Lessons from the History of Ivermectin and Other Antiparasitic Agents

17 Pages·2016·0.93 MB·English

by  William C. Campbell

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ANNUAL Further
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Antiparasitic Agents
g
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s. William C. Campbell
w
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eo ResearchInstituteforScientistsEmeriti,DrewUniversity,Madison,NewJersey07940;
ualruse email:[email protected]
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Biosci. ded by Annu.Rev.Anim.Biosci.2016.4:1–14 Keywords
v. Anim. cess provi FOicrtsotbpeurb2li9sh,e2d01o5nlineasaReviewinAdvanceon idviescrmoveecrtyin,macrocycliclactones,anthelmintics,antiparasitics,drug
ec TheAnnualReviewofAnimalBiosciencesisonlineat
RA
u. animal.annualreviews.org Abstract
n
An Thisarticle’sdoi: The twentieth century’s arsenal of chemical anthelmintics brought mani-
10.1146/annurev-animal-021815-111209
foldimprovementinhumanhealthand,moreabundantly,inanimalhealth.
Copyright(cid:2)c 2016byAnnualReviews. The benefits were not only in health per se but also in agricultural eco-
Allrightsreserved
nomics,livestockmanagement,andtheoverallproductionoffoodandfiber
tosupportexpandinghumanpopulations.Nevertheless,thereremains(due
inlargeparttodrugresistanceandpaucityofavailablevaccines)agreatneed
fornewmeansofcontrollingdiseasecausedbyparasiticworms.Prudence
should persuade us to look to our past for lessons that might help in our
questfornewdrugs.Thelessonssuggestedherederivefromthehistoryof
ivermectin and other anthelmintics. They deal with the means of finding
substanceswithusefulantiparasiticactivityandwithalternativeapproaches
todrugdiscovery.
1
AV04CH01-Campbell ARI 10January2016 8:3
INTRODUCTION
Thousands of years ago, Scribonius Largus taught that “medicine” is derived from the Latin
medicamentum,referringnarrowlytowhatwecallmedication.Althoughtheetymologyhasbeen
disputed,thereisnodoubtthatmedicineremainslargelyamatterofmedicines.Thequestionof
howweacquiremedicinesisthereforeworthsomeconsideration.
Itisapleasureonthisoccasiontocontemplatethesubjectinthecontextoftheanimalsciences,
formyownexperiencewithantiparasiticdrugshaskeptanimalscienceverymuchinmymind
formanydecades.Myruminationsaboutchemotherapymayhavesomerelevancetopathogens
otherthanparasites,butIdonotpresumetosuggestthattheywillbepertinentbeyondtheareaof
infectiousdiseases.Inanycase,thestudyofhost-pathogenrelationshipsmayeventuallybereduced
tobiophysics,thedistinctionbetweendrugsandvaccinesmaybeerased,andchemotherapistsmay
dieout.
Asachemotherapistofasort,Iwouldliketothinkthatthechemotherapycommunitywillnot
g
or faceextinctionanytimesoon.Idonotactuallythinkofmyselfasachemotherapist.Likemost
s.
ewy. practitionersoftheart,Iamachemotherapistsecondarilytobeingsomethingelse.Iamazoologist
evionl andparasitologist.Butthebulkofmyresearchoverfivedecadeshasbeendevotedtoantiparasitic
ualruse agents.Experimentalchemotherapyismyoccupationalcallingandismysubjecthere.Because
annnal biographicalnarrativehasbeenincorporatedinpreviousprefatoryarticlesintheAnnualReview
www.perso ofAnimalBiosciences,Ishouldperhapsexplainhowmyunplannedcareercameabout.
m or Myinterestinparasiticworms,ifnotactuallycongenital,waspresentatanearlyage.Growing
ded fro3/16. F uexppionsethdetsomtahleltiodweanooffRtraemateinltgondoinmreusrtaiclCanoiumnatlysDtooncuegrealt(hIereirladnidse),aIsews,ouanlddnIatruermalelmyhbaevrebbeeineng
a2
nlo11/ fascinatedasateenagerbyaleafletadvertisingImperialChemicalIndustries’hexachloroethane
wn for the treatment of liver-fluke disease in sheep and cattle. Whatever its origin, my interest in
oo
1-14. D195.89 parFasoitritchwisoprrmeoscwcauspsaotioonntwoitehxppaanrdasiintetso,tahleiflealtoenPgroocfecsuspoartJio.Dna.lSamdydtihctoiofnT.rinityCollege,Dublin
016.4:0.254. Uwansivtehresistoyr,tmoufsitnisnpilrairnggempaernttboerheevledryacsctuoduenntatbslheo.uTldhrhoauvgeh[oaustImwyausnpdrievriglergadeduattoeaycekanrso,wSlmedygthe
27
Biosci. ded by jpuasrtasbietofoloregyh,ebduitehde(1w)]a.sDtheesnmojunsdt bSemgyinthniwngoutoldggaoinoanttteontbieocnomfoer hanisidnitsecronvaetrioynoafltlhuemvinaalureyoinf
m. ovi semipermeabletubingintheinvitrocultureoftapeworms.Histechniqueisstillinuse.Itwas
Anis pr becauseofSmyththatIwasgivenanopportunitytodopostgraduatestudiesintheUnitedStates,
v. ces andbecauseofhimIwasemboldenedtosetofffortheUniversityofWisconsinandthelaboratory
ec
RA
u. ofthelateProfessorArlieTodd.
n
n Todd had an abiding interest in livestock farming and livestock parasitology, as well as a
A
warmheartedinterestinthewelfareofhisgraduatestudents.HislaboratorywasintheDepartment
of Veterinary Science, so the ambiance was conducive to research on the treatment of disease.
BythetimeIhadfinishedmygraduateprogram,Iwasready(tomysurprise)toacceptajobin
industry—adecisionmadeeasierbythenameoftheparticularbranchofthecompanyinquestion,
theMerckInstituteforTherapeuticResearch.
Companies vary in theirgeographical and organizational maps,intheir founding ethos and
the constancy of their mission, and in the liberality with which they view “basic” exploratory
research. All these things (even constancy) are liable to change over time. For these reasons,
I was fortunate in 1957 in joining the Merck organization as a member of the Department of
Parasitology, which was then led by Dr. Ashton Cuckler [whose marvelous flair for industrial
researchIhavesketchedelsewhere(2)].TheresearchdivisionofMerckturnedouttobeacauldron
ofintellectualstimulationandexcitementthatfarexceededmyexpectations.Myknowledgeof
2 Campbell
AV04CH01-Campbell ARI 10January2016 8:3
chemistrybeingmeager,Iwasfortunateindeedtofindmyselfsurroundedbybrilliantchemists
and (as projects matured) similarly impressive scientists of every description. I had decided to
try industrial research (as I had decided to try the United States) for a year—perhaps two at
most.IremainedatMerckfor33years(andmadetheUnitedStatesmyhome).Sinceretiring
fromindustry,Ihavebeenprivilegedtobeamember,forthepast25years,oftheincomparable
ResearchInstituteforScientistsEmeritiatDrewUniversityinMadison,NewJersey.Cantherebe
amoreperfectpostindustrial“retirement”thancontinuingtopursueone’sprofessionalinterests
withstudentcolleagues?Throughitall,chemotherapyhasremainedmyfocus—andmydelight.
For many years, it has seemed to me that there has been a glaring need for a more candid
appraisal of the role of scientific research in the acquisition and deployment of agents for the
controlofinfectiousdiseases(3).Throughoutthoseyears,valuablenewdrugshavecontinuedto
appear,sothatperhapstheneedIproclaimhasmoreofgossamerthanofglare.Yetwemustask
whetherthetherapeuticbenefitshavebeencommensuratewiththeinvestmentintime,treasure,
g andscientifictalentoftheperiod.Inaworldinwhichdemographicandclimatologicchangeadd
or
s. tothecomplexityofourstruggletocontroldisease,arewegettingenoughbangforourbuck?
w
ey.
vinl Forfoodandfiber,humanbeingsdependtoalargeextentonnonhumananimals,whichin
eo
ualruse turn depend on plants; our objective should be to control (but not eliminate) the pathogenic
nnal populationsofthoselife-forms.Thequestionishowbesttodothat.Thefollowingparagraphs
an
www.perso euxspeloourerrweshoeuthrceersrmecoernetecfhfeecmtivoethlye.rAapnedu,tfiocoelhxparedriyenthcoeuogfhfeirtsmanayyblees,saopnrsotphoastamliisgmhtadeenafobrletruysintog
om For outasomewhatdifferentapproachtothediscoveryofdrugsforinfectiousdiseases.
d fr16. Anoteforthenonbiologicalreader:Zoologically,theearlydevelopmentalstagesofnematode
ade23/ wormsare“juveniles,”not“larvae.”Popularusehas,asitsooftenhas,legitimizedmisuse,and
wnlon 11/ now “larva” and “larvae” are widely applied to juvenile nematodes even in the (nonzoological)
oo
D9 scientific literature. This lax convention is used here in the interest of making the text more
4:1-14. 4.195.8 rtheaoduiglyhuunndfaesrhstiooondab.lTehweotredrm“m“imcriocbroe.o”rganism”ishereusedinterchangeablywiththevenerable
016.0.25
27
Biosci. ded by LESSONSFROMTHEHISTORYOFIVERMECTIN
m. ovi Thebiggestsinglefactorleadingtothediscoveryofivermectinwassteadfastrelianceonempirical
Anis pr principles of drug discovery. The antiparasitic efficacy of the chemical class was revealed by a
v. ces processthatispartofalongtraditionandthathasmanycomponents.Itrepresentsanapproach
ec
RA todrugdiscoverythatismostly,butnotexclusively,concernedwiththetreatmentofinfectious
nu. disease.Itscoreelementisthetestingnotonlyofsubstancesthatmayreasonablybeexpected,on
n
A scientificgrounds,topossesstherapeuticactivity,butalso(andespecially)ofsubstancesforwhich
nosuchrationaleexists.
Ivermectin (Figure 1) was introduced commercially in 1981 (4) for the control of parasitic
worms(helminths)andectoparasiticarthropods.Earlyscientificreportsincludedadescriptionof
themicroorganismthatproducesivermectinanditsprecursorabamectin(avermectin)aswellas
evidenceoftheirexceptionalpotencyagainstintestinalandextraintestinalparasites(5–8).
Ivermectin and its microbial precursor abamectin (avermectin) were the first of the macro-
cycliclactoneantiparasitics.They,likeotheranthelminticclasses,proliferatedthroughstructural
modification,inresponsetomarketneedandcompetitivepressure.Eprinomycin,forexample,is
achemicalderivativeselectedonthebasisofpharmacologicaldistributionintreatedcattle.But
havingoriginatedasproductsofmicrobialfermentation,macrocycliclactoneswerealsoamenable
to modification through fermentation research. A mutant strain of Streptomyces avermitilis
was altered by changes in growth medium to produce doramectin; chemical derivitization of
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AV04CH01-Campbell ARI 10January2016 8:3
HC 23
3 O CH 22
3
HO O O O 25
13 O R
O 12
O O
O O
OH
22
22
22
O
H
OH
Figure1
Thestructureofivermectin(22,23-dihydroavermectin).R = CH3,ivermectinB1a;R = H,ivermectin
B1b.Thedrugmaycontainupto20%oftheB1bhomolog.
g
or
s.
w
ey.
vinl doramectin produced selamycin. A strain of Streptomyces cyanogriseus was found to produce a
eo
ualruse macrocyclic lactone that was developed as nemadectin; derivitization of nemadectin produced
nnal moxidectin. Milbemycin oxime was developed not from an avermectin but from an older
an
m www.or perso maspacercotscyocflitchelagcrtoounpehstarvuectbuereen,dmoiclbuemmeynctiend.iTnhceosmeparnedhemnsainvyemotuhletri-cbhaasipctearnrdevaipepwlsie(d9–s1c1ie)n.tific
oF Themacrocycliclactoneclasssooncametodominatethecontrolofparasiticdiseasesoflive-
ded fr3/16. stockandcompanionanimals.Becausethedrugsareeffectiveagainstbothendo-andectoparasites,
nloa11/2 theyareknownbytheinfelicitousbutconvenientterm“endecticides.”
wn Ivermectin is the macrocyclic lactone with which I am most familiar, and experience with
oo
D9 ivermectinisthemainsourceofthelessonsofferedhere.Itwasclearalmostfromthebeginning
1-14. 195.8 that, in financial terms, the potential value of ivermectin in the animal-health market was very
4:4. large and in the human-health market was very small. Nevertheless, the drug has been widely
016.0.25 distributednotonlyforlivestockandcompanionanimalsbutalso(inanon-marketcontext)for
27
Biosci. ded by humans.
m. ovi TheValueofCastingaWideNet
Anis pr
v. ces Thediscoveryofivermectinresultedfromscreeningmicrobialfermentationproductsforantipar-
ec
RA asiticactivity.Themicrobesthemselvesweregatheredfromawiderangeofsourcestoenhance
nu. the diversification of assay input. Among those sources was the Kitasato Institute in Tokyo. A
n
A groupofKitasatoscientists,ledbyProfessorSatoshiOmura,routinelyisolatedmicroorganisms
fromsoilbyallowingthemtogrowinlaboratoryculture,afterwhichtheyweretestedforactivity
againstavarietyofothermicroorganisms.UnderanagreementwithMerck&Co.,Inc.,isolated
microbesthatwereconsideredunusualinappearanceorculturalcharacteristicsweredispatched
toMercklaboratoriesintheUnitedStates.Whenabatchwassentin1974,itwasunderstoodby
MerckmicrobiologiststhatthoseisolateshadshownlittleantimicrobialactivityintheKitasato
tests, and that was in accord with the results of routine antimicrobial tests done at Merck. In
1975,theisolatesweretransferredfromtheMerckMicrobiologyDepartmenttotheParasitology
Department,wheretheycouldbetestedinanewassaythathadbeendevisedespeciallyfortesting
microbiologicalfermentationsforactivityagainstparasites.Oneofthoseisolates,whenregrown
intheMercklaboratoriesandtestedinthenewantiparasiticassay,yieldedapotentanthelmintic
substance.Thatsubstance(amixtureofabamectinandseveralrelatedstructuresknowncollec-
tivelyasavermectins)wouldbetheforerunnerofivermectinand,byextension,themacrocyclic
4 Campbell
AV04CH01-Campbell ARI 10January2016 8:3
lactoneclassofantiparasiticagents.Theself-evidentlessonisthatincreasingthemagnitudeand
diversityofscreeninginputwillenhancetheprobabilityofscreeningsuccess.
TheValueofManagersandTechnicians
Thoselookingfornewdrugstodayareunlikelytobesolitaryworkers;theyarelikelytodepend
on the assistance of technicians for the quality of their research and on managers or directors
forthecontinuedexistenceoftheresearch.Directorsandmanagersallocateresources,financial
andotherwise.Techniciansmaybescientificallytrained,butincasesinvolvingroutineprojects,
suchasscreeningoperations,manyofthetechnicianslearnonthejob.Managersandtechnicians
shouldofcoursebeaccordedduerecognitionandauthorshipforidentifiableandspecificscientific
contributions.Aswasevidentinthecaseofivermectin(12),theoutcomeofresearchwilloften
dependcruciallyonthewisejudgmentofthosewhodeterminestrategyandtactics,andonthe
g steadfastnessoftechnicianswhoremainalertthroughoutrepetitiveoperations.Whereauthorship
or
s. isnotappropriate,otherformsofrecognitionarewarranted.Tobendatropefromanillustrious
w
ey.
vinl nonscientist,theyalsoservewhoonlyhoveraboveorlaborbelow.
eo
ualruse Managers must confront imponderables. In developing a new chemical class intended for
nnal generaluse,howsoonshouldamemberoftheclassbesubjectedtoanAmes(teratogenicity)test?
an
www.perso Htootwurlnonugp(“aancdtivheosw”?mInucsuhc)hshcoirucludmrsetsaonucrecse,sabjeudingvmeestnetdminusstubpepomrtaidnegoannoasthsaeyrtthhaatnhsacsiefnatiilfiedc
om For grounds.Commonsenseandexperienceinexperimentalchemotherapywillhelp,butscienceitself
d fr16. willnot.
de3/
a2
nlo11/
owon TheValueofAssayInnovation
D9
4:1-14. 4.195.8 Terhyeodfispceonviecrilylionf,imvearnmyencetiwnraenstuibltieodticfrsowmeraessafoyuinndnobvyatsicorne.eInnintghefedremcaednetsatfioolnlopwriondgutchtesdagisacionvs-t
016.0.25 microorganismsinvitro.Yetitwouldappearthatnotasingleanthelminticwasfoundbythepri-
Biosci. 2ded by 7 mnoarsyuistcarbeleenainssgayofthfeernmeexnitsatetido.nTphreodburcetaskathgaroinusgthpatrhaastityiciewldoerdmisv.eTrmheacttwinaswparsesaunminabvlyivobeacsasuasye.
m. ovi Detailsoftheassayhavebeenpublished(4,7,11).Inessence,fermentationproductswerefed
Anis pr to worm-infected mice, which were later examined for evidence that the infection still existed.
v. ces Theempiricalnatureofthesystemisabundantlyevident:Themicewerefedanarbitrary,though
ec
RA standard,amountoffoodcontaininganunknownamountofanunknownsubstancethatmight
nu. notbethere(13).Thelessonisthatthewantofaknownassaytechnique(evenawantoflong
n
A standing)shouldnotdissuadeanyonefromtryingtocreateanassaytofittheneed.
Liabilities,RealandVirtual
Inthedevelopmentofabroad-spectrumanthelmintic,lackofefficacyagainstamajorpathogenic
speciesisgenerallyseenasaliability;itiscertainlynotlikelytobeareasonforcelebration.Yet
suchwasthecaseinthedevelopmentofivermectinforthe“worming”ofdogs.Therecalcitrant
parasiteinquestionistheadultheartworm,Dirofilariaimmitis.Itisnotkilledbyivermectineven
thoughthedrugreachesitthroughthehostbloodstream.Theadultstageistheprimarypathogen
indirofilariasis,buttreatmentofdogstogetridoftheadultishazardoustotheinfecteddogand
tothereputationoftheveterinarian.Itmustbedone,andcanbedonesafely,butitshouldnotbe
theincidentalresultofroutineworming.Ivermectinandothermacrocycliclactonesarecrucially
importantinthecontrolofheartwormdisease;yet,inanapparentlymade-to-orderanomaly,their
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AV04CH01-Campbell ARI 10January2016 8:3
successisduetotheirpotencyagainsttheessentiallynonpathogeniclarvae(microfilariae).Lack
ofefficacyagainsttheadult(pathogenic)stage,anapparentliability,isarealasset.
For the routine anthelmintic treatment of dairy cattle, lipophilicity in a drug is almost sure
tobealiabilitybecauselipophilicdrugsandtheirmetabolitesarelikelytobeeliminatedatleast
partlythroughthemilkoftreatedanimals.Theresultisthatthemilkmustbewithheldfromthe
marketuntilitisessentiallyfreeofdrugresidues(meetingtherequirementsofrelevantregulatory
agencies).Ivermectinisdistributedgenerouslyinthelipidcomponentoftreatedcattle,resulting
inanundesirablylong“withdrawalperiod”(14).Escapefromthisliabilitywasachievedbytesting
ivermectinderivativesinlactatingcattletodeterminetheirdistributioninmilk,andthentestingthe
morepromisingcandidatesforanthelminticefficacy.Inthiswayeprinomectinwassuccessfully
developed for use in dairy cattle as well as other livestock (15) and was later developed as an
exceptionallong-lastinginjectableproductforcattle(16).
It is clear that drug characteristics that are usually liabilities are not always liabilities. And
g things that really are liabilities may sometimes be converted into assets. There is, however, an
or
s. important warning to be made in respect to chemotherapeutic agents in general, a lesson that
w
ey.
vinl became apparent as a result of the unexpectedly wide spectrum of ivermectin: The wider the
eo
ualruse spectrumofefficacy,thegreatertheneedforenvironmentalvigilanceinthepracticalapplication
nnal oftheagent.
an
www.perso
m or ChallengingConventionalWisdom
oF
d fr16. When the antiparasitic efficacy of ivermectin was initially announced, there were experts who
ade23/ dismissed it as a curiosity on the grounds that it could never be produced economically in the
wnlon 11/ amounts needed for practical use. It was indeed true that large-scale synthetic production of
oo
D9 ivermectin was not feasible, but the filamentous bacterium Streptomyces avermitilis, the natural
4:1-14. 4.195.8 s(foruormcewohficthheivcehrmemecictianlicslamssa,dceobuyldchbeemciacjaollmedodinifitocaptiroond)u,csoinpgroudnuncattiuornalbyamfeormunetnstoatfioanbabmeceacmtine
016.0.25 standard.
Biosci. 2ded by 7 byAthneoirtheexrpleesrstiosneoinfwtohivcihewwiengarenerewmdinedveeldobpymtehnetisveinrmaecletisns-stahgaani-setxhpaetretxwpearyt.sAmtayabteimbeetrwahyeend
m. ovi a macrofilaricide (roughly “adulticide”) was a recognized need (as it still is) for the control of
Anis pr riverblindness,thevalueofamicrofilaricide(roughly“larvicide”)wasdoubtedeveninthefaceof
v. ces evidenceofitspotentialvalue.Thedanger,ofcourse,isthatmisplacedskepticismmightdiscourage
ec
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u.
n
n
A
TheValueofFlexibleClinicalObjectives
In disease control, causal prophylaxis would seem a priori to be the ideal. Yet clinical prophy-
laxis (prevention of disease without eradication of infection) is seen over and over again to be
ofgreatvalue.Indeed,clinicalprophylaxisisclosertothebiologicalideal.Undoubtedly,human
squeamishness (together, in some cases, with a modest health penalty) would be a deterrent to
clinicalprophylaxisinthehumancontext.Eveninthecaseofimmunologicalclinicalprophylaxis
ofhookwormdiseaseindogs,itwassociologicalandeconomicratherthanscientificfactorsthat
blockedtheacceptanceofavaccine(17).Theacceptanceofivermectininthecontrolofhuman
riverblindnessatteststothevalueofclinicalprophylaxis.
Riverblindness(onchocerciasis)isoneoftwomajortropicaldiseasescausedbyroundworms
knownasfilariae.Inriverblindness,thefirst-stagelarva(microfilaria)istheprimarypathogen.
In the other disease, lymphatic filariasis, the adult worm is the primary pathogen. In both,
6 Campbell
AV04CH01-Campbell ARI 10January2016 8:3
ivermectinislethaltothemicrofilariaebutdoesnotkilltheadultworms(thoughitmaysuppress
theirreproduction),andtransmissiontohumanscanbeblockedbyivermectintreatmentbecause
microfilariaewillnolongerbepresent(inskinorblood,respectively)tobepickedupbytheinsect
vector(blackflyormosquito,respectively).Inthecaseoflymphaticfilariasis,diethylcarbamazine
can similarly be used to kill microfilariae, and (whether diethylcarbamazine or ivermection is
usedasmicrofilaricide)albendazoleisadministeredconcurrentlytokilladultwormsandthereby
minimizemorbidity.
In several regions of Africa, Central America, and South America, river blindness has been
brought under control by repeated administration of ivermectin to kill the larval stage of the
parasite. Ecuador and Mexico are examples of countries in which the disease has actually been
eliminated (18,19). Forlymphatic filariasis,controlprograms have reached the pointat which
a plan has been developed for elimination of the disease from the vast tropical regions where
thediseaseisendemic.Thedonationofivermectinandthepracticalramificationsofitsusein
g internationalcontrolprogramshavebeendiscussedbymany(see,forexample,20).
or
s.
w
ey.
vinl
eo LESSONSFROMTHEGENERALHISTORYOFANTIPARASITICDRUGS
ualruse
nnal Patternsandapproachesinthefieldofnewdrugdiscoveryhavechangedinrecentyears(21–23),
an
www.perso yoeftthleesstownesnmtieatyhbceendteurriyve.dI hfraodmththeegogoendefroarlthuinsetotroyboefainnvtohlevlemdi,nitnicotnheerwapayyionrtahneotsheceor,nidnhtahlef
om For development not only of ivermectin but also of thiabendazole, cambendazole, rafoxanide, and
d fr16. clorsulon.Thelessonsofferedheremustowesomethingtothatexperienceandtheexperienceof
ade23/ workingonpotentialdrugsthatfailedtofulfilltheirpotential.Itisnotalwayspossibletolinka
wnlon 11/ particularlessontoaparticulardrug.
oo
D9 Thiabendazole was the first of the benzimidazoles, which were to become the blockbuster
4:1-14. 4.195.8 binroaasdsa-yspseccotrnudmucatnedthbelymminyticcoslolefathgeuierdDary.(2Jo4h).nItRsa.nEthgeelrmtoinn,tiucseifnfigcanceymwaatsoddiescloavrveareedinemviptirroicaanlldy
016.0.25 nematode infections in mice. Dr. Cuckler invited me to share the excitement of exploring its
Biosci. 2ded by 7 terfifcehctinsoosnisv.a(Triohuissmstaargkeesdotfhveabreioguinsnrionugnodfwaonremwsepreacfioers,mesepaetctihaellymTerreiclhyipneelrlsaosnpairlalleivs,etl.hIehaagdebneteonf
m. ovi lookingforwardtodevotingmycareertotheflukediseaseschistosomiasis,butIsoondiscovered
Anis pr thatbiodiversitycanbeappreciatedatthelaboratorybenchaswellasinthegreatoutdoors.)
v. ces
ec
RA
u. SomeChallengesofAssaySelection
n
n
A Inthefieldofinfectiousdiseases,infectedlaboratoryanimalshavelongbeenusedforthedetection
ofefficacioussubstances.Suchscreeningisprofligateoftestmaterialandlaboratoryresources,yet
invivoscreeninghastheadvantageofgivinghintsastothebioavailabilityandsafetyofanynew
activecompound,aswellastheadvantageoftestingcompoundsagainstallaccessiblebiological
systemsofthetargetparasiteatonce(unknownsystemsaswellasthoseknown).
AsEhrlichfoundmorethanacenturyago,adirectcorrelationbetweenefficacyinvivoandeffi-
cacyinvitrocannotbeassumed.Comprehensivecomparisonofinvivoandinvitroassayscannotbe
undertakenhere,noristhisaplacefordiscussionoftheethicalaspectsofinvivoexperimentation.
Oneparticularaspect,however,haspracticalimplicationsfortheexperimentalchemotherapist.
As ethical concern about the use of animals in research expands, the use of in vivo assays con-
tracts.Thismaythereforebeasuitabletimetoraisethequestionoftheappropriatesizeofgroups
ofexperimentalanimals.Muchattentionhasbeendevotedtothedesirabilityofreducinggroup
size. The ultimate reduction (a single animal per test substance) was pioneered by Ostlind in
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AV04CH01-Campbell ARI 10January2016 8:3
small-animal screening in the parasitology laboratories at Merck & Co. and was shown to be
perfectlysatisfactoryinseveralassays(25).Thekeytosuccessisnear-perfectcontroloftheexper-
imentalinfection,andthelessonisthis:Inawell-regulatedassay,oneissufficientandtwoasurfeit.
A much-neglected weakness of the in vitro assay is the inherent unpredictability of effects
associated with the ambient medium. The salinity of the medium, for example, may affect the
sensitivityoftheassay,andtheeffectisnotnecessarilyconsistentbetweendifferentdrugclasses
(26).
LessonsfromNewActives
Whenanactivecompoundisturnedupbyascreen,itsbiochemicalreactivityshouldbeconsidered.
Inthenormalcourseofevents,theextenttowhichthemodeofactionshouldbeexploredwillbe
determinedbyseveralfactors,includingtheperceivedpotentialofthecompoundforclinicalor
g commercialutility,thenovelty(patentabilityandprospectforbasicresearch)ofthecompound,and
or
s. thereputationofthatclassofcompoundfornastiness(isitknown,forexample,forteratogenicity
w
ey.
vinl orstench?).
eo
ualruse Iftheactivecompoundachievesthestatusofclinicalorcommercialutility,orevencomesclose,
nnal thenthemodeofactionwillnaturallysuggestitselfasausefultargetforfurtherdrugdiscovery.The
an
www.perso tTehmeprteaitsioanhtioghfopllroowbatbhiilsitlyintehaotfcthhoemugihcatlinsttrouacctutirveesrsehsaerairncghashgoivuelndbbieoacphpemroiaccahlemdewchitahncisamutiwoinll.
om For shareboththeassetsandtheliabilitiesoftheoriginal,sothatdrugssodevelopedwilllacktrue
d fr16. noveltyandwilllikelybesubjecttoresistancealreadyacquiredbytargetpathogens.Newactives,
ade23/ aswellasestablisheddrugsandabandonedleads,shouldberegardedaspotentialtoolsforbasic
wnlon 11/ researchaswellascandidatesfortherapeuticapplication(13).
oo
D9
1-14. 195.8 LessonsfromWeakorAbandonedActives
4:4.
016.0.25 Inempiricalrandomscreening,failuretofindanactivesubstanceisthedefaultday-to-dayout-
Biosci. 2ded by 7 coofmlitet.leCionntefirremstatbolethaceticvheesmaorethrearraep.iAstm.Honisgtotrhiecmallya,renesuitbhsetranacuethsothrsatn,odreespdiitteortshehiarvaecbtieveitny,parere-
m. ovi paredtoclutterupthepagesofjournalswithlistsofcompoundsthatwereinactiveormarginally
Anis pr activeinempiricalscreens,andtheproperdispositionofabandonedactivesremainsdifficultto
v. ces resolve. The enhancement of leads through molecular alteration or developmental research is
ec
RA anestablishedfeatureofexperimentalchemotherapy.Butinrandomscreening,aweakactiveis
nu. oftendismissedwithoutbeingaccordedthestatusoflead.Someofthemareabandonedbecause
n
A efficacy is weak at maximal tolerated dosage (though often it has not been feasible to test this
point adequately). Others are abandoned because they are found to be members of a chemical
classtaintedbytoxicity.Stillothersaredroppedfromconsiderationforreasonspeculiartothe
particularactivesubstance.Questionableleadscannotallbepursuedindiscriminately,butthey
shouldnotbediscardedhastily.
In the 1950s, a particular compound was weakly but reproducibly active against Schistosoma
mansoniwhengiventomiceaccordingtoaspecificprotocol(27).Ibelieveitwillnotbeabetrayal
of a significant secret to identify the substance, some half-century after the fact, as O-methyl
threonine.Thediscoverywasgreetedbymymoreexperiencedcolleagueswithastunninglackof
interest.Itdidnotseemworthwhiletopublishafindingsodestituteofpromise,buttothisdayI
wonderifthatbitofinformationmightsometimebeusefultosomeonesomewhere.
If there is a lesson here, it may be that actives of little apparent value should be published.
Indeed the suggestion has been made that inactives should be made public to reduce pointless
8 Campbell
AV04CH01-Campbell ARI 10January2016 8:3
screeningbyothers.Inthiseraofinformaticsandthepostingofmassiveamountsofinformation
insupplementaldepositories,thisquestionmightwarrantrenewedattention.
TheLessonof“Inactives”ThatAreActive
A parasitologist once told me with pride that he had tested thiabendazole against T. spiralis in
rodents before I had done it. He added that he had not taken the matter further because the
drughadbeeninactive.Heprobablythoughtitunnecessarytomentionthatwhathemeantto
say was that the drug had been inactive when given at the dosage and in the regimen he had
chosenforhisexperiment.Itisseldomunnecessarytomentionsuchinformation.(Theefficacy
of thiabendazole in rodent trichinellosis is widely recognized.) In random-screening assays the
vastmajorityoftestedsubstanceswillbe“inactives”eventhoughsomeofthemmighthavebeen
recorded as “active” if they had been tested at dosages higher than that selected arbitrarily for
g thatparticularassay.Screeningatmultipledosageswoulddomuchtooffsettheriskbutisalmost
s.or alwaysrejectedasprofligateofassayresources.Theriskof“falsenegatives”is,forthemostpart,
w
ey. acceptedasabuilt-indefectofrandomscreening.
vinl
eo
ualruse
nnal TheLessonofMultipleBiodynamicActions
an
www.perso Discoverythatananthelminticcausesaparticularbiochemicalactiononaparasitedoesnotmean
m or thatthemechanismofanthelminticactionhasbeenfound.Thiabendazole’sefficacyappearsto
oF
d fr16. resideinitsdisruptionofmicrotubuleformationratherthaninitsinhibitionoffumaratereductase.
de3/ Theparalyzingeffectofivermectinonnematodesappearstoresultfrombindingtoaneuronal
a2
nlo11/ glutamatereceptorratherthanfrompotentiationofthereleaseandbindingoftheneurotransmitter
wn GABA(gamma-aminobutyricacid).Othercompoundshavecholinergiceffectsthatmayormay
oo
1-14. D195.89 nleothtableepffreicmtaornyp(2ar8a)s.iDticrungesmmataoydaelssobuhtaavlesomhualstiapnltei-cilninfliacmalmefafteocrtys.aTcthiviaitbyeinndmazaomlemnaolts,oannldyihtahsaas
016.4:0.254. beenwidelyusedasanagriculturalfungicide.
27
Biosci. ded by TRADITIONALDISCOVERYSYSTEM
m. ovi GoodbutNotGreat
Anis pr
v. ces Throughoutthecenturies,drugswerediscoveredempirically—thatis,bychanceclinicalobser-
ec
RA vationortrial-and-errortesting.Itisimpossibletosaywhendeliberateattemptswerefirstmade
nu. to test a variety of substances in the hope that a particular thing might have a particular ther-
n
A apeutic effect, but Redi’s seventeenth-century experiments, in which he dunked parasitic (and
nonparasitic)wormsinto various liquids,may well bethe earliestonrecord(29). Aprehistoric
parentmighthavebeenthefirsttoplungeawormintoajarofalcoholtoreinforcethelesson
thatdrinkinghardliquorcanbelethaltothedrinker.Onlyaboldandastutechildwouldhave
counteredthatevidentlypeoplewithwormswouldbewelladvisedtodrinkalcohol.[Undervery
limitedcircumstancestheremaybeatraceoftruthinthat(30).]
Inbothveterinaryandhumanmedicine,anthelminticsareusedaschemoprophylacticagents—
providingclinicalprophylaxisbymeansofroutineherdtreatment.Somepoliticalandeconomic
aspectsofagriculturalusehavebeenreviewed(22).Theessentialgoalofthechemotherapistre-
mainsthediscoveryofsubstancesthatadverselyaffectparasiticworms(orrenderthemharmless,in
whichcasetheyarenolongerparasiticworms).Thetwotraditionalapproachestothisobjectiveare
empiricaltesting,asmentioned,andtheidentificationandexploitationofbiochemicaltargets.For
manydecadesitwasinculcatedinthemindsofmanythattheempiricalapproachwasintrinsically
www.annualreviews.org • AntiparasiticAgents 9