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PHAR6113

Lecture2:Therapeu.cDrugMonitoring

MasterofPharmacyProgram,UniversityofNewcastle

2011 edition

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.............................................................................Learning Outcomes: 3...........................................................................................Introduction 3

..............................Which drugs do we monitor using blood levels? 4...............................................What are indications for TDM testing? 7

.............................................................................How do we do TDM? 7.................................................................When do we take samples? 7

Measuring drug (and metabolite) concentrations: Assay techniques

...................................................................................commonly used: 8................................................................................HPLC and GC techniques: 8

....................................................................................................................FPIA: 9.....................................................................................................................EMIT 9

......................................................................................................................RIA: 9...............................................Comparing different assay methods: 10

EVALUATING THE ANALYTICAL TECHNIQUES USED AND

..............................................................COLLECTION TECHNIQUES

10...........................................................Containers used in collecting sample: 10

..................................................................................Processing and Storage 11.......................................................................................Sample identification 11

..........................................Free or total drug concentration measurements 11....................................................................Analytical technique evaluation: 12

........................................................................Sampling time in TDM 13.......Factors to consider when interpreting TDM results clinically 13

.....................Patient age, medical condition and other medication 13..............................Common Errors in TDM interpretation include: 14

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......................................................................Web resources to visit: 14.........................................................................................References: 14

.............................................................................Review Questions: 14LearningOutcomes:

Explainthedifferentwayswecanmonitordrugtherapy

Explainwhatmakesadrugagoodcandidatefortherapeu

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Letsconsidersomeclinicalscenarios.

Scenario1:

MrsSmithvisitsherPandherbloodpressureiselevateddespitelosingweightandregular

exercise.ThePisgoingtocommenceheronanan

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bymeasuringorobservingaclinicalresponse

byaninvitrotestthatindicatestherapeuGceffect

bymeasuringblood/plasmaconcentraGonandhavingatargetrange(TDM)

Whatsortofclinicalresponsescanwemeasure?

IfwehaveapaGentwhohaselevatedbloodpressure,wewillstartthemonanappropriate

anGhypertensivedrugandbymeasuringbloodpressurechangesandsideeffects,willalter

dosingofthedrugtoachieveopGmalbloodpressurereadings.ApaGentswithParkinsons

diseasemayrequiredrugssuchasacombinaGonofDOPA/Carbidopa.Wewouldcommencethe

paGentonthismedicaGonandwoulddecreasethedoseifwesawdyskinesiasorincreasethe

doseifweobserveporrcontrol.Ifweobservedconfusionordepression,thisisasignof

possibletoxicity.IfwewereusingadrugsuchasthiopentonetoanaestheiseapaGent,we

wouldincreasethedoseifwehaveinsufficientanaesthesiaanddecreasethedoseif

anaesthesiawastoodeep.Signoftoxicitywouldberespiratoryfailure.

Whenwouldweuseaninvitrotestoftherapeu.ceffect?Herearesomeexamplesofdrugswherewechangedosebasedonaninvitrotestof

therapeuGceffect:

Drug IndicaGon decreasedose increasedose toxicsigns

Warfarin TEdisease highINR lowINR Bleeding

Thyroxine Hypothyroidism lowTSH highTSH Hyperthyroidism

StaGn Raisedcholesterol raisedAST/CK highTC Myopathy

Whendoweusemeasurementoflevelsinblood/plasma?

WeareusingadrugtotreatpaGentsinaclinicalse]ng.Fromresearchstudies,ithasbeen

shownthatthereisagoodcorrelaGonbetweenconcentraGonofdruginplasmaandtheeffect

wewanttoachieve.FromthestudieshoweverweseethatthedrughasanarrowtherapeuGc

index.TheminimumeffecGveconcentraGonformostpaGentsis2mcg/mLandweseetoxicityin

manypaGentswhenwegoabove4mcg/mL.AddiGonally,thetoxiceffectsareseriousandcan

harmthepaGentsoavoidingtoxicityisessenGal.Tomakedosingmoredifficult,whenweadminister50mgdosestoallpaGentsweseedramaGcdifferencesinbloodlevelsachievedand

somepaGentsshownoresponse,someshowresponseandsomeshowsignsoftoxicity.We

havewideinterindividualvariabilityinbloodlevelsachievedwiththeSAMEdose.

Thisdrugisagoodcandidatefortherapeu

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AnexampleoftheuseofTDM:

PriortouseofTDM,phenytoinwasprescribedempiricallyasa300mgdailydose.PaGents

eitherresponded,showedsideeffectsordidnotrespond(sGllhadseizures).Withtheadventof

monitoring,itwasfoundthatwhenthisdoseisadministeredinalargegroupofpaGents,there

isagreaterthan10foldvariaGoninplasmadrugconcentraGonsobservedinthesepaGentsas

showninplotbelow.

WithTDM,itispossibletoindividualisedosesandnowdosesbetween150700mg/dayarenowused.Theadvantageofphenytoinmonitoringisbeingabletofinelyadjustdoseonanindividual

basisforadrugthatshowslargevariabilityinpharmacokineGcs.ThemeasureofeffecGveness

ofthisdrugiscessaGonoffi]ngnotaneasypharmacologicaleffecttoquanGfy.Researchwork

hasalsowellestablishedtheopGmumrangeofbloodconcentraGons(therapeuGcrange).This

drugisevenmorecomplicatedtoadjustdosethanusualasdrugshowsnonlinearkineGcs.We

willdiscussnonlinearkineGcsinfuturelectures.

RememberwithTDM,plasmaconcentraGonsalwaysactasaguideandareusedinconjuncGon

withclinicalobservaGonsandresponse.RememberthatasmallpercentageofpopulaGonare

likelytoshowadverseeffectswithinthetherapeuGcrangewhilesomemayrequirehigher

concentraGons.Manyresponsesaregradedeffecttoxiceffectsdonotjustmagicallyand

suddenlyappearusuallyadverseeffectsarealsoagradedresponse.

Tosummarise,monitoringisgenerallydonefordrugswhich: haveanarrowtherapeuGcindex, havelargeinterindividualvariaGoninpharmacokineGcs toxiceffectsaredramaGcand forwhichthereisgoodcorrelaGonbetweeneffectandconcentraGonandatherapeuGc

rangehasbeendetermined.

Wherewillyouseetherapeu0cdrugmonitoringusedineverydayprac0ce?

Herearesomecommonexamples

AminoglycosidesusedtotreatseriouslifethreateninginfecGons

Drugssuchasgentamicinmayproducenephrotoxicity(damageotkidneys)andototoxicity(loss

ofhearing).ThereisarelaGonshipbetwenconcentraGonsachievedandeffectandtoxicity.

An

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DigoxinisusedincardiaccondiGonssuchasheartfailureandatrialfibrillaGon.Thereisa

definedtherapeuGcrange.ThisdrugcanshowmarkedinterindividualvariaGoninbloodlevels

achievedatthesamedose.Signsoftoxicityincludenonspecificonessuchasnausea,vomiGng,

abdominalpainandconfusion,butcanalsohaveseriouscardiaceffect.

LithiumisusedintreaGngbipolardisorders.IthasanarrowtherapeuGcindex.Thereiswide

interindividualvariaGoninplasmalevelsachieved.Signsoftoxicitycanrangefromtremorto

ataxiaanddiarrhoeatoconvusionsandfits.

Nowtakeamomenttoconsiderthe4casescenariosatthebeginningofthislecture.Which

oneswouldrequiretherapeu

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bloodlevelsonconGnuedadministraGonunGlwegettoaplateau.Ideallywewantbloodlevels

toplateauinthetherapeuGcrange.

Measuringdrug(andmetabolite)concentra

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detectorwhichmeasurestheamountofdrugeluGng.Differenttypesofdetectorsmaybeused

includingUV,electrochemical,fluorescenceandMassspectrometry.

HerearesomelinkstousefulwebsitesonHPLC:Takeamomenttoviewthesepages

hp://www.waters.com/waters/nav.htm?cid=10048919&locale=en_US

hp://www.youtube.com/watch?v=ICdTU5X4HA

GasChromatography(GC):

Cisasimilarprinciplewiththemobilephasebeinggasnotaliquid.VerysensiGvetechniques

employmassspectrometryasdetector.

Hereisalinktoresourcesexplaininggaschromatography:

hp://www.youtube.com/watch?v=dffeiLgeKx 8

FPIA:(FPIA)isatechniquewhichtakesadvantageoftheincreasedpolarizaGonoffluorescentlight

emissionswhenafluorescentlylabeledanGgenisboundbyreagentanGbody.Thehigherthe

concentraGonofunlabeledpaGentanGgenpresentinthetestmixture,thelessbound

fluorescentanGgenispresentand,consequently,thelowerthepolarizaGonofthefluorescent

lightemission.

LabelledanGgen(inotherwordsdruglabelledwithafluorophore)isaddedtothesampleyou

wishtoassay.AnGbodyisalsoaddedtothesample.Thedruginthebiologicalsamplecompetes

withthelabelleddrugfortheanGbody.Ifthereisliledrugpresentinthebiologicalsample,

theaddedlabelleddrugwillbindtoanGbodyandtherewillbehigherpolarizaGon(higher

readings).Ifalotofdrugispresent,lileofthelabelleddrugwillgetaachedtoanGbodiesand

sowewillseealowerreading.Byusingsampleswithknownamountsofdrug,acalibraGon

curveisproducedandtheresultoftheclinicalsamplecanbereadoffthecalibraGoncurve.

Herearesomeusefullinks:

hp://www.boomer.org/c/p3/c03/c0309.html

EMIT

Thisisaprocedureinwhichthedrugbeingmeasuredcompetesforalimitednumberof

anGbodybindingsiteswithenzymelabelleddrug.Toabiologicalsampleisaddeddruglabelled

withenzymeandanGbodytothedrug.Onlyenzymewhichisnotlinkedtoboththedrugand

anGbodycanreactwithanaddedsubstrate.(TheanGbodyhastheabilitytoblockenzymaGc

acGvitywhenboundwiththereagentenzymedrugcomplexprevenGngit'sformaGonofproductinthepresenceofsubstrate).Asubstrate fortheenzymeisaddedandacolorchangeis

produced.ThiscolourchangeisproporGonaltotheconcentraGonofdrugpresentinthe

specimen.

RIA:

Thedrugsample,labelleddrugandanGbodytodrugaremixedandincubated.Thebounddrug

(bothlabelledandunlabelledfromsample)areseparatedandtheradioacGvitycounted.The

http://www.boomer.org/c/p3/c03/c0309.htmlhttp://www.youtube.com/watch?v=dffeiLgeKx8http://www.youtube.com/watch?v=I-CdTU5X4HAhttp://www.waters.com/waters/nav.htm?cid=10048919&locale=en_UShttp://www.boomer.org/c/p3/c03/c0309.htmlhttp://www.boomer.org/c/p3/c03/c0309.htmlhttp://www.youtube.com/watch?v=dffeiLgeKx8http://www.youtube.com/watch?v=dffeiLgeKx8http://www.youtube.com/watch?v=I-CdTU5X4HAhttp://www.youtube.com/watch?v=I-CdTU5X4HAhttp://www.waters.com/waters/nav.htm?cid=10048919&locale=en_UShttp://www.waters.com/waters/nav.htm?cid=10048919&locale=en_US

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moredrugintheactualsampleadded,thelesstheradioacGvityreadingaslessoftheadded

labelleddrugcanbindtotheanGbodiespresent

Comparingdifferentassaymethods:

TechniquesinvolvinguseofanGbodyanGgeninteracGonsmayshowcrossreacGvity.

Herearesomeusefulwebsitestolookat:

hp://www.clinchem.org/cgi/content/full/48/3/507

hp://books.google.com.au/books?id=1bv0cvkxklsC&pg=PA146&lpg=PA146&dq=digoxin+

+interference&source=bl&ots=vKs7bNdpZq&sig=0whutcgt8otFy

YPRQnVKquVg&hl=en&ei=F4eTYijJo3RccavmOgK&sa=X&oi=book_result&ct=result&resnum=7

&ved=0CEMQ6AEwBjge#v=onepage&q=digoxin%20%20interference&f=false

EALUATINTHEANALYTICALTECHNIQUESUSEDAND

COLLECTIONTECHNIQUES

Containersusedincollec

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problemswithassays.SometubescontaingelbarriersserumseparaGngtubesandthisgel

barrierhasbeenreportedtoadsorbquiteafewdrugs.Theamountofadsorbeddependedon

Gmeofcontact,theamountofsample,drugconcentraGonpresentinvial,temperatureand

lengthofstorage.Inotherinstances,chemicaladdiGvesaddedtoenhanceclo]ngorprevent

adsorpGonintotubesorplasGciserspresenthavebeenfoundtointerferewithanalyses.

ItisthereforeessenGalthatthesamplesarecollectedintothecorrecttypeoftube.Laboratorieswillusuallyspecifythetypeoftuberequired.

VisitthewebsitebelowforHAPSandseewhattheyspecifyfordrugssuchasdigoxin,lithium

andphenytoin

hp://www.haps.nsw.gov.au/Handbook/hh02.aspx?test=197

ProcessingandStorage

Itshouldneverbeassumedthatonceasampleiscollected,thedrugwillbestableinthetube.

Blood,plasmaandserumarecomplexmatricesandsamplesshouldbetestedassoonas

possibleaercollecGon.IftheycannotbeassayedrelaGvelyquickly,samplesshouldbeappropriatelyhandled(usuallycentrifugedtosparateplasma/serum)andthenrefrigeratedor

frozen.Asanexample,paGentsongentamicinareoenonotheranGbioGcssuchasbeta

lactams.WeavoidcomixingandadministraGonofthesedrugsbecausetheyinacGvateeach

other,sowhentheyareadministeredoneisgivenandthentheotherisgivensotheydonot

physicallymix.Forthelaboratorythismeansthatbothdrugsarepresentinthebloodsample.if

thebloodsampleisleatroomtemperatureandbothdrugsarepresentinthesample,then

theaminoglycosideconcentraGonwilldeclinesignificantlyataratedependentonthe

aminoglycosideandthebetalactampresentaswellasGmeandtemperatureofexposure.

Becausewedonotknowifbotharepresentandtowhatextent,thesesamplesshouldbe

separated(spindownincentrifuge)andanalysedassoonaspossibleorfreezesampleiftesGng

isdelayed.

Sampleiden

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OnetechniqueusedisultrafiltraGon.Withthistechnique,theplasmaorserumcontainingboth

freeandbounddrugisplacedinadevicewhichcontainsamembrane.Thedeviceiscentrifuged

foracertainperiodofGme.Themembranedoesnotallowplasmaproteintopassthroughbut

druginextracellularwater(unbound)cangothrough.Theultrafiltrateisthenassayedto

determinethefreedrugconcentraGon.

Analy

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performedandresultssenttoexternallab.Reportsreceivedbackassistindeterminingwhether

assayisperformingcorrectly.

Sampling

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CommonErrorsinTDMinterpreta

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5. Writeasummaryoftheadvantagesanddisadvantagesofeachofthesetechniques

listedabove

6. ExplainthedifferencebetweenmeasuringtotaldrugconcentraGoninasampleandfree

drugconcentraGon

7. DiscussfactorsthatneedtobeconsideredwhenevaluaGngtheassaytechniqueusedin

TDM

8. WhatdowemeanbyQualityControlsamples?

9. DiscussfactorsthatneedtobeconsideredwheninterpreGngTDMresultsclinically

10. Discusswhythewaythesampleiscollectedandhandledisimportant

11. ListsomedrugsforwhichTDMisperformed

SELFASSESSMENTTASK:

OOD FAIR POOR

ExplainingwhenweuseTDMandwhenwedo

not

Discussingfactorsthatmakedruggood

candidateforTDM

Describingprinciplesofdifferentassay

techniquesused

Discussingadvantagesanddisadvantagesof

differentassaytechniques(compare/contrast

methods)

DiscussfactorstoconsiderwhenevaluaGngthe

assaytechniqueused

DiscussfactorstoconsiderwheninterpreGng

resultsfromTDMlaboratory

DiscusssamplingGmesandsamplehandling

ListdrugsforwhichTDMiscommonlyused