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REGULATORYSCIENCEOFLIPOSOMEDRUGPRODUCTSDianeJ.Burgess,Ph.D. ProfessorofPharmaceuticsUniversityofConnecticutOfficeofTestingandResearchCDER,FDABurgess,June28,20011REGULATORYSCIENCEOFLIPOSOMEOutlineWhatareliposomes?Whataretheyusedfor?Whatdrugs?Whyliposomes?LiposomeformulationLiposomecharacterizationSafetyconcernsPerformanceconcernsInvitroreleasetestingstabilityBurgess,June28,20012OutlineWhatareliposomes?BurgOutlineContinuedPurposeofinvitroreleasetests?DesignofinvitroreleasetestAccelerated/stresstestsMethodvariablesaffectingreleaseMethodsunderdevelopmentInvivofactorsaffectingreleaseInvivodataandmodels?IVIVC?ResearchproposalBurgess,June28,20013OutlineContinuedPurposeofinLIPOSOMESLiposomesarecolloidal,lipidvesiclesconsistingofoneormoreself-assembledlipidbilayersenclosingasimilarnumberofaqueouscompartments.Lipids,suchaslecithin(diacylphosphatidylcholine),areamphiphilicmolecules.Duetothebulkynonpolarpartofthemoleculetheydonotpackintosphericalmicellesinaqueousphasebutratherself-assembleintobilayerswhichtendtoself-closeatlowconcentrationsintosphericalstructures.

Burgess,June28,20014LIPOSOMESLiposomesarecolloidLIPOSOMESContd.

Liposomescanbesubcategorizedinto:Smallunilamellarvesicles(SUV),25to100nminsize thatconsistofasinglelipidbilayerLargeunilamellarvesicles(LUV),100to400nminsize thatconsistofasinglelipidbilayerMultilamellarvesicles(MLV),200nmtoseveral microns,thatconsistoftwoormoreconcentric bilayersVesiclesabove1μmareknownasgiantvesicles.Burgess,June28,20015LIPOSOMESContd.LiposomescanLiposomesLocalizedandratecontrolleddelivery:

ImprovedtherapeuticresponseAchieveappropriatetissueorbloodlevelsReducedadversereactionsLessdrugadministeredTargeteddrugreleaseLowerdosingfrequency ImprovedpatientcomplianceSimplerdosingregimensLowercostperdoseUtilizationofotherwiseun-useablecompoundsPoorlysolubledrugsBurgess,June28,20016LiposomesLocalizedandratecoDrugCandidateSelectionKnowntherapeuticswithcleartoxicityandpharmacokineticprofilesPotentcompoundsNot“NarrowTherapeuticIndex”drugsProblemsassociatedwiththecurrentdosageformsFirstpasseffectsorpoorabsorptionGastricirritationRapidclearanceMedicalneedforimproveddeliveryDrugscompatiblewithmanufacturingconditionsBurgess,June28,20017DrugCandidateSelectionKnown.

APPROVEDLIPOSOMEPRODUCTS:Doxil Daunorubicin 1995Daunoxome Daunorubicin 1996Ambisome AmphotericinB 1997Depocyt Cytarabine 1999APPROVEDLIPIDCOMPLEXPRODUCTS:Ambelcet AmphotericinB 1995Amphotec AmphotericinB 1997Burgess,June28,20018. APPROVEDLIPOSOMEPRODUCTS:BSELECTIONOFDELIVERYSYSTEM

Liposomes–targeteddelivery.Theycandeliveragentsdirectlyintocells.Routes:i.v.,s.c.,i.m.,topical,pulmonary

Microspheres

-canprovidecontinuousdrugdeliveryoverperiodsofmonthstoyears.Systemicandlocalized.i.m.,s.c.,oral,pulmonary

Emulsions-canbeusedtomakehighlywaterinsolublecompoundsbioavailable.i.v.,oral,topicalBurgess,June28,20019SELECTIONOFDELIVERYSYSTEM LLIPOSOMEFORMULATION

LIPOSOMES

Liposomalcompositiondeterminestheproperties(e.g.surfacecharge,rigidityandstericinteractions)andtheinvitroandinvivoperformance. Bothwatersolubleandwaterinsolubledrugsmaybeencapsulated

Processingmethodsaffectparticlesize,percentagedrugentrapment,stabilityandreleaseratesBurgess,June28,200110LIPOSOMEFORMULATION LIPOSOMESLIPOSOMEFORMULATION

Processingmethods: Extrusion,ultrasonicationandmicrofluidizationforhydrophobicdrugsand Reversedphaseandfreeze-thawforhydrophilicdrugs.Burgess,June28,200111LIPOSOMEFORMULATION BurgessLiposomes:FactorsAffectingPerformance

ReleaseRateandStability

Phasetransitiontemperature(Tg)effectsmembranechangesfromorderedsolidtodisorderedfluidandisdependentonthelengthanddegreeofsaturationofthehydrocarbonchains.

Cholesterol-disorderingoftheorderedphaseandorderingofthedisorderedphaseeventuallyleadingtoaneliminationofthephasetransition.Highstabilityandlowleakage

Surfacechargeandstericinteraction:REStargeting/avoidingRESuptake

Burgess,June28,200112Liposomes:FactorsAffectingPTypesofLiposomes

ConventionalLiposomes

PreparedformnaturalneutralandanioniclipidsandhavenonspecificinteractionswiththeirenvironmentRelativelyunstable,havelowcarryingcapacities,andtendtobe“leaky”toentrappeddrugsubstancesMayliterallyfallapartoncontactwithplasma,particularlythoseofhighfluidity,CholeterolisoftenaddedtoincreaseplasmastabilityBurgess,June28,200113TypesofLiposomes ConventionTypesofLiposomes

Non-conventionalLiposomes

Smallsized(≤100nm),surfacemodifiedtoovercomesomeoftheshortcomingsofconventionalliposomesModifiedtoreducenegativecharge,decreasefluidityandcausesterichinderancetophagocytosisPropertiesaltered(e.g.byincorporationofcholesterol)Polymerizedliposomesmorestableandless“leaky”Polyetheyleneglycol,“pegylated”liposomes,avoiduptakebythemononuclearphagocyticcellsBurgess,June28,200114TypesofLiposomes Non-convenTYPESOFLIPOSOMES

Targetspecificligands,suchasantibodies,immunoglobulins,lectinsandoligosaccharidesattachedtothesurfacetoactivelytargettospecificsitesinthebodyTargetingviaparticlesizeLiposomespreparedwithcationicandfusogeniclipidsarecurrentlybeingutilizedingenetherapytodeliverDNAintotargetcellsBurgess,June28,200115TYPESOFLIPOSOMES Burgess,JTYPESOFLIPOSOMES

Highlyreactiveliposomes-readilyundergophasetransitioninparticularsituationsensitivetopH,ions,heatandlightForexample,pH-sensitiveliposomescanundergophasetransitioninacidicconditionsresultinginincreasedmembranefluidityandlossofencapsulatedmaterialsBurgess,June28,200116TYPESOFLIPOSOMES Burgess,JCRITICALFACTORSINLIPOSOMEPREPARATIONJ

Particlesize Methodofmanufacture LipidtypesPhasetransitiontemperature Polymerization Interfacialcharge Stericstabilization SterilizationBurgess,June28,200117CRITICALFACTORSINLIPOSOMEPLiposomes:FactorsAffectingPerformance

Liposomepreparationscanbestored:frozen,inliquidformandasafreezedriedpowder.

Reconstitutionofliposomesmayaffectparticlesizeandsizedistribution.

Burgess,June28,200118Liposomes:FactorsAffectingPSAFETYCONCERNS:LIPOSOMEFORMULATION

Lipidtoxicity(RBClysis)Typeandconcentration%Lyso-lipidsPresenceofproteinandlipoproteinfornaturallipidsResidualsolventOverloadofRESParticlesize(tailabove1um)-BlockageofcapillariesSizeaffectsRESuptakeandtissuetargetingStability:shelf-liveandinvivoDosedumping(viaproteinbinding)SterilityBurgess,June28,200119SAFETYCONCERNS:LIPOSOMEFORMLIPOSOMECHARACTERIZATION

StabilIty–DrugLipidsLiposomePhasetransitiontemperaturePercentdrugloadingPercentfreedrugDrugreleaserate/stabilityParticlesizeMorphology(lamellarity)SterilityBurgess,June28,200120LIPOSOMECHARACTERIZATION BurgSTERILITY

Terminalsterilization?AsepticprocessingMustconsiderbothinternalandexternalsterilityBurgess,June28,200121STERILITY Burgess,June28,20STABILITY

ActiveInactives(especiallythelipids)LiposomeasawholeneedAnychangeinparticlesizecanaffecttargeting,RESuptake,safetyandefficacy.Invivostabilityofwholeliposomeisparticularlyimportantfortargetedliposomes,sincetheyshouldremainstableintheplasmawithoutlossofcontentsuntiluptakeatthetargetsite.Burgess,June28,200122STABILITY Burgess,June28,20LIPOSOMEDESTABILIZATION

ProteinbindingMembranefusionBurgess,June28,200123LIPOSOMEDESTABILIZATION BurgeDrugReleasefromLiposomes

Releaseprofilesareapplicationdependent.

TargetedliposomesshouldremainintactuntildeliveryatsiteOther(shorttermCRandsolubilization)releaseduringappropriatetimescale.ReleasecontrolledbyFluidity/stability(lipids/co-lipids)ConditionsensitivityoflipidsSizeMLVoraSUVPhysicochemicalpropertiesofdrugDrug/lipidinteractionBurgess,June28,200124DrugReleasefromLiposomes BuInVitroDrugRelease

Apparatus?Media?Samplingmethods?Testingintervals?Totalpercentrelease?NostandardmethodatpresentBurgess,June28,200125InVitroDrugRelease Burgess,LiposomePerformance–InVitroReleaseandStabilitySeparationofliposomesfromdissolutionmediacomplicatestestingCurrentUSPmethodsdesignedfororalandtransdermalroutesInvitrotestsneedtotakeintoaccounttheexpectedinvivoperformanceofliposomesBurgess,June28,200126LiposomePerformance–InVitrLiposomePerformance–InVitroReleaseandStabilityReleasetestforatargetedliposomewouldneedtoshow1)liposomeisstableuntiluptakeatthesite2)liposomereleasesdrugatthesite(basedonthemechanismofreleaseinvivo).ReleasetestforanimmediatereleaseliposomewouldneedtoshowDrugisreleasedimmediatelyinconditionsmimickinghumanplasma.

Burgess,June28,200127LiposomePerformance–InVitrCurrentMethodsofInVitroTestingofLiposomeSystemsMembraneDiffusionTechniqueSampleandSeparateTechniqueInSituTechniqueContinuousFlowTechniqueBurgess,June28,200128CurrentMethodsofInVitroTeDevelopmentofInVitroReleaseandStabilityMethodsforLiposomesPurpose:methodstobeusedinsettingregulatoryspecificationsfortheseproductsforqualitycontrol(QC)purposestodifferentiatebetween“good”and“bad”batches.

TestsdesignwillvarydependingontheintendedinvivoperformanceofliposomesBurgess,June28,200129DevelopmentofInVitroReleasPurposeofInVitroReleaseTest?QualitycontrolandsafetyevaluationBatchtobatchManufacturingprocesschangesSubstantiationoflabelclaimsEvaluationofpotentialdosedumpingAssessmentofinvivostability“Realtime”vsaccelerated/stresstestInvitro-invivocorrelationBurgess,June28,200130PurposeofInVitroReleaseTeDesignofInVitroReleaseMethodSelectmediaandapparatustoachievereproducibleresultsAttempttoovercomelimitationsofexistingmethodsMiniaturizemethods

PrepareformulationvariantswithdifferentinvivoperformanceTestformulationvariantsinvitroandinvivoModifyinvitrotestifnotdiscriminatoryDetermineinvivofactorsthateffectreleaseModifyinvitromethodstoobtainIVIVrelationshipBurgess,June28,200131DesignofInVitroReleaseMetAcceleratedInVitroReleaseMethodsThesetestsshouldbepredictiveof“realtime”invitrotestsDrugreleasemechanismshouldnotbealteredAcceleratedtestshouldnotsimplydissolvetheliposomeBurgess,June28,200132AcceleratedInVitroReleaseMMediaandMethodsthatcanaffectReleaseSolventspHTemperatureAgitationEnzymesCellcultureSinkconditionsVolumeSampli