParasitology
TheBiologyofParasites
2012
2014
ISBN:978-0-8138-2053-8
ISBN:978-3-527-32848-2
Lamb, T.
Scott,I.,Sutherland, I.
ImmunitytoParasiticInfections
GastrointestinalNematodesof
2013 SheepandCattle
ISBN:978-0-470-97247-2 BiologyandControl
2009
ISBN:978-1-4051-8582-0
Edited by Timo Jäger, Oliver Koch, and Leopold Flohé
Trypanosomatid Diseases
Molecular Routes to Drug Discovery
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j
V
Contents
Foreword IX
Acknowledgment XV
Preface XVII
ListofContributors XIX
PartOne DiseaseBurden,CurrentTreatments,MedicalNeeds,
andStrategicApproaches 1
1 VisceralLeishmaniasis–CurrentTreatmentsandNeeds 3
PoonamSalotra,RuchiSingh,andKarinSeifert(cid:1)
2 ChemotherapyofLeishmaniasis:AVeterinaryPerspective 17
MaríaJesu(cid:1)sCorral-CaridadandJos(cid:1)eMaríaAlunda(cid:1)
3 PharmacologicalMetabolomicsinTrypanosomes 37
DarrenJ.Creek,IsabelM.Vincent,andMichaelP.Barrett(cid:1)
4 DrugDesignandScreeningbyInSilicoApproaches 57
MattiaMoriandMaurizioBotta(cid:1)
5 ComputationalApproachesandCollaborativeDrugDiscovery
forTrypanosomalDiseases 81
SeanEkins(cid:1)andBarryA.Bunin
PartTwo MetabolicPeculiaritiesintheTrypanosomatidFamily
GuidingDrugDiscovery 103
6 InteractionofLeishmaniaParasiteswithHostCellsanditsFunctional
Consequences 105
UtaSchurigt,AnitaMasic,andHeidrunMoll(cid:1)
j
VI Contents
7 FunctionofGlycosomesintheMetabolismofTrypanosomatidParasites
andthePromiseofGlycosomalProteinsasDrugTargets 121
MelisaGualdr(cid:1)on-L(cid:1)opez,PaulA.M.Michels(cid:1),WilfredoQuin~ones,AnaJ.C(cid:1)aceres,
LuisanaAvil(cid:1)an,andJuan-LuisConcepci(cid:1)on
8 GlyoxalaseEnzymesinTrypanosomatids 153
MartaSousaSilva,Ant(cid:1)onioE.N.Ferreira,RicardoGomes,AnaM.Tom(cid:1)as,
AnaPoncesFreire,andCarlosCordeiro(cid:1)
9 Trypanothione-BasedRedoxMetabolismofTrypanosomatids 167
MarceloA.Comini(cid:1)andLeopoldFloh(cid:1)e
10 ThiolPeroxidasesofTrypanosomatids 201
HelenaCastro(cid:1)andAnaM.Tom(cid:1)as
11 PeroxynitriteasaCytotoxicEffectorAgainstTrypanosomacruzi:
OxidativeKillingandAntioxidantResistanceMechanisms 215
MadiaTrujillo,MaríaNoelAlvarez,LucíaPiacenza,MartínHugo,
GonzaloPeluffo,andRafaelRadi(cid:1)
12 SelenoproteomeofKinetoplastids 237
AlexeiV.LobanovandVadimN.Gladyshev(cid:1)
13 ReplicationMachineryofKinetoplastDNA 243
RachelBezalel-Buch,NuritYaffe,andJosephShlomai(cid:1)
14 LifeandDeathofTrypanosomabrucei:NewPerspectivesforDrug
Development 261
TorstenBarth,JasminStein,StefanMogk,CarolineSch€onfeld,
BrunoK.Kubata,andMichaelDuszenko(cid:1)
PartThree ValidationandSelectionofDrugTargetsinKinetoplasts 279
15 RationalSelectionofAnti-MicrobialDrugTargets:Unique
orConserved? 281
BorisRodenkoandHarryP.deKoning(cid:1)
16 DrugTargetsinTrypanosomalandLeishmanialPentosePhosphate
Pathway 297
MarceloA.Comini(cid:1),CeciliaOrtíz,andJuanJos(cid:1)eCazzulo
17 GDP-Mannose:AKeyPointforTargetIdentificationandDrugDesign
inKinetoplastids 315
S(cid:1)ebastienPomel(cid:1)andPhilippeM.Loiseau
j
Contents VII
18 TransportersinAnti-ParasiticDrugDevelopmentandResistance 335
VincentDelespauxandHarryP.deKoning(cid:1)
19 PeptidasesinAutophagyareTherapeuticTargetsforLeishmaniasis 351
RoderickA.M.Williams(cid:1)
20 ProteasesofTrypanosomabrucei 365
DietmarSteverding(cid:1)
PartFour ExamplesofTarget-BasedApproachesandCompoundsUnder
Consideration 383
21 ScreeningApproachesTowardsTrypanothioneReductase 385
MathiasBeig,FrankOellien,R.LuiseKrauth-Siegel,andPaulM.Selzer(cid:1)
22 Redox-ActiveAgentsinReactionsInvolvingtheTrypanothione/Trypanothione
Reductase-basedSystemtoFightKinetoplastidalParasites 405
ThibaultGendron,DonAntoineLanfranchi,andElisabethDavioud-Charvet(cid:1)
23 InhibitionofTrypanothioneSynthetaseasaTherapeuticConcept 429
OliverKoch(cid:1),TimoJ€ager,LeopoldFloh(cid:1)e,andPaulM.Selzer
24 TargetingtheTrypanosomatidicEnzymesPteridineReductase
andDihydrofolateReductase 445
StefaniaFerrari,ValeriaLosasso,PuneetSaxena,andMariaPaolaCosti(cid:1)
25 ContributiontoNewTherapiesforChagasDisease 473
PatriciaS.Doyle(cid:1)andJuanC.Engel
26 ErgosterolBiosynthesisfortheSpecificTreatmentofChagasDisease:
FromBasicSciencetoClinicalTrials 489
JulioA.Urbina(cid:1)
27 NewDevelopmentsintheTreatmentofLate-StageHumanAfrican
Trypanosomiasis 515
CyrusJ.Bacchi,RobertT.Jacobs,andNigelYarlett(cid:1)
Index 531
j
IX
Foreword
DrugDiscoveryforNeglectedDiseases–
PastandPresentandFuture
The kinetoplastid diseases – sleeping sickness, the leishmaniases, and Chagas
disease – are “neglected diseases of poverty,” afflicting millions of people and
collectivelyresponsibleforover100000deathsperannum.Novaccinesareavailable
and current insect vector control methods and other public health measures are
insufficienttoeliminatethem.Thecurrentlyavailabledrugtherapiesarefarfrom
satisfactoryduetoissuessuchaspoorefficacy,toxicity,theneedforhospitalization,
therequirementforprolongedparenteraltreatment,andhighcost.Thisbookisa
timely attempt to address some of these unmet medical needs.
Itissomewhatironicthat,atthebeginningofthetwentiethcentury,manyofthe
ground-breaking developments in drug discovery were driven by economic and
colonialexpansioninAfricaandAsia.TheAfricantrypanosomeinparticularwasan
earlymodelforexperimentalchemotherapyalongtwomainlinesofinvestigation:
synthetic dyes, and organic arsenicals and antimonials (for further details, see
reviewsbyWilliamson[1]andSteverding[2]).Indeed,thefirstsyntheticcompound
tocureaninfectiousdiseaseinananimalmodelwasthedye,Trypanred(Ehrlich,
1904), which was a forerunner of suramin (1916–1920), the first effective
trypanocidal drug for human African trypanosomiasis (HAT). The demonstration
by Thomas and Breinl in 1905 of the trypanocidal activity in mice of atoxyl
(p-aminophenylarsonic acid) formed the basis of Ehrlich’s pioneering work on
organic arsenicals that culminated in the development of arsphenamine (606,
Salvarsan)forthetreatmentofsyphilis(EhrlichandHata,1910)andtryparsamide
for the treatment of HAT (Jacobs and Heidelberger, 1919). Tryparsamide, which
caused blindness in 10–20% of patients, was finally replaced by melarsoprol
(Friedheim, 1949). Trivalent antimony, in the form of tartar emetic, was shown
tobetrypanocidalinmice(PlimmerandThompson,1908),butlackedefficacyin
humans.However,potassiumantimonytartratewasfoundtohavesomeactivityin
the treatment of leishmaniasis (Vianna, 1912) and was the forerunner of the
1
pentavalent antimonial drugs, sodium stibogluconate (Pentostam ) and meglu-
1
mine antimonate (Glucantime ), both introduced in the 1940s. Along with the
diamidine, pentamidine (1937), all of these drugs are still in use today.
From the 1950s, subsequent drug treatments have largely been discovered by
serendipitythroughrepurposingofexistingdrugsusedforotherindications.The
nitrofuran, nifurtimox, and the nitroimidazole, benznidazole, used for the
j
X Foreword DrugDiscoveryforNeglectedDiseases–PastandPresentandFuture
treatmentofChagasdiseasearosefromresearchintonitro-compoundsasantibac-
terial agents. Amphotericin B, isolated in 1953, was originally developed for the
treatment of systemic mycoses, but later found use in the treatment of visceral
leishmaniasis in the 1990s as the expensive, but highly efficacious liposomal
1
formulation (AmBisome ) or as the cheaper, but more toxic amphotericin B
deoxycholate.BothformulationswereincludedintheWorldHealthOrganization’s
Essential Medicines List in 2009. The off-patent aminoglycoside, paromomycin,
originallydevelopedasanoraltreatmentforintestinalinfectionsinthe1960s,finally
gained approval as paromomycin intramuscular injection for the treatment of
visceral leishmaniasis in India in 2006. Two anticancer agents, the phospholipid
analog,miltefosine,registeredasthefirstoraltreatmentforvisceralleishmaniasis
in India in 2002, and eflornithine (now in combination with oral nifurtimox as
nifurtimox–eflornithine combination therapy (NECT), 2009) for the treatment of
HAT caused by Trypanosoma brucei gambiense complete the woefully inadequate
treatment options for these diseases. Notably, none of these newer developments
havecompletelydisplacedtheirforerunnersthatweredevelopedpriortothe1950s.
After nearly a century of research, only 10 novel chemical entities for three
diseases is a singularly unimpressive output by the pharmaceutical industry. The
reasonforthislamentableperformanceisnothardtofind.Pooreconomicreturnon
investment by pharma is a major factor, since these are diseases of poverty. The
thalidomide disaster of the late 1950s kick-started regulatory demands for greater
patient safety resulting in ever-increasing development costs. Blockbuster drugs
were“in;”smaller,lessprofitablemarketswere“out.”Ninetypercentofresearchand
developmentwasaimedat10%oftheworld’sunmetmedicalneed–theso-called
“10–90gap.”Thedriveforgreaterefficiencyandprofitabilitythroughmergersand
acquisitions resulted in the loss of parasitology expertise in most pharma compa-
nies. Medicinal chemists were seduced by combinatorial chemistry without due
regardforchemicalspaceanddrug-likeness.Miniaturizationofchemicalsynthesis
restricted the use of animal disease models and shifted emphasis towards target
screening. Intellectual property rights were increasingly used as an obstructive,
rather than an enabling tool.
Attheendofthetwentiethcentury,thesituationhadbecomesodirethataradical
new approach was required. One of the most encouraging developments was the
foundingof“public–privatepartnerships”(PPPs),suchastheDrugsforNeglected
Diseasesinitiative(DNDi)andMedicinesforMalariaVenture(MMV)–non-profit
organizations who strive to forge drug discovery partnerships between multiple
academic,biotech,andpharmapartnerswithfundingfromthegovernmentaland
charitablesector[3].PPPswereinitiallymetwithmuchskepticism,butby2005,an
analysisbyMaryMoranandcolleaguesconcludedthatPPPswereresponsiblefor
three-quarters of an expanded research and development portfolio for neglected
diseases [4]. Another important development was the publication of annotated
genomes for T. brucei, L. major, and T. cruzi in 2005 [5–7]. As Barry Bloom
optimistically prophesized 10 years earlier “Sequencing bacterial and parasitic
pathogens . . . could buy the sequence of every virulence determinant, every
protein antigen and every drug target . . . for all time” [8]. Certainly, pathogen
j
Foreword DrugDiscoveryforNeglectedDiseases–PastandPresentandFuture XI
genomes are proving to be a valuable resource for target discovery, but without a
deeperunderstandingofparasitebiologythefullpotentialofthesegenomeswillnot
berealized.Aboutthesametimeasgenomesequencingwasgettingunderway,C.C.
Wang threw down the gauntlet that academics needed genetic evidence of essen-
tiality to justify their claims of the therapeutic potential of their research field [9].
This dogma has now been refined and extended to include chemical evidence of
druggability,drivenbyadefinedtherapeuticproductprofile[10].Thesechallenges
haveencouragedsomeacademicstomoveoutoftheirtraditionalcomfortzonesto
fill the early-stage drug discovery gap in translational medicine not adequately
coveredbythePPPs[11].Theconceptof“onegene,onetarget,onedrug”hasbeen
very much at the forefront of current academic (and industry) thinking, with
structure-based design an important adjunct in this strategy. Thus, it is timely
thatmuchofthisbookisdevotedtotheidentificationofmetabolicpeculiaritiesin
thekinetoplastidsthatcanbechemicallyandgeneticallyvalidatedasdrugtargets.
However, what of the future? Experience in industry and in academia suggests
that the rate of validation of new targets is failing to keep pace with the rate of
attrition of currently validated targets. Despite initial promise, the target-based
approachhasyieldeddisappointingresultsinanti-bacterialdiscoveryinpharma[12]
and lessons need to be learned from this if we are to avoid making the same
mistakes.Rapidandrobustmethodsofgenetictargetvalidationarestillneededfor
parasitescausingvisceralleishmaniasisandChagasdisease,andweneedabetter
understanding of basic biology to understand why targets fail. Certainly, not all
targetsareequalfromamedicinalchemistrypointofview.Greaterattentionneeds
tobepaidtodruglikeness[13]andligandefficiency[14]forleadselection.Screening
offragmentlibrariesusingbiophysicalmethodsshouldhelptoweedout“undrug-
gable”targetswithoutrecoursetoexpensivehigh-throughputscreens[15].Froma
pharmacology perspective, cytocidal activity is much preferable to cytostatic, so
biologists should address this question early in discovery. Likewise, the potential
easeforresistancearisingasaresultofpointmutationsinasingle-targetstrategy
shouldbearesearchpriorityforbiologists.Systemsbiologysuggeststhatexquisitely
selective,single-targetcompoundsmayexhibitlowerthandesiredclinicalefficacy
comparedwithmultitargetdrugsduetotherobustnessofbiologicalnetworks[16].
Thus, polypharmacology (network pharmacology) is undergoing a resurgence of
interest. Giventhepaucity of validated druggable targets,phenotypic screening is
undergoingarevivalaidedbyaccesstolargecompoundcollectionsheldbypharma
andthedevelopmentofsuitableminiaturizedwhole-parasitescreensandmamma-
lian counter-screens. This approach has the advantage of addressing the key
druggability issues of cell permeability, desirable cytocidal activity, and a suitable
parasite–hostselectivitywindow.Phenotypicscreeningcanalsoidentifycompounds
hitting non-protein targets (e.g., amphotericin B) or compounds that act as pro-
drugs(e.g.,nitroimidazoles).However,thefuturechallengewillbetoidentifythe
oftencomplexmode(s)ofactionofsuchphenotypichits(targetdeconvolution),and
to use modern technologies to improve the potency and selectivity of these
molecules[17].Finally,weshouldaskourselveswhetherourcompoundcollections
are too “clean” in terms of chemical reactivity. After all, arsenicals, antimonials,
